U.S. patent application number 14/091192 was filed with the patent office on 2014-03-27 for methods of wound care and treatment.
This patent application is currently assigned to Revalesio Corporation. The applicant listed for this patent is Revalesio Corporation. Invention is credited to Gregory J. Archambeau, Richard L. Watson, Anthony B. Wood.
Application Number | 20140086963 14/091192 |
Document ID | / |
Family ID | 39325448 |
Filed Date | 2014-03-27 |
United States Patent
Application |
20140086963 |
Kind Code |
A1 |
Archambeau; Gregory J. ; et
al. |
March 27, 2014 |
METHODS OF WOUND CARE AND TREATMENT
Abstract
Particular embodiments disclosed herein relate to gas-enriched
fluids, methods of making the same, systems for making the same
and/or methods of treatment utilizing the gas-enriched fluids for
wound care related conditions and/or diseases. In certain
embodiments, the gas-enriched fluid is oxygen-enriched water.
Certain embodiments relate to cosmetic and/or therapeutic fluids
and/or methods of treatment utilizing the fluids in order to treat
a cosmetic and/or therapeutic symptom of wound care and/or increase
proper wound healing.
Inventors: |
Archambeau; Gregory J.;
(Puyallup, WA) ; Watson; Richard L.; (Ruston,
WA) ; Wood; Anthony B.; (Tacoma, WA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Revalesio Corporation |
Tacoma |
WA |
US |
|
|
Assignee: |
Revalesio Corporation
Tacoma
WA
|
Family ID: |
39325448 |
Appl. No.: |
14/091192 |
Filed: |
November 26, 2013 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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11978137 |
Oct 25, 2007 |
8597689 |
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14091192 |
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60862904 |
Oct 25, 2006 |
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60862953 |
Oct 25, 2006 |
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60862955 |
Oct 25, 2006 |
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60862959 |
Oct 25, 2006 |
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60982387 |
Oct 24, 2007 |
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Current U.S.
Class: |
424/400 ;
424/613 |
Current CPC
Class: |
A61K 45/06 20130101;
A61K 9/0014 20130101; B01F 3/04531 20130101; A61P 31/04 20180101;
A61K 9/0048 20130101; B01F 7/00816 20130101; A61P 27/02 20180101;
A61P 41/00 20180101; A61P 43/00 20180101; B01F 3/0807 20130101;
A61P 37/08 20180101; B01F 3/0853 20130101; B29D 11/00038 20130101;
A61K 33/14 20130101; A61P 27/04 20180101; B29D 11/00096 20130101;
A61P 17/02 20180101; A61K 33/00 20130101; A61K 33/00 20130101; A61K
2300/00 20130101; A61K 33/14 20130101; A61K 2300/00 20130101 |
Class at
Publication: |
424/400 ;
424/613 |
International
Class: |
A61K 33/00 20060101
A61K033/00 |
Claims
1. A method for treating a wound to a surface tissue, comprising
contacting a wound of a subject in need thereof with an effective
amount of an electrokinetically altered oxygenated aqueous fluid
comprising an ionic aqueous solution of charge-stabilized
oxygen-containing nanostructures having an average diameter of less
than 100 nanometers and stably configured in the ionic aqueous
fluid an amount sufficient to promote wound healing.
2. The method of claim 1 wherein the wound is selected from the
group consisting of: lacerations, abrasions, rupture, puncture
wounds, chemical, thermal, or radiation-induced burns, cuts,
scrapes, incisions, blisters, diabetic ulcers, bedsores or pressure
ulcers, skin grafts, and surgical wounds.
3. The method of claim 1, wherein promoting wound healing comprises
treatment of at least one wound healing property selected from the
group consisting of: epidmeral or dermal layering, cellular
migration, collagen deposition at the wound, neoangiogenesis at the
wound, elastin deposition at the wound, inflammation at the wound,
expression of proteoglycans or glycosaminoglycans, cellular
proliferation, and hyaluronic acid concentration at the wound, is
afforded.
4. The method of claim 3 wherein the electrokinetically altered
oxygenated aqueous fluid accelerates epidermal or dermal
layering.
5. The method of claim 3 wherein the electrokinetically altered
oxygenated aqueous fluid increases cellular migration of at least
one type of cell to the wound.
6. The method of claim 5 wherein the type of cellular migration or
proliferation comprises at least one cell selected from the group
consisting of: keratinocytes, fibroblasts, epidermal cells, dermal
cells, epithelial cells, mast cells, neutrophils, lymphocytes, and
macrophages.
7. The method of claim 3 wherein the electrokinetically altered
oxygenated aqueous fluid accelerates neoangiogenesis of blood
vessels or lymphatic vessels.
8. The method of claim 3 wherein the electrokinetically altered
oxygenated aqueous fluid increases collagen deposition at the
wound.
9. The method of claim 1, wherein the electrokinetically altered
oxygenated aqueous fluid is contacted to the wound by way of a
wound dressing.
10. A method for treating infection in a wound to a surface tissue,
comprising contacting a wound of a subject in need thereof with an
effective amount of an electrokinetically altered oxygenated
aqueous fluid comprising an ionic aqueous solution of
charge-stabilized oxygen-containing nanostructures predominantly
having an average diameter of less than 100 nanometers and stably
configured in the ionic aqueous fluid an amount sufficient to
reduce growth of at least one microbe in the wound.
11. The method of claim 10 wherein the microbe comprises
Pseudomonas.
12. A method for decreasing scarring in a wound to a surface tissue
comprising contacting a wound of a subject in need thereof with an
effective amount of an electrokinetically altered oxygenated
aqueous fluid comprising an ionic aqueous solution of
charge-stabilized oxygen-containing nanostructures predominantly
having an average diameter of less than 100 nanometers and stably
configured in the ionic aqueous fluid an amount sufficient to
reduce scarring.
13. The method of claim 12 wherein the electrokinetically altered
oxygenated aqueous fluid further increases the level of chemokines
at the wound.
14. A method for increasing or decreasing nitric oxide production
or degradation at a wound to a surface tissue comprising contacting
a wound of a subject in need thereof with an effective amount of an
electrokinetically altered oxygenated aqueous fluid comprising an
ionic aqueous solution of charge-stabilized oxygen-containing
nanostructures predominantly having an average diameter of less
than 100 nanometers and stably configured in the ionic aqueous
fluid an amount sufficient to promote increasing or decreasing
nitric oxide production or degradation at a wound, wherein an
increased or decreased level of nitric oxide is afforded.
15. The method of claim 12, wherein the electrokinetically altered
oxygenated aqueous fluid alters at least one wound healing property
selected from the group consisting of: cellular migration of at
least one type of cell to the wound, collagen deposition at the
wound, neoangiogenesis at the wound, elastin deposition at the
wound, expression of proteoglycans or glycosaminoglycans, and
hyaluronic acid concentration at the wound.
16. The method of any one of claims 1, 10, 12, and 14, wherein the
electrokinetically altered oxygenated aqueous fluid comprises
electrokinetically modified or charged oxygen species.
17. The method of claim 16, wherein the electrokinetically modified
or charged oxygen species are present in an amount of at least 0.5
ppm, at least 1 ppm, at least 3 ppm, at least 5 ppm, at least 7
ppm, at least 10 ppm, at least 15 ppm, or at least 20 ppm.
18. The method of any one of claims 1, 10, 12, and 14, wherein the
electrokinetically altered oxygenated aqueous fluid comprises
solvated electrons stabilized by molecular oxygen.
19. The method of claim 18, wherein the solvated electrons are
present in an amount of at least 0.01 ppm, at least 0.1 ppm, at
least 0.5 ppm, at least 1 ppm, at least 3 ppm, at least 5 ppm, at
least 7 ppm, at least 10 ppm, at least 15 ppm, or at least 20
ppm.
20. The method of claim 1, wherein the oxygen in the fluid is
present in an amount of at least 14 ppm, at least 15, ppm, at least
25 ppm, at least 30 ppm, at least 40 ppm, at least 50 ppm, or at
least 60 ppm oxygen at atmospheric pressure.
21. The method of claim 12, wherein the electrokinetically altered
oxygenated aqueous fluid is contacted to the wound by way of a
wound dressing.
22. The method of claim 1, wherein at least 90% of oxygen present
in the electrokinetically-altered aqueous fluid composition is in
the charge-stabilized oxygen-containing nanostructures.
23. The method of claim 10 wherein at least 90% of oxygen present
in the electrokinetically-altered aqueous fluid composition is in
the charge-stabilized oxygen-containing nanostructures.
24. The method of claim 12, wherein at least 90% of oxygen present
in the electrokinetically-altered aqueous fluid composition is in
the charge-stabilized oxygen-containing nanostructures.
25. The method of claim 14, wherein at least 90% of oxygen present
in the electrokinetically-altered aqueous fluid composition is in
the charge-stabilized oxygen-containing nanostructures.
26. The method of claim 1, wherein a majority of the
charge-stabilized oxygen-containing nanostructures have an average
diameter of less than 100 nanometers.
Description
CROSS-REFERENCE TO RELATED APPLICATIONS
[0001] This application is a continuation of U.S. patent
application Ser. No. 11/978,137, filed on Oct. 25, 2007 and
entitled METHODS OF WOUND CARE AND TREATMENT (issuing on Dec. 3,
2013 as U.S. Pat. No. 8,597,689), which claims priority to U.S.
Provisional Patent Application Ser. No. 60/862,904, filed Oct. 25,
2006 and entitled DIFFUSER/EMULSIFIER, No. 60/862,953, filed Oct.
25, 2006 and entitled METHOD OF THERAPEUTIC TREATMENTS OF EYES AND
OTHER HUMAN TISSUES USING AN OXYGENATED SOLUTION, No. 60/862,955,
filed Oct. 25, 2006 and entitled OXYGENATED SALINE SOLUTION, No.
60/862,959, filed Oct. 25, 2006 and entitled WOUND TREATMENT WITH
INFUSION MATERIALS and No. 60/982,387, filed Oct. 24, 2007 and
entitled MIXING DEVICE, all of which are incorporated by reference
herein in their entirety.
FIELD OF THE INVENTION
[0002] Disclosed herein are gas-enriched fluid compositions,
methods of making, and methods of using the same for general wound
care and treatment. In particular embodiments, the gas-enriched
fluid comprises an oxygen-enriched fluid.
SEQUENCE LISTING
[0003] A Sequence Listing comprising SEQ ID NOS:1-2, has been
provided in computer readable form (.txt) as part of this
application, and is incorporated by reference herein in its
entirety as part of this application.
BACKGROUND OF THE INVENTION
[0004] Wound care is desirable in order to improve health and
appearance of the outer epidermis, as well as underlying dermal and
other tissues. Wounds, either injury induced (such as cuts,
abrasions, blisters, etc.), or surgically induced (such as surgical
incisions, astomies, etc.) require localized treatment to remedy
the affected area and prevent further dermal damage. If wounds are
not properly treated, further dermal irritation can result,
including secondary infections and further discomfort to the
subject.
[0005] Improper wound care and/or wound healing result in greater
than 100 billion dollars per year, and may require antibiotic use,
hospitalizations, and great pain or discomfort to the subject.
Present methods of therapeutic wound care are inadequate and
routinely include washing with soap and water and/or applying an
antiseptic, but still may lead to infection and/or improper wound
healing and repair. Thus, novel therapeutic methods and
compositions for wound care and wound healing are needed.
SUMMARY OF THE INVENTION
[0006] Particular embodiments disclosed herein relate to methods
for treating a wound to a surface tissue, comprising contacting a
wound for a sufficient amount of time and with an effective amount
of a gas-enriched fluid comprising solvated electrons, wherein the
gas-enriched fluid alters a wound healing property selected from
the group consisting of: epidmeral or dermal layering, cellular
migration, collagen deposition at the wound, neoangiogenesis at the
wound, elastin deposition at the wound, chemokine levels at the
wound, inflammation at the wound, expression of proteoglycans or
glycosaminoglycans, cellular proliferation, and hyaluronic acid
concentration at the wound, when compared with an untreated wound
or a wound treated with non-gas enriched fluid. In particular
embodiments, the wound is selected from the group consisting of:
lacerations, abrasions, rupture, puncture wounds, chemical,
thermal, or radiation-induced burns, cuts, scrapes, incisions,
blisters, diabetic ulcers, bedsores or pressure ulcers, skin
grafts, and surgical wounds.
[0007] In certain embodiments, the gas-enriched fluid accelerates
epidermal or dermal layering compared with an untreated wound or a
wound treated with non-gas enriched fluid. In other embodiments,
the gas-enriched fluid increases cellular migration of at least one
type of cell to the wound. In still other embodiments, the type of
cellular migration or proliferation comprises at least one cell
selected from the group consisting of: keratinocytes, fibroblasts,
epidermal cells, dermal cells, epithelial cells, mast cells,
neutrophils, lymphocytes, and macrophages.
[0008] In some particular embodiments of the present invention, the
gas-enriched fluid accelerates neoangiogenesis of blood vessels or
lymphatic vessels compared with an untreated wound or wound treated
with non-gas enriched fluid. In still other embodiments, the
gas-enriched fluid increases collagen deposition at the wound
compared with an untreated wound or a wound treated with non-gas
enriched fluid.
[0009] Certain embodiments described herein relate to methods for
preventing infection in a wound to a surface tissue comprising
contacting a wound for a sufficient amount of time and with an
effective amount of a gas-enriched fluid comprising solvated
electrons, wherein the gas-enriched fluid reduces growth of at
least one microbe in the wound. In certain embodiments, the microbe
comprises Pseudomonas.
[0010] Other particular embodiments disclosed herein relate to
methods for decreasing scarring in a wound to a surface tissue
comprising contacting a wound for a sufficient amount of time and
with an effective amount of a gas-enriched fluid comprising
solvated electrons, wherein the gas-enriched fluid alters a wound
healing property selected from the group consisting of: cellular
migration of at least one type of cell to the wound, collagen
deposition at the wound, neoangiogenesis at the wound, elastin
deposition at the wound, expression of proteoglycans or
glycosaminoglycans, and hyaluronic acid concentration at the wound,
thereby decreasing scarring. In certain embodiments, the
gas-enriched fluid further increases the level of chemokines at the
wound.
[0011] Certain other embodiments relate to methods for increasing
or decreasing nitric oxide production or degradation at a wound to
a surface tissue comprising contacting a wound for a sufficient
amount of time and with an effective amount of a gas-enriched fluid
comprising solvated electrons, wherein the wound demonstrates an
increased or decreased nitric oxide level compared with an
untreated wound or a wound treated with deionized saline.
[0012] In any of the aforementioned embodiments, the gas-enriched
fluid may be contacted to the wound by way of a wound dressing.
Furthermore, in any of the aforementioned embodiments, the
gas-enriched fluid comprises oxygen-enriched fluid. In any of the
embodiments disclosed herein, the gas-enriched fluid contains
diffused or dissolved gas at a level of greater than about 15 parts
per million at atmospheric pressure, and further comprises solvated
electrons.
[0013] Certain embodiments herein relate to a therapeutic
composition comprising a gas-enriched fluid and a therapeutic agent
selected from the group consisting of: anti-microbial agent,
cytokines, salts, vasodilators, vasoconstrictors, lubricants,
chemokines, and any combination thereof.
[0014] Particular aspects provide a composition, comprising an
electrokinetically altered oxygenated aqueous fluid or solution,
wherein the oxygen in the fluid or solution is present in an amount
of at least 25 ppm, at least 30 ppm, at least 40 ppm, at least 50
ppm, or at least 60 ppm oxygen. In particular embodiments, the
electrokinetically altered oxygenated aqueous fluid or solution
comprises electrokinetically modified or charged oxygen species. In
certain aspects, the electrokinetically modified or charged oxygen
species are present in an amount of at least 0.5 ppm, at least 1
ppm, at least 3 ppm, at least 5 ppm, at least 7 ppm, at least 10
ppm, at least 15 ppm, or at least 20 ppm. In particular
embodiments, the electrokinetically altered oxygenated aqueous
fluid or solution comprises solvated electrons stabilized by
molecular oxygen. In certain aspects, the solvated electrons are
present in an amount of at least 0.01 ppm, at least 0.1 ppm, at
least 0.5 ppm, at least 1 ppm, at least 3 ppm, at least 5 ppm, at
least 7 ppm, at least 10 ppm, at least 15 ppm, or at least 20 ppm.
In particular embodiments, the fluid or solution facilitates
oxidation of pyrogallol to purpurogallin in the presence of
horseradish peroxidase enzyme (HRP) in an amount above that
afforded by a control pressure pot generated or fine-bubble
generated aqueous fluid or solution having an equivalent dissolved
oxygen level, and wherein there is no hydrogen peroxide, or less
than 0.1 ppm of hydrogen peroxide present in the electrokinetic
oxygen-enriched aqueous fluid or solution. In certain aspects, the
facilitation of oxidation of pyrogallol to purpurogallin persists
for at least three hours in an open container, or for at least two
months in a closed gas-tight container.
[0015] Additional aspects provide a composition, comprising an
electrokinetically altered oxygenated aqueous fluid or solution,
wherein the fluid or solution comprises at least 25 ppm, at least
30 ppm, at least 40 ppm, at least 50 ppm, or at least 60 ppm
oxygen, wherein the fluid or solution facilitates oxidation of
pyrogallol to purpurogallin in the presence of horseradish
peroxidase enzyme (HRP) in an amount above that afforded by a
control pressure pot generated or fine-bubble generated aqueous
fluid or solution having an equivalent dissolved oxygen level, and
wherein there is no hydrogen peroxide, or less than 0.1 ppm of
hydrogen peroxide present in the electrokenetic oxygen-enriched
aqueous fluid or solution. In particular embodiments, the
facilitation of oxidation of pyrogallol to purpurogallin persists
for at least three hours in an open container, or for at least two
months in a closed gas-tight container. In certain aspects, the
oxygenated aqueous fluid or solution comprises solvated electrons
stabilized by molecular oxygen. In particular embodiments, the
solvated electrons are present in an amount of at least 0.01 ppm,
at least 0.1 ppm, at least 0.5 ppm, at least 1 ppm, at least 3 ppm,
at least 5 ppm, at least 7 ppm, at least 10 ppm, at least 15 ppm,
or at least 20 ppm.
[0016] Further aspects provide a method of producing an
electrokinetically altered oxygenated aqueous fluid or solution,
comprising: providing a flow of a fluid material between two spaced
surfaces in relative motion and defining a mixing volume
therebetween, wherein the dwell time of a single pass of the
flowing fluid material within and through the mixing volume is
greater than 0.06 seconds or greater than 0.1 seconds; and
introducing oxygen (O.sub.2) into the flowing fluid material within
the mixing volume under conditions suitable to dissolve at least 20
ppm, at least 25 ppm, at least 30 ppm, at least 40 ppm, at least 50
ppm, or at least 60 ppm oxygen into the material, and
electrokinetically alter the fluid or solution. In certain aspects,
the oxygen is infused into the material in less than 100
milliseconds, less than 200 milliseconds, less than 300
milliseconds, or less than 400 milliseconds. In particular
embodiments, the ratio of surface area to the volume is at least
12, at least 20, at least 30, at least 40, or at least 50.
[0017] Yet further aspects, provide a method of producing an
electrokinetically altered oxygenated aqueous fluid or solution,
comprising: providing a flow of a fluid material between two spaced
surfaces defining a mixing volume therebetween; and introducing
oxygen into the flowing material within the mixing volume under
conditions suitable to infuse at least 20 ppm, at least 25 ppm, at
least 30 ppm, at least 40 ppm, at least 50 ppm, or at least 60 ppm
oxygen into the material in less than 100 milliseconds, less than
200 milliseconds, less than 300 milliseconds, or less than 400
milliseconds. In certain aspects, the dwell time of the flowing
material within the mixing volume is greater than 0.06 seconds or
greater than 0.1 seconds. In particular embodiments, the ratio of
surface area to the volume is at least 12, at least 20, at least
30, at least 40, or at least 50.
[0018] Additional embodiments provide a method of producing an
electrokinetically altered oxygenated aqueous fluid or solution,
comprising use of a mixing device for creating an output mixture by
mixing a first material and a second material, the device
comprising: a first chamber configured to receive the first
material from a source of the first material; a stator; a rotor
having an axis of rotation, the rotor being disposed inside the
stator and configured to rotate about the axis of rotation therein,
at least one of the rotor and stator having a plurality of
through-holes; a mixing chamber defined between the rotor and the
stator, the mixing chamber being in fluid communication with the
first chamber and configured to receive the first material
therefrom, and the second material being provided to the mixing
chamber via the plurality of through-holes formed in the one of the
rotor and stator; a second chamber in fluid communication with the
mixing chamber and configured to receive the output material
therefrom; and a first internal pump housed inside the first
chamber, the first internal pump being configured to pump the first
material from the first chamber into the mixing chamber. In certain
aspects, the first internal pump is configured to impart a
circumferential velocity into the first material before it enters
the mixing chamber.
[0019] Further embodiments provide a method of producing an
electrokinetically altered oxygenated aqueous fluid or solution,
comprising use of a mixing device for creating an output mixture by
mixing a first material and a second material, the device
comprising: a stator; a rotor having an axis of rotation, the rotor
being disposed inside the stator and configured to rotate about the
axis of rotation therein; a mixing chamber defined between the
rotor and the stator, the mixing chamber having an open first end
through which the first material enters the mixing chamber and an
open second end through which the output material exits the mixing
chamber, the second material entering the mixing chamber through at
least one of the rotor and the stator; a first chamber in
communication with at least a majority portion of the open first
end of the mixing chamber; and a second chamber in communication
with the open second end of the mixing chamber.
[0020] Additional aspects provide an electrokinetically altered
oxygenated aqueous fluid or solution made according to any of the
above methods.
BRIEF DESCRIPTION OF THE SEVERAL VIEWS OF THE DRAWING(S)
[0021] FIG. 1 is a partial cross-section, partial block diagram of
a prior art mixing device.
[0022] FIG. 2 is block diagram of an exemplary embodiment of a
mixing device.
[0023] FIG. 3 is an illustration of an exemplary system for
delivering a first material to the mixing device of FIG. 2.
[0024] FIG. 4 is a fragmentary partial cross-sectional view of a
top portion of the mixing device of FIG. 2.
[0025] FIG. 5 is a fragmentary cross-sectional view of a first side
portion of the mixing device of FIG. 2.
[0026] FIG. 6 is a fragmentary cross-sectional view of a second
side portion of the mixing device of FIG. 2.
[0027] FIG. 7 is a fragmentary cross-sectional view of a side
portion of the mixing device of FIG. 2 located between the first
side portion of FIG. 5 and the second side portion of FIG. 6.
[0028] FIG. 8 is a perspective view of a rotor and a stator of the
mixing device of FIG. 2.
[0029] FIG. 9 is a perspective view of an inside of a first chamber
of the mixing device of FIG. 2.
[0030] FIG. 10 is a fragmentary cross-sectional view of the inside
of a first chamber of the mixing device of FIG. 2 including an
alternate embodiment of the pump 410.
[0031] FIG. 11 is a perspective view of an inside of a second
chamber of the mixing device of FIG. 2.
[0032] FIG. 12 is a fragmentary cross-sectional view of a side
portion of an alternate embodiment of the mixing device.
[0033] FIG. 13 is a perspective view of an alternate embodiment of
a central section of the housing for use with an alternate
embodiment of the mixing device.
[0034] FIG. 14 is a fragmentary cross-sectional view of an
alternate embodiment of a bearing housing for use with an alternate
embodiment of the mixing device.
[0035] FIG. 15 is a cross-sectional view of the mixing chamber of
the mixing device of FIG. 2 taken through a plane orthogonal to the
axis of rotation depicting a rotary flow pattern caused by
cavitation bubbles when a through-hole of the rotor approaches (but
is not aligned with) an aperture of the stator.
[0036] FIG. 16 is a cross-sectional view of the mixing chamber of
the mixing device of FIG. 2 taken through a plane orthogonal to the
axis of rotation depicting a rotary flow pattern caused by
cavitation bubbles when the through-hole of the rotor is aligned
with the aperture of the stator.
[0037] FIG. 17 is a cross-sectional view of the mixing chamber of
the mixing device of FIG. 2 taken through a plane orthogonal to the
axis of rotation depicting a rotary flow pattern caused by
cavitation bubbles when a through-hole of the rotor that was
previously aligned with the aperture of the stator is no longer
aligned therewith.
[0038] FIG. 18 is a side view of an alternate embodiment of a
rotor.
[0039] FIG. 19 is an enlarged fragmentary cross-sectional view
taken through a plane orthogonal to an axis of rotation of the
rotor depicting an alternate configuration of through-holes formed
in the rotor and through-holes formed in the stator.
[0040] FIG. 20 is an enlarged fragmentary cross-sectional view
taken through a plane passing through and extending along the axis
of rotation of the rotor depicting a configuration of through-holes
formed in the rotor and through-holes formed in the stator.
[0041] FIG. 21 is an enlarged fragmentary cross-sectional view
taken through a plane passing through and extending along the axis
of rotation of the rotor depicting an alternate offset
configuration of through-holes formed in the rotor and
through-holes formed in the stator.
[0042] FIG. 22 is an illustration of a shape that may be used to
construct the through-holes of the rotor and/or the apertures of
the stator.
[0043] FIG. 23 is an illustration of a shape that may be used to
construct the through-holes of the rotor and/or the apertures of
the stator.
[0044] FIG. 24 is an illustration of a shape that may be used to
construct the through-holes of the rotor and/or the apertures of
the stator.
[0045] FIG. 25 is an illustration of a shape that may be used to
construct the through-holes of the rotor and/or the apertures of
the stator.
[0046] FIG. 26 is an illustration of an electrical double layer
("EDL") formed near a surface.
[0047] FIG. 27 is a perspective view of a model of the inside of
the mixing chamber.
[0048] FIG. 28 is a cross-sectional view of the model of FIG.
27.
[0049] FIG. 29 is an illustration of an experimental setup.
[0050] FIG. 30 illustrates dissolved oxygen levels in water
processed with oxygen in the mixing device of FIG. 2 and stored a
500 ml thin walled plastic bottle and a 1,000 ml glass bottle each
capped at 65.degree. Fahrenheit.
[0051] FIG. 31 illustrates dissolved oxygen levels in water
processed with oxygen in the mixing device of FIG. 2 and stored in
a 500 ml plastic thin walled bottle and a 1,000 ml glass bottle
both refrigerated at 39.degree. Fahrenheit.
[0052] FIG. 32 illustrates the dissolved oxygen levels in
GATORADE.RTM. processed with oxygen in the mixing device of FIG. 2
and stored in 32 oz. GATORADE.RTM. bottles having an average
temperature of 55.degree. Fahrenheit.
[0053] FIG. 33 illustrates the dissolved oxygen retention of a 500
ml braun balanced salt solution processed with oxygen in the mixing
device of FIG. 2.
[0054] FIG. 34 illustrates a further experiment wherein the mixing
device of FIG. 2 is used to sparge oxygen from water by processing
the water with nitrogen in the mixing device of FIG. 2.
[0055] FIG. 35 illustrates the sparging of oxygen from water by the
mixing device of FIG. 2 at standard temperature and pressure.
[0056] FIG. 36 is an illustration of a nanocage.
[0057] FIGS. 37A and B illustrate Rayleigh scattering effects of an
oxygen-enriched fluid;
[0058] FIG. 38 illustrates DNA thermostability of one embodiment of
the inventive fluid;
[0059] FIG. 39A illustrates the pyrogallol/horseradish peroxidase
reactivity test at initial point of fluid production, room
temperature;
[0060] FIG. 39B illustrates the pyrogallol/horseradish peroxidase
reactivity test at 30 minutes, room temperature;
[0061] FIG. 39C illustrates the pyrogallol/horseradish peroxidase
reactivity test at 2 hours, room temperature;
[0062] FIG. 39D illustrates the pyrogallol/horseradish peroxidase
reactivity test at 3 hours, room temperature;
[0063] FIG. 39E illustrates the pyrogallol/horseradish peroxidase
reactivity test with various gases enriched in the fluid, according
to particular embodiments;
[0064] FIG. 40 illustrates the results of the glutathione
peroxidase test, verifying the absence of hydrogen peroxide;
[0065] FIG. 41 illustrates the results of pyrogallol/horseradish
peroxidase reactivity assay with oxygen-enriched inventive fluid,
deionized water (-) control, and hydrogen peroxide (+) control;
[0066] FIG. 42 illustrates the cytokine profile of a mitogenic
assay in the presence of a gas-enriched fluid and deionized control
fluid;
[0067] FIG. 43 illustrates the difference in the growth rates of
Pseudomonas bacteria at various dissolved oxygen saturation
ratios;
[0068] FIGS. 44a and 44b illustrate in vitro healing of wounds
using an oxygen-enriched cell culture media and a non-gas-enriched
media;
[0069] FIGS. 45a through 45f show histological cross-sections of
dermal and epidermal in vivo wound healing;
[0070] FIG. 46 illustrates the expression of Hale's stain in
treated and control healing wounds, used to detect acid
mucopolysaccharides, such as hyaluronic acid;
[0071] FIG. 47 illustrates the expression of von Willebrand's
Factor stain used to detect angiogenesis in treated and control
healing wounds;
[0072] FIG. 48 illustrates the expression of Luna's stain used to
detect elastin in treated and control healing wounds;
[0073] FIG. 49 illustrates the number of mast cells per visual
field for treated and control healing wounds;
[0074] FIG. 50 illustrates the percentage of dead cells at separate
time points in a corneal fibroblast assay using inventive
gas-enriched culture media and control culture media,
[0075] FIG. 51 illustrates the shelf life of the inventive
gas-enriched fluid in a polymer pouch; and
[0076] FIG. 52 illustrates the results of contacting splenocytes
with MOG in the presence of pressurized pot oxygenated fluid (1),
inventive gas-enriched fluid (2), or control deionized fluid
(3).
DETAILED DESCRIPTION
[0077] The present invention includes compositions and methods for
promoting proper wound healing and/or wound care. The compositions
and/or methods disclosed herein may be used on a variety of wounds,
wherein a wound is a break in the skin or other tissue or organ,
such as lacerations, cuts, scrapes, blisters, abrasions, burns,
diabetic ulcers, bedsores, surgical wounds, and other wounds.
Healing is a biological response to the injury that occurs
typically with some scarring (with the exception of bone wounds).
Thus, one of the objects of proper wound care is to minimize the
possibility of infection and/or scarring.
[0078] The present medical practice is typically to wash or
irrigate the wound with a stream of sterile saline, soap and water,
providone/iodine, and/or other antiseptic antibiotic or anesthetic
solutions. Typically, irrigating fluid is splashed onto the wound
surface with an amount of force sufficient to dislodge dirt,
debris, bacteria or dead tissue. The goal of wound irrigation is to
remove debris and bacteria from the wound while minimizing injury
to the normal tissue around the wound. This is difficult for deep
penetrating wounds, such as those associated with a puncture wound
from a stick or those from an animal bite. In the case of deep
puncture wounds, and especially animal bites, it is especially
important to thoroughly clean the wound due to the debris and
microbial introduction deep into the tissue. These types of wounds
typically require daily care under the present modes of wound care.
Additionally, chronic wounds are common in subjects who are
confined to a bed, wheelchair, or are otherwise sedentary or
immobile, who have a nerve and/or blood circulation disorder or
condition that impairs wound healing (such as those suffering from
diabetes, cancer, HIV/AIDS, peripheral neuropathy,
immunosuppression, sickle cell anemia, those who are or have been
receiving steroids and/or chemotherapy drugs, those who are
advanced in age, and others).
[0079] In certain situations, it may be necessary to suture the
wound. If the wound requires sutures (stitches, staples, skin glue,
tape, etc.), it may be cleansed, sutured, and dressed with
bandages.
[0080] The present inventive compositions and/or methods may
further be utilized for dressing treatment and/or wound irrigation.
In certain embodiments, the wound irrigation may be applied under a
pressure. In certain embodiments the wound irrigation pressure
comprises about 2 psi, about 5 psi, about 8 psi, about 10 psi,
about 12 psi, about 15 psi, about 18 psi, about 20 psi, or any
value therebetween.
[0081] In certain embodiments, the gas-enriched fluid compositions
and/or methods of the present invention relate to treating wound
dressings. Wound dressings remove excess exudate from the wound,
keeps the wound moist, and prevents dehydration, which can increase
the rate of wound healing. In the treatment of wounds, antibiotics
and/or antiseptics may be incorporated into dressings. In some
particular embodiments, the wound dressing incorporates a
biologically and/or chemically active substance (such as a cosmetic
and/or therapeutic agent) in a separate phase which is disbursed in
a continuous phase upon applying and/or over time. In this manner,
the biologically and/or chemically active substance can be applied
directly to the wound by way of the dressing. Application of the
biologically and/or chemically active substance can be delivered
continuous, all at once, or intermittent over time by way of the
wound dressing.
[0082] In the case of occlusive dressings for wounds such as
pressure sores (including bedsores), ulcers, or other wounds that
may be persistent or chronic, the dressing may be applied that
contains both an inner contact layer and an outer protecting layer.
In certain embodiments, the inner layer or barrier layer, contains
water absorptive material such as hydrocolloids, which allows fluid
from the wound to be absorbed and making it possible to keep the
dressing in place for at least several days.
[0083] In certain embodiments, the gas-enriched fluid compositions
and/or methods of the present invention promote wound healing
and/or reduce scarring, and/or reduce infection in wounds. In
certain embodiments, the gas-enriched fluid compositions and/or
methods of the present invention increase extracellular matrix
deposition and/or production; increase collagen deposition and/or
production; hylauronic acid deposition and/or production; and/or
elastin deposition and/or production. In other embodiments, the
gas-enriched fluid compositions and/or methods of the present
invention promote proliferation, differentiation, and/or apoptosis
of epithelial cells, keratinocytes, connective tissue, and/or
muscle cells or other related cells of a wound.
[0084] The gas-enriched compositions and/or methods disclosed
herein may promote any one or more stages of wound healing,
including decreasing bleeding, increasing narrowing of blood
vessels and/or clot formation, release of various chemical
substances into the wound; increase angiogenesis, increase
epithelialization of the wound, increase production of proteins
related to wound healing, increase remodeling and/or proliferation
and/or migration of epithelial cells and/or keratinocytes.
[0085] Inventive Gas-Enriched Fluids and Solutions
[0086] Diffusing or enriching a fluid with another fluid may result
in a solution or suspension of the two fluids. In particular,
enriching a liquid with a gas (e.g., oxygen) may be beneficial for
certain applications, including therapeutic treatments. As utilized
herein, "fluid," may generally refer to a liquid, a gas, a vapor, a
mixture of liquids and/or gases, or any combination thereof, for
any particular disclosed embodiment. Furthermore, in certain
embodiments a "liquid" may generally refer to a pure liquid or may
refer to a gel, sol, emulsion, fluid, colloid, dispersion, or
mixture, as well as any combination thereof; any of which may vary
in viscosity.
[0087] In particular preferred embodiments, the dissolved gas
comprises oxygen. In other particular embodiments, the dissolved
gas comprises nitric oxide. In other particular embodiments, the
gas comprises ambient air.
[0088] There are several art-recognized methods of gas-enriching
fluids (such as oxygenating water). For example, a turbine aeration
system can release air near a set of rotating blades of an
impeller, which mixes the air or oxygen with the water, or water
can be sprayed into the air to increase its oxygen content.
Additionally, other systems on the market inject air or oxygen into
the water and subject the water/gas to a large scale vortex.
Naturally occurring levels of oxygen in water are typically no more
than 10 ppm, which is considered to be a level of 100% dissolved
oxygen. Tests on certain devices have shown that under ideal
conditions, the device can attain upwards of approximately 20 ppm
(parts per million)), or twice the natural oxygen levels of water.
However, the water loses that high level of dissolved oxygen very
rapidly, and within minutes the water returns to having the
baseline of about 10 ppm dissolved oxygen.
[0089] In certain embodiments disclosed herein, a gas-enriched
fluid of the present invention provides a cosmetic and/or
therapeutic wound care benefit. Certain embodiments disclosed
herein relate to a cosmetic and/or therapeutic composition
comprising a gas-enriched fluid of the present invention, and
optionally at least one additional therapeutic agent, such as a
pharmaceutical drug, a metal, a peptide, a polypeptide, a protein,
a nucleotide, a carbohydrate or glycosylated protein, a fat
(including oils or waxes), or other agent that prevents or
alleviates at least one symptom of a condition or disease
associated with wounds or wound healing.
[0090] Compositions and methods are disclosed for treating a wound
in any particular organ and/or tissues in need thereof by
administering an effective amount of a composition comprising a
gas-enriched fluid. As used herein, "treat," "treating,"
"treatment," and any and all derivations thereof refer to using the
compositions of the present invention either prophylactically to
conditions or diseases related to wounds and/or wound healing, or
cosmetically or therapeutically to ameliorate an existing condition
or disease related to improper wound healing, or to increase proper
wound healing. In one particular embodiment, the gas-enriched fluid
of the present invention inhibits microbial growth. In another
particular embodiment, the gas-enriched fluid of the present
invention promotes apoptosis.
[0091] Microbial infections, particularly of Staphlycoccus,
Streptococcus, yeast, Serratia, E. coli, Pseudomonas aeruginosa,
and other microbial infections, can cause devastating infections in
wounds. Thus, in certain embodiments, the gas-enriched fluid
compositions and/or methods of the present invention include
anti-microbial agents, such as antifungal, antibiotic, or other
anti-microbial agents. Some examples of anti-microbial agents that
may be utilized with the gas-enriched fluid compositions and/or
methods include, but are not limited to, amikacin, gentamicin,
kanamycin, neomycin, netilmicin, streptomycin, tobramycin,
paromomycin, geldanamycin, herimycin, loracarbef, ertapenem,
imipenem/cilastatin, meropenem, cefadroxil, cefazolin,
cefalotin/cefalothin, cephalexin, cefaclor, cefamandole, cefoxitin,
cefuroxime, cefixime, cefdinir, cefditoren, cefoperazone,
cefotaxime, cefpodoxime, ceftazidime, ceftibuten, ceftizoxime,
ceftriaxone, cefeprime, teicoplanin, vancomycin, azithromycin,
clarithromycin, dirithromycin, erythromycin, roxithromycin,
troleandomycin, telithromycin, spectinomycin, aztreonam,
amoxicillin, ampicillin, azlocillin, carbenicillin, cloxacillin,
dicloxacillin, flucloxacillin, mezlocillin, nafcillin, penicillin,
peperacillin, ticarcillin, bacitracin, colistin, polymyxin B,
ciprofloxacin, enoxacin, gatifloxacin, levofloxacin, lomefloxacin,
moxifloxacin, norfloxacin, ofloxacin, trovafloxacin, mafenide,
protosil, sulfacetamide, sulfamethizole, sulfanilamide,
sulfasalazine, sulfisoxazole, trimethoprim,
trimethoprim-sulfamethoxazole, demeclocycline, doxycycline,
minocycline, oxytetracycline, tetracycline, arsphenamine,
chloramphenicol, clindamycin, lincoamycin, ethambutol, fosfomycin,
fusidic acid, furazolidone, isoniazid, linezolid, metronidazole,
mupirocin, nitrofurantoin, platensimycin, pyrazinamide,
quinupristin/dalfopristin, rifampin/rifampicin, tinidazole,
miconazole, ketoconazole, clotrimazole, econazole, bifonazole,
butoconazole, fenticonazole, isoconazole, oxiconazole,
sertaconazole, sulconazole, tioconazole, fluconazole, itraconazole,
isavuconazole, ravuconazole, posaconazole, voriconazole,
teronazole, terbinafine, amorolfine, naftifine, butenafine,
anidulafungin, caspofungin, micafungin, ciclopirox, flucytosine,
griseofulvin, Gentian violet, haloprogin, tolnaftate, undecylenic
acid, and others.
[0092] In some embodiments, the composition further comprises an
inert and physiologically-acceptable carrier or diluent, enzymes,
anti-microbial agents (anti-bacterial agents, anti-fungal agents,
etc.), vasoconstrictors (such as epinephrine, naphazoline
hydrochloride, tetrahydrozoline, etc.), acids (such as boric acid,
hydrochloric acid, etc.), bases (such as sodium hydroxide, etc.),
salts (such as sodium, potassium, calcium, etc.), polymers,
alcohols (such as polyvinyl alcohol), cellulose or starch,
dextrose, mannose, sucrose, or other carbohydrates; glycoproteins,
proteins, polypeptides or peptides, colors, fragrances,
preservatives (such as edentate disodium, chlorhexidine gluconate,
etc.), or a mixture thereof. In other related embodiments, the
composition further comprises an active pharmaceutical drug or
therapeutic drug substance or an active cosmetic substance. In one
particular embodiment, the pharmaceutical or therapeutic drug
comprises an antihistamine, such as pheniramine maleate.
[0093] Gas-enriched fluids produced in accordance with the
disclosed invention may also be used to decontaminate or wash away
contaminants from a tissue, intact and/or ex vivo. In certain
embodiments, higher levels of gas (e.g. oxygen) in the fluid (e.g.
water) may provide for additional benefits when cleaning and/or
decontaminating wounds.
[0094] Particular embodiments provided herein relate to a
diffuser-processed gas-enriched fluid as defined herein,
comprising: a fluid host material; an infusion material diffused
into the host material; and optionally, at least one cosmetic
and/or therapeutic agent dispersed in the host material. In certain
embodiments, the infusion material comprises oxygen micro-bubbles
in the host fluid, wherein the majority of the micro-bubbles are
less than 0.2 microns, or preferably less than 0.1 microns in
size.
[0095] In certain embodiments, the dissolved oxygen level in the
infused fluid host material may be maintained at greater than 30
parts per million at atmospheric pressure for at least 13 hours. In
other particular embodiments, the dissolved oxygen level in the
infused fluid host material may be maintained at greater than 40
ppm at atmospheric pressure for at least 3 hours. In further
embodiments, the infused fluid host material maintains a dissolved
oxygen level of at least 20 ppm for a period of at least 100 days,
preferably 365 days within a sealed container at atmospheric
pressure. In certain embodiments, the infused fluid host material
may have a dissolved oxygen level of at least 50 parts per million
at atmospheric pressure.
[0096] In certain embodiments, the infused fluid host material
exhibits Rayleigh scattering for a laser beam shining therethrough
for a selected period of time after the oxygen has been diffused
therein.
[0097] By using the diffuser device described herein with respect
to exemplary embodiments illustrated in the corresponding figures,
an output fluid may be achieved having a gas diffused therein that
has a number of characteristics and provides a number of advantages
for use as a therapeutic composition. Solutions have been created
using freshwater, saline, distilled, deionized, double distilled,
and/or double distilled deionized water, oxygen, nitrogen and other
components. Experiments have indicated that oxygen bubbles produced
within saline solution are generally no greater than approximately
0.1 micron in size.
[0098] Wound Healing Activity of the Inventive Gas-Enriched Fluids
and Solutions:
[0099] According to certain aspects of the present invention, the
gas-enriched fluids and/or solutions disclosed herein have
anti-inflammatory properties and effects, and can be used as
anti-inflammatory agents for the treatment of subjects afflicted by
diseases or disorders relating to inflammation. FIG. 42 shows the
experimental results of cytokine profiles in stimulated lymphocytes
from a healthy blood donor. As can be seen in FIG. 42, the
inventive oxygen-enriched fluid (water) affected a down regulation
of particular cytokines, especially IL-6, IL-8, and IL-1.beta..
[0100] Increased production of pro-inflammatory cytokines has been
implicated in the pathogenesis of numerous inflammatory and
autoimmune diseases. Secretion of TNF.alpha. is a primary event in
the initiation of the inflammatory cascade (Brennan F. M., et. al.
Lancet, 1989, 2:244-7; Haworth C, et. al. Eur. J. Immunol. 1991,
21:2575-2579) and directly contributes to the initiation and
maintenance of inflammatory and autoimmune diseases. Other
pro-inflammatory cytokines also play a role, including interleukin
1.beta. (IL-1.beta.), IL-6, IL-8, IL-12 nitric oxide, IFN-.gamma.
and GM-CSF, while anti-inflammatory cytokines such as IL-10 may
reduce disease. Cells of the immune system, macrophages in
particular, secrete many of these cytokines in response to
activating stimuli.
[0101] A variety of cell types are involved in the inflammatory
process. Overproduction of TNF.alpha. by monocytes, macrophages and
other immune cells is a key element in the pathogenesis of a
multitude of diseases. Macrophages and T-cells in particular play a
central role in the initiation and maintenance of the immune
response.
[0102] Once activated by pathological or immunogenic stimuli,
macrophages respond by releasing a host of cytokines, including
TNF-.alpha., IL-1.beta., IL-8, IL-12, nitric oxide (NO), IL-6,
GM-CSF, G-CSF, M-CSF and others. T-cells release IL-2, IL-4,
INF-.gamma., and other inflammatory cytokines. These cytokines
activate other immune cells and some can also act as independent
cytotoxic agents. Excessive release of macrophage and T-cell
derived inflammatory mediators can particularly lead to damage of
normal cells and surrounding tissues.
[0103] Pro-inflammatory cytokines have been implicated in HIV-AIDS,
and other viral infections including the cytomegalovirus, influenza
virus and the herpes family of viruses. TNF.alpha. enhances the
basal activity of the major immediate early enhancer/promoter of
human cytomegalovirus and may play a role in reactivation of latent
HCMV infection in premonocytic cells (Prosch S., et. al. Virology
1995, 208:197-206).
[0104] Additionally, a number of inflammatory cytokines contribute
to mortality in patients suffering from sepsis or endotoxic shock.
For example, TNF.alpha. and IL-1.beta. have a well-established
central role in sepsis, septic shock and endotoxic shock. Increased
levels of these cytokines are associated with fever, hypotension
and shock (Smith J. W. et. al. J. Clin. Oncol. 1992, 10:1141-1152;
Chapman P. B., et. al. J. Clin. Oncol. 1987, 5:1942-1951) together
with the induction of gene expression for phospholipase A2 (Gronich
J., et. al. J. Clin. Invest. 1994, 93:1224-1233) and NO
synthase.
[0105] The induction of NO from smooth muscle cells mediates
decreased mean arterial pressure and systemic vascular resistance
during septic shock, suggesting a fundamental role for NO. Thus,
therapies that target downregulatory effects on IL-8, IL-1.beta.,
and NO could be beneficial in the treatment of inflammatory
diseases or disorders, including sepsis, septic shock, and
endotoxic shock.
[0106] Overproduction of TNF.alpha. contributes to the clinical
features of numerous autoimmune diseases such as diabetes and
rheumatoid arthritis. Systemic lupus erythematosus (SLE) is also
precipitated by increased IL-1.beta. and TNF.alpha. levels. Within
lupus patients, serum C-reactive protein, IL-1.beta and TNF.alpha.
levels were higher than in controls, suggesting that an increased
inflammatory response plays a role in the disease (Liou L. B. Clin.
Exp. Rheumatol. 2001, 19:515-523). A study of patients with one
form of SLE, neuropsychiatric lupus erythematosus (NPLE), showed
that the number of peripheral blood mononuclear cells expressing
mRNA for TNF.alpha. as well as the cerebrospinal fluid level of NO
metabolites correlated with NPLE disease severity (Svenungsson E.,
et al. Ann. Rheum. Dis. 2001, 60:372-9).
[0107] IL-1 and TNF.alpha. play a central role in various acute as
well as chronic responses in animal models. Additionally, IL-11,
IFN.alpha. and IFN.beta. may also up-regulate inflammatory
reactions. Conversely, several cytokines may be involved in
down-regulation of inflammatory responses (i.e. IL-4, IL-10, IL-13,
among others). As set forth in Example 1, cells contacted with the
inventive gas-enriched fluid showed an increase in IFN-.gamma.
levels with T3 antigen than in the control culture media with T3
antigen, while IL-8 was lower in the inventive gas-enriched culture
media with T3 antigen than in the control culture media with T3
antigen. Additionally, IL-6, IL-8, and TNF-.alpha. levels were
lower in the inventive gas-enriched media with PHA, than in the
control media with PHA, while IL-1.beta. levels were lower in the
inventive gas-enriched fluid with PHA when compared with control
media with PHA. In the inventive gas-enriched media alone,
IFN-.gamma. levels were higher than in control media. These results
are consistent with an anti-inflammatory microenvironment.
[0108] NO is recognized as a mediator and regulator of inflammatory
responses. It possesses cytotoxic properties toward pathogens, but
can also have deleterious effects on the subject's own tissues.
(Korhonen et al., Curr Drug Targets Inflamm Allergy 4(4): 471-9,
2005). NO reacts with soluble guanylate cyclase to form cyclic
guanosine monophosphate (cGMP), which mediates many of the effects
of NO. NO can also interact with molecular oxygen and superoxide
anion to produce reactive oxygen species that can modify various
cellular functions. These indirect effects of NO have a significant
role in inflammation, where NO is produced in high amounts by
inducible NO synthase (iNOS) and reactive oxygen species are
synthesized by activated inflammatory cells.
[0109] NO can be produced by keratinocytes, fibroblasts,
endothelial cells, and possibly others. Some of the vascular
actions of NO include vasodilation, inhibiting platelet adhesion to
the vascular endothelium, inhibiting leukocyte adhesion to the
vascular endothelium, and scavenging superoxides. (Shah et al.,
Env. Health Persp. v. 106 (5): 1139-1143.)
[0110] Furthermore, inhibition of NO synthesis has been shown to
delay wound contraction, alter collagen organization, and alter
neoepidermis thickness. (Amadeu and Costa, J. Cutan. Pathol. 33:
465-473, 2006.) Mast cell migration and angiogenesis in wounds is
also affected by inhibition of NO. (Id.) Without being bound to any
particular theory of mechanism, in certain embodiments the
inventive gas-enriched fluids may be modulating localized and/or
cellular NO production, or degradation, consistent with the
spectrum of wound healing effects illustrated in the Examples
section disclosed herein. Due to variable pathways of regulation,
in certain embodiments, the inventive gas-enriched fluid may
increase NO production and/or retard NO degradation, whereas in
other certain embodiments, the inventive gas-enriched fluid may
decrease NO production and/or hasten NO degradation.
[0111] Specifically, wounds treated with oxygen-enriched saline
solution showed an increase in wound healing at days 4 through 11,
and between days 3 and 11, the new epidermis in wounds treated with
the oxygen-enriched saline solution migrated at two to four times
as fast as the epidermis of the wounds treated with the normal
saline solution, as set forth in Example 9 herein. The study also
showed that between 15 and 22 days, wounds treated by the
oxygen-enriched saline solution differentiated at a more rapid rate
as evidenced by the earlier formation of more mature epidermal
layers. At all stages, the thickening that occurs in the epidermis
associated with normal healing did not occur within the wounds
treated by the oxygen-enriched saline solution.
[0112] Thus, in accordance with this spectrum of wound healing
effects, but without wishing to be bound by any particular theory,
it is believed that the oxygen-enriched saline solution may
modulate the localized and/or cellular level of NO within the
wounds. NO modulates growth factors, collagen deposition,
inflammation, mast cell migration, epidermal thickening, and
neovascularization in wound healing. Furthermore, nitric oxide is
produced by an inducible enzyme that is regulated by oxygen.
[0113] In the case of mast cell migration, differences also
occurred in early and late migration for the oxygen-enriched
solution. This is consistent with what is known in the art
regarding inhibition of NO synthesis (Amadeu and Costa, J. Cutan
Pathol 33: 465-473, 2006).
[0114] Referring now to FIG. 45a through FIG. 45h, various
illustrations compare the wound healing results of the porcine
epidermal tissues with or without oxygen-enriched saline solution.
As can be seen, the healing of the control wound and of the wound
using the oxygen-enriched saline solution was followed for days 1,
4 and 16.
[0115] FIG. 45a illustrates the wound healing for the control wound
on day 1. As can be seen, the wound shows epidermal/dermal
thickening and a loss of contour. FIG. 45b illustrates the wound
healing on day 1 for the wound treated using the oxygen-enriched
saline solution. The wound shows normal epidermal/dermal thickness
and normal contouring is typical on a new wound.
[0116] Referring now to FIGS. 45c and 45d, there are illustrated
the wound healing for the control wound on day 4 and the wound
healing for the wound treated with the oxygen-enriched saline
solution on day 4. For the control wound illustrated in FIG. 45c,
the wound shows a 600 micron epidermal spur. In the wound treated
with the oxygen-enriched saline solution in FIG. 45d, there is
illustrated a 1200 micron epidermal spur. Thus, in the first 4 days
of the experiment, the epidermal spur created in the wound treated
using the oxygen-enriched saline solution shows an epidermal growth
rate of twice of that of the wound that was not treated with the
oxygen-enriched saline solution.
[0117] Referring now to FIG. 45e, there is illustrated the control
wound at day 16. The wound shows less differentiated epidermis with
loss of epidermal/dermal contour than that illustrated by the wound
treated with the oxygen-enriched saline solution illustrated in
FIG. 45f. FIG. 45f shows more differentiated epidermis and more
normal epidermal/dermal contouring in the wound.
[0118] In the first two phases of the inflammatory process, the
foreign body is either destroyed, for example, if the foreign body
is an organism, or the tissue around it is loosened, for example,
if it is a splinter. In the healing phase, the inflammation begins
to subside; individual blood vessels and vascular patterns become
normal once again; and repair of the wound commences. The three
main events in the repair process are (1) formation of new
connective tissue by proliferating fibroblasts; (2) regeneration of
epithelium; and (3) outgrowth of new capillaries.
[0119] Even before the inflammation subsides, fibroblasts begin
moving into the injured area from the surrounding normal tissue,
where they usually exist in a dormant state. They migrate by an
amoeboid movement along strands of fibrin and distribute themselves
throughout the healing area. Once fixed into position in the
injured tissue, they begin to synthesize collagen and secrete this
protein, which arranges itself into fibers. The fibers orient
themselves with their longitudinal axes in the direction of the
greatest stress. As the collagen bundles grow in firmness, the
fibroblasts gradually degenerate and attach closely to the bundles,
and the injured area transforms into scar tissue.
[0120] Simultaneously with scar tissue formation, the intact
epidermal cells on the edge of the wound begin to proliferate and
move, as one sheet, toward the center of the injured area. As the
inflammation subsides, a need for a direct supply of blood arises,
and angiogenesis occurs at the wound site.
[0121] Inflammation is a complex process that involves multiple
cell types. For example, mast cells release mediators that trigger
an early phase of vasodilation, accompanied by the separation of
endothelial cells and exposure of collagen fibers in the
subendothelial layer. Fibers in the intercellular gaps that form in
blood vessels trap platelets and trigger the release of mediators
from these cells.
[0122] In addition to platelets, the exposed collagen fibers also
interact with proteins of the plasma that filter through the pores
of the dilated vessel wall, including the triggering factor of the
blood-clotting cascade, increased vasodilation, increased blood
vessel permeability, and chemotaxis.
[0123] Additionally, the complement cascade can be activated by
several stimuli: the injured blood vessels, the proteolytic enzymes
released by the damaged cells, the membrane components of any
participating bacteria, and antigen-antibody complexes. Some of the
activated complement components act as chemotactic factors,
responsible for the influx of leukocytes into the inflamed area,
while others facilitate phagocytosis and participate in cell
lysis.
[0124] In addition, it is believed that the inventive gas-enriched
fluids or solutions may also regulate at least one cytokine
involved in at least one aspect of inflammation, the cytokine(s)
including, but not limited to MAF (macrophage activating factor),
MMIF (macrophage migration inhibition factor), MCF (macrophage
chemotactic factor), LMIF (leukocyte migration inhibition factor),
HRFs (histamine releasing factors), TF (transfer factors),
interleukins (IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9,
IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, etc.), TNF-.alpha.,
TNF-.beta., interferons (IFN-.alpha., IFN-.beta., IFN-.gamma.,
IFN-.zeta., IFN-.delta., etc.), G-CSF (granulocyte colony
stimulating factor), GM-CSF (granulocyte-macrophage CSF), M-CSF
(macrophage CSF), multi-CSF (IL-3), fibroblast growth factor (aFGF,
bFGF), EGF (epidermal growth factor), NGF (nerve growth factor),
PDGF (platelet-derived growth factor), VEGF (vascular endothelial
growth factor), transforming growth factors (TGF-.alpha.,
TGF-.beta., etc.), NAP-2 (neutrophil-activating protein 2), PF-4
(platelet factor 4), thromboglobulin, MCP-1 (monocyte
chemoattractant protein 1), MCP-3, MIP-1.alpha., MIP-1.beta.-+
(macrophage inflammatory proteins), RANTES (regulated upon
activation normal T expressed and presumably secreted chemokine),
HSPs (heat shock proteins), GRPs (glucose-regulated proteins),
ubiquitin, and others.
[0125] Thus, in certain embodiments, the gas-enriched fluids and/or
therapeutic compositions may increase production and/or secretion
of anti-inflammatory molecules or cytokines or decrease the
degradation of anti-inflammatory molecules or cytokines, thereby
alleviating or preventing at least one symptom of inflammation. In
other embodiments, the gas-enriched fluids and/or therapeutic
compositions of the present invention may decrease production
and/or secretion of pro-inflammatory molecules or cytokines or
increase the degradation of pro-inflammatory molecules or
cytokines, thereby alleviating or preventing at least one symptom
of inflammation.
[0126] Previous studies had shown a critical role of anti-MOG
antibodies in augmentation of demyelination and worsening of EAE
(experimental autoimmune encephalomyelitis), an animal model system
for the human autoimmune disorder of rheumatoid arthritis.
(Linington, et al. 1992. J. Neuroimmunol. 40:219-224).
Additionally, antibodies against MOG have been implicated in the
pathogenesis of multiple sclerosis. (Berger et al. N. Engl. J. Med.
2003 Jul. 10. 349(2):139-45).
[0127] As set forth in FIG. 52 and Example 18, the inventive
gas-enriched fluid of the present invention amplifies the
lymphocyte response to an antigen for which an animal was
previously primed. As indicated in FIG. 52, lymphocyte
proliferation was greater for response to MOG challenge when
cultured in fluid reconstituted with the inventive gas-enriched
fluid comprising solvated electrons, when compared with
pressurized, oxygenated fluid (pressure pot) or control deionized
fluid.
Bubble Size Measurements
[0128] Experimentation was performed to determine a size of the
bubbles of gas diffused within the fluid by the mixing device 100.
While experiments were not performed to measure directly the size
of the bubbles, experiments were performed that established that
the bubble size of the majority of the gas bubbles within the fluid
was smaller than 0.1 microns. In other words, the experiments
determined a size threshold value below which the sizes of the
majority of bubbles fall.
[0129] This size threshold value or size limit was established by
passing the output material 102 formed by processing a fluid and a
gas in the mixing device 100 through a 0.22 filter and a 0.1 micron
filter. In performing these tests, a volume of the first material
110, in this case, a fluid, and a volume of the second material
120, in this case, a gas, were passed through the mixing device 100
to generate a volume of the output material 102 (i.e., a fluid
having a gas diffused therein). Sixty milliliters of the output
material 102 was drained into a 60 ml syringe. The DO level of the
fluid was measured via the Winkler Titration. The fluid within the
syringe was injected through a 0.22 micron filter into a 50 ml
beaker. The filter comprised the Milipor Millex GP50 filter. The DO
level of the material in the 50 ml beaker was then measured. The
experiment was performed three times to achieve the results
illustrated in Table II below.
TABLE-US-00001 TABLE II DO IN DO AFTER SYRINGE 0.22 MICRON FILTER
42.1 ppm 39.7 ppm 43.4 ppm 42.0 ppm 43.5 ppm 39.5 ppm
[0130] As can be seen, the DO levels measured within the syringe
and the DO levels measured within the 50 ml beaker were not changed
drastically by passing the output material 102 through the 0.22
micron filter. The implication of this experiment is that the
bubbles of dissolved gas within the output material 102 are not
larger than 0.22 microns otherwise there would be a significantly
greater reduction in the DO levels in the output material 102
passed through the 0.22 micron filter.
[0131] A second test was performed in which the 0.1 micron filter
was substituted for the 0.22 micron filter. In this experiment,
saline solution was processed with oxygen in the mixing device 100
and a sample of the output material 102 was collected in an
unfiltered state. The DO level of the unfiltered sample was 44.7
ppm. The output material 102 was filtered using the 0.1 micron
filter and two additional samples were collected. The DO level of
the first sample was 43.4 ppm. The DO level of the second sample
was 41.4 ppm. Then, the filter was removed and a final sample was
taken from the unfiltered output material 102. The final sample had
a DO level of 45.4 ppm. These results were consistent with those
seen using the Millipore 0.22 micron filter. These results lead to
the conclusion that there is a trivial reduction in the DO levels
of the output material 102 passed through the 0.1 micron filter
providing an indication that the majority of the bubbles in the
processed saline solution are no greater than 0.1 micron in
size.
[0132] As appreciated in the art, the double-layer (interfacial)
(DL) appears on the surface of an object when it is placed into a
liquid. This object, for example, might be that of a solid surface
(e.g., rotor and stator surfaces), solid particles, gas bubbles,
liquid droplets, or porous body. In the mixing device 100, bubble
surfaces represent a significant portion of the total surface area
present within the mixing chamber that may be available for
electrokinetic double-layer effects. Therefore, in addition to the
surface area and retention time aspects discussed elsewhere herein,
the relatively small bubble sizes generated within the mixer 100
compared to prior art devices 10, may also contribute, at least to
some extent, to the overall electrokinetic effects and output fluid
properties disclosed herein. Specifically, in preferred
embodiments, as illustrated by the mixer 100, all of the gas is
being introduced via apertures on the rotor (no gas is being
introduced through stator apertures. Because the rotor is rotating
at a high rate (e.g., 3,400 rpm) generating substantial shear
forces at and near the rotor surface, the bubble size of bubbles
introduced via, and adjacent to the spinning rotor surface
apertures would be expected to be substantially (e.g., 2 to 3-times
smaller) smaller than those introduced via and near the stationary
stator. The average bubble size of the prior art device 10 may,
therefore, be substantially larger because at least half of the gas
is introduced into the mixing chamber from the stationary stator
apertures. Because the surface area of a sphere surface varies with
r.sup.2, any such bubble component of the electrokinetic surface
area of the mixing device 100 may be substantially greater than
that of the prior art diffusion device 10.
[0133] Therefore, without being bound by theory, not only does the
mixing chamber of the mixing device 100 have (i) a substantially
higher surface to volume ratio than that of the prior art device 10
(the prior art device 10 has a ratio of surface to volume of 10.9,
whereas the present mixer 100 has a surface to volume ratio of
39.4), along with (ii) a 7-fold greater dwell-time, but (iii) the
unique properties of the current output solutions may additionally
reflect a contribution from the substantially larger bubble surface
area in the mixing device 100. These distinguishing aspects reflect
distinguishing features of the present mixer 100, and likely each
contribute to the unique electrokinetic properties of the inventive
output materials/fluids.
[0134] Compositions Comprising Hydrated (Solvated) Electrons
Imparted to the Inventive Compositions by the Inventive
Processes
[0135] In certain embodiments as described herein (see under
"Double-layer"), the gas-enriched fluid is generated by the
disclosed electromechanical processes in which molecular oxygen is
diffused or mixed into the fluid and may operate to stabilize
charges (e.g., hydrated (solvated) electrons) imparted to the
fluid. Without being bound by theory or mechanism, certain
embodiments of the present invention relate to a oxygen-enriched
fluid (output material) comprising charges (e.g., hydrated
(solvated) electrons) that are added to the materials as the first
material is mixed with oxygen in the inventive mixer device to
provide the combined output material. According to particular
aspects, these hydrated (solvated) electrons (alternately referred
to herein as `solvated electrons`) are stabilized in the inventive
solutions as evidenced by the persistence of assayable effects
mediated by these hydrated (solvated) electrons. Certain
embodiments may relate to hydrated (solvated) electrons and/or
water-electron structures, clusters, etc. (See, for example, Lee
and Lee, Bull. Kor. Chem. Soc. 2003, v. 24, 6; 802-804; 2003).
[0136] Novel HRP Based Assay.
[0137] Horseradish peroxidase (HRP) is isolated from horseradish
roots (Amoracia rusticana) and belongs to the ferroprotoporphyrin
group (Heme group) of peroxidases. HRP readily combines with
hydrogen peroxide or other hydrogen donors to oxidize the
pyrogallol substrate. Additionally, as recognized in the art, HRP
facilitates auto-oxidative degradation of indole-3-acetic acid in
the absence of hydrogen peroxide (see, e.g., Heme Peroxidases, H.
Brian Dunford, Wiley-VCH, 1999, Chapter 6, pages 112-123,
describing that auto-oxidation involves a highly efficient
branched-chain mechanism; incorporated herein by reference in its
entirety). The HRP reaction can be measured in enzymatic activity
units, in which Specific activity is expressed in terms of
pyrogallol units. One pyrogallol unit will form 1.0 mg
purpurogallin from pyrogallol in 20 sec at pH 6.0 at 20.degree. C.
This purpurogallin (20 sec) unit is equivalent to approx. 18 .mu.M
units per min at 25.degree. C.
##STR00001##
[0138] According to particular aspects of the present invention,
the oxygen-enriched inventive fluids (output materials) have been
described and disclosed herein to react with pyrogallol in the
presence of horseradish peroxidase. The reaction is most likely
based on an auto-oxidation of the pyrogallol, since no hydrogen
peroxide, superoxide, or other reactive oxygen species has been
detected in oxygen-enriched inventive fluid. The extent of this
reaction is greater than that of pressurized oxygen solutions
(pressure-pot oxygen solutions) and less than that of hydrogen
peroxide.
[0139] Specifically, the present applicants have determined that
while there is no hydrogen peroxide (none detected at a sensitivity
of 0.1 ppm), the inventive gas-enriched fluid may be consistently
characterized by its facilitation of the apparent auto-oxidation of
pyrogallol to purpurogallin in the presence of horseradish
peroxidase enzyme (HRP). That is, like the case of HRP facilitation
of the autoxidative degradation of indole-3-acietic acid in the
absence of hydrogen peroxide, applicants have discovered HRP
facilitation of the autoxidative degradation of pyrogallol in the
absence of hydrogen peroxide. According to particular aspects, the
presence and level of this activity are distinguishing features of
the inventive compositions in view of the prior art.
[0140] In certain embodiments, the inventive gas-enriched fluid
facilitates, in the presence of HRP and absence of hydrogen
peroxide, a pyrogallol auto-oxidation rate (under standard
conditions as defined herein under "Definitions") equivalent to
approximately 0.5 ppm of hydrogen peroxide, approximately 0.8 ppm
of hydrogen peroxide, approximately 1 ppm of hydrogen peroxide,
approximately 2 ppm of hydrogen peroxide, approximately 3 ppm of
hydrogen peroxide, approximately 4 ppm of hydrogen peroxide,
approximately 5 ppm of hydrogen peroxide, approximately 6 ppm of
hydrogen peroxide, approximately 7 ppm of hydrogen peroxide,
approximately 8 ppm of hydrogen peroxide, approximately 9 ppm of
hydrogen peroxide, approximately 10 ppm of hydrogen peroxide,
approximately 11 ppm of hydrogen peroxide, approximately 12 ppm of
hydrogen peroxide, approximately 20 ppm of hydrogen peroxide,
approximately 40 ppm of hydrogen peroxide, approximately 50 ppm of
hydrogen peroxide or any value therebetween or greater.
[0141] It is known that Horseradish peroxidase enzyme catalyzes the
auto-oxidation of pyrogallol by way of facilitating reaction with
the molecular oxygen in a fluid. (Khajehpour et al., PROTEINS:
Struct, Funct, Genet. 53: 656-666 (2003)). It is also known that
oxygen binds the heme pocket of horseradish peroxidase enzyme
through a hydrophobic pore region of the enzyme (between Phe68 and
Phe142), whose conformation likely determines the accessibility of
oxygen to the interior. Without being bound by mechanism, because
surface charges on proteins are known in the protein art to
influence protein structure, it is possible that the solvated
electrons present in the inventive gas-enriched fluid act to alter
the conformation of the horseradish peroxidase such that greater
oxygen accessibility results. The greater accessibility of oxygen
to the prosthetic heme pocket of the horseradish peroxidase enzyme
in turn would allow for increased reactivity with pyrogallol, when
compared with prior art oxygen-enriched fluids (pressure-pot,
fine-bubbled). Alternatively, the added or solvated electrons of
the present output compositions may be acting in other ways to
enable facilitation of the apparent auto-oxidation of pyrogallol to
purpurogallin in the presence of horseradish peroxidase enzyme
(HRP).
[0142] In any event, according to particular aspects, production of
output material using the inventive methods and devices comprises a
process involving: an interfacial double layer that provides a
charge gradient; movement of the materials relative to surfaces
pulling charge (e.g., electrons) away from the surface by virtue of
a triboelectric effect, wherein the flow of material produces a
flow of solvated electrons. Moreover, according to additional
aspects, and without being bound by mechanism, the orbital
structure of diatomic oxygen creates charge imbalances (e.g., the
two unpaired electrons affecting the hydrogen bonding of the water)
in the hydrogen bonding arrangement within the fluid material
(water), wherein electrons are solvated and stabilized within the
imbalances.
[0143] The inventive combination of oxygen-enrichment and solvated
electrons imparted by the double-layer effects and configuration of
the presently claimed devices facilitates the auto-oxidation of
pyrogallol in the presence of HRP, which is a distinguishing
feature of the present inventive output material compositions that
can be readily monitored and quantified by way of optical density.
Typically, the inventive oxygen-enriched compositions are
characterized in that they provide for about a 20% higher optical
density read-out in the standard assay compared to either
pressurized (pressure pot) or fine-bubbled control fluid have
equivalent dissolved oxygen concentrations.
[0144] Pyrogallol Reactivity Test
[0145] An aliquot of the inventive oxygen-enriched output material
was tested for peroxidase activity by using a commercially
available horseradish peroxidase and a pyrogallol assay (Sigma).
Briefly, pyrogallol stock solution was prepared with deionized
water. Pyrogallol measures peroxidase activity of the horseradish
peroxidase enzyme on the fluid as it reacts with a substrate (such
as hydrogen peroxide), to yield purpurogallin and water. Test fluid
with horseradish peroxidase, pyrogallol and the appropriate
potassium phosphate buffer were compared with other fluids.
Hydrogen peroxide served as the positive control. The other fluids
tested were water that was oxygen-enriched and pressurized in a
pressure pot, up to 100 psi to reach the desired dissolved oxygen
level (Pressure Pot), while the other fluid was oxygen-enriched
with an air stone in an open beaker (Fine Bubble). All fluids
tested were maintained at room temperature, and measured
approximately 55 ppm dissolved oxygen level (by FOXY probe). Water
samples were tested by adding the enzymatic reagents. Continuous
spectrophotometric rate determination was made at A.sub.420 nm, and
room temperature (25.degree. Celsius).
[0146] As indicated in FIGS. 39A-39E, the inventive oxygen-enriched
fluid tested positive for reactivity with horseradish peroxidase by
pyrogallol, while the pressure pot and fine bubbled water samples
had far less reactivity. As indicated in FIG. 39E, oxygen is
required for the reaction with pyrogallol in the presence of
horseradish peroxidase, as inventive fluid enriched with other
gases did not react in the same manner.
[0147] Several chemical tests of the inventive oxygen-enriched
fluid for the presence of hydrogen peroxide were conducted, as
described herein, and none of these tests were positive
(sensitivity of 0.1 ppm hydrogen peroxide). Thus, the inventive
oxygen-enriched fluid of the instant application provides for
peroxidase facilitated auto-oxidation activity in the absence of
hydrogen peroxide.
[0148] In particular embodiments, Applicants have determined that
the horseradish peroxidase effect remains at least up to seven
hours after opening of the bottle in which it is stored. In other
embodiments, Applicants have determined that the horseradish
peroxidase effect remains after opening of closed container after
105 days of storage in the closed container. By contrast, in other
embodiments, Applicants have determined that when testing
equivalent dissolved oxygen levels made with just pressurizing
fluid (pressure pot fluids), the decline of a background HRP effect
takes place rapidly, declining precipitously in under 4 hours.
[0149] Glutathione Peroxidase Study
[0150] The inventive oxygen-enriched output fluid material was
tested for the presence of hydrogen peroxide by testing the
reactivity with glutathione peroxidase using a standard assay
(Sigma). Water samples were tested by adding the enzymatic
reagents. Continuous spectrophotometric rate determination was made
at A.sub.340 nm, and room temperature (25.degree. Celsius). Samples
tested were: 1. deionized water (negative control), 2. inventive
oxygen-enriched fluid at low concentration, 3. inventive
oxygen-enriched fluid at high concentration, 4. hydrogen peroxide
(positive control). As illustrated in FIG. 40, the hydrogen
peroxide positive control showed a strong reactivity, while none of
the other fluids tested reacted with the glutathione.
[0151] Differential DNA Thermostability
[0152] Particular embodiments of the present invention provide
another distinguishing feature of the present inventive
compositions. Specifically, applicants have discovered that there
is a differential thermostability of nucleic acids associated with
the inventive output fluids compared to control fluids. For
example, the T7 promoter primer 5'-d(TAATACGACTCACTATAGGG)-3' (SEQ
ID NO:1) when measured in the inventive oxygen-enriched output
materials relative to non-enriched deionized water. As the
temperature of the water increases, the DNA oligomeric structure
performs a conformational change. As illustrated in FIG. 38,
consent with the art recognized melting temperature for this oligo
of about 48.degree. C., the T7 DNA begins to denature at about
50.degree. Celsius in the control (deionized water), whereas the
DNA in the oxygen-enriched inventive fluid remains intact until
about 60.degree. Celsius. Thus, the inventive oxygen-enriched fluid
comprising solvated electrons imparts a higher thermostability for
DNA when compared to control fluid, and provides a further
distinguishing feature of the present inventive output material
compositions that can be readily monitored and quantified by way of
optical density measurements.
[0153] Device for Generating Gas-Enriched Fluids or Solutions
[0154] Description of the Related Art
[0155] FIG. 1 provides a partial block diagram, partial
cross-sectional view of a prior art device 10 for diffusing or
emulsifying one or two gaseous or liquid materials ("infusion
materials") into another gaseous or liquid material ("host
material") reproduced from U.S. Pat. No. 6,386,751, incorporated
herein by reference in its entirety. The device 10 includes a
housing configured to house a stator 30 and a rotor 12. The stator
30 encompasses the rotor 12. A tubular channel 32 is defined
between the rotor 12 and the stator 30. The generally cylindrically
shaped rotor 12 has a diameter of about 7.500 inches and a length
of about 6.000 inches providing a length to diameter ratio of about
0.8.
[0156] The rotor 12 includes a hollow cylinder, generally closed at
both ends. A gap exists between each of the first and second ends
of the rotor 12 and a portion of the housing 34. A rotating shaft
14 driven by a motor 18 is coupled to the second end of the rotor
12. The first end of the rotor 12 is coupled to an inlet 16. A
first infusion material passes through the inlet 16 and into the
interior of the rotor 12. The first infusion material passes from
the interior of the rotor 12 and into the channel 32 through a
plurality of openings 22 formed in the rotor 12.
[0157] The stator 30 also has openings 22 formed about its
circumference. An inlet 36 passes a second infusion material to an
area 35 between the stator 30 and the housing 34. The second
infusion material passes out of the area 35 and into the channel 32
through openings 22.
[0158] An external pump (not shown) is used to pump the host
material into a single inlet port 37. The host material passes
through a single inlet port 37 and into the channel 32 where it
encounters the first and second infusion materials, which enter the
channel 32 through openings 22. The infusion materials may be
pressurized at their source to prevent the host material from
passing through openings 22.
[0159] The inlet port 37, is configured and positioned such that it
is located along only a relatively small portion (<about 5%) of
the annular inlet channel 32, and is substantially parallel to the
axis of rotation of the rotor 12 to impart an axial flow toward a
portion of the channel 32 into the host material.
[0160] Unfortunately, before entering the tubular channel 32, the
host material must travel in tortuous directions other than that of
the axial flow (e.g., including in directions substantially
orthogonal thereto) and down into and between the gap formed
between the first end of the rotor 12 and the housing 34 (i.e.,
down a portion of the first end of the rotor adjacent to the inlet
16 between the end of the rotor 12 and the housing 34). The
non-axial and orthogonal flow, and the presence of the host
material in the gap between the first end of the rotor 12 and the
housing 34 causes undesirable and unnecessary friction. Further, it
is possible for a portion of the host material to become trapped in
eddy currents swirling between the first end of the rotor and the
housing. Additionally, in the device 10, the host material must
negotiate at least two right angles to enter any aspect of the
annual of the annular inlet of the tubular channel 32.
[0161] A single outlet port 40 is formed in the housing 34. The
combined host material and infusion material(s) exit the channel 32
via the outlet 40. The outlet port 40, which is also located along
only a limited portion (<about 5%) of the annular outlet of
tubular channel 32, is substantially parallel to the axis of
rotation of the rotor 12 to impart or allow for an axial flow of
the combined materials away from the limited portion of the annular
outlet of tubular channel 32 into the outlet port 40. An external
pump 42 is used to pump the exiting fluid through the outlet port
40.
[0162] Unfortunately, before exiting the channel 32, a substantial
portion of the exiting material must travel in a tortuous direction
other than that of the axial flow (e.g., including in directions
substantially orthogonal thereto) and down into and between the gap
formed between the second end of the rotor 12 and the housing 34
(i.e., down a portion of the second end of the rotor adjacent to
the shaft 14 between the end of the rotor 12 and the housing 34).
As mentioned above, the non-axial and orthogonal flow, and the
presence of the host material in the other gap between the end (in
this case, the second end) of the rotor 12 and the housing 34
causes additional undesirable and unnecessary friction. Further, it
is possible for a portion of the host material to become trapped in
eddy currents swirling between the second end of the rotor and the
housing. Additionally, in the device 10, a substantial portion of
the exiting combined material must negotiate at least two right
angles as it exits form the annular exit of the tubular channel 32
into the outlet port 40.
[0163] As is apparent to those of ordinary skill in the art, the
inlet port 37 imparts only an axial flow to the host material. Only
the rotor 21 imparts a circumferential flow into the host material.
Further, the outlet port 40 imparts or provides for only an axial
flow into the exiting material. Additionally, the circumferential
flow velocity vector is imparted to the material only after it
enters the annular inlet 37 of the tubular channel 32, and
subsequently the circumferential flow vector must be degraded or
eliminated as the material enters the exit port 40. There is,
therefore, a need for a progressive circumferential acceleration of
the material as it passes in the axial direction through the
channel 32, and a circumferential deceleration upon exit of the
material from the channel 32. These aspects, in combination with
the tortuous path that the material takes from the inlet port 37 to
the outlet port 40, create a substantial friction and flow
resistance over the path that is accompanied by a substantial
pressure differential (26 psi, at 60 gallons/min flow rate) between
the inlet 37 and outlet 40 ports, and these factors, inter alia,
combine to reduce the overall efficiency of the system.
[0164] Electrokinetically Oxygen-Enriched Fluids and Solutions
[0165] FIG. 2 provides a block diagram illustrating some of the
components of a mixing device 100 and the flow of material into,
within, and out of the device. The mixing device 100 combines two
or more input materials to form an output material 102, which may
be received therefrom into a storage vessel 104. The mixing device
100 agitates the two or more input materials in a novel manner to
produce an output material 102 having novel characteristics. The
output material 102 may include not only a suspension of at least
one of the input materials in at least one of the other input
materials (e.g., emulsions) but also a novel combination (e.g.,
electrostatic combinations) of the input materials, a chemical
compound resulting from chemical reactions between the input
materials, combinations having novel electrostatic characteristics,
and combinations thereof.
[0166] The input materials may include a first material 110
provided by a source 112 of the first material, a second material
120 provided by a source 122 of the second material, and optionally
a third material 130 provided by a source 132 of the third
material. The first material 110 may include a liquid, such as
water, saline solution, chemical suspensions, polar liquids,
non-polar liquids, colloidal suspensions, cell growing media, and
the like. In some embodiments, the first material 110 may include
the output material 102 cycled back into the mixing device 100. The
second material 120 may consist of or include a gas, such as
oxygen, nitrogen, carbon dioxide, carbon monoxide, ozone, sulfur
gas, nitrous oxide, nitric oxide, argon, helium, bromine, and
combinations thereof, and the like. In preferred embodiments, the
gas is or comprises oxygen. The optional third material 130 may
include either a liquid or a gas. In some embodiments, the third
material 130 may be or include the output material 102 cycled back
into the mixing device 100 (e.g., to one or more of the pumps 210,
220 or 230, and/or into the chamber 310, and/or 330).
[0167] Optionally, the first material 110, the second material 120,
and the optional third material 130 may be pumped into the mixing
device 100 by an external pump 210, an external pump 220, and an
external pump 230, respectively. Alternatively, one or more of the
first material 110, the second material 120, and the optional third
material 130 may be stored under pressure in the source 112, the
source 122, and the source 132, respectively, and may be forced
into the mixing device 100 by the pressure. The invention is not
limited by the method used to transfer the first material 110, the
second material 120, and optionally, the third material 130 into
the mixing device 100 from the source 112, the source 122, and the
source 132, respectively.
[0168] The mixing device 100 includes a first chamber 310 and a
second chamber 320 flanking a mixing chamber 330. The three
chambers 310, 320, and 330 are interconnected and form a continuous
volume.
[0169] The first material 110 is transferred into the first chamber
310 and flows therefrom into the mixing chamber 330. The first
material 110 in the first chamber 310 may be pumped into the first
chamber 310 by an internal pump 410. The second material 120 is
transferred into the mixing chamber 330. Optionally, the third
material 130 may be transferred into the mixing chamber 330. The
materials in the mixing chamber 330 are mixed therein to form the
output material 102. Then, the output material 102 flows into the
second chamber 320 from which the output material 102 exits the
mixing device 100. The output material 102 in the mixing chamber
330 may be pumped into the second chamber 320 by an internal pump
420. Optionally, the output material 102 in the second chamber 320
may be pumped therefrom into the storage vessel 104 by an external
pump 430 (e.g., alone or in combination with the internal pump 410
and/or 420).
[0170] In particular aspects, a common drive shaft 500 powers both
the internal pump 410 and the internal pump 420. The drive shaft
500 passes through the mixing chamber 330 and provides rotational
force therein that is used to mix the first material 110, the
second material 120, and optionally, the third material 130
together. The drive shaft 500 is powered by a motor 510 coupled
thereto.
[0171] FIG. 3 provides a system 512 for supplying the first
material 110 to the mixing device 100 and removing the output
material 102 from the mixing device 100. In the system 512, the
storage vessel 104 of the output material 102 and the source 112 of
the first material 110 are combined. The external pump 210 is
coupled to the combined storage vessel 104 and source 112 by a
fluid conduit 514 such as hose, pipe, and the like. The external
pump 210 pumps the combined first material 110 and output material
102 from the combined storage vessel 104 and source 112 through the
fluid conduit 514 and into a fluid conduit 516 connecting the
external pump 210 to the mixing device 100. The output material 102
exits the mixing device 100 through a fluid conduit 518. The fluid
conduit 518 is coupled to the combined storage vessel 104 and
source 112 and transports the output material 102 exiting the
mixing device 100 to the combined storage vessel 104 and source
112. The fluid conduit 518 includes a valve 519 that establishes an
operating pressure or back pressure within the mixing device
100.
[0172] Referring to FIGS. 2 and 4-11, a more detailed description
of various components of an embodiment of the mixing device 100
will be provided. The mixing device 100 is scalable. Therefore,
dimensions provided with respect to various components may be used
to construct an embodiment of the device or may be scaled to
construct a mixing device of a selected size.
[0173] Turning to FIG. 4, the mixing device 100 includes a housing
520 that houses each of the first chamber 310, the mixing chamber
330, and the second chamber 320. As mentioned above, the mixing
device 100 includes the drive shaft 500, which rotates during
operation of the device. Therefore, the mixing device 100 may
vibrate or otherwise move. Optionally, the mixing device 100 may be
coupled to a base 106, which may be affixed to a surface such as
the floor to maintain the mixing device 100 in a substantially
stationary position.
[0174] The housing 520 may be assembled from two or more housing
sections. By way of example, the housing 520 may include a central
section 522 flanked by a first mechanical seal housing 524 and a
second mechanical seal housing 526. A bearing housing 530 may be
coupled to the first mechanical seal housing 524 opposite the
central section 522. A bearing housing 532 may be coupled to the
second mechanical seal housing 526 opposite the central section
522. Optionally, a housing section 550 may be coupled to the
bearing housings 530.
[0175] Each of the bearing housings 530 and 532 may house a bearing
assembly 540 (see FIGS. 5 and 6). The bearing assembly 540 may
include any suitable bearing assembly known in the art including a
model number "202SZZST" manufactured by SKF USA Inc., of
Kulpsville, Pa., operating a website at www.skf.com.
[0176] Seals may be provided between adjacent housing sections. For
example, o-ring 560 (see FIG. 5) may be disposed between the
housing section 550 and the bearing housing 530, o-ring 562 (see
FIG. 5) may be disposed between the first mechanical seal housing
524 and the central section 522, and o-ring 564 (see FIG. 6) may be
disposed between the second mechanical seal housing 526 and the
central section 522.
Mixing Chamber 330
[0177] Turning now to FIG. 7, the mixing chamber 330 is disposed
inside the central section 522 of the housing 520 between the first
mechanical seal housing 524 and the second mechanical seal housing
526. The mixing chamber 330 is formed between two components of the
mixing device 100, a rotor 600 and a stator 700. The rotor 600 may
have a sidewall 604 with an inside surface 605 defining a generally
hollow inside portion 610 and an outside surface 606. The sidewall
604 may be about 0.20 inches to about 0.75 inches thick. In some
embodiments, the sidewall 604 is about 0.25 inches thick. However,
because the mixing device 100 may be scaled to suit a particular
application, embodiments of the device having a sidewall 604 that
is thicker or thinner than the values provided are within the scope
of the present teachings. The sidewall 604 includes a first end
portion 612 and a second end portion 614 and a plurality of
through-holes 608 formed between the first end portion 612 and the
second end portion 614. Optionally, the outside surface 606 of the
sidewall 604 may include other features such as apertures,
projections, textures, and the like. The first end portion 612 has
a relieved portion 616 configured to receive a collar 618 and the
second end portion 614 has a relieved portion 620 configured to
receive a collar 622.
[0178] The rotor 600 is disposed inside the stator 700. The stator
700 has a sidewall 704 with an inside surface 705 defining a
generally hollow inside portion 710 into which the rotor 600 is
disposed. The sidewall 704 may be about 0.1 inches to about 0.3
inches thick. In some embodiments, the sidewall 604 is about 1.5
inches thick. The stator 700 may be non-rotatably coupled to the
housing 520 in a substantially stationary position. Alternatively,
the stator 700 may integrally formed with the housing 520. The
sidewall 704 has a first end portion 712 and a second end portion
714. Optionally, a plurality of apertures 708 are formed in the
sidewall 704 of the stator 700 between the first end portion 712
and the second end portion 714. Optionally, the inside surface 705
of the sidewall 704 may include other features such as
through-holes, projections, textures, and the like.
[0179] The rotor 600 rotates with respect to the stationary stator
700 about an axis of rotation ".alpha." in a direction indicated by
arrow "C3" in FIG. 9. Each of the rotor 600 and the stator 700 may
be generally cylindrical in shape and have a longitudinal axis. The
rotor 600 has an outer diameter "D1" and the stator 700 may have an
inner diameter "D2." The diameter "D1" may range, for example, from
about 0.5 inches to about 24 inches. In some embodiments, the
diameter "D1" is about 3.04 inches. In some embodiments, the
diameter "D1" is about 1.7 inches. The diameter "D2," which is
larger than the diameter "D1," may range from about 0.56 inches to
about 24.25 inches. In some embodiments, the diameter "D2" is about
4 inches. Therefore, the mixing chamber 330 may have a ring-shaped
cross-sectional shape that is about 0.02 inches to about 0.125
inches thick (i.e., the difference between the diameter "D2" and
the diameter "D1"). In particular embodiments, the mixing chamber
330 is about 0.025 inches thick. The channel 32 between the rotor
12 and the stator 34 of prior art device 10 (see FIG. 1) has a
ring-shaped cross-sectional shape that is about 0.09 inches thick.
Therefore, in particular embodiments, the thickness of the mixing
chamber 330 is less than about one third of the channel 32 of the
prior art device 10.
[0180] The longitudinal axis of the rotor 600 may be aligned with
its axis of rotation ".alpha.." The longitudinal axis of the rotor
600 may be aligned with the longitudinal axis of the stator 700.
The rotor 600 may have a length of about 3 inches to about 6 inches
along the axis of rotation ".alpha.." In some embodiments, the
rotor 600 may have a length of about 5 inches along the axis of
rotation ".alpha.." The stator 700 may have a length of about 3
inches to about 6 inches along the axis of rotation ".alpha.." In
some embodiments, the stator 700 may have a length of about 5
inches along the axis of rotation ".alpha.."
[0181] While the rotor 600 and the stator 700 have been depicted as
having a generally cylindrical shape, those of ordinary skill in
the art appreciate that alternate shapes may be used. For example,
the rotor 600 and the stator 700 may be conically, spherically,
arbitrarily shaped, and the like. Further, the rotor 600 and the
stator 700 need not be identically shaped. For example, the rotor
600 may be cylindrically shaped and the stator 700 rectangular
shaped or vice versa.
[0182] The apertures 708 of the stator 700 and the through-holes
608 depicted in FIGS. 4-7 are generally cylindrically shaped. The
diameter of the through-holes 608 may range from about 0.1 inches
to about 0.625 inches. The diameter of the apertures 708 may range
from about 0.1 inches to about 0.625 inches. One or more of
apertures 708 of the stator 700 may have a diameter that differs
from the diameters of the other apertures 708. For example, the
apertures 708 may increase in diameter from the first end portion
712 of the stator 700 to the second end portion 714 of the stator
700, the apertures 708 may decrease in diameter from the first end
portion 712 of the stator 700 to the second end portion 714 of the
stator 700, or the diameters of the apertures 708 may vary in
another manner along the stator 700. One or more of through-holes
608 of the rotor 600 may have a diameter that differs from the
diameters of the other through-holes 608. For example, the
through-holes 608 may increase in diameter from the first end
portion 612 of the rotor 600 to the second end portion 614 of the
rotor 600, the through-holes 608 may decrease in diameter from the
first end portion 612 of the rotor 600 to the second end portion
614 of the rotor 600, or the diameters of the through-holes 608 may
vary in another manner along the rotor 600.
[0183] As described below with reference to alternate embodiments,
the apertures 708 and the through-holes 608 may have shapes other
than generally cylindrical and such embodiments are within the
scope of the present invention. For example, the through-holes 608
may include a narrower portion, an arcuate portion, a tapered
portion, and the like. Referring to FIG. 7, each of the
through-holes 608 includes an outer portion 608A, a narrow portion
608B, and a tapered portion 608C providing a transition between the
outer portion 608A and the narrow portion 608B. Similarly, the
apertures 708 may include a narrower portion, an arcuate portion, a
tapered portion, and the like.
[0184] FIG. 8 provides a non-limiting example of a suitable
arrangement of the apertures 708 of the stator 700 and the
through-holes 608 of the rotor 600. The apertures 708 of the stator
700 may be arranged in substantially parallel lateral rows "SLAT-1"
through "SLAT-6" substantially orthogonal to the axis of rotation
".alpha.." The apertures 708 of the stator 700 may also be arranged
in substantially parallel longitudinal rows "SLONG-1" through
"SLONG-7" substantially parallel with the axis of rotation
".alpha.." In other words, the apertures 708 of the stator 700 may
be arranged in a grid-like pattern of orthogonal rows (i.e., the
lateral rows are orthogonal to the longitudinal rows) having the
longitudinal rows "SLONG-1" through "SLONG-7" substantially
parallel with the axis of rotation ".alpha.."
[0185] Like the apertures 708 of the stator 700, the through-holes
608 of the rotor 600 may be arranged in substantially parallel
lateral rows "RLAT-1" through "RLAT-6" substantially orthogonal to
the axis of rotation ".alpha.." However, instead of being arranged
in a grid-like pattern of orthogonal rows, the through-holes 608 of
the rotor 600 may also be arranged in substantially parallel rows
"RLONG-1" through "RLONG-7" that extend longitudinally along a
helically path. Alternatively, the through-holes 608 of the rotor
600 may also be arranged in substantially parallel rows "RLONG-1"
through "RLONG-7" that extend longitudinally at an angle other than
parallel with the axis of rotation ".alpha.."
[0186] The apertures 708 of the stator 700 and the through-holes
608 of the rotor 600 may be configured so that when the rotor 600
is disposed inside the stator 700 the lateral rows "SLAT-1" to
"SLAT-6" at least partially align with the lateral rows "RLAT-1" to
"RLAT-6," respectively. In this manner, as the rotor 600 rotates
inside the stator 700, the through-holes 608 pass by the apertures
708.
[0187] The through-holes 608 in each of the lateral rows "RLAT-1"
to "RLAT-6" may be spaced apart laterally such that all of the
through-holes 608 in the lateral row align, at least partially,
with the apertures 708 in a corresponding one of the lateral rows
"SLAT-1" to "SLAT-6" of the stator 700 at the same time. The
longitudinally extending rows "RLONG-1" through "RLONG-6" may be
configured such that the through-holes 608 in the first lateral row
"RLAT-1" in each of the longitudinally extending rows passes
completely by the apertures 708 of the corresponding lateral row
"SLAT-1" before the through-holes 608 in the last lateral row
"RLAT-6" begin to partially align with the apertures 708 of the
corresponding last lateral row "SLAT-6" of the stator 700.
[0188] While, in FIG. 8, six lateral rows and six longitudinally
extending rows have been illustrated with respect to the rotor 600
and six lateral rows and seven longitudinally extending rows have
been illustrated with respect stator 700, it is apparent to those
of ordinary skill in the art that alternate numbers of lateral rows
and/or longitudinal rows may be used with respect to the rotor 600
and/or stator 700 without departing from the present teachings.
[0189] To ensure that only one pair of openings between
corresponding lateral rows will be coincident at any one time, the
number of apertures 708 in each of the lateral rows "SLAT-1" to
"SLAT-6" on the stator 700 may differ by a predetermined number
(e.g., one, two, and the like) the number of through-holes 608 in
each of the corresponding lateral rows "RLAT-1" to "RLAT-6" on the
rotor 600. Thus, for example, if lateral row "RLAT-1" has twenty
through-holes 608 evenly spaced around the circumference of rotor
600, the lateral row "SLAT-1" may have twenty apertures 708 evenly
spaced around the circumference of stator 700.
[0190] Returning to FIG. 7, the mixing chamber 330 has an open
first end portion 332 and an open second end portion 334. The
through-holes 608 formed in the sidewall 604 of the rotor 600
connect the inside portion 610 of the rotor 600 with the mixing
chamber 330.
[0191] The rotor 600 is rotated inside the stator 700 by the drive
shaft 500 aligned with the axis of rotation ".alpha." of the rotor
600. The drive shaft 500 may be coupled to the first end portion
612 and the second end portion 614 of the rotor 600 and extend
through its hollow inside portion 610. In other words, a portion
720 of the drive shaft 500 is disposed in the hollow inside portion
610 of the rotor 600.
[0192] The collar 618 is configured to receive a portion 721 of the
drive shaft 500 disposed in the hollow inside portion 610 and the
collar 622 is configured to receive a portion 722 of the drive
shaft 500 disposed in the hollow inside portion 610.
[0193] The portion 721 has an outer diameter "D3" that may range
from about 0.5 inches to about 2.5 inches. In some embodiments, the
diameter "D3" is about 0.625 inches. The portion 722 has an outer
diameter "D4" that may be substantially similar to the diameter
"D3," although, this is not required. The diameter "D4" may range
from about 0.375 inches to about 2.5 inches.
[0194] The rotor 600 may be non-rotationally affixed to the portion
721 and the portion 722 of the drive shaft 500 by the collar 618
and the collar 622, respectively. By way of example, each of the
collars 618 and 622 may be installed inside relieved portions 616
and 620, respectively. Then, the combined rotor 600 and collars 618
and 622 may be heated to expand them. Next, the drive shaft 500 is
inserted through the collars 618 and 622 and the assembly is
allowed to cool. As the collars 618 and 622 shrink during cooling,
they tighten around the portions 722A and 722B of the drive shaft
500, respectively, gripping it sufficiently tightly to prevent the
drive shaft 500 from rotating relative to the rotor 600. The collar
618, which does not rotate with respect to either the portion 721
or the relieved portion 616, translates the rotation of the drive
shaft 500 to the first end portion 612 the rotor 600. The collar
622, which does not rotate with respect to either the portion 722
or the relieved portion 620, translates the rotation of the drive
shaft 500 to the second end portion 614 of the rotor 600. The drive
shaft 500 and the rotor 600 rotate together as a single unit.
[0195] The drive shaft 500 may have a first end portion 724 (see
FIG. 5) and a second end portion 726 (see FIG. 6). The first end
portion 724 may have a diameter "D5" of about 0.5 inches to about
1.75 inches. In particular embodiments, the diameter "D5" may be
about 1.25 inches. The second end portion 726 may have a diameter
"D6" that may be substantially similar to diameter "D5."
[0196] The second material 120 may be transported into the mixing
chamber 330 through one of the first end portion 724 and the second
end portion 726 of the rotating drive shaft 500. The other of the
first end portion 724 and the second end portion 726 of the drive
shaft 500 may be coupled to the motor 510. In the embodiment
depicted in FIGS. 5 and 6, the second material 120 is transported
into the mixing chamber 330 through the first end portion 724 and
the second end portion 726 of the drive shaft 500 is coupled to the
motor 510.
[0197] Turning to FIG. 5, the drive shaft 500 may have a channel
728 formed therein that extends from first end portion 724 into the
portion 720 disposed in the inside portion 610 of the rotor 600.
The channel 728 has an opening 730 formed in the first end portion
724. When the mixing device 100 is operating, the second material
120 is introduced into the channel 728 through the opening 730.
[0198] A valve 732 may be disposed inside a portion of the channel
728 located in the first end portion 724 of the drive shaft 500.
The valve 732 may restrict or otherwise control the backward flow
of the second material 120 from inside the hollow inside portion
610 through the channel 728 and/or the forward flow of the second
material 120 into the channel 728. The valve 732 may include any
valve known in the art including a check valve. A suitable check
valve includes a part number "CKFA1876205A," free flow forward
check valve, manufactured by The Lee Company USA having an office
in Bothell, Wash. and operating a website at www.theleeco.com.
[0199] The drive shaft 500 may include an aperture 740 located in
the inside portion 610 of the rotor 600 that connects the channel
728 with the inside portion 610 of the rotor 600. While only a
single aperture 740 is illustrated in FIG. 5, it is apparent to
those of ordinary skill in the art that multiple apertures may be
used to connect the channel 728 with the inside portion 610 of the
rotor 600.
[0200] Referring to FIG. 2, optionally, the external pump 220 may
pump the second material 120 into the mixing device 100. The pump
220 may include any suitable pump known in the art. By way of
non-limiting example, the pump 220 may include any suitable pump
known in the art including a diaphragm pump, a chemical pump, a
peristaltic pump, a gravity fed pump, a piston pump, a gear pump, a
combination of any of the aforementioned pumps, and the like. If
the second material 120 is a gas, the gas may be pressurized and
forced into the opening 730 formed in the first end portion 724 of
the drive shaft 500 by releasing the gas from the source 122.
[0201] The pump 220 or the source 122 is coupled to the channel 728
by the valve 732. The second material 120 transported inside the
channel 728 exits the channel 728 into the inside portion 610 of
the rotor 600 through the aperture 740. The second material 120
subsequently exits the inside portion 610 of the rotor 600 through
the through-holes 608 formed in the sidewall 608 of the rotor
600.
[0202] Referring to FIG. 5, the mixing device 100 may include a
seal assembly 750 coupled to the first end portion 724 of the drive
shaft 500. The seal assembly 750 is maintained within a chamber 752
defined in the housing 520. The chamber 752 has a first end portion
754 spaced across the chamber from a second end portion 756. The
chamber 752 also includes an input port 758 and an output port 759
that provide access into the chamber 752. The chamber 752 may be
defined by housing section 550 and the bearing housing 530. The
first end portion 754 may be formed in the housing section 550 and
the second end portion 756 may be adjacent to the bearing housing
530. The input port 758 may be formed in the bearing housing 530
and the output port 759 may be formed in the housing section
550.
[0203] The seal assembly 750 includes a first stationary seal 760
installed in the first end portion 754 of the chamber 752 in the
housing section 550 and the bearing housing 530. The first
stationary seal 760 extends around a portion 762 of the first end
portion 724 of the drive shaft 500. The seal assembly 750 also
includes a second stationary seal 766 installed in the second end
portion 756 of the chamber 752 in the bearing housing 530. The
second stationary seal 766 extends around a portion 768 of the
first end portion 724 of the drive shaft 500.
[0204] The seal assembly 750 includes a rotating assembly 770 that
is non-rotatably coupled to the first end portion 724 of the drive
shaft 500 between the portion 762 and the portion 768. The rotating
assembly 770 rotates therewith as a unit. The rotating assembly 770
includes a first seal 772 opposite a second seal 774. A biasing
member 776 (e.g., a spring) is located between the first seal 772
and the second seal 774. The biasing member 776 biases the first
seal 772 against the first stationary seal 760 and biases the
second seal 774 against the second stationary seal 766.
[0205] A cooling lubricant is supplied to the chamber 752 and
around rotating assembly 770. The lubricant enters the chamber 752
through the input port 758 and exits the chamber 752 through output
port 759. The lubricant may lubricate the bearing assembly 540
housed by the bearing housing 530. A chamber 570 may be disposed
between the bearing housing 530 and the mechanical seal housing
524. The bearing housing 530 may also include a second input port
759 connected to the chamber 570 into which lubricant may be
pumped. Lubricant pumped into the chamber 570 may lubricate the
bearing assembly 540. The seal assembly 750 may significantly, if
not greatly, reduce frictional forces within this portion of the
device caused by the rotation of the rotor 600 and may increase the
active life of the seals 770. The seals may include surfaces
constructed using silicon carbide.
[0206] Referring to FIG. 9, as the rotor 600 rotates about the axis
of rotation ".alpha." in the direction indicated by arrow "C1," the
rotor expels the second material 120 into the mixing chamber 330.
The expelled bubbles, droplets, particles, and the like of the
second material 120 exit the rotor 600 and are imparted with a
circumferential velocity (in a direction indicated by arrow "C3")
by the rotor 600. The second material 120 may be forced from the
mixing chamber 330 by the pump 220 (see FIG. 2), the centrifugal
force of the rotating rotor 600, buoyancy of the second material
120 relative to the first material 110, and a combination
thereof.
Motor 510
[0207] Returning to FIG. 6, the second end portion 726 of the drive
shaft 500 may be coupled to a rotating spindle 780 of a motor 510
by a coupler 900. The spindle 780 may have a generally circular
cross-sectional shape with a diameter "D7" of about 0.25 inches to
about 2.5 inches. In particular embodiments, the diameter "D7" may
be about 0.25 inches to about 1.5 inches. While in the embodiment
depicted in FIG. 6, the diameter "D5" of the first end portion 724
of the drive shaft 500 is substantially equal to the diameter "D7"
and the spindle 780, embodiments in which one of the diameter "D5"
and the diameter "D7" is larger than the other are within the scope
of the present invention.
[0208] Referring also to FIG. 4, it may be desirable to cover or
shield the coupler 900. In the embodiment illustrated in FIGS. 4
and 6, a drive guard 910 covers the coupler 900. The drive guard
910 may be generally U-shaped having a curved portion 914 flanked
by a pair of substantially linear portions 915 and 916. The distal
end of each of the substantially linear portions 915 and 916 of the
drive guard 910 may have a flange 918 and 919, respectively. The
drive guard 910 may be fastened by each of its flanges 918 and 919
to the base 106.
[0209] The motor 510 may be supported on the base 106 by a support
member 920. The support member 920 may be coupled to the motor 510
near the spindle 780. In the embodiment depicted, the support
member 920 includes a through-hole through which the spindle 780
passes. The support member 920 may be coupled to the motor 510
using any method known in the art, including bolting the support
member 920 to the motor 510 with one or more bolts 940.
[0210] The coupler 900 may include any coupler suitable for
transmitting a sufficient amount of torque from the spindle 780 to
the drive shaft 500 to rotate the rotor 600 inside to the stator
700. In the embodiment illustrated in FIGS. 4 and 6, the coupler
900 is a bellows coupler. A bellows coupler may be beneficial if
the spindle 780 and the drive shaft 500 are misaligned. Further,
the bellows coupler may help absorb axial forces exerted on the
drive shaft 500 that would otherwise be translated to the spindle
780. A suitable bellows coupler includes a model "BC32-8-8-A,"
manufactured by Ruland Manufacturing Company, Inc. of Marlborough,
Mass., which operates a website at www.ruland.com.
[0211] The motor 510 may rotate the rotor 600 at about 0.1
revolutions per minute ("rpm") to about 7200 rpm. The motor 510 may
include any motor suitable for rotating the rotor 600 inside to the
stator 700 in accordance with the present teachings. By way of
non-limiting example, a suitable motor may include a one-half
horsepower electric motor, operating at 230/460 volts and 3450 per
minute ("rpm"). A suitable motor includes a model "C4T34NC4C"
manufactured by LEESON Electric Corporation of Grafton, Wis., which
operates a website at www.leeson.com.
First Chamber 310
[0212] Turning to FIGS. 4 and 7, the first chamber 320 is disposed
inside the central section 522 of the housing 520 between the first
mechanical seal housing 524 and the first end portions 612 and 712
of the rotor 600 and the stator 700, respectively. The first
chamber 310 may be annular and have a substantially circular
cross-sectional shape. The first chamber 310 and the mixing chamber
330 form a continuous volume. A portion 1020 of the drive shaft 500
extends through the first chamber 310.
[0213] As may best be viewed in FIG. 4, the first chamber 310 has
an input port 1010 through which the first material 110 enters the
mixing device 100. The first material 110 may be pumped inside the
first chamber 310 by the external pump 210 (see FIG. 2). The
external pump 210 may include any pump known in the art for pumping
the first material 110 at a sufficient rate to supply the first
chamber 310.
[0214] The input port 1010 is oriented substantially orthogonally
to the axis of rotation ".alpha.." Therefore, the first material
110 enters the first chamber 310 with a velocity tangential to the
portion 1020 of the drive shaft 500 extending through the first
chamber 310. The tangential direction of the flow of the first
material 110 entering the first chamber 310 is identified by arrow
"T1." In the embodiment depicted in FIGS. 4 and 7, the input port
1010 may be offset from the axis of rotation ".alpha.." As is
apparent to those of ordinary skill in the art, the direction of
the rotation of the drive shaft 500 (identified by arrow "C1" in
FIG. 9), has a tangential component. The input port 1010 is
positioned so that the first material 110 enters the first chamber
310 traveling in substantially the same direction as the tangential
component of the direction of rotation of the drive shaft 500.
[0215] The first material 110 enters the first chamber 310 and is
deflected by the inside of the first chamber 310 about the portion
1020 of the drive shaft 500. In embodiments wherein the first
chamber 310 has a substantially circular cross-sectional shape, the
inside of the first chamber 310 may deflect the first material 110
in a substantially circular path (identified by arrow "C2" in FIG.
9) about the portion 1020 of the drive shaft 500. In such an
embodiment, the tangential velocity of the first material 110 may
cause it to travel about the axis of rotation ".alpha." at a
circumferential velocity, determined at least in part by the
tangential velocity.
[0216] Once inside the first chamber 310, the first material 110
may be pumped from the first chamber 310 into the mixing chamber
330 by the pump 410 residing inside the first chamber 310. In
embodiments that include the external pump 210 (see FIG. 2), the
external pump 210 may be configured to pump the first material 110
into the first chamber 310 at a rate at least as high as a rate at
which the pump 410 pumps the first material 110 from the first
chamber 310.
[0217] The first chamber 310 is in communication with the open
first end portion 332 of the mixing chamber 330 and the first
material 110 inside the first chamber 310 may flow freely into the
open first end portion 332 of the mixing chamber 330. In this
manner, the first material 110 does not negotiate any corners or
bends between the mixing chamber 330 and the first chamber 310. In
the embodiment depicted, the first chamber 310 is in communication
with the entire open first end portion 332 of the mixing chamber
330. The first chamber 310 may be filled completely with the first
material 110.
[0218] The pump 410 is powered by the portion 1020 of the drive
shaft 500 extending through the first chamber 310. The pump 410 may
include any pump known in the art having a rotating pump member
2022 housed inside a chamber (i.e., the first chamber 310) defined
by a stationary housing (i.e., the housing 520). Non-limiting
examples of suitable pumps include rotary positive displacement
pumps such as progressive cavity pumps, single screw pumps (e.g.,
Archimedes screw pump), and the like.
[0219] The pump 410 depicted in FIGS. 7 and 9, is generally
referred to as a single screw pump. In this embodiment, the pump
member 2022 includes a collar portion 2030 disposed around the
portion 1020 of the drive shaft 500. The collar portion 2030
rotates with the portion 1020 of the drive shaft 500 as a unit. The
collar portion 2030 includes one or more fluid displacement members
2040. In the embodiment depicted in FIGS. 7 and 9, the collar
portion 2030 includes a single fluid displacement member 2040
having a helical shape that circumscribes the collar portion 2030
along a helical path.
[0220] Referring to FIG. 9, the inside of the first chamber 310 is
illustrated. The pump 410 imparts an axial flow (identified by
arrow "A1" and arrow "A2") in the first material 110 inside the
first chamber 310 toward the open first end portion 332 of the
mixing chamber 330. The axial flow of the first material 110
imparted by the pump 410 has a pressure that may exceed the
pressure obtainable by the external pump of the prior art device 10
(see FIG. 1).
[0221] The pump 410 may also be configured to impart a
circumferential flow (identified by arrow "C2") in the first
material 110 as it travels toward the open first end portion 332 of
the mixing chamber 330. The circumferential flow imparted in the
first material 110 before it enters the mixing chamber 330 causes
the first material 110 to enter the mixing chamber 330 already
traveling in the desired direction at an initial circumferential
velocity. In the prior art device 10 depicted in FIG. 1, the first
material 110 entered the channel 32 of the prior art device 10
without a circumferential velocity. Therefore, the rotor 12 of the
prior art device 10 alone had to impart a circumferential flow into
the first material 110. Because the first material 110 is moving
axially, in the prior art device 10, the first material 110
traversed at least a portion of the channel 32 formed between the
rotor 12 and the stator 30 at a slower circumferential velocity
than the first material 110 traverses the mixing chamber 330 of the
mixing device 100. In other words, if the axial velocity of the
first material 110 is the same in both the prior art device 10 and
the mixing device 100, the first material 110 may complete more
revolutions around the rotational axis ".alpha." before traversing
the axial length of the mixing chamber 330, than it would complete
before traversing the axial length of the channel 32. The
additional revolutions expose the first material 110 (and combined
first material 110 and second material 120) to a substantially
larger portion of the effective inside surface 706 (see FIG. 7) of
the stator 700.
[0222] In embodiments including the external pump 210 (see FIG. 2),
the circumferential velocity imparted by the external pump 210
combined with the input port 1010 being oriented according to the
present teachings, may alone sufficiently increase the revolutions
of the first material 110 (and combined first material 110 and
second material 120) about the rotational axis ".alpha.." Further,
in some embodiments, the circumferential velocity imparted by the
pump 210 and the circumferential velocity imparted by the pump 410
combine to achieve a sufficient number of revolutions of the first
material 110 (and combined first material 110 and second material
120) about the rotational axis ".alpha.." As is appreciated by
those of ordinary skill in the art, other structural elements such
as the cross-sectional shape of the first chamber 310 may
contribute to the circumferential velocity imparted by the pump
210, the pump 410, and a combination thereof.
[0223] In an alternate embodiment depicted in FIG. 10, the pump 410
may include one or more vanes 2042 configured to impart a
circumferential flow in the first material 110 as it travels toward
the open first end portion 332 of the mixing chamber 330.
Second Chamber 320
[0224] Turning now to FIGS. 4 and 7, the second chamber 320 is
disposed inside the central section 522 of the housing 520 between
the second mechanical seal housing 526 and the second end portions
614 and 714 of the rotor 600 and the stator 700, respectively. The
second chamber 320 may be substantially similar to the first
chamber 310. However, instead of the input port 1010, the second
chamber 320 may include an output port 3010. A portion 3020 of the
drive shaft 500 extends through the second chamber 320.
[0225] The second chamber 320 and the mixing chamber 330 form a
continuous volume. Further, the first chamber 310, the mixing
chamber 330, and the second chamber 320 form a continuous volume.
The first material 110 flows through the mixing device 100 from the
first chamber 310 to the mixing chamber 330 and finally to the
second chamber 320. While in the mixing chamber 330, the first
material 110 is mixed with the second material 120 to form the
output material 102. The output material 102 exits the mixing
device 100 through the output port 3010. Optionally, the output
material 102 may be returned to the input port 1010 and mixed with
an additional quantity of the second material 120, the third
material 130, or a combination thereof.
[0226] The output port 3010 is oriented substantially orthogonally
to the axis of rotation ".alpha." and may be located opposite the
input port 1010 formed in the first chamber 310. The output
material 102 enters the second chamber 320 from the mixing chamber
330 having a circumferential velocity (in the direction indicated
by arrow "C3" in FIG. 9) imparted thereto by the rotor 600. The
circumferential velocity is tangential to the portion 3020 of the
drive shaft 500 extending through the second chamber 320. In the
embodiment depicted in FIGS. 4, 6, and 7, the output port 3010 may
be offset from the axis of rotation ".alpha.." The output port 3010
is positioned so that the output material 102, which enters the
second chamber 320 traveling in substantially the same direction in
which the drive shaft 500 is rotating (identified in FIG. 9 by
arrow "C1"), is traveling toward the output port 3010.
[0227] The output material 102 enters the second chamber 320 and is
deflected by the inside of the second chamber 320 about the portion
3020 of the drive shaft 500. In embodiments wherein the second
chamber 320 has a substantially circular cross-sectional shape, the
inside of the second chamber 320 may deflect the output material
102 in a substantially circular path about the portion 3020 of the
drive shaft 500.
[0228] Referring to FIG. 2, optionally, the output material 102 may
be pumped from inside the second chamber 320 by the external pump
430. The external pump 430 may include any pump known in the art
for pumping the output material 102 at a sufficient rate to avoid
limiting throughput of the mixing device 100. In such an
embodiment, the external pump 430 may introduce a tangential
velocity (in a direction indicated by arrow "T2" in FIGS. 4 and 11)
to at least a portion of the output material 102 as the external
pump 430 pumps the output material 102 from the second chamber 320.
The tangential velocity of the portion of the output material 102
may cause it to travel about the axis of rotation ".alpha." at a
circumferential velocity, determined in part by the tangential
velocity.
Pump 420
[0229] Turning to FIGS. 6 and 7, the pump 420 residing inside the
second chamber 320 may pump the output material 102 from the second
chamber 320 into the output port 3010 and/or from the mixing
chamber 330 into the second chamber 320. In embodiments that
include the external pump 430, the external pump 430 may be
configured to pump the output material 102 from the second chamber
320 at a rate at least as high as a rate at which the pump 420
pumps the output material 102 into the output port 3010.
[0230] The second chamber 320 is in communication with the open
second end portion 334 of the mixing chamber 330 and the output
material 102 inside the mixing chamber 330 may flow freely from the
open second end portion 334 into the second chamber 320. In this
manner, the output material 102 does not negotiate any corners or
bends between the mixing chamber 330 and the second chamber 320. In
the embodiment depicted, the second chamber 320 is in communication
with the entire open second end portion 334 of the mixing chamber
330. The second chamber 320 may be filled completely with the
output material 102.
[0231] The pump 420 is powered by the portion 3020 of the drive
shaft 500 extending through the second chamber 320. The pump 420
may be substantially identical to the pump 410. Any pump described
above as suitable for use as the pump 410 may be used for the pump
420. While the pump 410 pumps the first material 110 into the
mixing chamber 330, the pump 420 pumps the output material 102 from
the mixing chamber 330. Therefore, both the pump 410 and the pump
420 may be oriented to pump in the same direction.
[0232] As is appreciated by those of ordinary skill in the art, the
first material 110 may differ from the output material 102. For
example, one of the first material 110 and the output material 102
may be more viscous than the other. Therefore, the pump 410 may
differ from the pump 420. The pump 410 may be configured to
accommodate the properties of the first material 110 and the pump
420 may be configured to accommodate the properties of the output
material 102.
[0233] The pump 420 depicted in FIGS. 6 and 7, is generally
referred to as a single screw pump. In this embodiment, the pump
member 4022 includes a collar portion 4030 disposed around the
portion 3020 of the drive shaft 500. The collar portion 4030
rotates with the portion 3020 of the drive shaft 500 as a unit. The
collar portion 4030 includes one or more fluid displacement members
4040. The collar portion 4030 includes a single fluid displacement
member 4040 having a helical shape that circumscribes the collar
portion 4030 along a helical path.
[0234] Referring to FIG. 11, the inside of the second chamber 320
is illustrated. The pump 420 imparts an axial flow (identified by
arrow "A3" and arrow "A4") in the output material 102 inside the
second chamber 320 away from the open second end portion 334 of the
mixing chamber 330.
[0235] The pump 420 may be configured to impart a circumferential
flow (identified by arrow "C4") in the output material 102 as it
travels away from the open second end portion 334 of the mixing
chamber 330. The circumferential flow imparted in the output
material 102 may help reduce an amount of work required by the
rotor 600. The circumferential flow also directs the output
material 102 toward the output port 3010.
[0236] In an alternate embodiment, the pump 420 may have
substantially the same configuration of the pump 410 depicted in
FIG. 10. In such an embodiment, the one or more vanes 2042 are
configured to impart a circumferential flow in the output material
102 as it travels away from the open second end portion 334 of the
mixing chamber 330.
[0237] As is apparent to those of ordinary skill, various
parameters of the mixing device 100 may be modified to obtain
different mixing characteristics. Exemplary parameters that may be
modified include the size of the through-holes 608, the shape of
the through-holes 608, the arrangement of the through-holes 608,
the number of through-holes 608, the size of the apertures 708, the
shape of the apertures 708, the arrangement of the apertures 708,
the number of apertures 708, the shape of the rotor 600, the shape
of the stator 700, the width of the mixing chamber 330, the length
of the mixing chamber 330, rotational speed of the drive shaft 500,
the axial velocity imparted by the internal pump 410, the
circumferential velocity imparted by the internal pump 410, the
axial velocity imparted by the internal pump 420, the
circumferential velocity imparted by the internal pump 420, the
configuration of disturbances (e.g., texture, projections,
recesses, apertures, and the like) formed on the outside surface
606 of the rotor 600, the configuration of disturbances (e.g.,
texture, projections, recesses, apertures, and the like) formed on
the inside surface 706 of the stator 700, and the like.
Alternate Embodiment
[0238] Referring to FIG. 12, a mixing device 5000 is depicted. The
mixing device 5000 is an alternate embodiment of the mixing device
100. Identical reference numerals have been used herein to identify
components of the mixing device 5000 that are substantially similar
corresponding components of the mixing device 100. Only components
of the mixing device 5000 that differ from the components of the
mixing device 100 will be described.
[0239] The mixing device 5000 includes a housing 5500 for housing
the rotor 600 and the stator 5700. The stator 5700 may be
non-rotatably couple by its first end portion 5712 and its second
end portion 5714 to the housing 5500. A chamber 5800 is defined
between the housing 5500 and a portion 5820 of the stator 5700
flanked by the first end portion 5712 and the second end portion
5714. The housing 5500 includes an input port 5830 which provides
access into the chamber 5800. The input port 5830 may be oriented
substantially orthogonally to the axis of rotation ".alpha.."
however, this is not a requirement.
[0240] The stator 5700 includes a plurality of through-holes 5708
that connect the chamber 5800 and the mixing chamber 330 (defined
between the rotor 600 and the stator 5700). An external pump 230
may be used to pump the third material 130 (which may be identical
to the second material 120) into the chamber 5800 via the input
port 5830. The third material 130 pumped into the chamber 5800 may
enter the mixing chamber 330 via the through-holes 5708 formed in
the stator 5700. The third material 130 may be forced from the
channel 5800 by the pump 230, buoyancy of the third material 130
relative to the first material 110, and a combination thereof. As
the rotor 600 rotates, it may also draw the third material 130 from
the channel 5800 into the mixing chamber 330. The third material
130 may enter the mixing chamber 330 as bubbles, droplets,
particles, and the like, which are imparted with a circumferential
velocity by the rotor 600.
Alternate Embodiment
[0241] An alternate embodiment of the mixing device 100 may be
constructed using a central section 5900 depicted in FIG. 13 and a
bearing housing 5920 depicted in FIG. 14. FIG. 13 depicts the
central section 5900 having in its interior the stator 700 (see
FIG. 7). Identical reference numerals have been used herein to
identify components associated with the central section 5900 that
are substantially similar corresponding components of the mixing
device 100. Only components of the central section 5900 that differ
from the components of the central section 522 will be described.
The central section 5900 and the stator 700 are both constructed
from a conductive material such as a metal (e.g., stainless steel).
The input port 1010 and the output port 3010 are both constructed
from a nonconductive material such as plastic (e.g., PET, Teflon,
nylon, PVC, polycarbonate, ABS, Delrin, polysulfone, etc.).
[0242] An electrical contact 5910 is coupled to the central section
5900 and configured to deliver a charge thereto. The central
section 5900 conducts an electrical charge applied to the
electrical contact 5910 to the stator 700. In further embodiments,
the central section 5900 may be constructed from a nonconductive
material. In such embodiments, the electrical contact 5910 may pass
through the central section 5900 and coupled to the stator 700. The
electric charge applied by the electrical contact 5910 to the
stator 700 may help facilitate redox or other chemical reactions
inside the mixing chamber 330.
[0243] Optionally, insulation (not shown) may be disposed around
the central section 5900 to electrically isolate it from the
environment. Further, insulation may be used between the central
section 5900 and the first and second mechanical seals 524 and 526
that flank it to isolate it electrically from the other components
of the mixing device.
[0244] Turning now to FIG. 14, the bearing housing 5920 will be
described. The bearing housing 5920 is disposed circumferentially
around the portion 726 of the drive shaft 500. An electrical
contact 5922 is coupled to the bearing housing 5920. A rotating
brush contact 5924 provides an electrical connection between the
drive shaft 500 and the electrical contact 5922.
[0245] In this embodiment, the drive shaft 500 and the rotor 600
are both constructed from a conductive material such as a metal
(e.g., stainless steel). The bearing housing 5920 may be
constructed from either a conductive or a nonconductive material.
An electrical charge is applied to the drive shaft 500 by the
electrical contact 5922 and the rotating brush contact 5924. The
electrical charge is conducted by the drive shaft 500 to the rotor
600.
[0246] The alternate embodiment of the mixing device 100
constructed using the central section 5900 depicted in FIG. 13 and
the bearing housing 5920 depicted in FIG. 14 may be operated in at
least two ways. First, the electrical contacts 5910 and 5922 may be
configured not to provide an electrical charge to the stator 700
and the rotor 600, respectively. In other words, neither of the
electrical contacts 5910 and 5922 are connected to a current
source, a voltage source, and the like.
[0247] Alternatively, the electrical contacts 5910 and 5922 may be
configured to provide an electrical charge to the stator 700 and
the rotor 600, respectively. For example, the electrical contacts
5910 and 5922 may be coupled to a DC voltage source (not shown)
supplying a steady or constant voltage across the electrical
contacts 5910 and 5922. The negative terminal of the DC voltage
source may be coupled to either of the electrical contacts 5910 and
5922 and the positive terminal of the DC voltage source may be
coupled to the other of the electrical contacts 5910 and 5922. The
voltage supplied across the electrical contacts 5910 and 5922 may
range from about 0.0001 volts to about 1000 volts. In particular
embodiments, the voltage may range from about 1.8 volts to about
2.7 volts. By way of another example, a pulsed DC voltage having a
duty cycle of between about 1% to about 99% may be used.
[0248] While the above examples of methods of operating the mixing
device apply a DC voltage across the electrical contacts 5910 and
5922, as is apparent to those of ordinary skill in the art, a
symmetrical AC voltage or non symmetrical AC voltage having various
shapes and magnitudes may be applied across the electrical contacts
5910 and 5922 and such embodiments are within the scope of the
present invention.
Mixing Inside the Mixing Chamber 330
[0249] As mentioned above, in the prior art device 10 (shown in
FIG. 1), the first material 110 entered the channel 32 between the
rotor 12 and the stator 30 via a single limited input port 37
located along only a portion of the open second end of the channel
32. Likewise, the output material 102 exited the channel 32 via a
single limited output port 40 located along only a portion of the
open first end of the channel 32. This arrangement caused
undesirable and unnecessary friction. By replacing the single
limited inlet port 37 and the single limited outlet port 40 with
the chambers 310 and 320, respectively, friction has been reduced.
Moreover, the first material 110 does not negotiate a corner before
entering the mixing chamber 330 and the output material 102 does
not negotiate a corner before exiting the mixing chamber 330.
Further, the chambers 310 and 320 provide for circumferential
velocity of the material prior to entering, and after exiting the
channel 32.
[0250] Accordingly, pressure drop across the mixing device 100 has
been substantially reduced. In the embodiments depicted in FIGS. 2,
4-9, and 11, the pressure drop between the input port 1010 and the
output port 3010 is only approximately 12 psi when the mixing
device 100 is configured to produce about 60 gallons of the output
material 102 per minute. This is an improvement over the prior art
device 10 depicted in FIG. 1, which when producing about 60 gallons
of output material per minute was at least 26 psi. In other words,
the pressure drop across the mixing device 100 is less than half
that experienced by the prior art device 10.
[0251] According to additional aspects, the inclusion of pumps 410
and 420, which are powered by the drive shaft 500, provides a
configuration that is substantially more efficient in mixing
materials and that requires less energy than the external pumps
used in the prior art.
Micro-Cavitation
[0252] During operation of the mixing device 100, the input
materials may include the first material 110 (e.g., a fluid) and
the second material 120 (e.g., a gas). The first material 110 and
the second material 120 are mixed inside the mixing chamber 330
formed between the rotor 600 and the stator 700. Rotation of the
rotor 600 inside the stator 700 agitates the first material 110 and
the second material 120 inside the mixing chamber 330. The
through-holes 608 formed in the rotor 600 and/or the apertures 708
formed in the stator 700 impart turbulence in the flow of the first
material 110 and the second material 120 inside the mixing chamber
330.
[0253] Without being limited by theory, the efficiency and
persistence of the diffusion of the second material 120 into the
first material 110 is believed to be caused in part by
micro-cavitation, which is described in connection with FIGS.
15-17. Whenever a material flows over a smooth surface, a rather
laminar flow is established with a thin boundary layer that is
stationary or moving very slowly because of the surface tension
between the moving fluid and the stationary surface. The
through-holes 608 and optionally, the apertures 708, disrupt the
laminar flow and can cause localized compression and decompression
of the first material 110. If the pressure during the decompression
cycle is low enough, voids (cavitation bubbles) will form in the
material. The cavitation bubbles generate a rotary flow pattern
5990, like a tornado, because the localized area of low pressure
draws the host material and the infusion material, as shown in FIG.
15. When the cavitation bubbles implode, extremely high pressures
result. As two aligned openings (e.g., one of the apertures 708 and
one of the through-holes 608) pass one another, a succussion (shock
wave) occurs, generating significant energy. The energy associated
with cavitation and succussion mixes the first material 110 and the
second material 120 together to an extremely high degree, perhaps
at the molecular level.
[0254] The tangential velocity of the rotor 600 and the number of
openings that pass each other per rotation may dictate the
frequency at which the mixing device 100. It has been determined
that operating the mixing device 100 within in the ultrasonic
frequency range can be beneficial in many applications. It is
believed that operating the mixing device 100 in the ultrasonic
region of frequencies provides the maximum succession shock energy
to shift the bonding angle of the fluid molecule, which enables it
to transport an additional quantity of the second material 120
which it would not normally be able to retain. When the mixing
device 100 is used as a diffuser, the frequency at which the mixing
device 100 operates appears to affect the degree of diffusion,
leading to much longer persistence of the second material 120
(infusion material) in the first material 110 (host material).
[0255] Referring now to FIG. 15, an alternate embodiment of the
rotor 600, rotor 6000 is provided. The cavitations created within
the first material 110 in the mixing chamber 330 may be configured
to occur at different frequencies along the length of the mixing
chamber 330. The frequencies of the cavitations may be altered by
altering the number and/or the placement of the through-holes 6608
along the length of the rotor 600. Each of the through-holes 6608
may be substantially similar to the through-holes 608 (discussed
above).
[0256] By way of non-limiting example, the rotor 6000 may be
subdivided into three separate exemplary sections 6100, 6200, and
6300. The through-holes 6608 increase in density from the section
6100 to the section 6200, the number of holes in the section 6100
being greater than the number of holes in the section 6200. The
through-holes 6608 also increase in density from the section 6200
to the section 6300, the number of holes in the section 6200 being
greater than the number of holes in the section 6300. Each of the
sections 6100, 6200, and 6300 create succussions within their
particular area at a different frequency due to the differing
numbers of through-holes 6608 formed therein.
[0257] By manufacturing the rotor 6000 with a desired number of
through-holes 6608 appropriately arranged in a particular area, the
desired frequency of the succussions within the mixing chamber 330
may be determined. Similarly, the desired frequency of the
cavitations may be determined by a desired number of apertures 708
appropriately arranged in a particular area upon the stator 700
within which the rotor 600 rotates. Further, the desired frequency
(or frequencies) of the succussions within the mixing chamber 330
may be achieved by selecting both a particular number and
arrangement of the apertures 708 formed in the stator 700 and a
particular number and arrangement of the through-holes 608 formed
in the rotor 600.
[0258] FIGS. 19-21, depict various alternative arrangements of the
apertures 708 formed in the stator 700 and the through-holes 608
formed in the rotor 600 configured to achieve different results
with respect to the cavitations created. FIG. 19 illustrates a
configuration in which the apertures 708 and the through-holes 608
are aligned along an axis 7000 that is not parallel with any line
(e.g., line 7010) drawn through the axis of rotation ".alpha." of
the rotor 600. In other words, if the rotor 600 has a cylindrical
shape, the axis 7000 does not pass through the center of the rotor
600. Thus, the first material 110 within the mixing chamber 330
will not be oriented perpendicularly to the compressions and
decompressions created by the apertures 708 and the through-holes
608. The compressions and decompressions will instead have a force
vector that has at least a component parallel to the
circumferential flow (in the direction of arrow "C3" of FIG. 9) of
first material 110 within the mixing chamber 330.
[0259] Relative alignment of the apertures 708 and the
through-holes 608 may also affect the creation of cavitations in
the mixing chamber 330. FIG. 20 illustrates an embodiment in which
the apertures 708 are in registration across the mixing chamber 330
with the through-holes 608. In this embodiment, rotation of the
rotor 600 brings the through-holes 608 of the rotor into direct
alignment with the apertures 708 of the stator 700. When in direct
alignment with each other, the compressive and decompressive forces
created by the apertures 708 and the through-holes 608 are directly
aligned with one another.
[0260] In the embodiment depicted in FIG. 21, the apertures 708 and
the through-holes 608 are offset by an offset amount "X" along the
axis of rotation ".alpha..". By way of non-limiting example, the
offset amount "X" may be determined as a function of the size of
the apertures 708. For example, the offset amount "X" may be
approximately equal to one half of the diameter of the apertures
708. Alternatively, the offset amount "X" may be determined as a
function of the size of the through-holes 608. For example, the
offset amount "X" may be approximately equal to one half of the
diameter of the through-holes 608. If features (e.g., recesses,
projections, etc.) other than or in addition to the through-holes
608 and the apertures 708 are included in either the rotor 600 or
the stator 700, the offset amount "X" may be determined as a
function of the size of such features. In this manner, the
compressive and decompressive forces caused by the apertures 708 of
the stator 700 and the through-holes 608 of the rotor 600 collide
at a slight offset causing additional rotational and torsional
forces within the mixing chamber 330. These additional forces
increase the mixing (e.g., diffusive action) of the second material
120 into the first material 110 within the mixing chamber 330.
[0261] Referring now to FIGS. 22-25, non-limiting examples of
suitable cross-sectional shapes for the apertures 708 and the
through-holes 608 are provided. The cross-sectional shape of the
apertures 708 and/or the through-holes 608 may be square as
illustrated in FIG. 22, circular as illustrated in FIG. 23, and the
like.
[0262] Various cross-sectional shapes of apertures 708 and/or the
through-holes 608 may be used to alter flow of the first material
110 as the rotor 600 rotates within the stator 700. For example,
FIG. 24 depicts a teardrop cross-sectional shape having a narrow
portion 7020 opposite a wide portion 7022. If the through-holes 608
have this teardrop shape, when the rotor 600 is rotated (in the
direction generally indicated by the arrow "F"), the forces exerted
on the first material 110, the second material 120, and optionally
the third material 130 within the mixing chamber 330 increase as
the materials pass from the wide portion 7022 of the teardrop to
the narrow portion 7020.
[0263] Additional rotational forces can be introduced into the
mixing chamber 330 by forming the apertures 708 and/or the
through-holes 608 with a spiral configuration as illustrated in
FIG. 25. Material that flows into and out of the apertures 708
and/or the through-holes 608 having the spiral configuration
experience a rotational force induced by the spiral configuration.
The examples illustrated in FIGS. 25-28 are provided as
non-limiting illustrations of alternate embodiments that may be
employed within the mixing device 100. By application of ordinary
skill in the art, the apertures 708 and/or the through-holes 608
may be configured in numerous ways to achieve various succussive
and agitative forces appropriate for mixing materials within the
mixing chamber 330.
Double Layer Effect
[0264] The mixing device 100 may be configured to create the output
material 102 by complex and non-linear fluid dynamic interaction of
the first material 110 and the second material 120 with complex,
dynamic turbulence providing complex mixing that further favors
electrokinetic effects (described below). The result of these
electrokinetic effects may be observed within the output material
102 as charge redistributions and redox reactions, including in the
form of solvated electrons that are stabilized within the output
material.
[0265] Ionization or dissociation of surface groups and/or
adsorption of ions from a liquid cause most solid surfaces in
contact with the liquid to become charged. Referring to FIG. 26, an
electrical double layer ("EDL") 7100 forms around exemplary surface
7110 in contact with a liquid 7120. In the EDL 7100, ions 7122 of
one charge (in this case, negatively charged ions) adsorb to the
surface 7120 and form a surface layer 7124 typically referred to as
a Stern layer. The surface layer 7124 attracts counterions 7126 (in
this case, positively charged ions) of the opposite charge and
equal magnitude, which form a counterion layer 7128 below the
surface layer 7124 typically referred to as a diffuse layer. The
counterion layer 7128 is more diffusely distributed than the
surface layer 7124 and sits upon a uniform and equal distribution
of both ions in the bulk material 7130 below. For OH-- and H+ ions
in neutral water, the Gouy-Chapman model would suggest that the
diffuse counterion layer extends about one micron into the
water.
[0266] According to particular aspects, the electrokinetic effects
mentioned above are caused by the movement of the liquid 7120 next
to the charged surface 7110. Within the liquid 7120 (e.g., water,
saline solution, and the like), the adsorbed ions 7122 forming the
surface layer 7124 are fixed to the surface 7120 even when the
liquid 7120 is in motion (for example, flowing in the direction
indicated by arrow "G"); however, a shearing plane 7132 exists
within the diffuse counterion layer 7128 spaced from the surface
7120. Thus, as the liquid 7120 moves, some of the diffuse
counterions 7126 are transported away from the surface 7120, while
the absorbed ions 7122 remain at the surface 7120. This produces a
so-called `streaming current.`
[0267] Within the mixing chamber 330, the first material 110, the
second material 120, and optionally, the third material 130 are
subject to an electromagnetic field created by the inside surface
705 of the stator 700 and/or the outside surface 606 of the rotor
600, a voltage between the inside surface 705 and the outside
surface 606, and/or an electrokinetic effect (e.g., streaming
current) caused by at least one EDL formed in the first material
110. The at least one EDL may be introduced into the first material
110 by at least one of the inside surface 705 of the stator 700 and
the outside surface 606 of the rotor 600.
[0268] Movement of the first material 110 through the mixing
chamber 330 relative to surface disturbances (e.g., the
through-holes 608 and apertures 708) creates cavitations in the
first material 110 within the mixing chamber 330, which may diffuse
the second material 120 into the first material 110. These
cavitations may enhance contact between of the first material 110
and/or the second material 120 with the electric double layer
formed on the inside surface 705 of the stator 700 and/or the
electric double layer formed on the outside surface 606 of the
rotor 600. Larger surface to volume ratios of the mixing chamber,
an increased dwell time of the combined materials within the mixing
chamber, and further in combination with a smaller average bubble
size (and hence substantially greater bubble surface area) provide
for effectively imparting EDL-mediated effects to the inventive
output materials.
[0269] In embodiments in which the inside surface 705 and the
outside surface 606 are constructed from a metallic material, such
as stainless steel, the motion of the liquid 7120 and/or the
streaming current(s) facilitate redox reactions involving H.sub.2O,
OH--, H+, and O.sub.2 at the inside surface 705 and the outside
surface 606.
[0270] Referring to FIG. 27, without being limited by theory, it is
believed a section 7140 of the mixing chamber 330 between the
inside surface 705 and the outside surface 606 the may be modeled
as a pair of parallel plates 7142 and 7144. If the first material
110 is a liquid, the first material 110 enters the section 7140
through an inlet "IN" and exits the section 7140 through an outlet
"OUT." The inlet "IN" and the outlet "OUT" restrict the flow into
and out of the section 7140.
[0271] Referring to FIG. 28, the area between the parallel plates
7142 and 7144 has a high surface area to volume ratio. Hence, a
substantial portion of the counterion layer 7128 (and counterions
7126) may be in motion as the first material 110 moves between the
plates 7142 and 7144. The number of counterions 7126 in motion may
exceed the number allowed to enter the section 7140 by the inlet
"IN" and the number allowed to exit the section 7140 by the outlet
"OUT." The inlet "IN" and the outlet "OUT" feeding and removing the
first material 110 from the section 7140, respectively, have far
less surface area (and a lower surface area to volume ratio) than
the parallel plates 7142 and 7144 and thereby reduce the portion of
the counterions 7126 in motion in the first material 110 entering
and leaving the section 7140. Therefore, entry and exit from the
section 7140 increases the streaming current locally. While a
background streaming current (identified by arrow "BSC") caused by
the flowing first material 110 over any surface is always present
inside the mixing device 100, the plates 7142 and 7144 introduce an
increased "excess" streaming current (identified by arrow "ESC")
within the section 7140.
[0272] Without a conductive return current (identified by arrow
"RC") in the plates 7142 and 7144 in the opposite direction of the
flow of the first material 110, an excess charge 7146 having the
same sign as the adsorbing ions 7122 would accumulate near the
inlet "IN," and an excess charge 7148 having the same sign as the
counterion 7126 would accumulate near the at outlet "OUT." Because
such accumulated charges 7146 and 7148, being opposite and
therefore attracted to one another, cannot build up indefinitely
the accumulated charges seek to join together by conductive means.
If the plates 7142 and 7144 are perfectly electrically insulating,
the accumulated charges 7146 and 7148 can relocate only through the
first material 110 itself. When the conductive return current
(identified by arrow "RC") is substantially equivalent to the
excess streaming current (identified by arrow "ESC") in the section
7140, a steady-state is achieved having zero net excess streaming
current, and an electrostatic potential difference between the
excess charge 7146 near the inlet "IN," and the excess charge 7148
near the outlet "OUT" creating a steady-state charge separation
therebetween.
[0273] The amount of charge separation, and hence the electrostatic
potential difference between the excess charge 7146 near the inlet
"IN," and the excess charge 7148 near the outlet "OUT," depends on
additional energy per unit charge supplied by a pump (e.g., the
rotor 600, the internal pump 410, and/or the external pump 210) to
"push" charge against the opposing electric field (created by the
charge separation) to produce the a liquid flow rate approximating
a flow rate obtainable by a liquid without ions (i.e., ions 7122
and 7126). If the plates 7142 and 7144 are insulators, the
electrostatic potential difference is a direct measure of the EMF
the pump (e.g., the rotor 600, the internal pump 410 and/or the
external pump 210) can generate. In this case, one could measure
the electrostatic potential difference using a voltmeter having a
pair of leads by placing one of the leads in the first material 110
near the inlet "IN," and the other lead in the first material 110
near the outlet "OUT."
[0274] With insulating plates 7142 and 7144, any return current is
purely an ion current (or flow of ions), in that the return current
involves only the conduction of ions through the first material
110. If other conductive mechanisms through more conductive
pathways are present between the excess charge 7146 near the inlet
"IN," and the excess charge 7148 near the outlet "OUT," the return
current may use those more conductive pathways. For example,
conducting metal plates 7142 and 7144 may provide more conductive
pathways; however, these more conductive pathways transmit only an
electron current and not the ion current.
[0275] As is appreciated by those of ordinary skill, to transfer
the charge carried by an ion to one or more electrons in the metal,
and vice versa, one or more oxidation-reduction reactions must
occur at the surface of the metal, producing reaction products.
[0276] Assuming the first material 110 is water (H.sub.2O) and the
second material 120 is oxygen (O.sub.2), a non-limiting example of
a redox reaction, which would inject negative charge into the
conducting plates 7142 and 7144 includes the following known
half-cell reaction:
O.sub.2+H.sub.2O.fwdarw.O.sub.3+2H.sup.++2e.sup.-,
[0277] Again, assuming the first material 110 is water (H.sub.2O)
and the second material 120 is oxygen (O.sub.2), a non-limiting
example of a redox reaction includes the following known half-cell
reaction, which would remove negative charge from the conducting
plates 7142 and 7144 includes the following known half-cell
reaction:
2H.sup.++e.sup.-H.sub.2,
[0278] With conducting metal plates 7142 and 7144, most of the
return current is believed to be an electron current, because the
conducting plates 7142 and 7144 are more conductive than the first
material 110 (provided the redox reactions are fast enough not to
be a limiting factor). For the conducting metal plates 7142 and
7144, a smaller charge separation accumulates between the inlet
"IN" and the outlet "OUT," and a much smaller electrostatic
potential exists therebetween. However, this does not mean that the
EMF is smaller.
[0279] As described above, the EMF is related to the energy per
unit charge the pump provides to facilitate the flow of the first
material 110 against the opposing electric field created by the
charge separation. Because the electrostatic potential is smaller,
the pump may supply less energy per unit charge to cause the first
material 110 to flow. However, the above example redox reactions do
not necessarily occur spontaneously, and thus may require a work
input, which may be provided by the pump. Therefore, a portion of
the EMF (that is not reflected in the smaller electrostatic
potential difference) may be used to provide the energy necessary
to drive the redox reactions.
[0280] In other words, the same pressure differentials provided by
the pump to push against the opposing electric field created by the
charge separation for the insulating plates 7142 and 7144, may be
used both to "push" the charge through the conducting plates 7142
and 7144 and drive the redox reactions.
[0281] Referring to FIG. 29, an experimental setup for an
experiment conducted by the inventors is provided. The experiment
included a pair of substantially identical spaced apart 500 ml
standard Erlenmeyer flasks 7150 and 7152, each containing a volume
of deionized water 7153. A rubber stopper 7154 was inserted in the
open end of each of the flasks 7150 and 7152. The stopper 7154
included three pathways, one each for a hollow tube 7156, a
positive electrode 7158, and a negative electrode 7160. With
respect to each of the flasks 7150 and 7152, each of the hollow
tube 7156, the positive electrode 7158, and the negative electrode
7160 all extended from outside the flask, through the stopper 7154,
and into the deionized water 7153 inside the flask. The positive
electrode 7158 and the negative electrode 7160 were constructed
from stainless steel. The hollow tubes 7156 in both of the flasks
7150 and 7152 had an open end portion 7162 coupled to a common
oxygen supply 7164. The positive electrode 7158 and the negative
electrode 7160 inserted into the flask 7152 where coupled to a
positive terminal and a negative terminal, respectively, of a DC
power supply 7168. Exactly the same sparger was used in each
flask.
[0282] Oxygen flowed through the hollow tubes 7156 into both of the
flasks 7150 and 7152 at a flow rate (Feed) of about 1 SCFH to about
1.3 SCFH (combined flow rate). The voltage applied across the
positive electrode 7158 and the negative electrode 7160 inserted
into the flask 7152 was about 2.55 volts. This value was chosen
because it is believed to be an electrochemical voltage value
sufficient to affect all oxygen species. This voltage was applied
continuously over three to four hours during which oxygen from the
supply 7164 was bubbled into the deionized water 7153 in each of
the flasks 7150 and 7152.
[0283] Testing of the deionized water 7153 in the flask 7150 with
HRP and pyrogallol gave an HRP-mediated pyrogallol reaction
activity, consistent with the properties of fluids produced with
the alternate rotor/stator embodiments described herein. The HRP
optical density was about 20% higher relative to pressure-pot or
fine-bubbled solutions of equivalent oxygen content. The results of
this experiment indicate that mixing inside the mixing chamber 330
involves a redox reaction. According to particular aspects, the
inventive mixing chambers provide for output materials comprising
added electrons that are stabilized by either oxygen-rich water
structure within the inventive output solutions, or by some form of
oxygen species present due to the electrical effects within the
process.
[0284] Additionally, the deionized water 7153 in both of the flasks
7150 and 7152 was tested for both ozone and hydrogen peroxide
employing industry standard colorimetric test ampules with a
sensitivity of 0.1 ppm for hydrogen peroxide and 0.6 ppm for ozone.
There was no positive indication of either species up to the
detection limits of those ampules.
[0285] Referring now to FIG. 30, there is illustrated the DO levels
in water enriched with oxygen in the mixing device 100 and stored
in a 500 ml thin-walled plastic bottle and a 1000 ml glass bottle
out to at least 365 days. Each of the bottles was capped and stored
at 65.degree. Fahrenheit. As can be seen in the figure, the DO
levels of the oxygen-enriched fluid remained fairly constant out to
at least 365 days.
[0286] Referring to FIG. 31, there is illustrated the DO levels in
water enriched with oxygen in the mixing device 100 and stored in a
500 ml plastic thin-walled bottle and a 1000 ml glass bottle. Both
bottles were refrigerated at 39.degree. Fahrenheit. Again, DO
levels of the oxygen-enriched fluid remained steady and decreased
only slightly out to at least 365 days.
[0287] Referring now to FIG. 32, there is illustrated the dissolved
oxygen levels in GATORADE.RTM. enriched with oxygen in the mixing
device 100 and stored in 32 oz. GATORADE.RTM. bottles having an
average temperature of 55.degree. Fahrenheit at capping. The
GATORADE.RTM. bottles were subsequently refrigerated at 38.degree.
Fahrenheit between capping and opening. During the experiment, a
different bottle was opened at 20, 60, and 90 days, respectively,
to measure the DO levels of the GATORADE.RTM. stored therein. Line
8102 represents the DO level of normal (i.e., unprocessed)
GATORADE.RTM. at 38.degree. Fahrenheit which is slightly less than
10 ppm.
[0288] The GATORADE.RTM. within a first group of GATORADE.RTM.
bottles was processed with oxygen in the mixing device 100 at
approximately 56.degree. Fahrenheit. The DO levels of the
GATORADE.RTM. at bottling were approximately 50 ppm as indicated by
point 8104. A first bottle was opened at approximately 20 days, and
the DO level of the GATORADE.RTM. was determined to be
approximately 47 ppm as indicated by point 8106. A second bottle
was then opened at 60 days, and the DO level of the GATORADE.RTM.
was measured to be approximately 44 ppm as indicated by point 8108.
Finally, a third bottle was opened at 90 days, and the DO level of
the GATORADE.RTM. was determined to be slightly below 40 ppm as
indicated by point 8110.
[0289] The GATORADE.RTM. within a second group of GATORADE.RTM.
bottles was processed with oxygen in the mixing device 100 at
approximately 52.degree. Fahrenheit. The initial DO level for
GATORADE.RTM. stored in this group of bottles was 45 ppm as
illustrated by point 8112. The GATORADE.RTM. in the bottle opened
at 20 days had a DO level of only slightly lower than 45 ppm as
indicated by point 8114. The second bottle of GATORADE.RTM. was
opened at 60 days and the GATORADE.RTM. therein had a DO level of
slightly more than 41 ppm. Finally, a third bottle of GATORADE.RTM.
was opened at 90 days and the GATORADE.RTM. therein had a DO level
of approximately 39 ppm as shown by point 8116. As before, with
respect to the water test in the plastic and glass bottles (see
FIG. 31), it can be seen that the DO levels remain at relatively
high levels over the 90 day period and substantially higher than
those levels present in normal (unprocessed) GATORADE.RTM. stored
in 32 oz. GATORADE.RTM. bottles. Point 8010 is the level
corresponding to inventive output fluid in a covered PET
bottle.
Dwell Time
[0290] Dwell time is an amount of time the first material 110, the
second material 120, and optionally the third material 130 spend in
the mixing chamber 330. The ratio of the length of the mixing
chamber 330 to the diameter of the mixing chamber 330 may
significantly affect dwell time. The greater the ratio, the longer
the dwell time. As mentioned in the Background Section, the rotor
12 of the prior art device 10 (see FIG. 1) had a diameter of about
7.500 inches and a length of about 6.000 inches providing a length
to diameter ratio of about 0.8. In contrast, in particular
embodiments, the length of the mixing chamber 330 of the mixing
device 100 is about 5 inches and the diameter "D1" of the rotor 600
is about 1.69 inches yielding a length to diameter ratio of about
2.95.
[0291] Dwell time represents the amount of time that the first
material 110, the second material 120, and optionally the third
material 130 are able to interact with the electrokinetic phenomena
described herein. The prior art device 10 is configured to produce
about 60 gallons of the output material 102 per minute and the
mixing device 100 is configured to produce about 0.5 gallons of the
output material 102 per minute, the prior art device 10 (see FIG.
1) had a fluid dwell time of about 0.05 seconds, whereas
embodiments of the mixing device 100 have a substantially greater
(about 7-times greater) dwell time of about 0.35 seconds. This
longer dwell time allows the first material 110, the second
material 120, and optionally the third material 130 to interact
with each other and the surfaces 606 and 705 (see FIG. 7) inside
the mixing chamber 330 for about 7 times longer than was possible
in the prior art device 10.
[0292] With reference to Table I below, the above dwell times were
calculated by first determining the flow rate for each device in
gallons per second. In the case of the prior art device 10 was
configured to operate at about 60 gallons of output material per
minute, while the mixing device 100 is configured to operate over a
broader range of flow rate, including at an optimal range of about
0.5 gallons of output material per minute. The flow rate was then
converted to cubic inches per second by multiplying the flow rate
in gallons per second by the number of cubic inches in a gallon
(i.e., 231 cubic inches). Then, the volume (12.876 cubic inches) of
the channel 32 of the prior art device 10 was divided by the flow
rate of the device (231 cubic inches/second) to obtain the dwell
time (in seconds) and the volume (0.673 cubic inches) of the mixing
chamber 330 of the mixing device 100 was divided by the flow rate
(1.925 cubic inches/second) of the device (in cubic inches per
second) to obtain the dwell time (in seconds).
TABLE-US-00002 TABLE I Table 1. Inventive device can accommodate a
range of dwell times, including a substantially increased (e.g.,
7-times) dwell time relative to prior art devices. Volume Flow
Mixing Rate Flow Rate Flow Rate Chamber Dwell Gallons/ Gallons/
Cubic Inches/ (Cubic Time Device Minute Second Second Inches)
(Seconds) Prior art 60 1.000 231.000 12.876 0.056 device 10 Mixing
2 0.033 7.700 0.673 0.087 device 100 Mixing 0.5 0.008 1.925 0.673
0.350 device 100
Rate of Infusion
[0293] Particular aspects of the mixing device 100 provide an
improved oxygen infusion rate over the prior art, including over
prior art device 10 (see FIG. 1). When the first material 110 is
water and the second material 120 is oxygen, both of which are
processed by the mixing device 100 in a single pass (i.e., the
return block of FIG. 2 is set to "NO") at or near 20.degree.
Celsius, the output material 102 has a dissolved oxygen level of
about 43.8 parts per million. In certain aspects, an output
material having about 43.8 ppm dissolved oxygen is created in about
350 milliseconds via the inventive flow through the inventive non
pressurized (non-pressure pot) methods. In contrast, when the first
material 110 (water) and the second material 120 (oxygen) are both
processed in a single pass at or near 20.degree. Celsius by the
prior art device 10, the output material had dissolved oxygen level
of only 35 parts per million in a single pass of 56
milliseconds.
Output Material 102
[0294] When the first material 110 is a liquid (e.g., freshwater,
saline, GATORADE.RTM., and the like) and the second material 120 is
a gas (e.g., oxygen, nitrogen, and the like), the mixing device 100
may diffuse the second material 120 into the first material 110.
The following discusses results of analyses performed on the output
material 102 to characterize one or more properties of the output
material 102 derived from having been processed by the mixing
device 100.
[0295] When the first material 110 is saline solution and the
second material 120 is oxygen gas, experiments have indicated that
a vast majority of oxygen bubbles produced within the saline
solution are no greater than 0.1 micron in size.
Decay of Dissolved Oxygen Levels
[0296] Referring now to FIG. 30, there is illustrated the DO levels
in water processed with oxygen in the mixing device 100 and stored
in a 500 ml thin-walled plastic bottle and a 1000 ml glass bottle.
Each of the bottles was capped and stored at 65.degree. Fahrenheit.
Point 7900 is the DO level at bottling. Line 7902 illustrates the
Henry's Law equilibrium state (i.e., the amount of dissolved oxygen
that should be within the water at 65.degree. Fahrenheit), which is
a DO level of slightly less than 10 ppm. Points 7904 and 7906
represent the DO levels within the water in the plastic bottle at
65 days and 95 days respectively. As can be seen at point 7904,
when the plastic bottle is opened approximately 65 days after
bottling, the DO level within the water is approximately 27.5 ppm.
When the bottle is opened approximately 95 days after bottling, as
indicated at point 7906, the DO level is approximately 25 ppm.
Likewise, for the glass bottle, the DO level is approximately 40
ppm at 65 days as indicated at point 7908 and is approximately 41
ppm at 95 days as illustrated at point 7910. Thus, FIG. 30
indicates the DO levels within both the plastic bottle and the
glass bottle remain relatively high at 65.degree. Fahrenheit.
[0297] Referring to FIG. 31, there is illustrated the DO levels in
water processed with oxygen in the mixing device 100 and stored in
a 500 ml plastic thin-walled bottle and a 1000 ml glass bottle.
Both bottles were refrigerated at 39.degree. Fahrenheit. The
Henry's Law equilibrium for water at 39.degree. Fahrenheit,
illustrated as line 8002, is approximately 14 ppm. The DO levels
observed in the water at bottling were slightly less than 40 ppm as
illustrated generally by point 8004. At approximately 30 days
between bottling and opening, the DO level of the water in the
plastic bottle has dropped slightly as indicated by point 8006 to
approximately 38 ppm. The DO level of the water in the glass bottle
has dropped slightly less as indicated generally by point 8012. At
approximately 65 days between bottling and opening, the DO level in
the plastic bottle has dropped to nearly 35 ppm as illustrated by
point 8008, and the DO level within the glass bottle as illustrated
generally by point 8014 has maintained a relatively constant value
slightly below 40 ppm. At just over 90 days between bottling and
opening, the DO level within the plastic bottle remains at
approximately 38 ppm while the DO level within the glass bottle has
risen to approximately 42 ppm as indicated by point 8016. Thus,
FIG. 31 illustrates that, at lower temperature levels, the DO
levels may be maintained at a high constant level in both the glass
bottle and the plastic bottle for a long period of time. Point 8010
is the level corresponding to inventive output fluid in a PET
bottle.
[0298] Referring now to FIG. 32, there is illustrated the dissolved
oxygen levels in GATORADE.RTM. processed with oxygen in the mixing
device 100 and stored in 32 oz. GATORADE.RTM. bottles having an
average temperature of 55.degree. Fahrenheit at capping. The
GATORADE.RTM. bottles were subsequently refrigerated at 38.degree.
Fahrenheit between capping and opening. During the experiment, a
different bottle was opened at 20, 60, and 90 days, respectively,
to measure the DO levels of the GATORADE.RTM. stored therein.
[0299] The GATORADE.RTM. within a first group of GATORADE.RTM.
bottles was processed with oxygen in the mixing device 100 at
approximately 56.degree. Fahrenheit. The DO levels of the
GATORADE.RTM. at bottling were approximately 50 ppm as indicated by
point 8104. A first bottle was opened at approximately 20 days, and
the DO level of the GATORADE.RTM. was determined to be
approximately 47 ppm as indicated by point 8106. A second bottle
was then opened at 60 days, and the DO level of the GATORADE.RTM.
was measured to be approximately 44 ppm as indicated by point 8108.
Finally, a third bottle was opened at 90 days, and the DO level of
the GATORADE.RTM. was determined to be slightly below 40 ppm as
indicated by point 8110.
[0300] The GATORADE.RTM. within a second group of GATORADE.RTM.
bottles was processed with oxygen in the mixing device 100 at
approximately 52.degree. Fahrenheit. The initial DO level for
GATORADE.RTM. stored in this group of bottles was 45 ppm as
illustrated by point 8112. The GATORADE.RTM. in the bottle opened
at 20 days had a DO level of only slightly lower than 45 ppm as
indicated by point 8114. The second bottle of GATORADE.RTM. was
opened at 60 days and the GATORADE.RTM. therein had a DO level of
slightly more than 41 ppm. Finally, a third bottle of GATORADE.RTM.
was opened at 90 days and the GATORADE.RTM. therein had a DO level
of approximately 39 ppm as shown by point 8116. As before, with
respect to the water test in the plastic and glass bottles (see
FIG. 30), it can be seen that the DO levels remain at relatively
high levels over the 90 day period and substantially higher than
those levels present in normal (unprocessed) GATORADE.RTM. stored
in 32 oz. GATORADE.RTM. bottles. Point 8010 is the level
corresponding to inventive output fluid in a covered PET
bottle.
[0301] FIG. 33 illustrates the DO retention of 500 ml of balanced
salt solution processed with oxygen in the mixing device 100 and
kept at standard temperature and pressure in an amber glass bottle.
The DO level of the solution before processing is 5 ppm. After
processing in the mixing device 100, the DO level was increased to
approximately 41 ppm (illustrated as point 8202). An hour after
processing, the DO level dropped to approximately 40 ppm as
indicated by point 8204. Two hours after processing, the DO level
dropped to approximately 36 ppm as indicated by point 8206. The DO
level dropped to approximately 34 ppm three hours after processing
as indicated by point 8208. At approximately four and a half hours
after processing, the DO level within the salt solution dropped to
slightly more than 30 ppm. The final measurement was taken shortly
before six hours after processing whereat the DO level had dropped
to approximately 28 ppm. Thus, each of the experiments illustrated
in FIGS. 30-33 illustrate that that the DO levels remain at
relatively high levels over extended periods.
[0302] Because the output material 102 may be consumed by human
beings, the materials used to construct the mixing device 100
should be suitable for food and/or pharmaceutical manufacture. By
way of non-limiting example, the housing 520, the housing 5520, the
rotor 600, the stator 700, and the stator 5700 may all be
constructed from stainless steel.
Molecular Interactions
[0303] A number of physicists have begun to describe the quantum
properties of water. Conventionally, quantum properties are thought
to belong to elementary particles of less than 10.sup.-10 meters,
while the macroscopic world of our everyday life is referred to as
classical, in that it behaves according to Newton's laws of motion.
Between the macroscopic classical world and the microscopic quantum
world is the mesoscopic domain, where the distinction between
macroscopic and microscopic is becoming increasingly blurred.
Indeed, physicists are discovering quantum properties in large
collections of atoms and molecules in the nanometer to micrometer
range, particularly when the molecules are packed closely together
in a liquid phase.
[0304] Recently, chemists have made a surprising discovery that
molecules form clusters that increase in size with dilution. These
clusters measure several micrometers in diameter. The increase in
size occurs non-linearly with dilution and depends on history,
flying in the face of classical chemistry. Indeed, there is yet no
explanation for this phenomena. It may well be yet another
reflection of the strangeness of water that depends on its quantum
properties.
[0305] In the mid 1990's, quantum physicist del Giudice and
Preparata and other colleagues at the University of Milan, in
Italy, argued that quantum coherent domains measuring 100
nanometers in diameter could arise in pure water. They show how the
collective vibrations of water molecules in the coherent domain
eventually become phase locked to the fluctuations of the global
electromagnetic field. In this way, long lasting, stable
oscillations could be maintained in water.
[0306] One way in which memory might be stored in water is through
the excitation of long lasting coherent oscillations specific to
one or more substances (such as a therapeutic agent) dissolved in
the water. Interactions between the water molecules and the
molecules of the substances dissolved in the water change the
collective structure of the water, which would in turn determine
the specific coherent oscillations that develop. If these
oscillations become stabilized and maintained by phase coupling
between the global field and the excited molecules, then, even when
the dissolved substances are diluted away, the water may still
carry the coherent oscillations that can seed other volumes of
water on dilution.
[0307] The discovery that dissolved substances form increasingly
large clusters is compatible with the existence of a coherent field
in water that can transmit attractive resonance between molecules
when the oscillations are in phase leading to clumping in dilute
solutions. As a cluster of molecules increases in size, its
electromagnetic signature is correspondingly amplified, reinforcing
the coherent oscillations carried by the water.
[0308] One should expect changes in some physical properties in
water that could be detectible. Unfortunately, all attempts to
detect such coherent oscillations by usual spectroscopic and
nuclear magnetic resonance methods have yielded ambiguous results.
This is not surprising in view of the finding that cluster size of
the dissolved molecules depends on the precise history of dilution
rather than concentration of the molecules.
[0309] It is possible that despite variations in the cluster size
of the dissolved molecules and detailed microscopic structure of
the water, a specificity of coherent oscillations may nonetheless
exist. Usual detection methods fail because they depend upon using
the microscopic particles of individual molecules, or of small
aggregates.
[0310] Instead, what is needed is a method of detecting collective
global properties over many, many molecules. Some obvious
possibilities that suggest themselves are the measurements of
freezing points and boiling points, viscosity, density,
diffusivity, and magnet properties. One possibility for detecting
changes in collective global properties of water is by means of
crystallization. Crystals are formed from macroscopic collections
of molecules. Like other measurements that depend on global
properties, crystals simplify the subtle changes in the individual
molecules that would have been undetectable otherwise.
[0311] With reference to FIG. 36, a simplified protonated water
cluster forming a nanoscale cage 8700 is shown. A protonated water
cluster typically takes the form of H.sup.+(H.sub.20).sub.n. Some
protonated water clusters occur naturally, such as in the
ionosphere. Without being bound by any particular theory, and
according to particular aspects, other types of water clusters or
structures (clusters, nanocages, etc.) are possible, including
structures comprising oxygen and stabilized electrons imparted to
the inventive output materials. Oxygen atoms 8704 may be caught in
the resulting structures 8700. The chemistry of the semi-bound
nanocage allows the oxygen 8704 and/or stabilized electrons to
remain dissolved for extended periods of time. Other atoms or
molecules, such as medicinal compounds, can be caged for sustained
delivery purposes. The specific chemistry of the solution material
and dissolved compounds depend on the interactions of those
materials.
[0312] Fluids processed by the mixing device 100 have been shown
via experiments to exhibit different structural characteristics
that are consistent with an analysis of the fluid in the context of
a cluster structure.
[0313] Water processed through the mixing device 100 has been
demonstrated to have detectible structural differences when
compared with normal unprocessed water. For example, processed
water has been shown to have more Rayleigh scattering than is
observed in unprocessed water. In the experiments that were
conducted, samples of processed and unprocessed water were prepared
(by sealing each in a separate bottle), coded (for later
identification of the processed sample and unprocessed sample), and
sent to an independent testing laboratory for analysis. Only after
the tests were completed were the codes interpreted to reveal which
sample had been processed by the mixing device 100.
[0314] At the laboratory, the two samples were placed in a laser
beam having a wavelength of 633 nanometers. The fluid had been
sealed in glass bottles for approximately one week before testing.
With respect to the processed sample, Sample B scattered light
regardless of its position relative to the laser source. However,
Sample A did not. After two to three hours following the opening of
the bottle, the scattering effect of Sample B disappeared. These
results imply the water exhibited a memory causing the water to
retain its properties and dissipate over time. These results also
imply the structure of the processed water is optically different
from the structure of the unprocessed fluid. Finally, these results
imply the optical effect is not directly related to DO levels
because the DO level at the start was 45 ppm and at the end of the
experiment was estimated to be approximately 32 ppm.
[0315] The presently disclosed system and methods allow gas (e.g.,
oxygen) to be enriched stably at a high concentration with minimal
passive loss. This system and methods can be effectively used to
enrich a wide variety of gases at heightened percentages into a
wide variety of fluids. By way of example only, deionized water at
room temperature that typically has levels of about 7-9 ppm (parts
per million) of dissolved oxygen can achieve levels of dissolved
oxygen ranging from about 8-70 ppm using the disclosed systems
and/or methods. In accordance with a particular exemplary
embodiment, oxygen-enriched water may be generated with levels of
about 30-60 ppm of dissolved oxygen.
[0316] Table 1 illustrates various partial pressure measurements
taken in a healing wound treated with an oxygen-enriched saline
solution and in samples of the gas-enriched oxygen-enriched saline
solution of the present invention.
TABLE-US-00003 TABLE 1 TISSUE OXYGEN MEASUREMENTS Probe Z082BO In
air: 171 mmHg 23.degree. C. Column Partial Pressure (mmHg) B1 32-36
B2 169-200 B3 20-180* B4 40-60 *wound depth minimal, majority
>150, occasional 20 s
[0317] Using a gas-enriching diffusion system in accordance with
the present disclosure it is possible to significantly increase the
amount of dissolved gas in most liquids. The system and method
allows gas (e.g., oxygen) to be dissolved stably at a high
concentration with minimal passive loss. This system and method can
be effectively used to incorporate a wide variety of gases at
heightened percentages into a wide variety of fluids. By way of
example only, a de-ionized water sample at room temperature that
typically has levels of about 2-3 ppm (parts per million) of
dissolved oxygen can achieve levels of dissolved oxygen ranging
from about 8-70 ppm using the disclosed gas-enriching system and/or
methods. In accordance with a disclosed embodiment, a gas-enriched
fluid may be generated with levels of about 30-60 ppm of dissolved
gas (e.g., oxygen).
Cosmetic and/or Therapeutic Application and Administration
[0318] In particular exemplary embodiments, the gas-enriched fluid
of the present invention may function as a cosmetic and/or
therapeutic composition alone or in combination with another
cosmetic and/or therapeutic agent such that the therapeutic
composition prevents or alleviates at least one symptom of a
wound-related disease or condition, or to increase proper wound
healing. The therapeutic compositions of the present invention
include compositions that are able to be administered to a subject
in need thereof. As used herein, "subject," may refer to any living
creature, preferably an animal, more preferably a mammal, and even
more preferably a human.
[0319] In certain embodiments, the composition formulation may also
comprise at least one additional agent selected from the group
consisting of: carriers, adjuvants, emulsifying agents, suspending
agents, sweeteners, flavorings, perfumes, and binding agents.
[0320] As used herein, "pharmaceutically acceptable carrier" and
"carrier" generally refer to a non-toxic, inert solid, semi-solid
or liquid filler, diluent, encapsulating material or formulation
auxiliary of any type. Some non-limiting examples of materials
which can serve as pharmaceutically acceptable carriers are sugars
such as lactose, glucose and sucrose; starches such as corn starch
and potato starch; cellulose and its derivatives such as sodium
carboxymethyl cellulose, ethyl cellulose and cellulose acetate;
powdered tragacanth; malt; gelatin; talc; excipients such as cocoa
butter and suppository waxes; oils such as peanut oil, cottonseed
oil; safflower oil; sesame oil; olive oil; corn oil and soybean
oil; glycols; such as propylene glycol; esters such as ethyl oleate
and ethyl laurate; agar; buffering agents such as magnesium
hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water;
isotonic saline; Ringer's solution; ethyl alcohol, and phosphate
buffer solutions, as well as other non-toxic compatible lubricants
such as sodium lauryl sulfate and magnesium stearate, as well as
coloring agents, releasing agents, coating agents, sweetening,
flavoring and perfuming agents, preservatives and antioxidants can
also be present in the composition, according to the judgment of
the formulator. In particular aspects, such carriers and excipients
may be gas-enriched fluids or solutions of the present
invention.
[0321] The pharmaceutically acceptable carriers described herein,
for example, vehicles, adjuvants, excipients, or diluents, are
well-known to those who are skilled in the art. Typically, the
pharmaceutically acceptable carrier is chemically inert to the
therapeutic agents and has no detrimental side effects or toxicity
under the conditions of use. The pharmaceutically acceptable
carriers can include polymers and polymer matrices, nanoparticles,
microbubbles, and the like.
[0322] In addition to the therapeutic gas-enriched fluid of the
present invention, the therapeutic composition may further comprise
inert diluents such as additional non-gas-enriched water or other
solvents, solubilizing agents and emulsifiers such as ethyl
alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl
alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol,
dimethylformamide, oils (in particular, cottonseed, groundnut,
corn, germ, olive, castor, and sesame oils), glycerol,
tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid
esters of sorbitan, and mixtures thereof. As is appreciated by
those of ordinary skill, a novel and improved formulation of a
particular therapeutic composition, a novel gas-enriched
therapeutic fluid, and a novel method of delivering the novel
gas-enriched therapeutic fluid may be obtained by replacing one or
more inert diluents with a gas-enriched fluid of identical,
similar, or different composition. For example, conventional water
may be replaced or supplemented by a gas-enriched fluid produced by
infusing oxygen into water or deionized water to provide
gas-enriched fluid.
[0323] Certain embodiments provide for therapeutic compositions
comprising a gas-enriched fluid of the present invention, a
pharmaceutical composition or other therapeutic agent or a
pharmaceutically acceptable salt or solvate thereof, and at least
one pharmaceutical carrier or diluent. These pharmaceutical
compositions may be used in the prophylaxis and treatment of the
foregoing diseases or conditions and in therapies as mentioned
above. Preferably, the carrier must be pharmaceutically acceptable
and must be compatible with, i.e., not have a deleterious effect
upon, the other ingredients in the composition. The carrier may be
a solid or liquid and is preferably formulated as a unit dose
formulation, for example, a tablet which may contain from 0.05 to
95% by weight of the active ingredient.
[0324] While the compositions and/or methods disclosed herein
generally relate to topical application, the most suitable means of
administration for a particular subject will depend on the nature
and severity of the disease or condition being treated or the
nature of the therapy being used, as well as the nature of the
therapeutic composition or additional therapeutic agent.
[0325] In addition to the therapeutic gas-enriched fluid of the
present invention, the therapeutic composition may further comprise
inert diluents such as additional non-gas-enriched water or other
solvents, solubilizing agents and emulsifiers such as ethyl
alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl
alcohol, benzyl benzoate, propylene glycol, 1,3-butylene glycol,
dimethylformamide, oils (in particular, cottonseed, groundnut,
corn, germ, olive, castor, and sesame oils), glycerol,
tetrahydrofurfuryl alcohol, polyethylene glycols and fatty acid
esters of sorbitan, and mixtures thereof. As is appreciated by
those of ordinary skill, a novel and improved formulation of a
particular therapeutic composition, a novel gas-enriched
therapeutic fluid, and a novel method of delivering the novel
gas-enriched therapeutic fluid may be obtained by replacing one or
more inert diluents with a gas-enriched fluid of identical,
similar, or different composition. For example, conventional water
may be replaced or supplemented by a gas-enriched fluid produced by
infusing oxygen into water or deionized water to provide
gas-enriched fluid.
[0326] In certain embodiments, the inventive gas-enriched fluid may
be combined with one or more therapeutic agents and/or used alone.
In particular embodiments, incorporating the gas-enriched fluid may
include replacing one or more solutions known in the art, such as
deionized water, saline solution, and the like with one or more
gas-enriched fluid, thereby providing an improved therapeutic
composition for delivery to the subject.
[0327] Certain embodiments provide for therapeutic compositions
comprising a gas-enriched fluid of the present invention, a
pharmaceutical composition or other therapeutic agent or a
pharmaceutically acceptable salt or solvate thereof, and at least
one pharmaceutical carrier or diluent. These pharmaceutical
compositions may be used in the prophylaxis and treatment of the
foregoing diseases or conditions and in therapies as mentioned
above. Preferably, the carrier must be pharmaceutically acceptable
and must be compatible with, i.e., not have a deleterious effect
upon, the other ingredients in the composition. The carrier may be
a solid or liquid and is preferably formulated as a unit dose
formulation, for example, a tablet which may contain from 0.05 to
95% by weight of the active ingredient.
[0328] Possible administration routes include oral, sublingual,
buccal, parenteral (for example subcutaneous, intramuscular,
intra-arterial, intraperitoneally, intracisternally,
intravesically, intrathecally, or intravenous), rectal, topical
including transdermal, intravaginal, intraoccular, intraotical,
intranasal, inhalation, and injection or insertion of implantable
devices or materials.
[0329] Formulations
[0330] Most suitable means of administration for a particular
subject will depend on the nature and severity of the disease or
condition being treated or the nature of the therapy being used, as
well as the nature of the therapeutic composition or additional
therapeutic agent. In certain embodiments, oral or topical
administration is preferred.
[0331] Formulations suitable for oral administration may be
provided as discrete units, such as tablets, capsules, cachets,
syrups, elixirs, chewing gum, "lollipop" formulations,
microemulsions, solutions, suspensions, lozenges, or gel-coated
ampules, each containing a predetermined amount of the active
compound; as powders or granules; as solutions or suspensions in
aqueous or non-aqueous liquids; or as oil-in-water or water-in-oil
emulsions.
[0332] Formulations suitable for transmucosal methods, such as by
sublingual or buccal administration include lozenges patches,
tablets, and the like comprising the active compound and, typically
a flavored base, such as sugar and acacia or tragacanth and
pastilles comprising the active compound in an inert base, such as
gelatin and glycerine or sucrose acacia.
[0333] Formulations suitable for parenteral administration
typically comprise sterile aqueous solutions containing a
predetermined concentration of the active gas-enriched fluid and
possibly another therapeutic agent; the solution is preferably
isotonic with the blood of the intended recipient. Additional
formulations suitable for parenteral administration include
formulations containing physiologically suitable co-solvents and/or
complexing agents such as surfactants and cyclodextrins.
Oil-in-water emulsions may also be suitable for formulations for
parenteral administration of the gas-enriched fluid. Although such
solutions are preferably administered intravenously, they may also
be administered by subcutaneous or intramuscular injection.
[0334] Formulations suitable for urethral, rectal or vaginal
administration include gels, creams, lotions, aqueous or oily
suspensions, dispersible powders or granules, emulsions,
dissolvable solid materials, douches, and the like. The
formulations are preferably provided as unit-dose suppositories
comprising the active ingredient in one or more solid carriers
forming the suppository base, for example, cocoa butter.
Alternatively, colonic washes with the gas-enriched fluids of the
present invention may be formulated for colonic or rectal
administration.
[0335] Formulations suitable for topical, intraoccular, intraotic,
or intranasal application include ointments, creams, pastes,
lotions, pastes, gels (such as hydrogels), sprays, dispersible
powders and granules, emulsions, sprays or aerosols using flowing
propellants (such as liposomal sprays, nasal drops, nasal sprays,
and the like) and oils. Suitable carriers for such formulations
include petroleum jelly, lanolin, polyethyleneglycols, alcohols,
and combinations thereof. Nasal or intranasal delivery may include
metered doses of any of these formulations or others. Likewise,
intraotic or intraocular may include drops, ointments, irritation
fluids and the like.
[0336] Formulations of the invention may be prepared by any
suitable method, typically by uniformly and intimately admixing the
gas-enriched fluid optionally with an active compound with liquids
or finely divided solid carriers or both, in the required
proportions and then, if necessary, shaping the resulting mixture
into the desired shape.
[0337] For example a tablet may be prepared by compressing an
intimate mixture comprising a powder or granules of the active
ingredient and one or more optional ingredients, such as a binder,
lubricant, inert diluent, or surface active dispersing agent, or by
molding an intimate mixture of powdered active ingredient and a
gas-enriched fluid of the present invention.
[0338] Suitable formulations for administration by inhalation
include fine particle dusts or mists which may be generated by
means of various types of metered dose pressurized aerosols,
nebulisers, or insufflators. In particular, powders or other
compounds of therapeutic agents may be dissolved or suspended in a
gas-enriched fluid of the present invention.
[0339] For pulmonary administration via the mouth, the particle
size of the powder or droplets is typically in the range 0.5-10
.mu.M, preferably 1-5 .mu.M, to ensure delivery into the bronchial
tree. For nasal administration, a particle size in the range 10-500
.mu.M is preferred to ensure retention in the nasal cavity.
[0340] Metered dose inhalers are pressurized aerosol dispensers,
typically containing a suspension or solution formulation of a
therapeutic agent in a liquefied propellant. In certain
embodiments, as disclosed herein, the gas-enriched fluids of the
present invention may be used in addition to or instead of the
standard liquefied propellant. During use, these devices discharge
the formulation through a valve adapted to deliver a metered
volume, typically from 10 to 150 .mu.L, to produce a fine particle
spray containing the therapeutic agent and the gas-enriched fluid.
Suitable propellants include certain chlorofluorocarbon compounds,
for example, dichlorodifluoromethane, trichlorofluoromethane,
dichlorotetrafluoroethane and mixtures thereof.
[0341] The formulation may additionally contain one or more
co-solvents, for example, ethanol surfactants, such as oleic acid
or sorbitan trioleate, anti-oxidants and suitable flavoring agents.
Nebulisers are commercially available devices that transform
solutions or suspensions of the active ingredient into a
therapeutic aerosol mist either by means of acceleration of a
compressed gas (typically air or oxygen) through a narrow venturi
orifice, or by means of ultrasonic agitation. Suitable formulations
for use in nebulisers consist of another therapeutic agent in a
gas-enriched fluid and comprising up to 40% w/w of the formulation,
preferably less than 20% w/w. In addition, other carriers may be
utilized, such as distilled water, sterile water, or a dilute
aqueous alcohol solution, preferably made isotonic with body fluids
by the addition of salts, such as sodium chloride. Optional
additives include preservatives, especially if the formulation is
not prepared sterile, and may include methyl hydroxy-benzoate,
anti-oxidants, flavoring agents, volatile oils, buffering agents
and surfactants.
[0342] Suitable formulations for administration by insufflation
include finely comminuted powders which may be delivered by means
of an insufflator or taken into the nasal cavity in the manner of a
snuff. In the insufflator, the powder is contained in capsules or
cartridges, typically made of gelatin or plastic, which are either
pierced or opened in situ and the powder delivered by air drawn
through the device upon inhalation or by means of a
manually-operated pump. The powder employed in the insufflator
consists either solely of the active ingredient or of a powder
blend comprising the active ingredient, a suitable powder diluent,
such as lactose, and an optional surfactant. The active ingredient
typically comprises from 0.1 to 100 w/w of the formulation.
[0343] In addition to the ingredients specifically mentioned above,
the formulations of the present invention may include other agents
known to those skilled in the art, having regard for the type of
formulation in issue. For example, formulations suitable for oral
administration may include flavoring agents and formulations
suitable for intranasal administration may include perfumes.
[0344] The therapeutic compositions of the invention can be
administered by any conventional method available for use in
conjunction with pharmaceutical drugs, either as individual
therapeutic agents or in a combination of therapeutic agents.
[0345] The dosage administered will, of course, vary depending upon
known factors, such as the pharmacodynamic characteristics of the
particular agent and its mode and route of administration; the age,
health and weight of the recipient; the nature and extent of the
symptoms; the kind of concurrent treatment; the frequency of
treatment; and the effect desired. A daily dosage of active
ingredient can be expected to be about 0.001 to 1000 milligrams
(mg) per kilogram (kg) of body weight, with the preferred dose
being 0.1 to about 30 mg/kg.
[0346] Dosage forms (compositions suitable for administration)
contain from about 1 mg to about 500 mg of active ingredient per
unit. In these pharmaceutical compositions, the active ingredient
will ordinarily be present in an amount of about 0.5-95% weight
based on the total weight of the composition.
[0347] Ointments, pastes, foams, occlusions, creams and gels also
can contain excipients, such as starch, tragacanth, cellulose
derivatives, silicones, bentonites, silica acid, and talc, or
mixtures thereof. Powders and sprays also can contain excipients
such as lactose, talc, silica acid, aluminum hydroxide, and calcium
silicates, or mixtures of these substances. Solutions of
nanocrystalline antimicrobial metals can be converted into aerosols
or sprays by any of the known means routinely used for making
aerosol pharmaceuticals. In general, such methods comprise
pressurizing or providing a means for pressurizing a container of
the solution, usually with an inert carrier gas, and passing the
pressurized gas through a small orifice. Sprays can additionally
contain customary propellants, such as nitrogen, carbon dioxide, or
other inert gases. In addition, microspheres or nanoparticles may
be employed with the gas-enriched therapeutic compositions or
fluids of the present invention in any of the routes required to
administer the therapeutic compounds to a subject.
[0348] The injection-use formulations can be presented in unit-dose
or multi-dose sealed containers, such as ampules and vials, and can
be stored in a freeze-dried (lyophilized) condition requiring only
the addition of the sterile liquid excipient, or gas-enriched
fluid, immediately prior to use. Extemporaneous injection solutions
and suspensions can be prepared from sterile powders, granules, and
tablets. The requirements for effective pharmaceutical carriers for
injectable compositions are well known to those of ordinary skill
in the art. See, for example, Pharmaceutics and Pharmacy Practice,
J. B. Lippincott Co., Philadelphia, Pa., Banker and Chalmers, Eds.,
238-250 (1982) and ASHP Handbook on Injectable Drugs, Toissel, 4th
ed., 622-630 (1986).
[0349] Formulations suitable for topical administration include
lozenges comprising a gas-enriched fluid of the invention and
optionally, an additional therapeutic and a flavor, usually sucrose
and acacia or tragacanth; pastilles comprising a gas-enriched fluid
and optional additional therapeutic agent in an inert base, such as
gelatin and glycerin, or sucrose and acacia; and mouth washes or
oral rinses comprising a gas-enriched fluid and optional additional
therapeutic agent in a suitable liquid carrier; as well as creams,
emulsions, gels and the like.
[0350] Additionally, formulations suitable for rectal
administration may be presented as suppositories by mixing with a
variety of bases such as emulsifying bases or water-soluble bases.
Formulations suitable for vaginal administration may be presented
as pessaries, tampons, creams, gels, pastes, foams, or spray
formulas containing, in addition to the active ingredient, such
carriers as are known in the art to be appropriate.
[0351] Suitable pharmaceutical carriers are described in
Remington's Pharmaceutical Sciences, Mack Publishing Company, a
standard reference text in this field.
[0352] The dose administered to a subject, especially an animal,
particularly a human, in the context of the present invention
should be sufficient to affect a therapeutic response in the animal
over a reasonable time frame. One skilled in the art will recognize
that dosage will depend upon a variety of factors including a
condition of the animal, the body weight of the animal, as well as
the condition being treated. A suitable dose is that which will
result in a concentration of the therapeutic composition in a
subject which is known to affect the desired response.
[0353] The size of the dose also will be determined by the route,
timing and frequency of administration as well as the existence,
nature, and extent of any adverse side effects that might accompany
the administration of the therapeutic composition and the desired
physiological effect.
[0354] Formulations suitable for topical application include
liquids (aqueous or oil based), ointments, creams, lotions, pastes,
gels (such as hydrogels), sprays, dispersible powders and granules,
emulsions, sprays or aerosols using flowing propellants (such as
liposomal sprays, nasal drops, nasal sprays, and the like) and
oils. Suitable carriers for such formulations include petroleum
jelly, lanolin, polyethylene glycol (such as PEG 3000, PEG 5000, or
other), alcohols, and combinations thereof.
[0355] Formulations of the invention may be prepared by any
suitable method, typically by uniformly and intimately admixing the
gas-enriched fluid optionally with an active compound with liquids
or finely divided solid carriers or both, in the required
proportions and then, if necessary, shaping the resulting mixture
into the desired shape.
[0356] Ointments, pastes, foams, occlusions, creams, gels, sols,
suspensions, and patches also can contain excipients, such as
starch, tragacanth, cellulose derivatives, silicones, bentonites,
silica acid, and talc, or mixtures thereof. Powders and sprays also
can contain excipients such as lactose, talc, silica acid, aluminum
hydroxide, and calcium silicates, or mixtures of these substances.
Solutions of nanocrystalline antimicrobial metals can be converted
into aerosols or sprays by any of the known means routinely used
for making aerosol pharmaceuticals. In general, such methods
comprise pressurizing or providing a means for pressurizing a
container of the fluid, usually with an inert carrier gas, and
passing the pressurized gas through a small orifice. Sprays can
additionally contain customary propellants, such as nitrogen,
carbon dioxide, or other inert gases. In addition, microspheres or
nanoparticles may be employed with the gas-enriched therapeutic
compositions or fluids of the present invention in any of the
routes required to administer the compounds to a subject.
[0357] The topical formulations can be presented in unit-dose or
multi-dose sealed containers, such as ampules and vials, and can be
stored in a freeze-dried (lyophilized) condition requiring only the
addition of the sterile liquid excipient, or gas-enriched fluid,
immediately prior to use. Extemporaneous fluids, and/or suspensions
can be prepared from sterile powders, granules, and tablets.
[0358] The dose administered to a subject, especially an animal,
particularly a human, in the context of the present invention
should be sufficient to affect a cosmetic and/or therapeutic
response in the animal over a reasonable time frame. One skilled in
the art will recognize that dosage will depend upon a variety of
factors including a condition of the animal, the body weight of the
animal, the desired outcome, as well as the condition being
treated. A suitable dose is that which will result in a
concentration of the cosmetic and/or therapeutic composition in a
subject which is known to affect the desired response.
[0359] The size of the dose also will be determined by the route,
timing and frequency of administration as well as the existence,
nature, and extent of any adverse side effects that might accompany
the administration of the therapeutic composition and the desired
physiological effect.
[0360] The gas-enriched fluids of the present invention may be used
to improve existing drug delivery compositions and methods. The
diffuser processed fluids may be formulated, alone or together with
one or more therapeutic agents, in suitable dosage unit
formulations. In various embodiments, these formulations may
include conventional non-toxic pharmaceutically acceptable
carriers, adjuvants, emulsifying and suspending agents, sweetening,
flavoring, perfuming agents, and vehicles appropriate for the
particular route taken into the body of the subject.
[0361] As used herein, the terms "pharmaceutically acceptable
carrier" and "carrier" mean a non-toxic, inert solid, semi-solid or
liquid filler, diluent, encapsulating material or formulation
auxiliary of any type. Some non-limiting examples of materials
which can serve as pharmaceutically acceptable carriers are sugars
such as lactose, glucose and sucrose; starches such as corn starch
and potato starch; cellulose and its derivatives such as sodium
carboxymethyl cellulose, ethyl cellulose and cellulose acetate;
powdered tragacanth; malt; gelatin; talc; excipients such as cocoa
butter and suppository waxes; oils such as peanut oil, cottonseed
oil; safflower oil; sesame oil; olive oil; corn oil and soybean
oil; glycols; such as propylene glycol; esters such as ethyl oleate
and ethyl laurate; agar; buffering agents such as magnesium
hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water;
isotonic saline; Ringer's solution; ethyl alcohol, and phosphate
buffer solutions, as well as other nontoxic compatible lubricants
such as sodium lauryl sulfate and magnesium stearate, as well as
coloring agents, releasing agents, coating agents, sweetening,
flavoring and perfuming agents, preservatives and antioxidants can
also be present in the composition, according to the judgment of
the formulator. In particular aspects, such carriers and excipients
or are themselves diffuser processed fluids.
[0362] In certain embodiments, the gas-enriched fluid disclosed
herein may comprise a wetting, irrigation or soaking fluid to apply
to organs, such as the skin, or into which organs and/or tissues
may be placed. One or more therapeutic agents may be dissolved in
the gas-enriched fluid or placed in the tissue before wetting,
irrigating or soaking it in the gas-enriched fluid. The
gas-enriched fluid may also be applied to the skin or other organ
or tissue in combination with a preexisting medication, thereby
creating a gas-enriched therapeutic fluid, to increase the efficacy
of the medication. The gas-enriched fluid may also be used to
dissolve a powder, thereby creating a gas-enriched therapeutic
fluid and drug delivery method. Alternatively, the inventive
gas-enriched fluid may comprise infused ingredients of these
patches, gels, creams, lotions, ointments, pastes, solutions,
sprays, aqueous or oily suspensions, emulsions, and the like.
[0363] Topical drug delivery, including drug delivery by
administration to the skin or mucosal membranes is also within the
scope of this invention. Particular aspects provide for therapeutic
wound care products for delivery to the skin or to a portion
thereof, or to any bodily location of a wound. For example, the
gas-enriched diffuser processed fluid may comprise ointments,
drops, therapeutic solutions, irrigation fluids, pharmaceuticals
(prescription and over the counter drops and other medications,
steroid drops, carteol, a non-selective beta blocker, vitamins),
and the like. For example, the gas-enriched fluid may replace
common carriers such as saline and water.
[0364] Enhancing Biological Tissue Growth:
[0365] In other related embodiments, the gas-enriched fluid
compositions and/or methods described herein can be used in
relation to enhancing biological cell and/or tissue growth of cell
(including stem cells), tissue and organ cultures, whether
naturally occurring, or genetically modified. In addition,
gas-enriched fluid compositions and/or methods disclosed herein can
be used for artificial blood and surgical procedures requiring
artificial blood (such as coronary bypass surgery and shock-trauma
procedures). Similarly, oxygen-rich solutions may be used to
perfuse one or more solid organs, such as liver, kidney, heart,
eye, hand, foot, brain, and others after harvesting and prior to
(e.g., in transit) and during transplantation. Use of gas-enriched
(particularly oxygen-enriched) fluids in accordance with the
disclosed embodiments may lead to longer storage time and more
successful transplant rates.
[0366] In particular embodiments, the gas-enriched fluid may be
used in certain embodiments, storing, transporting, infusing,
delivering, and or transplanting organs with the gas-enriched fluid
of the invention may provide for more successful preservation
and/or transplantation of organs and/or tissues, due to decreased
inflammation, decreased necrosis, and or increased cellular
function. In addition, the gas-enriched fluid of the present
invention may be delivered intravenously to a subject, including a
human patient. It is possible to oxygenate plasma for use in a
subject (human body or other animal) which may have application in
the treatment of cancer or other medical conditions or disorders.
It may also be useful to oxygenate plasma to preserve it when
stored for extended periods of time.
[0367] In other embodiments, hydrotherapy or topical application of
gas-enriched or oxygen-enriched solution, it is possible for the
subject to receive an increased blood oxygen level and would
thereby lead to a healthier, subject with increased energy and
vitality.
Example 1
Decayed Oxygen Content in Balanced Salt Solution
[0368] FIG. 33 illustrates the dissolved oxygen retention of a 500
ml Braun balanced salt solution that originally had a dissolved
oxygen level of 5 ppm. Following enrichment of the solution at
standard temperature and pressure with the diffuser of the present
invention, the dissolved oxygen level was approximately 41 ppm at
point 2202. The solution was kept in an amber glass bottle. After
an hour, the dissolved oxygen level was 40 ppm as indicated at
point 2204; 36 ppm after two hours; 34 ppm after three hours; and
slightly more than 30 ppm after approximately four and a half
hours. The final measurement was taken shortly before six hours, at
which point the dissolved oxygen level was approximately 28
ppm.
Example 2
Microbubble Size
[0369] Experiments were performed with a gas-enriched fluid by
using the diffuser of the present invention in order to determine a
gas microbubble size limit. The microbubble size limit was
established by passing the gas enriched fluid through 0.22 and 0.1
micron filters. In performing these tests, a volume of fluid passed
through the diffuser of the present invention and generated a
gas-enriched fluid. Sixty milliliters of this fluid was drained
into a 60 ml syringe. The dissolved oxygen rate of the fluid within
the syringe was then measured using Winkler titration. The fluid
within the syringe was injected through a 0.22 micron Millipore
Millex GP50 filter and into a 50 ml beaker. The dissolved oxygen
rate of the material in the 50 ml beaker was then measured. The
experiment was performed three times to achieve the results
illustrated in Table 3 below.
TABLE-US-00004 TABLE 3 DO AFTER 0.22 MICRON DO IN SYRINGE FILTER
42.1 ppm 39.7 ppm 43.4 ppm 42.0 ppm 43.5 ppm 39.5 ppm
[0370] As can be seen, the dissolved oxygen levels that were
measured within the syringe and the dissolved oxygen levels within
the 50 ml beaker were not significantly changed by passing the
diffused material through a 0.22 micron filter, which implies that
the microbubbles of dissolved gas within the fluid are not larger
than 0.22 microns.
[0371] A second test was performed in which a batch of saline
solution was enriched with the diffuser of the present invention
and a sample of the output solution was collected in an unfiltered
state. The dissolved oxygen level of the unfiltered sample was 44.7
ppm. A 0.1 micron filter was used to filter the oxygen-enriched
solution from the diffuser of the present invention and two
additional samples were taken. For the first sample, the dissolved
oxygen level was 43.4 ppm. For the second sample, the dissolved
oxygen level was 41.4 ppm. Finally, the filter was removed and a
final sample was taken from the unfiltered solution. In this case,
the final sample had a dissolved oxygen level of 45.4 ppm. These
results were consistent with those in which the Millipore 0.2
micron filter was used. Thus, the majority of the gas bubbles or
microbubbles within the saline solution are approximately less than
0.1 microns in size.
Example 3
Sparging Effects
[0372] FIGS. 34 and 35 illustrate the sparging effects of the
diffuser of the present invention on a fluid passing therethrough.
The sparging of oxygen-enriched water was conducted in an 8 gallon
tank at standard temperature and pressure. As indicated, initially
the oxygen-enriched water had a dissolved oxygen level of
approximately 42 ppm. After 2 minutes of running through the
diffuser, the nitrogen had sparged the oxygen-enriched water such
that the dissolved oxygen level was then slightly more than 20 ppm.
At 6 minutes, the dissolved oxygen level was approximately 6 ppm.
The dissolved oxygen level of the oxygen-enriched water reached a
minimum value slightly greater than zero (0) at approximately 14
minutes after the beginning of the process. These figures
illustrate the manner in which nitrogen may be diffused into water
to sparge the oxygen from the water. However, any gas could be used
within any fluid to sparge one gas from the other and diffuse the
other gas into the fluid. The same experiment could utilize any
host fluid material, and any fluid infusion material.
Example 4
Rayleigh Effects
[0373] Fluids processed through the diffuser device described
herein exhibit differences within the structure of the water when
compared with normal unprocessed water. Gas-enriched water made by
embodiments disclosed herein has been shown to have more Rayleigh
scattering effect compared to water that is not gas-enriched
according to the present invention.
[0374] In experiments conducted, samples of gas-enriched and
non-enriched water were prepared and sent for optical analysis. The
purpose of these tests was to determine whether there are any gross
optical differences between normal (unprocessed) deionized water
and water enriched by the diffuser device of the present
invention.
[0375] The two samples, were coded to maintain their identities in
secrecy, and only after the tests were completed were the samples
identified. The two samples were placed in a laser beam of 633
nanometers according to the diagram illustrated in FIG. 37A. Sample
B, which was gas-enriched fluid according to certain embodiments
disclosed herein, exhibited scattered light regardless of its
position relative to the laser source. The Sample B fluid had been
sealed in glass bottles for approximately one week. After two to
three hours of opening the bottle, the scattering effect
disappeared. Thus, the structure of the gas-enriched fluid is
optically different from the structure of the unprocessed fluid.
The optical effect is not directly related to dissolved oxygen
levels since the dissolved oxygen level at the start was
approximately 45 ppm and at the end of the experiment was estimated
to be approximately 32 ppm. The results are illustrated in FIG.
37B.
Example 5
Generation of Solvated Electrons
[0376] Additional evidence has also suggested that the diffusion
process generated by the diffuser device of the present invention
result in solvated electrons within the gas-enriched solution. Due
to the results of the polarographic dissolved oxygen probes, it is
believed that the diffused fluid exhibits an electron capture
effect and thus the fluid may include solvated electrons within the
gas-enriched material.
[0377] There are two fundamental techniques for measuring dissolved
oxygen levels electrically: galvanic measuring techniques and
polarographic measurements. Each process uses an electrode system
wherein the dissolved oxygen levels within the solution being
tested react with a cathode of the probe to produce a current.
Dissolved oxygen level sensors consist of two electrodes, an anode
and a cathode, which are both immersed in electrolyte within the
sensor body. An oxygen permeable membrane separates the anode and
cathode from the solution being tested. Oxygen diffuses across the
membrane and interacts with the internal components of the probe to
produce an electrical current. The cathode is a hydrogen electrode
and carries negative potential with respect to the anode. The
electrolyte solution surrounds the electrode pair and is contained
by the membrane. When no oxygen is present, the cathode is
polarized by hydrogen and resists the flow of current. When oxygen
passes through the membrane, the cathode is depolarized and
electrons are consumed. The cathode electrochemically reduces the
oxygen to hydroxyl ions according to the following equation:
O.sub.2+2H.sub.2O+4E.sup.-=4OH.sup.-
[0378] When performing dissolved oxygen level measurements of a
gas-enriched solution according to the systems of the present
invention, an overflow condition has been repeatedly experienced
wherein the dissolved oxygen meter displays a reading that is
higher than the meter is capable of reading. However, evaluation of
the gas-enriched solution by Winkler Titration indicates lower
dissolved oxygen (DO) level for the solution than indicated by the
probe. Typically, in a DO probe (such as the Orion 862 used in
these experiments) has a maximum reading of 60 ppm. However, when
the meter is left in gas-enriched water of the present invention,
it overflows.
[0379] Without wishing to be bound by any particular mechanism of
action, the mechanism of the meter responds to electrons where the
oxygen reacts. Further, since electron spin resonance indicates the
fluid is free of ions, it follows that there is solvated electrons
stabilized by the oxygen levels present in the gas-enriched
fluid.
Example 6
Improved Wound Healing
[0380] A study was performed to determine the improved healing
characteristics of wounds which were exposed to an oxygen-enriched
saline solution that was processed according to embodiments
disclosed herein. In this experiment, bandages were placed on
porcine dermal excision biopsy wounds. The bandages soaked in
oxygen-enriched saline solution or a control group of bandages
soaked in a saline solution that was not oxygen-enriched.
Microscopically, several factors were evaluated by the study
including: 1) epidermalization; 2) neovascularization; 3) epidermal
differentiation; 4) mast cell migration; and 5) mitosis.
[0381] Externally, the wounds appeared to heal at varying rates.
The wounds treated with the oxygen-enriched saline solution showed
an increase in wound healing at days 4 through 11. However, both
wounds seemed to complete healing at approximately the same time.
The study showed that between days 3 and 11, the new epidermis in
wounds treated with the oxygen-enriched saline solution migrated at
two to four times as fast as the epidermis of the wounds treated
with the normal saline solution. The study also showed that between
15 and 22 days, the wound treated by the oxygen-enriched saline
solution differentiated at a more rapid rate as evidenced by the
earlier formation of more mature epidermal layers. At all stages,
the thickening that occurs in the epidermis associated with normal
healing did not occur within the wounds treated by the
oxygen-enriched saline solution.
[0382] Without wishing to be bound by any particular theory, it is
believed that the oxygen-enriched saline solution may increase the
level of NO within the wounds. NO modulates growth factors,
collagen deposition, inflammation, mast cell migration, epidermal
thickening, and neovascularization in wound healing. Furthermore,
NO is produced by an inducible enzyme that is regulated by oxygen.
Thus, while not wishing to be bound to any particular theory, the
inventive gas-enriched fluid may stimulate NO production, in
accordance with the spectrum of wound healing effects as seen in
these experiments.
[0383] The epidermis of the healing pigs experienced earlier
differentiation in the oxygen-enriched saline group at days 15
through 22. In the case of mast cell migration, differences also
occurred in early and late migration for the oxygen-enriched
solution. A conclusive result for the level of mitosis was
unascertainable due to the difficulty in staining.
[0384] Referring now to FIG. 45a through 45h, various illustrations
compare the wound healing results of the porcine epidermal tissues
with or without oxygen-enriched saline solution. Thus, the healing
of the control wound and of the wound using the oxygen-enriched
saline solution was followed for days 1, 4 and 16. FIG. 45a
illustrates the wound healing for the control wound on day 1. As
can be seen, the wound shows epidermal/dermal thickening and a loss
of contour. FIG. 45b illustrates the wound healing on day 1 for the
wound treated using the oxygen-enriched saline solution. The wound
shows normal epidermal/dermal thickness and normal contouring is
typical on a new wound.
[0385] Referring now to FIGS. 45c and 45d, there are illustrated
the wound healing for the control wound on day 4 and the wound
healing for the wound treated with the oxygen-enriched saline
solution on day 4. For the control wound illustrated in FIG. 45c,
the wound shows a 600 micron epidermal spur. In the wound treated
with the oxygen-enriched saline solution in FIG. 45d, there is
illustrated a 1200 micron epidermal spur. Thus, in the first 4 days
of the experiment, the epidermal spur created in the wound treated
using the oxygen-enriched saline solution shows an epidermal growth
rate of twice of that of the wound that was not treated with the
oxygen-enriched saline solution.
[0386] Referring now to FIG. 45e, there is illustrated the control
wound at day 16. The wound shows less differentiated epidermis with
loss of epidermal/dermal contour than that illustrated by the wound
treated with the oxygen-enriched saline solution illustrated in
FIG. 45f. FIG. 45f shows more differentiated epidermis and more
normal epidermal/dermal contouring in the wound.
[0387] Thus, as illustrated with respect to FIGS. 45a through 45f,
the wound treated with the inventive oxygen-enriched saline
solution shows much greater healing characteristics than the
untreated wound and shows a greater differentiated epidermis with
more normal epidermal/dermal contour.
[0388] Furthermore, as illustrated in FIGS. 46-49, the wounds
treated with the inventive gas-enriched saline solution exhibited
increased levels at multiple time points of mast cells, elastin,
angiogenesis, and proteoglycan/glycosaminoglycan expression, as
detected by various specific stains. Accordingly, the gas-enriched
fluid does not have deleterious effects on cutaneous wound healing,
and indeed provides an environment of increased angiogenesis, and
enhanced deposition of glycosaminoglycans and elastin fractions of
the dermis during repair of full-thickness wounds.
Example 7
In Vitro Wound Healing
[0389] The effects of a gas-enriched fluid (enriched with oxygen)
were tested for the ability of cultured human epidermal
keratinocytes to seal a wound.
[0390] Human epidermal keratinocytes were isolated from neonatal
foreskins that were obtained from routine circumcision and
de-identified. Foreskins were washed twice in PBS and incubated in
2.4 U/mL Dispase II in order to separate the dermis from the
epidermis. The epidermis was incubated with 0.25% trypsin/1 mM
EDTA, neutralized with soy bean trypsin inhibitor, agitated, and
passed through a 70 um sieve to separate the cells. Next, the cell
suspension was centrifuged and resuspended in cell culture medium
(M154) supplemented with 0.07 mM CaCl.sub.2, and human keratinocyte
growth supplements (0.2% hydrocortisone, 0.2 ng/mL human epidermal
growth factor) and penicillin/streptomycin, amphoteracin antibiotic
cocktail. The keratinocyte cell suspensions were plated onto
uncoated 12-well culture dishes and the medium replaced after 24
hours, and every 48 hours after the initial seeding.
[0391] Upon reaching cellular confluence, linear scratches were
made with a sterile p1000 pipette tip, which resulted in a uniform
cell-free wound. The monolayers were washed several times with
Dulbecco's PBS in order to remove any cellular debris. The wound
monolayers were then incubated in the following media: i) the
complete growth media (as described above in this Example); ii) the
complete growth media diluted 1:1 with a sheared version of saline
without oxygen (control fluid that was processed using the
disclosed diffuser device but without adding a gas); and iii) the
complete growth media diluted 1:1 with oxygen-enriched saline. Each
study was done in triplicate.
[0392] Prior to incubation, the wells were filled with the
respective media and sealed by placing a 25.times.25 mm glass
coverslip on top of each well. At 6, 12, 24, and 48 hours
post-wounding, oxygen measurements were made, and cultures were
imagined.
[0393] Six hours post-wounding, the edges of the wounds in the
saline and gas-enriched media were more ruffled than those in the
media control that was processed with the diffuser device disclosed
herein, but without the addition of a gas. Twelve hours
post-wounding the edges of the wounds in all three media appeared
uneven, with keratinocytes along the borders migrating toward the
center of the wounds. Quantification of migrating keratinocytes
revealed approximately the same level of keratinocyte migration in
the saline and gas-enriched media. Results of the experiment are
shown in FIGS. 44a and 44b.
Example 8
Corneal Fibroblast Proliferation
[0394] Sterile gas-enriched water (with oxygen content of
approximately 50 ppm), or standard sterile deionized water was
utilized in preparing cell culture media. A human corneal stromal
fibroblast cell line was plated in 24 well tissue culture plates at
a density of 1.times.10.sup.5 cells/cm.sup.2 and cells were tested
for viability using a standard Live/Dead assay from Molecular
Probes, Inc. Viability tests were conducted at 1 or 3 days of
continuous culture with media changes twice per day. Viable and
non-viable cells were counted in 10 random 20.times. fields (0.2
mm.sup.2 area/field) in each well. The percentage of dead cells was
then calculated. Cells in the inventive gas-enriched media had
fewer dead cells than in standard culture media. Results are shown
in FIG. 50.
Example 9
Pseudomonas Inhibition, Plates
[0395] Applying gas-enriched saline solution of the present
invention limits Pseudomonas growth. Testing performed using a
gas-enriched saline solution has indicated a reduction in
Pseudomonas using water gas-enriched with oxygen.
[0396] Two test strains of Pseudomonas (ATCC accession no. 10145
and ATCC accession no. 27853) were prepared from fresh 24 hour
cultures to a McFarland 1 concentration (approximately
3.times.10.sup.8 microorganisms/mL). Each of the bacterial aliquots
(1 mL) was serially diluted in 10 fold dilutions in 9 mL of
broth-saline made from 1 part TSB broth and 9 parts sterile saline.
The bacterial concentrations tested were 10.sup.7, 10.sup.6,
10.sup.5, 10.sup.4, 10.sup.3, and 10.sup.2. A negative control tube
(no bacteria and no gas-enriched fluid of the present invention)
was prepared. The positive control tubes (containing no
gas-enriched fluid, normal saline and each of the 6 bacterial
concentrations) were included in each set of tubes for testing of
each bacterial strain.
[0397] Using the 10 fold dilutions of each bacterial strain, a set
of 36 tubes was inoculated as follows:
[0398] 1. 6 tubes received 1 mL of each of the dilutions; then 4 mL
of each of the test gas-enriched fluids were added to each tube
(bringing the total volume to 5 mL). The gas-enriched fluids were
labeled as 50 ppm, 40 ppm, 30 ppm, 20 ppm, 10 ppm, and normal
saline. The positive control was normal saline, against which all
of the other tubes were measured for growth.
[0399] All tubes of each 36 tube experiment set for each
Pseudomonas species were measured serially at 2 hour intervals
within a 24 hour period after initial incubation at 35.degree. C.
Between readings of each set, the tubes were returned to 35.degree.
C. for continued incubation. Readings were performed using a
calibrated spectrophotometer set at OD.sub.540.
[0400] The results of these tests indicate that the gas-enriched
fluid of the present invention positively inhibits Pseudomonas
strains at several bacterial dilutions within 4 to 12 hours after
the start of incubation at 35.degree. C., as well as later (16-24
hours) for both strains tested.
[0401] Specifically, the highest positive inhibition was found
during 16 to 24 hours for both strains, at 30 or 50 concentration.
The positive inhibition varies slightly depending on the
concentration of test solutions, and concentration of bacterial
samples tested.
TABLE-US-00005 TABLE 4 Time at positive inhibition of Pseudomonas
Time (hours) 2 4 6 8 10 12 16 18 2 22 24 ATCC 27853 0 1 3 1 2 2 5 0
5 7 6 % + -- 3 10 3 6 6 16 -- 1 23 20 ATCC 10145 0 4 3 4 3 3 2 1 5
6 8 % + -- 13 10 13 10 10 6 3 1 20 26 % + = number +/30 (6
bacterial concentrations .times. 5 solution concentrations)
Example 10
MIC Studies on Pseudomonas Inhibition, Tubes
[0402] Minimum inhibitory concentration (MIC) test solutions in
two-fold dilutions were prepared in a base of broth-saline mixture
made from 1 part TSB broth and 4 parts CFU-NS (normal saline). The
negative control tube (containing no bacteria and no gas-enriched
fluid) and the positive control tube (containing no gas-enriched
fluid and bacteria) were included in each set of tubes tested. The
fluid dilutions were: 50 ppm, 25 ppm, 12.5 ppm, 6.25 ppm, 3.12 ppm,
1.55 ppm, and 0.7 ppm. Following preparation of the tubes for each
bacterial strain, pH was measured on samples from the two
solutions. CFU-gas enriched fluid pH was about 6.8-7.2, while
CFU-NS pH was about 6.2.
[0403] Following preparation of the 2 sets of covered tubes at
35.degree. C. for 18 hours, visual inspection of all tubes revealed
that the first tube labeled 50 ppm in each set showed no growth and
all other tubes (except the negative control tube) showed moderate
growth.
[0404] The tubes labeled 50 ppm and showed no growth upon visual
inspection from the MIC studies were further tested for minimum
bactericidal concentrations (MBC). Samples were taken from each of
these two tubes using sterile calibrated disposable loops of 0.1 mL
capacity and were immediately streaked on BA plates to detect
growth or inhibition of growth on the challenge bacteria. Following
18 hours incubation at 35.degree. C., samples collected from each
of the tubes showing no visual growth (tube 50 from ATCC 10145 and
tube 50 from ATCC 27853) both showed growth at very high levels
when streaked and plated on BA plates. The numbers of colonies were
too numerous to count. Thus, a slight delay in growth was
experienced with the tube 50 gas-enriched fluids.
Example 11
Pseudomonas Inhibition, Dressings
[0405] Aquacel dressings were tested dry and hydrated with either
test fluids 52 (pH 7.2-7.8), 50 (pH 6.0-6.2), 42 (pH 7.2), 34 (pH
7.2), 25 (pH 6.8), and 10 (pH 6.2), or normal saline (pH 6.2),
against Pseudomonas strains ATCC 10145 and ATCC 27853 A first
application of the gas-enriched test fluids or normal saline fluid
to the dressings (0.4 mL) was followed by a second application 12
hours later (0.25 mL).
[0406] Results revealed a 3-4 mm clear area of inhibition around
one of the three dressing pieces (1 cm square) treated with sterile
25 fluid applied to the three Pseudomonas ATCC accession no. 10145
seeded plates, and a 3-4 mm clear area of inhibition around one of
the three dressing pieces (1 cm square) treated with sterile 25
fluid applied to one of the three Pseudomonas ATCC accession no.
27853 seeded plates. A clear area of less than 1-2 mm of inhibition
was around 2 sides of the dry dressing on both strains. No zones of
inhibition were detected around the other test or control
dressings.
Example 12
Microbial Inhibition, Dressings
[0407] Blood Agar plates were used and wound dressings used were
Promogram Prisma matrix dressing (A) and hydrofiber dressing (B).
The sterile dressings were tested with microorganisms from a 24
hour culture at a cell density with McFarland equivalence turbidity
standards of 3.0.times.10.sup.8/mL.
[0408] Test microorganisms were Staphlycoccus aureaus,
Staphylococcus epidermidis, Pseudomonas aeruginosa, E. coli, and
Candida albicans.
[0409] Dressings of each type were hydrated with approximately 0.8
mL of either gas-enriched fluid or normal saline, and left for
approximately 30 minutes. Dry dressings were used as controls.
Plates were then grown for 24 hours, and results recorded.
[0410] As viewed in the cell plates, the hydrofiber dressing (B)
had no effect on growth of test microorganisms whether dry or
hydrated with either fluid. The Prisma dressing (A) hydrated with
gas-enriched fluid showed a zone of partial inhibition with
colonies for Staph. aureaus, a 1 mm zone of inhibition for Staph.
epidermidis, a 2-3 mm zone of inhibition with an added halo effect
extending out an additional 2-3 mm for Pseudomonas, a 2 mm zone of
inhibition for Candida, and no effect for E. coli.
[0411] The dry control hydrofiber dressing (A) had a zone of 2-3 mm
of inhibition for Staph. auereaus, Staph. epidermidis, Pseudomonas,
and a partial inhibition of about 1-2 mm for E. coli and Candida,
with break-through colonies observed.
[0412] The normal saline control hydrofiber dressing (A) showed a 2
mm zone of partial inhibition for Staph. auereaus, Staph.
epidermidis, and Pseudomonas. Colonies were observed grown into E.
coli and Candida test strains.
Example 13
MIC/MBC Testing with Tobramycin
[0413] The gas-enriched fluids of the present invention were tested
randomly against three Pseudomonas strains, PA 01, PA 14, and PA K.
Each bacterial culture was prepared from fresh 24 hour cultures to
McFarland 1 concentration (approximately 3.times.10.sup.8
microorganisms/mL). Each of the bacterial samples (1 mL) were then
serially diluted in 10 fold dilutions to 9 mL of a broth-saline
mixture made from 1 part MH broth and 9 parts sterile saline. The
bacterial concentrations tested were 10.sup.5, 10.sup.4, 10.sup.3,
and 10.sup.2. Negative control tubes (containing no bacteria and no
gas-enriched saline solution, and no Tobramycin), as well as
positive control tubes (containing test bacteria concentrations and
normal saline but no Tobramycin, and no gas-enriched fluid), were
run side-by-side with the test solutions. Cells were incubated at
35.degree. C. Results are shown in Table 5 below. Sheared normal
saline was passed through the gas-enriching diffuser device of the
present invention without the addition of a gas to the fluid.
TABLE-US-00006 TABLE 5 Better kill rate with Tobramycin and
Gas-enriched fluid than Tobramycin Alone Dissolved Gas Fluid
Description (Oxygen, ppm) PA01 PA14 PAK A Gas-enriched Saline 33.4
Yes Yes No B Gas-enriched saline 23.8 No Yes No C Gas-enriched
saline 49.6 Yes Yes Yes D Normal saline 9.6 No No No E Gas-enriched
saline 42.8 Yes Yes Yes F Sheared normal saline (no 14.1 Yes Yes
Yes gas enrichment)
Example 14
DNA Thermostability
[0414] The thermostability of T7 DNA oligonucleotide in one
embodiment of the inventive oxygen-enriched water, was compared
with non-enriched deionized water. As the temperature of the water
increases, the DNA undergoes conformational changes and "melts." As
illustrated in FIG. 38, the DNA oligonucleotide begins to denature
at about 50.degree. Celsius in the control (deionized water),
whereas the DNA oligonucleotide in the oxygen-enriched inventive
fluid remains intact until about 60.degree. Celsius. Indeed, based
on known thermodynamic principles, and G-C/A-T content of the
oligo, the "melting" temperature of the T7 primer is estimated to
be about 47.7.degree. C. Thus, the inventive oxygen-enriched fluid
imparts a higher thermostability to the DNA, and different
conformational change and denaturation temperature when compared to
control fluid.
Example 15
Pyrogallol Reactivity Test
[0415] An aliquot of the inventive oxygen-enriched water was tested
for peroxidase activity by using a commercially available
horseradish peroxidase and pyrogallol assay (Sigma). Briefly,
pyrogallol stock solution was prepared with deionized water.
Pyrogallol measures peroxidase activity of the horseradish
peroxidase enzyme on the fluid as it reacts with a substrate (such
as hydrogen peroxide or other electron acceptor), to yield
purpurogallin and water. Test fluid with horseradish peroxidase,
pyrogallol and the appropriate potassium phosphate buffer were
compared with other fluids. Hydrogen peroxide served as the
positive control. The other fluids tested were water that was
oxygen-enriched and pressurized in a pressure pot, up to 100 psi to
reach the desired dissolved oxygen level (Pressure Pot), while the
other fluid was oxygen-enriched with an air stone in an open beaker
(Fine Bubble). All fluids tested were maintained at room
temperature, and measured approximately 55 ppm dissolved oxygen
level (by FOXY probe).
[0416] The reaction was carried out at 20.degree. Celsius, pH 6.0,
A.sub.420 nm light path at 1 cm with continuous spectrophotometric
rate determination.
[0417] Reagents were as follows:
[0418] 100 mM potassium phosphate buffer, pH 6.0, 20.degree.
Celsius
[0419] 5% (w/v) pyrogallol solution
[0420] 0.4-0.7 unit/mL peroxidase enzyme in cold buffer (made
fresh).
[0421] The test sample contained 2.10 mL of inventive fluid, 0.32
mL buffer, and 0.32 mL pyrogallol, and 0.10 mL enzyme solution. The
control sample contained 2.10 mL of inventive fluid, 0.32 mL
buffer, 0.32 mL pyrogallol, and an additional 0.10 buffer added for
equivalent volume with test sample. The reaction was immediately
mixed by inversion and recorded increase in A.sub.420 nm for
approximately 6 minutes. When the test solution is 0.50% (w/v)
hydrogen peroxide, then one unit will form 1.0 mg of purpurogallon
from pyrogallol in 20 seconds at pH 6.0 at 20.degree. Celsius. The
purpurogallon unit is equivalent to approximately 18 .mu.M units
per minute at 25.degree. Celsius. Thus, in a 3.00 mL reaction mix,
the final concentration are 14 .mu.M potassium phosphate, 0.27%
(w/w) hydrogen peroxide, 0.5% (w/v) pyrogallol and 0.04-0.07 unit
peroxidase.
[0422] As indicated in FIGS. 39A-39E, the inventive oxygen-enriched
fluid tested positive for reactivity with horseradish peroxidase by
pyrogallol, indicating the presence of an electron acceptor, while
the pressure pot and fine bubbled water samples had far less
reactivity.
[0423] Several chemical tests for the presence of hydrogen peroxide
were conducted, as described herein, and none of these tests were
positive. Thus, the inventive oxygen-enriched fluid of the instant
application provides for peroxidase activity in the absence of
hydrogen peroxide.
Example 16
Glutathione Peroxidase Study
[0424] The inventive oxygen-enriched fluid was tested for the
presence of hydrogen peroxide by testing the reactivity with
glutathione peroxidase using a standard assay (Sigma). Briefly,
glutathione peroxidase enzyme cocktail was constituted in deionized
water and the appropriate buffers. Water samples were tested by
adding the enzymatic reagents. Continuous spectrophotometric rate
determination was made at A.sub.340 nm, and room temperature
(25.degree. Celsius). Samples tested were: 1. deionized water
(negative control), 2. inventive oxygen-enriched fluid at low
concentration, 3. inventive oxygen-enriched fluid at high
concentration, 4. hydrogen peroxide (positive control). As
illustrated in FIG. 40, the hydrogen peroxide positive control
showed a strong reactivity, while none of the other fluids tested
reacted with the glutathione peroxidase.
Example 17
Cytokine Profile
[0425] Mixed lymphocytes were obtained from a single healthy
volunteer donor. Buffy coat samples were washed according to
standard procedures to remove platelets. Lymphocytes were plated at
a concentration of 2.times.10.sup.6 per plate in RPMI media (+50 mm
HEPES) diluted with either inventive gas-enriched fluid or
distilled water (control). Cells were stimulated with 1
microgram/mL T3 antigen, or 1 microgram/mL phytohemagglutinin (PHA)
lectin (pan-T cell activator), or unstimulated (negative control).
Following 24 hour incubation, cells were checked for viability and
the supernatants were extracted and frozen.
[0426] The supernatants were thawed, centrifuged, and tested for
cytokine expression using a XMAP.RTM. (Luminex) bead lite protocol
and platform. Results are shown in FIG. 42. Notably, IFN-gamma
level was higher in the inventive gas-enriched culture media with
T3 antigen than in the control culture media with T3 antigen, while
IL-8 was lower in the inventive gas-enriched culture media with T3
antigen than in the control culture media with T3 antigen.
Additionally, IL-6, IL-8, and TNF-alpha levels were lower in the
inventive gas-enriched media with PHA, than in the control media
with PHA, while IL-1b levels were lower in the inventive
gas-enriched fluid with PHA when compared with control media with
PHA. In gas-inventive media alone, IFN-gamma levels were higher
than in control media.
[0427] Two million cells were plated into 6 wells of a 24-well
plate in full RPMI+50 mm Hepes with either inventive
oxygen-enriched fluid (water) (wells 1, 3, and 5) or distilled
water (2, 4 and 6) (10.times.RPMI diluted into water to make 1 x).
Cells were stimulated with 1 ug/ml T3 antigen (wells 1 and 2) or
PHA (wells 3 and 4). Control wells 5 and 6 were not stimulated.
After 24 hours, cells were checked for viability and supernatants
were collected and frozen. Next, the supernatants were thawed and
spun at 8,000 g to pellet. The clarified supernatants were assayed
for the cytokines listed using a LUMINEX BEAD LITE protocol and
platform. The numerical data is tabulated in Table 1, and the
corresponding bar graphs are depicted in FIG. 42.
TABLE-US-00007 TABLE 1 Sample IFN IL-10 IL-12p40 IL-12p70 II-2 IL-4
IL-5 IL-6 IL-8 IL-ib IL-10 TNFa 1 0 0 0 2.85 0 0 7.98 20.3 1350
7.56 11500 15.5 2 0 0 0 3.08 0 0 8 15.2 8940 3.68 4280 7.94 3 0 581
168 3.15 0 0 8 16400 2200 3280 862 13700 4 0 377 56.3 4.22 0 0 8.08
23800 22100 33600 558 16200 5 0 0 0 2.51 0 0 7.99 24 1330 7.33 5900
8.55 6 0 0 0 2.77 0 0 8 5.98 3210 4.68 3330 0
Example 18
Myelin Oligodendrocyte Glycoprotein (MOG)
[0428] As set forth in FIG. 52, lymphocyte proliferation in
response to MOG antigenic peptide was increased when cultured in
the presence of the inventive gas-enriched fluid when compared to
pressurized, oxygenated fluid (pressure pot) or deionized control
fluid. Thus, the inventive gas-enriched fluid amplifies the
lymphocyte proliferative response to an antigen to which the cells
were previously primed.
[0429] Myelin oligodendrocyte glycoprotein peptide 35-55 (MOG
35-55) (M-E-V-G-W-Y-R-S-P-F-S-R-O-V-H-L-Y-R-N-G-K) (SEQ ID NO:2)
corresponding to the known mouse sequence was synthesized. Next,
5.times.10.sup.5 spleen cells were removed from MOG T cell receptor
transgenic mice previously immunized with MOG, and were cultured in
0.2 ml TCM fluid reconstituted with inventive gas-enriched fluid,
pressurized oxygenated water (pressure pot water) or with control
deionized water. Splenocytes were cultured with MOG p35-55 for 48
or 72 hours, respectively. Cultures were pulsed with 1Ci
[3H]-thymidine and harvested 16 hours later. Mean cpm of [3H]
thymidine incorporation was calculated for triplicate cultures.
Results are shown in FIG. 52.
Sequence CWU 1
1
2120DNAArtificial SequenceOligonucleotide 1taatacgact cactataggg
20221PRTArtificial SequenceMOG 35-55 2Met Glu Val Gly Trp Tyr Arg
Ser Pro Phe Ser Arg Xaa Val His Leu1 5 10 15 Tyr Arg Asn Gly Lys
20
* * * * *
References