U.S. patent application number 09/878656 was filed with the patent office on 2002-06-06 for method and apparatus for reducing fouling of injection and recovery wells.
Invention is credited to Perriello, Felix Anthony.
Application Number | 20020066566 09/878656 |
Document ID | / |
Family ID | 23127621 |
Filed Date | 2002-06-06 |
United States Patent
Application |
20020066566 |
Kind Code |
A1 |
Perriello, Felix Anthony |
June 6, 2002 |
Method and apparatus for reducing fouling of injection and recovery
wells
Abstract
A method and apparatus are disclosed in which alkane-utilizing
bacteria are used to reduce fouling of injection and recovery
wells. Fouling materials such as combinations of bacteria and metal
oxides that would otherwise clog the wells are prevented from
depositing on the wells. In a preferred embodiment, a butane
substrate and an oxygen-containing gas are injected near a well
inlet or outlet to stimulate the growth of butane-utilizing
bacteria which are effective at reducing or eliminating fouling of
the well.
Inventors: |
Perriello, Felix Anthony;
(Norwood, MA) |
Correspondence
Address: |
Pietragallo, Bosick & Gordon
One Oxford Centre, 38th Floor
301 Grant Street
Pittsburgh
PA
15219
US
|
Family ID: |
23127621 |
Appl. No.: |
09/878656 |
Filed: |
June 11, 2001 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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09878656 |
Jun 11, 2001 |
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09293088 |
Apr 16, 1999 |
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09293088 |
Apr 16, 1999 |
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09275320 |
Mar 24, 1999 |
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09275320 |
Mar 24, 1999 |
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08767750 |
Dec 17, 1996 |
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Current U.S.
Class: |
166/304 ;
166/246; 166/312; 166/403 |
Current CPC
Class: |
C09K 8/52 20130101 |
Class at
Publication: |
166/304 ;
166/246; 166/312; 166/403; 210/747 |
International
Class: |
E21B 037/06 |
Claims
What is claimed is:
1. A method of reducing well fouling comprising: stimulating growth
of alkane-utilizing bacteria adjacent to the well; and reducing
deposition of fouling material on the well with the
alkane-utilizing bacteria.
2. The method of claim 1, wherein the alkane-utilizing bacteria
comprise butane-utilizing bacteria.
3. The method of claim 2, wherein the butane-utilizing bacteria
comprises at least one bacterium selected from the group consisting
of Pseudomonas, Variovorax, Nocardia, Chryseobacterium, Comamonas,
Acidovorax, Rhodococcus, Aureobacterium, Micrococcus, Aeromonas,
Stenotrophomonas, Sphingobacterium, Shewanella, Phyllobacterium,
Clavibacter, Alcaligenes, Gordona, Corynebacterium and
Cytophaga.
4. The method of claim 2, wherein the butane-utilizing bacteria
comprises at least one bacterium selected from the group consisting
of putida, rubrisubalbicans, aeruginosa, paradoxus, asteroides,
brasiliensis, restricta, globerula, indologenes, meningosepticum,
acidovorans, delafieldii, rhodochrous, erythropolis, fascians,
barkeri, esteroaromaticum, saperdae, varians, kristinae, caviae,
maltophilia, thalpophilum, spiritivorum, putrefaciens B,
myrsinacearum, michiganense, xylosoxydans, terrae, aquaticum B and
johnsonae.
5. The method of claim 2, wherein the butane-utilizing bacteria
comprises at least one bacterium selected from the group consisting
of Pseudomonas rubrisubalbicans, Pseudomonas aeruginosa, Variovorax
paradoxus, Nocardia asteroides, Nocardia restricta,
Chryseobacterium indologenes, Comamonas acidovorans, Acidovorax
delafieldii, Rhodococcus rhodochrous, Rhodococcus erythropolis,
Aureobacterium esteroaromaticum, Aureobacterium saperdae,
Micrococcus varians, Micrococcus kristinae, Aeromonas caviae,
Stenotrophomonas maltophilia, Sphingobacterium thalpophilum,
Clavibacter michiganense, Alcaligenes xylosoxydans, Corynebacterium
aquaticum B and Cytophaga johnsonae.
6. The method of claim 1, wherein the fouling material comprises at
least one metal oxide.
7. The method of claim 1, wherein the fouling material comprises
bacteria.
8. The method of claim 1, wherein the fouling material comprises a
combination of at least one metal oxide and bacteria.
9. The method of claim 8, wherein the at least one metal oxide is
hydrated.
10. The method of claim 8, wherein the at least one metal oxide
comprises an oxide of iron, manganese, lead, arsenic, nickel,
mercury, molybdenum, cadmium, copper, chromium, silver, zinc,
potassium or combinations thereof.
11. The method of claim 8, wherein the at least one metal oxide
comprises ferric oxide.
12. The method of claim 1, wherein the well comprises a recovery
well.
13. The method of claim 12, wherein the recovery well comprises a
water recovery well.
14. The method of claim 12, wherein the recovery well comprises an
oil recovery well.
15. The method of claim 1, wherein the well comprises an injection
well.
16. The method of claim 15, wherein the injection well comprises an
in-situ bioremediation well.
17. A method of reducing well fouling comprising: introducing at
least one alkane and oxygen to a region of the well susceptible to
fouling; and stimulating growth of alkane-utilizing bacteria which
reduce deposition of fouling material on the well.
18. The method of claim 17, wherein the at least one alkane
comprises a butane substrate.
19. The method of claim 18, wherein the butane substrate comprises
at least about 10 weight percent butane.
20. The method of claim 18, wherein the butane substrate comprises
at least about 50 weight percent butane.
21. The method of claim 18, wherein the butane substrate comprises
at least about 90 weight percent butane.
22. The method of claim 18, wherein the butane substrate comprises
at least about 99 weight percent n-butane.
23. The method of claim 17, further comprising introducing the at
least one alkane to the well continuously.
24. The method of claim 17, further comprising introducing the at
least one alkane to the well periodically.
25. The method of claim 17, wherein the oxygen is introduced in the
form of an oxygen-containing gas.
26. The method of claim 25, wherein the oxygen-containing gas
comprises air.
27. The method of claim 25, further comprising introducing the
oxygen-containing gas to the well continuously.
28. The method of claim 25, further comprising introducing the
oxygen-containing gas to the well periodically.
29. The method of claim 17, further comprising introducing
butane-utilizing bacteria to the site.
30. The method of claim 17, wherein the well comprises a recovery
well.
31. The method of claim 30, wherein the recovery well comprises a
water recovery well.
32. The method of claim 30, wherein the recovery well comprises an
oil recovery well.
33. The method of claim 32, further comprising injecting a
pressurized fluid adjacent to the oil recovery well.
34. The method of claim 33, wherein the pressurized fluid comprises
water.
35. The method of claim 34, wherein at least a portion of the
oxygen is introduced to the well by mixing an oxygen-containing gas
with the water.
36. The method of claim 33, wherein the pressurized fluid comprises
sea water.
37. The method of claim 17, wherein the well comprises an injection
well.
38. The method of claim 37, wherein the injection well comprises an
in-situ bioremediation well.
39. Apparatus for reducing well fouling comprising means for
stimulating growth of alkane-utilizing bacteria adjacent to the
well to thereby reduce deposition of fouling material on the
well.
40. Apparatus for reducing well fouling comprising: a source of an
alkane substrate; a source of an oxygen-containing gas; and at
least one injector in flow communication with the source of alkane
substrate and the source of oxygen-containing gas having a distal
end located in proximity to at least a portion of the well that is
susceptible to fouling.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application is a continuation-in-part of U.S. patent
application Ser. No. 09/275,320 filed Mar. 24, 1999, which is a
continuation-in-part of U.S. patent application Ser. No. 08/767,750
filed Dec. 17, 1996, now U.S. Pat. No. 5,888,396, which are
incorporated herein by reference.
FIELD OF THE INVENTION
[0002] The present invention relates to anti-fouling of injection
and recovery wells, and more particularly relates to a method and
apparatus for reducing or eliminating fouling of injection and
recovery wells with alkane-utilizing bacteria.
BACKGROUND INFORMATION
[0003] Many different types of injection wells and recovery wells
are widely used. Typical injection wells are used for water supply,
groundwater control, solution mining, waste disposal, geothermal
energy and to increase oil production. Typical recovery wells are
used for groundwater control, capture and treatment, municipal
water supplies, domestic water supplies and in the oil and
geothermal industries.
[0004] Fouling at injection and recovery wells is a major problem
worldwide. Chemical and biological incrustation are major causes of
decreased well performance and eventual failure. Material
comprising bacteria and metal oxides accumulates and clogs the
wells. Iron and manganese fouling at well screens is a global
problem for water supply and production wells. In addition, metals
fouling is a problem at most wastewater treatment plants. For
example, iron and manganese bacteria occur widely in wells open to
the atmosphere when sufficient iron and/or manganese are present in
the groundwater. Such bacteria plug wells by enzymatically
catalyzing the oxidation of metals, using the energy to promote the
growth of threadlike slimes, and accumulating large amounts of
metal hydroxides, such as ferric hydroxide, in the slime. For
instance, the bacteria may obtain their energy by oxidizing ferrous
ions to ferric ions, which are then precipitated as hydrated ferric
hydroxide on or in their mucilaginous sheaths. Iron bacteria
produce accumulations of slimy material that have a gelatinous
consistency. In addition, they precipitate dissolved iron and
manganese. The dual effect of the growing bacteria and
precipitating minerals occurs. Precipitation of the iron and rapid
growth of the bacteria create a voluminous material that quickly
plugs the screen pores of the sediment surrounding the well bore.
The explosive growth rates of iron bacteria can render a well
useless within a matter of months.
[0005] Other forms of iron bacteria induce the precipitation of
iron through nonenzymatic means. These bacteria promote
precipitation by mechanisms such as: raising pH; changing the redox
potential of the water by algal photosynthesis; and liberating
chelated iron. Some forms of iron bacteria can reduce iron to a
ferrous state under anaerobic conditions.
[0006] A conventional method for reducing the amount of iron
incrusting materials reaching production well screens, called the
Vyredox System, uses a series of injection wells located in a
circle around a production well. Oxygenated water is injected into
the wells to oxidize iron in solution and promote the growth of
iron bacteria so that little iron reaches the production well.
[0007] In many wells, incrusting iron cannot be removed before
reaching a production well. In these cases, caustic chemicals are
added to the well to clear biofouling and free the production well
screen. These practices are costly, time consuming and require the
production well to be brought off-line, thus disrupting service.
Furthermore, the chemicals and their toxic fumes may cause serious
injury to the technicians injecting them into the production
well.
[0008] Other methods conventionally used to control iron bacteria
are heat, explosives, ultrasonics, radiation and anoxic blocks.
[0009] Despite the above-noted efforts, need exists for the
effective reduction or elimination of fouling at various types of
injection and recovery wells. The present invention has been
developed in view of the foregoing, and to remedy other
deficiencies of the prior art.
SUMMARY OF THE INVENTION
[0010] In accordance with the present invention, a method and
apparatus are provided for anti-fouling of well inlets and outlets.
Alkanes such as a butane substrate are used in combination with
oxygen to stimulate the growth of microorganisms near the well
inlets and/or outlets. The present alkane/oxygen injection method
is a simple and cost effective treatment process to abate and
prevent metal fouling at wellheads and other industrial
applications by oxidizing dissolved metal concentrations and
immobilizing them from the production and supply routes.
[0011] An aspect of the present invention is to provide a method of
reducing well fouling. The method includes the steps of stimulating
growth of alkane-utilizing bacteria adjacent to the well, and
reducing the deposition of fouling material on the well with the
alkane-utilizing bacteria.
[0012] Another aspect of the present invention is to provide a
method of reducing well fouling, including the steps of introducing
at least one alkane and oxygen to a region of the well susceptible
to fouling, and stimulating growth of alkane-utilizing bacteria
which reduce the deposition of fouling material on the well.
[0013] Another aspect of the present invention is to provide an
apparatus for reducing well fouling. The apparatus includes a
source of a butane substrate, a source of an oxygen-containing gas,
and at least one injector in flow communication with the source of
butane substrate and the source of oxygen-containing gas having a
distal end located in proximity to at least a portion of the well
that is susceptible to fouling.
[0014] These and other aspects of the present invention will become
more apparent from the following description.
BRIEF DESCRIPTION OF THE DRAWINGS
[0015] FIG. 1 is a schematic illustration of an in-situ butane
injection system which may be used for reducing or eliminating
fouling of injection or recovery wells in accordance with an
embodiment of the present invention.
[0016] FIG. 2 is a schematic illustration of an in-situ air
injection system which may be used for reducing or eliminating
fouling of injection or recovery wells in accordance with an
embodiment of the present invention.
[0017] FIG. 3 is a partially schematic illustration of a butane
injection well which may be used for reducing or eliminating
fouling of injection or recovery wells in accordance with an
embodiment of the present invention.
[0018] FIG. 4 is a partially schematic illustration of an air
injection well which may be used for reducing or eliminating
fouling of injection or recovery wells in accordance with an
embodiment of the present invention.
[0019] FIG. 5 is a plan view of a hazardous waste site contaminated
with 1,1,1-TCA which was treated with an in-situ bioremediation
system, and which exhibited reduced recovery well fouling in
accordance with an embodiment of the present invention.
[0020] FIG. 6 is a plan view of the site shown in FIG. 5,
illustrating a zone of butane influence in accordance with an
embodiment of the present invention.
[0021] FIG. 7 is a plan view of the site shown in FIG. 5,
illustrating a zone of dissolved oxygen influence in accordance
with an embodiment of the present invention.
[0022] FIG. 8 is a schematic illustration of a recovery well and
adjacent alkane/oxygen injection wells for reducing fouling of the
recovery well in accordance with an embodiment of the present
invention.
[0023] FIG. 9 is a schematic illustration of a recovery well and an
adjacent alkane/oxygen injection well in accordance with an
embodiment of the present invention.
[0024] FIG. 10 is a schematic illustration of a recovery well, a
pressurized fluid injection well, and an alkane/oxygen injection
well for reducing fouling of the recovery well and pressurized
fluid well in accordance with an embodiment of the present
invention.
[0025] FIG. 11 is a schematic plan view of an oil recovery well,
water injection wells, and butane/air injection wells for reducing
fouling of the oil recovery well and water injection wells in
accordance with an embodiment of the invention.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0026] The present invention relates to a method and apparatus for
reducing or eliminating fouling of injection and recovery wells. An
alkane substrate and an oxygen-containing gas are injected into a
well site to stimulate the growth of microorganisms which act to
reduce or eliminate fouling at well inlets and/or outlets.
[0027] In accordance with the present invention, alkanes are used
to stimulate the growth of alkane-utilizing bacteria which are
effective in reducing or eliminating fouling of injection and
recovery wells. Suitable alkane substrates include methane, ethane,
propane, butane and mixtures thereof. For example, natural gas may
be used as the alkane source. Alkanes and mixtures thereof having
relatively high water solubility are preferred for many
applications. Preferably, the solubility of the alkanes in water at
17.degree. C. is greater than about 5 ml per 100 ml of water, more
preferably greater than about 10 ml per 100 ml of water. Butane is
a particularly preferred alkane for use in accordance with the
present invention. The butane may be provided in the form of a
butane substrate.
[0028] As used herein, the term "butane substrate" includes liquids
and gases in which butane is present in sufficient amounts to
stimulate substantial growth of butane-utilizing bacteria. Butane
is preferably the most prevalent compound of the butane substrate
on a weight percent basis, and typically comprises at least about
10 weight percent of the butane substrate. The other constituents
of the butane substrate may include any suitable compounds,
including inert gases and/or other alkanes such as methane, ethane
and propane. The butane substrate preferably comprises at least
about 50 weight percent butane. More preferably, the butane
substrate comprises at least about 90 weight percent butane. In a
particular embodiment, the butane substrate comprises at least
about 99 weight percent n-butane. The butane may contain straight
(n-butane) and/or branched chain compounds. While the use of a
butane substrate is primarily described herein, it is to be
understood that the use of other alkanes, alone or in combination,
is within the scope of the present invention.
[0029] As used herein, the term "oxygen-containing gas" means gases
which comprise oxygen, including pure oxygen as well as mixtures of
oxygen with other gases. For example, the oxygen-containing gas may
comprise air, pure oxygen, or oxygen blended with inert gases such
as helium, argon, nitrogen, carbon monoxide or the like.
[0030] As used herein, the term "well" means any injection or
recovery structure that may experience fouling during operation.
Typical injection wells are used for water supply, groundwater
control, solution mining, waste disposal, geothermal energy and to
increase oil production. Typical recovery wells are used for
groundwater control, capture and treatment, municipal water
supplies, domestic water supplies and in the oil and geothermal
industries.
[0031] As used herein, the term "fouling" means the deposition of
fouling material on at least a portion of a well. Fouling can
result in well clogging and failure due to the growth of, e.g.,
iron and manganese bacteria, including oxide and hydroxide
deposition and incrustation caused by the activities of the iron
and manganese bacteria.
[0032] The term "fouling material" means a material comprising
bacteria and metal oxides (including hydroxides). The metal
component of the fouling material may include, e.g., iron,
manganese, lead, arsenic, nickel, mercury, molybdenum, cadmium,
copper, chromium, silver, zinc and potassium. As a particular
example of the formation of a fouling material, iron bacteria, such
as Crenothrix, Leptothrix, Gallionella, Clonothiix, and
Pseudomonas, can change dissolved iron to insoluble ferric iron,
which is subsequently deposited in the sheaths of the bacterial
cells. The sheaths create a gel-like slime that eventually clog
well screen slots.
[0033] The phenomenon of well screen fouling may be caused by iron
and manganese bacteria, and the incrustations they deposit on the
well screens. The fouling is caused when the solubility of iron and
manganese changes due to, e.g., microbial activities, pressure
reduction, pH fluctuations and other chemical and physical
alterations. The following equations depict the chemical and
solubility changes that lead up to well screen incrustation.
Fe(HCO.sub.3).sub.2=Fe(OH).sub.2+2CO.sub.2
[0034] Solubility of ferrous hydroxide (Fe(OH).sub.2) is less than
20 mg/l (ppm).
4Fe(OH).sub.2=2H.sub.2O+O.sub.2=4Fe(OH).sub.3
[0035] Solubility of ferric hydroxide (4Fe(OH).sub.3) is less than
0.01 mg/l.
[0036] Further oxidation of the hydroxides of iron and manganese
causes the formation of hydrated oxides. Ferrous iron in solution
can react with oxygen to form ferric oxide.
[0037] Soluble manganese becomes insoluble in the same manner as
iron.
2Mn(HCO.sub.3).sub.2+O.sub.2+2H.sub.2O=2Mn(OH).sub.4+4CO.sub.2
[0038] Processes that may be used in accordance with the present
invention include the use of indigenous alkane-utilizing
microorganisms and/or the injection of non-indigenous
alkane-utilizing microorganisms into the well treatment area.
Preferably, indigenous microorganisms are be stimulated to flourish
by the addition of the butane substrate, oxygen-containing gas and,
optionally, bacterial nutrients that may be limited in the system
under scrutiny. Suitable bacterial nutrients include
nitrogen-containing compounds and phosphorous-containing compounds.
For example, the bacterial nutrients may comprise ammonia, nitrate,
ammonium chloride and sodium orthophosphate salts, and combinations
thereof.
[0039] FIG. 1 schematically illustrates an in-situ butane injection
system in which fouling of injection and recovery wells may be
substantially reduced in accordance with an embodiment of the
present invention. The butane injection system is contained within
a shed 10 which houses a butane cylinder 12. The butane cylinder 12
rests on a scale 13 which is used to measure the amount of butane
contained within the cylinder 12. The cylinder 12 is connected to a
dual port valve 14. A helium cylinder 16 is also contained within
the shed 10. The helium cylinder 16 is connected through a
regulator 18 and a gate valve 20 to the dual port valve 14. A check
valve 22 is positioned between a single line from the butane
cylinder 12 and two branched lines leading to solenoid valves 24
and 25. A digital timer 26 controls the solenoid valve 24, while
another digital timer 27 controls the solenoid valve 25. Gate
valves 28 and 29 are positioned downstream from the solenoid valves
24 and 25, respectively. The gate valve 28 communicates with a
butane injection well line 30. The gate valve 29 communicates with
another butane injection well line 31. A power reset assembly 32 is
connected between the digital timers 26 and 27 and an electrical
power source 34 such as a GFCI receptacle (120 VAC).
[0040] The flow of the butane substrate from the cylinder 12
through the butane injection well lines 30 and 31 is controlled by
the in-situ butane injection system shown in FIG. 1. The flow of
the butane substrate to the injection well lines 30 and 31 may be
constant or may be pulsed. In one embodiment, the butane substrate
is supplied periodically to the lines 30 and 31 at desired
intervals. For example, butane pulses may be supplied from 0.01
second to several minutes per hour at any suitable flow rate.
[0041] FIG. 2 schematically illustrates an in-situ air injection
system in which fouling of injection and recovery wells may be
substantially reduced in accordance with an embodiment of the
present invention. The air injection system is housed within the
shed 10. An air compressor 40 supplies air through a system gauge
41 to gate valves 42 and 43. A flow gauge 44 is located downstream
from the gate valve 42, while another flow gauge 45 is located
downstream from the gate valve 43. The air compressor 40 is
electrically connected through a fuse panel disconnect system 46 to
an electrical power supply 47 such as a 220 volt AC power supply. A
dilution valve 48 is connected to the line between the system gauge
and the gate valves 42 and 43. The dilution valve 48 is connected
to a vent 49. An air injection well line 50 communicates with the
gate valve 42, while another air injection well line 51
communicates with the gate valve 43. The gate valves 42 and 43 are
used to equalize the air flow to each of the air injection well
lines 50 and 51.
[0042] The flow of air from the compressor 40 through the air
injection well lines 50 and 51 is controlled by the in-situ air
injection system shown in FIG. 2. The flow of air or other types of
oxygen-containing gases to the injection well lines 50 and 51 may
be constant or may be pulsed. The oxygen-containing gas may be
supplied periodically to the lines 50 and 51 at desired intervals.
For example, air may be supplied from 0.1 second to 50 minutes per
hour at any suitable flow rate.
[0043] FIG. 3 illustrates a butane injection well 60 that may be
used to prevent fouling in accordance with an embodiment of the
present invention. The butane well injection line 30 shown in FIG.
1 is connected to the butane injection well 60 by a horizontal pipe
61 which is positioned a distance D below finish grade 62. The
distance D is preferably at least 3 feet. The horizontal pipe 61 is
connected by an elbow 63 to a vertical pipe 64. The vertical pipe
64 may have any suitable diameter and length. For example, the
vertical pipe 64 may comprise a 1 inch outside diameter iron pipe
having a length of from about 1 to about 100 or 500 feet or more. A
fitting 66 is connected to the end of the vertical pipe 64. For
example, the fitting 66 may be a 1 inch by 11/4 inch increaser
fitting. A well point 68 is connected to the distal end of the
vertical pipe 64 by the fitting 66. The well point 68 may be of any
suitable construction which adequately permits dispersion of the
butane into the treatment site. For example, the well point 68 may
comprise a slotted stainless steel tube having an outside diameter
of 11/4 inch and a length of 2 feet. Butane supplied from the
butane injection well line 30 to the butane injection well 60 is
introduced via the well point 68 into the treatment site at the
desired location.
[0044] FIG. 4 illustrates an air injection well 70 that may be used
to prevent fouling in accordance with an embodiment of the present
invention. The air injection well line 50 shown in FIG. 2 is
connected to a horizontal pipe 71 of the air injection well 70. The
horizontal pipe 71 is located a distance D below the finish grade
62, which is preferably at least 3 feet. A tee joint 73 connects
the horizontal pipe 71 to a vertical pipe 74. For example, the tee
joint 73 may have outside dimensions of 2 inch by 2 inch by 2 inch.
The vertical pipe 74 may have any suitable diameter and length. For
example, the vertical pipe 74 may comprise a 2 inch outside
diameter PVC pipe having a length of from about 1 to about 100 or
500 feet or more, depending upon the desired depth of the air
injection well 70. A fitting 75 connects the distal end of the
vertical pipe 74 to a well screen 76. The fitting 75 may, for
example, comprise a 2 inch by 2 inch coupler fitting. The well
screen 76 may be of any suitable construction which adequately
allows dispersion of air or other oxygen-containing gases into the
treatment zone. For example, the well screen 76 may comprise a
slotted PVC tube having an inside diameter of 2 inches and a length
of 2 feet. Air or another oxygen-containing gas supplied from the
air injection well line 50 to the air injection well 70 is
dispersed via the well screen 46 at the desired location within the
contaminated site. A road box 77 including a cap 78 is connected to
the tee joint 73 in order to protect the top of the well 70 and to
allow access to the well 70 for sampling purposes. In addition, the
road box 77 allows access to the well 70 for manual or automatic
addition of non-indigenous bacteria and/or bacterial nutrients such
as nitrogen-containing compounds and phosphorous-containing
compounds, if desired.
[0045] Although the butane injection well 60 and the air injection
well 70 shown in FIGS. 3 and 4 are separate from each other, both
butane and air could be supplied through the same injection well.
Although two butane injection well lines 30 and 31 and two air
injection well lines 50 and 51 are shown in FIGS. 1 and 2, single
lines or any suitable number of multiple injection well lines may
be used. In addition, the lines may be connected to the butane and
air injection wells by any suitable manifold system.
[0046] The following example illustrates various aspects of the
present invention, and is not intended to limit the scope of the
invention.
[0047] A subsurface investigation was performed at a hazardous
waste site in Massachusetts. A plan view of the site is shown in
FIG. 5. The investigation indicated the presence of high
concentrations of chlorinated volatile organic compounds (VOCs) in
the soil and, to a greater extent, in the groundwater. Further
assessment indicated the presence of chlorinated VOCs in the
shallow aquifer. The soils and groundwater at the site had elevated
concentrations of 1,1,1-TCA at dissolved-phase concentrations up to
900 mg/L. Given the extent of VOC contamination at the site, mass
excavation and disposal was not deemed an acceptable remediation
option. A conventional groundwater pump and treat (GP&T) system
using standard air-stripping and granular activated carbon
technology was initially installed at the site. This system
included three recovery wells screened in the shallow aquifer in
the vicinity of the highest VOC concentrations encountered during
the subsurface investigations. Periodic groundwater quality
monitoring revealed that the groundwater pump and treat system was
having a limited impact on groundwater conditions in the shallow
aquifer, and 1,1,1-TCA concentrations remained at levels above
approximately 150,000 ppb at a particular monitoring well located
near the source of the contamination. In an effort to accelerate
remediation of the VOC plume identified at the site, an in-situ
bioremediation system was installed.
[0048] An eighteen month in-situ field demonstration was conducted
at the hazardous waste site shown in FIG. 5. Butane was injected
into the subsurface using a gas delivery system of the present
invention, as illustrated in FIGS. 1-4. Monitoring for fugitive
butane emissions was performed daily by automated instrumentation.
Butane gas was pulsed into the subsurface at a rate that resulted
in full microbial oxidation within the groundwater, capillary
fringe and vadose zones. The total dissolved-phase concentrations
of 1,1,1-TCA in a single monitoring well located within the
biotreatment zone was 150,000 ppb shortly after the biotreatment
began, 2,500 ppb eight months after the treatment began, and 580
ppb fourteen months after the biotreatment began. Pilot testing of
this technology has shown that chloride ion and carbon dioxide
concentrations and cell densities (in groundwater) increased by
several orders of magnitude over background in the butane
biostimulation zone.
[0049] The vapor pressure of butane is normally about 16 psi. As
the ambient temperature drops to 40.degree. F., the pressure inside
a butane cylinder drops to a vacuum (negative). Therefore, an
injection system was designed that would operate in extreme
conditions of heat and cold while effectively and safely injecting
butane into targeted areas in the subsurface. This was accomplished
by pressurizing the butane cylinder with a helium feed through a
dual port valve. The helium maintains a constant pressure (e.g., 50
psi) inside the butane cylinder. An internal dip tube located
within the butane cylinder insured that the helium only pushed
liquid butane out of the tank. Alternatively, the butane cylinder
could be heated to a sufficient temperature to feed gaseous butane
from the cylinder.
[0050] The electrical requirement for the system was a 120-V outlet
to operate two digital timers (NEMA-4 and explosion proof) and two
electric solenoid valves (NEMA-4 and explosion proof) that
regulated the introduction of the cometabolic butane substrate into
the VOC-impacted areas. Liquid butane was injected each hour from a
120-pound cylinder for 0.5 seconds using a helium pusher set at 50
psi for a total of approximately 24 cubic feet per day. Oxygen in
the form of air was supplied by a five-horsepower rotary-vane air
compressor (220-V). The system components, including the air
compressor, butane and helium cylinders, injection system timers
and valves, and ancillary equipment were located in a storage shed,
as illustrated in FIGS. 1 and 2. The storage shed was equipped with
a Lower Explosive Limit (LEL) monitor.
[0051] A hollow-stem-auger drill rig was used to install the two
butane injection wells as shown in FIG. 3 about 20 feet below the
surface of the ground. The butane injection wells comprised 11/4
inch outside diameter black iron pipe fitted with a 2-foot slotted
stainless steel well point, advanced from the ground surface to a
depth above the silt layer. Clean sand was placed around and up to
two feet above the top of the well point, with a grout seal placed
form the top of the sand pack to the ground surface using a tremie
pipe, thus sealing the borehole and preventing short-circuits to
the surface via the well annular space.
[0052] Each butane injection well was programmed to deliver two
pounds of liquid butane into the subsurface daily. Butane was
pulsed for a period of 0.5 seconds every hour utilizing helium as a
pusher gas (approximately 24 ft.sup.3 total of butane gas per
day).
[0053] Butane injection at each well was controlled by a digital
timer/intervalometer (NEMA-4 and explosion proof). Both timers
operated on a GFCI circuit. The digital timers operated normally
closed electric solenoid valves (one per well) designed for
operation with flammable gases and liquids (NEMA-4 and explosion
proof). The timers were programmed to open the solenoid valves for
0.5 seconds every hour to regulate the introduction of the
cometabolic butane substrate into the VOC-impacted areas. Both
solenoid valves operated on a GPCI circuit.
[0054] The butane injection system was equipped with an automatic
flow sensor programmed to interrupt the power supply to the
solenoid timers if a sudden release of butane gas was detected
anywhere within the system in excess of two seconds. The
de-energized solenoid valves returned to a normally closed
position.
[0055] The two air injection wells, as illustrated in FIG. 4, were
installed using a hollow-stem auger drill rig to a depth of about
20 feet. The air injection wells were constructed of two-inch
inside-diameter schedule 40 PVC slotted well screens two feet long.
Filter sand was placed in the annular space around the well screen
to approximately two feet above the screen/riser interface. The
two-foot well screen was installed at a depth above the silt layer
in close proximity to a butane injection well with sand pack placed
two feet above the top of the screened interval and grout seal
placed form the top of the sand pack to the ground surface. The
ground was placed in the annular space using a tremie pipe. A
watertight road box was cemented flush to the ground surface over
each well.
[0056] Each air injection well was equipped with a pressure gauge
to monitor the injection pressure at each well point. The air
compressor was set to deliver 5 to 10 cfm per well at less than one
breakout pressure in order to minimize VOC volatilization.
[0057] Periodic monitoring at the site consisted of in-situ
measurement of dissolved oxygen, carbon dioxide and chloride ion
concentration using colorimetric methods. Dissolved butane
concentrations were quantified using a portable gas chromatograph.
Serial dilutions and plating were conducted to enumerate viable
cell counts in groundwater. Monitoring for fugitive butane
emissions was conducted daily during the pilot study, and no
evidence of breakthrough to the ambient air or other potential
receptors was noted during the field study. Groundwater quality
samples collected from the on-site monitoring wells were analyzed
at a certified laboratory for VOCs referencing EPA method 8260.
[0058] Prior to the above-noted in-situ field study, dissolved iron
at the site caused fouling of the recovery wells and monitoring
well screens. Such iron fouling is a common problem for most water
supply and production wells. The dissolved iron concentration in
the groundwater at the site was initially approximately 55 to 60
ppm. However, after operating for eighteen months, the butane
injection wells showed no indication of fouling at the well
screens. In addition, a shallow recovery well at the site which
previously required frequent treatment to clean iron precipitation,
no longer became clogged after implementation of the butane
sparging program. Since implementation of the study, the fouling,
e.g., dissolved metals precipitation, was no longer a problem at
the site in the butane biotreatment zone. The dissolved iron
concentration around the butane injection wells dropped to 2 to 5
ppm.
[0059] The data in Table 1 summarizes the analytical data acquired
at the site during the field study. Monitoring wells IND-1, IND-2
and IND-3 were inside the biotreatment zone. Monitoring well
RIZ-21S served as a control outside the butane biotreatment zone.
Analytical testing of the iron concentration was conducted using
Chemetrics titration cells, colorimetric analysis.
1TABLE 1 Iron Concentration Iron Concentration After Monitoring
Well Prior to Butane 8 Months of Butane Locations Injection
Injection IND-1 55 ppm 2.0 ppm IND-2 58 ppm 3.5 ppm IND-3 57 ppm
5.0 ppm RIZ-21S 55 ppm 50 ppm
[0060] While not intending to be bound by any particular theory,
butane oxidation may not only cometabolize chlorinated solvents,
but may also be used to oxidize metals which would otherwise
promote fouling. Butane injection in accordance with the present
invention is a simple and cost effective treatment system to abate
or prevent metal fouling at wellheads and other industrial
applications by, e.g., oxidizing dissolved metal concentrations and
immobilizing them from the production and supply routes.
[0061] In an embodiment of the present invention, in-situ injection
wells for the bioremediation of trichloroethene (TCE) and
trichbroethane (TCA) pollutants as disclosed in U.S. patent
application Ser. No. 09/275,320 may be prevented from fouling in
accordance with the method and apparatus of the present invention.
It has been discovered that butane-utilizing bacteria which are
extremely effective at degrading pollutants such as low molecular
weight halogenated aliphatic hydrocarbons including TCE and TCA,
may also be used to reduce or eliminate fouling of injection and
recovery wells. In accordance with an embodiment of the present
invention, the same in-situ injection wells may be used for both
bioremediation of pollutants and reducing fouling of the in-situ
injection wells themselves, as well as other injection and/or
recovery wells in the treatment zone. Alternatively, different
injection wells may be provided for bioremediation and
anti-fouling.
[0062] The present system may also be used to prevent fouling of
in-situ injection wells for remediating methyl tertiary butyl ether
(MTBE) pollutants, as described in U.S. patent application Ser. No.
09/275,840, which is incorporated herein by reference.
[0063] The present system may further be used to prevent fouling of
in-situ injection wells for remediating polychlorinated biphenyl
(PCB) pollutants, as described in U.S. patent application Ser. No.
09/275,324, which is incorporated herein by reference.
[0064] In addition, the present system may be used to prevent
fouling of in-situ injection wells for remediating petroleum
pollutants, as described in U.S. patent application Ser. No.
09/275,381, which is incorporated herein by reference.
[0065] FIG. 8 schematically illustrates an anti-fouling system in
accordance with an embodiment of the present invention. A recovery
well 80 extends from a surface 82 to a distal inlet end 84. A
recovered fluid 86 travels from the distal end 84 of the recovery
well 80 through the surface 82. The surface 82 may be, for example,
a ground surface or a water surface such as an ocean, lake, etc.
The recovered fluid 86 may be in the form of a liquid, gas,
fluidized solid or the like. For example, the recovered liquid 86
may be groundwater, wastewater, oil, a mixture of oil and water, or
the like.
[0066] As shown in FIG. 8, in order to reduce fouling at the inlet
end 84 of the recovery well 80, alkane/oxygen injection wells 90
and 91 are provided. The injection well 90 includes a distal outlet
end 94, while the injection well 91 includes a distal outlet end
95. An injection fluid 96 travels through the injection well 90,
while an injection fluid 97 travels through the injection well 91.
The injection fluids 96 and 97 may each comprise a mixture of
alkanes and oxygen-containing gas in accordance with an embodiment
of the present invention. Alternatively, the injection fluid 96 may
be the alkane substrate while the injection fluid 97 may be the
oxygen-containing gas, or vice versa. The flow of the alkane
substrate and/or oxygen-containing gas through the injection well
90 may be continuous or discontinuous, e.g., pulsed. The alkane
substrate and oxygen-containing gas supplied from the outlet ends
94 and 95 of the injection wells 90 and 91 stimulate growth of
alkane-utilizing bacteria in a zone around the inlet end 84 of the
recovery well 80, to thereby reduce fouling of the well. For
example, where the recovered fluid 86 is groundwater, injection of
a butane substrate and an oxygen-containing gas via the injection
wells 90 and 91 may be used to reduce the deposition of fouling
material on the inlet end 84 of the recovery well 80.
[0067] FIG. 9 schematically illustrates the treatment of a recovery
well in accordance with another embodiment of the present
invention. A recovery well 100 extends from a surface 102 to a
distal inlet end 104. The surface 102 may be, for example, a ground
surface or a water surface such as an ocean, lake, etc. A recovered
fluid 106 such as oil, groundwater, a mixture of oil and water,
etc. travels from the inlet end 104 through the recovery well 100.
An alkane/oxygen injection well 110 is located adjacent to the
recovery well 100. The injection well 110 includes an outlet end
114 through which an alkane/oxygen fluid 116 passes. The fluid 116
preferably comprises a butane substrate and oxygen-containing gas.
The butane substrate and oxygen-containing gas may be supplied as a
mixture through the injection well 110, or may be supplied
separately. The flow of the butane substrate and/or
oxygen-containing gas through the injection well 110 may be
continuous or discontinuous, e.g., pulsed. As a particular example,
the recovery well 100 may be an oil recovery well with a well head
located at the distal end 104 thereof. In this case, the
alkane/oxygen fluid 116, e.g., butane substrate and
oxygen-containing gas, are injected near the distal end 104 of the
recovery well 100 in order to reduce fouling of the oil well
head
[0068] FIG. 10 schematically illustrates another alkane/oxygen
treatment system of the present invention. A recovery well 120
extends from a surface 122 to a distal inlet end 124. The surface
122 may be, for example, a ground surface or a water surface such
as an ocean, lake, etc. A recovered fluid 126 travels from the
inlet 124 through the recovery well 120. A pressurized fluid
injection well 130 extends from the surface 122 to a distal outlet
end 134 which is located in proximity to the inlet end 124 of the
recovery well 120. A pressurized fluid 136 flows through the
injection well 130. The pressurized fluid 136 may comprise a liquid
and/or gas, such as water, sea water, air, natural gas or the like.
Although the pressurized fluid injection well 130 shown in FIG. 10
extends from the surface 122, a submerged pump (not shown) could
alternatively be used to supply the pressurized fluid 136, e.g.,
sea water, to the zone surrounding the inlet end 124 of the
recovery well 120.
[0069] As shown in FIG. 10, in order to reduce fouling at the inlet
end 124 of the recovery well 120, an alkane/oxygen injection well
140 is provided. Although a single alkane/oxygen injection well 140
is shown in FIG. 10, multiple injection wells may be used. The
injection well 140 includes a distal outlet end 144 located in
proximity to the inlet end 124 of the recovery well 120. An
alkane/oxygen fluid 146 travels through the injection well 140. The
alkane/oxygen fluid 146 preferably comprises a butane substrate and
an oxygen-containing gas. The butane substrate and
oxygen-containing gas may be supplied as a mixture through the
injection well 140, or may be supplied separately. The flow of the
butane substrate and/or oxygen-containing gas through the injection
well 140 may be continuous or discontinuous, e.g., pulsed. In
accordance with the present invention, the alkane/oxygen fluid 146
stimulates the growth of alkane-utilizing bacteria in a zone
surrounding the inlet end 24 of the recovery well 20 and the outlet
end 134 of the pressurized fluid injection well 130. Such
alkane-utilizing bacteria, e.g., butane-utilizing bacteria,
effectively reduce fouling of the recovery well 120 and pressurized
fluid injection well 130.
[0070] In accordance with an embodiment of the present invention, a
system as schematically illustrated in FIG. 10 may be used to
reduce fouling of oil recovery wells. In many oil drilling
operations, sea water is pumped to a zone near the inlet end of the
oil recovery well. During such oil recovery operations, the inlet
end of the oil recovery well becomes fouled. In addition, the
outlet end of the sea water injection system may also be fouled. By
providing at least one alkane/oxygen injection well near the oil
well head, fouling of the inlet end of the oil recovery well and
outlet end of the pressurized sea water injection well may be
reduced substantially. As a result, oil production rates may be
increased significantly.
[0071] One major application for anti-fouling technology is the
petroleum industry. Most of the U.S. oil wells are currently in
secondary recovery, that is, the oil no longer flows up a recovery
well under natural pressure and energy. It is the energy in natural
gas and/or salt water occurring under high pressures with the oil
that furnishes the requisite energy to displace the oil in the
subsurface into the recovery or production wells. Currently, oil
production includes the recovery of oil and salt water through
forced pumping activities or artificial lift methods. The salt
water is separated from the recovered oil and is re-injected into
the oil-bearing formation through a series of injection wells. The
re-injected salt water also assists in pushing or herding oil
remaining in the subsurface toward recovery wells where it is then
pumped back to the surface through the forced pumping
activities.
[0072] When the water is re-injected, increased fouling is observed
at the injection wells screens probably due to the growth of iron
and manganese oxidizing bacteria and associated oxide and hydroxide
incrustations.
[0073] FIG. 11 is a schematic plan view depicting a method of
reducing the fouling observed at salt water injection wells and oil
recovery wells. If a series of butane and air injection wells are
installed in strategic positions around the injection and recovery
wells, the activities of butane-oxidizing bacteria enhance iron and
manganese oxidation within the oil-bearing reservoir and in the
salt water. Since butane has the highest solubility of any of the
gaseous hydrocarbons, butane enrichment enhances microbial activity
by increasing substrate availability. By oxidizing the metals,
lower concentrations of dissolved metals will be available for
transport through the reservoir to the recovery wells where it is
then pumped to the surface. By progressively lowering the iron and
manganese concentrations in the recovered salt water, fouling at
the injection and recovery well screens will diminish over
time.
[0074] The butane-utilizing bacteria used in accordance with a
preferred embodiment of the present invention preferably produce
oxygenase enzymes and are capable of metabolizing butane. The
operative enzymes may include extracellular enzymes, intracellular
enzymes and cell-bound enzymes. The butane-utilizing bacteria
typically produce butane monoxygenase and/or butane dioxygenase
enzymes.
[0075] The butane-utilizing bacteria may contain gram negative and
gram positive aerobic rods and cocci, facultative anaerobic gram
negative rods, non-photosynthetic, non-fruiting gliding bacteria
and irregular non-sporing gram positive rods.
[0076] Of the Pseudomonadaceae family comprising gram-negative
aerobic rods and cocci, species of the following genera may be
suitable: Pseudomonas; Variovorax; Chryseobacterium; Comamonas;
Acidovorax; Stenotrophomonas; Sphingobacterium; Xanthomonas;
Frateuria; Zoogloea; Alcaligenes; Flavobacterium; Derxia;
Lampropedia; Brucella; Xanthobacter; Thermus; Thermomicrobium;
Halomonas; Alteromonas; Serpens; Janthinobacterium; Bordetella;
Paracoccus; Beijerinckia; and Francisella.
[0077] Of the Nocardioform Actinomycetes family comprising
gram-positive Eubacteria and Actinomycetes, the following genera
may be suitable: Nocardia; Rhodococcus; Gordona; Nocardioides;
Saccharopolyspora; Micropolyspora; Promicromonospora;
Intrasporangium; Pseudonocardia; and Oerskovia.
[0078] Of the Micrococcaceae family comprising gram-positive cocci,
the following genera may be suitable: Micrococcus; Stomatococcus;
Planococcus; Staphylococcus; Aerococcus; Peptococcus;
Peptostreptococcus; Coprococcus; Gemella; Pediococcus; Leuconostoc;
Ruminococcus; Sarcina; and Streptococcus.
[0079] Of the Vibrionaceae family comprising facultative anaerobic
gram-negative rods, the following genera may be suitable:
Aeromonas; Photobacterium; Vibrio; Plesiomonas; Zymomonas;
Chromobacterium; Cardiobacterium; Calymmatobacterium;
Streptobacillus; Eikenella; and Gardnerella.
[0080] Of the Rhizobiaceae family comprising gram-negative aerobic
rods and cocci, the following genera may be suitable:
Phyllobacterium; Rhizobium; Bradyrhizobium; and Agrobacterium.
[0081] Of the Cytophagaceae family comprising non-photosynthetic,
gliding bacteria, non-fruiting, the following genera may be
suitable: Cytophaga; Flexibacter; Saprospira; Flexithrix;
Herpetosiphon; Capnocytophaga; and Sporocytophaga.
[0082] Of the Corynebacterium family comprising irregular,
non-sporing gram-positive rods, the following genera may be
suitable: Aureobacterium; Agromyces; Arachnia; Rothia;
Acetobacterium; Actinomyces; Arthrobactera: Arcanobacterium;
Lachnospira; Propionibacterium; Eubacterium; Butyrivibria;
Brevibacterium; Bifidobacterium; Microbacterium; Caseobacter; and
Thermoanaerobacter.
[0083] The following isolation techniques were used for obtaining
pure and mixed cultures of various methane-, propane- and
butane-utilizing bacteria. Enrichment procedures were used to
increase bacterial population for a given growth substrate. Soil
samples collected from a variety of sites underwent enrichment
transfers weekly for a period of one year. The methods and
materials used for the enrichment studies are described below.
[0084] Soil samples were collected with a stainless-steel hand
auger at depths that varied between one to two feet. The soils
samples were stored in dedicated glass containers and moistened
with sterile deionized/distilled water for transport to the
laboratory. The hand auger was decontaminated between sampling
locations with three Alconox soap/distilled water rinses. Soil
samples used as inocula were collected from the locations
summarized in Table 2.
2TABLE 2 Sample Number/Matrix Sample Location 1/soil Landfill cell
2/soil #2 fuel oil impacted soil 3/soil Landfill cell 4/soil
Gasoline and waste oil impacted soils 5/soil Shallow freshwater
lagoon 6/soil Salt marsh 7/soil Industrial outfall 8/soil #2 fuel
oil impacted soil
[0085] Cultures were transferred weekly for a period of one year in
liquid media to increase the relative numbers of methane-, propane-
and butane-utilizing bacteria. The liquid media was a mineral salts
media (MSM) prepared from the following chemicals:
3 MgSO.sub.4--7H.sub.2O 1.0 g; CaCl.sub.2 0.2 g; NH.sub.4Cl 0.5 g;
FeCl.sub.3--6H.sub.2O 4.0 mg; Trace elements solution 0.5 ml; and
Distilled water 900 ml.
[0086] A trace elements solution, which provides micronutrients for
bacterial growth, was prepared comprising the following
ingredients:
4 ZnCl.sub.2 5.0 mg; MnCl.sub.2--4H.sub.2O 3.0 mg; H.sub.3BO.sub.4
30.0 mg; NiCl.sub.2--6H.sub.2O 2.0 mg;
(NH.sub.4).sub.6Mo.sub.7O.sub.24--4H.sub.2O 2.25 mg; and Distilled
water 1000 ml.
[0087] The pH of the MSM was adjusted to 6.8 before autoclaving (20
min at 121 degree C) with 20.0 ml of a phosphate buffer solution
comprising 3.6 g of Na.sub.2HPO.sub.4 and 1.4 g of KH.sub.2PO.sub.4
in 100 ml of distilled water. After autoclaving the MSM and the
buffer solution, another 2.0 ml of the buffer solution was added to
the MSM when the temperature of the media reached 60 degree C. The
MSM cocktail was completed with the addition of 4.0 mg of casamino
acids and 4.0 mg of yeast (Difco) dissolved in 100 ml of distilled
water. The nutrient solution was filter sterilized by vacuum
filtration through a 0.2 micron filter (Gelman) prior to addition
to the MSM.
[0088] Prior to the first enrichment transfer, the inocula from the
eight sampling locations summarized in Table 2 underwent a series
of pre-treatments. The first two pre-treatments were conducted on
the original soil materials used as inocula. The last two
treatments were applied as MSM alterations during the weekly
transfers. The pre-treatments consisted of the following: (1) 30%
ethanol saturation rinse followed by a sterile phosphate buffer
rinse (ethanol); (2) 60.degree. C. water bath for 15 minutes
(heat); (3) no treatment (no-treat); (4) MSM containing 10% aqueous
solution of sodium chloride (10% NaCl); and (5) MSM with pH of 2.0
(pH of 2). Treatment Nos. (4) and (5) were employed in an attempt
to locate extreme halophiles and acidophiles capable of utilizing
hydrocarbons as a growth substrate.
[0089] The first enrichment transfers for each sample series were
conducted in 72 ml serum bottles (Wheaton) with 20 ml of MSM and
1.0 g of inocula. Subsequent culture transfers (5.0 ml) were
conducted with sterilized plastic syringes (B&D). The bottles
were capped with red rubber plugs and crimped with aluminum seals
(Wheaton). Each sample was handled aseptically and all glassware,
materials and supplies were sterilized by autoclaving. Table 3
summarizes the enrichment transfer schedule and the concentration
of methane or propane replaced in the headspace of each serum
bottle using a dedicated gas tight syringe (Hamilton) with a Fisher
Scientific inert sampling valve (on/off lever) to control gas loss
from the needle tip between each transfer.
5TABLE 3 Sample No. Pre-Treatment Food Source Sample ID 1 ethanol
methane 2EM 1 heat methane 1HM 1 no-treat methane 1NM 1 10% NaCl
methane 1SM 1 pH of 2.0 methane 1AM 1 ethanol propane 1EP 1 heat
propane 1HP 1 no-treat propane 1NP 1 10% NaCl propane 1SP 1 pH of
2.0 propane 1AP
[0090] The amount of oxygen required for mineralization of methane,
propane and butane can be derived from the following equations.
CH.sub.4+2O.sub.2=CO.sub.2+2H.sub.2O 2:1
C.sub.3H.sub.8+5O.sub.2=3CO.sub.2+4H.sub.2O 5:1
C.sub.4H.sub.10+6.5O.sub.2=4CO2+5H.sub.2O 6.5:1
[0091] Table 3 summarizes the entire set of enrichment transfers
prepared for Sample No. 1. The first sample series did not include
a butane treatment. The remaining seven samples were prepared in
identical fashion and, in addition, contained a butane treatment
series, as shown in Tables 4 through 10. A control (serum bottle
with sterilized MSM only) was maintained for each sample
series.
[0092] All hydrocarbon gases described herein were research grade
quality (Scott Specialty Gases). Methane was added at a
concentration of 27% (vol/vol), propane at 10% and butane at 6%.
After the first six months of enrichment transfers, the
concentrations were reduced to 13% for methane and 9% for propane.
The concentration of butane remained the same at 6%.
6TABLE 4 Sample No. Pre-Treatment Food Source Sample ID 2 ethanol
methane 2EM 2 heat methane 2HM 2 no-treat methane 2NM 2 10% NaCl
methane 2SM 2 pH of 2.0 methane 2AM 2 ethanol propane 2EP 2 heat
propane 2HP 2 no-treat propane 2NP 2 10% NaCl propane 2SP 2 pH of
2.0 propane 2AP 2 ethanol butane 2EB 2 heat butane 2HB 2 no-treat
butane 2NB 2 10% NaCl butane 2SB 2 pH of 2.0 butane 2AB
[0093]
7TABLE 5 Sample No. Pre-Treatment Food Source Sample ID 3 ethanol
methane 3EM 3 heat methane 3HM 3 no-treat methane 3NM 3 10% NaCl
methane 3SM 3 pH of 2.0 methane 3AM 3 ethanol propane 3EP 3 heat
propane 3HP 3 no-treat propane 3NP 3 10% NaCl propane 3SP 3 pH of
2.0 propane 3AP 3 ethanol butane 3EB 3 heat butane 3HB 3 no-treat
butane 3NB 3 10% NaCl butane 3SB 3 pH of 2.0 butane 3AB
[0094]
8TABLE 6 Sample No. Pre-Treatment Food Source Sample ID 4 ethanol
methane 4EM 4 heat methane 4HM 4 no-treat methane 4NM 4 10% NaCl
methane 4SM 4 pH of 2.0 methane 4AM 4 ethanol propane 4EP 4 heat
propane 4HP 4 no-treat propane 4NP 4 10% NaCl propane 4SP 4 pH of
2.0 propane 4AP 4 ethanol butane 4EB 4 heat butane 4HB 4 no-treat
butane 4NB 4 10% NaCl butane 4SB 4 pH of 2.0 butane 4AB
[0095]
9TABLE 7 Sample No. Pre-Treatment Food Source Sample ID 5 ethanol
methane 5EM 5 heat methane 5HM 5 no-treat methane 5NM 5 10% NaCl
methane 5SM 5 pH of 2.0 methane 5AM 5 ethanol propane 5EP 5 heat
propane 5HP 5 no-treat propane 5NP 5 10% NaCl propane 5SP 5 pH of
2.0 propane 5AP 5 ethanol butane 5EB 5 heat butane 5HB 5 no-treat
butane 5NB 5 10% NaCl butane 5SB 5 pH of 2.0 butane 5AB
[0096]
10TABLE 8 Sample No. Pre-Treatment Food Source Sample ID 6 ethanol
methane 6EM 6 heat methane 6HM 6 no-treat methane 6NM 6 10% NaCl
methane 6SM 6 pH of 2.0 methane 6AM 6 ethanol propane 6EP 6 heat
propane 6HP 6 no-treat propane 6NP 6 10% NaCl propane 6SP 6 pH of
2.0 propane 6AP 6 ethanol butane 6EB 6 heat butane 6HB 6 no-treat
butane 6NB 6 10% NaCl butane 6SB 6 pH of 2.0 butane 6AB
[0097]
11 TABLE 9 Sample No. Pre-Treatment Food Source Sample ID 7 ethanol
methane 7EM 7 heat methane 7HM 7 no-treat methane 7NM 7 10% NaCl
methane 7SM 7 pH of 2.0 methane 7AM 7 ethanol propane 7EP 7 heat
propane 7HP 7 no-treat propane 7NP 7 10% NaCl propane 7SP 7 pH of
2.0 propane 7AP 7 ethanol butane 7EB 7 heat butane 7HB 7 no-treat
butane 7NB 7 10% NaCl butane 7SB 7 pH of 2.0 butane 7AB
[0098]
12 TABLE 10 Sample No. Pre-Treatment Food Source Sample ID 8
ethanol methane 8EM 8 heat methane 8HM 8 no-treat methane 8NM 8 10%
NaCl methane 8SM 8 pH of 2.0 methane 8AM 8 ethanol propane 8EP 8
heat propane 8HP 8 no-treat propane 8NP 8 10% NaCl propane 8SP 8 pH
of 2.0 propane 8AP 8 ethanol butane 8EB 8 heat butane 8HB 8
no-treat butane 8NB 8 10% NaCl butane 8SB 8 pH of 2.0 butane
8AB
[0099] After the first two weeks of enrichment transfers, all
liquid suspensions, with the exception of the 10% NaCl treatments,
the 2.0 pH treatments and the controls, demonstrated a significant
increase in turbidity.
[0100] After conducting the enrichment transfers for 25 weeks,
morphological descriptions and direct cell counts were compiled for
all isolates. Morphological descriptions of the isolates were
compiled using an Olympus BH-2 Phase Contrast Microscope. In
addition, a Bright Line Hemacytometer (Fisher Scientific) was used
to enumerate cell densities by the direct count method. Table 11
summarizes the descriptions and cell density enumerations. Serum
bottles of sterilized MSM were maintained as controls.
13TABLE 11 Enumeration Sample ID Morphology (cells/ml) 1EM cocci
2.5E8 1HM cocci/bacilli 1.8E8 1NM bacilli 1.3E8 1SM cocci 5.8E6 1AM
cocci 3.8E6 1EP bacilli 4.0E6 1HP cocci 1.3E7 1NP bacilli 9.8E6 1SP
diplococci 4.0E6 1AP bacilli (variable) 1.5E6 2EM cocci/bacilli
1.2E8 2HM cocci/bacilli 7.3E7 2NM streptococci/bacilli 1.1E8 2SM
comma-shaped 6.6E7 2AM comma-shaped 8.3E6 2EP bacilli 1.2E8 2HP
bacilli/comma-shaped 1.8E8 2NP bacilli (variable) 1.1E8 2SP cocci
7.0E6 2AP cocci 3.3E6 2EB cocci/bacilli 2.1E8 2HB bacilli
(variable) 2.5E8 2NB cocci/comma-shaped 1.9E8 2SB bacilli 2.5E6 2AB
cocci 3.0E6 3EM cocci/bacilli 1.4E8 3HM cocci 1.2E8 3NM cocci 5.8E7
3SM cocci 7.5E5 3AM cocci 7.5E5 3EP bacilli 7.8E7 3HP bacilli 3.0E7
3NP bacilli 7.1E7 3SP cocci 1.0E6 3AP bacilli 2.5E5 3EB bacilli
(variable) 1.5E8 3HB cocci/bacilli 3.1E7 3NB cocci 3.1E8 3SB cocci
(irregular) 1.7E7 3AB cocci/bacilli 2.5E5 4EM cocci (variable)
1.6E8 4HM diplococci 3.1E8 4NM cocci 1.6E8 4SM cocci 1.3E6 4AM
bacilli 2.5E5 4EP bacilli (variable) 1.0E8 4HP bacilli (variable)
2.2E8 4NP cocci 1.3E8 4SP cocci 1.5E6 4AP cocci/bacilli 6.5E6 4EB
bacilli 3.6E8 4HB bacilli (variable) 4.8E8 4NB bacilli 2.6E8 4SB
comma-shaped 1.3E6 4AB cocci 1.0E6 5EM cocci (variable) 1.3E8 5HM
cocci 1.4E8 5NM cocci 2.4E8 5SM no cells 0.0 5AM no cells 0.0 5EP
cocci (variable) 5.1E7 5HP bacilli 3.2E7 5NP streptococci 2.1E8 5SP
cocci (variable) 2.8E6 SAP bacilli 5.0E5 5EB bacilli 3.1E8 5HB
cocci 3.2E7 5NB cocci 1.6E8 5SB bacilli 1.0E6 5AB cocci 2.5E6 6EM
bacilli (variable) 1.7E8 6HM cocci 2.6E8 6NM cocci/spirochetes
1.3E8 6SM cocci (variable) 1.3E6 6AM cocci (variable) 2.0E6 6EP
bacilli 2.8E7 6HP bacilli 1.3E8 6NP bacilli/spirochetes 2.0E8 6SP
cocci (variable) 3.5E6 6AP cocci (variable) 5.0E5 6EB cocci 3.5E7
6HB bacilli 1.3E8 6NB bacilli 4.8E7 6SB cocci 2.3E6 6AB cocci 3.3E6
7EM streptococci 1.3E8 7HM staphylococci 3.2E7 7NM cocci/bacilli
3.1E8 7SM cocci (variable) 3.0E6 7AM cocci (variable) 4.0E6 7EP
bacilli 1.4E8 7HP bacilli 4.1E8 7NP bacilli 3.5E8 7SP cocci
(variable) 1.2E7 7AP cocci (variable) 1.5E6 7EB bacilli (variable)
1.6E8 7HB bacilli (variable) 3.9E8 7NB bacilli 4.2E8 7SB cocci
(variable) 4.3E6 7AB cocci (variable) 2.8E6 8EM cocci 5.6E7 8HM
cocci 6.1E7 8NM cocci 5.7E7 8SM cocci (variable) 5.3E6 8AM bacilli
2.3E6 8EP bacilli 1.4E8 8HP cocci 3.8E8 8NP cocci 2.9E8 8SP
square-shaped 6.5E6 8AP cocci (variable) 3.8E6 8EB bacilli 1.3E8
8HB bacilli/streptococci 9.8E7 8NB bacilli (variable) 1.2E8 8SB
bacilli (variable) 2.0E6 8AB cocci (variable) 2.8E6 Control-1 no
cells 0.0 Control-2 no cells 0.0 Control-3 no cells 0.0
[0101] Sample ID strains 3NB and 6NB were placed on deposit with
the American Type Culture Collection (ATCC), Rockville, Md. on Aug.
22, 1996, under ATCC designation numbers 55808 and 55809,
respectively.
[0102] As a food source for microbial consumption, butane has been
found to be a preferred substrate to methane or propane due to its
solubility factor. Methane and propane are characterized as
slightly soluble in water, while butane is characterized as very
soluble in water. At 17 degrees centigrade, 3.5 ml of methane and
6.5 ml of propane dissolves in 100 ml of water. In contrast, 15 ml
of butane dissolves in 100 ml of water. Such higher solubility
increases microbial access to the growth substrate for metabolism.
Butane is thus approximately four times more soluble in groundwater
than methane. In accordance with the present invention, butane
injection results in large radii of influence at injection
wellheads.
[0103] Various propane-utilizing and butane-utilizing bacteria were
characterized as follows. Microorganism identification is based on
the Similarity Index. The Similarity Index in the Microbial
Identification System (MIS) is a numerical value which expresses
how closely the fatty acid composition of an unknown sample
compares with the mean fatty acid methyl ester composition of the
strains used to create the library entry listed as its match. The
database search presents the best matches and associated similarity
indices. An exact match of the fatty acid make-up of the unknown
sample to the mean of a library entry results in a similarity index
of 1.000. The similarity index will decrease as each fatty acid
varies from the mean percentage. Strains with a similarity of 0.500
or higher and with a separation of 0.100 between first and second
choice are good matches (good or excellent). A similarity index
between 0.300 and 0.500 may be a good match but would indicate an
atypical strain (OK). Values lower than 0.300 suggest that the
species is not in the database but those listed provide the most
closely related species (weak or poor).
[0104] In the cases where a strain remained unidentified after
fatty acid analysis, the Biolog system was employed where
microorganisms are identified by comparing substrate utilization
characteristics of the unknown isolate to the Biolog database.
[0105] The following isolates were chosen for identification at two
independent laboratories: propane-utilizers 2EP, 3EP, 4HP, 6HP, 6NP
and 8NP; and butane-utilizers 2EB, 2HB, 3EB, 3NB, 4EB, 4HB, 4NB,
SEB, 6HB, 6NB and 7NB.
[0106] The majority of the propane-utilizers and butane-utilizers
were characterized as different genera/species by both laboratories
for the comparison-pair isolates 2EP-2EB, 3EP-3EB, 4HP-4HB,
6HP-6HB, and 6NP-6NB, thus indicating that the butane-utilizers are
a distinct class of microorganism from the propane degraders. Since
methane-utilizing bacteria are obligate methane oxidizers, no
isolates from the methane microcosms were submitted for laboratory
analysis. Most isolates from the microcosms were mixed. Between
both laboratories. 59 genus/specie were identified with "good or
excellent" precision, 14 with "OK" precision (atypical strains) and
22 with "weak" precision (species not in database and remain as
unknowns). A summary of the butane-utilizers that have demonstrated
the ability to degrade TCE are identified in Table 12.
14TABLE 12 Sample ID Genus Species 2HB* Pseudomonas putida 2EB
Pseudomonas rubrisubalbicans 3EB Pseudomonas rubrisubalbicans 5EB
Pseudomonas aeruginosa 6NB Pseudomonas aeruginosa 2EB Variovorax
paradoxus 2HB Variovorax paradoxus 3EB Variovorax paradoxus 3NB
Variovorax paradoxus 4HB Variovorax paradoxus 4NB Variovorax
paradoxus 5EB* Variovorax paradoxus 6HB Variovorax paradoxus 2EB
Variovorax paradoxus** 6NB Variovorax paradoxus*** 7NB Nocardia
asteroides 2HB Nocardia asteroides*** 3EB Nocardia asteroides***
4HB* Nocardia asteroides*** 4NB Nocardia asteroides*** 7NB Nocardia
asteroides*** 5EB* Nocardia brasiliensis 2EB Nocardia restricta 2HB
Nocardia globerula 2HB Chryseobacterium indologenes 4HB
Chryseobacterium indologenes 7NB Chryseobacterium indologenes 5EB
Chryseobacterium meningosepticum 2EB Comamonas acidovorans 3NB
Comamonas acidovorans 6HB Comamonas acidovorans 6NB Comamonas
acidovorans 4EB Acidovorax delafieldii 4NB Acidovorax delafieldii
6NB Acidovorax delafieldii 4NB Rhodococcus rhodochrous 7NB
Rhodococcus rhodochrous 2EB Rhodococcus erythropolis 3EB
Rhodococcus erythropolis 6HB Rhodococcus erythropolis 4EB*
Rhodococcus fascians 5EB* Rhodococcus fascians 4NB Aureobacterium
barkeri 4HB Aureobacterium esteroaromaticum 4NB Aureobacterium
esteroaromaticum 6HB Aureobacterium saperdue 5EB Micrococcus
varians 7NB Micrococcus varians 7NB Micrococcus kristinae 6HB
Aeromonas caviae 6NB Aeromonas caviae 2EB Stenotrophomonas
maltophilia 3EB Stenotrophomonas maltophilia 4EB Stenotrophomonas
maltophilia 5EB Stenotrophomonas maltophilia 6HB Stenotrophomonas
maltophilia 6NB Stenotrophomonas maltophilia 4EB Sphingobacterium
thalpophilum 4NB* Sphingobacterium spiritivorum 4NB Shewanella
putrefaciens B 3NB* Phyllobacterium myrsinacearum 6HB Clavibacter
michiganense 6HB Clavibacter michiganense**** 6NB Alcaligenes
xylosoxydans 7HB* Gordona terrae 7NB Corynebacterium aquaticum B
7NB Cytophaga johnsonae * = low similarity index indicating a poor
match with the fatty-acid database. In these cases, the species in
the consortia listed was matched to a database testing substrate
utilization and remained unidentified. The (*) best describes an
unknown genera/species. ** = GC Subgroup A subspecies *** = GC
Subgroup B subspecies **** = tessellarius subspecies
[0107] Whereas particular embodiments of this invention have been
described above for purposes of illustration, it will be evident to
those skilled in the art that numerous variations of the details of
the present invention may be made without departing from the
invention as defined in the appended claims.
* * * * *