U.S. patent application number 10/434309 was filed with the patent office on 2004-07-01 for reducing hair growth, hair follicle and hair shaft size and hair pigmentation.
Invention is credited to Liu, Jue-Chen, Miller, jonathan Davis, Seiberg, Miri, Shapiro, Stanley S..
Application Number | 20040126353 10/434309 |
Document ID | / |
Family ID | 26843282 |
Filed Date | 2004-07-01 |
United States Patent
Application |
20040126353 |
Kind Code |
A1 |
Seiberg, Miri ; et
al. |
July 1, 2004 |
Reducing hair growth, hair follicle and hair shaft size and hair
pigmentation
Abstract
The present invention utilizes natural and/or synthetic serine
protease inhibitory agents or botanical extracts containing serine
protease inhibitory activity, with or without the addition of one
or more isoflavones and/or additional natural extracts containing
one or more isoflavones, and their ability to affect changes in
mammalian hair growth, hair follicle and hair shaft size and hair
pigmentation.
Inventors: |
Seiberg, Miri; (Princeton,
NJ) ; Shapiro, Stanley S.; (Roseland, NJ) ;
Liu, Jue-Chen; (Belle Mead, NJ) ; Miller, jonathan
Davis; (Lawrenceville, NJ) |
Correspondence
Address: |
PHILIP S. JOHNSON
JOHNSON & JOHNSON
ONE JOHNSON & JOHNSON PLAZA
NEW BRUNSWICK
NJ
08933-7003
US
|
Family ID: |
26843282 |
Appl. No.: |
10/434309 |
Filed: |
December 1, 2003 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10434309 |
Dec 1, 2003 |
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09621565 |
Jul 21, 2000 |
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60145774 |
Jul 27, 1999 |
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Current U.S.
Class: |
424/74 ; 424/750;
424/757; 424/758 |
Current CPC
Class: |
A61K 8/9794 20170801;
A61P 17/14 20180101; A61K 8/9789 20170801; A61K 2800/782 20130101;
A61Q 5/08 20130101; A61Q 7/02 20130101 |
Class at
Publication: |
424/074 ;
424/750; 424/757; 424/758 |
International
Class: |
A61K 035/78; A61K
007/06 |
Claims
We claim:
1. A method for effecting changes in mammalian hair appearance,
hair growth, hair pigmentation and hair follicle and hair shaft
size, comprising topical application to the skin of a mammal an
effective amount of a topically active composition comprising one
or more compounds derived from one or more of the botanical
families leguminosae, solanaceae, gramineae and cucurbitaceae.
2. The method of claim 1 wherein said extract contains at least one
serine protease inhibitory activity.
3. The method of claim 2 wherein said at least one serine protease
inhibitory active agent is a serine protease inhibitor.
4. The method of claim 3 wherein said at least one serine protease
inhibitor is present in an amount, based upon the total volume of
the topically active composition, of from about 0.0001% (w/v) to
20% (w/v).
5. The method of claim 4 wherein the at least one serine protease
inhibitor is present in an amount, based upon the total volume of
the topically active composition, of from about 0.001% (w/v) to
about 5% (w/v).
6. The method of claim 1 wherein one of said changes is a delay in
hair growth, reduced hair follicle and hair shaft size and reduced
hair pigmentation.
7. The method of claim 1 wherein said topically active composition
further comprises a pharmaceutically or cosmetically acceptable
vehicle.
8. The method of claim 1 wherein said topically active composition
further comprises one or more isoflavones.
9. The method of claim 8 wherein said topically active composition
further comprises a pharmaceutically or cosmetically acceptable
vehicle.
10. The method of claim 1 wherein said topically active composition
further comprises natural extracts containing one or more
isoflavones.
11. The method of claim 1 wherein said composition is applied
topically in conjunction with one or more products whose purpose is
to either facilitate the removal of hair or actually remove hair or
reduce hair visibility or improve hair style or improve hair
management.
12. The method of claim 1 wherein said composition is applied
topically before or following hair removal.
13. The method of claim 1 wherein said composition is applied
topically during hair removal.
14. The method of claim 1 wherein said composition is applied
topically in conjunction with one or more of the group consisting
of: depilatory agents, shampoo, hair conditioner, styling gel, hair
care products, waxing products, shaving products, hair-removal
products, after-shave products, deodorant, anti-perspirant,
electrolysis, laser hair removal, light-induced hair removal, mask,
bath additives.
15. The method of claim 1 further comprising leaving said
composition on said skin for a period sufficient to effect said
changes.
16. The method of claim 1 wherein said period is daily treatment
for at least about four weeks.
17. A method according to claim 16 wherein said composition is
applied daily for at least eight weeks.
18. A method according to claim 1 wherein said composition is
applied daily to reduce or prevent pseudofolliculitis barbae.
19. A method according to claim 1 wherein said composition is
applied daily to the axilliary area to reduce hair growth.
20. A method according to claim 1 wherein said composition is
applied daily to the scalp to style and improve management of
African type hair.
21. A method according to claim 1 wherein said composition is added
daily to a bath.
22. A method according to claim 1 wherein said composition is used
daily on facial or body parts for delaying hair growth and reducing
hair visibility in sufferers of hirsutism.
23. A composition for affecting changes in mammalian hair growth,
hair pigmentation and hair shaft and follicle size, comprising
topically applying to skin of a mammal an effective amount of a
topically active composition comprising one or more compounds
derived from one or more of the botanical families leguminosae,
solanaceae, gramineae and cucurbitaceae.
24. The composition of claim 23 wherein the extract contains at
least one serine protease inhibitory activity.
25. The composition of claim 24 wherein the at least one serine
protease inhibitory active agent is present in an amount, based
upon the total volume of the topically active composition, of from
about 0.0001% (w/v) to about 20% (w/v).
26. The composition of claim 25 wherein the at least one protease
inhibitory active agent is present in an amount, based upon the
total volume of the topically active composition, of from about
0.001% (w/v) to about 5% (w/v).
27. The composition of claim 23 wherein the composition further
comprises one or more isoflavones.
28. The composition of claim 23 wherein the composition further
comprises one or more natural extracts containing one or more
isoflavones.
29. The composition of claim 23 wherein the composition further
comprises surfactants, moisturizers, humectants, foaming agents,
cosmetic adjuvants, buffering agents, preservatives, anti-oxidants,
conditioners, depigmenting agents, hair removal agents, anti-aging
agents, sunscreens, fragrances, colorants, or mixtures thereof.
30. The composition of claim 23 wherein said composition is in the
form of liquid, cream, gel, paste, powder, essence, mousse,
emulsion, film patch or spray.
31. The composition of claim 23 wherein said composition further
comprises one or more products whose purpose is to either
facilitate the removal of hair or actually remove hair or reduce
hair visibility or improve hair management.
32. The composition of claim 23 wherein said composition further
comprises one or more of the group consisting of: depilatory cream,
shampoo, hair conditioner, styling gel, hair care products, waxing
products, shaving products, hair removal products, after-shave
products, deodorant, anti-perspirant, pre- or post -electrolysis
products, pre- or post-laser hair removal products, pre- or
post-light-induced hair removal products, mask, bath additives.
33. A composition according to claim 23 wherein said compound is
derived from legumes.
34. A composition according to claim 33 wherein said compound is
derived from soy, lima and/or black beans.
35. A composition according to claim 34 wherein said compound is
selected from the group consisting of soybean milk, limabean milk,
blackbean milk, soybean extract, limabean extract, blackbean
extract, soybean paste, limabean paste, blackbean paste, soybean
powder, blackbean powder, limabean powder, soymilk powder,
blackbean milk powder and limabean milk powder and mixtures
thereof.
36. A composition according to claim 35 wherein said compound is a
fraction of soybean milk, soybean extract, soybean paste, limabean
milk, limabean extract, limabean paste, blackbean milk, blackbean
extract, blackbean paste, soybean powder, blackbean powder,
limabean powder, soymilk powder, blackbean milk powder and limabean
milk powder and mixtures thereof.
37. A composition according to claim 35 comprising bean milk in an
amount of from about 1 to about 99% by weight.
38. A composition according to claim 35 comprising soybean powder,
soymilk powder or a mixture thereof in an amount of from about 1 to
about 99% by weight.
39. A composition according to claim 36 comprising soybean trypsin
inhibitor, limabean trypsin inhibitor or blackbean trypsin
inhibitor in an amount of from about 0.0001 to about 20% by
weight.
40. A composition according to claim 33 said composition further
comprising one or more isoflavones.
41. A composition according to claim 33 said composition further
comprising one or more natural extracts containing one or more
isoflavones.
42. A composition according to claim 40 wherein said one or more
isoflavones are present in said composition in amount of from about
0.00001 to about 0.1% by weight.
43. A composition according to claim 41 wherein said one or more
natural extract containing said one or more isoflavones are present
in said composition in amount of from about 0.00001 to about 0.1%
by weight.
44. A cosmetic composition according to claim 23, said composition
further comprising a cosmetically-acceptable vehicle.
45. A composition according to claim 23 wherein said composition
further comprises liposomes.
46. A composition according to claim 45 wherein said composition
comprises glycerol dilaurate, cholesterol,
polyoxyethylene-10-stearyl ether and polyoxyethylene-9-lauryl
ether.
47. A composition according to claim 23 wherein said composition
comprises from about 1 to about 99% bean milk, from about 0.1 to
about 20% emulsifier and preservatives in an effective amount.
48. A composition according to claim 23 wherein said composition
comprises from about 0.5 to about 20% soybean powder or soymilk
powder, from about 0.1 to about 20% emulsifier and preservatives in
an effective amount.
49. A composition according to claim 47 wherein said composition
further comprises one or more isoflavones in an amount of from
about 0.00001 to about 0.1%.
50. A composition according to claim 47 wherein said composition
further comprises one or more natural extracts comprising one or
more isoflavones in an amount of from about 0.0001 to about
0.1%.
51. A composition according to claim 23 wherein said composition
further comprises one or more isoflavones in an amount of from
about 0.0001 to about 0.1%.
52. A composition according to claim 48 wherein said composition
further comprises one or more natural extracts comprising one or
more isoflavones in an amount of from about 0.0005 to about 0.05%.
Description
FIELD OF THE INVENTION
[0001] This invention is related to methods and compositions
effective for reducing hair growth. More specifically, the present
invention is directed to methods for changing the rate of hair
growth, reducing the size of the hair follicle and the hair shaft,
and reducing hair shaft pigmentation, by topical application of
either botanical extracts containing serine protease inhibitory
activity and in particular soybean extracts such as soymilk, or
mixtures and formulations of the above, combined with other active
ingredients such as isoflavones.
BACKGROUND OF THE INVENTION
[0002] One main function of mammalian hair is to provide
environmental protection. However, that function has largely been
lost in humans, in whom hair is kept or removed essentially for
social and cosmetic purposes.
[0003] Many procedures are used to remove unwanted hair including
shaving, electrolysis, plucking, laser and light therapies and
injection of therapeutic antiandrogens. These conventional methods
are not without their shortcomings. Shaving, for instance, may
result in nicks and cuts in the skin's surface, may leave a
perception of an increase in the rate of hair regrowth, and may
also leave undesirable stubble. While electrolysis may keep an area
free of unwanted hair for a prolonged period of time, the process
is often expensive and painful and may further result in scarring.
Not only may plucking cause pain and discomfort, but it often
result in a poor removal of short hair. Several unwanted side
effects, such as effects on muscularity, often accompany the use of
antiandrogens. For these reasons, better methods for reducing hair
growth are needed.
[0004] Pseudofolliculitis barbae is an inflammatory hair disorder,
most commonly found on the beard area. Inflammatory follicular
papules result when hair tips penetrate into the skin rather than
passing through the follicular orifice. This process is extremely
common in black men because their hairs are frequently curly,
exiting the skin at an acute angle. Close shaves, particularly with
a razor blade, predispose them to pseudofolliculitis barbae. The
most effective treatment available is to allow the hairs to grow
well beyond the skin surface. Such a treatment is often not
desired.
[0005] Hirsutism is a relatively frequent condition affecting about
4% of women. Facial hirsutism often interferes with personal and
work activities, and temporary hair removal is a major component in
the management of hirsute patients. Shaving is the most frequently
used temporary method for facial hair, as plucking, waxing and
depilatories are more difficult to tolerate and care must be taken
to avoid folliculitis, pigmentation, and scarring. Cosmetic
cover-ups are usually used to hide cuts and stubble and
electrolysis and thermolysis may be used for permanent hair removal
when affordable.
[0006] An alternative or complementary desired approach to hair
removal, would be a method to reduce hair growth, reduce hair
follicle and hair shaft size and reduce hair shaft pigmentation.
Such an approach could reduce the visibility of existing hair,
making it softer and lighter. When combined with other methods of
hair removal such a method could enhance and prolong the removal
effect, and reduce the need and frequency of hair removal. Long
term use of such an approach could lead to attenuated, soft,
pigmentation-reduced hair growth, that is less visible and does not
require the use of other removal methods.
[0007] Reduced hair growth is desired in the axilla area (fossa
axillaris), where deodorants and anti-perspirants are used to
control odor trapped within the axillary hairs. It would be desired
to have products for under-arm use, which combine deodorant or
anti-perspirant activities with reduced hair growth activity.
[0008] African type hair is unique in its morphology--a kinky hair
shaft with variations in diameter. This complex shaft structure
creates the need for specialized grooming products and procedures
to ensure that the African type hair maintains cosmetic desired
properties. It is desired to have products that reduce this
complexity and make the African type hair more manageable,
improving its appearance.
[0009] It would be desirable to provide a method for chemically or
naturally affecting hair growth, hair follicle and hair shaft size
and hair shaft pigmentation, which does not cause unwanted side
effects to the user.
SUMMARY OF THE INVENTION
[0010] In accordance with the present invention, we have found
compositions and methods for affecting changes in mammalian hair
growth, hair follicle and hair shaft size and hair pigmentation by
topically applying to the skin of a mammal an effective amount of a
topically active composition comprising protease inhibitors,
botanical extracts, and in particular legume extracts including,
but not limited to, soymilk, for a period of time sufficient to
affect hair growth, hair follicle and hair shaft size and hair
shaft pigmentation. Such topically active compositions may be
further combined with other active ingredients including, but not
limited to, synthetic or naturally occurring isoflavones, to
enhance the desired effects on hair growth and pigmentation.
[0011] The compositions and methods of this invention provide a
unique, convenient means for delaying hair growth, reducing hair
follicle and hair shaft size and hair shaft pigmentation, by using
serine protease inhibitors, botanical extracts of the legume
family, and in particular, but not limited to, soymilk, containing
serine protease inhibitory activity, and their combinations with
isoflavones.
BRIEF DESCRIPTION OF THE DRAWINGS
[0012] The file of this patent contains at least one drawing
executed in color. Copies of this patent with color drawing(s) will
be provided by the Patent and Trademark Office upon request and
payment of the necessary fee.
[0013] The invention will be more fully understood and further
advantages will become apparent when reference is made to the
following detailed description of the invention and the
accompanying drawings in which:
[0014] FIG. 1: A photograph of control and soymilk treated C57Bl/6
mouse hair (high magnification).
[0015] FIG. 2: Histological sections of control and soymilk-treated
C57Bl/6 mouse hair follicles at day four of the hair cycle.
[0016] FIG. 3: Histological sections of control and soymilk-treated
C57Bl/6 mouse hair follicles at day seven of the hair cycle, high
and low magnifications.
[0017] FIG. 4: Histological sections of control and soymilk-treated
C57Bl/6 mouse hair follicles at day 18 of the hair cycle.
[0018] FIG. 5: Histological sections of control and soymilk-treated
C57Bl/6 mouse hair follicles at day 21 of the hair cycle.
[0019] FIG. 6: Histological sections of control, soymilk, and
soymilk-derived proteins-treated C3H mouse hair follicles (High
magnification) at day seven of the hair cycle.
[0020] FIG. 7: Histological sections of control, soymilk, and
soymilk-derived proteins-treated C3H mouse hair follicles (lower
magnifications) at day seven of the hair cycle.
[0021] FIG. 8: Histological sections of control and soymilk-treated
C3H mouse hair follicles at day 21 of the hair cycle.
[0022] FIG. 9: A photograph of control and soymilk-derived proteins
treated C3H mouse hair (high magnification).
[0023] FIG. 10: Histological sections of control and
soymilk-derived proteins-treated C57Bl/6 mouse hair follicles (High
magnification) at day eight of the hair cycle.
[0024] FIG. 11: A graph demonstrating the trypsin inhibitory
activity of soymilk.
[0025] FIG. 12: Western blot of C57Bl/6 mouse skins throughout the
hair cycle, demonstrating reduced tyrosinase and TRP-1 protein
levels following soymilk treatment.
[0026] FIG. 13: Photographs of untreated and soymilk treated sides
of human face, treated with soymilk daily for four weeks.
[0027] FIG. 14: Quantitative analysis of hair follicle dimensions
with and without soymilk treatment.
[0028] FIG. 15: Photographs of human leg hair following five weeks
of soymilk treatment on one leg.
[0029] FIG. 16: Photographs of control, soymilk, and
isoflavone-enriched soymilk treated C57Bl/6 mouse hair (high
magnification).
[0030] FIG. 17: Histological sections of control, soymilk, and
isoflavone-enriched soymilk treated C57Bl/6 mouse skins at day 15
of the hair cycle, documenting the thickness and color of the hair
shafts.
[0031] FIG. 18: Photograph of C57Bl/6 mouse hair after three weeks
of treatment with various soymilk and isoflavones formulations.
DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS
[0032] As used herein, "mammal" shall mean any member "of the
higher vertebrate animals comprising the class Mammalia," as
defined in Webster's Medical Desk Dictionary 407 (1986), and
includes but is not limited to humans. As used herein "(%, w/v)"
shall mean grams of a given component per 100 ml of the total
composition.
[0033] Topically active agents suitable for use in the composition
of the present invention include protease inhibitors and natural
plant extracts having protease inhibitory activity and mixtures
thereof. Preferred protease inhibitors are serine protease
inhibitors, and in particular Soybean Trypsin Inhibitor ("STI") and
the soybean-derived Bowman Birk Inhibitor ("BBI). Preferred
botanical extracts are of the legume family and in particular bean
extracts, such as soymilk. Preferably, the protease inhibitors are
present in an amount, based upon the total volume of the
composition of the present invention, of from about 0.0001% (w/v)
to about 20% (w/v), and more preferably from about 0.001% (w/v) to
about 5% (w/v). Preferably, botanical aqueous extracts such as
soymilk are present in an amount of 10-99% (v/v), and more
preferably from 50-99% (v/v).
[0034] We have unexpectedly found that when topically active agents
such as described above, and in particular soymilk or soymilk
containing formulations, are enriched with isoflavones, and in
particular soybean-derived isoflavones, the inhibitory effect on
hair growth, hair dimensions and hair pigmentation is enhanced.
Preferably, the isoflavones are present in the botanical aqueous
extracts such as soymilk in an amount of 0.000005-15% (v/v), and
more preferably from 0.00001-10% (v/v).
[0035] If the delivery parameters of the topically active
pharmaceutical or cosmetic agent so require, the topically active
composition of the present invention may be further comprised of a
pharmaceutically or cosmetically acceptable vehicle capable of
functioning as a delivery system to enable the penetration of the
topically active agent into the hair follicle and the skin.
[0036] The pharmaceutical or cosmetic composition may be optionally
combined with other ingredients such as moisturizers, cosmetic
adjuvants, anti-oxidants, depigmenting agents, anti-aging agents,
hair removal agents, hair styling agents, sunscreens surfactants,
foaming agents, conditioners, humectants, fragrances, colorants,
viscosifiers, buffering agents, preservatives, and the like and
mixtures thereof. These will be combined in an amount which will
not affect the serine protease inhibitory activity, in order to
produce cosmetic or pharmaceutical products such as,
non-exclusively, essences, creams, lotions, pastes, gels, powders,
patches or injectables and the like for the reduction of hair
growth, hair size and hair pigmentation.
[0037] The compositions of this invention may be applied prior to,
concurrently with or after other active ingredients or compositions
to enhance their effect. For example, the compositions of this
invention may be applied in conjunction with one or more products
whose purpose is to facilitate the removal of hair to to actually
remove hair, reduce hair visibility, improve hair style or improve
hair management. The compositions of this invention may be applied
topically prior to, during or following hair removal. They may be
applied topically concurrently with one or more of the following
group: depilatory agents, shampoo, hair conditioner, styling gel,
hair care products, waxing products, shaving products, hair-removal
products, after-shave products, deodorant, anti-perspirant,
electrolysis, laser hair removal, light-induced hair removal, mask
or bath additives.
[0038] The compositions of this invention may be applied daily for
at least four to eight weeks, by which an effect upon the
appearance of hair should be observed. Application may be continued
as long as desired to maintain the condition of the hair. Daily
application to the face may mitigate the condition of
pseudofolliculitis barbae and/or hirsutism; application to the
axillary area may reduce hair growth under the arms and application
to the scalp and hair may assist in managing and styling
African-type hair.
[0039] The topically active pharmaceutical or cosmetic composition
should be applied in an amount effective to effect changes in
mammalian hair growth, hair follicle and hair shaft size and hair
shaft pigmentation. As used herein "amount effective" shall mean an
amount sufficient to cover the region of skin surface where a delay
in hair growth and hair pigmentation and reduced hair size are
desired. Preferably, the composition is applied to the skin surface
such that, based upon a square cm of skin surface, from about 2
.mu.l/cm.sup.2 to about 500 .mu.l/cm.sup.2 of topically active
agent is present when a delay in hair growth, hair size and hair
pigmentation is desired.
[0040] We have unexpectedly found that when topically active
agents, such as soymilk, or isoflavone-enriched soymilk are
topically applied to an animal's skin, a significant delay in hair
growth, hair follicle and hair shaft size and hair shaft
pigmentation was achieved. We further believe that since the hair
growth cycle for humans is often slower than that for mice, it is
further likely that the hair growth delay in humans would be
considerably longer than in mice.
[0041] The invention illustratively disclosed herein suitably might
be practiced in the absence of any component, ingredient, or step
which is not specifically disclosed herein. Several examples are
set forth below to further illustrate the nature of the invention
and the manner of carrying it out. However, the invention should
not be considered as being limited to the details thereof.
EXAMPLES
Example 1
[0042] Depilation of Test Subjects in the Mouse System
[0043] C57BI/6 or C3H mice (male and female) were obtained from
Charles River (Kingston, N.Y.), at 8 -10 weeks of age and were in
the resting (telogen) phase of their respective hair cycle. Hair
growth was induced by wax depilation (plucking) of each respective
animal's back fur according to the procedure set forth in Stenn, et
al., "Glucocorticoid Effect on Hair Growth Initiation: A
Reconsideration," 6 Skin Pharmacol. , 125-134 (1993) In C57Bl/6 and
C3H mice, 8-10 weeks old, the growth phase (anagen) starts
synchronously in all hair follicles at the time of depilation. As
illustrated in Table 1, the following observations were noticed at
the induction site:
1TABLE 1 Observations at Induction Site Morphological and
Histological Days Post-Induction Observations at the induction site
1-2 (early anagen) new follicle starts to grow 3 to 4 hair
follicles were fully developed, but the hair shafts were not yet
visible 7 to 8 (late anagen) each mouse had very dark skin; their
hair shafts are histologically visible 11-12 the hair shafts
started to penetrate through the epidermis. 14 each mouse was
covered with short hairs 19 the regression of the follicle
(catagen) was observed histologically 21 to 25 the hair follicle is
back to resting phase.
[0044] As shown in Table 1, the hair growth was visible several
days after depilation as the pink skin of the animal began to
darken. This is likely due to hair pigmentation in the shaft since
the C57BI/6 and C3H mice contained melanocytes only in the hair
follicles and not in the dorsal epidermis. Similar hair growth
pattern was documented in our international application No. PCT/US
97/11033, when chemical depilation using commercially available
products was performed.
[0045] Since the murine hair cycle varies not only between strains,
but also amongst individual animals, the status of the hair cycle
was analyzed in each animal on study. A 2 cm by 1 cm skin sample
was isolated from each mouse with scissors, fixed with a 10%
buffered formalin solution having a pH of about 6.9-7.1 at
25.degree. C. (Stephens Scientific), and then formed into a
paraffin block according to well-known procedures. The block was
then microtomed, and sections were stained with H&E or
Fontana-Mason stain. Sections were examined histologically in order
to verify the phase of the hair cycle, the size of the hair
follicle and hair shaft and the level of hair pigmentation, using
procedures well known in the art. Hair length was assessed
visually, and by using a low magnification (.times.8) dissecting
microscope.
[0046] This Example, as well as the one described in our
international application No. PCT/US 97/11033, shows that the hair
growth cycle for C57BI/6 and C3H mice averaged about 25 days and
reports similar timing of hair follicle and shaft development
regardless of the method used for depilation.
Example 2
[0047] Preparation of Soymilk and Soymilk Formulations
[0048] One way to make soymilk is to soak the soybeans in deionized
or purified water for several hours, and grind them after they were
fully hydrated, with the addition of small quantities of water.
(The grinding process allows the soybean milk to be extracted).
After collection, the soybean milk may be filtered to remove any
residual parts of the bean husk. The soymilk used in the
formulations described below can be fresh soymilk as described
above, or may be made from soybean powder and water. The soybean
powder is milled from soybeans and may also be lyophilized, spray
dried, or freeze-dried and the resulting soymilk may or may not be
filtered. Such prepared soymilk may have from about 1 to about 90%
by weight dry soybean powder. Another example is the use of soymilk
powder, made from lyophilized, spray dried or freeze-dried soymilk,
with the addition of water and finished with or without filtration
or homogenization. Other methods of soybean extraction could also
be used to create the active ingredients in the formulations
described below. For example, the active ingredients could be
extracted from ground soybeans using ethanol/water mixtures,
followed by the removal of the ethanol from the extract, in such
ways that the serine protease inhibitory activity of the soybean
will be retained, and preferably that the protein STI will remain
intact.
[0049] The soy products useful in this invention may be produced
from all soybean species, regardless of their geographic origin,
sun exposure, harvest time and the like. However, specific strains,
geographic origins or growth conditions might be preferred. For
example, but not limiting to, soybean strains particularly rich in
its Soybean Trypsin Inhibitor (STI) content or in isoflavone
content, or growth conditions that result in STI or isoflavone
enrichment in the bean, might be preferred. It should be noted that
the soy products useful in the compositions of this invention have
a distinctive odor, which may be tolerable in some cultures, but is
undesired in others. If necessary, the odor of the compositions of
this invention may be reduced by using soybean products derived
from specific strains of soybeans known to produce reduced-odor,
including, but not limited to, lipoxygenase-2-deficient beans and
those having modified sugar profile, and the like. A process to
reduce oxygen levels in the formulation may also reduce the odor.
Various masking agents or fragrances may also be used to mask the
odor.
[0050] The compositions of this invention may further comprise
surfactants, moisturizers, humectants, conditioners, fragrances,
colorants, preservatives, anti-oxidants, depigmenting agents, hair
removal agents, anti-aging agents, sunscreens, foaming agents,
cosmetic adjuvants, buffering agents or mixtures thereof.
[0051] The compositions of this invention may be left on the skin
for a period sufficient to effect changes. For example, the
compositions of this invention may be applied to the skin daily
treatment for at least about four weeks, more preferably, the
composition should applied daily for at least eight weeks.
[0052] Another method according to this invention is a method to
reduce or prevent pseudofolliculitis barbae. Daily application of
the compositions of this invention may reduce or prevent this
condition. The compositions of this invention may also be applied
daily to the axilliary area to reduce hair growth. Furthermore, the
compositions of this invention may be applied daily to the scalp to
style and improve management of African type hair.
[0053] As shown in our co-pending U.S. patent application (Attorney
Docket No. JBP 430), numerous soymilk-based formulations could be
used to reduce pigmentation. All these formulations could also be
used to reduce hair growth. Some particularly preferred examples of
soymilk formulations and soymilk formulations containing
isoflavones are shown in table 2 below. An example for an
isoflavones preparation that could be used in this invention is
Flavosterone SE from Ichimaru, Japan, which contains about 0.1%
pure isoflavones. In all these formulations, soymilk could be
replaced with the appropriate quantities of soybean powder or
soymilk powder and water.
2TABLE 2 Soymilk Essence formulations: Soybean Essences 1 6 8 21 22
23 24 25 26 Soymilk 87.42% 89.04% 96.09% 96.05% 96.05% 95.70%
94.40% 94.40% 92.40% Phenoxyethanol 0.73% Phenoxyethanol and 1.00%
1.00% 1.00% 1.00% 1.00% 1.00% 1.00% 1.00% Parabens Glycerin 2.50%
2.50% Cyclomethicone 2.00% Aluminum Starch 0.75% Ocetyl Succinate
Sucrose Cocoate 1.00% 1.00% PEG-6 3.00% 3.00% Capric/Caprylic
Triglycerides Disodium EDTA 0.10% 0.10% 0.05% 0.05% 0.05% 0.05%
Polyacrylamide/Laur 2.50% 2.75% 2.90% 2.90% 2.90% 3.20% 3.50% 3.50%
3.50% eth-7/C.sub.13-14 Isoparrafin Ascorbic Acid 0.01% 1.00%
Butylated 0.10% 0.01% 0.05% 0.05% 0.05% 0.05% 0.05% 0.05%
Hydroxytoluene Polysorbate 20 0.50% Lactoferrin 1.00% 1.00% 1.00%
Tocopherol 1.00% TOTAL 100.00% 100.00% 100% 100% 100% 100% 100%
100% 100% 27 28 29 30 31 32 33 34 35 Soymilk 90.70% 94.70% 85.70%
90.70% 93.70% 94.70% Phenoxyethanol and 1.00% 1.00% 1.00% 1.00%
1.00% 1.00% 1.00% 1.00% 1.00% Parabens Glycerin 5.00% Disodium EDTA
0.05% 0.05% 0.05% 0.05% 0.05% 0.05% 0.05% 0.05% 0.05%
Polyacrylamide/Laur 3.20% 3.20% 3.20% 3.20% 3.20% 3.20% 3.20% 3.20%
3.20% eth-7/C.sub.13-14 Isoparrafin Ascorbic Acid Butylated 0.05%
0.05% 0.05% 0.05% 0.05% 0.05% 0.05% 0.05% 0.05% Hydroxytoluene
Deionized Water 90.70% 90.70% 85.70% Dow Corning 200 1.00% Fluid
Flavosterone SE 10.00% 5.00% 2.00% 1.00% Soymilk Powder 5.00%
Soybean Extract 5% 10% using Ethanol/Water Mixture TOTAL 100% 100%
100% 100% 100% 100% 100% 100% 100%
Example 3
[0054] Preparation of Topically Active Compositions Containing
Soybean Derived Protease Inhibitors
[0055] Soybean trypsin inhibitor (STI) and Bowman-Birk inhibitor
(BBI), from Sigma-Aldrich Corporation were mixed into a 0.1M
phosphate buffered saline (PBS, Gibco-BRL, Gaithersburg, Md.), pH
7.4, in concentrations of 1% to 0.001% (w/v). Four volumes of the
resulting solutions were then mixed with 1 volume of (100 mg/ml)
liposomes vehicle, which was prepared by the methods described in
Niemiec et. al, in order to yield the topically active composition.
Non-ionic liposomes preparations, such as those disclosed in
Niemiec et al., "Influence of Nonionic Liposomal Composition On
Topical Delivery of Peptide Drugs Into Pilosebaceous Units: An In
Vivo Study Using the Hamster Ear Model," 12 Pharm. Res. 1184-88
(1995) ("Niemiec"), which is incorporated by reference herein in
its entirety, are well known in the art, and are described our U.S.
patent application (Attorney Docket No. JBP 430). GDL liposomes
were prepared as set forth in Niemiec, et al., above, with the
exception of the following changes: the non-ionic liposomal
formulation contained glycerol dilaurate (Emulsynt GDL, ISP Van
Dyk)/cholesterol (Croda)/polyoxyethylene-10-stearyl ether (Brij76,
ICI)/polyoxyethylene-9-- lauryl ether, as at ratio of
37.5:12.5:33.3:16.7. Either PBS or Hepes buffer, 0.05M, pH 7.4
(Gibco-BRL of Gaithersburg, Md.) were used as the aqueous phase in
the preparation of the liposomes.
Example 4
[0056] Soymilk Delays Hair Growth and Reduce Hair Follicle and Hair
Shaft Size and Hair Shaft Pigmentation
[0057] C57Bl/6 mice were induced for a new hair cycle as described
in Example 1, and treated daily with soymilk. Animals were observed
daily for their hair growth pattern, and skin biopsies were taken
at important time points of the hair cycle. As a result of soymilk
treatment the hair growth of the treated mice was delayed, and
their hairs were visibly thinner, and smoother to touch. Treated
mice did not show skin darkness at days 7-8 of the hair cycle, as
expected, and hair shafts were not visible at days 11-12 as in the
control animals. In average, the hair cycle of the soymilk treated
mice was delayed by 3-6 days. FIG. 1 is a picture of the mice fur,
showing the difference in hair appearance, color, size and
thickness following soymilk treatment.
[0058] Histological examination of the biopsied skin samples
confirmed these observations. As shown in FIG. 2 by Fontana-Mason
(F&M) staining, at day four of the hair cycle the untreated
hair follicle is fully developed, as expected, containing all the
cellular layers and pigment deposition. In contrast, the soymilk
treated sample, (shown at same magnification), shows a smaller and
not as fully developed hair follicle, with no pigment
deposition.
[0059] FIG. 3 shows two sets of histological sections stained with
F&M, of lower and higher magnification. These sections are from
day seven of the hair cycle. The upper panel shows that soymilk
treated skin has smaller, shorter, and less pigmented hair
follicles than the untreated control. The lower panel shows a
higher magnification of the follicles, further demonstrating the
difference in hair follicle and hair shaft size and pigmentation
following soymilk treatment.
[0060] FIG. 4 shows low magnification of F&M stained skin
sections at day 18 of the hair cycle. At this magnification it is
obvious that soymilk treatment results in reduced hair follicle
size, which leads to reduced hair shaft length and thickness, and
reduced total pigment deposition within the treated follicles.
[0061] FIG. 5 shows skin sections at day 21 of the hair cycle, with
two magnifications. The upper panel demonstrates that the control
animals were in the catagen stage, when hair follicles are
regressing. Soymilk treated follicles, on the other hand, had
already completed the catagen stage, as they are shown in telogen,
the resting stage. This indicates that not only the hair cycle was
delayed following soymilk treatment, it was also prematurely
terminated. The lower panel demonstrates the catagen control
follicle and the shorter, telogen (resting) soymilk-treated
follicle using higher magnification.
Example 5
[0062] The Effects of Soymilk on Hair Growth, Size and Pigmentation
are Reproducible in C3H Mice
[0063] In order to verify that the effect of soymilk on hair growth
is not specific to C57Bl/6 mice, we repeated the experiment
described in Example 4 using the brown haired (Agouti) C3H mice.
The results of these experiments were similar both visually and
histologically. Soymilk delayed hair growth and reduced hair
follicle and hair shaft size and pigment deposition in the C3H
mice.
[0064] Histological analysis confirmed these visual observations.
As shown in the upper panel of FIG. 6, using F&M staining, at
day seven of the hair cycle soymilk treated follicles are smaller
and accumulate less pigment than untreated controls. The upper
panel of FIG. 7 (F&M staining) shows a lower magnification of
the same skin sections, demonstrating the thinner and less
pigmented follicles following soymilk treatment.
[0065] FIG. 8 shows F&M stained skin sections at day 21 is of
the hair cycle. As shown for the C57Bl/6 mice, following soymilk
treatment the hair cycle terminates prematurely. Soymilk treated
follicles are in the resting state, while untreated control
follicles are still in catagen.
Example 6
[0066] The Effects of Soymilk and Soybean Derived Serine Protease
Inhibitors on Hair Growth, Size and Pigmentation
[0067] In search for a mechanism to explain the effect of soymilk
on hair growth, we tested the effect of the soymilk-derived serine
protease inhibitors, STI and BBI. We had shown earlier that these
proteins induce depigmentation in skin, by affecting the PAR-2
pathway (U.S. patent application, Attorney Docket No. JBP 430).
[0068] The experiments described in Example 4 were repeated using
STI, BBI, and soymilk. STI and BBI were used in a PBS-liposome
vehicle as described in Example 3. Visual observations throughout
the hair cycle confirmed that both STI and BBI could delay hair
growth and reduce hair follicle and hair shaft size, similar to
soymilk (see hair pictures in FIG. 9). Using high concentrations of
STI or BBI, the effect on hair growth and pigmentation was
substantial.
[0069] Histological analysis confirmed these finding. As shown in
FIG. 6, at day seven of the hair cycle 1% of STI and 1% of BBI
reduce hair follicle and hair shaft size and hair shaft
pigmentation in C3H mice. FIG. 7 shows lower magnification sections
of the same day into the hair cycle, demonstrating smaller hair
follicles and hair shafts and reduced pigmentation, relative to
untreated control, with soymilk, STI or BBI treatment. FIG. 10
shows that STI and BBI have the same effect in C57Bl/6 mice too,
demonstrating smaller and less pigmented follicles. Taken together,
this example shows that STI and BBI are soybean-derived serine
protease inhibitors, found in soymilk, that could delay hair
growth, reduce hair follicle and hair shaft size and reduce hair
pigmentation. STI and BBI could represent a part of the soymilk
ingredients that affects hair growth.
[0070] In order to support the hypothesis that STI and BBI in
soymilk are involved in the hair growth effects described above, we
tested soymilk for its serine protease inhibitory activity. An
enzymatic assay was performed using "Enzchek", a protease digestion
fluorescent test system made by Molecular Probes of Eugene, Oreg.
Using 100 units of trypsin (from Sigma chemicals, St. Louis Mo.)
the test system produced fluorescence reading of about 1100 units.
This reaction was inhibited with increasing concentrations of STI,
as expected from a known trypsin inhibitor. Serial dilutions of
soymilk were tested in this assay, and found to inhibit trypsin
activity. As shown in FIG. 11, soymilk exerts trypsin inhibitory
activity similar to about 0.2% of pure STI. This suggests that
soymilk could exert its hair growth effect, at least in part, by
STI and BBI.
Example 7
[0071] Soymilk Induces Changes in Tyrosinase and TRP-1 Protein
Expression
[0072] The histological analyses of soymilk treated skin samples
described in the examples above show dramatic reduction in pigment
deposition within the hair follicle. To further understand the
mechanism of soymilk-induced depigmentation, we studied tyrosinase,
the key enzyme in melanogenesis and Tyrosinase-Related Protein-1
(TRP-1), the enzyme that stabilizes tyrosinase. C57Bl/6 and C3H
mice were treated as described above, and samples were collected
throughout the study for protein analysis. Protein extraction and
Western blot analysis were performed using standard procedures,
such as the one described in Current Protocols in Cell Biology,
Edited by Juan S. Bonifacino et al. Chapter 6: Electrophoresis and
Immunoblotting. Copyright 1999 by John Wiley & Sons, Inc.,
which is incorporated herein by reference in its entirety. An
example of one such study is shown in FIG. 12.
[0073] Equal amounts of skin-extracted proteins were probed with
the anti-tyrosinase antibody "anti PEP1", and with the anti-TRP-1
antibody "anti PEP7" which are described in Jimenez, M., Kameyama,
K., Maloy, W L, Tomita Y., and Hearing, V. Mammalian tyrosinase:
biosynthesis, processing and modulation by melanocyte stimulating
hormone. Proc. Natl. Acad. Sci. USA (1988), 85:3830-34, and
Jimenez, M., K., Maloy, W L, and Hearing, V. Specific
identification of an authentic tyrosinase clone. J. Biol. Chem.
(1989) 264:3397-3403, which are incorporated herein by reference in
their entirety.
[0074] As shown in FIG. 12, The expression of Tyrosinase and TRP-1
proteins is dramatically affected by soymilk treatment. Tyrosinase
and TRP-1 levels are reduced, and the duration of the expression is
shortened. These two factors affect overall hair pigmentation,
which is reduced due to the reduced level and shorter duration of
melanogenesis.
Example 8
[0075] Soymilk Reduces Human Facial Hair Length and Thickness
[0076] An individual male with dark facial hair who shaves daily
was treating the right side of his face with soymilk, immediately
after shaving, for five weeks. By the third week, and more
noticeably by the forth week, the hair of the treated side was
visually lighter and felt smoother to touch. Digital pictures at
different magnifications were taken throughout the treatment
period, using Hi-Scope. These pictures clearly demonstrate the
reduced size and thickness of the hair shafts at the treated side.
An example of such pictures is shown in FIG. 13, demonstrating the
difference in hair shaft thickness and density at four weeks of
treatment. Since both sides of the face were shaved at the same
time, and pictures of both sides were taken at the same time, the
difference in length of the facial hair indicates slower growth
rate at the treated area.
[0077] FIG. 14 shows a computerized image analysis of the facial
hair length, thickness and total area, following four weeks of
soymilk treatment. All images were analyzed with Image Pro Plus 3.0
software (Media Cybernetics, Silver Spring, Md.). Data are
presented as average of 180 hair shafts of each side of the face,
with standard deviation (SigmaPlot.RTM. 5.0, SPSS Science, Chicago,
Ill.). Statistical analysis was performed using SigmaStat.RTM. 2.0
(SPSS Science) software, demonstrating a statistical significant
difference in all measured parameters, following soymilk
treatment.
Example 9
[0078] Soymilk Reduces Human Legs Hair length and Thickness
[0079] Hair was wax-depilated of two symmetrical areas of the
medial part of the legs, below the knee, in one individual. One leg
was treated daily, for four weeks, with soymilk. Visual
observations indicate slower hair growth on the treated site. Hair
shafts were reduced in number and were shorter and thinner than
those of the untreated site, as shown in FIG. 15. These
observations further confirm the effect of soymilk on hair growth.
Examples 8 and 9 together confirm that the effect of soymilk on
human hair growth is not related to the method of hair removal or
to the body part being treated.
Example 10
[0080] Soymilk Formulations Enriched With Isoflavones are Preferred
to Soymilk Formulations in Reducing Hair Growth and
Pigmentation.
[0081] The experiments described in Example 4 were repeated, using
two formulations described in Table 2 above, Soymilk Essence 23
which is a soymilk-based formulation, and soymilk Essence 30 which
is identical to Soymilk Essence 23 except the addition of 5% of a
0.1% isoflavones extract. As shown in FIG. 16, mice treated with
Soymilk essence 23 show reduced hair growth and nicer hair
appearance. This effect was more pronounced with the use of soymilk
Essence 30, demonstrating that isoflavone-enriched soymilk
formulations are superior to soymilk formulations in reducing
hair-growth. FIG. 17 shows histological skin sections of the
treated mice, at day 15 of the treatment. The hair shafts
documented in these sections clearly demonstrate the reduction in
hair shaft dimensions, the reduced level of pigmentation within the
hair shaft, and the increased smoothness of the hair shaft
following the Soymilk Essence treatments.
Example 11
[0082] Soymilk Formulations Enriched With Isoflavones are Preferred
to Isoflavone Formulations Which are Effective in Reducing Hair
Growth and Pigmentation.
[0083] The experiments described in Example 4 were repeated, using
formulations described in Table 2 above, of soymilk essence with or
without increasing concentrations of isoflavones. These Soy Essence
formulations were compare to similar formulations, where the
soymilk component only was replaced with water. These three sets of
formulations (Soy Essence, isoflavones, Soy Essence containing
additional isoflavones) were prepared to test the possibility that
isoflavones might be sufficient for the effect observed on hair
growth. FIG. 18 shows the C57Bl/6 mouse hair following three weeks
of topical treatment, as described in example 4. Both untreated
control mice and placebo treated mice have long and less "ordered"
hair. Soymilk Essence 23 reduces hair growth and leads to a nicer
appearance, as described earlier in this application. Soymilk
Essence formulations containing 1, 5 and 10% of a 0.1% isoflavones
containing extract result in a superior effect on hair growth.
However, formulations containing isoflavones but no soymilk
demonstrate milder, and not as superior effect as when combined
with soymilk. This example demonstrates that soymilk formulations
containing isoflavone could reduce hair growth. This example
further demonstrates that soymilk formulations containing
isoflavones reduce hair growth to a higher degree than formulations
containing isoflavones alone.
Example 12
[0084] Soy Essence Formulations Affect Human Hair Growth
[0085] The efficacy and irritancy potential of Soymilk Essence 23
and 30 compared to a placebo formulation were examined in a blinded
placebo-controlled four-week test with twelve pre-menopausal female
panelists ages 29 to 45 by evaluations by the study investigator,
self-assessment by panelists and Hi-scope image analysis. Panelists
signed an Informed Consent and were instructed about study
procedures and expectations and were asked to shave that night. At
the baseline visit the following day, two test lotions were
distributed to each panelist (Day 1), a placebo lotion and either
soymilk Essence 23 or 30. The lotions were randomly assigned to
either the right leg or left leg. The test lotions and placebo were
used on the respective legs for the duration of the study with no
other lotions used on the lower legs. Panelists were instructed to
apply the test lotions twice daily, morning and evenings and were
also instructed to try to refrain from shaving their lower legs
until after each weekly evaluation. On evaluation days, the
investigator visually inspected the panelists' legs for any
clinical signs of irritation and compared legs for hair growth
attenuation. Self-assessment questionnaires were completed by
panelists at each evaluation time point (Weeks 1, 2, 3 and 4). In
addition, Hi-scope images (2.5 cm in diameter for each image,
KH-2400R, Hirox) were obtained at each time point using a MX-MACROZ
lens (Hirox).
[0086] No panelists dropped from the study for any product-related
reason. No signs of irritation were seen in any of the study
panelists at any time point, nor was any irritation reported when
self-assessed by panelists at any time point during the study.
[0087] For the purpose of hair counts all hair, including
"stubble", were counted in the given 2.5 cm field for each panelist
at each time point. Results showed a decrease in lower leg hair
counts by week 5 for Soymilk Essence 30 and by week 4 for Soymilk
Essence 23. The placebo treated legs did not show a change in mean
leg hair counts throughout the study although the standard
deviations were large. The growth rate was calculated by dividing
the length of time (in days) since the panelist last shaved, by the
average length of leg hairs for that panelist, which was calculated
from the hi-scope images using Image Pro Plus analysis for each
panelist at each test site. The results are documented in Table 3,
demonstrating that both Soymilk Essences 23 and 30 treatments
resulted in reduced hair growth rate compared to placebo.
3TABLE 3 Mean leg hair growth rates following Soymilk Essence or
placebo treatment Soymilk Soymilk Week Location Essence 30 Essence
23 Placebo 0 Upper 0.211 (.+-.0.07) 0.184 (.+-.0.07) 0.243
(.+-.0.07) (baseline) Lower 0.248 (.+-.0.08) 0.191 (.+-.0.06) 0.235
(.+-.0.09) 1 Upper 0.216 (.+-.0.05) 0.153 (.+-.0.03) 0.211
(.+-.0.06) Lower 0.178 (.+-.0.08) 0.213 (.+-.0.03) 0.188 (.+-.0.07)
2 Upper 0.232 (.+-.0.11) 0.181 (.+-.0.04) 0.221 (.+-.0.07) Lower
0.236 (.+-.0.12) 0.195 (.+-.0.07) 0.217 (.+-.0.09) 3 Upper 0.241
(.+-.0.06) 0.185 (.+-.0.09) 0.285 (.+-.0.13) Lower 0.213 (.+-.0.10)
0.147 (.+-.0.04) 0.253 (.+-.0.12) 4 Upper 0.234 (.+-.0.07) 0.209
(.+-.0.11) 0.211 (.+-.0.09) Lower 0.194 (.+-.0.05) 0.208 (.+-.0.07)
0.220 (.+-.0.04)
[0088] Results from panelists' self-assessment questionnaires
showed that panelists felt that the test lotions attenuated hair
growth and softened the feel of leg hair, compared to the placebo
lotion. Panelists felt that the hair felt less coarse and less
stubbley. The majority of the panelists believed that the test
lotions were attenuating leg hair growth or altering the texture of
the hair so that it felt smoother and less coarse. Hi Scope
analysis further demonstrated that the hair re-growth following
treatment with Soymilk Essence 23 or 30 seemed to be growing in the
same direction and was more uniform in shape, texture and
appearance. In contrast, the hair that re-grew on the
placebo-treated legs grew in different directions, differing in
length, angle of growth and thickness.
[0089] This Example clearly demonstrate the effect of soymilk
formulations in delaying and reducing hair growth, and enabling the
growth of softer, less coarse and more managed and
directionally-organized hair.
* * * * *