U.S. patent application number 13/921123 was filed with the patent office on 2014-02-13 for polyphenol/theanine compositions.
The applicant listed for this patent is Jack F. Bukowski, Jeffrey Walters. Invention is credited to Jack F. Bukowski, Jeffrey Walters.
Application Number | 20140045928 13/921123 |
Document ID | / |
Family ID | 39201267 |
Filed Date | 2014-02-13 |
United States Patent
Application |
20140045928 |
Kind Code |
A1 |
Walters; Jeffrey ; et
al. |
February 13, 2014 |
POLYPHENOL/THEANINE COMPOSITIONS
Abstract
Disclosed herein are compositions comprising tea derived
components which are useful in reducing the incidence of cold
and/or flu in a subject. Specifically exemplified herein are
compositions comprising predetermined amounts of L-theanine and
EGCG, and methods of using same.
Inventors: |
Walters; Jeffrey; (Ft.
Pierce, FL) ; Bukowski; Jack F.; (Marlboro,
FL) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Walters; Jeffrey
Bukowski; Jack F. |
Ft. Pierce
Marlboro |
FL
FL |
US
US |
|
|
Family ID: |
39201267 |
Appl. No.: |
13/921123 |
Filed: |
June 18, 2013 |
Related U.S. Patent Documents
|
|
|
|
|
|
Application
Number |
Filing Date |
Patent Number |
|
|
11672542 |
Feb 8, 2007 |
8465782 |
|
|
13921123 |
|
|
|
|
60826451 |
Sep 21, 2006 |
|
|
|
Current U.S.
Class: |
514/456 |
Current CPC
Class: |
A61P 31/12 20180101;
A61K 36/82 20130101; A61K 31/353 20130101; A61P 37/00 20180101;
A61K 31/198 20130101; A61K 9/48 20130101; A61P 31/16 20180101; A61P
31/00 20180101; A61P 11/00 20180101; A61P 43/00 20180101; A61P
37/04 20180101 |
Class at
Publication: |
514/456 |
International
Class: |
A61K 31/353 20060101
A61K031/353; A61K 31/198 20060101 A61K031/198 |
Claims
1. A method of reducing the incidence of colds and/or flu in a
subject comprising orally administering a therapeutically effective
amount of a composition comprising an admixture of an L-theanine
composition comprising at least 10-100 percent L-theanine and a tea
polyphenol composition comprising at least 10-100 percent EGCG,
wherein said composition is administered according to a dosage
amounting to at least 25 mg of L-theanine and at least 25 mg of
EGCG per day.
2. The method of claim 1, wherein said composition is administered
according to a dosage amounting to at least 100 mg of L-theanine
per day.
3. The method of claim 1, wherein said composition is administered
according to a dosage amounting to at least 150 mg L-theanine and
at least 125 mg of EGCG per day.
4. The method of claim 1 wherein said composition is administered
according to a dosage amounting to between 150 to 600 mg of
L-theanine and between 150 to 600 mg of EGCG per day.
5. The method of claim 1, wherein said composition comprises at
least 200 mg of an admixture of an L-theanine composition and a tea
polyphenol composition disposed in a capsule, wherein said
L-theanine composition comprises at least 80 percent L-theanine and
said decaffeinated tea polyphenol composition comprises at least 40
percent EGCG.
6. The method of claim 5, wherein said capsule comprises 300 mg of
said admixture.
7. The method of claim 5, wherein said L-theanine composition
comprises at least 90 percent L-theanine.
8. The method of claim 5, wherein said polyphenol composition
comprises at least 45 percent EGCG.
9. The method of claim 5, wherein said composition is administered
as two of said capsules per day.
10. The method of claim 9, wherein said capsules each contain at
least 300 mg of said admixture.
11. A method of reducing the severity of cold and/or flu symptoms
in a subject experiencing cold and/or flu, said method comprising
orally administering a therapeutically effective amount of a
composition comprising an admixture of an L-theanine composition
comprising 10-100 percent L-theanine and a tea polyphenol
composition comprising 10-100 percent EGCG, wherein said
composition is administered according to a dosage amounting to at
least 25 mg of L-theanine and at least 25 mg of EGCG per day.
12. The method of claim 11, wherein said composition is
administered according to a dosage amounting to at least 100 mg of
L-theanine per day.
13. The method of claim 11, wherein said composition is
administered according to a dosage amounting to at least 150 mg
L-theanine and at least 125 mg of EGCG per day.
14. The method of claim 11 wherein said composition is administered
according to a dosage amounting to between 150 to 600 mg of
L-theanine and between 150 to 600 mg of EGCG per day.
15. The method of claim 11, wherein said composition comprises at
least 200 mg of an admixture of an L-theanine composition and a tea
polyphenol composition disposed in a capsule, wherein said
L-theanine composition comprises at least 80 percent L-theanine and
said decaffeinated tea polyphenol composition comprises at least 40
percent EGCG.
16. The method of claim 15, wherein said capsule comprises 300 mg
of said admixture.
17. The method of claim 15, wherein said L-theanine composition
comprises at least 90 percent L-theanine.
18. The method of claim 15, wherein said polyphenol composition
comprises at least 45 percent EGCG.
19. The method of claim 15, wherein said composition is
administered as two of said capsules per day.
20. The method of claim 19, wherein said capsules each contain at
least 300 mg of said admixture.
21. A capsule or tablet comprising at least 50 mg of an admixture
of an L-theanine composition comprising at least 10-100 percent
L-theanine and a tea polyphenol composition comprising at least
10-100 percent EGCG.
22. The capsule or table of claim 21, wherein said capsule or
tablet comprises at least 200 mg of an admixture of an L-theanine
composition and a tea polyphenol composition, wherein said
L-theanine composition comprises at least 80 percent L-theanine and
said decaffeinated tea polyphenol composition comprises at least 40
percent EGCG.
23. The capsule or tablet of claim 22, wherein said capsule or
tablet comprises 300 mg of said admixture.
24. The capsule or tablet of claim 22, wherein said L-theanine
composition comprises at least 90 percent L-theanine or at least 95
percent L-theanine.
25. The capsule or tablet of claim 22, wherein said polyphenol
composition comprises at least 45 percent EGCG or at least 50
percent EGCG.
26. The capsule or tablet of claim 21, wherein said capsule or
tablet comprises at least 50 mg of L-theanine and at least 50 mg of
EGCG.
26. A composition comprising an admixture of an L-theanine
composition and a decaffeinated tea polyphenol composition, wherein
said L-theanine composition comprises 10-100 percent L-theanine and
said tea polyphenol composition comprises 10-100 percent EGCG,
wherein said L-theanine composition and tea polyphenol composition
are present in a ratio of 2:1, 1.9:1, 1.8:1, 1.7:1, 1.6:1, 1.5:1,
1.4:1, 1.3:1, 1.2:1, 1:1, 1:1.1, 1:1.2, 1:1.3, 1:1.4, 1:1.5, 1:1.6,
1:1.7, 1:1.8, 1:1.9, 1:2, or 1:3 respective to each other.
27. A method of enhancing immune function by .gamma..delta. T cells
in a subject comprising orally administering a therapeutically
effective amount of a composition comprising L-theanine and EGCG,
wherein said composition is administered according to a dosage
amounting to at least 25 mg of L-theanine and at least 25 mg of
EGCG per day.
28. The method of claim 27 wherein said composition comprises an
admixture of an L-theanine composition comprising at least 10-100
percent L-theanine and a tea polyphenol composition comprising at
least 10-100 percent EGCG
29. The method of claim 27, wherein said composition is
administered according to a dosage amounting to at least 100 mg of
L-theanine per day.
30. The method of claim 27, wherein said composition is
administered according to a dosage amounting to at least 150 mg
L-theanine and at least 125 mg of EGCG per day.
31. The method of claim 27 wherein said composition is administered
according to a dosage amounting to between 150 to 600 mg of
L-theanine and between 150 to 600 mg of EGCG per day.
32. The method of claim 27, wherein said composition comprises at
least 50 mg of an admixture of an L-theanine composition and a
decaffeinated tea polyphenol composition disposed in a capsule,
wherein said L-theanine composition comprises at least 80 percent
L-theanine and said tea polyphenol composition comprises at least
40 percent EGCG.
33. The method of claim 1, wherein said composition is administered
orally in a beverage or food product.
34. The method of claim 1, wherein said composition comprises
between 10 percent to 80 percent L-theanine, by weight, and between
10 percent to 80 percent EGCG, by weight.
35. The method of claim 33, wherein said composition comprises
between 20 percent to 40 percent L-theanine, by weight, and between
20 percent to 40 percent EGCG, by weight.
36. The composition of claim 26, wherein said composition comprises
between 10 percent to 80 percent L-theanine, by weight, and between
10 percent to 80 percent EGCG, by weight.
37. The composition of claim 36, wherein said composition comprises
between 20 percent to 40 percent L-theanine, by weight, and between
20 percent to 40 percent EGCG, by weight.
37. A method of reducing the incidence of cold and/or flu
comprising orally administering the composition of claim 35 in
therapeutically effective daily dosages over the course of at least
12 weeks, at least 18 weeks, 24, weeks, 30 weeks, 36, weeks, 42
weeks, 48 weeks, and 52 weeks.
38. A method of reducing the length of cold and/or flu symptoms in
a subject experiencing cold and/or flu, said method comprising
orally administering a therapeutically effective amount of a
composition comprising an admixture of an L-theanine composition
comprising 10-100 percent L-theanine and a tea polyphenol
composition comprising 10-100 percent EGCG, wherein said
composition is administered according to a dosage amounting to at
least 25 mg of L-theanine and at least 25 mg of EGCG per day.
39. The method of claim 38, wherein said composition is
administered according to a dosage amounting to at least 125 mg of
L-theanine and at least 125 mg of EGCG per day.
40. An article of manufacture comprising between 25 ml to 5000 ml
of a beverage comprising water, an amount of an EGCG composition,
an amount of a L-theanine composition and a container in which the
beverage is disposed, said article of manufacture comprising at
least 25 mg of L-theanine and at least 25 mg of EGCG.
41. The article of manufacture of claim 40, wherein said EGCG
composition comprises at least 40 percent EGCG composition and said
L-theanine composition comprises at least 80 percent
L-theanine.
42. The article of manufacture of claim 40, wherein the article of
manufacture comprises at least 50 mg of L-theanine and at least 50
mg of EGCG.
43. The article of manufacture of claim 40, wherein said article of
manufacture comprises at least 75 mg, 100 mg, 125 mg, 150 mg, 175
mg, 200 mg 225 mg or 250 mg of EGCG and at least 75 mg, 100 mg, 125
mg, 150 mg, 175 mg, 200 mg 225 mg or 250 mg mg of L-theanine.
44. The article of manufacture of claim 40, further comprising
instructions to drink said beverage to reduce the incidence of
colds and/or flu, or to boost immune function.
45. A method of reducing incidence of cold and/or flu in a subject
comprising providing the article of manufacture of claim 40 and
instructing said subject to drink said beverage in said article of
manufacture at a dosage of at least once daily.
46. A method of boosting immune function in a subject comprising
providing the article of manufacture of claim 40 and instructing
said subject to drink said beverage in said article of manufacture
at a dosage of at least once daily.
47. An article of manufacture comprising at least 25 mg of a food
product, an amount of an EGCG composition, an amount of a
L-theanine composition and a container in which said food product
is disposed, said article of manufacture comprising at least 25 mg
of L-theanine and at least 25 mg of EGCG.
48. A method of reducing incidence of cold and/or flu in a subject
comprising providing the article of manufacture of claim 47 and
instructing said subject to eat said food product in said article
of manufacture at a dosage of at least once daily.
48. A method of boosting immune function in a subject comprising
providing the article of manufacture of claim 47 and instructing
said subject to eat said food product in said article of
manufacture at a dosage of at least once daily.
49. A method of reducing the severity of cold and/or flu symptoms
in a subject experiencing cold and/or flu, comprising providing the
article of manufacture of claim 40 and instructing said subject to
drink said beverage in said article of manufacture at a dosage of
at least once daily.
50. A method of reducing the severity of cold and/or flu symptoms
in a subject experiencing cold and/or flu providing the article of
manufacture of claim 47 and instructing said subject to eat said
food product in said article of manufacture at a dosage of at least
once daily.
51. The method of claims 1, 5, 11, 15, 28 and 32 wherein said tea
polyphenol composition is decaffeinated.
52. The capsule of claims 21 and 22 wherein said tea polyphenol
composition is decaffeinated.
53. The composition of claim 26, wherein said tea polyphenol
composition is decaffeinated.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] This application is related to U.S. Provisional Patent
Application 60/826,451; filed Sep. 21, 2006 to which priority is
claimed under 35 USC .sctn.119. This application is a continuation
of U.S. application Ser. No. 11/672,542 filed Feb. 8, 2007 and U.S.
application Ser. No. 11/672,547 filed Feb. 8, 2007. The foregoing
applications incorporated herein.
INTRODUCTION
[0002] For centuries, tea beverage has been linked to good health.
Most studies have been observational, showing benefit in
cardiovascular, anti-aging, neurodegenerative, anti-cancer, and
bone areas [1-3]. There are numerous other studies consistent with
no health benefits [4, 5]. These studies, both positive and
negative, are fraught with confounding variables that are inherent
in observational studies employing free-living human beings as
subjects. Such studies are further complicated by the nature of tea
beverage. There are hundreds of varieties of the tea species,
Camellia sinensis, and numerous ways to process tea that can lead
to different components in a cup of tea [6, 7]. Storing and brewing
methods lead to further variability that is difficult to
standardize. Another obvious source of conflict is that there is no
general agreement on what quantity constitutes a cup of tea. Thus,
it is not surprising that the results of many published trials
conflict with one another.
[0003] Whereas most clinical tea research has focused on cardiac
and cancer outcomes, there are no available clinical human data on
prevention of cold and flu symptoms.
[0004] These symptoms are the most common afflictions in humans,
resulting in misery, loss of productivity, and absence from work
and school [7]. Further, recent studies that call in to question
the efficacy of zinc-based nasal sprays and lozenges [8], and
Echinacea [9] make new research into cold and flu more important.
Undoubtedly, there has been a long unmet need for a cold and flu
preventive therapy that is safe and effective. Any advancement in
developing a cold and/or flu preventive therapeutic product, either
a pharmaceutical type product or natural product, would provide a
dramatic benefit to the millions of people who suffer from colds
and flu each year.
SUMMARY
[0005] The subject invention is based on the inventors surprising
discovery that administration of composition containing
standardized amounts of certain tea components, namely L-theanine
and EGCG, enhances systemic immunity, and reduces the risk of onset
of cold and flu symptoms in humans. Through a randomized,
double-blind, placebo-controlled interventional study using
compositions containing these key tea components (Camellia sinensis
composition; CSC) with a primary endpoint of reduction in the
number of subjects who developed cold and flu symptoms, the
inventors have discovered that L-theanine and EGCG work together to
generate surprising, unexpected, results of reducing the risk of
cold and flu, and reducing the severity of symptoms of subject who
contract colds and/or flu. The inventors present herein results
showing that CSC reduces by about one third the number of subjects
who develop cold and flu symptoms, while enhancing .gamma..delta. T
cell proliferation and IFN-.gamma. secretion in response to
microbial antigen.
BRIEF DESCRIPTION OF THE DRAWINGS
[0006] FIG. 1 Shows a graph representing the percentage of 56 CSC
subjects and 55 placebo subjects who had at least one symptom
during the 12-week study period.
[0007] FIG. 2. Mean number of days each subject had cold and flu
symptoms over the 12 week study period.
[0008] FIG. 3. Percentage of subjects seeking medical
treatment.
[0009] FIG. 4. Percentage of subject having individual symptoms.
One of 56 (1.79%) CSF subjects had diarrhea; 3/55 (5.45%) placebo
subjects had diarrhea (67.1% difference). Three of 56 (5.35%) CSF
subjects had fever; 7/55 (12.7%) placebo subjects had fever (57.8%
difference). Sixteen of 56 (28.6%) CSF subjects had sore throat;
25/55 (45.4%) placebo subjects had sore throat (37.0% difference).
Fourteen of 56 (23.2%) CSF subjects had headache; 15/55 (27.2%)
placebo subjects had headache (1.47% difference). Two of 56 (3.57%)
CSF subjects had nausea; 4/55 (7.27%) placebo subjects had nausea
(50.9% difference). Eleven of 56 (19.6%) CSF subjects had cough;
16/55 (29.1%) placebo subjects had cough (32.6% difference). Twenty
of 56 (35.7%) CSF subject had runny nose; (29/56) (51.8%) placebo
subjects had runny nose (31.1% difference). Nineteen of 56 (33.9%)
CSF subjects had stuffy nose; 21/55 (38.2%) placebo subjects had
stuffy nose (11.2% difference).
DETAILED DESCRIPTION
[0010] L-theanine, abundant in tea, is catabolized in the body to
yield ethylamine, a gamma delta T cell antigen, which appears in
the blood of rats given L-theanine, and has been shown to appear in
the urine of tea drinkers. Tea extraction procedures normally
exclude amino acids, including L-theanine. Compositions taught
herein are unique in that they intentionally combine L-theanine and
an EGCG in enriched amounts.
[0011] The EGCG composition is procured from tea and comprises
between 10-100 percent EGCG, by weight. In optimal embodiments, the
EGCG comprises at least 40 to 50 percent EGCG, by weight. It is
known that EGCG procured from tea can contain caffeine. U.S. Pat.
No. 7,012,149 is cited for background on procuring EGCG containing
composition. Also commercially available products such as
SUNPHENON.RTM. line of products from Taiyo International,
Minneapolis, Minn. offer EGCG compositions may be used in
accordance with the teachings herein.
[0012] In preferred embodiments caffeine is reduced or eliminated
from the EGCG composition either before or after procurement of the
EGCG composition. The inventors have found that reducing the
caffeine amount provides a novel composition with increased
compliance. Caffeine intake can cause a number of adverse side
effects, including, but not limited to, excitability in children,
constipation, nervousness, dizziness, hypertension, and arythmias.
For certain embodiments, the inventors have realized that
compositions may be particularly marketed to schools for use in
beverages and food served to students. Schools can be a primary
source epicenter of virus origination and distribution. The
administration of compositions taught herein will serve to reduce
or slow the spread of microbial infections. A composition
containing caffeine would not be appropriate for such
application.
[0013] The procurement of L-theanine is separate from tea
polyphenols. Accordingly, an L-theanine composition is obtained
from tea where the L-theanine composition comprises between 10-100
percent L-theanine. This L-theanine composition may be admixed with
the EGCG containing tea polyphenol composition. See U.S. Pat. No.
6,831,103 for background on procurement of L-theanine.
[0014] Theanine may be a glutamic acid derivative
(.gamma.-glutamylethylamide), which is an amino acid component
naturally contained largely in tea-leaves. Methods for preparing
theanine used in the present invention may include, for instance, a
method of extracting from tea-leaves; an organic synthesis method
[Chem. Pharm. Bull, 19(7), 1301-1307 (1971)]; a method of treating
a mixture of glutamine and ethylamine with glutaminase (Japanese
Unexamined Patent Publication No. Hei 7-55154); a method comprising
culturing cultured cells of tea in a medium containing ethylamine,
thereby achieving growth promotion of the cultured cells while
increasing the cumulative amount of theanine in the cultured cells
(Japanese Patent Laid-Open No. Hei 5-123166); modification methods
in which ethylamine is substituted by an ethylamine derivative such
as ethylamine hydrochloride in the methods disclosed in Japanese
Unexamined Patent Publication No. Hei 7-55154 or Japanese Patent
Laid-Open No. Hei 5-123166; and the like, and any of the methods
may be used. The above-mentioned "tea-leaves" include green
tea-leaves, oolong tea-leaves, black tea-leaves, and the like.
[0015] Theanine can be used as any of L-theanine, D-theanine and
DL-theanine. Among them, the L-form is preferred in the present
invention, because the L-form is approved as a food additive, and
is economically utilizable. In addition, theanine used in the
present invention may be in any forms, such as purified products,
crudely purified products and extracts. Also, a commercially
available product [SUNTHEANINE (registered trade mark),
manufactured by Taiyo Kagaku Co., Ltd.] may be used.
[0016] According to one embodiment, the invention pertains to a
method of reducing the incidence of colds and/or flu comprising
orally administering a composition comprising L-theanine and EGCG,
wherein said composition is administered according to a dosage
amounting to at least 25 mg of L-theanine and 25 mg EGCG per day.
In a specific embodiment at least 200 mg of L-theanine is
administered per day. In a further embodiment at least 150 mg of
L-theanine is co-administered with at least 125 mg of EGCG per day.
In a particular embodiment, between 180 to 220 mg of L-theanine and
between 180 to 220 mg of EGCG is administered per day. In
alternative embodiments, the daily amount of EGCG that is used to
combine with L-theanine is from about 200 mg to 450 mg. In specific
embodiment EGCG is provided at up to 800 mg of 45 percent EGCG
composition.
[0017] According to another embodiment, the invention pertains to a
capsule or tablet or liquid suspension comprising a combination of
L-theanine and EGCG, wherein the capsule or table comprises between
10 percent to 100 percent, by weight, L-theanine and between 10
percent to 100 percent, by weight EGCG. In a specific embodiment,
the capsule or tablet comprises between 15 percent to 65 percent
L-theanine and 15 percent to 65 percent EGCG. In a further
embodiment, the capsule or tablet or suspension comprises between
30 percent to 60 percent, by weight, tea polyphenols. In a specific
embodiment, the capsule or table or suspension is decaffeinated. In
an alternative embodiment, the capsule or tablet may contain
sulphoraphane (from 5-50 mg per day).
[0018] The tablet or capsule may be, but is not limited to, from 10
to 500 mg total weight. In specific embodiments, the capsules are
25, 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350,
375, 400, 425, 450, 475, or 500 mg, total individual weight.
[0019] According to another embodiment, the invention pertains to
reducing the severity of symptoms of cold and/or flu comprising
orally administering a composition comprising L-theanine and EGCG,
wherein said composition is administered according to a dosage
amounting to at least 25 mg of L-theanine a day. In a specific
embodiment at least 100 mg of L-theanine is administered per day.
In a further embodiment at least 115 mg of L-theanine is
co-administered with at least 100 mg of EGCG per day. In a
particular embodiment, between 150 to 600 mg. of L-theanine and
between 150 to 600 mg of EGCG is administered per day.
[0020] According to another embodiment, the invention pertains to
reducing the severity of symptoms of cold and/or flu comprising
orally administering a beverage comprising water, flavoring, EGCG
composition, and L-theanine, wherein said beverage is administered
according to a dosage amounting to at least 25 mg of L-theanine a
day and at least 25 mg of EGCG composition a day. In a specific
embodiment at least 100 mg of L-theanine is administered per day.
In a further embodiment at least 120 mg of L-theanine is
co-administered with at least 125 mg of EGCG per day. In a specific
embodiment, between 150 to 600 mg. of L-theanine and between 150 to
600 mg of EGCG is administered per day.
[0021] According to another embodiment, the subject invention
pertains to a method of enhancing immune function by .gamma..delta.
T cells in a subject comprising orally administering a
therapeutically effective amount of a composition comprising
L-theanine and EGCG, wherein said composition is administered
according to a dosage amounting to at least 25 mg of L-theanine and
at least 25 mg of EGCG per day. In a more specific embodiment,
composition comprises an admixture of an L-theanine composition
comprising at least 10-100 percent L-theanine and a decaffeinated
tea polyphenol composition comprising at least 10-100 percent
EGCG
[0022] For certain method embodiments, the dosage of L-theanine is
at least 25 mg per day. The dosage of L-theanine is typically
between 25-600 mg of L-theanine a day, but may be higher than 600
mg. In certain embodiments, the dosage of L-theanine is or about
25, 50, 75, 100, 125, 150, 160, 170, 180, 190, 200, 210, 220, 230,
240, 260, 280, 300, 320, 340, 360, 380, 400, 420, 440, 460, 480,
500, 520, 540, 560, 580 or 600 mg of L-theanine a day. In a
specific embodiment, the dosage is such that about 200 mg
L-theanine is provided per day. The dosage of EGCG is at least 25
mg per day. The dosage of EGCG is typically between 25-600 mg of
EGCG a day. In certain embodiments, the dosage of L-theanine is or
about 25, 50, 75, 100, 125, 150, 160, 170, 180, 190, 200, 210, 220,
230, 240, 260, 280, 300, 320, 340, 360, 380, 400, 420, 440, 460,
480, 500, 520, 540, 560, 580 or 600 mg of L-theanine a day. In a
specific embodiment, the dosage is such that about 200 mg
L-theanine is provided per day.
[0023] In another embodiment, the subject invention is directed a
composition comprising an admixture of an L-theanine composition
and a decaffeinated tea polyphenol composition, wherein said
L-theanine composition comprises at least 50 percent L-theanine and
said decaffeinated tea polyphenol composition comprises at least 30
percent EGCG, wherein said L-theanine composition and tea
polyphenol composition are present in a ratio of 2:1, 1.9:1, 1.8:1,
1.7:1, 1.6:1, 1.5:1, 1.4:1, 1.3:1, 1.2:1, 1:1, 1:1.1, 1:1.2, 1:1.3,
1:1.4, 1:1.5, 1:1.6, 1:1.7, 1:1.8, 1:1.9, 1:2, or 1:3 respective to
each other. In a related embodiment, the composition is used as a
food additive. In a specific embodiment, the composition comprises
between 10 percent to 80 percent, by weight, L-theanine and between
10 percent to 80 percent, by weight EGCG. In a specific embodiment,
the capsule or tablet comprises between 15 percent to 65 percent
L-theanine and 15 percent to 65 percent EGCG. In a more specific
embodiment, the composition comprises between 20 percent to 40
percent L-theanine and 20 percent to 40 percent EGCG.
[0024] In yet another embodiment, the subject invention pertains to
a food product comprising a therapeutically effective amount of
L-theanine and EGCG. In a specific embodiment, the invention
relates to a food product comprising at least 25 mg of a L-theanine
and tea polyphenol composition, such as, but not limited to, the
composition described in the preceding paragraph. Examples of food
products may include, but are not limited to, energy bars, sauces,
salad dressings, frozen dinners, chips, canned soups, yogurt,
cereals, bread, flour and grains. In an alternative but related
embodiment, the subject invention pertains to a packaged food
product having a total weight of between 25 mg to 50 kg. The
packaged food product comprises at least 0.1 percent by weight
L-theanine and at least 0.1 percent by weight EGCG. In a specific
embodiment, the food product comprises between about 0.1-10
percent, by weight, L-theanine and between about 0.1 to 10 percent,
by weight, EGCG. In other embodiments, the amount of L-theanine and
EGCG are at least 25 mg per serving.
[0025] Not to be bound by any particular mechanistic theory, it is
the belief of the inventors that the unexpected antiviral effects
of certain embodiments is due to a one-way synergy between
L-theanine and EGCG. In particular, the inventors believe that
there is one-way synergy, in two dimensions: in the first
dimension, the L-theanine catabolite ethylamine binds to
.gamma..delta. T cells to prime them for activation (proliferation
and IFN-.gamma. secretion) by cold and flu viruses and IL-12. EGCG
serves to enhance the secretion of IL-12, which is essential for
.gamma..delta. T cell activation and protection against cold and
flu (refs 18 and 20). The IFN-.gamma. is what acts against viruses.
However, as an unwanted side effect, this IFN-.gamma. acts on
macrophages, causing the release of oxygen free radicals, and
cytokines such as IL-1 and TNF-.alpha. that are toxic to cells of
the lung, nose, and throat. These toxicities create symptoms of
cold and flu. In the second dimension, EGCG mimimizes this damage
created by L-theanine's action on .gamma..delta. T cells by
decreasing the synthesis of these toxins and also protecting the
lung, nose, and throat cells against the damaging effects of these
toxins. (REFS 20, 22-25)
[0026] In certain embodiments, the composition will generally be
used in an amount effective to achieve the intended purpose. For
use to treat or prevent the above diseases or disorders the CSC is
administered or applied in a therapeutically effective amount.
[0027] The amount of a L-theanine and EGCG, and/or CSC including
such compounds, that will be effective in the treatment of a
particular disorder or condition disclosed herein will depend on
the nature of the disorder or condition, and can be determined by
standard clinical techniques known in the art and by doctors
skilled in treating or preventing a particular disease or disorder.
In addition, in vitro or in vivo assays may optionally be employed
to help identify optimal dosage ranges. The amount of a compound of
the invention and/or pharmaceutical composition thereof
administered will, of course, be dependent on, among other factors,
the subject being treated, the weight of the subject, the severity
of the affliction, the manner of administration and the judgment of
the prescribing physician.
[0028] For example, the dosage may be delivered in a a single
administration, by multiple applications or controlled release. In
one embodiment, the compounds of the invention are delivered by
oral sustained release administration. Preferably, in this
embodiment, the compounds of the invention are administered twice
per day (more preferably, once per day). Dosing may be repeated
intermittently, may be provided alone or in combination with other
drugs and may continue as long as required for effective treatment
of the disease state or disorder.
[0029] Suitable dosage ranges for oral administration are generally
about 0.0001 mg to about 2000 mg of a compound of the invention per
kilogram body weight. In one embodiment, the dosage range is
between about 0.1 mg/kg to about 5 mg/kg. Dosage ranges may be
readily determined by methods known to the artisan of ordinary
skill. Effective doses may be extrapolated from dose-response
curves derived from in vitro or animal model test systems. Such
animal models and systems are well-known in the art.
[0030] The compounds of the invention are preferably assayed in
vitro and in vivo, for the desired therapeutic or prophylactic
activity, prior to use in humans. For example, in vitro assays can
be used to determine whether administration of a specific compound
of the invention or a combination of compounds of the invention is
preferred. The compounds of the invention may also be demonstrated
to be effective and safe using animal model systems.
[0031] Preferably, a therapeutically effective dose of a compound
of the invention and/or pharmaceutical composition thereof
described herein will provide therapeutic benefit without causing
substantial toxicity. Toxicity of compounds of the invention and/or
pharmaceutical compositions thereof may be determined using
standard pharmaceutical procedures and may be readily ascertained
by the skilled artisan. The dose ratio between toxic and
therapeutic effect is the therapeutic index. A compound of the
invention and/or pharmaceutical composition thereof will preferably
exhibit particularly high therapeutic indices in treating disease
and disorders. The dosage of a compound of the invention and/or
pharmaceutical composition thereof described herein will preferably
be within a range of circulating concentrations that include an
effective dose with little or no toxicity.
[0032] Unless defined otherwise, all technical and scientific terms
used herein have the same meaning as commonly understood by one of
ordinary skill in the art to which the present invention belongs.
Although any methods and materials similar or equivalent to those
described can be used in the practice or testing of the present
invention, the preferred methods and materials are now
described.
[0033] Those skilled in the art of medicinal chemistry and
pharmaceutical formulations will appreciate that other formulations
can be devised for appropriate oral, parenteral or other
administration. U.S. Pat. Nos. 6,821,532 and 7,157,493 are cited
for general background of pharmaceutical formulations.
Example 1
EGCG/L-Theanine Formulation Embodiment
[0034] According to a specific embodiment, the invention pertains
to a capsule comprising an admixture of or about 25-300 mg of an at
least 95 percent L-theanine containing composition and of or about
50-400 mg of a decaffeinated tea polyphenol composition containing
at least 45 percent EGCG. In a more specific embodiment, the
subject invention pertains to a capsule comprising 300 mg of an
admixture comprising 100 mg of a 98 percent L-theanine containing
composition and 200 mg of a 50 percent EGCG containing tea
polyphenol composition.
Example 2
Cold and/or Flu Treatment/Prevention Beverage
[0035] According to another embodiment, the invention pertains to
an article of manufacture comprising a beverage comprising water,
flavoring, EGCG composition, and L-theanine and a container in
which the beverage is disposed. In certain embodiments, the
container volume may be in the range of between 25 ml to 5000 ml.
The article of manufacture comprises at least 25 mg of L-theanine
and at least 25 mg of EGCG. In a preferred embodiment, the article
of manufacture comprises at least 50 mg of L-theanine and at least
50 mg of EGCG. In a specific embodiment, the article of manufacture
comprises at least 75 mg, 100 mg, 125 mg, 150 mg, 175 mg, 200 mg
225 mg or 250 mg of EGCG and at least 75 mg, 100 mg, 125 mg, 150
mg, 175 mg, 200 mg 225 mg or 250 mg mg of L-theanine. The EGCG
composition is procured from tea and comprises between 10-100
percent EGCG, by weight. In optimal embodiments, the EGCG comprises
at least 40 to 50 percent EGCG, by weight. It is known that EGCG
procured from tea can contain caffeine. In preferred embodiments
caffeine is reduced or eliminated from the EGCG composition either
before or after procurement of the EGCG composition. The
procurement of L-theanine is separate from tea polyphenols.
Accordingly, an L-theanine composition is obtained from tea where
the L-theanine composition comprises between 10-100 percent
L-theanine. This is admixed with the EGCG containing tea polyphenol
composition. Flavorings pertain to any substance that is used to
modify, enhance or mask flavors. Flavors can include but are not
limited to sweeteners, salts, flavor substances, acidulents.
Sweetener includes both natural and artificial sweeteners.
Sweeteners include, but are not limited to, sucralose, acesulfame
potassium, aspartame, saccharin, sucrose, glucose, fructose, high
fructose corn syrup, invert sugars, sugar alcohols including
sorbitol, mannitol and mixtures thereof. As used herein, the term
"acidulents" includes, but is not limited to, citric acid, lactic
acid, malic acid, sodium citrate, potassium citrate. As used
herein, the term "preservatives/antimicrobial agents" includes, but
is not limited to sodium benzoate, potassium benzoate, benzoic
acid, ethylparaben, methylparaben, propylparaben, sorbic acid.
[0036] Examples of beverages include, but are not limited to, fruit
juice mixed with the L-theanine/EGCG composition, a "smoothie"
(fruit juice and blended fruit) with L-theanine/EGCG composition
provided therein, fitness drinks, such as GATORADE.RTM.,
POWERADE.RTM., etc. with composition provided therein, tea drinks
with composition added therein, and sodas with composition added
therein.
Example 3
Double Blind Placebo Study
Materials and Methods
[0037] Subjects.
[0038] Healthy men (n=52) and women (n=72) between 21 and 70
(mean=29) years of age were recruited to participate in a 12-wk
randomized, double-blind placebo controlled parallel study.
Subjects were recruited from the University of Florida campus, and
the Gainesville, Fla. community, during January of 2006. The
University of Florida Institutional Review Board approved the study
protocol, and informed written consent was obtained from each
subject. Screening for the study occurred by telephone and/or
personal interviews. Exclusion criteria consisted of the following:
had not had a cold in the past two years; vegetarian diet;
steroids; chemotherapy or other immune suppressing therapy within
the last year; chronic antibiotics or other infectious disease
preventative; chronic illness; recent surgery or illness; pregnant
and/or lactating females. Also excluded were those who daily
consumed: greater than one cup (250 mL) of tea; an average of seven
or more servings of fruits and vegetables; herbal supplements,
vitamins other than a multivitamin or vitamin D, or osteoporosis
medicine or medications containing bisphosphonates, that are
activators of .gamma..delta. T cells [10]. The study was conducted
from January through May of 2006. Participants were in contact with
the enrolling research assistant by e-mail and telephone throughout
the study, and returned to fill out an exit questionnaire upon
study completion. Overall study compliance was monitored through
the exit questionnaire and by enumeration of remaining capsules in
returned bottles at the end of then study [11].
[0039] Study Protocol.
[0040] The study was conducted from January through May of 2006, to
coincide with normal cold and flu season. The CDC weekly report of
influenza activity in Florida was as follows: January-March
regional to widespread, March-April widespread to local activity,
April-May local activity to sporadic.
[0041] Subjects were randomly assigned to supplement and placebo
groups. Both subjects and investigators were blinded as to the
treatments. Human Health Biotech Institute (Naples, Fla.) provided
CSC and placebo capsules. The CSC was decaffeinated and comprised a
mixture of L-theanine (Suntheanine.RTM., standardized at 99%; Taiyo
International, Minneapolis, Minn.), and epigallocatechin gallate
(EGCG; Sunphenon.RTM., Taiyo International, Minneapolis, Minn.,
standardized at 50%). The placebo capsules contained
microcrystalline cellulose, dextrose, dicalcium phosphate,
magnesium stearate, silicon dioxide, FD&C red #40, yellow #6,
and blue #1. Each participant was given a bottle containing 180
capsules and was instructed to take 2 capsules every day (one in
the morning and one in the evening, preferably with meals) for 12
weeks.
[0042] Subjects were given an illness log to record any cold and
flu symptoms during the 12 week experimental period. Primary
outcomes were defined as the number of subjects experiencing any
symptoms, and the mean number of days they self-reported symptoms.
The symptoms assessed were: runny nose, congested or stuffy nose,
headache, cough, sore throat, fever, nausea/vomiting, and diarrhea.
Subjects were also asked to report if they sought medical treatment
and were prescribed any medications as a result of seeking
treatment. The exit questionnaire included questions: to determine
if subjects experienced any side effects and/or experienced any
changes in feelings of stress or anxiety, or took any additional
dietary supplements during the study. Finally, subjects were asked
to report whether they thought they had taken the active or the
placebo capsules.
[0043] Blood Collection.
[0044] Blood was obtained from fasting subjects on Days 0
(baseline), and 21. Blood was collected into one 10 mL sodium
heparin tubes for peripheral blood mononuclear cell (PBMC)
separation, and one 10 mL SST.TM. tube (Vacutainer, Becton
Dickinson, Franklin Lakes, N.J.) for serum. Tubes for PBMC were
maintained at room temperature (RT), while tubes for serum were
kept at 4.degree. C. All tubes were processed within 1 hr of blood
collection. Blood cell separation and culture procedures were
carried out under sterile conditions.
[0045] Serum Collection and Treatment.
[0046] Serum was removed from SST.TM. tubes after centrifugation
(1000 g, 10 min, 4.degree. C.) and frozen at -80.degree. C.
[0047] Blood Cell Separation.
[0048] Whole blood was diluted and placed on a gradient to separate
PBMC. Briefly, 7 mL of whole, anti-coagulated blood (RT) was
diluted 1:1 with 0.9% NaCl. Diluted blood (6 mL) was layered over 3
mL of Nycoprep 1-Step.TM. 1.077 (Axis-Shield, Oslo, Norway) and
centrifuged (800 g, 20 min, 20.degree. C.). The mononuclear cell
layer was removed, washed twice with 10% FBS-RPMI 1640 (Cellgro;
Mediatech, Herndon, Va.) complete (100 U/mL Penicillin; 100
.mu.g/mL Streptomycin; 0.25 .mu.g/mL Fungizone; 50 .mu.g/mL
Gentamycin; 2 mM 1-glutamine; 25 mM HEPES) by centrifugation (400
g, 10 min, 4.degree. C.). Individual cell pellets were resuspended
in 2 mL RPMI 1640 complete, without serum, and counted on a Z-2
Coulter Counter (3.8.mu. setting for the 100 .mu.m aperture tube,
Beckman Coulter, Inc., Fullerton, Calif.).
[0049] Culture of Peripheral Blood Mononuclear Cells (PBMC) for
.gamma..delta.T-Cell Expansion and Cytokine Production.
[0050] On Day 0, 1.0.times.10.sup.6 PBMC in 10% FBS-RPMI 1640
complete medium containing 50 .mu.M 2-ME, were seeded into each of
two wells of duplicate 24-well tissue culture plates (Costar,
Corning, N.Y.). Ethylamine (1 mM/mL, Sigma) in the same medium was
added to one set of wells on each plate, while the other set was
mock treated with 1 mL medium. The plate was incubated in a
humidified 5% CO.sub.2 atmosphere at 37.degree. C. On Day 1, cells
and supernatant fluids from one plate were harvested and
centrifuged (1500, 10 min, 4.degree. C.). The supernatant fluids
were removed and frozen at -80.degree. C. for cytokine analysis,
while the cell pellets were re-suspended in 0.1 mL PBS and 0.5 mL
RNAlater.TM. (Ambion, Austin, Tex.) and frozen at -80.degree. C. On
Day 3, 30 U/mL of recombinant human IL-2 (BD Biosciences, San
Diego, Calif.) were added to all wells of the remaining plate,
which was incubated until Day 10 when cells were harvested. Total
cell counts were obtained on a Coulter Counter (4.5.mu. setting)
and dead cells enumerated via trypan blue exclusion on a
hemocytometer, and cells processed for flow cytometry.
[0051] Flow Cytometry.
[0052] The ratio of .alpha..beta. and .gamma..delta.T-cells in PBMC
suspensions and cultures was determined by flow cytometry, using
cell surface markers for identification on Days 0 and 10 of
culture.
[0053] Antibodies:
[0054] Phycoerythrin (PE)-conjugated anti-human CD3.sup.+, and
fluorescein isothiocyanate (FITC)-conjugated anti-human T-cell
surface markers .alpha..beta.TCR and .gamma..delta.TCR
(eBioscience, San Diego, Calif.) were used to stain PBMC.
[0055] Staining/Fixation:
[0056] PBMC in staining buffer (PBS+0.1% NaN.sub.3+2% FBS) were
stained on ice in the dark for 30 min, with PE anti-human CD3.sup.+
antibody and one of the two FITC-conjugated TCR specific
antibodies. Cells were washed (PBS+0.1% NaN.sub.3) by
centrifugation (1000 g, 10 min, 20.degree. C.). Cells were fixed
with 1% paraformaldehyde in PBS+0.1% NaN.sub.3. Cells were analyzed
within 24 hours on a FACScan (Becton-Dickinson, San Jose, Calif.).
Data was analyzed using WinMDI Software (Scripps Institute, build
1301-19-2000). Data collected was from gated CD3.sup.+ cells
(events). Final data is reported as the mean percentage of cells
expressing the specific cell surface marker, .+-.the standard
deviation (SD). Cytokine Level Determination in Supernatants from
Cell Proliferation.
[0057] Levels of human IFN-.gamma. from PBMC culture supernatants
were quantified using enzyme linked-immunosorbent assay (ELISA)
kits, according to the manufacturer's directions (BD Biosciences
Pharmingen, San Diego, Calif.). Human cytokine standards provided
with each kit were used for a standard curve. Supernatants were
analyzed undiluted for IFN-.gamma..gamma.. Plates were read at 450
nm (with .lamda., correction of 570 nm) on a SPECTRAmax 340PC plate
reader (Molecular Devices, Menlo Park, Calif.). Cytokine
concentrations were calculated as the mean value obtained for
values within the range of the standard curve. The limits of
detection of the cytokine ELISA kits were: IFN.gamma.--300
pg/mL.
[0058] Statistical Analysis.
[0059] For analysis describing the percentage of subjects
experiencing symptoms, the two sample t test between percents was
used. For analysis describing symptom days, the independent groups
t test for means was used. Two-way ANOVA was used for
.gamma..delta. T cell proliferation studies and IFN-.gamma.
secretion.
Results
Study Subjects
[0060] One hundred and twenty-four subjects were enrolled in the
study. Six subjects withdrew from the study (three CSC, and three
placebo). Adverse events were mild, infrequent, and transient.
Bloating, gastric upset, dizziness, skin rash, and constipation
were reported, and were not different between experimental and
control groups. Two subjects taking CSC withdrew with mild skin
rashes. One subject said she thought the rash was related to a
seafood allergy, and the other thought it might be from the
capsule. Neither subject sought medical attention. Such food
allergies for green tea have been described previously, mostly in
tea factory workers, and the causative agent in green tea is EGCG
[12]. However, other reports in the literature describe EGCG as
beneficial for asthma and atopic dermatitis [13, 14].
[0061] One placebo subject withdrew due to an unrelated urinary
tract infection. One placebo individual withdrew because traveling
interfered. One placebo individual withdrew because he could not
return. One placebo subject withdrew because he just stopped taking
the capsules (Table 1). Five more subjects (two CSC and three
placebo) were excluded from all data because they were less than
70% compliant. Two more subjects (one CSC and one placebo) were
excluded when they reported in their logs that they were ill when
the study began, which was an exclusion criterion. Fifty-six
subjects in the supplement group and 55 subjects in the placebo
group completed the study with 70% or greater compliance, and their
data were included in all results and analyses.
Demographics
[0062] Average age, gender and BMI values did not differ between
experimental and control groups. Subjects were adequately blinded,
as there was no difference between CSC and placebo in the
percentage of subjects who guessed which treatment to which they
were assigned (Table 2).
[0063] Ingestion of Camellia sinensis Composition (CSC) Reduced the
Number of Subjects Having Cold and Flu Symptoms.
[0064] Throughout the three month study, subjects were asked to
keep an illness log to report daily cold and flu symptoms,
including fever, runny nose, stuffy nose, sore throat, cough,
headache, diarrhea, and nausea. Thirty-six of 55 (65.5%) placebo
subjects, but only 25 of 56 (44.6%) CSC subjects had at least one
symptom during the 12 week study. Thus, 31.9% fewer subjects taking
CSC experienced cold and flu symptoms as compared to subjects
taking placebo (P<0.029; FIG. 1).
[0065] Fifty-six CSC subjects had a total of 412 symptom days
(mean, 7.37+/-13.95), and 55 placebo subjects had a total of 595
symptom days (mean, 10.8+/-14.98). This represents a 30.2%
reduction in the number of symptoms days for CSC subjects as
compared to the placebo group (P<0.199; FIG. 2). Three of 56
(5.3%) CSC subjects, and six of 55 (12.7%) placebo subjects sought
medical treatment (58.3% difference; P<0.175; FIG. 3). We
analyzed the subjects in each group with regard to eight individual
symptoms. There were 67.1, 57.8, 50.9, 37, 32.6, 31.1, 11.2, 1.47
percent fewer CSC subjects experiencing diarrhea, fever, nausea,
sore throat, cough, runny nose, stuffy nose, and headache,
respectively, as compared to placebo subjects, but differences
within each individual symptom did not reach statistical
significance (FIG. 4 and Table 5).
[0066] Taken together, these results show that CSC is effective in
preventing cold and flu symptoms in this group of healthy
volunteers. The data suggest that CSC may also decrease the
duration of symptoms and the need for medical treatment.
Ingestion of CSC Enhanced Proliferation and IFN-.gamma. Secretion
by .gamma..delta. T Cells.
[0067] Our previous study showed that PBMC taken from subjects who
started drinking daily five to six cups of tea, containing
L-theanine, for as little as one week secreted significantly more
IFN-.gamma. in response to .gamma..delta. T cell antigens, as
compared to before [20]. To determine if ingestion of CSC,
containing L-theanine, could enhance IFN-.gamma. production, we
isolated PBMC taken from subjects three weeks after they started
taking CSC or placebo, and cultured them for 24 hours with media
alone, or with media containing ethylamine, which is a
.gamma..delta. T cell antigen that is a byproduct of L-theanine
catabolism (Table 3). It is important to note that only
.gamma..delta. T cells respond to ethylamine [19, 15], and that CSC
ingestion had no effect on IFN-.gamma. secretion or T cell
proliferation in response to a non-specific T cell mitogen,
phytohemagglutinin. Also, absolute numbers of .alpha..beta. T cells
did not change when cultured in media alone, or with ethylamine
(data not shown). PBMC from CSC placebo subjects secreted only 2.8
and 2.3 ng/mL IFN-.gamma., respectively. As expected, PBMC from
placebo subjects secreted more IFN-.gamma. in response to 5 mM
ethylamine as compared to media alone (13.6 ng/mL). However PBMC
from CSC subjects secreted 18.4 ng/mL IFN-.gamma. in response to
ethylamine. This response was 26% higher than that from placebo
PMBC (P=0.046).
[0068] We established separate 10-day PBMC cultures from the
subjects to assess by flow cytometry the proliferative responses of
.alpha..beta. and .gamma..delta. T cells (Table 4). In the presence
of ethylamine, .gamma..delta. T cells from CSC subjects expanded to
28% of CD3+ cells, as compared to 20.3% from placebo subjects
(P=0.017). Thus, CSC consumption was associated with a significant
increase in the capacity of .gamma..delta. T cells to secrete
IFN-.gamma. and to proliferate in response to antigenic
challenge.
Discussion
[0069] Fewer than 5% of Americans eat the nine servings per day of
fruits and vegetables suggested by the latest USDA guidelines
released in 2005 [16]. Only 20% of Americans drink any tea at all,
while those who do only drink an average of one to two cups per
day. Unfortunately, most health benefits from tea are associated
with higher amounts of daily consumption [1]. Tea is a vegetable
infusion, containing antioxidants and other beneficial nutrients
such as L-theanine. Numerous observational studies suggest that tea
drinking is beneficial to health, but negative studies have
introduced controversy surrounding its health benefits. Though any
two studies can yield different results, the conflict between
negative and positive studies is likely due in large measure to the
observational nature of the studies, and differences in tea
preparations (see introduction). We have conducted a randomized,
double blind placebo-controlled study using a Camellia sinensis
composition (CSC) with defined amounts of L-theanine and EGCG that
as closely as possible approximates the ingestion of 10 cups of
green tea per day. The strength of this design lies in elimination
of subject selection bias inherent in observational studies.
Another strength is the elimination of the variability that can be
associated with tea varieties and tea beverage preparations.
[0070] The results show that the ingestion of two capsules daily
decreased by about a third the number of subjects who experienced
cold and flu symptoms. Also seen were decreases in the number of
days subjects had symptoms, the number of subjects who had specific
cold and flu symptoms, and the number of subjects needing medical
treatment. The CSC benefits seen were not related to sex, gender,
race, or BMI. The magnitude of reduction of these symptoms in a
preventive manner has enormous implications for public health.
[0071] In addition to its effect on the incidence of cold and flu
symptoms, CSC ingestion enhanced proliferation of, and IFN-.gamma.
secretion by, .gamma..delta. T cells challenged in vitro with
ethylamine. These findings are consistent with our previous study
showing that subjects who drank tea containing L-theanine, but not
coffee, increased their .gamma..delta. T cell function by up to
15-fold [20]. It is important to point out that .gamma..delta. T
cell numbers or serum IFN-.gamma. levels were not increased in
subjects taking CSC, but that their .gamma..delta. T cells were
primed in vivo by CSC to respond more vigorously to antigenic
challenge. Constitutively increased in .gamma..delta. T cell
numbers and serum IFN-.gamma. titers in the absence of pathogen
exposure may actually increase symptoms due to inflammation. Such
increased symptoms occur after ingestion of high doses of
nitrogen-containing bisphosphonates (which activate .gamma..delta.
T cells) for treatment of osteoporosis [17].
[0072] The mechanism of action of symptom reduction has not been
determined with certainty. L-theanine is known to activate human
.gamma..delta. T lymphocytes to secrete IFN-.gamma., a powerful
antimicrobial cytokine [19, 20]. Without being bound to any
particular mechanistic theory, the inventors proposed that in the
early stages of infection, EGCG may aid in the activation of
.gamma..delta. T cells by enhancing secretion of IL-12, a very
important factor in .gamma..delta. T cell activation [20, 18]. If
infection does establish itself by evading the first-line
.gamma..delta. T cell response, EGCG may decrease cold and flu
symptoms due its anti-inflammatory activity [21]. See also above.
Studies in our laboratory are now underway to address these
mechanistic questions.
[0073] This study has several limitations. Symptom data were
collected by self-reporting, so an actual medical diagnosis of cold
or flu was not possible. Other illnesses, such as pneumonia,
bronchitis, or allergy might have caused similar symptoms. The
study was comprised of only healthy adult subjects, so it was not
possible to assess the effect of CSC on children, or subjects with
chronic illnesses who have increased susceptibility to acute
illness.
[0074] Cold and flu symptoms can be perennial sources of misery and
lost productivity for most healthy adults, and the introduction of
a safe, effective, and natural capsule that can prevent such
symptoms represents a significant breakthrough in preventive
medicine. The foregoing study demonstrates that compositions
containing L-theanine in combination with EGCG are a safe and
effective preventative for cold and flu symptoms, and for enhancing
the innate immune response. Widespread use of CSC could have
enormous beneficial effects in decreasing morbidity in healthy
populations.
ABBREVIATIONS
[0075] CSC, Camellia sinensis composition; EGCG, epigallocatechin
gallate, PBMC, peripheral blood mononuclear cells
REFERENCES
[0076] 1. Cabrera C, Artacho R, Gimenez R: Beneficial effects of
green tea--a review. J Am Coll Nutr 25(2): 79-99; 2006 [0077] 2.
Chung F L, Schwartz J, Herzog C R, Yang Y M: Proceedings of the
third international scientific symposium on tea and human health:
Role of flavonoids in the diet. J. Nutr. 33: 3268S-32744S; 2003
[0078] 3. Chung F L, Schwartz J, Herzog C R, Yang Y M: Tea and
cancer prevention: studies in animals and humans. J Nutr 133(10):
3268S-3274S; 2003 [0079] 4. Michels K B, Willett W C, Fuchs C S,
Giovannucci E: Coffee, tea, and caffeine consumption and incidence
of colon and rectal cancer. J Natl Cancer Inst 97(4): 282-292; 2005
[0080] 5. Jordan S J, Purdie D M, Green A C, Webb P M: Coffee, tea
and caffeine and risk of epithelial ovarian cancer. Cancer Causes
Control 15(4): 359-365; 2004 [0081] 6. Graham H N: Green tea
composition, consumption, and polyphenol chemistry. Preventive
Medicine 21(3): 334-350; 1992 [0082] 7. Nichol K L, D'Heilly S,
Ehlinger E: Colds and influenza-like illnesses in university
students: impact on health, academic and work performance, and
health care use. Clin Infect Dis 40(9): 1263-1270; 2005 [0083] 8.
Eby G A, Halcomb W W: Ineffectiveness of zinc gluconate nasal spray
and zinc orotate lozenges in common-cold treatment: a double-blind,
placebo-controlled clinical trial. Altern Ther Health Med 12(1):
34-38; 2006 [0084] 9. Linde K, Barrett B, Wolkart K, Bauer R,
Melchart D: Echinacea for preventing and treating the common cold.
Cochrane Database Syst Rev (1): CD000530; 2006 [0085] 10. Das H,
Wang L, Kamath A, Bukowski J: V.gamma.2 and V.delta.2 T cell
receptor-mediated recognition of aminobisphosphonates. Blood (in
press) 2001 [0086] 11. Nantz M P, Rowe C A, Nieves C, Jr., Percival
S S: Immunity and antioxidant capacity in humans is enhanced by
consumption of a dried, encapsulated fruit and vegetable juice
concentrate. J Nutr 136(10): 2606-2610; 2006 [0087] 12. Shirai T,
Hayakawa H, Akiyama J, et al.: Food allergy to green tea. J Allergy
Clin Immunol 112(4): 805-806; 2003 [0088] '3. Kim S H, Park H J,
Lee C M, et al.: Epigallocatechin-3-gallate protects toluene
diisocyanate-induced airway inflammation in a murine model of
asthma. FEBS Lett 580(7): 1883-1890; 2006 [0089] 14. Hisano M,
Yamaguchi K, Inoue Y, et al.: Inhibitory effect of catechin against
the superantigen staphylococcal enterotoxin B (SEB). Arch Dermatol
Res 295(5): 183-189; 2003 [0090] 15. Wang L, Das H, Kamath A,
Bukowski J: Human Vg2Vd2 T cells produce IFN-g and TNF-a with an
on/off/on cycling pattern in response to live bacterial products.
J. Immunol. 167: 6195-6201; 2001 [0091] 16. Committee DGA: Dietary
guidelines for Americans, 2005. Washington, D.C.: USDA, 2005.
[0092] 17. Reginster J Y, Adami S, Lakatos P, et al.: Efficacy and
tolerability of once-monthly oral ibandronate in postmenopausal
osteoporosis: 2 year results from the MOBILE study. Ann Rheum Dis
65(5): 654-661; 2006 [0093] 18. Katiyar S K: Skin photoprotection
by green tea: antioxidant and immunomodulatory effects. Curr Drug
Targets Immune Endocr Metabol Disord 3(3): 234-242; 2003 [0094] 19.
Bukowski J F, Morita C T, Brenner M B: Human gd T cells recognize
alkylamines derived from microbes, edible plants, and tea:
implications for innate immunity. Immunity 11(1): 57-65; 1999
[0095] 20. Kamath A B, Wang L, Das H, Li L, Reinhold V N, Bukowski
J F: Antigens in tea-beverage prime human Vgamma 2Vdelta 2 T cells
in vitro and in vivo for memory and nonmemory antibacterial
cytokine responses. Proc Natl Acad Sci USA 100(10): 6009-6014; 2003
[0096] 21. Handa O, Naito Y, Takagi T, et al.: Inhibitory effects
of catechins on neutrophil-dependent gastric inflammation. Redox
Rep 7(5): 324-328; 2002 [0097] 22. August D A, Landau J, Caputo D,
Hong J, Lee M J, Yang C S: Ingestion of green tea rapidly decreases
prostaglandin E2 levels in rectal mucosa in humans. Cancer
Epidemiol Biomarkers Prey 8(8): 709-713; 1999 [0098] 23. Yuan G J,
Gong Z J, Sun X M, Zheng S H, Li X: Tea polyphenols inhibit
expressions of iNOS and TNF-alpha and prevent
lipopolysaccharide-induced liver injury in rats. Hepatobiliary
Pancreat Dis Int 5(2): 262-267; 2006 [0099] 24. Kawai K, Tsuno N H,
Kitayama J, et al.: Epigallocatechin gallate attenuates adhesion
and migration of CD8+ T cells by binding to CD11b. J Allergy Clin
Immunol 113(6): 1211-1217; 2004 [0100] 25. Singh R, Ahmed S,
Malemud C J, Goldberg V M, Haqqi T M: Epigallocatechin-3-gallate
selectively inhibits interleukin-1beta-induced activation of
mitogen activated protein kinase subgroup c-Jun N-terminal kinase
in human osteoarthritis chondrocytes. J Orthop Res 21(1): 102-109;
2003
TABLE-US-00001 [0100] TABLE 1 Study withdrawals Capsules Subject
Consumed # Treatment For Reason for Withdrawal 2 CSC 11 d Facial
& chest rash, itching, red/watery eyes, puffy eyes, congestion)
30 placebo ? Unable to return 43 CSC On & off Hives 3x 20 dy
(?) 45 placebo 32 dy Traveling interfered 60 placebo 1 mo (?)
Urinary tract infection 116 placebo ? Discontinued taking capsules
119 placebo 1 mo (?) Capsules upsetting stomach
TABLE-US-00002 TABLE 2 Demographics of the study population CSC
Placebo P value Age 28.9 .+-. 1.07 30.3 .+-. 1.5 0.65 Gender male
20 23 Gender female 32 33 Height (m) 1.7 .+-. 0.01 1.7 .+-. 0.01
0.60 Weight (kg) 74.9 .+-. 1.7 73.3 .+-. 2.0 0.67 BMI 25.4 .+-. 0.6
24.3 .+-. 0.8 0.94 Compliance (%) 93% .+-. 7 93% .+-. 7 0.56
Blinding (% 50 48 1.0 guessed correctly)
TABLE-US-00003 TABLE 3 Interferon-.gamma. (ng/mL) secreted into the
culture medium after 24 h Placebo CSC n = 52) (n = 56) p values 0
mM ethylamine 2.8.sup.C 2.3.sup.C Ethylamine level = 0.001 1 mM
ethylamine 3.8.sup.C 2.1.sup.C Treatment = 0.046 5 mM ethylamine
18.4.sup.A 13.6.sup.B Interaction = 0.084 Pooled SEM 3.9 Repeated
measures two-way ANOVA. Means with different superscripts are
statistically different using a post hoc SNK test.
TABLE-US-00004 TABLE 4 % .gamma..delta.-T cells in baseline blood
sample and after 10 d of culture Placebo CSC n = 52) (n = 56) p
values Baseline (no culture) 13.2 .+-. 4.6 14.3 .+-. 6.6 0.270 0
ethylamine, 10 d 20.8 .+-. 7.0 21.9 .+-. 5.7 0.298 1 mM ethylamine,
28.0 .+-. 5.0 20.3 .+-. 4.0 0.017 10 d Repeated measures two-way
ANOVA; means .+-. SEM.
TABLE-US-00005 TABLE 5 1/56 (1.79%) CSC subjects had diarrhea; 3/55
(5.45%) placebo subjects had diarrhea (67.1%) 3/56 (5.35%) CSC
subjects had fever; 7/55 (12.7%) placebo subjects had fever (57.8%)
This represents the % reduction in subjects experiencing these
symptoms, compared to placebo 16/56 (28.6%) CSC subjects had sore
throat; 25/55 (45.4%) placebo subjects has sore throat (37.0%)
14/56 (23.2%) CSC subjects had headache; 15/55 (27.2%) placebo
subjects had headache (1.47%) 2/56 (3.57%) CSC subjects had nausea;
4/55 (7.27%) placebo subjects had nausea (50.9%) 11/56 (19.6%) CSC
subjects had cough; 16/55 (29.1%) placebo subjects had cough
(32.6%) 20/56 (35.7%) CSC subject had runny nose; (29/56) (51.8%)
placebo subjects had runny nose (31.1%) 19/56 (33.9%) CSC subjects
had stuffy nose; 21/55 (38.2%) placebo subjects had stuffy nose
(11.2%)
[0101] While the principles of the invention have been made clear
in illustrative embodiments, there will be immediately apparent to
those skilled in the art, in view of the teachings herein, many
modifications of structure, arrangement, proportions, the elements,
materials, and components used in the practice of the invention,
and otherwise, which are particularly adapted to specific
environments and operative requirements without departing from
those principles. The appended claims are intended to cover and
embrace any and all such modifications, with the limits only of the
true purview, spirit and scope of the invention.
[0102] The references referred to herein are incorporated herein in
their entirety to the extent they are not inconsistent with the
teachings herein.
* * * * *