FIELD OF INVENTION
[001] The present disclosure relates to the field of cosmetic formulations in general and compositions for the inhibition of skin aging in particular. There is provided a composition comprising phlorizin, L-citrulline, and an amino acid, and the uses thereof to address the problem of skin aging.
BACKGROUND OF THE INVENTION
[002] Skin aging is a multifactorial process that is marked by reduced synthesis of extracellular matrix (ECM) proteins like collagen and elastin and increase in protease activity leading to degradation of ECM and decrease in epidermal turnover. Collagen is a major component of the ECM and one of the primary connective tissues found in the skin and other parts of the body. The protein is found in the form of helical fibrils within the ECM. The degradation of these fibrils occurs primarily by the activity of enzymes such as Matrix Metalloproteinase (MMP) enzymes, which cleave the fibrils into fragments. Studies suggest that these enzymes are activated by UV, which then subsequently leads to the aggregation of collagen fibres causing loss in skin elasticity (www.thedermalinstitute.com).
[003] Skin aging manifests in the form of fine lines and wrinkles on the facial skin and other parts of the body. As skin ages, due to changes in the desquamation pattern, the epidermal barrier is disrupted resulting in the loss of natural moisturizing factors. Uncontrolled dehydration severely affects the ECM maintenance and continuity as a result of which skin becomes dry and thin. A literature survey reveals (MinaYaar et al., 2002, J. Invest. Dermatol. Symp. Proc; 7:51-58; Ganceviciene R, 2012; Dermatoendocrinol; 4(3):308-319) that active research is being carried out to increase the levels of collagen by boosting collagen synthesis, which can eventually decrease facial fine lines.
[004] Prior art documents disclosing various compounds/compositions aimed at ameliorating the problem of skin aging are found ubiquitously. These include compounds, such as, retinol, epidermal growth factors, ascorbic acid and argireline,

among others. However, the problem of skin aging has still not been adequately addressed and therefore there is a need to identify new formulations.
SUMMARY OF INVENTION
[005] In an aspect of the present disclosure there is provided a composition
comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-
citrulline to L-histidine w/w ratio is said composition is in the range of 1:1:0.8- 1:
1.3:5.
[006] In an aspect of the present disclosure there is a provided a method of
preparing a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-
histidine, said method comprising:
(a) obtaining phlorizin;
(b) obtaining L-citrulline;
(c) obtaining L-histidine; and
(d) contacting phlorizin, L-citrulline and L-histidine to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio is said composition is in the range of 1:1:0.8- 1: 1.3:5.
[007] In an aspect of the present disclosure there is a provided a method of preparing a composition comprising: (i) phlorizin; (ii) L-citrulline; (iii) L-histidine; and (iv) at least one carrier, at least one diluent, and at least one excipient, said method comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline;
(c) obtaining L-histidine; and
(d) obtaining at least one carrier, at least one diluent and at least one excipient; and
(e) contacting phlorizin, L-citrulline, L-histidine and at least one carrier, at least one
diluent and at least one excipient to obtain said composition,
wherein phlorizin to L-citrulline to L-histidine w/w ratio is said composition is in the range of 1:1:0.8- 1: 1.3:5.

[008] In an aspect of the present disclosure there is provided a method of inhibiting skin aging, said method comprising:
(a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-histidine, having phlorizin to L-citrulline to L-histidine in a w/w ratio of 1:1:0.8- 1: 1.3:5; and
(b) contacting said composition with skin.
[009] These and other features, aspects, and advantages of the present subject matter will be better understood with reference to the following description and appended claims. This summary is provided to introduce a selection of concepts in a simplified form. This summary is not intended to identify key features or essential features of the claimed subject matter, nor is it intended to be used to limit the scope of the claimed subject matter.
BRIEF DESCRIPTION OF ACCOMPANYING DRAWINGS
[0010] The following drawings form a part of the present specification and are
included to further illustrate aspects of the present disclosure. The disclosure may be
better understood by reference to the drawings in combination with the detailed
description of the specific embodiments presented herein.
[0011] Figure 1 shows the effect of the combination of phlorizin (P) and L-citrulline
(C) on the expression of collagen proteins, in accordance with an embodiment of the
present disclosure.
[0012] Figure 2 shows the effect of the combination of phlorizin (P), L-citrulline (C),
and L-histidine (H) on the expression of collagen proteins at a 1:1:2.4 w/w ratio of
phlorizin to L-citrulline to L-histidine, in accordance with an embodiment of the
present disclosure.
[0013] Figure 3 shows the effect of the combination of phlorizin (P), L-citrulline (C),
and L-histidine (H) on the expression of collagen proteins at a 1:1:4.7 w/w ratio of

phlorizin to L-citrulline to L-histidine, in accordance with an embodiment of the present disclosure.
[0014] Figure 4 shows the effect of the combination of phlorizin (P), L-citrulline (C), and L-histidine (H) on the expression of collagen proteins at a 1:1:3 w/w ratio of phlorizin to L-citrulline to L-histidine, in accordance with an embodiment of the present disclosure.
[0015] Figure 5 shows the effect of the combination of phlorizin (P), L-citrulline (C), and oleanolic acid (OA) on the expression of collagen proteins at a 1:1:0.008 and 1:1:0.016 w/w ratio of phlorizin to L-citrulline to oleanolic acid, in accordance with an embodiment of the present disclosure.
[0016] Figure 6 shows the effect of the combination of phlorizin (P), L-citrulline (C), and oleanolic acid (OA) on the expression of collagen proteins at a 1:1:0.006 and 1:1:0.012 w/w ratio of phlorizin to L-citrulline to oleanolic acid, in accordance with an embodiment of the present disclosure.
[0017] Figure 7 shows the cytotoxic effects of phlorizin at w/w concentrations ranging from 0.0008% to 0.1%, in accordance with an embodiment of the present disclosure.
[0018] Figure 8 shows the cytotoxic effects of L-citrulline at w/w concentrations ranging from 0.008%) to 1%, in accordance with an embodiment of the present disclosure.
[0019] Figure 9 shows the cytotoxic effects of L-histidine at w/w concentrations ranging from 0.00002% to 0.039% L-histidine, in accordance with an embodiment of the present disclosure.
[0020] Figure 10 shows the cytotoxic effects of a combination of phlorizin and L-citrulline (PC) at w/w concentrations ranging from 0.00005%) to 0.1%> PC, in accordance with an embodiment of the present disclosure.

DETAILED DESCRIPTION OF THE INVENTION
[0021] The present disclosure relates to a composition comprising phlorizin, L-
citrulline and L-histidine, which at particular w/w concentrations can promote
collagen protein synthesis and therefore delay the process of skin aging.
[0022] Those skilled in the art will be aware that the present disclosure is subject to
variations and modifications other than those specifically described. It is to be
understood that the present disclosure includes all such variations and modifications.
The disclosure also includes all such steps, features, compositions, and compounds
referred to or indicated in this specification, individually or collectively, and any and
all combinations of any or more of such steps or features.
Definitions
[0023] For convenience, before further description of the present disclosure, certain
terms employed in the specification, and examples are delineated here. These
definitions should be read in the light of the remainder of the disclosure and
understood as by a person of skill in the art. The terms used herein have the meanings
recognized and known to those of skill in the art, however, for convenience and
completeness, particular terms and their meanings are set forth below.
[0024] The articles "a", "an" and "the" are used to refer to one or to more than one
(i.e., to at least one) of the grammatical object of the article.
[0025] The terms "comprise" and "comprising" are used in the inclusive, open sense,
meaning that additional elements may be included. It is not intended to be construed
as "consists of only".
[0026] Throughout this specification, unless the context requires otherwise the word
"comprise", and variations such as "comprises" and "comprising", will be understood
to imply the inclusion of a stated element or step or group of element or steps but not
the exclusion of any other element or step or group of element or steps.
[0027] The term "including" is used to mean "including but not limited to".
"Including" and "including but not limited to" are used interchangeably.

[0028] Phlorizin is a naturally occurring flavonoid found in bark of apple and many fruit bearing trees. It belongs to group of dihydrochalcones (Silke et al., Dermatoendocrinol., 4(3), 298-307, 2012). Phlorizin is a sodium glucose co-transporter inhibitor. Type IV and type VI collagen synthesis is normalized by phlorizin with decreasing glucose consumption under high glucose conditions (Wakisaka et al., Biochimica et Biophysica Acta (BBA), 1362 (1), 87-96, 1997). [0029] L-citrulline is an alpha-amino acid and is found in many fruits particularly in watermelon which is a major source. In the body, it can be metabolized to L-arginine and it is involved in the maintenance of vascular function (Kobayashi, Yoshinori et al.; Pharmacognosy Research 6.4: 297-302, 2014).
[0030] L-histidine is an essential amino acid and is positively charged at physiological pH, thus classifying it as a basic amino acid. It is not synthesized de novo in humans and therefore must be supplemented in the diet. Major sources of the amino acid include legumes, seafood and dairy products.
[0031] 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 this disclosure belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the disclosure, the preferred methods, and materials are now described. All publications mentioned herein are incorporated herein by reference.
[0032] Skin aging is a long-term degenerative process that involves, among others, the degradation of proteins that act as the connective tissue of the skin. Collagen and elastin among the primary proteins involved, and their degradation results in the observable effects of skin ageing such as the appearance of fine lines and wrinkles. Although several compositions are available to address this problem, no solution has been found. The invention disclosed herein provides a solution to address skin aging by promoting collagen synthesis using a composition of phlorizin, L-citrulline and L-histidine. It provides a formulation and the best method to prepare the same.

[0033] The present disclosure is not to be limited in scope by the specific embodiments described herein, which are intended for the purposes of exemplification only. Functionally-equivalent products, compositions, and methods are clearly within the scope of the disclosure, as described herein.
[0034] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0035] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5, wherein said composition enhances collagen synthesis. [0036] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0037] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of 0.02-0.1%; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0038] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline having a weight percentage of 0.0165%; and (c) L-histidine having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.

[0039] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of 0.02-0.1%; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4. [0040] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline having a weight percentage of 0.0165%; and (c) L-histidine having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0041] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of 0.02-0.1%; (b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5. [0042] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8-1:1.3:5.
[0043] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8-1:1.3:5.
[0044] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of 0.02-0.1%; (b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.

[0045] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight
percentage in the range of 0.02-0.1%; and (c) L-histidine, wherein phlorizin to L-
citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0046] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine
having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-
citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0047] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%;
(b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine
w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0048] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight
percentage of 0.0165%; and (c) L-histidine, wherein phlorizin to L-citrulline to L-
histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0049] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine
having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine
w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0050] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%;
(b) L-citrulline; and (c) L-histidine, wherein phlorizin to L-citrulline to L-histidine
w/w ratio in said composition is 1:1:2.4.
[0051] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight
percentage of 0.0165%; and (c) L-histidine, wherein phlorizin to L-citrulline to L-
histidine w/w ratio in said composition is 1:1:2.4.

[0052] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine
having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine
w/w ratio in said composition is 1:1:2.4.
[0053] In an embodiment of the present disclosure, there is provided a composition
as described herein, wherein said composition can be for topical application.
[0054] In an embodiment of the present disclosure, there is provided a composition
as described herein, wherein said composition can be in the form of an aerosol.
[0055] In an embodiment of the present disclosure, there is provided a composition
as described herein, wherein said composition can be in the form of a lotion.
[0056] In an embodiment of the present disclosure, there is provided a composition
as described herein, wherein said composition can be in form of an oil.
[0057] In an embodiment of the present disclosure, there is provided a composition
as described herein, wherein said composition can be in the form of a serum.
[0058] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; (c) L-histidine; (d) at
least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein
phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8-
1:1.3:5.
[0059] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of
0.02-0.1%; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; and
(c) L-histidine having a weight percentage in the range of 0.01-0.1%; (d) at least one
carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-
citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8-
1:1.3:5.
[0060] In an embodiment of the present disclosure, there is provided a composition to
inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of

0.02-0.1%; (b) L-citrulline; (c) L-histidine; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0061] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) L-histidine; (d) at least one carrier; (e) at least one diluent, and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0062] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; (c) L-histidine having a weight percentage in the range of 0.01-0.1%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0063] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; (c) L-histidine; (d) at least one carrier; (e) at least one diluent, and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4. [0064] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of 0.02-0.1%>; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) L-histidine having a weight percentage in the range of 0.01-0.1%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0065] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage in the range of 0.02-0.1%>; (b) L-citrulline; (c) L-histidine; (d) at least one carrier, (e) at least one diluent, and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.

[0066] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) L-histidine; (d) at least one carrier, (e) at least one diluent, and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0067] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; (c) L-histidine having a weight percentage in the range of 0.01-0.1%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0068] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline having a weight percentage of 0.0165%; (c) L-histidine having a weight percentage of 0.039%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0069] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline; (c) L-histidine; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0070] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight percentage of 0.0165%; (c) L-histidine; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0071] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; (c) L-histidine having a

weight percentage of 0.039%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0072] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline having a weight percentage of 0.0165%; (c) L-histidine having a weight percentage of 0.039%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0073] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline; (c) L-histidine; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0074] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline having a weight percentage of 0.0165%; (c) L-histidine; (d) at least one carrier; (e) at least one diluent, and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0075] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; (c) L-histidine having a weight percentage of 0.039%; (d) at least one carrier; (e) at least one diluent; and (f) at least one excipient, wherein phlorizin to L-citrulline to L-histidine in said composition is 1:1:2.4.
[0076] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging, said method comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine; and (d) contacting phlorizin, L-citrulline and L-histidine to obtain said composition.

[0077] In an embodiment of the present disclosure, there is provided a method of
preparing a composition to inhibit skin aging comprising the steps of: (a) obtaining
phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine; and (d) contacting
phlorizin, L-citrulline, and L-histidine to obtain said composition, wherein phlorizin
to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8-
1:1.3:5.
[0078] In an embodiment of the present disclosure, there is provided a method of
preparing a composition to inhibit skin aging comprising the steps of: (a) obtaining
phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine; and (d) contacting
phlorizin, L-citrulline, and L-histidine to obtain said composition, wherein phlorizin
to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0079] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)
L-citrulline having a weight percentage of 0.02-0.1%; (b) phlorizin; and (c) L-
histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition
is in the range of 1:1:0.8- 1:1.3:5.
[0080] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)
L-citrulline having a weight percentage in the range of 0.02-0.1%; (b) phlorizin; and
(c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said
composition is 1:1:2.4.
[0081] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)
L-citrulline having a weight percentage of 0.0165%; (b) phlorizin; and (c) L-histidine,
wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the
range of 1:1:0.8- 1:1.3:5.
[0082] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)

L-citrulline having a weight percentage of 0.0165%; (b) phlorizin; and (c) L-histidine,
wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is
1:1:2.4.
[0083] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)
L-citrulline; (b) phlorizin having a weight percentage in the range of 0.02-0.1%; and
(c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said
composition is in the range of 1:1:0.8- 1:1.3:5.
[0084] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)
L-citrulline; (b) phlorizin having a weight percentage in the range of 0.02-0.1%; and
(c) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said
composition is 1:1:2.4.
[0085] In an embodiment of the present disclosure, there is provided a method of
preparing a composition as described herein, wherein said composition comprises: (a)
L-citrulline; (b) phlorizin having a weight percentage of 0.0165%; and (c) L-histidine;
wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the
range of 1:1:0.8- 1:1.3:5.
[0086] In an embodiment of the present disclosure, there is provided a method of
preparing a composition described herein, wherein said composition comprises: (a) L-
citrulline; (b) phlorizin having a weight percentage of 0.0165%; and (c) L-histidine,
wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is
1:1:2.4.
[0087] In an embodiment of the present disclosure, there is provided a method of
preparing a composition described herein, wherein said composition comprises: (a)
L-citrulline; (b) phlorizin; and (c) L-histidine having a weight percentage in the range
of 0.01-0.1%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said
composition is in the range of 1:1:0.8- 1:1.3:5.

[0088] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) L-citrulline; (b) phlorizin; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0089] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) L-citrulline; (b) phlorizin; and (c) L-histidine having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0090] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) L-citrulline; (b) phlorizin; and (c) L-histidine having a weight percentage of 0.039%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0091] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) L-citrulline having a weight percentage in the range of 0.02-0.1%; (b) phlorizin having a weight percentage in the range of 0.02-0.1%; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5. [0092] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) L-citrulline having a weight percentage in the range of 0.02-0.1%; (b) phlorizin having a weight percentage in the range of 0.02-0.1%; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.

[0093] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline having a weight percentage of 0.0165%; and (c) L-histidine having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0094] In an embodiment of the present disclosure, there is provided a method of preparing a composition as described herein, wherein said composition comprises: (a) phlorizin having a weight percentage of 0.0165%; (b) L-citrulline having a weight percentage of 0.0165%; and (c) L-histidine having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[0095] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition.
[0096] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5. [0097] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at

least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio is 1:1:2.4.
[0098] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin having a weight percentage in the range of 0.02-0.1%; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[0099] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin having a weight percentage in the range of 0.02-0.1%; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[00100] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin having a weight percentage of 0.0165%; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.

[00101] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin having a weight percentage of 0.0165%; (b) obtaining L-citrulline; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[00102] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[00103] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[00104] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline having a weight percentage of 0.0165%; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient,

and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[00105] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline having a weight percentage of 0.0165%; (c) obtaining L-histidine; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition 1:1:2.4. [00106] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine having a weight percentage in the range of 0.01-0.1%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8-1:1.3:5.
[00107] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine having a weight percentage in the range of 0.01-0.1%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said

composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[00108] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine having a weight percentage of 0.039%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[00109] In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin; (b) obtaining L-citrulline; (c) obtaining L-histidine having a weight percentage of 0.039%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[OOllOJIn an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin having a weight percentage in the range of 0.02-0.1%; (b) obtaining L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) obtaining L-histidine having a weight percentage in the range of 0.01-0.1%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.

[OOlllJIn an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging comprising: (a) obtaining phlorizin having a weight percentage in the range of 0.02-0.1%; (b) obtaining L-citrulline having a weight percentage in the range of 0.02-0.1%; (c) obtaining L-histidine having a weight percentage in the range of 0.01-0.1%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.
[00112]In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging said method comprising: (a) obtaining phlorizin having a weight percentage of 0.0165%; (b) obtaining L-citrulline phlorizin having a weight percentage of 0.0165%; (c) obtaining L-histidine having a weight percentage of 0.039%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
[00113]In an embodiment of the present disclosure, there is provided a method of preparing a composition to inhibit skin aging, said method comprising: (a) obtaining phlorizin having a weight percentage of 0.0165%; (b) obtaining L-citrulline phlorizin having a weight percentage of 0.0165%; (c) obtaining L-histidine having a weight percentage of 0.039%; (d) obtaining at least one carrier, at least one diluent, at least one excipient, and combinations thereof; and (e) contacting phlorizin, L-citrulline, L-histidine and carrier, diluent, excipient and combinations thereof, to obtain said composition, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4.

[00114]In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin.
[00115] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin.
[00116] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage in the range of 0.02-0.1%; (ii) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (iii) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin.
[00117] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage of 0.0165%; (ii) L-citrulline having a weight percentage of 0.0165%; and (iii) L-histidine having a weight percentage of 0.039%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin. [00118] In an embodiment of the present disclosure there is provided, a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage in the range of 0.02-0.1%; (ii) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (iii) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-

histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin.
[00119] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage of 0.0165%; (ii) L-citrulline having a weight percentage of 0.0165%; and (iii) L-histidine having a weight percentage of 0.039%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin.
[00120] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage in the range of 0.02-0.1%; (ii) L-citrulline; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin.
[00121] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin.
[00122] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-histidine having a weight percentage in the range of 0.01-0.1%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin.
[00123] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i)

phlorizin having a weight percentage in the range of 0.02-0.1%; (ii) L-citrulline; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin. [00124] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline having a weight percentage in the range of 0.02-0.1%; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin. [00125] In an embodiment of the present disclosure, there is provided a composition to inhibit skin aging comprising: (a) phlorizin; (b) L-citrulline; and (c) L-histidine having a weight percentage in the range of 0.01-0.1%, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4. [00126] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage of 0.0165%; (ii) L-citrulline; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin. [00127] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline having a weight percentage of 0.0165%; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin. [00128] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-histidine having a weight percentage of 0.039%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5; and (b) contacting said composition with skin.

[00129] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin having a weight percentage of 0.0165%; (ii) L-citrulline; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin.
[00130] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline having a weight percentage of 0.0165%; and (iii) L-histidine, wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin.
[00131] In an embodiment of the present disclosure, there is provided a method to inhibit skin aging, comprising the steps of: (a) obtaining a composition comprising: (i) phlorizin; (ii) L-citrulline; and (iii) L-histidine having a weight percentage of 0.039%), wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is 1:1:2.4; and (b) contacting said composition with skin.
[00132]Although the subject matter has been described with reference to specific embodiments, this description is not meant to be construed in a limiting sense. Various modifications of the disclosed embodiments, as well as alternate embodiments of the subject matter, will become apparent to persons skilled in the art upon reference to the description of the subject matter. It is therefore contemplated that such modifications can be made without departing from the spirit or scope of the present subject matter as defined.
[00133]Although the subject matter has been described with reference to specific embodiments, this description is not meant to be construed in a limiting sense. Various modifications of the disclosed embodiments, as well as alternate embodiments of the subject matter, will become apparent to persons skilled in the art upon reference to the description of the subject matter. It is therefore contemplated

that such modifications can be made without departing from the spirit or scope of the present subject matter as defined.
EXAMPLES
[00134] The disclosure will now be illustrated with working examples, which is intended to illustrate the working of disclosure and not intended to take restrictively to imply any limitations on the scope of the present disclosure. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art to which this disclosure belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice of the disclosed methods and compositions, the exemplary methods, devices and materials are described herein. It is to be understood that this disclosure is not limited to particular methods, and experimental conditions described, as such methods and conditions may vary
Example 1
Material and Methods
[001351 Testing cellular uptake of compositions: Human Adult dermal fibroblasts (Himedia, India) were seeded in 12- well plates (Costar) in fibroblast growth medium (LONZA). The culture was incubated in 5% CO2 incubator and was allowed to reach confluency to form a fibroblast bed. HaCaT keratinocytes (NCCS, India) at a seeding density of 0.1 million cells per well was seeded on the fibroblast bed. Medium 154CF/PRF (GIBCO) containing 0.03mM calcium chloride was used while seeding HaCaT keratinocytes. After 24 hrs, medium was replenished with actives and incubated in 5% CO2 incubator. Cells were harvested after 48 hrs for total collagen estimation. Sircol assay was performed as per Biocolor life science assays (SI000) protocol.

[00136]Phlorizin (CAS Number 7061-54-3), L-citrulline (CAS number 372-75-8) and L-histidine (CAS number 71-00-1) were procured from SIGMA. Oleanolic acid (80%) was obtained from Sami labs.
[00137]Preparation of phlorizin and L-citrulline (PC) combination: PC combination contained 2.5% of phlorizin and 2.5% of L- citrulline. The total stock concentration of PC was taken as 5% and the tested final concentrations were calculated accordingly. L-Histidine was prepared at a stock concentration of 1.55% in distilled water and added to the cell culture treatment medium accordingly.
Example 2
Effect of a combination of phlorizin and L-citrulline on collagen protein
expression
[00138] The combination of phlorizin (P) and citrulline (C) at different concentrations was first tested to determine the effect of this composition on collagen expression. PC at the concentrations of 0.1%, 0.033% and 0.011% was tested as per the protocol mentioned in Example 1. It was found that PC at 0.033%) had slightly more collagen expression at approximately 170%), than PC at 0.1% or 0.011% concentrations (Figure 1). This concentration was therefore chosen for further experiments.
Example 3
Effect of a combination of phlorizin, L-citrulline and L-histidine on collagen
protein expression
[00139]Phlorizin and citrulline at 0.033%) was determined as the working concentration that stimulated collagen protein expression. Compared to the individual actives, this combination showed a marginal increase in collagen protein expression. To this combination, L-histidine was then added to ascertain whether this could enhance collagen synthesis in cells.
[00140]Remarkably, it was found that the addition of 0.039%) (w/w) L-histidine to the combination of 0.033% of phlorizin and L-citrulline combined, significantly enhanced

collagen protein expression as compared to the individual actives (Table 1, Figure 2). This represents a 1:1:2.4 w/w ratio of phlorizin: L-citrulline: L-histidine. This specific combination of phlorizin, L-citrulline and L-histidine together showed enhanced protein expression at 117%, which was approximately 10-20% higher than the individual actives or the combination of phlorizin and L-citrulline.
[00141]Further, synergism is evidenced by the observation that the individual actives demonstrate very similar expression values, at around 100%) total collagen content. Thus, the increased collagen protein expression observed is more than what is expected from a mere admixture of the three components together. [00142] The synergistic composition of phlorizin, L-citrulline and L-histidine was then tested at different concentrations to determine whether synergism occurred across a range of concentrations. A 1:1:4.7 ratio of phlorizin: L-citrulline: L-histidine at 0.033% phlorizin and L-citrulline combined, and 0.078% L-histidine did not demonstrate any synergism (Table 2, Figure 3). The individual actives and the composition showed very similar expression values.

Additionally, the combination of phlorizin, L-citrulline and L-histidine was tested at a w/w ratio of 1:1:3 at 0.025% phlorizin and L-citrulline, and 0.039% L-histidine. This specific combination also did not demonstrate any synergism in its activity and as compared to the individual actives, this composition showed no marked improvement in collagen protein expression, rather as compared to some actives, such as histidine alone, the percent of total collagen of a 1:1: 3 showed a marginal decrease (Table 3, Figure 4).
[00143] Thus, from these data it is inferred that within a 1:1:2.4 w/w ratio the combination of phlorizin: L-citrulline: L-histidine, demonstrates distinct synergism in enhancing collagen protein expression.
[00144] The examples listed below further demonstrate that the specific effect of this composition and thereby its use to inhibit skin aging, is a property specific to this combination of constituents and cannot be obtained by interchanging any of the constituents.

[00145] A composition comprising phlorizin, L-citrulline and oleanolic acid at two different concentrations was administered to cells, as in the previous experiments, to determine if the composition could influence collagen protein expression. It was found that this composition did not have any effect on collagen expression as compared to the individual actives (Figure 5 and 6, Table 4 and 5). Using either 0.0001% or 0.0002% oleanolic acid with 0.025% and 0.033% phlorizin and L-citrulline combination, did not demonstrate any significant effect in stimulating collagen gene expression.

Cytotoxicity of phlorizin, L-citrulline, L-histidine and the combination thereof
[00146] Figures 7, 8, 9, 10 depict the concentrations of phlorizin, L-citrulline and L-
histidine and the combination of these three constituents at which cell toxic effects are
observed.
[00147] Figure 7 shows cytotoxicity data for phlorizin. Phlorizin does not show any
distinct toxic effects between the ranges of 0.0008%-0.05%. At 0.1% phlorizin, the
cell viability shows a significant reduction at 80%.
[00148] Figure 8 shows the cell toxic effects of L-citrulline. Within the ranges of
0.008%) to 1%>, L-citrulline does not have any toxic effects and cells show 100%)
viability at all the concentrations chosen within this range.
[00149]Figure 9 shows the cell toxic effects of L-histidine. Concentrations between
0.00002% to 0.0014% do not show any cell toxic effects. Above 0.004%, the L-
histidine demonstrates marginal toxic effects, however, these are not at significant
levels.
[00150] Figure 10 illustrates the cytotoxicity data for the combination of phlorizin and
citrulline within the range of 0.000005% to 0.1%. The combination of phlorizin and
L-citrulline does not show any significant loss to cell viability within this range. At
0.033%, the concentration at which the combination of phlorizin and L-citrulline is
synergistic with L-histidine, no cell toxicity is observed.
[00151] Overall, the present disclosure provides a composition of phlorizin, L-
citrulline and L-histidine, which exhibits a surprising and unexpected synergistic
effect on collagen protein expression and thus the skin aging process. This apparent
synergism of the said combination is not readily apparent at all concentration
combinations (as discussed previously), suggesting that the results are not a result of
mere admixture of the three or routine in nature. Further, the identification of this
novel combination to reduce skin again cannot be arrived at without undue
experimentation, and is not obvious to a person skilled in the art.

[00152]Although the subject matter has been described with reference to specific embodiments, this description is not meant to be construed in a limiting sense. Various modifications of the disclosed embodiments, as well as alternate embodiments of the subject matter, will become apparent to persons skilled in the art upon reference to the description of the subject matter. It is therefore contemplated that such modifications can be made without departing from the spirit or scope of the present subject matter as defined.

1. A composition comprising:
(a) phlorizin;
(b) L-citrulline; and
(c) L-histidine,
wherein phlorizin to L-citrulline to L-histidine w/w ratio in said composition is in the range of 1:1:0.8- 1:1.3:5.
2. The composition as claimed in claim 1, wherein phlorizin to L-citrulline to L-histidine w/w ratio is 1:1:2.4.
3. The composition as claimed in claim 1, wherein
(a) phlorizin weight percentage in said composition is in the range of 0.02-0.1%;
(b) L-citrulline weight percentage in said composition is in the range of 0.02-0.1%;
and
(c) L-histidine weight percentage in said composition is in the range of 0.01-0.1%).
4. The composition as claimed in any of the claims 1-3, wherein:
(a) phlorizin weight percentage in said composition is 0.0165%>;
(b) L-citrulline weight percentage in said composition is 0.0165%>; and
(c) L-histidine weight percentage in said composition is 0.039%).
5. The composition as claimed in claim 1, further comprising at least one carrier; at
least one diluent; and at least one excipient.
6. A method of preparing a composition as claimed in claim 1, said method
comprising:
(a) obtaining phlorizin;
(b) obtaining L-citrulline;
(c) obtaining L-histidine; and
(d) contacting phlorizin, L-citrulline and L-histidine to obtain said composition.

7. A method of preparing a composition as claimed in claim 1, said method
comprising:
(a) obtaining phlorizin;
(b) obtaining L-citrulline;
(c) obtaining L-histidine;
(d) obtaining at least one carrier, at least one diluent and at least one excipient; and contacting phlorizin, L-citrulline, L-histidine and at least one carrier, at least one diluent and at least one excipient to obtain said composition.
8. A method of inhibiting skin aging, said method comprising:
(a) obtaining a composition as claimed in claim 1; and
(b) contacting said composition with skin.
9. The composition as claimed in any of the claims 1-5, wherein said composition
inhibits appearance of skin aging by enhancing collagen synthesis.