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 xanthohumol and N6-benzyladenine and the uses thereof to address the problem of skin aging.
BACKGROUND OF THE INVENTION
[002] Skin aging is a complex biological process marked by characteristic features of thinning, fine wrinkling, and reduced elasticity. Both intrinsic as well as extrinsic factors are responsible for the process. Intrinsic factors include the natural course of aging along with the genetic predisposition, whereas extrinsic factors are mainly concerned with environmental factors and personal lifestyle. Dermal matrix which is continuously subjected to degradation and renewal cycle majorly constitutes of collagen. The protein is widely distributed throughout the body and provides strength and structure to the skin. In due course of time the accumulation of the degraded collagen fragments not only results in prevention of neocollagenesis (synthesis of new collagen) but also deteriorates the skin by accelerating the degradation process. One of the study reveals that the overall collagen content per unit area of the skin surface is known to decline approximately 1%/year (Shuster S. et al. Br J Dermatol. 1975; 93:639-43.)
[003] Various efforts are being continuously made towards developing strategies that involves the exploitation of plant metabolites for impeding the process of skin aging. According to a review by Ram Kumar Sahu et al. secondary plant metabolites such as quercetin, polyphenol, flavonoid and flavonol, ellagic acids, provoke the free radical scavenging activity, antagonize the UV signalling pathway, inhibition of elastase activity and matrix metalloproteinase expression, and increase of expression of procollagen type 1.

[004] WO2009126320A1 features a composition comprising xanthohumol for
treatment of skin disorders and diseases.
[005] US5371089A describes compositions containing 6- (substituted amino) purine
cytokinins for countering the effects of skin aging.
[006] Although advances have been made in the direction of reducing the effects of
skin aging there still remains the scope of discovering and identifying compositions
that result in better prevention, regeneration and delaying of the process of skin aging.
SUMMARY OF THE INVENTION
[007] In an aspect of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8.
[008] In an aspect of the present disclosure, there is provided a method of preparing
a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method
comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c)
contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein
xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of
1:2-1:8.
[009] 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)
xanthohumol; and (ii) N-6 benzyladenine, wherein xanthohumol to N-6 benzyladenine
w/w ratio is in a range of 1:2-1:8; and (b) contacting said composition with skin,
wherein said composition inhibits skin aging.
[0010] 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
[0011] 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.
[0012] Figure 1 shows the effect of the combination of xanthohumol (A), and N-6 benzyladenine (K) on the expression of collagen proteins at a 1:6.8 w/w ratio of xanthohumol to N-6 benzyladenine, in accordance with an embodiment of the present disclosure.
[0013] Figure 2 shows the effect of the combination of xanthohumol (A), and N-6 benzyladenine (K) on the expression of collagen proteins at a 1:1.3 w/w ratio of xanthohumol to N-6 benzyladenine, in accordance with an embodiment of the present disclosure.
[0014] Figure 3 shows the cytotoxic effects of xanthohumol at w/w concentrations ranging from 0.0000125% to 0.0016%, in accordance with an embodiment of the present disclosure.
[0015] Figure 4 shows the cytotoxic effects of N-6 benzyladenine at w/w concentrations ranging from 0.00004% to 0.001%, in accordance with an embodiment of the present disclosure.
DETAILED DESCRIPTION OF THE INVENTION
[0016] 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
[0017] 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.
[0018] 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.
[0019] 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".
[0020] 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.
[0021] The term "including" is used to mean "including but not limited to".
"Including" and "including but not limited to" are used interchangeably.
[0022] Xanthohumol is a prenylated chalconoid found in hops and beer. It is a
bioactive flavonoid molecule with strong antioxidant properties. Consumption of
xanthohumol by diabetic animals consistently decreases inflammation and oxidative
stress allowing neovascularization control and improving diabetic wound healing
(Raquel Costa et al.; J.Nat.Prod.76(ll): 2047-2053, 2013).
[0023] N-6 benzyladenine is a first generation synthetic cytokinin that elicits plant
growth and development responses. It shows antioxidant properties and is a

potentially useful agent in the treatment and prevention of many diseases connected with oxidative stress in skin (Jablonska et al. Mol Cell Biochem, 2016, 413: 97-107). [0024] Carriers are substances that serve as mechanisms to improve the delivery and the effectiveness of the drug.
[0025] A diluent (also referred to as filler, dilutant, or thinner) is a diluting agent. [0026] An excipient is an inactive substance that serves as a vehicle or medium for a drug or other active substance. Excipients include colouring agents, humectants, preservatives, emollients and combinations thereof.
[0027] 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.
[0028] Skin aging is a complex process leading to biologic debilitation at tissue and cellular level, that is exhibited in many ways. Over a period of time due to the senescence visible effects of aging, such as sagging and wrinkling, are observed. One of the major proteins responsible for skin elasticity is collagen which undergoes continuous degeneration due to release of MMPs (matrix metalloproteinases). The composition disclosed herein provides a solution to address skin aging by promoting collagen synthesis using a composition of xanthohumol and N-6 benzyladenine. It provides a formulation and a method to prepare the same.
[0029] 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.

[0030] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is in the range of 1:2- 1:8.
[0031] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is in the range of 1:6.5- 1:7.1.
[0032] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is 1:6.8
[0033] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol
weight percentage in said composition is in the range of 0.00001%-0.00008%; N-6
benzyladenine weight percentage in said composition is in the range of 0.00025%-
0.00038%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in
the range of 1:2-1:8.
[0034] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol
weight percentage in said composition is in the range of 0.00001%>-0.00008%), N-6
benzyladenine weight percentage in said composition is in the range of 0.00025%>-
0.00038%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in
the range of 1:6.5-1:7.1.
[0035] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol
weight percentage in said composition is in the range of 0.00001%>-0.00008%), N-6
benzyladenine weight percentage in said composition is in the range of 0.00025%>-
0.00038%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is
1:6.8.

[0036] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol weight percentage in said composition is 0.00005%, N-6 benzyladenine weight percentage in said composition is 0.00034%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0037] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8. [0038] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-l :7.1. [0039] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0040] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%), N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%)-0.00038%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8.
[0041] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%>-0.00008%), N-6

benzyladenine weight percentage in said composition is in the range of 0.00025%-0.00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1.
[0042] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%), N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%)-0.00038%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0043] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is 0.00005%), N-6 benzyladenine weight percentage in said composition is 0.00034%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0044] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8.
[0045] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1.

[0046] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8. [0047] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-0.00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8.
[0048] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%), N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-0.00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5:1:7.1.
[0049] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%)-0.00008%), N-6 benzyladenine weight percentage in said composition is in

the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0050] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; and (c) contacting xanthohumol, and N-6 benzyladenine to obtain said composition, wherein xanthohumol weight percentage in said composition is 0.00005%, N-6 benzyladenine weight percentage in said composition is 0.00034%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0051] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8.
[0052] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1.

[0053] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0054] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition, wherein xanthohumol w/w percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8. [0055] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition, wherein xanthohumol weight percentage in said composition is in the

range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in the said composition is in the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-l :7.1. [0056] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0057] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient, said method comprising: (a) obtaining xanthohumol; (b) obtaining N-6 benzyladenine; (c) obtaining the at least one carrier; (d) obtaining the at least one diluent; (e) obtaining the at least one excipient; and (f) contacting xanthohumol, N-6 benzyladenine, the at least one carrier, the at least one diluent and, the at least one excipient to obtain said composition, wherein xanthohumol weight percentage in said composition is 0.00005%), N-6 benzyladenine weight percentage in said composition is 00034%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8. [0058] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to

obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1.2-1:8.
[0059] In an embodiment of the present disclosure, there is provided a method of preparing a composition to comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1.6.5-1:7.1.
[0060] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0061] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8. [0062] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said

composition is in the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-l :7.1. [0063] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8.
[0064] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein xanthohumol weight percentage in said composition is 0.00005%, N-6 benzyladenine weight percentage in said composition is 0.00034%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8. [0065] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein said solution A is prepared in an organic solvent. [0066] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein said solution A is prepared in an inorganic solvent.

[0067] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein said solution B is prepared in an organic solvent. [0068] In an embodiment of the present disclosure, there is provided a method of preparing a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine, said method comprising: (a) obtaining a solution A of xanthohumol; (b) obtaining a solution B of N-6 benzyladenine; and (c) contacting solution A and solution B to obtain said composition, wherein said solution B is prepared in an inorganic solvent. [0069] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said composition inhibits skin aging.
[0070] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio is in the range of 1:6.5-1:7.1, and said composition inhibits skin aging.
[0071] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol; and (ii) N-6 benzyladenine; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging.
[0072] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol having a weight percentage in said composition in the range of

0.00001%-0.00008%; and (ii) N-6 benzyladenine having a weight percentage in said composition in the range of 0.00025%-00038%; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-l :8, and said composition inhibits skin aging. [0073] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol having a weight percentage in said composition in the range of 0.00001%-0.00008%; and (ii) N-6 benzyladenine having a weight percentage in said composition in the range of 0.00025%-00038%; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-l :7.1, and said composition inhibits skin aging. [0074] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol having a weight percentage in said composition in the range of 0.00001%-0.00008%>; and (ii) N-6 benzyladenine having a weight percentage in said composition in the range of 0.00025%>-00038%>; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging.
[0075] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol having a weight percentage in said composition of 0.00005%>; and (ii) N-6 benzyladenine having a weight percentage in the said composition of 0.00034%>; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6. 8, and said composition inhibits skin aging.
[0076] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent;

and (v) at least one excipient; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said composition inhibits skin aging.
[0077] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1, and said composition inhibits skin aging.
[0078] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol; (ii) N-6 benzyladenine; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging.
[0079] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%; (ii) N-6 benzyladenine weight percentage in said composition is in the range of 0.000025%-0.000038%; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said composition inhibits skin aging.
[0080] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%>; (ii) N-6 benzyladenine weight percentage in said composition is in the range of 0.000025%>-0.000038%>; (iii) at least one carrier; (iv) at least one diluent; and

(v) at least one excipient, and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1, and said composition inhibits skin aging.
[0081] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%; (ii) N-6 benzyladenine weight percentage in said composition is in the range of 0.000025%-0.000038%; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging.
[0082] In an embodiment of the present disclosure, there is provided a method of inhibiting skin aging comprising: (a) obtaining a composition comprising: (i) xanthohumol weight percentage in said composition is 0.00005%; (ii) N-6 benzyladenine weight percentage in said composition is 0.000034%; (iii) at least one carrier; (iv) at least one diluent; and (v) at least one excipient; and (b) contacting said composition with skin, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging.
[0083] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said composition inhibits skin aging by enhancing collagen synthesis. [0084] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1, and said composition inhibits skin aging by enhancing collagen synthesis. [0085] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-

6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits
skin aging by enhancing collagen synthesis.
[0086] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol having a weight percentage in said composition in the
range of 0.00001%-0.00008%; and (b) N-6 benzyladenine having a weight percentage
in said composition in the range of 0.00025%-0.00038%, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said
composition inhibits skin aging by enhancing collagen synthesis.
[0087] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol having a weight percentage in said composition in the
range of 0.00001%-0.00008%; and (b) N-6 benzyladenine having a weight percentage
in said composition in the range of 0.00025%-0.00038%, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1, and said
composition inhibits skin aging by enhancing collagen synthesis.
[0088] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol having a weight percentage in said composition in the
range of 0.00001%-0.00008%; and (b) N-6 benzyladenine having a weight percentage
in said composition in the range of 0.00025%-0.00038%, wherein xanthohumol to N-
6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits
skin aging by enhancing collagen synthesis.
[0089] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol having a weight percentage in said composition of
0.00005%>; and (b) N-6 benzyladenine having a weight percentage in said composition
of 0.00034%), wherein xanthohumol to N-6 benzyladenine w/w ratio in said
composition is 1:6.8, and said composition inhibits skin aging by enhancing collagen
synthesis.
[0090] In an embodiment of the present disclosure, there is provided a composition
comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at

least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said composition inhibits skin aging by enhancing collagen synthesis. [0091] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1, and said composition inhibits skin aging by enhancing collagen synthesis. [0092] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging by enhancing collagen synthesis.
[0093] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol having a weight percentage in said composition in the range of 0.00001%-0.00008%; (b) N-6 benzyladenine having a weight percentage in said composition in the range of 0.00025%-0.00038%; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:2-1:8, and said composition inhibits skin aging by enhancing collagen synthesis. [0094] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol having a weight percentage in said composition in the range of 0.00001%-0.00008%; (b) N-6 benzyladenine having a weight percentage in said composition in the range of 0.00025%-0.00038%; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.5-1:7.1, and said composition inhibits skin aging by enhancing collagen synthesis.

[0095] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol having a weight percentage in said composition in the range of 0.00001%-0.00008%; and (b) N-6 benzyladenine having a weight percentage in said composition in the range of 0.00025%-0.00038%; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging by enhancing collagen synthesis.
[0096] In an embodiment of the present disclosure, there is provided a composition comprising: (a) xanthohumol weight percentage in said composition is 0.00005%; (b) N-6 benzyladenine weight percentage in said composition is 0.00034%; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8, and said composition inhibits skin aging by enhancing collagen synthesis. [0097] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1.2-1.8.
[0098] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1.6.5-1.7.1.
[0099] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said composition comprising: (a) xanthohumol; and (b) N-6 benzyladenine, wherein xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.8.
[00100] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said method comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one

excipient, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%, N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-0.00038%; and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1.2-1.8.
[00101] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said method comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%-0.00008%), N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-0.00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1.6.5-1.7.1.
[00102] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said method comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is in the range of 0.00001%)-0.00008%), N-6 benzyladenine weight percentage in said composition is in the range of 0.00025%-0.00038%, and xanthohumol to N-6 benzyladenine w/w ratio in said composition is in the range of 1:6.8.
[00103] In an embodiment of the present disclosure, there is provided a composition for use in preparing formulations, said method comprising: (a) xanthohumol; (b) N-6 benzyladenine; (c) at least one carrier; (d) at least one diluent; and (e) at least one excipient, wherein xanthohumol weight percentage in said composition is 0.00005%), N-6 benzyladenine weight percentage in said composition is 0.00034%), and xanthohumol to N-6 benzyladenine w/w ratio in said composition is 1:6.8. [00104] In an embodiment of the present disclosure, there is provided a composition as described herein, wherein said composition can be for topical application. [00105] 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.

[00106] 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. [00107] 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. [00108] 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. [00109] 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
[00110] 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
[00111] To determine percentage collagen release in co-culture model: Normal Human dermal fibroblasts (Lonza) were seeded in 12- well plates (Costar) in Dulbecco's Modified Eagle's Medium High Glucose (DMEM-Gibco) medium containing 10% fetal bovine serum (Gibco). The culture was incubated at 37°(7 5% CO2 incubator and was allowed to reach 100% confluency. HaCaT keratinocytes (NCCS - India) at a density of 0.9x105cells per ml were seeded on the fibroblast bed in 154CF medium (Gibco-Ml 54500) containing 0.03mM calcium chloride. Post 24 hrs, medium was replenished with specific concentrations of xanthohumol and N-6 benzyladenine and incubated in 37°(75% CO2 incubator. Cells were harvested after 48 hrs for total collagen estimation using Sircol assay from Biocolor life sciences, UK. All the steps were performed as per the manufacturer's protocol. [00112] To determine the cytotoxicity: Normal human Dermal fibroblasts were seeded onto a 96 well plate (Costar) in Dulbecco's Modified Eagle's Medium supplemented with 10% FBS. The plate was incubated in a 37°(75% CO2 incubator and was allowed to reach 85% confluency post which the medium was replaced with serum free DMEM medium for serum starvation for another 24 hours. The media was then replenished with different concentrations of xanthohumol and N-6 benzyladenine individually and plates were incubated in 37°(75% CO2 incubator for 48hours. Post 48 hours, cells were washed with IX PBS and WST reagent (prepared in DMEM media in a ratio of 1:10) was added to each of the wells. Plates were incubated in 37°(75% CO2 incubator for 2 hours for colour development. After 2 hours of incubation the plate was read at 450nm against blank (only media). The percentage viability of the cells was calculated relative to the control wells that were treated with vehicle. [00113] Xanthohumol from Humulus Lupulus (CAS No. 6754-58-1) and N-6 benzyladenine (CAS no. 1214-39-7) were procured from Sigma Aldrich. [00114] Xanthohumol(A) and N-6 benzyladenine(K) were prepared in DMSO.

Example 2
Effect of combination of xanthohumol and N-6 benzyladenine on collagen
protein expression
[00115] Xanthohumol(A) and N-6 benzyladenine(K) were tested individually and in combination at different concentrations to determine the effect on collagen expression. The protocol followed for the experiment was same as mentioned in Example 1. The concentrations tested are mentioned in the table below:

[00116] When the individual actives were tested individually at a concentration of 0.00005% xanthohumol and 0.00034% N-6 benzyladenine, only basal level of collagen protein production was observed and there was no significant increase in the level of collagen protein and at these concentrations. However, a remarkable increase of greater than 20% was observed when the actives were tested at the same concentrations but in combination [(AK) 0.00005%+0.00034%]. Thus, xanthohumol and N-6 benzyladenine show distinct synergy at the concentrations of 0.00005%) xanthohumol and 0.00034% N-6 benzyladenine, in combination. This represents a w/w ratio of 1:6.8.
[00117] Additionally, the synergistic effect of xanthohumol and N-6 benzyladenine in enhancing the levels of collagen is only apparent at this ratio, as there was no increase in the levels of collagen when the AK was tested in a w/w ratio of 1:1.3 (0.00015%)+0.00017%)) (Figure 2). Thus, a mere admixture of the two actives does not result in a synergistic enhancement of collagen levels, and therefore this combination of xanthohumol and N-6 benzyladenine cannot be arrived at without undue experimentation.

Example 3
Cytotoxicity of xanthohumol and N-6 benzyladenine.
[00118] Figure 3 and 4 depicts the cytotoxic effects of different concentrations of
xanthohumol and N-6 benzyladenine respectively on Normal Human Dermal
Fibroblasts.
[00119] According to Figure 3 xanthohumol is toxic to the cells in a concentration
range of 0.0016%-0.0002%. However, 100% viability was observed at concentrations
equal to and lower than 0.0001%. It was thus confirmed that 0.00005%) of
xanthohumol is non-toxic to the cells.
[00120] According to Figure 4 N-6 benzyladenine did not show any cytotoxic effect
across the range of concentration tested (0.001%>-0.00004%)) thus confirming that a
concentration of 0.00034 can be employed safely for the composition.
[00121] Thus, the present disclosure provides a novel and synergistic composition of
xanthohumol and N-6 benzyladenine, which exhibits an enhancement in the level of
collagen protein at a 1:6.8 ratio of xanthohumol to N-6 benzyladenine. This
combination can thus aid in the inhibition of 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 two 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.

I/We Claim:
1. A composition comprising:
(a) xanthohumol; and
(b) N6- benzyladenine,
wherein xanthohumol to N6- benzyladenine w/w ratio in said composition is 1:2- 1:8.
2. The composition as claimed in claim 1, wherein xanthohumol to N6-benzyladenine w/w ratio in said composition is 1:6.5- 1:7.1.
3. The composition as claimed in any of the claims 1-2, further comprising:

(a) at least one carrier;
(b) at least one diluent; and
(c) at least one excipient.
4. A method of preparing a composition as claimed in any of the claims 1-2, said
method comprising:
(a) obtaining xanthohumol;
(b) obtaining N6- benzyladenine; and
(c) contacting xanthohumol and N6- benzyladenine to obtain said composition.
5. A method of preparing a composition as claimed in any of the claims 1-3, said
method comprising:
(a) obtaining xanthohumol;
(b) obtaining N6- benzyladenine;
(c) obtaining the at least one carrier;
(d) obtaining the at least one diluent;

(e) obtaining the at least one excipient; and
(f) contacting xanthohumol, N6- benzyladenine, the at least one carrier, the at least one diluent, and the at least one excipient to obtain said composition.
6. A method of inhibiting skin aging, said method comprising:
(a) obtaining a composition as claimed in any of the claims 1-3; and
(b) contacting said composition with skin, wherein said composition inhibits skin aging.
7. The composition as claimed in any of the claims 1-3, wherein said composition
inhibits skin aging by enhancing collagen synthesis.