hereinafter , the present invention will be described in further detail with reference to examples . it is to be understood , however , that these examples are illustrative only and the scope of the present invention is not limited thereto . 120 mice were divided into 6 groups , each consisting of 20 mice . group 1 was untreated as a control , group 2 was treated only with 12 - o - tetradecanoylphorbol - 13 - acetate ( tpa ) as a positive control , group 3 was treated with physiogel ™ containing physiologic lipid mixture ( manufactured by “ a ” company ) as a comparative example , and groups 4 , 5 and 6 were treated with multi - lamellar emulsion ( mle ), mle containing aforementioned pc - 9s ( 0 . 6 %) ( mle / pc - 9s 1 ) and mle containing aforementioned pc - 9s ( 1 . 2 %) ( mle / pc - 9s 2 ), respectively . as used herein , the term multi - lamellar emulsion ( mle ) means the emulsions with multi - layered structures and expresses the “ maltese cross ” structure under cross - polarized microscope . first , mice were sensitized by topically applying tpa ( 1 μg / 20 μl acetone ) to the ear . at 6 days from the next day , 20 μl of tpa was topically applied to the same site of mouse ear . at 1 hour and 8 hours after application of tpa , 20 μl of each of physiogel ™, mle , mle / pc - 9s 1 and mle / pc - 9s 2 were topically applied to the same site in which tpa has been applied . at day 7 , each of physiogel ™, mle , mle / pc - 9s 1 and mle / pc - 9s 2 was topically applied to the mouse ear at 1 hour after application of 20 μl tpa . after 3 hours , the ear thickness of the mice and the number of mast cells in the mouse ear were measured , and the measurement results are shown in fig1 and 2 . as can be seen in fig1 and 2 , the compound according to the present invention showed significant anti - inflammatory effects in the tpa - induced inflammation model , compared with negative control groups and physiogel ™ applied groups . 140 hairless mice ( skh ) were divided into 7 groups , each consisting of 20 mice . group 1 was untreated as a control , group 2 was treated with acetone , group 3 was treated with the ppar - α activator wy14 , 643 , group 4 was treated with physiogel ™ containing physiologic lipid mixture ( manufactured “ a ” company ) as a comparative example , and groups 5 , 6 and 7 were treated with mle , mle / pc - 9s 1 and mle / pc - 9s 2 , respectively . 100 μl of 1 % oxazolone ( 4 - ethoxymethylene - 2 - phenoxazol - 5 - one , oxa ) was topically applied to both flanks of the hairless mice , and after 7 days , 100 μl of 0 . 1 % oxa was topically applied to the same site at 2 - day intervals for 10 days . after atopic induction in the mice , each of the above - described treating agents was applied to the mouse groups twice a day for 4 days . during the application of each treating agent , the transepidermal water loss ( tewl ) and fold thickness of the mouse skin were measured , and the measurement results are shown in fig3 and 4 . the above results demonstrate that the topical formulations containing the compound according to the present invention shows significant effects of restoring damaged skin barrier function . also , it could be observed that the skin thickening by the inflammation became normalized by the treatment with the compound of the present invention . atopic model mice induced by oxazolone ( 4 - ethoxymethylene - 2 - phenoxazol - 5 - one , oxa ) were applied with the treating agents described in example 2 , and then the number of epidermal mast cells in the mice was measured in order to measure the inflammation - alleviating effects of the treating agents . the measurement results are shown in fig5 . as can be seen in fig5 , the compound of the present invention showed a significant inflammation - alleviating effect in the atopic model mice . in order to examine whether the results of example 2 induce the expression of antimicrobial peptides , the degrees of epidermal expression of mouse beta defensin - 3 ( mbd - 3 ) and cathelicidin - related murine antimicrobial peptide ( cramp ) were measured . the skin was collected from the treated sites of the atopic model mice tested in example 2 , and then total rna was harvested from the collected skin . in order to harvest the total rna , 1 ml of trizol reagent was added to the collected skin . after the skin tissue was allowed to react with the trizol reagent at room temperature for 15 seconds , 200 μl of chloroform was added thereto . then , the tissue was centrifuged at 13 , 000 rpm for 10 minutes , and the supernatant was transferred into another tube . then , 500 μl of isopropanol was added thereto , and the tissue solution was centrifuged again at 13 , 000 rpm for 10 minutes . the precipitated rna was washed twice with 70 % ethanol , and then diluted in triple - distilled water . the diluted rna was quantitatively analyzed at a wavelength between 260 nm and 280 nm . the rna thus obtained was subjected to reverse transcriptase polymerase chain reaction ( rt - pcr ), in order to quantitate the messenger rna expressions . for rt - pcr , 2 μl mgcl 2 , 1 μl rt buffer , 1 μl dntp mix , 0 . 25 μl rnase inhibitor , 0 . 5 μl rtase , 0 . 5 μl oligo dt , 3 . 75 μl distilled water and 2 μg rna were placed in a tube , and then subjected to an rt - pcr reaction . the rt - pcr reaction was performed at 45 ° c . for 1 hour and 95 ° c . for 5 minutes . for the qualitative analysis of gapdh ( glyceraldehyde 3 - phosphate dehydrogenase ) and ppar - α , pcr was performed using the following primers : for pcr , 12 . 5 μl pcr premix , 1 μl primer sense ( 10 μm ), 2 μl primer anti - sense ( 10 μm ), 1 . 5 μl cdna and 7 μl distilled water were placed in a tube , and then subjected to a pcr reaction . the pcr of ppar - α was performed in the following conditions : 30 cycles of 94 ° c . for 30 sec , 59 ° c . for 30 sec and 72 ° c . for 30 sec , followed by 72 ° c . for 10 minutes . after the pcr amplification , the resulting products were mixed with each other , and the final solution was loaded onto 1 . 5 % agarose gel stained with a nucleic acid - specific fluorescent compound ( such as , ethidium bromide ) allowing visualization under uv light . the sample on the gel was visualized under uv light in a dark room and photographed with a computer - coupled camera . the gel photograph was analyzed in image processing software allowing band strength to be quantified . to further clarify the results , real - time pcr was performed . 5 μl cybergreen , 2 μl primer sense ( 10 μm ), 2 μl primer anti - sense ( 10 μm ), 1 μl cdna and 1 μl distilled water were placed in a tube , and then subjected to real - time pcr . the pcr results were analyzed using a program , and the analysis results are shown in fig6 . as can be seen in fig6 , it was found that the compound according to the present invention significantly activated the antimicrobial peptides . in order to examine whether the results of example 2 induce the expression of an antimicrobial peptide , the expression of cramp in the skin was measured . the skin was collected from the treated sites of the atopic model mice prepared in example 2 , and then made into a paraffin block . the tissue was attached to a slide using a paraffin microtome , and 500 μl of peroxidase blocking reagent was loaded onto the slide , and then incubated for 30 minutes . then , the slide was washed three times with pbs buffer . 500 μl of protein blocking reagent was loaded onto the slide , and then incubated for 15 minutes . the tissue on the slide was incubated with primary goat anti - mouse cramp at 25 ° c . for 30 minutes and incubated with donkey anti - goat horse reddish peroxidase as a secondary antibody at 25 ° c . for 30 minutes . the tissue on the slide was incubated with the color development reagent dab at 25 ° c . for 10 minutes . after completion of the incubation , the expression of cramp in the horny layer of the skin was measured with a microscope . it was measured that the expression of cramp in the skin horny layer of the mice applied with the compound of the present invention significantly increased , and the measurement results are shown in fig7 . in order to examine whether the results of example 2 are attributable to the epidermal activation of ppar - α , β / δ , the activation of ppar - α , β / δ in the skin was measured using the following primers : real - time pcr was performed . for this purpose , 5 μl cybergreen , 2 μl primer sense ( 10 μm ), 2 μl primer anti - sense ( 10 μm ), 1 μl cdna and 1 μl distilled water were placed in a tube , and then subjected to real - time pcr . the pcr results were analyzed using a program , and the analysis results are shown in fig8 . as can be seen in fig8 , it was found that the compound according to the present invention activated ppar - α , β / δ . in order to examine whether the results of example 2 induce the activation of spt that promotes the in situ synthesis of ceramide , the activation of spt in the skin was measured using the following primers : real - time pcr was performed . for this purpose , 5 μl cybergreen , 2 μl primer sense ( 10 μm ), 2 μl primer anti - sense ( 10 μm ), 1 μl cdna and 1 μl distilled water were placed in a tube , and then subjected to a pcr reaction . the pcr results were analyzed using a program , and the analysis results are shown in fig9 . the obtained results revealed that the compound according to the present invention significantly activates spt . in order to examine whether the results of example 2 induce the terminal differentiation of the epidermal keratinocytes , keratinocyte differentiation in the skin was measured . the skin was collected from the treated sites of the atopic model mice prepared in example 2 , and then made into a paraffin block . the skin tissue was attached to a slide using a paraffin microtome , and 500 μl of peroxidase blocking reagent was loaded onto the slide , and then incubated for 30 minutes . then , the slide was washed three times with pbs buffer at a 5 - minute interval . 500 μl of protein blocking reagent was loaded onto the slide , and then incubated for 15 minutes . the tissue on the slide was incubated with primary anti - mouse filaggin , loricrin and involucrin at 25 ° c . for 30 minutes and incubated with donkey anti - rabbit hrp as a secondary antibody at 25 ° c . for 30 minutes . the tissue on the slide was incubated with the color development reagent dab at 25 ° c . for 10 minutes . after completion of the incubation , the expression of each of the differentiation markers in the skin horny layer was measured with a microscope . as a result , it was measured that differentiation in the skin horny layer of the mice applied with the pseudo - ceramide compound of the present invention increased , and the measurement results are shown in fig1 . in order to evaluate the safety of the ppar - α activator compound according to the present invention for application as a skin external preparation , an animal toxicity test and a human patch test were performed . to perform this evaluation , a single dose oral toxicity test using rats , a skin irritation test using rabbits , a skin sensitization test using guinea pigs and an ophthalmic mucous membrane irritation test using rabbits were performed as toxicity tests for pc - 9s in animals . those tests were performed based on “ toxicity test standards for pharmaceutical products ” disclosed by the korea food & amp ; drug administration . additionally , a human patch test for pc - 9s was carried out on 30 subjects ( average age : 24 . 1 ). as a result , as shown in table 1 below , a slight skin irritation was observed only in the skin irritation test using rabbits . however , when evaluating the overall results obtained from the other toxicity tests and the human patch test , it was considered that the pseudo - ceramide compound would not pose a safety problem . a moisturizing agent was added to purified water and heated to 70 ° c . pc - 9s and oil phase components were dissolved by heating , and an emulsifier , a preservative , or the like were added thereto , followed by heating to 70 ° c . the oil phase was added to the above aqueous phase . then , the emulsified particles were homogenized with a homomixer , followed by debubbling , filtration and cooling . pc - ps and oil phase components were dissolved by heating , and an emulsifier , a preservative , or the like was added thereto , followed by adjustment of the temperature to 70 ° c . the resultant mixture was mixed homogeneously in a homomixer , followed by debubbling , filtration and cooling . although the preferred embodiment of the present invention has been described for illustrative purposes , those skilled in the art will appreciate that various modifications , additions and substitutions are possible , without departing from the scope and spirit of the invention as disclosed in the accompanying claims .
disclosed herein is the use of a non - natural compound represented by the following formula 1 as an activator of the peroxisome proliferator activated receptor - α which shows an anti - inflammatory effect on inflammatory skin diseases . the ppar - α activator increases the expression of ppar - α in the skin to enhance various physiological effects of ppar - α , that is , the anti - inflammatory effect caused by inhibiting the activity of transcription factors such as the nuclear factor κk that induces inflammatory reactions , the effect of improving the epidermal permeability barrier function of the skin , and the effect of promoting the terminal differentiation of epidermal keratinocytes . thus , the ppar - α activator is useful for the treatment , alleviation or amelioration of skin diseases such as acne , seborrheic dermatitis , viral skin diseases , urticaria , pruritus , viral infectious diseases and contact dermatitis . wherein r 1 and r 2 are the same or different and each represents a linear or branched c 1 - 22 alkyl group , a phenyl group or a benzene group .