Experience The Magic of Science. DermaPep UL. Multi-functional whitening active. Experience the magic of science
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1 Experience The Magic of Science Multi-functional whitening active DermaP ep Experience the magic of science
2 Anti-aging DermaPep A35 DermaPep A42 DermaPep A44 DermaPep A53 Whitening DermaPep A35 DermaPep UL DermaPep W22 DermaPep W411 Anti-inflammatory DermaPep A35 DermaPep A44 DermaPep A53 DermaPep UL DermaPep W22 DermaPep W411
3 Multi-functional agent for skin whitening Introduction Alpha-MSH, melanocyte stimulating hormone, is best known as a major factor in the regulation of pigmentation through its unique mechanism. Alpha- MSH involves in the control of tyrosinase activity, melanin synthesis and melanosome transfer. Methyl undecenoyl leucinate interacts with MC1R, a receptor for alpha-msh, competes against its natural ligand, alpha-msh and thereby prevents any further activation of melanogenic genes such as tyrosinase, TRP1, TRP2 (DCT), MITF and POMC and finally blocks melanin synthesis. In vitro and in vivo study results show that Methyl undecenoyl leucinate is far more effective than other known whitening ingredients such as kojic acid, arbutin and hydroquinone. Function Whitening for face and body Inhibition of melanogenic pathway Providing exceptional whitening benefits Anti-inflammation effect against environmental stimulus such as UV irradiation Applications can be incorporated in cosmetic formulations such as emulsions, oily sera, gels and creams for skin-whitening and anti-inflammatory purposes. Formulation Dermatological tolerance : Standard testing has been performed on which has showed neither cytotoxic effects nor any irritation or sensitization reaction in healthy volunteers with an occlusive single patch test. Recommended concentration : 2-4 % Product Information Appearance : Transparent solution INCI/CTFA-Declaration :, (and) Glycerin (and) Butylene Glycol Active ingredient content : 1 % (CAS No ) Purity : 95 % up Preservative : None
4 DCT TRP1 Tyrosine metabolism Melanosome TYR Mechanism of whitening action General Melanogenesis Process Alpha-melanocyte stimulating hormone ( -MSH) through the cyclic AMP pathway, plays a pivotal role in melanocyte differentiation and in the regulation of melanogenesis. On binding to the MC1R, one of the five subtypes of G-protein coupled receptors, -MSH activates adenylate cyclase (AC) which, in turn, causes an increase in intracellular camp. This cascade involves the activation of protein kinase A (PKA) and CREB transcription factor, leading to the up-regulation of the expression of Microphthalmia associated transcription factor (MITF). MITF then binds and activates melanogenic gene promoters, thereby increasing their expression which results in an elevated melanin synthesis., MC1R antagonist Methyl undecenoyl leucinate binds strongly the MC1R receptor, thereby antagonizing the -MSH effects. Through the cyclic AMP pathway, Methyl undecenoyl leucinate inhibits the -MSH-induced AC and PKA activation, down-regulates melanogenic gene expressions such as MITF, tyrosinase, TRP-1 and TRP-2 (DCT) and finally suppresses melanin synthesis., Mechanism of Whitening Action Affinity for -MSH receptor Inhibition of camp Inhibition of tyrosinase Inhibition of melanogenesis Anti-inflammatory effect Mechanism of Whitening Action MSH Melanin Synthesis MC1R AC Ca 2 camp tyrosine PKA MITF TYR TRP1 DCT melanin
5 Cell viability Safety Cell Cytotoxicity Test Human primary melanocyte HEMn-LP cells were seeded into 96-well plates (2 x 1 3 ) and treated with different concentrations of Methyl undecenoyl leucinate for 48 hours. Cell viability was assessed by MTT assay and was shown relative to untreated control. Absorbance was measured by an ELISA reader at 54 nm. Skin Irritation Test Skin irritation test was performed on 31 healthy Asian volunteers (Spincontrol Asia (study report number IR- 6Q1-MW-DE8), Bangkok, Thailand). Evaluation and grading of skin irritation Erythema (no redness = to very strong redness = 5) After 24 hrs, mean score =. (max = ) After 48 hrs, mean score =. (max = ) Oedema (no oedema = to very strong oedema = 5) After 24 hrs, mean score =. (max = ) After 48 hrs, mean score =. (max = ) Scaling (no scaling = to very strong scaling = 5) After 24 hrs, mean score =. (max = ) After 48 hrs, mean score =. (max = ) In vitro Ocular Irritation Test at 3 different concentrations (2, 4, 8 %) and butylene glycol with glycerin were evaluated with the ocular irritection assay system (CA, USA). MTT Assay The ocular results demonstrated that the DermaPep TM UL was classified as mild irritants as well as butylene glycol with glycerin Concentration
6 Melanin contents Melanin contents In vitro efficacy Inhibition of Melanogenesis Melanoma B16F1 or human primary melanocyte HEMn-LP cells were pre-treated with arbutin, kojic acid or various concentrations of Methyl undecenoyl leucinate for 3 minutes and then stimulated with or without 1 ppm α-msh. After 48 hours, the cells were washed with PBS and dissolved in 1 l of 2 N NaOH containing 5 % DMSO at 6 C and absorbance was measured at 475 nm using an ELISA reader. Methyl undecenoyl leucinate inhibits melanin synthesis stimulated by α-msh. In addition, Methyl undecenoyl leucinate shows far better depigmenting effect than other known whitening agents such as arbutin and kojic acid. Inhibition of Melanogenesis A B C D B16F1 A : Not Treated Control B : -MSH C : Undecylenoyl Phenylalanine 3 ppm -MSH D : Methyl undecenoyl leucinate 3 ppm -MSH A1 A2 A3 A4 A1 : Not Treated Control A2 : -MSH A3 : Methyl undecenoyl leucinate 15 ppm -MSH A4 : Methyl undecenoyl leucinate 3 ppm -MSH Melanin Contents Assay -MSH Arbutin Kojic acid B16F1-78 % Arbutin Kojic acid % HEMn-LP - -MSH -MSH * Arbutin 2 M ( 5.44 ppm) Kojic acid 2 M ( ppm)
7 Tyrosinase activity Tyrosinase activity In vitro efficacy Inhibition of Tyrosinase Activity Tyrosinase activity was examined in melanoma B16F1 and human primary melanocyte HEMn-LP cells. After treatment with arbutin, kojic acid or various concentrations of Methyl undecenoyl leucinate, the cells were stimulated with or without 1 ppm α-msh. After 72 hours, cells were removed from culture dishes and washed with PBS. Then, the cells were lysed and the L-Dopa oxidation activity of tyrosinase was measured at 492 nm by spectrophotometer. Methyl undecenoyl leucinate inhibits endogenous tyrosinase activity induced by α-msh. The efficacy of Methyl undecenoyl leucinate is far better than that of the reference ingredients. Tyrosinase Activity Assay B16F1-57 % % HEMn-LP - -MSH -MSH 13 -MSH Arbutin Kojic acid Arbutin Kojic acid * Arbutin 2 M ( 5.44 ppm) Kojic acid 2 M ( ppm)
8 Inhibition of camp In vitro efficacy Comparison of IC 5 value IC 5 represents the concentration of actives that reduces the cellular melanin content to the half of that of non-treated control. Methyl undecenoyl leucinate is 2 times stronger whitening agent than arbutin and kojic acid. Inhibition of Cellular camp Level To examine the inhibitory effect of Methyl undecenoyl leucinate on camp activation, the intracellular level of camp was measured by direct ELISA. Melanoma B16F1 cells were exposed to arbutin, kojic acid, undecylenoyl phenylalanine or Methyl undecenoyl leucinate for 2 min and were lysed. The intracellular camp was measured at 45 nm by spectrophotometer. Methyl undecenoyl leucinate blocks camp production which is direct down-steam target of MC1R. IC5 Value camp Level (ELISA) 15 ppm Und-Phenylalanine 3 ppm Kojic acid Arbutin 28 ppm 3 ppm % -1% % -2% -3% -4% -32% -3% -37% -29%
9 Melanin contents In vitro efficacy Methyl undecenoyl leucinate, as a MC1R-specific Antagonist To determine whether Methyl undecenoyl leucinate is a MC1R-specific antagonist, B16F1 were activated with several melanogenenic stimulators. -MSH, forskolin, 8-bromo-cAMP, PMA, and 8-bromo-cGMP were used for activating MC1R, AC, camp, PKC and PKG respectively. After 48 hours of incubation with 3ppm of undecylenoyl phenylalanine or Methyl undecenoyl leucinate, the melanin contents were measured by an ELISA reader. Melanins produced only through the activation by - MSH are significantly suppressed by Methyl undecenoyl leucinate. Down-Regulation of Melanogenic Genes Melanoma B16F1 cells were treated with various concentrations of undecylenoyl phenylalanine or Methyl undecenoyl leucinate and then stimulated with or without 1ppm -MSH. After 48 hours, total cellular RNA was extracted and reverse-transcription PCR was performed to determine tyrosinase, POMC, MITF, TRP1 and TRP2. -actin mrna level was used for sample standardization. Methyl undecenoyl leucinate significantly downregulates the expression of melanogenic genes. Signaling Study ISA) Melanogenic Genes Expression (RT-PCR) -MSH (1ppm) 6 5 NT Undphenylalanine Tyrosinase 3 2 POMC 1 MITF NT -MSH Forsk olin 8-brcAMP 8-brcGMP PMA TRP1 TRP2 Control Und-phenylalanine -actin
10 IL-1 expression IL-1 expression In vitro efficacy Methyl undecenoyl leucinate, as an anti-inflammatory agent Proinflammatory Cytokines Level (RT-PCR) Human keratinocyte HaCaT cells were treated with Methyl undecenoyl leucinate and then stimulated with or without UVB 75 mj/cm 2. After 48 hours, total cellular RNA was extracted and reverse-transcription PCR was performed to determine IL-1, IL-1β, IL-6 and IL-8. GAPDH mrna level was used for sample standardization. NT UVB 75mJ/cm 2 Methyl Undecenoyl 15 3 IL-1 The production of IL-1 and IL-6 were determined by enzyme-linked immunosorbent assay (ELISA). Human keratinocyte HaCaT cells were pre-treated with Methyl undecenoyl leucinate or undecylenoyl phenylalanine and then stimulated with or without UVB 75 mj/cm 2. After 48 hours, cell lysates were transferred into immuno-well plates and analyzed. The absorbance was measured at 45 nm by an ELISA reader. IL-1β IL-6 IL-8 Gapdh Methyl undecenoyl leucinate significantly inhibits UVinduced expression and production of proinflammatory cytokines. Proinflammatory Cytokines Level (ELISA) HaCaT -139 % HaCaT -246 % UVB -MSH - UVB -MSH - Und-phenylalanine 15 3 Und-phenylalanine
11 In vivo efficacy Whitening Efficacy on Asian Skin Type In a clinical study with 3 Asian healthy female volunteers (Bangkok), aged 18 t 52, was found to have a significant skin-lightening effect..3 % formula containing has been applied twice daily during 8 weeks on the face area. Evaluation was performed by chromameter (Chromameter CR-3, Minolta, Japan). The degree of whitening effect was estimated by following parameters : L* = Luminosity (Lightness) ITA = Arc tan ((L*-5)/b*) 18π Comparison to Other Whitening Ingredients L* parameter evolution in comparison to D DermaPep UL α- arbutin.6 Kojic acid.2 - arbutin.26 Hydro quinone Increase of L* value in 8% of volunteers Increase of L* and ITA indicates a decrease in skin pigmentation. Lightening Effect of Skin Tone L* parameter evolution in comparison to D Increase of L* value in 8% of volunteers ITA % variation in comparison to D Increase of ITA value in 8% of volunteers %.4 D D56.2 D.46 D28 D L* is luminosity which represents relative brightness from total darkness (L*=) to absolute white (L*=1) A whitening agent must increase Individual Typological Angle ITA parameter.
12 Personal Care Ingredients Business Unit dermapep@mwc.co.kr Last revised on Jan 17, 213 MIWON COMMERCIAL CO., LTD MONGRAE-DONG, DANWON-GU, ANSAN-SHI, GYEONGGI, 425-1, KOREA PHONE FAX
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