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Of Saururus Chinensis Extract” “Whitening, Anti-wrinkle and Anti-inflammatory effect Of Saururus chinensis Extract” Dae Woo Lee1,2, Dong Wook Kim1, Tae Seon Baek1, Han Na Oh2,3, Won Sang Seo3, Sung Kyu Choi1, Sang Yo Byun2, Jung Wook Ha1 1R&D Center, Dermalab Ltd., 231 Munmakgongdan-gil, Munmak-eup, Wonsu-si, Gangwon-do, Korea 2Cosmetic Science Major, Graduate School, Ajou University, Korea, San 5 Woncheon Dong Yeongtong Gu, Suwon, , Korea 3R&D Center, Charmzone Ltd., 58 Taejanggongdan-gil, Wonsu-si, Gangwon-do, Korea To whom correspondence should be addressed. E-mail: [email protected] Abstract In this study, in our continuing effort to develop active cosmetic ingredients from natural resource, we continuously screened about Saururus chinensis from South Korea. The alcoholic extract of Saururus chinensis whole plants and its fractions (separated with Column Chromatography) were evaluated for their effects against Whitening, Anti-wrinkle and Anti-inflammatory. Effect of Tyrosianse, MITF, MMP-1, Tropoelastin and TNF-α exhibited Isoquercetin fraction isolated isolated from Saururus chinensis whole plants. The results of this study suggest that Isoquercetin fraction from Saururus chinensis may be beneficial as whitening, Anti-wrinkle and Anti-inflammatory compound in functional cosmetics resource. Materials Results 160 140 Saururus chinensis (Saururaceae) is a perennial 140 120 Herbaceous plant that has been used in the treatment 120 100 of various diseases such as edema, jaundice, gonorrhea, 100 80 80 fever, and inflammation in Korean folk medicine. 60 60 40 40 MITF mRNA Expression(%) mRNA MITF Chemical studies of the genus Saururus have shown the presence of Expression(%) mRNA Tyrosinase 20 20 0 0 lignans, 2-6 aristolactams, flavonoids, anthraquinones, and Blank Control 1 5 10 50 Arubtin Blank Control 1 5 10 50 Arubtin 50 50 fruanoditerpenes, 7-10 some of which exhibited neuroleptic, 11 Concentration(ppm) Concentration(ppm) Fig 4. Effect of Isoquercetin fractions on the Fig 5. Effect of Isoquercetin fractions on the mRNA hepatoprotective, 12 antifeedant 13 and antioxidant activities. mRNA expression of Tyrosinase in B16F10 expression of MITF in B16F10 melanoma cells melanoma cells stimulated with α-MSH. Cells stimulated with α-MSH. Cells were pretreated were pretreated with different concentrations with different concentrations of Isoquercetin of Isoquercetin fraction and Arubutin fraction and Arubutin concentration (50ppm) Identification and isolation of Saurus chinensis concentration (50ppm) for 72 h. for 72 h. Saururus chinensis (50g) Extraction with 70% Ethanol 35℃, 24h 120 160 140 Ethanol Extract 100 (8.58%) 120 80 100 Solvent soluble fraction 60 80 (n-Hexane, CH2Cl2, EtOAc, n-BuOH) 60 mRNA Expression(%) mRNA 1 40 - 40 MMp 20 Tropoelastin mRNA Expression(%) mRNA Tropoelastin 20 n-Hexane Aqueous layer (14.45%) 0 0 Blank Control 1 5 10 50 Ade 50 Blank 1 5 10 50 Ade 50 Concentration(ppm) Concentration(ppm) CH2Cl2 Aqueous layer (22.54%) Fig 6. Effect of Isoquercetin fractions on the Fig 7. Effect of Isoquercetin fractions on the mRNA expression of MMP-1 in CCD-986sk cells mRNA expression of Tropoelastin in CCD-986sk stimulated with UV. Cells were pretreated with cells stimulated with UV. Cells were pretreated EtOAc Aqueous layer different concentrations of Isoquercetin with different concentrations of Isoquercetin (18.41% fraction and Adenosine concentration fraction and Adenosine concentration ) (50ppm). (50ppm). n-BuOH Aqueous layer (8.84%) 120 100 HPLC (C18 ODS column) 80 mRNA 60 Isoquercetin fraction α - 40 TNF Fig 1. Procedure for extraction and fractionation of Saururus chinensis. Expression(%) 20 0 C:\Xcalibur\Company\DML\20150421\S_03 4/21/2015 4:54:26 PM 1 Control 1 5 10 50 OH (A) RT: 0.00 - 74.99 Concentration(ppm) OH 5.38 58.92 NL: 100 57.74 8.23E6 4.99 68.31 7.67 61.34 69.11 TIC F: MS 13.47 21.08 51.23 HO O 65.60 S_03 9.04 18.09 24.76 70.72 50 43.80 25.21 47.52 3.63 34.21 35.36 52.84 Fig 8. Effect of Isoquercetin fractions on the mRNA expression of TNF-α in Raw 274.6 cells stimulated O-Glc 0 57.71 NL: OH O 58.87 1.77E6 with LPS(10ng/mL). Cells were pretreated with different concentrations of Isoquercetin fraction. 1500000 Channel A UV S_03 1000000 51.15 uAU 500000 20.70 0.51 4.93 11.19 16.56 21.53 28.13 33.17 35.48 43.63 47.54 56.13 63.91 67.47 69.85 0 58.92 NL: 100 57.74 5.68E6 51.28 m/z= 298.00- 50 299.00 F: 47.62 MS S_03 2.53 5.00 10.66 17.20 20.86 23.49 24.03 33.11 39.88 45.58 57.43 63.95 68.21 68.73 0 0 5 10 15 20 25 30 35 40 45 50 55 60 65 70 Conclusions Time (min) (B) S_03 #1104 RT: 13.47 AV: 1 NL: 1.74E6 T: + c Q3MS [ 150.00-500.00] 166.72 100 The results of this study suggest that Isoquercetin fraction 90 80 70 from Saururus chinensis may be beneficial as whitening, Anti- 60 50 40 wrinkle and Anti-inflammatory compound in functional Relative Relative Abundance 30 304.82 258.75 20 206.02 10 287.56 cosmetics resource. And the Saururus chinensis was 190.70 207.99 247.27 322.07 353.54 371.94 409.12 431.37 453.58 466.95 494.00 0 150 200 250 300 350 400 450 500 m/z investigated for Tyrosinase, MITF, MMP-1 mRNA expression Fig 2. The Isoquercetin fraction was purified Fig 3. Identification of HPLC LC-MS/MS of the inhibitory activity, Tropoelastin mRNA expression activity and using the C18 ODS column for EtOAc fraction (B). purified Isoquercetin fraction. Isoquercetin Standard (A). TNF-α mRNA expression inhibitory activity compound. Acknowledgement “This study was supported by the Traditonal Korean Medicine R&D Program funded by the Ministry of Health & Welfare through the Korea Health Industry Development Institute (KHIDI) (HI14C-0779-020014)” “Isoquercitrin” Fig. 2. Structure assign of compound 1 isolated from Saururus chinensis .
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