Journal of Life Science (생명과학회지)

Korean Society of Life Science (한국생명과학회)
권호 표지
DOI QR코드

DOI QR Code

Effect of Ethanolic Extracts Mixed with Grains and Fallopia multiflora on Melanogenesis

곡물과 적하수오를 혼합한 에탄올 추출물이 멜라닌 합성에 미치는 영향

  • Lee, Eunbeen (Department of Chemistry, Dong-Eui University) ;
  • Kim, Moon-Moo (Department of Applied Chemistry, Dong-Eui University)
  • Received : 2018.04.04
  • Accepted : 2019.03.04
  • Published : 2019.04.30
Download PDF
⟨ Previous Next ⟩

Abstract

The aim of this study was to investigate the effect of a mixture of ethanol extracts of Black oryzasativa, Sesamum indicum, Oryza sativa, Rhynchosia Nulubilis, and Polygoni multiflori radix (MIXEE) on melanogenesis to develop a natural product for black hair growth. An accumulation of hydrogen peroxide ($H_2O_2$) in hair follicles, which reduces melanin synthesis, is responsible for hair graying. In the present study, MIXEE showed scavenging activity against DPPH radicals and reducing power. In addition, it reduced the cellular $H_2O_2$ level, indicating that it could inhibit oxidation and promote melanin synthesis, which was decreased by $H_2O_2$. On the other hand, it did not affect tyrosinase activity in vitro but promoted the turnover of L-DOPA into melanin. MIXEE promoted melanin synthesis at the cellular level in B16F1 cells. Furthermore, MIXEE increased the expression levels of superoxide dismutase 2 (SOD2) and SOD3 in western blot analysis. In addition, MIXEE increased the expression levels of tyrosinase-related protein (TRP)-2, which promoted melanin synthesis from L-DOPA. The results suggested that MIXEE could promote melanogenesis. Therefore, MIXEE may have potential as a natural product for promotion of melanin production and reversal of gray hair to black hair.

본 연구의 목적은 흑모 성장과 관련 있는 천연물을 개발하기 위하여 흑미, 흑임자, 현미, 서목태 및 적하수오의 에탄올 추출혼합물의 melanin 합성에 대한 효능을 조사한 것이다. 흑모가 백모로 변화되는 원인으로 $H_2O_2$가 모낭 주위에 축적되어 melanin 합성이 저하되어 백모가 유발되는 것으로 보고된 바 있다. MIXEE는 DPPH radical에 대한 소거능력과 활성산소에 대한 환원력이 있는 것으로 관찰되었다. 뿐만 아니라 세포내 $H_2O_2$ 수준을 감소시켰는데, 이러한 결과는 MIXEE는 산화를 억제하여 $H_2O_2$에 의하여 감소된 melanin 합성을 촉진 시킬수 있다는 것을 나타내었다. 한편 in vitro에서 MIXEE는 tyrosinase 활성에는 영향을 주지 않았지만 L-dopa에서 melanin 합성을 촉진하였다. 더욱이, B16F1세포에서 MIXEE는 melanin 생성을 세포수준에서 촉진하였다. Western blot을 이용한 단백질 발현 분석에서 MIXEE는 SOD-2 및 SOD-3. 특히 DOPA에서 melanin 이 생성되는 과정에 관여하는 효소인 TRP-2 발현에 긍정적인 영향을 미치는 것으로 입증되었다. 이상의 결과는 MIXEE가 백발화과정을 가역화시키는 과정에서 주요한 역할을 하는 melanin 생성을 촉진하는 하나의 약학 소재로 응용될 수 있다는 것을 시사하고 있다.

Keywords

SMGHBM_2019_v29n4_461_f0001.png 이미지

Fig. 1. Effect of MIXEE on DPPH radical Scavenging activity and reducing power. (A) Scavenging activity of MIXEE on DPPH radical. Vitamin C (Vit C) at 100 μg/ml was used as a positive control. (B) Reducing power of MIXEE. Vitamin C (Vit C) at 10 μg/ml was used as a positive control. Data are given as means of values ± SD from three independent experiments. Level of significance was identified statistically (* p<0.05, ** p<0.01, *** p< 0.001) using Student’s t test.

SMGHBM_2019_v29n4_461_f0002.png 이미지

Fig. 2. Effect of MIXEE on tyrosinase activity and melanin synthesis in vitro. (A) Effect of MIXEE on tyrosinase activity. (B) Effect of MIXEE on melanin synthesis. Vitamin C at 1,000 μg/ml was used as a negative control in these experiments. Data are given as means of values ± S.D. from three independent experiments. Level of significance was identified statistically (* p<0.05, ** p<0.01, *** p<0.001) using Student’s t test.

SMGHBM_2019_v29n4_461_f0003.png 이미지

Fig. 3. Effect of MIXEE on viability in B16F1 cells. The cells were treated with MIXEE at 1, 2, 4, 8, 16, 32 and 64 μg/ml. Cell viability was determined by MTT assay after 24 hr of incuvation. Data are given as means of values ± S.D. from three independent experiments. Level of significance was identified satistically (** p<0.01) using Student’s t test.

SMGHBM_2019_v29n4_461_f0004.png 이미지

Fig. 4. Scavenging effect of MIXEE on intracellular H2O2. B16F1 cells were loaded with 20 μM DCFH-DA for 20 minutes. The cells were pre-treated with MIXEE at concentrations indicated for 1 hr, and were exposed to H2O2 in HBSS buffer for 1 hr. The fluorescence intensities were measured with 488 nm and 530 nm of excitation and emission frequencies, respectively. Level of significance was identified statistically (** p<0.01, *** p<0.001) using Student’s t test.

SMGHBM_2019_v29n4_461_f0005.png 이미지

Fig. 5. Effect of MIXEE on melanin production in B16F1 treated without or with 250 μM H2O2. (A) The amount of itracellular melanine were analyzed in B16F1 cells treated with MIXEE. (B) The amount of itracellular melanine were analyzed in B16F1 cells treated with MIXEE after H2O2 stimulation. Vit.C 1,000 μg/ml was used as a positive control. In these experiments Data are given as means of values ± S.D. from three independent experiments. Level of significance was identified statistically (** p< 0.01, *** p<0.001) using Student’s t test.

SMGHBM_2019_v29n4_461_f0006.png 이미지

Fig. 6. Effect of MIXEE on protein expressions of SOD-2, SODD-3, catalase in B16F1 cells. The cells were treated with MIXEE at 4, 8 and 16 μg/ml. Western blot analysis of cell lysates was performed using antibodies as indicated. The expression of β-actin was used as a control for normalization of target proteins. Level of significance was identified statistically (* p<0.05, ** p<0.01, *** p<0.001) using Student’s t test

SMGHBM_2019_v29n4_461_f0007.png 이미지

Fig. 7. Effect of MIXEE on protein expressions of TRP-1, TRP-2, tyrosinase in B16F1 cells. The cells were treated with MIXEE at 4, 8, 16 and 32 μg/ml. Western blot analysis of cell lysates was performed using antibodies as indicated. The expression of β-actin was used as a control for normalization of target proteins. Level of significance was identified statistically (* p<0.05) using Student’s t test.

References

  1. Akiba, Y. and Matsumoto, T. 1980. Effects of several types of dietary fibers on lipid content in liver and plasma, nutrient retentions and plasma transaminase activities in force-fed growing chicks. J. Nutr. 110, 1112-1121. https://doi.org/10.1093/jn/110.6.1112
  2. Bohm, M. and Hill, H. Z. 2016. Ultraviolet B, melanin and mitochondrial DNA: Photo-damage in human epidermal keratinocytes and melanocytes modulated by alpha-melanocyte-stimulating hormone. F1000Res. 5, 881. https://doi.org/10.12688/f1000research.8582.1
  3. Chan, T., Chang, C., Koonchanok, N. and Geahlen, R. 1993. Selective inhibition of the growth of ras-transformed human bronchial epithelial cells by emodin, a protein-tyrosine kinase inhibitor. Biochem. Biophys. Res. Commun. 193, 1152-1158. https://doi.org/10.1006/bbrc.1993.1746
  4. Chang, C. H., Lin, C. C., Yang, J. J., Namba, T. and Hattori, M. 1996. Anti-inflammatory effects of emodin from Ventilago leiocarpa. Am. J. Chin. Med. 24, 139-142. https://doi.org/10.1142/S0192415X96000189
  5. Choi, H. R., Kang, Y. A., Lee, H. S. and Park, K. C. 2016. Disulfanyl peptide decreases melanin synthesis via receptor-mediated ERK activation and the subsequent down-regulation of MITF and tyrosinase. Int. J. Cosmet. Sci. 38, 279-285. https://doi.org/10.1111/ics.12291
  6. Fardet, A., Rock, E. and Remesy, C. 2008. Is the in vitro antioxidant potential of whole-grain cereals and cereal products well reflected in vivo? J. Cereal Sci. 48, 258-276. https://doi.org/10.1016/j.jcs.2008.01.002
  7. Feng, G. Y., Shi, L. Y., Cui, B. D., Li, D. K. and Sun, Z. X. 2015. Effect of water extract and main monomer components of processed Polygonum multiflorum on intracellular tyrosinase and their antioxidantactivity. Nat. Prod. Res. Dev. 4, 007.
  8. Furumoto, T. and Nishimoto, K. 2016. Identification of a characteristic antioxidant, anthrasesamone F, in black sesame seeds and its accumulation at different seed developmental stages. Biosci. Biotechnol. Biochem. 80, 350-355. https://doi.org/10.1080/09168451.2015.1091717
  9. Gan, E., Haberman, H. and Menon, I. 1974. Oxidation of NADH by melanin and melanoproteins. Biochim. Biophys. Acta (BBA)-Enzymol. 370, 62-69. https://doi.org/10.1016/0005-2744(74)90031-X
  10. Hachinohe, M. and Matsumoto, H. 2005. Involvement of reactive oxygen species generated from melanin synthesis pathway in phytotoxicty of L-DOPA. J. Chem. Ecol. 31, 237-246. https://doi.org/10.1007/s10886-005-1338-9
  11. Han, K. H., Oh, J. C. and Ryu, C. H. 2004. A study on the optimization for preparation conditions of germinated brown rice gruel. J. Kor. Soc. Food Sci. Nutr. 33, 1735-1741. https://doi.org/10.3746/jkfn.2004.33.10.1735
  12. Han, S. H. and Shin, M. K. 2002. Effects of methanol extracts from Rhynchosia nulubilis on serum lipid concentrations in rats fed high fat and high cholesterol diet. J. Kor. Soc. Food Cul. 17, 64-69.
  13. Han, S. J., Ryu, S. N. and Kang, S. S. 2004. A new 2-arylbenzofuran with antioxidant activity from the black colored rice (Oryza sativa L.) bran. Chem. Pharm. Bull. 52, 1365-1366. https://doi.org/10.1248/cpb.52.1365
  14. Hasan, S., Dinh, K., Lombardo, F., Dawkins, F. and Kark, J. 2003. Hypopigmentation in an African patient treated with imatinib mesylate: a case report. J. Natl. Med. Assoc. 95, 722.
  15. Hatano, T., Uebayashi, H., Ito, H., Shiota, S., Tsuchiya, T. and Yoshida, T. 1999. Phenolic constituents of Cassia seeds and antibacterial effect of some naphthalenes and anthraquinones on methicillin-resistant Staphylococcus aureus. Chem. Pharm. Bull. 47, 1121-1127. https://doi.org/10.1248/cpb.47.1121
  16. Herrling, T., Jung, K. and Fuchs, J. 2008. The role of melanin as protector against free radicals in skin and its role as free radical indicator in hair. Spectrochim. Acta A. Mol. Biomol. Spectrosc. 69, 1429-1435. https://doi.org/10.1016/j.saa.2007.09.030
  17. Hong, S. H., Sim, M. J. and Kim, Y. C. 2016. Melanogenesis-promoting effects of Rhynchosia nulubilis and Rhynchosia volubilis ethanol extracts in melan-a cells. Toxicol. Res. 32, 141. https://doi.org/10.5487/TR.2016.32.2.141
  18. Jang, J. Y., Kim, H. N., Kim, Y. R., Kim, B. W., Choi, Y. H. and Choi, B. T. 2010. Studies of inhibitory mechanism on melanogenesis by partially purified Asiasari radix in ${\alpha}$-MSH stimulated B16F10 melanomac cells. J. Life Sci. 20, 1617-1624. https://doi.org/10.5352/JLS.2010.20.11.1617
  19. Jie, Y., Jie, X., Xiao-Jian, M., Hua, W., Sheng-Lan, Z., Ya-Ge, M., Na, L. and Rong-Hua, Z. 2012. Comparison of laxative and antioxidant activities of raw, processed and fermented Polygoni Multiflori radix. Chin. J. Nat. Med. 10, 63-67. https://doi.org/10.3724/SP.J.1009.2012.00063
  20. Kang, S. and Han, J. 2004. Acetylcholinesterase inhibiting effect and free radical scavenging effect of soybean (Glycine max) and Yak-Kong (Rhynchosia nolubilis). J. East. Asian Soc. Diet Life 14, 64-69.
  21. Kim, K. S., Kim, J. A., Eom, S. Y., Lee, S. H., Min, K. R. and Kim, Y. 2006. Inhibitory effect of piperlonguminine on melanin production in melanoma B16 cell line by down-regulation of tyrosinase expression. Pigment Cell Res. 19, 90-98. https://doi.org/10.1111/j.1600-0749.2005.00281.x
  22. Kim, M. K. and Won, E. J. 1984. Effects of feeding polished or brown rice diet with different kinds of lipids on the lipid metabolism in rats. J. Nutr. Health 17, 154-162.
  23. Kim, S. R., Ahn, J. Y., Lee, H. Y. and Ha, T. Y. 2004. Various properties and phenolic acid contents of rices and rice brans with different milling fractions. Kor. J. Food Sci. Technol. 36, 930-936.
  24. Kong, S. Y., Choi, Y. M., Lee, S. M. and Lee, J. S. 2008. Antioxidant compounds and antioxidant activities of the methanolic extracts from milling fractions of black rice. J. Kor. Soc. Food Sci. Nutr. 37, 815-819. https://doi.org/10.3746/jkfn.2008.37.7.815
  25. Kritchevsky, D. 1982. Dietary fiber and disease. Bull. N. Y. Acad. Med. 58, 230.
  26. Lee, S. C., Chen, C. H., Yu, C. W., Chen, H. L., Huang, W. T., Chang, Y. S., Hung, S. H. and Lee, T. L. 2016. Inhibitory effect of Cinnamomum osmophloeum Kanehira ethanol extracts on melanin synthesis via repression of tyrosinase expression. J. Biosci. Bioeng. 123, 263-269.
  27. Musa, A. S., Umar, M. and Ismail, M. 2011. Physicochemical properties of germinated brown rice (Oryza sativa L.) starch. Afr. J. Biotechnol. 10, 6281-6291.
  28. Park, J. D., Cho, B. K., Kum, J. S. and Lee, H. Y. 2005. Quality properties of cooked germinated-brown rice. Kor. J. Food Preserv. 12, 101-106.
  29. Park, J. D., Choi, B. K., Kum, J. S. and Lee, H. Y. 2006. Physicochemical properties of brown rice flours produced under different drying and milling conditions. Kor. J. Food Sci. Technol. 38, 495-500.
  30. Park, Y. J., Min, S. K., Min, S. I., Kim, S. J. and Ha, J. 2015. Effect of imatinib mesylate and rapamycin on the preformed intimal hyperplasia in rat carotid injury model. Ann. Surg. Treat. Res. 88, 152-159. https://doi.org/10.4174/astr.2015.88.3.152
  31. Park, Y. S., Kim, S. J. and Chang, H. I. 2008. Isolation of anthocyanin from black rice (Heugjinjubyeo) and screening of its antioxidant activities. Kor. J. Microbiol. Biotechnol. 36, 55-60.
  32. Ramarathnam, N., Osawa, T., Ochi, H. and Kawakishi, S. 1995. The contribution of plant food antioxidants to human health. Trends Food Sci. Technol. 6, 75-82. https://doi.org/10.1016/S0924-2244(00)88967-0
  33. Seagle, B.-L. L., Rezai, K. A., Gasyna, E. M., Kobori, Y., Rezaei, K. A. and Norris, J. R. 2005. Time-resolved detection of melanin free radicals quenching reactive oxygen species. J. Am. Chem. Soc. 127, 11220-11221. https://doi.org/10.1021/ja052773z
  34. Shim, Y., Cha, G. and Sin, J. 1995. Studies on the experimental cookery and the preservation of the Hugimja Dashik. J. Nat. Sci. Inst. Seoul Worman's Univ. 6, 13-26.
  35. Shyu, Y. S. and Hwang, L. S. 2002. Antioxidative activity of the crude extract of lignan glycosides from unroasted Burma black sesame meal. Food Res. Int. 35, 357-365. https://doi.org/10.1016/S0963-9969(01)00130-2
  36. Stickel, F., Seitz, H., Hahn, E. and Schuppan, D. 2001. [Liver toxicity of drugs of plant origin]. Z. Gastroenterol. 39, 225-232, 234-237. https://doi.org/10.1055/s-2001-11772
  37. Trowell, H. 1975. Letters to the editor: Coronary heart disease and dietary fiber. Am. J. Clin. Nutr. 28, 798-800. https://doi.org/10.1093/ajcn/28.8.798
  38. Wood, J. M., Decker, H., Hartmann, H., Chavan, B., Rokos, H., Spencer, J., Hasse, S., Thornton, M., Shalbaf, M. and Paus, R. 2009. Senile hair graying: $H_2O_2$-mediated oxidative stress affects human hair color by blunting methionine sulfoxide repair. FASEB J. 23, 2065-2075. https://doi.org/10.1096/fj.08-125435
  39. Yang, Z., Nakabayashi, R., Mori, T., Takamatsu, S., Kitanaka, S. and Saito, K. 2016. Metabolome analysis of Oryza sativa (Rice) using liquid chromatography-mass spectrometry for characterizing organ secificity of favonoids with ati-inflammatory and anti-oxidant activity. Chem. Pharm. Bull. 64, 952-956. https://doi.org/10.1248/cpb.c16-00180
  40. Yen, G. C., Chen, H. W. and Duh, P. D. 1998. Extraction and identification of an antioxidative component from Jue Ming Zi (Cassia tora L.). J. Agric. Food Chem. 46, 820-824. https://doi.org/10.1021/jf970690z
  41. Zhaohuan, L., Guiyuan, L. V. and Jingjing, Y. U. 2014. Review on the research of the components, pharmacological actions and toxicity of Polygonum multiflorum Thunb (Heshouwu). J. Zhejiang Chinese Medical University 4, 495-500.
  42. Kang, K. K., Lee, Y. E., Woo, W. H. and Mun, Y. Z. 2014. Commelina communis Ledeb inhibits melanin synthesis in alpha-MSH-stimulated B16F10 cells. J. Pathool. Pathol. Kor. Med. 28, 506-511.