裂葉月見草萃取物於抗氧化及抑制黑色素生合成效果之評估

Antioxidant and anti-melanogenic effects of the extracts of Oenothera laciniata Hill

張佑慈(Yeo-Tzu Chang)

指導教授 ： 柯宏慧

高雄醫學大學/藥學院/香粧品學系碩士班/碩士(2012年)

摘要

裂葉月見草為多年生草本植物被用於治療多種發炎性疾病。例如
：紅斑性狼瘡、僵直性脊椎炎及類風溼性關節炎等。然而其抗氧化及抑制黑色素生成的相關活性尚未釐清。本研究中利用裂葉月見草甲醇萃取物(OLM)、乙酸乙酯萃取層(OLEF)、正丁醇萃取層(OLBF)及水萃取層(OLWF)，在細胞外的試驗，藉由DPPH、ABTS、超氧陰離子自由基清除、還原力及亞鐵離子螯合方法，評估裂葉月見草甲醇萃取物及其劃分層的抗氧化能力；並評估各萃取層抑制蘑菇酪胺酸酶的活性。在B16-F10黑色素腫瘤細胞中，針對黑色素含量及酪胺酸酶活性進行評估並利用西方點墨法分析各種與黑色素生成相關之蛋白質表現，如tyrosinase、TRP-1、TRP-2、 MITF、p-CREB、p-ERK及p-JNK。結果顯示，OLM及其劃分層具有良好的抗氧化及抑制酪胺酸酶的能力與總酚含量有關。在B16-F10黑色素腫瘤細胞中，OLM及其劃分層可抑制黑色素含量及酪胺酸酶的活性，並具有劑量正相關的現象。此外，本研究亦證實OLM及其劃分層被證實能藉由抑制CREB磷酸化進而減少MITF的表現，並且調控下游酪胺酸酶及TRP-2在B16F10細胞內的蛋白質表現；但是，裂葉月見草萃取物不會抑制TRP-1的表現。此外，結果顯示OLM、OLBF及OLWF可藉由增加ERK的磷酸化來達到減少黑色素生合成的效果，而OLEF則是藉由誘導JNK的磷酸化來抑制黑色素生合成。這些結果顯示裂葉月見草具有發展為化粧品中抗氧化劑及抗黑色素增生劑之潛能。

關鍵字

裂葉月見草 ； 抗氧化 ； 抗黑色素生成

並列摘要

Oenothera laciniata Hill, a perennial herb, is used to treat various inflammatory diseases such as systemic lupus erythematosus, ankylosing spondylitis, and rheumatoid arthritis. However, the antioxidant and antimelanogenic activities of O. laciniata are still not clear. The aim of the study compared the antioxidant activity of methanolic extract (OLM) from O. laciniata and its different fractions, including ethyl acetate (OLEF), n-butanol (OLBF), and water (OLWF), by determing the total phenolic content, the redical scavenging effects of DPPH, ABTS, and superoxide anion radical (O2˙−) scavenging, reducing power, and metal chelating ability. In addition, the anti-melanogenesis effect was determined by cell-free tyrosinase assay, cellular melanin content and tyrosinase activity in B16F10 melanoma cell. The melanin synthesis proteins were also quantified by western blot assay, including tyrosinase, tyrosinase related protein-1 (TRP-1), tyrosinase related protein-2 (TRP-2), microphthalmia-associated transcription factor (MITF), phospho-cAMP- response element binding protein (p-CREB), phospho-extracellular-signal -regulated kinase (ERK), and phospho-c-jun N-terminal kinase (JNK). The results showed that OLM and its fractions exhibited strong antioxidant and antityrosinase activities and there is a correlation with the total phenolic content. In B16F10 melanoma cells, OLM and its fractions decreased melanin production and inhibited tyrosinase activity in a dose-dependent manner. Moreover, OLM and its fractions demonstrated that induced down-regulation of melanogenesis via inhibiting p-CREB and subsequently reducing the expression of MITF, which in turn down-regulated tyrosinase and TRP-2 activities at the protein levels, but not inhibit TRP-1 expression in B16F10 cell. The results also indicated that OLM, OLBF, and OLWF reduced melanogenesis through an increase in ERK phosphorylation, but OLEF inhibited melanogenesis by inducing JNK phosphorylation. These results suggested that O. laciniata has great potential for becoming antioxidative and anti-melanogenesis agents in cosmetics.

並列關鍵字

Oenothera laciniata ； antioxidant ； anti-melanogenesis

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