中藥水芫花粗萃物抑制黑色素生成之機轉研究

Inhibitory effect of a water extract from Pemphis acidula on melanogenesis in mouse B16 melanoma cells.

葉品萱(Pin-Hsuan Yeh)

指導教授 ： 戴守谷

共同指導教授 ： 張德生

國立虎尾科技大學/文理學院/生物科技研究所/碩士(2010年)

摘要

近年來美白化妝品之開發，最主要的作用機理是透過抑制黑色素細胞中tyrosinase之活性而達到美白的效果。然而目前臨床醫療及化妝品市場中所使用的美白有效成分大多具有生理毒性(如熊果素、對苯二酚與麴酸等)，在使用上有所限制。因此，本論文即以中草藥為對象，進行黑色素抑制劑篩選與機轉研究。本實驗利用老鼠B16黑色素瘤細胞及斑馬魚胚胎進行體外及體內美白測試，透過斑馬魚胚胎之藥物初步篩選美白效果分析，發現水芫花粗萃物(Pemphis acidula extract, PAE)對黑色素生成有明顯的抑制效果，故進一步進行細胞美白測試。首先水芫花對B16老鼠黑色素瘤細胞之細胞毒性分析，測試細胞存活率(MTT法)，結果顯示水芫花粗萃物之細胞毒性IC50值為211μg/ml。然而以水芫花粗萃物之3種不同濃度，分別為50μg/ml、25μg/ml、12.5μg/ml處理B16細胞後，發現水芫花之粗萃物可有效地降低細胞黑色素的含量且對處理劑量有相對應之效果(dose-dependent)，且其抑制黑色素生成IC50值為33.3 μg/ml。實驗結果顯示，水芫花萃取物比熊果素具有更強的抑制活性，在更一步的研究中，分別偵測PAE對細胞內及細胞外tyrosinase活性的抑制、以西方點墨法偵測PAE處理後細胞內tyrosinase蛋白質含量及以定量PCR (qRT-PCR)偵測PAE處理後細胞tyrosinase及MITF之RNA表現量，實驗結果證實PAE抑制黑色素生成之機轉乃是透過抑制酪胺酸?上游基因MITF轉錄因子之活性來達成。

關鍵字

水芫花 ； 酪胺酸酵素 ； 黑色素 ； 抑制 ； 美白

並列摘要

Mouse B16 melanoma cells and zebrafish were used as an in vitro and an in vivo system to evaluate the depigmenting activity of tested drugs. Among the screened crude extracts from selected Chinese herbs, a crude extract from Pemphis acidula was found to show strong depigmenting activity on melanogenesis of both B16 cells and zebrafish embryos. The inhibitory effects of the crude extract on melanogenesis were investigated in mouse B16 melanoma cells. The cytotoxicity IC50 value of the extract toward B16 cells was determined to be 211 μg/ml by a MTT method. In the other hand, the extract inhibited melanogenesis of B16 cells in a dose-dependent manner with an IC50 value of 33.3 μg/ml. Moreover, the inhibitory activity of the extract from Pemphis acidula also showed more potent than that of arbutin, which is a well-known commercial skin-whitening agent. The inhibitory mechanism of the extract on melanogenesis was to decrease the expression of tyrosinase by MITF down-regulation in the B16 cell.

並列關鍵字

Pemphis acidula ； tyrosinase ； melanin ； inhibition ； skin-whitening

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