| 摘要: | 痤瘡 (Acne) 俗稱青春痘或是面皰是一種常見的皮膚問題，通常出現在青春期的青少年和年輕成人荷爾蒙分泌失調、細菌感染、食物或是化妝品的應用上。到手香 [Plectranthus amboinicus (Lour.) Spreng] 是唇形花科 (Lamiaceae) 的植物，含豐富的精油，民間用藥用於治療喉嚨腫痛和消炎殺菌。許多研究文獻資料證明，到手香葉片萃取之香芹酚 (Carvacrol) 及百里香酚 (Thymol) 具有較強的抗發炎及鎮痛作用；本論文摒除精油的萃取，著重於到手香葉片多酚類的獲得和痤瘡桿菌的抑制作用。研究結果顯示，到手香葉片乙醇提取物之甲醇分配萃取層的總多酚含量最高 (167.79 ± 12.67 微克沒食子酸/毫克 乾重)，其也同樣具有最佳的 DPPH 清除活性，其 IC50 為 27±2 mg/mL，清除 ABTS 陽離子的能力 (IC50 為 21±2 mg/mL)和 Trolox 抗氧化當量 (Trolox Equivalent Antioxidant Capacity, TEAC) 為 0.38 μg TE /μg。而另外正丁醇和正己烷分配萃取物除了多酚含量僅為甲醇分配萃取物的七分之一，相對 DPPH 的 IC50 也不如甲醇分配萃取物。至於水層其多酚萃出量最低，亦無抗氧化效果。因此後續研究我們僅以甲醇分配萃取物作為抗痤瘡桿菌 (Propionibacterium acnes ) 功能性的研究。首先我們評估乙醇提取物及甲醇分配萃取物抑制痤瘡相關微生物的能力，紙錠擴散法 (Disk diffusion method) 的研究結果顯示，5 mg 甲醇分配萃取物可抑制痤瘡桿菌的生長，其抑制圈大小為 23 mm；至於乙醇提取物則無法抗痤瘡桿菌。另外容易出現在痤瘡膿腫傷口的金黃色葡萄球菌，5 mg 甲醇分配萃取物其抑制圈大小為 24 mm ；至於乙醇提取物為 10 mm。本實驗利用肉湯微量稀釋法 (Broth microdilution) 測試最小抑制濃度 (Minimum inhibitory concentration, MIC)，結果顯示甲醇分配萃取物之最小抑制金黃色葡萄球菌和痤瘡桿菌的濃度，皆為 0.3 mg/mL，而乙醇提取物則分別為 2.5 和 2 mg/mL，其中甲醇分配萃取物抑菌效果明顯優於乙醇提取物，尤其在抗金黃色葡萄球菌部分。再者，我們研究了乙醇提取物及甲醇分配萃取物對於LPS 所誘導的小鼠巨噬細胞(RAW264.7)其一氧化氮 (NO) 的產生量的影響，在劑量上，甲醇分配萃取物在 25 和 50 μg/mL 都有顯著的抑制 NO 的產生 (*p<0.05 和 **p<0.01)，且有劑量的依賴性，而乙醇提取物則無明顯功效。兩者皆不會影響細胞的存活率。至於甲醇分配萃取物的相關功能性機轉研究，則發現其會抑制誘發型一氧化氮合成? (inducible NO synthase, iNOS）和第二型環氧化?-2（cyclooxygenase-2,COX-2）的產生，但會促進第一型血紅素氧化酵素 (Heme oxygenase-1, HO-1) 的產生。甲醇分配萃取物應可成為青春痘的治療替代方式，相關機轉仍待持續研究中。
Acne is a common skin problem and usually appears in many teenagers and young adults with hormone imbalance, bacterial infection, food or cosmetic application. Plectranthus amboinicus is a plant of the Lamiaceae family. The heavy and strong scent of P. amboinicus has been used for centuries in perfumes, and more recently in insect repellents, and alternative medicines. Most of pharmacological studies indicated that the essential oil from P. amboinicus has modulatory effect on gastrointestinal tract, antibacterial activity, anti-inflammatory and analgesic effect. In this research, we aimed to isolate the polyphenols, instead of the essential oil, from the leaves of P. amboinicus and to test its therapeutic effects on acne. The total polyphenol concentrations and in vitro antioxidant activity in crude ethanol extract, n-hexane layer, methanol layer, n-butanol layer, and water layer, were investigated. We found the methanol layer, exhibited the highest total phenol content (167.79 ± 12.67 mg of gallic acid/mg of dry weight), DPPH and ABTS+ radical cation scavenging activities (IC50=27 ± 2 mg/mL and 21 ± 2 mg/mL, respectively) and Trolox equivalent antioxidant capacity (TEAC=0.38 μg TE /μg). However, the other three layers showed no or little activity to scavenge DPPH and ABTS. Furthermore, the crude ethanol extract and the methanol layer were tested for anti-bacterial activity against Propionibacterium acnes and Staphylococcus aureus, which have been recognized as pus-forming bacteria triggering an inflammation in acne. Disc diffusion and broth microdilution were employed. Disc diffusion method showed that the 5 mg methanol layer exhibited 23 mm of inhibition zones against P. acnes, respectively. Crude ethanol extract did not show clear inhibition zone. The MIC values against Staphylococcus aureus for crude ethanol extract and methanol layer were 2.5 mg/mL and 0.3 mg/mL, respectively. On the other hand, the MIC values against Propionibacterium acnes were about 2 mg/mL and 0.3 mg/mL for crude ethanol extract and methanol layer, respectively. The anti-inflammatory effects of crude ethanol extract and the methanol layer were examined in LPS-induced RAW264.7 cells. The methanol layer with the highest phenolic content, exhibited dose-dependent inhibitory effect against LPS-mediated nitric oxide (NO) release, without affecting cell viability in RAW264.7 macrophages. The methanol layer also significantly downregulated the expression of LPS-induced protein expression of iNOS (inducible NO synthase) and COX-2 (cyclooxygenase-2); while upregulated HO-1 (heme oxygenase-1) expression. To sum up, the methanol layer of P. amboinicus may serve as an attractive candidate in complementary and alternative medicine for the treatment of acne. |
