台灣筋骨草葉萃取物之抗氧化及抗發炎活性之探討

發炎反應在許多疾病的發生有著重要作用，而發炎和氧化之間有著密不可分的關係，故許多研究致力於尋找具抗氧化效果及抑制發炎介質生成且無副作用的天然物質成分。台灣筋骨草 (Ajuga taiwanensis Nakai ex Murata)為唇形科 (Lamiaceae)筋骨草屬 (Ajuga)植物，廣泛分布於琉球、台灣及廣東，其含有許多生物活性物質，如萜類、類黃酮等。許多文獻指出筋骨草屬植物具有抗腫瘤、抗菌、抗發炎等功效。本研究以95℃熱水及不同濃度乙醇 (25%、50%及75%)為溶劑萃取台灣筋骨草葉，分析其萃取物之抗氧化能力及對脂多醣誘發RAW 264.7小鼠巨噬細胞發炎反應之影響。實驗結果顯示，總酚及類黃酮含量以75%乙醇萃取物為最高，分別為79.46 mg/g及53.46 mg/g；DPPH自由基清除能力以75%乙醇萃取有最佳的清除效果，於濃度 400 μg/mL下，抑制率可達85%；TEAC總抗氧化能力及螯合亞鐵能力皆以95℃熱水萃取的效果最佳，分別為93%與83%。而台灣筋骨草葉萃取物經HPLC鑑定其酚類化合物，經比對滯留時間發現台灣筋骨草葉萃取物皆含有沒食子酸、兒茶素、綠原酸及槲皮素。以液相層析質譜儀 (LC-MS)鑑定波峰面積最大者，其為芹菜素。MTT結果顯示，95℃熱水萃取物及25%乙醇萃取物濃度為400 μg/mL、50%乙醇萃取物濃度為200 μg/mL、75%乙醇萃取物濃度為100 μg/mL，均不影響細胞存活率，添加LPS (100 ng/mL)共處理24小時也不具細胞毒性。以LPS (100 ng/mL)誘導RAW 264.7巨噬細胞發炎，測定台灣筋骨草葉萃取物對其NO生成量之影響，結果顯示台灣筋骨草葉50%乙醇萃取物具有較佳之抑制NO生成之效果。台灣筋骨草葉50%乙醇萃取物亦可抑制TNF-α、IL-6之生成及iNOS蛋白質表現量。本研究結果顯示台灣筋骨草葉萃取物具抗氧化能力及抑制發炎介質生成之效果，未來可應用於保健食品之開發。

關鍵字

台灣筋骨草葉；抗氧化；巨噬細胞；脂多醣

並列摘要

Inflammation plays an important role in the initiation of many diseases. There is an inseparable relationship between inflammation and oxidation. Many studies have focused on searching natural ingredients that can inhibit the inflammation mediators and have antioxidant activity without side effects. Ajuga taiwanensis Nakai ex Murata is a plant belonging to genus Ajuga (Lamiaceae) which is distributed in Ryukyu, Taiwan and Guangdong. It contains many biologically active substances such as terpenoids and flavonoids. In the present study, A. taiwanensis Nakai ex Murata leaves was extracted with 95℃ hot water and various concentrations (25%, 50% and 75%) of ethanol. The extracts were investigated in antioxidant activity and the effect on LPS-induced inflammatory response in mouse RAW 264.7 macrophages. The results showed that A. taiwanensis Nakai ex Murata leaves extracted with 75% ethanol indicated the highest total phenolic and total flavonoid content to be 79.46 mg/g and 53.46 mg/g, respectively. The best DPPH radical scavenging ability reached 85% at concentration of 400μg/mL by 75% ethanol extract. The greatest Trolox-Equivalent antioxidant capacity (TEAC) and chelating ferrous iron capacity was found in hot water extract: TEAC exhibited 93% at 4 mg/mL concentration and the chelating ferric ability was 83%. Active ingredients, gallic acid, catechin, chlorogenic acid and quercetin, analyzed by HPLC were identified. The highest content of 75% ethanol extract identified by LC-MS is apigenin. The MTT cell viability assay was employed to evaluate the toxicity of various extracts and the results suggested little effect on cell viability at 400 μg/mL, 400 μg/mL, 200 μg/mL, 100 μg/mL was investigated, respectively. Using LPS (100 ng/mL) to induce inflammation in RAW 264.7 macrophages and the effect of A. taiwanensis Nakai ex Murata leaf extracts on NO production was measured. The results demonstrated that 50% ethanol extract from A. taiwanensis Nakai ex Murata leaf exhibited the best inhibiting NO production. The 50% ethanol extract from A. taiwanensis Nakai ex Murata leaves indicating inhibition on the production of TNF-α, IL-6 and iNOS protein expressssion. In summary, A. taiwanensis Nakai ex Murata leaves not only illustrated a reliable antioxidant ability but also inhibited the production of inflammatory mediators, showed the potentiality to be developed as a nutracieutical product.