探討台灣紫金牛科草藥之抗氧化活性與成分分析

過去數年，許多化學合成的抗氧化劑，如Butylated hydroxytoluene (BHT)和Butylated hydroxyanisole (BHA)，已被廣泛用於食品、藥品和化妝品。這些化學抗氧化劑被發現易引起過敏反應和致癌性，因此尋求有效的天然抗氧化劑是一個重要的問題。天然草藥已被發現具有多種生物活性，也擁有低副作用、高穩定性和低毒性的特徵。在本研究中，使用五種不同的傳統溶劑與超臨界CO2萃取台灣原住民紫金牛科草藥，並比較其抗氧化活性。其中，五種傳統溶劑包括:甲醇、乙醇、乙酸乙酯、乙醚和水，體外抗氧化分析包括三種測試: DPPH自由基清除能力，Trolox當量抗氧化能力和還原能力。此外，利用光譜定量分析各種萃出物之總酚與類黃酮含量，並進行抗氧化能力與總酚和類黃酮含量之相關性分析。抗氧化分析結果顯示，超臨界CO2萃取物(SFE fraction 2) 的 DPPH自由基清除能力最好，其IC50值為164.5±3.38 ppm；傳統溶劑乙酸乙酯萃取物的Trolox當量抗氧化能力(30.83±0.25 g Trolox/ 100 g Dry Weight; DW)最佳；還原能力(0.5±0.01 abs / 10-3 ppm)也是傳統溶劑乙酸乙酯萃取物最好。各種萃出物中，總酚含量最高的是傳統溶劑乙酸乙酯萃取物，含量為4.27±0.03 g Gallic acid /100 g DW；類黃酮含量最高的則為超臨界CO2萃取物(SFE fraction 2)，含量為4.82±0.01 g Quercetin /100 g DW。以GC/MS分析各萃出物成分，含量最高的三個分別為，乙酸乙酯萃出物含有主要成分2,4-Di-tert-butylphenol，其次為Stigmasterol ，第三為Campesterol。乙醇萃出物含有主要成分5-Hydroxymethylfurfural，其次為Stigmasterol，第三為n-Hexadecanoic acid。乙醚萃出物含有主要成分Stigmasterol，其次為endo-Borneol，第三為Campesterol。超臨界萃出物SFE fraction 1和SFE fraction 2含有主要成分n-Hexadecanoic acid，其次為Stigmasterol，第三為(Z,Z)-9,12-Octadecadienoic acid。依據文獻資料，這些成分中可能提供抗氧化活性的成分分別為2,4-Di-tert-butylphenol, Campesterol, Stigmasterol, 5-Hydroxymethylfurfural, n-Hexadecanoic acid, (Z,Z)-9,12-Octadecadienoic acid。根據本研究所得結果，證實以超臨界CO2萃取的SFE fraction 2和傳統溶劑乙酸乙酯萃取物的抗氧化能力最好，可以深入研發成天然抗氧化劑，取代現有合成的抗氧化劑。

關鍵字

紫金牛科草藥；抗氧化；超臨界流體萃取；傳統溶劑萃取

並列摘要

Over the past few years, many chemical synthetic antioxidants, such as Butylated hydroxytoluene (BHT) and Butylated hydroxyanisole (BHA), has been widely used in food, pharmaceuticals and cosmetics. These chemical antioxidants were found easy to cause allergic reactions and carcinogenicity, and therefore the search for effective natural antioxidants is an important issue. Natural herbs have been found to have a variety of biological activity, but also has low side effects, high stability and low toxicity characteristics. In the present study, five different traditional solvents and supercritical CO2 fluid were used to extract Taiwan aboriginal herbal medicine, Myrsinaceae, and compared their antioxidant activities. Five of the traditional solvents included methanol, ethanol, ethyl acetate, ethyl ether and water. The antioxidant activity was determined by three methods: DPPH radical scavenging ability, Trolox equivalent antioxidant capacity and reducing power. In addition, Spectrophotometric assays were used to determine the total phenolic and flavonoid contents for each of the various extracts. The antioxidant activities and their total phenolic and flavonoid contents were evaluated for the correlation analysis. The results of antioxidant activity assay showed that the supercritical CO2 extract (SFE fraction 2) revealed a highest DPPH radical scavenging ability with an IC50 value of 164.5 ± 3.38 ppm; the ethyl acetate extract revealed a highest Trolox equivalent antioxidant capacity (30.83 ± 0.25 g Trolox / 100 g DW) and reducing power (0.5 ± 0.01 abs / 10-3 ppm). Among the various extracts, the ethyl acetate extract also possessed the highest total phenol content (TPC) with a TPC value of 4.27 ± 0.03 g Gallic acid / 100 g DW. However, the supercritical CO2 extract (SFE fraction 2) showed the highest total flavonoid content (TFC) with a TFC value of 4.82 ± 0.01 g Quercetin / 100 g DW. In the analysis of GC-MS, three of the major components found in each extracts were 2,4-Di-tert-butylphenol, Stigmasterol and Campesterol in the ethyl acetate extracts; 5-Hydroxymethylfurfural, Stigmasterol and n-Hexadecanoic acid in the ethanol extract; Stigmasterol, endo-Borneol and Campesterol in the ether extracts; n-Hexadecanoic acid, Stigmasterol and, (Z, Z) –9,12-Octadecadienoic acid in the Supercritical CO2 extracts (SFE1 and SFE2). Based on the previous literature reports, the composition of 2,4-Di-tert-butylphenol, Campesterol, Stigmasterol, 5-Hydroxymethylfurfural, n-Hexadecanoic acid, and (Z, Z)-9,12-Octadecadienoic acid might be the active substances that contribute the antioxidant activity. According to the results obtained in this study, the supercritical CO2 extract (SFE fraction 2) possesses significant DPPH free radical scavegening ability and the ethyl acetate extract revealed significant total antioxidant activity and reducing power, which showed positive correlation with their total phenolic and flavonoid contents. In conclusion, the extracts obtained from Taiwan aboriginal medicine, Myrsinaceae, has the potential to be developed as a natural antioxidant agent.