山藥(Dioscorea)自古以來即是藥用及保健用食材重要之原料,對人體有益腸胃、滋補、降血糖及調養婦科疾病等多種效能,目前也是本省大力推廣種植之保健食材。現代研究發現某些山藥品系含有獨特的皂甘、多糖、多酚化合物及其他抗氧化物質,這些成分都可能與山藥所具有之保健功能有密切相關。但這些分子多數都仍未被清楚了解及定義,而這些分子是否確為其食用山藥後所發揮保健功能之有效成分或這些具生物活性分子之間是否具有其交互作用則更不得而知,再加上山藥品系繁多,其中組成也常因品系、栽培之地理氣候因素而有極大差異,因此深入了解山藥中具生物活性之成分及與其保健功能間之相關性,並依此建立山藥品系間機能性之評估標準,對未來山藥的推廣種植是極為重要的工作。目前眾多的醫學報告幾乎已確定生物氧化物與抗氧化系統間之失調是萬病之因。因此,本研究計畫即以抗氧化活性作為其功能性指標,藉此找尋並特性化山藥仿食用方式萃取所得冷凍乾燥樣品中所具有抗氧化活性之主要成分,了解這些分子在細胞或動物之生理功能,並建立未來評估山藥不同品系之機能性指標是本研究計畫及後續計畫之長遠目標。本期計畫完成之重要成果如下:(1)臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品具有明顯保護硫酸銅與過氧化氫合併處理造成牛胸腺DNA的傷害。(2)臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品可能具與銅離子鍵結之活性。(3)臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品中具有明顯保護硫酸銅與過氧化氫合併處理造成牛胸腺DNA傷害的主要有效分子其分子量分佈在30kDa以下範圍。(4)臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品中具有明顯保護硫酸銅與過氧化氫合併處理造成牛胸腺DNA傷害的主要有效成分對熱呈不敏感性。甚至加熱有促進其抗氧化活性。(5)利用慧星分析(Comet assay)發現:臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品可有效保護過氧化氫、硫酸鐵在人類淋巴母細胞株所造成的核DNA傷害。但對硫酸鐵所造成的核DNA傷害則不具保護性。(6)利用過氧化氫螢光探針Amplex red測量臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品都不具顯著分解過氧化氫之能力,推測本研究中所製備之山藥萃取樣本不含過氧化氫酵素(Catalase)及過氧化酵素(Peroxidase)。(7)以質體DNA構形改變(Conformational change)的分析發現:臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品不具抗游離輻射(X-ray)之活性。(8)以Bio-Rad試劑測量發現:30%酒精當作萃取溶劑可明顯增加蛋白質之含量約1.5(不加熱)至2.0倍(加熱)。臺農二號山藥之水、30%酒精及30%酒精加熱萃取之冷凍乾縮樣品中含有一主要蛋白分子量約36.8kDa。(9)臺農二號山藥之水萃物再經煮沸30分鐘後,顯著改變其對溶劑之溶解能力,不溶於一般之極性溶劑如水、Dimethyl sulfoxide、乙醚等,但可溶解於鹼性溶劑如NaOH,以0.2N NaOH溶解之山藥水萃物亦不具保護硫酸銅與過氧化氫合併處理造成牛胸腺DNA傷害之活性。
The tubers of Discorea are mainly used as a health food and for extraction of diogenin, sapogenin and its glycoside dioscin for use as steroid intermediate in the pharmaceutical industry in China. Free radicals have been related with aging and diseases, such as cancer, diabetes and especially in neurological disorders. A diet including variable antioxidant foods may therefore help to prevent these illnesses. In this experiment, we evaluated the in vitro antioxidant activities of four lyophilized extracts obtained from Discorea alata L. tubers: (1) aqueous extract, AE (2) 30% ethanolic extract, EE (3) aqueous extract boiling for 30 minutes, ABE and (4) 30% ethanolic extract boiling for 30 minutes, EBE. Protective effect against H/sub 2/O/sub 2/-CuSO/sub 4/ induced DNA damages on calf thymus DNA damages were detected by ethidium bromide binding assay as a model system. Except the lyophilized aqueous extract for boiling 30 minutes, three other different extracts were significantly reduced the H/sub 2/O/sub 2/-CuSO/sub 4/ induced DNA damages on calf thymus DNA. The ID50 value for lyophilized aqueous extract, 30% ethanolic extract and 30% ethanolic extract boiling for 30 mnutes was 0.14, 0.22, and 0.17 mg/mL, respectively. These results suggest that the Discorea tubers extracts have strong antioxidant actions and that it may be prophylactic food against the age related and neurological diseases.