金線連水萃取物中的醣類與有機酸之分析

The analysis of carbohydrates and organic acids in water extract of Anoectochilus formosanus Hayata

林佳潔(Chia-Chieh Lin)

指導教授 ： 呂廷璋

共同指導教授 ： 許順堯

國立臺灣大學/生物資源暨農學院/食品科技研究所/碩士(2009年)

https://doi.org/10.6342/NTU.2009.10052

摘要

臺灣金線連(Anoectochilus formosanus Hayata)為台灣原生種蘭科植物，為民間珍貴草藥，被稱為藥王或藥虎等。金線連水萃取為常用的製備方式，本研究針對水萃物中之醣類與有機酸進行篩檢與特性分析。熱水萃取與乙醇沉澱可獲得金線連之粗多醣含量為全草乾重之14.81%，其中α-D-Glucan含量約60-66 %，與碘液反應不呈現藍紫色，呈現紫色至紅棕色，但可被α-amylase與amyloglucosidase水解，故推測其為一高度分支之α- (1, 4；1, 6) phytoglycogen。不可消化之多醣部份，其單醣組成主要為阿拉伯醣、半乳醣、甘露醣與半乳糖醛酸。利用四級銨鹽選擇性錯和沉降、DEAE陰離子交換層析法、β-galactosidase酵素水解與銅離子錯和沉澱法，確認金線連多醣中含有半乳聚醣、甘露聚醣與低陰電荷的阿拉伯半乳聚糖，與高陰電荷的果膠多醣。其中半乳聚醣分子量約為116kDa，單醣組成中半乳糖莫耳比例為82.65 %；甘露聚醣分子量為844kDa，單醣組成中甘露糖莫耳比例為94.02 %；低陰電荷的阿拉伯半乳聚醣為一複雜混合物，分子量分佈範圍廣(2~817kDa)，平均分子量為359kDa，單醣組成主要為阿拉伯糖與半乳糖，莫耳比為22.38與40.95%，且可與Yariv antigen專一性結合產生明顯紅色沉澱反應，確定此為type-II Arabinogalactan (AG-II)果膠多醣，此類多醣已被證實具有免疫調節的功效；而高陰電荷之果膠多醣(AFP-P)，平均分子量為290kDa。在多醣各區分的活性試驗，金線連粗多醣與阿拉伯半乳聚醣具有明顯免疫活性刺激功效，都有明顯促進Raw 264.7 細胞產生NO與TNF-α。在金線連另含有豐富游離糖主要為葡萄糖、果糖與少量半乳醣和甘露糖，佔金線連乾草10.86%。水萃物中另含有機酸，佔金線連酒精上清液固型物的6.12±0.09 %，主要為蘋果酸、檸檬酸、草酸以及奎寧酸，仍然還有三種酸性物質，其結果仍需後續確認。

關鍵字

金線連 ； 阿拉伯半乳聚糖 ； 甘露聚醣 ； 半乳聚醣 ； 果膠多醣 ； 有機酸

並列摘要

Anoectochilus formosanus Hayata is an original species plant used as folk medicine in Taiwan. Previous research has shown many biological functional attributes of its water-extracts such as anti-hyperliposis, anti-inflammatory, antioxidation, anti-hyperglycemic effects. The polysaccharides of Anoectochilus formosanus Hayata have been associated with its immuno-regulatory effects. This study focused on screening and properties analysis of carbohydrates and organic acids in water-extracts of the whole herb, the most popular way to consume this plant. The content of crude polysaccharide obtained through hot-water extraction and ethanol precipitation was 14.81% of the dried whole plant. The content of α-D-glucans was 60 ~ 66% in crude polysaccharide and the α-D-glucans could be digested by α-amylase and amyloglucosidase and showed red-brown color as stained with iodine solution, not typical blue or purple color of starch. The results indicated that the α-D-glucan was a highly branched phytoglycogen with α-(1, 4; 1, 6)-linkages. The major sugar composition of non-digestible polysaccharide in crude polysaccharide was arabinose, galactose, mannose and galacturonic acid. One galactan, one mannan, one low anionic charged arabinogalactan with and one pectic polysaccharide fractions were obtained through quaternary ammonium salt selective precipitation, copper ion complexing, β-galactosidase digestion and DEAE-anionic column chromatographic methods. The galactan was composed of 82.65% of galactose with molecular-weight in the range of 116 kDa. The mannan was composed of 94.02% mannose with molecular-weight in the range of 844 kDa. The low anionic charged arabionogalactan was a complex polysaccharide and was composed of galactose arabinose, rhamnose, mannose, glucose and galacturonic and glucuronic acids. It could reacted with artificial β-D-glucosyl Yariv antigen to form red color precipitates indicating that it was a rhamnogalacturonan I carrying type II arabinogalactan ramified branches which was an bioactive polysaccharide found in many medical herbs. The bioactivity was confirmed on an in vitro test indicating that the arabinogalactan was the major component to stimulate RAW 264.7, a rodent macrophage cell line, to produce NO and TNF-α. The weight-average molecular-weight of the arabinogalactan fraction was 359 k Da with widely distributed range of 2-817 kDa. The pectic polysaccharide fraction was also a complex polysaccharide group which was composed of sugar components similar to arabinogalactan with different ratio and with molecular-weight in the range of 290 kDa. There were substantial amount (10.86%) of free sugars in dry A. formosanus plant including glucose and fructose and relative small amount of galactose and mannose. The content of organic acid was 6.12% on dry basis of EtOH suspensition in water extract including malic, citric, oxalic and quinic acids with three unknown acidic compounds remaining to be further elucidated.

並列關鍵字

Anoectochilus formosanus Hayata ； arabinogalactan ； mannan ； galactan ； pectic polysaacharides ； organic acids

參考文獻

阮昌廷。臺灣金線連水溶性多醣分子結構特徵。臺灣大學碩士論文。台北。2005。

劉新裕、張同吳、王昭月、張愛華、王順成。1998。金線連之品種特性與急毒性研究。中華農業研究。43(3):242~258。

蕭翌柱、高瑞隆、賴瑞聲、胡敏夫。2007。台灣金線連之產品開發與利用。行政院農業委員會農業試驗所生物技術組。林業研究專訊14(4):34-36。

Albersheim, P.; Nevins, D.J.; English, P. D. Karr A. A method for the analysis of sugars in plant cell-wall polysaccharides by gas-liquid chromatography. Carbohydr. Res. 1967, 5, (3), 340-345.

Aspinall G. O. 1959. Structural chemistry of the hemicelluloses. Adv. Carbohydr. Chem. 14:429-468.