Fractionation and characterization of water-solubleolysaccharides from Codonopsis pilosula ( Franch.) Nannf.
|Keywords:||黨參;果聚醣;果膠多醣;阿拉伯半乳聚醣;同質半乳醛酸聚醣;鼠李半乳醛酸聚醣-Ⅰ;Codonopsis pilosula;Pectic polysaccharides;fructan;Arabinogalactan;homogalacturon;RhamnogalacturonanⅠ||Issue Date:||2009||Abstract:||
中藥黨參為桔梗科植物黨參 (Codonopsis pilosula (Franch.) Nannf.) 根之乾燥物，其多醣被認為與免疫調節活性有關，本研究之目的在探討黨參水溶性多醣組成並經劃分後研究其化學性質。根據溶解度與多醣帶電荷性的不同，作為基本的分離技術，並針對果聚醣與果膠多醣作為研究的目標物。果聚醣方面以鍵結與分子量分布為目標，果膠多醣則針對所含之鼠李半乳醛酸聚醣Rhamnogalacturonan-Ⅰ結構做探討。中藥黨參含有26.65%果寡醣與果聚醣(或稱菊糖)，其重量平均分子量為6.42kDa (DP = 40)，聚合度分佈範圍為DP = 7 ~ 112，可被80度熱水所萃出，萃取時增加乙醇濃度或降低萃取溫度則降低果寡醣的萃取率。黨參飲片用熱水萃取物並經酒精沉降所得之粗多醣產率為12.86%，選用四級銨塩選擇性錯和沉澱具有高陰電荷性的多醣，於上清液中留下中性與低電荷多醣其回收率為10.28%(佔水溶性多醣之79.94%)。高陰電荷性沉澱區分之回收率為黨參飲片之2.58% (佔水溶性多醣之20.06%)，其單糖主要組成為半乳糖醛酸(69.91%)為主，另含有較少量之半乳糖、阿拉伯糖與鼠李糖，重量平均分子量為240.46 kDa，為一主體為homogalacturonan (HG or pectic acid)之果膠多醣中。上清液區分中含有不帶陰電荷之果聚醣收率為粗多醣之48.15%，依照fructanase酵素之降解可判斷此果聚醣具有(2,1)-β-鍵結之骨幹；其重量平均分子量為3.04 kDa (DP = 19)。不與四級銨塩選擇性錯和沉澱之上清液區分中另含有兩個富含阿拉伯糖與半乳糖的多醣區分，能以DEAE陰離子交換樹脂區分開來，其一為不帶電荷的Arabinogalactan區分(DEAE-Fraction I)與帶低量醛醣酸區分。Arabinogalactan區分單醣組成以半乳糖(62.95%)為主，阿拉伯糖與鼠李糖(26.89%)次之，分子量為369.5kDa；帶低醛醣酸區分為一分子量分布很廣的混合多醣，除含有半乳糖、阿拉伯糖與鼠李糖外，另有顯著含量的半乳糖醛酸，此區分可與人工抗原Yariv antigen產生專一性結合，出現紅色的沉澱。此區分之單醣組成和人工抗體反應與由其他中草藥中分離出之具有免疫調節與補體活化功效具有arabinogalactan 分支之Rhamnogalacturonan I有相似之分子特徵。
Dangshen, dried roots of Codonopsis pilosula ( Franch.) Nannf., is an important traditional Chinese medicine. Hot-water extractable polysaccharides of dangsen have been associated with its attributes of immunomodulatory activities. The main goal of this research was focusing on profiling and characterizing water-soluble polysaccharides of dangsen. Solubility and charge selective precipitation were employed to separate different polysaccharide fractions. Fructo-oligosacchride and fructan (or known as Inulin) and pectic polysaccharides have been selected as major targets because they are play important roles as active components in many herbal medicines. The fructan analysis was focused on linkage determination and molecular weight distribution. The study of pectic polysaccharides was focused on structural characteristics of rhamnogalacturonan I. The content of fructo-oligosaccharide and furctan of dangshen was found to be 26.65% and the weight-average molecular weight was 6.42 kDa (DP = 40) with degree of polymerization in the range of DP = 7 ~ 112. The fructo-oligosaccharides and fructans were hot-water extractable at 80C. Increasing ethanol concentration and decreasing extracting temperature decreased the extraction ratio. Yield of crude polysaccharides from sliced and dried dangsen, a common consuming form, was 12.86% by weight. Quaternary ammonium salt effectively separated high anionic polysaccharides through selective precipitation. The yield of polysaccharide precipitate and supernatant was 2.58% and 10.28%, repectively. The major sugar composition of the polysaccharide precipitate was galactouronic acid (69.91% molar ratio) with relative small amount of galactose, arabinose and rhamnose. It is a homogalacuronan (HG or pectic acid) component of pectic polysaccharide with weight-average molecular weight of 240.46 kDa. The fructan, remaining in the supernatant of quaternary ammonium reacted solution, was found to be the most abundant component in the crude polysaccharides with the ratio of 56.81％ and the weight-average molecular weight was 3.04 kDa (DP= 19). According to its susceptibility to inulinase and fructanase, the backbone of the fructan was found to be (2,1)-β-linkage. Other components found in the supernatant were two arabinogalactan enriched fractions, one neutral (DEAE-FractionI) and one low anionic charged, which could be further separated by using DEAE-anion exchange chromatography. The neutral arabinogalactan fraction was composed of galactose (62.95%), arabinose and rhamnose with molecular-weight of 369.5 kDa. The low anionic charged polysaccharide fraction was compose of galacturonic acid、arabinose、rhamnose and galactose and could reacted with artificial β-D-glucosyl Yariv antigen to form pink color indicating that it was a rhamnogalacturonan I carrying type II arabinogalactan ramified branches which was an bioactive polysaccharide found in many medical herbs.
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