藥用真菌多醣體成份的分離方法之研究

Abstract

關於藥用真菌的傳統性味功效，早在許多古老的中藥典籍內便有詳細 的記載，近年來更利用現代科學技術的深入分析、臨床研究，證實這些珍 貴的傳統中藥材，像冬蟲夏草、靈芝、雲芝等，的確具有相當重要的生物 藥理活性，而這些藥用真菌的有效成份大部份是屬於多醣體，即所謂的生 物反應調節劑(Biological ResponseModifer, BRM)，能提供訊號起動原 有的生物能力，符合傳統中醫藥『扶正固本』的說法，且其為無毒或低毒 性的免疫促進劑，具有調節免疫機能、抗腫瘤等活性，因此，這些藥用真 菌對於未來在醫藥領域上的應用，極具開發潛力。 由於天然物成份相 當複雜，純化不易。回顧以往的研究，通常是使用這些藥用 真菌的粗抽 提物進行藥理活性研究，至於主要的藥效成份仍有待進一步的瞭解；因此 本論文便是針對，如何將粗抽多醣體內的有效成份分離，以利未來藥性成 份的深入研究。 我們利用酒精沉澱、熱水萃取及冷凍乾燥等步驟所得 的棉絮狀粗抽多醣體，以Gel Filtration Chromatography進行進一步的 純化，經由Anthrone Method及Bradford Assay分別測定其醣類及蛋白質 的含量，然而以此種方式所得的分離純化效果不佳；所以，再將原來的棉 絮狀粗抽多醣體，先以超音波振盪，分別處理15、30、60、90分鐘後，再 以Gel Permeation Chromatography進行成份的分析，於適當濃度的有機 移動相下，可使其各成份有效的被區別開來，顯示超音波振盪及有機移動 相效應，有助於粗抽成份內多醣體及蛋白質的純化；此結果對於日後探討 中藥有效成份的快速分離及結構分析，甚至對於有效成份的確認及其作用 機制的研究，將有很大的助益。 The fruiting bodies of a variety of fungi are used in traditional Chinesemedicines. Cordyceps sinensis, Ganoderma lucidum and Coriolus versicolor arethree medically important fungi which have attracted great attention in recentyears for their biological and pharmacological activities. There have been in the literature many studies on the relationship between the function and structure of polysaccharides. Herbal fungi polysaccharides, which are known asbiological response modifers( BRM), can modulate the recipient's immunological activities, thus presenting a great potential in the treatment of cancers and other diseases. The polysaccharides extracted from medicinal fungi is usually a mixture of proteins and polysaccharides. At this stage, the compnents are generally not yet purified, therefore the pharmacologically active compounds are not identifiable. The objecttive of this research is to develop an effective and convenient method so as to further purify the biologically active components for pharmacokinetic and clinical studies in the future. We have extracted crude polysaccharides from the above three species, and then treated the extracts with ultrasonication for different durations. The water-soluble suspension is analyzed by using the Gel Permeation Chromatographywith various methanol concentrations. The results showed sharp resolution. In summary, our results showed that the ultrasonication and organic mobile phase in couple with the GPC analysis can greatly enhance the resolution, hence the biologically active compounds can be better purified for strcutural identification and for further pharmacokinetic and clinical studies.