摘要： 本實驗以板栗殼為原料，用水煮醇沉法提取了板栗殼粗多糖，測得其提取率為1.3%；用考馬斯亮藍G-250法對提取的板栗殼粗糖蛋白的蛋白質含量作了測定，結果為2.73%；得到的粗糖蛋白用酶+Sevag法脫蛋白，處理了12次后無明顯的蛋白層；紫外光譜檢測脫蛋白后的板栗殼粗多糖，顯示在280nm處已無吸收峰，說明蛋白質已經基本除盡；脫蛋白后得到的板栗殼粗多糖用苯酚-硫酸法測糖含量，測得粗多糖中己糖含量為74%。
用DEAE-Sepadax Fast Flow陰離子交換層析柱對板栗殼粗多糖進行了分離純化，經蒸餾水和不同濃度的NaCl溶液梯度洗脫后得到兩個組分cnps1，cnps2，用Sephacryl 100 HR層析柱對組分cnps2作了純度檢驗，結果顯示cnps2并不是單一的均多糖，表明經一次陰離子交換柱層析后所得到的板栗殼多糖并不是一個單一的均多糖，需要調整分離條件，或改變填料類型，或改變洗脫劑種類進行進一步的純化研究。通過紅外光譜檢測層析前后的板栗殼多糖的組成變化，發現雖然分離純化沒有得到精多糖，但是粗多糖中的某些組分已經被分離并富集，從而產生了不同的吸收峰。
最后，采用Fenton法對板栗殼粗多糖進行了抗氧化活性的研究，結果顯示板栗殼粗多糖在劑量為1-20mg/ml范圍內，對羥基自由基具有一定的清除效果。但在同等劑量下弱于陽性對照Vc，20 mg/ml的板栗粗多糖的清除率為35.1%，而陽性對照Vc(20mg/ml)的清除率為66.08%。
關鍵詞：板栗殼；多糖；提取；分離純化；抗氧化活性
Study on the extraction, purification and antioxidant activity of polysaccharide in Castanea mollissima shell
Abstract: In this study, the Castanea mollissima shell was extracted with water and Castanea mollissima shell crude polysaccharide was obtained with a productivity of 1.3%. The protein content of the Castanea mollissima shell crude polysaccharide measured with Coomassie brilliant blue G-250 method was 2.73%. The crude polysaccharide was deproteined with enzyme and Sevag, after handling 12 times there was no obvious protein layer left. UV detection showed that there was no absorption peak at 280 nm, that is to say ,the protein in the crude polysaccharide had been removed entirely. The polysaccharide content of the deproteined crude polysaccharide was measured with phenol and sulfuric acid, hexose had a content of 74%.
Then using DEAE - Sepadax Fast Flow anion exchange chromatography column to purify chestnut shell crude polysaccharides. Two components cnps1, cnps2 were abtained with the mobile phase distilled water and NaCl solution of different concentration. The purity of components cnps2 was measured by differential refractive index detector with Sephacryl 100 HR column, results showed that it was not uniform. That is to say, the crude polysaccharidea handled one time by anion-exchange column chromatography was not uniform. Thus, separation conditions, the filling or the eluting agent is need to change for further purification. Characterized the Chestnut polysaccharide before and after the separation and purification by infrared spectroscopy, we found that though uniform polysaccharide could not been abtained, some components of the crude polysaccharides had been isolated and enriched, thereby different absorption signal was observed.
Finally, the antioxidant activity of chestnut shell crude polysaccharide was studied by Fenton reaction. The results proved that chestnut shell crude polysaccharide in the dose range from 1mg/ml to 20mg/ml had scavenging effects on hydroxyl radical. However, it is weaker than the positive control Vitamin C in the same dose , the clearance rate of the former in the dose of 20 mg / ml was 35.1%, while that of the later was 66.08%.
Key Words：Castanea mollissima shell; Polysaccharides; Extraction; Purification; Antioxidant activity
