可溶性猪软骨Ⅱ型胶原蛋白的提取与鉴定  

Isolation and identification of soluable collagen typeⅡ from porcine articular cartilage

作　　者：宁志刚[1] 王富友[1] 崔运利[1] 熊娟[1] 杨柳[1] 

机构地区：[1]第三军医大学西南医院关节中心,重庆400038

出　　处：《重庆医学》2011年第10期954-955,958,共3页Chongqing medicine

基　　金：国家863计划基金资助项目(2006AA02A125)

摘　　要：目的从猪膝关节软骨中提取纯化Ⅱ型胶原蛋白,并对其进行鉴定。方法选择猪膝关节软骨为提取原料,用盐酸胍去除蛋白多糖、胃蛋白酶两步消化、氯化钠盐析、超纯水透析、离心浓缩等方法提取纯化Ⅱ型胶原蛋白;采用SDS-PAGE、氨基酸成分分析对提取的Ⅱ型胶原蛋白进行鉴定;利用冻干的方法计算提取的浓度。结果 SDS-PAGE电泳结果显示提取的Ⅱ型胶原蛋白分子量约120 kD;氨基酸成分分析显示甘氨酸、脯氨酸和丙氨酸含量最高,并且可以检出羟脯氨酸、羟赖氨酸,符合Ⅱ型胶原特征;该法提取的Ⅱ型胶原蛋白浓度为66 mg/mL。结论从猪关节软骨提取的Ⅱ型胶原蛋白纯度高,方法简便,提取的胶原蛋白浓度更高。Objective To isolate and purify collagen type Ⅱ from porcine articular cartilage and identify its purity. Methods The porcine articular cartilage was selected as raw material. Guanidine hydrochloride was chosen to withdraw the proteoglyeans. Twostep digestion of pepsin,salting of sodium chloride,dialyzing of ultrapure water,centrifuge enrichment were applied for extracting; The identification was done by SDS-PAGE and amino acid analysis ; The concentration of extracted collagen type Ⅱ was measured by cool-dry. Results It was found that the molecular weight of extracted collagen type Ⅱ was about 120KD by SDS-PAGE;The content of GLY,PRO and ALA were highest by amino acid analysis,and Hypro, Hylsy can be detected,which accorded with the characteristics of collagen type Ⅱ ;the concentration of extracted collagen type Ⅱ was 66mg/ml by cool-dry. Conclusion The results suggest that the extracted collagen type Ⅱ has the high purity and concentration with a more convenient method.

关 键 词：Ⅱ型胶原蛋白 软骨 电泳 聚丙烯酰氨凝胶 

分 类 号：R593.22[医药卫生—内科学]