摘要
目的用柱层析法分离纯化人血清白蛋白降解片段,优选最佳工艺。方法首先采用葡聚糖凝胶Sephadex G-100和Superdex 75对人血清白蛋白降解产物进行分离纯化,再用羧甲基纤维素和DEAE-纤维素在不同pH的磷酸盐缓冲液中下进一步纯化,通过分离度、蛋白回收率优选最佳工艺。结果 Superdex 75比Sephadex G-100分离效果好,将人血清白蛋白降解产物分离纯化为粗品Ⅰ和Ⅱ,蛋白回收率为61.82%±2.01%;羧甲基纤维素较DEAE-纤维素分离效果好,将粗品Ⅰ分离纯化为人血清白蛋白多肽片段A299-585,蛋白回收率为62.07%±1.12%;将粗品Ⅱ分离纯化为人血清白蛋白多肽片段A1-123和A124-298,蛋白回收率为66.29%±1.79%。结论 Superdex 75和CM-Sepharose FF不仅对人血清白蛋白的降解产物具有良好的分离度,而且有较好的蛋白回收率。
OBJECTIVE To study on the separation and purification of degradation fragment of human serum albumin（HSA） in the chromatography column,and to optimize the process condition. METHODS Firstly, Sephadex G - 100 and Superdex 75 were used respectively to separate and purify the degradation products of HSA. Secondly, the carboxymethylcellulose （CM - Sepharose FF） and DEAE - cellulose（ DEAE - Sepharose FF） were used respectively under the different pH of phosphate buffered saline for further purification. The process condition was optimized by the resolution and protein recovery. RESULTS The separating effect of Superdex 75 was better than that of Sephadex G- 100, and the degradation products were separated into two parts Ⅰ and Ⅱ , and protein recovery was 61.82% ± 2.01%. CM -Sepharose FF was superior to DEAE -Sepharose FF,part Ⅰ was fractionated into HSA polypeptide fragment A299_585 and protein recovery was 62.07% ± 1.12%. Part Ⅱ was fractionated into HSA polypeptide fragment At -123 and A124 -298 ＇ and protein recovery was 66.29% ± 1.79%. CONCLUSION It is experimentally shown that high resolution and protein recovery of the degradation products of HSA are obtained in fixed bed chromatography with Superdex 75 and CM - Sepharose FF.
出处
《华西药学杂志》
CAS
CSCD
北大核心
2013年第5期476-478,共3页
West China Journal of Pharmaceutical Sciences
基金
国家自然科学基金资助项目(批准号:81202489)
关键词
人血清白蛋白
多肽片段
分离纯化
柱层析法
分离度
Human serum albumin
Polypeptide fragment
Separation and purification
Column chromatography method
Resolution
