一种基于胰蛋白酶酶解BSA获得小多肽库的新型手性固定相制备(英文)

Preparation of A Novel Chiral Stationary Phase Based on Library of Small Peptides Obtained by Digestion of Bovine Serum Albumin with Trypsin

介绍了一种基于小多肽库的高效液相色谱手性固定相(CSP)的制备.胰蛋白酶酶解牛血清白蛋白(BSA)的优化条件为:在pH8.2,37℃下,酶解6 h,所得到的小多肽库以最大截留分子量(MWCO)为6 000的超滤膜过滤,滤液为含分子量小于6000的多肽分子库,以羰基咪唑法将该小多肽库键合至氨基化硅胶上。与完整蛋白-CSP相比,该多肽-CSP拥有更高的多肽键合密度以及更好的稳定性。对流动相组成以及pH对保留和立体选择性的影响也进行了考察,在优化的条件下,D,L-色氨酸、D,L-酪氨酸以及手性药物R,S-盐酸苯海索在该新型多肽-CSP上均得到了很好的分离。

A novel chiral stationary phase（CSP） based on ＇library of small peptides for high performance liquid chromatography（HPLC） was prepared. Tile conditions for digestion of bovine serum albumin （BSA） with trypsin were optimized, and the results of which was ： enzymolysis time 6 h, enzymolysis pH 8.2, enzymolysis temperature 37℃. The obtained library of small peptides, which was ultrafihrated with ultrafihration membrane（ MWCO = 6 000） to discard Mw 〉 6 000 molecules, was bonded to aminopropyl - silica gels activated with 1,1＇ - carbonyldiimidazole （CDI）. The proposed CSP held higher density of peptides _ligand and was more steady than intact protein - CSP. D, L - tryptophan and D, L - tyrosine were well separated on the novel peptides - CSP. Chiral drug R, S - trihexypheni- dyl hydrochloride was also separated. Effects of the mobile phase composition and pH on retention and stereoselectivity were investigated too.