Alhama J, López-Barea J, Toribio F
Departamento de Bioquímica y Biología Molecular, Facultad de Veterinaria, Universidad de Córdoba, Spain.
J Chromatogr. 1991 Nov 8;586(1):51-9. doi: 10.1016/0021-9673(91)80024-b.
An epoxy-activated silica column (50 cm x 0.45 cm I.D.) was derivatized with 8-[6-aminohexyl)amino]-2'-phosphoadenosine-5'-diphosphoribose; the bound ligand concentration was 11.4 mumol/g of dry silica, and the useful loading capacity was 2.3 mg of glutathione reductase. The new high-performance liquid chromatographic column specifically retained NADP(+)-dependent enzymes, which were quantitatively eluted specifically by NADP+ or, with better resolution, by potassium chloride. The new high-performance liquid chromatographic support was applied to the purification of glutathione reductase and glucose-6-phosphate dehydrogenase from cell-free extracts of baker's yeast, fish liver and rabbit hemolysates, with high recoveries and excellent purification factors.
一个环氧活化硅胶柱（内径50 cm×0.45 cm）用8-[(6-氨基己基)氨基]-2'-磷酸腺苷-5'-二磷酸核糖进行衍生化；结合配体浓度为11.4 μmol/g干硅胶，有效负载量为2.3 mg谷胱甘肽还原酶。这种新型高效液相色谱柱能特异性保留依赖NADP(+)的酶，这些酶可用NADP+进行定量洗脱，或者用氯化钾洗脱，分辨率更高。这种新型高效液相色谱载体用于从面包酵母、鱼肝和兔溶血产物的无细胞提取物中纯化谷胱甘肽还原酶和葡萄糖-6-磷酸脱氢酶，回收率高，纯化因子优异。
