牛胰岛素的酰胺化反应及其六酰胺产物的分离及鉴定

Total Amidation of Insulin and Characterization of Insulin Hexamide

牛胰岛素用1％三氟化硼甲醇溶液反应后所得六甲酯与饱和氨甲醇溶液反应,在室温下约经144小时可完全酰胺化。反应过程用红外光谱及酰胺含量分析测定。所得酰胺化粗产品经Sephadex G-75凝胶层析可以将胰岛素六酰胺化合物从其聚合物中分离提纯。由红外光谱、紫外光谱及圆二色散光谱测定表明:牛胰岛素的六酰胺化合物的构象与胰岛素有很大不同,其生物活性也丧失殆尽。

Total amidation of six carboxyl groups in bovine zinc insulin has been achieved by esterification in methanol containing 1% boronfluoride followed by reaction with ammonia. Amidation was found to occur most effectively in the methanol solution saturated with ammonia and went to completion in 144 hrs at room temperature, as determined by the amide analysis of the product. Insulin hexamide was separated from its polymeric by-products by Sephadex G-75 gel filtration in 1% formic acid. Infra red spectroscopy of the product after different times of amidation indicated the progressive loss of free carboxyl groups in insulin, and confirmed the complete amidation after 144 brs. Ultra-violet and circular dichroism spectra of insulin hexamide in a physiological solution indicated a profound change in the conformation from its parent protein, insulin.