Ladrón de Guevara O, Estrada G, Antonio S, Alvarado X, Guereca L, Zamudio F, Bolívar F
J Chromatogr. 1985 Dec 6;349(1):91-8. doi: 10.1016/s0021-9673(00)90637-x.
A method for the isolation, identification and quantification of human insulin A and B chains by high-performance liquid chromatography (HPLC) is described. These chains were isolated from a peptide mixture produced by E. coli with modified genes obtained by genetic engineering. The method is based on the use of hydrophilic reagents, forming ion pairs in a reversed-phase column. Because some undesirable effects resulting from the use of phosphoric acid were observed, especially with the B chain, a new HPLC method was developed for each of the two human insulin chains. The use of trifluoroacetic acid as a counter ion for the A chain and of formic acid for the B chain led to the rapid isolation and purification of each chain by HPLC. The advantage of this method is that it provides a highly pure product, which was identified by polyacrylamide gel electrophoresis and amino acid analysis.
描述了一种通过高效液相色谱（HPLC）分离、鉴定和定量人胰岛素A链和B链的方法。这些链是从大肠杆菌产生的肽混合物中分离出来的，该混合物是通过基因工程获得的修饰基因产生的。该方法基于使用亲水性试剂，在反相柱中形成离子对。由于观察到使用磷酸会产生一些不良影响，尤其是对B链，因此针对两条人胰岛素链分别开发了一种新的HPLC方法。使用三氟乙酸作为A链的抗衡离子，使用甲酸作为B链的抗衡离子，通过HPLC实现了每条链的快速分离和纯化。该方法的优点是提供了高纯度的产物，通过聚丙烯酰胺凝胶电泳和氨基酸分析对其进行了鉴定。
