Bertucci C, Bartolini M, Gotti R, Andrisano V
Dipartimento di Scienze Farmaceutiche, Università di Bologna, Via Belmeloro 6, 40126 Bologna, Italy.
J Chromatogr B Analyt Technol Biomed Life Sci. 2003 Nov 25;797(1-2):111-29. doi: 10.1016/j.jchromb.2003.08.033.
This review addresses the use of high-performance liquid chromatography (HPLC) and capillary electrophoresis (CE) as affinity separation methods to characterise drugs or potential drugs-bio-polymer interactions. Targets for the development of new drugs such as enzymes (IMERs), receptors, and membrane proteins were immobilized on solid supports. After the insertion in the HPLC system, these immobilized bio-polymers were used for the determination of binding constants of specific ligands, substrates and inhibitors of pharmaceutical interest, by frontal analyses and zonal elution methods. The most used bio-polymer immobilization techniques and methods for assessing the amount of active immobilized protein are reported. Examples of increased stability of immobilized enzymes with reduced amount of used protein were shown and the advantages in terms of recovery for reuse, reproducibility and on-line high-throughput screening for potential ligands are evidenced. Dealing with the acquisition of relevant pharmacokinetic data, examples concerning human serum albumin binding studies are reviewed. In particular, papers are reported in which the serum carrier has been studied to monitor the enantioselective binding of chiral drugs and the mutual interaction between co-administered drugs by CE and HPLC. Finally CE, as merging techniques with very promising and interesting application of microscale analysis of drugs' binding parameters to immobilized bio-polymers is examined.
本综述探讨了使用高效液相色谱（HPLC）和毛细管电泳（CE）作为亲和分离方法来表征药物或潜在药物与生物聚合物的相互作用。将诸如酶（IMERs）、受体和膜蛋白等新药开发靶点固定在固体支持物上。插入HPLC系统后，这些固定化生物聚合物通过前沿分析和区域洗脱方法用于测定具有药学意义的特定配体、底物和抑制剂的结合常数。报告了最常用的生物聚合物固定化技术以及评估固定化活性蛋白量的方法。展示了使用减少量的蛋白质使固定化酶稳定性增加的实例，并证明了在回收再利用、重现性以及对潜在配体进行在线高通量筛选方面的优势。在获取相关药代动力学数据方面，综述了有关人血清白蛋白结合研究的实例。特别报道了一些论文，其中通过CE和HPLC研究了血清载体，以监测手性药物的对映体选择性结合以及联合给药药物之间的相互作用。最后，探讨了CE作为一种融合技术在对固定化生物聚合物进行药物结合参数的微尺度分析方面非常有前景且有趣的应用。
