Large Scale Displacement High Performance Liquid Chromatography for Separation of Peptides and Proteins

本研究计划的目标是开发高效、经济、高纯度的蛋白质和多肽置换层析回收系统。为了满足人类治疗用途的纯度要求，生物技术行业采用了借用蛋白质化学的最先进的分离技术。在分辨率最高的分离过程中是高效液相色谱(HPLC)，它已经在大规模纯化蛋白质方面找到了有限的用途。除了柱和设备的高成本之外，阻碍更广泛地接受过程规模的高效液相纯化的主要特征是较大的溶剂体积，柱容量低，生物活性丧失，尤其是对于在该浓度下不可逆变性或聚集的蛋白质，以及在典型的层析操作过程中遇到的条件下。置换层析是一种可获得更高吞吐量、更有效地利用固定相和流动相以及高分辨率的替代方法。此外，置换层析可在常规离子交换色谱柱和反相色谱柱上进行，不需要像亲和分离技术那样生产昂贵的外来柱试剂，它涉及的操作步骤很容易放大。

The goal of this research program is to develop systems for efficient, economical, high-purity recovery of proteins and peptides by displacement chromatography. In order to meet the purity requirements for human therapeutic use, the biotechnology industry employs state-of-the-art separation techniques borrowed from protein chemistry. Among the highest resolution separation processes is high performance liquid chromatography (HPLC), which has found a limited use on a large scale for purification of proteins. The main features impeding broader acceptance of process scale HPLC purification, in addition to the high cost of columns and equipment, are the large solvent volumes, low column capacity and loss of biological activity, particularly for proteins that irreversibly denature or aggregate at the concentrations, and under the conditions encountered during typical chromatographic operations. Displacement chromatography is an alternative to yield much higher throughput, more efficient utilization of stationary and mobile phases, and high resolution. In addition, displacement chromatography can be performed on conventional ion exchange and reversed phase chromatography columns, does not require the expensive production of exotic column reagents as do affinity separation techniques, and it involves operating steps that are readily scaled up.