Development of simultaneous purification methodology for multiple synthetic peptides by reversed-phase sample displacement chromatography
Development of simultaneous purification methodology for multiple synthetic peptides by reversed-phase sample displacement chromatography
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DOI：
10.1016/s0021-9673(00)00751-2
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发表时间：
2000-09-29
影响因子：
4.1
通讯作者：
Hodges, RS
中科院分区：
文献类型：
--
作者：
Husband, DL;Mant, CT;Hodges, RS
我们开发了一种低压方案，旨在作为一种快速、简单且经济有效的程序，用于高效、平行纯化多种肽混合物。这是通过将我们新颖的反相样品置换色谱 (SDC) 方法（其中主要分离过程在没有有机改性剂的情况下进行）适应模块化固相萃取 (SPE) 技术来实现的。因此，粗制肽样品在过载条件下应用到由含有二氧化硅基反相填料的 SPE 管组成的萃取柱。通过施加真空将溶液抽过填料，在两级纯化装置中实现产物与疏水性和亲水性杂质的分离：短的预柱用作疏水性杂质的捕集器，而第二个较长的SPE柱用作产物分离柱。因此，在理想的 SDC 条件下，用 100% 水性溶剂洗涤将实现疏水性杂质保留在捕集器上。随着产物和亲水性杂质从捕集器转移到产物分离塔：亲水性杂质因此从产物分离器转移到废物中，仅留下产物保留在主柱上。在此初步评估中，该纯化系统在代表粗制肽混合物的模型合成肽系统的各种肽疏水性和粗制组成范围内，以良好的收率证明了产品与亲水性和疏水性杂质的出色分离。 (C) 2000 Elsevier Science B.V. 保留所有灯光。
We have developed a low-pressure protocol, designed as a rapid, simple and cost-effective procedure for the efficient and parallel purification of multiple peptide mixtures. This was achieved through adaptation of our novel reversed-phase sample displacement chromatography (SDC) method, where the major separation process takes place in the absence of organic modifier, to modular solid-phase extraction (SPE) technology. Thus, crude peptide sample is applied at overload conditions to extraction columns consisting of SPE tubes containing silica-based reversed-phase packing. By applying a vacuum to draw the solution through the packing, product separation from hydrophobic and hydrophilic impurities is accomplished in a two-stage purification unit: a short pre-column functions as a trap for hydrophobic impurities, while a second, longer SPE column is used as a product isolation column. Thus, under ideal SDC conditions, washing with a 100% aqueous solvent will achieve retention of hydrophobic impurities on the trap. with displacement of product and hydrophilic impurities from the trap to the product isolation column: hydrophilic impurities are thus displaced off the product isolation to waste, leaving only product retained on the main column. In this initial evaluation, this purification system has demonstrated excellent separation of product, in good yield, from both hydrophilic and hydrophobic impurities over a wide range of peptide hydrophobicity and crude composition for model synthetic peptide systems representing crude peptide mixtures. (C) 2000 Elsevier Science B.V. All lights reserved.