IEX/RP二维液相色谱中弱保留组分收集模式的构建及系统评价

以十通阀和捕集柱接口形式,构建了弱阴离子交换/反相(WAX/RP)二维液相色谱分离系统.通过将第一维离子交换色谱分析中的前部集中洗脱出的弱保留组分收集后单独分析,剩余样品进一步采用二维液相色谱分析,可以有效避免第二维色谱柱对特殊样品局部集中流出导致的第二维分离超柱容量问题,提高了系统的整体分离能力.使用4种蛋白胰蛋白酶酶解后的多肽样品评价该系统,在不分流的系统中共检测到115个峰.对第一维分离的前15 min分流后得到的组分单独分析,共识别出58个峰,后续的二维分离中共得到78个峰.2种方法相比,第二种方法检测峰数增加了21个,一些低丰度的组分在弱保留组分收集后被识别.

Construction and Evaluation of Weak Retention Components Collection Mode-IEX/RP Two-Dimensional Liquid Chromatography System

An ion-exchange/reversed phase liquid chromatography(2D-LC) system was developed based on trap column and valve interface. Weak anion exchange ( WAX ) chromatography mode was used in the first dimension and reversed-phase(RP) chromatography mode was used in the second di-mension. The two dimensions were connected by a 10-port two-position valve and two trap columns. The weak retained components of the first-dimensional were collected and analyzed separately, and then the remained components were further separated by 2D-LC system. The overloading of the sec-ond-dimensional column was effectively avoided and the separation power of the 2D-LC system was improved greatly. Tryptic digest of four proteins were separated on this system. 115 peaks were identified in the traditional 2D-LC separation mode. However, in the weak retention components collection mode, 58 peaks were detected in the front 15 min elution of the first dimension. 78 peaks were detected in the subsequent 2D-LC analysis. Compared with that of traditional 2D-LC, another 21 peaks were identified and some low abundance components were identified after weak retained components were collected.