Development of comprehensive two-dimensional HPLC for high-performance separations in the future

开发用于未来高性能分离的综合二维 HPLC

基本信息

批准号：
14340234
负责人：
TANAKA Nobuo
金额：
$ 7.55万
依托单位：
Kyoto Institute of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2002
资助国家：
日本
起止时间：
2002 至 2004
项目状态：
已结题

项目摘要

The objective of this research project was to increase peak capacity of HPLC by developing simple and comprehensive two dimensional (2D) HPLC. By employing a monolithic silica-C18 column at the second dimension (2nd-D), we were able to obtain the following results.(1) We proved the concept of simple and comprehensive 2D-HPLC in a reversed-phase-reversed-phase mode by using a particle-packed column of fluoroalkyl-bonded silica as a first-dimension (1st-D) column, and C18- or pentabromobenzyloxypropyl-bonded monolithic silica as a 2nd-D column.(2) By utilizing the difference in selectivity provided by organic solvents, 2D-HPLC was carried out using monolithic silica C18 columns both at 1st-D and at 2nd-D.(3) 2D-HPLC was carried out by using an ion-exchange column packed with polymer particles at 1st-D and a monolithic silica-C18 column or a monolithic silica capillary column at 2nd-D for peptide separations.The use of these systems produced (1)peak capacity of up to 1100 in methanol-water mobile phase, (2)peak capacity of 700 or greater for hydrocarbons and benzene derivatives, respectively. The ion-exchange-reversed-phase 2D-HPLC system provided (3)peak capacity of 700 or greater for BSA digest with a short monolithic silica column at 2nd-D, and peak capacity of about 500 for BSA digest by using a 0.2 mmlD, 10 cm long capillary column at 2nd-D, respectively. Optimization of 2D-HPLC conditions were attempted to show the possibility of achieving peak capacity of greater than 2000 in one hour. The flow rate, retention factors of solutes, and column length at 2nd-D were shown to dominate the peak capacity of a 2D-HPLC system. Monolithic silica columns were found to be best suited for 2D-HPLC based on their high permeability and high performance at high linear velocity of mobile phase.

该研究项目的目标是通过开发简单且全面的二维 (2D) HPLC 来提高 HPLC 的峰容量。通过使用二维（2nd-D）整体硅胶-C18柱，我们得到了以下结果。（1）我们在反相中证明了简单而全面的2D-HPLC的概念-反相模式，使用氟烷基键合硅胶颗粒填充柱作为第一维 (1st-D) 色谱柱，使用 C18- 或五溴苄氧基丙基键合整体硅胶作为2nd-D 柱。(2) 利用有机溶剂提供的选择性差异，在 1st-D 和 2nd-D 均使用整体式硅胶 C18 柱进行 2D-HPLC。(3) 进行 2D-HPLC通过使用一维填充聚合物颗粒的离子交换柱和二维整体硅胶-C18柱或整体硅胶毛细管柱进行肽分离。这些系统 (1) 在甲醇-水流动相中产生高达 1100 的峰容量，(2) 对于烃类和苯衍生物分别产生 700 或更高的峰容量。离子交换反相 2D-HPLC 系统 (3) 在二维短整体硅胶柱上提供了 700 或更高的 BSA 消解峰容量，并使用 0.2 mmlD 的 BSA 消解提供了约 500 的峰容量, 10 cm 长的毛细管柱在 2nd-D 处分别。尝试优化 2D-HPLC 条件以表明在一小时内实现大于 2000 峰容量的可能性。结果显示，二维 HPLC 系统的峰容量由流速、溶质保留因子和二维柱长决定。人们发现整体硅胶柱最适合 2D-HPLC，因为它具有高渗透性和流动相高线速度下的高性能。