High-Throughput Parallel LC⧸UV⧸MS Analysis of Combinatorial Libraries

Abstract

This chapter discusses the optimization of an eight-channel parallel liquid chromatography (LC)/UV/mass spectrometry (MS) system for high-throughput LC/UV/MS analysis of large combinatorial libraries. Because the LC gradient is divided into eight LC columns by a simple splitter, the flow fluctuation has been continuously monitored and minimized using a standard mixture during analysis to ensure performance consistency among the eight channels. To preserve the separation integrity in the total ion chromatogram, the zero dead volume T-joint used to split the flow (after UV detection) should be best placed as close to the eight-way MUX inlet as possible. A flow rate of 12 ml/min on eight 2.1×50 mm Polaris C18 columns was optimal for general purposes in an investigation presented in the chapter. This system could analyze more than 3000 compounds per day for a gradient separation with a cycle time of 3.5 min. The chapter also presents the results of several LC/UV/MS analyses carried out in 15 months using two eight-channel parallel LC/UV/MS systems. It was found that it was beneficial to evaluate a few representative compounds from each library and optimize ion optics to make product identification simple and reliable. This parallel system has enabled simultaneous evaluation of eight compounds and significantly improved the speed of optimization. The identity and purity of every single product could be obtained from OpenLynx in 10 96-well plate per batch process and transferred into an Excel spreadsheet for the entire library.