Lim H K, Chan K W, Sisenwine S, Scatina J A
Drug Safety and Metabolism, Wyeth-Ayerst Research, Monmouth Junction, New Jersey 08852, USA.
Anal Chem. 2001 May 1;73(9):2140-6. doi: 10.1021/ac001112b.
A LC-LC/MS/MS method has been developed that significantly increases the throughput in metabolism screening of drug candidates during lead optimization in discovery. This was accomplished by the reduction of sample preparation time through an on-line extraction of a drug and its metabolites from microsomal proteins using turbulent flow chromatography. Following its injection onto a column at turbulent flow, the drug and its metabolites are backwashed onto a reverse-phase column via on-line column switching and resolved chromatographically at a laminar flow of 2 mL/min. This tandem turbulent-laminar flow chromatographic system in a total cycle time of 8 min can achieve adequate separation of isomeric metabolites of venlafaxine, haloperidol, or adatanserin. Further improvement in throughput can be achieved by multiplexing both microsomal stability assessment and metabolite profiling into a single analysis. This is made possible by the ability of the ion-trap mass spectrometer to perform simultaneously multiple-reaction monitoring for microsomal stability and data-dependent multiple-stage mass spectrometric analysis for metabolite profiling within a single LC analysis. Such a LC-LC/MS/MS approach can dramatically shorten the time for providing metabolism feedback to the drug discovery process.
已开发出一种液相色谱-串联质谱（LC-LC/MS/MS）方法，该方法可显著提高在发现阶段先导化合物优化过程中药物候选物代谢筛选的通量。这是通过使用湍流色谱法从微粒体蛋白中在线提取药物及其代谢物来减少样品制备时间实现的。在湍流条件下将其注入色谱柱后，药物及其代谢物通过在线柱切换反冲至反相柱上，并在流速为2 mL/min的层流条件下进行色谱分离。这种串联的湍流-层流色谱系统在总循环时间为8分钟内，能够实现文拉法辛、氟哌啶醇或阿坦色林的同分异构代谢物的充分分离。通过将微粒体稳定性评估和代谢物谱分析复用至单次分析中，可进一步提高通量。这是通过离子阱质谱仪在单次液相色谱分析中同时进行微粒体稳定性的多反应监测和代谢物谱分析的数据依赖多级质谱分析的能力实现的。这种LC-LC/MS/MS方法可大幅缩短为药物发现过程提供代谢反馈的时间。
