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Liquid chromatography/electrospray ionization-tandem mass spectrometry (LC/ESI-MS/MS) enables the accurate and precise quantification of various analytes at very low concentrations, but it has room for improvement in analysis throughput. Multiplexing of samples in the same injection could be a promising procedure for enhancing the analysis throughput. This could be achieved by derivatization of the multiple samples with multiple isotopologous reagents. In this study, a sample-multiplexed LC/ESI-MS/MS assay using the 1-[(4-dimethylaminophenyl)carbonyl]piperazine (DAPPZ) isotopologues ( 2 H 0 -, 2 H 3 -, and 2 H 6 -forms) was developed and validated for the simultaneous determination of primary bile acids in three different plasma samples in a single run. The developed method had satisfactory intra- and inter-assay precisions (≤ 2.3 and ≤ 4.2%, respectively) and accuracy (99.0 - 100.3%), and could reduce the total LC/ESI-MS/MS run time by more than 60% for 42 samples compared to the conventional method. Thus, the derivatization with the DAPPZ isotopologues worked well for enhancing the throughput of the LC/ESI-MS/MS assay of the bile acids.

Enhancing LC/ESI-MS/MS Throughput for Plasma Bile Acid Assay by Derivatization-based Sample-Multiplexing