Liquid chromatography-mass spectrometry (LC-MS) is preferred for analyzing small molecules such as polar metabolites, complex lipids, and drugs in biofluids [1]. However, capturing the full range of these compounds often requires multiple extraction methods or platforms [2]. We optimized an LC-MS workflow for extracting polar metabolites, complex lipids, and drugs from human plasma, followed by rapid LC-MS analysis. The extraction process utilized a bi-phase approach with a methanol/methyl tert-butyl ether mixture and water, enabling separate analyses of the organic phase (for complex lipids) and aqueous phase (for polar metabolites) for untargeted metabolomics and lipidomics [3]. To address the wide range of drug polarities, an additional extraction step using a methanol/ethanol mixture was performed. Using MS-DIAL software and combined MS/MS spectral libraries, we annotated over 600 complex lipids and polar metabolites from plasma samples of type 2 diabetes (T2D) patients and non-T2D controls. Targeted drug analysis quantified more than 40 drugs and selected metabolites at concentrations as low as 1–10 ng/mL, providing sufficient sensitivity for monitoring drug adherence. This study highlights metabolome and lipidome alterations in non-adherent T2D patients and discusses the tools available for data interpretation.

[1] Rakusanova S, Cajka T. Trend Anal Chem (2024) 180:117940.
[2] Rakusanova S, Cajka T. Trend Anal Chem (2023) 158:116831.
[3] Cajka T, et al. Int J Mol Sci (2023) 24(3) 1987.

IPHYS contact person: Tomáš Čajka, tomas.cajka@fgu.cas.cz