Conclusion
In our sample preparation protocol, the drying-reconstitution step was identified as the main cause of metabolite losses or increased data variability during metabolomics analysis by LC-HRMS. Furthermore, PE was affected by the type of matrix. Isotopologue internal standards fully compensate losses or enhancements.
Methods
The sample preparation strategies were evaluated by calculating the sample preparation effect, matrix factor, and process efficiency (PE) for 16 tobacco exposition-related metabolites, including nicotine, nicotine-derived nitrosamine ketone, their major metabolites, and glutathione, using isotopically-labelled internal standards. Samples were analyzed by liquid chromatography (LC) coupled to high-resolution mass spectrometry (HRMS).
Results
A sample drying step increased losses or variability for some selected metabolites. By avoiding evaporation, good sample preparation recovery was obtained for these compounds. For some metabolites, the cell or culture type impacted PE and matrix factor.