Strategies for using post-column infusion of standards to correct for matrix effect in LC-MS-based quantitative metabolomics

Matrix effect limits the accuracy of quantitation of the otherwise popular metabolomics technique liquid chromatography coupled to mass spectrometry (LC-MS). The gold standard to correct for this phenomenon, whereby compounds co-eluting with the analyte-of-interest cause ionization enhancement or suppression, is to quantify an analyte based on the peak area ratio with an isotopologue added to the sample as an internal standard. However, these stable isotopes are expensive and sometimes unavailable. Here we describe an alternative approach: matrix effect correction and quantifying analytes using signal ratio with a post-column infused standard (PCIS). Using an LC-MS/MS method for 8 endocannabinoids and related metabolites in plasma, we provide strategies to select, optimize and evaluate PCIS candidates. Based on 7 characteristics, the structural endocannabinoid analogue arachidonoyl-2'-fluoroethylamide was selected as a PCIS. Three methods to evaluate the PCIS correction vs. no correction showed that PCIS correction improved values for matrix effect, precision and dilutional linearity of at least six of the analytes to within acceptable ranges. PCIS correction also resulted in parallelization of calibration curves in plasma and neat solution, for 6 of 8 analytes even with higher accuracy than peak area ratio correction with their stable isotope labeled internal standard, i.e. the gold standard. This enables quantification based on neat solutions, which is a significant step towards absolute quantification. We conclude that PCIS has great, but so far under-appreciated, potential in accurate LC-MS quantification.