Strategies for Using Postcolumn Infusion of Standards to Correct for Matrix Effect in LC-MS-Based Quantitative Metabolomics

The 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 coeluting 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 a signal ratio with a postcolumn infused standard (PCIS). Using an LC-MS/MS method for eight endocannabinoids and related metabolites in plasma, we provide strategies to select, optimize, and evaluate PCIS candidates. Based on seven 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 the 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 six of eight 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 toward absolute quantification. We conclude that PCIS has great, but so far underappreciated, potential in accurate LC-MS quantification.