Rapid, easy analysis of urinary vanillylmandelic acid for diagnostic testing of pheochromocytoma by liquid chromatography tandem mass spectrometry.

Vanillylmandelic acid (VMA), as one of the most important catecholamine metabolites, is commonly used to aid in diagnosis of pheochromocytoma. This study develops a rapid and simple high-throughput LC-MS/MS method for the measurement of urinary VMA. Without sample pretreatment, the urine specimens were mixed with internal standard (IS) solution for direct analysis by LC-MS/MS in two minutes. VMA and VMA-d3 were detected in the multiple-reaction monitoring mode using the specific transitions m/z 197.0→137.0 and 200.0→140.0, respectively. This method was validated for consistent linearity from 1.0 to 250.0μM with CVs≤3.12%, excellent recovery, good stability and low carryover. The lowest limit of quantification (LLOQ) was 0.125μmol/L for VMA with CV of 14.1%, and the lower limit of detection (LOD) was 0.025μmol/L. Intra-assay and inter-assay imprecision values (CVs) for VMA were all below 2.11%. Dilution linearity was investigated with a satisfied mean accordance of 105%. Method comparison of LC-MS/MS and microcolumn chromatography in our lab was performed and the reference interval was established in agreement with that of the Mayo Clinic. All these results demonstrate that this validated LC-MS/MS approach shows improved accuracy and reproducibility compared with microcolumn chromatography. The low sample volume, simplicity, rapidity, and robustness of the method make it suitable for use as a high-throughput assay in routine clinical biochemistry laboratories.