Simultaneous determination of VD2, VD3, 25(OH) D2, and 25(OH) D3 in human plasma using electrospray LC–MS/MS as well as its application to evaluate VD plasma levels in depressive,schizophrenic patients and healthy individuals

Vitamin D measurements in biological fluids by liquid chromatography–tandem mass spectrometry (LC–MS/MS) have been widely used but remain challenging at very low concentration levels. Rapid, high recovery, sensitive and reliable measurements of vitamin D, as well as its primary metabolites using LC–MS/MS are urgently needed for a routine clinical laboratory. Herein, we reported a novel electrospray LC–MS/MS method for determining vitamin D and its primary metabolites using the supported liquid extraction method to achieve higher recoveries, with optimized pH values to achieve optimal derivatization efficiency for higher sensitivity and selected chromatographic conditions to shorten the separation time. The method has been validated with respect to selectivity, recovery, matrix effects, accuracy and precision, stabilities, carryover and dilution effects. The method has been successfully applied to quantify the VD plasma concentrations of depressive, schizophrenic patients and healthy individuals. The result showed that there were significant differences in plasma VD levels between mental disorder patients with healthy individuals, and the total VD levels in mental disorder patients were much higher than healthy individuals, which might require larger clinical samples for validation.