Application of 4-(4-nitrophenyl)-1,2,4-triazoline-3,5-(dione to analysis of 25-hydroxyvitamin D3 in human plasma by liquid chromatography/electron capture atmospheric pressure chemical ionization-mass spectrometry

The utility of 4-(4-nitrophenyl)-1,2,4-triazoline-3,5-dione (NPTAD) as a derivatization reagent in the analysis of 25-hydroxyvitamin D-3 [25(OH)D-3] using liquid chromatography/electron capture atmospheric pressure chemical ionization-mass spectrometry (LC/ECAPCI-MS) was examined. The derivatives of 25(OH)D-3 with NPTAD underwent electron capture in the APCI source in the negative-ion mode and provided 30-fold higher sensitivity compared to an intact compound. This derivatization-LC/ECAPCI-MS method was applied to a plasma 25-hydroxyvitamin D-3 assay, and gave satisfactory results in sensitivity, specificity, measurable range and throughput of the analysis.