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Determination of vitamins D2 and D 3 in selected food matrices by online high-performance liquid chromatography–gas chromatography–mass spectrometry (HPLC-GC-MS)
- Nestola, Marco, Thellmann, Andrea
- Analytical and bioanalytical chemistry 2015 v.407 no.1 pp. 297-308
- cholecalciferol, detection limit, ergocalciferol, food matrix, high performance liquid chromatography, ions, isomers, isotope labeling, margarine, mass spectrometry, spectrometers
- An online normal-phase liquid chromatography–gas chromatography–mass spectrometry (HPLC-GC-MS) method was developed for the determination of vitamins D₂and D₃in selected food matrices. Transfer of the sample from HPLC to GC was realized by large volume on-column injection; detection was performed with a time-of-flight mass spectrometer (TOF-MS). Typical GC problems in the determination of vitamin D such as sample degradation or sensitivity issues, previously reported in the literature, were not observed. Determination of total vitamin D content was done by quantitation of its pyro isomer based on an isotopically labelled internal standard (ISTD). Extracted ion traces of analyte and ISTD showed cross-contribution, but non-linearity of the calibration curve was not determined inside the chosen calibration range by selection of appropriate quantifier ions. Absolute limits of detection (LOD) and quantitation (LOQ) for vitamins D₂and D₃were calculated as approximately 50 and 150 pg, respectively. Repeatability with internal standard correction was below 2 %. Good agreement between quantitative results of an established high-performance liquid chromatography with UV detection (HPLC-UV) method and HPLC-GC-MS was found. Sterol-enriched margarine was subjected to HPLC-GC-MS and HPLC-MS/MS for comparison, because HPLC-UV showed strong matrix interferences. HPLC-GC-MS produced comparable results with less manual sample cleanup. In summary, online hyphenation of HPLC and GC allowed a minimization in manual sample preparation with an increase of sample throughput.