Simultaneous quantitative analysis of eight vitamin D analogues in milk using liquid chromatography–tandem mass spectrometry

Development of an assay for total vitamin D content in milk and application to study the effect of pasteurisation in breastmilk

“…Although there is a wide range of derivatising reagents, 4-phenyl-1,2,4-triazole-3,5-dione (PTAD) is the most popular due to its ease of availability (93). While it has been successfully applied to enhance the detectability of vitamin D compounds in MS/MS detections, the derivatisation reaction with PTAD has not been systematically optimised (93).…”

“…Chemical derivatisation has been used to address this problem and it has been shown to be a valuable strategy to enhance vitamin D compound ionisation efficiencies in API sources; Cooksontype reagents are the most common derivatives for this class of compounds (93,123,171).…”

“…It has been clearly demonstrated that when isotopically labelled standards are used as internal standards, there is no significant difference in the results when calibration standards are prepared either in mobile phase solvent or in matrix matched medium (119,139,150). The samples as well as calibration solutions were therefore prepared in mobile phase (85% B, being the mobile phase composition at the beginning of the gradient).…”

“…For the analysis of derivatised forms of the standards and samples, a previously developed method in our laboratory was used (150). The derivatised and underivatised forms were prepared to obtain the same final concentrations.…”

“…The milk samples were processed and analysed according to an established LC-MS/MS method for the analysis of a range of vitamin D analogues in milk (150).…”