Determination of the native forms of vitamin B₁ in bovine milk using a fast and simplified UHPLC method

The aim of this study was to develop a high-throughput UHPLC method for the determination vitamin B₁ active compounds; thiamin, thiamin monophosphate and thiamin diphosphate in bovine milk. In order to sustain the native vitamin B₁ phosphorus esters, sample preparation is crucial. Various acids as well as commonly used enzymes and their enzyme mixtures were compared. Method accuracy was confirmed using certified reference material as well as comparison with the corresponding CEN method, and was found to be satisfactory. Studied milk samples showed significant amounts of thiamin monophosphate, which can make up to 53.9% of the total vitamin B₁ content in commercial milk, and up to 78% in raw milk. Moreover, a tremendous variation of the total content of vitamin B₁ was observed between single cows, which ranged from 0.24mg/L up to 0.54mg/L of total vitamin B₁.