Fast and simple liquid chromatographic determination of nonphosphorylated thiamine in infant formula, milk, and other foods.

A very fast and simple method for determination of nonphosphorylated thiamine in infant formula products, milk, and other nonfortified foods using reverse-phase ion-pairing liquid chromatography (LC) has been developed. Sample preparation consists of merely acid treatment to precipitate protein, followed by gravity filtration. No concentration, extraction, derivatization, or preliminary column cleanup is necessary. The chromatography is done on muBondapack C18 with an aqueous mobile phase containing 0.15% sodium hexane sulfonate, 20% MeOH, 1.5% HOAc, and 0.1% EDTA at a flow rate of 2.5 mL/min. Ultraviolet detection at 248 nm is used. A typical run takes 7 min, and 60 samples can be processed in 4 h. Results average from 96 to 104% of theory for the infant formula products analyzed. A 99 to 103% recovery of spike has been demonstrated. Method precision is good (2 to 4% RSD, short-term, and 2 to 5% RSD, long-term, depending on sample type). Peak separation from thiamine phosphate esters is achieved. Specificity is demonstrated by UV spectral scan and absorbance ratios. Equivalency to a microbial method (validated against the official AOAC fluorometric method) was established. The method is used for high-volume quality control testing of milk-based infant formula products in the ready-to-use, concentrate, or powder form.