HPLC-ESI-MS analysis of Vitamin B12 in food products and in multivitamins-multimineral tablets

Methods for the determination of Vitamin B12 remain limited due to their low sensitivity and poor selectivity. In the present work, a simple and sensitive HPLC-ESI-MS method for determining Vitamin B12 in food products and in multivitamin-multimineral tablets was developed. Vitamin B12 was extracted from food products with 50 mM sodium acetate buffer (pH 4.0) in the presence of sodium cyanide. Total Vitamin B12 content in food product can be obtained by efficient enzymatic hydrolysis to release the bound Vitamin B12. Vitamin B12 was quantified with ginsenoside Re as internal standard (I.S.) after their separations on a C18 column with a gradient of mobile phase made of water and acetonitrile. MS with SIR mode at m/z 930.8 was used for Vitamin B12 quantification. The calibration graphs plotted with five concentrations of Vitamin B12 was linear with a regression coefficient r2 = 0.9994. The intra-assay R.S.D. and the inter-assay R.S.D. were 2.6% (n = 5) and 3.5% (n = 6), respectively. The recoveries evaluated at spiking three different concentrations on fortified products were above 93%. The method has been applied successfully in the determination of Vitamin B12 in fortified food products and multivitamin-multimineral tablets.