Development of a validated liquid chromatography method for the simultaneous determination of water soluble vitamins in food supplements and pharmaceutical dosage Forms with ultra-violet detection

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In the present work a fast, simple and reliable method for the simultaneous isolating and determining of the vitamins [thiamin (B1), riboflavin (B2), nicotinamide (PP), pyridoxine (B6), folic acid (B9) and ascorbic acid (C)] in food supplements has been developed. The chromatographic separation is carried out on a reversed phase ACE 5 C18 with eluting at a flow rate of 1 ml/ min using a linear gradient with phosphate buffer- methanol, starting with 0.05 M phosphate buffer pH 2.5 for 3 min, changing to 45% methanol in 13 min, changing to 80% methanol in 3 min with equilibrating for ten min. The separation was achieved within 25 min and the detection was performed at 270 nm. The calibration graphs plotted with six concentrations of each vitamin were linear with a regression coefficient R2>0.996. The statistical evaluation of the method was examined performing intra-day (n=6) and inter day calibration which found to be satisfactory, with high accuracy and precision results. The average coefficients of variation of intra and inter-day assays were 1.2, 2.7 for thiamine hydrochloride, 2.4, 3.4 for riboflavin, 1.5, 3.3 for pyridoxine hydrochloride, 0.9, 2.3 for nicotinamide, 2.5, 2.9 for folic acid, and 2.1, 3.1 for ascorbic acid. The average of recoveries of thiamine hydrochloride, riboflavin, pyridoxine hydrochloride, nicotinamide, folic acid and ascorbic acid were % 100±1.4, %100±1.5, %104±1.1, %102±3.2, %100±0.8 and %99±2.1 respectively. The simplicity of the method should make it highly valuable for quality control of premixes and food supplements in pharmaceutical and food industries.

water soluble vitamins; pharmaceutical formulation; liquid chromatography