Development of High-Performance Liquid Chromatographic Method for Vitamin D3 Analysis in Pharmaceutical Preparation

In the present study, a rapid, simple and economical reversed phase-hplc procedure has been developed for the determination of vitamin d3 and vitamin d2 in pharmaceutical preparations. the vitamins were separated isocratically on a teknokroma c18 column (150~4.6mm, 3.5ƒêm particle size) with a mobile phase consisting of isocratic acetonitrile and methanol (75:25, v/v) operated at 40oc with retention times less than 12 min. the eluted vitamins were identified and monitored on a diode-array detector (dad) at 265nm. the linearity of the method was excellent (r2 > 0.999), over the concentration range of 0-10 µg/ml. the statistical evaluation of the method was examined performing intra-day and inter-day calibrations and were found to be satisfactory, with 2.600 ± 0.078% and 6.267 ± 0.050% accuracy and 0.001± 0.002% to 2.144 ± 0.050% precision results. mean recovery of vitamin d3 was 98.2 ± 0.349% for different spiking levels. detection limit was found to be 25ng/ml. no interference was found from other fat soluble vitamins (vitamin a, vitamin e and vitamin e acetate) that are commonly presents with vitamin d pharmaceutical preparation. the method was applied for the analysis of sixteen solid samples of pharmaceutical preparation (calcium with vitamin d). the results show only 31.25% of the tested samples give satisfactory agreement with the declared values.