Design of HPLC method and stability studies of cyanocobalamin injection

Thumbnail Image
Journal Title
Journal ISSN
Volume Title
An accurate and selective HPLC with ultraviolet absorbance detection was developed for the quantification of vitamin B12 (cyanocobalamin) in vitamin B12 injections. The method developed involves an isocratic mode of elution of cyanocobalamin from a reversed-phase 150mm, 4.6mm, 5μm hypersil base-deactivated (BDS) C-18 column. Cyanocobalamin was quantitatively analysed using the HPLC method comprising methanol/ phosphate buffer pH7 (30 / 70 v/v) using ascorbic acid as an internal standard. Separation for quantitative determination was achieved within 6 minutes at a flow rate of 1.4ml/min .The absorbance of the drugs was monitored at 278nm at a detector sensitivity of 0.03. The standard calibration curves were linear with correlation coefficient (R2) of 0.996. The limit of quantification (LOQ) and detection (LOD) were 0.069 ug/ml and 0.0228ug/ml respectively. Four brands of cyanocobalamin injections sampled from some pharmacies and health facilities were analysed with the method and then compared with the standard UV method. The mean percentage recovery of the stated content of four batches of three brands of the injections were 99.00 ± 3.95, 96.05 ± 0.85, 96.00 ± 5.23, 94.36 ±8.73 as against a standard UV method va1ues of 100.33 ± 4.67, 96.97 ± 0.50, 96.22 ± 6.20, 94.47 ± 10.64 respectively. Moreover, within and between run relative standard deviations were 1.682% and 1.656% respectively for cyanocobalamin. These values were all less than the maximum threshold value of 2% for a method to be good as a precise method according to the ICH guidelines . All the validation parameters were assessed statistically and proven to be valid.
A Thesis submitted to the Department of Pharmaceutical Chemistry, Kwame Nkrumah University of Science and Technology In partial fulfillment of the requirements for the degree of MASTER OF PHILOSOPHY, 2012