Voltammetric behaviour of vitamins D2 and D3 at a glassy carbon electrode and their determination in pharmaceutical products by using liquid chromatography with amperometric detection

Abstract

Cyclic voltammetry was used to study the oxidation of vitamin D₂(ergocalciferol) and vitamin D₃(cholecalciferol) at a planar glassy carbon electrode. The electrode reaction for cholecalciferol was found to be dependent on the apparent pH between 4.95 and 6.10, and pH independent between pH 6.10 and 8.65 when the solutions contained 90% methanol; this suggested a pK_a value of 6.10 for vitamin D₃. Similar behaviour was exhibited by ergocalciferol, and a pK_a value of 6.35 was found. The peak currents for both vitamins were found to be dependent on the methanol concentration of the supporting electrolyte. The peak currents were also found to be dependent on the ionic strength of the acetate buffer (pH 6.0) over the range 0.1–1.0 mol dm^–3. Both substances were oxidized in one step, which was found to be an irreversible reaction; the final product for vitamin D₂ can undergo absorption at the electrode surface. The parent compounds could be undergoing oxidation at the triene moieties. The optimum mobile phase for liquid chromatography with amperometric detection was found to be 95% methanol–0.05 mol dm^–3 acetate buffer (pH 6.0); the detector was operated at a potential of + 1.3 V (versus Ag—AgCl), and a linear response was obtained for vitamin D₃ over the range from 10 to 100 ng injected; for vitamin D₂ the response was linear from 20 to 200 ng injected. Extracts of pharmaceutical products were separated on reversed-phase columns prior to amperometric detection of the vitamins. Cholecalciferol was successfully determined in a multivitamin table, and ergocalciferol in a multivitamin liquid preparation.