Improved analysis of related substances of cobamamide and the impurities research / 中国生化药物杂志

ABSTRACT

Objective To improve the related substances analysis method of cobamamide.And to further research light degradation characteristics of cobamamide.Methods Determined the related substances of solid and aqueous solution of cobamamide after light degradation by high performance liquid chromatography(HPLC) conditions,which is Merck Hibar C18 column (4.6 mm ×250 mm, 5μm), 0.05 mol/L KH2PO4 solutionacetonitrile (9010) -80% acetonitrile as mobile by gradient elution and detection wavelength 260 nm.And compared their light stability.The main three kinds of light degradation impurities were determinate from LC-MS.Results Gradient elution made the light degradation impurities separate better.The results of precision and reproducibility tests increased to RSD =0.2% (n=6) and RSD =8% (n=5) from RSD =3.8% (n =6) and RSD=38%(n=5). Cobamamide solution was very sensitive to light, the preparation should be strict dark operation.Two of the light degradation impurities were adenosine and hydroxycobalamin, with the relative response factor 2.5 and 0.7.Conclusion New method is specific, durable and reproducible, which can be used for quality control of cobamamide.