Determination of cobalamins and cobinamides by microbore reversed-phase HPLC with spectrophotometric, ion-spray ionization MS and inductively coupled plasma MS detection

Microbore reversed-phase HPLC was optimized for the separation of cyanocobalamin, 5′-desoxyadenosylcobalamin (coenzyme B12), methylcobalamin, hydroxocobalamin, aquocobalamin and cobinamide dicyanide isoforms. Various detection modes: UV at 278 nm, ion-spray (IS)-MS and direct-injection nebulization ICP-MS were examined and compared. IS-MS allows one to identify, unambiguously, the eluted compounds, whereas ICP MS was found to be more sensitive than any other on-line detection techniques described for the cobalamin analogues, so far allowing to achieve detection limits down to ca. 10 ng ml−1. The usefulness of the methods developed was demonstrated for the determination of cobalamins present in pharmaceutical preparations.