Speciation of vitamin B12 analogues by liquid chromatography with flame atomic absorption spectrometric detection

Vitamin B12 analogues and aqueous cobalt are separated by liquid chromatography using a reversed-phase column (Spherisorb ODS-2) and a mobile phase consisting of a linear gradient from a 25:75 (vv) mixture of methanol and 0.085 M phosphoric acid solution buffered at pH 5.2 with triethanolamine to a 60:40 mixture over 5 min. The vitamins are selectively detected by measuring cobalt with an atomic absorption spectrometer directly interfaced to the Chromatograph. A simple T-piece is used to compensate with air the difference between the nebulizer uptake rate and the Chromatographic flow-rate (1.5 ml min−1). Recoveries of the components from spiked preparations ranged from 90.8 to 108%. The method is applicable to the analysis of pharmaceuticals for cobalamins.