Separation and study of corrinoid cobalt-ligand isomers by high-performance liquid chromatography

Vitamin B12 belongs to a group of complex organo-cobalt compounds, the corrinoids. "Complete" corrinoids, such as B12, contain a nucleotide as the cobalt alpha-(lower)ligand. This nucleotide is also connected to the periphery of the corrin ring. The "incomplete" corrinoids, in contrast, contain simple cobalt alpha-ligands, such as water or cyanide. Using analytical reversed-phase and anion-exchange high-performance liquid chromatography (HPLC), we have been able to study the behavior of several aquocyano-"incomplete" corrinoids: three isomeric cobinic acid pentaamides, cobinamide, and cobyric acid, all of which exist as thermally unstable isomers, which separate during HPLC. All but one of these corrinoids gave isomer mixtures of 1:1, the exception giving mixtures of 2:1 to 3:1. The separated stereoisomers had different retention times and were collected from analytical columns for further study.