Identification of the product of heme degradation catalyzed by the heme oxygenase system as biliverdin IX alpha by reversed-phase high-performance liquid chromatography.

Biliverdins formed from heme by a microsomal preparation and a reconstituted heme oxygenase system were each converted to their dimethyl esters and analyzed for isomeric composition by reversed-phase high-performance liquid chromatography, using a column of mu Bondapak C18 (Waters Associates); on this column, the dimethyl esters of four biliverdin IX isomers, that is IX alpha, IX beta, IX gamma, and IX delta, have been shown to be eluted separately in the order IX alpha, IX beta, IX delta, and IX gamma, when developed with methanol/water. The analysis indicated that the enzymatically formed biliverdins were exclusively IX alpha; the elution profile exhibited no other significant elution peak due to other biliverdin isomers. It was concluded that the heme oxygenase system cleaves the heme ring specifically at the alpha-methene bridge to yield biliverdin IX alpha.