Detection of cobamide coenzymes in microorganisms by the ionophoretic-bioautographic method

The ionophoretic-bioautographic method of detecting cobinamide and cobamide vitamins in millimicrogram quantities has been modified and applied to the detection of cobamide coenzymes in microbial extracts. For the detection of cobamide coenzymes, the cells must be extracted with a neutral buffer in the dark. When cells are extracted with either an alkaline or acid cyanide solution or when the extract is exposed to light, only the cobamide vitamins and other coenzyme decomposition products are observed. Cobamides with the ionophoretic mobility of the cobamide coenzymes have been shown to be present in extracts of Escherichia coli B, E. coli 113-3, Bacillus megatherium, Streptomyces fradiae, Propionibacterium arabinosum, P. shermanii, and Clostridium tetanomorphum. The coenzymes present in several of these organisms have been specifically identified. E. coli 113-3 was shown to convert vitamin B 12 to the 5,6-dimethylbenzimidazolylcobamide coenzyme (coenzyme B 12). When grown with cobinamide and benzimidazole, the benzimidazolylcobamide coenzyme was formed.