Abstract
This paper deals with studies on natural forms of vitamin B_<12> active substances in the cells of thermophylic methane bacteria. A purified extract was obtained by an extraction with hot alcohol and subsequent phenol treatment, instead of the usual extraction method with cyanide solution in order to avoid the transformation of cyanide-labile vitamin B_<12> analogues. The extract exhibited the coenzyme actvity in Abeles・Lee's intramolecular oxidation-reduction system. By paper ionophoresis performed in 0.5N acetic acid, five active zones on E.coli vitamin B_<12>-less mutant were observed ; three of them (tentatively named P1-P3 in order of mobolities) moved toward the cathode, one (P4) stayed near the origin, and the fifth (P5) migrated toward the anode. By cyanide treatment, disappearances of P1 and P2 and simultaneous increase in area of P4 were observed. In bioautogram using Ochromonas malhamensis, growth zones corresponding to P2 and P3 were not detected. From these results, it seems appropriate to conclude as follows : P1 is DBCC ; P2,non-cyano type B_<12> analogue lacking 5,6-dimethylbenzimidazolyl cobamide structure ; P3,pseudocobalamin ; P4,cyanocobalamin and P5,an unidentified protein-bound form with anionic character.
