Application of Chromatography. XXXVI. Biosynthesis of Riboflavin. (2). Riboflavin-synthesizing Enzyme extracted from Eremothecium ashbyii.

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The crude enzyme solution, prepared by extracting the ground mycelium of Er. ashbyii with a phosphate buffer and dialyzing the extract against the same buffer solution, was incubated with 6,7-dimethylribolumazine at 37°. Paper partition chromatography of the reaction mixture gave a yellow and a purple fluorescent spot. The former spot (Rf 0.3 ; solvent-system : EtOH・BuOH・H_2O) was cut out and extracted with hot water, and the ultraviolet spectrum of the extract was measured, showing that it is identical with that of riboflavin. The riboflavin was determined by bioassay with L. casei and by various chemical methods such as the lumiflavin-fluorescence method and others. Next, investigation was made on relationship between quantity of the resulting riboflavin and reaction time, quantity of the crude enzyme solution added, and concentration of the substrate, and further on the optimal pH and temperature for the action of the enzyme solution. Lastly, the above purple-fluorescent spot (Rf 0.23 ; solvent-system : EtOH・BuOH・H_2O) was proved to be that of 6-methyl-7-hydroxyribolumazine, and a new route was proposed for the formation of this compound. Further, the fact that the lumazine derivative was not affected by the crude enzyme solution made more certain the authors'previous assumption that the compound is a final product in the metabolism by Er. ashbyii.
社団法人日本薬学会の論文
1958-12-15

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