PREPARATION OF RIBOFLAVIN SPECIFICALLY LABELED IN THE 5'-HYDROXYMETHYL TERMINUS USING A VITAMIN-B-2-ALDEHYDE-FORMING ENZYME FROM SCHIZOPHYLLUM-COMMUNE

A method is described for synthesis of riboflavin selectively labeled in the hydrogens at the 5'-hydroxymethyl position. In this method, a v itamin B-2-aldehyde-forming enzyme from Schizophyllum commune is used to specifically and completely oxidize the 5'-hydroxymethyl of ribofla vin to the 5'-aldehyde. This reaction is monitored spectrophotometrica lly by the reduction of 2,6-dichlorophenolindophenol at 600 nm. Appear ance of aldehyde product was directly quantitated by reverse-phase hig h-performance liquid chromatography. Product is extracted from the inc ubation mixture by phenol after saturation with (NH4)(2)SO4 and then f urther purified by benzyl alcohol extraction. The 5'-aldehyde is reduc ed with appropriately labeled sodium borohydride to yield the vitamin specifically labeled in the 5'-hydroxymethyl group.