2 METHYLENEDIOXY BRIDGE FORMING CYTOCHROME-P-450 DEPENDENT ENZYMES ARE INVOLVED IN (S)-STYLOPINE BIOSYNTHESIS

The enzymatic synthesis of the alkaloids (S)-[10-(OCH3)-H-3]scoulerine and (S)-[3-(OCH3)-H-3]cheilanthifoline is described. A microsomal preparation from Eschscholtzia californica cell suspension cultures in the presence of NADPH and O2 catalyses the formation of two methylenedioxy bridges in two consecutive steps: (S)-cheilanthifoline from (S)-scoulerine and (S)-stylopine from (S)-cheilanthifoline. Nandinine was not found to be an intermediate. The release of (HOH)-H-3 into the aqueous incubation mixture from the radiolabelled substrates served as an assay for both enzyme activities. The enzymes were characterized as to their substrate specificity, cofactor requirements and inhibition by cytochrome P-450 specific inhibitors. Both enzymes were drastically induced (12- and 5-fold) 20 hr after challenging the cell suspension culture with elicitor. The mechanism of enzymic methylenedioxy bridge group formation from an o-methoxyphenol function is discussed.