Biochemical studies in tobacco plants: IV. N-malonylmethionine, metabolite of d-methionine in Nicotiana rustica

The metabolism of d-methionine-methyl- 14C in intact Nicotiana rustica plant has been studied, and the radioactive ninhydrin-negative spots from the acidic fraction of methanolic plant extracts were investigated. The bulk of the fraction activity was associated with a thioether-positive spot; the corresponding sulfoxide is believed to be an artifact. For comparison, 2,4-dinitrophenylhydrazones of α-keto-γ-methyl-thiobutyric acid (I), α-keto-γ-methylsulfinylbutyric acid (II), and α-keto-γ-methyl-sulfonylbutyric acid (III) were synthesized; the resolution of geometrical isomers of I- and II-hydrazones succeeded by chromatographic methods. By isotope dilution technique it has been established that only a negligible part of d-methionine-methyl- 14C was converted into I. The purification of the thioether-positive peak was performed by ion-exchange chromatography, ethyl acetate extraction, and preparative paper chromatography; as the final step, fractionation on Sephadex G-25 was used. Acid hydrolysis, and hydrazinolysis of the thioether-positive peak, its electrophoretic mobility, and the fact that the compound decarboxylated on heating gave evidence for N-malonylmethionine structure. This was confirmed by comparison with synthetically prepared N-malonylmethionine (V) and crystallization with the radioactive metabolite to constant activity.