Octanoate-thiokinase, an enzyme from liver mitochondria, was found to catalyze the formation of auxin-CoA esters with several different auxins in the presence of adenosinetriphosphate and coenzyme-A. Evidence was provided to show that indoleacetyl-adenosinemonophosphate was an intermediate in the formation of indoleacetyl-CoA. This intermediate was supplied to the enzyme as the synthetic anhydride, and could lead either to the formation of indoleacetyl-CoA when supplied with CoA or to the formation of indoleacetic acid plus adenosinetriphosphate when supplied with pyrophosphate. Indoleacetyl-CoA was shown to be the intermediary product in the enzymatic formation of indoleacetyl-glycine. 2.4-Dichlorphenoxyacetic acid and α-naphthylacetic acid were not measurably conjugated with glycine under the same conditions. The results are discussed as to their implications in auxin metabolism in plants.
Coenzyme A Thiol Esters, Cinnamic Acids Acyl-CoA derivatives of several hydroxylated cinnamic acids have been synthesized in 30 to 50% yield via a. acyl phenyl thiol esters, b. acyl N-hydroxysuccinimide esters, and c. glucocinnamoyl derivatives. Properties of the intermediates have been determined. The cinnamyol-CoA thiol esters were characterized by their chromatographic behaviour and UV spectra. The molar extinction coefficients of these important intermediates in plant phenylpropane metabolism have been unequivocally determined. Recently published values13 for the molar extinction coefficients of these derivatives are incorrect; the methodological reason for this error has been established.
The biosynthesis of the anthraquinones, alizarin I and purpurin II, in Rubia tinctorum (madder) has been investigated. Acetate- [2-14C] proved to be a precursor only of ring C and in part of the keto groups of the quinone ring. While phenylalanine- [U-14C] was not incorporated into I and II, shikimic acid was found to be a good precursor of these compounds. The radioactivity of shikimic acid-[1,2-14C] could be localized only in ring A; shikimic acid-[U-14C] however was incorporated in toto; the ring being transformed into ring A of the anthraquinone, and the carboxyl group of this acid into one of the keto groups of ring B.
These findings demonstrate the existence of a biosynthetic pathway for the formation of anthraquinones in Nature, which is entirely different from the polyacetate route.
The biosynthesis of 5-hydroxy-1,4-naphthoquinone (juglone) was studied by supplying radioactive precursors to leaves of Juglans regia plants. A chemical degradation of the juglone molecule was devised (Fig. 1). With these methods it was shown that the ring atoms of shikimic acid are incorporated into the benzene ring of the quinone, while the carboxyl group of this acid is transformed to 50% into each of the keto groups of the quinone ring (C-atoms 1 and 4 of juglone. Tab. 3). This suggested a symmetrical molecule to be an intermediate in the formation of juglone — most probable 1,4-naphthoquinone. This compound was synthetized with 14C in the positions 2, 3, 9, and 10 and was found to be a good precursor of juglone in Juglans as well as for 2-hydroxy-1,4-naphthoquinone in Impatiens plants (Tab. 4) . 3,4-Dihydroxybenzaldehyde (Tab. 2) and chorismic acid (Tab. 7) which have been suggested previously as intermediates in the biosynthesis of naphthoquinones are no precursors of juglone. The source of three carbon atoms of the quinone nucleus remains to be determined; one or two of these carbon atoms (C2 and/or C3 of juglone) are formed from the methylen carbon of malonate (Tab. 5 and 6); surprisingly, however, the carboxyl carbons of malonic acid are not incorporated. The substitution of shikimic acid occures in the position 6 of this acid as could be judged from the degradation of juglone labelled with shikimic acid [ 1,2-14C] (Fig. 2; Tab. 3). 1,4-Naphthoquinone (or naphthohydroquinone) is postulated as an important intermediate in the biosynthesis of naphthoquinone derivatives in higher plants.
Feeding experiments with glucose- (2-14C), phenylalanine- (3-14C), tyrosine- (3-14C) and p-coumaric acid- (3-14C) showed that the latter three substances are incorporated in good yields into p-hydroxybenzoic acid in leaves of Catalpa ovata. Kinetic experiments showed that p-hydroxybenzoic acid is formed from phenylalanine via p-coumaric acid and the subsequent β-oxidation of the side chain. p-Hydroxybenzoic acid can also be synthetised by hydroxylation of benzoic acid, but this does not seem to be the biosynthetic route in Catalpa.
Phenylalanine- (3-14C) is also incorporated into benzoic acid, protocatechuic acid, and vanillic acid by different plants; the radioactivity of the β-C atom of the amino acid was found in each case to be located in the carboxyl group of the C6 — C1 acid. This suggests that in higher plants the benzoic acids are formed from the corresponding cinnamic acids via β-oxidation.