3 -hydroxy-4-methylanthranilic acid

Actinomydn.—The phenoxazinone skeleton of the actinomycins (181) has been shown to derive from two molecules of tryptophan via 3-hydroxy-4-methylanthranilic acid (185),methionine serving as the source of the aromatic methyl groups.Depression of the incorporation of radioactive tryptophan into actinomycin by kynurenine (182) and 3-hydroxy-4-methylkynurenine (184) indicated that (182) and (184) could well be biosynthetic intermediates. In contrast 3-hydroxykynurenine (183) was found to have little or no effect on tryptophan incorporation, thus casting doubt on its participation in actinomycin biosynthesis. However, both [ Hjkynurenine [as (182)] and [ H]-3-hydroxykynurenine [as (183)] have recently been shown to be efficient precursors for actinomycin in Streptomyces antibioticus. 3-Hydroxykynurenine (183) is therefore implicated in actinomycin biosynthesis and further substantive evidence is provided for the pathway tryptophan —> (182) — (183) — (184) —> (185) —> actinomycin (181). 

Birch (1958) had suggested that acetate rather than tryptophan may play an important role for s mthesis of the actinomycin chromophore. He postulated that the 3-hydroxy-4-methylanthranilic acid precursor might arise from four acetate molcules through a process of oxidation, reduction and amination. If acetate constituted the source of the anthranilate precursor, eight moles of acetate would be required for synthesis of one mole of the actinomycin chromophore and it would be expected that the actinocin formed would possess appreciable C-label. The results of an experiment with sodium acetate-2- C indicate that acetate is not directly involved as a precursor of the actinomycin chromophore (Sivak, Meloni, Nobili and Katz, I962). Of the total radioactivity incorporated into actinomycin by 5. antihioticus following addition of acetate- C, only 4% was present in the chromophore. On the other hand, the amino acid, proline, in the peptide portion of the actinomycin molecule possessed appreciable radioactivity. 

The enzymatic synthesis of the actinomycin chromophore also has been described (Weissbach and Katz, 1961 Katz and Weissbach 1962 Salzman, Weissbach and Katz, unpublished results). An enzyme, designated phenoxa-zinone synthetase, was obtained from sonic extracts of S. antihioticus and was shown to catalyze the oxidative condensation of o-aminophenols to the corresponding phenoxazinone. When 3-hydroxy-4-methylanthranilic acid (4MHAA) was used as substrate, actinocin was synthesized. The enzyme has been purified approximately 25- to 30-fold from sonic or French pressure cell extracts of the organism by means of standard protein fractionation procedures (ammonium... 

The oxidative condensation of 3-hydroxy-4-methylanthranilic acid toactinocin is represented by the following equation ... 

Hydroxy-3-methylanthranil is methylated normally with methyl iodide and sodium carbonate to give the 7-methoxy derivative.135 Anthranil carboxylic acids are converted into their acid chlorides by thionyl chloride,208 and their methyl esters by diazomethane.169 The acid chloride of 5-chloro-3-phenylanthranil-6-carboxylic acid participates in the Friedel-Crafts acylation of benzene to give 6-benzoyl-5-chloro-3-phenylanthranil (67%).208... 

Similar hydrogen bonding is also obs ved in o>-0 -tolyl)acstophenone enol (3) and AT-methylanthranilic add, while in 2-hydroxy-3-methyl-benzoic acid there exists an internal hydrogen bond between the adjacent hydroxy- and carbojqr-groups. The cisoid relationship of the amino- and hydroxy-groups on the side-chain of isoproterenol sulphate (4) results in... 

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