A Study of the Para-Aminobenzoic Acid Requirement of Clostridium Acetobutylicum.Application to Assay Procedure

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Abstract
Conditions were described by which turbidity and acid production of C. acetobutylicum can be correlated with the PAB content of the medium. Quantities of PAB as low as 0.0004 [gamma] per ml. may be detected by this method. Growth of the organism in aerobic tubes was secured by use of certain reducing agents and agar, but this method was not as satisfactory as the use of anaerobic jars for estimating accurately the amts. of growth in relation to added PAB. Ethanol- and pyridine-extractable factors found in agar supported growth of the test organism in the absence of biotin and PAB. Detailed studies have been made on both specific and non-specific factors affecting growth and metabolism of the test organisms. The activity of 15 compounds structurally related to PAB was studied. Several of these compounds, which presumably may be converted to PAB, function as growth factors for this organism. A change in position of the COOH or NH2 group from the para to the ortho or meta position in the benzene ring resulted in great loss in activity. All related compounds exhibiting high activity had their substituents arranged in the para position. Methyl and ethyl-p-aminobenzoate, procaine hydro-chloride, p-njtrobenzoic acid, p-aminobenzoylglycine, p-nitrobenzoylglycine, and p-chloroacetylbenzoylglycine, in which the substituents occupy the 1:4 position, were found to be from 10 to 100% as active as PAB. Adenine sulfate, guanine hydrochloride, xanthine and uracil, in the absence of PAB, supported growth in cones, of from 10 to 100 [gamma] per ml. 5 [gamma] per ml of each of the 3 purines together stimulated growth in the presence of small amts. of PAB. It is uncertain whether this effect is due to the purines per se. Tryptophane did not support growth of the test organism through conversion to anthranilic acid. An assay procedure based on these studies gave reasonably accurate recovery values on peptone with known amts. of added PAB. A sulfonamide-resistant Staphylococcus aureus produced from 10 to 100 times the amt. of PAB produced by its parent (non-resistant) strain. The PAB synthesis of this organism may be increased by lengthening the incubation period or by increasing its resistance to sulfonamides through continued exposure to increasing amts. of the drug. A sulfonamide-resistant pneumococcus and a sulfonamide-resistant dysentery bacillus did not show increased PAB synthesis compared with the original non-resistant strains. These results confirm those of Landy and associates with Acetobacter suboxydans as the test organism.