Gladiolic Acid, an Antibiotic Substance Produced by Penicillium gladioli McCull. & Thom

SUMMARY: Gladiolic acid is produced when Penicillium gladioli is grown on a wide range of culture media. The main factor influencing its production and accumulation is the pH drift of the medium, high yields being associated with a characteristic pH drift, consisting of an initial fall to about pH 4·0 followed by a steady but not too rapid rise. Continued low pH is unfavourable; too rapid a rise of pH is also unfavourable since gladiolic acid tends to disappear from the medium when pH 6·0 is reached, the disappearance being very rapid at pH 7·0 or above. The effects of variation of the initial pH of the medium, of glucose concentration, of variation of nitrogen source and of additions of certain organic acids are all explicable in terms of their effect on pH drift.

The antibiotic is best extracted from culture filtrates by treatment with activated charcoal after adjustment to pH 4·0, elution of the charcoal with ether, and re-crystallization from water after evaporation of the ether. Yields of the order of 300 mg./l. are obtained.

Gladiolic acid is highly fungistatic if tested at low pH; the toxic effect is due to the undissociated molecules only and at pH 7·0, when dissociation is virtually complete, gladiolic acid is almost inactive. At pH 3·5, the least concentration inhibiting germination of fungus spores varies from 0·9 µg./ml. for Fusarium graminearum to 250 µg./ml. for Trichoderma viride. It is not highly antibacterial in broth, many organisms growing freely in the presence of 500 µg./ml. This low activity is thought to be due in part to the dissociation of gladiolic acid at pH 7·0 and in part to inactivation by certain broth constituents. This view is supported by the observation that bacterial cells suspended in gladiolic acid solutions (100 µg./ml.) in buffer at pH 4·0 are rapidly killed. This bactericidal effect occurs with both Gram-positive and Gram-negative organisms.

Gladiolic acid in solution is relatively stable in the range pH 3·0–8·0. In the presence of ammonium salts or certain amino-acids, notably p-aminobenzoic acid, it is rapidly inactivated, coloured complexes being formed. The reaction with ammonium salts is dependent on pH, not proceeding at pH 3·5 but proceeding rapidly at pH 7·0. The rapid disappearance of gladiolic acid in culture when the pH rises above 6·0 is possibly associated with this type of reaction.