Antimicrobial activity of patulin derivatives: a preliminary report.

Preliminary results are reported of antimicrobial assays conducted with chemically synthesized derivatives of patulin, a mycotoxin produced by several species of Penicillia and Aspergilli. Patulin (Fig. 1) has been shown to be an antibiotic1) and a carcinogen2); it is considered too toxic for humans or animals3) and is inhibitory to plants4). It is an a,R-unsaturated r-lactone with two conjugated double bonds and a very reactive hemiacetal group that racemizes rapidly in aqueous media, precluding isolation of the (+) and (-) optical isomers. TANABE and SUZUKI5) reduced the hemiacetal group of patulin to form ascladiol, a compound reported to be 25 % as toxic as the parent compound although it has the same unsaturated bonds and 1-lactone structure. No details of their toxicity tests were given. LIEU and BULLERMAN6) observed decreased activity against Bacillus subtilis, brine shrimp larvae, and chick egg embryos when patulin reacted with sulfhydryl (-SH) groups of cysteine or glutathione. Although postulated to be the result of MICHAEL addition of sulfhydryl to the double bonds of patulin, such groups can also react with the hemiacetal moiety, and this may explain the decrease in toxicity of the reaction products. Some beneficial uses of patulin are mentioned in the report by SINGH4) of the work Of OOSTERHUIS, who used patulin for curing clinical dermatomycoses, and in a patent issued to VERMEUIEN and LE DRUTT7), who prepared patulin thiosemicarbazone and found it to be a bacteriolytic agent against KOCH bacilli without the expected bacteriostatic activity of patulin against Gram-positive and Gram-negative bacilli. Because of the solubility of patulin thiosemicarbazone in lipids, the derivative readily resorbed via the lymphatic passages and gave favorable results in the treatment of pulmonary and several other forms of tuberculosis. The lack of information on the biological activity of patulin derivatives, as well as the work cited above, led us to prepare thirteen compounds for evaluation of their toxicity and teratogenicity relative to patulin. The following derivatives (Fig. 1), prepared by established procedures, were characterized on the basis of their physical constants and spectral properties: lactone 28), acetate 3 and benzoate 49), oximes 5, 6, 7 and 810), semicarbazone 9 and thiosemicarbazone 107) hydrazones 11 and 120), dimethone anhydride 139), and octahydroxanthene 1411). Preparation of derivatives utilized the free hydroxyl group of patulin to give a lactone (oxidation of hydroxyl) or esters (acylation of hydroxyl). In solution, the hemiacetal reacted as an aldehyde with appropriate reactants to give the other derivatives reported here. As a preliminary step, a series of disc assays was run and the results are reported here. These assays may aid in the discovery of new products of pharmacological interest; subsequent toxicity tests to be conducted later will utilize egg embryos and small animals to help determine the role in living systems of the reactive moieties of patulin, such as the unsaturated bonds and hemiacetal group. The derivatives were tested at a concentration of 100 ug/paper disc for activity against six microorganisms. This concentration assumes an average take-up of 0.15 ml of test solution per disc, from a solution containing 2 mg test compound in 3 ml of solvent. All compounds were dissolved in methanol (non-inhibitory solvent) except patulin and the free oximes, which were soluble in water. Seeded agar plates were prepared as described by PRIDHAM et al.12). The microorganisms used were: Bacillus subtilis NRRL B-765, Micrococcus luteus NRRL B-1018, Escherichia coli NRRL B-766, Saccharomvices cerevisiae NRRL Y-139, Candida albicans NRRL Y-477, and Mucor ramannianus NRRL 1839, representing, respectively, two Gram-positive bacteria, one Gram-negative bacterium, two yeasts and a mold. Thus, patulin derivatives were tested for their ability to inhibit spore germi* The mention of firm names or trade products