Fungal transformations of antihistamines : metabolism of cyproheptadine hydrochloride by Cunninghamella elegans

Abstract

1. Metabolites formed during incubation of the antihistamine cyproheptadine hydrochloride with the zygomycete fungus Cunninghamella elegans in liquid culture were determined. The metabolites were isolated by hplc and identified by mass spectrometric and proton nmr spectroscopic analysis. Two C. elegans strains, ATCC 9245 and ATCC 36112, were screened and both produced essentially identical metabolites. 2. Within 72 h cyproheptadine was extensively biotransformed to at least eight oxidative phase-1 metabolites primarily via aromatic hydroxylation metabolic pathways. Cyproheptadine was biotransformed predominantly to 2-hydroxycyprohep tadine. Other metabolites identified were 1- and 3-hydroxycyprohep tadine, cyproheptadine 10,11epoxide, N-desmethylcyproh eptadine, N-desmethyl- 2-hydroxycyprohep tadine, cyproheptadine N-oxide, and 2-hydroxycyprohep tadine N-oxide. Although a minor fungal metabolite, cyproheptadine 10,11- epoxide represents the first stable epoxide isolated from the microbial biotransformation of drugs. 3. The enzymatic mechanism for the formation of the major fungal metabolite, 2hydroxycyprohep tadine, was investigated. The oxygen atom was derived from molecular oxygen as determined from ¹⁸O-labelling experiments. The formation of 2-hydroxycyproheptadine was inhibited 35, 70 and 97% by cytochrome P450 inhibitors metyrapone, proadifen and 1-aminobenzotriazole respectively. Cytochrome P450 was detected in the microsomal fractions of C. elegans. In addition, 2-hydroxylase activity was found in cellfree extracts of C. elegans. This activity was inhibited by proadifen and CO, and was inducible by naphthalene. These results are consistent with the fungal epoxidation and hydroxylation reactions being catalysed by cytochrome P450 monooxygenases. 4. The effects of types of media on the biotransformation of cyproheptadine were investigated. It appears that the glucose level significantly affects the biotransformation rates of cyproheptadine; however, it did not change the relative ratios between metabolites produced.