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Abstract: Fexofenadine, an antihistamine drug used in allergic rhinitis treatment, can be produced by oxidative biotransformation of terfenadine by Streptomyces platensis, which involves three consecutive oxidation reactions. We report here the purification and identification of the enzyme responsible for the first step, a cytochrome P450 (P450)-dependent monooxygenase. The corresponding P450, designated P450 terf , was found to catalyze the hydroxylation of the t-butyl group of terfenadine and exhibited UV–Vis characteristics of a P450. Its interaction with terfenadine led to a shift of its Soret peak from 418 to 390nm, as expected for the formation of a P450–substrate complex. In combination with spinach ferredoxin:NADP(+) oxidoreductase and ferredoxin, and in the presence of NADPH, it catalyzed the hydroxylation of terfenadine and some of its analogues, such as terfenadone and ebastine, with km values at the μM level, and kcat values around 30min−1. Sequencing of the p450 terf gene led to a 1206bp sequence, encoding for a 402 aminoacid polypeptide exhibiting 56–65% identity with the P450s from the 107L family. These results confirmed that P450s from Streptomyces species are interesting tools for the biotechnological production of secondary metabolites, such as antibiotics or antitumor compounds, and in the oxidative biotransformation of xenobiotics, such as drugs. [Copyright &y& Elsevier]

Abstract: The actinomycete Streptomyces platensis grown in culture medium containing soybean peptones can transform terfenadine, an antihistamine drug, into its active metabolite fexofenadine. The microbial oxidation of methyl group of terfenadine into carboxylic acid could be an alternative to chemical ways to produce fexofenadine. This bioconversion requires three oxidation steps: a hydroxylation of one methyl group followed by the oxidation of the corresponding alcohol into the aldehyde and finally its oxidation into the carboxylic acid. The oxidation reaction of each step has been studied. Terfenadine and intermediates incubated with whole cells were not oxidized under argon whereas their biotransformation under 18O2-enriched atmosphere gave labeled fexofenadine. P450 inhibitors, such as clotrimazole or fluconazole, inhibited oxidation activity of each step. While the two last steps could be catalyzed by dehydrogenases or oxidases, this study strongly demonstrates the role of at least one, or possibly several cytochromes P450, in the oxidation of terfenadine into fexofenadine by S. platensis cells. To our knowledge, this is one of the few examples of involvement of P450s in such three steps oxidation of a xenobiotic. [ABSTRACT FROM AUTHOR]

The biotransformation of terfenadine into a primary alcohol, hydroxyterfenadine, followed by its oxidation to an acid, fexofenadine, was investigated using Streptomyces platensis cells. Time-courses of metabolite formation were established, and the results underlined the modulation of the alcohol to acid formation ratio according to culture conditions. Optimization of the hydroxylation step (pH, temperature, culture medium composition) led to the preparation of hydroxyterfenadine with a good yield (51%) using cells grown in culture medium without soybean peptone. In contrast, when incubations were performed with cells cultured in a medium containing soybean peptone, the alcohol to acid formation ratio decreased. The efficiency of the conversion to fexofenadine was shown to depend on the age of the cells, thus suggesting the induction of an oxidative activity. Both the hydroxylation reaction and the following two-oxidation steps leading to the acid seemed to depend on oxygen. [ABSTRACT FROM AUTHOR]