Mechanistic explanation to the catalysis by pyrazinamide and ethambutol of reaction between rifampicin and isoniazid in anti-TB FDCs

Rifampicin and isoniazid are known to interact with each other in solid formulation environment to yield isonicotinyl hydrazone (HYD). In earlier studies, this reaction was indicated to be catalyzed by pyrazinamide and ethambutol hydrochloride, the two other co-drugs present in oral anti-tuberculosis fixed-dose combination (FDC) formulations. Accordingly, the present study was carried out to understand the catalytic role of pyrazinamide and ethambutol hydrochloride on the reaction between rifampicin and isoniazid. For the purpose, organic bases and amides similar in structure to pyrazinamide and ethambutol hydrochloride were combined individually with rifampicin and isoniazid. The compounds employed were pyrazine, piperdine, pyrollidine, pyridine, triethylamine, diisopropylethylamine, picolinamide, benzamide, ethylenediamine, ethanolamine, diethanolamine, and triethanolamine. An additional study was also carried out in the presence of free base of ethambutol. The mixtures were exposed to accelerated stability test condition of 40 °C/75% RH for 15 d. The nature of the products formed and the changes in relative concentrations of the drugs and products were followed by HPLC. The drugs showed different extent of degradation, yielding HYD, and in some cases degradation products of rifampicin. The results confirmed the catalytic role of pyrazinamide and ethambutol hydrochloride. The catalysis is postulated to involve intra-molecular proton transfer during transhydrazone formation process, entailing a tetrahedral mechanism.