Overestimation of rifampicin during colorimetric analysis of anti-tuberculosis products containing isoniazid due to formation of isonicotinyl hydrazone.

When present together in fixed-dose combinations (FDC) of anti-tuberculosis drugs, rifampicin (R) and isoniazid (H) interact with each other to form isonicotinyl hydrazone (HYD). In a preliminary study, this product was found to possess similar colorimetric spectrum to that of rifampicin. Therefore, an investigation was undertaken to determine interference of HYD during analysis of rifampicin in FDC products by colorimetry. For the purpose, standard plots were constructed for rifampicin and HYD at 475 nm, the wavelength maximum for both the compounds. The plots were linear in the range of 10-100 microg/ml. Molar absorptivity values for rifampicin and HYD were 15279 and 5034, respectively. It indicated that HYD possessed one-third absorptivity to that of rifampicin. The analysis of combinations of rifampicin and HYD revealed that rifampicin could be overestimated to a maximum extent of 33%, while interference varied at other relative ratios of the two compounds. This was also confirmed by colorimetric and HPLC analysis of a degraded marketed product and samples from a dissolution study. Thus this investigation suggests that any method devoid of interference of HYD should be preferred for analysis of rifampicin, whenever it is present along with isoniazid.