Improved Stability of Rifampicin in the Presence of Gastric-Resistant Isoniazid Microspheres in Acidic Media

The degradation of rifampicin (RIF) in an acidic medium to form 3-formyl rifamycin SV, a poorly absorbed compound, is accelerated in the presence of isoniazid, contributing to the poor bioavailability of rifampicin. This manuscript presents a novel approach in which isoniazid is formulated into gastric-resistant sustained-release microspheres and RIF into microporous floating sustained-release microspheres to reduce the potential for interaction between RIF and isoniazid (INH) in an acidic environment. Hydroxypropyl methylcellulose acetate succinate and Eudragit® L100 polymers were used for the manufacture of isoniazid-loaded gastric-resistant sustained-release microspheres using an o/o solvent emulsification evaporation approach. Microporous floating sustained-release microspheres for the delivery of rifampicin in the stomach were manufactured using emulsification and a diffusion/evaporation process. The design of experiments was used to evaluate the impact of input variables on predefined responses or quality attributes of the microspheres. The percent degradation of rifampicin following 12 h dissolution testing in 0.1 M HCl pH 1.2 in the presence of isoniazid gastric-resistant sustained-release microspheres was only 4.44%. These results indicate that the degradation of rifampicin in the presence of isoniazid in acidic media can be reduced by encapsulation of both active pharmaceutical ingredients to ensure release in different segments of the gastrointestinal tract, potentially improving the bioavailability of rifampicin.


Supplementary Materials: Improved Stability of Rifampicin in the Presence of Gastric-Resistant Isoniazid Microspheres in Acidic Media
Chiluba Mwila and Roderick B. Walker

Differential Scanning Calorimetry
The DSC thermograms for isoniazid (INH), 1:1 mixtures of INH with hydroxypropyl methylcellulose acetate succinate (HPMC-AS) or Eudragit ® L100 are depicted in Figures S1-S3.    slight broadening at the base of the peak, indicating a slight reduction in the crystallinity of INH. These results do not however, suggest a potential or significant incompatibility between INH and the excipient tested, However, real-time long-term stability studies would be required to ensure that this is indeed the case.

FT Raman Spectroscopy
The FT Raman spectra for INH alone and in microspheres in the presence of all excipients are depicted in Figure S4. The comparative frequencies for important functional groups of INH and the microspheres, in relation to what has been reported [2] are summarised in Table S1.  The FT Raman spectroscopic study of the microspheres confirmed the presence of all relevant functional groups for INH and the absence of potential detrimental interactions between INH when incorporated into a microsphere formulation in combination with HPMC-AS and Eudragit ® L100. However, real-time long-term stability studies would be required to ensure that this is indeed the case.

Differential Scanning Calorimetry
The DSC thermograms for rifampicin (RIF) is depicted in Figure S5. The DSC thermogram for RIF was characterized by a melting endotherm between 183.32-203.39 °C, immediately followed by an exothermic recrystallisation event that is a characteristic of a solidliquid-solid transition, and finally a decomposition event at 240.78-275.58 °C. These results were similar to those in which the form II polymorph of RIF exhibited a melting endotherm at between 180-197 °C that was immediately followed by recrystallisation to the form I polymorph with an exothermic peak at between 197-223 °C and a final decomposition event that was observed between 247-266 °C [3].
The DSC thermogram for a 1:1 mixture of RIF and d-glucose is depicted in Figure S6. The thermogram for the 1:1 mixture of RIF and d-glucose revealed a melting endotherm for RIF between 210.04-231.77 °C, no recrystallisation exothermic event and decomposition between 231.77-279.94°C. This thermal behaviour suggested a more rapid transition from polymorphic form II to form I, in the presence of d-glucose, however, this phenomenon has no biopharmaceutical significance but can be attributed to a decrease in melting point of RIF in the presence of d-glucose.
The thermogram for the 1:1 mixture of RIF and ethyl cellulose (EC) revealed a melting endotherm for RIF between 184.02-202.66 °C, immediately followed by a recrystallisation event at 202.66-239.69 °C and decomposition between 239.69-271.92 °C and is depicted in Figure S7.   Figure S8. Figure S8. Typical DSC thermogram for a 1:1 mixture of rifampicin (RIF) and Eudragit ® RLPO generated at a heating rate of 10 °C/min over the temperature range 30-400 °C.
The thermogram for the 1:1 mixture of RIF and sodium bicarbonate is depicted in Figure S9 and revealed overlapping endothermic peaks at 137.82-194.66 °C which were identified as melting endotherms for sodium bicarbonate and polymorph form II of RIF. The overlapping endothermic peaks were immediately followed by a period of recrystallisation at between 194.66-240.59 °C and decomposition at 240.59-273.46 °C. The thermogram for sodium bicarbonate alone has been reported to depict two endothermic events, one at 96 °C and one at 150 °C that is evidence of thermal decomposition of NaHCO3 generating H2O and CO2 and sodium carbonate and commences at approximately 50 °C and ends at 170 °C. The thermogram for the 1:1 mixture of RIF and anhydrous citric acid is depicted in Figure S10   The overlapping peaks in the thermogram for the 1:1 mixture of RIF and sodium bicarbonate, in addition to the interference with the melting endotherm for RIF from possible degradation products of anhydrous citric acid in the thermogram of the 1:1 mixture of RIF and citric acid makes the analysis cumbersome and suggests that other analytical techniques would be more useful in determining the compatibility of RIF with these excipients and that long term stability of the manufactured product containing RIF, sodium bicarbonate and citric acid should be assessed. These results suggest that the intended excipients were compatible with RIF and may be used in the same formulation composition, however, real-time long-term stability studies would be required to confirm this is indeed, the case.

FTIR Spectroscopy
The FTIR spectra of 1:1 mixtures of RIF and EC, d-glucose, Eudragit ® RLPO, sodium bicarbonate and citric acid are depicted in Figures S11-S15.     The infrared absorption spectra for the 1:1 mixtures do not reveal any significant shifts in the frequency of resonation for RIF and the minor shifts in the FTIR frequencies of resonance observed for the functional groups of RIF (Table S2) were attributed to potential weak hydrogen bonding, further confirming the absence of potential interactions between RIF and the excipients to be used for the manufacture of the floating microspheres. However, real-time long-term stability studies would be necessary to ensure this is, indeed the case.