Development and Validation of Stability Indicating RP-HPLC Method for Simultaneous Determination of Doxycycline and Rifampicin in Polymeric Nanoparticles

Abstract

Background: The combination of doxycycline (DOXY) and rifampicin (RIF) is recom-mended as a treatment therapy for brucellosis by the World Health Organization. Objective: The aim of the current study was to develop and validate the stability-indicating reverse-phase high-performance liquid chromatography (RP-HPLC) method for the analysis of a combina-tion of doxycycline & rifampicin. Methods: The RP-HPLC method was developed and validated to estimate the doxycycline and ri-fampicin combination as per ICH guidelines. The drug combination solution was exposed to differ-ent stress conditions: acidic, basic, photo-oxidation, and oxidation. Results: The method was found linear in the range of 2 -10μg/mL for both the drugs with a reten-tion time of 3.5 min for doxycycline and 6.5 min for rifampicin at lambda maximum of 350 nm. The RP-HPLC method was precise and accurate with %RSD < 2%. The intra-day and inter-day precisions were calculated and found within the acceptable range of 5%. Both drugs demonstrated good stability in the mobile phase after 6 hours. The LOD and LOQ of doxycycline and rifampicin were 100 ng/mL & 200ng/mL and 150ng/mL & 500ng/mL, respectively. The forced degradation of the combination of drug solutions was performed. The degraded drug peaks were well-resolved from the peaks of drugs. The percentage encapsulation efficiency of doxycycline and rifampicin in the nanoparticle system was assessed by utilizing the validated RP-HPLC method and found >60% (DOXY) and >70% (RIF). Conclusion: The developed RP-HPLC method of DOXY-RIF combination was rapid, accurate, precise, and stability-indicating. The method can be appropriately applied todetecte drugs in the na-noparticulate system.