Development Of Hplc Method For Stress Testing Of Combination Of Two Drugs Using Design Of Experiment Concept

ABSTRACT

This study was conducted to develop, an High Performance Liquid Chromatography using photodiode array detector (HPLC-PDA) method to analyse the samples generated by the stress testing of antifilarial combination (albendazole and diethylcarbamazine citrate) in the solution state. The concept of Quality by Design (Design of Experiment, DoE) approach was used for the development. For the separation of the drugs and its degradation products (DPs), DoE was applied in two stages, i.e., primary parameter stage where factors having major effect were selected. This stage gives us CQA (Critical Quality Attribute) which along with minor factors affecting were varied to get the secondary design. For each of the stage a different design was selected; for primary stage IV optimal design (Response Surface Method) was selected whereas for secondary stage, Taguchi orthogonal array design was selected. The major primary parameters affecting the HPLC method as screened by preliminary studies were the buffer pH, organic modifier (methanol or acetonitrile), initial hold time (start of gradient) and gradient time. The primary stage was completed successfully. The results were compiled in form of resolution of peak from next peak and analysed by DoE. The process fixed the values for buffer pH (4.38), organic modifier (acetonitrile) and gradient time (30 min). The CQA from primary run was initial hold time. This parameter along with other parameters initial and final concentration of organic modifier, buffer type (phosphate or acetate), buffer strength (mM) and oven temperature were further varied and samples withdrawn were analysed. The data of secondary design was compiled in the form of resolution (R), analysed by Design Expert and final value for secondary parameter for HPLC method were fixed. The resolution of the peaks for some secondary runs was sufficient reflecting some type of interaction between the drugs and/or degradation products.