Article 4

DOE-BASED OPTIMIZATION STRATEGY IN MULTICOMPONENT HPLC ANALYSIS FOR DETERMINATION OF ANTI-VIRAL DRUGS IN COMBINED DOSAGE FORMS

Author (s): Prafulla Kumar Sahu


Abstract: The complexities of reverse phase high performance liquid chromatography (RP-HPLC) require a deeper understanding of input method variability in order to achieve the desired separation. During the method optimization, a statistical relationship among the input variables and the chromatographic responses is to be recognised with the help of a suitable design of experiment (DoE). The present study demonstrated the use of a full factorial design (FFD) in a RP-HPLC method development to ascertain the significant variables’ effects that impact the separation of five anti viral drugs i.e.; Lamividine (LAM), Stavudine (STA), Atazanavir (ATA), Ritonavir (RIT), and Abacavir (ABA). Influence variables including mobile phase flow rate, buffer composition (ratio of acetic acid and triethylamine) and organic modifier concentration (acetonitrile), each at two levels were investigated to achieve the best desirability criteria. Critical responses such as RSLAM-STA, RSATA-RIT, TLAM, TSTA, TATA and TRIT that may violate the validity of the method were considered as the measure of performance index of the separation process. The six responses were simultaneously optimized with the aid of a multiple response algorithm using a statistical tool (Design Expert 12.0). 50mM acetic acid and 50mM triethylamine (75:25 %v/v) buffer and acetonitrile (52: 58 v/v) at a flow rate of 0.25ml/min was found to be the optimum condition to produce the desirable responses. Validation of the developed method was furnished  and compliance to the current regulatory requirements is ascertained. The RP-HPLC method provides a rapid and well resolved analysis of the five analytes.


Keywords: RP-HPLC, Design of Experiment, optimisation, separation efficiency, validation