African Journal of Chemistry

African Journal of Chemistry ISSN 4391-3199 Vol. 5 (9), pp. 357-364, September, 2018. © International Scholars Journals

Full Length Research Paper

Development and validation of LC method for the determination of leflunomide in pharmaceutical formulations using an experimental design

Srinivas Rao V2, Sunanda K. K3, Narasimha Rao M.1, Allam Appa Rao1, Maheswari IL1 and Srinubabu G1*

1Center for Biotechnology, Andhra University College of Engineering (A), Visakhapatnam- 530003, India.

2St John's Pharmacy College, Vijayanagar, Bangalore-40, India.

3Department of Microbiology, Andhra University College of Science and Technology, Visakhapatnam-530003, India.

Accepted 06 June, 2018


 A rapid and sensitive RP-HPLC method with UV detection (260 nm) for routine analysis of leflunomide in a pharmaceutical formulation was developed. Chromatography was performed with mobile phase containing a mixture of acetonitrile and phosphate buffer (60:40, v/v) with flow rate of 0.8 ml min−1. Quantitation was accomplished with internal standard method. The procedure was validated for linearity (correlation coefficient = 0.9999), accuracy, robustness and intermediate precision. Experimental design was used for validation of robustness and intermediate precision. Plackett-Burman design was used to screen the essential factors for optimization. To test robustness, three factors were considered; percentage v/v of acetonitrile in mobile phase, flow rate and pH; an increase in the flow rate results in a decrease of the drug found concentration, while the percentage of organic modifier and pH have no important effect on the response. For intermediate precision measure the variables considered were: analyst, equipment and number of days. The RSD value (0.93%, n = 24) indicated a good precision of the analytical method. The proposed method was simple; highly sensitive, precise, accurate and retention time less than 6 min indicating that the method is useful for routine quality control.

Key words: Leflunomide, HPLC, validation, Robustness testing, experimental design.