Development and Validation of Ultrasound Assisted and Dispersive Liquid-Liquid Microextractions Combined with HPLC-UV Method for Determination of Bosentan in Human Plasma and Urine

Abstract

A feasible, rapid, sensitive and reliable dispersive liquid-liquid microextraction (DLLME) and ultrasonic assisted liquid-liquid microextraction (USAEME) combined with high-performance liquid chromatography (HPLC) method with UV detection was developed and validated for analysis of bosentan in human plasma and urine samples. All validation tests were reported according to FDA guidelines. Since chromatographic methods are precise, accurate and provide excellent separation are increasingly being used in determination of trace amount of drugs in biological goals. So the optimized experimental extraction factors from our previous work were used to evaluate HPLC-UV method for decreasing the limit of quantifications (LOQs) to cover the therapeutic range. Simple centrifugation of urine samples was sufficient prior to microextraction procedure, while acetonitrile protein precipitation provides sample clean up for plasma samples. The suggested method was linear (correlation coefficient R-2 > 0.998) over a range of 0.1-10 mu g mL(-1) which covers the therapeutic range. Proposed methods are practical for quantitative analysis of bosentan in real samples, and could be used in routine clinical analyses.