Abstract

BIOANALYTICAL METHOD DEVELOPMENT AND VALIDATION OF SOLIFENACIN SUCCINATE AND SILODOSIN USING LC-MS/MS WITH ALFUZOSIN AS INTERNAL STANDARD

N. Ravindra Babu, K. V. Padmavathi, M. Sridhar, Mannam Subbarao*

ABSTRACT

The present study aimed to develop and validate a rapid, reliable, and reproducible bioanalytical method for the simultaneous determination of Solifenacin Succinate and Silodosin in rat plasma. The method employed Liquid Chromatography–Tandem Mass Spectrometry (LC-MS/MS) using Alfuzosin as the internal standard (IS).Chromatographic separation was achieved on a Luna Phenyl Hexyl column (250 × 4.6 mm, 5 μm) with an isocratic mobile phase of 0.1% formic acid and acetonitrile (50:50, %v/v) at a flow rate of 1.0 mL/min. The injection volume was 10 μL, and the total run time was 10 minutes. Detection was performed on a QTRAP 5500 triple quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source operating in positive ion mode. The monitored mass transitions were m/z 481.1235 → 119.9866 for Solifenacin Succinate, m/z 496.1023 → 120.0313 for Silodosin, and m/z 390.0947 → 112.2369 for Alfuzosin (IS). The calibration curves showed excellent linearity over the range of 2.5–100.0 ng/mL for Solifenacin Succinate and 4.0–160.0 ng/mL for Silodosin. Recovery studies at multiple quality control levels demonstrated consistent recoveries of 94.10–98.12% for Solifenacin Succinate and 95.43–98.31% for Silodosin. Matrix effect evaluation confirmed that ion suppression/enhancement was within acceptable limits. Precision and accuracy values met USFDA bioanalytical validation guidelines, with %CV below 15% (20% for LLOQ). Stability experiments, including freeze–thaw, bench-top, autosampler, and long-term stability, indicated that both analytes remained stable under all tested conditions. The validated LC-MS/MS method fulfilled all essential bioanalytical validation parameters, including selectivity, linearity, accuracy, precision, recovery, matrix effect, and stability. This method is suitable for pharmacokinetic studies of Solifenacin Succinate and Silodosin in rats and can be applied to future preclinical and clinical investigations.

Keywords: Solifenacin Succinate, Silodosin, Alfuzosin, LC-MS/MS, Rat Plasma, USFDA Guidelines.