Analytical Method Validation of Vilazodone with Spectrofluorimetric Detection in Rabbit Plasma

Abstract

Vilazodone (VLZ) is an antidepressant agent approved in 2011 by Food and Drug Administration (FDA) for the treatment of major depressive syndromes. Its chemical structure derives from trazodone which is used as an off-label drug for dogs and cats in order to induce sedation and reduce anxiety, behavioural and pre-and post-operative stress. VLZ could be a potentially useful compound in reducing severe anxiety responses in pet animals. The aim of the present research was to develop and validate a method to quantify VLZ in rabbit plasma. A 500 μL aliquot of rabbit control plasma was added to 50 µL of IS (0.1 ug/mL) and 100 µL of NaOH 0.1M. Then 1 mL of Et₂O:EtOAc (70:30 v/v) was added and the sample, shaken and centrifuged. The organic layer was transferred into a clean polypropylene vial. This procedure was repeated twice. The combined supernatants were evaporated under nitrogen at 40°C and reconstituted with 100 µL of mobile phase. 50 µL of this latter solution was injected onto HPLC-FL. The mobile phase consisted of Na₂HPO₄ (0.02M, pH 5):ACN (67:33 v/v) at a flow rate of 1 mL/min. The analytical column (C18) was maintained at 40°C. Excitation and emission wavelengths were set at 353 and 486 nm, respectively. Times of retention of VLZ and IS were 3.26±0.09 and 4.70±0.08 minutes, respectively. The recovery of VLZ was about 83%. Limits of quantification and detection were 0.005 µg/mL and 0.001 µg/mL, respectively. This method was verified by determining VLZ concentration in rabbit plasma after a 1 mg/kg single IV administration. The analysis of samples allowed the calculation of the main pharmacokinetic parameters. In conclusion, the present research was able to quantify VLZ in rabbit plasma after VLZ administration. This method might have application for pharmacokinetic or toxicological studies.