DEVELOPMENT AND VALIDATION OF AN RP-HPLC BIOANALYTICAL METHOD FOR QUANTIFICATION OF RIZATRIPTAN BENZOATE IN RAT PLASMA AND BRAIN HOMOGENATE: APPLICATION TO PHARMACOKINETIC AND BRAIN-TARGETING STUDIES

Abstract

Purpose This study was conducted to establish and validate a new, rapid, specific, sensitive and reproducible reverse-phase high performance liquid chromatographic (RP-HPLC) method for the determination of Rizatriptan Benzoate (RZT) in rat plasma and brain homogenate. The validated method was then used to assess the PK profile and brain targeting ability of intranasally administered RZT formulations. Methods The chromatographic separation was performed on a C18 analytical column with isocratic mobile phase in the ratio from the acetonitrile and phosphate buffer (pH 3.0) optimal flow rate of approximately 0.4 ml/min. Detection was conducted at the selected wavelength with a UV detector. Biological sample preparation by a protein precipitation method with acetonitrile This analytical method was validated based on selectivity, linearity, precision, accuracy, recovery data and the matrix effect across their corresponding ranges to comply with US Food and Drug Administration (FDA) and International Council for Harmonisation (ICH M10) guidelines. The approved method was used for the analysis of RZT in rat plasma/brain homogenate after nasal administration of RZT solution, solid lipid nanoparticles (SLNs) and SLN loaded locally gel. Results A good chromatographic performance of the developed RP-HPLC method was observed with a retention time of approximately 3.5 min. The calibration curve showed wonderful linearity in the evaluated focus vary (R² = 0.9994). Precision and accuracy reported were within the regulatory guideline limits. The extraction recovery was greater than 85% for all three level of quality control samples in plasma or brain homogenate. The matrix factors were between 0.992 and 0.997(preparing method reduces or removes interference related to matrix). RZT was shown to be stable under all investigated storage and processing conditions in stability studies. Implementation of a validated method showed relevant brain accumulation as well as an enhanced pharmacokinetic profile for SLN-loaded in situ gel phase compared to the conventional route used for RZT solution. Conclusion The developed RP-HPLC method is simple, reproducible, accurate and appropriate for bioanalytical routine quantification of Rizatriptan Benzoate in biological matrices. The validated methodology can be easily useful on pharmacokinetic studies and bioavailability of nasally delivered drugs in the CNS.