Comparative Evaluation of HPLC and Capillary Electrophoresis in Intestinal Permeability of Oseltamivir Phosphate via Single-pass IntestinalPerfusion (SPIP) Method

Introduction: Oseltamivir phosphate (OP), a widely used antiviral prodrug for influenza, requires accurate Biopharmaceutics Classification System (BCS) classification to inform formulation and bioequivalence decisions. Despite its high solubility, the permeability characteristics of OP remain inadequately defined. Methods: This study evaluated the intestinal permeability of OP using the in situ Single-Pass Intestinal Perfusion (SPIP) model in male Sprague Dawley rats. Two validated analytical methods - High- Performance Liquid Chromatography (HPLC) and Capillary Zone Electrophoresis (CZE) were developed and compared. Metoprolol tartrate served as the high-permeability reference standard, and phenol red was used to monitor water flux. Permeability coefficients (Peff) were calculated after validation of analytical methods per ICH guidelines. Results: The HPLC and CZE methods demonstrated excellent linearity (R² ≥ 0.999), precision (RSD < 0.9%), and accuracy (99.3–101.7%). The Peff of OP was significantly lower than that of metoprolol, with the values of 0.14 ± 0.06 × 10⁻⁴ cm/s (HPLC) compared to 0.55 ± 0.08 × 10⁻⁴ cm/s, and 0.30 ± 0.18 × 10⁻⁴ cm/s (CZE) compared to 0.41 ± 0.11 × 10⁻⁴ cm/s, respectively. No statistical difference was found between the two methods (p > 0.05). HPLC provided superior sensitivity, while CZE offered operational and environmental advantages. Discussion: OP showed low intestinal permeability, supporting its BCS Class III classification and aligning with values for other low-permeability drugs. Both HPLC and CZE yielded consistent results, with HPLC providing greater sensitivity while CZE offered greener and cost-effective advantages. These findings confirm the utility of SPIP and highlight complementary roles of the two methods in permeability profiling. Conclusion: OP exhibited low permeability in rat jejunum, consistent with BCS Class III classification. The validated HPLC and CZE methods are effective for intestinal permeability studies, with HPLC recommended for regulatory purposes and CZE for green analytical applications.