Optimization and Validation of a High‐Performance Liquid Chromatography (HPLC‐UV) Method for Quantification of α‐Lipoic Acid in Cyclodextrins Complex for Ocular Delivery

Alpha-lipoic acid (ALA) is a natural antioxidant with therapeutic potential, but no method currently quantifies ALA in ocular structures. This study utilizes beta-cyclodextrins (β-CD) and (2-hydroxypropyl)-β-cyclodextrins (HP-β-CD) to enhance ALA solubility and stability in 1:1 M ratio. This study aims to optimize and validate a method for quantifying ALA in CD complexes using high-performance liquid chromatography with ultraviolet detection (HPLC-UV) and to perform a diffusion study to evaluate ALA's permeability in ocular tissue. The HPLC method employed a C18 column (150 × 4.6 mm, 5 μm particle size) with an isocratic mobile phase of acetonitrile and water (50:50, v/v) at pH 4.0, a flow rate of 1 mL/min, detected ALA after 4.8 min at a wavelength of 334 nm. Validation of the method was conducted according to the ICH guideline M10 on bioanalytical method validation. The stability study revealed that better stability was obtained for ALA in β-CD inclusion in comparison with HP-β-CD. The ex vivo study using Franz diffusion cells was performed to confirm that ALA was able to permeate through the cornea and sclera via ocular administration. This accurate and precise HPLC-UV method paves the way for future research into ALA's potential to alleviate ocular conditions. SUMMARY: This study optimized and validated a precise and accurate HPLC-UV method for quantifying ALA in CD complexes, following ICH guideline M10 on bioanalytical method validation. Stability studies revealed that β-CD complexes provided better thermal and solubility stability for ALA compared to HP-β-CD complexes. The ex vivo diffusion study confirmed that ALA successfully permeated the cornea and sclera via ocular administration. The validated method lays the groundwork for future research on ALA's potential to alleviate complications associated with ocular diseases.