Simultaneous determination of chondroitin sulfate sodium,allantoin and pyridoxine hydrochloride in pharmaceutical eye drops by an ion-pair high-performance liquid chromatography

An ion-pair high-performance liquid chromatography (HPLC) method has been developed for the simultaneous determination of chondroitin sulfate sodium (CSS), allantoin and pyridoxine hydrochloride (VB₆) in a commercial eye drops dosage form. An Alltima C₁₈ column (250 mm × 4.6 mm i.d., 5 μm) was used for the separation at room temperature, with 25 mM ammonium dihydrogen phosphate (containing 0.01% heptanesulfonic acid sodium salt) and acetonitrile (95:5, v/v) as the mobile phase at the flow rate of 0.5 mL min⁻¹. The detection wavelength for CSS, allantoin and VB₆ was 195 nm, 215 nm and 291 nm, respectively. The method showed good linearity for CSS, allantoin and VB₆, with correlation coefficients greater than 0.9996, in the range of 203.96–815.84 mg L⁻¹, 371.16–1488.64 mg L⁻¹, and 23.32–93.28 mg L⁻¹, respectively. The instrumental and method precisions were adequate with all relative standard deviations lower than 2.0%. The accuracy of this method, measured by the recovery of three compounds from spiked placebo solutions, was from 99.01% to 101.92%. The three components, CSS, allantoin and VB₆ were well separated from other ingredients and degradation products. This method is fast, simple, and can be used for direct and simultaneous determination of CSS, allantoin and VB₆ in the pharmaceutical preparation.