Development of micellar electrokinetic chromatography method for the determination of three defined impurities in indomethacin
Abstrakt
A micellar electrokinetic chromatography method for the determination of indomethacin impurities (4-chlorobenzoic acid, 5-methoxy-2-methyl-3-indoleacetic acid, and 3,4-dichloroindomethacin) has been developed. A 64.5/56-cm fused silica 50 mu m id capillary with extended light path (150 mu m id) detection window was used.
Internal standard was 1-naphthylacetic acid. The analytes were separated at 30 kV with DAD detection at 224 nm.
A central composite face-centered design was applied for the optimization of the separation conditions. The effect of SDS concentration, content of methanol, concentration of phosphate buffer, and pH of the buffer were studied at three levels.
The optimized background electrolyte was 20 mmol/L phosphate buffer (pH 7.57) containing 58 mmol/L SDS and 0% MeOH. Sufficient resolution of all compounds with Rs >= 3.5 was achieved within 10 min.
The method was validated for a range of 1.25-80 mu g/mL of each impurity corresponding to 0.05-3.2% relative to the concentration of indomethacin (2.5 mg/mL). The calibration curves were rectilinear with correlation coefficients r(2) exceeding 0.9994.
The lower limit of quantification was 0.05% or 1.25 mu g/mL that complies with the reporting limits regarding the ICH Q3A guideline. The method was applied to purity assay of indomethacin in both bulk drug and gel.