A validated RP-HPLC method for quantitative determination of related impurities of ursodeoxycholic acid (API) by refractive index detection.

ABSTRACT

An isocratic RP-HPLC method was developed and validated for quantitative determination of ursodeoxycholic acid (UDCA) and its related impurities. Considering the lower molecular absorptivity of UDCA, refractive index detector was used to detect the impurities on a Phenomenex Luna C(18), 150 mm × 4.6 mm, 5 µm column. The mobile phase was 0.1% acetic acid/methanol (3070, v/v) and flow rate was 0.8 ml/min. The detector and column temperature was maintained at 40°C. The method is linear over a range of 0.25-3.5 µg/ml for all impurities and coefficient of correlation (r(2)) was ≥0.9945. The accuracy of method demonstrated at three levels in the range of 50-150% of the specification limit and recoveries were found to be in the range of 97.11-100.75%. The precision for all related impurities was below 3.5% R.S.D. The method was applied to commercial bulk drug sample for assay purpose.