Determination of Zoledronic Acid and Its Related Substances by High Performance Liquid Chromatography with Evaporative Light Scattering Detection.

A method has been developed and validated for analysis of zoledronic acid (ZOL) and its related substances by ion-pair reversed-phase high performance liquid chromatography with evaporative light scattering detection (ELSD). Chromatographic separation was achieved with gradient elution by using a C18 column, mobile phase containing 12 mM ammonium acetate buffer and 35 mM n-pentylamine, whose pH value is 7.0, and 5% acetonitrile. The mobile-phase flow rate was 1.0 mL/min. The calibration plot was linear in the range from 0.4 mg/mL to 6.0 mg/mL for ZOL and from 6.25 μg/mL to 100 μg/mL for its related substances. ZOL and its related substances, namely imidazole-1-yl-acetic acid, phosphate, phosphite and degradation products did not interfere with each other. The method was rapid, linear, accurate and reproducible. The high performance liquid chromatographic method that has been developed to determine the related substances and assay of ZOL can be used simultaneously to evaluate the quality of regular samples. It can be also used to test the stability samples of ZOL.