Determination of roxithromycin and its active metabolite in human urine by liquid chromatography-electro-chemical detection and liquid chromatography-mass spectrometry

OBJECTIVE: To establish a sensitive method for the determination of roxithromycin (E-Rox) and its active metabolite, (9Z)-epimeric isomer (Z-Rox). METHOD: E-Rox and Z-Rox were determined by using HPLC-ECD in combination with HPLC-MS. Quantitative analyze of E-Rox was performed with HPLC-ECD after liquid-liquid extraction. Afterwards, the isomeric ratios of Z-Rox and E-Rox were determined by using HPLC-MS in the selected reaction monitoring (SRM) mode via electrospray ionization (ESI). The SRM transition m/z 837 to m/z 679 was monitored for detection of Z-Rox and E-Rox isomers. RESULT: The calibration curves were over the concentration range of 0.1 to approximately 10 mg·L-1 for E-Rox and 0.05 to approximately 5 mg·L-1 for Z-Rox. The limits of quantification were found to be 0.1 mg·L-1 for E-Rox and 0.05 mg·L-1 for Z-Rox. The intra-and inter-day RSD were within 12.5% form each of analytes. The combined method was applied to the human urine samples from 4 healthy volunteers after oral administration of 150 mg E-Rox. CONCLUSION: The combined method proved to be suitable for studies on the metabolism of E-Rox in human. With the method, the concentrations of the active metabolite Z-Rox in human urine were for the first time investigated.