Trace analysis of tetracycline antibiotics in human urine using UPLC–QToF mass spectrometry

A rapid, sensitive and specific ultra-performance liquid chromatography–quadrupole time-of-flight mass spectrometry (UPLC–QToF-MS) method has been developed and validated for simultaneous determination of three tetracyclines (TCs) including oxytetracycline, tetracycline, and chlortetracycline in human urine. Human urine sample preparation involves pH adjustment to 4 with hydrochloric acid, pre-concentration, and cleanup by solid phase extraction (SPE) process. The chromatographic separation of all TCs was achieved in less than 5 min using UPLC and ESI-QToF-MS was successfully employed for the identification and quantification. The accurate masses of the product ions were calculated using lock mass correction, and were deviated from the theoretical masses by 0.5–1.9 mDa and 1.1–4.5 ppm, respectively. The developed method was validated in terms of linearity, sensitivity, selectivity, decision limit, detection capability, accuracy, and precision. Limit of detection and limit of quantitation for all TCs were estimated in the range of 0.089–0.138 ng mL− 1 and 0.294–0.455 ng mL− 1, respectively. High overall recoveries of greater than 90% were achieved with linear responses over the 0.5–2 ng mL− 1 range for all TCs in urine samples.