UHPLC-MS/MS method for the determination of azithromycin in human plasma of pediatric patients

The presence of azithromycin in the human plasma of pediatric patients was determined with a UHPLC-MS/MS assay. Sample preparation was done by protein precipitation, and the separation was achieved on a C18 column by the gradient mixture of the mobile phase A (0.1% acetic acid and 3 mM ammonium acetate in water) and the mobile phase B (0.1% acetic acid and 3 mM ammonium acetate in the solution of acetonitrile, methanol, and water, 47.5/47.5/5, V/V/V). The multiple reaction monitoring mode was adopted to monitor the precursor-to-product ion transitions of m/z 749.6 → m/z 591.5 for azithromycin and m/z 753.6 → m/z 595.5 for azithromycin-¹³ C-d3 (the internal standard) at the positive ionization mode. The calibration curve ranged from 0.5 to 500.0 ng·mL^-1 , and the correlation coefficient was greater than 0.99. The intra- and inter-batch precision was less than 13.7%. Accuracy determined at four concentrations ranged from 99.5 to 110.8%. The extraction recoveries were more than 95%, and the matrix effects were 98-100%. The stability under various conditions was acceptable with the accuracy deviation within 9.2%. In conclusion, our method was simple, sensitive, and reliable for quantifying azithromycin in plasma among pediatric patients.