Rapid determination of tetracycline antibiotics in serum by reversed-phase high-performance liquid chromatography with fluorescence detection.

A rapid and accurate determination of tetracycline antibiotics in human serum by reversed-phase high-performance liquid chromatography with fluorescence detection has been developed, based on protein precipitation in serum. Various reagents for precipitation were investigated, and 24% trichloroacetic acid in methanolic solution gave the maximum recovery (at least 94.3%) and interference-free chromatograms of different three tetracyclines. At a concentration of 0.5 micrograms/ml, the precision (relative standard deviation) ranged from 1.12 to 1.94%. In the range 0.04-10.0 micrograms/ml for oxytetracycline and chlorotetracycline and 0.01-10.0 micrograms/ml for tetracycline, linear responses were observed. The detection limits of this method were 10-35 ng/ml for all three antibiotics. The proposed method was applied to the determination of serum concentrations in subjects receiving tetracycline antibiotics.