High Performance Liquid Chromatographic Determination of Clarithromycin in Lymphocytes Using a Post-column with Tris(2,2-bipyridine) Ruthenium (III) Chemiluminescence Detection

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The concentration of antibiotics in cells must be determined for effective treatment of infectious diseases caused by obligate intracellular parasites, such as Chlamydia trachomatis and Legionella pneumophila. We confirmed the usefulness of high performance liquid chromatography (HPLC), which is already commonly used in medical facilities, for the measurement of antibiotics in cells. The measurement was carried out using a post-column with tris(2,2-bipyridine) ruthenium (III) chemiluminescence detection. Clarithromycin (CAM) was used as the model antibiotic. The retention time of CAM on the column was 7.6±0.4 min and the detection limit was 2.0×10-3 ng on column. The linearity of the calibration curve was guaranteed until 2.0 ng on column. This level of performance is comparable to that of liquid chromatography-tandem mass spectrometry. The system was able to monitor changes in the concentration of CAM in lymphocytes. These findings suggest that HPLC, a general-purpose, already widely used detection system, could also contribute to the development of effective individual antibiotic treatment regimens at a wide variety of medical facilities.