High-performance liquid chromatography measurement of antimicrobial concentrations in polymorphonuclear leukocytes

Abstract

High-performance liquid chromatography was used to determine the penetration of 19 antimicrobial agents into human polymorphonuclear leukocytes. The ratios of the intracellular concentration to the extraceHlular concentration of ampicillin, piperacillin, cefazolin, ceftizoxime, cefpimizole, and ceftazidime were all less than 0.6. Lincomycin showed a high intracellular-to-extraceliular ratio (3.0), while clindamycin achieved a ratio of 15.5, which was the highest ratio of all of the 19 tested antibiotics. Ratios for rifampin, isoniazid, chloramphenicol, and trimethoprim were 8.2, 1.1, 9.6, and 6.1, respectively. Six quinolone-class antimicrobial agents had ratios from 2.2 to 8.2. Flucytosine showed a penetration ratio of 4.6. Clindamycin uptake was temperature dependent and occurred best with live polymorphonuclear leukocytes; sodium fluoride, adenosine, and puromycin were inhibitory. The results obtained in this study correlate well with the results of other studies involving radioisotopic methods. This indicates that high-performance liquid chromatography is a useful method for determining the intracellular penetration of antimicrobial agents.

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

“…The injected sample flowed with the mobile phase through the separation column and to the mixer. At the mixer, the separated sample solution was mixed with bipyridine complex of ruthenium ((Ru(bpy) 3 ) 3+ ) solution, which was prepared by oxidation of (Ru(bpy) 3 ) 2+ . The reaction between (Ru(bpy) 3 ) 3+ and compounds with the alicyclic tertiary amine structure produced chemiluminescence, the intensity of which was measured and digitized.…”

“…10,11) Thus, in the present study, we examined whether LC using a tris(2,2 -bipyridine) ruthenium (III) chemiluminescence detection system would be suitable for the quantitative analysis of macrolide antibiotics in cells. The ruthenium complex (Ru(bpy) 3 ) 3+ has specific binding activity to the alicyclic tertiary amine structure found in macrolide antibiotics, and is currently used for chemiluminescence analysis. [10][11][12][13][14][15] Clarithromycin (CAM) was used in the present study as the representative macrolide antibiotic since it has been widely used for a long time in Japan.…”

“…Thermostatic oven of Comet-2000 was also set at 40 • C. Under controlled current electrolysis, the current was maintained at 100 µA. The reagent solution was dissolved to 0.25 mM of Ru(bpy) 3 Cl 2 in 10 mM sulfuric acid. The mobile phase was a mixture of acetonitrile, buffer solution (acetate or phosphate) and methanol.…”

“…Accordingly, it is important to estimate the concentration of the antibiotic in the cell rather than in the blood stream when the appropriate dosing regimens are being determined. 2,3) To this end we need to establish a system to measure the * To whom correspondence should be addressed: Showa Macrolide antibiotics are the drug group of choice to treat patients with chlamydial infections and legionella pneumonia. 4,5) Some authors have suggested analytical methods such as the bioassay method and liquid scintillation counter method to determine the concentration of antibiotics in cells.…”

Lincomycin Protects Mice from Septic Shock in .BETA.-Glucan-Indomethacin Model

“…26) CLDM/LCM enhanced the cytoplasmic function of polymorphonuclear leukocytes, accelerated phagocytosis and promoted antibody production by lymphocytes. [27][28][29] It is also reported that subminimal inhibitory concentrations of CLDM influenced bacterial viability, toxin production, and host response. 30) As shown in Fig.…”