Automated analytical systems for drug development studies. Part IV. A microdialysis system to study the partitioning of lomefloxacin across an erythrocyte membrane in vitro.

An automated system utilizing microdialysis sampling, intermittent dosing, and liquid chromatographic analysis was assembled in order to study the partitioning of lomefloxacin, a fluoroquinolone antimicrobial, into human erythrocytes in vitro. The apparent erythrocyte:buffer partition coefficient was found to be approximately 2.0 with this system and by a manual method. The value was concentration-dependent; lower partition coefficients were observed at lomefloxacin concentrations less than 1 microgram ml-1. At all concentrations, values obtained by microdialysis were statistically indistinguishable from those obtained by a conventional manual method. The results indicate that erythrocyte partition coefficients can be measured successfully with the microdialysis system. Furthermore, microdialysis sampling eliminates the tedious methodology associated with traditional erythrocyte partitioning studies, including sample clean-up. Due to automated intermittent dosing and on-line LC analysis, the system operates unattended.

A 19F NMR study of lomefloxacin in human erythrocytes and its interaction with hemoglobin.

19F NMR spectroscopy of a model fluoroquinolone, lomefloxacin, in an erythrocyte suspension showed separate resonances for the intra- and extra-cellular compartments. The intra-cellular peak revealed significant line broadening of the fluorine signals of lomefloxacin. Line broadening also occurred in the presence of oxyhemoglobin (HbO2), hematin, globin and iron. This evidence indicated that lomefloxacin interacted with these compounds; however, ultrafiltration experiments indicated that there was only weak binding (5%) of lomefloxacin to HbO2. 19F and 31P NMR spectroscopy revealed that lomefloxacin may compete with 2,3-diphosphoglycerate for its binding site on HbO2. An apparent partition coefficient of 1.90 +/- 0.15 was observed for lomefloxacin in human erythrocytes, utilizing LC analysis.

The effect of protein binding on ivermectin uptake by bovine brain microvessel endothelial cells.

The effect of albumin binding on ivermectin uptake and transfer across the endothelial component of the blood-brain barrier (BBB) was determined with an in vitro model comprised of bovine brain microvessel endothelial cell (BMEC) monolayers. Cellular uptake of ivermectin was limited in the absence of albumin and 90% inhibited in the presence of 10% albumin. Cell membrane association of ivermectin, as followed by fluorescent probe labelling, was observed only at high (micromolar) concentrations of the drug. Membrane association was about 75% inhibited in the presence of albumin. Similarly, transfer across BMEC monolayers was restricted, equivalent to that of BBB impermeant markers. Unlike the uptake studies, however, albumin had little effect on the transfer of ivermectin across BMEC monolayers. These results support recent in vivo findings on the distribution of ivermectin into the brain and suggest that ivermectin has only a limited affinity for the endothelial component of the normal BBB.