Simultaneous determination of unbound ropivacaine in rat blood and brain using microdialysis

To investigate the pharmacokinetics of ropivacaine in rat blood and brain, a sensitive HPLC method and microdialysis were developed for the simultaneous determination of unbound ropivacaine in rat blood and brain. Adult, male Sprague-Dawley rats (290-350 g) were anesthetized with sodium pentobarbital (50 mg/kg, i.p.). Two microdialysis probes were inserted, one into the jugular vein toward right atrium, and one into the brain striatum of rats. Ropivacaine (5 mg/kg, i.v.) was then administered via the femoral vein. Blood and brain dialysates were collected and eluted with a mobile phase containing methanol-acetonitrite-20 mM monosodium phosphoric acid (pH 5.5) (10:40:50, v/v/v) in a liquid chromatographic system. Separation of ropivacaine was achieved by a CN column (Phenomenex Luna, 250×4.6 mm, particle size 5 μm; Torrance, CA, USA) within 10 min. The UV detector wavelength was set at 205 nm and the detection limit of ropivacaine was 20 ng/ml. The intra- and inter-day accuracy and precision of the analyses were less than 10% in the ranges of 0.02-5 μg/ml. The pharmacokinetic data were calculated from the individual animal measurements of dialysate concentration versus time. This method exhibits no endogenous interference and its sensitivity is sufficient for the determination of biological samples. The present results confirm that microdialysis sampling followed by LC separation with UV detection represents a viable approach for the measurement of free ropivacaine in rat brain and plasma.