Large Doses Of Ropivacaine For Sciatic Nerve Block In Dogs The Pharmacokinetics And Tolerability Study

Part 1The Feasibility of Reversed-Phase High Performance Liquid Chromatography for Determination of the Concentration of Ropivacaine in Dog PlasmaBackgroundRopivacaine, an aminoamide derivative, unlike bupivacaine, ropivacaine is exclusively the (S)-enantiomer. ropivacaine potentially provides lower toxicity on heart and central nervous system compared with bupivacaine. Ropivacaine has an advantage over lidocaine in the drug metabolism. Lidocaine produces the toxic metabolites,monoethyl-glycinexylidide and glycinexylidide, in its hepatic metabolism, which are circulating with higher concentration in blood. Blood concentrations of ropivacaine metabolites, 3-hydroxy-ropivacaine, have known to be very low (undetectable levels), even though they are pharmacologically active. These characteristics of ropivacaine may be suitable for nerve block. So we evaluated a rapid and sensitive method for quantitating plasma ropivacaine in dog.Materials and methods1. HPLC Apparatus and Analytical ConditionsHPLC system used for present study consisted of a pump (waters 610,Waters, america), an UV detector (UV-996, PDA)and an integrator (CAY-1, CHANGAN, Beijing). Intertsil C18 column (4.6 i.d.₂50 mm,) as the analytical column was maintained at ambient temperature. The detection wavelength was set at 215 nm. The mobile phase solvent consisted of methanol and water, 0.05 acetic acid buffer adjusted to pH 3.5 (50∶50, v/v) was pumped at a flow rate of 1.4 ml/min.2. Preparation of Standard SolutionsThe control plasma was prepared by using an alternative dog plasma. Standard solution of ropivacaine hydrochloride was prepared by diluting the stock solution with dog plasma to make the concentration of 0.1,0.5,1.0,2.5,5.0,10.0 and25.0μg/ml and stored at ₂0℃until use. The IS was prepared as the 1.0 mg/ml solution in distilled water and stored at 4℃.3. Assay ProceduresTo 500μl plasma sample in a 10 ml siliconized test tube, 20μl IS solution (10mg/ml) and 500μl acetonitrile were added. The mixture was centrifuged at 3000g for 10min. The upper organic phase was transferred to another tube, and then the plasma sample was treated by further 500μl acetonitrile. The upper organic phase was transferred to the tube again. The combined organic phase was evaporated to dryness under a stream of nitrogen at 40℃. The residue was dissolved in 200μl of the mobile phase solvent and the 10μl aliquot was injected into the HPLC column.Results1. Typical typical chromatograms for the blank plasma and the plasma spiked with ropivacaine and internal stand bupivacaine were observed at the retention times of 9. 805 and 14. 988min, respectively. The detection limit ofropivacaine in plasma was 0.05μg/ml.2. The calibration curve for determining plasma ropivacaine was linear at the concentration of 0.1—25μg.ml^-1(r=0.9992). The equation of the calibration curve calculated by regression analysis for ropivacaine was Y=0.1794X＋0.3806, where Y was the plasma ropivacaine concentration and X, the peak area ratio of ropivacaine to bupivacaine.3. The recovery of ropivacaine from dog plasma examined at concentrations of 0.5,2.5,25μg/ml were 91.2%,92.4%,93.6%, respectively, with RSD less than 3.40%.. The assay precision of ropivacaine was evaluated by the intra-day and inter-day validation with the values at 1.35～2.88% and 1.80～3.76%, respectively.Conclusions1. The method of Reversed-Phase high performance liquid chromatography can be used in determining the concentration of ropivacaine in dog plasma;2. The method is simple,rapid,accurate and convenient for scrutinizing of the concentration of ropivacaine in dog plasma. Part 2The Pharmacokinetics and Tolerance Study of High Dose Ropivacaine for Dog Single Sciatic Nerve BlockBackgroundIt is taughted that peripheral nerve block has some advantages over intrathecal and general anesthesia. However, larger dose of local anesthetic is usually needed for peripheral nerve block, such as sciatic nerve block. Unfortunately, this may cause the high incidence of toxicity of local anesthetics. An ideal local anesthetic should be lower toxicity, faster onset, providing complete sensory and motor block, and more satisfactory analgesia as well.Ropivacine is a new, long-acting aminoamide local anesthetic. It is a pure S-(-) enantimer monohydrate of the hydrochloride salt. This contribute to its decreased central nervous system toxicity and cardiotoxity when compared to that of bupivacaine. In contrast to bupivacaine, it has shown fewer inherent vasodilator properities.The pharmacodynamics and pharmacokinetics of ropivacaine used for epidural,brachial plexus and caudal block had been studied, but was not found in sciatic nerve block. This study was to investigate the pharmacokinetics and tolerability of high dose ropivacaine for single sciatic nerve block in dogs.Materials and MethodsHealthy adult mongrel dogs were randomly assigned to receive 10mg/kg or 20mg/kg ropivacaine for single sciatic nerve block. Animal preparation more than 48h before nerve block, anesthesia was induced with 30mg/kg of IM pentobarbital sodium. Both femoral arterials and veins were cannulated for arterial pressure monitoring and sampling, for drug and fluid infusion, respectively. The catheter was inserted into endolemmal of sciatic nerve. The adverse reactions,blood pressure and electrocardiogram were recorded during anesthesia, before or after ropivacaine injection0,10,20,30,40,60,90,120,150,180,240,360,720min, 3ml femoral blood was sampled to receive reverse-phase high performance liquid chromatography respectively at the indicated time. The samples of the femoral blood were also taken and determined if the adverse reactions were found.The pharmacokinetics parameters were determined from concentration-time data with DAS 1.0 software package. Other parmmeters were analysed with SPSS 13.0 software package. The value of p＜0.05 was considered statistically significant.Results1. Time to peak plasma(Tmax) after 10mg/kg ropivacaine sciatic nerve block was 0.39±0.09h, t₁/2。was 0.34±0.09h, t_1/2 was 4.42±0.90h, T_1/2ka was 0.09±0.09h clearance (CL) was 0.43±0.09h^-1;2. The results of 20mg/kg ropivacaine for sciatic nerve block were 0.36±0.07h, 0.29±0.27, 5.37±1.89h, 0.04±1.38h^-1, respectively.3. The peak plasma concentration of total ropivacaine and area under curve(0-12h) in group 10mg/kg were significantly lower than that with ropivacaine 20mg/kg. Other parameters were no significant difference between two groups.4. Convulsion was observed in two dogs after ropivacaine 20mg/kg administration for sciatic nerve block, at which average plasma concentration was measured around 13.12mg/l.Conclusion1. The concentration-time curve in the two groups was adequately matched to two-compartment open model.2. The plasma concentration of ropivacaine was fairly relative to the doses given for sciatic nerve block.3.10mg/kg ropivacaine can be safely used for single sciatic nerve block and no systemic toxic reactions were found in this dog study setting. Cmax of ropivacaine 20mg/kg is significantly higher than that of ropivacaine 10mg/kg for sciatic nerve block. However, 20mg/kg ropivacaine for sciatic nerve block was hardly tolerated by dogs.