Kleef U W, Proost J H, Roggeveld J, Wierda J M
Research Group for Experimental Anesthesiology and Clinical Pharmacology, University Hospital, Groningen, Netherlands.
J Chromatogr. 1993 Nov 17;621(1):65-76. doi: 10.1016/0378-4347(93)80077-h.
A sensitive and selective HPLC method was developed for the quantification of the neuromuscular blocking agent rocuronium and its putative metabolites (the 17-desacetyl derivative and the N-desallyl derivative of rocuronium) in plasma, urine, bile, tissue homogenates and stoma fluid. Samples were prepared by extraction of the biological matrix with dichloromethane, after mixing with a KI-glycine buffer. After evaporation of the organic solvent the samples were chromatographed on a reversed-phase HPLC column, using an aqueous buffer-dioxane (84:16, v/v) as the mobile phase. The aqueous buffer consisting of 0.1 M sodium dihydrogen phosphate, 0.11 mM 9,10-dimethoxyanthracene-2-sulphonate (DAS), 0.11 mM 1-heptane-sulfonic acid, was adjusted to pH 3 with orthophosphoric acid. After separation, the eluent was extracted with dichloroethane, and the organic phase was led to a fluorimetric detector, operating at 385 nm (excitation) and 452 nm (emission). The method was validated for the assay in plasma, urine, bile, tissue homogenates and stoma fluid, by determination of the repeatability, reproducibility, accuracy, lower limit of quantification, lower limit of detection, extraction recovery, effect of sample volume, and stability in the biological matrix. The method was found to be sensitive (lower limit of quantification for rocuronium in plasma is 10 ng/ml) and accurate. The interference of concomitant drugs with the assay of rocuronium and its putative metabolites has been studied extensively. In order to confirm the identity of rocuronium and its putative metabolites, a TLC method was developed. The method has been applied successfully in several pharmacokinetic studies with rocuronium.
建立了一种灵敏且具选择性的高效液相色谱法，用于定量测定血浆、尿液、胆汁、组织匀浆和造口液中的神经肌肉阻滞剂罗库溴铵及其假定代谢物（罗库溴铵的17-去乙酰衍生物和N-去烯丙基衍生物）。样品通过与KI-甘氨酸缓冲液混合后，用二氯甲烷萃取生物基质来制备。有机溶剂蒸发后，样品在反相高效液相色谱柱上进行色谱分析，使用水缓冲液-二氧六环（84:16，v/v）作为流动相。水缓冲液由0.1 M磷酸二氢钠、0.11 mM 9,10-二甲氧基蒽-2-磺酸盐（DAS）、0.11 mM 1-庚烷磺酸组成，用正磷酸调节至pH 3。分离后，洗脱液用二氯乙烷萃取，有机相导入荧光检测器，在385 nm（激发）和452 nm（发射）下运行。通过测定重复性、重现性、准确性、定量下限、检测下限、萃取回收率、样品体积的影响以及在生物基质中的稳定性，对该方法在血浆、尿液、胆汁、组织匀浆和造口液中的测定进行了验证。发现该方法灵敏（血浆中罗库溴铵的定量下限为10 ng/ml）且准确。已广泛研究了伴随药物对罗库溴铵及其假定代谢物测定的干扰。为了确认罗库溴铵及其假定代谢物的身份，开发了一种薄层色谱法。该方法已成功应用于多项罗库溴铵的药代动力学研究。
