Allopurinol is the most commonly used drug for the treatment of hyperuricemia in people, and in view of the risks of fatal hypersensitivity in patients with renal dysfunction, doses based on the glomerular filtration rate are proposed. In veterinary medicine, allopurinol is used in the treatment of canine leishmaniasis (CanL) caused by Leishmania infantum owing to the drug action of inhibiting the parasite's RNA synthesis. However, renal dysfunction frequently ensues from disease progression in dogs. The purpose of the present study was to standardize and validate a sensitive high-performance liquid chromatography-mass spectrometric (HPLC-MS/MS) method to determine the concentration of allopurinol and its active metabolite oxypurinol in canine urine for clinical pharmacokinetic investigation. Urine samples of eleven (11) dogs with naturally occurring CanL and in the maintenance phase of the treatment with alopurinol were used. For the chromatographic analysis of urine, the mobile phase consisted of a solution of 0.1 % formic acid (88 %) in 10 mM ammonium acetate. Separation of allopurinol and oxypurinol occurred in a flow of 0.8 mL/min on a C8 reverse phase column 5 μm, and acyclovir was the internal standard. The HPLC-MS/MS method was validated by reaching the limits of detection and quantification, reproducibility and linearity. The lower limit of quantification achieved by the method was 10 μg/mL for both allopurinol and oxypurinol. Calibration curves were prepared in blank urine added with allopurinol at concentrations of 10-1000 μg/mL, and oxypurinol at 10-200 μg/mL. Coefficients of variation of less than 15 % between intracurrent and intercurrent accuracy values were observed for both allopurinol and oxypurinol. Urine test samples remained stable after being subjected to freeze-thaw cycles and remaining at room temperature for 4 h. The method proved to be adequate to quantify allopurinol and oxypurinol in urine samples from dogs under treatment.

中文翻译：

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通过HPLC / MS-MS检测犬尿中的别嘌呤醇和羟嘌呤醇：专注于兽医临床药代动力学。

别嘌醇是最常用于治疗人高尿酸血症的药物，鉴于肾功能不全患者发生致命性超敏反应的风险，提出了基于肾小球滤过率的剂量。在兽医学中，别嘌呤醇由于抑制寄生虫RNA合成的药物作用，被用于治疗婴儿利什曼原虫引起的犬利什曼病（CanL）。然而，狗的疾病进展常常导致肾功能不全。本研究的目的是标准化和验证灵敏的高效液相色谱-质谱（HPLC-MS / MS）方法，以确定犬尿中别嘌醇及其活性代谢物氧嘌呤醇的浓度，以进行临床药代动力学研究。使用十一只（11）具有自然存在的CanL的狗的尿液样本，并在维持期用金嘌呤醇治疗期间。对于尿液进行色谱分析，流动相由0.1％甲酸（88％）的10 mM乙酸铵溶液组成。在5μm的C8反相色谱柱上，别嘌呤醇和羟嘌呤醇的分离以0.8 mL / min的流速发生，而阿昔洛韦是内标。HPLC-MS / MS方法通过达到检测和定量，重现性和线性度的极限而得到验证。该方法达到的定量下限为别嘌呤醇和羟嘌呤醇均为10μg/ mL。在空白尿中添加浓度为10-1000μg/ mL的别嘌呤醇和氧嘌呤醇为10-200μg/ mL的校正曲线。对于别嘌呤醇和羟嘌呤醇，在电流内和电流间的准确度值之间的变化系数均小于15％。尿液测试样品经过冻融循环后保持稳定，并在室温下放置4小时。事实证明该方法足以定量治疗狗的尿液中的别嘌醇和羟嘌呤。