【摘要】 目的:建立并验证一种简便快捷的超高效液相色谱(UPLC)测定癫痫患儿血浆中氯巴占的药物浓度方法。方法:采用Waters ACQUITY UPLC BEH C18色谱柱(100 mm×2.1 mm×1.7μm)。流动相为乙腈-10 mmol/L磷酸二氢铵溶液(体积比45∶55,甲酸调至pH 4.0),流速0.15 mL/min,进样量2μL,检测波长230 nm,柱温30℃。分别精密吸取血浆样品100μL和蛋白沉淀剂200μL涡旋混合后以12 000 r/min离心10 min,将上清液过滤(0.22μm针头式过滤器)后进样测定。结果:氯巴占的保留时间为2.8 min。氯巴占的血药浓度在10～600 ng/mL范围内呈现出良好的线性关系(Y=21 922X+143 806,r=0.999 8)。检测下限为10 ng/mL,定量下限为30 ng/mL。该方法的定量下限(LLOQ,30 ng/mL)、低浓度质控(LQC,90 ng/mL)、中浓度质控(MQC,240 ng/mL)和高浓度质控(HQC,480 ng/mL)4个质控浓度的日内、日间精密度良好,相对标准偏差(RSD)<15.0%,提取回收率为91.18%～99.92%,稳定性良好(RSD<15.0%)。32例癫痫患儿服用氯巴占10～12 h后氯巴占平均血药浓度为52.10 ng/mL,治疗有效率为65.62%,不良反应发生率为31.25%。年龄>3～6岁组患儿的氯巴占血药浓度高于年龄>14～18岁组(P<0.05);体质量指数(BMI)18.5～<24.0 kg/m~2组患儿的氯巴占血药浓度高于BMI≥24.0 kg/m~2组(P<0.05)。氯巴占的稳态血药浓度参考区间为33.80～339.50 ng/mL。结论:本研究建立的UPLC法能在癫痫患儿在使用氯巴占过程中实时准确快速地检测血药浓度。更多还原

【Abstract】 Objective: To establish and validate a simple and rapid ultra-high performance liquid chromatography(UPLC) method for the determination of clobazam blood concentration in children with epilepsy. Methods: The chromatographic separation was performed on the Waters ACQUITY UPLC BEH C18 column(100 mm×2. 1 mm×1. 7 μm) with a mobile phase consisting of acetonitrile and 10 mmol/L ammonium dihydrogen phosphate solution(45 ∶ 55, adjusted to p H 4. 0 with formic acid). The flow rate was 0. 15 mL/min, the injection volume was 2 μL, the detection wavelength was set at 230 nm, and the column temperature was maintained at 30 ℃. Totally 100 μL plasma sample and 200 μL protein precipitant were vortex mixed and centrifuged at 12 000 r/min for 10 min. The supernatant was filtered through a 0. 22 μm syringe filter and determined by injection. Results: The retention time of clobazam was 2. 8 min. The method demonstrated good linearity over the concentration range of 10 to 600 ng/mL(Y= 21, 922X+143, 806, r= 0. 999,8). The detection limit of the method was 10 ng/mL, and the quantitative limit was 30 ng/mL. The lower limit of quantification(LLOQ, 30 ng/mL), low concentration quality control(LQC, 90 ng/mL), medium concentration quality control(MQC, 240 ng/mL) and high concentration quality control(HQC, 480 ng/mL) of the method had good intra-and inter-day precision(relative standard deviation, RSD<15. 0%), the extraction recovery was from 91. 18% to 99. 92%, and the stability was good(RSD<15. 0%). The average blood concentration of clobazam in 32 cases of children with epilepsy measured 10 to 12 hours after administration was 52. 10 ng/mL, the efficacy rate of clobazam treatment was 65. 62%, and the incidence of adverse drug reactions was 31. 25%. The blood concentration of clobazam in children aged from 3 to 6 years was significantly higher than those aged from 14 to 18 years(P< 0. 05). The blood concentration of clobazam in children with body mass index(BMI) of 18. 5 to <24. 0 kg/m~2 was higher than that in children with BMI of ≥24. 0 kg/m~2(P<0. 05). The reference range for the steady-state blood concentration of clobazam was from 33. 80 to 339. 50 ng/mL. Conclusion: The UPLC method established in this study can enable real-time, accurate, and rapid detection of the blood concentration of children with epilepsy during the clobazam treatment.更多还原