HPLC法测定人尿液中尼非卡兰浓度

Determination of the concentration of nifekalant in human urine by HPLC

目的建立人尿液中尼非卡兰浓度测定的HPLC法,并研究健康志愿者给予注射用盐酸尼非卡兰后尼非卡兰原形经尿液的累积排泄量。方法采用内标法,以乙酸乙酯进行提取,提取液吹干,残渣用流动相溶解后进行HPLC法检测。色谱柱为μ-BondapakC_(18)(150mm×4.6mm,10μm);流动相为0.05 mol·L^(-1)磷酸二氢铵(pH 5.6)-甲醇-乙腈(60:20:20,V/V/V);流速1.0mL·min^(-1);紫外检测波长270 nm;内标为卡马西平。测定了16名健康志愿者先后给予2次负荷剂量并经连续静滴持续给药6h后,收集不同时间段的尿液样品,同时记录不同时间段的尿液体积,以测定尿药浓度后计算累积排泄量。结果尼非卡兰尿样质量浓度测定的线性范围为20～5000μg·L^(-1),最低定量限20μg·L^(-1);高、中、低3个QC(质量浓度分别为20、300和2500μg·L^(-1))的日内精密度分别为6.4%、5.8%和6.9%,日间精密度分别为8.2%、5.9%和10.9%,回收率分别为(83.0±18.9)%、(85.0±26.2)%和(74.4±4.8)%。结论该方法简便,准确,专属性强,可用于人尿液中尼非卡兰的含量测定。

AIM To develop a HPLC method for the determination of the concentration of nifekalant in human urine and to study its cumulative excretion amounts from urine in Chinese healthy volunteers.METHODS Chromatography was carried out on aμ-Bondapak C（18） column（150 mm×4.6 mm,10μm）,using the mobile phase of 0.05 mol·L^-1 NH4H2PO4（pH was adjusted to 5.6 with 25%ammonium hydrous）-methanol-acetonitrile（60：20：20,V/V/V） at a flow rate of 1.0 mL·min^（-1）.The UV detection wavelength was set at 270 nm,and the internal standard was carbamazepine. Sixteen qualified healthy volunteers were involved in this study.After two intravenous bolus injections and a continuous intravenous injection of nifekalant within 6 h,urine samples at different intervals were collected.RESULTS The urine standard curve was linear within the range of 20 - 5 000μg·L^-1 with the correlation coefficient more than 0.996.The lower limit of quantification（LLOQ） was 20μg·L^-1.The precisions（RSD） of intra-day and inter-day at low,medium and high concentrations（QCs） were less than 10.9%.The average recoveries of nifekalant at low,medium and high concentrations（QCs） were（83.0±18.9）%,（85.0±26.2）%and（74.4±4.8）%,respectively.CONCLUSION The established method is simple,accurate and specific.It has been successfully used for the analysis of nifekalant in urine.