超高效液相色谱-串联质谱法测定人尿中硫酸吲哚酚和硫酸对甲酚的含量  

UPLC-MS/MS Determination of Indoxyl Sulfat and p-Cresyl Sulfate in Human Urine

作　　者：肖永华[1] 许开冰 王胜[1] 毛翔[1] 卢圆圆 XIAO Yonghua;XU Kaibing;WANG Sheng;MAO Xiang;LU Yuanyuan(Wuhan Centers for Disease Prevention and Control,Wuhan 430022,China;Tongji Hospital,Tongji Medical College of Huazhong University of Science and Technology,Wuhan 430030,China)

机构地区：[1]武汉市疾病预防控制中心,武汉430022 [2]华中科技大学同济医学院附属同济医院,武汉430030

出　　处：《理化检验（化学分册）》2019年第10期1151-1156,共6页Physical Testing and Chemical Analysis(Part B:Chemical Analysis)

基　　金：国家自然科学基金项目(31700294)

摘　　要：选取武汉市中心及周边城区监测对象的尿样418份(其中儿童295份,孕妇123份),分别操作如下:取尿样100μL置于离心管中,加入硫酸吲哚酚-d4及硫酸对甲酚-d7内标溶液各100μL,再加入乙腈700μL,涡旋振荡1min并高速离心10min,使样品中杂质沉淀析出,取上清液100μL,加水定容至1.0mL,混匀,作为分析试液。选择ACQUITY UPLC■ HSS T3色谱柱(100mm×2.1mm,1.8μm)为固定相,进样量为10μL,用由5mmol·L^-1乙酸铵溶液和甲醇按5%~95%之间变动的体积比组成流动相,在0~5.0min之间分4个时段进行梯度洗脱,使硫酸吲哚酚和硫酸对甲酚分离。质谱测定中采用电喷雾离子源,负离子扫描模式和多反应监测模式。结果表明:硫酸吲哚酚和硫酸对甲酚的质量浓度在相同的范围(5.00~500μg·L^-1)内与其对应的峰面积与各自内标的峰面积的比值之间呈线性关系。以3倍信噪比计算两者的检出限(3S/N)依次为0.5,0.1μg·L^-1。取实际样品作基体,在3个浓度水平上加入两组分的混合标准溶液进行精密度和回收试验,测得回收率在93.0%~98.4%之间,测得相对标准偏差(n=6)在4.2%~5.3%之间。所测得大量样品的数据可为评价学龄儿童和孕妇肾功能情况提供依据。418 urine samples were collected from 295 school-children and 123 pregnant women residing in the central area and at suburbs of Wuhan municipality.100μL of the urine sample together with 100μL of each of the internal standard solution of indoxyl sulfate-d4 and of p-cresyl sulfate-d7,and 700μL of acetonitrile were added into a centrifuging tube and after swirling for 1 min,the mixed solution was centrifuged at high rate for 10 min,to separate the impurities in the sample by precipitation.100μL of the supernatant were taken and used as testing solution for analysis.ACQUITY UPLC■ HSS T3 chromatographic column(100 mm×2.1 mm,1.8μm)was used as stationary column,and an aliquot of 10μL of the testing solution were introduced.Mixtures of 5 mmol·L^-1 NH4 OAc solution and methanol,with their volumic ratio varied in the range of 5%-95%in 4 time intervals within5 min,were used as mobile phases in the gradient elution,to have the indoxyl sulfate and p-cresyl sulfate well separated.In MS/MS determination,ESI-with mode of negative ion scanning and the mode of MRM were adopted.As shown by the result that linear relationships between values of ratios of peak areas of indoxyl sulfate and p-cresyl sulfate to the peak areas of their respective internal standards and values of corresponding mass concentration of indoxyl sulfate and p-cresyl sulfate were found in the same range of 5.00-500μg·L^-1.Values of detection limits(3 S/N)were found 0.5μg·L^-1(for indoxyl sulfate)and 0.1μg·L^-1(for p-cresyl sulfate).Tests for recovery and precision were made by addition of standard solutions of indoxyl sulfate and p-cresyl sulfate at 3 concentration levels to a real sample as matrixes.Values of average recovery were found in the range of 93.0%-98.4%and RSDs(n=6)were found in the range of 4.2%-5.3%.It was shown that the data of urine analysis of the 418 samples provided a reliable basis in judging of function of school-children and pregnant women.

关 键 词：超高效液相色谱-串联质谱法 硫酸吲哚酚 硫酸对甲酚 人尿 

分 类 号：O657.63[理学—分析化学]