离子色谱-三重四极杆质谱联用法测定血浆和尿液中3-硝基丙酸

Determination of 3-nitropropionic acid in plasma and urine by ion chromatography-triple quadrupole mass spectrometry

目的建立离子色谱-三重四极杆质谱联用技术测定血浆和尿液中3-硝基丙酸的方法。方法血浆和尿液标本经高氯酸水溶液提取3-硝基丙酸,pH=1~2条件下经固相支持液液萃取法净化,叔丁基甲醚作为萃取剂,萃取液氮吹浓缩后溶于水。以Ionpac AS 19型阴离子色谱柱(250 mm×2 mm,7.5μm)作为分析柱,氢氧化钾作为淋洗液进行梯度分离,电喷雾负离子多反应监测(MRM)模式检测,基质加标标准曲线外标法定量。结果 3-硝基丙酸的线性范围为0.1~250μg/L,相关系数优于0.999,在0.30、1.00、25.0和200μg/L的浓度水平下进行加标回收试验,血浆和尿液中3-硝基丙酸的平均加标回收率为92.0%~114%,相对标准偏差在2.0%~12%之间(n=6),方法的检出限(信噪比为3)均为0.1μg/L。结论本方法简单、灵敏、准确,可用于血浆和尿液中3-硝基丙酸的检测。

Objective A simple and sensitive method for the determination of 3-nitropropionic acid(3-NPA) in plasma and urine by ion chromatography-triple quadrupole mass spectrometry(IC-MS/MS) was established. Methods Plasma and urine samples were extracted with aqueous solution containing 3% perchloric acid, and then the extract was centrifuged to remove the protein and lipids. The target analyte in the extract was cleaned-up by solid supported liquid/liquid extraction under pH=1-2,and methyl tert-butyl ether(MTBE) was used as eluent. After the MTBE was removed by nitrogen,the 3-NPA in the residues was dissolved into water. The separation of 3-NPA was carried out on a Dionex Ionpac AS19 analytic column(250 mm×2 mm, 7.5 μm) with gradient elution using KOH solution electrolytically generated from on-line eluent generation cartridge. Before the eluent flow entered the ion source of mass spectrometer,the potassium ion in the eluent was removed by an in-line suppressor. The 3-NPA was detected by negative electrospray ionization triple quadrupole mass spectrometry in the multiple reaction monitoring(MRM) mode. Results The result showed that the correlation coefficients of linear calibration cuve of 3-NPA were above 0.999 at the corresponding concentration ranges of 0.1-250 μg/L. The average recoveries were 92.0%-114% for 3-NPA in plasma and urine with relative standard deviations of 2.0%-12%(n=6). The limits of detection of 3-NPA(S/N=3)were 0.1 μg/L.Conclusion The method is simple, sensitive and accurate, and can be used for the determination of 3-NPA in plasma and urine.