UPLC-MS/MS测定大鼠尿液中柠檬酸、琥珀酸、延胡索酸、乳酸、丙酮酸、苹果酸和果糖

UPLC-MS/MS Simultaneous Determination of Citric Acid,Succinic Acid,Fumaric Acid,Lactic Acid,Pyruvic Acid,Malic Acid and Fructose in Rat Urine and Its Application

目的建立同时测定大鼠尿液中柠檬酸、琥珀酸、延胡索酸、乳酸、丙酮酸、苹果酸和果糖的高效液相色谱-串联质谱(UPLC-MS/MS)检测方法。方法尿液样品用乙腈沉淀法去除蛋白,色谱分离采用UPLC HSS C18(2.1mm×100mm,1.7μm)柱,流动相为甲醇-水(含0.3%甲酸)梯度洗脱,采用电喷雾多反应监测模式定量。结果日间和日内精密度小于12%,准确度范围从92.1%到108.7%。标准曲线在0.05~20μg·mL^-1范围内呈线性,相关系数r大于0.995。结论该方法可用于亚健康大鼠模型氨基酸含量测定。结果表明柠檬酸、乳酸、丙酮酸、苹果酸在亚健康组有所升高,而北美海蓬子干预后有降低。

OBJECTIVE To establish a method for simultaneous determination of citric acid,succinic acid,fumaric acid,lactic acid,pyruvic acid,malic acid and fructose in urine by ultra-high performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS).METHODS The urine samples were prepared by acetonitrile precipitation.The chromatographic separation was performed on a UPLC HSS C18(2.1 mm×100 mm,1.7μm) column using a gradient elution of methanol-water(containing 0.3% formic acid) as mobile phase.The multiple reactions monitoring in electrospray was for quantitative determination.RESULTS The intra-day and inter-day precision were less than 12%,with accuracy ranging from 92.1% to 108.7%.The standard curve was linear in the range of 0.05-20μg·mL^-1 with a correlation coefficient r greater than 0.995.CONCLUSION This method could be used to determine the amino acid content of sub-healthy rat model.The results showed that citric acid,lactic acid,pyruvic acid,malic acid increased in the sub-health group,while decreased after intervention by SalicorniabigeloviiTorr.