超高效液相色谱-串联质谱法快速测定大鼠血清中8种神经递质

Simultaneous Determination of Eight Kinds of Neurotransmitters in Rat Blood Serum by Ultra-high Performance Liquid Chromatograph-Tandem Mass Spectrometry

建立了同时测定8种神经递质(5-HT、GABA、Glu、ACH、NE、DA、5-HIAA和HVA)的超高效液相色谱-三重四极杆质谱方法(UHPLC-MS/MS),并用于大鼠血清样品的测定。大鼠血清经含0.1%甲酸-乙腈沉淀蛋白处理后,选用Waters ACQUITY UPLC BEH C18色谱柱(100 mm×2.1 mm,1.7μm),以0.1%甲酸-乙腈为流动相梯度洗脱进行分离;采用电喷雾离子源(ESI),在正离子扫描下,采用多反应监测模式(MRM)对8种神经递质及内标化合物进行检测。结果表明,8种神经递质可在10 min内准确测定;定量限为1.8 ng/m L,且线性良好,相关系数均大于0.994,日内、日间精密度(n=6)≤9.2%,稳定性、加标回收率和基质效应等均符合分析要求。本方法分析时间短、准确度好、灵敏度高、专属性强、稳定性好、基质效应小,适用于血清中3类(单胺类、氨基酸类和乙酰胆碱)共8种神经递质的同时定量检测。

An ultra-high performance liquid chromatography tandem mass spectrometric(LC-MS/MS)method for simultaneous determination of8kinds of neurotransmitters(5-HT,GABA,Glu,ACH,NE,DA,5-HIAA,HVA)in rat serum was developed.The blood samples were extracted by0.1%formic acid in acetonitrile and separated on a Waters ACQUITY UPLC BEH C18(2.1mm伊100mm,1.7um)using gradient elution,with the mobile phase consisting of acetonitrile and0.1%formic acid in water.The samples were then ionized with positive electrospray(ESI+),and detected under multiple reaction monitoring(MRM)mode.As a result,8kinds of neurotransmitters were determined accurately in10min with a limit of quantitation of1.8ng/mL and intra-day and inter-day precisions of≤9.2%(n=6).This method showed a good linearity in detection of neurotransmitters and the linear correlation coefficients were greater than0.994.Also the stability,recovery and matrix effect were eligible for the analysis.This method showed high accuracy,sensitivity,strong specificity,good stability,small matrix effect and short time for analysis,and was suitable for the quantitative determination of monoamine,amino acids and acetylcholine neurotransmittes in rat serum.