摘要
目的建立同时测定人血清中13种氨基酸的超高效液相色谱-串联质谱(UPLC-MS/MS)法。方法以稳定同位素标记的氨基酸作为内标,血清样本经沉淀蛋白后,取上清液进行UPLC-MS/MS分析,色谱柱为Acquity BEH amide色谱柱(2.1 mm×100 mm,1.7μm),流动相为乙腈-水-20 m mol·L^(-1)甲酸铵-0.15%甲酸溶液,用甲酸调p H至3.0,梯度洗脱。结果基质效应不影响测定结果,保留时间(tR)均在11 min内,精氨酸、谷氨酸、脯氨酸、缬氨酸、亮氨酸、异亮氨酸、苯丙氨酸、赖氨酸和苏氨酸的线性范围为1.0~200.0μmol·L^(-1),酪氨酸、丝氨酸的线性范围为5.0~200.0μmol·L^(-1),丙氨酸、甘氨酸的线性范围为10.0~200.0μmol·L^(-1),各氨基酸在标准曲线范围内线性关系良好(r>0.9971),加标回收率均≥89.6%,批内精密度(RSD)均≤3.2%,批间精密度(RSD)均≤4.1%,在测定过程中稳定性符合要求。结论该方法操作简便、专属性强,可同时测定人血清中13种内源性氨基酸的含量。
Objective To establish an ultra high performance liquid chromatography-tandem mass spectrometry（ UPLC-MS/MS） method for the simultaneous determination of 13 kind of amino acids in human serum. Methods The 13 corresponding stable isotope-labeled amino acids were used as the internal standards. For serum samples,acetonitrile was used as the precipitant. Chromatographic separation carried on a acquity BEH amide column（ 2. 1 mm × 100 mm,1. 7 μm）. The gradient elution mobile phase was consisted of acetonitrile-H2 O-20 mmol·L^-1 ammonium formate-formic acid（ p H 3. 0）. Results Assays were validated according to current bioanalytical guidelines,with good matrix effect and linearity（ r〉 0. 9971）. Retention time（ tR） was within 11 min. The linear range of argine, glutamic acid, proline,valine, leucine, isoleucine, phenylalanine, tyrosine and lysine is1. 0-200. 0 μmol·L^-1,of threonine and serine is 5. 00-200. 0μmol·L^-1,of alanine and glycine is 10. 0-200. 0 μmol · L^-1. The spiked recovery rates were more than 89. 6%. The RSDs of intra-day and inter-day precision were less than 3. 2% and 4. 1%. Analytes were stable during the determination. Conclusion A simple and selective method was developed for the simultaneous determination of 13 kinds of amino acids in human serum.
作者
时正媛
王淑梅
鄢丹
SHI Zheng - yuan, WANG Shu - mei, YAN Dan(Department of Pharmacy, Beijing Shijitan Hospital, Capital Medical University ,Beijing 100038, Chin)
出处
《中国临床药理学杂志》
CAS
CSCD
北大核心
2018年第9期1108-1110,1118,共4页
The Chinese Journal of Clinical Pharmacology
基金
G20工程支撑保障基金资助项目(Z161100001816025)
北京市自然科学基金资助项目(7162092)
北京世纪坛医院中青年学科骨干培养专项基金资助项目(2017-QB01)
关键词
串联质谱法
稳定同位素标记
氨基酸
人血清
tandem mass spectrometry
stable isotope label
amino acids
human serum
