超高效液相色谱-串联质谱法同时测定高脂血症患者血浆中 5 种必需氨基酸
Simultaneous determination of five essential amino acids in plasma of Hyperlipidemic subjects by UPLC-MS/MS.
机构信息
Department of Nephrology, The Affiliated Yueqing Hospital of Wenzhou Medical University, Yueqing, Zhejiang, China.
School of Medicine, Zhejiang University, Zhejiang, Hangzhou, China.
出版信息
Lipids Health Dis. 2020 Mar 23;19(1):52. doi: 10.1186/s12944-020-01216-8.
BACKGROUND
Millions of adults have been reported with hyperlipemia in the world. It is still unclear whether the plasma level of essential amino acids (AAs) will be influenced by the hyperlipemia. This study was aimed to investigate the AAs levels and the underlying metabolic relationship in hyperlipidemic subjects.
METHODS
An ultra-high performance liquid chromatography-tandem mass spectrometric (UPLC-MS/MS) method was developed for the determination of phenylalanine (Phe), valine (Val), histidine (His), tryptophan (Trp), and methionine (Met). Plasma samples (100 μL) were precipitated by acetonitrile (300 μL) and analyzed on a BEH C18 (2.1 mm × 100 mm, 1.7 μm) column at 40 °C by gradient elution. The mobile phase composed of 0.1% formic acid and acetonitrile was used with flow rate at 0.2-0.4 ml/0-3 min. Five AAs were determined at positive electrospray ionization (ESI+) at m/z 118.1/72.1 (Val), 150.12/104.02(Met), 156.06/110.05(His), 166.1/120.1(Phe), and 205.2/188.02 (Trp). A total of 75 healthy subjects and 83 hyperlipidemic subjects, who had blood routine test and plasma lipid test were determined by developed UPLC-MS/MS.
RESULTS
It was shown that there was good linearity for Val, Met, His, Phe, and Trp within 1-100 μg/mL. The relative standard deviations of precision and accuracy were all within 15%. The level of Val, Phe, Trp, His, and Met were 35.34 ± 15.64, 22.72 ± 9.13, 17.23 ± 4.94, 16.78 ± 13.64, and 6.24 ± 1.97 μg/mL in healthy subjects, while they were 38.04 ± 16.70, 22.41 ± 8.45, 15.62 ± 5.77, 18.35 ± 14.49, and 6.21 ± 1.97 μg/mL in hyperlipidemic subjects respectively. The Spearman's correlations analysis showed that there were high correlations between Val, Phe, Trp, His, Met and triglyceride in healthy subjects. While, those correlations decreased in hyperlipemia cases.
CONCLUSION
A convenient and sensitive method for simultaneous determination of Val, Phe, Trp, His, and Met in human plasma was developed. There was a high correlation between Val, Phe, Trp, His, Met and triglyceride. Hyperlipemia influences the metabolic balance of His, Phe, Trp, Met and Val.
背景
据报道，全世界有数百万成年人患有高脂血症。目前尚不清楚高脂血症是否会影响必需氨基酸（AA）的血浆水平。本研究旨在调查高脂血症患者 AA 水平及其潜在的代谢关系。
方法
建立了一种超高效液相色谱-串联质谱（UPLC-MS/MS）法测定苯丙氨酸（Phe）、缬氨酸（Val）、组氨酸（His）、色氨酸（Trp）和蛋氨酸（Met）的方法。用乙腈（300μL）沉淀 100μL 血浆样品，在 40°C 下用 BEH C18（2.1mm×100mm，1.7μm）柱进行梯度洗脱。流动相由 0.1%甲酸和乙腈组成，流速为 0.2-0.4ml/0-3min。在正电喷雾电离（ESI+）下，在 m/z 118.1/72.1（Val）、150.12/104.02（Met）、156.06/110.05（His）、166.1/120.1（Phe）和 205.2/188.02（Trp）处测定 5 种 AA。共对 75 名健康受试者和 83 名高脂血症受试者进行了血液常规检查和血浆脂质检查，采用所建立的 UPLC-MS/MS 进行了测定。
结果
结果表明，Val、Met、His、Phe 和 Trp 在 1-100μg/mL 范围内具有良好的线性关系。精密度和准确度的相对标准偏差均在 15%以内。健康受试者中 Val、Phe、Trp、His 和 Met 的水平分别为 35.34±15.64μg/mL、22.72±9.13μg/mL、17.23±4.94μg/mL、16.78±13.64μg/mL 和 6.24±1.97μg/mL，而高脂血症患者中 Val、Phe、Trp、His 和 Met 的水平分别为 38.04±16.70μg/mL、22.41±8.45μg/mL、15.62±5.77μg/mL、18.35±14.49μg/mL 和 6.21±1.97μg/mL。Spearman 相关分析表明，健康受试者中 Val、Phe、Trp、His 和 Met 与甘油三酯之间存在高度相关性。然而，在高脂血症病例中，这些相关性降低。
结论
建立了一种同时测定人血浆中 Val、Phe、Trp、His 和 Met 的灵敏、简便的方法。Val、Phe、Trp、His、Met 与甘油三酯之间存在高度相关性。高脂血症影响 His、Phe、Trp、Met 和 Val 的代谢平衡。
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