Quantification of trimethylamine-N-oxide (TMAO) and its main related trimethylammonium-containing compounds in human plasma by LC-MS/MS

IF 3.2 3区医学 Q2 MEDICAL LABORATORY TECHNOLOGY

Clinica Chimica Acta Pub Date : 2025-04-11 DOI:10.1016/j.cca.2025.120294

Salvatore Sotgia

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引用次数: 0

Abstract

Background

Trimethylammonium-containing compounds, including choline (CHOL), carnitine (CAR), trimethylglycine (TMG), ergothioneine (ERT), Nε,Nε,Nε-trimethyllysine (TML), γ-butyrobetaine (gBB), and dimethylglycine (DMG) contribute to trimethylamine N-oxide (TMAO) production, a metabolite linked to cardiovascular, renal, and metabolic diseases. An LC-MS/MS method has been established for their simultaneous measurement in human plasma, as an accurate quantification of TMAO and its precursors is crucial for understanding its clinical relevance.

Methods

Blood samples from ten healthy male volunteers were processed using acetonitrile protein precipitation. Analysis was performed using a HILIC column and an isocratic methanol-formic acid mobile phase, achieving a total run time of less than 6 min. Linearity was adequate for all analytes (R² > 0.995), with mean intra- and inter-assay variation coefficients of 2.88 % and 4.23 %, respectively. Recoveries ranged from 95 % to 101 %, limits of detection from 0.009 to 0.068 µmol/L, and limits of quantification from 0.031 to 0.187 µmol/L. Plasma mean concentrations were 3.18 ± 0.73 µmol/L for TMAO, 3.99 ± 0.65 µmol/L for DMG, 9.84 ± 2.08 µmol/L for CHOL, 24.22 ± 6.19 µmol/L for TMG, 0.54 ± 0.22 µmol/L for gBB, 57.29 ± 8.89 µmol/L for CAR, 1.10 ± 0.42 µmol/L for ERT, and 0.40 ± 0.11 µmol/L for TML. Significant inter-individual variability (mean RSD% of 26 %) was observed.

Conclusion

The developed LC-MS/MS method enables rapid, sensitive, and selective quantification of TMAO and its precursors in human plasma. The analytical performance supports its application in clinical and metabolomic studies, contributing to a better understanding the role of TMAO in disease states.

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LC-MS/MS定量测定人血浆中三甲胺- n -氧化物（TMAO）及其主要相关含三甲胺化合物

背景含三甲铵的化合物，包括胆碱（CHOL）、肉碱（CAR）、三甲基甘氨酸（TMG）、麦角硫因（ERT）、Nε,Nε,Nε-三甲基甘氨酸（TML）、γ-丁基甜菜碱（gBB）和二甲基甘氨酸（DMG）有助于产生三甲胺N-氧化物（TMAO），TMAO是一种与心血管、肾脏和代谢疾病相关的代谢物。准确定量 TMAO 及其前体对于了解其临床意义至关重要，因此我们建立了一种 LC-MS/MS 方法，用于同时测量人体血浆中的 TMAO 及其前体。使用 HILIC 色谱柱和甲醇-甲酸等度流动相进行分析，总运行时间小于 6 分钟。所有分析物的线性关系良好（R2 >0.995），测定内和测定间的平均变异系数分别为 2.88 % 和 4.23 %。回收率为 95 % 至 101 %，检测限为 0.009 至 0.068 µmol/L，定量限为 0.031 至 0.187 µmol/L。血浆中 TMAO 的平均浓度为 3.18 ± 0.73 µmol/L，DMG 为 3.99 ± 0.65 µmol/L，CHOL 为 9.84 ± 2.08 µmol/L，TMG 为 24.22 ± 6.19 µmol/L，gBB 为 0.54 ± 0.22 µmol/L，CAR 为 57.29 ± 8.89 µmol/L，ERT 为 1.10 ± 0.42 µmol/L，TML 为 0.40 ± 0.11 µmol/L。结论所开发的 LC-MS/MS 方法可快速、灵敏、选择性地定量检测人体血浆中的 TMAO 及其前体。其分析性能支持其在临床和代谢组学研究中的应用，有助于更好地了解 TMAO 在疾病状态中的作用。

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来源期刊

Clinica Chimica Acta 医学-医学实验技术

CiteScore

10.10

自引率

2.00%

发文量

1268

审稿时长

23 days

期刊介绍： The Official Journal of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC) Clinica Chimica Acta is a high-quality journal which publishes original Research Communications in the field of clinical chemistry and laboratory medicine, defined as the diagnostic application of chemistry, biochemistry, immunochemistry, biochemical aspects of hematology, toxicology, and molecular biology to the study of human disease in body fluids and cells. The objective of the journal is to publish novel information leading to a better understanding of biological mechanisms of human diseases, their prevention, diagnosis, and patient management. Reports of an applied clinical character are also welcome. Papers concerned with normal metabolic processes or with constituents of normal cells or body fluids, such as reports of experimental or clinical studies in animals, are only considered when they are clearly and directly relevant to human disease. Evaluation of commercial products have a low priority for publication, unless they are novel or represent a technological breakthrough. Studies dealing with effects of drugs and natural products and studies dealing with the redox status in various diseases are not within the journal''s scope. Development and evaluation of novel analytical methodologies where applicable to diagnostic clinical chemistry and laboratory medicine, including point-of-care testing, and topics on laboratory management and informatics will also be considered. Studies focused on emerging diagnostic technologies and (big) data analysis procedures including digitalization, mobile Health, and artificial Intelligence applied to Laboratory Medicine are also of interest.