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Metabolomics is developing rapidly. In medical science research, it is hoped that analysis to identify abnormal metabolic pathways and characteristic biomarkers closely related to diseases will provide a basis for further elucidation of disease pathogenesis.

Metabolomics Research Methods

Metabolomics collects samples through established metabolomics research methods, and subsequently uses techniques to obtain metabolites, determine whether there are statistical differences in metabolites between groups and perform further analysis. There are two types of metabolomics research methods: untargeted metabolomics and targeted metabolomics.

• Untargeted metabolomics analyzes the full metabolite profile without making presuppositions about specific metabolites and is relatively less reproducible and targeted. Often referred to as hypothesis generation or discovery phase experiments, they are characterized by the generation of large amounts of complex and multi-effective data and therefore require high-performance bioanalytical tools. Untargeted metabolomics are relatively open-ended analyses that are often used to discover previously unknown physiological patterns and provide direction for the discovery of new metabolites and metabolic pathways.

• Targeted metabolomics studies validate previous scientific hypotheses or possible biomarkers and perform more targeted studies with high precision, reproducibility and selectivity, also known as hypothesis-driven experiments. Targeted metabolomics is a relatively closed analysis suitable for identifying novel biomarkers, studying metabolite functions and pathways, and further revealing associations between metabolites and diseases.

Targeted metabolomics has two limitations in its application: 1) there may be metabolites that go undetected, thus reducing the chance of discovery; 2) a large amount of data collection from non-targeted studies is required before using the method.

Techniques of Metabolomics

• Nuclear magnetic resonance (NMR) technology

NMR technology is a spectroscopic technique that uses the absorption of radiation by different nuclei to produce different resonance frequencies, and translates these resonance frequencies into molecular chemical and structural information. Different target nuclei to which a magnetic field is applied produce different metabolomics data. Hydrogen is the most common target nucleus (¹H-NMR), while other atoms such as carbon (¹³C-NMR) and phosphorus (³¹P-NMR) can also be used to obtain information on specific metabolite types by NMR techniques. Currently, ¹H-NMR spectroscopy, ²H-NMR spectroscopy, and high-resolution magic angle rotation (HRMAS-NMR) spectroscopy are common. HRMAS-NMR spectroscopy can be used for metabolic profiling of intact tissues without any sample pre-treatment and has been used to study metabolomic analysis of small intact tissue samples including brain, kidney, liver, etc.

NMR spectroscopy does not require chromatographic processing. Sample preparation is simple and can be repeated multiple times. It is a rapid and efficient non-targeted analytical technique for qualitative and quantitative studies of metabolites. It has been widely used in many fields such as biological structure, biology and biochemistry, and is one of the main analytical methods in metabolomics. However, low sensitivity has been an inherent drawback of NMR spectroscopy and a challenge in its application in biomedical research.

• Mass spectrometry (MS)

MS analysis uses electric and magnetic fields to separate and detect moving ions by mass-to-charge ratio (m/z). Current mass spectrometry techniques include liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS), and capillary electrophoresis-mass spectrometry (CE-MS).

LC-MS has the advantage of high sensitivity and high resolution and is capable of detecting more (200 to 500) metabolites. It is probably the most widely used mass spectrometry technique today. In LC-MS, several types of atmospheric pressure ionization methods are used for the ionization of different classes of metabolites: the most commonly used electrospray ionization is used for the initial screening of unknown metabolites; atmospheric pressure chemical ionization and atmospheric pressure photoelectron ionization are suitable for the detection of non-polar metabolites and have been widely used in lipidomics studies.

GC-MS has high separation efficiency and good sensitivity. However, only volatile or derivatizable volatile compounds can be detected, and some metabolites that cannot be ionized cannot be detected.

Reference

1. Jacob, M., Lopata, A. L., Dasouki, M., & Abdel Rahman, A. M. (2019). Metabolomics toward personalized medicine. Mass spectrometry reviews, 38(3), 221-238.