Metabolomics techniques overview
July 22, 2019METABOLOMICS TECHNIQUES
There are multiple techniques are used in for metabolites for metabolite profiling. Each technique has its advantages and drawbacks. Thus, a combination of different analytical technologies must be used to gain a broad perspective of the metabolome of a tissue. NMR, gas chromatography–mass spectrometry (GC–MS) , liquid chromatography–mass spectrometry (LC–MS) and capillary electrophoresis–mass spectrometry (CE–MS) are most often used techniques.
To metabolomics studies, NMR is well-suited as it can uniquely identify and simultaneously quantify a wide range of organic compounds in the micro-molar range. Unlike MS, the samples can continue for further analysis as NMR is non-destructive. Sample preparation for NMR is straightforward and largely automated. However, the analysis of NMR spectra of complex mixtures has traditionally been fraught with difficulty. NMR has been extensively used for metabolite fingerprinting, profiling and metabolic flux analysis. Low sensitivity, making it inappropriate for the analysis of a large number of low-abundance metabolites become a major limitation of NMR for comprehensive metabolites profiling.
MS has become the technique of choice in many metabolomics studies, as it has high sensitivity and wide range of covered metabolites. Via direct-injection MS or following chromatographic or electrophoretic separation, MS can be used to analyze biological samples. Recent developments of new mass analyzers as well as improvements in mass accuracy dramatically expanded the range of metabolites that can be analyzed by MS and improved the accuracy of compound identification. Recent developments of new mass analyzers as well as improvements in mass accuracy dramatically expanded the range of metabolites that can be analysed by MS and improved the accuracy of compound identification. When using a high-resolution mass spectrometer in direct-injection MS, it provides a very rapid technique to analyse a large number of metabolites, and therefore is extensively used for metabolic fingerprinting and metabolite profiling.
Gas chromatography coupled with electron impact (EI) quadrupole or time-of-flight (TOF) MS (GC–MS) is currently the most mature technology for rapid metabolite profiling. Over 25 years ago, the concept of automated GC–MS metabolic profiling was developed and was later adopted as a major technology for metabolomics. Using this approach, it is possible to simultaneously profile several hundred chemically diverse compounds including organic acids, most amino acids, sugars, sugar alcohols, aromatic amines and fatty acids. By GC–MS, volatile metabolites can be separated and quantified directly. For others, chemical derivatization is required to make them amenable for GC–MS analysis. Although chemical derivatization provides significant improvement in the GC separation of many compounds, it also can introduce artifacts due to the derivatization process itself.
Due to its high sensitivity and a range in analyte polarity and molecular mass wider than GC–MS, LC–MS is being increasingly used in metabolomics applications. Liquid chromatography and specifically high-performance liquid chromatography (HPLC) known as a mature technique that combines high resolution and analytical flexibility as it can be tailored for the analysis of a specific metabolite or class of compounds. Numerous applications for a broad range of metabolites already exist or can be converted for using LC in combination with MS.
In fact that CE–MS is a relatively recent technique compared with GC–MS and LC–MS, it is becoming one of the major analytical techniques for metabolomics. It has very high resolving power with plate numbers of 100 000–1 000 000, very small sample requirement with average injection volume ranging from 1 to 20 nl, and short analysis time. For both targeted and non-targeted analysis of metabolites, CE has been used. Including, analysis of inorganic ions , organic acids, amino acids, nucleotides and nucleosides , vitamins , thiols , carbohydrates and peptides.
References:
Xie, J., Zhang, A., & Wang, X. (2017). Metabolomic applications in hepatocellular carcinoma: toward the exploration of therapeutics and diagnosis through small molecules. RSC Advances, 7(28), 17217-17226.
https://en.wikipedia.org/wiki/Metabolomics
