# Skin Cancer Metabolic Profile Assessed by Different Analytical Platforms
## 5 Analytical Platform in Metabolomics Workflow
### Mass Spectrometry-Based Metabolomics  
Mass spectrometry (MS), a potent analytical tool, has important uses and wide a range of applications nowadays, as it measures the mass of molecules with a high level of sensitivity [34]. It allows the analysis of hundreds to thousands of metabolites on a routine basis [33]. The implementation of Mass spectrometry has taken so long and was not used on a larger scale until the 1990s, because it analyzes gaseous, charged molecules. Although biomolecules are polar and large, they cannot be easily shifted to the ionized gas phase. Currently, Mass spectrometry is also a vital component of the metabolomics and proteomics field [34].  
Mass spectrometry-based metabolomics approaches are characterized by their capability to distinguish and quantify thousands of metabolites simultaneously. However, these tasks can be complex and are associated with major challenges, due to metabolites diversity, including dynamic range and chemical complexity. Moreover, metabolite quantification within complex combinations can be difficult due to isomers, ion suppression, and fragmentation. At present, the most comprehensive combinations of analytical methods can detect and quantify around 8000 of the 114,100 metabolites predicted to exist in humans. However, they cannot specify a fixed upper limit for metabolite amounts [35]. In mass spectrometry analysis, metabolites are first exposed to chromatographic separation techniques such as gas chromatography (GC) or liquid chromatography (LC), before detection [33].  
In GC-MS and LC-MS-based metabolomic approaches, relative quantification is carried out, which determines fold changes in expression between two different samples, with one being a reference sample, so metabolomic data are detected as relative quantities. In contrast, the NMR based-approach uses absolute quantification, providing absolute concentrations that are directly proportional to peak intensities. The exact amount of the template used for the curve is determined in NMR-based studies.  
The first and vital step is the fast enzyme quenching of the metabolic process and terminating all enzyme activities, to get a stable extract of metabolites that quantitively reflects the levels of endogenous metabolite in the original cells and tissues. This reflection is less important in the case of biofluids such as urine, plasma, and serum samples. Indeed, for each tissue type or biofluid, there is a specific method for sampling and extraction. The second step is metabolite separation on a chromatographic column, such as LC, GC, or capillary electrophoresis (CE) columns.  
In this review we focus on GC and LC techniques only. The third step is metabolite ionization using an ion source, followed by ion separation in a mass spectrometer, according to their mass-to-charge m/z ratio. Several ionization types are used, such as electrospray ionization (ESI), electron impact (EI), and atmospheric pressure chemical ionization (APCI). Furthermore, mass analyzers used can be time of flight (TOF), Orbitrap, quadrupole (Q), and ion trap (IT). Finally, metabolites are detected and identified through their MS signature and retention time (RT) [35].