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Analytical chemistry studies and uses instruments and methods to separate, identify, and quantify matter. In practice, separation, identification or quantification may constitute the entire analysis or be combined with another method. Separation isolates analytes. Qualitative analysis identifies analytes, while quantitative analysis determines the numerical amount or concentration.
High-performance liquid chromatography (HPLC), formerly referred to as high-pressure liquid chromatography, is a technique in analytical chemistry used to separate, identify, and quantify specific components in mixtures. The mixtures can originated from food, chemicals, pharmaceuticals, biological, environmental and agriculture, etc, which have been dissolved into liquid solutions. It relies on high pressure pumps, which deliver mixtures of various solvents, called the mobile phase, which flows through the system, collecting the sample mixture on the way, delivering it into a cylinder, called the column, filled with solid particles, made of adsorbent material, called the stationary phase. Each component in the sample interacts slightly different with the adsorbent material, causing different migration rate for the each component, leading to their separation, as they flow out of the column into a specific detector. The output of the detector is a graph, called Chromatogram. Chromatograms are graphical representations of the signal intensity versus time or volume, showing peaks, which represent components of the sample, each appears in its respective time, called Retention Time, having area proportional to its amount.
Mass spectrometry (MS) is an analytical technique that is used to measure the mass-to-charge ratio of ions. The results are presented as a mass spectrum, a plot of intensity as a function of the mass-to-charge ratio. Mass spectrometry is used in many different fields and is applied to pure samples as well as complex mixtures.
An ion source is a device that creates atomic and molecular ions. Ion sources are used to form ions for mass spectrometers, optical emission spectrometers, particle accelerators, ion implanters and ion engines.
Electron ionization is an ionization method in which energetic electrons interact with solid or gas phase atoms or molecules to produce ions. EI was one of the first ionization techniques developed for mass spectrometry. However, this method is still a popular ionization technique. This technique is considered a hard ionization method, since it uses highly energetic electrons to produce ions. This leads to extensive fragmentation, which can be helpful for structure determination of unknown compounds. EI is the most useful for organic compounds which have a molecular weight below 600. Also, several other thermally stable and volatile compounds in solid, liquid and gas states can be detected with the use of this technique when coupled with various separation methods.
Gas chromatography–mass spectrometry (GC–MS) is an analytical method that combines the features of gas-chromatography and mass spectrometry to identify different substances within a test sample. Applications of GC–MS include drug detection, fire investigation, environmental analysis, explosives investigation, food and flavor analysis, and identification of unknown samples, including that of material samples obtained from planet Mars during probe missions as early as the 1970s. GC–MS can also be used in airport security to detect substances in luggage or on human beings. Additionally, it can identify trace elements in materials that were previously thought to have disintegrated beyond identification. Like liquid chromatography–mass spectrometry, it allows analysis and detection even of tiny amounts of a substance.
Metabolomics is the scientific study of chemical processes involving metabolites, the small molecule substrates, intermediates, and products of cell metabolism. Specifically, metabolomics is the "systematic study of the unique chemical fingerprints that specific cellular processes leave behind", the study of their small-molecule metabolite profiles. The metabolome represents the complete set of metabolites in a biological cell, tissue, organ, or organism, which are the end products of cellular processes. Messenger RNA (mRNA), gene expression data, and proteomic analyses reveal the set of gene products being produced in the cell, data that represents one aspect of cellular function. Conversely, metabolic profiling can give an instantaneous snapshot of the physiology of that cell, and thus, metabolomics provides a direct "functional readout of the physiological state" of an organism. There are indeed quantifiable correlations between the metabolome and the other cellular ensembles, which can be used to predict metabolite abundances in biological samples from, for example mRNA abundances. One of the ultimate challenges of systems biology is to integrate metabolomics with all other -omics information to provide a better understanding of cellular biology.
![<span class="mw-page-title-main">Chemical ionization</span> Ionization technique used in mass [[spectroscopy]]](https://upload.wikimedia.org/wikipedia/commons/thumb/7/7b/Chemical_Ionization.png/320px-Chemical_Ionization.png)
Chemical ionization (CI) is a soft ionization technique used in mass spectrometry. This was first introduced by Burnaby Munson and Frank H. Field in 1966. This technique is a branch of gaseous ion-molecule chemistry. Reagent gas molecules are ionized by electron ionization to form reagent ions, which subsequently react with analyte molecules in the gas phase to create analyte ions for analysis by mass spectrometry. Negative chemical ionization (NCI), charge-exchange chemical ionization, atmospheric-pressure chemical ionization (APCI) and atmospheric pressure photoionization (APPI) are some of the common variants of the technique. CI mass spectrometry finds general application in the identification, structure elucidation and quantitation of organic compounds as well as some utility in biochemical analysis. Samples to be analyzed must be in vapour form, or else, must be vapourized before introduction into the source.
Ion chromatography is a form of chromatography that separates ions and ionizable polar molecules based on their affinity to the ion exchanger. It works on almost any kind of charged molecule—including small inorganic anions, large proteins, small nucleotides, and amino acids. However, ion chromatography must be done in conditions that are one unit away from the isoelectric point of a protein.
In mass spectrometry, matrix-assisted laser desorption/ionization (MALDI) is an ionization technique that uses a laser energy-absorbing matrix to create ions from large molecules with minimal fragmentation. It has been applied to the analysis of biomolecules and various organic molecules, which tend to be fragile and fragment when ionized by more conventional ionization methods. It is similar in character to electrospray ionization (ESI) in that both techniques are relatively soft ways of obtaining ions of large molecules in the gas phase, though MALDI typically produces far fewer multi-charged ions.
Liquid chromatography–mass spectrometry (LC–MS) is an analytical chemistry technique that combines the physical separation capabilities of liquid chromatography with the mass analysis capabilities of mass spectrometry (MS). Coupled chromatography - MS systems are popular in chemical analysis because the individual capabilities of each technique are enhanced synergistically. While liquid chromatography separates mixtures with multiple components, mass spectrometry provides spectral information that may help to identify each separated component. MS is not only sensitive, but provides selective detection, relieving the need for complete chromatographic separation. LC–MS is also appropriate for metabolomics because of its good coverage of a wide range of chemicals. This tandem technique can be used to analyze biochemical, organic, and inorganic compounds commonly found in complex samples of environmental and biological origin. Therefore, LC–MS may be applied in a wide range of sectors including biotechnology, environment monitoring, food processing, and pharmaceutical, agrochemical, and cosmetic industries. Since the early 2000s, LC–MS has also begun to be used in clinical applications.
Atmospheric pressure chemical ionization (APCI) is an ionization method used in mass spectrometry which utilizes gas-phase ion-molecule reactions at atmospheric pressure (105 Pa), commonly coupled with high-performance liquid chromatography (HPLC). APCI is a soft ionization method similar to chemical ionization where primary ions are produced on a solvent spray. The main usage of APCI is for polar and relatively less polar thermally stable compounds with molecular weight less than 1500 Da. The application of APCI with HPLC has gained a large popularity in trace analysis detection such as steroids, pesticides and also in pharmacology for drug metabolites.
In a chemical analysis, the internal standard method involves adding the same amount of a chemical substance to each sample and calibration solution. The internal standard responds proportionally to changes in the analyte and provides a similar, but not identical, measurement signal. It must also be absent from the sample matrix to ensure there is no other source of the internal standard present. Taking the ratio of analyte signal to internal standard signal and plotting it against the analyte concentrations in the calibration solutions will result in a calibration curve. The calibration curve can then be used to calculate the analyte concentration in an unknown sample.
In mass spectrometry, direct analysis in real time (DART) is an ion source that produces electronically or vibronically excited-state species from gases such as helium, argon, or nitrogen that ionize atmospheric molecules or dopant molecules. The ions generated from atmospheric or dopant molecules undergo ion-molecule reactions with the sample molecules to produce analyte ions. Analytes with low ionization energy may be ionized directly. The DART ionization process can produce positive or negative ions depending on the potential applied to the exit electrode.
Desorption electrospray ionization (DESI) is an ambient ionization technique that can be coupled to mass spectrometry (MS) for chemical analysis of samples at atmospheric conditions. Coupled ionization sources-MS systems are popular in chemical analysis because the individual capabilities of various sources combined with different MS systems allow for chemical determinations of samples. DESI employs a fast-moving charged solvent stream, at an angle relative to the sample surface, to extract analytes from the surfaces and propel the secondary ions toward the mass analyzer. This tandem technique can be used to analyze forensics analyses, pharmaceuticals, plant tissues, fruits, intact biological tissues, enzyme-substrate complexes, metabolites and polymers. Therefore, DESI-MS may be applied in a wide variety of sectors including food and drug administration, pharmaceuticals, environmental monitoring, and biotechnology.
Sample preparation for mass spectrometry is used for the optimization of a sample for analysis in a mass spectrometer (MS). Each ionization method has certain factors that must be considered for that method to be successful, such as volume, concentration, sample phase, and composition of the analyte solution. Quite possibly the most important consideration in sample preparation is knowing what phase the sample must be in for analysis to be successful. In some cases the analyte itself must be purified before entering the ion source. In other situations, the matrix, or everything in the solution surrounding the analyte, is the most important factor to consider and adjust. Often, sample preparation itself for mass spectrometry can be avoided by coupling mass spectrometry to a chromatography method, or some other form of separation before entering the mass spectrometer. In some cases, the analyte itself must be adjusted so that analysis is possible, such as in protein mass spectrometry, where usually the protein of interest is cleaved into peptides before analysis, either by in-gel digestion or by proteolysis in solution.
Capillary electrophoresis–mass spectrometry (CE–MS) is an analytical chemistry technique formed by the combination of the liquid separation process of capillary electrophoresis with mass spectrometry. CE–MS combines advantages of both CE and MS to provide high separation efficiency and molecular mass information in a single analysis. It has high resolving power and sensitivity, requires minimal volume and can analyze at high speed. Ions are typically formed by electrospray ionization, but they can also be formed by matrix-assisted laser desorption/ionization or other ionization techniques. It has applications in basic research in proteomics and quantitative analysis of biomolecules as well as in clinical medicine. Since its introduction in 1987, new developments and applications have made CE-MS a powerful separation and identification technique. Use of CE–MS has increased for protein and peptides analysis and other biomolecules. However, the development of online CE–MS is not without challenges. Understanding of CE, the interface setup, ionization technique and mass detection system is important to tackle problems while coupling capillary electrophoresis to mass spectrometry.
Ambient ionization is a form of ionization in which ions are formed in an ion source outside the mass spectrometer without sample preparation or separation. Ions can be formed by extraction into charged electrospray droplets, thermally desorbed and ionized by chemical ionization, or laser desorbed or ablated and post-ionized before they enter the mass spectrometer.
Bioanalysis is a sub-discipline of analytical chemistry covering the quantitative measurement of xenobiotics and biotics in biological systems.
Atmospheric pressure photoionization (APPI) is a soft ionization method used in mass spectrometry (MS) usually coupled to liquid chromatography (LC). Molecules are ionized using a vacuum ultraviolet (VUV) light source operating at atmospheric pressure, either by direct absorption followed by electron ejection or through ionization of a dopant molecule that leads to chemical ionization of target molecules. The sample is usually a solvent spray that is vaporized by nebulization and heat. The benefit of APPI is that it ionizes molecules across a broad range of polarity and is particularly useful for ionization of low polarity molecules for which other popular ionization methods such as electrospray ionization (ESI) and atmospheric pressure chemical ionization (APCI) are less suitable. It is also less prone to ion suppression and matrix effects compared to ESI and APCI and typically has a wide linear dynamic range. The application of APPI with LC/MS is commonly used for analysis of petroleum compounds, pesticides, steroids, and drug metabolites lacking polar functional groups and is being extensively deployed for ambient ionization particularly for explosives detection in security applications.
References
- ↑ Johnson RC, Cooks RG, Allen TM, Cisper ME, Hemberger PH (2000). "Membrane introduction mass spectrometry: trends and applications". Mass Spectrometry Reviews. 19 (1): 1–37. Bibcode:2000MSRv...19....1J. doi:10.1002/(SICI)1098-2787(2000)19:1<1::AID-MAS1>3.0.CO;2-Y. PMID 10715830.
- ↑ Demeestere K, Dewulf J, De Witte B, Van Langenhove H (2007). "Sample preparation for the analysis of volatile organic compounds in air and water matrices". Journal of Chromatography A. 1153 (1–2): 130–44. doi:10.1016/j.chroma.2007.01.012. PMID 17258752.
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