A mass chromatogram is a representation of mass spectrometry data as a chromatogram, where the x-axis represents time and the y-axis represents signal intensity. [1] The source data contains mass information; however, it is not graphically represented in a mass chromatogram in favor of visualizing signal intensity versus time. The most common use of this data representation is when mass spectrometry is used in conjunction with some form of chromatography, such as in liquid chromatography–mass spectrometry or gas chromatography–mass spectrometry. In this case, the x-axis represents retention time, analogous to any other chromatogram. The y-axis represents signal intensity or relative signal intensity. There are many different types of metrics that this intensity may represent, depending on what information is extracted from each mass spectrum.
The total ion current (TIC) chromatogram represents the summed intensity across the entire range of masses being detected at every point in the analysis. [2] [3] The range is typically several hundred mass-to-charge units or more. In complex samples, the TIC chromatogram often provides limited information as multiple analytes elute simultaneously, obscuring individual species.
The base peak chromatogram is similar to the TIC chromatogram, however it monitors only the most intense peak in each spectrum. [4] [3] This means that the base peak chromatogram represents the intensity of the most intense peak at every point in the analysis. Base peak chromatograms often have a cleaner look and thus are more informative than TIC chromatograms because the background is reduced by focusing on a single analyte at every point.
In an extracted-ion chromatogram (EIC or XIC), also called a reconstructed-ion chromatogram (RIC), one or more m/z values representing one or more analytes of interest are recovered ('extracted') from the entire data set for a chromatographic run. [3] The total intensity or base peak intensity within a mass tolerance window around a particular analyte's mass-to-charge ratio is plotted at every point in the analysis. The size of the mass tolerance window typically depends on the mass accuracy and mass resolution of the instrument collecting the data. This is useful for re-examining data to detect previously-unsuspected analytes, to highlight potential isomers, resolve suspected co-eluting substances, or to provide clean chromatograms of compounds of interest. An extracted-ion chromatogram is generated by separating the ions of interest from a data file containing the full mass spectrum over time after the fact; this is different from selected-ion chromatograms, discussed below, in which data is collected only for specific m/z values. A closely related term is extracted-compound chromatogram (ECC).
A selected-ion monitoring (SIM) chromatogram is similar to an EIC/XIC, with the exception that the mass spectrometer is operated in SIM mode, such that only preselected m/z values are detected in the analysis. [5] SIM experiments can be performed using mass spectrometry (MS) or tandem mass spectrometry (MS/MS) instruments. They are more common on MS instruments. This differs significantly from the extracted-ion chromatogram mentioned above in that only data for the ion(s) of interest are collected in a SIM experiment; for extracted-ion chromatograms (EIC or XIC), data for an entire mass range are collected during the run and then examined for analytes of interest after the completion of the run.
The selected-reaction monitoring (SRM) experiment is very similar to the SIM experiment except that tandem mass spectrometry is used and a specific product ion of a specific parent ion is detected. [3] The mass of the parent analyte is first selected while other ions are filtered away. The parent analyte ion is then fragmented in the gas phase and a specific fragment ion is monitored. This experiment has very high specificity because the SRM chromatogram represents only ions of a particular mass that fragment in a manner that produce a very specific product mass. This type of experiment can only be performed using tandem mass spectrometry. The technology progress in the MS/MS area lead to the development of MRM, Multiple Reaction Monitoring, which allows simultaneous detection of several coeluting analytes with different parent and/or product ions.
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 each component in a mixture. It relies on pumps to pass a pressurized liquid solvent containing the sample mixture through a column filled with a solid adsorbent material. Each component in the sample interacts slightly differently with the adsorbent material, causing different flow rates for the different components and leading to the separation of the components as they flow out of the column.
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.
A mass spectrum is a histogram plot of intensity vs. mass-to-charge ratio (m/z) in a chemical sample, usually acquired using an instrument called a mass spectrometer. Not all mass spectra of a given substance are the same; for example, some mass spectrometers break the analyte molecules into fragments; others observe the intact molecular masses with little fragmentation. A mass spectrum can represent many different types of information based on the type of mass spectrometer and the specific experiment applied. Common fragmentation processes for organic molecules are the McLafferty rearrangement and alpha cleavage. Straight chain alkanes and alkyl groups produce a typical series of peaks: 29 (CH3CH2+), 43 (CH3CH2CH2+), 57 (CH3CH2CH2CH2+), 71 (CH3CH2CH2CH2CH2+) etc.
Gas chromatography (GC) is a common type of chromatography used in analytical chemistry for separating and analyzing compounds that can be vaporized without decomposition. Typical uses of GC include testing the purity of a particular substance, or separating the different components of a mixture. In preparative chromatography, GC can be used to prepare pure compounds from a mixture.
Tandem mass spectrometry, also known as MS/MS or MS2, is a technique in instrumental analysis where two or more mass analyzers are coupled together using an additional reaction step to increase their abilities to analyse chemical samples. A common use of tandem MS is the analysis of biomolecules, such as proteins and peptides.
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, 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.
![<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.
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.
XIC can stand for:
Mass spectrometry imaging (MSI) is a technique used in mass spectrometry to visualize the spatial distribution of molecules, as biomarkers, metabolites, peptides or proteins by their molecular masses. After collecting a mass spectrum at one spot, the sample is moved to reach another region, and so on, until the entire sample is scanned. By choosing a peak in the resulting spectra that corresponds to the compound of interest, the MS data is used to map its distribution across the sample. This results in pictures of the spatially resolved distribution of a compound pixel by pixel. Each data set contains a veritable gallery of pictures because any peak in each spectrum can be spatially mapped. Despite the fact that MSI has been generally considered a qualitative method, the signal generated by this technique is proportional to the relative abundance of the analyte. Therefore, quantification is possible, when its challenges are overcome. Although widely used traditional methodologies like radiochemistry and immunohistochemistry achieve the same goal as MSI, they are limited in their abilities to analyze multiple samples at once, and can prove to be lacking if researchers do not have prior knowledge of the samples being studied. Most common ionization technologies in the field of MSI are DESI imaging, MALDI imaging and secondary ion mass spectrometry imaging.
Quantitative proteomics is an analytical chemistry technique for determining the amount of proteins in a sample. The methods for protein identification are identical to those used in general proteomics, but include quantification as an additional dimension. Rather than just providing lists of proteins identified in a certain sample, quantitative proteomics yields information about the physiological differences between two biological samples. For example, this approach can be used to compare samples from healthy and diseased patients. Quantitative proteomics is mainly performed by two-dimensional gel electrophoresis (2-DE) or mass spectrometry (MS). However, a recent developed method of quantitative dot blot (QDB) analysis is able to measure both the absolute and relative quantity of an individual proteins in the sample in high throughput format, thus open a new direction for proteomic research. In contrast to 2-DE, which requires MS for the downstream protein identification, MS technology can identify and quantify the changes.
Two-dimensional chromatography is a type of chromatographic technique in which the injected sample is separated by passing through two different separation stages. Two different chromatographic columns are connected in sequence, and the effluent from the first system is transferred onto the second column. Typically the second column has a different separation mechanism, so that bands that are poorly resolved from the first column may be completely separated in the second column. Alternately, the two columns might run at different temperatures. During the second stage of separation the rate at which the separation occurs must be faster than the first stage, since there is still only a single detector. The plane surface is amenable to sequential development in two directions using two different solvents.
In mass spectrometry, resolution is a measure of the ability to distinguish two peaks of slightly different mass-to-charge ratios ΔM, in a mass spectrum.
Electron capture ionization is the ionization of a gas phase atom or molecule by attachment of an electron to create an ion of the form . The reaction is
Selected ion monitoring (SIM) is a mass spectrometry scanning mode in which only a limited mass-to-charge ratio range is transmitted/detected by the instrument, as opposed to the full spectrum range. This mode of operation typically results in significantly increased sensitivity. Due to their inherent nature, this technique is most effective—and therefore most common—on quadrupole mass spectrometers and Fourier transform ion cyclotron resonance mass spectrometers.
A direct electron ionization liquid chromatography–mass spectrometry interface is a technique for coupling liquid chromatography and mass spectrometry (LC-MS) based on the direct introduction of the liquid effluent into an electron ionization (EI) source. Library searchable mass spectra are generated. Gas-phase EI has many applications for the detection of HPLC amenable compounds showing minimal adverse matrix effects. The direct-EI LC-MS interface provides access to well-characterized electron ionization data for a variety of LC applications and readily interpretable spectra from electronic libraries for environmental, food safety, pharmaceutical, biomedical, and other applications.
The component detection algorithm (CODA) is a name for a type of LC-MS and chemometrics software algorithm focused on detecting peaks in noisy chromatograms (TIC) often obtained using the electrospray ionization technique.
In chromatography, resolution is a measure of the separation of two peaks of different retention time t in a chromatogram.