Information from Mass Spectra

The DENDRAL project initiated in 1964 at Stanford was the prototypical application of artificial intelligence techniques - or what was understood at that time under this name - to chemical problems. Chemical structure generators were developed and information from mass spectra was used to prune the chemical graphs in order to derive the chemical structure associated with a certain mass spectrum. 

When extracting sequence information from mass spectra, not only is the m/z value at which the ions occur of importance since these provide an indication of the amino acid composition of the peptide giving rise to the ion, but so is the mass difference between adjacent ions. This indicates the particular amino acid residue that has been lost and thus provides the sequence information required. The mass differences arising from each of the amino acids are shown in Table 5.6. 

Upon extrapolation to larger ketones one can expect to observe larger acylium and alkyl fragments. The occurrence of series of homologous ions is a feature that can be very helpful to deduce structural information from mass spectra. Ions such as the acylium ion series and the carbenium ion series are also known as characteristic ions. Learning the nominal masses of the first members of each series by heart is useful (Tables 6.2 and 6.3). 

Deduce structural information from mass spectra... 

This section concerns the use of non-heuristic programs to deduce structural information from mass spectra. Automated systems were proposed as early as 1966 for compound-t5q)e identification 24,25) nd as an aid in the inter-... 

Factor analysis has been used for extracting information from mass spectra recorded in a TG-MS analysis of the styrene-isoprene mixture. Statheropoulos and co-workers... 

The signal intensity of spectral features contains quantitative information but it is highly sensitive to changes in experimental conditions. Therefore, special precautions need to be taken to extract quantitative information from mass spectra. Discussion of quantitative data treatment is also included in Chapter 8. In this section, we will focus on the basic features of mass spectra that carry information on ion abundances. 

Extracting structural Information from mass spectra - tools and rules Interpreting mass spectra - systematic approaches... 

The kind of information available from mass spectrometry falls into two categories. First, the m/z value for the molecular ion provides information useful in calculating the molecular formula of the molecule. Second, the lower molecular weight fragments that appear in the mass spectrum contain clues concerning structural features of the molecule in question. Be sure that you understand how to extract these kinds of information from mass spectral data. 

Figure 14 Additive analysis results. The chromatogram is an overlay of the MS and PDA responses. Each response curve contains extraordinary amounts of information. The mass spectrum, upper left, and a spectral measure of peak purity, upper right, have been extracted from the chromatogram. — TMD total ion chromatogram from 100 to 700 amu.. .. PDA extracted chromatogram at 230 nm. 

In contrast to IR and NMR spectroscopy, the principle of mass spectrometry (MS) is based on decomposition and reactions of organic molecules on theii way from the ion source to the detector. Consequently, structure-MS correlation is basically a matter of relating reactions to the signals in a mass spectrum. The chemical structure information contained in mass spectra is difficult to extract because of the complicated relationships between MS data and chemical structures. The aim of spectra evaluation can be either the identification of a compound or the interpretation of spectral data in order to elucidate the chemical structure [78-80],... 

Nd in samples. Unfortunately, mass spectrometry is not a selective technique. A mass spectrum provides information about the abundance of ions with a given mass. It cannot distinguish, however, between different ions with the same mass. Consequently, the choice of TIMS required developing a procedure for separating the tracer from the aerosol particulates. 

In GC-MS effluent from the column is introduced directly into the mass spectrometer s ionization chamber in a manner that eliminates the majority of the carrier gas. In the ionization chamber all molecules (remaining carrier gas, solvent, and solutes) are ionized, and the ions are separated by their mass-to-charge ratio. Because each solute undergoes a characteristic fragmentation into smaller ions, its mass spectrum of ion intensity as a function of mass-to-charge ratio provides qualitative information that can be used to identify the solute. 

When a mass spectrum has been acquired by the spectrometer/computer system, it is already in digital form as m/z values versus peak heights (ion abundances), and it is a simple matter for the computer to compare each spectrum in the library with that of the unknown until it finds a match. The shortened search is carried out first, and the computer reports the best fits or matches between the unknown and spectra in the library. A search of even 60,000 to 70,000 spectra takes only a few seconds, particularly if transputers are used, thus saving the operator a great deal of time. Even a partial match can be valuable because, although the required structure may not have been found in the library, it is more than likely that some of the library compounds will have stractural pieces that can be recognized from a partial fit and so provide information on at least part of the structure of the unknown. 

However, interpretation of, or even obtaining, the mass spectrum of a peptide can be difficult, and many techniques have been introduced to overcome such difficulties. These techniques include modifying the side chains in the peptide and protecting the N- and C-terminals by special groups. Despite many advances made by these approaches, it is not always easy to read the sequence from the mass spectrum because some amide bond cleavages are less easy than others and give little information. To overcome this problem, tandem mass spectrometry has been applied to this dry approach to peptide sequencing with considerable success. Further, electrospray ionization has been used to determine the molecular masses of proteins and peptides with unprecedented accuracy. 

Special isotope ratio mass spectrometers are needed to measure the small variations, which are too small to be read off from a spectrum obtained on a routine mass spectrometer. Ratios of isotopes measured very accurately (usually as 0/00, i.e., as parts per 1000 [mil] rather than parts per 100 [percent]) give information on, for example, reaction mechanisms, dating of historic samples, or testing for drugs in metabolic systems. Such uses are illustrated in the main text. 

What kinds of information can we get from a mass spectrum Certainly the most obvious information is the molecular weight, which in itself can be invaluable. For example, if we were given samples of hexane (MW = 86), 1-hexene (MW = 84), and 1-hexyne (MW = 82), mass spectrometry would easily distinguish them. 

Some detectors can give additional information about the elutes (the eluted solutes). One example is the gas chromatograph—mass spectrometer (GC-MS), which produces a mass spectrum of each compound as well as its mass and location in the chromatogram. This powerful means of detection can be used when standard samples are not available to help determine the identities of the solutes. A beam of ions bombards each compound as it emerges from the chromatograph. The compound breaks up into ions of different masses, providing a spread of narrow peaks instead of one peak for each compound. The relative amount of each fragment is determined and used to help identify the compound. 

The mass spectrum produced should provide unambiguous molecular weight information from the wide range of compounds amenable to analysis by HPLC, including biomolecules with molecular weights in excess of 1000 Da. The study of these types of molecule by mass spectrometry may be subject to limitations associated with their ionization and detection and the mass range of the instrument being used. 

A more definitive identification may be obtained by combining retention characteristics with more specific information from an appropriate detector. Arguably, the most information-rich HPLC detectors for the general identification problem are the diode-array UV detector, which allows a complete UV spectrum of an analyte to be obtained as it elutes from a column, and the mass spectrometer. The UV spectrum often allows the class of componnd to be determined but the... 

Very rarely, however, will a single mass spectrum provide us with complete analytical information for a sample, particularly if mass spectral data from a chromatographic separation, taking perhaps up to an hour, is being acquired. The mass spectrometer is therefore set up to scan, repetitively, over a selected m jz range for an appropriate period of time. At the end of each scan, the mass spectrum obtained is stored for subsequent manipulation before a further spectrum is acquired. 

The mass spectrum produced from an analyte, in terms of the m/z range of the ions observed and their relative intensities, depends upon a number of factors and spectra obtained using different experimental conditions may therefore differ considerably in appearance. This may or may not have implications on the analytical investigation being nndertaken, although the molecular weight information that may be extracted from these spectra, however, is independent of these differences. 

MS-MS is a term that covers a number of techniques in which two stages of mass spectrometry are used to investigate the relationship between ions found in a mass spectrum. In particular, the product-ion scan is used to derive structural information from a molecular ion generated by a soft ionization technique such as electrospray and, as such, is an alternative to CVF. The advantage of the product-ion scan over CVF is that it allows a specific ion to be selected and its fragmentation to be studied in isolation, while CVF bring about the fragmentation of all species in the ion source and this may hinder interpretation of the data obtained. 

The general procednre is to nse reconstrncted ion chromatograms at appropriate m/z values in an attempt to locate componnds of interest and then look at the mass spectrum of the unknown to determine its molecnlar weight. MS-MS can then be employed to obtain spectra from this and related compounds to find ions that are common to both and which may therefore contain common stmctmal features. Having the same m/z value does not necessarily mean the ions are identical and further MS-MS data or the elemental composition may be required. If these data do not allow unequivocal structure identification, then further MS" information may be required. 

The central engine of this data workflow is the process of spectral deconvolution. During spectral deconvolution, sets of multiply charged ions associated with particular proteins are reduced to a simplified spectrum representing the neutral mass forms of those proteins. Our laboratory makes use of a maximum entropy-based approach to spectral deconvolution (Ferrige et al., 1992a and b) that attempts to identify the most likely distribution of neutral masses that accounts for all data within the m/z mass spectrum. With this approach, quantitative peak intensity information is retained from the source spectrum, and meaningful intensity differences can be obtained by comparison of LC/MS runs acquired and processed under similar conditions. 

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