Matrix-assisted laser desorption ionisation MALDI

Fast atom bombardment (FAB) Plasma desorption (PD) Liquid secondary-ion mass spectrometry (LSIMS) Thermospray (TSP)/plasmaspray (PSP) Electrohydrodynamic ionisation (EHI) Multiphoton ionisation (MPI) Atmospheric pressure chemical ionisation (APCI) Electrospray ionisation (ESI) Ion spray (ISP) Matrix-assisted laser desorption/ionisation (MALDI) Atmospheric pressure photoionisation (APPI) Triple quadrupole (QQQ) Four sector (EBEB) Hybrid (EBQQ) Hybrid (EB-ToF, Q-ToF) Tandem ToF-ToF Photomultiplier... 

Matrix-assisted laser desorption ionisation (MALDI) MH+ (M - H) ToF, IT, FTMS Polar and some nonpolar biopolymers, synthetic polymers >250000... 

As evident from Scheme 7.13, most modern ionisation techniques have been used for TLC-MS, and no single ionisation method is used exclusively with TLC-MS. Various ionisation methods may be applied that avoid the need to evaporate the sample into an El or Cl source these are based in particular on sputtering (FAB, SIMS) or laser desorption. Several sputtering methods of ionisation do not require the use of a liquid matrix, e.g. TLC-SIMS [797], Recent developments include the use of matrix-assisted laser desorption ionisation (MALDI) and surface-assisted laser desorption ionisation (SALDI). It is obvious that TLC-MS is complemented with TLC-MS11 [800] and TLC-HRMS techniques. Table 7.82 lists the general characteristics of TLC-MS. 

For the quick characterisation of polydisperse surfactants with relative high molecular weight distributions matrix-assisted laser desorption/ionisation (MALDI)-time of flight (TOF)-MS represented an interesting alternative since low mass compounds did not interfere with the mass spectrometric detection of the compounds of interest. For example, the mass spectrum of C12-APG (Fig. 2.7.8) exhibited equally spaced signals with Am/z 162 corresponding to sodiated adduct ions of the mono- (m/z 371) to heptaglucosides (m/z 1343) [7]. 

Matrix assisted laser desorption ionisation (MALDI) and ESI-MS spectra of non-ionic surfactant blends of AE obtained after positive ionisation were compared [28]. Both the ionisation procedures, which produce [M + Na]+ ion clusters, were very useful for this purpose, but the ESI spectra generated were more complex, whereas MALDI ionisation led to simpler spectra that can be interpreted more easily [28]. 

For the characterisation of the biodegradation intermediates of C12-LAS, metabolised in pure culture by an a-proteobacterium, Cook and co-workers [23] used matrix-assisted laser desorption/ionisation (MALDI)-time of flight (TOF)-MS as a complementary tool to HPLC with diode array detection and 1H-nuclear magnetic resonance. The dominating signal in the spectrum at m/z 271 and 293 were assigned to the ions [M - H] and [M - 2H + Na]- of C6-SPC. Of minor intensity were the ions with m/z 285 and 299, interpreted to be the deprotonated molecular ions of C7- and C8-SPC, respectively. 

While fast atom bombardment (FAB) [66] and TSI [25] built up the basis for a substance-specific analysis of the low-volatile surfactants within the late 1980s and early 1990s, these techniques nowadays have been replaced successfully by the API methods [22], ESI and APCI, and matrix assisted laser desorption ionisation (MALDI). In the analyses of anionic surfactants, the negative ionisation mode can be applied in FIA-MS and LC-MS providing a more selective determination for these types of compounds than other analytical approaches. Application of positive ionisation to anionics of ethoxylate type compounds led to the abstraction of the anionic moiety in the molecule while the alkyl or alkylaryl ethoxylate moiety is ionised in the form of AE or APEO ions. Identification of most anionic surfactants by MS-MS was observed to be more complicated than the identification of non-ionic surfactants. Product ion spectra often suffer from a reduced number of negative product ions and, in addition, product ions that are observed are less characteristic than positively generated product ions of non-ionics. The most important obstacle in the identification and quantification of surfactants and their metabolites, however, is the lack of commercially available standards. The problems with identification will be aggravated by an absence of universally applicable product ion libraries. 

Matrix assisted laser desorption ionisation (MALDI)... 

Two techniques that have become preffered for ionisation of proteins/peptides is electrospray ionisation (ESI) and matrix-assisted laser desorption/ionisation (MALDI). Although different combinations of ionisation techniques and mass analyser exist, MALDI usually is coupled with a time-of-flight (TOF) (Figure 7) tube as a mass analyser while ESI is tradionally combined with quadrupole mass analysers. Instruments capable of MS/MS have the ability to select ions of particular m/z ratio from a mixture, to fragment selected ions and to record the precise masses of the resulting fragment ions. If this process is applied to the analysis of peptide ions, in principle the amino acid sequence of the peptide can be deduced. 

Mass spectrometry has assumed great importance in determinations of the molar masses of biological macromolecules, even quite large ones. This is due to developments such as electrospray ionisation (ESI) and matrix assisted laser desorption/ ionisation (MALDI), which have made it possible to determine the molar masses of biopolymers up to several 100 kDa (Pitt 1996 Kellner et al. 1999 Snyder 2000). The combination of MALDI techniques with time-of-flight mass spectrometers (MALDI-TOF) is of particular significance for determination of the molar masses of proteins with high sensitivity (typically pmol quantities, although exceptionally fmol) and precision (proteins up to 100 kDa with precision of about 0.01 %). Mass spectrometry can provide very accurate measurements of protein molar mass that can yield information about even minor structural modifications not readily accessible by other means. 

The classical area of application of mass spectrometry has been with small volatile compounds, although non-volatile samples could be analysed if they were suitably derivatised. The application of mass spectrometry to large complex molecules like proteins has been made possible by the development of novel ionisation techniques which enable large molecules (> 200 kDa) to be introduced into the mass spectrometer in an intact form suitable for analysis (Siuzdak 1996 Dass 2000). Of the various techniques that have been developed, electrospray ionisation (ESI) and matrix-assisted laser desorption ionisation (MALDI) are the ones best suited for use with macromolecules and macromolecular complexes. 

Kaufmann, R., Kirsch, D. and Spengler, B. (1994) Sequencing of peptides in a time-of-flight mass spectrometer evaluation of postsource decay following matrix-assisted laser desorption ionisation (MALDI). Int. J. Mass Spectrom. Ion Process. 131, 355-385. 

Matrix assisted laser desorption ionisation (MALDI) is an important technique for characterisation and fingerprinting of large molecules. However, solvents used at sample preparation have a negative effect on the quahty of the fingerprint. Work is therefore performed on development of solvent free sample preparation methods for characterisation of synthetic polymers [61]. 

Three soft ionisation methods are in use for earbohydrates, fast atom bombardment (FAB), eleetrospray ionisation (ESI) and matrix-assisted laser desorption/ionisation (MALDI). FAB is the oldest and involves directing a high-energy beam of Cs" ions or Xe atoms at the sample dissolved in a nonvolatile solvent such as m-nitrobenzyl alcohol. The atoms sputter the sample and matrix [M + H] or [M + Na]" ions are commonly observed. With an upper limit of M of about 2000, FAB is not that soft, and is usually used for small oligosaccharides it has the further disadvantage that the sample is prepared and then directly introduced into the mass spectrometer, so that it cannot be combined with liquid chromatography. 

Matrix-assisted laser desorption ionisation (MALDI) can analyse molecules with large mass >10 000 Da. The technique is similar to FAB and is also considered a soft ionisation process. The analyte is mixed with a matrix such as sinapinic acid or 2,5-dihydroxybenzoic acid that strongly absorbs UV radiation. A few microlitres of this analyte/matrix mixture is deposited on the sample plate and dried. The plate is inserted into the source region... 

Figure 9.5 Schematic of Matrix Assisted Laser Desorption Ionisation (MALDI). The laser is fired at the sample analyte of interest admixed with a crystalline matrix that readily absorbs laser energy and allows the sputtering of analyte into the vapour phase in association with matrix molecules of solvation. Desolvation and proton transfer leads to naked molecular ion species, ready for mass analysis.

Alternatively, the protein can be partially digested by an enzyme like trypsin. The fragments of this tryptic digest can then be separated and their molecular weights can be measured with a mass spectrometer. Methods available for such analysis are matrix assisted laser desorption ionisation (MALDI) time of flight (TOE)... 

The sample molecules can be ionised by one of several techniques. In electron impact ionisation, (El), electrons are fired at the sample molecules, whereas in chemical ionisation, (Cl), the sample molecules are collided with a reactive gas. The sample can also be bombarded with argon atoms (fast atom bombardment, FAB) or the dissolved sample can be sprayed into an electric field electrospray ionisation, ESI). Furthermore, the sample can be co-crystallised with a matrix and then ions can be generated by exposure to photons (matrix assisted laser desorption ionisation, MALDI). 

Koichi Tanaka presented experiments for soft laser desorption ionisation (SLD) of proteins in 1987. However, the predominant and most widely used version of SLD, matrix assisted laser desorption ionisation, MALDI, was introduced shortly afterwards by Michael Karas and Eranz Hillenkamp. Tanaka was awarded the Nobel Prize for his cornerstone invention in 2002. Prior to that, no method was available to transfer large biomolecules with molecular weights of more than... 

Matrix assisted laser desorption/ionisation (MALDI) For laser desorption methods a pulsed laser is used to desorb species from a target surface. Therefore, a mass analyser compatible with pulsed ionisation methods has to be used. Typically, time-offlight (TOF) analysers are employed, but several hybrid systems (Q-TOF) and, recently, high resolution Fourier transform ion cyclotron resonance (FT-ICR) analysers have been successfully adapted (see Section 10.2.4). Direct laser desorption rehes on the very rapid heating of the sample or sample substrate to vapourise molecules without decomposition. The more recent development of MALDI relies on the absorption of laser energy by a solid, microcrystalline matrix compound such as a-cyano-4-hydroxy ciimamic acid or sinapinic acid [8, 34]. MALDI has become an extremely popular method for the rapid and sensitive analysis of high-molecular-weight compounds [4]. 

Giannakopulos, A. E., Bahir, S., and Derrick, P. J., Comment Reproducibility of Spectra and Threshold Fluence in Matrix-assisted Laser Desorption/Ionisation (MALDI) of Polymers, Eur. Mass, Spectrom., 4, 127, 1998. 

Li, C.-Z., Herod, A.A., John, R, Johnson, C.A.F., Parker, J.E., Smith, G.P, Humphrey, R, Chapman, J.R., Rahman, M.,Kinghorn,R.R.F.,Kandiyoti,R. (1994) Characterisation of kerogens by matrix assisted laser desorption ionisation (MALDI) mass spectroscopy. Rapid Commun. Mass Spectrom., 8,823-828. 

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