Applications of MALDI

MALDI is mainly used for the analysis of proteins and peptides and their mixtures. It is possible to determine the molecular weights, to obtain structural information 

The strong points of MALDl include the very low amount of sample necessary for analysis, a few fmol are sufficient. Unlike other ionisation methods, MALDl tolerates moderate concentrations of buffer and salts in the analyte mixture. Sample preparation is relatively easy and the spectra obtained are simple, so that even mixtures can be analysed without the need to separate the components prior to MALDl analysis. However, in contrary to ESI, MALDl cannot be directly coupled to liquid chromatography (LC) or capillary electrophoresis (CE) as it is not a continuous but a batch ionisation method. 

A typical MALDl spectrum is shown in Pig. 4.9. It is the spectrum of r-hirudin, a protein consisting of 64 amino acids. As little or no fragmentation occurs, only oriQ major peak, [M- -H]+, and two minor peaks, [M- -2H] + and the agglomerate [2M- -H]+ are observed. This peak-pattern is very typical of MALDI-TOF spectra. Depending on the salt concentrations in the buffers used for sample preparation, peaks like [M -f Na]+ and [M -f K]+ are also observed. 

The spectrum in Fig. 4.9 demonstrates that MALDI-TOF is a powerful method for accurate molecular weight determination of peptides and proteins. As there is almost no fragmentation, mixtures of peptides and proteins can be analysed without having to separate the compounds prior to analysis. In this respect, MALDI-TOF has to be regarded as a very fast separation method and is in many ways more powerful than chromatography or electrophoresis. In Fig. 4.10, a spectrum of low fat bovine milk is shown. The milk sample was added to the matrix without any pre-treatment and the different components present in the sample are resolved in the obtained mass spectrum. 

MALDI is more reliable for protein identification than other, commonly used methods, such as identification due to migration patterns in 2D gel electrophoresis 

MALDI applications are growing at a rapid pace with over 3000 publications annually since 2006 (Fig. 11.18). A single book chapter thus can impossibly cover all aspects of these developments. However, there are several excellent monographs on different aspects of MALDI which are highly recommended to anyone intending to pursue further studies of MALDI-MS [15,23,143-145]. The following section will merely outline some flagship applications of MALDI-MS. 

---

Two relatively new techniques, matrix assisted laser desorption ionization-lime of flight mass spectrometry (MALDI-TOF) and electrospray ionization (FS1), offer new possibilities for analysis of polymers with molecular weights in the tens of thousands. PS molecular weights as high as 1.5 million have been determined by MALDI-TOF. Recent reviews on the application of these techniques to synthetic polymers include those by Ilantoif54 and Nielen.555 The methods have been much used to provide evidence for initiation and termination mechanisms in various forms of living and controlled radical polymerization.550 Some examples of the application of MALDI-TOF and ESI in end group determination are provided in Table 3.12. The table is not intended to be a comprehensive survey. 

Table 3.12 Application of MALDI-TOF or ESI Mass Spectrometry to Polymers Prepared by Radical Polymerization...

Lasers have advanced the analytical use of mass spectrometers to characterise additives in polymers, and routine application of MALDI is no longer limited to high molecular masses only. MALDI can now clearly produce isotopically resolved mass spectra of small molecules (<800 Da) in an L-ToF instrument, which can be used successfully for the characterisation of molecules of different chemical classes. High mass resolution with an improvement of mass accuracy to... 

Most often proteins are the bacterial biopolymers studied using MALDI MS either from fractions or whole cells. They are not the only isolated cellular biopolymers studied by MALDI, nor the first. Very soon after the introduction of MALDI there were a few reports of the analysis of bacterial RNA or DNA from bacterial fractions. One of the first applications of MALDI to bacteria fractions involved analysis of RNA isolated from E. coli,4 Other studies included analysis of PCR-amplified DNA,5 6 DNA related to repair mechanisms7 and posttranscriptional modification of bacterial RNA.8 While most MALDI studies involve the use of UV lasers, IR MALDI has been reported for the analysis of double stranded DNA from restriction enzyme digested DNA plasmids, also isolated from E. coli.9... 

The investigations directed at the synthesis of thymine-substituted polymers demonstrate that the type of functional groups displayed by nucleic acid bases are compatible with ROMP. Moreover, the application of MALDI-TOF mass spectrometry to the analysis of these polymers adds to the battery of tools available for the characterization of ROMP and its products. The utility of this approach for the creation of molecules with the desired biological properties, however, is still undetermined. It is unknown whether these thymine-substituted polymers can hybridize with nucleic acids. Moreover, ROMP does not provide a simple solution to the controlled synthesis of materials that display specific sequences composed of all five common nucleic acid bases. Nevertheless, the demonstration that metathesis reactions can be conducted with such substrates suggests that perhaps neobiopolymers that function as nucleic acid analogs can be synthesized by such processes. 

Jackson, A.T. Yates, H.T. Scrivens, J.H. Critchley, G. Brown, J. Green, M.R. Bateman, R.H. The Application of MALDI Combined With Collision-Induced Dissociation to the Analysis of Synthetic Polymers. Rapid Commun. Mass Spectrom. 1996,10, 1668-1674. 

In 2006, Greis and co-workers reported on the application of MALDI-TOF MS as a tool for rapid inhibitor screening [10]. Different kinases (protein kinase C-a, cAMP-dependent protein kinase) in combination with their substrates were assayed, and the inhibitory potencies of staurosporine and three novel compounds were determined. For all four compounds, IC50 values could be determined, and... 

Kinsel, G. CAREER Analytical Applications of MALDI Mass Spectrometry to the Study of Surface-Protein Interactions, 1999 (NSF CHE 9876249). 

HPLC and the extensive complexity of ESI and MALDI spectra for multicomponent polymers with molar mass over about 10 g mol. Some applications of MALDI MS in polymer HPLC can be found for example in [300-303],... 

Bucknall, M., Fung, K. Y. C., and Duncan, M. W. (2002). Practical quantitative biomedical applications of MALDI-TOF mass spectrometry. J. Amer. Soc. Mass Spectrom. 13 1015-1027. 

Levery.193 Illustrated here are typical examples of the application of MALDI and ESI to glycolipid research. 

The successful application of MALDI imaging in clinical proteomics is now being extended to lipids and metabolites in order to make similar discoveries. Some first publications in lipid analysis indicate that similar information can be obtained, although the number of involved patients is still very low (45,70,71). 

---