Fixed-charge derivatization

SCHEME 4.2 Fixed-charge derivatization and selective gas-phase fragmentation of methionine fixed-charge sulfonium ion derivatized peptides. (Reproduced from Amunugama, M. Roberts, K.D. Reid, G.E., J. Am. Soc. Mass Spectrom. 2006,17,1631-1642. With permission from the American Society for Mass Spectrometry, published by Elsevier Inc.)... 

Chamot-Rooke, J. van der Rest, G. Dalleu, A. Bay, S. Lemoine, J. The combination of electron capture dissociation and fixed charge derivatization increases sequence coverage for O-glycosylated and O-phosphorylated peptides. J. Am. Soc. Mass Spectrom. 2007,18,1405-1413. 

ESI is best described as a mixed-mode ionization, where various processes contribute to the final result. The soft-desolvation and lEV models indicate the importance of generating preformed analyte ions in solution. For most analytes with basic functions, e.g., amines and amides, or acidic functions, e.g., carboxylic acid or aromatic phenols, preformed ions can be produced by the selection of an appropriate pH of the mobile phase. For basic compounds, acidic mobile-phase conditions ate selected, and basic conditions for acidic compounds. Analyte derivatization has been described to introduce a basic site or a fixed charge from a quaternary ammonium group in analytes that lack such properties. 

In order to enhance the sensitivity in steroid analysis, a number of derivatization procedures have been proposed. In most cases, the aim of the derivatization is an increase of the proton affinity of the compound, although the introduction of a fixed charge on a quaternary ammonium or a group with high electron affinity has been described as well. An overview of the derivatives generated is given in Figme 13.4. Most derivatization procedures are applied in combination with ESI-MS. 

The introduction of a quaternary ammonium cation to the hydroxyl moieties of DAG species, using A-chlorobetainyl chloride, was reported [12]. This derivatization not only affords an increased ionization efficiency of two orders compared to their underivatized sodium adducts but also makes the ionization efficiency independent of FA chains present in DAG species because of the existence of a fixed charge site after derivatization. The fragmentation pattern of A-chlorobetainyl-derivatized DAG species is essentially identical to that of PC species (see Chapter 7). 

Gel permeation chromatographic (GPC) analysis was not possible directly on the primary-amine-functionalized oligomers because of their tendency to be adsorbed on the GPC columns rather than to elute. A method has been developed in our laboratories (9, 10) for derivatization of the amine end groups with benzophenone to form an imine functionality. This method allows FPC (fixed-partial-charge) analysis of the oligomers. 

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