The results of previous works that have claimed to detect cyclodextrin inclusion complexes via the "soft" ionization technique of electrospray ionization mass spectrometry are revisited. A more extensive study of cyclodextrin mixtures with amino acids and small peptides demonstrates that amino acid and peptide "complexes" are detected by electrospray mass spectrometry regardless of the presence (or not) of an aromatic moiety on the side chain. Amino acids that may be least likely to form hydrophobic inclusion complexes with cyclodextrin in solution generally show the most intense complex ions. The data suggest that these "complexes" are, in all likelihood, electrostatic adducts formed during the electrospray process. Systematic controls are suggested to ensure that "false positives" do not negate many of the claims concerning the detection of solution-derived noncovalent compounds.
A method has been developed that takes advantage of the formation of noncovalent compounds in electrospray mass spectrometry. Mixtures of proteins and peptides are shown to produce an intense ion that corresponds to a 1:1 complex with a crown ether (18-crown-6). Although the crown ether may be added directly to the solution, for the current experiments it is introduced via the methanol liquid sheath. The spacing of these complexed species in the mass spectrum allows unambiguous determination of the charge state of the ions and their actual mass. Through constant neutral loss scans, charge state may be determined, mass assigned, spectra simplified, and chemical noise may be reduced for the analysis of complex peptide samples without Chromatographic separation. Finally, the prevalence of single complexation permits mass assignments based on the mass difference of a single protein ion and its complexed form at any charge state. In essence, the method performs a separation based on charge state. It can be used to complement Chromatographic separation and deconvolution algorithms for the electrospray mass spectrometry analysis of peptide-protein mixtures.
The introduction of 18-crown-6 into the liquid sheath during the electrospray ionization mass spectrometry (ESI-MS) of peptides leads to the formation of crown ethedpeptide complexes. The nominal mass spacing of the peaks from these complexes allows an unambiguous determination of the charge state of the ions and thus of their actual mass. The 264 Da mass shift is much larger than what is achieved by either Na+ or Cu2+ adduction, and is thus potentially more useful for the determination of charge states, especially for multiply charged highmolecular-weight species. The addition of an uncharged crown ether does not appear to interfere with the ESI process. Because the crown ether is added to the liquid sheath and not to the original solution, the method should be particularly amenable to chromatographic or electrophoretic techniques because it is essentially a postcolumn noncovalent derivatization procedure. Finally, because the complex formation constants are relatively large, the peptide/crown ether complex yields high intensity ions, normally the base peak in the spectrum when analyzing pure peptide samples. Finally, as the crown ether is neutral and noncovalently complexed, a constantneutral-loss scan of their collision-induced dissociation spectra yields a simplified spectrum that is free of chemical noise.With the advent of electrospray ionization' mass spectrometry (ESI-MS), there has come a revolution in the manner in which polar aqueous samples are analyzed and this is particularly true for the analysis of biologically important peptides and proteins. Two basic strengths of the ESI technique lie in its ability to detect large polar molecules within a relatively small m/z charge range via multi-charging and in its ability to couple the ionization process with existing separation methods. A drawback to the technique lies in the potential difficulty in determining the charge states (and thus the actual masses) of the observed ions.Recently, methods have been developed that have had some success in determining charge states and the actual mass of electrosprayed peptides. Neubauer and Anderegg used the addition of sodium acetate to the mobile phase during an experiment in which a liquid chromatograph (LC) was coupled to ESI-MS,' whereas Senko et al. utilized the addition of Cu2+ ions3 Both techniques determine the charge state of ions by observation of the mass shift from the metal catiodpeptide adduct. For example, a single Na+ adduct will show a mass shift of 23 Da, indicating a charge of l + . A mass shift of 11.5 Da indicates a charge state of 2+, etc. A major advantage of Cu'+ over Na' adduction lies in its larger mass, which provides for a greater mass shift than for sodiated peaks. A larger nominal mass shift is especially important when multiple charges effectively reduce the observed mass shift. If the number of charges is sufficiently large, then the observed mass shift could fall below the mass resolution capability of the mass spectrometer.In our current study, we have investigated the adduction of...
The liquid secondary ionization mass spectra of crown ether solutions and crown ether solutions containing alkali metal cations were generated. Cesium cations acted as both the primary ion beam and as a competing gas-selvedge-phase reactant. The data suggest that crown ether complexes formed in the condensed phase survive intact the fast ion bombarding event and the transition into the gas phase. The data further suggest that crown ether complexes formed in the condensed phase predominate in the ion spectrum over the corresponding complexes formed in the selvedge.
Static liquid secondary ion mass spectra were generated for a variety of dipeptides dissolved in common liquid secondary ion mass spectrometric matrices. The relationship of the ion intensities of the monomer and non-covalent dimers was examined in order to determine the origin of non-covalent dimerization. The results demonstrate that the ion intensities are independent of solution equilibria and are indicative of first-order gas-phase kinetics.
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