Post-source decay time-of-flight study of fragmentation mechanisms of protonated synthetic polymers under matrix-assisted laser desorption/ionization conditions

Post-source decay (PSD) of three different nylon oligomers desorbed under matrix-assisted laser desorption/ionization (MALDI) conditions was studied and their fragmentation pathways were investigated. The fragmentation of the protonated oligomers is very similar to that of peptides. The b(n)(+), y(n)(+) and z(n)(+) series of ions were observed in abundance in the PSD spectrum. The end groups and the length of the spacer in the repeating unit influence the fragmentation of the different polyamides and the relative abundances of the product ions. Competitive dehydration and deamination reactions were observed, and depend on the nature of the end groups and the repeating units. The PSD spectra are very similar to collision-induced dissociation (CID) spectra obtained under low-energy conditions, implying that the selected precursor ions possess similar average internal energies. All the peaks observed in the PSD spectrum can be rationalized by reasonable fragmentation mechanisms.     

Simultaneous detection of N-terminal fragment ions in a protein mixture using a ruthenium(II) complex

Use of a bis(terpyridine)ruthenium(II) derivative as an N-terminal labeling reagent resulted in the simultaneous detection and individual determination of all the N-terminal fragments of the proteins in a mixture without requiring any separation. All of the N-termini of the guanidinated proteins were labeled selectively by the ruthenium complex (-CO-labeling). After chymotryptic digestion, the fragments were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and post-source decay (PSD). The -CO moiety exclusively enhanced N-terminal fragment ions in mass spectra and enabled easy N-terminal sequencing. In a mixture containing three different proteins (lysozyme, ubiquitin, and insulin), all of the N-terminal fragment ions labeled with the ruthenium complex were found to produce uniformly intense peaks without the detection of the other unlabeled fragments. The N-terminal sequences of these ions were determined individually by PSD analysis. Application to unknown proteins from Thermus thermophilus HB8 with two-dimensional electrophoretic separation resulted in the successful determination of the N-terminal sequence and easy identification of the target protein.     

Derivatisation of peptides with osmium tetroxide, 2,2′-bipyridine: capillary electrophoretic and MALDI-TOF mass spectrometric study

Site-specific chemical modification is a useful technology in characterisation of proteins, but the number of chemical probes of the protein structure reacting with proteins under mild conditions in aqueous solutions is rather limited. Here we studied the reaction of osmium tetroxide, 2,2′-bipyridine (Os,bipy) with several peptides using capillary zone electrophoresis (CZE) and matrix-assisted laser desorption-ionisation-time-of-flight mass spectrometry (MALDI-TOF MS). Both techniques showed formation of a stable complex of Os,bipy with tryptophan residues. In CZE peaks with different migration times and UV-Vis spectra were observed. MALDI-TOF MS showed the formation of a product with characteristic isotopic pattern corresponding to the presence of osmium atom. Oxidation of cysteine and methionine side chains to cysteic acid and methionine sulfone by Os,bipy was detected by CZE and confirmed by MALDI-TOF and post-source decay (PSD) mass spectra. PSD showed specific shifts of molecular weights of the peptides and their fragments after the derivatisation. We believe that Os,bipy may become a useful agent in the characterisation of proteins.     

Improved PSD and CID on a MALDI TOFMS

The influence of several instrument-operating parameters on the product-ion resolution and mass accuracy in matrix-assisted laser desorption/ionization (MALDI) time-of-flight (TOF) post-source decay (PSD) and collision-induced dissociation (CID) experiments is reported. Voltages commonly applied to the reflectron for PSD and CID experiments were found to be non-ideal; optimization of these voltages resulted in better resolution across each segment of the measured PSD spectrum. Mass resolution, calculated as M/DeltaM (FWHM) for the product-ion peaks, was as high as 2500. Additionally, precursor-ion selection and segment mass range setup were each found to have dramatic influences on product-ion mass accuracy. An understanding of the influence of these variables aided in the interpretation of (a-NH3) and (b - NH3) ions observed in the PSD/CID spectra of a number of peptides. In addition, product ions resulting from coincidence peaks in the precursor-ion selection window were found to be a general problem. With the improvements to resolution and optimization of these mass accuracy variables, the mass accuracy of product ions from MALDI TOF PSD and CID experiments was tested with several reference materials, including the peptides Substance P, bradykinin, angiotensin I, and angiotensin II and the synthetic polymers poly(methyl methacrylate) and polystyrene. The absolute error (Da) for each test material was, on average, below 0.1 Da, demonstrating a significant improvement in mass accuracy using the improved operational parameters and an extension of the use of poly(ethylene glycol) (PEG) as a mass calibrant for the PSD/CID spectra.     

Identification of non-peptide species in selenized yeast by MALDI mass spectrometry using post-source decay and orthogonal Q-TOF detection

The potential of tandem mass spectrometry following matrix-assisted laser desorption ionization (MALDI) was studied for speciation of selenium. Non-peptide selenium-containing compounds were isolated from a selenized yeast aqueous extract by size-exclusion chromatography. Post-source decay (PSD) was compared with orthogonal quadrupole collision cell dissociation for the purpose of obtaining fragmentation and structural information. In the PSD mode, the use of ion gate covering the whole isotopic cluster of the parent compound allowed the immediate recognition of fragments containing Se and those in which this element was absent. The tandem mass spectra obtained by orthogonal MALDI Q-TOF were equally informative in terms of the number of fragments but suffered from a poorer sensitivity. The mass accuracy was ca. 20 times better in the oMALDI configuration than in the PSD mode. An unknown selenium compound with an m/z 388 was detected with a mass accuracy of 3 ppm according to the proposed empiric formula.     

A deflection system to reduce the interference from post-source decay product ions in photodissociation tandem time-of-flight mass spectrometry

A deflection system consisting of four deflectors was designed and used to reduce the interference from post-source decay (PSD) product ions in photodissociation (PD) tandem time-of-flight (TOF) mass spectrometry. For simple protonated peptides generated by matrix-assisted laser desorption/ionization, the presence of PSD product ions at the laser irradiation spot was found to noticeably alter the minor peaks in the PD spectra even though the major ones were hardly affected. Other benefits from the use of the deflection system such as the improvement in the resolving power in PSD tandem mass spectra are discussed.     

Long-lived coherent oscillations of the femtosecond transients in cyanobacterial photosystem I

The pulsed excitation of electronic levels coupled to specific nuclear modes by a 26 fs laser pulse at 706 nm creates a wavepacket in the nuclear space of photopystem I (PS I) of Synechocystis sp. strain PCC 6803 both in the ground state and in the one-exciton manifold. Fourier transform of transient decay curves shows several low frequency peaks. The most prominent Power Spectral Density (PSD) peaks are at omega = 49 cm(-1) and omega = 88 cm(-1). The peculiarity of the coherent wavepacket in the PS I of S. sp. strain PCC 6803 is the unique, long-lived 49 cm(-1) and 88 cm(-1) oscillations with decay times up to 10 ps. It was suggested that such a long-lived coherence is determined by a contribution of the ground state wavepacket. The dependence of these two PSD peaks on the probe wavelength resembles the profile of the transient absorption spectra of PS I. The pump-probe signal in the Soret region reflects the dynamics of the ground state wavepacket created by pulsed excitation of the Q(y)-band. It was shown that the multimode Brownian oscillator model allows a quantitative fit of the oscillatory patterns of the pump-probe signal to be obtained.     

Matrix-Assisted Laser Desorption/Ionization Mass Spectrometric study of bis(imidazole-1-carboxylate) endfunctionalized polymers,

A detailed Matrix-Assisted Laser Desorption/Ionization Mass Spectrometric (MALDI-TOF MS) investigation of polyethylene glycol (PEG), polypropylene glycol (PPG) and polyisobutylene (PIB) bis(imidazole-1-carboxylate) esters is reported. The MS spectra of PPG and PIBb is (imidazole-1-carboxylate) esters recorded in the reflectron mode showed the presence of two additional series of peaks compared to those recorded in the linear mode, while in the case of PEG bis(imidazole-1-carboxylate) only one additional peak series appeared in the reflectron MS spectra. These additional series were attributed to the formation of fragment ions by the loss of one and two end groups in the first field-free region of the instrument. The neutral losses for the three polymers were also supported by using the post-source decay method (PSD). The observation of decreasing mass difference between the adjacent peaks from two series of polymeric ions with the increase of oligomer mass indicates the presence of PSD ions in a reflectron MALDI spectrum. The relationship between the mass of the precursor ions and those of the PSD ions and neutral loss are also discussed.     

Sequencing wasp venom peptides by endopeptidase digestion and nested collision-induced dissociation/post-source decay methods

A method incorporating nested collision-induced dissociation/post-source decay (CID/PSD) combined with endopeptidase digestion is described as an approach to determine the sequence of N-terminally modified peptides. The information from immonium and related ions observed in the CID/PSD spectrum was used for the selection of a suitable endopeptidase for the digestion of peptides. Rapid and reliable assignment of peptide sequence was performed by the comparison of CID/PSD spectra of both intact and endopeptidese-digested peptide fragments, since the assignments of the observed fragment ions to either N- or C-terminal ions can thus be carried out unambiguously. This nested CID/PSD method was applied to the sequence determination of two peptides from the solitary wasps Anoplius samariensis and Batozonellus maculifrons (pompilid wasps), which could not be sequenced by the Edman method due to N-terminal modification.     

Field ionization mass spectrometry—I. Nucleosides

The field ionization mass spectra of a series of nucleosides are reported, and compared with spectra obtained by conventional electron-impact ionization. The latter are complex, with structurally significant molecular ion and sugar cleavage peaks often of low intensity or completely absent. In contrast, the field ion spectra are extremely simple, with all except guanosine (highest mass peak [M — 18]) showing intense molecular ion peaks, and the characteristic sugar (S) and base (B + H) cleavage products as the only other important fragments.     

Comparison of the secondary-ion and electron-impact mass spectra of alkaloids

An advantage of using secondary-ion spectra for determining the molecular masses of quaternary bases and bases the molecular ions of which are unstable to electron impact is shown. The LSIMS spectra of four groups of diterpene alkaloids include the 100% peaks of the (M + H)⁺ ions and the peaks of the main fragments present in the EI spectra. The B/E = const. spectra of the key ions obtained by the different methods are compared. The closeness of the values of A of analogous transitions calculated from the B/E and MD spectra is confirmed.     

Investigation of VUV Photodissociation Propensities Using Peptide Libraries

PSD does not usually generate a complete series of y-type ions, particularly at high mass, and this is a limitation for de novo sequencing algorithms. It is demonstrated that b(2) and b(3) ions can be used to help assign high mass x(N-2) and x(N-3) fragments that are found in vacuum ultraviolet (VUV) photofragmentation experiments. In addition, v(N)-type ion fragments with side chain loss from the N-terminal residue often enable confirmation of N-terminal amino acids. Libraries containing several thousand peptides were examined using photodissociation in a MALDI-TOF/TOF instrument. 1345 photodissociation spectra with a high S/N ratio were interpreted.     

Laser Desorption Ionization Versus Electrospray Ionization Mass Spectrometry: Applications in the Analysis of Cluster Anions

Mass spectra of transition metal carbonyl cluster anions were recorded using laser desorption ionization time-of-flight (LDI-TOF) and electrospray ionization (ESI) techniques. The LDI spectra generally contain peaks corresponding the intact cluster together with extensive CO loss fragments ions whereas the ESI spectra exhibit peaks corresponding the intact cluster together with few (if any) CO loss fragment ions. The parameters of both techniques can be modified to vary the extent of fragmentation. In all cases no fragmentation of the metal core is observed. Overall, ESI is a more informative method for the analysis of these types of cluster anions.     

Direct mass spectrometric analysis of Bacillus spores

Spores from the Bacillus species, B. cereus, B. anthracis, B. thuringensis, B. lichenformis, B. globigi, and B. subtilis, were examined by direct probe mass spectrometry using electron ionization (EI) and positive and negative chemical ionization (CI). Molecular ions from free fatty acids and nucleic acids were observed in the 70eV spectra as were fragments from glycerides. Spectra obtained with isobutane positive chemical ionization (CI(+)) were dominated by ions associated with pyranose compounds such as N-acetylglucosamine (NAG). Unlike the positive ion spectra, the negative ion spectra of the spores were very simple and contained few peaks. The M(-.) ion from dipicolinic acid (DPA) was the base peak in the negative ion spectra of all spore species except those from B. lichenformis. The negative ion of DPA produced such a strong signal that 10(8) colony forming units (CFUs) of B. cereus spores could be detected directly in 0.5 g of ground rice. Principal component analysis (PCA) of the spectra revealed that only CI(+) spectra contained differences that could be used to identify the spectra by species. Differentiation of the CI(+) spectra by PCA was attributed to variances in the peaks associated with the bacterial polymer poly(3-hydroxybutyrate) (PHB) and NAG. Similar differences in PHB and NAG peaks were detected in the CI(+) spectra of a suite of vegetative Bacillus stains grown with various media.     

Mechanism of the Anionic Ring-Opening Oligomerization of Propylene Carbonate Initiated by the tert-Butylphenol/KHCO3 System

The ring-opening oligomerization reaction of propylene carbonate in the presence of the tert-butylphenol/KHCO₃ initiating system was studied by means of Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS) and Electrospray Ionization Time of Flight Mass Spectrometry (ESI-TOF MS). According to the MS spectra obtained, different series of peaks were identified. The MS spectra clearly showed that besides the chain-extension reaction yielding oligomers with all propylene oxide units, the formation of oligomers containing carbonate linkages in the chain, and condensation reaction between the latter two also took place. The structure of the oligomers carrying carbonate linkages was determined by the post-source decay (PSD) MALDI-TOF MS/MS method. Based on the MS results, a mechanism for the oligomerization reaction is proposed.     

Chemical characterisation of different separation media based on agarose by static time-of-flight secondary ion mass spectrometry

In this paper, the novel application of time-of-flight secondary ion mass spectrometry (TOF-SIMS) for qualitative and semi-quantitative investigation of the surface chemistry of separation media based on beaded agarose is reported. Five different media were studied: DEAE Sepharose Fast Flow, Q Sepharose Fast Flow, SP Sepharose Fast Flow, Phenyl Sepharose Fast Flow at ligand densities between 7 and 33% (w/w) and the base matrix Sepharose 6 Fast Flow. The obtained TOF-SIMS spectra reveal significant chemical information regarding the ligands (DEAE, Q, SP and Phenyl) which are covalently attached to the agarose-based matrix Sepharose 6 Fast Flow. For the anion-exchange media (DEAE and Q Sepharose Fast Flow), the positive TOF-SIMS spectra yielded several strong characteristic fragment peaks from the amine ligands. Structural information was obtained, e.g. from the peak at m/z 173.20, originating from the ion structure [(C2H5)2NCH2CH2NH(C2H5)2l+, which shows that the ligand in DEAE Sepharose Fast Flow is composed of both tertiary and quaternary amines. The positive spectrum of Phenyl Sepharose Fast Flow contained major fragments both from the base matrix and the ligand. The cation-exchanger (SP Sepharose Fast Flow) gave rise to a positive spectrum resembling that of the base matrix (Sepharose 6 Fast Flow) but with a different intensity pattern of the matrix fragments. In addition, peaks with low intensity at m/z 109.94, 125.94 and 139.95 corresponding to Na2SO2+, Na2SO3+ and Na2SO3CH2+, respectively, were observed. The positive TOF-SIMS spectrum of Sepharose 6 Fast Flow contains a large number of fragments in the mass range up to m/z 200 identified as CxHyOz and CxHy structures. The results clearly show that positive TOF-SIMS spectra of different media based on Sepharose 6 Fast Flow are strongly influenced by the ligand coupled to the matrix. The negative TOF-SIMS spectra contained several ligand-specific, characteristic peaks for the cation-exchanger, having sulphonate as the ion-exchange group. Negative fragments such as S-, SO-, SO2-, SO3-, C2H3SO3-, C3H5SO3- and OC3H5SO3- were observed. Phenyl Sepharose Fast Flow, which has an uncharged group (Phenyl) coupled to the agarose matrix yielded one ligand-related peak corresponding to the C6H5O- fragment. DEAE and Q ligands could only be identified by the appearance of the fragments CN- and CNO- in the negative spectrum. However, a strong peak corresponding to the counter ion (Cl-) was observed. TOF-SIMS analysis can also be used for the investigation of residues from the coupling procedure that bonds the ligands to the matrix. One example is the observation of bromine peaks in the negative spectrum of Q Sepharose Fast Flow. Furthermore, it has also been shown that different ligand concentrations of Phenyl Sepharose Fast Flow can easily be detected by TOF-SIMS analysis. Information regarding the difference between the ligand density on the surface of the beads and in the bulk can also be obtained. However, spectra registered on the outermost surface and on the pore surface (crushed beads) of DEAE Sepharose Fast Flow clearly show that the agarose and the DEAE groups are homogeneously distributed in the beads.     

Mass spectrometry of the trimethylsilyl derivatives of some cardiac aglycones and monoglycosides

Mass spectra of the trimethylsilyl (TMS) ethers of digitoxigenin, digoxigenin, digitoxigenin monodigitoxoside, digoxigenin monodigitoxoside and peruvoside were obtained. Ions appearing at m/e 170 and m/e 183 are proposed to be indicative of the enol TMS ether moiety as confirmed by appropriate shifts in the mass spectrum of digitoxigenin derivatized with perdeuterotrimethylsilylating reagent. The TMS ether derivatives of the monoglycosides produced abundant molecular ions and the base peaks in the spectra represent fragments of the sugar portion of the molecule.     

Vacuum-ultraviolet photoionization and mass spectrometric characterization of lignin monomers coniferyl and sinapyl alcohols

The fragmentation mechanisms of monolignols under various energetic processes are studied with jet-cooled thermal desorption molecular beam (TDMB) mass spectrometry (MS), 25 keV Bi(3)(+) secondary ion MS (SIMS), synchrotron vacuum-ultraviolet secondary neutral MS (VUV-SNMS) and theoretical methods. Experimental and calculated appearance energies of fragments observed in TDMB MS indicate that the coniferyl alcohol photoionization mass spectra contain the molecular parent and several dissociative photoionization products. Similar results obtained for sinapyl alcohol are also discussed briefly. Ionization energies of 7.60 eV ± 0.05 eV for coniferyl alcohol and <7.4 eV for both sinapyl and dihydrosinapyl alcohols are determined. The positive ion SIMS spectrum of coniferyl alcohol shares few characteristic peaks (m/z = 137 and 151) with the TDMB mass spectra, shows extensive fragmentation, and does not exhibit clear molecular parent signals. VUV-SNMS spectra, on the other hand, are dominated by the parent ion and main fragments also present in the TDMB spectra. Molecular fragmentation in VUV-SNMS spectra can be reduced by increasing the extraction delay time. Some features resembling the SIMS spectra are also observed in the desorbed neutral products. The monolignol VUV-SNMS peaks shared with the TDMB mass spectra suggest that dissociative photoionization of ion-sputtered neutral molecules predominate in the VUV-SNMS mass spectra, despite the extra internal energy imparted in the initial ion impact. The potential applications of these results to imaging MS of biomolecules are discussed.     

Comparisom of electron impact and field desorption mass spectra I : Substituted sultams

The low and the high resolution electron impact (EI) and field desorption (FD) mass spectra (MS) of substituted 5- or 6-membered ring sultams are given and the main fragmentation pathways are interpreted. Sultams carrying polar substituents, such as dicarboxylic acids and their mono- and di-amides show either small or no molecular ions with EI-MS. Their existence, however, can be indirectly inferred by metastable defocusing. In contrast to these findings FD-MS display in all our examples the quasimolecular ions (M + 1)⁺ as the base peaks of the spectra. While the pattern for the EI fragmentations follows the generally known rules very closely, five general aspects for the fragmentation pathway of FD mass spectra have been established which demonstrate the characteristics of this modern analytical method. The use of high temperature activated emitters enables a choice to be made between obtaining only molecular peaks and producing significant fragments via a controlled thermal degradation. This is the first time that the complementary and different characters of the information obtained with the two ionization modes EI and FD have been demonstrated on a homologous series of compounds.     

Soft x-ray photoreactions of CF3Cl adsorbed on Si(111)-7x7 studied by continuous-time photon-stimulated desorption spectroscopy near F(1s) edge

The continuous-time core-level photon-stimulated desorption (PSD) spectroscopy was employed to monitor the monochromatic soft x-ray-induced reactions of CF3Cl adsorbed on Si(111)-7x7 near the F(1s) edge (681-704 eV). Sequential F+ PSD spectra were measured as a function of photon exposure at the CF3Cl-covered surface (dose=0.3x10(15) molecules/cm2, approximately 0.75 ML). The F+ PSD and total electron yield (TEY) spectra of molecular solid CF3Cl near the F(1s) edge were also measured. Both F+ PSD and TEY spectra show two features at the energy positions of 690.2 and 692.6 eV, and are attributed to the excitations of F(1s) to 11a1[(C-Cl)*] and (8e+12a1)[(C-F)*] antibonding orbitals, respectively. Following Auger decay, two holes are created in the F(2p) lone pair and/or C-F bonding orbitals forming the 2h1e final state which leads to the F+ desorption. This PSD mechanism, which is responsible for the F+ PSD of solid CF3Cl, is employed to interpret the first F+ PSD spectrum in the sequential F+ PSD spectra. The variation of spectrum shapes in the sequential F+ PSD spectra indicates the dissipation of adsorbed CF3Cl molecules and the formation of surface SiF species as a function of photon exposure. From the sequential F+ PSD spectra the photolysis cross section of the adsorbed CF3Cl molecules by photons with varying energy (681-704 eV) is determined to be approximately 1.0x10(-17) cm2.