Quantitative analysis of low molecular weight polar compounds by continuous flow liquid secondary ion tandem mass spectrometry

Quantitative analyses of low molecular weight (100-200) polar compounds [1-methyl-4-phenylpyridine (MPP+), 2-amino-3-(methylamino)propanoic acid (synonyms, beta-(methyl-amino)-L-alanine or BMAA), and tryptophan] were conducted on a triple-stage quadrupole mass spectrometer configured for continuous flow liquid secondary ion mass spectrometry ionization (CF L-SIMS). It is shown that quantification by CF L-SIMS at subnanogram sensitivity can be precise (correlation coefficients greater than 0.99), accurate, specific, and routine for compounds not measurable by static L-SIMS. Successful analyses, however, are strongly dependent upon the stability of the film formed by the mobile phase on the probe tip. In our system, film stability is affected by mobile phase composition and flow rate, ion source and probe tip temperature, probe-tip and capillary alignment, film thickness, and sample composition.     

Fast screening of low molecular weight compounds by thin-layer chromatography and "on-spot" MALDI-TOF mass spectrometry

Fast screening of low-MW compounds is performed by thin-layer chromatography (TLC) followed by direct on-spot matrix-assisted laser desorption/ionization time-of-flight mass spectrometry identification with nearly "matrix-free" mass spectra using an UV-absorbing ionic liquid matrix. Owing to minimal background ions from the proton donor triethylamine/alpha-cyano-4-hydroxycinnamic acid ionic liquid matrix, three arborescidine alkaloids, the anesthesics levobupivacaine and mepivacaine, and the antibiotic tetracycline were readily characterized most frequently by the MS detection of their protonated molecules. The technique is fast and sensitive, requires little sample preparation and manipulation, and is therefore suitable for fast screening with TLC separation and MS identification of low-MW compounds, with potential applications in areas such as phytochemistry, synthetic chemistry, and product manufacturing quality monitoring.     

Metal-assisted secondary ion mass spectrometry: influence of Ag and Au deposition on molecular ion yields

A series of organic dyes and a pharmaceutical are used to study secondary ion yield enhancement by metal deposition. The molecules were dissolved in methanol and spin-casted on silicon substrates. Subsequently, silver or gold was evaporated onto the samples to produce a very thin coating. The coated samples, when measured with TOF-SIMS, showed a considerable increase in characteristic secondary ion intensity. Gold evaporated samples appeared to exhibit the highest signal enhancement. A major advantage of the metallization technique is that it can be used on real world samples. In particular, additive containing organic crystals could be studied; however, the observed signal increase does not occur at any given moment. The time between metal deposition on the sample surface and the measurement of the sample with TOf-SIMS appears to have an important influence on the enhancement of the secondary ion intensities. Therefore, the metal-coated samples were measured at different times after sample preparation. The results show that, depending on the sample and the metal deposited, the secondary ion signals reach their maximum at different times. Further study will be necessary to reveal the mechanism responsible for the observed enhancement effect.     

Reversed-phase membrane inlet mass spectrometry applied to the real-time monitoring of low molecular weight alcohols in chloroform.

Reversed-phase membrane inlet mass spectrometry incorporating a hollow-fiber Nafion membrane has been evaluated for the determination of low molecular weight alcohols in chloroform. The hydrophilic Nafion membrane preferentially transports methanol and ethanol, allowing percentage concentrations of the alcohols to be determined in a chloroform matrix. A linear response was observed for ethanol over the working range 0.5-2.5%, with a limit of detection of 0.1%. The application of reversed-phase membrane inlet mass spectrometry using a Nafion membrane to the monitoring of a chloroform recovery process has been investigated using a residual gas analyzer. Evolving methanol and ethanol concentrations were determined in real time and compared favorably with off-line determinations by gas chromatography.     

Instrumental aspects of secondary ion mass spectrometry and secondary ion imaging mass spectrometry

A short survey of the varieties of the Secondary Ion Mass Spectrometry (SIMS) known at present is given. The principle of quantitative analysis with respect to thin film analysis is discussed. The properties of SIMS and SIIMS (Secondary Ion Imaging Mass Spectrometry) are compared with those of Electron Microprobe Analysis. Results of an analysis of a thin film of titanium oxide and of an FeMn ferrite by means of SIMS and SIIMS are given.     

Observation of low molecular weight poly(methylsilsesquioxane)s by graphite plate laser desorption/ionization time-of-flight mass spectrometry

Mass spectra of polystyrene and poly(methylsilsesquioxane)(PMSSQ) derived from methyltriethoxysilane(MTES) were obtained in the 100-1,000 Da range by laser desorption/ionization time-of-flight mass spectrometry using a graphite plate without a matrix. Clean mass spectra were obtained without interference from carbon clusters or other low molecular weight compounds. Initial reaction products derived from condensation of partially hydrolyzed MTES were observed. Upon 30 min of heating at 30 degrees C, the ethoxy groups were fully hydrolyzed to hydroxy groups. Many PMSSQ species consistent with predictable polymerization reaction pathways involving intermolecular condensation and intramolecular dehydration were observed. Thus, laser desorption/ionization time-of-flight mass spectrometry using a graphite plate, without added matrix materials, is shown to provide valuable information on low molecular weight polymer not available by MALDI-TOF-MS.     

Cluster secondary ion mass spectrometry and the temperature dependence of molecular depth profiles

The quality of molecular depth profiles created by erosion of organic materials by cluster ion beams exhibits a strong dependence upon temperature. To elucidate the fundamental nature of this dependence, we employ the Irganox 3114/1010 organic delta-layer reference material as a model system. This delta-layer system is interrogated using a 40 keV C(60)(+) primary ion beam. Parameters associated with the depth profile such as depth resolution, uniformity of sputtering yield, and topography are evaluated between 90 and 300 K using a unique wedge-crater beveling strategy that allows these parameters to be determined as a function of erosion depth from atomic force microscope (AFM) measurements. The results show that the erosion rate calibration performed using the known Δ-layer depth in connection with the fluence needed to reach the peak of the corresponding secondary ion mass spectrometry (SIMS) signal response is misleading. Moreover, we show that the degradation of depth resolution is linked to a decrease of the average erosion rate and the buildup of surface topography in a thermally activated manner. This underlying process starts to influence the depth profile above a threshold temperature between 210 and 250 K for the system studied here. Below that threshold, the process is inhibited and steady-state conditions are reached with constant erosion rate, depth resolution, and molecular secondary ion signals from both the matrix and the Δ-layers. In particular, the results indicate that further reduction of the temperature below 90 K does not lead to further improvement of the depth profile. Above the threshold, the process becomes stronger at higher temperature, leading to an immediate decrease of the molecular secondary ion signals. This signal decay is most pronounced for the highest m/z ions but is less for the smaller m/z ions, indicating a shift toward small fragments by accumulation of chemical damage. The erosion rate decay and surface roughness buildup, on the other hand, exhibit a rather sudden delayed onset after erosion of about 150 nm, indicating that a certain damage level must be reached in order to influence the erosion dynamics. Only after that onset does the depth resolution become compromised, indicating that the temperature reduction does not significantly influence parameters like ion-beam mixing or the altered-layer thickness. In general, the wedge-crater beveling protocol is shown to provide a powerful basis for increased understanding of the fundamental factors that affect the important parameters associated with molecular depth profiling.     

Characterization of phosphoantigens by high-performance anion-exchange chromatography-electrospray ionization ion trap mass spectrometry and nanoelectrospray ionization ion trap mass spectrometry

New phosphorylated microbial metabolites referred to as phosphoantigens activate immune responses in humans. Although these molecules have leading applications in medical research, no direct method allows their rapid and unambiguous structural identification. Here, we interfaced online HPAEC (high performance anion-exchange chromatography) with ESI-ITMS (electrospray ionization ion trap mass spectrometry) to identify such pyrophosphorylated molecules. A self-regenerating anion suppressor located upstream of electrospray ionization enabled the simultaneous detection of pyrophosphoester by conductimetry, UV and MS. By HPAEC-ITMS and HPAEC-ITMS2, a single run permitted characterization of reference phosphoantigens and of related structures. Although all compounds were resolved by HPAEC, MS enabled their detection and identification by [M-H]- and fragment ions. Isobaric phosphoantigen analogues were also separated by HPAEC and distinguished by MS2. The relevance of this device was demonstrated for phosphoantigens analysis in human urine and plasma. Furthermore, identification of natural phosphoantigens by automatically generated 2D mass spectra from nano-ESI-ITMS is presented. This last technique permits the simultaneous performance of molecular screening of natural phosphoantigen extracts and their identification.     

Determination of molecular weight distributions of tert-octylphenol ethoxylate surfactant polymers by laser desorption Fourier transform ion cyclotron resonance mass spectrometry and high-performance liquid chromatography

Triton polymers are commercial surfactants whose molecular weight distributions are conventionally determined by means of high-performance liquid chromatography (HPLC). However, in the case of the important octylphenol ethoxylates [p-C8H17-C6H4-O-(CH2CH2O)n-H], HPLC cannot resolve individual oligomers of high molecular weight Triton surfactants (e.g., greater than 2000 u or so; u = unified atomic mass unit). In this paper, we show that laser desorption Fourier transform ion cyclotron resonance mass spectrometry (LD/FT/ICR/MS) provides a simple and accurate measure of such Triton surfactant molecular weight distributions up to at least 3500 u, based on a single-shot laser pulse measurement of a few seconds duration. Comparison of LD/FT/ICR/MS and HPLC molecular weight distributions of low molecular weight surfactants shows that laser desorption/ionization produces minimal fragmentation and thus offers an accurate measure of the relative abundances of the neutral oligomers, without the need for prior chromatographic separation of the components. Moreover, for all Triton polymer molecular weight distributions (700-3000 u), LD/FT/ICR/MS provides much more highly resolved profiles of oligomer relative abundances. Finally, LD/FT/ICR/MS reveals the presence of poly(ethylene oxide) side products of the polymerization process, which are not observed by HPLC with conventional ultraviolet absorption detection.     

Metal salts for molecular ion yield enhancement in organic secondary ion mass spectrometry: a critical assessment

In a search for molecular ion signal enhancement in organic SIMS, the efficiency of a series of organic and inorganic salts for molecular cationization has been tested using a panel of nonvolatile molecules with very different chemical characteristics (leucine enkephalin, Irganox 1010, tetraphenylnaphthalene, polystyrene). The compounds used for cationization include alkali bromide and group Ib metal salts (XBr with X = Li, Na, K; CF3CO2Ag; AgNO3; [CH3COCH=C(O-)CH3]2Cu; AuCl3). Alkali ions, very good for polar molecule cationization, prove to be of limited interest for nonpolar molecules such as polystyrene. Silver trifluoroacetate displays excellent results for all the considered molecules, except for leucine enkephalin (which might be due to the use of different solvents for the analyte and the salt). Instead, silver nitrate mixed with leucine enkephalin in an ethanol solution provides intense molecular signals. The influence of the respective concentrations of analyte and salt in solution, of the silver trifluoroacetate solution stability, and of the sample microstructure on the secondary ion intensities are also investigated. The results of other combinations of analyte and salts are reported. Finally, the use of salts is critically compared to other sample preparation procedures previously proposed for SIMS analysis of large organic molecules.     

Negative ion mass spectrometry of sialylated carbohydrates: discrimination of N-acetylneuraminic acid linkages by MALDI-TOF and ESI-TOF mass spectrometry

Negative ion MALDI and electrospray fragmentation spectra were recorded from 12 sialylated carbohydrates ranging from trisaccharides to biantennary N-linked glycans. D-Arabinosazone was found to be the most satisfactory MALDI matrix for these compounds. Fragmentation mechanisms were investigated with the aid of several synthesized analogues of the sugars labeled with 13C and 2H. The substitution position of the sialic acid (alpha2-->3 or alpha2-->6) was found to have a dramatic effect on the overall fragmentation pattern of these compounds, and several features of the spectra were identified that allowed the substitution pattern to be determined. In particular, the appearance of an ion at m/z 306 appeared to be diagnostic of the presence of an alpha2-->6-linked sialic acid. Selection and further fragmentation of the in-source (conevoltage) fragment ion corresponding to the trisaccharide Neu5Acalpha2-->3(or 6)Galbeta1-->4GlcNAc from larger, N-linked glycans, ionized by electrospray, gave fragmentation patterns identical to those of the reference trisaccharides, thus providing a method for confirming the sialic acid linkage.     

Analysis of phases in the Y-Ba-Cu-O system by lims technique laser-induced ion mass spectrometry

Laser-induced ion mass spectrometry (LIMS) was used for the analysis of oxide phases in the Y-Ba-Cu-O system. The formation of polynuclear ions (clusters) as a result of laser sputtering of oxides was examined. The possibility of quantitative phase analysis of inhomogeneous oxide materials in the Y-Ba-Cu-O system by the LIMS technique is demonstrated.     

Protein dynamics in iron-starved Mycobacterium tuberculosis revealed by turnover and abundance measurement using hybrid-linear ion trap-Fourier transform mass spectrometry

To study the proteome response of Mycobacterium tuberculosis H37Rv to a change in iron level, iron-starved late-log-phase cells were diluted in fresh low- and high-iron media containing [ (15)N]-labeled asparagine as the sole nitrogen source for labeling the proteins synthesized upon dilution. We determined the relative protein abundance and protein turnover in M. tuberculosis H37Rv under these two conditions. For measurements, we used a high-resolution hybrid-linear ion trap-Fourier transform mass spectrometer coupled with nanoliquid chromatography separation. While relative protein abundance analysis shows that only 5 proteins were upregulated by high iron, 24 proteins had elevated protein turnover for the cells in the high-iron medium. This suggests that protein turnover is a sensitive parameter to assess the proteome dynamics. Cluster analysis was used to explore the interconnection of protein abundance and turnover, revealing coordination of the cellular processes of protein synthesis, degradation, and secretion that determine the abundance and allocation of a protein in the cytosol and the extracellular matrix of the cells. Further potential utility of the approach is discussed.     

Molecular weight distributions of asphaltenes and deasphaltened oils studied by laser desorption ionization and ion mobility mass spectrometry

Laser desorption ionization (LDI) and ion mobility mass spectrometry (IM-MS) are applied to study molecular weight distribution and cross sections of petroleum asphaltene (ASPH) and deasphaltened oils (DAO). Ion mobility data confirmed the presence of gas-phase aggregation in LDI experiments. Most of the molecules with MW > 3000 g/mol in LDI result from gas-phase aggregation. Two-dimensional (2D) IM-MS trend lines are compared with model polymer systems to confirm the order of cross sections (polywax > polystyrene > DAO > ASPH > fullerenes), and these data illustrate that ASPH has a more condensed average structure than DAO.     

A C60 primary ion beam system for time of flight secondary ion mass spectrometry: its development and secondary ion yield characteristics

A buckminsterfullerene (C60)-based primary ion beam system has been developed for routine application in TOF-SIMS analysis of organic materials. The ion beam system is described, and its performance is characterized. Nanoamp beam currents of C60+ are obtainable in continuous current mode. C60(2+) can be obtained in pulsed mode. At 10 keV, the beam can be focused to less than 3 microm with 0.1 nA currents. TOF-SIMS studies of a series of molecular solids and a number of polymer systems in monolayer and thick film forms are reported. Very significant enhancement of secondary ion yields, particularly at higher mass, were observed using 10-keV C60+ for all samples other than PTFE, as compared to those observed from 10 keV Ga+ primary ions. Three materials (PS2000, Irganox 1010, PET) were studied in detail to investigate primary ion-induced disappearance (damage) cross sections to determine the increase in secondary ion formation efficiency. The C60 disappearance cross sections observed from monolayer film PS2000 and self-supporting PET film are close to those observed from Ga+. The resulting C60 efficiencies are 30-100 times those observed from gallium. The cross sections observed from C60 bombardment of multilayer molecular solids are approximately 100 times less, such that essentially zero damage sputtering is possible. The resulting efficiencies are > 10(3) greater than from gallium. It is also shown that C60 primary ions do not generate any more low-mass fragments than any other ion beam system does. C60 is shown to be a very favorable ion beam system for TOF-SIMS, delivering high yield, close to 10% total yield, favoring high-mass ions, and on thick samples, offering the possibility of analysis well beyond the static limit.     

Solvent extraction, ion chromatography, and mass spectrometry of molybdenum isotopes

A procedure was developed that allows precise determination of molybdenum isotope abundances in natural samples. Purification of molybdenum was first achieved by solvent extraction using di(2-ethylhexyl) phosphate. Further separation of molybdenum from isobar nuclides was obtained by ion chromatography using AG1-X8 strongly basic anion exchanger. Finally, molybdenum isotopic composition was measured using a multiple collector inductively coupled plasma hexapole mass spectrometer. The abundances of molybdenum isotopes 92, 94, 95, 96, 97, 98, and 100 are 14.8428(510), 9.2498(157), 15.9303(133), 16.6787(37), 9.5534(83), 24.1346(394), and 9.6104(312) respectively, resulting in an atomic mass of 95.9304(45). After internal normalization for mass fractionation, no variation of the molybdenum isotopic composition is observed among terrestrial samples within a relative precision on the order of 0.00001-0.0001. This demonstrates the reliability of the method, which can be applied to searching for possible isotopic anomalies and mass fractionation.