Ultratrace analysis and isotope ratio measurements of long-lived radioisotopes by resonance ionization mass spectrometry (RIMS)

Resonance Ionization Mass Spectrometry (RIMS) is a sensitive and selective method for ultratrace analysis of long-lived radioisotopes and isotope ratio measurements. It provides extremely high isobaric suppression and good overall efficiency. The experimental limits of detection are as low as 10⁶ atoms per sample and isotopic selectivities of 5×10¹² have been obtained. The widespread potential of RIMS, using different experimental arrangements, is demonstrated for the determination of the radiotoxic isotopes Pu-238 to Pu-244 and Sr-89/Sr-90 in various environmental samples as well as for Ca-41 in nuclear reactor components and biomedical samples.     

Electrospray ionisation Fourier-transform ion cyclotron resonance mass spectrometry of dynamic combinatorial libraries

The use of electrospray ionisation Fourier-transform ion cyclotron resonance tandem mass spectrometry (ESI-FTICR-MS/MS) for the analysis of dynamic combinatorial libraries (DCLs) of pseudo-peptide macrocyclic hydrazone oligomers is presented. The design of library building blocks results in mixtures of compounds with greater diversity than libraries generated by conventional combinatorial chemistry and so presents increased demands for analysis. The extended capabilities of the FTICR technique, specifically selective ion trapping, sensitivity, high resolution and mass accuracy over a broad mass range, are compatible with these increased demands and, most importantly, without the need for chromatography. Preliminary studies on the sequencing of cyclic oligomers and confirmation of the presence of sequence isomers are presented. These studies highlight the potential of FTICR-MS as a superior technique for the analysis of combinatorially generated compounds.     

Homogeneous sample preparation for automated high throughput analysis with matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

This work presents a simple method for obtaining homogeneous sample surfaces in matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) for the automated analysis of peptides and proteins. The sample preparation method is based on applying the sample/matrix mixture onto a pre-deposited highly diluted matrix spot. The pre-deposited crystals act as seeds for the new sample containing crystals which become much smaller in size and more evenly distributed than with conventional methods. This 'seed-layer' method was developed, optimised and compared with the dried-droplet method using peptides and proteins in the 1000-20,000 Da range. The seed-layer method increases the surface homogeneity, spot to spot reproducibility and sample washability as compared with the commonly used dried-droplet method. This methodology is applicable to alpha-cyanohydroxycinnamic acid, sinapinic acid and ferulic acid, which all form homogeneous crystal surfaces. Within-spot variation and between-spot variation was investigated using statistics at a 95% confidence level (n = 36). The statistical values were generated from more than 5000 data points collected from 500 spectra. More than 90% of the sample locations results in high intensity spectra with relatively low standard deviations (RSDs). Typically obtained data showed an RSD of 19-35% within a sample spot as well as in-between spots for proteins, and an RSD of < or = 50% for peptides. Linear calibration curves were obtained within one order of magnitude using internal calibration with a point-RSD of 3% (n = 10). The sample homogeneity allows mass spectra (average of 16 laser shots) to be obtained on each individual sample within 15 sec, whereby a 100 spot target plate can be run in 25 min. High density target plates using the seed-layer method were prepared by spotting approximately 100 picoliter droplets onto the target, resulting in sample spots < or = 500 microns in diameter using a flow-through piezo-electric micro-dispenser. By using this automated sample preparation step lower standard deviations are obtained in comparison to manually prepared samples.     

Characterisation of sulphoxides by atmospheric pressure ionisation mass spectrometry

An observation that a series of proprietary compounds containing a methyl thiophenyl group all underwent metabolic S-oxidation, and that the product ion spectra of the resulting S-oxides showed methyl radical loss under low-energy atmospheric pressure ionisation tandem mass spectrometry (API-MS/MS) conditions, has led to an investigation of the fragmentation of commercially available sulphoxides. The phenyl methyl sulphoxides studied do lose methyl radicals under MS/MS conditions on triple quadrupole mass spectrometers. In addition, the phenyl sulphoxides, with simple substituents other than a methyl group, also showed a tendency to lose the substituent as a radical. It was concluded that radical loss from these simple sulphoxides was characteristic of S-oxidation of these molecules. Radical losses, such as those reported here, are used in-house to distinguish S-oxidation from N- and C-oxidation in metabolism studies.     

Structure analysis of naturally occurring alkyl porphyrins by hydrogen chemical ionisation mass spectrometry

Chemical ionisation (CI) mass spectrometry with H₂ as reagent gas has been used to obtain spectra of a number of demetallated alkyl porphyrins from an oil shale and a bitumen. The spectra give more structural informa- tion than the electron impact spectra, by way of odd electron dipyrrolic fragment ions. Thus, the structural possibilities for a novel /c30 aetio porphyrin have been narrowed to two, although¹HNMR only reduced the possibilities to six. The spectra of four types of components containing a five, six, seven or methyl-substituted six membered exocyclic ring show, for example, that three out of four C₃₂ isomers can be distinguished. The technique offers promise as a tool in structural elucidation of other novel naturally occurring porphyrins.     

Matrix-free mass spectrometric imaging using laser desorption ionisation Fourier transform ion cyclotron resonance mass spectrometry

Mass spectrometry imaging (MSI) is a powerful tool in metabolomics and proteomics for the spatial localization and identification of pharmaceuticals, metabolites, lipids, peptides and proteins in biological tissues. However, sample preparation remains a crucial variable in obtaining the most accurate distributions. Common washing steps used to remove salts, and solvent-based matrix application, allow analyte spreading to occur. Solvent-free matrix applications can reduce this risk, but increase the possibility of ionisation bias due to matrix adhesion to tissue sections. We report here the use of matrix-free MSI using laser desorption ionisation performed on a 12 T Fourier transform ion cyclotron resonance (FTICR) mass spectrometer. We used unprocessed tissue with no post-processing following thaw-mounting on matrix-assisted laser desorption ionisation (MALDI) indium-tin oxide (ITO) target plates. The identification and distribution of a range of phospholipids in mouse brain and kidney sections are presented and compared with previously published MALDI time-of-flight (TOF) MSI distributions.     

Characterisation of nicotine and related compounds using electrospray ionisation with ion trap mass spectrometry and with quadrupole time-of-flight mass spectrometry and their detection by liquid chromatography/electrospray ionisation mass spectrometry

Electrospray ionisation ion trap mass spectrometry (ESI-MS(n)) has been used to study the fragmentation patterns of nicotine and nine of its related compounds. From this study certain characteristic fragmentations are apparent with generally the pyrrolidine or piperidine ring being subject to chemical modifications. The structures of the product ions proposed for the ESI-MS(n) study have been supported by results from electrospray ionisation quadrupole time-of-flight mass spectrometry (ESI-QTOF-MS). Compounds with pyrrolidine and piperidine rings that possess an unsubstituted N atom have been shown to lose NH(3) at the MS(2) stage. Those compounds with N-methyl groups lose CH(3)NH(2) at the MS(2) stage. The loss of NH(3) or CH(3)NH(2) leaves the corresponding rings opened and this is followed by ring closure at the pyridine-2 carbon atom. Mono-N-oxides fragment in a similar way but the di-N-oxide can also fragment by cleavage of the bond between the pyridine and pyrrolidine rings. Cotinine also can undergo cleavage of this bond between the rings.This data therefore provides useful information on how substituents and the nature of the non-pyridine ring can affect the fragmentation patterns of nicotine and its related compounds. This information can be used in the characterisation of these compounds by liquid chromatography/electrospray ionization mass spectrometry (LC/ESI-MS) which results in the separation of nicotine and its related compounds with limits of detection (LODs) ranging from 15 to 105 ng/mL. The use of LC/ESI-MS to study nicotine-containing samples resulted in the simultaneous and unambiguous identification of seven of the compounds discussed in this paper: cotinine identified at retention time 12.5 min (with its [M+H](+) ion at m/z 177), nornicotine 16.0 min (m/z 149), anatabine 18.0 min (m/z 161), myosmine 18.5 min (m/z 147), anabasine 20.4 min (m/z 163), nicotine 22.2 min (m/z 163), and nicotyrine 31.4 min (m/z 159). For quality control of nicotine replacement therapy products, these nicotine impurities can be readily identified and determined at levels up to 0.3% for single impurities and up to 1.0% for total impurities.     

Investigation of drift gas selectivity in high resolution ion mobility spectrometry with mass spectrometry detection

Recent studies in electrospray ionization (ESI)/ion mobility spectrometry (IMS) have focussed on employing different drift gases to alter separation efficiency for some molecules. This study investigates four structurally similar classes of molecules (cocaine and metabolites, amphetamines, benzodiazepines, and small peptides) to determine the effect of structure on relative mobility changes in four drift gases (helium, nitrogen, argon, carbon dioxide). Collision cross sections were plotted against drift gas polarizability and a linear relationship was found for the nineteen compounds evaluated in the study. Based on the reduced mobility database, all nineteen compounds could be separated in one of the four drift gases, however, the drift gas that provided optimal separation was specific for the two compounds.     

Field ionisation mass spectrometry of acetylpyridine isomers

An investigation of the metastable transition in the field ionisation mass spectra of acetylpyridine isomers has established that the reaction pathway is greatly influenced by the ring position of the substituent. It is suggested that the pyridine nitrogen atom induces a rearrangement process in the case of 2-acetylpyridine. In other acetylpyridines it has a little effect and the cleavage reaction occured prior to rearrangement.     

Ambient ionisation mass spectrometry for in situ analysis of intact proteins

Ambient surface mass spectrometry is an emerging field which shows great promise for the analysis of biomolecules directly from their biological substrate. In this article, we describe ambient ionisation mass spectrometry techniques for the in situ analysis of intact proteins. As a broad approach, the analysis of intact proteins offers unique advantages for the determination of primary sequence variations and posttranslational modifications, as well as interrogation of tertiary and quaternary structure and protein‐protein/ligand interactions. In situ analysis of intact proteins offers the potential to couple these advantages with information relating to their biological environment, for example, their spatial distributions within healthy and diseased tissues. Here, we describe the techniques most commonly applied to in situ protein analysis (liquid extraction surface analysis, continuous flow liquid microjunction surface sampling, nano desorption electrospray ionisation, and desorption electrospray ionisation), their advantages, and limitations and describe their applications to date. We also discuss the incorporation of ion mobility spectrometry techniques (high field asymmetric waveform ion mobility spectrometry and travelling wave ion mobility spectrometry) into ambient workflows. Finally, future directions for the field are discussed.     

The characterisation of selected antidepressant drugs using electrospray ionisation with ion trap mass spectrometry and with quadrupole time-of-flight mass spectrometry and their determination by high-performance liquid chromatography/electrospray ionisation tandem mass spectrometry

The electrospray ionisation ion trap tandem mass spectrometry (ESI-MS(n)) of selected antidepressant drugs, i.e., citalopram, fluoxetine, mirtazapine, paroxetine, sertraline, and venlafaxine, has been investigated. Sequential product ion fragmentation experiments (MS(n)) have been performed in order to elucidate the degradation pathways for the [M+H](+) ions and their predominant product ions. These MS(n) experiments show certain characteristic fragmentations in that functional groups are generally cleaved from the ring systems as molecules such as H(2)O, amines and phenols. When an aromatic entity is present in a drug molecule together with a nitrogen-containing saturated ring structure as with mirtazapine, fragmentation initially occurs at the latter ring with the former being predictably resistant to fragmentation. Also, when an amine-containing drug molecule such as fluoxetine also contains a functional group, which liberates a phenol with a significantly lower DeltaH(f) (0) value than that of the corresponding amine, the phenol is preferentially liberated. The structures of product ions proposed for ESI-MS(n) can be supported by electrospray ionisation quadrupole-time-of-flight tandem mass spectrometry (ESI-QToF-MS/MS). These molecules can be identified and determined in mixtures at low ng/mL concentrations by the application of high-performance liquid chromatography/electrospray ionisation tandem mass spectrometry (HPLC/ESI-MS(2)), which can also be used for their analysis in hair samples.     

Analysis of oak tannins by liquid chromatography-electrospray ionisation mass spectrometry

Extractable tannins were analysed by liquid chromatography-electrospray ionisation mass spectrometry in two oak species, North American white oak (Quercus alba) and European red oak (Quercus robur). They mainly included various glucose gallic and ellagic acid esters. The structures were partially determined, and they included grandinin/roburin E, castalagin/vescalagin, gallic acid, valoneic acid bilactone, monogalloyl glucose, digalloyl glucose, trigalloyl glucose, ellagic acid rhamnose, quercitrin and ellagic acid.     

Measurement of deamidation of intact proteins by isotopic envelope and mass defect with ion cyclotron resonance Fourier transform mass spectrometry

After synthesis and folding, proteins undergo many post-synthetic modifications, including cleavage, oxidation, glycosylation, methylation, racemization, phosphorylation, and deamidation. Of these modifications, non-enymatic deamidation is the most prevalent. Each asparaginyl and glutaminyl residue in a protein is a miniature molecular clock that deamidates with a genetically determined half-time. These half-times vary from a few hours to more than a century, depending on a primary, secondary, tertiary, and quaternary structure near the amide residue. It has been suggested that these clocks regulate many biological processes. A few such processes have been discovered. These discoveries have been difficult because deamidation is inconvenient to measure. While most post-synthetic changes are easily measured by mass spectrometry, deamidation increases molecular mass by only one nominal Dalton, so the deamidated isotopic envelope overlaps the undeamidated isotopic envelope. While peptide deamidation rate determination through deconvolution of these envelopes has been accomplished for several hundred peptides, deconvolution becomes more difficult as the molecular weight increases. In high-resolution mass spectrometers, this deconvolution is possible for larger molecules and an alternative method based on the 19 mDa mass defect between the deamidated envelope and the isotopic envelope of protein fragments can also be utilized. We herein report a comparison of the envelope deconvolution and the mass defect methods for measurement of deamidation in human eye lens crystallins, with special emphasis on betaB2 crystallin and gammaS crystallin. Measurement of extent of deamidation of betaB2 crystallin in a 7 Tesla ion cyclotron resonance Fourier transform mass spectrometer is found to be accurate to a relative standard deviation in a single measurement of about 4% for each method. The envelope deconvolution method is further illustrated by detection of deamidation in intact gammaS crystallin, a 20 904 Da protein, and discovery of the principal gammaS deamidation site.     

Electrospray ionisation tandem mass spectrometry of poly

Evaluation of polymer end-capping reactions with the aid of electrospray ionisation tandem mass spectrometry techniques (ESI-MS(n)) allows characterisation of novel poly[(R, S)-3-hydroxybutanoic acid]-(a-PHB) telechelics, containing primary hydroxyl groups at both polymer chain ends. The chemical structures of individual mass-selected macromolecules of the well-defined a-PHB telechelics have been defined in this way, and fragmentation mechanisms have been proposed. Copyright 1999 John Wiley & Sons, Ltd.     

The formation of complexes by tetraethylresorcarene with planar guest ions studied by electrospray ionisation Fourier transform ion cyclotron resonance mass spectrometry

Resorcarenes have become a popular subject of study in the field of supramolecular chemistry. In this work the formation of host-guest complexes between a synthetic macrocyclic host, tetraethylresorcarene, and various eligible planar guests, was studied by mass spectrometric methods. The size and nature of the guest ion strongly influenced the complex formation. Collision-induced dissociation experiments revealed the fragmentation patterns of the resorcarene skeleton and the differences in fragmentation induced by the guest ions.     

Peptide polarity and the position of arginine as sources of selectivity during positive electrospray ionisation mass spectrometry

Electrospray ionisation (ESI) is a selective process and, for similar sized analytes, the intrinsic properties of the molecules affect the ionisation process and their response. This research sets out to determine the effect of some of these properties in peptides: peptide polarity and the presence of arginine at positions 1 and 4 in the amino acid sequence on the ESI response. Six peptides; molecular mass ranges 1.3-1.6 kDa; substance P (SP) and glutamate fibrinopeptide (GFP) and 3.2-3.7 kDa; calcitonin gene-related peptide (CGRP), vasoactive intestinal peptide (VIP), glucagon-like peptide 1 (GLP1) and defensin human neutropeptide 2 (DHNP2), were investigated. We have demonstrated that in positive ESI, for similar sized peptides and the same charge state, the responsiveness is in the order: Peptides with N or C terminal arginine > most non-polar peptides > least non-polar peptides. Therefore, arginine at the terminal position is a source of selectivity. Data from matrix-assisted laser desorption ionisation (MALDI) analysis supports that of the ESI experiments: Peptides with a terminal arginine residue generated higher signal intensities. Our observations extend our understanding of the ESI process and provide a rational approach to optimising sensitivity of electrospray conditions where a narrow mass range of peptides are poorly chromatographically resolved. This information will provide for a more effective method development process, especially during label-free quantitative determination of peptides extracted in solution.