Application of liquid chromatography/quadrupole time-of-flight mass spectrometry (LC-QqTOF-MS) in the environmental analysis

This paper gives an overview of the potentials of liquid chromatography-hybrid quadrupole time-of-flight mass spectrometry (LC-QqTOF) in the environmental analysis. Examples of applications of QqTOF instruments for target analysis of pharmaceuticals and pesticides are presented and discussed, as well as applications aimed on the identification of unknown compounds present in environmental waters or on the elucidation of structures of biodegradation and photodegradation products. Specific issues such as uncertainty of mass measurement and quantitative performances are discussed in details.     

Rapid identification of differentiation markers from whole epithelial cells by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry and statistical analysis

Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry (MALDI-TOFMS) was applied to identify markers for cellular differentiation. The differentiation of a human colon epithelial carcinoma T84 cell line was monitored over a period of 28 days by transepithelial electrical resistance (TER) measurements, alkaline phosphatase (AP) assay, and MALDI-TOF mass spectral fingerprints combined with statistical analysis. MALDI-MS generated specific mass spectral fingerprints characteristic of cell differentiation. Twenty-two ions were selected as diagnostic signals of fully differentiated T84 cells. Ten protein ion signals, detected by MALDI-MS and validated by statistical analysis, were proposed as T84 cell differentiation markers. Among these signals, ubiquitin was identified as a T84 cell differentiation marker by nanospray liquid chromatography/tandem mass spectrometry (nanoLC/MS/MS). Moreover, depending on the concentration of the cells seeded on the growth support, it was possible to predict the timing of the exponential phase and of cellular differentiation by MALDI-MS-derived marker ions. MALDI-TOFMS was compared to other methods for the determination of cellular differentiation: TER measurements are rapid but yield limited information as to the cellular differentiation state. AP assays are more specific for the differentiation state but take more time. By contrast, MALDI-MS has been found to be a fast, sensitive and precise method for cell differentiation assessment and provides the opportunity for multiplexing and high throughput. Moreover, the consumable costs per assay are very low.     

Characterisation of derivatised monomeric and prepolymeric isocyanates by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry and structural elucidation by tandem mass spectrometry

Isocyanates are an important class of compounds in occupational hygiene monitoring due mainly to their behaviour as respiratory sensitisers. Here, we demonstrate the application of matrix-assisted laser desorption/ionisation mass spectrometry (MALDI-MS) and MALDI tandem mass spectrometry (MS-MS) to the analysis of derivatised isocyanate monomers and prepolymers. The aim of the work has been to gauge the selectivity obtainable from the direct analysis of isocyanate mixtures without prior separation. Monomeric and prepolymeric isocyanate mixtures were analysed as their 1-(2-methoxyphenyl) piperazine derivatives and the potential of MALDI time-of-flight (ToF)-MS for an NCO monitoring program was assessed. The results obtained demonstrated the possibility of direct mixture analysis by this method. MALDI-MS-MS was used for the elucidation of fragment structures in the prepolymer samples. The developed methodology was then applied to the analysis of swabs from an occupational hygiene monitoring scheme and enabled the identification of the isocyanate species detected.     

Matrix-assisted laser desorption and ionisation time-of-flight mass spectrometry of Pt(II) and Pd(II) complexes

In this work, we have analysed matrix-assisted laser desorption and ionisation time-of-flight (MALDI-TOF) mass spectra of [PtCl₂(en)], [PtCl₂(dach)] and [PdCl(dien)]Cl acquired either with 2,5-dihydroxybenzoic acid (DHB) or α-cyano-hydroxycinnamic acid (CHCA) as matrices. For certain experiments, small amounts of trifluoro acetic acid (TFA) or higher concentration of inorganic salts (NaCl or KCl) was added to the matrix solution. The majority of peaks arising from the Pt(II) and Pd(II) complexes could be identified, but certain ions detectable in the spectra were generated upon ligand loss. Additionally, the analysis of Pt(II) complexes was also possible in the presence of a higher salt content, which is a commonly used analysis condition for the samples of biological origin. While DHB appears to be the best suited for MALDI-TOF mass spectrometric analysis of Pt(II) complexes, CHCA seems to be a better matrix for Pd(II) complex used in this study. On the other hand, small amounts of TFA improve the spectra quality of Pt(II) complexes, but lead most probably to the degradation of Pd(II) complex. Taken together, we have demonstrated that the analysis of metallo-drugs using MALDI-TOF MS, though accompanied with certain identification problems, is easy and reliable. On the other hand, having in mind that some complexes (i.e. a combination of a particular transition metal/ligand) cannot be analysed under conditions usually applied for others, we deem it necessary to find out the best conditions for MALDI-TOF MS analysis of each metal complex.     

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.     

The effect of solvent and matrix combinations on the analysis of bacteria by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

The ability to rapidly identify the taxonomic class of the wide variety of microorganisms involved in human and animal disease is becoming increasingly important, especially with the increasing development of resistance to the antibiotics which form the main defence against them. A number of groups have recognised the utility of matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry MALDI-TOF in the analysis of these microorganisms. However, no consistent methodology has been developed which is in general use. In particular the use of different solvent extraction systems and mass spectrometric matrices can have significant effects on the quality of the data obtained. We have now studied a number of the commonly used matrices and a range of solvent systems of widely varying polarity in an attempt to devise an optimum analytical strategy for the rapid characterisation of these organisms by MALDI-TOFMS. The E. coli ATCC 9637 organisms were initially washed to remove growth medium contaminants, followed by extraction with one of a range of solvents prior to admixing with a number of different single matrices or binary and ternary combinations of these matrices. The results obtained indicate that a binary combination of 2-(4-hydroxyphenylazo)benzoic acid and 2-mercaptobenzothiazole (1:1) as matrix provides the best data after the proteinaceous material from the organism cell surface was extracted with 17% formic acid, 33% isopropyl alcohol and 50% water, (solvent 2 in this work).     

Detection of organic products of polymer pyrolysis by thermogravimetry-supersonic jet-skimmer time-of-flight mass spectrometry (TG-Skimmer-SPI-TOFMS) using an electron beam pumped rare gas excimer VUV-light source (EBEL) for soft photo ionisation

A commercial thermogravimetry—supersonic jet-skimmer quadrupole mass spectrometer system (TG-Skimmer-QMS, Netzsch GmbH, Germany) was successfully converted for soft single photon ionisation time-of-flight mass spectrometric (SPI-TOFMS) detection of organic compounds. VUV light for SPI was generated by an electron beam pumped argon excimer light source (EBEL; E _photon = 9.8 eV). Furthermore, the versatility of the system was conserved, as high temperature TG and DSC measurements as well as electron ionisation mass spectrometry for the detection of inorganic compounds are still possible. The new system was tested with two polymers and a hydrocarbon mixture (diesel). It was demonstrated that aliphatic and aromatic organic compounds can be detected without fragmentation. Thus the system allows the recording of a readily interpretable organic signature of, e.g. thermal polymer decomposition. The thermal degradation of polystyrene shows a rich signature of the monomer, some oligomers and minor products of irregular cleavings of carbon chains. Polycarbonate exhibits a thermal decomposition fingerprint which is dominated by products of bisphenol A. The bisphenol A monomer, however, is also detectable.     

Absolute and relative quantification of sheep brain prion protein (PrP) allelic variants by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

Transmissible spongiform encephalopathies (TSEs) are characterised by the accumulation in the tissues of affected individuals of an abnormal form (PrP(Sc)) of a protein naturally produced by the host, the cellular prion protein (PrP(C)). In sheep, susceptibility to TSEs is tightly controlled by polymorphism at positions 136 (A or V), 154 (R or H) and 171 (R or Q) of the Prnp gene encoding the prion protein (PrP). Quantification of PrP variants at positions 136, 154 and 171 can be achieved by matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectrometric analysis of the respective peptides 114-139, 152-159 and 160-171 obtained after tryptic digestion of the PrP protein. In this study we quantified the tryptic peptide 114-139 containing the first polymorphic site. Quantification was either relative, between variants of this peptide, or absolute with respect to the C-terminally (18)O-labelled peptide obtained by hydrolysing known amounts of recombinant protein with trypsin in H(2) (18)O. After purification of PrP(C) and PrP(Sc) from the brain of two heterozygous sheep carrying either the ARQ/VRQ or ARR/VRQ genotypes, the proportion of each variant was measured. In the ARQ/VRQ animal, while both variants were equally represented in the normal isoform, the VRQ variant was predominantly found in the abnormal PrP protein, suggesting dissimilar behaviour of the two variants in the pathological process. The situation was even more contrasted in the ARR/VRQ animal where PrP(Sc) was solely composed of the VRQ variant. These two examples clearly illustrate the value of MALDI-TOF analysis, combined with appropriate immunopurification techniques, in seeking a precise understanding of the influence of PrP polymorphisms on TSE pathogenesis.     

Analysis of 8-aminonaphthalene-1,3,6-trisulfonic acid labelled N-glycans by matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry

Fluorophore-assisted carbohydrate electrophoresis (FACE) is a fast and efficient analytical method which is now widely used in glycobiology for the separation and quantification of free or glycoprotein-released oligosaccharides. However, since identification by FACE of N-glycan structures is only based on their electrophoretic mobility after labelling with 8-aminonaphthalene-1,3, 6-trisulfonic acid (ANTS), co-migration of derived glycans on gel could occur which may result in erroneous structural assignments. As a consequence, a protocol was developed for the fast and efficient matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) mass spectrometric analysis of ANTS-labelled N-glycans. N-Glycans were isolated from plant and mammalian glycoproteins, reductively aminated with the charged fluorophore 8-aminonaphthalene-1, 3, 6-trisulfonic acid (ANTS) and separated using high resolution polyacrylamide gel electrophoresis. The ANTS-labelled glycans were eluted from FACE gel slices and then analysed by MALDI-TOF mass spectrometry in negative ion mode. Using 3-aminoquinoline containing 2.5 mM citrate NH(4)(+) as matrix, neutral N-linked N-glycans, as well as labelled sialylated oligosaccharides, were found to be easily detected in the 2-10 picomole range giving rise to ?M - H(-) ions.     

Puff-by-puff resolved characterisation of cigarette mainstream smoke by single photon ionisation (SPI)-time-of-flight mass spectrometry (TOFMS): comparison of the 2R4F research cigarette and pure Burley, Virginia, Oriental and Maryland tobacco cigarettes

Soft single photon ionisation (SPI)-time-of-flight mass spectrometry (TOFMS) is applied for the characterisation and comparison of puff-by-puff resolved and total yields of cigarette mainstream smoke from single tobacco type cigarettes (Virginia, Oriental, Burley, and Maryland) and the 2R4F University of Kentucky research cigarette. Puff-by-puff characteristics of various smoke components within one cigarette type as well as between different cigarette types can differ tremendously. This is demonstrated by means of a few selected compounds. Puff yields vary between 15 and 106 μm for acetaldehyde, 6 and 57 μm for NO, and between 1 and 8 μm for butadiene. Thereby, cigarettes containing 100% Oriental and Burley tobacco exhibit a very unique behaviour for the first and last puff. Different cultivation and processing methods as well as burning characteristics are most likely responsible for this. Since the 2R4F cigarette contains all four tobacco types it combines features of all of them. However, for some smoke constituents, smoking of the 2R4F reference cigarette results in exceptionally high yields which might not be attributable to the four pure tobacco types, but to other factors. In addition, comparison of the different cigarettes was also carried out by normalising the yields to puff resolved particulate matter. This procedure minimises effects caused by unequal smoke formation and represents another approach in evaluating the data.     

Application of capillary electrochromatography (CEC) for the analysis of phenols in mainstream and sidestream tobacco smoke

Summary This paper describes the application of capillary electrochromatography (CEC) to the rapid quantitative analysis of individual mono- and dihydroxy phenols in tobacco smoke. The method derived provides reproducible and quantitative analysis of real samples, in significantly shorter times than are achieved by current GC and HPLC methods.     

Analysis of bacterial lipodepsipeptides by matrix-assisted laser desorption/ionisation time-of-flight and high-performance liquid chromatography with electrospray mass spectrometry

Strains of certain plant pathogenic bacteria, in particular several pathovars of Pseudomonas syringae, are known to produce cyclic lipodepsipeptides (LDPs) endowed with peculiar structural features and noticeable biological activities. In this study, a mass spectrometry procedure is proposed for screening LDP-producing bacterial strains and for identifying and assessing individual LDPs. After matrix-assisted laser desorption/ionisation time-of-flight (MALDI-TOF) screening of thirteen P. syringae strains for LDP production, the extracts from culture filtrates of eight positive strains were subjected to electrospray mass spectrometry for the identification of LDPs. Five strains were found to produce two forms of syringomycins (SR-E and SR-G) and two forms of syringopeptin 25 (SP25A and SP25B); two strains produced SR-E, SR-G and a new form of SP22; one strain produced syringotoxin (ST) and syringostatin A (SS-A) in addition to SP25A and SP25B. The yield in culture of two major LPDs: SR-G (3.2-13.8 mg x L(-1)) and SP25A (41.6-231.5 mg x L(-1)) was assessed by and high-performance liquid chromatography with electrospray mass spectrometry (HPLC/ESI-MS) in both scan and single ion monitoring (SIM) modes. Results of this investigation showed that the mass spectrometry protocol developed here is a precise and reliable method for screening bacterial strains for LDP production and for assessing the amount of each metabolite under various culture conditions. This could be of practical value in view of potential applications, e.g. biocontrol of post-harvest fungal diseases.     

Real-time analysis of aromatics in combustion engine exhaust by resonance-enhanced multiphoton ionisation time-of-flight mass spectrometry (REMPI-TOF-MS): a robust tool for chassis dynamometer testing

Resonance-enhanced multiphoton ionisation time-of-flight mass spectrometry (REMPI-TOF-MS) is a robust method for real-time analysis of monocyclic and polycyclic aromatic hydrocarbons in complex emissions. A mobile system has been developed which enables direct analysis on site. In this paper, we utilize a multicomponent calibration scheme based on the analytes' photo-ionisation cross-sections relative to a calibrated species. This allows semi-quantification of a great number of components by only calibrating one compound of choice, here toluene. The cross-sections were determined by injecting nebulised solutions of aromatic compounds into the TOF-MS ion source with the help of a HPLC pump. Then, REMPI-TOF-MS was implemented at various chassis dynamometers and test cells and the exhaust of the following vehicles and engines investigated: a compression ignition light-duty (LD) passenger car, a compression ignition LD van, two spark ignition LD passenger cars, 2 two-stroke mopeds, and a two-stroke engine of a string gas trimmer. The quantitative time profiles of benzene are shown. The results indicate that two-stroke engines are a significant source for toxic and cancerogenic compounds. Air pollution and health effects caused by gardening equipment might still be underestimated.     

Identification of sulfoglycolipids from the alga Porphyridium purpureum by matrix-assisted laser desorption/ionisation quadrupole ion trap time-of-flight mass spectrometry

Sulfoglycolipids, isolated from different phototrophic organisms, particularly plants and algae, have already been identified as bioactive compounds. In addition to their antiviral activity their influence on the immune response in mammalian cells is the focus of many studies. For the first time it has been possible to investigate purified sulfoquinovosyldiacylglycerols (SQDGs) from the microalga Porphyridium purpureum by matrix-assisted laser desorption/ionisation (MALDI) in the negative ion reflectron mode. Thereby, different solid and ionic liquid matrices have been tested to improve signal intensity during the laser ionisation. By using the MALDI Trap time-of-flight (ToF) multiple-stage (MS(n)) hybrid mass spectrometer the fatty acid compositions of the SQDGs were analysed by MS, and confirmed by MS(2) and MS(3) experiments. Thereby, hexadecanoic acid (C16:0), octadecadienoic acid (C18:2), eicosatetraenoic acid (C20:4), and eicosapentaenoic acid (C20:5) were detected in the purified fraction of SQDGs. The localisation of hexadecanoic acid (C16:0) at the sn-2 position, and unsaturated fatty acids at the sn-1 position of the SQDGs, determined by specific enzymatic hydrolysis, marks a procaryotic biosynthesis of SQDGs in the eucaryotic alga cells.     

Sequence determination in aliphatic poly(ester amide)s by matrix-assisted laser desorption/ionization time-of-flight and time-of-flight/time-of-flight tandem mass spectrometry

Poly(ester amide)s from dimethyl sebacate or sebacic acid and 2-aminoethanol or 4-amino-1-butanol were characterized by post-source decay matrix-assisted laser desorption/ionization time-of-flight (PSD-MALDI-TOF) and time-of-flight/time-of-flight tandem mass spectrometry (MALDI-TOF/TOF-MS/MS). Sodiated oligomers were selected as precursor ions for dissociation studies. PSD analysis was performed on dimethyl sebacate, dicarboxylic, carboxylic and amino alcohol, and diamino alcohol terminated oligomers. PSD-MALDI-TOF mass spectra yielded information on the fragmentation mechanisms of the poly(ester amide) chains, showing that the main cleavages proceed through a beta-hydrogen transfer rearrangement. MALDI-TOF/TOF-MS/MS provided structural information concerning ester/amide sequences in the polymer chains. As expected, together with the ions appearing in the PSD-MALDI mass spectrum, several new abundant fragment ions in the low-mass range are present in MALDI-TOF/TOF-MS/MS spectra. These new product ions proved to be diagnostic and made it possible to establish the presence of random sequences of ester and amide bonds in the poly(ester amide)s samples.     

Comparison of comprehensive two-dimensional gas chromatography/time-of-flight mass spectrometry and gas chromatography-mass spectrometry for the analysis of tobacco essential oils

A sample of tobacco essential oil was analyzed using gas chromatography-mass spectrometry (GC/MS) and comprehensive two-dimensional gas chromatography coupled to a time-of-flight mass spectrometry (GC × GC/TOFMS), respectively. In the GC/MS analysis, serially coupled columns were used. By comparing the GC/MS results with GC × GC/TOFMS results, many more components in the essential oil could be found within the two-dimensional separation space of GC × GC. The quantitative determination of components in the essential oil was performed by GC × GC with flame ionization detection (FID), using a method of multiple internal standards calibration.     

Characterisation of the intact rainbow trout vitellogenin protein and analysis of its derived tryptic and cyanogen bromide peptides by matrix-assisted laser desorption/ionisation time-of-flight-mass spectrometry and electrospray ionisation quadrupole/time-of-flight mass spectrometry

Vitellogenin (VTG) is a protein produced by the liver of oviparous animals. It is being used as a biomarker for exposure to endocrine disruptors in many species. Rainbow trout Vtg has recently been sequenced by the conventional cDNA nucleotide approach. We focused on protein characterization of the intact protein and its derived tryptic and cyanogen bromide peptides by matrix-assisted laser desorption/ionisation and electrospray ionisation mass spectrometry. The molecular mass of the intact protein was found to be 183127 Da. A large number of unidentified peptide ions encourage further structural analysis to propose possible sequence variants and post-translational modifications.     

Selection of internal standards for quantitative analysis by matrix-assisted laser desorption-ionization (MALDI) time-of-flight mass spectrometry

 The selection of an appropriate internal standard (IS) for quantification by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is critical for the successful application of quantitative MALDI. Selection of the IS depends on the chemical similarity of the analyte and IS and the mass separation of the analyte and IS as a function of instrumental peak resolution. For the quantification of bovine insulin, a series of internal standards including horse heart cytochrome C, bovine insulin chain B, des-pentapeptide human insulin, and des-octapeptide porcine insulin was investigated. Des-pentapeptide human insulin was found to be the most appropriate internal standard (relative standard deviation of the standard curve slope=2.99%, correlation coefficient=0.988 in the range of 0.5–0.4 μmol/L). Two methods for measuring of the MALDI signal intensity were evaluated, direct peak integration following subtraction of a linear background and non-linear least squares curve fitting. The results obtained with these methods were equivalent. Received: 10 November 1995 / Revised: 4 March 1996 / Accepted: 6 March 1996     

光电离质谱法测定卷烟抽吸过程中烟气成分在人体呼吸道的截留

采用光电离质谱法测定卷烟抽吸过程中烟气成分在人体呼吸道的截留.将烟气采样系统与光电离质谱相结合,通过取样毛细管的长度优化、烟气成分的光电离质谱图定性分析、吸入和呼出的流量数据优化,实现吸入和呼出烟气的直接采样和质谱检测,以计算卷烟抽吸过程中烟气成分的人体呼吸道截留率.结果表明,吸烟者经口腔循环抽吸卷烟时,烟气成分在人体...