Elimination of isocyanate and isothiocyanate molecules at the electrospray ionization ion trap, electrospray ionization quadrupole time-of-flight and matrix-assisted laser desorption/ionization time-of-flight tandem mass spectrometry fragmentation of sodium cationized brassitin, brassinin and their glycosides

Fragmentation mechanisms of phytoalexin analogs, including brassitin and brassinin and their glucosylated analogs, have been studied by electrospray (ESI) ion trap (IT) multistage (MS(n), n = 1-4) mass spectrometry, matrix-assisted laser desorption/ionization time-of-flight (MALDI ToF/ToF) and ESI-Q/ToF tandem mass spectrometry techniques. At the fragmentation of sodium adducts a hitherto not described process has been elucidated The proposed mechanism of this process includes cyclization of the brassitin and brassinin cationized adducts through a six-membered cycle of the molecules and the elimination of isocyanate or isothiocyanate from the thio- or dithiocarbamate moiety, giving rise to [M + Na - 43](+) or [M + Na - 59](+) adducts. The elimination of NH=C=O or NH=C=S molecules has been confirmed by the high resolution measurement of the elemental composition of the ions produced and quantum-chemical calculations of the six-membered transition state. Other fragmentation routes include cleavage of an alkane linker, while numerous characteristic hexopyranose pathways are taking place in the glucosylated compounds. The presented theoretical data on the ESI and MALDI behavior of the saccharidic, as well as of the indole aglycon parts, can facilitate structural elucidation of the analogous compounds.     

Metabolomics for unknown plant metabolites

In this article we discuss current trends in the techniques available for plant metabolomics. Chemical assignment of unknown metabolites leads to understanding of biosynthetic mechanisms at the gene level for genome-sequenced plants. Metabolomics using mass spectrometry has achieved innovative results in phytochemical genomics for primary and secondary metabolism in the model plant Arabidopsis thaliana by using publicly and commercially available information and standard compounds. However, finding a consolidated analytical technique for elucidation of structural information (e.g., elemental composition and structure) remains challenging. Recently, hyphenated analytical techniques and computer-assisted structural analysis with high-throughput and high-accuracy have been developing. Metabolite-driven approaches using such technology will be of central importance in phytochemical genomics.     

Rapid structural elucidation of composite bacterial hopanoids by atmospheric pressure chemical ionisation liquid chromatography/ion trap mass spectrometry

Bacteriohopanepolyols (BHPs) are membrane lipids produced by a wide range of eubacteria. Their use, however, as molecular markers of bacterial populations and processes has until recently been hampered by the lack of a suitable rapid method for fingerprinting their composition in complex environmental matrices. New analytical procedures employing ion trap mass spectrometry now allow us to investigate the occurrence of BHPs in diverse biological and environmental samples including bacterial cultures, soils, and recent and ancient sediments. Here, we describe the structural characterisation using atmospheric pressure chemical ionisation liquid chromatography/ion trap mass spectrometry (APCI-LC/MS(n)) of a number of previously identified but less commonly occurring BHPs such as adenosylhopane and ribonylhopane. Many of the structures described here have previously only been reported in one or just a small number of cultured organisms having been isolated from large amounts of cellular mass (4-26 g) and identified by nuclear magnetic resonance (NMR) techniques after purification of individual compounds. Now, having established characteristic APCI fragmentation patterns, it is possible to rapidly screen many more bacterial cultures using only small amounts of material (<50 mg) as well as environmental samples for these atypical structures and a rapidly growing suite of novel structures.     

Control of the Properties of Carbon Nanotubes Synthesized by CVD for Application in Electrochemical Biosensors

Interest in carbon nanotubes (CNT) has grown at a very rapid rate in the last decade. Their interesting physical and chemical properties open attractive possibilities in many application areas. These properties depend on the process conditions during synthesis and on subsequent purification steps. Recent studies have demonstrated that CNT can promote the electron transfer of biomolecules. These exceptional properties make them attractive for use in electrochemical biosensors. Multi walled nanotubes have been synthesized by the Chemical Vapor Deposition (CVD) method using methane as a carbon source and Ni–Al₂O₃–SiO₂ as the catalyst. The influence of the variation of certain reaction parameters such as feed gas composition, catalyst mass, temperature and reaction time in the yield of the CVD process has been established. In addition, the structural and chemical characteristics of the CNTs have been studied and a purification process to eliminate the catalyst and amorphous carbon has been developed that involves a gaseous oxidative process and acid treatment. The efficiency of the purification step has been determined by analytical techniques. Atomic force microscopy, Raman scattering, thermogravimetric analysis, inductively coupled plasma atomic spectroscopy are the characterization techniques employed in this work.     

Analysis of Organotin Compounds via a Thermabeam® LC–MS Interface

Selected organotin compounds, relating to antifouling paints, have been analysed using a particle beam interface system designed for use on liquid chromatography–mass spectrometry (LC–MS) instruments. The resultant mass spectra matched those obtained from conventional electron-impact (EI) techniques, and consistent data over several injections and different elution times were obtained. Data obtained from tributyltin, dibutyltin, monobutyltin, triphenyltin and diphenyltin (each as the chlorides) are presented. This interface has been shown to maintain sample and therefore spectral integrity for these compounds and is of potential use in further investigations relating to organotin environmental pollution.     

Synthesis and comparative structural investigation of crystalline and liquid-crystalline phases of low molecular mass compounds and side group polymers I. Low molecular mass compounds

Low molecular mass compounds and analogous side group polymers with a thermotropic phase behaviour crystalline-smectic liquid-crystalline-isotropic have been synthesized for a comparative structural study. As characteristic features of the compounds a biphenyl group has been chosen for the mesogenic core and alkoxy parts of various lengths as terminal and spacer groups. The phase behaviour has been studied with differential calorimetric (DSC) measurements and polarization microscopic observations. The low molecular mass compounds form crystalline phases at room temperature and exhibit predominantly mosaic textures in the polarization microscope at elevated temperatures indicating high order of the packing of the molecules. The arrangements of the molecules in the crystalline and liquid-crystalline phase can best be described as layered structures according to X-ray diffraction measurements. A structural analysis of a solution grown single crystal provides valuable information on the conformation and packing of the compounds investigated in this study.     

Relative molecular mass information from aliphatic nitro compounds: A chemical ionization study

The mass spectra of a number of aliphatic nitro compounds have been studied using electron Ionization (EI) and a variety of chemical Ionization (CI) techniques in attempts to obtain relative molecular mass information. The use of positive ion ammonia chemical Ionization techniques gave very satisfactory results, providing abundant [M + NH4]⁺ ions, not only from both primary and secondary nitro compounds, but also from the much more labile tertiary nitro compounds. However, the use of methane and isobutane positive ion CI or EI conditions resulted in facile fragmentation with little relative molecular mass information being made available. Negative ion CI using methane, isobutane or ammonia as moderating gases all gave abundant [M ? 1]^? ions with primary and secondary nitro compounds but at much reduced sensitivity.     

Identification of the major metabolites of tectorigenin in rat bile by liquid chromatography combined with time-of-flight and ion trap tandem mass spectrometry

A novel methodology for the identification of tetorigenin and its metabolites in rat bile has been created using liquid chromatography (LC) combined with time-of-flight (TOF) and ion trap multiple mass spectrometry (IT-MSn). As a means to discriminate amongst unknown organic compounds in complex biological matrices, the proposed methodology relies upon the combination of LC/TOF-MS to provide accurate mass measurements in generating a molecular formula while benefiting from the complementary structural information provided by the LC/IT-MSn. In this study, the combined approach has been applied to the metabolic fingerprinting chromatograms of rat bile samples before and after tectorigenin administration. All possible metabolites are investigated based on accurate mass data and isotope function using LC/TOF-MS and structural confirmation using LC/IT-MSn. Seven phase II metabolities of tectorigenin in rat bile have been successfully elucidated using this novel LC approach and are being reported for the first time.     

Identification of unknown compounds using data bases and computer simulation of mass spectra

The possibility of identification based on the comparison of experimental electron-ionization mass spectrum of an unknown (in our case, model) compound with the mass spectra of the candidate compounds generated by the Mass Frontier software has been demonstrated by the example of three model compounds. The structural isomers of the identified substances found in the ChemExper database have been used as the candidate compounds. The candidate substances have been ranged by the degree of similarity between their simulated mass spectra and the experimental mass spectrum of the unknown compound. The mass spectra have been compared on the basis of the algorithm used in the NIST MS Search standard search system. In all three cases, the sought-after structure has been indicated as the most probable one of all the candidate structures.     

Temperature-Controlled Electrospray Ionization: Recent Progress and Applications

Native electrospray ionization (ESI) and nanoelectrospray ionization (nESI) allow researchers to analyze intact biomolecules and their complexes by mass spectrometry (MS). The data acquired using these soft ionization techniques provide a snapshot of a given biomolecules structure in solution. Over the last thirty years, several nESI and ESI sources capable of controlling spray solution temperature have been developed. These sources can be used to elucidate the thermodynamics of a given analyte, as well as provide structural information that cannot be readily obtained by other, more commonly used techniques. This review highlights how the field of temperature-controlled mass spectrometry has developed.     

Flow injection spectroscopic analysis of model drugs using on-line UV-diode array, FT-infrared and 1H-nuclear magnetic resonance spectroscopy and time-of-flight mass spectrometry

A prototype flow injection analysis (FIA) system for the characterisation of compounds via a combination of diode array UV, 1H NMR, FT-IR spectroscopy and time-of-flight (TOF) mass spectrometry has been investigated using a number of pharmaceuticals and related compounds as model compounds. This combination of spectrometers allowed the on-flow collection of UV, 1H NMR, IR and mass spectra together with atomic composition data, enabling almost complete structural characterisation to be performed. Practical detection limits with the current system were in the region of 50 micrograms, however, the use of state of the art spectrometers would result in a significant reduction in the amount of material required.     

The application of gas chromatography/atmospheric pressure chemical ionisation time‐of‐flight mass spectrometry to impurity identification in Pharmaceutical Development

Accurate mass measurement (used to determine elemental formulae) is an essential tool for impurity identification in pharmaceutical development for process understanding. Accurate mass liquid chromatography/mass spectrometry (LC/MS) is used widely for these types of analyses; however, there are still many occasions when gas chromatography (GC)/MS is the appropriate technique. Therefore, the provision of robust technology to provide accurate mass GC/MS (and GC/MS/MS) for this type of activity is essential. In this report we describe the optimisation and application of a newly available atmospheric pressure chemical ionisation (APCI) interface to couple GC to time‐of‐flight (TOF) MS. To fully test the potential of the new interface the APCI source conditions were optimised, using a number of standard compounds, with a variety of structures, as used in synthesis at AstraZeneca. These compounds were subsequently analysed by GC/APCI‐TOF MS. This study was carried out to evaluate the range of compounds that are amenable to analysis using this technique. The range of compounds that can be detected and characterised using the technique was found to be extremely broad and include apolar hydrocarbons such as toluene. Both protonated molecules ([M + H]⁺) and radical cations (M^+.) were observed in the mass spectra produced by APCI, along with additional ion signals such as [M + H + O]⁺. The technique has been successfully applied to the identification of impurities in reaction mixtures from organic synthesis in process development. A typical mass accuracy of 1–2 mm/zunits (m/z 80–500) was achieved allowing the reaction impurities to be identified based on their elemental formulae. These results clearly demonstrate the potential of the technique as a tool for problem solving and process understanding in pharmaceutical development. The reaction mixtures were also analysed by GC/electron ionisation (EI)‐MS and GC/chemical ionisation (CI)‐MS to understand the capability of GC/APCI‐MS relative to these two firmly established techniques. Copyright © 2010 John Wiley & Sons, Ltd.     

Mass spectrometry and combinatorial chemistry: a short outline.

The rapid evolution of combinatorial chemistry in recent years has led to a dramatic improvement in synthetic capabilities. The goal is to accelerate the discovery of molecules showing affinity against a target, such as an enzyme or a receptor, through the simultaneous synthesis of a great number of structurally diverse compounds. This is done by generating combinatorial libraries containing as many as hundreds or thousands of compounds. The need to test all these compounds led to the development of high-throughput screening (HTS) techniques, and also high-throughput analytical techniques capable of assessing the occurrence, structure and purity of the products. In order to be applied effectively to the characterization of combinatorial libraries, an analytical technique must be adequately sensitive (to analyse samples which are typically produced in nanomole amounts or less), fast, affordable and easy to automate (to minimize analysis time and operator intervention). Although no method alone can meet all the analytical challenges underlying this task, the recent progress in mass spectrometric (MS) instrumentation renders this technique an essential tool for scientists working in this area. We describe here relevant aspects of the use of MS in combinatorial technologies, such as current methods of characterization, purification and screening of libraries. Some examples from our laboratory deal with the analysis of pooled oligomeric libraries containing n x 324(n = 1, 2) compounds, using both on-line high-performance liquid chromatography/MS with an ion trap mass spectrometer, and direct infusion into a triple quadrupole instrument. In the first approach, MS and product ion MS/MS with automatic selection of the precursor were performed in one run, allowing library confirmation and structural elucidation of unexpected by-products. The second approach used MS scans to characterize the entire library and also precursor ion and neutral loss scans to detect selectively components with given structural characteristics.     

Determination of alkyl benzyl and dialkyl dimethyl quaternary ammonium biocides in occupational hygiene and environmental media by liquid chromatography with electrospray ionisation mass spectrometry and tandem mass spectrometry

A new method for the simultaneous qualitative and quantitative determination of alkyl benzyl and dialkyl quaternary ammonium compounds (QACs) has been developed. Analysis is by reversed-phase high-performance liquid chromatography coupled with electrospray ionisation mass spectrometry. QACs are extremely amenable to the electrospray ionisation technique (limit of detection of BAC C12 homologue 3 ng ml(-1)). The selectivity of mass spectrometric detection allows simultaneous determination of benzyl and dialkyl dimethyl ammonium compounds. The method was successfully applied to the analysis of real samples (occupational hygiene sampling devices, products and swimming pool water). Structural information was obtained by MS-MS and cone voltage ion dissociation techniques. Ion dissociation enabled the structural elucidation of an unknown quaternary ammonium compound present in a commercial formulation.     

Thin layer chromatography/mass spectrometry

Thin layer chromatography (TLC)--a simple, cost-effective, and easy-to-operate planar chromatographic technique--has been used in general chemistry laboratories for several decades to routinely separate chemical and biochemical compounds. Traditionally, chemical and optical methods are employed to visualize the analyte spots on the TLC plate. Because direct identification and structural characterization of the analytes on the TLC plate through these methods are not possible, there has been long-held interest in the development of interfaces that allow TLC to be combined with mass spectrometry (MS)--one of the most efficient analytical tools for structural elucidation. So far, many different TLC-MS techniques have been reported in the literature; some are commercially available. According to differences in their operational processes, the existing TLC-MS systems can be classified into two categories: (i) indirect mass spectrometric analyses, performed by scraping, extracting, purifying, and concentrating the analyte from the TLC plate and then directing it into the mass spectrometer's ion source for further analysis; (ii) direct mass spectrometric analyses, where the analyte on the TLC plate is characterized directly through mass spectrometry without the need for scraping, extraction, or concentration processes. Conventionally, direct TLC-MS analysis is performed under vacuum, but the development of ambient mass spectrometry has allowed analytes on TLC plates to be characterized under atmospheric pressure. Thus, TLC-MS techniques can also be classified into two other categories according to the working environment of the ion source: vacuum-based TLC-MS or ambient TLC-MS. This review article describes the state of the art of TLC-MS techniques used for indirect and direct characterization of analytes on the surfaces of TLC plates.     

LC-MS analysis in the aquatic environment and in water treatment technology – a critical review

Environmental contaminants of recent concern are pharmaceuticals, estrogens and other endocrine disrupting chemicals (EDC) such as degradation products of surfactants, algal and cyanobacterial toxins, disinfection by-products (DBPs) and metalloids. In addition, pesticides (especially their transformation products), microorganisms, and humic substances (HS), in their function as vehicles for contaminants and as precursors for by-products in water treatment, traditionally play an important role. The present status of the application of LC-MS techniques for these water constituents are discussed and examples of application are given. Solid-phase extraction with various non-selective materials in combination with liquid chromatography (LC) on reversed-phase columns have been the most widely used methods for sample preconcentration and separation for different compound classes like pesticides, pharmaceuticals or estrogens. Electrospray ionization (ESI) and atmospheric pressure ionization (APCI) are the most frequently used ionization techniques for polar and ionic compounds, as well as for less polar non-ionic ones. The facilities of LC-MS have been successfully demonstrated for different compound classes. Polar compounds from pharmaceuticals used as betablockers, iodinated X-ray contrast media, or estrogens have been determined without derivatization down to ultratrace concentrations. LC-MS can be viewed as a prerequisite for the determination of algal and cyanobacterial toxins and the homologues and oligomers of alkylphenol ethoxylates and their metabolites. Tandem mass spectrometric techniques and the use of diagnostic ions reveal their usefulness for compound-class specific screening and unknown identification, and are also valid for the analysis of pesticides and especially for their transformation products. Structural information has been gained by the application of LC-MS methods to organometallic species. New insights into the structural variety of humic substances have been made possible by FT-ICR-MS due to its ultrahigh mass resolution. Finally, exciting possibilities for rapid detection and identification of microorganisms have been made possible by MALDI and LC-MS methods.     

Distinguishing N-oxide and hydroxyl compounds: impact of heated capillary/heated ion transfer tube in inducing atmospheric pressure ionization source decompositions

In the pharmaceutical industry, a higher attrition rate during the drug discovery process means a lower drug failure rate in the later stages. This translates into shorter drug development time and reduced cost for bringing a drug to market. Over the past few years, analytical strategies based on liquid chromatography/mass spectrometry (LC/MS) have gone through revolutionary changes and presently accommodate most of the needs of the pharmaceutical industry. Among these LC/MS techniques, collision induced dissociation (CID) or tandem mass spectrometry (MS/MS and MS(n)) techniques have been widely used to identify unknown compounds and characterize metabolites. MS/MS methods are generally ineffective for distinguishing isomeric compounds such as metabolites involving oxygenation of carbon or nitrogen atoms. Most recently, atmospheric pressure ionization (API) source decomposition methods have been shown to aid in the mass spectral distinction of isomeric oxygenated (N-oxide vs hydroxyl) products/metabolites. In previous studies, experiments were conducted using mass spectrometers equipped with a heated capillary interface between the mass analyzer and the ionization source. In the present study, we investigated the impact of the length of a heated capillary or heated ion transfer tube (a newer version of the heated capillary designed for accommodating orthogonal API source design) in inducing for-API source deoxygenation that allows the distinction of N-oxide from hydroxyl compounds. 8-Hydroxyquinoline (HO-Q), quinoline-N-oxide (Q-NO) and 8-hydroxyquinoline-N-oxide (HO-Q-NO) were used as model compounds on three different mass spectrometers (LCQ Deca, LCQ Advantage and TSQ Quantum). Irrespective of heated capillary or ion transfer tube length, N-oxides from this class of compounds underwent predominantly deoxygenation decomposition under atmospheric pressure chemical ionization conditions and the abundance of the diagnostic [M + H - O](+) ions increased with increasing vaporizer temperature. Furthermore, the results suggest that in API source decompostion methods described in this paper can be conducted using mass spectrometers with non-heated capillary or ion transfer tube API interfaces. Because N-oxides can undergo in-source decomposition and interfere with quantitation experiments, particular attention should be paid when developing API based bioanalytical methods.     

Characterization and differentiation of heterocyclic isomers. Part 2. Mass spectrometry and molecular orbital calculations on pyrrolo[1,2-a][1,4]benzodiazepin-4-one, -6-one,and -4,6-dione

Pyrrolo[1,2-a][1,4]benzodiazepin-4-one (1), -6-one (2), and -4,6-dione (3), which are starting materials for the synthesis of pharmacologically interesting compounds that are active as neurotropic agents, have been characterized in the gas phase. The application of different mass spectrometric techniques, such as electron ionization, high-resolution, and tandem mass spectrometry, has allowed the structural characterization and differentiation of their molecular ions and most abundant fragment ions formed in the source. In particular, the two positional isomers 1 and 2 produce quite different mass spectra, and their molecular and the most intense fragment ions yield different metastable mass-analyzed ion kinetic energy spectra. Furthermore, high-resolution mass spectrometry and accurate mass measurements have revealed different elemental compositions and abundances for isobaric fragment ions produced by isomers 1 and 2. From these data and from the comparison with those relevant to compound 3, it has been possible to evaluate the influence of the position of the carbonyl group on the fragmentation pathways. Semiempirical molecular orbital calculations carried out by both the modified neglect of differential overlap and Austin 1 methods have provided useful information on the characterization of the neutrals as well as the molecular ions of compounds 1–3.     

A comparison of accurate mass techniques for the structural elucidation of fluconazole

Mass spectrometry plays a major role in the structural elucidation and characterisation of drug candidates and related substances. Accurate mass data allow the mathematical prediction of molecular formula of both precursor and fragment ions. In this paper, a comparison of the accurate mass data obtained for the fragmentation of fluconazole, an antifungal drug, by three different methods is made: electron ionisation (EI) using a magnetic sector instrument; electrospray ionisation (ES) using a Fourier transform ion cyclotron mass spectrometer (FTICRMS); and ES using a quadrupole-time-of-flight mass spectrometer (Q-ToF). It is clear from the data obtained that mass accuracy is not simply a function of instrument resolution. The subtle differences observed between collisionally activated dissociation (CAD) and sustained off-resonance collisionally activated dissociation (SORI-CAD) spectra are explained as a consequence of the excitation process. The advantages and disadvantages of the three techniques are discussed within the context of structural elucidation.     

Thermal properties of different transition metal forms of montmorillonite intercalated with mono-, di-, and triethanolammonium compounds

In the present study, different transition metal forms of montmorillonite have been intercalated with mono-, di-, and triethanolammonium cations via d coordination mechanism to investigate their thermal behavior, structural characteristics, surface properties, and elemental composition using TG, XRD, BET, and CHNS techniques. Thermogravimetric analysis showed two thermal transition steps for transition metal-exchanged montmorillonites, which attributed to desorption of the physically adsorbed water and hydrated water, and dehydroxylation of the structural water; whereas for ammonium-montmorillonite complexes, the TG curves showed three thermal transition steps which attributed to desorption of the adsorbed water and dehydration, decomposition of the ammonium cations in the interlayer space of montmorillonite, and the dehydroxylation of the structural water. The thermal analysis of ammonium-montmorillonites affirmed that the molar mass of amine compounds used affects both desorption temperature (position) and the amount of the adsorbed water (intensity). XRD results revealed that the molar mass of amine used has linear relation with the basal spacings of the corresponding ammonium-montmorillonites, indicating structural changes. BET results showed that the molar mass of amines has an inverse effect on the surface area of the studied samples. CHNS analysis for the studied samples quantitatively confirmed the intercalation of ammonium cations into the interlayer space of montmorillonite.