Analysis of Volatile and Oxidation Sensitive Compounds Using a Cold Inlet System and Electron Impact Mass Spectrometry

This video presents a protocol for the mass spectrometrical analysis of volatile and oxidation sensitive compounds using electron impact ionization. The analysis of volatile and oxidation sensitive compounds by mass spectrometry is not easily achieved, as all state-of-the-art mass spectrometric methods require at least one sample preparation step, e.g., dissolution and dilution of the analyte (electrospray ionization), co-crystallization of the analyte with a matrix compound (matrix-assisted laser desorption/ionization), or transfer of the prepared samples into the ionization source of the mass spectrometer, to be conducted under atmospheric conditions. Here, the use of a sample inlet system is described which enables the analysis of volatile metal organyls, silanes, and phosphanes using a sector field mass spectrometer equipped with an electron impact ionization source. All sample preparation steps and the sample introduction into the ion source of the mass spectrometer take place either under air-free conditions or under vacuum, enabling the analysis of compounds highly susceptible to oxidation. The presented technique is especially of interest for inorganic chemists, working with metal organyls, silanes, or phosphanes, which have to be handled using inert conditions, such as the Schlenk technique. The principle of operation is presented in this video.     

Selective determination of haloperidol in human serum: surface ionization mass spectrometry and gas chromatography with surface ionization detection

Surface ionization organic mass spectrometry (SIOMS) has been performed on the clinically important drug haloperidol using quadrupole mass spectrometry in which the thermal ion source has a rhenium oxide emitter. The surface ionization (SI) mass spectrum is presented, interpreted in a purely empirical way by means of evidence from previous investigations, and then compared to results from conventional electron impact (EI) ionization. An approach to detection of this drug in serum by gas chromatography (GC) with a surface ionization detector (SID) and GC-SIOMS is described. This approach demonstrates that (a) haloperidol is efficiently surface-ionized, giving a unique SI mass spectrum, (b) experimental results rationalize the combined sensitivity and selectivity of the GC-SID for the examined drug, (c) the detection limit for haloperidol in serum is 1.1 ng/ml (S/N = 3) by GC-SID (the coefficients of variation of the assay are generally low, i.e. below 8.5%) and (d) the GC-SIOMS coupling can be used for sensitive and selective detection of haloperidol in serum.     

Ultra-performance liquid chromatography–tandem mass spectrometry analysis of the bioactive components and their metabolites of Shaofu Zhuyu decoction active extract in rat plasma

A rapid, sensitive and selective ultra-performance liquid chromatography–electrospray ionization tandem mass spectrometric (UPLC–ESI-MS/MS) method was developed for analysis and identification of the bioactive components and their metabolites in rat plasma following oral administration Shaofu Zhuyu decoction active extract. The analysis was carried out on an AcQuity? UPLC chromatographic instrument and a QTOF mass spectrometer using positive and negative electrospray ionization (ESI), respectively. The results showed that sixteen peaks were detected and twelve peaks, including flavones, organic acids and terpene glycosides, were identified by comparing with reference compounds. Furthermore, nine metabolites, including quercetin glucuronide sulfates, quercetin diglucuronides, isorhamnetin sulfates, isorhamnetin glucosides, and isorhamnetin glucuronides were detected and identified in rat plasma based on the mass fragmentation behaviors and literature reports. These results provided a meaningful basis for evaluating the bioactive components and their action mechanisms of complex traditional Chinese medicines (TCMs).     

Desorption/ionization on silicon for small molecules:a promising alternative to MALDI TOF

A method has been developed for laser desorption/ionization of catecholamines from porous silicon. This methodology is particularly attractive for analysis of small molecules. MALDI TOF mass spectrometry, although a very sensitive technique, utilizes matrices that need to be mixed with the sample prior to their analysis. Each matrix produces its own background, particularly in the low-molecular mass region. Therefore, detection and identification of molecules below 400 Da can be difficult. Desorption/ionization of samples deposited on porous silicon does not require addition of a matrix, thus, spectra in the low-molecular mass region can be clearly readable. Here, we describe a method for the analysis of catecholamines. While MALDI TOF is superior for proteomics/peptidomics, desorption/ionization from porous silicon can extend the operating range of a mass spectrometer for studies on metabolomics (small organic molecules and their metabolites, such as chemical neurotransmitters, prostaglandins, steroids, etc.).     

Determination of long-chain fatty acid acyl-coenzyme A compounds using liquid chromatography-electrospray ionization tandem mass spectrometry

Acyl-CoAs have important role in fat and glucose metabolism of the cells. In this study we have developed an on-line HPLC-ESI-MS/MS method for determination of long-chain acyl-CoA compounds in rat liver samples. Six long-chain acyl-CoAs (C16:0, C16:1, C18:0, C18:1, C20:0 and C20:4) were separated with a C4 reversed-phase column using triethylamine acetate and acetonitrile gradient. Negative electrospray ionization is very suitable for acyl-CoA compounds and excellent MS/MS spectra for long-chain acyl-CoAs can be obtained. MS/MS method with an ion trap mass spectrometer makes it possible to identify and quantitate individual acyl-CoAs simultaneously. The method proved to be sensitive enough for determination of all compounds of interest using 0.4-0.7 g of tissue and was validated in the range of 0.1-15.0 pmol/microl.     

Mass spectrometric localization of carbon-carbon double bonds: A critical review of recent methods

Current methods are reviewed for determining the position of double bonds in fatty acids, and other unsaturated organic compounds, using mass spectrometry. ‘On-site’ and ‘remote-site’ derivatization methods are described, and their advantages and disadvantages for mass spectrometric analysis discussed. Chemical transformation of double bonds by methoxylation, silyloxylation or deuteration, together with electron impact (EI), chemical ionization (CI) or collisionally induced decomposition (CID) techniques in combination with fast atom bombardment (FAB) or CI, are found to be most suitable for polyunsaturated fatty acids (PUFA): for the analysis of less unsaturated compounds on a submicrogram scale those methods are most promising which either do not involve derivatization of the double bonds, or give derivatives in quantitative yields. Effects of mass spectrometer geometry and operating conditions are also considered.     

Quantification of zolmitriptan in plasma by high-performance liquid chromatography-electrospray ionization mass spectrometry

A sensitive and specific liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) method has been developed and validated for the identification and quantification of zolmitriptan in human plasma. After the addition of the internal standard (IS) and 1.0 M sodium hydroxide solution, plasma samples were extracted with methylene chloride:ethyl acetate mixture (20:80, v/v). The organic layer was evaporated under a stream of nitrogen at 40 degrees C. The residue was reconstituted with 100 microl mobile phase. The compounds were separated on a prepacked Lichrospher CN (5 microm, 150 mm x 2.0 mm) column using a mixture of methanol:water (10 mM NH(4)AC, pH 4.0) = 78:22 as mobile phase. Detection was performed on a single quadrupole mass spectrometer by selected ion monitoring (SIM) mode via electrospray ionization (ESI) source. The method was proved to be sensitive and specific by testing six different plasma batches. Linearity was established for the range of concentrations 0.30-16.0 ng/ml with a coefficient of determination (r) of 0.9998 and good back-calculated accuracy and precision. The intra- and inter-day precision (R.S.D.%) were lower than 15% and accuracy ranged from 85 to 115%. The lower limit of quantification was identifiable and reproducible at 0.30 ng/ml. The proposed method enables the unambiguous identification and quantification of zolmitriptan for pharmacokinetic, bioavailability or bioequivalence studies.     

Chemical characterization of dissolved organic matter in moist acidic tussock tundra soil using ultra-high resolution 15T FT-ICR mass spectrometry

Global warming is considered one of the most serious environmental issues, substantially mediating abrupt climate changes, and has stronger impacts in the Arctic ecosystems than in any other regions. In particular, thawing permafrost in the Arctic region with warming can be strongly contributing the emission of greenhouse gases (CO₂ and CH₄) that are produced from microbial decomposition of preserved soil organic matter (SOM) or are trapped in frozen permafrost soils, consequently accelerating global warming and abrupt climate changes. Therefore, understanding chemical and physical properties of permafrost SOM is important for interpreting the chemical and biological decomposability of SOM. In this study, we investigated dissolved organic matter (DOM) along the soil depth profile in moist acidic tussock tundra to better understand elemental compositions and distributions of the arctic SOM to evaluate their potential decomposability under climate change. To achieve ultra-high resolution mass profiles, the soil extracts were analyzed using a 15 Tesla Fourier transform ion cyclotron resonance mass spectrometer in positive and negative ion modes via electrospray ionization. The results of this analysis revealed that the deeper organic soil (2Oe1 horizon) exhibits less CHON class and more aromatic class compounds compared to the surface organic soils, thus implying that the 2Oe1 horizon has undergone a more decomposition process and consequently possessed the recalcitrant materials. The compositional features of DOM in the Arctic tundra soils are important for understanding the changes in biogeochemical cycles caused from permafrost changes associated with global warming and climate change.     

Trace determination and isotopic analysis of the elements in life sciences by mass spectrometry

The main ionization methods in a mass spectrometer for isotope ratio determinations of the elements are discussed in this review. These methods are thermal ionization, spark source, electron impact, inductively coupled plasma and field desorption. As concerns thermal ionization, electron impact and field desorption, a survey of the possibilities of isotope analyses in the periodic table of the elements is given. Besides kinetic studies, trace element determination by isotope dilution technique is the main application for isotope ratio measurements of the elements. The definitive method, isotope dilution mass spectrometry, is discussed as a potential tool for achieving accurate and precise trace analyses. Using field desorption mass spectrometry, one example of calcium kinetics in man and one example of thallium trace determination in an animal tissue are given. Other metal trace analyses with the isotope dilution technique are presented for biological and medical samples using positive thermal ionization mass spectrometry. Negative thermal ions are formed for the mass spectrometric analysis of non-metals and non-metal compounds in food samples, e.g. for iodine and nitrate in milk powder. Preliminary results with the isotope dilution technique are presented for a new quadrupole thermal ionization mass spectrometer which is a low-cost instrument and can be easily handled.     

Direct protein detection from biological media through electrospray-assisted laser desorption ionization/mass spectrometry

We report here using a novel technology-electrospray-assisted laser desorption ionization (ELDI)/mass spectrometry-for the rapid and sensitive detection of the major proteins that exist in dried biological fluids (e.g., blood, tears, saliva, serum), bacterial cultures, and tissues (e.g., porcine liver and heart) under ambient conditions. This technique required essentially no sample pretreatment. The proteins in the samples were desorbed using a pulsed nitrogen laser without the assistance of an organic matrix. The desorbed protein molecules were then post-ionized through their fusion into the charged solvent droplets produced from the electrospray of an acidic methanol solution; electrospray ionization (ESI) proceeded from the newly formed droplets to generate the ESI-like protein ions. This new ionization approach combines some of the features of electrospray ionization with those of matrix-assisted laser desorption ionization (MALDI), that is, sampling of a solid surface with spatial resolution, generating ESI-like mass spectra of the desorbed proteins, and operating under ambient conditions.     

Pyrolysis experiment of simulated exogenous complex organics synthesized from the gas mixtures of CO, NH3, and H2O by 3 MeV proton irradiation.

High molecular weight organic matter synthesized from mixtures of carbon monoxide, ammonia and water gases similar to those found in the interstellar medium were irradiated with a 3 MeV proton beam and analyzed by Curie point pyrolysis with detection by gas chromatograph and mass spectrometer (Pyr-GC-MS). A wide variety of organic compounds, not only a number of amide compounds, but also heterocyclic and polycyclic aromatic hydrocarbons (PAHs), were detected among the products of the pyrolysis. The present data shows that primary and primitive organic matter serving as precursors to bioorganic compounds such as amino acids, nucleic acid bases and sugar might have been formed in a gaseous mixture of similar composition to that of the interstellar dust environment.     

Analysis of the Proteinaceous Components of the Organic Matrix of Calcitic Sclerites from the Soft Coral Sinularia sp.

An organic matrix consisting of a protein-polysaccharide complex is generally accepted as an important medium for the calcification process. While the role this “calcified organic matrix” plays in the calcification process has long been appreciated, the complex mixture of proteins that is induced and assembled during the mineral phase of calcification remains uncharacterized in many organisms. Thus, we investigated organic matrices from the calcitic sclerites of a soft coral, Sinularia sp., and used a proteomic approach to identify the functional matrix proteins that might be involved in the biocalcification process. We purified eight organic matrix proteins and performed in-gel digestion using trypsin. The tryptic peptides were separated by nano-liquid chromatography (nano-LC) and analyzed by tandem mass spectrometry (MS/MS) using a matrix-assisted laser desorption/ionization (MALDI) – time-of-flight-time-of-flight (TOF-TOF) mass spectrometer. Periodic acid Schiff staining of an SDS-PAGE gel indicated that four proteins were glycosylated. We identified several proteins, including a form of actin, from which we identified a total of 183 potential peptides. Our findings suggest that many of those peptides may contribute to biocalcification in soft corals.     

Metabolic profiling by ion mobility mass spectrometry (IMMS)

Ion Mobility Mass Spectrometry (IMMS) was evaluated as an analytical method for metabolic profiling. The specific instrument used in these studies was a direct infusion (DI)-electrospray ionization (ESI)—ambient pressure ion mobility spectrometer (APIMS) coupled to a time-of-flight mass spectrometer (TOFMS). The addition of an ion mobility spectrometer to a mass spectrometer had several advantages over direct infusion electrospray mass spectrometry alone. This tandem instrument (ESI-IMMS) added a rapid separation step with high-resolution prior to mass spectrometric analysis of metabolite mixtures without extending sample preparation time or reducing the high through put potential of direct mass spectrometry. Further, IMMS also reduced the baseline noise common with ESI-MS analyses of complex samples and enabled rapid separation of isobaric metabolites. IMMS was used to analyze the metabolome of Escherichia coli (E. coli), containing a collection of extremely diverse chemical compounds including hydrophobic lipids, inorganic ions, volatile alcohols and ketones, amino and non-amino organic acids, and hydrophilic carbohydrates. IMMS data were collected as two-dimensional spectra showing both mobility and mass of each ion detected. Using direct infusion ESI-IMMS of a non-derivatized methanol extract of an E. coli culture, more than 500 features were detected, of which over 200 intracellular metabolites were tentatively assigned as E. coli metabolites. This analytical method also allowed simultaneous separation of isomeric metabolic features.     

Applications of forensic chemistry to environmental work

Recent trends in environmental remediation have increasingly employed the use of environmental chemistry techniques to decipher the source(s) and fate of the contaminants and, in some cases, to determine their age or apportion them to sources. An extensive database of pyrogenic and petrogenic 'chemical fingerprints' has been constructed by the Gas Technology Institute (GTI) and META Environmental, Inc. using gas chromatography coupled with a flame ionization detector (GC/FID) or with a mass spectrometer (GC/MS). The use of these chemical fingerprinting techniques have been highly successful in discerning wastes from wholly different sources as well as among Manufactured Gas Plant (MGP)-type wastes from different plant operations. However, these techniques have been limited when low-level polycyclic aromatic hydrocarbon (PAH) discernment is required. Specifically, these techniques often do not provide data with sufficient conclusive discriminating power between the 'urban background'PAH sources and those from MGP-operations, which is pertinent for meeting low-level, stringent site-cleanup standards. GTI has been developing a new analytical method for the measurement of 'urban background' PAH contamination. This method measures the compound-specific isotope ratio (CSIR) carbon with a GC/IRMS (isotope ratio mass spectrometer). The GC/IRMS technique is a relatively new analytical tool that has great potential as an environmental forensic method at former MGP sites. This paper focuses on the applications of both chemical and isotopic analysis of samples to discern PAH contamination in the environment.     

Surface-assisted laser desorption/ionization mass spectrometry

Laser desorption/ionization mass spectrometry (MS) is rapidly growing in popularity as an analytical characterization method in several fields. The technique shot to prominence using matrix-assisted desorption/ionization for large biomolecules (>700 Da), such as proteins, peptides and nucleic acids. However, because the matrix, which consists of small organic molecules, is also ionized, the technique is of limited use in the low-molecular-mass range (<700 Da). Recent advances in surface science have facilitated the development of matrix-free laser desorption/ionization MS approaches, which are referred to here as surface-assisted laser desorption/ionization (SALDI) MS. In contrast to traditional matrix-assisted techniques, the materials used for SALDI-MS are not ionized, which expands the usefulness of this technique to small-molecule analyses. This review discusses the current status of SALDI-MS as a standard analytical technique, with an emphasis on potential applications in proteomics.     

Surface-assisted laser desorption/ionization mass spectrometry

Laser desorption/ionization mass spectrometry (MS) is rapidly growing in popularity as an analytical characterization method in several fields. The technique shot to prominence using matrix-assisted desorption/ionization for large biomolecules (>700 Da), such as proteins, peptides and nucleic acids. However, because the matrix, which consists of small organic molecules, is also ionized, the technique is of limited use in the low-molecular-mass range (<700 Da). Recent advances in surface science have facilitated the development of matrix-free laser desorption/ionization MS approaches, which are referred to here as surface-assisted laser desorption/ionization (SALDI) MS. In contrast to traditional matrix-assisted techniques, the materials used for SALDI-MS are not ionized, which expands the usefulness of this technique to small-molecule analyses. This review discusses the current status of SALDI-MS as a standard analytical technique, with an emphasis on potential applications in proteomics.     

Organic compounds in re-circulated leachates of aerobic biological treated municipal solid waste

Biodegradation of organic matter is required to reduce the potential of municipal solid waste for producing gaseous emissions and leaching contaminants. Therefore, we studied leachates of an aerobic-treated waste from municipal solids and a sewage sludge mixture that were re-circulated to decrease the concentration of biodegradable organic matter in laboratory-scale reactors. After 12 months, the total organic C and biological and chemical oxygen demands were reduced, indicating the biodegradation of organic compounds in the leachates. Curie-point pyrolysis-gas chromatography/mass spectrometry (Py-GC/MS) and pyrolysis-field ionization mass spectrometry (Py-FIMS) revealed that phenols, alkylaromatic compounds, N-containing compounds and carbohydrates were the predominate compounds in the leachates and solid waste. Leachate re-circulation led to a higher thermal stability of the residual organic matter as indicated by temperature-resolved Py-FIMS. Admixture of sewage sludge to solid waste was less effective in removing organic compounds from the leachates. It resulted in drastic higher and more bio-resistant loads of organic matter in the leachates and revealed increased proportions of alkylaromatic compounds. The biodegradation of organic matter in leachates, re-circulated through municipal solid waste, offers the potential for improved aerobic waste treatments and should be investigated on a larger scale.     

Simultaneous determination of clozapine, olanzapine, risperidone and quetiapine in plasma by high-performance liquid chromatography-electrospray ionization mass spectrometry

Clozapine (CLZ), olanzapine (OLZ), risperidone (RIP) and quetiapine (QTP) have been widely used in the treatment of schizophrenia. However, no study (or little study) has been conducted to determine the four drugs simultaneously by the use of high-performance liquid chromatography-electrospray ionization mass spectrometry (HPLC-MS/ESI). OBJECTIVE: To develop a sensitive method for simultaneous determination of CLZ, OLZ, RIP and QTP in human plasma by HPLC-MS/ESI. METHODS: The analytes were extracted twice by ether after samples had been alkalinized. The HPLC separation of the analytes was performed on a MACHEREY-NAGEL C(18) ( [Formula: see text] mm, 3 microm, Germany) column, using water (formic acid: 2.70 mmol/l, ammonium acetate: 10 mmol/l)-acetonitrile (53:47) as mobile phase, with a flow-rate of 0.16 ml/min. The compounds were ionized in the electrospray ionization (ESI) ion source of the mass spectrometer and were detected in the selected ion recording (SIR) mode. RESULTS: The calibration curves were linear in the ranges of 20-1000 ng/ml for CLZ and QTP, 1-50 ng/ml for OLZ and RIP, respectively. The average extraction recoveries for all the four analysts were at least above 80%. The methodology recoveries were higher than 91% for the analysts. The intra- and inter-day R.S.D. were less than 15%. CONCLUSION: The method is accurate, sensitive and simple for routine therapeutic drug monitoring (TDM) and for the study of the pharmacokinetics of the four drugs.     

Determination of the post mortem interval in skeletal remains by the comparative use of different physico-chemical methods: Are they reliable as an alternative to 14C?

The determination of the post-mortem interval (PMI) of skeletal remains is a challenging aspect in the forensic field. Previous studies focused their attention on different macroscopic and morphological aspects but a thorough and complete evaluation of the potential of chemical and physical analyses in this field of research has not been performed. In addition to luminol test and Oxford histology index (OHI) reported in a recent paper, widely spread and accessible methods based on physical aspect and chemical characteristics of skeletal remains have been investigated as potential alternatives to dating by determination of ¹⁴C.The investigation was performed on a total of 24 archeological and forensic bone samples with known PMI, with inductively coupled plasma optical emission spectrometer (ICP-OES), inductively coupled plasma quadruple mass spectrometry (ICP-MS), Fourier transform infrared (FT-IR) spectroscopy, energy dispersive X-ray analysis (EDX), powder X-ray diffraction analysis (XRPD) and scanning electron microscopy (SEM). Finally, the feasibility of such alternative methods was discussed. Some results such as carbonates/phosphates ratio from FT-IR, the amounts of organic and inorganic matter by EDX, crystallite sizes with XRPD, and surface morphology obtained by SEM, showed significant trends along with PMI. Though, from a chemical point of view cut-off values and gold-standard methods still present challenges, and rather different techniques together can provide useful information toward the assessment of the PMI of skeletal remains. It is however clear that in a hypothetical flowchart those methods may be placed practically at the same level and a choice should always consider the evaluation of results by each technique, execution times and a costs/benefits relationship.     

Frontal affinity chromatography-mass spectrometry

Frontal affinity chromatography (FAC) is a biophysical method for the discovery and characterization of molecular interactions in a flow-based system. Several different modes of analysis are possible by interfacing to the mass spectrometer, including robust single-compound characterizations as well as high-throughput screening of over 1,000 compounds per run. The method supports thermodynamic and kinetic characterization of interactions for a wide range of molecular species and possesses similarities to flow-based biosensors such as surface plasmon resonance. It offers sensitive detection of ligands present well below their respective dissociation constants, and can be assembled from readily available laboratory components. Direct coupling of the FAC cartridge to the mass spectrometer is useful for the interrogation of single compounds or mixtures of limited complexity. An offline fractionation schema is more appropriate for discovery-mode applications. A high-performance FAC system enabling both modes can be assembled in 2-3 h. Measurements of dissociation constants can be made with such a system in 0.5-3 h, and the system supports higher-throughput screening modes at a rate of 10,000 compounds d(-1).