Optimization of the Ion Source-Mass Spectrometry Parameters in Non-Steroidal Anti-Inflammatory and Analgesic Pharmaceuticals Analysis by a Design of Experiments Approach

The flow rates of drying and nebulizing gas, heat block and desolvation line temperatures and interface voltage are potential electrospray ionization parameters as they may enhance sensitivity of the mass spectrometer. The conditions that give higher sensitivity of 13 pharmaceuticals were explored. First, Plackett-Burman design was implemented to screen significant factors, and it was concluded that interface voltage and nebulizing gas flow were the only factors that influence the intensity signal for all pharmaceuticals. This fractionated factorial design was projected to set a full 2(2) factorial design with center points. The lack-of-fit test proved to be significant. Then, a central composite face-centered design was conducted. Finally, a stepwise multiple linear regression and subsequently an optimization problem solving were carried out. Two main drug clusters were found concerning the signal intensities of all runs of the augmented factorial design. p-Aminophenol, salicylic acid, and nimesulide constitute one cluster as a result of showing much higher sensitivity than the remaining drugs. The other cluster is more homogeneous with some sub-clusters comprising one pharmaceutical and its respective metabolite. It was observed that instrumental signal increased when both significant factors increased with maximum signal occurring when both codified factors are set at level +1. It was also found that, for most of the pharmaceuticals, interface voltage influences the intensity of the instrument more than the nebulizing gas flowrate. The only exceptions refer to nimesulide where the relative importance of the factors is reversed and still salicylic acid where both factors equally influence the instrumental signal. Graphical Abstract ᅟ.

Analysis of organic aerosols collected on filters by Aerosol Mass Spectrometry for source identification

Aerosol mass spectrometers (AMS) are powerful tools in the analysis of the chemical composition of airborne particles, particularly organic aerosols which are gaining increasing attention. However, the advantages of AMS in providing on-line data can be outweighed by the difficulties involved in its use in field measurements at multiple sites. In contrast to the on-line measurement by AMS, a method which involves sample collection on filters followed by subsequent analysis by AMS could significantly broaden the scope of AMS application. We report the application of such an approach to field studies at multiple sites. An AMS was deployed at 5 urban schools to determine the sources of the organic aerosols at the schools directly. PM1 aerosols were also collected on filters at these and 20 other urban schools. The filters were extracted with water and the extract run through a nebulizer to generate the aerosols, which were analysed by an AMS. The mass spectra from the samples collected on filters at the 5 schools were found to have excellent correlations with those obtained directly by AMS, with r2 ranging from 0.89 to 0.98. Filter recoveries varied between the schools from 40 -115%, possibly indicating that this method provides qualitative rather than quantitative information. The stability of the organic aerosols on Teflon filters was demonstrated by analysing samples stored for up to two years. Application of the procedure to the remaining 20 schools showed that secondary organic aerosols were the main source of aerosols at the majority of the schools. Overall, this procedure provides accurate representation of the mass spectra of ambient organic aerosols and could facilitate rapid data acquisition at multiple sites where AMS could not be deployed for logistical reasons.

Advantages of using nested collision induced dissociation/post-source decay with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: sequencing of novel peptides from wasp venom

We have examined the applicability of the 'nested' collision induced dissociation/post-source decay (CID/PSD) method to the sequencing of novel peptides from solitary wasps which have neurotoxic venom for paralyzing other insects. The CID/PSD spectrum of a ladder peptide derived from an exopeptidase digest was compared with that of the intact peptide. The mass peaks observed only in the CID/PSD spectrum of a ladder peptide were extracted as C-terminal fragment ions. Assignment of C-terminal fragment ions enabled calculation of N-terminal fragment masses, leading to differentiation between N-terminal fragment ions and internal fragment ions. This methodology allowed rapid and sensitive identification by removing ambiguity in the assignment of the fragment ions, and proved useful for sequencing unknown peptides, in particular those available as natural products with a limited supply. Copyright (C) 2000 John Wiley & Sons, Ltd.

Aircraft-based in-situ aerosol mass spectrometry: Chemical characterization and source identification of submicron particulate matter in the free and upper troposphere and lower stratosphere

The composition of the atmosphere is frequently perturbed by the emission of gaseous and particulate matter from natural as well as anthropogenic sources. While the impact of trace gases on the radiative forcing of the climate is relatively well understood the role of aerosol is far more uncertain. Therefore, the study of the vertical distribution of particulate matter in the atmosphere and its chemical composition contribute valuable information to bridge this gap of knowledge. The chemical composition of aerosol reveals information on properties such as radiative behavior and hygroscopicity and therefore cloud condensation or ice nucleus potential. rnThis thesis focuses on aerosol pollution plumes observed in 2008 during the POLARCAT (Polar Study using Aircraft, Remote Sensing, Surface Measurements and Models, of Climate, Chemistry, Aerosols, and Transport) campaign over Greenland in June/July and CONCERT (Contrail and Cirrus Experiment) campaign over Central and Western Europe in October/November. Measurements were performed with an Aerodyne compact time-of-flight aerosol mass spectrometer (AMS) capable of online size-resolved chemical characterization of non-refractory submicron particles. In addition, the origins of pollution plumes were determined by means of modeling tools. The characterized pollution episodes originated from a large variety of sources and were encountered at distinct altitudes. They included pure natural emissions from two volcanic eruptions in 2008. By the time of detection over Western Europe between 10 and 12 km altitude the plume was about 3 months old and composed to 71 % of particulate sulfate and 21 % of carbonaceous compounds. Also, biomass burning (BB) plumes were observed over Greenland between 4 and 7 km altitude (free troposphere) originating from Canada and East Siberia. The long-range transport took roughly one and two weeks, respectively. The aerosol was composed of 78 % organic matter and 22 % particulate sulfate. Some Canadian and all Siberian BB plumes were mixed with anthropogenic emissions from fossil fuel combustion (FF) in North America and East Asia. It was found that the contribution of particulate sulfate increased with growing influences from anthropogenic activity and Asia reaching up to 37 % after more than two weeks of transport time. The most exclusively anthropogenic emission source probed in the upper troposphere was engine exhaust from commercial aircraft liners over Germany. However, in-situ characterization of this aerosol type during aircraft chasing was not possible. All long-range transport aerosol was found to have an O:C ratio close to or greater than 1 implying that low-volatility oxygenated organic aerosol was present in each case despite the variety of origins and the large range in age from 3 to 100 days. This leads to the conclusion that organic particulate matter reaches a final and uniform state of oxygenation after at least 3 days in the free troposphere. rnExcept for aircraft exhaust all emission sources mentioned above are surface-bound and thus rely on different types of vertical transport mechanisms, such as direct high altitude injection in the case of a volcanic eruption, or severe BB, or uplift by convection, to reach higher altitudes where particles can travel long distances before removal mainly caused by cloud scavenging. A lifetime for North American mixed BB and FF aerosol of 7 to 11 days was derived. This in consequence means that emission from surface point sources, e.g. volcanoes, or regions, e.g. East Asia, do not only have a relevant impact on the immediate surroundings but rather on a hemispheric scale including such climate sensitive zones as the tropopause or the Arctic.

Characterization and source apportionment of organic aerosol using offline aerosol mass spectrometry

Field deployments of the Aerodyne Aerosol Mass Spectrometer (AMS) have significantly advanced real-time measurements and source apportionment of non-refractory particulate matter. However, the cost and complex maintenance requirements of the AMS make its deployment at sufficient sites to determine regional characteristics impractical. Furthermore, the negligible transmission efficiency of the AMS inlet for supermicron particles significantly limits the characterization of their chemical nature and contributing sources. In this study, we utilize the AMS to characterize the water-soluble organic fingerprint of ambient particles collected onto conventional quartz filters, which are routinely sampled at many air quality sites. The method was applied to 256 particulate matter (PM) filter samples (PM1, PM2:5, and PM10, i.e., PM with aerodynamic diameters smaller than 1, 2.5, and 10 μm, respectively), collected at 16 urban and rural sites during summer and winter. We show that the results obtained by the present technique compare well with those from co-located online measurements, e.g., AMS or Aerosol Chemical Speciation Monitor (ACSM). The bulk recoveries of organic aerosol (60–91 %) achieved using this technique, together with low detection limits (0.8 μg of organic aerosol on the analyzed filter fraction) allow its application to environmental samples. We will discuss the recovery variability of individual hydrocarbon ions, ions containing oxygen, and other ions. The performance of such data in source apportionment is assessed in comparison to ACSM data. Recoveries of organic components related to different sources as traffic, wood burning, and secondary organic aerosol are presented. This technique, while subjected to the limitations inherent to filter-based measurements (e.g., filter artifacts and limited time resolution) may be used to enhance the AMS capabilities in measuring size-fractionated, spatially resolved longterm data sets.

Source apportionment of PM2.5 in Cork Harbour, Ireland using a combination of single particle mass spectrometry and quantitative semi-continuous measurements

An aerosol time-of-flight mass spectrometer (ATOFMS) was deployed for the measurement of the size resolved chemical composition of single particles at a site in Cork Harbour, Ireland for three weeks in August 2008. The ATOFMS was co-located with a suite of semi-continuous instrumentation for the measurement of particle number, elemental carbon (EC), organic carbon (OC), sulfate and particulate matter smaller than 2.5 μm in diameter (PM2.5). The temporality of the ambient ATOFMS particle classes was subsequently used in conjunction with the semi-continuous measurements to apportion PM2.5 mass using positive matrix factorisation. The synergy of the single particle classification procedure and positive matrix factorisation allowed for the identification of six factors, corresponding to vehicular traffic, marine, long-range transport, various combustion, domestic solid fuel combustion and shipping traffic with estimated contributions to the measured PM2.5 mass of 23%, 14%, 13%, 11%, 5% and 1.5% respectively. Shipping traffic was found to contribute 18% of the measured particle number (20–600 nm mobility diameter), and thus may have important implications for human health considering the size and composition of ship exhaust particles. The positive matrix factorisation procedure enabled a more refined interpretation of the single particle results by providing source contributions to PM2.5 mass, while the single particle data enabled the identification of additional factors not possible with typical semi-continuous measurements, including local shipping traffic.

Preliminary results on the characterisation of organic aerosols in urban schools by Aerosol Mass Spectrometry

Vehicle emissions have been linked to detrimental health effects with children thought to be more susceptible (See e.g., Ryan et al 2005). In an urban environment a major source of organic aerosols (OA) are vehicle emissions. The ambient concentration of OA is dynamic in nature and the use of an aerosol mass spectrometer can achieve the necessary temporal resolution to capture the daily variation of OA (Jimenez et al 2009). Currently there is a limited understanding of effects of long term exposure to traffic emissions on children’s health. In the present study, we used an aerosol mass spectrometer to monitor OA and determine children’s potential exposure at school to traffic emissions.In this paper, we present the preliminary results of this investigation. The study is a part of a larger project aimed at gaining a holistic picture of the exposure of children to traffic related pollutants, known as UPTECH (www.ilaqh.qut.edu.au/Misc/ UPTECH%20Home.htm).

Elucidation of double bond position in unsaturated lipids by ozone electrospray ionization mass spectrometry

The position(s) of carbon-carbon double bonds within lipids can dramatically affect their structure and reactivity and thus has a direct bearing on biological function. Commonly employed mass spectrometric approaches to the characterization of complex lipids, however, fail to localize sites of unsaturation within the molecular structure and thus cannot distinguish naturally occurring regioisomers. In a recent communication \[Thomas, M. C.; Mitchell, T. W.; Blanksby, S. J. J. Am. Chem. Soc. 2006, 128, 58-59], we have presented a new technique for the elucidation of double bond position in glycerophospholipids using ozone-induced fragmentation within the source of a conventional electrospray ionization mass spectrometer. Here we report the on-line analysis, using ozone electrospray mass spectrometry (OzESI-MS), of a broad range of common unsaturated lipids including acidic and neutral glycerophospholipids, sphingomyelins, and triacylglycerols. All lipids analyzed are found to form a pair of chemically induced fragment ions diagnostic of the position of each double bond(s) regardless of the polarity, the number of charges, or the adduction (e.g., \[M - H](-), \[M - 2H](2-), \[M + H](+), \[M + Na](+), \[M + NH4](+)). The ability of OzESI-MS to distinguish lipids that differ only in the position of the double bonds is demonstrated using the glycerophosphocholine standards, GPCho(9Z-18:1/9Z-18:1) and GPCho(6Z-18:1/6Z-18:1). While these regioisomers cannot be differentiated by their conventional tandem mass spectra, the OzESI-MS spectra reveal abundant fragment ions of distinctive mass-to-charge ratio (m/z). The approach is found to be sufficiently robust to be used in conjunction with the m/z 184 precursor ion scans commonly employed for the identification of phosphocholine-containing lipids in shotgun lipidomic analyses. This tandem OzESI-MS approach was used, in conjunction with conventional tandem mass spectral analysis, for the structural characterization of an unknown sphingolipid in a crude lipid extract obtained from a human lens. The OzESI-MS data confirm the presence of two regioisomers, namely, SM(d18:0/15Z-24:1) and SM(d18:0/17Z-24:1), and suggest the possible presence of a third isomer, SM(d18:0/19Z-24:1), in lower abundance. The data presented herein demonstrate that OzESI-MS is a broadly applicable, on-line approach for structure determination and, when used in conjunction with established tandem mass spectrometric methods, can provide near complete structural characterization of a range of important lipid classes. As such, OzESI-MS may provide important new insight into the molecular diversity of naturally occurring lipids.

Identification of phospholipids in human meibum by nano-electrospray ionisation tandem mass spectrometry

Meibum is believed to be the major source of tear film lipids, which are vital in the prevention of excess evaporation of the aqueous phase. The complete lipid composition of meibum has yet to be established. While earlier studies reported the presence of phospholipids in human meibum, recent mass spectrometric studies have not detected them. In this study we use electrospray ionisation tandem mass spectrometry to investigate the presence of phospholipids in meibum and provide comparison to the phospholipid profile of tears.Lipids were extracted from human meibum and tear samples using standard biphasic methods and analysed by nano-electrospray ionisation tandem mass spectrometry using targeted ion scans. A total of 35 choline-containing phospholipids were identified in meibum and the profile of these was similar to that observed in tears, suggesting tear lipids are derived from meibum. The results shown here highlight the need for a combination of optimised techniques to enable the identification of the large range of lipid classes in meibum. © 2011 Elsevier Ltd.

Imaging of human lens lipids by desorption electrospray ionization mass spectrometry

The lipid composition of the human lens is distinct from most other tissues in that it is high in dihydrosphingomyelin and the most abundant glycerophospholipids in the lens are unusual 1-O-alkyl-ether linked phosphatidylethanolamines and phosphatidylserines. In this study, desorption electrospray ionization (DESI) mass spectrometry-imaging was used to determine the distribution of these lipids in the human lens along with other lipids including, ceramides, ceramide-1-phosphates, and lyso 1-O-alkyl ethers. To achieve this, 25 μm lens slices were mounted onto glass slides and analyzed using a linear ion-trap mass spectrometer equipped with a custom-built, 2-D automated DESI source. In contrast to other tissues that have been previously analyzed by DESI, the presence of a strong acid in the spray solvent was required to desorb lipids directly from lens tissue. Distinctive distributions were observed for [M + H]+ ions arising from each lipid class. Of particular interest were ionized 1-O-alkyl phosphatidylethanolamines and phosphatidylserines, PE (18:1e/18:1), and PS (18:1e/18:1), which were found in a thin ring in the outermost region of the lens. This distribution was confirmed by quantitative analysis of lenses that were sectioned into four distinct regions (outer, barrier, inner, and core), extracted and analyzed by electrospray ionization tandem mass spectrometry. DESI-imaging also revealed a complementary distribution for the structurally-related lyso 1-O-alkyl phosphatidylethanolamine, LPE (18:1e), which was localized closer to the centre of the lens. The data obtained in this study indicate that DESI-imaging is a powerful tool for determining the spatial distribution of human lens lipids. © 2010 American Society for Mass Spectrometry.

Analysis of the extreme diversity of salivary alpha-amylase isoforms generated by physiological proteolysis using liquid chromatography-tandem mass spectrometry

Saliva is a crucial biofluid for oral health and is also of increasing importance as a non-invasive source of disease biomarkers. Salivary alpha-amylase is an abundant protein in saliva, and changes in amylase expression have been previously associated with a variety of diseases and conditions. Salivary alpha-amylase is subject to a high diversity of post-translational modifications, including physiological proteolysis in the oral cavity. Here we developed methodology for rapid sample preparation and non-targeted LC-ESI-MS/MS analysis of saliva from healthy subjects and observed an extreme diversity of alpha-amylase proteolytic isoforms. Our results emphasize the importance of consideration of post-translational events such as proteolysis in proteomic studies, biomarker discovery and validation, particularly in saliva. (C) 2012 Elsevier B.V. All rights reserved.

Heated Nebulizer Microchips for Mass Spectrometry

Miniaturized analytical devices, such as heated nebulizer (HN) microchips studied in this work, are of increasing interest owing to benefits like faster operation, better performance, and lower cost relative to conventional systems. HN microchips are microfabricated devices that vaporize liquid and mix it with gas. They are used with low liquid flow rates, typically a few µL/min, and have previously been utilized as ion sources for mass spectrometry (MS). Conventional ion sources are seldom feasible at such low flow rates. In this work HN chips were developed further and new applications were introduced. First, a new method for thermal and fluidic characterization of the HN microchips was developed and used to study the chips. Thermal behavior of the chips was also studied by temperature measurements and infrared imaging. An HN chip was applied to the analysis of crude oil – an extremely complex sample – by microchip atmospheric pressure photoionization (APPI) high resolution mass spectrometry. With the chip, the sample flow rate could be reduced significantly without loss of performance and with greatly reduced contamination of the MS instrument. Thanks to its suitability to high temperature, microchip APPI provided efficient vaporization of nonvolatile compounds in crude oil. The first microchip version of sonic spray ionization (SSI) was presented. Ionization was achieved by applying only high (sonic) speed nebulizer gas to an HN microchip. SSI significantly broadens the range of analytes ionizable with the HN chips, from small stable molecules to labile biomolecules. The analytical performance of the microchip SSI source was confirmed to be acceptable. The HN microchips were also used to connect gas chromatography (GC) and capillary liquid chromatography (LC) to MS, using APPI for ionization. Microchip APPI allows efficient ionization of both polar and nonpolar compounds whereas with the most popular electrospray ionization (ESI) only polar and ionic molecules are ionized efficiently. The combination of GC with MS showed that, with HN microchips, GCs can easily be used with MS instruments designed for LC-MS. The presented analytical methods showed good performance. The first integrated LC–HN microchip was developed and presented. In a single microdevice, there were structures for a packed LC column and a heated nebulizer. Nonpolar and polar analytes were efficiently ionized by APPI. Ionization of nonpolar and polar analytes is not possible with previously presented chips for LC–MS since they rely on ESI. Preliminary quantitative performance of the new chip was evaluated and the chip was also demonstrated with optical detection. A new ambient ionization technique for mass spectrometry, desorption atmospheric pressure photoionization (DAPPI), was presented. The DAPPI technique is based on an HN microchip providing desorption of analytes from a surface. Photons from a photoionization lamp ionize the analytes via gas-phase chemical reactions, and the ions are directed into an MS. Rapid analysis of pharmaceuticals from tablets was successfully demonstrated as an application of DAPPI.

Deriving structural information from non-coding ribonucleic acids by mass spectrometry

The purpose of this study is to describe the development of application of mass spectrometry for the structural analyses of non-coding ribonucleic acids during past decade. Mass spectrometric methods are compared of traditional gel electrophoretic methods, the characteristics of performance of mass spectrometric, analyses are studied and the future trends of mass spectrometry of ribonucleic acids are discussed. Non-coding ribonucleic acids are short polymeric biomolecules which are not translated to proteins, but which may affect the gene expression in all organisms. Regulatory ribonucleic acids act through transient interactions with key molecules in signal transduction pathways. Interactions are mediated through specific secondary and tertiary structures. Posttranscriptional modifications in the structures of molecules may introduce new properties to the organism, such as adaptation to environmental changes or development of resistance to antibiotics. In the scope of this study, the structural studies include i) determination of the sequence of nucleobases in the polymer chain, ii) characterisation and localisation of posttranscriptional modifications in nucleobases and in the backbone structure, iii) identification of ribonucleic acid-binding molecules and iv) probing of higher order structures in the ribonucleic acid molecule. Bacteria, archaea, viruses and HeLa cancer cells have been used as target organisms. Synthesised ribonucleic acids consisting of structural regions of interest have been frequently used. Electrospray ionisation (ESI) and matrix-assisted laser desorption ionisation (MALDI) have been used for ionisation of ribonucleic analytes. Ammonium acetate and 2-propanol are common solvents for ESI. Trihydroxyacetophenone is the optimal MALDI matrix for ionisation of ribonucleic acids and peptides. Ammonium salts are used in ESI buffers and MALDI matrices as additives to remove cation adducts. Reverse phase high performance liquid chromatography has been used for desalting and fractionation of analytes either off-line of on-line, coupled with ESI source. Triethylamine and triethylammonium bicarbonate are used as ion pair reagents almost exclusively. Fourier transform ion cyclotron resonance analyser using ESI coupled with liquid chromatography is the platform of choice for all forms of structural analyses. Time-of-flight (TOF) analyser using MALDI may offer sensitive, easy-to-use and economical solution for simple sequencing of longer oligonucleotides and analyses of analyte mixtures without prior fractionation. Special analysis software is used for computer-aided interpretation of mass spectra. With mass spectrometry, sequences of 20-30 nucleotides of length may be determined unambiguously. Sequencing may be applied to quality control of short synthetic oligomers for analytical purposes. Sequencing in conjunction with other structural studies enables accurate localisation and characterisation of posttranscriptional modifications and identification of nucleobases and amino acids at the sites of interaction. High throughput screening methods for RNA-binding ligands have been developed. Probing of the higher order structures has provided supportive data for computer-generated three dimensional models of viral pseudoknots. In conclusion. mass spectrometric methods are well suited for structural analyses of small species of ribonucleic acids, such as short non-coding ribonucleic acids in the molecular size region of 20-30 nucleotides. Structural information not attainable with other methods of analyses, such as nuclear magnetic resonance and X-ray crystallography, may be obtained with the use of mass spectrometry. Sequencing may be applied to quality control of short synthetic oligomers for analytical purposes. Ligand screening may be used in the search of possible new therapeutic agents. Demanding assay design and challenging interpretation of data requires multidisclipinary knowledge. The implement of mass spectrometry to structural studies of ribonucleic acids is probably most efficiently conducted in specialist groups consisting of researchers from various fields of science.