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In cartography, the combination of colour and contour lines is used to express a three-dimensional landscape on a two-dimensional map. We transferred this concept to the analysis of mass spectrometry imaging (MSI) data and developed a collection of R scripts for the efficient evaluation of .imzML archives in a four-step strategy: (1) calculation of the density distribution of mass-to-charge ratio (m/z) signals in the .imzML file and assembling of a pseudo-master spectrum with peak list, (2) automated generation of mass images for a defined scan range and subsequent visual inspection, (3) visualisation of individual ion distributions and export of relevant .mzML spectra and (4) creation of overlay graphics of ion images and photographies. The use of a Hue-Chroma-Luminance (HCL) colour model in MSI graphics takes into account the human perception for colours and supports the correct evaluation of signal intensities. Further, readers with colour blindness are supported. Contour maps promote the visual recognition of patterns in MSI data, which is particularly useful for noisy data sets. We demonstrate the scalability of MSI.R scripts by running them on different systems: on a personal computer, on Amazon Web Services (AWS) instances and on an institutional cluster. By implementing a parallel computing strategy, the execution speed for .imzML data scanning with image generation could be improved by more than an order of magnitude. Applying our MSI.R scripts ( http://www.bioprocess.org/MSI.R ) to low-temperature plasma (LTP)-MSI data shows the localisation of volatile and semi-volatile compounds in the cross-cut of a chilli (Capsicum annuum) fruit. The subsequent identification of compounds by gas and liquid chromatography coupled to mass spectrometry (GC-MS, LC-MS) proves that LTP-MSI enables the direct measurement of volatile organic compound (VOC) distributions from biological tissues.

One of the initial and critical procedures for the analysis of metabolomics data using liquid chromatography and mass spectrometry is feature detection. Feature detection is the process to detect boundaries of the mass surface from raw data. It consists of detected abundances arranged in a two-dimensional (2D) matrix of mass/charge and elution time. MZmine 2 is one of the leading software environments that provide a full analysis pipeline for these data. However, the feature detection algorithms provided in MZmine 2 are based mainly on the analysis of one-dimension at a time. We propose GridMass, an efficient algorithm for 2D feature detection. The algorithm is based on landing probes across the chromatographic space that are moved to find local maxima providing accurate boundary estimations. We tested GridMass on a controlled marker experiment, on plasma samples, on plant fruits, and in a proteome sample. Compared with other algorithms, GridMass is faster and may achieve comparable or better sensitivity and specificity. As a proof of concept, GridMass has been implemented in Java under the MZmine 2 environment and is available at http://www.bioinformatica.mty.itesm.mx/GridMass and MASSyPup. It has also been submitted to the MZmine 2 developing community., (Copyright © 2015 John Wiley & Sons, Ltd.)

Mass spectrometry imaging (MSI) is of high and growing interest in life science research, but the investment for necessary equipment is often prohibitive for small research groups. Therefore, we developed a basic MSI system from low cost 'Plug and Play' components, which are connected to the Universal Serial Bus (USB) of a standard computer. Our open source software OpenMZxy (http://www.bioprocess.org/openmzxy) enables automatic and manual sampling, as well as the recording of position data. For ionization we used a low-temperature plasma probe (LTP), coupled to a quadrupole mass analyzer. The current set-up has a practical resolution of 1mm, and a sampling area of 100×100mm, resulting in up to 10,000 sampling points. Our prototype is easy and economical to adopt for different types of mass analyzers. We prove the usability of the LTP-MSI system for macroscopic samples by imaging the distribution of metabolites in the longitudinal cross-cut of a chili (Capsicum annuum, 'Jalapeño pepper') fruit. The localization of capsaicin in the placenta could be confirmed. But additionally, yet unknown low molecular weight compounds were detected in defined areas, which underline the potential of LTP-MSI for the imaging of volatile and semi-volatile metabolites and for the discovery of new natural products. Biological significance Knowledge about the spatial distribution of metabolites, proteins, or lipids in a given tissue often leads to novel findings in medicine and biology. Therefore, mass spectrometry based imaging (MSI) is becoming increasingly popular in life science research. However, the investment for necessary equipment is often prohibitive for small research groups. We built a prototype with an ambient ionization source, which is easy and economical to adopt for different types of mass analyzers. Therefore, we hope that our system contributes to a broader use of mass spectrometry imaging for answering biological questions., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Rationale: The direct detection of organic molecules by mass spectrometry requires ionization methods which are compatible with ambient conditions. A relatively new strategy is the use of a free low-temperature plasma beam for ionization. The objective is to design a safe and adjustable plasma beam to enable optimal ionization and desorption parameters for specific molecules., Methods: A plasma probe based on a dielectric barrier discharge was designed, where the plasma is guided through an internal second tube. This setup permits different beam diameter settings and the control of the plasma temperature. The ionization and desorption of pure organic compounds, as well as their direct detection from roasted coffee beans, were tested., Results: The presented plasma probe provides improved safety with respect to arcing, ozone generation and electric shock, compared with conventional designs. The functionality of previously reported devices is expanded. A defined plasma diameter can be set by choosing the appropriate insert, while the input voltage controls the plasma temperature. The variation of measurement parameters enables the optimized direct detection of target compounds from roasted coffee beans, such as caffeine, guaiacol and vanillin., Conclusions: The presented low-temperature plasma probe allows the fine-tuning of ionization and desorption parameters, according to the target molecules. Possible applications include: (1) The ambient ionization and desorption of organic compounds with different volatility and (2) The direct analysis of food products such as roasted coffee beans., (Copyright © 2013 John Wiley & Sons, Ltd.)

Visualization of proteins and MS-based analyses are elemental tasks in modern biochemistry. Nevertheless, reports about covalent protein dyes and their suitability for subsequent MS experiments remain scarce. In a recent work, we demonstrated that covalent prestaining of proteins with Uniblue A drastically speeds up proteomic workflows. The present study introduces dabsyl chloride as another truly MS-compatible protein stain. Remarkably, although Uniblue A and dabsyl chloride employ different nucleophilic reaction mechanisms, both are highly specific for lysine residues. The predictable peptide modifications allow easy integration into state-of-the-art bioinformatic workflows. Further, lysine-directed derivatizations with hydrophobic reagents such as dabsyl chloride complement the cysteine-directed ALiPHAT strategy for increasing the sensitivity of peptide identifications., (© 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Background: The identification of proteins by mass spectrometry is a standard method in biopharmaceutical quality control and biochemical research. Prior to identification by mass spectrometry, proteins are usually pre-separated by electrophoresis. However, current protein staining and de-staining protocols are tedious and time consuming, and therefore prolong the sample preparation time for mass spectrometry., Methodology and Principal Findings: We developed a 1-minute covalent pre-gel staining protocol for proteins, which does not require de-staining before the mass spectrometry analysis. We investigated the electrophoretic properties of derivatized proteins and peptides and studied their behavior in mass spectrometry. Further, we elucidated the preferred reaction of proteins with Uniblue A and demonstrate the integration of the peptide derivatization into typical informatics tools., Conclusions and Significance: The Uniblue A staining method drastically speeds up the sample preparation for the mass spectrometry based identification of proteins. The application of this chemo-proteomic strategy will be advantageous for routine quality control of proteins and for time-critical tasks in protein analysis.

Early differential diagnosis of systemic inflammatory reactions in critically ill patients is essential for timely implementation of lifesaving therapies. Despite many efforts made, reliable biomarkers to discriminate between infectious and noninfectious causes of systemic inflammatory response syndrome (SIRS) are currently not available. Recent advances in mass spectrometry-based methods have raised hopes that identification of spectral patterns from serum/plasma samples can be instrumental in this context. We compared protein expression patterns from patients with SIRS of infectious and noninfectious origin. Plasma samples from 166 patients obtained under rigorously standardized preanalytical conditions were applied to Q10 and CM10 ProteinChips. Protein profiles were used to train and develop decision tree classification algorithms. Discriminatory peaks were isolated and identified. Classification trees distinguished patients with noninfectious SIRS with organ dysfunction following open heart surgery using cardiopulmonary bypass from those with severe sepsis or septic shock with distinct sensitivities and specificities. Results were validated in a blinded test set in two independent experiments and in a second independently collected test set. Discriminatory peaks at 13.8 and 55.7 kd were identified as transthyretin and α1-antitrypsin; the third protein at m/z 4,798 was assigned to a proteolytic fragment of α1-antitrypsin. Taken together, our data demonstrate that plasma protein profiling allows reproducible discrimination between patients with infectious and noninfectious SIRS with high sensitivity and specificity. However, rigorous standardization as well as considering drug-related interferences is essential when interpreting protein profiling studies. Identification of discriminatory proteins suggests a direct link between infectious-related protease activity and a sepsis-specific diagnostic pattern for discrimination of patients with SIRS.

Soldiers of the Mexican Army with obesity were subjected to an intense 60-day weight-loss course consisting of a controlled diet, daily physical training, and psychological sessions. The nutritional treatment followed the European Society of Cardiology (ESC) recommendations, incorporating elements of the traditional milpa diet in the nutritional intervention. The total energy intake was reduced by 200 kcal every 20 days, starting with 1,800 kcal and ending with 1,400 kcal daily. On average, the participants reduced their body weight by 18 kg. We employed an innovative approach to monitor the progress of the twelve soldiers who completed the entire program. We compared the untargeted metabolomics profiles of their urine samples, taken before and after the course. The data obtained through liquid chromatography and high-resolution mass spectrometry (LC-MS) provided insightful results. Classification models perfectly separated the profiles pre and post-course, indicating a significant reprogramming of the participants' metabolism. The changes were observed in the C1-, vitamin, amino acid, and energy metabolism pathways, primarily affecting the liver, biliary system, and mitochondria. This study not only demonstrates the potential of rapid weight loss and metabolic pathway modification but also introduces a non-invasive method for monitoring the metabolic state of individuals through urine mass spectrometry data. [ABSTRACT FROM AUTHOR]

Vitamaize lines (VMLs) were created by backcrossing the pigmented aleurone trait into Centro Internacional de Mejoramiento de Maíz y Trigo (CIMMYT) maize lines (CMLs). This study evaluates metabolic differences between the VMLs and their original CMLs. Direct infusion mass spectrometry (DIMS) analyses, carotenoid profiling, total anthocyanins content (TAC) determination, and biochemical evaluation of the quality protein maize (QPM) endosperm trait allowed a comprehensive chemical characterization of the maize lines. DIMS data indicate higher hexoses and trigonelline content for most VMLs; the carotenoid profile revealed a decrease in β-cryptoxanthin to less than half of the original parent content for two VMLs but an augmentation for one VML. The pigmented aleurone VMLs did not inherit the complex QPM endosperm trait of the QPM CMLs. Except for anthocyanin accumulation, no other metabolites were consistently modified across all the backcross-generated maize lines with a pigmented aleurone trait. These findings suggest using genetic or metabolic markers rather than morphological or visual traits for future breeding programs. [ABSTRACT FROM AUTHOR]

Rationale: Ambient ionization mass spectrometry (AIMS) delivers realistic data from samples in their native state. In addition, AIMS methods reduce time and costs for sample preparation and have less environmental impact. However, AIMS data are often complex and require substantial processing before interpretation. Methods: We developed an interactive R script for guided mass spectrometry (MS) data processing. The "MQ_Assistant" is based on MALDIquant, a popular R package for MS data processing. In each step, the user can try and preview the effect of chosen parameters before deciding on the values with the best result and proceeding to the next stage. The outcome of the MQ_Assistant is a feature matrix that can be further analyzed in R and statistics tools such as MetaboAnalyst. Results: Using 360 AIMS example spectra, we demonstrate the step‐by‐step processing for creating a feature matrix. In addition, we show how to visualize the results of three biological replicates of a plant–microbe interaction between Arabidopsis and Trichoderma as a heatmap using R and upload them to MetaboAnalyst. The final parameter set can be saved for reuse in MALDIquant workflows of similar data. Conclusions: The MQ_Assistant helps novices and experienced users to develop workflows for (AI)MS data processing. The interactive procedure supports the quick finding of appropriate settings. These parameters can be exported and reused in future projects. The stepwise operation with visual feedback also suggests the use of the MQ_Assistant in education. [ABSTRACT FROM AUTHOR]

Thin-layer chromatography (TLC) is a classic method for the separation and analysis of complex mixtures. Biological assays, chemical derivatisation and spectroscopy techniques are compatible with TLC and provide extra information about isolated compounds. However, coupling TLC to mass spectrometry is hampered by the difficulty to desorb the analytes from the silica surfaces. In this study, we used a multimodal ion source for laser desorption (LD) and low-temperature plasma (LTP) post-ionisation. Efficient desorption was reached by covering the TLC plates with activated carbon. Regions of interest can be analysed by spots, by lines or by area. We show the separation of methylxanthines from coffee, tea and cocoa preparations by TLC, with subsequent mass spectrometry imaging (MSI). Using a lateral resolution of 400 μm × 400 μm, allowed the acquisition of 21 895 spectra in 2.4 h (2.5 pixels per s). Further, we demonstrate the possibility of direct mass fragmentation studies and quantification. We mounted the system on an Open LabBot with a theoretical lateral resolution of 12.5 μm and performed the visualisation of ions of interest and the pixel-wise review of mass spectra with our free software RmsiGUI (https://bitbucket.org/lababi/rmsigui). This non-proprietary and modular platform enables the cost-efficient adaption of the system and further development by the community. [ABSTRACT FROM AUTHOR]

Main conclusion: The method introduced here to grow F. hygrometrica in high concentrations of D O is an excellent alternative to produce highly deuterated metabolites with broad applications in metabolic studies. Our mass spectrometry experiments strongly indicate the successful incorporation of deuterium into organic compounds. Deuterated metabolites are useful tracers for metabolic studies, yet their wide utilization in research is limited by the multi-step total synthesis required to produce them in the laboratory. Alternatively, deuterated metabolites can be obtained from organisms grown in DO or deuterated nutrients. This approach also has limitations as DO in high concentrations negatively affects the survival of most organisms. Here we report the moss Funaria hygrometrica as an unusual high tolerant to DO in liquid culture. We found that this moss is able to grow in up to 90% DO, a condition lethal for many eukaryotes. Mass spectrometric analyses of F. hygrometrica extracts showed a strong deuteration pattern. The ability to tolerate high concentrations of DO together with the development of a rich molecular toolbox makes F. hygrometrica an ideal system for the production of valuable deuterated metabolites. [ABSTRACT FROM AUTHOR]

Mass spectrometry has evolved to a key technology in the areas of metabolomics and proteomics. Centralized facilities generate vast amount of data, which frequently need to be processed off-site. Therefore, the distribution of data and software, as well as the training of personnel in the analysis of mass spectrometry data, becomes increasingly important. Thus, we created a comprehensive collection of mass spectrometry software which can be run directly from different media such as DVD or USB without local installation. MASSyPup is based on a Linux Live distribution and was complemented with programs for conversion, visualization and analysis of mass spectrometry (MS) data. A special emphasis was put on protein analysis and proteomics, encompassing the measurement of complete proteins, the identification of proteins based on Peptide Mass Fingerprints (PMF) or LC-MS/MS data, and de novo sequencing. Another focus was directed to the study of metabolites and metabolomics, covering the detection, identification and quantification of compounds, as well as subsequent statistical analyses. Additionally, we added software for Mass Spectrometry Imaging (MSI), including hardware support for self-made MSI devices. MASSyPup represents a 'ready to work' system for teaching or MS data analysis, but also represents an ideal platform for the distribution of MS data and the development of related software. The current Live DVD version can be downloaded free of charge from http://www.bioprocess.org/massypup. Copyright © 2014 John Wiley & Sons, Ltd. [ABSTRACT FROM AUTHOR]

Ambient ionisation mass spectrometry (AIMS) enables studying biological systems in their native state and direct high-throughput analyses. The ionisation occurs in the physical conditions of the surrounding environment. Simple spray or plasma-based AIMS devices allow the desorption and ionisation of molecules from solid, liquid and gaseous samples. 3D printing helps to implement new ideas and concepts in AIMS quickly. Here, we present examples of 3D printed AIMS sources and devices for ion transfer and manipulation. Further, we show the use of 3D printer parts for building custom AIMS sampling robots and imaging systems. Using 3D printing technology allows upgrading existing mass spectrometers with relatively low cost and effort. [ABSTRACT FROM AUTHOR]

Abstract: In an approach to gain insight into the sequence-dependent dephosphorylation of multiple phosphotyrosyl-containing peptides by the phosphatases SHP-1 and PTP1B, we applied a chromatographic technique for the analysis of the dephosphorylation products. Mono-, bi- and triphosphorylated reference peptides corresponding to positions 1999–2014 in the activation loop of the receptor tyrosine kinase Ros were first analyzed by reversed-phase HPLC and MALDI-TOF/TOF mass spectrometry. Then, the respective products from enzymatic treatment were investigated by HPLC and compared to the standard peptides. The results obtained in this study emphasize the advantage of monitoring phosphatase reactions for mono- and biphosphorylated peptides using the described procedure rather than spectrophotometric and fluorimetric methods that do not allow for a clear identification of the products formed. [Copyright &y& Elsevier]

Background: different Solanaceae and Erythroxylaceae species produce tropane alkaloids. These alkaloids are the starting material in the production of different pharmaceuticals. The commercial demand for tropane alkaloids is covered by extracting them from cultivated plants. Datura stramonium is cultivated under greenhouse conditions as a source of tropane alkaloids. Here we investigate the effect of different levels of water availability in the soil on the production of tropane alkaloids by D. stramonium. Methods: We tested four irrigation levels on the accumulation of tropane alkaloids. We analyzed the profile of tropane alkaloids using an untargeted liquid chromatography/mass spectrometry method. Results: Using a combination of informatics and manual interpretation of mass spectra, we generated several structure hypotheses for signals in D. stramonium extracts that we assign as putative tropane alkaloids. Quantitation of mass spectrometry signals for our structure hypotheses across different anatomical organs allowed us to identify patterns of tropane alkaloids associated with different levels of irrigation. Furthermore, we identified anatomic partitioning of tropane alkaloid isomers with pharmaceutical applications. Conclusions: Our results show that soil water availability is an effective method for maximizing the production of specific tropane alkaloids for industrial applications. [ABSTRACT FROM AUTHOR]

Low-temperature plasma (LTP) is capable of ionizing a broad range of organic molecules at ambient conditions. The coupling of LTP to a mass analyzer delivers chemical profiles from delicate objects. To investigate the suitability of LTP ionization for mass spectrometry (MS) based in vivo studies, we monitored the auxin-regulated nicotine biosynthesis in tobacco ( Nicotiana tabacum ) and evaluated possible biological effects. The measured nicotine concentrations in different experiments were comparable to literature data obtained with conventional methods. The observed compounds suggest the rupture of trichomes, and cell damage was observed on the spots exposed to LTP. However, the lesions only affected a negligible proportion of the leaf surface area and no systemic reaction was noted. Thus, our study provides the proof-of-concept for measuring the biosynthetic activity of plant surfaces in vivo . [ABSTRACT FROM AUTHOR]

Low-temperature plasma (LTP) ionization represents an emerging technology in ambient mass spectrometry. LTP enables the solvent-free direct detection of a broad range of molecules and mass spectrometry imaging (MSI). The low energy consumption and modest technical requirements of these ion sources favors their employment in mobile applications and as a means to upgrade existing mass analyzers. However, the broad adoption of LTP is hindered by the lack of commercial devices, and constructing personal devices is tricky. Improper setup can result in equipment malfunction or may cause serious damage to instruments due to strong electromagnetic fields or arcing. With this in mind, we developed a reproducible LTP probe, which is designed exclusively from commercial and 3D printed components. The plasma jet generated by the device has a diameter of about 200 Î?m, which is satisfactory for the ambient imaging of macroscopic samples. We coupled the 3D-LTP probe to an ion trap analyzer and demonstrated the functionality of the ion source by detecting organic and chemical compounds from pure reference standards, biological substances, and pharmaceutical samples. Molecules were primarily detected in their protonated form or as water/ammonium adducts. The identification of compounds was possible by standard collision-induced dissociation (CID) fragmentation spectra. The files necessary to reproduce the 3D parts are available from the project page (http://lababi.bioprocess.org/index.php/3d-ltp) under a dual license model, which permits reproduction of the probe and further community-driven development for noncommercial use (“peer productionâ€?). Our reproducible probe design thus contributes to a facilitated adaption and evolution of low-temperature plasma technologies in analytical chemistry. [ABSTRACT FROM AUTHOR]

• Modular Open LabBot for sampling/ imaging based on 3D-printer components. • RmsiGUI software for robot control and mass spectrometry imaging. • Example 1: High-throughput analysis of essential oils with a PlasmaChip. • Example 2: Ambient mass spectrometry imaging of a lime slice. Building novel analytical systems requires various elements: Sensors, a sampling system and data evaluation software. For emerging technologies, such as ambient ionization mass spectrometry (AIMS), the assembly of a functional prototype is often more time-consuming than developing the actual probes. The modification of commercial platforms for testing hardware is usually not possible due to technical and legal restrictions. Therefore, we designed a robotic platform, which is based on RepRap 3D-printer components, and therefore easy to adopt for custom set-ups. Adjustable rails in an aluminium frame construction facilitate the mounting of additional parts such as sensors, probes, or optical components. The Open LabBot uses industry-standard G-code for its control. Its simple syntax facilitates the programming of the movement. We developed two programs for the robot control and mass spectrometry imaging: "LABI-Imaging" for synchronizing the robot movements and MS data acquisition and "RmsiGUI", a software solution for the complete AIMS workflow. We proved the functionality of the system by the automated sampling and classification of essential oils with a PlasmaChip probe. Further, we performed an AIMSI imaging experiment of a lime slice with laser desorption low-temperature plasma (LD-LTP) ionization, demonstrating the integration of the complete workflow in RmsiGUI. The design of the Open LabBot and the software are released under open licenses to promote their use and adoption in the instrument developers' community. [ABSTRACT FROM AUTHOR]