Screening and Quantification of Pesticides in Water Using a Dual-Function Graphitized Carbon Black Disk

A simple platform for combining solid phase extraction (SPE) and surface-assisted laser desorption ionization mass spectrometry (SALDI-MS) of extracted analytes, using disks prepared by embedding graphitized carbon black (GCB-4) particles in a network of polytetrafluoroethylene (PTFE), is presented. The system provides a convenient approach for rapid SALDI-MS screening of substances in aqueous samples, which can be followed by robust quantitative and/or structural analyses by liquid chromatography (LC)/MS/MS of positive samples. The extraction discs are easily transferred between gaskets where the sample extraction and desorption of selected samples is performed and the mass spectrometer. The SPE and SALDI properties of the new GCB-4 disc have been characterized for 15 pesticides with varying chemical properties, and the screening strategy has been applied to the analysis of pesticides in agricultural drainage water. Atrazine and atrazine-desethyl-2-hydroxy were detected in the sampled water by SALDI-MS screening and subsequently confirmed and quantified using LC/MS/MS

A Space Efficient Direct Access Data Compression Approach for Mass Spectrometry Imaging

Advances in mass spectrometry imaging that improve both spatial and mass resolution are resulting in increasingly larger data files that are difficult to handle with current software. We have developed a novel near-lossless compression method with data entropy reduction that reduces the file size significantly. The reduction in data size can be set at four different levels (coarse, medium, fine, and superfine) prior to running the data compression. This can be applied to spectra or spectrum-by-spectrum, or it can be applied to transpose arrays or array-by-array, to efficiently read the data without decompressing the whole data set. The results show that a compression ratio of up to 5.9:1 was achieved for data from commercial mass spectrometry software programs and 55:1 for data from our in-house developed msIQuant program. Comparing the average signals from regions of interest, the maximum deviation was 0.2% between compressed and uncompressed data sets with coarse accuracy for the data entropy reduction. In addition, when accessing the compressed data by selecting a random <i>m</i>/<i>z</i> value using msIQuant, the time to update an image on the computer screen was only slightly increased from 92 (±32) ms (uncompressed) to 114 (±13) ms (compressed). Furthermore, the compressed data can be stored on readily accessible servers for data evaluation without further data reprocessing. We have developed a space efficient, direct access data compression algorithm for mass spectrometry imaging, which can be used for various data-demanding mass spectrometry imaging applications

msIQuant – Quantitation Software for Mass Spectrometry Imaging Enabling Fast Access, Visualization, and Analysis of Large Data Sets

This paper presents msIQuant, a novel instrument- and manufacturer-independent quantitative mass spectrometry imaging software suite that uses the standardized open access data format imzML. Its data processing structure enables rapid image display and the analysis of very large data sets (>50 GB) without any data reduction. In addition, msIQuant provides many tools for image visualization including multiple interpolation methods, low intensity transparency display, and image fusion. It also has a quantitation function that automatically generates calibration standard curves from series of standards that can be used to determine the concentrations of specific analytes. Regions-of-interest in a tissue section can be analyzed based on a number of quantities including the number of pixels, average intensity, standard deviation of intensity, and median and quartile intensities. Moreover, the suite’s export functions enable simplified postprocessing of data and report creation. We demonstrate its potential through several applications including the quantitation of small molecules such as drugs and neurotransmitters. The msIQuant suite is a powerful tool for accessing and evaluating very large data sets, quantifying drugs and endogenous compounds in tissue areas of interest, and for processing mass spectra and images

Molecular imaging identifies age-related attenuation of acetylcholine in retrosplenial cortex in response to acetylcholinesterase inhibition

The neurotransmitter of the cholinergic system, acetylcholine plays a major role in the brain's cognitive function and is involved in neurodegenerative disorders. Here, we present age-related alterations of acetylcholine levels after administration of the acetylcholinesterase inhibitor drug tacrine in normal mice. Using a quantitative, robust and molecular-specific mass spectrometry imaging method we found that tacrine administration significantly raised acetylcholine levels in most areas of sectioned mice brains, inter alia the striatum, hippocampus and cortical areas. However, acetylcholine levels in retrosplenial cortex were significantly lower in 14-month-old than in 12-week-old animals following its administration, indicating that normal aging affects the cholinergic system's responsivity. This small brain region is interconnected with an array of brain networks and is involved in numerous cognitive tasks. Simultaneous visualization of distributions of tacrine and its hydroxylated metabolites in the brain revealed a significant decrease in levels of the metabolites in the 14-month-old mice. The results highlight strengths of the imaging technique to simultaneously investigate multiple molecular species and the drug-target effects in specific regions of the brain. The proposed approach has high potential in studies of neuropathological conditions and responses to neuroactive treatments

Controlled-pH Tissue Cleanup Protocol for Signal Enhancement of Small Molecule Drugs Analyzed by MALDI-MS Imaging

The limit of detection of low-molecular weight compounds in tissue sections, analyzed by matrix assisted laser desorption ionization mass spectrometry imaging (MALDI-MSI), was significantly improved by employing sample washing using a pH-controlled buffer solution. The pH of the washing solutions were set at values whereby the target analytes would have low solubility. Washing the tissue sections in the buffered solution resulted in removal of endogenous soluble ionization-suppressing compounds and salts, while the target compound remained in situ with minor or no delocalization during the buffered washing procedure. Two pharmaceutical compounds (cimetidine and imipramine) and one new protease inhibitor compound were successfully used to evaluate the feasibility of the pH-controlled tissue washing protocol for MALDI-MSI. Enhancement in signal-to-noise ratio was achieved by a factor of up to 10

Cross-validated Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging Quantitation Protocol for a Pharmaceutical Drug and Its Drug-Target Effects in the Brain Using Time-of-Flight and Fourier Transform Ion Cyclotron Resonance Analyzers

Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) is an established tool in drug development, which enables visualization of drugs and drug metabolites at spatial localizations in tissue sections from different organs. However, robust and accurate quantitation by MALDI-MSI still remains a challenge. We present a quantitative MALDI-MSI method using two instruments with different types of mass analyzers, i.e., time-of-flight (TOF) and Fourier transform ion cyclotron resonance (FTICR) MS, for mapping levels of the in vivo-administered drug citalopram, a selective serotonin reuptake inhibitor, in mouse brain tissue sections. Six different methods for applying calibration standards and an internal standard were evaluated. The optimized method was validated according to authorities' guidelines and requirements, including selectivity, accuracy, precision, recovery, calibration curve, sensitivity, reproducibility, and stability parameters. We showed that applying a dilution series of calibration standards followed by a homogeneously applied, stable, isotopically labeled standard for normalization and a matrix on top of the tissue section yielded similar results to those from the reference method using liquid chromatography-tandem mass spectrometry (LC-MS/MS). The validation results were within specified limits and the brain concentrations for TOF MS (51.1 +/- 4.4 pmol/mg) and FTICR MS (56.9 +/- 6.0 pmol/mg) did not significantly differ from those of the cross-validated LC-MS/MS method (55.0 +/- 4.9 pmol/mg). The effect of in vivo citalopram administration on the serotonin neurotransmitter system was studied in the hippocampus, a brain region that is the principal target of the serotonergic afferents along with the limbic system, and it was shown that serotonin was significantly increased (2-fold), but its metabolite 5-hydroxyindoleacetic acid was not. This study makes a substantial step toward establishing MALDI-MSI as a fully quantitative validated method

Simultaneous mass spectrometry imaging of multiple neuropeptides in the brain and alterations induced by experimental parkinsonism and L-DOPA therapy

Neuropeptides are important signalling molecules in the brain and alterations in their expression levels have been linked to neurological disorders such as Parkinson's disease. It is challenging to map neuropeptide changes across and within brain regions because of their low in vivo concentrations and complex post-translational processing. Consequently, the role of neuropeptides in Parkinson's disease is not well understood. Thus, we have developed and evaluated a method to image multiple neuropeptides simultaneously in both rat and primate brain tissue sections by matrix-assisted laser desorption/ionisation mass spectrometry imaging at high lateral resolution. Using a unilateral 6-hydroxydopamine rat model of Parkinson's disease, we imaged changes in enkephalins, dynorphins, tachykinins and neurotensin associated with the dopaminergic denervation and L-DOPA treatment in multiple brain regions. L-DOPA administration significantly affected neuropeptides in the globus pallidus, while neuropeptides in the caudate-putamen were mostly affected by dopamine depletion. Using high lateral resolution imaging, we observed an increase of neurotensin in the dorsal sub-region of the globus pallidus after dopamine depletion. This study highlights the capacity of mass spectrometry imaging to elucidate the dynamics of neuropeptide signalling during Parkinson's disease and its treatment

TAAR1-Dependent and-Independent Actions of Tyramine in Interaction With Glutamate Underlie Central Effects of Monoamine Oxidase Inhibition

BACKGROUND: Monoamine oxidase inhibitors (MAOIs) exert therapeutic actions by elevating extracellular levels of monoamines in the brain. Irreversible MAOIs cause serious hypertensive crises owing to peripheral accumulation of tyramine, but the role of tyramine in the central effects of MAOIs remains elusive, an issue addressed herein. To achieve robust inhibition of MAOA/B, the clinically used antidepressant tranylcypromine (TCP) was employed. METHODS: Behavioral, histological, mass spectrometry imaging, and biosensor-mediated measures of glutamate were conducted with MAOIs in wild-type and TAAR1-knockout (KO) mice. RESULTS: Both antidepressant and locomotion responses to TCP were enhanced in TAAR1-KO mice. A recently developed fluoromethylpyridinium-based mass spectrometry imaging method revealed robust accumulation of striatal tyramine on TCP administration. Furthermore, tyramine accumulation was higher in TAAR1-KO versus wild type mice, suggesting a negative feedback mechanism for TAAR1 in sensing tyramine levels. Combined histoenzymological and immunohistological studies revealed hitherto unknown TAAR1 localization in brain areas projecting to the substantia nigra/ventral tegmental area. Using an enzyme-based biosensor technology, we found that both TCP and tyramine reduced glutamate release in the substantia nigra in wild-type but not in TAAR1-KO mice. Moreover, glutamate measures in freely moving animals treated with TCP demonstrated that TAAR1 prevents glutamate accumulation in the substantia nigra during hyperlocomotive states. CONCLUSIONS: These observations suggest that tyramine, in interaction with glutamate, is involved in centrally mediated behavioral, transcriptional, and neurochemical effects of MAOIs