9 research outputs found
Analysis of Ammonium Nitrate/Urea Nitrate with Crown Ethers and Sugars as Modifiers by Electrospray Ionization-Mass Spectrometry and Ion Mobility Spectrometry
Ammonium nitrate
(AN) and urea nitrate (UN) are commonly used materials
in improvised explosive devices (IEDs). Detection by mass spectrometry
(MS) and/or ion mobility spectrometry (IMS) is traditionally difficult.
The major challenges of detecting these species arise from their ionic
nature and their low mass (for MS detection) and size (for IMS detection).
Although AN and UN both produce characteristic higher mass (and size)
cluster ions when ionized by electrospray ionization (ESI), detection
of AN/UN using cluster ions poses difficulty at trace levels because
their formation is concentration-dependent. The addition of modifiers
to the ESI process is demonstrated here to overcome some of these
challenges for the detection of AN and UN using MS and/or IMS
Precast Gelatin-Based Molds for Tissue Embedding Compatible with Mass Spectrometry Imaging
Preparation
of tissue for matrix-assisted laser desorption ionization mass spectrometry
imaging (MALDI-MSI) generally involves embedding the tissue followed
by freezing and cryosectioning, usually between 5 and 25 Ī¼m
thick, depending on the tissue type and the analyte(s) of interest.
The brain is approximately 60% fat; it therefore lacks rigidity and
poses structural preservation challenges during sample preparation.
Histological sample preparation procedures are generally transferable
to MALDI-MSI; however, there are various limitations. Optimal cutting
temperature compound (OCT) is commonly used to embed and mount fixed
tissue onto the chuck inside the cryostat during cryosectioning. However,
OCT contains potential interferences that are detrimental to MALDI-MSI,
while fixation is undesirable for the analysis of some analytes either
due to extraction or chemical modification (i.e., polar metabolites).
Therefore, a method for both fixed and fresh tissue compatible with
MALDI-MSI and histology is desirable to increase the breadth of analyte(s),
maintain the topographies of the brain, and provide rigidity to the
fragile tissue while eliminating background interference. The method
we introduce uses precast gelatin-based molds in which a whole mouse
brain is embedded, flash frozen, and cryosectioned in preparation
for mass spectrometry imaging (MSI)
Characterization of Phosphatidylcholine Oxidation Products by MALDI MS<sup><i>n</i></sup>
Phospholipid oxidation has been implicated
in the pathogenesis
and progression of numerous age-related and neurodegenerative diseases.
Despite these implications, this broad class of biomolecules remains
poorly characterized. In this work, the fragmentation patterns of
[M + H]<sup>+</sup> and [M + Na]<sup>+</sup> ions of intact phosphatidylcholine
oxidation products (OxPCs) were characterized by matrix-assisted laser
desorption/ionization tandem mass spectrometry (MALDI MS<sup><i>n</i></sup>, <i>n</i> = 2, 3, and 4). MS<sup>2</sup> of both the [M + H]<sup>+</sup> and [M + Na]<sup>+</sup> ions of
short-chain OxPCs yielded product ions related to the PC headgroup
and the fatty acid substituents. MS<sup>3</sup> of the [M + Na ā
NĀ(CH<sub>3</sub>)<sub>3</sub>]<sup>+</sup> ions yielded fragmentation
indicative of the OxPC modification; specifically, a product ion corresponding
to the neutral loss of CO<sub>2</sub> (NL of 44) was observed for
OxPCs containing a terminal carboxylic acid rather than an aldehyde.
Furthermore, MS<sup>4</sup> of the [M + Na ā HPO<sub>4</sub>(CH<sub>2</sub>)<sub>2</sub>NĀ(CH<sub>3</sub>)<sub>3</sub>]<sup>+</sup> ions resulted in fragmentation pathways dependent on the <i>sn</i>-2 fatty acid chain length and type of functional group(s).
Specifically, CHO-containing OxPCs with palmitic acid esterified to
the <i>sn</i>-1 position of the glycerol backbone yielded
a NL of 254, 2 u less than the nominal mass of palmitic acid, whereas
the analogous terminal COOH-containing OxPCs demonstrated a NL of
256. Finally, the presence of a Ī³-ketone relative to the terminal
carboxyl group resulted in CāC bond cleavages along the <i>sn</i>-2 substituent, providing diagnostic product ions for
keto-containing OxPCs. This work illustrates the enhanced selectivity
afforded by MS<sup><i>n</i></sup> on the linear ion trap
and develops a method for the identification of individual products
of PC oxidation
lāCarnitine Inhibits Lipopolysaccharide-Induced Nitric Oxide Production of SIM-A9 Microglia Cells
Microglia
are the resident immune effector cells of the central
nervous system. They account for approximately 10ā15% of all
cells found in the brain and spinal cord, acting as macrophages, sensing
and engaging in phagocytosis to eliminate toxic proteins. Microglia
are dynamic and can change their morphology in response to cues from
their milieu. Parkinsonās disease is a neurodegenerative disease,
associated with reactive gliosis, neuroinflammation, and oxidative
stress. It is thought that Parkinsonās disease is caused by
the accumulation of abnormally folded alpha-synuclein protein, accompanied
by persistent neuroinflammation, oxidative stress, and subsequent
neuronal injury/death. There is evidence in the literature for mitochondrial
dysfunction in Parkinsonās disease as well as fatty acid beta-oxidation,
involving l-carnitine. Here we investigate l-carnitine
in the context of microglial activation, suggesting a potential new
strategy of supplementation for PD patients. Preliminary results from
our studies suggest that the treatment of activated microglia with
the endogenous antioxidant l-carnitine can reverse the effects
of detrimental neuroinflammation in vitro
Isotopic Ratio Outlier Analysis of the <i>S. cerevisiae</i> Metabolome Using Accurate Mass Gas Chromatography/Time-of-Flight Mass Spectrometry: A New Method for Discovery
Isotopic ratio outlier analysis (IROA)
is a <sup>13</sup>C metabolomics
profiling method that eliminates sample to sample variance, discriminates
against noise and artifacts, and improves identification of compounds,
previously done with accurate mass liquid chromatography/mass spectrometry
(LC/MS). This is the first report using IROA technology in combination
with accurate mass gas chromatography/time-of-flight mass spectrometry
(GC/TOF-MS), here used to examine the <i>S. cerevisiae</i> metabolome. <i>S. cerevisiae</i> was grown in YNB media,
containing randomized 95% <sup>13</sup>C, or 5%<sup>13</sup>C glucose
as the single carbon source, in order that the isotopomer pattern
of all metabolites would mirror the labeled glucose. When these IROA
experiments are combined, the abundance of the heavy isotopologues
in the 5%<sup>13</sup>C extracts, or light isotopologues in the 95%<sup>13</sup>C extracts, follows the binomial distribution, showing mirrored
peak pairs for the molecular ion. The mass difference between the <sup>12</sup>C monoisotopic and the <sup>13</sup>C monoisotopic equals
the number of carbons in the molecules. The IROA-GC/MS protocol developed,
using both chemical and electron ionization, extends the information
acquired from the isotopic peak patterns for formulas generation.
The process that can be formulated as an algorithm, in which the number
of carbons, as well as the number of methoximations and silylations
are used as search constraints. In electron impact (EI/IROA) spectra,
the artifactual peaks are identified and easily removed, which has
the potential to generate ācleanā EI libraries. The
combination of chemical ionization (CI) IROA and EI/IROA affords a
metabolite identification procedure that enables the identification
of coeluting metabolites, and allowed us to characterize 126 metabolites
in the current study
LipidQC: Method Validation Tool for Visual Comparison to SRM 1950 Using NIST Interlaboratory Comparison Exercise Lipid Consensus Mean Estimate Values
As
advances in analytical separation techniques, mass spectrometry
instrumentation, and data processing platforms continue to spur growth
in the lipidomics field, more structurally unique lipid species are
detected and annotated. The lipidomics community is in need of benchmark
reference values to assess the validity of various lipidomics workflows
in providing accurate quantitative measurements across the diverse
lipidome. LipidQC addresses the harmonization challenge in lipid quantitation
by providing a semiautomated process, independent of analytical platform,
for visual comparison of experimental results of National Institute
of Standards and Technology Standard Reference Material (SRM) 1950,
āMetabolites in Frozen Human Plasmaā, against benchmark
consensus mean concentrations derived from the NIST Lipidomics Interlaboratory
Comparison Exercise
Isotopic Ratio Outlier Analysis Global Metabolomics of Caenorhabditis elegans
We demonstrate the global metabolic
analysis of Caenorhabditis elegans stress
responses using a mass-spectrometry-based
technique called isotopic ratio outlier analysis (IROA). In an IROA
protocol, control and experimental samples are isotopically labeled
with 95 and 5% <sup>13</sup>C, and the two sample populations are
mixed together for uniform extraction, sample preparation, and LC-MS
analysis. This labeling strategy provides several advantages over
conventional approaches: (1) compounds arising from biosynthesis are
easily distinguished from artifacts, (2) errors from sample extraction
and preparation are minimized because the control and experiment are
combined into a single sample, (3) measurement of both the molecular
weight and the exact number of carbon atoms in each molecule provides
extremely accurate molecular formulas, and (4) relative concentrations
of all metabolites are easily determined. A heat-shock perturbation
was conducted on C. elegans to demonstrate
this approach. We identified many compounds that significantly changed
upon heat shock, including several from the purine metabolism pathway.
The metabolomic response information by IROA may be interpreted in
the context of a wealth of genetic and proteomic information available
for C. elegans. Furthermore, the IROA
protocol can be applied to any organism that can be isotopically labeled,
making it a powerful new tool in a global metabolomics pipeline
Isotopic Ratio Outlier Analysis Global Metabolomics of Caenorhabditis elegans
We demonstrate the global metabolic
analysis of Caenorhabditis elegans stress
responses using a mass-spectrometry-based
technique called isotopic ratio outlier analysis (IROA). In an IROA
protocol, control and experimental samples are isotopically labeled
with 95 and 5% <sup>13</sup>C, and the two sample populations are
mixed together for uniform extraction, sample preparation, and LC-MS
analysis. This labeling strategy provides several advantages over
conventional approaches: (1) compounds arising from biosynthesis are
easily distinguished from artifacts, (2) errors from sample extraction
and preparation are minimized because the control and experiment are
combined into a single sample, (3) measurement of both the molecular
weight and the exact number of carbon atoms in each molecule provides
extremely accurate molecular formulas, and (4) relative concentrations
of all metabolites are easily determined. A heat-shock perturbation
was conducted on C. elegans to demonstrate
this approach. We identified many compounds that significantly changed
upon heat shock, including several from the purine metabolism pathway.
The metabolomic response information by IROA may be interpreted in
the context of a wealth of genetic and proteomic information available
for C. elegans. Furthermore, the IROA
protocol can be applied to any organism that can be isotopically labeled,
making it a powerful new tool in a global metabolomics pipeline
Expanding Per- and Polyfluoroalkyl Substances Coverage in Nontargeted Analysis Using Data-Independent Analysis and IonDecon
Per- and polyfluoroalkyl substances (PFAS) are widespread,
persistent
environmental contaminants that have been linked to various health
issues. Comprehensive PFAS analysis often relies on ultra-high-performance
liquid chromatography coupled with high-resolution mass spectrometry
(UHPLC HRMS) and molecular fragmentation (MS/MS). However, the selection
and fragmentation of ions for MS/MS analysis using data-dependent
analysis results in only the topmost abundant ions being selected.
To overcome these limitations, All Ions fragmentation (AIF) can be
used alongside data-dependent analysis. In AIF, ions across the entire m/z range are simultaneously fragmented;
hence, precursorāfragment relationships are lost, leading to
a high false positive rate. We introduce IonDecon, which filters All
Ions data to only those fragments correlating with precursor ions.
This software can be used to deconvolute any All Ions files and generates
an open source DDA formatted file, which can be used in any downstream
nontargeted analysis workflow. In a neat solution, annotation of PFAS
standards using IonDecon and All Ions had the exact same false positive
rate as when using DDA; this suggests accurate annotation using All
Ions and IonDecon. Furthermore, deconvoluted All Ions spectra retained
the most abundant peaks also observed in DDA, while filtering out
much of the artifact peaks. In complex samples, incorporating AIF
and IonDecon into workflows can enhance the MS/MS coverage of PFAS
(more than tripling the number of annotations in domestic sewage).
Deconvolution in complex samples of All Ions data using IonDecon did
retain some false fragments (fragments not observed when using ion
selection, which were not isotopes or multimers), and therefore DDA
and intelligent acquisition methods should still be acquired when
possible alongside All Ions to decrease the false positive rate. Increased
coverage of PFAS can inform on the development of regulations to address
the entire PFAS problem, including both legacy and newly discovered
PFAS