8 research outputs found
Isotope Cluster-Based Compound Matching in Gas Chromatography/Mass Spectrometry for Non-Targeted Metabolomics
Gas chromatography coupled to mass
spectrometry (GC/MS) has emerged
as a powerful tool in metabolomics studies. A major bottleneck in
current data analysis of GC/MS-based metabolomics studies is compound
matching and identification, as current methods generate high rates
of false positive and false -negative identifications. This is especially
true for data sets containing a high amount of noise. In this work,
a novel spectral similarity measure based on the specific fragmentation
patterns of electron impact mass spectra is proposed. An important
aspect of these algorithmic methods is the handling of noisy data.
The performance of the proposed method compared to the dot product,
the current gold standard, was evaluated on a complex biological data
set. The analysis results showed significant improvements of the proposed
method in compound matching and chromatogram alignment compared to
the dot product
Fragment Formula Calculator (FFC): Determination of Chemical Formulas for Fragment Ions in Mass Spectrometric Data
The accurate determination
of mass isotopomer distributions (MID)
is of great significance for stable isotope-labeling experiments.
Most commonly, MIDs are derived from gas chromatography/electron ionization
mass spectrometry (GC/EI-MS) measurements. The analysis of fragment
ions formed during EI, which contain only specific parts of the original
molecule can provide valuable information on the positional distribution
of the label. The chemical
formula of a fragment ion is usually applied to derive the correction
matrix for accurate MID calculation. Hence, the correct assignment
of chemical formulas to fragment ions is of crucial importance for
correct MIDs. Moreover, the positional distribution of stable isotopes
within a fragment ion is of high interest for stable isotope-assisted
metabolomics techniques. For example, <sup>13</sup>C-metabolic flux
analyses (<sup>13</sup>C-MFA) are dependent on the exact knowledge
of the number and position of retained carbon atoms of the unfragmented
molecule. Fragment ions containing different carbon atoms are of special
interest, since they can carry different flux information. However,
the process of mass spectral fragmentation is complex, and identifying
the substructures and chemical formulas for these fragment ions is
nontrivial. For that reason, we developed an algorithm, based on a
systematic bond cleavage, to determine chemical formulas and retained
atoms for EI derived fragment ions. Here, we present the fragment
formula calculator (FFC) algorithm that can calculate chemical formulas
for fragment ions where the chemical bonding (e.g., Lewis structures)
of the intact molecule is known. The proposed algorithm is able to
cope with general molecular rearrangement reactions occurring during
EI in GC/MS measurements. The FFC algorithm is able to integrate stable
isotope labeling experiments into the analysis and can automatically
exclude candidate formulas that do not fit the observed labeling patterns. We applied the FFC algorithm to create a fragment
ion repository that contains the chemical formulas and retained carbon
atoms of a wide range of trimethylsilyl and <i>tert</i>-butyldimethylsilyl
derivatized compounds. In total, we report the chemical formulas and
backbone carbon compositions for 160 fragment ions of 43 alkylsilyl-derivatives
of primary metabolites. Finally, we implemented the FFC algorithm
in an easy-to-use graphical user interface and made it publicly available
at http://www.ffc.lu
Levels of intracellular metabolite pools in aerosol-exposed macrophages.
<p>Pools of (a) lactic acid, (b) succinic acid, and itaconic acid were measured. Levels represent the mean of 9 biological replicates. *** = p < 0.001. Error bars represent s.e.m.</p
HFO particles induce activation of immune response in RAW 264.7 macrophages.
<p>(a) The Gene Ontology term GO:0006955, corresponding to activation of immune response, was found to be significantly up-regulated in HFO-treated samples (p = 0.059) and not regulated in the DF-treated samples. (b) Model of how the regulated proteins found in this study affect the NF-kB immune response pathway in the cell. Stimulation of the toll-like receptor (TLR2) leads to activation of NF-kB. Tumor necrosis factor alpha-induced protein 8-like protein 2 (TNFAIP8L2) acts as a negative regulator of TLR2, preventing hyperresponsiveness of the immune system, and inhibiting NF-kappa-B activation. Peroxiredoxin 2 (Pdrx2) reduces hydrogen peroxide, inhibiting NF-kappa-B activation.</p
Summary of the main HFO- and DF-particle exposure effects.
<p>The arrows indicate the direction of regulation for cellular functions derived from the most statistically significant enriched Gene Ontology terms from the transcriptome, proteome, and metabolome (details in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0126536#pone.0126536.s012" target="_blank">S2 Table</a>).</p><p><sup>x</sup> BEAS-2B up, A549 down</p><p>* BEAS-2B down, A549 up</p><p>Summary of the main HFO- and DF-particle exposure effects.</p
Experimental set-up and global omics analyses.
<p>(A) An 80 KW common-rail-ship diesel engine was operated with heavy fuel oil (HFO) or refined diesel fuel (DF). The exhaust aerosols were diluted and cooled with clean air. On-line real-time mass spectrometry, particle-sizing, sensor IR-spectrometry and other techniques were used to characterise the chemical composition and physical properties of the particles and gas phase. Filter sampling of the particulate matter (PM) was performed to further characterise the PM composition. Lung cells were synchronously exposed at the air-liquid-interface (ALI) to aerosol or particle-filtered aerosol as a reference. The cellular responses were characterised in triplicate at the transcriptome (BEAS-2B), proteome and metabolome (A549) levels with stable isotope labelling (SILAC and <sup>13</sup>C<sub>6</sub>-glucose). (B) Heatmap showing the global regulation of the transcriptome, proteome and metabolome.</p
Effects of shipping particles on lung cells.
<p>The net effects from the particles were referenced against the gaseous phase of the emissions. (A) Number of the regulated components in the transcriptome shows more genes regulated by the DF than the HFO particles (in BEAS-2B cells). Similar results were observed for the proteome (B) and metabolome (C) (in A549 cells). (D) Meta-analyses for the transcriptome and proteome using the combined Gene Ontology (GO) term analysis of the 10% most regulated transcripts and proteins. Individual GO terms are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0126536#pone.0126536.s012" target="_blank">S2 Table</a>; the hierarchical pathways are indicated on the right. (E) Gene regulation of Wiki-pathway bioactivation; (F) gene regulation of Wiki-pathway inflammation; g, secreted metabolites; and h, metabolic flux measurements using <sup>13</sup>C-labelled glucose. For all experiments, n = 3.</p
Chemical and physical aerosol characterisation.
<p>(A) The ship diesel engine was operated for 4 h in accordance with the IMO-test cycle. (B) Approximately 28 ng/cm<sup>2</sup> and 56 ng/cm<sup>2</sup> were delivered to the cells from DF and HFO, respectively, with different size distributions. The HFO predominantly contained particles <50 nm, and the DF predominantly contained particles >200 nm, both in mass and number. (C) Number of chemical species in the EA particles. (D) Transmission electron microscope (TEM) images and energy-dispersive X-ray (EDX) spectra of DF-EA and HFO-EA; heavy elements (black speckles, arrow); and contributions of the elements V, P, Fe and Ni in the HFO particles using EDX (* = grid-material). (E) Exemplary EA concentrations (right) and concentration ratios (left) for particulate matter-bound species. For all experiments, n = 3.</p