Volatile breath metabolites serve as potential disease biomarkers. Online mass spectrometry (MS) presents real-time quantification of breath volatile organic compounds (VOCs). The study aims to assess the relationship between two online analytical mass spectrometry techniques in the quantification of target breath metabolites: selected ion flow tube mass spectrometry (SIFT-MS) and proton-transfer-reaction time-of-flight mass spectrometry (PTR-ToF-MS). The two following techniques were employed: (i) direct injection with bag sampling using SIFT-MS and PTR-ToF-MS and (ii) direct injection and thermal desorption (TD) tube comparison using PTR-ToF-MS. The concentration of abundant breath metabolites, acetone and isoprene, demonstrated a strong positive linear correlation between both mass spectrometry techniques (r = 0.97, r = 0.89, respectively; p < 0.001) and between direct injection and TD tube (r = 0.97, r = 0.92, respectively; p < 0.001) breath sampling techniques. This was reflected for the majority of short chain fatty acids and alcohols tested (r > 0.80, p < 0.001). Analyte concentrations were notably higher with the direct injection of a sampling bag compared to the TD method. All metabolites produced a high degree of agreement in the detection range of VOCs between SIFT-MS and PTR-ToF-MS, with the majority of compounds falling within 95% of the limits of agreement with Bland-Altman analysis. The cross platform analysis of exhaled breath demonstrates strong positive correlation coefficients, linear regression, and agreement in target metabolite detection rates between both breath sampling techniques. The study demonstrates the transferability of using data outputs between SIFT-MS and PTR-ToF-MS. It supports the implementation of a TD platform in multi-site studies for breath biomarker research in order to facilitate sample transport between clinics and the laboratory.
- MeSH
- aceton analýza MeSH
- butadieny analýza MeSH
- dechové testy přístrojové vybavení metody MeSH
- dospělí MeSH
- hemiterpeny analýza MeSH
- hmotnostní spektrometrie přístrojové vybavení metody MeSH
- lidé MeSH
- těkavé organické sloučeniny analýza MeSH
- Check Tag
- dospělí MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
Pumping toxic substances through a cytoplasmic membrane by protein transporters known as efflux pumps represents one bacterial mechanism involved in the stress response to the presence of toxic compounds. The active efflux might also take part in exporting low-molecular-weight alcohols produced by intrinsic cell metabolism; in the case of solventogenic clostridia, predominantly acetone, butanol and ethanol (ABE). However, little is known about this active efflux, even though some evidence exists that membrane pumps might be involved in solvent tolerance. In this study, we investigated changes in overall active efflux during ABE fermentation, employing a flow cytometric protocol adjusted for Clostridia and using ethidium bromide (EB) as a fluorescence marker for quantification of direct efflux. A fluctuation in efflux during the course of standard ABE fermentation was observed, with a maximum reached during late acidogenesis, a high efflux rate during early and mid-solventogenesis and an apparent decrease in EB efflux rate in late solventogenesis. The fluctuation in efflux activity was in accordance with transcriptomic data obtained for various membrane exporters in a former study. Surprisingly, under altered cultivation conditions, when solvent production was attenuated, and extended acidogenesis was promoted, stable low efflux activity was reached after an initial peak that appeared in the stage comparable to standard ABE fermentation. This study confirmed that efflux pump activity is not constant during ABE fermentation and suggests that undisturbed solvent production might be a trigger for activation of pumps involved in solvent efflux. KEY POINTS: • Flow cytometric assay for efflux quantification in Clostridia was established. • Efflux rate peaked in late acidogenesis and in early solventogenesis. • Impaired solventogenesis led to an overall decrease in efflux.
- MeSH
- aceton MeSH
- butanoly MeSH
- Clostridium beijerinckii * MeSH
- Clostridium MeSH
- ethanol MeSH
- fermentace MeSH
- Publikační typ
- časopisecké články MeSH
We report for the first time the efficient use of accelerated solvent extraction (ASE) for extraction of ricin to analytical purposes, followed by the combined use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), and MALDI-TOF MS/MS method. That has provided a fast and unambiguous method of ricin identification for in real cases of forensic investigation of suspected samples. Additionally, MALDI-TOF MS was applied to characterize the presence and the toxic activity of ricin in irradiated samples. Samples containing ricin were subjected to ASE, irradiated with different dosages of gamma radiation, and analyzed by MALDI-TOF MS/MS for verification of the intact protein signal. For identification purposes, samples were previously subjected to SDS-PAGE, for purification and separation of the chains, followed by digestion with trypsin, and analysis by MALDI-TOF MS/MS. The results were confirmed by verification of the amino acid sequences of some selected peptides by MALDI-TOF MS/MS. The samples residual toxic activity was evaluated through incubation with a DNA substrate, to simulate the attack by ricin, followed by MALDI-TOF MS/MS analyses.
- MeSH
- aceton chemie MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- hexany chemie MeSH
- peptidy analýza chemie MeSH
- ricin analýza chemie MeSH
- rozpouštědla chemie MeSH
- sekvence aminokyselin MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice MeSH
- tandemová hmotnostní spektrometrie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Gas sensitive cerium oxide-tungsten oxide core-shell nanowires are synthesized and integrated directly into micromachined platforms via aerosol assisted chemical vapor deposition. Tests to various volatile organic compounds (acetone, ethanol, and toluene) involved in early disease diagnosis demonstrate enhanced sensitivity to acetone for the core-shell structures in contrast to the non-modified materials (i.e., only tungsten oxide or cerium oxide). This is attributed to the high density of oxygen vacancy defects at the shell, as well as the formation of heterojunctions at the core-shell interface, which provide the modified nanowires with 'extra' chemical and electronic sensitization as compared to the non-modified materials.
- MeSH
- aceton metabolismus MeSH
- cer chemie MeSH
- nanodráty chemie MeSH
- oxidy chemie MeSH
- těkavé organické sloučeniny metabolismus MeSH
- wolfram chemie MeSH
- Publikační typ
- časopisecké články MeSH
BACKGROUND: Dietary restriction together with alteration of the gastrointestinal tract results in major metabolic changes and significant weight loss in patients undergoing bariatric surgery. Current methods of measuring these changes are often inaccurate and lack a molecular basis. The objective of this study was to determine the role of exhaled ketones as non-invasive markers of nutritional status in patients undergoing surgical treatment of obesity. METHODS: Patients at different stages of treatment for obesity were recruited to this single-centre cross-sectional study. The sample time points were as follows: (i) at the time of initial attendance prior to dietary or surgical interventions, (ii) on the day of surgery following a low carbohydrate diet, and (iii) > 3 months after either Roux-en-Y gastric bypass or sleeve gastrectomy. The concentrations of ketones within breath samples were analysed by selected ion flow tube mass spectrometry. RESULTS: Forty patients were recruited into each of the three study groups. Exhaled acetone concentrations increased significantly following pre-operative diet (1396 ppb) and bariatric surgery (1693 ppb) compared to the start of treatment (410 ppb, P < 0.0001). In comparison, concentrations of heptanone (6.5 vs. 4.1 vs. 1.4 ppb, P = 0.021) and octanone (3.0 vs. 1.4 vs. 0.7 ppb, P = 0.021) decreased significantly after dieting and surgical intervention. Exhaled acetone (ρ - 0.264, P = 0.005) and octanone (ρ 0.215, P = 0.022) concentrations were observed to correlate with excess body weight at the time of sampling. Acetone and octanone also correlated with neutrophil and triglyceride levels (P < 0.05). CONCLUSION: Findings confirm breath ketones, particularly acetone, to be a potentially clinically useful method of non-invasive nutritional assessment in obese patients.
- MeSH
- aceton * analýza MeSH
- bariatrická chirurgie MeSH
- biologické markery MeSH
- dechové testy MeSH
- dieta MeSH
- dospělí MeSH
- gastrektomie * metody MeSH
- hmotnostní úbytek MeSH
- ketony * analýza MeSH
- lidé středního věku MeSH
- lidé MeSH
- morbidní obezita * chirurgie MeSH
- nutriční stav MeSH
- obezita chirurgie MeSH
- prospektivní studie MeSH
- průřezové studie MeSH
- žaludeční bypass * metody MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
The production of acetone, butanol and ethanol by fermentation of renewable biomass has potential to become a valuable industrial process. Mechanisms of solvent production and sporulation involve some common regulators in some ABE-producing clostridia, although details of the links between the pathways are not clear. In this study, we compare a wild-type (WT) Clostridium beijerinckii NRRL B-598 with its mutant strain OESpo0A, in which the gene encoding Spo0A, an important regulator of both sporulation and solventogenesis, is overexpressed in terms of solvent and acid production. We also compare morphologies during growth on two different media: TYA broth, where the WT culture sporulates, and RCM, where the WT culture does not. In addition, RT-qPCR-based analysis of expression profiles of spo0A, spoIIE, sigG, spoVD, ald and buk1 genes involved in sporulation or solvent production in these strains, were compared. The OESpo0A mutant did not produce spores and butanol titre was lower compared to the WT, but increased amounts of butyric acid and ethanol were produced. The gene spo0A had high levels of expression in the WT under non-sporulating culture conditions while other selected genes for sporulation factors were downregulated significantly. Similar observations were obtained for OESpo0A where spo0A overexpression and downregulation of other sporulation genes were demonstrated. Higher expression of spo0A led to higher expression of buk1 and ald, which could confirm the role of spo0A in activation of the solventogenic pathway, although solvent production was not affected significantly in the WT and was weakened in the OESpo0A mutant.
- MeSH
- aceton metabolismus MeSH
- bakteriální proteiny genetika MeSH
- butanoly metabolismus MeSH
- Clostridium beijerinckii genetika metabolismus MeSH
- ethanol metabolismus MeSH
- fermentace MeSH
- mutace MeSH
- polymerázová řetězová reakce MeSH
- regulace genové exprese u bakterií MeSH
- rozpouštědla metabolismus MeSH
- spory bakteriální růst a vývoj MeSH
- transkripční faktory genetika MeSH
- Publikační typ
- časopisecké články MeSH
- srovnávací studie MeSH
The ability of a bubble column reactor (BCR) to biodegrade a mixture of styrene and acetone vapors was evaluated to determine the factors limiting the process efficiency, with a particular emphasis on the presence of degradation intermediates and oxygen levels. The results obtained under varied loadings and ratios were matched with the dissolved oxygen levels and kinetics of oxygen mass transfer, which was assessed by determination of kLa coefficients. A 1.5-L laboratory-scale BCR was operated under a constant air flow of 1.0 L.min-1, using a defined mixed microbial population as a biocatalyst. Maximum values of elimination capacities/maximum overall specific degradation rates of 75.5 gC.m-3.h-1/0.197 gC.gdw-1.h-1, 66.0 gC.m-3.h-1/0.059 gC.gdw-1.h-1, and 45.8 gC.m-3.h-1/0.027 gC.gdw-1.h-1 were observed for styrene/acetone 2:1, styrene-rich and acetone-rich mixtures, respectively, indicating significant substrate interactions and rate limitation by biological factors. The BCR removed both acetone and styrene near-quantitatively up to a relatively high organic load of 50 g.m-3.h-1. From this point, the removal efficiencies declined under increasing loading rates, accompanied by a significant drop in the dissolved oxygen concentration, showing a process transition to oxygen-limited conditions. However, the relatively efficient pollutant removal from air continued, due to significant oxygen mass transfer, up to a threshold loading rate when the accumulation of acetone and degradation intermediates in the aqueous medium became significant. These observations demonstrate that oxygen availability is the limiting factor for efficient pollutant degradation and that accumulation of intermediates may serve as an indicator of oxygen limitation. Microbial (activated sludge) analyses revealed the presence of amoebae and active nematodes that were not affected by variations in operational conditions.
A 2:1 (w/w) mixture of styrene (STY) and acetone (AC) was subjected to lab-scale biofiltration under varied loading in both a trickle bed reactor (TBR) and biofilter (BF) to investigate substrate interactions and determine the limits of biofiltration efficiency of typical binary air pollutant mixtures containing both hydrophobic and polar components. A comparison of the STY/AC mixture degradation in the TBR and BF revealed higher pollutant removal efficiencies and degradation rates in the TBR, with the pollutant concentrations increasing up to the overloading limit. The maximum styrene degradation rates were 12 and 8 gc m(-3) h(-1) for the TBR and BF, respectively. However, the order of performance switched in favor of the BF when the loading was conducted by increasing air flow rate while keeping the inlet styrene concentration (Cin) constant in contrast to loading by increasing Cin. This switch may be due to a drastic difference in the effective surface area between these two reactors, so the biofilter becomes the reactor of choice when the rate-limiting step switches from biochemical processes to mass transfer by changing the loading mode. The presence of acetone in the mixture decreased the efficiency of styrene degradation and its degradation rate at high loadings. When the overloading was lifted by lowering the pollutant inlet concentrations, short-term back-stripping of both substrates in both reactors into the outlet air was observed, with a subsequent gradual recovery taking several hours and days in the BF and TBR, respectively. Removal of excess biomass from the TBR significantly improved the reactor performance. Identification of the cultivable strains, which was performed on Day 763 of continuous operation, showed the presence of 7 G(-) bacteria, 2 G(+) bacteria and 4 fungi. Flies and larvae of Lycoriella nigripes survived half a year of the biofilter operation by feeding on the biofilm resulting in the maintenance of a nearly constant pressure drop.
- MeSH
- aceton chemie MeSH
- bioreaktory * MeSH
- filtrace metody MeSH
- látky znečišťující vzduch chemie MeSH
- lidé MeSH
- styren chemie MeSH
- znečištění ovzduší prevence a kontrola MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
Flow cytometry, in combination with fluorescent staining, was used to evaluate population heterogeneity in acetone-butanol-ethanol fermentation that was carried out with type strain Clostridium beijerinckii NCIMB 8052 and non-type C. pasteurianum NRRL B-598. A combination of propidium iodide (PI) and carboxyfluorescein diacetate (CFDA), PI plus Syto-9 and bis-oxonol (BOX) alone were employed to distinguish between active and damaged cells together with simultaneous detection of spores. These strategies provided valuable information on the physiological state of clostridia. CFDA and PI staining gave the best separation of four distinct subpopulations of enzymatically active cells, doubly stained cells, damaged cells and spores. Proportional representation of cells in particular sub-regions correlated with growth characteristics, fermentation parameters such as substrate consumption and product formation in both species under different cultivation conditions.
- MeSH
- aceton metabolismus MeSH
- biomasa MeSH
- bioreaktory MeSH
- butanoly metabolismus MeSH
- Clostridium fyziologie MeSH
- energetický metabolismus * MeSH
- ethanol metabolismus MeSH
- fermentace * MeSH
- glukosa metabolismus MeSH
- mikrobiální viabilita MeSH
- průtoková cytometrie MeSH
- spory bakteriální * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
In vitro impacts of five organic solvents on cytochrome P450 (CYP450) enzyme activity were investigated using hepatic microsomes of rainbow trout. The rates of several CYP450-mediated reactions were investigated at solvent concentrations ranging from 0.01% to 3%. The solvents greatly affected all tested reactions. In at least 0.8% ethanol, 2% methanol or acetone, 1% acetonitrile or 3% dimethyl sulfoxide (DMSO), 7-ethoxyresorufin-O-deethylase (EROD) activity decreased and at 3% acetonitrile or ethanol, it was undetected. At 3%, all tested solvents except methanol reduced 7-benzyloxy-4-trifluoromethylcoumarin-O-debenzylase (BFCOD) activity, but at low concentrations of ethanol (2% and lower) or DMSO (1% and lower), it was induced. This was not seen with the inclusion of a pre-incubation step. p-Nitrophenolhydroxylase (PNPH) activity was not affected at concentrations below 1% DMSO, and at 2% acetonitrile it was reduced, as it was above 1% methanol or 0.5% ethanol. Acetone did not affect PNPH activity with or without a pre-incubation step. In general, the degree of inhibition was similar with and without the pre-incubation step. We conclude that the concentration of organic solvent for solubilizing the substrate and inhibitor in in vitro microsomal studies should be minimized.
- MeSH
- aceton farmakologie MeSH
- acetonitrily farmakologie MeSH
- dimethylsulfoxid farmakologie MeSH
- ethanol farmakologie MeSH
- izoenzymy MeSH
- jaterní mikrozomy účinky léků enzymologie MeSH
- játra účinky léků enzymologie MeSH
- kinetika MeSH
- methanol farmakologie MeSH
- Oncorhynchus mykiss metabolismus MeSH
- rozpouštědla farmakologie MeSH
- substrátová specifita MeSH
- systém (enzymů) cytochromů P-450 metabolismus MeSH
- vztah mezi dávkou a účinkem léčiva MeSH
- zvířata MeSH
- Check Tag
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
