| 1. |
- Ademark, Pia, et al.
(author)
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Softwood hemicellulose-degrading enzymes from Aspergillus niger: Purification and properties of a β-mannanase
- 1998
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In: Journal of Biotechnology. - 1873-4863. ; 63:3, s. 199-210
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Journal article (peer-reviewed)abstract
- The enzymes needed for galactomannan hydrolysis, i.e. β-mannanase, α-galactosidase and β-mannosidase, were produced by the filamentous fungus Aspergillus niger. The β-mannanase was purified to electrophoretic homogeneity in three steps using ammonium sulfate precipitation, anion-exchange chromatography and gel filtration. The purified enzyme had an isoelectric point of 3.7 and a molecular mass of 40 kDa. Ivory nut mannan was degraded mainly to mannobiose and mannotriose when incubated with the β-mannanase. Analysis by 1H NMR spectroscopy during hydrolysis of mannopentaose showed that the enzyme acts by the retaining mechanism. The N-terminus of the purified A. niger β-mannanase was sequenced by Edman degradation, and comparison with Aspergillus aculeatus β-mannanase indicated high identity. The enzyme most probably lacks a cellulose binding domain since it was unable to adsorb on cellulose.
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| 2. |
- Varga, Arthur, et al.
(author)
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Formation of phosphatidylethanol in vitro in red blood cells from healthy volunteers and chronic alcoholics.
- 2002
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In: Journal of Laboratory and Clinical Medicine. - : Elsevier BV. - 0022-2143. ; 140:2, s. 79-83
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Journal article (peer-reviewed)abstract
- Phosphatidylethanol (PEth) is an abnormal phospholipid, formed only in the presence of ethanol via a transphosphatidylation reaction of phospholipase D (PLD). PEth in blood is a promising new marker of alcohol abuse. Blood PEth is found almost exclusively in red cells. This study was performed to investigate a possible PEth formation in human red cells from alcoholics and healthy individuals, at physiologically relevant ethanol concentrations. Blood was drawn from six healthy volunteers (controls) and six chronic inpatient alcoholics. Hematological analyses were performed, and red blood cells were separated and incubated in plasma with ethanol to study PEth formation. Lipids were extracted and PEth analyzed with high pressure liquid chromatography and evaporative light-scattering detection. Incubation of red cells in 50 mM ethanol yielded detectable PEth after 12 hours. Formation of PEth was concentration dependent at 10 to 50 mM ethanol. In vitro formation of PEth was significantly higher (P <.001) in red cells from alcoholics (5.2 +/- 1.1 micromol/l) compared to controls (2.4 +/- 0.6 micromol/l) (mean +/- SD). A significant correlation (P <.01) was observed between initial mean corpuscular volume and accumulated PEth. This study demonstrates that PEth is formed in human red cells at physiologically relevant ethanol concentrations. Alcoholics accumulate about twice as much PEth than controls. The accumulation rate of PEth is slower in red cells compared to rates reported for other tissues.
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| 3. |
- Varga, Arthur, et al.
(author)
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Nonaqueous capillary electrophoresis for analysis of the ethanol consumption biomarker phosphatidylethanol.
- 2008
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In: Electrophoresis. - : Wiley. - 0173-0835 .- 1522-2683. ; 29:8, s. 1667-1671
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Journal article (peer-reviewed)abstract
- Nonaqueous CE (NACE) methodology was developed for the separation and determination of phosphatidylethanol (Peth), a new biomarker of ethanol intake. Peth is an abnormal phospholipid formed in cell membranes only in the presence of ethanol, via the transphosphatidylation reaction of phospholipase D. The NACE separation medium consisted of 80 mM ammonium acetate in 50% ACN, 33% 2-propanol, 12% hexane and 5% methanol. A stacking effect was obtained by reducing the concentration of ammonium acetate in the separation medium for all injected samples. The LOD was estimated to 1 muM (5.6 fmol) of Peth with conventional UV detection, equalling 0.4 mumol/L blood. Peth was successfully determined in extracts of human blood samples. Separation of Peth from other blood lipids in the lipid extract sample was performed in 5 min. The method facilitates smaller sample volumes and performs about ten times faster compared to earlier chromatographical methods.
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| 4. |
- Varga, Arthur, et al.
(author)
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Optimization of steam pretreatment of corn stover to enhance enzymatic digestibility
- 2004
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In: Applied Biochemistry and Biotechnology. - 1559-0291. ; 114:1-3, s. 509-523
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Journal article (peer-reviewed)abstract
- Among the available agricultural byproducts, corn stover, with its yearly production of 10 million t (dry basis), is the most abundant promising raw material for fuel ethanol production in Hungary. In the United States, more than 216 million t of corn stover is produced annually, of which a portion also could possibly be collected for conversion to ethanol. However, a network of lignin and hemicellulose protects cellulose, which is the major source of fermentable sugars in corn stover (approx 40% of the dry matter [DM]). Steam pretreatment removes the major part of the hemicellulose from the solid material and makes the cellulose more susceptible to enzymatic digestion. We studied 12 different combinations of reaction temperature, time, and pH during steam pretreatment. The best conditions (200degreesC, 5 min, 2% H2SO4) increased the enzymatic conversion (from cellulose to glucose) of corn stover more then four times, compared to untreated material. However, steam pretreatment at 190degreesC for 5 min with 2% sulfuric acid resulted in the highest overall yield of sugars, 56.1 g from 100 g of untreated material (DM), corresponding to 73% of the theoretical. The liquor following steam explosion was fermented using Saccharomyces cerevisiae to investigate the inhibitory effect of the pretreatment. The achieved ethanol yield was slightly higher than that obtained with a reference sugar solution. This demonstrates that baker's yeast could adapt to the pretreated liquor and ferment the glucose to ethanol efficiently.
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| 5. |
- Varga, Arthur
(author)
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Phosphatidylethanol in blood as a marker of alcohol abuse
- 2001
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Doctoral thesis (other academic/artistic)abstract
- Biological markers offer a way of assessing ethanol intake and determining whether a health problem is alcohol related. This study investigated the use of phosphatidylethanol (PEth) in blood as a new marker of alcohol abuse. PEth is an abnormal phospholipid, formed only in the presence of ethanol via the transphosphatidylation reaction of phospholipase D (PLD). A new HPLC-evaporative light scattering detector (ELSD) method was developed for the analysis of PEth in blood. The method has high precision and analytical specificity with few interferents and a very high biological specificity. Inmoderate drinking (>50 g ethanol/day for several weeks) is required to give detectable PEth. A dose-response between alcohol intake and level of PEth seems to occur. PEth is almost completely compartmentalized in the red blood cell (RBC) fraction of human blood. Chronic alcoholics display a one-compartment elimination of PEth in blood, with a half-life of 4 days and a normalization that takes up to 14 days after end of drinking. PEth is formed in human RBC at physiologically relevant ethanol concentrations (50 mM), with an accumulation rate that is slower compared to rates in other tissues. RBC with high MCV from alcoholics seem to accumulate twice as much PEth compared to RBC with normal MCV from healthy control individuals. Increased PEth formation by phorbol ester TPA and 1-octanol in RBC indicate involvement of protein kinase C (PKC) in PLD-formation of PEth. Enzyme inhibitor studies indicate that PEth in human RBC is metabolized by phosphatidic acid phosphohydrolase. These results support the use of PEth in blood as a reliable long-time state marker of alcohol abuse.
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| 6. |
- Vialas, Vital, et al.
(author)
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A multicentric study to evaluate the use of relative retention times in targeted proteomics
- 2017
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In: Journal of Proteomics. - : Elsevier BV. - 1874-3919. ; 152, s. 138-149
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Journal article (peer-reviewed)abstract
- Despite the maturity reached by targeted proteomic strategies, reliable and standardized protocols are urgently needed to enhance reproducibility among different laboratories and analytical platforms, facilitating a more widespread use in biomedical research. To achieve this goal, the use of dimensionless relative retention times (iRT), defined on the basis of peptide standard retention times (RT), has lately emerged as a powerful tool. The robustness, reproducibility and utility of this strategy were examined for the first time in a multicentric setting, involving 28 laboratories that included 24 of the Spanish network of proteomics laboratories (ProteoRed-ISCIII). According to the results obtained in this study, dimensionless retention time values (iRTs) demonstrated to be a useful tool for transferring and sharing peptide retention times across different chromatographic set-ups both intra- and inter-laboratories. iRT values also showed very low variability over long time periods. Furthermore, parallel quantitative analyses showed a high reproducibility despite the variety of experimental strategies used, either MRM (multiple reaction monitoring) or pseudoMRM, and the diversity of analytical platforms employed. Biological significance From the very beginning of proteomics as an analytical science there has been a growing interest in developing standardized methods and experimental procedures in order to ensure the highest quality and reproducibility of the results. In this regard, the recent (2012) introduction of the dimensionless retention time concept has been a significant advance. In our multicentric (28 laboratories) study we explore the usefulness of this concept in the context of a targeted proteomics experiment, demonstrating that dimensionless retention time values is a useful tool for transferring and sharing peptide retention times across different chromatographic set-ups.
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| 7. |
- Wurst, FM, et al.
(author)
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Ethyl glucuronide discloses recent covert alcohol use not detected by standard testing in forensic psychiatric inpatients
- 2003
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In: Alcoholism: Clinical and Experimental Research. - 0145-6008. ; 27:3, s. 471-476
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Journal article (peer-reviewed)abstract
- Background: Considerable lives and money could be saved if one could detect early stages of lapsing/ relapsing behavior in addicted persons (e.g., in safety-sensitive workplaces) and could disclose harmful drinking in social drinkers. Due to the serious public health problem of alcohol use and abuse worldwide, markers of alcohol use have been sought. Both ethyl glucuronide (EtG) and phosphatidyl ethanol (PEth) appear to have high sensitivity and specificity and a time frame of detection that may elucidate alcohol use not detected by standard testing. Our aim was to assess their potential for detecting recent covert alcohol use under controlled conditions. Methods: Thirty-five forensic psychiatric inpatients in a closed ward who had committed a substance-related offense (xi64 StGB), were followed for 12 months. The complete time spectrum of possible alcohol consumption was covered by the complementary use of breath and urinary ethanol (hours), urinary EtG (days), %carbohydrate-deficient transferrin (CDT)/PEth (weeks), and gamma-glutamyltranspeptidase (GGT)/ mean corpuscular volume (MCV) (weeks-mouths). Results: Fourteen of the 146 urine samples examined were positive for EtG. In all EtG-positive cases, patients reported alcohol consumption of between 40 and 200 g of ethanol 12-60 hr prior to testing. Urinary and breath ethanol were positive in only one case. In the blood samples, PEth was not positive in any case and %CDT did not exceed the reference value. Isoelectric focusing showed no abnormal Tf subtypes. Conclusions: The findings emphasize the diagnostic and therapeutic usefulness, specificity, and sensitivity of EtG as a marker of recent alcohol use. Such a test is needed in numerous settings, including alcohol and drug treatment (to detect lapse/relapse), in safety-sensitive work settings where use is dangerous or in other settings where use may be inappropriate (e.g., such as driving, workplace, pregnancy, or monitoring physicians or other professionals who are in recovery and working), or for testing other groups (such as children or those with medical problems) where alcohol use would be unhealthy or unsafe. The health, social and socioeconomic benefits arising from the future use of these markers is hard to overestimate.
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| 8. |
- Tabiri, S, et al.
(author)
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- 2021
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swepub:Mat__t
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