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At least 595 records · Page 33Linked to original sources
PubMed2004

Rapid and sensitive automated method for glucose monitoring in wine processing.

A rapid and sensitive automated method for glucose monitoring that might be employed during wine fermentation and processing was developed. A flow injection (FI) system coupled with an automated dilutor and the "redox-versatile" modified electrode were used to directly measure glucose in wine. To avoid interferences during wine analysis, different formulations of enzymatically modified carbon paste electrodes (CPE) were used and evaluated in oxidation and reduction mode. The best selectivity and sensitivity for glucose monitoring in real samples was obtained in cathodic mode at a fixed potential of 0 V versus Ag/AgCl using a CPE modified with glucose oxidase, horseradish peroxidase, and ferrocene as redox mediator. A total linear range of 0.02-50 g/L glucose was covered using this automated system and allowed the measurement of glucose in dry, medium, and sweet white or red wines without any sample pretreatment. The results showed a good correlation with the standard method, and the proposed method is very rapid, simple, and reliable and does not need skilled operators.

Autoanalysis↗
PubMed1999

Morphological phenotypes and functional capabilities of submandibular parenchymal cells of the ferret investigated by protein, mucosubstance and enzyme histochemistry.

Submandibular glands obtained post-mortem from mature ferrets of both sexes were examined with the use of light microscopical histochemical methods for proteins, mucosubstances and enzymes associated with cell functions or organelles. Demilunar cells showed carboxylated mucosubstances that were mainly non-sulphated, and diffuse activity for peroxidase, E600-sensitive esterase and acid phosphatase. Thiol groups were also detected in these cells. Central acinar cells showed sulphated mucosubstances, disulphides and reticular staining for thiamine pyrophosphatase. Intercalary ducts showed diffuse activity for NADH and NAD(P)H dehydrogenases. Striated ducts contained protein, tryptophan, disulphides, neutral mucosubstances and E600-sensitive esterase periluminally. Basally, the striated ductal cells showed variable activity for peroxidase, cytochrome oxidase, succinate dehydrogenase and acid phosphatase. Basolateral plasma membranes of these cells exhibited ouabain-sensitive Na,K-ATPase activity. The collecting ducts were characterized by variable periluminal staining for acid phosphatase, beta-glucuronidase, acid beta-galactosidase and E600-resistant esterase. The results suggest that the histological appearances of the acini of the submandibular gland of the ferret are dependent on the synthesis of secretory acid glycoproteins, that the striated ducts are involved with the secretion of tryptophan-rich product comprising neutral glycoproteins and showing esterase activity and with marked transport of ions and that the collecting ducts are involved with absorption.

Animals↗
PubMed2000

Secreted proteins of tobacco cultured BY2 cells: identification of a new member of pathogenesis-related proteins.

Cultured cells of tobacco BY2 secrete more than 100 proteins into culture medium. Six major proteins were purified, and partial protein sequences were determined. Five of them were found to be similar to an ascorbic acid oxidase, three peroxidase isozymes and a beta-1,3-exoglucanase, respectively. A cDNA clone encoding the remaining polypeptide, whose amino acid sequence showed no similarity with earlier reported proteins, was isolated. It encoded a putative 27 kDa protein of 242 amino acids with resemblance to WCI-5, a wheat protein induced by benzo(1,2,3)thiadiazole-7-carbothioic acid S-methyl ester (BTH) which activates genes involved in systemic acquired resistance. Transcripts of this clone accumulated upon tobacco mosaic virus infection, mechanical wounding and drought treatment, an induction profile that satisfies the definition of pathogenesis-related (PR) proteins by van Loon et al. (Plant Mol. Biol. Rep. 12 (1994) 245). No similar PR proteins have so far been reported, and therefore our newly designated NtPRp27 points to the existence of a novel PR protein family in tobacco plants.

Abscisic Acid↗
PubMed2003

On-line monitoring of the methanol concentration in Pichia pastoris cultures producing an heterologous lipase by sequential injection analysis.

An automated sequential injection analysis (SIA) system using stop-flow technique was developed to determine methanol concentration by means of the enzymatic reactions of alcohol oxidase and peroxidase. Its application as an on-line device for monitoring Pichia pastoris fermentations producing an heterologous protein was demonstrated. Linear response, observed up to 2 g l(-1), was reached by including a dilution chamber in the SIA manifold. The sampling frequency was 7 analyses per hour with a relative standard deviation lower than 4%.

Bioreactors↗
PubMed2002

Differences in the accumulation of ascorbic acid in normal, myeloperoxidase deficient and NADPH-oxidase deficient granulocytes.

Granulocytes contain large quantities of ascorbic acid (AA). The uptake mechanism is mainly restricted to the accumulation of the oxidized form, dehydroascorbate (DHA). We investigated the uptake of ascorbic acid and dehydroascorbate of normal, myeloperoxidase (MPO)-deficient, and NADPH-oxidase-deficient granulocytes. The accumulation of ascorbic acid was increased in all types of granulocytes after stimulation with phorbol-myristate-acetate, whereas the NADPH-oxidase-deficient cells showed a decreased uptake compared to normal and MPO-deficient cells. The intracellular concentration of ascorbic acid was further enhanced after incubation of granulocytes with DHA, most prominently in NADPH-oxidase-deficient granulocytes. MPO-deficient granulocytes are not able to produce HOCl after activation. The granulocytes of one individual with total MPO deficiency accumulated ascorbate in higher concentrations than did cells with partial MPO deficiency, indicating that HOCl is of minor importance for the oxidation of ascorbate. Since the ability of MPO-deficient cells to kill microorganisms is pronounced in contrast to NADPH-oxidase-deficient cells, effective mechanisms of compensating for the absence of HOCl must exist. We hypothesize that the enhanced uptake of ascorbic acid combined with an enhanced superoxide anion production may favor the generation of OH radicals via the Fenton reaction.

Antioxidants↗
PubMed1996

Sensitive and specific electrochemical sensors for charge-diffuse cations: use of lipophilic cyclodextrins and an enzyme relay for the determination of acetylcholine.

Lipophilic, alkylated cyclodextrins (CDs) are aromatic hosts which are excellent ionophores for reversibly binding size-matched, charge-diffuse cations. The application of lipophilic beta-CDs as ionophores for sensing acetylcholine chloride using a potentiometric ISE and an amperometric biosensor is described. Potentiometric ISEs, using 2,6-didodecyl-beta-CD as ionophore, showed a Nernstian response with a limit of detection -log[C] = 5.0 and selectivity coefficient -logKijPot = 4.2 (in serum levels of Na+, K+ and Ca2+). A Nernstian response is maintained in the presence of bovine serum albumin for a PVC- and a polyurethane-based electroactive membrane on initial contact. However, on prolonged contact, the polyurethane-based membranes showed a lower shift in Einitial0 (the bias potential on initial contact with analyte solution) than PVC. Amperometric biosensors were assembled by modifying screen-printed electrodes with a ferrocenyl charge shuttle, enzymes (horseradish peroxidase, choline oxidase and/or acetylcholine esterase) and a thin film comprising a polyurethane matrix, 2,3,6-triethyl-beta-CD, a plasticizer and a large anionic salt. The resulting sensor, which was capable of detecting subpicomolar levels of acetylcholine, was highly specific and was stable to storage in air and in solution. Interference from endogenous electroactive compounds was minimal.

Acetylcholine↗
PubMed2001

Cyclodextrin-modified biosensors: comparision of cyclodextrin-linked ferrocenes as mediators in sol-gel and screen-printed formats for sensing acetylcholine.

This work reports the comparison of a sol-gel and a screen-printed biosensor format using new mediators and a sensitive thin-film (ref. 1: P. Kataky and D. Parker, Analyst, 1996, 121, 1829) to enhance the sensitivity and stability of biosensors. The new mediators were per-alkylated cyclodextrin linked ferrocenes and a control, ferrocene aminocarboxylic acid. The thin film comprised a cocktail with polyurethane, plasticiser, lipophilic anion and a perethylated beta-CD. The analyte targeted was acetylcholine using the established horseradish peroxidase-choline-oxidase-acetylcholine esterase relay. The screen-printed electrode format showed a marked decrease in oxidation potential, the magnitude of the shift depended on the structure of the mediator and the membrane covering. Although lower oxidation-potentials were observed with the sol-gel format sensors, their response was more akin to aqueous solution behaviour. Electrocatalytic currents were observed suggesting a highly efficient electron transfer process.

Journal Article↗
PubMed1992

Chitin-binding proteins in potato (Solanum tuberosum L.) tuber. Characterization, immunolocalization and effects of wounding.

Tubers of potato (Solanum tuberosum L.) contain a number of chitin-binding proteins which have possible functions in defence against pathogens. A major protein of the tuber is the chitin-binding lectin which has been further characterized with respect to its antigenicity and N-terminal amino acid sequence. By using an antiserum monospecific for tuber lectin in unwounded potato the protein was found in the cytoplasm and vacuole, unusually for a hydroxyproline-rich glycoprotein, but consistent with its soluble nature in subcellular extracts. Little increased synthesis of the lectin precursor or the post-translationally modified form could be demonstrated in excised potato tuber discs. However, after wounding there is increased synthesis of another hydroxyproline-containing glycoprotein of Mr 57,000, which binds to chitin and shares common epitopes with the lectin. In comparison with the tuber lectin, this novel glycoprotein contains less hydroxyproline, but from its overall composition it is clearly not an underhydroxylated form of the tuber lectin. It differed in its N-terminal amino acid sequence and was much less glycosylated, although arabinose was still present. Synthesis of the Mr-57,000 polypeptide began after the initial burst of protein synthesis and increased, reaching a peak at 24 h after wounding. The protein was produced with its enzymes of post-translational modification, prolyl hydroxylase and arabinosyltransferase, concomitantly with the marker enzymes for wounding, phenylalanine ammonia-lyase and membrane-bound phenol oxidase and peroxidase.

Amino Acid Sequence↗
PubMed1985

Electron-paramagnetic-resonance studies of structure and function of the two-haem enzymes Pseudomonas cytochrome c peroxidase and beef heart cytochrome c oxidase.

Beef heart cytochrome c oxidase contains two cytochromes, a and a3, and Pseudomonas aeruginosa cytochrome c peroxidase has one high- and one low-potential c haem, cHP and cLP. The parallelism in co-ordination and spin states between cytochrome a and haem cHP on the one hand and between cytochrome a3 and haem cLP on the other is illustrated. The two latter haems become accessible to cyanide, when the former are reduced. Such reduction also leads to an activation of the enzymes. Mechanisms are presented in which ferryl forms of cytochromes a3 and haem cLP take part. The enzymes reach an oxidation state, formally the same as resting enzyme, but with different properties.

Animals↗
PubMed2004

The effect of oxygen on the growth and physiology of Porphyromonas gingivalis.

Oxygen constitutes a constant challenge for the survival of strict anaerobes in the oral environment. The aim of this study was to investigate the effect of oxygen on the physiology and growth of Porphyromonas gingivalis in a continuous culture system when grown under conditions of hemin limitation and excess. Results showed that, when grown in the presence of hemin at 0.5 mg/l, P. gingivalis could tolerate low levels of oxygen, being able to reach steady-state when 6% oxygen was present in the incoming gas mixture. When the hemin concentration was increased to 5 mg/l, the culture tolerated 10% oxygen. Anaerobically-grown cells were coccoid in shape, whereas those grown in the presence of oxygen were bacillary. Acetate was the predominant end-product in cultures grown in the presence of oxygen or in cultures hemin-limited. Despite some changes in the activity of Arg- and Lys-gingipain, most of the proteolytic activity was retained in the presence of oxygen. Activity of each of the three anti-oxidant enzymes tested (NADH oxidase, NADH peroxidase and SOD) was detected under all conditions and usually increased under oxygenated environments. Higher activities were also seen in the hemin-limited cultures. These results show some of the changes that occur in the physiology of P. gingivalis as a result of oxidative stress and confirm that hemin has a protective effect on the growth of the microorganism in the presence of oxygen.

Acetates↗
PubMed2002

Critical role of myeloperoxidase and nicotinamide adenine dinucleotide phosphate-oxidase in high-burden systemic infection of mice with Candida albicans.

Oxygen metabolites generated by myeloperoxidase (MPO) and nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase contribute to microbial killing by phagocytes. To compare the importance of the 2 enzymes for host defense, MPO-deficient (MPO(-/-)) mice and NADPH-oxidase-deficient mice with chronic granulomatous disease (CGD mice) were intraperitoneally infected with 3 different doses of Candida albicans, and their infection severity was analyzed. CGD mice had increased mortality and exhibited increased tissue fungal burden in a dose-dependent manner, whereas normal mice showed no symptoms. Of interest, at the highest dose, the mortality of MPO(-/-) mice was comparable to that of CGD mice, but at the lowest dose, it was the same as that of normal mice. At the middle dose, the number of fungi disseminated into various organs of the MPO(-/-) mice was comparable to that of the CGD mice at day 6 of infection, but it was significantly lower at day 14. These results suggest that MPO and NADPH-oxidase are equally important for early host defense against a large inoculum of Candida.

Animals↗
PubMed2003

Enzyme-linked sensitive fluorometric imaging of glutamate release from cerebral neurons of chick embryos.

This paper describes a method for imaging the endogenous release of glutamate from cerebral neurons. This method is based on the reactions of glutamate oxidase and peroxidase, and on the detection of hydrogen peroxide by a fluorescent substrate of peroxidase. Glutamate has been sensitively measured in vitro in the range of 20 nM to 1 microM. We used two types of Ca(2+) channel inhibitors, MK-801 and omega-Conotoxin GVIA, which act to suppress Ca(2+) transport at postsynaptic and presynaptic neurons, respectively. MK-801 did not inhibit the increase in glutamate release after KCl stimulation, while there was no increase in glutamate release after KCl stimulation when omega-Conotoxin GVIA was used, probably due to the inhibition of voltage-activated Ca(2+) channels in the presynapse. Glutamate release and Ca(2+) flow in the synaptic regions were imaged using a laser confocal fluorescence microscope. KCl-evoked glutamate release was localized around cell bodies linked to axon terminals. This procedure allows imaging that can be sensitively detected by the fluorometric enzymatic assay of endogenous glutamate release in synapses.

Animals↗
PubMed2006

Upregulation of jasmonate-inducible defense proteins and differential colonization of roots of Oryza sativa cultivars with the endophyte Azoarcus sp.

The endophyte Azoarcus sp. strain BH72 expresses nitrogenase (nif) genes inside rice roots. We applied a proteomic approach to dissect responses of rice roots toward bacterial colonization and jasmonic acid (JA) treatment. Two sister lineages of Oryza sativa were analyzed with cv. IR42 showing a less compatible interaction with the Azoarcus sp. resulting in slight root browning whereas cv. IR36 was successfully colonized as determined by nifHi::gusA activity. External addition of JA inhibited colonization of roots and caused browning in contrast to the addition of ethylene, applied as ethephon (up to 5 mM). Only two of the proteins induced in cv. IR36 by JA were also induced by the endophyte (SalT, two isoforms). In contrast, seven JA-induced proteins were also induced by bacteria in cv. IR42, indicating that IR42 showed a stronger defense response. Mass spectrometry analysis identified these proteins as pathogenesis-related (PR) proteins (Prb1, RSOsPR10) or proteins sharing domains with receptorlike kinases induced by pathogens. Proteins strongly induced in roots in both varieties by JA were identified as Bowman-Birk trypsin inhibittors, germinlike protein, putative endo-1,3-beta-D-glucosidase, glutathion-S-transferase, and 1-propane-1-carboxylate oxidase synthase, peroxidase precursor, PR10-a, and a RAN protein previously not found to be JA-induced. Data suggest that plant defense responses involving JA may contribute to restricting endophytic colonization in grasses. Remarkably, in a compatible interaction with endophytes, JA-inducible stress or defense responses are apparently not important.

Azoarcus↗
PubMed2026

Biocontrol Potential and Mechanism of Endophytic Bacillus velezensis WSR1 Against Rubber Tree Anthracnose.

Fungal leaf anthracnose, caused by Colletotrichum species, is a major leaf disease of rubber trees, significantly reducing global natural rubber yields. To explore sustainable and safe biological control strategies, eight bacterial strains were isolated from rubber tree tissues, demonstrating antagonistic activity against Colletotrichum pathogens (C. siamense and C. australisinense). Among these, WSR1 exhibited the most pronounced antifungal effect, with inhibition rates of 87.64 and 89.03% against C. siamense and C. australisinense, respectively. Genomic analysis identified WSR1 as Bacillus velezensis. In pot experiments, WSR1 exhibited preventive efficacy of 77.24 and 73.42% for C. siamense- and C. australisinense-induced anthracnose, respectively, with therapeutic efficacy of 42.28 and 45.57%. WSR1 compromised the integrity of the cell walls and membranes of both C. siamense and C. australisinense, while inducing reactive oxygen species accumulation within the hyphae. Additionally, WSR1 enhanced rubber tree resistance to anthracnose by activating defense-related enzymes, including phenylalanine ammonia-lyase, polyphenol oxidase, and peroxidase. Plate assays and genomic analysis revealed that WSR1 secretes fungal cell wall-degrading enzymes (cellulases, pectinases, and proteases) and siderophores. Furthermore, liquid chromatography-mass spectrometry and gene cluster analysis confirmed the synthesis of antagonistic secondary metabolites, such as surfactin, macrolactin H, and fengycin. This study represents the first identification of B. velezensis as a potential biocontrol agent against rubber tree anthracnose, offering a promising candidate for the eco-friendly management of rubber tree diseases.

C. australisinense↗
PubMed1985

Assay of serum cholinesterase with succinylcholine and propionylthiocholine as substrates.

Serum cholinesterase (E.C. 3.1.1.8) was assayed with succinylcholine as a substrate. The reaction was coupled with choline oxidase and peroxidase in the presence of 4-aminoantipyrine and phenol to produce a red quinone dye that was measured spectrophotometrically. The method requires 25 microliter of sample in a total volume of 1.0 ml. The mean activity for 35 adults of the usual genotype was 74.4 +/- 28 U/l (range 24-125 U/l). Succinylcholine-sensitive individuals had activities below 18 U/l. The same serum samples also were assayed with propionylthiocholine as a substrate. Activities with the two substrates showed a coefficient of correlation of 0.980 (n = 68). However, the method using propionylthiocholine showed more overlap between the activities of succinylcholine-sensitive and insensitive individuals. Assay with succinylcholine thus may offer a more effective method of screening for sensitive individuals, since some escape detection by conventional genotyping with dibucaine and fluoride.

Alcohol Oxidoreductases↗