1,720,983 research outputs found
Improved chromatographic purification of peroxidase and beta–glucosidase from Hordeum vulgare seedlings
No full textPeroxidases (E.C. 1.11.1.7., hydrogen donor oxidoreductase) utilize hydrogen peroxide or substituted peroxides for the oxidation of a large number of substrates. Peroxidases are widely distributed and have been isolated from many higher plants (1). The wide distribution of the enzyme suggests that it could be of great biological importance, but the physiological functions and metabolic control of these enzymes are still poorly understood. The simultaneous presence of amine oxidase and peroxidase in cell walls suggests that the peroxide generated on oxidation of the amines could be utilized by the peroxidase (2,3). Recently we have purified an amine oxidase from Hordeum vulgare (4) and we have attempted to purify the peroxidase in order to study in vitro the reconstituted coupled system. Beta-glucosidase (beta-D-glucoside glucohydrolase E.C. 3.2.1.21.) is capable of transforming glucosides in glucose and the corresponding aglycone or disaccarides as cellobiose, sophorose, gentiobiose. This enzyme is widely distributed in plants, fungi, bacteria, yeasts and animals (5, 6). In the homogenate of Hordeum vulgare seedlings we also found beta-glucosidase activity and also attempted to purify beta-glucosidase. This enzyme copurified whit peroxidase up to the last step. We report here the isolation of peroxidase and Beta-glucosidase from Hordeum vulgare seedlings: some molecular and kinetic properties are given
Properties of Thermus aquaticus b-NADH oxidase immobilised on various supports
No full textbeta-NADH oxidase purified from Thermus aquaticus was covalently immobilised on various solid supports. The preparations obtained were compared with the soluble enzyme for activity and kinetic properties. Activated glutaryl-PVA was found to be the best support. The immobilised enzyme was less stable at high temperatures than the soluble enzyme. No differences could be detected in the presence of organic solvents.beta-NADH oxidase purified from Thermus aquaticus was covalently immobilised on various solid supports. The preparations obtained were compared with the soluble enzyme for activity and kinetic properties. Activated glutaryl-PVA was found to be the best support. The immobilised enzyme was less stable at high temperatures than the soluble enzyme. No differences could be detected in the presence of organic solvents
3-Hydroxykynurenine as a substrate/activator for mushroom tyrosinase
No full text3-Hydroxykynurenine is a tryptophan metabolite with an o-aminophenol structure. It is both a tyrosinase activator and a substrate, reducing the lag phase, stimulating the monophenolase activity, and being oxidized to xanthommatin. In the early stage of monophenol hydroxylation, catechol accumulation takes place, whereas 3-hydroxykynurenine is substantially unchanged and no significant amounts of the o-quinone are produced. These results suggest an activating action of 3-hydroxykynurenine toward o-hydroxylation of monophenols. 3-Hydroxykynurenine could therefore well act as a physiological device to control phenolics metabolism to catechols and quinonoids. (C) 2003 Elsevier Science (USA). All rights reserved
Photometric assay for polyphenol oxidase activity in olives, olive pastes, and virgin olive oils
No full textA photometric method is proposed that allows the determination of phenolase activity in olive fruits, olive pastes, and virgin olive oil. The method can also be used to quantify partially purified phenolase from olives, and is based on the coupling between 4-methyl-o-benzoquinone, the reaction product of phenolase toward its substrate 4-methylcatechol, and the aromatic amine 4-amino-N,N-diethylaniline. The deep-blue adduct arising from this reaction has been characterized by means of nuclear magnetic resonance and mass spectrometric techniques and identified as 4-(4'-diethylamino-phenylimino)-2-hydroxy-5-methyl-cyclohexa-2,5-dienone . This compound shows an absorption band, centered (in dichloromethane) at 617 nm, with an c of 11,080 M(-1)cm(-1). The main advantage of the proposed method resides in the high absorption coefficient of the adduct and its ultraviolet/visible absorption pattern, with a gimel(max) in a spectral region void of significant interferences by the pigments that ultimately will probably be present in the extracts to be tested by this proposed method. The method has proven to be sensitive, specific, and reliable
A dyed substrate for the assay of endo-1,4-b-glucanases
No full textA new dyed substrate was prepared for the rapid determination of endo-1,4-beta-glucanases. Carboxymethylcellulose was coupled with Ruthenium red to obtain a violet powder, which was stable enough to allow for reproducible assays. The spontaneous reaction is based on ionic interactions between the negatively charged carboxymethylcellulose and the complex cation of the dye. The enzymic assay is based on spectrophotometric measurement at 535 nm of the enzyme-released dyed fragments with low-molecular-weight filterable through a 0.22 mu m filter. The release of coloured fragments from this dyed substrate was proportional to its solubilization. The absorbance was directly proportional to the enzyme amount in the range 0.5-5.5 mU enzyme (A(535) = 0.1-0.95). This enzymic assay is advantageous for both rapid time of analysis and sensitivity
Note. Xylose production from durum wheat bran: enzymic versus chemical methods
No full textBoth acidic and enzymic hydrolytic processes of durum wheat bran to obtain high xylose syrups are discussed in terms of yields and operative conditions. Mild sulphuric hydrolysis gave the best results (similar to400 mg of aldopentoses per gram of pre-extracted wheat bran within 4-6 h) whereas hydrochloric and especially phosphoric acids led to low yields. Preliminary work with a highly active recombinant xylanase (endoxylanase) led to deep solubilisation of arabinoxylans, mainly leading to oligosaccharides and their acyl (essentially feruloyl) derivatives. Mild sulphuric hydrolysis still seems to be the treatment of choice for xylose recovery from durum wheat bran
Biodegradation of polycyclic aromatic hydrocarbons by pleurotus sajor-caju
No full textPolycyclic aromatic hydrocarbons (PAHs) are considered priority pollutants because they have potentially dangerous effects on the environment and human health. Bioremediation has recently become attractive to restore polluted sites, because it is not expensive and fulfils the most important properties required by the current regulations. In this work the capability of a white-rot fungi, Pleurotus sajor-caju, was assessed for degradation of pyrene and chrysene. The main objective was to investigate the effects of pollutant concentration on the mycelium growth and find the conditions that can enhance the microorganism tolerance when exposed to pyrene and chrysene. The tests carried out in Petri dishes showed that chrysene inhibited mycelium growth, whereas pyrene was well tolerated. Experiments in liquid medium evidenced that the mycelium was able to degrade pyrene with a removal efficienty greater than 90%
Complete and efficient enzymic hydrolysis of pretreated wheat straw
No full textA fractionation of wheat straw components in a two-step chemical pretreatment is proposed. Hemicelluloses were hydrolysed
by dilute sulphuric acid, allowing a substantial recovery of crystalline xylose. Lignin was removed by means of a mild
alkaline/oxidative solubilisation procedure, involving no sulphite or chlorine and its derivatives. The use of diluted reagents and
relatively low temperatures, was both cheap and environmentally friendly. The pretreated material was nearly pure cellulose,
whose enzymic hydrolysis proceeded fast and with high yields, leading to high glucose syrups of remarkable purity
Pyrene and chrysene tolerance and biodegradation capability of pleurotus sajor-caju
Full text linkIntroduction: The present work focused on the biodegradation capability of a white-rot fungus, the Pleurotus sajor-caju, when exposed to polycyclic aromatic hydrocarbons. Methods: The research was carried out by using in vitro systems developed on Petri dishes, to evaluate the fungal tolerance to pyrene and chrysene, followed by experiments in liquid medium. The first experimental campaign was necessary to evaluate the conditions promoting fungal growth and tolerance (presence of surfactants, peptone, copper sulphate and lecithin) and it was designed and analysed using statistical techniques. Results: It was found that the fungal population growth is strongly inhibited by chrysene presence. On the other hand, pyrene had a mild negative impact on the mycelia growth, which seemed to be positively influenced by the presence of Tween 80 and copper sulphate. Starting from these results, the behaviour of Pleurotus sajor-caju in presence of pyrene was investigated in liquid medium. Results showed that the depletion of pyrene was evident during a period of 20 days, and removal efficiency was greater than 90%
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