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  • 1980-1984  (7)
  • 1
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 26 (1984), S. 560-563 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Additional Material: 4 Ill.
    Type of Medium: Electronic Resource
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  • 2
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 22 (1980), S. 2251-2272 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: A method of enzyme immobilization by graft copolymerization on polysaccharides is reported. Glycidylmethacrylate was used as a vinylating reagent and the reaction product with enzymes (HRP, GOD, Am, ChT) was copolymerized with different matrices (cellulose, Sepharose, Sephadex, Starch). Various factors affect the final activity of copolymers; these include the redox system, the type of support, and the quantity and type of vinyl monomer added. Using a fixed quantity of enzyme and support (3 mg enzyme, 100 mg support), the coupling efficiency varied from 2 to 50%. The most important characteristics in these immobilized systems were tested (stability in continuous washing, kinetic characteristics, storage, thermal, and lyophilization stability). Immobilized-enzyme graft copolymers have very similar kinetic behavior to that of the free enzyme. Diffusion is not seriously limited, as shown by kinetic parameters and energy activation values, and this indicates that the immobilization reaction does not alter the enzymatic activity.
    Additional Material: 10 Ill.
    Type of Medium: Electronic Resource
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  • 3
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 24 (1982), S. 207-216 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The performance of a new method of enzyme immobilization based on photochemically initiated direct graft copolymerization was recently investigated. The immobilization reaction can be carried out in a simple way and by carefully selecting the reaction conditions, the enzyme-graft copolymer can be obtained as the main reaction product. Coupling efficiency of glucose oxidase has been found to depend only on the amount of photocatalyst (FeCl3) fixed on Sepharose used as polysaccharide support. Small quantities of glycidymethacrylate (GMA) (0.25 g/g dry Sepharose) are sufficient but necessary to achieve the best enzyme coupling efficiency (20-40%). Enzyme immobilization occurs very rapidly and the entire reaction occurs within 60 min. Reaction patterns and physicochemical characteristics of the obtained enzyme-graft copolymers exclude the glucose oxidase entrapment: therefore a covalent attachment mechanism may be proposed. The kinetic parameters of immobilized glucose oxidase (Km′ = 2.0 × 10-2M) are quite similar to those of free enzyme (Km = 1.93 × 10-2M), and no diffusion limitation phenomena are evidenced in samples having different enzyme or polymer content. Lyophilization, thermostability, and long-term continuous operation also have been investigated. The advantages of this method over that using vinylenzyme copolymerization are discussed.
    Additional Material: 3 Ill.
    Type of Medium: Electronic Resource
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  • 4
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 25 (1983), S. 735-744 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The characteristics of horseradish peroxidase (HRP) immobilized onto Sepharose by a photochemical-initiated graft copolymerization are presented. Active copolymers were synthesized using different amounts of glycidylmethacrylate (GMA), bisacryloylpiperazine (BAP), or 1,3,5-hexhydrotriacryloyl-s-triazine (HTsT) as functional monomer. The activities, the K′m values (pGMA) copolymers: 0.53-0.76 × 10-4M; pBAP copolymers: 0.90-1.4 × 10-4; pHTsT copolymers: 1.8-2.6 × 10-4M and the thermal stabilities of the enzyme copolymers were strictly connected to the type of polymer. By varying the polymer amount present in a given copolymer, significant differences were found in the thermostability properties of pBAP and pHTsT copolymers both when checked in water or in phosphate buffer. No differences were found for pGMA copolymers. The samples in which there are the lowest pBAP or pHTsT content resulted the most stable. The activity retained after 240 min at 60°C by free HRP and pGMA-HRP was 30% whereas by pBAP-HRP and pHTsT-HRP it was 50 and 75% of the original. Operational stability of the materials was in agreement with thermostability data. These results are discussed in terms of enzyme microenvironment which is strongly affected by the different network of the three polymers.
    Additional Material: 7 Ill.
    Type of Medium: Electronic Resource
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  • 5
    Electronic Resource
    Electronic Resource
    New York, NY [u.a.] : Wiley-Blackwell
    Biotechnology and Bioengineering 24 (1982), S. 1-7 
    ISSN: 0006-3592
    Keywords: Chemistry ; Biochemistry and Biotechnology
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Biology , Process Engineering, Biotechnology, Nutrition Technology
    Notes: The effect of several factors on the activity and stability of alcohol dehydrogenase, glyceraldehyde-3-phosphate dehydrogenase, and 20β-hydroxysteroid dehydrogenase, both free and immobilized on CNBr-activated Sepharose 4B, was investigated. Enzymes were im- mobilized under different conditions including various degrees of matrix activation, variable amounts of protein, in the presence, or in the absence of, additives (coenzymes, dithioth- reitol, salts). Activity recovery was in general satisfactorily high with 20β-hydroxysteroid dehydrogenase, low with glyceraldehyde-3-phosphatedehydrogenase, and markedly linked to the concentration of immobilized protein with alcohol dehydrogenase. In the latter case the advantageous stabilizing effect of high enzyme concentrations was notably diminished by the parallel decrease of the effectiveness factor. The effect of high concentrations of anions of the Hofmeister series was examined. It was found that 1M phosphate and 0.5M sulfate dramatically stabilize both free and immobilized enzymes against inactivation by temperature and urea. Km, values of apolar substrates were considerably lowered by the two anions while Km values of polar substrates were not affected. In some cases Vmax values also were influenced by high concentrations of these anions. The present results appear of interest particularly in view of enzyme utilization for analytical as well as for preparative purposes.
    Additional Material: 1 Ill.
    Type of Medium: Electronic Resource
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  • 6
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    Angewandte Makromolekulare Chemie 91 (1980), S. 161-178 
    ISSN: 0003-3146
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Physics
    Description / Table of Contents: Es wird über vergleichende Ergebnisse bei der Herstellung und Charakterisierung von Enzymen berichtet, die mittels Reaktion von Glycidylmethacrylat-(PGMA)-Polymeren mit verschiedenen Polysaccharidmatrizen immobilisiert wurden. Es wurden Sepharosecopolymere, die 25 bis 50% synthetisches Polymers enthielten, verwendet, um die besten Immobilisierungsbedingungen (pH-Wert, Behandlungsdauer und -temperatur) für Meerrettich-Peroxidase (HPR) und Glucoseoxidase (GOD) zu ermitteln. Die spezifische Aktivität, das Aufzieh- und Kupplungsvermögen von HRP und GOD auf verschiedenen Matrizen sind weitgehend von deren Art abhängig, wäh-rend der Gehalt an Polymerem eine untergeordnete Rolle spielt. Die HRP-Muster ergaben ein Aufzieh- und Kupplungsvermögen zwischen 0.8 und 1.5%, wärend bei GOD-Mustern ein dreimal höheres Aufziehvermögen festgestellt wurde. Immobilisierte HRP- und GOD-Muster wiesen höhere Km′-Werte als die entsprechenden freien Enzyme auf, was auf Erscheinungen von Diffusionsbegrenzung hindeutet.Es wurden ebenfalls die Lagerungs-, Wärme und Arbeitsbestlndigkeit untersucht. Die Lagerungsbestädigkeit kann im allgemeinen als zufriedenstellend angesehen werden (50% Restaktivität nach 360 Tagen). Sepharose- und Stärke-HRP-Copolymere besaßen höhere Wärmebestlndigkeit als das freie Enzym. Die mittels kontinuierlichen Tests an immobilisierten HRP-Mustern festgestellte Restaktivität war vom Trägermaterial abhängig. HRP-PGMA-Cellulosemuster zeigten die besten Ergebnisse (50% Restaktivitat nach 16 Tagen). PGMA-Pfropfcopolymere wurden ebenfalls zur Immobilisierung anderer Enzyme, wie Amylase, Chymotrypsin und Cellulase herangezogen.
    Notes: Comparative results obtained in preparing and characterizing samples of enzymes immobilized by reaction with polyglycidylmethacrylate (PGMA) copolymers with different polysaccharide matrices are reported. Sepharose copolymers having between 25 and 50% synthetic polymer were used to find the best immobilization conditions of horseradish peroxidase (HRP) and glucose-oxidase (GOD) (pH, time, temperature, enzyme cncentration). Activity, enzyme loading and coupling efficiency of immobilized HRP and GOD are greatly dependent on the type of matrix while the polymer content is less important. Coupling efficiencies between 0.8 and 1.5% have been obtained for HRP samples, whereas for GOD samples coupling efficiencies three times greater were obtained. HRP and GOD immobilized samples show Km′ values greater than those of corresponding free enzymes and this indicates diffusion limitation phenomena.Storage, thermal and operational stability were also studied. In general the storage stability could be considered satisfactory (50% residual activity after 360 days). Sepharose and starch HRP-copolymers had an improved thermal stability compared with that of free enzyme. Residual activity found in continuous operation tests carried out on HRP-immobilized samples turned out to be dependent on support. HRP-PGMA-Cellulose sample gave the best results (50% residual activity after 16 days). PGMA-graft-copolymers have also been used to immobilize other enzymes such as α-amylase, α-chymotrypsin and cellulase.
    Additional Material: 8 Ill.
    Type of Medium: Electronic Resource
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  • 7
    Electronic Resource
    Electronic Resource
    Weinheim : Wiley-Blackwell
    Angewandte Makromolekulare Chemie 104 (1982), S. 129-143 
    ISSN: 0003-3146
    Keywords: Chemistry ; Polymer and Materials Science
    Source: Wiley InterScience Backfile Collection 1832-2000
    Topics: Chemistry and Pharmacology , Physics
    Description / Table of Contents: Es wird über ein Verfahren zur Immobilisierung von Enzymen mittels Pfropf-copolymerisation auf Polysacchariden berichtet. Als Vinylierungsreagens wurde Bisacryloylpiperazin verwendet. Das Reaktionsprodukt wurde mit verschiedenen Enzymen (HRP, GOD, Am, ChT, Cel) auf verschiedenen Trägern (Cellulose, Sepharose, Sephadex, Stärke) copolymerisiert. Die Immobilisierungsparameter, welche die Aktivität des Copolymeren beeinflussen, wurden auf die Möglichkeit, die Unlöslichkeit der Meerrettichperoxidase auf Cellulose zu erreichen, untersucht. Es handelte sich dabei um folgende Parameter: pH, Zeit und Temperatur der Aktivierungsreaktion des Enzyms mit Bisacryloylpiperazin. Unter optimalen Immobilisierungsbedingungen zeigte die Aktivität des Copolymeren eine lineare Abhängigkeit von der Enzymkonzentration. Die Wirksamkeit der Bindung wird von der Art des Enzyms bestimmt und liegt zwischen 7 und 20%.Die wichtigsten Eigenschaften (Festigkeit gegen kontinuierliches Waschen, kinetisches Verhalten, Lagerungs-, Wärme- und Gebrauchsbeständigkeit) dieses immobilisierten Enzymsystems wurden untersucht und mit den Eigenschaften ähnlicher glycidylmethacrylat-aktivierter Enzymsysteme verglichen. Immobilisierte Pfropfcopolymere zeigen ein dem freien Enzym ähnliches Verhalten. Die Diffusion bleibt weitgehend unbeschräkt, da die Immobilisierungsreaktion die enzymatische Aktivität nicht beeinträchtigt. Mit Hilfe des Bisacryloylpiperazins war es mbglich, mit Chymotrypsin, besonders was die Wirksamkeit der Bindung und die Beständigkeit bei kontinuierlichem Einsatz betrifft, bessere Ergebnisse als die vordem erreichten zu erhalten.
    Notes: A method of enzyme immobilization by graft-copolymerization onto polysaccharides is reported. Bisacryloylpiperazine has been used as a vinylating reagent and the reaction product with several enzymes (HRP, GOD, Am, ChT, Cel) was copolymerized onto different matrices (cellulose, Sepharose, Sephadex, starch). Immobilization parameters which influence the copolymer activity have been studied for the insolubilization of horseradish peroxidase onto cellulose. These parameters are pH, time, and temperature of bisacryloylpiperazine enzyme activation reaction.Under the best immobilization conditions copolymer activity linearly depends on enzyme concentration. Enzyme coupling efficiency depends on the type of enzyme and it ranges from 7 to 20%.The most important characteristics of these immobilized enzyme systems were tested and compared with those of similar systems obtained by glycidylmethacrylate enzyme activation (stability in continuous washing, kinetic characteristics, and storage, thermal, and operational stability). Immobilized enzyme graft copolymers have kinetic behaviour very close to that of the free enzymes. Diffusion is not seriously limited because immobilization reaction does not alter the enzymatic activity. By means of bisacryloylpiperazine it was possible to immobilize chymotrypsin with better results than those previously obtained, particularly coupling efficiency and long term continuous working.
    Additional Material: 6 Ill.
    Type of Medium: Electronic Resource
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