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ABSTRACT: Microbial β-glucosidases have been used for the enhancement of wine aroma. Nevertheless, few enzymes are active in the conditions of winemaking. In this work, the production of a β-glucosidase by an Aureobasidium pullulans strain (Ap-β-gl) isolated from grape ecosystems was evaluated. The maximum enzymatic synthesis using submerged fermentation was after 96 h of growth in complex media containing 20 g/L of cellobiose as the sole carbon source. The crude enzyme (Ap-β-gl) showed optimal pH at 5.5 and two peaks of optimum temperature (at 45 and 70 °C). It showed a wide range of pH stability, stability at low temperatures, and tolerance to ethanol, showing suitable characteristics for winemaking conditions. The hydrolysis of glycosidic terpenes by Ap-β-gl was studied, and its ability to efficiently release free terpenols was demonstrated by gas chromatography/mass spectrometry. The enzymatic treatment notably increased the amount of monoterpenes, showing good prospects for its potential application for the development of aroma in wines.
Applied biochemistry and biotechnology 12/2012; · 1.94 Impact Factor
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ABSTRACT: Wood-rotting fungi have the ability to degrade lignin by secreting ligninases, a promising enzyme for degradation of environmental
pollutants. Nine basidiomycete strains collected just outside the city of São José do Rio Preto, upstate São Paulo, Brazil,
were evaluated for their manganese peroxidase (MnP), lignin peroxidase (LiP) and laccase production by solid-state fermentation
on wheat bran.Datronia caperata SP381992,Polyporus tenuiculus SP381977 andPycnoporus sanguineus SP381968 were the highest producers of laccase, whilePolyporus tenuiculus SP381971,Datronia caperata SP381992,Coriolopsis polyzona SP381989 andHexagonia hirta SP382026 produced the most MnP and LiP activity. The majority of strains secreted laccase with optimum activity at 70 °C
and, when maintained at 60 °C, in the absence of substrate, the crude enzymes preserved 100% of their initial for periods
of 30 min up to 8 h. Enzymes fromD. caperata SP381992,P. tenuiculus SP381977,P. sanguineus SP381968 andH. hirta SP382026 were tested for activity on the azo dye orange II and afforded 96–100% decolourisation of the dye in 1 to 48 h.
Since this reaction depended on the presence of ABTS and there was no decolourisation when H2O2 or MnSO4 was present, it was attributed to the laccase activity.
Annals of Microbiology 04/2012; 58(3):427-432. · 0.69 Impact Factor
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ABSTRACT: The aim of this study was the detailed characterization of the phenolic composition and the determination of the antioxidant activity of the Bordô grape (Vitis labrusca) cultivated in South Brazil. The edible parts of Bordô grapes (flesh and skin) contained 1130 mg/kg of total phenolic compounds (as gallic acid), mainly located in the skins. Anthocyanin content in the skins was high, largely as 3,5-diglucosides (1359 mg/kg, as malvidin 3,5-diglucoside). Total flavonols accounted for 154 μmol/kg, mainly located in the skins and with myricetin 3-glucoside as the principal flavonol in both grape parts. Very low amounts of flavan-3-ol monomers and dimers and low amounts of polymeric proanthocyanidins, with a composition similar to that reported for V. vinifera grape varieties, were found in Bordô grape skins. Hydroxycinnamic acid derivatives mainly derived from caffeic acid and were found in the skins in high amounts, ten times higher than in the flesh (total amount: 483 μmol/kg). Finally, the Bordô grape cultivar can be considered a high resveratrol producer (10.91 mg/kg) and also exhibited a high value of total antioxidant capacity (37.6 ± 1.0 mmol/kg, as Trolox).
Journal of Agricultural and Food Chemistry 11/2011; 59(24):13136-46. · 2.82 Impact Factor
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ABSTRACT: The seedless grapes BRS Clara and BRS Morena, developed in Brazil, are currently growing in popularity due to their premium texture and taste. However, there are no reports on the polyphenoloxidase (PPO) from these cultivars. In this paper, active and latent PPO from BRS Clara and BRS Morena seedless grapes were extracted using the non-ionic detergents Triton-X-100 (active) and Triton-X-114 (latent), and their catecholase activities were characterized. The PPO extracted using Triton-X-110 exhibited maximum activities at pH 6.0 and at 25 °C. Above 30 °C, a gradual decline in activities was noted, with complete inactivation at 60 °C. The PPO from grapes extracted with Triton-X-114 was activated with 0.2% of the ionic detergent sodium dodecyl sulfate (SDS), and exhibited maximum activities at pH 5.5 and at 30 °C. It was stable until the temperature reached 60 °C.
Plant Physiology and Biochemistry 11/2011; 49(11):1251-8. · 2.84 Impact Factor
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ABSTRACT: This work is an exploratory study of the possibility of promoting the consumption of Syzygium cumini fruit by adding its extract to orange juice making good use of its functional (antioxidant) properties. S. cumini fruit extract was characterized in terms of its anthocyanin content (2.11 g/100 g expressed in cyanidine-3-glucoside equivalents), total phenolic compounds (360 mg/100 g expressed in gallic acid equivalents) and antioxidant capacity evaluated by the 2,2-diphenyl-1-picrylhydrazyl free radical scavenging method. The effects of the addition of S. cumini fruit crude extract as well as its chromatographic fractions on the juice were assessed chemically by headspace solid-phase micro-extraction and gas chromatography coupled with a mass spectrometry detector. Only six compounds had their chromatographic peak intensities clearly changed and the results are discussed in terms of the inhibition of the formation of 2-octanone, hexanol, α-copaene, and α-panasinsene and the conservation of octyl acetate and p-menth-1-en-9-ol. Sensory evaluation of orange juice with and without S. cumini crude extract addition did not show any significant differences in the sensorial profile, discriminative and acceptance tests.
International Journal of Food Sciences and Nutrition 10/2011; 63(3):273-7. · 1.15 Impact Factor
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08/2011; , ISBN: 978-953-307-447-4
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ABSTRACT: For the first time, the production of an extracellular β-glucosidase (Sp-β-gl) by a Sporidiobolus pararoseus yeast strain is reported. The Sp-β-gl activity was quantified, characterized, and assessed for its efficiency in releasing aroma-enhancing compounds in wines. The maximum enzymatic synthesis was after 72 h of growth in a complex media with 20 g/L of cellobiose. The optimal pH and temperature were 5.5 and at 50 °C, respectively. It showed a wide range of pH stability and exhibited quite high thermostability at low temperatures. In addition, this β-glucosidase revealed tolerance to wine-associated inhibitory compounds (sugars and ethanol), showing suitable characteristics for all the stages of alcoholic fermentation. The hydrolysis of the glycosidic terpenes by Sp-β-gl was studied by gas chromatography, and its ability to efficiently release free terpenols has been demonstrated. The concentrations of geraniol, linalool, α-terpineol, and nerol were significantly increased in treated wines. These results suggest the potential application of this new yeast β-glucosidase as an aroma-enhancing enzyme in winemaking. PRACTICAL APPLICATION: The search for new β-glucosidase from yeast sources is important to improve the quality of wines. In this work, an S. pararoseus yeast strain has shown to be capable to produce a β-glucosidase with suitable combination of properties for functionality in wines and with potential to increase the concentration of free aroma compounds, showing good prospects for an industrial application.
Journal of Food Science 08/2011; 76(7):C997-1002. · 1.66 Impact Factor
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ABSTRACT: For the first time, the production of an extracellular β-glucosidase (Sp-β-gl) by a Sporidiobolus pararoseus yeast strain is reported. The Sp-β-gl activity was quantified, characterized, and assessed for its efficiency in releasing aroma-enhancing compounds in wines. The maximum enzymatic synthesis was after 72 h of growth in a complex media with 20 g/L of cellobiose. The optimal pH and temperature were 5.5 and at 50 °C, respectively. It showed a wide range of pH stability and exhibited quite high thermostability at low temperatures. In addition, this β-glucosidase revealed tolerance to wine-associated inhibitory compounds (sugars and ethanol), showing suitable characteristics for all the stages of alcoholic fermentation. The hydrolysis of the glycosidic terpenes by Sp-β-gl was studied by gas chromatography, and its ability to efficiently release free terpenols has been demonstrated. The concentrations of geraniol, linalool, -terpineol, and nerol were significantly increased in treated wines. These results suggest the potential application of this new yeast β-glucosidase as an aroma-enhancing enzyme in winemaking.Practical Application: The search for new β-glucosidase from yeast sources is important to improve the quality of wines. In this work, an S. pararoseus yeast strain has shown to be capable to produce a β-glucosidase with suitable combination of properties for functionality in wines and with potential to increase the concentration of free aroma compounds, showing good prospects for an industrial application.
Journal of Food Science 08/2011; 76(7):C997 - C1002. · 1.66 Impact Factor
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ABSTRACT: The detailed phenolic composition (anthocyanins, flavonols, hydroxycinnamic acid derivatives, stilbenes, and flavan-3-ols) in the skin and flesh of the new BRS Clara and BRS Morena seedless table grapes has been studied using HPLC-DAD-ESI-MS/MS. The two grapes, especially BRS Morena, contained high amounts of phenolic compounds, mainly located in their skins and qualitatively not different from those found in Vitis vinifera grapes. In addition, BRS Morena (a teinturier variety) showed qualitatively different phenolic compositions in its skin and flesh, mainly affecting the anthocyanin and flavonol profiles. Consistent with high phenolic contents, high antioxidant capacity values were registered for both grape varieties, especially for BRS Morena. Proanthocyanidins and hydroxycinnamoyl-tartaric acids were the major phenolic compounds found in BRS Clara and were also important in BRS Morena, although anthocyanins were the main phenolic compounds in the latter case. These results suggest that the entire grapes, including the skin, may potentially possess properties that are beneficial to human health. In this context, the BRS Morena grape can be considered as a high resveratrol producer.
Journal of Agricultural and Food Chemistry 06/2011; 59(15):8314-23. · 2.82 Impact Factor
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ABSTRACT: Resumo Os oligossacarídeos participam da constituição da fibra alimentar e são principalmente utilizados como prebióticos. Esta revisão apresenta as vias de obtenção destes açúcares, os quais podem ser produzidos por síntese (química ou enzimática) ou da despolimerização de polissacarídeos (física, química ou enzimática), como descrito na presente revisão. Os oligossacarídeos vêm sendo utilizados comercialmente como ingredientes de cosméticos, medicamentos, produtos agrícolas e principalmente na indústria alimentícia. O potencial de aplicações dos oligossacarídeos nessas áreas nas mais diversas áreas tais como alimentos, rações animais, fármacos e cosméticos tem contribuído para o aumento das investigações científicas. O uso de oligossacarídeos como agentes imunomoduladores e modificadores de respostas biológicas foi recentemente descrito, assim como seus efeitos como compostos antiinflamatórios e hipocolesterolêmicos. Uma visão geral sobre as diversas funções nutracêuticas e biológicas destes oligômeros de carboidratos visando benefícios para a saúde humana também foi reportada. Palavras-chave: Oligossacarídeos. Fibras da dieta. Saúde humana. Abstract Oligosaccharides participate in the formation of dietary fiber and are mainly used as prebiotic agents. This review presents ways of obtaining these sugars, which can be produced by synthesis (chemical or enzymatic), or through depolymerization of polysaccharides (physical, chemical or enzymatic). Oligosaccharides have also been used commercially as an ingredient in cosmetics, pharmaceuticals, agricultural products and especially in the food industry because of their physical properties. The potential applications of oligosaccharides in several areas such as food, animal feed, pharmaceuticals and cosmetics have contributed to the increase in scientific research on these carbohydrates. The use of oligosaccharides as immuno-modulatory agents and biological response modifiers has been recently described, and their effects as anti-inflammatory and in reducing cholesterol. An overview of the various nutraceutical and biological functions of these carbohydrates in order to benefit human health is also reported.
Semina 01/2011;
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ABSTRACT: In recent years, the baking industry has focused its attention on substituting several chemical compounds with enzymes. Enzymes that hydrolyze nonstarch polysaccharides, such as xylanase, lead to the improvement of rheological properties of dough, loaf specific volume, and crumb firmness. The purpose of this study was to find a better solid-state fermentation substrate to produce high levels of xylanase and low levels of protease and amylase, which are enzymes involved in bread quality, from Thermoascus aurantiacus CBMAI 756. Wheat bran, corncob, and corn straw were used as energy sources. The enzyme extract of corncob showed high xylanase activity (130 U/mL) and low amylase and protease activity (<1 and 15 U/mL, respectively). This enzyme profile may be more profitable for the baking industry, because it results in a slower degradation of gluten. Our results confirm this finding, because the enzyme obtained by fermentation in corncob resulted in a gluten with a higher specific volume than all the other substrates that were tested. The crude xylanase presented maximum activity at a pH of 5, and the optimum temperature was 75 °C. It was stable up to 70 °C for an hour and at a pH range from 4 to 10.
Journal of Food Science 09/2010; 75(7):C588-94. · 1.66 Impact Factor
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ABSTRACT: An alpha-glucosidase enzyme produced by the fungus Thermoascus aurantiacus CBMAI 756 was purified by ultra filtration, ammonium sulphate precipitation, and chromatography using Q Sepharose, Sephacryl S-200, and Superose 12 columns. The apparent molecular mass of the enzyme was 83 kDa as determined in gel electrophoresis. Maximum activity was observed at pH 4.5 at 70 degrees C. Enzyme showed stability stable in the pH range of 3.0-9.0 and lost 40% of its initial activity at the temperatures of 40, 50, and 60 degrees C. In the presence of ions Na(+), Ba(2+), Co(2+), Ni(2+), Mg(2+), Mn(2+), Al(3+), Zn(2+), Ca(2+) this enzyme maintained 90-105% of its maximum activity and was inhibited by Cr(3+), Ag(+), and Hg(2+). The enzyme showed a transglycosylation property, by the release of oligosaccharides after 3 h of incubation with maltose, and specificity for short maltooligosaccharides and alpha-PNPG. The K(m) measured for the alpha-glucosidase was 0.07 microM, with a V(max) of 318.0 micromol/min/mg.
The Journal of Microbiology 08/2010; 48(4):452-9. · 1.10 Impact Factor
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ABSTRACT: Three mutations, Ser54→Pro, Thr314→Ala, and His415→Tyr, were identified in Aspergillus awamori glucoamylase gene expressed by Saccharomyces cerevisiae. The mutant glucoamylase (GA) was substantially more thermostable than a wild-type GA at 70 °C, with a 3.0 KJ mol(-1) increase in the free energy of thermo-inactivation. The effect of starch from different botanical sources on the production of this GA was measured in liquid fermentation using commercial soluble starch, cassava, potato, and corn as the carbon source. The best substrate for GA production was the potato starch showing an enzymatic activity of 6.6 U/mL. The commercial soluble starch was also a good substrate for the enzyme production with 6.3 U/mL, followed by cassava starch and corn starch with 5.9 and 3.0 U/mL, respectively. These results showed a significant difference on GA production related to the carbon source employed. The mutant GA was purified by acarbose-Sepharose affinity chromatography; the estimated molecular mass was 100 kDa. The mutant GA exhibited optimum activity at pH 4.5 and an optimum temperature of 65 °C.
Applied biochemistry and biotechnology 04/2010; 163(1):14-24. · 1.94 Impact Factor
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ABSTRACT: Pectate lyase (PL) was produced by the filamentous fungus Penicillium viridicatum RFC3 in solid-state cultures of a mixture of orange bagasse and wheat bran (1 : 1 w/w), or orange bagasse, wheat bran and sugarcane bagasse (1 : 1 : 0.5 w/w), and in a submerged liquid culture with orange bagasse and wheat bran (3%) as the carbon source. PL production was highest (1,500 U mL(-1) or 300 Ug(-1) of substrate) in solid-state fermentation (SSF) on wheat bran and orange bagasse at 96 hours. PL production in submerged fermentation (SmF) was influenced by the initial pH of the medium. With the initial pH adjusted to 4.5, 5.0, and 5.5, the peak activity was observed after 72, 48, and 24 hours of fermentation, respectively, when the pH of the medium reached the value 5.0. PL from SSF and SmF were loaded on Sephadex-G75 columns and six activity peaks were obtained from crude enzyme from SSF and designated PL I, II, III, IV, V, and VI, while five peaks were obtained from crude enzyme from SmF and labeled PL I', II', III', IV', and VII'. Crude enzyme and fraction III from each fermentative process were tested further. The optimum pH for crude PL from either process was 5.5, while that for PL III was 8.0. The maximum activity of enzymes from SSF was observed at 35 degrees C, but crude enzyme was more thermotolerant than PL III, maintaining its maximum activity up to 45 degrees C. Crude enzyme from SmF and PL III' showed thermophilic profiles of activity, with maximum activity at 60 and 55 degrees C, respectively. In the absence of substrate, the crude enzyme from SSF was stable over the pH range 3.0-10.0 and PL III was most stable in the pH range 4.0-7.0. Crude enzyme from SmF retained 70%-80% of its maximum activity in the acid-neutral pH range (4.0-7.0), but PIII showed high stability at alkaline pH (7.5-9.5). PL from SSF was more thermolabile than that from SmF. The latter maintained 60% of its initial activity after 1 h at 55 degrees C. The differing behavior of the enzymes with respect to pH and temperature suggests that they are different isozymes.
International Journal of Microbiology 01/2010; 2010.
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ABSTRACT: Protease production was carried out in solid state fermentation. The enzyme was purified through precipitation with ethanol at 72% followed by chromatographies in columns of Sephadex G75 and Sephacryl S100. It was purified 80-fold and exhibited recovery of total activity of 0.4%. SDS-PAGE analysis indicated an estimated molecular mass of 24.5 kDa and the N-terminal sequence of the first 22 residues was APYSGYQCSMQLCLTCALMNCA. Purified protease was only inhibited by EDTA (96.7%) and stimulated by Fe(2+) revealing to be a metalloprotease activated by iron. Optimum pH was 5.5, optimum temperature was 75 degrees C, and it was thermostable at 65 degrees C for 1 h maintaining more than 70% of original activity. Through enzyme kinetic studies, protease better hydrolyzed casein than azocasein. The screening of fluorescence resonance energy transfer (FRET) peptide series derived from Abz-KLXSSKQ-EDDnp revealed that the enzyme exhibited preference for Arg in P(1) (k(cat)/K(m) = 30.1 mM(-1) s(-1)).
Journal of Agricultural and Food Chemistry 09/2009; 57(19):9210-7. · 2.82 Impact Factor
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ABSTRACT: An exo-PG obtained from Penicillium viridicatum in submerged fermentation was purified to homogeneity. The apparent molecular weight of the enzyme was 92 kDa, optimum pH and temperature for activity were pH 5 and 50-55 degrees C. The exo-PG showed a profile of an exo-polygalacturonase, releasing galacturonic acid by hydrolysis of pectin with a high degree of esterification (D.E.). Ions Ca(2+) enhanced the stability of enzyme and its activity by 30%. The K(m) was 1.30 in absence of Ca(2+) and 1.16 mg mL(-1) in presence of this ion. In relation to the V(max) the presence of this ion increased from 1.76 to 2.07 mumol min(-1)mg(-1).
International Journal of Microbiology 01/2009; 2009:631942.
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ABSTRACT: A comparative study was carried out to evaluate protease production in solid-state fermentation (SSF) and submerged fermentation (SmF) by nine different thermophilic fungi--Thermoascus aurantiacus Miehe, Thermomyces lanuginosus, T. lanuginosus TO.03, Aspergillus flavus 1.2, Aspergillus sp. 13.33, Aspergillus sp. 13.34, Aspergillus sp. 13.35, Rhizomucor pusillus 13.36 and Rhizomucor sp. 13.37--using substrates containing proteins to induce enzyme secretion. Soybean extract (soybean milk), soybean flour, milk powder, rice, and wheat bran were tested. The most satisfactory results were obtained when using wheat bran in SSF. The fungi that stood out in SSF were T. lanuginosus, T. lanuginosus TO.03, Aspergillus sp. 13.34, Aspergillus sp. 13.35, and Rhizomucor sp. 13.37, and those in SmF were T. aurantiacus, T. lanuginosus TO.03, and 13.37. In both fermentation systems, A. flavus 1.2 and R. pusillus 13.36 presented the lowest levels of proteolytic activity.
Applied biochemistry and biotechnology 04/2008; 146(1-3):223-30. · 1.94 Impact Factor
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ABSTRACT: A cyclomaltodextrin glucanotransferase (E.C. 2.4.1.19) from a newly isolated alkalophilic and moderately thermophilic Paenibacillus campinasensis strain H69-3 was purified as a homogeneous protein from culture supernatant. Cyclomaltodextrin glucanotransferase was produced during submerged fermentation at 45 degrees C and purified by gel filtration on Sephadex G50 ion exchange using a Q-Sepharose column and ion exchange using a Mono-Q column. The molecular weight of the purified enzyme was 70 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and the pI was 5.3. The optimum pH for enzyme activity was 6.5, and it was stable in the pH range 6.0-11.5. The optimum temperature was 65 degrees C at pH 6.5, and it was thermally stable up to 60 degrees C without substrate during 1 h in the presence of 10 mM CaCl(2). The enzyme activity increased in the presence of Co(2+), Ba(2+), and Mn(2+). Using maltodextrin as substrate, the K(m) and K(cat) were 1.65 mg/mL and 347.9 micromol/mg x min, respectively.
Applied biochemistry and biotechnology 05/2007; 137-140(1-12):41-55. · 1.94 Impact Factor
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ABSTRACT: An extracellular polygalacturonase was isolated from 5-day culture filtrates of Thermoascus aurantiacus CBMAI-756 and purified by gel filtration and ion-exchange chromatography. The enzyme was maximally active at pH 5.5 and 60-65 degrees C. The apparent K (m) with citrus pectin was 1.46 mg/ml and the V (max) was 2433.3 micromol/min/mg. The apparent molecular weight of the enzyme was 30 kDa. The enzyme was 100% stable at 50 degrees C for 1 h and showed a half-life of 10 min at 60 degrees C. Polygalacturonase was stable at pH 5.0-5.5 and maintained 33% of initial activity at pH 9.0. Metal ions, such as Zn(+2), Mn(+2), and Hg(+2), inhibited 50, 75 and 100% of enzyme activity. The purified polygalacturonase was shown to be an endo/exo-enzyme, releasing mono, di and tri-galacturonic acids within 10 min of hydrolysis.
Antonie van Leeuwenhoek 05/2007; 91(3):291-9. · 2.09 Impact Factor
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ABSTRACT: This article investigates a strain of the yeast Aureobasidium pullulans for cellulase and hemicellulase production in solid state fermentation. Among the substrates analyzed, the wheat bran culture presented the highest enzymatic production (1.05 U/mL endoglucanase, 1.3 U/mL beta-glucosidase, and 5.0 U/mL xylanase). Avicelase activity was not detected. The optimum pH and temperature for xylanase, endoglucanase and beta-glucosidase were 5.0 and 50, 4.5 and 60, 4.0 and 75 degrees C, respectively. These enzymes remained stable between a wide range of pH. The beta-glucosidase was the most thermostable enzyme, remaining 100% active when incubated at 75 degrees C for 1 h.
Applied biochemistry and biotechnology 05/2007; 137-140(1-12):281-8. · 1.94 Impact Factor
