Characterization of a cellulase-free xylanase producing Bacillus sp for biobleaching of kraft pulp

Application of the xylanase in the pulp bleaching process has been shown to be effective in decreasing the amount of chlorinating agents in the process and improving the brightness of the pulp. The use of thermostable cellulase-free xylanase might enhance both the technical and economic feasibility of the process. In this work an alkalophylic strain of Bacillus sp 77-2, was isolated which showed a high production of xylanase and free cellulases. The xylanase of Bacillus sp displayed an optimum pH of 6.0 (with 70% activity at pH 9.0), all optimum temperature of 60 degrees C, pH stability in the range 5-10 and thermal stability of 50 degrees C. These characteristics are important to the kraft pulp bleaching because they are similar to those found in the industrial paper environment.

Xylanase production by Aspergillus awamori under solid state fermentation conditions on tomato pomace

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Ribonuclease production by Aspergillus species

Ribonuclease production by Aspergillus flavipes. A sulphureus and A. fischeri in semisynthetic medium, after 24-144 hours at 30 degrees C under shaking, was studied. After cultivation, the medium was separated from micelia by filtration and the resultant solution was used as enzymatic extract. The highest amount of biomass and RNase was obtained after 96 hours of cultivation. The enzymes produced by three species presented similar characteristics, with optimum temperature at 55 degrees C and two peaks of activity at pH 4.5 and 7.0. A. flavipes RNases were more sensitive to temperature: 50% of the initial activity was lost after 1 hour at 70 degrees C. After this heat treatment, RNase of A. sulphureus lost 30% of this activity and that of A. fischeri only 16%. The nucleotides released by enzimatic hydrolysis of RNA were separated by ion exchange chromatography in a AG-1X8-formiate column and identified by paper chromatography. This procedure indicated that the raw enzymatic extract of Aspergillus flavipes is able to hydrolyze RNA, releasing 3'-nucleotides monophosphate at pH 4.5 and 3' and 5'-nucleotides monophosphate at pH 7.0 and 8.5. This result suggests that this strain produces two different types of RNase, one acidic and other alcaline, with different specificities.

Enrichment of a continuous culture of Saccharomyces cerevisiae with the yeast Issatchenkia orientalis in the production of ethanol at increasing temperatures

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Seed germination in Miconia theaezans (Bonpl.) Cogniaux (Melastomataceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Quimiometria como ferramenta analitica para definição das condiçoes de ensaio da enzima peroxidase de soja

Enzimas Peroxidases são heme-proteínas encontradas nos diferentes organismos vivos, especialmente vegetais, apresentam importante papel fisiológico/bioquímico como proteção contra microorganismos invasores. A soja, um dos mais importantes produtos para o agronegócio brasileiro apresenta na casca de suas sementes (subproduto) alta atividade de peroxidase, denominada soybean peroxidase,com potencial de utilização em métodos analíticos clínicos. A proposta do trabalho foi aplicar o planejamento fatorial para otimização das condições extração da enzima, definição das condições ótimas de atividade (pH e temperatura), utilizando metodologia de superfície de resposta. Os dados obtidos com clara definição foram: i) extração em pó cetonico, ii) meio reacional: pH 3,3, volume da amostra contendo a enzima 330 µL - 340 µL, peróxido de hidrogênio 4,2 mmol.L-1 150 µL, tempo de reação 20 segundos, temperatura 50º C, substrato guaiacol 30mmol.L-1 300 µL, e 0,1 mol.L-1 de NaCl. O uso da dessa metodologia para definição das condições de extração e estudos cinético-enzimáticos da peroxidase de soja foram eficientes e mais precisos, comparado a metodologia de variações/repetições (tentativa e erro).

Germinação de sementes e emergência de plântulas de Tabebuia rosea (Bertoloni) a.p. de Candolle (Bignoniaceae), uma espécie exótica com potencial invasor

Este estudo teve por objetivo determinar os efeitos da temperatura e da luz na germinação de sementes de Tabebuia rosea, bem como avaliar a influência dos ambientes de pleno sol e sob dossel na emergência de suas plântulas. Foram testadas temperaturas constantes de 10 a 45 ºC com intervalos de 5 ºC, sob luz branca e escuro. Verificou-se que a faixa ótima de temperatura para a germinação foi de 20 a 40 ºC na luz e de 20 a 35 ºC no escuro. As sementes apresentaram maior sincronização da germinação a 25 ºC na luz e 30 ºC no escuro. Os resultados indicam que as sementes de Tabebuia rosea podem germinar em ampla faixa de temperatura, tanto em ambientes abertos, com disponibilidade de luz, quanto em locais com ausência de luz, enterradas no solo ou, mesmo, no interior das florestas. Essa germinação, bem como o recrutamento das plântulas, tanto a pleno sol quanto sob a sombra da vegetação, indica o potencial invasor da espécie.

Germinação de sementes de Dyckia tuberosa (Vell.) Beer (Bromeliaceae) sob diferentes temperaturas em luz e escuro

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Efeito da temperatura e a participação do fitocromo no controle da germinação de sementes de embaúba

A embaúba é considerada uma espécie pioneira que ocorre na mata Atlântica, principalmente em borda da mata ou em matas secundárias. O presente trabalho teve como objetivo a análise da influência da temperatura e do modo de ação da luz através de curva de fluência resposta, para a melhor compreensão do comportamento das sementes desta espécie. Através de incubações isotérmicas foi determinada que a temperatura ótima de germinação de sementes de Cecropia glaziovi, situa-se entre 25 e 30ºC e a saturação da indução da germinação com luz branca mediada pelo fitocromo ocorreu com 1W.m-2. Embora as sementes de embaúba necessitem de luz de alta razão de V:VE para a indução do processo, a germinação ocorreu em fluência baixa de luz branca, indicando alta sensibilidade dessas sementes ao ambiente aberto, como borda de matas e pequenas clareiras. Estas características indicam a participação do fitocromo B no controle da germinação de sementes nesta espécie.

Aspectos morfológicos de frutos, sementes, germinação e plântulas de Hymenolobium petraeum

Hymenolobium petraeum Ducke é uma espécie arbórea pertencente à família Leguminosae conhecida popularmente por angelim-pedra. Apresenta alto valor comercial, com madeira muito utilizada na construção civil e marcenaria. Este trabalho teve por objetivo descrever morfologicamente o fruto, a semente e as plântulas, assim como, determinar as temperaturas cardeais para a germinação de sementes de angelim-pedra. Foram determinados o comprimento, a largura e a massa fresca de frutos e sementes. Para os testes de germinação foram utilizadas três repetições de 50 sementes, colocadas em placas de Petri e mantidas em germinadores nas temperaturas de 15, 20, 25, 30, 35 e 40°C e fotoperíodo de 12 horas. Os frutos são legumes-samaróides, indeiscentes, oblongos e unicarpelares. As sementes são de coloração castanho-escura, oblongas, estenospérmicas, exalbuminosas e com plúmula inconspícua. A raiz primária é branca e pubescente na região próxima ao colo; a parte aérea das plântulas possui pilosidade branca, protófilos compostos imparipinados e com inserção oposta, epicótilo verde, ereto, cilíndrico e piloso e os metáfilos imparipinados e com inserção alterna-espiralada. A germinação é semi-hipógea criptocotiledonar. Para de sementes de angelim-pedra as temperaturas máximas de germinação estão acima de 35°C e a mínima abaixo de 15°C, enquanto a faixa de temperatura ótima para germinação está entre 25 e 35°C.

Thermostable conidial and mycelial alkaline phosphatases from the thermophilic fungus Scytalidium thermophilum

An extracellular (conidial) and an intracellular (mycelial) alkaline phosphatase from the thermophilic fungus Scytalidium thermophilum were purified by DEAE-cellulose and Concanavalin A-Sepharose chromatography. These enzymes showed allosteric behavior either in the presence or absence of MgCl2, BaCl2, CuCl2, and ZnCl2. All of these ions increased the maximal velocity of both enzymes. The molecular masses of the conidial and mycelial enzymes, estimated by gel filtration, were 162 and 132 kDa, respectively. Both proteins migrated on SDS-PAGE as a single polypeptide of 63 and 58.5 kDa, respectively, suggesting that these enzymes were dimers of identical subunits. The best substrate for the conidial and mycelial phosphatases was p-nitrophenylphosphate, but,beta -glycerophosphate and other phosphorylated compounds also served as substrates. The optimum pH for the conidial and mycelial alkaline phosphatases was 10.0 and 9.5 in the presence of AMPOL buffer, and their carbohydrate contents were about 54% and 63%, respectively. The optimum temperature was 70-75 degreesC for both activities. The enzymes were fully stable up to 1 h at 60 degreesC. These and other properties suggested that the alkaline phosphatases of S. thermophilum might be suitable for biotechnological applications.

Acerola's clones of industrial interest

In order to know which clone of acerola is better for acerola industrialization, we studied the pectin methylesterase (PME) specific activity, pectin content and vitamin C content in five different clones of acerola. The pectin yield varied from 1.37 to 2.99% and the highest content of pectin occurred in clones 3 and 5. Ascorbic acid varied significantly from 1157.5 to 1735.5 mg/100 g of pulp in the five clones. The highest content of vitamin C occurred in clone 4. The PME specific activity varied from 0.79 to 2.92 units g(-1)/g of pulp and the highest values occurred in clone 2. We also studied the optimum temperature and the optimum pH of this enzyme. Clones 1, 2, 4 and 5 showed optimum temperature at 90C. Clone 3 showed practically the same specific activity at all temperatures studied. Clones 1 and 4 showed an optimum pH of 9.0 and clone numbers 2, 3 and 5 showed a pH optimum at 8.5.

Partial purification, heat stability and kinetic characterization of the pectinmethylesterase from Brazilian guava, Paluma cultivars

Pectinmethylesterase (PME) was extracted from guava fruit (Psidium guajava L.), cultivar Paluma, by 70% ammonium sulphate saturation and partially purified by gel filtration on Sephadex G100. Gel filtration showed PME isoenzymes with different values of molecular mass. Two samples were examined: concPME (70% saturation by ammonium sulphate) and Iso4 PME (one of the isoforms from gel filtration with the greatest specific activity). Optimum pH of the enzyme (for both samples) was 8.5 and optimum temperature ranged from 75 and 85 degrees C. The optimum sodium chloride concentration was 0.15 M. The K-M and V-max ranged from 0.32 to 0.23 mg m1(-1) and 244 to 53.2 mu mol/min, respectively, for concPME and Iso4PME. The activation energies (E-a) were 64.5 and 103 kJ/mol, respectively, for concPME and Iso4PME. Guava PME, cv Paluma, is a very thermostable enzyme, showing great heat stability at all temperatures studied. (c) 2005 Elsevier Ltd. All rights reserved.

GROWTH AND FERMENTATION CHARACTERISTICS OF NEW SELECTED STRAINS OF SACCHAROMYCES AT HIGH-TEMPERATURES AND HIGH CELL DENSITIES

Maintenance of high cell viability was the main characteristic of our new strains of thermotolerant Saccharomyces. Total sugar conversion to ethanol was observed for sugarcane juice fermentation at 38-40-degrees-C in less than 10 h and without continuous aeration of the culture. Invertase activity differed among the selected strains and increased during fermentation but was not dependent on cell viability. Invertase activity of the cells and optimum temperature for growth, as well as velocity of ethanol formation, were dependent on medium composition and the type of strain used. At high sugarcane syrup concentrations, the best temperature for ethanol formation by strain 781 was 35-degrees-C. Distinct differences among the velocities of ethanol production using selected strains were also observed in sugarcane syrup at 35-38-degrees-C.

Production and characterization of alpha-amylase from Rhizopus sp

A strain of Rhizopus sp. screened among more than 800 filamentous fungi showed great ability to produce a thermostable alpha-amylase by solid state fermentation. The best production was obtained with a bran moisture content of 40% when the enzyme activity reached 60 EU/g. of medium. During the purification procedures, a column of DEAE-Sephadex A-50 separated the enzyme in two fractions and the larger (85% of the total activity) showed optimum pH in a range from 4.0 to 5.6. Optimum temperature was found at 60-65 degrees C and in this range no loss of activity was observed after 60 min. of treatment in pH 5,0. Its K-m and V-m are, respectively, of 5.0 mg/ml of starch and 10,01 uMol of reducing sugar/min./mg. of protein. Its molecular weight was calculated in 64.000 by gel filtration in Sephadex G-200. The dextrinization power of the enzyme was observed preferentialy on substrates compound by chains with higher ramifications, that is: amylopectin > starch > amylose. Other aspects of the enzyme pattern action are also discussed.