Base de dados : LILACS
Pesquisa : G02.494 [Categoria DeCS]
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Id: biblio-1022834
Autor: Espinosa Pérez, Raúl; García Suárez, José; Narciandi Diaz, Emilio; Silva Rodríguez, Ricardo; Caballero Menéndez, Evelin; Díaz Balaguer, Héctor; Musacchio Lasa, Alexis.
Título: Scaling-up fermentation of Escherichia coli for production of recombinant P64k protein from Neisseria meningitidis
Fonte: Electron. j. biotechnol;33:29-35, May. 2018. tab, graf, ilus.
Idioma: en.
Resumo: Background: P64k is a Neisseria meningitidis high molecular weight protein present in meningococcal vaccine preparations. The lpdA gene, which encodes for this protein, was cloned in Escherichia coli and the P64k recombinant protein was expressed in E. coli K12 GC366 cells under the control of a tryptophan promoter. P64k was expressed as an intracellular soluble protein about 28% of the total cellular protein. Several scale-up criteria of fermentation processes were studied to obtain the recombinant P64k protein at the pilot production scale. Results: The best operational conditions at a larger scale production of P64k recombinant protein were studied and compared using the four following criteria: Constant Reynold's number (Re constant), Constant impeller tip speed (n di constant), Constant power consumption per unit liquid volume (P/V constant) and Constant volumetric oxygen transfer coefficients (KLa/k constant). The highest production of the recombinant protein was achieved based on the constant KLa/k scale-up fermentation criterion, calculating the aeration rate (Q) and the impeller agitation speed (n) by iterative process, keeping constant the KLa/k value from bench scale. The P64k protein total production at the 50 l culture scale was 546 mg l -1 in comparison with the 284 mg l -1 obtained at 1.5 l bench scale. Conclusions: The methodology described herein, for the KLa/k scale-up fermentation criterion, allowed us to obtain the P64k protein at 50 l scale. A fermentation process for the production of P64k protein from N. meningitidis was established, a protein to be used in future vaccine formulations in humans.
Descritores: Proteínas da Membrana Bacteriana Externa/biossíntese
Proteínas Recombinantes/biossíntese
Escherichia coli/metabolismo
Neisseria meningitidis/metabolismo
-Triptofano
Vacinas Meningocócicas
Fermentação
Peso Molecular
Responsável: CL1.1 - Biblioteca Central


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Id: biblio-1283494
Autor: Ponce, Belén; Urtuvia, Viviana; Maturana, Nataly; Peña, Carlos; Díaz-Barrera, Álvaro.
Título: Increases in alginate production and transcription levels of alginate lyase (alyA1) by control of the oxygen transfer rate in Azotobacter vinelandii cultures under diazotrophic conditions
Fonte: Electron. j. biotechnol;52:35-44, July. 2021. tab, ilus.
Idioma: en.
Projeto: ANID-Chile.
Resumo: BACKGROUND: Alginates are polysaccharides used in a wide range of industrial applications, with their functional properties depending on their molecular weight. In this study, alginate production and the expression of genes involved in polymerization and depolymerization in batch cultures of Azotobacter vinelandii were evaluated under controlled and noncontrolled oxygen transfer rate (OTR) conditions. RESULTS: Using an oxygen transfer rate (OTR) control system, a constant OTR (20.3 ± 1.3 mmol L 1 h 1 ) was maintained during cell growth and stationary phases. In cultures subjected to a controlled OTR, alginate concentrations were higher (5.5 ± 0.2 g L 1 ) than in cultures under noncontrolled OTR. The molecular weight of alginate decreased from 475 to 325 kDa at the beginning of the growth phase and remained constant until the end of the cultivation period. The expression level of alyA1, which encodes an alginate lyase, was more affected by OTR control than those of other genes involved in alginate biosynthesis. The decrease in alginate molecular weight can be explained by a higher relative expression level of alyA1 under the controlled OTR condition. CONCLUSIONS: This report describes the first time that alginate production and alginate lyase (alyA1) expression levels have been evaluated in A. vinelandii cultures subjected to a controlled OTR. The results show that automatic control of OTR may be a suitable strategy for improving alginate production while maintaining a constant molecular weight.
Descritores: Polissacarídeo-Liases/metabolismo
Transferência de Oxigênio
Azotobacter vinelandii/metabolismo
-Oxigênio/metabolismo
Expressão Gênica
Reação em Cadeia da Polimerase
Azotobacter vinelandii/genética
Alginatos/metabolismo
Fermentação
Peso Molecular
Responsável: CL1.1 - Biblioteca Central


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Toledo, Vicente de Paulo Coelho Peixoto de
Cunha, Mariem Rodrigues Ribeiro
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Id: biblio-838784
Autor: Sugimoto, Michelle A A; Toledo, Vicente de Paulo Coelho Peixoto de; Cunha, Mariem Rodrigues Ribeiro; Carregal, Virginia M; Jorge, Rodrigo; Leão, Pedro; Fialho, Sílvia Ligorio; Silva-Cunha, Armando.
Título: Quality of bevacizumab (Avastin®) repacked in single-use glass vials for intravitreal administration / Avaliação da qualidade do bevacizumabe (Avastin®) fracionado em frascos de vidro de dose única para administração intravítrea
Fonte: Arq. bras. oftalmol;80(2):108-113, Mar.-Apr. 2017. tab, graf.
Idioma: en.
Resumo: ABSTRACT Purpose: Avastin® (bevacizumab) is an anti-vascular endothelial growth factor (VEGF) monoclonal antibody given as an off-label drug by intravitreal administration for treatment of ocular diseases. The drug's clinical application and its cost-benefit profile has generated demand for its division into single-use vials to meet the low volume and low-cost doses necessary for intraocular administration. However, the safety of compounding the drug in single-use vials is still under discussion. In this study, the stability and efficacy of Avastin® repacked in individual single-use glass vials and glass ampoules by external compounding pharmacies were evaluated. Methods: Polyacrylamide gel electrophoresis (PAGE), size-exclusion chromatography (SEC), dynamic light scattering (DLS), and turbidimetry were selected to detect the formation of aggregates of various sizes. Changes in bevacizumab biological efficacy were investigated by using an enzyme-linked immunosorbent assay (ELISA). Results: Repacked and reference bevacizumab showed similar results when analyzed by PAGE. By SEC, a slight increase in high molecular weight aggregates and a reduction in bevacizumab monomers were observed in the products of the three compounding pharmacies relative to those in the reference bevacizumab. A comparison of repacked and reference SEC chromatograms showed that the mean monomer loss was ≤1% for all compounding pharmacies. Protein aggregates in the nanometer- and micrometer-size ranges were not detected by DLS and turbidimetry. In the efficacy assay, the biological function of repacked bevacizumab was preserved, with <3% loss of VEGF binding capacity relative to that of the reference. Conclusion: The results showed that bevacizumab remained stable after compounding in ampoules and single-use glass vials; no significant aggregation, fragmentation, or loss of biological activity was observed.

RESUMO Objetivos: Avastin® (bevacizumabe) é um anticorpo monoclonal inibidor do fator de crescimento endotelial de vasos (VEGF) utilizado "off-label" por meio de administração intravítrea para o tratamento de doenças oculares. A sua aplicação clínica associada ao custo-benefício do medicamento gerou uma demanda para seu fracionamento em frascos de dose única para utilização pela via intraocular. No entanto, a segurança do fracionamento do anticorpo em frascos de dose única ainda é alvo de discussão. Neste trabalho, a estabilidade e a eficácia do Avastin® fracionado em frascos ou ampolas de vidro de dose unitária por farmácias de manipulação do mercado foram avaliadas. Métodos: As técnicas de eletroforese em gel de poliacrilamida (PAGE), cromatografia por exclusão de tamanho (SEC), espalhamento dinâmico da luz (DLS) e turbidimetria foram empregadas para avaliar a formação de agregados de diferentes tamanhos. Alterações na atividade biológica do bevacizumabe foram estudadas utilizando ELISA. Resultados: Amostras referência e do bevacizumabe fracionado apresentaram resultados semelhantes quando analisado por gel de poliacrilamida. Por cromatografia por exclusão de tamanho, um pequeno aumento na quantidade de agregados de alta massa molar seguido de uma redução nos monômeros do bevacizumabe foram observados para as amostras das três farmácias de manipulação quando comparado ao referência. A comparação dos cromatogramas mostrou uma quantidade de redução do monômero inferior a 1% para todas as amostras fracionadas. Por espalhamento dinâmico da luz e turbidimetria, não foram detectados agregados de proteína na faixa de tamanho de micrômetro e nanômetro. No ensaio de eficácia, o bevacizumabe fracionado preservou sua função biológica pois apresentou menos de 3% de perda na capacidade de ligação ao VEGF quando comparado ao referência. Conclusão: Este estudo sugere que o bevacizumabe se mantem estável após fracionamento em ampolas e frascos de vidro de dose unitária pois não foram observadas agregação e/ou fragmentação de proteínas e perda de atividade biológica em quan tidades significativas.
Descritores: Controle de Qualidade
Inibidores da Angiogênese/química
Embalagem de Medicamentos
Bevacizumab/química
-Ensaio de Imunoadsorção Enzimática/métodos
Cromatografia em Gel/métodos
Inibidores da Angiogênese/análise
Fator A de Crescimento do Endotélio Vascular/análise
Estabilidade de Medicamentos
Eletroforese em Gel de Poliacrilamida/métodos
Injeções Intravítreas
Bevacizumab/análise
Difusão Dinâmica da Luz/métodos
Peso Molecular
Nefelometria e Turbidimetria/métodos
Responsável: BR1.1 - BIREME


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Id: biblio-1040224
Autor: Carvalho, Luana Dutra de; Peres, Bernardo Urbanetto; Maezono, Hazuki; Shen, Ya; Haapasalo, Markus; Jackson, John; Carvalho, Ricardo M; Manso, Adriana P.
Título: Doxycycline release and antibacterial activity from PMMA/PEO electrospun fiber mats
Fonte: J. appl. oral sci;27:e20180663, 2019. tab, graf.
Idioma: en.
Resumo: Abstract Objective: To investigate the use of polymethyl methacrylate (PMMA) electrospun fiber mats containing different amounts of polyethylene oxide (PEO) as a doxycycline delivery system and to test antibacterial activity against an oral pathogen. Methodology: PMMA powders or PEO (mol wt 200 Kd) (10,20,30% w/w/) were dissolved in N, N-dimethylformamide (DMF) to obtain a final polymer concentration of 15% in DMF (w/v). 2% Doxycycline monohydrate was added to the solutions and submitted to vortex mixing. The solution was transferred to a plastic syringe and fit into a nanofiber electrospinning unit. The parameters applied were: voltage at 17.2 kV; distance of 20 cm between the needle tip and the collector plate; target speed at 2 m/min; and transverse speed at 1cm/min. Syringe pump speed was 0.15 mm/min. The drug release analysis was performed by removing aliquots of the drug-containing solution (in PBS) at specific periods. Doxycycline release was quantified using RP-HPLC. Fiber mats from all groups had their antibacterial action tested against S. mutans based on inhibition halos formed around the specimens. The experiments were performed in triplicate. Gravimetric analysis at specific periods was performed to determine any polymer loss. Morphological characterization of the electrospun fibers was completed under an optical microscope followed by SEM analysis. Results: The addition of PEO to the PMMA fibers did not affect the appearance and diameter of fibers. However, increasing the %PEO caused higher doxycycline release in the first 24 h. Fibers containing 30% PEO showed statistically significant higher release when compared with the other groups. Doxycycline released from the fibers containing 20% or 30% of PEO showed effective against S. mutans. Conclusion: The incorporation of PEO at 20% and 30% into PMMA fiber mat resulted in effective drug release systems, with detected antibacterial activity against S. mutans.
Descritores: Polietilenoglicóis/farmacocinética
Doxiciclina/farmacocinética
Polimetil Metacrilato/farmacocinética
Nanofibras/química
Antibacterianos/farmacocinética
-Polietilenoglicóis/química
Streptococcus mutans/efeitos dos fármacos
Fatores de Tempo
Água/química
Microscopia Eletrônica de Varredura
Reprodutibilidade dos Testes
Análise de Variância
Cromatografia Líquida de Alta Pressão/métodos
Doxiciclina/química
Polimetil Metacrilato/química
Imersão
Antibacterianos/química
Peso Molecular
Responsável: BR28.1 - Serviço de Biblioteca e Documentação Professor Doutor Antônio Gabriel Atta


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Id: biblio-886635
Autor: SHI, CHEN-SHAN; SANG, YA-XIN; SUN, GUI-QING; LI, TIAN-YE; GONG, ZHENG-SI; WANG, XIANG-HONG.
Título: Characterization and bioactivities of a novel polysaccharide obtained from Gracilariopsis lemaneiformis
Fonte: An. acad. bras. ciênc;89(1):175-189, Jan,-Mar. 2017. graf.
Idioma: en.
Resumo: ABSTRACT Gracilariopsis lemaneiformis is a type of red alga that contains seaweed polysaccharide agar. In this study, a novel non-agar seaweed polysaccharide fraction named GCP (short of crude polysaccharide obtained from Gracilariopsis lemaneiformis) was isolated from Gracilariopsis lemaneiformis. Structural analysis showed that GCP shows triple helical chain conformation when dissolved in water and has many branches and long side chains. Also, 1→3 linkage is the major linkage and the sugar structures are galactopyranose configurations linked by β-type glycosidic linkages. Two macromolecular substance fractions (GCP-1 and GCP-2) were purified by DEAE Sepharose Fast Flow column chromatography. Moreover, a splenocyte damage assay and splenocyte proliferation assay were used to analyse the bioactivities of GCP, GCP-1 and GCP-2. It was demonstrated that polysaccharides could protect splenocyte damaged by H2O2; GCP-2 shows a greatest protection rate, that is, 92.8%, which significantly enhanced the splenocyte proliferation, and GCP showed the highest proliferation rate, 9.30%. The results suggested that this type of novel non-agar polysaccharide displayed remarkable antioxidant and immunomodulatory activities and early alkali treatment could decrease the activities. It may represent a potential material for health food and clinical medicines.
Descritores: Polissacarídeos/química
Alga Marinha/química
Rodófitas/química
-Polissacarídeos/isolamento & purificação
Valores de Referência
Ensaio de Imunoadsorção Enzimática
Linfócitos/efeitos dos fármacos
Microscopia Eletrônica de Varredura
Espectroscopia de Ressonância Magnética
Estrutura Molecular
Cromatografia Líquida de Alta Pressão
Espectroscopia de Infravermelho com Transformada de Fourier
Ácido Periódico/química
Proliferação de Células/efeitos dos fármacos
Peso Molecular
Limites: Animais
Ratos
Responsável: BR1.1 - BIREME


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Id: lil-103059
Autor: Ori, Maria; Seguro, Antonio Carlos; Rocha, Antonino dos Santos.
Título: Efeito inibidor do soro de ratos com insuficiência renal aguda e crônica sobre a atividade fagocitária "in vitro" / Inhibitory effect of serum from rats with acute and chronic renal insufficiency on phagocytic activity in vitro
Fonte: Rev. Inst. Med. Trop. Säo Paulo;32(6):409-13, nov.-dez. 1990. tab.
Idioma: pt.
Resumo: O efeito do soro de ratos com insuficiência renal aguda e crônica sobre a atividade fagocitária foi estudado "in vitro" em cultura de macrófagos. A atividade eritrofagocitária destas células foi menor na presença de soro de animais com insuficiência renal aguda e crônica quando comparados ao plasma normal. Esta inibiçäo persistiu quando estes soros foram filtrados através de membranas PM-30, sendo porém abolida quando filtrados através de membrana PM-10. Estes dados sugerem a presença de um fator no soro urêmico que inibe a fagocitose, cujo peso molecular varia entre 10.000 e 30.000 daltons
Descritores: Fagocitose
Eritrócitos/fisiologia
Injúria Renal Aguda/sangue
Insuficiência Renal Crônica/sangue
Técnicas In Vitro
Macrófagos/fisiologia
-Células Cultivadas
Peso Molecular
Limites: Animais
Masculino
Ratos
Responsável: BR1.1 - BIREME


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Tuan, Roseli
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Id: lil-282858
Autor: Tuan, Roseli; Bortolato, Paula Cristina.
Título: Genetic markers from Biomphalaria tenagophila (Gastropoda: Pulmonata: Planorbidae) obtained by the double stringency polymerase chain reaction technique
Fonte: Mem. Inst. Oswaldo Cruz;96(3):435-436, Apr. 2001. ilus.
Idioma: en.
Resumo: Biomphalaria tenagophila, one of the intermediate hosts of the trematoda Schistosoma mansoni, is a simultaneous hermafrodite snail species. In order to analyse the genetic structure of these populations, we performed a double-stringency PCR technique to obtain genetic markers with microsatellites and arbitrary primers in a single reaction
Descritores: Biomphalaria/genética
DNA/análise
Marcadores Genéticos
Reação em Cadeia da Polimerase/métodos
-Polimorfismo Genético
Peso Molecular
Limites: Animais
Responsável: BR1.1 - BIREME


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Id: biblio-994930
Autor: Vasquez Mejia, Sandra Milena; De Francisco, Alicia; Manique Barreto, Pedro Luiz; Mattioni, Bruna; Zibetti, André Wüst; Molognoni, Luciano; Daguer, Heitor.
Título: Physicochemical comparison of commercial VS. extracted ß-glucans and structural characterization after enzymatic purification / Comparación fisicoquímica entre ß-glucanos comerciales y extraídos, y caracterización estructural después de purificación enzimática
Fonte: Vitae (Medellín);25(1):26-36, 2018. Ilustraciones.
Idioma: en.
Resumo: Background: ß-glucans (1-3: 1-4) are soluble fibers applied to foods due to their technological properties (water binding capacity, viscosity, emulsification and stabilization) and their beneficial effects on health. The functional properties of ß-glucans can be lost during the extraction and purification processes. The high viscosity of ß-glucans is related to a high molecular weight and its physiological properties in the intestine. Therefore, to characterize the fiber after its extraction and purification is fundamental to understand its possible applications in foods. Objectives: characterize ß-glucans extracted (EßG) and compare them with three commercial ß-glucans (CßG-A, CßG-B and CßG-C) to identify its possible applications in foods and to evaluate if enzymatic purification affects molecular and structurally the ß-glucans. Methods: barley ß-glucans were extracted (EßG), characterized by chemical analyzes, rheological behavior, and color, and compared to three commercial ß-glucans samples. Then, the extract was purified and its structural and molecular characteristics were calculated. Results: EßG contained 64.38 ± 3.54% of ß-glucans, high starch contamination (12.70 ± 1.73%), high content of calcium (8894 mg/kg), pseudoplastic behavior, and dark color (L* = 52.77 ± 0.7). All commercial samples showed low starch contamination, lighter color, and Newtonian behavior. After purification starch and protein contamination decreased (0.85 ± 0.46% and 5.50 ± 0.12% respectively), increased the content of ßG (69.45 ± 0.81%) and increased brightness (L* = 92.60 ± 1.70). Purified ß-glucans (PßG) showed a molar weight of 690 ± 1.6 kDa and species with degree polymerization 3 (DP3) to 11 (DP11) were identified on the structure. Conclusions: EßG extracts before the purification presented a high viscosity and contamination. The enzymatic purification process was effective and allowed to maintain a high molar mass of PßG and its distinctive molecular structures (species with DP3 and DP4). The commercial samples CßG-A and CßG-B showed a low content of ß-glucans. Finally, CßG-C presented the best physicochemical and rheological properties for its subsequent application in food.

Antecedentes: los ß-glucanos (1-3: 1-4) son fibras solubles aplicadas a los alimentos debido a sus propiedades tecnológicas (capacidad de retención de agua, viscosidad, emulsificación y estabilización) y a sus efectos beneficiosos en la salud. Las propiedades funcionales de los ß-glucanos pueden perderse durante los procesos de extracción y purificación. La alta viscosidad de los ß-glucanos está relacionada con un alto peso molecular y con sus propiedades fisiológicas en el intestino. Por lo tanto, caracterizar la fibra después de su extracción y purificación es fundamental para comprender sus posibles aplicaciones en alimentos. Objetivos: caracterizar ß-glucanos extraídos (EßG) y compararlos con tres marcas comerciales (CßG-A, CßG-B y CßG-C) para identificar su futura aplicación en alimentos y evaluar si la purificación enzimática afecta molecular y estructuralmente los ß-glucanos. Métodos: se extrajeron ß-glucanos de cebada (EßG), caracterizados por análisis químicos, comportamiento reológico y color, y se compararon con tres muestras comerciales. Posteriormente, el extracto (EßG) se purificó y se identificaron sus características estructurales y su peso molecular. Resultados: EßG contenía 64.38 ± 3.54% de ß-glucanos, alta contaminación con almidón (12.70 ± 1.73%), alto contenido de calcio (8894 mg / kg), comportamiento pseudoplástico y color oscuro (L* = 52.77 ± 0.7). Todas las muestras comerciales mostraron una baja contaminación con almidón, color más claro y comportamiento newtoniano. Después de la purificación de EßG, la contaminación con almidón y proteína disminuyó (0.85 ± 0.46% y 5.50 ± 0.12%, respectivamente), aumentó el contenido de ßG (69.45 ± 0.81%) y aumentó su luminosidad (L* = 92.60 ± 1.70). Los ß-glucanos purificados (PßG) mostraron un peso molar de 690 ± 1,6 kDa y se identificaron en la estructura especies con grado de polimerización desde 3 (GP3) hasta 11 (GP11). Conclusiones: los EßG antes de la purificación presentaron alta viscosidad y contaminación. El proceso de purificación enzimática fue efectivo y permitió mantener una alta masa molar de la fibra y sus estructuras moleculares características (especies con GP3 y GP4). Las muestras comerciales CßG-A y CßG-B mostraron un bajo contenido de ß-glucanos. Finalmente, la CßG-C presentó las mejores propiedades fisicoquímicas y reológicas para su posterior aplicación en alimentos.
Descritores: beta-Glucanas
-Viscosidade
Fibras na Dieta
Alimentos Integrais
Peso Molecular
Limites: Humanos
Tipo de Publ: Artigo Clássico
Responsável: CO56.3 - Biblioteca


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Id: biblio-951792
Autor: Moghannem, Saad A. M; Farag, Mohamed M. S; Shehab, Amr M; Azab, Mohamed S.
Título: Exopolysaccharide production from Bacillus velezensis KY471306 using statistical experimental design
Fonte: Braz. j. microbiol;49(3):452-462, July-Sept. 2018. tab, graf.
Idioma: en.
Resumo: Abstract Exopolysaccharide (EPS) biopolymers produced by microorganisms play a crucial role in the environment such as health and bio-nanotechnology sectors, gelling agents in food and cosmetic industries in addition to bio-flocculants in the environmental sector as they are degradable, nontoxic. This study focuses on the improvement of EPS production through manipulation of different culture and environmental conditions using response surface methodology (RSM). Plackett-Burman design indicated that; molasses, yeast extract and incubation temperature are the most effective parameters. Box-Behnken RSM indicated that; the optimum concentration for each parameter was 12% (w/v) for molasses, 6 g/L yeast extract and 30 °C for incubation temperature. The most potent bacterial isolate was identified as Bacillus velezensis KY498625. After production, EPS was extracted, purified using DEAE-cellulose, identified using Fourier transform infrared (FTIR), gel permeation chromatography (GPC) and gas chromatography-mass spectroscopy (GC-MS). The result indicated that; it has molecular weight 1.14 × 105 D consisting of glucose, mannose and galactose.
Descritores: Polissacarídeos Bacterianos/metabolismo
Bacillus/metabolismo
-Polissacarídeos Bacterianos/isolamento & purificação
Polissacarídeos Bacterianos/química
Bacillus/química
Microbiologia Industrial
Espectroscopia de Infravermelho com Transformada de Fourier
Meios de Cultura/metabolismo
Meios de Cultura/química
Peso Molecular
Responsável: BR1.1 - BIREME


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Id: lil-788974
Autor: Gururaj, P; Ramalingam, Subramanian; Devi, Ganesan Nandhini; Gautam, Pennathur.
Título: Process optimization for production and purification of a thermostable, organic solvent tolerant lipase from Acinetobacter sp. AU07
Fonte: Braz. j. microbiol;47(3):647-657, July-Sept. 2016. tab, graf.
Idioma: en.
Projeto: Department of Biotechnology, Government of India.
Resumo: ABSTRACT The purpose of this study was to isolate, purify and optimize the production conditions of an organic solvent tolerant and thermostable lipase from Acinetobacter sp. AU07 isolated from distillery waste. The lipase production was optimized by response surface methodology, and a maximum production of 14.5 U/mL was observed at 30 ºC and pH 7, using a 0.5% (v/v) inoculum, 2% (v/v) castor oil (inducer), and agitation 150 rpm. The optimized conditions from the shake flask experiments were validated in a 3 L lab scale bioreactor, and the lipase production increased to 48 U/mL. The enzyme was purified by ammonium sulfate precipitation and ion exchange chromatography and the overall yield was 36%. SDS-PAGE indicated a molecular weight of 45 kDa for the purified protein, and Matrix assisted laser desorption/ionization time of flight analysis of the purified lipase showed sequence similarity with GDSL family of lipases. The optimum temperature and pH for activity of the enzyme was found to be 50 ºC and 8.0, respectively. The lipase was completely inhibited by phenylmethylsulfonyl fluoride but minimal inhibition was observed when incubated with ethylenediaminetetraacetic acid and dithiothreitol. The enzyme was stable in the presence of non-polar hydrophobic solvents. Detergents like SDS inhibited enzyme activity; however, there was minimal loss of enzyme activity when incubated with hydrogen peroxide, Tween 80 and Triton X-100. The kinetic constants (Km and Vmax) revealed that the hydrolytic activity of the lipase was specific to moderate chain fatty acid esters. The Vmax, Km and Vmax/Km ratio of the enzyme were 16.98 U/mg, 0.51 mM, and 33.29, respectively when 4-nitrophenyl palmitate was used as a substrate.
Descritores: Compostos Orgânicos
Solventes
Proteínas de Bactérias/isolamento & purificação
Proteínas de Bactérias/biossíntese
Acinetobacter/enzimologia
Lipase/isolamento & purificação
Lipase/biossíntese
-Compostos Orgânicos/química
Solventes/química
Especificidade por Substrato
Temperatura
Proteínas de Bactérias/química
Estabilidade Enzimática
Cinética
Cromatografia por Troca Iônica
Ativação Enzimática
Espaço Extracelular/enzimologia
Concentração de Íons de Hidrogênio
Íons
Lipase/química
Lipólise
Metais
Peso Molecular
Responsável: BR1.1 - BIREME



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