Despirogenação de soro antiofidico : adsorção seletiva em membrana de quitosana

Orientador: Sonia Maria Alves BuenoDissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia QuimicaResumo: A indústria farmacêutica enfrenta dificuldades com relação a susceptibilidade de seus produtos à contaminação por endotoxinas (ETs). Estas moléculas são lipopolissacarídeos presentes na parede externa de bactérias Gram negativas e devido apresentar alta toxicidade a sua remoção é essencial para administração parenteral segura. Este trabalho visou a remoção de ETs de soro antiofidico, produto farmacêutico que consiste na única forma de tratamento de vítimas de acidentes antiofidicos. A técnica utilizada foi a filtração em membrana reticulada de quito sana por esta demonstrar potencial para a interação com endotoxinas. A membrana de quito sana foi sintetizada em nosso laboratório e apresentou valores de vazão, tensão e área de adsorção (13 rnL.min°1, 0,81 MPa e 15,9 cm2, respectivamente) adequados para a execução dos experimentos. Inicialmente, investigou-se a existência de adsorção de anticorpos do soro na membrana e notou-se que em soluções tamponantes acetato de sódio pH 5,0 e Tris-HCI pH 7,0 (4,25% e 4,67%, respectivamente) obteve-se uma menor adsorção. Em seguida, estudou-se a adsorção de ETs em membranas a partir das soluções tamponantes definidas anteriormente. Neste estudo verificou-se que em tampão Tris-HCI pH 7,0 foram alcançados maiores porcentagens de remoção (97,7% e 99,0% para soluções contaminadas nas concentrações de 150 EU.rnL-1e 1000 EU.rnL-t, respectivamente). Na despirogenação de soluções de soro antiofidico contendo baixa concentração de anticorpos (1 mg.rnL-1) altos valores para a remoção de endotoxinas e recuperação de anticorpos foram observados, chegando a apresentar 1 % de endotoxinas remanescentes e 94,6% de proteína recuperada no produto final inicialmente contaminado com 1000 EU.rnL-1. Os resultados obtidos para a remoção de ETs de soluções de soro contendo alta concentração de anticorpos (cerca de 15 mg.rnL-1), também foram significativos quanto a eficiência de remoção de ETs e recuperação de anticorpos para soluções inicialmente contaminadas com 1000 EU.rnL-1 (98,7% e 82% de ET removida e de proteína recuperada, respectivamente). Além disso, notou-se também que o aumento na concentração de anticorpos resultou em uma maior dificuldade na recuperação destes, sem contudo representar uma dificuldade ao processo de remoção por adsorçãoAbstract: Contamination of therapeutic solutions by endotoxins (ET's) are ofmajor concem in the pharmaceutical industry. The ET's are lipopolyscaccharides found in the outer cell membranes of Gram-negative bacteria and are known to cause strong reactions in man after intravenous application. For this reason, the removal of ET's ftom therapeutic solutions is a major challenge in downstream processing. In this work, we studied the removal of ET' s ftom snake antivenom serum, the therapeutic employed on the treatment of snake bite accidents. The technique studied for the this removal was the membrane filtration using cross-linked chitosan membranes. We investigated the chitosan membrane synthesis that presented appropriated features of flowrate, resistance to stress and adsorption area (13 rnL.min-l, 0,81 MPa and 15,9 cm2, respectively). The adsorption of proteins (antibodies) and ET's on the membrane was also investigated. The influence of three different buffer systems (Tris-HCI, sodium acetate and phosphate) on the adsorption of antibodies was studied. The best conditions found (lower antibody adsorption) were: pH 5,0 using acetate buffer, and pH 7,0 using Tris-HCI buffer (4,25% and 4,67% of antibody adsorption, respectively). The adsorption of ET's on the membrane when using the selected conditions was studied. Higher ET clearances were obtained using the Tris-HCI pH 7,0 buffer (97,7% and 99,0% for solutions containing initial ET concentrations of 150 and 1000 EU.rnL-MestradoDesenvolvimento de Processos BiotecnologicosMestre em Engenharia Químic

Enhancing acetic acid and 5-hydroxymethyl furfural tolerance of C. saccharoperbutylacetonicum through adaptive laboratory evolution

Supplementary material related to this article can be found, in the online version, at doi:https://doi.org/10.1016/j.procbio.2020.11.013.In this study, adaptive laboratory evolution (ALE) was applied to isolate four strains of Clostridium saccharoperbutylacetonicum able to grow in the presence of hemicellulosic hydrolysate inhibitors unsupported by the parental strain. Among them, isolate RAC-25 presented the best fermentative performance, producing 22.1g/L of ABE and 16.7g/L of butanol. Genome sequencing revealed a deletion in the arabinose transcriptional repressor gene (araR) and a mutation in the anti-sigma factor I that promoted a downregulation of sigI. Gene expression analysis indicated high expression of genes related to H+-pumps (ATP synthases), proline biosynthesis (gamma phosphate reductase) and chaperonins (Grol), suggesting an integrated mechanism that is probably coordinated by the repression of sigI. Therefore, in addition to highlighting the power of ALE for selecting robust strains, our results suggest that sigI and araR may be interesting gene targets for increased tolerance toward inhibitor compounds relevant for lignocellulosic biofuels production.The authors would like to thank the Brazilian Center for Research in Energy and Materials (CNPEM) for providing access to the bioprocess facility of the Brazilian Biorenewables National Laboratory, and CNPq (400803/2013-5), FCT (UID/BIO/04469), BioTecNorte Operation (NORTE-01-0145-FEDER-000004) and Portuguese Biological Data Network” (ref. LISBOA-01-0145-FEDER-022231) for financial support.info:eu-repo/semantics/publishedVersio

Transcriptome Profile of Trichoderma harzianum IOC-3844 Induced by Sugarcane Bagasse

Profiling the transcriptome that underlies biomass degradation by the fungus Trichoderma harzianum allows the identification of gene sequences with potential application in enzymatic hydrolysis processing. in the present study, the transcriptome of T. harzianum IOC-3844 was analyzed using RNA-seq technology. the sequencing generated 14.7 Gbp for downstream analyses. de novo assembly resulted in 32,396 contigs, which were submitted for identification and classified according to their identities. This analysis allowed us to define a principal set of T. harzianum genes that are involved in the degradation of cellulose and hemicellulose and the accessory genes that are involved in the depolymerization of biomass. An additional analysis of expression levels identified a set of carbohydrate-active enzymes that are upregulated under different conditions. the present study provides valuable information for future studies on biomass degradation and contributes to a better understanding of the role of the genes that are involved in this process.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Univ Campinas UNICAMP, CBMEG, Campinas, SP, BrazilBrazilian Ctr Res Energy & Mat CNPEM, Brazilian Bioethanol Sci & Technol Lab CTBE, Campinas, SP, BrazilUniv São Paulo, Phys Inst Sao Carlos, Sao Carlos, SP, BrazilFed Univ São Paulo UNIFESP, Inst Sci & Technol, Sao Jose Dos Campos, SP, BrazilUniv Campinas UNICAMP, Dept Plant Biol, Inst Biol, Campinas, SP, BrazilFed Univ São Paulo UNIFESP, Inst Sci & Technol, Sao Jose Dos Campos, SP, BrazilWeb of Scienc

Improvement On Sugar Cane Bagasse Hydrolysis Using Enzymatic Mixture Designed Cocktail.

The aim of this work was to study cocktail supplementation for sugar cane bagasse hydrolysis, where the enzymes were provided from both commercial source and microorganism cultivation (Trichoderma reesei and genetically modified Escherichia coli), followed by purification. Experimental simplex lattice mixture design was performed to optimize the enzymatic proportion. The response was evaluated through hydrolysis microassays validated here. The optimized enzyme mixture, comprised of T. reesei fraction (80%), endoglucanase (10%) and β-glucosidase (10%), converted, theoretically, 72% of cellulose present in hydrothermally pretreated bagasse, whereas commercial Celluclast 1.5L converts 49.11%±0.49. Thus, a rational enzyme mixture designed by using synergism concept and statistical analysis was capable of improving biomass saccharification.187173-18

Improvement on sugar cane bagasse hydrolysis using enzymatic mixture designed cocktail

The aim of this work was to study cocktail supplementation for sugar cane bagasse hydrolysis, where the enzymes were provided from both commercial source and microorganism cultivation (Trichoderma reesei and genetically modified Escherichia coli), followed by purification. Experimental simplex lattice mixture design was performed to optimize the enzymatic proportion. The response was evaluated through hydrolysis microassays validated here. The optimized enzyme mixture, comprised of T. reesei fraction (80%), endoglucanase (10%) and β-glucosidase (10%), converted, theoretically, 72% of cellulose present in hydrothermally pretreated bagasse, whereas commercial Celluclast 1.5 L converts 49.11% ± 0.49. Thus, a rational enzyme mixture designed by using synergism concept and statistical analysis was capable of improving biomass saccharification187173181FUNDAÇÃO DE AMPARO À PESQUISA DO ESTADO DE SÃO PAULO - FAPESP2008/57873-8; 2010/08089-2; 2012/23223-

MOESM3 of Techno-economic analysis of the industrial production of a low-cost enzyme using E. coli: the case of recombinant β-glucosidase

Additional file 3. Detailed simulation data—optimized scenarios. This file lists a compilation of the results for cellulase production found in the literature, including results of this work. There is also a list of 25 different simulation scenarios for the recombinant β-glucosidase process generated in this work

MOESM2 of Techno-economic analysis of the industrial production of a low-cost enzyme using E. coli: the case of recombinant β-glucosidase

Additional file 2. Cost data and economic indices. This file lists the costs of raw materials, utilities, labor, financing, as well as price indices used in the economic analysis