ABSTRACT Due to its zootechnical importance, the gene encoding growth hormone (GH) from several fish species has been isolated and expressed in heterologous systems. In this study, we isolated the cDNA sequence of GH of tambaqui (Colossoma macropomum), a species native to the Amazon basin and considered promising for Brazilian fish farming, and expressed it in the yeast Komagataella phaffii GS115. Heterologous expression was regulated by the AOX1 promoter and recombinant tambaqui GH (rtGH) secreted to the culture supernatant via the alpha factor of Saccharomyces cerevisiae. The rtGH was successfully expressed in shaken flask and bioreactor systems and purified using affinity chromatography. In the bioreactor, the production reached 250 mg L-1. This is the first report of the heterologous expression, purification and production of rtGH in a bioreactor. Our results contribute to making tambaqui a target species for future biotechnological innovations based on the significant levels of rtGH obtained in the bioreactor and the future zootechnical applications of this protein, which can be exploited in aquaculture. importance (GH study Colossoma macropomum, macropomum , macropomum) farming GS115 GS AOX (rtGH cerevisiae chromatography 25 L1. L1 L 1. 1 L-1 protein aquaculture GS11 2 L- GS1

RESUMO Devido à sua importância zootécnica, o gene que codifica o hormônio de crescimento (GH) de diversas espécies de peixes tem sido isolado e expresso em sistemas heterólogos. Neste estudo, isolamos a sequência de cDNA do GH do tambaqui (Colossoma macropomum), espécie nativa da bacia amazônica e considerada promissora para a piscicultura brasileira, e a expressamos na levedura Komagataella phaffii GS115. A expressão heteróloga foi regulada pelo promotor AOX1 e GH de tambaqui recombinante (rtGH) foi secretado para o sobrenadante da cultura através do fator alfa de Saccharomyces cerevisiae. O rtGH foi expresso com sucesso em frascos agitados e sistemas de biorreatores e purificado por cromatografia de afinidade. No biorreator a produção atingiu 250 mg L-1. Este é o primeiro relato da expressão heteróloga, purificação e produção de rtGH em biorreator. Nossos resultados contribuem para tornar o tambaqui uma espécie alvo para futuras inovações biotecnológicas baseadas nos níveis significativos de rtGH obtidos no biorreator e nas futuras aplicações zootécnicas desta proteína, que pode ser explorada na aquicultura. zootécnica (GH heterólogos estudo Colossoma macropomum, macropomum , macropomum) brasileira GS115 GS AOX (rtGH cerevisiae afinidade 25 L1. L1 L 1. 1 L-1 proteína aquicultura GS11 2 L- GS1

Os precursores mais empregados para obtenção da fibra de carbono são a poliacrilonitrila, o piche e o rayon. Com o aumento da demanda de materiais compósitos leves e de alta resistência, a produção da fibra de carbono tem aumentado exponencialmente e novos precursores têm sido requisitados. A lignina, um material orgânico natural, tem sido estudada como possível candidata para a produção das fibras. Além de ser rejeito industrial e de biorrefinarias, as fibras de carbono obtidas com a lignina podem ter um custo mais vantajoso quando comparado com os demais precursores comerciais. Em decorrência da biodiversidade de vegetais na natureza, a lignina pode apresentar diferentes estruturas. O método de recuperação também é parâmetro que influencia diretamente na sua composição. Como as propriedades térmicas para extrusão são fortemente dependentes dessas e de muitas outras variáveis, muitas pesquisas têm sido reportadas utilizando lignina pura de diversas fontes, lignina modificada e com adição de plastificantes. As etapas de pré-tratamento, termoestabilização e carbonização/grafitização também têm sido avaliadas para conferir melhores propriedades mecânicas ao produto final. Não obstante serem muitas as condições para a produção da fibra de carbono a partir da lignina, resultados bastante promissores têm sido alcançados. As aplicações em diversas áreas estão diretamente relacionadas com o conjunto de propriedades mecânicas inerentes a ela. A fibra de carbono proveniente da lignina vem contribuir com inúmeros empregos no mercado, de maneira a baratear sua produção, minimizar a dependência de materiais fósseis e tornar sua manufatura ecologicamente sustentável.

The most applied precursors to obtain carbon fiber are polyacrylonitrile, pitch and rayon. With the increased demand of lightweight composites and high strength, carbon fiber manufacting has increased exponentially and new precursors are been request. Lignin, a natural organic material, has been studied as possible candidate. Besides being an industrial and biorrefinary waste, lignin-based carbon fibers have a lower cost compared to other commercial precursors. Because of the biodiversity of plants in nature, lignin can present different structures. The lignin extraction method can directly influence the composition. As thermic extrusion properties are hardly dependent on this and other properties, many researches have been reported using pure lignin from different sources, modified lignin and plasticizers addiction. Steps of pretreatment, thermostabilization and carbonization/graphitization have also been evaluated to give better mechanical properties to the final product. Nevertheless many conditions required to manufacture lignin-based carbon fiber, promising results have been achieved. The set of mechanical properties inherent to the fiber allows its application to different areas. Lignin-based carbon comes to collaborate on cheapening its production, minimizing the dependence of fossil materials and making its manufacture process ecologically sustainable.

The growing concerns regarding the future of global energy supplies have encouraged R&D in alternative sources to complement the global energy matrix. Brazil has earned notoriety as one of the largest producers of ethanol and biomass. This has aroused other countries’ interest in Brazil’s capabilities. However, the technologies for converting biomass into ethanol are not sufficiently mature, and have been developed in a decentralized manner. The lignocellulosic ethanol technologies can be divided into the following three groups: pretreatment, enzymes and ethanol production. One of the most efficient methods for mapping such technologies is through patent applications because the applications provide important information on trends in long-term technological development. Among all the patent applications deposited in the Brazilian database, pretreatment technologies were the most exploited followed by enzymes and ethanol production. The large number of documents from USA and European countries efforts the importance of Brazil in this technological scenario.

The use of enzymes in the pulp and paper industry was introduced in the 1986. However, their use has been relatively minor. This prospective study aims at enhancing the understanding of the most important advances regarding the use of enzymes in this industry and to identify the future trends of this technology. Information gathered from the Web of Science shows a growing number of papers published on this topic indicating an increased interest in this issue. A study on patents also displayed a high number documents related to this technology. Cellulase, xylanase, laccase and lipase are the most important enzymes that can be used in the pulp and paper processes. Furthermore, the key objectives of enzymes development have been in the bleaching boosting with xylanases and fiber modification with cellulases. The current and future trends on the development of enzymes are focused on increasing their thermostability and their alkalinity strength.

The influence of different nutrients on biosurfactant production by Rhodococcus erythropolis was investigated. Increasing the concentration of phosphate buffer from 30 up through 150 mmol/L stimulated an increase in biosurfactant production, which reached a maximum concentration of 285 mg/L in shaken flasks. Statistical analysis showed that glycerol, NaNO3,MgSO4 and yeast extract had significant effects on production. The results were confirmed in a batchwise bioreactor, and semi-growth-associated production was detected. Reduction in the surface tension, which indicates the presence of biosurfactant, reached a value of 38 mN/m at the end of 35 hours. Use of the produced biosurfactant for washing crude oil-contaminated soil showed that 2 and 4 times the critical micellar concentration (CMC) were able to remove 97 and 99% of the oil, respectively, after 1 month of impregnation.

Flocculent Zymomonas mobilis breaks down in smaller flocs and individual cells when centrifuged. The main consequence of it is an increase in the dispersion of the sample, suggesting that the influence of the centrifugal force on the aggregation of cells is worth to study. The experiments showed that the degree of dispersion varied between 30% and 100% when the centrifugal acceleration changed from 25 g to 2500 g. Observation under the electronic microscope showed that a slimy material covered the cells recovered by gentle gravitational settling and, that the centrifuged cells presented a bare cell wall.

O cultivo de Aspergillus awamori em bagaço de cana, finamente dividido e sem tratamento prévio, favoreceu a produção de níveis elevados de endo-xilanase (100 U/ml) e beta-xilosidase (3.5 U/ml) extracelulares. A caracterização parcial das enzimas xilanolíticas permitiu estabelecer os parâmetros cinéticos aparentes, sendo Km igual a 3.12 ( 0.05 mg/ml para endox-xilanase (xilana birchwood) e 0.45 ( 0.05 mM para beta-xylosidase (p-nitrofenil beta-D-xilanopiranosideo). Os correspondentes valores de Vm foram 6.63 ( 0.02 e 0.078 ( 0.02 mimol/min. Observou-se uma repressão gradual da atividade endo-xilanásica quando concentrações crescentes de glicose e xilose (1, 2, 4, 6 e 8 g de carboidrato / 4 g de bagaço de cana) foram adicionadas aos meios de produção. Contrariamente, a atividade beta-xilosidásica foi estimulada quando níveis baixos de açúcares simples foram utilizados (1 g de xilose ou glicose/ 4 g de bagaço de cana).

Aspergillus awamori showed high extracellular endoxylanase (100 U/ml) and beta-xylosidase activities (3.5 U/ml) when grown on milled sugar cane bagasse as the principal carbon source without treatment. Partial characterization of xylanases showed that the apparent values of Km were 3.12 ± 0.05 mg/ml for endoxylanase (in birchwood xylan) and 0.45 ± 0.05 mM for beta-xylosidase (in p -nitrophenyl beta-D-xylanopiranoside). Corresponding values of Vmax were 6.63 ± 0.02 and 0.078 ± 0.02 mumol/min. Gradual repression of endoxylanase activity was observed when increasing concentrations of glucose and xylose (1, 2, 4, 6 and 8 g of carbohydrate / 4 g of sugar cane bagasse) were added to production media. In contrast, beta-xylosidase activity was stimulated using low levels of carbohydrates (1 g xylose or glucose/ 4 g of sugar cane bagasse).

O presente trabalho trata do estudo do efeito da temperatura na atividade e estabilidade de endo-xilanase e beta-xilosidase produzidas, extracelularmente, por Aspergillus awamori. O cultivo deste microrganismo, em bagaço de cana finamente dividido, produziu predominantemente endo-xilanases (30 U/ml) e menores atividades de beta-xilosidase (1,3 U/ml); esta última exibiu considerável estabilidade em faixa de temperatura variando de 35 a 55°C, por outro lado verificou-se uma menor termo estabilidade para a endoenzima. A estabilidade térmica de endo-xilanase foi aumentada consideravelmente através da adição de polióis, principalmente xilitol e sorbitol em concentração de 2,0 M. No que concerne à estocagem a baixa temperatura (-4°C), observou-se uma estabilidade particular na atividade endo-xilanásica (100%), durante 165 dias, porém, um decréscimo de aproximadamente 20% na atividade beta-xilosidásica foi verificado após os primeiros 15 dias de armazenamento nas mesmas condições, mantendo-se aproximadamente em 75% da atividade inicial no mesmo período de tempo.

The effect of temperature on the activity and stability of endoxylanase and beta-xylosidase from Aspergillus awamori was investigated. The growth of A. awamori in milled sugar cane bagasse produced predominantly extracellular endoxylanase (30 U/ml) and lower amounts of beta-xylosidase (1.3 U/ml). Grown in sugar cane bagasse as the principal carbon source, the microorganism produced a quite stable beta-xylosidase in a temperature range of 35-55°C, but it exhibited a lower thermostable endoxylanase. The thermostability of endoxylanase was enhanced through addition of polyhydric alcohols, mainly 2 M xylitol and sorbitol solutions. Particular stability upon storage (100%) was found for endoxylanase at -4°C for 165 days. Yet for beta-xylosidase, an activity decrease of approximately 20% was observed during the first 15 days of storage, maintaining roughly 75% of initial activity until the end of the experiment.