Produção e caracterização de exopolissacarídeo (EPS) bacteriano
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Data
2022-02-08
Autores
Minari, Guilherme Deomedesse [UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Os EPS são polímeros naturais, não tóxicos, biodegradáveis e desempenham um papel importante em uma variedade de mecanismos biológicos. São macromoléculas secretadas por microrganismos compostas por resíduos de monossacarídeos e derivados de açúcares divididos em dois grupos de acordo com sua composição: homopolissacarídeos e heteropolissacarídeos, sendo a diferença entre estes dois grupos os tipos de açúcares que os compõem. A produção de EPS via lactobacilus tem basicamente glicose e lactose como fontes de carbono e, neste estudo, foi demonstrada a possibilidade de utilizar resíduos como melaço de cana-de-açúcar e glicerol como fonte de carbono para produção deste polímero. Para tal, um screening entre 22 cepas de Lactobacillus foi realizado a fim de se determinar a cepa que apresentasse a maior produção, com base no peso seco a 37 ºC, de EPS produzido a partir da composição do meio contendo glicerol como principal fonte de carbono. O processo fermentativo para produção de EPS foi realizado em triplicata em 3 L de meio semi definido (SDM) modificado em bioreator (Zeta). O EPS obtido foi extraído, dialisado, liofilizado e purificado. A quantidade de EPS foi mensurada através da concentração de açúcares totais presente nas amostras por meio do método do fenol ácido sulfúrico e o teor de proteínas foi quantificado de acordo com o método de Lowry. O Lactobacillus casei foi capaz de crescer em meios utilizando fontes de carbono alternativas e capaz de produzir EPS. A caracterização da molécula foi realizada em HPLC, cromatografia por exclusão de tamanho, GC-MS, FT-IR e RMN 1H13C. Os resultados obtidos sugerem que o EPS obtido utilizando glicerol e glicose como fonte de carbono é classificado como heteropolissacarideo composto por glicose e manose com 106 g.mol-1 , e 39,4% de carboidratos e 18% de proteínas, enquanto o EPS obtido utilizando melaço como fonte de carbono é caracterizado como heteropolissacarídeo composto por glicose, galactose e arabinose com 81 g.mol-1 , 52,9% de carboidratos e 11,69% de proteínas. A existência dos polissacarídeos foi comprovada pelas análises de FT-IR e RMN.
EPS are natural, non-toxic, biodegradable polymers and play an important role in a variety of biological mechanisms. They are macromolecules secreted by microorganisms composed of monosaccharide residues and sugar derivatives divided into two groups according to their composition: homopolysaccharides and heteropolysaccharides, the difference between these two groups being the types of sugars that compose them. The production of EPS via lactobacillus basically has glucose and lactose as carbon sources and, in this study, the possibility of using residues such as sugarcane molasses and glycerol as a carbon source for the production of this polymer was demonstrated. For this, a screening between 22 strains of Lactobacillus was carried out in order to determine the strain that presented the highest production, based on dry weight at 37 ºC, of EPS produced from the composition of the medium containing glycerol as the main carbon source. The fermentation process for the production of EPS was carried out in triplicate in 3 L of semi-defined medium (SDM) modified in a bioreactor (Zeta). The EPS obtained was extracted, dialyzed, lyophilized and purified. The amount of EPS was measured through the concentration of total sugars present in the samples using the phenol sulfuric acid method and the protein content was quantified according to the Lowry method. Lactobacillus casei was able to grow in media using alternative carbon sources and was able to produce EPS. The characterization of the molecule was performed using HPLC, size exclusion chromatography, GC-MS, FT-IR and 1H13C NMR. The results obtained suggest that the EPS obtained using glycerol and glucose as carbon source is classified as a heteropolysaccharide composed of glucose and mannose with 106 g.mol-1, and 39.4% of carbohydrates and 18% of proteins, while the EPS obtained using molasses as a carbon source is characterized as a heteropolysaccharide composed of glucose, galactose and arabinose with 81 g.mol-1, 52.9% of carbohydrates and 11.69% of proteins. The existence of polysaccharides was confirmed by FT-IR and NMR analyses.
EPS are natural, non-toxic, biodegradable polymers and play an important role in a variety of biological mechanisms. They are macromolecules secreted by microorganisms composed of monosaccharide residues and sugar derivatives divided into two groups according to their composition: homopolysaccharides and heteropolysaccharides, the difference between these two groups being the types of sugars that compose them. The production of EPS via lactobacillus basically has glucose and lactose as carbon sources and, in this study, the possibility of using residues such as sugarcane molasses and glycerol as a carbon source for the production of this polymer was demonstrated. For this, a screening between 22 strains of Lactobacillus was carried out in order to determine the strain that presented the highest production, based on dry weight at 37 ºC, of EPS produced from the composition of the medium containing glycerol as the main carbon source. The fermentation process for the production of EPS was carried out in triplicate in 3 L of semi-defined medium (SDM) modified in a bioreactor (Zeta). The EPS obtained was extracted, dialyzed, lyophilized and purified. The amount of EPS was measured through the concentration of total sugars present in the samples using the phenol sulfuric acid method and the protein content was quantified according to the Lowry method. Lactobacillus casei was able to grow in media using alternative carbon sources and was able to produce EPS. The characterization of the molecule was performed using HPLC, size exclusion chromatography, GC-MS, FT-IR and 1H13C NMR. The results obtained suggest that the EPS obtained using glycerol and glucose as carbon source is classified as a heteropolysaccharide composed of glucose and mannose with 106 g.mol-1, and 39.4% of carbohydrates and 18% of proteins, while the EPS obtained using molasses as a carbon source is characterized as a heteropolysaccharide composed of glucose, galactose and arabinose with 81 g.mol-1, 52.9% of carbohydrates and 11.69% of proteins. The existence of polysaccharides was confirmed by FT-IR and NMR analyses.
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Palavras-chave
Polímero extracelular, Glicose, Manose, Galactose, Arabinose, Biodegradáveis, Lactobacillus, Extracellular polymer, Glucose, Mannose, Galactose, Arabinose, Biodegradable