Biosurfactants synthesized by endophytic Bacillus strains as control of Moniliophthora perniciosa and Moniliophthora roreri

ABSTRACT: Ecuador is responsible for up to 70 % of the worlds production of fine flavor cocoa. Although fungal diseases have reduced cocoa production, ecofriendly alternatives have been implemented to control the spread of phytopathogens. This study aimed to explore the potential of endophytic strains isolated from fine flavor Nacional cocoa trees to synthetize compounds with antimicrobial and surface active properties. It was determined that under in vitro conditions, seven strains (DS03, DS07, DS18, DS23, DS31, DS34 and DS50) exhibited antifungal activity that inhibited the mycelial growth of Moniliophthora roreri and Moniliophthora perniciosa. The strains were identified based on their morphological (pattern forming Bacilli) and genetic characteristics using the 16S SSU rRNA gene sequences. Furthermore, we determined that the endophytic strains contained genes that encoded the lipopeptides Fengycin (fenA, fenB, fenC, fenE), Iturin (ituB, ituC, ituD), and Bacyllomicin D (bamC, bamD). The production of biosurfactant compounds was determined by a reduction in surface tension in the mineral medium. Additionally, the biosurfactant crude extract (BCE) was tested for its inhibitory activity against M. perniciosa and M. roreri. The minimal inhibitory concentration (MIC) was determined to be 0.07 and 0.035 mg mL1, respectively, in both cases in a fungistatic action mode. The swelling granulation and fragmentation of both phytopathogens hyphae were observed using the BCE treatment. These results suggested the biocontrol capacity of the endophytic strains, which under in vitro conditions, have the ability to synthesize antifungal and surface active compounds that might be used as an ecological alternative in cocoa plants to prevent infection by phytopathogens.

Biosurfactants synthesized by endophytic Bacillus strains as control of Moniliophthora perniciosa and Moniliophthora roreri

Ecuador is responsible for up to 70 % of the world’s production of fine flavor cocoa. Although fungal diseases have reduced cocoa production, eco–friendly alternatives have been implemented to control the spread of phytopathogens. This study aimed to explore the potential of endophytic strains isolated from fine flavor Nacional cocoa trees to synthetize compounds with antimicrobial and surface active properties. It was determined that under in vitro conditions, seven strains (DS03, DS07, DS18, DS23, DS31, DS34 and DS50) exhibited antifungal activity that inhibited the mycelial growth of Moniliophthora roreri and Moniliophthora perniciosa. The strains were identified based on their morphological (pattern forming Bacilli) and genetic characteristics using the 16S SSU rRNA gene sequences. Furthermore, we determined that the endophytic strains contained genes that encoded the lipopeptides Fengycin (fenA, fenB, fenC, fenE), Iturin (ituB, ituC, ituD), and Bacyllomicin D (bamC, bamD). The production of biosurfactant compounds was determined by a reduction in surface tension in the mineral medium. Additionally, the biosurfactant crude extract (BCE) was tested for its inhibitory activity against M. perniciosa and M. roreri. The minimal inhibitory concentration (MIC) was determined to be 0.07 and 0.035 mg mL−1, respectively, in both cases in a fungistatic action mode. The swelling granulation and fragmentation of both phytopathogens’ hyphae were observed using the BCE treatment. These results suggested the biocontrol capacity of the endophytic strains, which under in vitro conditions, have the ability to synthesize antifungal and surface active compounds that might be used as an ecological alternative in cocoa plants to prevent infection by phytopathogens.

Biosurfactants synthesized by endophytic Bacillus strains as control of Moniliophthora perniciosa and Moniliophthora roreri

Ecuador is responsible for up to 70 % of the worlds production of fine flavor cocoa. Although fungal diseases have reduced cocoa production, eco–friendly alternatives have been implemented to control the spread of phytopathogens. This study aimed to explore the potential of endophytic strains isolated from fine flavor Nacional cocoa trees to synthetize compounds with antimicrobial and surface active properties. It was determined that under in vitro conditions, seven strains (DS03, DS07, DS18, DS23, DS31, DS34 and DS50) exhibited antifungal activity that inhibited the mycelial growth of Moniliophthora roreri and Moniliophthora perniciosa. The strains were identified based on their morphological (pattern forming Bacilli) and genetic characteristics using the 16S SSU rRNA gene sequences. Furthermore, we determined that the endophytic strains contained genes that encoded the lipopeptides Fengycin (fenA, fenB, fenC, fenE), Iturin (ituB, ituC, ituD), and Bacyllomicin D (bamC, bamD). The production of biosurfactant compounds was determined by a reduction in surface tension in the mineral medium. Additionally, the biosurfactant crude extract (BCE) was tested for its inhibitory activity against M. perniciosa and M. roreri. The minimal inhibitory concentration (MIC) was determined to be 0.07 and 0.035 mg mL−1, respectively, in both cases in a fungistatic action mode. The swelling granulation and fragmentation of both phytopathogens hyphae were observed using the BCE treatment. These results suggested the biocontrol capacity of the endophytic strains, which under in vitro conditions, have the ability to synthesize antifungal and surface active compounds that might be used as an ecological alternative in cocoa plants to prevent infection by phytopathogens.(AU)

Green (Detox) juice physicochemical properties and stabilization effect of naturals emulsifiers / Propriedades físico-químicas do suco verde (detox) e efeito de estabilização de emulsificantes naturais

Green or “detox” juice is a mixture of fruit juice with vegetables, which has been used intensively by consumers seeking for healthy food. Physicochemical properties of Green juice were accessed in the present research, which brings new insights for the use of this beverage in human diet. A total phenolic content of 2833.60 mg GAE (Gallic acid equivalent)/ g of juice and a Total Antioxidant Capacity by FRAP of 323.62 µM Fe2SO4 / g of juice and by ABTS•+ of 333.11 µM Trolox/ g of juice, indicated good antioxidant properties. Low energy and reducing sugar content indicate its use for low calorie diet, but low carbohydrate and protein content prove that Green juice cannot be used as meal replacement. The addition of a microbial biosurfactant (YlBio) and chia gel as bioemulsifiers was tested in the Green juice formulation to reduce solid decantation and increase consistency. YlBio and chia gel were able to change the Newtonian behavior of the Green juice to a Pseudoplastic behavior due to stabilization properties and also increase consistency, without the need to add synthetic stabilizers.(AU)

O suco verde ou “detox” é uma mistura de suco de frutas com vegetais que tem sido intensamente utilizado por consumidores que buscam alimentos saudáveis. As propriedades físico-químicas do suco verde foram avaliadas no presente trabalho, o que traz novas perspectivas para o uso dessa bebida na dieta humana. Um conteúdo fenólico total de 2833,60 mg de EAG (equivalente em ácido gálico) / g de suco) e uma capacidade antioxidante total por FRAP de 323,62 µM de Fe2SO4 / g de suco e por ABTS•+ de 333,11 µM de Trolox / g de suco, indicam boas propriedades antioxidantes. Um baixo teor de energia e açúcar redutor indica seu uso em dietas de baixa caloria, mas o baixo teor de carboidratos e proteínas prova que o suco verde não pode ser usado como substituto de refeição. A adição de um biossurfactante microbiano (YlBio) e do gel de chia no suco foi testada na formulação do suco verde, para reduzir a decantação de sólidos e aumentar a consistência. YlBio e o gel de chia foram capazes de mudar o comportamento do suco de fluido Newtoniano para um fluido pseudoplástico devido às propriedades estabilizantes, e também aumentaram a consistência do suco, sem a necessidade de adição de estabilizantes sintéticos.(AU)

Isolation and characterization of hydrocarbon-degrading bacteria from gas station leaking-contaminated groundwater in the Southern Amazon, Brazil / Isolamento e caracterização de bactérias degradadoras de hidrocarbonetos de água subterrânea contaminada por vazamentos em posto de combustível no sul da Amazônia, Brasil

Twenty-three hydrocarbon-degrading bacteria strains were isolated from gas station leaking-contaminated groundwater located in the Southern Amazon, Brazil. Based on hydrocarbon (diesel, hexadecane, benzene, toluene and xylene) degradation ability, two strains were selected for further study. The amplification and sequencing of the 16S rRNA gene showed that these two strains belonged to the genus Bacillus (Bacillus sp. L26 and Bacillus sp. L30). GC-MS analysis showed that strain L30 was the most effective in degrading n-alkane (C10-C27) from diesel after 7 days of cultivation in mineral medium. Both strains produced biosurfactants and showed emulsification activity, specially the strain L30. Alkane hydroxylase gene (group III), which is important for alkane biodegradation, was present in strains. As a result, this study indicated that these bacteria could have promising applications in hydrocarbon bioremediation.(AU)

Vinte e três linhagens bacterianas degradadoras de hidrocarbonetos foram isoladas de água subterrânea contaminada por vazamento em posto de combustível no sul da Amazônia, Brasil. Com base na habilidade de degradar hidrocarbonetos (diesel, hexadecano, benzeno, tolueno e xileno), duas linhagens foram selecionadas para estudos posteriores. A amplificação e sequenciamento do gene 16S rRNA demonstrou que essas linhagens pertencem ao gênero Bacillus (Bacillus sp. L26 and Bacillus sp. L30). Análises de GC-MS mostraram que a linhagem L30 foi mais eficiente em degradar n-alcanos (C10-C27) presentes no diesel, após 7 dias de cultivo em meio mineral. Ambas as linhagens produziram biossurfactantes e apresentaram atividade emulsificante, especialmente a linhagem L30. O gene alcano hidroxilase (grupo III), o qual é importante para degradação de alcanos, foram detectados nas linhagens. Como resultado, este estudo indicou que essas linhagens bacterianas podem ser promissoras se aplicadas em processos de biorremediação.(AU)

Biosurfactant production by Phialemonium sp. using agroindustrial wastes: influence of culture conditions

Biosurfactant are surface active compounds with emulsifying capacity and are produced by microorganisms, and they may be affected by factors related to microbial cultivation, such as pH, salinity, incubation time, carbon and nitrogen sources. The aim of this work was to study the influence of the culture conditions on the production of biosurfactants by Phialemonium sp using agroindustrial wastes. The processing parameters of temperature, humidity and pH produced the most significant effects on the production of biosurfactant and emulsifying activity. The maximum concentration of biosurfactant obtained in this study was equivalent to a surface tension reduction of 8.5 mg L-1 surfactin commercial solution using wheat bran, pH of 4.5, and 0.5% of soybean oil added at 30°C. Under these conditions, 83.4 EU g-1 of emulsifying activity, 16.4 g L-1 of emulsifier index and 18.3 U g-1 lipolytic capacity were obtained.

Biosurfactant production by Phialemonium sp. using agroindustrial wastes: influence of culture conditions

Biosurfactant are surface active compounds with emulsifying capacity and are produced by microorganisms, and they may be affected by factors related to microbial cultivation, such as pH, salinity, incubation time, carbon and nitrogen sources. The aim of this work was to study the influence of the culture conditions on the production of biosurfactants by Phialemonium sp using agroindustrial wastes. The processing parameters of temperature, humidity and pH produced the most significant effects on the production of biosurfactant and emulsifying activity. The maximum concentration of biosurfactant obtained in this study was equivalent to a surface tension reduction of 8.5 mg L-1 surfactin commercial solution using wheat bran, pH of 4.5, and 0.5% of soybean oil added at 30°C. Under these conditions, 83.4 EU g-1 of emulsifying activity, 16.4 g L-1 of emulsifier index and 18.3 U g-1 lipolytic capacity were obtained.(AU)

Isolation and characterization of hydrocarbon-degrading bacteria from gas station leaking-contaminated groundwater in the Southern Amazon, Brazil

Abstract Twenty-three hydrocarbon-degrading bacteria strains were isolated from gas station leaking-contaminated groundwater located in the Southern Amazon, Brazil. Based on hydrocarbon (diesel, hexadecane, benzene, toluene and xylene) degradation ability, two strains were selected for further study. The amplification and sequencing of the 16S rRNA gene showed that these two strains belonged to the genus Bacillus (Bacillus sp. L26 and Bacillus sp. L30). GC-MS analysis showed that strain L30 was the most effective in degrading n-alkane (C10-C27) from diesel after 7 days of cultivation in mineral medium. Both strains produced biosurfactants and showed emulsification activity, specially the strain L30. Alkane hydroxylase gene (group III), which is important for alkane biodegradation, was present in strains. As a result, this study indicated that these bacteria could have promising applications in hydrocarbon bioremediation.

Resumo Vinte e três linhagens bacterianas degradadoras de hidrocarbonetos foram isoladas de água subterrânea contaminada por vazamento em posto de combustível no sul da Amazônia, Brasil. Com base na habilidade de degradar hidrocarbonetos (diesel, hexadecano, benzeno, tolueno e xileno), duas linhagens foram selecionadas para estudos posteriores. A amplificação e sequenciamento do gene 16S rRNA demonstrou que essas linhagens pertencem ao gênero Bacillus (Bacillus sp. L26 and Bacillus sp. L30). Análises de GC-MS mostraram que a linhagem L30 foi mais eficiente em degradar n-alcanos (C10-C27) presentes no diesel, após 7 dias de cultivo em meio mineral. Ambas as linhagens produziram biossurfactantes e apresentaram atividade emulsificante, especialmente a linhagem L30. O gene alcano hidroxilase (grupo III), o qual é importante para degradação de alcanos, foram detectados nas linhagens. Como resultado, este estudo indicou que essas linhagens bacterianas podem ser promissoras se aplicadas em processos de biorremediação.

Composição de ácidos graxos de biossurfactantes produzidos por fungos filamentosos / Composition of fatty acids of biosurfactant produced by filamental fungi

Biossurfactantes são compostos com características tensoativas, produzidos por diversas espécies de microrganismos. Nesse estudo, a composição de ácidos graxos de biossurfactantes produzidos por fungos filamentosos endofíticos foram analisados. Para a produção dos biossurfactantes, empregou-se fontes de carbono (óleo vegetal de milho) e nitrogênio (ureia) de baixo custo no meio de cultivo. A composição de ácidos graxos foi determinada a partir da cromatografia gasosa. Foram identificados seis picos de ésteres metílicos de ácidos graxos, com predominância do ácido linoleico (C18:2n-6c) e ácido oleico (C18:1n-9c). Tanto os ácidos graxos saturados quanto os insaturados foram encontrados na estrutura dos biossurfactantes. Estes resultados demonstraram a importância destes compostos para potenciais aplicações em várias áreas industriais.(AU)

Composição de ácidos graxos de biossurfactantes produzidos por fungos filamentosos / Composition of fatty acids of biosurfactant produced by filamental fungi

Biossurfactantes são compostos com características tensoativas, produzidos por diversas espécies de microrganismos. Nesse estudo, a composição de ácidos graxos de biossurfactantes produzidos por fungos filamentosos endofíticos foram analisados. Para a produção dos biossurfactantes, empregou-se fontes de carbono (óleo vegetal de milho) e nitrogênio (ureia) de baixo custo no meio de cultivo. A composição de ácidos graxos foi determinada a partir da cromatografia gasosa. Foram identificados seis picos de ésteres metílicos de ácidos graxos, com predominância do ácido linoleico (C18:2n-6c) e ácido oleico (C18:1n-9c). Tanto os ácidos graxos saturados quanto os insaturados foram encontrados na estrutura dos biossurfactantes. Estes resultados demonstraram a importância destes compostos para potenciais aplicações em várias áreas industriais.

First report of the production of a potent biosurfactant with ,-trehalose by Fusarium fujikuroi under optimized conditions of submerged fermentation

Abstract Biosurfactants have many advantages over synthetic surfactants but have higher production costs. Identifying microorganisms with high production capacities for these molecules and optimizing their growth conditions can reduce cost. The present work aimed to isolate and identify a fungus with high biosurfactant production capacity, optimize its growth conditions in a low cost culture medium, and characterize the chemical structure of the biosurfactant molecule. The fungal strain UFSM-BAS-01 was isolated from soil contaminated with hydrocarbons and identified as Fusarium fujikuroi. To optimize biosurfactant production, a PlackettBurman design and a central composite rotational design were used. The variables evaluated were pH, incubation period, temperature, agitation and amount of inoculum in a liquid medium containing glucose. The partial structure of the biosurfactant molecule was identified by nuclear magnetic resonance spectrometry. F. fujikuroi reduced surface tension from 72 to 20 mN m1 under the optimized conditions of pH 5.0, 37 °C and 7 days of incubation with 190 rpm agitation. The partial identification of the structure of the biosurfactant demonstrated the presence of an ,-trehalose. The present study is the first report of the biosynthesis of this compound by F. fujikuroi, suggesting that the biosurfactant produced belongs to the class of trehalolipids.

First report of the production of a potent biosurfactant with alfa, beta-trehalose by Fusarium fujikuroi under optimized conditions of submerged fermentation

Biosurfactants have many advantages over synthetic surfactants but have higher production costs. Identifying microorganisms with high production capacities for these molecules and optimizing their growth conditions can reduce cost. The present work aimed to isolate and identify a fungus with high biosurfactant production capacity, optimize its growth conditions in a low cost culture medium, and characterize the chemical structure of the biosurfactant molecule. The fungal strain UFSM-BAS-01 was isolated from soil contaminated with hydrocarbons and identified as Fusarium fujikuroi. To optimize biosurfactant production, a Plackett–Burman design and a central composite rotational design were used. The variables evaluated were pH, incubation period, temperature, agitation and amount of inoculum in a liquid medium containing glucose. The partial structure of the biosurfactant molecule was identified by nuclear magnetic resonance spectrometry. F. fujikuroi reduced surface tension from 72 to 20 mN m−1 under the optimized conditions of pH 5.0, 37 °C and 7 days of incubation with 190 rpm agitation. The partial identification of the structure of the biosurfactant demonstrated the presence of an α,β-trehalose. The present study is the first report of the biosynthesis of this compound by F. fujikuroi, suggesting that the biosurfactant produced belongs to the class of trehalolipids.(AU)

Biosurfactant production by fungi as a sustainable alternative / Produção de biossurfactantes por fungos como uma alternativa sustentável

A wide variety of bacteria is far more exploited than fungi as biosurfactants (BS) or bioemulsifiers (BE), using renewable sources. BS are considered to be environmentally safe and offer advantages over synthetic surfactants. However, the BS yield depends largely on the metabolic pathways of the microorganisms and the nutritional medium. The production of BS or BE uses several cultural conditions, in which a small change in carbon and nitrogen sources affects the quantity of BS or BE produced. The type and quantity of microbial BS or BE produced depend mainly on the producer organism, and factors such as carbon and nitrogen sources, trace elements, temperature and aeration. The diversity of BS or BE makes it interesting to apply them in the pharmaceutical and cosmetics industries, agriculture, public health, food processes, detergents, when treating oily residues, environmental pollution control and bioremediation. Thus, this paper reviews and addresses the biotechnological potential of yeasts and filamentous fungi for producing, characterizing and applying BS or BE.(AU)

Uma grande variedade de espécies bacterianas é bem mais explorada que os fungos como agentes biossurfactantes (BS) ou bioemulsificantes (BE), usando fontes renováveis. Os BS são considerados ecologicamente seguros e oferecem vantagens sobre os surfactantes sintéticos. Entretanto o rendimento de BS depende grandemente das vias metabólicas dos micro-organismos e do meio nutricional. A produção de BS ou BE utiliza várias condições culturais, em que uma pequena alteração nas fontes de carbono e nitrogênio afeta a produção de BS. O tipo e a quantidade de BS ou BE microbianos produzidos dependem principalmente do organismo produtor e de fatores como fontes de carbono e nitrogênio, oligoelementos, temperatura e aeração. A diversidade de BS ou BE torna-os interessantes para aplicação nos campos farmacêutico, cosmético, da agricultura, da saúde pública, em processos alimentares, detergentes, no tratamento de resíduos oleosos, no controle de poluição ambiental e na biorremediação. Assim, a presente revisão aborda o potencial biotecnológico de leveduras e fungos filamentosos para produção, caracterização e aplicações de BS ou BE.(AU)

Biosurfactant production by fungi as a sustainable alternative / Produção de biossurfactantes por fungos como uma alternativa sustentável

A wide variety of bacteria is far more exploited than fungi as biosurfactants (BS) or bioemulsifiers (BE), using renewable sources. BS are considered to be environmentally safe and offer advantages over synthetic surfactants. However, the BS yield depends largely on the metabolic pathways of the microorganisms and the nutritional medium. The production of BS or BE uses several cultural conditions, in which a small change in carbon and nitrogen sources affects the quantity of BS or BE produced. The type and quantity of microbial BS or BE produced depend mainly on the producer organism, and factors such as carbon and nitrogen sources, trace elements, temperature and aeration. The diversity of BS or BE makes it interesting to apply them in the pharmaceutical and cosmetics industries, agriculture, public health, food processes, detergents, when treating oily residues, environmental pollution control and bioremediation. Thus, this paper reviews and addresses the biotechnological potential of yeasts and filamentous fungi for producing, characterizing and applying BS or BE.(AU)

Uma grande variedade de espécies bacterianas é bem mais explorada que os fungos como agentes biossurfactantes (BS) ou bioemulsificantes (BE), usando fontes renováveis. Os BS são considerados ecologicamente seguros e oferecem vantagens sobre os surfactantes sintéticos. Entretanto o rendimento de BS depende grandemente das vias metabólicas dos micro-organismos e do meio nutricional. A produção de BS ou BE utiliza várias condições culturais, em que uma pequena alteração nas fontes de carbono e nitrogênio afeta a produção de BS. O tipo e a quantidade de BS ou BE microbianos produzidos dependem principalmente do organismo produtor e de fatores como fontes de carbono e nitrogênio, oligoelementos, temperatura e aeração. A diversidade de BS ou BE torna-os interessantes para aplicação nos campos farmacêutico, cosmético, da agricultura, da saúde pública, em processos alimentares, detergentes, no tratamento de resíduos oleosos, no controle de poluição ambiental e na biorremediação. Assim, a presente revisão aborda o potencial biotecnológico de leveduras e fungos filamentosos para produção, caracterização e aplicações de BS ou BE.(AU)

Effect of Synthetic Emulsifier and Natural Biosurfactant on Feed Process and Quality of Pelletized Feed in Broiler Diet

A feed production trial was conducted to study the effect of synthetic emulsifier and natural biosurfactant the process and quality of pelletized broiler feed. A corn-soy based broiler diet was formulated with fixed ratio 2:1 of oil-to-water with two types of emulsifiers, namely glyceryl polyethylene glycol ricinoleate synthetic emulsifier and lysophosphatidylcholine natural biosurfactant. T1: Basal diet with no water and no emulsifier; T2: Basal diet with water and no emulsifier; T3: Basal diet with water and synthetic emulsifier glyceryl polyethylene glycol ricinoleate; T4: Basal diet with water and a natural biosurfactant lysophosphatidylcholine as comparative treatment. The treatment diets were manufactured by a commercial feed mill. The electricity cost and meal temperature were measured during the process of milling. Composite samples were collected from different processed points, tested for physical properties, chemical stability and biostability of pelletized feed. Pellet quality of emulsifier supplemented diets was significantly (p 0.05) improved in crumble and pellet intact form. Correlation between emulsifier and pelletize processed cost was not observed in this present study. No deteriorate effect was observed in hydrolytic rancidity (AV), oxidation rancidity (PV), mold count, moisture content and water activity. However, percentage of starch gelatinization on pelletized feed was significantly (p 0.0001) improved in both types of emulsifier treated diets. These results demonstrated that the addition of emulsifier to broiler diet improved pellet quality to some extent although significant difference between synthetic emulsifier and natural biosurfactant was not observed. Abbreviations: AV, acid value; PV, peroxide value; amp, ampere; vol, voltage; T/h, tonne per hour; kW, kilowatt; Aw, water activity; PDI, pellet durability index(AU)

Effect of Synthetic Emulsifier and Natural Biosurfactant on Feed Process and Quality of Pelletized Feed in Broiler Diet

A feed production trial was conducted to study the effect of synthetic emulsifier and natural biosurfactant the process and quality of pelletized broiler feed. A corn-soy based broiler diet was formulated with fixed ratio 2:1 of oil-to-water with two types of emulsifiers, namely glyceryl polyethylene glycol ricinoleate synthetic emulsifier and lysophosphatidylcholine natural biosurfactant. T1: Basal diet with no water and no emulsifier; T2: Basal diet with water and no emulsifier; T3: Basal diet with water and synthetic emulsifier glyceryl polyethylene glycol ricinoleate; T4: Basal diet with water and a natural biosurfactant lysophosphatidylcholine as comparative treatment. The treatment diets were manufactured by a commercial feed mill. The electricity cost and meal temperature were measured during the process of milling. Composite samples were collected from different processed points, tested for physical properties, chemical stability and biostability of pelletized feed. Pellet quality of emulsifier supplemented diets was significantly (p 0.05) improved in crumble and pellet intact form. Correlation between emulsifier and pelletize processed cost was not observed in this present study. No deteriorate effect was observed in hydrolytic rancidity (AV), oxidation rancidity (PV), mold count, moisture content and water activity. However, percentage of starch gelatinization on pelletized feed was significantly (p 0.0001) improved in both types of emulsifier treated diets. These results demonstrated that the addition of emulsifier to broiler diet improved pellet quality to some extent although significant difference between synthetic emulsifier and natural biosurfactant was not observed. Abbreviations: AV, acid value; PV, peroxide value; amp, ampere; vol, voltage; T/h, tonne per hour; kW, kilowatt; Aw, water activity; PDI, pellet durability index

Isolation and characterization of hydrocarbon-degrading bacteria from gas station leaking-contaminated groundwater in the Southern Amazon, Brazil

Abstract Twenty-three hydrocarbon-degrading bacteria strains were isolated from gas station leaking-contaminated groundwater located in the Southern Amazon, Brazil. Based on hydrocarbon (diesel, hexadecane, benzene, toluene and xylene) degradation ability, two strains were selected for further study. The amplification and sequencing of the 16S rRNA gene showed that these two strains belonged to the genus Bacillus (Bacillus sp. L26 and Bacillus sp. L30). GC-MS analysis showed that strain L30 was the most effective in degrading n-alkane (C10-C27) from diesel after 7 days of cultivation in mineral medium. Both strains produced biosurfactants and showed emulsification activity, specially the strain L30. Alkane hydroxylase gene (group III), which is important for alkane biodegradation, was present in strains. As a result, this study indicated that these bacteria could have promising applications in hydrocarbon bioremediation.

Resumo Vinte e três linhagens bacterianas degradadoras de hidrocarbonetos foram isoladas de água subterrânea contaminada por vazamento em posto de combustível no sul da Amazônia, Brasil. Com base na habilidade de degradar hidrocarbonetos (diesel, hexadecano, benzeno, tolueno e xileno), duas linhagens foram selecionadas para estudos posteriores. A amplificação e sequenciamento do gene 16S rRNA demonstrou que essas linhagens pertencem ao gênero Bacillus (Bacillus sp. L26 and Bacillus sp. L30). Análises de GC-MS mostraram que a linhagem L30 foi mais eficiente em degradar n-alcanos (C10-C27) presentes no diesel, após 7 dias de cultivo em meio mineral. Ambas as linhagens produziram biossurfactantes e apresentaram atividade emulsificante, especialmente a linhagem L30. O gene alcano hidroxilase (grupo III), o qual é importante para degradação de alcanos, foram detectados nas linhagens. Como resultado, este estudo indicou que essas linhagens bacterianas podem ser promissoras se aplicadas em processos de biorremediação.

Biosurfactants produced by Microbacterium sp., isolated from aquatic macrophytes in hydrocarbon-contaminated area in the Rio Negro, Manaus, Amazonas / Biossurfactantes produzidos por Microbacterium sp., isolada de macrófita aquática em áreaimpactada por hidrocarbonetos no rio Negro, Manaus, Amazonas

Endophytic bacteria isolated from Eichhornia crassipes (Mart) Solms., collected in oil contaminated wastewater of effluent generated by Petrobras refinery in Manaus were investigated to determine their potential for producing biosurfactants. Assay with 2.6-dichlorophenol indophenol (DCPIP) indicator to verify hydrocarbon biodegradation activity; oil emulsification test; drop-collapse method; surface tension and growth curve of biosurfactant production. The M87 Microbacterium sp. strain chosen for this work was identified by the sequencing of the rDNA region and the chemical characterization was performed by FTIR, UFLC/MS and 1H RMN techniques. The selected bacterial isolate provided 3g L-1 of biosurfactant, using diesel oil as sole carbon source, being efficient in biodegrading oil as demonstrated by the DCPIP test. Fractions obtained by column chromatography were efficient in reducing water surface tension around 40 mN m-1, especially fraction 1, which reduced it to 34.17 mN m-1. The different techniques of chemical analysis used for the identification of the biosurfactant isolate indicated that this is probably a long - chain fatty acid lipid type, which may be used in the future as both biosurfactant in decontamination processes of hydrocarbon-polluted areas or as bioemulsifier in countless processes, since it exhibited no toxicity as determined by Alamar Blue assay. 

Foram investigadas bactérias endofíticas isoladas de Eichhornia crassipes (Mart.) Solms., coletadas em águas contaminadas com resíduos de petróleo em um afluente da refinaria da Petrobrás/Manaus, para valiação da produção de biossurfactantes. Para selecionar o micro-organismos à produção e caracterização de biossurfactantes, foram realizados os seguintes testes: a descoloração do indicador 2,6 indofenol (DCPIP), emulsificação do diesel, colapso da gota, tensão superficial e curva de produção. A caracterização química foi realizada por meio das técnicas de FT-IR, UFLC/MS e RMN1H. A bactéria M87 Microbacterium neste estudo, foi identificada pelo sequenciamento da região rDNA e produziu 3g L-1 de biossurfactantes utilizando o diesel como fonte de carbono, mostrando-se eficiente na ação biodegradadora do petróleo, por meio do teste de Indofenol (DCPIP). As frações obtidas, mostraram-se eficazes na redução da tensão superficial da água abaixo de 40 mN m-1, com destaque para a fração 1 que reduziu a tensão superficial para 34,17 mN m-1. Pelas análises química utilizadas, pode-se inferir que, provavelmente, se trata de um ácido graxo de cadeia longa, que pode ser utilizado futuramente tanto como biossurfactante em processos de descontaminação de ambientes impactados por hidrocarbonetos, assim como bioemulsificante em inúmeros processos uma vez que não apresentou toxicidade por meio do teste realizado.(AU)

Characterization of dioxygenases and biosurfactants produced by crude oil degrading soil bacteria

ABSTRACT Role of microbes in bioremediation of oil spills has become inevitable owing to their eco friendly nature. This study focused on the isolation and characterization of bacterial strains with superior oil degrading potential from crude-oil contaminated soil. Three such bacterial strains were selected and subsequently identified by 16S rRNA gene sequence analysis as Corynebacterium aurimucosum, Acinetobacter baumannii and Microbacterium hydrocarbonoxydans respectively. The specific activity of catechol 1,2 dioxygenase (C12O) and catechol 2,3 dioxygenase (C23O) was determined in these three strains wherein the activity of C12O was more than that of C23O. Among the three strains, Microbacterium hydrocarbonoxydans exhibited superior crude oil degrading ability as evidenced by its superior growth rate in crude oil enriched medium and enhanced activity of dioxygenases. Also degradation of total petroleum hydrocarbon (TPH) in crude oil was higher with Microbacterium hydrocarbonoxydans. The three strains also produced biosurfactants of glycolipid nature as indicated d by biochemical, FTIR and GCMS analysis. These findings emphasize that such bacterial strains with superior oil degrading capacity may find their potential application in bioremediation of oil spills and conservation of marine and soil ecosystem.(AU)

Lipopeptide production by Bacillus subtilis R1 and its possible applications

The possible application of a bacterial strain - Bacillus subtilis R1, isolated from an oil contaminated desert site in India, as biocontrol agent and its biosurfactant in microbial enhanced oil recovery are discussed. The biosurfactant production in minimal medium was carried out at different temperatures and salt concentrations, where it produced an efficient biosurfactant at 30-45 °C and in presence of up to 7% salt. It significantly reduced the surface tension from 66 ± 1.25 mN/m to 29 ± 0.85 mN/m within 24 h. In order to enhance the biosurfactant production, random mutagenesis of B. subtilis R1 was performed using chemical mutagen - ethyl methanesulfonate. Majority of the isolated 42 mutants showed biosurfactant production, but the difference was statistically insignificant as compared with parent strain R1. Therefore none of the mutants were selected for further study, and only parent strain R1 was studied. The biosurfactant was quite stable under harsh conditions for up to 10 days. The biosurfactant was extracted and characterized as similar to the lipopeptide group - surfactins and fengycin. The crude oil displacement experiments using biosurfactant broth in sand pack glass columns showed 33 ± 1.25% additional oil recovery. The strain also showed inhibition of various plant pathogenic fungi on potato dextrose agar medium.(AU)