The Emergence of Different Functionally Equivalent PAH Degrading Microbial Communities from a Single Soil in Liquid PAH Enrichment Cultures and Soil Microcosms Receiving PAHs with and without Bioaugmentation.

Polycyclic aromatic hydrocarbon (PAHs) are common soil contaminants of concern due to their toxicity toward plants, animals and microorganisms. The use of indigenous or added microbes (bioaugmentation) is commonly used for bioremediation of PAHs. In this work, the biodegradation rates and changes in the bacterial community structure were evaluated. The enrichment culture was useful for unambiguously identifying members of the soil bacterial community associated with PAH degradation and yielded a low diversity community. No significant difference in the rate of PAH degradation was observed between the microcosm receiving only PAHs or PAHs and bioaugmentation. Moreover, identical matches to the bioaugmentation inoculum were only observed at the initial stages of PAH degradation on day 8. After 22 days of incubation, the substantial degradation of all PAHs had occurred in both microcosms and the PAH contaminated soil had statistically significant increases in Alphaproteobacteria. There were also increases in Betaproteobacteria. In contrast, the PAH contaminated and bioaugmented soil was not enriched in PAH degrading Proteobacteria genera and, instead, an increase from 1.6% to 8% of the population occurred in the phylum Bacteroidetes class Flavobacteria, with Flavobacterium being the only identified genus. In addition, the newly discovered genus Ohtaekwangia increased from 0% to 3.2% of the total clones. These results indicate that the same soil microbial community can give rise to different PAH degrading consortia that are equally effective in PAH degradation efficiency. Moreover, these results suggest that the lack of efficacy of bioaugmentation in soils can be attributed to a lack of persistence of the introduced microbes, yet nonetheless may alter the microbial community that arises in response to PAH contamination in unexpected ways.

Production and characterization of surface-active compounds from Gordonia amicalis

Two methods were used to make crude preparations of surface-active compounds (SACs) produced by Gordonia amicalis grown on the medium containing 1% diesel oil. Using a 2:1 (v/v) solution of chloroform:methanol for extraction, Type I SACs were isolated and shown to produce oil in water (O/W) emulsions. Type II SACs were isolated by precipitation with ammonium sulfate and produced predominantly water in oil emulsions (W/O). The crude Type I and II preparations were able to produce a significant reduction in the surface tension of water; however, the crude Type II preparation had 10-25 fold higher emulsification activity than the Type I preparation. Both SAC preparations were analyzed by the TLC and each produced two distinct bands with Rf 0.44 and 0.62 and Rf 0.52 and 0.62, respectively. The partially purified SACs were characterized by the ESI(+)-MS, FT-IR and NMR. In each one of these fractions, a mixture of 10 oligomers was found consisting of a series of compounds, with masses from 502 to 899, differing in molecular mass by a repeating unit of 44 Daltons. The mass spectra of these compounds did not appear to match other known biosurfactants and could represent a novel class of these compounds.

Phenol degradation by halophilic bacteria isolated from hypersaline environments.

Phenol is a toxic aromatic compound used or produced in many industries and as a result a common component of industrial wastewaters. Phenol containing waste streams are frequently hypersaline and therefore require halophilic microorganisms for efficient biotreatment without dilution. In this study three halophilic bacteria isolated from different saline environments and identified as Halomonas organivorans, Arhodomonas aquaeolei and Modicisalibacter tunisiensis were shown to be able to grow on phenol in hypersaline media containing 100 g/L of total salts at a concentration of 3 mM (280 mg/L), well above the concentration found in most waste streams. Genes encoding the aromatic dioxygenase enzymes catechol 1,2 dioxygenase and protocatechuate 3,4-dioxygenase were present in all strains as determined by PCR amplification using primers specific for highly conserved regions of the genes. The gene for protocatechuate 3,4-dioxygenase was cloned from the isolated H. organivorans and the translated protein was evaluated by comparative protein sequence analysis with protocatechuate 3,4-dioxygenase proteins from other microorganisms. Although the analysis revealed a wide range of sequence divergence among the protocatechuate 3,4-dioxygenase family, all of the conserved domain amino acid structures identified for this enzyme family are identical or conservatively substituted in the H. organivorans enzyme.

Decolorization and biodegradation of reactive sulfonated azo dyes by a newly isolated Brevibacterium sp. strain VN-15.

Azo dyes constitute the largest and most versatile class of synthetic dyes used in the textile, pharmaceutical, food and cosmetics industries and represent major components in wastewater from these industrial dying processes. Biological decolorization of azo dyes occurs efficiently under low oxygen to anaerobic conditions. However, this process results in the formation of toxic and carcinogenic amines that are resistant to further detoxification under low oxygen conditions. Moreover, the ability to detoxify these amines under aerobic conditions is not a wide spread metabolic activity. In this study we describe the use of Brevibacterium sp. strain VN-15, isolated from an activated sludge process of a textile company, for the sequential decolorization and detoxification of the azo dyes Reactive Yellow 107 (RY107), Reactive Black 5 (RB5), Reactive Red 198 (RR198) and Direct Blue 71 (DB71). Tyrosinase activity was observed during the biotreatment process suggesting the role of this enzyme in the decolorization and degradation process, but no-activity was observed for laccase and peroxidase. Toxicity, measured using Daphnia magna, was completely eliminated.

Purification and structural characterization of fengycin homologues produced by Bacillus subtilis LSFM-05 grown on raw glycerol.

Raw glycerol is a byproduct of biodiesel production that currently has low to negative value for biodiesel producers. One option for increasing the value of raw glycerol is to use it as a feedstock for microbial production. Bacillus subtilis LSFM 05 was used for the production of fengycin in a mineral medium containing raw glycerol as the sole carbon source. Fengycin was isolated by acid precipitation at pH 2 and purified by silica gel column chromatography and characterized using electrospray ionization (ESI) Fourier transform ion cyclotron resonance mass spectrometry (ESI FT-ICR MS) with collision-induced dissociation (CID). The mass spectrum revealed the presence of the ions of m/z 1,435.7, 1,449.9, 1,463.8, 1,477.8, 1,491.8 and 1,505.8, which were further fragmented by ESI-MS/MS. The CID profile showed the presence of a series of ions (m/z 1,080 and 966) and (m/z 1,108 and 994) that represented the different fengycin homologues A and B, respectively. Fengycin homologues A and B are variants that differ at position 6 of the peptide moiety, having either Ala or Val residues, respectively. Mass spectrometry analyses identified four fengycin A and three fengycin B variants with fatty acid components containing 14-17 carbons. These results demonstrate that raw glycerol can be used as feedstock to produce fengycin, and additional work should focus on the optimization of process conditions to increase productivity.

Interaction of curcumin and bixin with ß-cyclodextrin: complexation methods, stability, and applications in food.

This work aimed to compare methods for the formation of complexes of bixin and curcumin with ß-cyclodextrin (ß-CD) and to evaluate the stability of the complexes formed by these methods and their food applications. The stoichiometric relationship between curcumin and ß-CD was 1:2 and that between bixin and ß-CD was 1:1. Curcumin-ß-CD and bixin-ß-CD complexes formed by kneading, coprecipitation, and simple mixing were evaluated by differential scanning calorimetry (DSC), thermogravimetry analysis (TGA), or nuclear magnetic resonance (NMR-H). For both curcumin and bixin, the best method of complexation was coprecipitation. Complexation of colorants with ß-CD promoted an intensification of color and increased water solubility; however, stabilization in the presence of light occurred only for bixin. Application of curcumin-ß-CD in cheese and yogurt and bixin-ß-CD in the curd did not alter the initial characteristics of the products, which were sensorialy well accepted. Therefore, the complexation of these natural colorants with ß-CD favors their use in low-fat foods, broadening the field of industrial application.

Degradation of lignosulfonic and tannic acids by ligninolytic soil fungi cultivated under icroaerobic conditions

Soil fungi were evaluated regarding their ability to degrade lignin-related compounds by producing the ligninolytic enzymes. Lignosulfonic and tannic acids were used as sole carbon sources during 30 days under microaerobic and very-low-oxygen conditions. The fungi produced lignin-peroxidase, manganese-peroxidase and laccase . Expressive degradations was observed by C18 reversed-phase HPLC, indicating the biodegradation potential of these fungi, showing more advantages than obligate anaerobes to decontaminate the environment when present naturally.

Fungos isolados de solo foram avaliados quanto à habilidade em degradarem compostos derivados de lignina pela produção de enzimas ligninolíticas. Os ácidos lignosulfônicos e tânico foram usados separadamente como única fonte de carobono para cultivo dos fungos em 30 dias sob condições microaeróbias. Os fungos foram capazes de crescer e usar tais compostos como fonte de carbono e mostraram produção de lignina-peroxidase, manganês-peroxidase e lacase. Degradações expressivas dos ácidos lignosulfônico e tânico foram verificadas por Cromatografia Liquida de Alta Eficiência (CLAE), indicando grande potencial de uso em processos de biorremediação de macromoléculas aromáticas similares à lignina em ambientes naturais sob condições baixas de oxigenação.

Application of molecular fingerprinting for analysis of a PAH-contaminated soil microbiota growing in the presence of complex PAHs / Aplicação de técnica molecular para análise da microbiota de solo contaminado em misturas complexas de HPAs

Polycyclic aromatic hydrocarbons (PAHs) constitute a group of priority pollutants which are present at high concentrations in the soils of many industrial contaminated sites. Pollution by these compounds may stimulate growth of organisms able to live in these environments causing changes in the structure of the microbial community due to some cooperative process of metabolization of toxic compounds. A long-term PAH-contaminated soil was stored for several years and used to analyze the native microbiota regarding their ability to grow on pyrene, benzo[a]pyrene, as well as in mixtures of LMW- and HMW-PAHs. Molecular profiles of the microbial community was assessed by PCR-DGGE of 16S rRNA gene, and the number of bands observed in DGGE analyses was interpreted as dominant microbial members into the bacterial community. Results of PAH-contaminated soil microorganisms showed different profiles in the degradative dynamics when some nutrients were added. Predominant species may play a significative role while growing and surviving on PAHs, and some other metabolically active species have emerged to interact themselves in a cooperative catabolism of PAHs.

Os hidrocarbonetos poliaromáticos (HPAs) são considerados poluentes prioritários presentes em expressiva concentração no solo contaminado com derivados de petróleo. A poluição por esses compostos estimula o crescimento de microrganismos capazes de sobreviverem nestes ambientes contaminados, causando alterações na estrutura da comunidade microbiana do solo pelo processo de cooperação metabólica entre as populações. Um solo contaminado por um longo período de tempo foi coletado de uma área industrial (Port Melbourne, Austrália) e utilizado para análise da capacidade da comunidade microbiana em crescer em HPAs isolados e/ou em misturas como únicas fontes de carbono e energia. Os perfis moleculares foram obtidos por PCR-DGGE do fragmento da subunidade 16S do DNA ribossomal, sendo o número de bandas presentes nos géis de eletroforese interpretado como populações dominantes dentro da comunidade do solo. Resultados demonstraram diferentes perfis quando vitaminas e micronutrientes fizeram parte do meio líquido de crescimento da microbiota, além da presença dos HPAs. As espécies predominantes podem ter papel significativo na degradação desses compostos no solo a ser remediado, enquanto sobrevivem e interagem com outras populações por meio de mecanismos de catabolismo cooperativo.

Purification and partial characterization of manganese peroxidase from Bacillus pumilus AND Paenibacillus sp

The production of manganese peroxidase (MnP) from Bacillus pumilus and Paenibacillus sp. was studied under absence and presence of the inducers indulin AT, guayacol, veratryl alcohol, lignosulfonic acid and lignosulfonic acid desulfonated. Indulin AT increased the activity of B. pumilus MnP up to 31.66 U/L after 8 h, but no improve was observed for Paenibacillus sp., which reached maximum activity (12.22 U/L) after 20 h. Both MnPs produced by these microorganisms were purified in phenyl sepharose resin and the proteins from crude extracts were eluted in two fractions. However, only the first fraction of each extract exhibited MnP activities. Tests in different pH and temperature values, from pH 5.0 to pH 10.0 and 30 ºC to 60 ºC, respectively, were carried out with the purified MnP. The maximum activity reached for B. pumilus and Paenibacillus sp. MnPs were 4.3 U/L at pH 8.0 and 25 ºC and 11.74 U/L at pH 9.0 and 35 ºC, respectively. The molar masses determined by SDS-PAGE gel eletrophoresis were 25 kDa and 40 kDa, respectively, for the purified enzyme from B. pumilus and Paenibacillus sp.

Reducing COD level on oily effluent by utilizing biosurfactant-producing bacteria

Two bacteria isolated from crude oil contaminated soil, Pantoea agglomerans and Planococcus citreus, produced biosurfactants utilizing 1.5 percent of kerosene and olive oil as the sole carbon sources, respectively. The bacteria and the biosurfactants produced were introduced to oily effluent, arising from margarine and soap industry. Emulsification activities were determined by increases in the absorbance of the oil-in-water emulsions at 610 nm, whereas the water-in-oil emulsions were expressed as the height (cm) of the emulsion layers formed. The 72 h incubation experiment resulted in a COD (Chemical Oxygen Demand) reduction of 76 percent with Planococcus citreus strain and 70 percent with Pantoea agglomerans.The COD reduction with bacterial biosurfactants was over 50 percent in 24 h of incubation. The COD reduction showed that these strains and the surfactants produced could be used in bioremediation processes.

Duas bactérias isoladas de solo contaminado com derivados de petróleo, Pantoea agglomerans e Planococcus citreus, produzem biosurfactantes utilizando respectivamente 1.5 por cento de querosene e óleo de oliva como únicas fontes de carbono. As bactérias e os biosurfactantes produzidos foram adicionados a um efluente oleoso obtido de uma indústria nacional de sabão e margarina. As atividades de emulsificação foram determinadas pelo aumento da absorbância das emulsões óleo em água a 610 nm, enquanto que as emulsões do tipo água em óleo foram expressas em centímetros, pela altura do halo de espumas formado. A redução da demanda química de oxigênio (COD) mostra que as linhagens e os biosurfactantes produzidos podem ser utilizados em processos de biorremediação.

Use of Bacillus pumilus CBMAI 0008 and Paenibacillus sp. CBMAI 868 for colour removal from paper mill effluent / Emprego de Bacillus pumilus CBMAI 0008 e Paenibacillus sp. CBMAI 868 para remoção da cor do efluente da indústria papeleira

Bacillus pumilus and Paenibacillus sp. were applied on the paper mill effluent to investigate the colour remotion. Inocula were individually applied in effluent at pH 7.0, 9.0 and 11.0. The real colour and COD remotion after 48h at pH 9.0 were, respectively, 41.87% and 22.08% for B. pumilus treatment and 42.30% and 22.89% for Paenibacillus sp. Gel permeation chromatography was used to verify the molar masses of compounds in the non-treated and treated effluent, showing a decrease in the compounds responsible for the paper mill effluent colour.

Bacillus pumilus e Paenibacillus sp. foram aplicados separadamente no efluente da indústria papeleira a pH 7,0, 9,0 e 11,0, para verificação da remoção da cor e da DQO. As remoções da cor real e DQO após 48h a pH 9,0 foram, respectivamente, de 41,87% e 22,08% após o tratamento com B. pumilus e 42,30% e 22,89% após tratamento com Paenibacillus sp. As massas molares dos compostos presentes no efluente não tratado e tratado foram determinadas por cromatografia de permeação em gel. O emprego dos microrganismos reduziu os compostos responsáveis pela cor do efluente da indústria papeleira.

Bioremediation of a polyaromatic hydrocarbon contaminated soil by native soil microbiota and bioaugmentation with isolated microbial consortia.

Biodegradation of a mixture of PAHs was assessed in forest soil microcosms performed either without or with bioaugmentation using individual fungi and bacterial and a fungal consortia. Respiratory activity, metabolic intermediates and extent of PAH degradation were determined. In all microcosms the low molecular weight PAH's naphthalene, phenanthrene and anthracene, showed a rapid initial rate of removal. However, bioaugmentation did not significantly affect the biodegradation efficiency for these compounds. Significantly slower degradation rates were demonstrated for the high molecular weight PAH's pyrene, benz[a]anthracene and benz[a]pyrene. Bioaugmentation did not improve the rate or extent of PAH degradation, except in the case of Aspergillus sp. Respiratory activity was determined by CO(2) evolution and correlated roughly with the rate and timing of PAH removal. This indicated that the PAHs were being used as an energy source. The native microbiota responded rapidly to the addition of the PAHs and demonstrated the ability to degrade all of the PAHs added to the soil, indicating their ability to remediate PAH-contaminated soils.

Use of Bacillus pumilus CBMAI 0008 and Paenibacillus sp. CBMAI 868 for colour removal from paper mill effluent.

Bacillus pumilus and Paenibacillus sp. were applied on the paper mill effluent to investigate the colour remotion. Inocula were individually applied in effluent at pH 7.0, 9.0 and 11.0. The real colour and COD remotion after 48h at pH 9.0 were, respectively, 41.87% and 22.08% for B. pumilus treatment and 42.30% and 22.89% for Paenibacillus sp. Gel permeation chromatography was used to verify the molar masses of compounds in the non-treated and treated effluent, showing a decrease in the compounds responsible for the paper mill effluent colour.

Purification and Partial characterization of manganese peroxidase from Bacillus pumilus AND Paenibacillus sp.

The production of manganese peroxidase (MnP) from Bacillus pumilus and Paenibacillus sp. was studied under absence and presence of the inducers indulin AT, guayacol, veratryl alcohol, lignosulfonic acid and lignosulfonic acid desulfonated. Indulin AT increased the activity of B. pumilus MnP up to 31.66 U/L after 8 h, but no improve was observed for Paenibacillus sp., which reached maximum activity (12.22 U/L) after 20 h. Both MnPs produced by these microorganisms were purified in phenyl sepharose resin and the proteins from crude extracts were eluted in two fractions. However, only the first fraction of each extract exhibited MnP activities. Tests in different pH and temperature values, from pH 5.0 to pH 10.0 and 30 °C to 60 °C, respectively, were carried out with the purified MnP. The maximum activity reached for B. pumilus and Paenibacillus sp. MnPs were 4.3 U/L at pH 8.0 and 25 °C and 11.74 U/L at pH 9.0 and 35 °C, respectively. The molar masses determined by SDS-PAGE gel eletrophoresis were 25 kDa and 40 kDa, respectively, for the purified enzyme from B. pumilus and Paenibacillus sp.

Xilooligossacarídeos: produção, aplicações e efeitos na saúde humana / Xylooligosaccharides: production, applications and effects on human health

Os xilooligossacarídeos são açúcares não-convencionais (oligômeros formados por unidades de xilose), não-calóricos e não são metabolizados pelo organismo humano. São considerados prebióticos, uma vez que promovem seletivamente o crescimento de probióticos como Lactobacillus sp. e Bifidobacterium bifidum, promovendo uma série de benefícios à saúde humana, como a redução da constipação intestinal, a promoção da digestão e a absorção de nutrientes, a prevenção de infecções gastrintestinais e a inibição do crescimento de microrganismos patogênicos. Este artigo de revisão demonstra a produção de xilooligossacarídeos, tendo como fonte materiais lignocelulósicos, através de métodos químicos e enzimáticos, e também a sua aplicação como ingredientes em alimentos, salientando os efeitos benéficos à saúde provindos desses tipos de compostos.

Xylooligosaccharides are non-conventional sugars (sugar oligomers made of xylose units), non-calorics and non-digestible by humans. They are recognyzed as prebiotics once there are non digestible food ingredient that stimulate selectively probiotic growth like Lactobacillus sp. and Bifidobacterium bifidum, promoting several benefices to the human health as the reduction of the gut constipation, the increase of the digestion and nutrient absorption, prevention of gastrointestinal infections, and the inhibition of pathogenic microorganisms. This article reviews the production of xylooligosaccharides from lignocellulosic materials (by chemical or enzymatic methods) and its end products, as well as their application as food ingredients, with special attention to the beneficial effects caused on health by these types of compounds.