The influence of Tween 80 on laccase production by Pleurotus sajor-caju and the efficiency of crude enzyme broth in the removal of bisphenol-A / Influência do Tween 80 na produção de lacases de Pleurotus sajor-caju e eficiência do caldo enzimático bruto na remoção de bisfenol-A

Bisphenol-A is currently considered an environmental pollutant, capable of interfering in the endocrine system of organisms and causing alterations in its development and reproductive system. An alternative method to the chemical treatment of this pollutant has been the use of oxidative enzymes, especially laccases produced by fungi. In order to reduce production costs, agro-industrial waste can be used in the culture medium composition. Nonionic surfactants, which are only slightly toxic to biological membranes, can be applied, as well as Tween 80, to facilitate the excretion of these enzymes into the culture medium. The objectives of this work were: a) characterize the immersion water of banana straw used in the formulation of the culture medium; b) evaluate laccase production by Pleurotus sajor-caju in culture medium with and without addition of Tween 80, through shaken flasks; c) evaluate the efficiency of the crude enzyme broth in degrading bisphenol-A. The shaken flasks were incubated at 30°C for 12 days. The immersion water had a C:N ratio of 13.8, ash percentage of 28.6%, and pH close to neutrality. The addition of Tween 80 on the culture medium (7.5%, m/v) yielded laccase activity and productivity values equal to 3,016.47 U L-1 and 502.7 U L-1 day-1, respectively. These values were 50 and 33.5 times higher than those obtained in the culture medium without addition of Tween 80 for laccase activity and productivity, respectively. The crude enzyme broth degraded 100% of bisphenol-A after 48 hours, regardless of concentration (500, 750 and 1,000 mg L-1).(AU)

O bisfenol-A é considerado um poluente ambiental capaz de interferir no sistema endócrino dos organismos, ocasionando alterações em seu desenvolvimento e sistema de reprodução. Um método alternativo ao tratamento químico desse tipo de poluente tem sido a utilização de enzimas oxidativas, especialmente as lacases, produzidas por fungos. A fim de diminuir custos de produção, resíduos agroindustriais podem compor o meio de cultivo. Assim, surfactantes não iônicos e pouco tóxicos para as membranas biológicas, como o Tween 80, podem ser utilizados para facilitar a excreção dessas enzimas para o meio de cultivo. Os objetivos deste trabalho foram: caracterizar quimicamente o resíduo água de imersão de palha de bananeira, usado na formulação do meio de cultivo; avaliar a produção de lacase por Pleurotus sajor-caju em meio de cultivo líquido (frascos Erlenmeyer) com e sem adição de Tween 80; e avaliar a eficiência do caldo enzimático bruto em degradar bisfenol-A. Os frascos foram incubados a 30°C, por 12 dias. A água de imersão apresentou relação C:N 13,8, percentual de cinzas 28,6% e pH próximo da neutralidade. O cultivo adicionado de Tween 80 (7,5%, m/v) propiciou valores de atividade e produtividade em lacase iguais a 3.016,47 U L-1 e 502,7 U L-1 dia-1, respectivamente. Esses valores são 50 e 33,5 vezes maiores que os obtidos no cultivo sem adição de Tween 80, para atividade e produtividade em lacase, respectivamente. O caldo enzimático bruto degradou 100% do bisfenol-A após 48 horas, independentemente da concentração (500, 750 e 1.000 mg L-1).(AU)

Maintenance culture medium and inoculum based on peach palm leaves for Pleurotus spp. production / Meio de manutenção e inóculo a partir de folhas de pupunheira para a produção de Pleurotus spp

This study evaluated the use of immersion water from peach palm leaves (PPLDA) as a component of the culture medium for the maintenance of Pleurotus spp. and the use of agricultural waste, peach palm leaves, as inoculum support for the fungi. The performance of the inoculum based on peach palm leaves (PPL) for the production of Pleurotus spp. fruiting bodies was compared with that using wheat grains (WG) as inoculum support. PPLDA culture medium (immersion water of peach palm leaves, dextrose, and agar) showed a lower radial velocity of mycelial growth for both fungi than that obtained with the culture medium WDA (wheat extract, dextrose and agar), commonly used as maintenance medium for Pleurotus spp. However, the type of inoculum support does not significantly influence the linear velocity of P. ostreatus mycelial growth, reaching 6.71 mm/day on wheat grains and 6.18 mm/day on peach palm leaves. Thus, when the inoculum based on peach palm leaves is utilized, the immersion water used for preparing this support can be used for preparing the PPLDA maintenance culture medium, diminishing the production costs of Pleurotus mushrooms. Data also showed that when Pleurotus sajor-caju was cultivated on peach palm leaves, using PPL as inoculum support, the fruiting bodies production parameters (Y = 47%, BE = 3% and Pr = 0.2 g/day) did not differ from that obtained using WG.(AU)

Este trabalho avaliou a utilização da água de imersão das folhas de pupunheira como componente do meio de cultivo para manutenção da cepa fúngica e a produção de inóculo de Pleurotus spp. utilizando como suporte folhas de pupunheira, um resíduo agrícola. Ainda, para verificar a capacidade do inóculo à base de folhas de pupunheira (FP) de produzir corpos frutíferos, foi realizado um experimento comparando FP com o inóculo à base de grãos de trigo (GT). Para tanto, realizou-se a comparação das velocidades de crescimento radial entre o meio de cultivo TDA (extrato de trigo, dextrose e ágar) utilizado para a manutenção de fungos do gênero Pleurotus e o meio FPDA (água de imersão de folhas de pupunheira, dextrose e ágar), e a comparação das velocidades de crescimento linear entre os grãos de trigo, utilizados como suporte para o inóculo de fungos do gênero Pleurotus e as folhas de pupunheira. O meio de cultivo TDA apresentou velocidade radial superior ao meio FPDA para as duas espécies, no entanto, o tipo de suporte não influenciou significativamente a velocidade de crescimento linear de P. ostreatus , que apresentou 6,71 mm/dia em grãos de trigo e 6,18 mm/dia em folhas de pupunheira. Assim, quando o inóculo à base de folhas de pupunheira é utilizado, a água de imersão deste suporte pode ser aproveitada para a confecção do meio de cultivo de manutenção (FPDA), diminuindo os custos produção. Observou-se que Pleurotus sajor-caju , quando cultivado em folhas de pupunheira utilizando inóculo FP, apresentou cerca de 47% de rendimento, 3% de eficiência biológica e 0,2 g/dia de produtividade, sem diferença significativa com o inóculo GT.(AU)

Antitumor activity of Pleurotus ostreatus polysaccharide fractions on Ehrlich tumor and Sarcoma 180.

The medicinal properties of fungi of the genus Pleurotus have attracted great interest due to their therapeutic properties. Polysaccharides synthesized by Pleurotus, including the ß-glucans are considered the main responsible for its therapeutic properties. This study aimed to evaluate the efficacy of polysaccharidic fractions extracted from mycelial biomass of Pleurotus ostreatus DSM 1833 in inhibiting the development of Ehrlich Tumor (ET) and Sarcoma 180 (S-180). FC, FI and FII fractions provided 60.6, 76.5 and 73.6% of ET inhibition, respectively (mean value of about 70%) while FS, FIII-1 and FIII-2 showed no inhibition against ET. FII and FIII-2 resulted in 85.6 and 93.6% (mean value of about 90%) while FIII-1, FC and FS resulted in 54.1 and 0%, respectively, of S-180 inhibition. The yields of the fractions FS, FI, FII, FIII-1 and FIII-2 obtained from P. ostreatus mycelial biomass were 11.6, 1.3, 0.4, 0.65 and 0.35%, respectively. FII fraction (30mg/kg) apparently had no toxic effect on healthy animals, since no difference between the body weights of animals in substance control (SC) and negative control (NC) groups was observed.

Extracellular polysaccharide production by a strain of Pleurotus djamor isolated in the south of Brazil and antitumor activity on Sarcoma 180

Polysaccharides with medicinal properties can be obtained from fruiting bodies, mycelium and culture broth of several fungus species. This work was carried out in batch culture using a stirred tank reactor with two different initial glucose concentrations (40-50 g/L) and pH values (3.0-4.0) to enhance extracellular polysaccharides production by Pleurotus djamor UNIVILLE 001 and evaluate antitumor effect of intraperitonial administration of Pleurotus djamor extract on sarcoma 180 animal model. According to factorial design, the low pH value (pH 3.0) led to a gain of 1.6 g/L on the extracellular polysaccharide concentration, while glucose concentration in the tested range had no significant effect on the concentration of polysaccharide. With 40 g/L initial glucose concentration and pH 3.0, it was observed that yield factor of extracellular polysaccharide on substrate (Y P/S = 0.072) and maximum extracellular polysaccharide productivity (Q Pmax = 11.26 mg/L.h) were about 188% and 321% respectively higher than those obtained in the experiment performed at pH 4.0. Under these conditions, the highest values of the yield factor of biomass on substrate (Y X/S = 0.24) and maximal biomass productivity (Q Xmax = 32.2 mg/L.h) were also reached. In tumor response study, mean tumor volume on the 21th day was 35.3 cm³ in untreated group and 1.6 cm³ in treated group (p = 0.05) with a tumor inhibition rate of 94%. These impressive results suggests an inhibitory effect of P.djamor extract on cancer cells.

Production of bioactive compounds with antitumor activity against sarcoma 180 by Pleurotus sajor-caju.

This work studied the influence of culture medium composition and pH on exopolysaccharide (EPS) production by Pleurotus sajor-caju and validates the antitumor activity of the produced EPSs and of the mycelial biomass (intracellular polysaccharides [IPSs]) against Sarcoma 180 (S180) cells. The effect of the initial concentrations of (NH4)2SO4, yeast extract and soy peptone on EPS production by P. sajor-caju was studied in shake flasks. A bioreactor was used to evaluate the pH values and the initial CaCO3 and glucose concentrations. Extracts of EPSs (PE1) and IPSs obtained through two different separation processes (PM1 and PM2) were tested on mice inoculated with S180 cells. A medium containing 2.5, 1.0, and 1.0 g/L of (NH4)2SO4, yeast extract and soy peptone, respectively, provided the highest EPS concentration (0.6 g/L). The use of pH 4.0, 1.0 g/L CaCO3 and 20 g/L initial glucose concentration enhanced EPS productivity (3.84 g/L per hour). The PE1 extract promoted the highest reduction of S180 growth (86%), followed by the PM2 extract (80%); growth reduction was dose-independent for both substances. This work provides information about culture medium and conditions that enhanced the production of extracellular polysaccharides by P. sajor-caju. The results can contribute to the search for new bioactive products bringing novel aspects to the medical and pharmaceutical areas.

Extracellular polysaccharide production by a strain of Pleurotus djamor isolated in the south of Brazil and antitumor activity on Sarcoma 180.

Polysaccharides with medicinal properties can be obtained from fruiting bodies, mycelium and culture broth of several fungus species. This work was carried out in batch culture using a stirred tank reactor with two different initial glucose concentrations (40-50 g/L) and pH values (3.0-4.0) to enhance extracellular polysaccharides production by Pleurotus djamor UNIVILLE 001 and evaluate antitumor effect of intraperitonial administration of Pleurotus djamor extract on sarcoma 180 animal model. According to factorial design, the low pH value (pH 3.0) led to a gain of 1.6 g/L on the extracellular polysaccharide concentration, while glucose concentration in the tested range had no significant effect on the concentration of polysaccharide. With 40 g/L initial glucose concentration and pH 3.0, it was observed that yield factor of extracellular polysaccharide on substrate (YP/S = 0.072) and maximum extracellular polysaccharide productivity (Q(Pmax) = 11.26 mg/L.h) were about 188% and 321% respectively higher than those obtained in the experiment performed at pH 4.0. Under these conditions, the highest values of the yield factor of biomass on substrate (YX/S = 0.24) and maximal biomass productivity (Q(Xmax) = 32.2 mg/L.h) were also reached. In tumor response study, mean tumor volume on the 21th day was 35.3 cm(3) in untreated group and 1.6 cm(3) in treated group (p = 0.05) with a tumor inhibition rate of 94%. These impressive results suggests an inhibitory effect of P.djamor extract on cancer cells.

Laccase production by the aquatic ascomycete Phoma sp. UHH 5-1-03 and the white rot basidiomycete Pleurotus ostreatus DSM 1833 during submerged cultivation on banana peels and enzyme applicability for the removal of endocrine-disrupting chemicals.

This work aimed to study the production of laccase from Pleurotus ostreatus DSM 1833 and Phoma sp. UHH 5-1-03 using banana peels as alternative carbon source, the subsequent partial purification and characterization of the enzyme, as well the applicability to degrade endocrine disruptors. The laccase stability with pH and temperature, the optimum pH, the K (m) and V(max) parameters, and the molar mass were determined. Tests were conducted for assessing the ability of degradation of the endocrine disruptors t-nonylphenol, bisphenol A, and 17α-ethinylestradiol. Laccase production of 752 and 1,117 U L⁻¹ was obtained for Phoma sp. and P. ostreatus, respectively. Phoma sp. laccase showed higher stability with temperature and pH. The laccase from both species showed higher affinity by syringaldazine. The culture broth with banana peels induced the production of two isoforms of P. ostreatus (58.7 and 21 kDa) and one of Phoma sp. laccase (72 kDa). In the first day of incubation, the concentrations of bisphenol A and 17α-ethinylestradiol were reduced to values close to zero and after 3 days the concentration of t-nonylphenol was reduced in 90% by the P. ostreatus laccase, but there was no reduction in its concentration by the Phoma sp. laccase.

Production of bio-inseticide Bacillus thuringiensis var. israelensis in semicontinuous processes combined with batch processes for sporulation

The aim of this work was to study the production of the bioinsecticide using the aerobic cultivation. Two-stage cultivation process was used: aerobic vegetative growth and end-phase of sporulation (with aeration and without aeration). The cuts of working volume evaluated were 20, 40, 60 and 80 percent. The highest value of spore concentration (2.17x10(9) CFU/mL) and spore productivity (7.2x10(7) CFU/mL.h) were observed using SP60. The lethal concentration (CL50) obtained through the aeration process and non-aeration process were 0.21 and 0.05 mg/L, respectively. The specific potential of the bioinsecticide produced without aeration (0.025 mg/10-12 CFU) was two times higher than that obtained with aeration (0.05 mg/10-12 CFU).

Cultivation of Agaricus blazei on Pleurotus spp. spent substrate

The aim of this work was the use of Pleurotus ostreatus and Pleurotus sajor-caju for the previous lignocellulolytic decomposition of banana tree leaf straw and the further use of the degraded straw as substrate for the culture of Agaricus blazei. For optimising the production of A. blazei in terms of yield (Y percent) and biological efficiency (BE percent), adjustments to the composition of the substrate were evaluated in a 2(5) experimental design. The following components were tested in relation to percent of substrate dry mass: urea (1 and 10 percent), rice bran (10 or 20 percent) or ammonium sulphate (0 or 10 percent), inoculum (10 or 20 percent) and the casing material (subsoil or burned rice husks). The best results (79.71 Y percent and 6.73 BE percent) were found when the substrate containing 10 percent of rice bran, without ammonium sulphate, inoculated with 20 percent and covered with subsoil was used.

O cultivo de fungos comestíveis e medicinais utilizando resíduos da agroindústria vem se apresentando como uma alternativa econômica para o pequeno produtor rural, favorecendo a agricultura familiar do nordeste catarinense. Este trabalho avaliou o fungo Pleurotus para a decomposição lignocelulolítica de palha de folhas de bananeira e a utilização da palha residual como substrato para o cultivo de Agaricus blazei. Ajustes na composição do substrato residual de Pleurotus, tais como o tipo e a concentração da fonte de nitrogênio, a porcentagem de inóculo e a camada de cobertura, foram avaliadas. O substrato residual que mais favoreceu a produção de A. blazei em Eficiência Biológica (6,73 por cento), Rendimento (79,71 por cento) e menor tempo para emissão do primeiro primórdio (27 dias) foi o substrato residual de P. ostreatus inoculado com 20 por cento de inóculo (ms), 10 por cento de farelo de arroz (ms), sem sulfato de amônio e utilizando terra de subsolo como camada de cobertura.

Drying and rehydration of oyster mushroom

Dehydration and rehydration processes of Pleurotus ostreatus fruiting bodies were investigated in this work. Mushroom samples were dehydrated at 40, 50 and 60 ºC, using drying air with relative humidity of 75 percent. The rehydration was investigated at different temperatures of immersion water (25, 55 and 85 ºC) and different immersion times (30, 75 and 120 minutes). The best rehydration occurred for the samples dried at 40 ºC. The rehydration could be done in water at room temperature, during 30 minutes. Water sorption isotherms of samples were determined at 30, 40 and 50 ºC. Both GAB and BET models satisfactorily represented the experimental data of moisture sorption of dried mushrooms.

Processos de desidratação e de rehidratação de cogumelos da espécie Pleurotus ostreatus foram avaliados neste trabalho. Os cogumelos foram desidratados a 40, 50 e 60 ºC, com umidade relativa do ar de 75 por cento. O processo de rehidratação foi avaliado para diferentes temperaturas de água de imersão (25, 55 e 85 ºC) e diferentes tempos de imersão (30, 75 e 120 minutos). A melhor temperatura de secagem foi 40 ºC, levando em consideração a melhor rehidratação dos cogumelos desidratados nesta temperatura. A rehidratação pode ser feita em água a temperatura ambiente, por 30 minutos. Isotermas de sorção de umidade de amostras foram determinadas a 30, 40 e 50 ºC.Tanto o modelo de GAB quanto o de BET representaram satisfatoriamente os dados experimentais de isoterma de sorção de umidade.

Characterization and antineoplasic effect of extracts obtained from Pleurotus sajor-caju fruiting bodies.

Fungi of the Pleurotus genus present a great industrial interest due to their possibility of producing pharmacological compounds, pigments, aromas, organic acids, polysaccharides, enzymes, vitamins, amino acids, etc. Among the therapeutic products, we can highlight those with antineoplasic activity, attributed to the fungi cell wall components. Based on this, the objective of this work was to study the antineoplasic capacity of the polysaccharidic fractions obtained from Pleurotus sajor-caju fruiting bodies. Female Swiss mice were inoculated with the Ehrlich ascitic tumor (5 x 10(6) cells/animal) in ascitic form. The polysaccharidic fractions were administered intraperitoneally, during a 6-day period. Fractions FI and FII presented a lower volume of ascitic liquid (3.1 and 1.8 mL, respectively) and a higher reduction in the number of neoplasic cells present in the ascitic liquid (86.2% and 85%, respectively), when compared to the positive control (group inoculated with the tumor but without treatment). These fractions were characterized in terms of monosaccharide composition. Glucose was the major component detected, followed by galactose and mannose. The anomeric carbon configuration of the beta-glucan was confirmed by the (13)C NMR (delta 103.7). Substituted and free C3 and C6 were also detected. Protein bands were confirmed through infrared analysis.

Biodegradation of 2, 4 dichlorophenol by Pleurotus ostreatus DSM 1833

This work aimed to investigate the capacity of Pleurotus ostreatus DSM 1833 to degrade 2,4-dichlorophenol, important pollutant found in the wastewaters of the paper and cellulose industry. Using a factorial design 2², the concentrations of glucose and 2,4-dichlorophenol varied between 0 and 10g.L-1 and 5 and 30mg.L-1, respectively. The best global biodegradation rate was obtained using 30 mg.L-1 of 2,4- dichlorophenol in the absence of glucose. This culture medium was used for scaling up the process, resulting in a global biodegradation rate of 0.47mg.L-1.h-1. A comparative test between an inoculated medium and an abiotic control demonstrated that 54.1 percent of 2,4- dichlorophenol degradation could be attributed to the presence of P. ostreatus.

A indústria de papel e celulose contribui para a contaminação ambiental devido aos resíduos gerados, especialmente, no processo de branqueamento da polpa Kraft, realizada com cloro. Basideomicetos saprófitas têm a capacidade de degradar compostos organoclorados como cloroligninas e clorofenóis. Este trabalho teve como objetivo investigar a capacidade de Pleurotus ostreatus DSM 1833 em degradar 2,4-diclorofenol, importante poluente encontrado nos efluentes da indústria de papel e celulose. Utilizando um planejamento fatorial 2², as concentrações de glicose e de 2,4-diclorofenol variaram entre 0 e 10 g.L-1 e 5 e 30 mg.L-1, respectivamente. A melhor taxa global de degradação foi obtida usando-se 30 mg.L-1 de 2,4-diclorofenol na ausência de glicose. Este meio de cultura foi utilizado para a ampliação da escala do processo, resultando em uma taxa global de biodegradação de 0,47 mg.L-1.h-1. Um teste comparativo entre o meio inoculado e o controle abiótico demonstrou que 54,1 por cento da degradação do 2,4-diclorofenol pode ser atribuída à presença de Pleurotus ostreatus.

Alternative medium for production of Pleurotus ostreatus biomass and potential antitumor polysaccharides.

Pleurotus species are recognized for producing beta-glucans with important medicinal properties as a constituent of the cellular wall of the fruiting body or of the mycelium. The aims of this work were to select a culture medium that maximized the production of biomass and polysaccharides produced by Pleurotus ostreatus DSM 1833 and to evaluate the selected medium in two values of initial oxygen transfer rate -K(L)a (10.2 and 19.3 h(-1)). A 2* *4 factorial design was constructed to evaluate the supplementation of wheat extract with corn steep liquor--CSL (10 or 20 g L(-1)), yeast extract--YE (2 or 5gL(-1)), ammonium sulfate--AS (0 or 5 g L(-1)) and glucose (20 or 40 g L(-1)). In terms of maximum productivity in biomass and global productivity in polysaccharides, the best values were obtained when 5 g L(-1) of YE and 40 g L(-1) of glucose were used. In terms of maximum concentration of biomass, the best results were obtained when 20 g L(-1) of CSL and 40 g L(-1) of glucose were used. The best results in terms of production of biomass and polysaccharides were achieved when lower initial K(L)a (10.2 h(-1)) was used.