Alkaline Cellulase Production by Penicillium mallochii LMB-HP37 Isolated from Soils of a Peruvian Rainforest.

Alkaline cellulases are demanded by the textile industry for several purposes but commercial preparations showing activity at alkaline conditions are very scarce. Aim: To characterize a Penicillium strain isolated form soils of a Peruvian rainforest showing alkaline cellulase activity that may be useful for the textile industry. Methodology: The molecular identification was based on the DNA sequence of its ITS region using ITS1 and ITS4 primers after PCR amplification. Cellulase production was evaluated in shaken flasks by using either lactose or microcrystalline cellulose. Total cellulase (as FPA) and endoglucanase activities were evaluated by the standard methods at several pH levels. Also, the cellulase activity of culture filtrates was tested for antipilling activity as compared to a commercial neutral cellulase preparation. Results: After raw data of ITS DNA sequence was processed, multiple alignment and phylogenetic analysis confirmed that our strain can be named as Penicillium mallochii LMB-HP37. Higher activity was attained for neutral total cellulase on lactose (3371±108 U/l at pH 7.4) and alkaline cellulases attained similar activity levels than the acid cellulase (2978±151 U/l at pH 8.4 and 2910±42 U/l at pH 9.4). FPA and endoglucanase activities were produced at high volumetric (46.8±1.5 and 13.5±1.0 U/l.h, respectively) and specific (32.9±1.1 and 9.5±0.7 U/gbiomass.h, respectively) productivities at the same pHs which indicate that this strain may be suitable for commercial development. The enzyme of P. mallochii LMB-HP37 had slightly better results than the commercial enzyme as an anti-pilling agent even though is a crude preparation. Conclusion: Penicillium mallochii LMB-HP37 produced high total cellulase activity on lactose which compares to well-known cellulase producers but at neutral to alkaline pH levels. Data obtained reveal that the crude enzyme is suitable for anti-pilling process (biopolishing) and may be also useful for biostoning.

Efecto de diámetro de esfera y densidad celular en la producción de etanol con levadura inmovilizada en alginato / Effect of pellet diameter and cell density on ethanol production with alginate immobilized yeast

Las células inmovilizadas tienen aplicación potencial en la producción de biocombustibles posibilitando la reutilización de biomasa, el empleo de diversas configuraciones de reactores y sistemas de cultivo, el manejo de altas densidades celulares alcanzando altas productividades volumétricas, y la simplificación de operaciones de procesamiento de salida. El objetivo del presente estudio fue evaluar la influencia del diámetro de las perlas y la densidad celular en la producción de etanol con Saccharomyces uvarum inmovilizada en alginato al 2% (p/v). Para ello se evaluaron tres diámetros de perlas de 2, 2,5 y 3 mm. Las células inmovilizadas fueron cultivadas en medio con 12% (p/v) de glucosa en biorreactores de columna sin agitación a 28 ºC, y se operaron cuatro lotes consecutivos de 48 horas cada uno. En cada lote se cuantificó el consumo de glucosa y se determinó la cantidad de etanol producido. Los rendimientos máximos de etanol para las esferas de 2, 2,5 y 3 mm de diámetro fueron 81, 83 y 97% del rendimiento teórico. La máxima productividad volumétrica de etanol fue 1,2 g/L-1/h-1 con un consumo de glucosa de 99,8% al término del lote, correspondiente a las columnas con perlas de 3 mm y con una producción de 0,017 g de etanol por esfera. La producción de etanol acumulada en cada sistema fue 178, 189 y 200 g/L-1 para 2, 2,5 y 3 mm respectivamente, encontrándose una relación directa con el diámetro de perla e inversa respecto a la densidad celular. Los rendimientos de etanol obtenidos son superiores a los reportados para la misma especie.

Immobilized cells have a potential use in biofuel production. They also allow re-using biomass, using diverse reactor configurations and culture systems, handling high cell densities to obtain high volumetric productivities and to simplify the downstream processing. The purpose of this work was to evaluate the influence of bead diameter and cell density on ethanol production using immobilized Saccharomyces uvarum in 2% (w/v) alginate. For that, three bead diameters (2, 2.5 and 3 mm) were evaluated. Immobilized cells were cultured on a 12% (w/v) glucose medium in column bioreactors without agitation at 28 °C for four 48 h–repeated batches. For each batch, both glucose consumption and ethanol produced were measured. Maximum yields for 2, 2.5 and 3 mm bead diameters were 81, 83 and 97% of theoretical yield. Maximum volumetric productivity of ethanol was 1.2 g/L-1/h-1 with 99.8% glucose consumption at the end of the batch, corresponding to the 3 mm bead diameter and the ethanol production per bead was 0.017 g. Accumulated ethanol production for each system was 178, 189 and 200 g/L-1 for 2, 2.5 y 3 mm bead diameter, respectively, being this directly related to bead diameter and inversely related to cell density. Ethanol yields were higher than those reported for the same species.

Influencia de la concentración de inóculo en la producción de celulasa y xilanasa por aspergillus niger / Influence of inoculum concentration on the cellulase and xylanase production by aspergillus niger

Existe un gran interés por el uso de enzimas lignocelulolíticas en varias industrias, y en la biodegradación de biomasa para la producción de biocombustibles y otras aplicaciones. Entre las fuentes microbianas de enzimas, Aspergillus niger es uno de los microorganismos más utilizados en la producción de enzimas industriales, debido a sus niveles altos de secreción de proteína y a su condición GRAS (generally regarded as safe). El objetivo del presente estudio fue evaluar la influencia de la concentración de inóculo en la morfología y producción de celulasas y xilanasas con A. niger en cultivo sumergido. Para ello, fueron inoculados matraces de 250 mL con 40 mL de medio con 3% (v/v) de una suspensión de 104 o 108 esporas por mililitro e incubados a 28 ºC y 175 rpm durante 120 horas. Se utilizaron 10 g*L-1 de lactosa como fuente de carbono. En cada caso se determinó la cantidad de biomasa, la proteína extracelular soluble, lactosa residual, actividad celulasa total y xilanasa cada 24 horas. Aunque no hubo un efecto notorio en la morfología de crecimiento, salvo en el color y el diámetro de pellets obtenidos, sí se afectó la µmax (0,06 y 0,03 h-1 para 104 y 108 esporas*mL-1, respectivamente) y la concentración máxima de biomasa. Además, mientras que las productividades volumétricas de celulasa (ΓFPA) (8,2 y 8,0 UI.*L-1*h-1 para 104 y 108 esporas*mL-1, respectivamente) fueron similares para ambos inóculos, la productividad de xilanasa (ΓXIL) fue mayor para el inóculo más concentrado (29,7 y 33,4 UI¨*L-1*h-1 para 104 y 108 esporas*mL-1, respectivamente). Los resultados indican que la productividad de celulasas y xilanasas está estrechamente relacionada con la concentración de inóculo.

There is a great interest for the use of lignocellulolytic enzymes in several industries and in biomass degradation for production of biofuels and other applications. Among the microbial sources of enzymes, Aspergillus niger is one of the most used microorganisms in the production of industrial enzymes due to its high levels of protein secretion and its GRAS (generally regarded as safe) condition. The aim of the present study was to evaluate the influence of A. niger inoculum concentration in the morphology and production of cellulases and xylanases in submerged cultures. For this, 250 mL flasks containing 40 mL culture medium were inoculated with a 3% (v/v) of either 104 or 108 spores per milliliter suspension and incubated at 28 º C and 175 rpm during 120 hours. Lactose (10 g*L-1) was used as the carbon source. In each case, the amount of biomass, the extracellular soluble protein, residual lactose, total celullase activity and xylanase activity were determined every 24 hours. Even thought there was not a notorious effect on the growth morphology, except in color and diameter of pellets; µmax was affected (0.06 and 0.03 h-1 for 104 and 108 spores*mL-1, respectively) as well as maximum biomass concentration. In addition, while the volumetric productivity of cellulase (8.2 and 8.0 UI*L-1*h-1 for 104 and 108 spores*mL-1, respectively) were similar for both inocula, the productivity of xylanase was greater for the more concentrated inoculum (29.7 and 33.4 UI*L-1*h-1 for 104 and 108 spores*mL-1, respectively).The results show that cellulase and xylanase productivities are closely related to the inoculum concentration.

Production of lignocellulolytic enzymes by Aspergillus niger biofilms at variable water activities

Lignocellulolytic enzyme production by Aspergillus niger was compared both in submerged fermentation (SF) and biofilm fermentation (BF) at varying water activities. Maximal filter paper activity, endoglucanase and xylanase activities were much higher in BF (2.96, 4.7 and 4.61 IU ml-1, respectively) than in SF cultures (1.71, 1.31 and 2.3 IU ml-1, respectively) but biomass yields were lower in BF than in SF (0.338 g g-1 and 0.431 g g-1, respectively). In the presence of 20 percent ethylene glycol (a w = 0.942) the enzyme activities decreased in both systems but BF still had higher levels (1.0, 1.0 and 2.6 IU ml-1, respectively) than SF cultures (0.6, 0.7 and 1.5 IU ml-1, respectively). An increase in xylanase specific activity of more than 2 fold (from 4.2 to 10.2 IU mg-1 biomass) was observed in the presence of 20 percent ethylene glycol, suggesting differential regulatory mechanisms in biofilm fermentation related to cell adhesion.