A. Sanromán

Improving laccase production by employing different lignocellulosic wastes in submerged cultures of Trametes versicolor.

Laccase production by the white-rot fungus Trametes versicolor (CBS100.29) grown in submerged cultures was studied. Addition of different insoluble lignocellulosic materials into the culture medium in order to enhance laccase production was investigated. The lignocellulosic materials were grape seeds, grape stalks and barley bran, selected because of their availability and low cost, since they are agro-industrial wastes abundant in most countries. Barley bran gave the highest activities, a maximum value of 639U/l, which was 10 times the value attained in the cultures without lignocellulosics addition. The decolourisation of a model dye, Phenol Red, by the ligninolytic fluids obtained in the above-mentioned cultures was investigated. Grape stalk and barley bran cultures showed the highest ability to decolourise the dye, attaining a percentage of decolourisation of around 60% in 72 h.

Enhanced ligninolytic enzyme production and degrading capability of Phanerochaete chrysosporium and Trametes versicolor

Ligninolytic enzyme production by the white-rot fungi Phanerochaete chrysosporium and Trametes versicolor precultivated with different insoluble lignocellulosic materials (grape seeds, barley bran and wood shavings) was investigated. Cultures of Phanerochaete chrysosporium precultivated with grape seeds and barley bran showed maximum lignin peroxidase (LiP) and manganese-dependent peroxidase (MnP) activities (1000 and 1232 U/l, respectively). Trametes versicolor precultivated with the same lignocellulosic residues showed the maximum laccase activity (around 250 U/l). For both fungi, the ligninolytic activities were about two-fold higher than those attained in the control cultures. In vitro decolorization of the polymeric dye Poly R-478 by the extracellular liquid obtained in the above-mentioned cultures was monitored in order to determine the respective capabilities of laccase, LiP and MnP. It is noteworthy that the degrading capability of LiP when P. chrysosporium was precultivated with barley bran gave a percentage of Poly R-478 decolorization of about 80% in 100 s, whereas control cultures showed a lower percentage, around 20%, after 2 min of the decolorization reaction.

Investigation of several bioreactor configurations for laccase production by Trametes versicolor operating in solid-state conditions

Abstract In the present paper, the production of laccase by Trametes versicolor (CBS 100.29) in laboratory-scale bioreactors, operating in semi-solid-state conditions, was studied. Three bioreactor configurations were investigated in order to determine the most suitable one for laccase production: immersion, expanded-bed and tray. In addition, the nature of support employed (inert or non-inert) on laccase production was also evaluated. According to the results attained in the previous work by our research group, nylon sponge and barley bran was employed as an inert and as a lignocellulosic support, respectively. Higher laccase activities were produced operating with barley bran than with nylon sponge as a support in all the configurations tested. As regards bioreactor design, the tray configuration led to the highest laccase activities especially operating with barley bran as a support, where activities of about 10-fold higher than that found in the corresponding cultivation with nylon sponge were attained. Therefore, it could be asserted that the tray bioreactor is a very appropriate bioreactor configuration to produce laccase by T. versicolor in solid-state conditions operating with lignocellulosic supports.

Photocatalytic degradation of dyes in aqueous solution operating in a fluidised bed reactor

This work reports a preliminary design of a new photochemical reactor and its application to photochemical degradation of two dyes, Crystal Violet and Azure B, operating in both batch and continuous processes. A novel kind of photocatalyst, consisting of ZnO immobilised in alginate gel beads, which is able to photodegrade organic dyes effectively, has been employed in the present study. When this photocatalyst, at a concentration of 1 g of ZnO per litre of alginate gel at 3%, was employed in batch process, almost total decolourisation of Crystal Violet in reaction times lower than 120 min was observed. Operating in continuous process at different residence times, it was possible to achieve a total decolourisation of both Crystal Violet and Azure B. Moreover, the total organic carbon content (TOC) was reduced to 90% in the former and to 52% in the latter. These results indicated that the photoreactor developed in the present work was able to degrade effectively dyes of different structures, revealing the non-specificity of the system.

Ligninolytic enzyme production and the ability of decolourisation of Poly R-478 in packed-bed bioreactors by Phanerochaete chrysosporium

Abstract  The production of ligninolytic enzymes by the fungus Phanerochaetechrysosporium BKM-F-1767 (ATCC 24725) in packed-bed tubular bioreactors, operating in semi-solid-state conditions, was studied. Three types of carriers were assayed: cubes of polyurethane foam, cubes of nylon sponge and chopped corncob, in order to determine the more suitable one to produce ligninolytic enzymes by this fungus. The cultivations were carried out in discontinuous and in continuous mode.For discontinuous cultivation, maximum individual manganese-dependent peroxidase (MnP) activities of 1593, 1371 and 346 U/l were achieved in the bioreactors filled with cubes of nylon sponge, cubes of polyurethane foam and with corncob, respectively. On the other hand, lignin peroxidase (LiP) activities about 100 U/l were found in the two former and around 200 U/l in the latter. Moreover, laccase, was detected in all cultures, with average values of 30 U/l. Nonetheless, continuous mode cultivation led to lower ligninolytic enzyme activities than those produced in discontinuous, except in the case of the corncob.Furthermore, the decolourisation of the dye Poly R-478 by the above-mentioned cultures was investigated. The percentage of biological decolourisation reached was about 70% in the bioreactor filled with cubes of nylon sponge whereas it was rather low in the others (around 30%).

Utilisation of lignocellulosic wastes for lignin peroxidase production by semi-solid-state cultures of Phanerochaete chrysosporium.

In the present work, the production of ligninolytic enzymes by semi-solid-statecultures of Phanerochaete chrysosporium BKM-F-1767 (ATCC 24725),employing different lignocellulosic wastes as support, was investigated. Thewaste materials employed were grape seeds, wheat straw and wood shavings.Maximum lignin peroxidase activities of 1620 ± 123 U/l, 364 ± 35 U/l and 571 ± 42 U/l were attained, respectively. Nevertheless, lowmanganese-dependent peroxidase activities were found, being insignificantin the grape seed cultures. Moreover, the in vivo decolourisation of a model dye compound, the polymeric dye Poly R-478 (polyvinylamine sulfonateanthrapyridone), by the above-mentioned cultures was monitored to assessthe degrading capability of the extracellular liquid secreted by such cultures.The percentage of biological decolourisation attained by grape seed and woodshaving cultures was around 74% and 63%, respectively, whereas it was ratherlow (40%) in the wheat straw ones.

Comparison between the protease production ability of ligninolytic fungi cultivated in solid state media

Solid state cultures of two white-rot fungi, Phanerochaete chrysosporium and Phlebia radiata, have been carried out, using an inert support (nylon sponge) and a support-substrate (corncob). The suitable medium and culture conditions have been chosen to favour the secretion of ligninolytic enzymes. The production of manganese peroxidase, lignin peroxidase, laccase and proteases has been monitored during the cultures, in an attempt to investigate the possible effect of the latter on the integrity of ligninolytic enzymes. The higher the protease concentration in the culture medium, the more irregular the profiles of ligninolytic enzyme activity. P. chrysosporium secretes proteolytic enzymes mainly during primary metabolism, while P. radiata produced these at the onset of secondary metabolism. Furthermore, different types of proteases produced were identified, P. chrysosporium secreted mainly thiol and acidic proteases, while P. radiata cultures contained thiol-, serin- and metalloproteases.

Performance of a solid-state immersion bioreactor for ligninolytic enzyme production: evaluation of different operational variables

The production of ligninolytic enzymes by the white-rot fungus Phanerochaete chrysosporium BKM-F-1767 in a solid-state immersion bioreactor, employing cubes of nylon sponge as a support, was studied. Cultivation was carried out in both batch and continuous mode, and the effect of some operational variables (aeration level, pH) was investigated. Batch operation at an aeration level of 0.5 vvm led to maximum manganese-dependent peroxidase (MnP) and lignin peroxidase (LiP) activities of 574 and 116 U l−1, respectively. The results were compared with those obtained at a higher aeration level (1 vvm), reported in a previous work, and it appeared that LiP productivity increased with aeration rate, while MnP was not significantly affected. Continuous operation showed much lower MnP activities (around 60 U l−1) and similar LiP activities (about 132 U l−1). However, in terms of productivity the difference between batch and continuous operation for MnP was less remarkable (239 and 150 U day−1, respectively), whereas LiP productivity was ten-fold higher in continuous operation than in batch mode. This could be attributed to the influence of operation pH on ligninolytic enzyme activities. The study of the kinetic characteristics of the biocatalysts supported this hypothesis.

Stimulation of ligninolytic enzyme production and the ability to decolourise Poly R-478 in semi-solid-state cultures of Phanerochaete chrysosporium

Abstract The effect of adding some activators of ligninolytic enzyme production, Tween 80, veratryl alcohol and manganese (IV) oxide, to semi-solid-state cultures of Phanerochaete chrysosporium BKM-F-1767 (ATCC 24725) was studied. As carrier, cubes of polyurethane foam were employed. Supplementing the cultures with Tween 80, a maximum manganese-dependent peroxidase (MnP) activity of 350 U/l was achieved. This value was about 3-fold higher than that obtained in the control cultures (without Tween 80). When not only Tween 80 but also veratryl alcohol or manganese (IV) oxide were incorporated into the culture medium, the effect was greater and led to MnP activities of about 600 and 1100 U/l with alcohol or oxide, respectively. These activities were notably higher than those obtained in the control cultures (about 3-fold and 7-fold, respectively). The decolourisation of the dye Poly R-478 by the above-mentioned cultures was investigated. The percentage of biological decolourisation reached at the end of all the cultivations was higher than 95%.

Production of manganese-dependent peroxidase in a new solid-state bioreactor by Phanerochœte chrysosporium grown on wood shavings. Application to the decolorization of synthetic dyes

The production of manganese-dependent peroxidase (MnP) byPhanerochœte chrysosporium in a new solid-state bioreactor, the immersion bioreactor, operating with lignocellulosic waste, such as wood shavings, was investigated. Maximum MnP and lignin peroxidase (LiP) activity of 13.4 and 8.48 μkat/L were obtained, respectively. Thein vitro decolorization of several synthetic dyes by the extracellular liquid produced in the above-mentioned bioreactor (containing mainly MnP) was carried out and its degrading ability was assessed. The highest decolorization was reached with Indigo Carmine (98%) followed by Bromophenol Blue (56%) and Methyl Orange (36%), whereas Gentian Violet was hardly decolorized (6%).