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The ability of the multispecies biofilm membrane reactors (MBM reactors) to provide distinguished niches for aerobic and anaerobic microbes at the same time was used for the investigation of the consolidated bioprocessing of cellulose to short chain fatty acids (SCFAs). A consortium based consolidated bioprocess (CBP) was designed. The rumen microbiome was used as the converting microbial consortium, co-cultivated with selected individual aerobic fungi which formed a biofilm on the tubular membrane flushed with oxygen. The beneficial effect of the fungal biofilm on the process yields and productivities was attributed to the enhanced cellulolytic activities compared with those achieved by the rumen microbiome alone. At 30 °C, the MBM system with Trichoderma reesei biofilm reached a concentration 39% higher (7.3 g/L SCFAs), than the rumen microbiome alone (5.1 g/L) using 15 g/L crystalline cellulose as the substrate. Fermentation temperature was crucial especially for the composition of the short chain fatty acids produced. The temperature increase resulted in shorter fatty acids produced. While a mixture of acetic, propionic, butyric, and caproic acids was produced at 30 °C with Trichoderma reesei biofilm, butyric and caproic acids were not detected during the fermentations at 37.5 °C carried out with Coprinopsis cinerea as the biofilm forming fungus. Apart from the presence of the fungal biofilm, no parameter studied had a significant impact on the total yield of organic acids produced, which reached 0.47 g of total SCFAs per g of cellulose (at 30 °C and at pH 6, with rumen inoculum to total volume ratio equal to 0.372). Electronic supplementary material The online version of this article (10.1007/s00253-019-09706-1) contains supplementary material, which is available to authorized users.

A co-fermentation process involving Saccharomyces cerevisiae and Fusarium oxysporum was studied, using hydrothermally pretreated wheat straw as substrate. In the first step of the study, we examined liquefaction of the material in a free-fall reactor. Both the enzyme loading and the dry matter content affected severely the liquefaction efficiency. In the second step (simultaneous saccharification and fermentation (SSF) experiments), we found that the enzymatic system of F. oxysporum contributed significantly to substrate hydrolysis, while its metabolic system played a secondary role in fermentation. SSF in the presence of F. oxysporum cells and enzymes gave 62 g L −1 ethanol. In the third step of the study, a semi-consolidated bioprocess was designed in which F. oxysporum culture (submerged or solid-state) was added at the SSF stage along with S. cerevisiae . The addition of solid F. oxysporum culture increased ethanol production by 19%, leading to a final ethanol concentration of 58 g L −1 . The present study proposes a semi-consolidated process combining two microorganisms for the fermentation at high solids concentration of a liquefied material using an in house free fall mixing reactor. The semi-consolidated process proposed not only increased the ethanol yields significantly, but could also lead to lower overall cost of the process by incorporating in-situ enzyme production.

High-gravity (HG) technology aims at generating final ethanol concentrations above 50 kg m(-3) in order to reduce the cost of the distillation step. The generation of higher amounts of inhibitors during the pretreatment step is one of the challenges that accompany the increase in initial dry matter. Detoxification of spruce hydrolysate, adaptation of the cells before fermentation, supplementation with nutrients, and washing of solids were the strategies compared in this study. They represent different approaches to cope with the inhibitory effects, and we compared their efficiencies using a thermotolerant strain of Saccharomyces cereuisiae at temperatures from 30 degrees C up to 40 degrees C. The dilute acid-pretreated spruce used as substrate in this study was not fermentable under HG conditions (200 g kg(-1) water-insoluble solids) when no improvement method was applied. In HG simultaneous saccharification and fermentation at 30 degrees C combined with a 24 h pre-hydrolysis step, the detoxification of pretreated spruce with reducing agent (Na2S2O4) gave the best result with an ethanol yield of 57% (on total sugars) of the maximum theoretical and a volumetric productivity of 1.58 g dm(-3) h(-1). In HG separate hydrolysis and fermentation, nutrients supplementation gave better final ethanol yields than detoxification of the material, reaching an ethanol yield of about 60% of the theoretical (on total sugars). The results obtained, showed an increase in severity of inhibitory effects with temperature increase. Improved cell viability was observed when detoxified material was used and also when yeast extract addition was coupled with adaptation of the cells to the hydrolysate. (C) 2014 Published by Elsevier Ltd.

BACKGROUND: During the pretreatment of lignocellulosic materials, molecules such as carboxylic acids, furan derivatives and phenolic compounds, which inhibit the growth and ethanol fermentation by bacteria, fungi and yeasts, are produced. The present work determines the tolerance levels of the C5, C6 fermenting fungus Fusarium oxysporum, towards individual model inhibitory compounds on aerobic growth, on lignocellulolytic activities and on fermentative performance. RESULTS During the growth stage, maximum biomass production was more affected than the specific growth rate by the presence of inhibitors. The presence of high concentrations of inhibitors resulted, in most cases, in prolongation of the lag phase. The fermentative performance of F. oxysporum was significantly inhibited by carboxylic acids, while the lignocellulolytic activities were affected to a lesser extent. CONCLUSION: The toxicity tolerance of fF. oxysporum was high enough for aerobic growth in the presence of significant concentrations of inhibitors, which in most cases were higher than those generated from various treatments of lignocellulosic materials, while its fermentative performance was relatively more affected by the presence of inhibitors. The decrease of ethanol production in the presence of weak organic acids could be the main obstacle to the application of F. oxysporum in large-scale bioconversion processes of hydrolysates containing inhibitors. Copyright © 2010 Society of Chemical Industry

In this study, the ethanol production by the mesophilic fungus Neurospora crassa from BG was studied and optimized concerning the induction of lignocellulose degrading enzymes and the production phase as well. The production of cellulolytic and hemicellulolytic enzymes was studied under solid-state cultivation (SSC). SSC in a laboratory horizontal bioreactor using the optimized medium, WS and BG in the ratio 1:1 and initial moisture level 61.5%, allowed the large scale production of the multienzymatic system. Similar yields with those from flasks experiments, as high as 1073,56,4.2,1.6,3.1,5.7 and 0.52 U g(-1) carbon source of xylanase, endoglucanase, cellobiohydrolase, beta-glucosidase, alpha-l-arabinofuranosidase, acetyl esterase and feruloyl esterase, respectively, were obtained. Chromogenic (fluorogenic) 4-methylumbelliferyl substrates were used to characterize the major activities of the multienzyme component, after the separation by isoelectric focusing (IEF) electrophoresis. Alkali pre-treated BG was used for ethanol production. A yield of about 74 g of ethanol kg(-1) dry BG (5,6 g L(-1)) was obtained under optimum conditions (aeration 0.1 vvm, pre-treatment with 1g NaOH 10 g(-1)dry BG).

Background Fusarium oxysporum is a filamentous fungus which has attracted a lot of scientific interest not only due to its ability to produce a variety of lignocellulolytic enzymes, but also because it is able to ferment both hexoses and pentoses to ethanol. Although this fungus has been studied a lot as a cell factory, regarding applications for the production of bioethanol and other high added value products, no systematic study has been performed concerning its ethanol tolerance levels. Results In aerobic conditions it was shown that both the biomass production and the specific growth rate were affected by the presence of ethanol. The maximum allowable ethanol concentration, above which cells could not grow, was predicted to be 72 g/L. Under limited aeration conditions the ethanol-producing capability of the cells was completely inhibited at 50 g/L ethanol. The lignocellulolytic enzymatic activities were affected to a lesser extent by the presence of ethanol, while the ethanol inhibitory effect appears to be more severe at elevated temperatures. Moreover, when the produced ethanol was partially removed from the broth, it led to an increase in fermenting ability of the fungus up to 22.5%. The addition of F. oxysporum’s system was shown to increase the fermentation of pretreated wheat straw by 11%, in co-fermentation with Saccharomyces cerevisiae. Conclusions The assessment of ethanol tolerance levels of F. oxysporum on aerobic growth, on lignocellulolytic activities and on fermentative performance confirmed its biotechnological potential for the production of bioethanol. The cellulolytic and xylanolytic enzymes of this fungus could be exploited within the biorefinery concept as their ethanol resistance is similar to that of the commercial enzymes broadly used in large scale fermentations and therefore, may substantially contribute to a rational design of a bioconversion process involving F. oxysporum. The SSCF experiments on liquefied wheat straw rich in hemicellulose indicated that the contribution of the metabolic system of F. oxysporum in a co-fermentation with S. cerevisiae may play a secondary role.

In this study, the factors affecting ferulic acid (FA) release from Brewer’s spent grain (BSG), by the crude enzyme extract of Fusarium oxysporum were investigated. In order to evaluate the importance of the multienzyme preparation on FA release, the synergistic action of feruloyl esterase (FAE, FoFaeC-12213) and xylanase (Trichoderma longibrachiatum M3) monoenzymes was studied. More than double amount of FA release (1 mg g−1 dry BSG) was observed during hydrolytic reactions by the crude enzyme extract compared to hydrolysis by the monoenzymes (0.37 mg g−1 dry BSG). The protease content of the crude extract and the inhibitory effect of FA as an end-product were also evaluated concerning their effect on FA release. The protease treatment prior to hydrolysis by monoenzymes enhanced FA release about 100%, while, for the first time in literature, FA in solution found to have a significant inhibitory effect on FAE activity and on total FA release.

Background Brewer's spent grain (BG), a by-product of the brewing process, is attracting increasing scientific interest as a low-cost feedstock for many biotechnological applications. BG in the present study is evaluated as a substrate for lignocellulolytic enzyme production and for the production of ethanol by the mesophilic fungus Fusarium oxysporum under submerged conditions, implementing a consolidated bioconversion process. Fermentation experiments were performed with sugar mixtures simulating the carbohydrate content of BG in order to determine the utilization pattern that could be expected during the fermentation of the cellulose and hemicellulose hydrolysate of BG. The sugar mixture fermentation study focused on the effect of the initial total sugar concentration and on the effect of the aeration rate on fermenting performance of F. oxysporum. The alkali pretreatment of BG and different aeration levels during the ethanol production stage were studied for the optimization of the ethanol production by F. oxysporum. Results Enzyme yields as high as 550, 22.5, 6.5, 3225, 0.3, 1.25 and 3 U per g of carbon source of endoglucanase, cellobiohydrolase, β-D-glucosidase, xylanase, feruloyl esterase, β-D-xylosidase and α-L-arabinofuranosidase respectively, were obtained during the growth stage under optimized submerged conditions. An ethanol yield of 109 g ethanol per kg of dry BG was obtained with alkali-pretreated BG under microaerobic conditions (0.01 vvm), corresponding to 60% of the theoretical yield based on total glucose and xylose content of BG. Conclusion The enzymatic profile of the extracellular extract from F. oxysporum submerged cultures using BG and corn cob as the carbon source was proved efficient for a successful hydrolysis of BG. The fermentation study carried out using sugar mixtures simulating BG's carbohydrates content and consecutively alkali-pretreated and untreated BG, indicates that BG hydrolysis is the bottleneck of the bioconversion process. However, a considerable bioconversion yield was achieved (60% of the theoretical) making this bioprocess worthy of further investigation for a potential commercial application.