Bioethanol Production from Brewers Spent Grains Using a Fungal Consolidated Bioprocessing (CBP) Approach.
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ABSTRACT:
SUBMITTER: Wilkinson S
PROVIDER: S-EPMC7114960 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Brewing spent grains (BSGs) are the main by-product from breweries and they are rich of proanthocyanidins, among other phenolic compounds. However, literature on these compounds in BSGs is scarce. Thus, this research focuses on the establishment of ultrasound-assisted extraction of proanthocyanidin compounds in brewing spent grains using a sonotrode. To set the sonotrode extraction up, response surface methodology (RSM) was used to study the effects of three factors, namely, solvent composition, time of extraction, and ultrasound power. Qualitative and quantitative analyses of proanthocyanidin compounds were performed using HPLC coupled to fluorometric and mass spectrometer detectors. The highest content of proanthocyanidins was obtained using 80/20 acetone/water (v/v), 55 min, and 400 W. The established method allows the extraction of 1.01 mg/g dry weight (d.w.) of pronthocyanidins from BSGs; this value is more than two times higher than conventional extraction.
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Project description:Fusarium oxysporum is among the few filamentous fungi capable of fermenting ethanol directly from lignocellulose biomass (LCB). It has the essential enzymatic toolbox to disintegrate LCB to its monosaccharides, which subsequently fermented to ethanol under anaerobic and micro-aerobic conditions. However, the structural complexity of LCB and modest performances of wild fungi are major limitations for application in local biorefineries. This study assessed the potential of the locally isolated Fusarium oxysporum for the production of bioethanol from Ficus fruits (Ficus cunia) using Consolidated Bioprocessing (CBP). The maximum ethanol concentration achieved was at 5% substrate loadings with pH 6 irrespective of temperature variance, attaining a concentration of 3.54 g/L and 3.88 g/L at 28 °C and 32 °C, respectively. The monitoring of analytes (glucose, arabinose, cellobiose, xylose, acetic acid, ethanol, furfural, and HMF) in this study suggests the utilization of an array of sugars released from Ficus fruits, irrespective of the difference in the process parameters. This study also shows that CBP of freshly grounded Ficus fruits was feasible employing a mild hydrothermal pretreatment (autoclaved at 121 °C for 30 min in 1:10 w/v) and without supplementing any extraneous enzymes.Supplementary informationThe online version contains supplementary material available at 10.1007/s13205-022-03234-y.
| S-EPMC9294110 | biostudies-literature
Project description:Brewers' spent grain (BSG) is the major by-product of the brewing industry which remain largely unutilized despite its nutritional quality. In this study, the effects of fermentation on BSG antioxidant potential were analyzed. A biotechnological protocol including the use of xylanase followed by fermentation with Lactiplantibacillus plantarum (Lactobacillus plantarum) PU1, PRO17, and H46 was used. Bioprocessed BSG exhibited enhanced antioxidant potential, characterized by high radical scavenging activity, long-term inhibition of linoleic acid oxidation and protective effect toward oxidative stress on human keratinocytes NCTC 2544. Immunolabelling and confocal laser microscopy showed that xylanase caused an extensive cell wall arabinoxylan disruption, contributing to the release of bound phenols molecules, thus available to further conversion through lactic acid bacteria metabolism. To clarify the role of fermentation on the antioxidant BSG potential, phenols were selectively extracted and characterized through HPLC-MS techniques. Novel antioxidant peptides were purified and identified in the most active bioprocessed BSG.
| S-EPMC7411387 | biostudies-literature
Project description:Brewers' spent grains (BSG) were fermented with Rhizopus oligosporus and up to 15% of original protein was hydrolysed. Fermented BSG was then subjected to an ethanolic-alkali extraction and isolated fractions contained 61-66% protein. An evaluation of functional properties suggested that fermented extracts presented superior emulsifying abilities (15-34 m2/g of activity and 16-42 min of stability), foaming properties (16-30% capacity and 7-14% stability), and water/oil binding capacities (0.41 g/g and 0.24 g/g, respectively). They also showed significantly higher ABTS inhibition and stronger reducing power than unfermented ones, indicating that fermented BSG protein extract had greater antioxidant activities. No cytotoxic effect was detected in the range of 2-10 mg/mL. When applied in a mayonnaise formulation, fermented hydrolysates demonstrated better emulsion stability in terms of creaming, microstructure and viscosity. Thus, fermented BSG protein is a potential plant-based emulsifier for food, pharmaceutical and cosmetic applications.
| S-EPMC8666519 | biostudies-literature
Project description:Natural yeast with superior fermentative traits can serve as a platform for the development of recombinant strains that can be used to improve the sustainability of bioethanol production from starch. This process will benefit from a consolidated bioprocessing (CBP) approach where an engineered strain producing amylases directly converts starch into ethanol. The yeast Saccharomyces cerevisiae L20, previously selected as outperforming the benchmark yeast Ethanol Red, was here subjected to a comparative genomic investigation using a dataset of industrial S. cerevisiae strains. Along with Ethanol Red, strain L20 was then engineered for the expression of α-amylase amyA and glucoamylase glaA genes from Aspergillus tubingensis by employing two different approaches (delta integration and CRISPR/Cas9). A correlation between the number of integrated copies and the hydrolytic abilities of the recombinants was investigated. L20 demonstrated important traits for the construction of a proficient CBP yeast. Despite showing a close relatedness to commercial wine yeast and the benchmark Ethanol Red, a unique profile of gene copy number variations (CNVs) was found in L20, mainly encoding membrane transporters and secretion pathway proteins but also the fermentative metabolism. Moreover, the genome annotation disclosed seven open reading frames (ORFs) in L20 that are absent in the reference S288C genome. Genome engineering was successfully implemented for amylase production. However, with equal amylase gene copies, L20 proved its proficiency as a good enzyme secretor by exhibiting a markedly higher amylolytic activity than Ethanol Red, in compliance to the findings of the genomic exploration. The recombinant L20 dT8 exhibited the highest amylolytic activity and produced more than 4 g/L of ethanol from 2% starch in a CBP setting without the addition of supplementary enzymes. Based on the performance of this strain, an amylase/glucoamylase ratio of 1:2.5 was suggested as baseline for further improvement of the CBP ability. Overall, L20 showed important traits for the future construction of a proficient CBP yeast. As such, this work shows that natural S. cerevisiae strains can be used for the expression of foreign secreted enzymes, paving the way to strain improvement for the starch-to-bioethanol route.
| S-EPMC8815085 | biostudies-literature
Project description:BACKGROUND: The recalcitrant nature of hemicellulosic materials and the high cost in depolymerization are the primary obstacles preventing the use of xylan as feedstock for fuel and chemical production. Consolidated bioprocessing, incorporating enzyme-generating, biomass-degrading and bioproduct-producing capabilities into a single microorganism, could potentially avoid the cost of the dedicated enzyme generation in the process of xylan utilization. In this study, we engineered Escherichia coli strains capable of exporting three hemicellulases to the broth for the succinate production directly from beechwood xylan. RESULTS: Xylanases were extracellular environment-directed by fusing with OsmY. Subsequently, twelve variant OsmY fused endoxylanase-xylosidase combinations were characterized and tested. The combination of XynC-A from Fibrobacter succinogenes S85 and XyloA from Fusarium graminearum which appeared to have optimal enzymatic properties was identified as the best choice for xylan hydrolysis (0.18 ± 0.01 g/l protein in the broth with endoxylanase activity of 12.14 ± 0.34 U/mg protein and xylosidase activity of 92 ± 3 mU/mg protein at 8 h after induction). Further improvements of hemicellulases secretion were investigated by lpp deletion, dsbA overexpression and expression level optimization. With co-expression of ?-arabinofuranosidase, the engineered E. coli could hydrolyze beechwood xylan to pentose monosaccharides. The hemicellulolytic capacity was further integrated with a succinate-producing strain to demonstrate the production of succinate directly from xylan without externally supplied hydrolases and any other organic nutrient. The resulting E. coli Z6373 was able to produce 0.37 g/g succinate from xylan anaerobically equivalent to 76% of that from xylan acid hydrolysates. CONCLUSIONS: This report represents a promising step towards the goal of hemicellulosic chemical production. This engineered E. coli expressing and secreting three hemicellulases demonstrated a considerable succinate production on the released monosaccharides from xylan. The ability to use lower-cost crude feedstock will make biological succinate production more economically attractive.
| S-EPMC3340313 | biostudies-literature
Project description:The constraints and advantages in cellulolytic enzymes production by fungal biofilms for a consolidated bioconversion process were investigated during this study. The biofilm cultivations were carried out in reactors designed for consolidated bioprocessing Multispecies Biofilm Membrane reactors, (MBM) where an aerobic fungal biofilm produces the lignocellulolytic enzymes while a fermenting microorganism forms the fermentation product at anaerobic conditions. It was shown that although mycelial growth was limited in the MBM reactors compared to submerged cultivations, the secretion of cellulolytic enzymes per cell dry weight was higher. When Trichoderma reesei was used as the sole enzyme producer, cellobiose accumulated in the liquid medium as the result of the deficiency of β-glucosidase in the fungal secretome. To enhance β-glucosidase activity, T. reesei was co-cultivated with A. phoenicis which is a β-glucosidase overproducer. The two fungi formed a multispecies biofilm which produced a balanced cellulolytic cocktail for the saccharification of plant biomass. The mixed biofilm reached a 2.5 fold increase in β-glucosidase production, compared to the single T. reesei biofilm. The enzymatic systems of single and mixed biofilms were evaluated regarding their efficiency on cellulosic substrates degradation. Washed solids from steam pretreated beechwood, as well as microcrystalline cellulose were used as the substrates. The enzymatic system of the multispecies biofilm released four times more glucose than the enzymatic system of T. reesei alone from both substrates and hydrolyzed 78 and 60% of the cellulose content of washed solids from beechwood and microcrystalline cellulose, respectively.
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Project description:Vermicomposting is a cost-effective biotechnology for the management of organic wastes that relies on the activity of earthworms and their associated microbiota. Here, the microbiotas of the earthworm Eisenia fetida fed with brewers' spent grains (FBSG), cow manure (FCM) and a mix of brewers' spent grains/cow manure (FMIX), were identified by high-throughput DNA sequencing (16S rRNA). Bacterial community variance was correlated with the pH and the organic carbon content of the rearing substrates. FBSG microbiota was enriched in Paenibacillaceae, Enterobacteriaceae, Chitinophagaceae and Comamonadaceae. In addition, FBSG microbiota had a predicted higher abundance of genes involved in cellulose degradation as well as in the nitrogen cycle and showed higher utilization of ammonia and nitrate. Results obtained will allow to optimize the vermicomposting of brewers' spent grains and to evaluate the effect of vermicompost addition on nutrient dynamics in soil.
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Project description:A system for itaconic acid synthesis from cellulose by Neurospora crassa was established, resulting in the highest yield of itaconic acid was 354.08 + 35.99 mg/L. Meanwhile, cellulase activity increased significantly, without any strain modifications for improved cellulase production. Multi-omics analyses showed that itaconic acid synthesis reduced energy production, leading to decreases in trehalose, cell wall, fatty acids synthesis and downregulations in MAPK signaling pathway, cell cycle and meiosis. More importantly, the low-energy environment enhanced the energy-efficient cellobionic acid/gluconic acid pathway, and the cellulase composition also changed significantly, manifested as the up-regulation of LPMOs and the down-regulation of β-glucosidases. Enhancing LPMOs-cellobionic acid/gluconic acid system has the potential to reduce energy consumption of the consolidated bioprocessing. These findings offer an overview of resource allocations by N. crassa in response to itaconic acid synthesis and highlight a series of intriguing connections between itaconic acid synthesis and cellulase synthesis in consolidated bioprocessing.
| S-EPMC9922455 | biostudies-literature
Project description:BACKGROUND:Yarrowia lipolytica, one of the most widely studied "nonconventional" oleaginous yeast species, is unable to grow on cellulose. Recently, we identified and overexpressed two endogenous ?-glucosidases in Y. lipolytica, thus enabling this yeast to use cello-oligosaccharides as a carbon source for growth. Using this engineered yeast platform, we have now gone further toward building a fully cellulolytic Y. lipolytica for use in consolidated bioprocessing of cellulose. RESULTS:Initially, different essential enzyme components of a cellulase cocktail (i.e,. cellobiohydrolases and endoglucanases) were individually expressed in Y. lipolytica in order to ascertain the viability of the strategy. Accordingly, the Trichoderma reesei endoglucanase I (TrEG I) and II (TrEG II) were secreted as active proteins in Y. lipolytica, with the secretion yield of EG II being twice that of EG I. Characterization of the purified His-tagged recombinant EG proteins (rhTrEGs) revealed that rhTrEG I displayed higher specific activity than rhTrEG II on both cellotriose and insoluble cellulosic substrates, such as Avicel, ?-1, 3 glucan, ?-1, 4 glucan, and PASC. Similarly, cellobiohydrolases, such as T. reesei CBH I and II (TrCBH I and II), and the CBH I from Neurospora crassa (NcCBH I) were successfully expressed in Y. lipolytica. However, the yield of the expressed TrCBH I was low, so work on this was not pursued. Contrastingly, rhNcCBH I was not only well expressed, but also highly active on PASC and more active on Avicel (0.11 U/mg) than wild-type TrCBH I (0.065 U/mg). Therefore, work was pursued using a combination of NcCBH I and TrCBH II. The quantification of enzyme levels in culture supernatants revealed that the use of a hybrid promoter instead of the primarily used TEF promoter procured four and eight times more NcCBH I and TrCBH II expressions, respectively. Finally, the coexpression of the previously described Y. lipolytica ?-glucosidases, the CBH II, and EG I and II from T. reesei, and the N. crassa CBH I procured an engineered Y. lipolytica strain that was able to grow both on model cellulose substrates, such as highly crystalline Avicel, and on industrial cellulose pulp, such as that obtained using an organosolv process. CONCLUSIONS:A Y. lipolytica strain coexpressing six cellulolytic enzyme components has been successfully developed. In addition, the results presented show how the recombinant strain can be optimized, for example, using artificial promoters to tailor expression levels. Most significantly, this study has provided a demonstration of how the strain can grow on a sample of industrial cellulose as sole carbon source, thus revealing the feasibility of Yarrowia-based consolidated bioprocess for the production of fuel and chemical precursors. Further, enzyme and strain optimization, coupled to appropriate process design, will undoubtedly lead to much better performances in the future.
| S-EPMC5438512 | biostudies-literature