Solid-state fermentation of Ulva Rigida for production of cellulases, xylanases and ß-glucosidase

Seaweeds are important organisms in marine ecosystems, are included on blue biorefinery category and they are rich in different polysaccharides, that can induce enzyme production by solid-state fermentation (SSF). Ulva rigida was used as substrate in SSF to produce cellulases and xylanases and to increase its protein content. The SSF were performed with previously washed and with unwashed seaweed by Aspergillus ibericus. Xylanase and cellulase activities were higher in fermented unwashed seaweed, correspondingly 359.8±6.1 U/g dry substrate and 59.9±2.4 U/g, respectively. In fermented washed U. rigida, these values achieved 212.4±17.7 U/g and 43.6±3.7 U/g, respectively. -glucosidase activity was similar in washed seaweed (6.94±0.21 U/g) and in unwashed seaweed (6.87±0.18 U/g). After SSF, protein content of unwashed seaweed was increased by 14%. Unwashed U. rigida resulted in better enzymatic activities, being a promising option to produce these commercially-valued enzymes.Helena Fernandes was supported by PhD grant SFRH/BD/131219/2017, funded by the Portuguese Foundation for Science and Technology (FCT). José M. Salgado was supported by grant CEB/N2020 – INV/01/2016 from Project “BIOTECNORTE - Underpinning Biotechnology to foster the north of Portugal bioeconomy” (NORTE-01- 0145-FEDER-000004). This study was partially supported by FCT under the scope of the strategic funding of UID/BIO/04469/2013 unit and COMPETE 2020 (POCI-01-0145- FEDER-006684) and BioTec-Norte operation (NORTE-01-0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte and by the project SPO3 (Development of innovative sustainable protein and omega-3 rich feedstuffs for aquafeeds, from local agro-industrial by-products reference POCI-01-0145-FEDER-030377 funded by FEDER-Operational Programme Competitiveness and Internationalization and FCT.info:eu-repo/semantics/publishedVersio

Biorefinery of brewery spent grain by solid-state fermentation and ionic liquids

Novel environmentally friendly pretreatments have been developed in recent years to improve biomass fractionation. Solid-state fermentation (SSF) and treatment with ionic liquids show low environmental impact and can be used in biorefinery of biomass. In this work, these processes were assessed with brewery spent grain (BSG). First, BSG was used as a substrate to produce cellulases and xylanases by SSF with the fungi Aspergillus brasiliensis CECT 2700 and Trichoderma reesei CECT 2414. Then, BSG was pretreated with the ionic liquid [N1112OH][Gly] and hydrolyzed with the crude enzymatic extracts. Results showed that SSF of BSG with A. brasiliensis achieved the highest enzyme production; meanwhile, the pretreatment with ionic liquids allowed glucan and xylan fractions to increase and reduce the lignin content. In addition, a mixture of the extracts from both fungi in a ratio of 2.5:0.5 Aspergillus/Trichoderma (v/v) efficiently hydrolyzed the BSG previously treated with the ionic liquid [N1112OH][Gly], reaching saccharification percentages of 80.68%, 54.29%, and 19.58% for glucan, xylan, and arabinan, respectively. In conclusion, the results demonstrated that the BSG biorefinery process developed in this work is an effective way to obtain fermentable sugar-containing solutions, which can be used to produce value-added products.São Paulo Research Foundation | Ref. 2018/25511-1São Paulo Research Foundation | Ref. 2021/15138-4National Council for Scientific and Technological Development | Ref. 408783/2021-4National Council for Scientific and Technological Development | Ref. 312923/2020-1Xunta de Galicia | Ref. GPC-ED431B 2021/23Ministerio de Ciencia e Innovación | Ref. PID2020-115879RB-I0

Bio-enrichment of oilseed cakes by Mortierella alpina under solid-state fermentation

"Available online 3 August 2020"Oilseed cakes have potential for new applications as substrates for solid-state fermentation (SSF), to increase their nutritional value by increasing its polyunsaturated fatty acids (PUFAs) or protein content. In this sense, it was performed a screening of oilseed cakes to be used as substrates for the production of PUFAs by Mortierella alpina Peyronel MUM 9412. Of all by-products tested, linseed cake (LSC) was the oilseed cake that achieved the highest production of total PUFAs: 153.09 ± 2.25 mg/g. Overall, the PUFAs and protein contents of fermented LSC increased 33% and 11%, respectively. Further, supplementation of the rapeseed cake with linseed oil, prior to the SSF, proved to increase the PUFAs production in about 26%. This study demonstrated the potential of SSF for improving linseed and linseed cakes nutritional composition and the positive effect of linseed oil as inductor to improve the PUFAs production by M. alpina.José Manuel Salgado was supported by grant CEB/N2020 – INV/01/ 2016 from Project “BIOTECNORTE - Underpinning Biotechnology to foster the north of Portugal bioeconomy” (NORTE-01-0145-FEDER000004). This work was supported by the project “Development of innovative sustainable protein and omega-3 rich feedstuffs for aquafeeds, from local agro-industrial by-products”, reference POCI-01-0145- FEDER-030377, funded by European Regional Development Fund (ERDF). This study was supported by the Portuguese Foundation for Science and Technology (FCT) under the scope of the strategic funding of UID/BIO/04469/2019 unit and BioTecNorte operation (NORTE-01- 0145-FEDER-000004) funded by the European Regional Development Fund under the scope of Norte2020 - Programa Operacional Regional do Norte.info:eu-repo/semantics/publishedVersio

Biorefinery of brewery spent grain to obtain bioproducts with high value-added in the market

The brewery industry is under economic and environmental pressure to minimize residual management costs, particularly brewery spent grain (BSG), which accounts for 80–85% (w/w) of the total by-products generated. BSG is a lignocellulosic material primarily composed of carbohydrates, proteins and lipids. Developing a biorefinery model for conversion of BSG into value-added products is a plausible idea. Previous work optimized the pretreatment of BSG with the ionic liquid [N1112OH][Gly] and further release of fermentable sugar-containing solutions by enzymatic hydrolysis, using an enzymatic cocktail obtained by solid-state fermentation of BSG with Aspergillus brasiliensis CECT 2700 and Trichoderma reesei CECT 2414. The current work ends the biorefinery process, studying the fermentation of these culture media with two LAB strains, Lactobacillus pentosus CECT 4023 and Lactobacillus plantarum CECT 221, from which the production of organic acids, bacteriocins, and microbial biosurfactants (mBS) was obtained. In addition to the bacteriocin activity observed, a mass balance of the whole biorefinery process quantified the production of 106.4 g lactic acid and 6.76 g mBS with L. plantarum and 116.1 g lactic acid and 4.65 g mBS with L. pentosus from 1 kg of dry BSG. Thus, BSG shows a great potential for waste valorization, playing a major role in the implementation of biomass biorefineries in circular bioeconomy.Agencia Estatal de Investigación | Ref. PID2020–115879RB-I00São Paulo Research Foundation | Ref. 2018/25511–1São Paulo Research Foundation | Ref. 2023/09256–0Conselho Nacional de Desenvolvimento Científico e Tecnológico (Brasil) | Ref. 408783/2021–4Conselho Nacional de Desenvolvimento Científico e Tecnológico (Brasil) | Ref. 312923/2020–1Xunta de Galicia | Ref. GPC-ED431B 2021/23Universidade de Vigo/CISU

Production of enzymatic extracts for aquafeeds by solid-state fermentation with Aspergillus ibericus of winery and olive mill wastes

[Excerpt] Introduction The replacement of fishmeal by plant ingredients in aquafeeds imposed new formulation strategies to overcome some nutritional restrictions associated with these alternative ingredients. Thus, supplementation of plant-based diets with feed additives, as exogenous enzymes and antioxidants compounds, has recently attracted increasing interest to improve feed utilization and to promote animal health. Solid state fermentation (SSF) of agro-industrial wastes has a high potential for the production of these additives, being a practical, economical, and environmentally-friendly process. Olive mill and winery wastes have valuable compounds that may be valorized thought SSF and that may be used as additives for aquafeeds. This study was conducted to optimize the production of non-starch carbohydrases through the SSF of the olive mill and winery wastes and to test its efficacy to improve the release of pentoses during digestion of a plant-based diet in European seabass.[...]Work supported by FCT PhD grant SFRH/BD/131219/2017, SFRH/BDP/114942/2016, IJFCT-POCI 01-0145-FEDER-030377 and MAR-02.01.01-FEAMP-0111info:eu-repo/semantics/publishedVersio

In vitro evaluation of the interaction between exogenous carbohydrases produced by solid-state fermentation of brewers’ spent grain and digestive enzymes

[Excerpt] Introduction The presence of non-starch polysaccharides (NSP) in most plant feedstuffs (PF) has been associated with adverse effects on carnivorous fish nutrient digestibility and, ultimately, on growth and health. The lack of enzymatic machinery and well-developed microbiota in the digestive tract of carnivorous fish for processing NSP are possible causes for such effects. A promising nutritional strategy to improve nutrient digestibility of plant-based diets is the use of exogenous enzymes (e.g. carbohydrases, proteases) as feed additives. Solid-state fermentation (SSF) is an eco-friendly and cost-effective biotechnology process that allows converting inexpensive agro-industrial by-products into added-value products such as carbohydrases. As a low-cost and lignocellulosic by-product of the brewing industry, brewery spent grain (BSG) is an attractive substrate for microbial enzyme production by SSF. The present study was designed to assess the potential of a carbohydrase enzyme extract obtained by SSF to release total amino acids (AA) and monosaccharides (pentoses) from plant-based diets in an in vitro gastrointestinal model with enzyme extracts of European sea bass (Dicentrarchus labrax).[...]Supported by POCI-01-0145-FEDER-030377 and MAR-02.01.01-FEAMP-0111. HF and CC supported by grants SFRH/BD/131219/2017; SFRH/BPD/114942/2016, respectively.info:eu-repo/semantics/publishedVersio

Novel fish feed supplements: giving value to agroindustrial wastes

Sustainable aquaculture requires the reduction of fish meal use in diets, by replacing it with ecofriendly ingredients, as that vegetal feedstuff not desirable for human consumption. However, these pose challenges due to the presence of antinutritional factors. The supplementation with some supplements, like exoenzymes and antioxidants, has shown potential to reduce their nutritional drawbacks. The solid-state fermentation (SSF) of agro-industrial wastes may be used to obtain such compounds. The goal of this work was to test the applicability of an SSF-extract, obtained through the SSF of an optimized mixture of three agro-industrial wastes with Aspergilus ibericus (30% EGM, 36% VTS, 34% EOP), determined previously with a simplex-centroid mixture design. The activity of this extract was measured to be xylanase: 1866.9 U/g (lyophilized extract); cellulose: 1563.6 U/g; β-glucosidase: 736.455 U/g, and total antioxidants: 439.4 μmol Trolox equivalents/g lyophilized extract; total phenols: 28.1 mg caffeic acid equivalents/g lyophilized extract. The efficiency of this extract as a feed additive in plant-based diets evaluated by an in vitro digestibility trial, analyzing its efficiency to improve the release of pentoses and antioxidants during digestion. The extract was used to supplement diets for Dicentrarchus labrax, with increasing levels of extract: 0, 4 and 8 U cellulase/g diet. Dietary supplementation with the extract led to the increase of pentose release, during alkaline digestion. The effect of the supplementation with the SSF extract on the release of antioxidants and phenols compounds are being evaluated and will be presented.Project SPO3 (ref. POCI-01-0145-FEDER-030377; FEDER-Operational Programme Competitiveness and Internationalization and FCT); project InovFeed (ref. MAR-02.01.01-FEAMP0111; Programa Operacional Mar2020); BioTecNorte (NORTE-01-0145-FEDER-000004; ERDF under the scope of Norte2020-Programa Operacional Regional do Norte)info:eu-repo/semantics/publishedVersio

Evaluation of biotechnological processing through solid-state fermentation of oilseed cakes on extracts bioactive potential

Oilseed cakes (OC) are natural sources of lignocellulosic biomass, produced every year in large amounts. In addition to their main applications as animal feed, plant or soil fertilizer, and compost, they present enormous potential for being used in biotechnological processes for the obtainment and extraction of valuable bioactive compounds. This work evaluated the effect of solid-state fermentation on the bioactive properties of extracts obtained from the bioprocessing of OC and evaluated the effect of solvents on the recovery of compounds with higher bioactive potential. A general decrease of EC50 values was observed for fermented extracts obtained using a mixture of water/methanol (1:1) as extraction solvent. A decrease in the minimum inhibitory concentration was observed for fermented water extracts compared to non-fermented. Additionally, growth inhibition of Listeria monocytogenes was observed when using aqueous methanolic fermented extracts. These extracts also exhibited a higher percentage of growth reduction against phytopathogenic fungi, and some extracts exhibited increased protection against genotoxic agents such as camptothecin and bisphenol A. It was demonstrated that bioprocessing of OC through SSF is an effective approach to obtaining valuable compounds with bioactive properties for use in the food, pharmaceutical or cosmetic industries.Open access funding provided by FCT|FCCN (b-on). The authors thank the Portuguese Foundation for Sci ence and Technology (FCT) under the scope of the strategic funding of UIDB/BIO/04469/2020 and UIDB/04033/2020 units. Daniel Sousa acknowledges the fnancial support pro vided by national funds through FCT(PD/BD/135328/2017), under the Doctoral Program “Agricultural Production Chains – from fork to farm” (PD/00122/2012) and from the European Social Funds and the Regional Operational Programme Norte 2020.info:eu-repo/semantics/publishedVersio

Valorization of brewers spent grain using biological treatments and its application in feeds for European seabass (Dicentrarchus labrax)

Brewers spent grain (BSG) is the main brewery industry by-product, with potential applications in the feed and food industries due to its carbohydrate composition. In addition, the lignocellulosic nature of BSG makes it an adequate substrate for carbohydrases production. In this work, solid-state fermentation (SSF) of BSG was performed with Aspergillus ibericus, a non-mycotoxin producer fungus with a high capacity to hydrolyze the lignocellulosic matrix of the agro-industrial by-products. SSF was performed at different scales to produce a crude extract rich in cellulase and xylanase. The potential of the crude extract was tested in two different applications: -(1) - the enzymatic hydrolysis of the fermented BSG and (2) - as a supplement in aquafeeds. SSF of BSG increased the protein content from 25% to 29% (w/w), while the fiber content was reduced to 43%, and cellulose and hemicellulose contents were markedly reduced to around 15%. The scale-up of SSF from 10 g of dry BSG in flasks to 50 g or 400 g in tray-type bioreactors increased 55% and 25% production of cellulase and xylanase, up to 323 and 1073 U g1 BSG, respectively. The optimum temperature and pH of maximal activities were found to be 55°C and pH 4.4 for xylanase and 50°C and pH 3.9 for cellulase, cellulase being more thermostable than xylanase when exposed at temperatures from 45°C to 60°C. A BoxBehnken factorial design was applied to optimize the hydrolysis of the fermented BSG by crude extract. The crude extract load was a significant factor in sugars release, highlighting the role of hydrolytic enzymes, while the load of fermented BSG, and addition of a commercial -glucosidase were responsible for the highest phenolic compounds and antioxidant activity release. The lyophilized crude extract (12,400 and 1050 U g1 lyophilized extract of xylanase and cellulase, respectively) was also tested as an enzyme supplement in aquafeed for European seabass (Dicentrarchus labrax) juveniles. The dietary supplementation with the crude extract significantly improved feed and protein utilization. The processing of BSG using biological treatments, such as SSF with A. ibericus, led to the production of a nutritionally enriched BSG and a crude extract with highly efficient carbohydrases capable of hydrolyzing lignocellulosic substrates, such as BSG, and with the potential to be used as feed enzymes with remarkable results in improving feed utilization of an important aquaculture fish species.This study was supported by the project “SPO3-Development of innovative sustainable protein and omega-3 rich feedstuffs for aquafeeds, from local agro-industrial by-products”, reference POCI-01-0145-FEDER-030377, funded by the European Regional Development Fund (ERDF) and Portuguese Foundation for Science and Technology (FCT) and by the strategic funding of the UIDB/04469/2020 unit. HF and CC were supported by grants SFRH/BD/131219/2017 and SFRH/BPD/114942/2016, respectively, from FCT, MCTES, FSE, and UE under the North Portugal Regional Operational Program (NORTE 2020). José Manuel Salgado was supported by the grant CEB/N2020—INV/01/2016 from Project “BIOTECNORTE—Underpinning Biotechnology to foster the north of Portugal bioeconomy” (NORTE-01-0145-FEDER-000004).info:eu-repo/semantics/publishedVersio

Effect of biological and physical pre-treatments of Ulva rigida in the quality of on-growing European seabass Dicentrarchus labrax

[Excerpt] Content A growth trial with European seabass was performed to assess the effect of the dietary inclusion of Ulva rigida as is or with technological pre-treatment with ultrasounds (US) or solid-state fermentation (SSF) with Aspergillus ibericus. Promising results of the pre-treatment of U. rigida prior to dietary inclusion on growth performance and feed utilization efficiency of seabass were obtained, however, the effect of these treatments on fillet nutritional and sensory quality was not yet evaluated.[...]Supported by PhD grant by FCT ref.SFRH/BD/131219/2017, IJFCT-POCI-01-0145-FEDER-030377 and MAR-02.01.01-FEAMP-0111info:eu-repo/semantics/publishedVersio