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.

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.

Enhancing the saccharification of pretreated chestnut burrs to produce bacteriocins.

The present study aims to valorize chestnut burrs, an important lignocellulosic waste, through a biorefinery concept. A solid residue rich in glucan (41.36 ± 0.59 %) and lignin (39.06 ± 0.01 %) obtained from a previous process of pre-hydrolysis was subjected to four treatments with water or NaOH to enhance enzymatic hydrolysis. Saccharification was performed using different ratios of commercial cellulases and ß-glucosidases and at controlled pH 4.8 or 6.0 (with citrate buffer) or uncontrolled pH. Carbohydrate-rich solutions with or without nutrients were used to produce bacteriocins by Lactobacillus plantarum CECT 211. The use of NaOH at high temperatures (120 and 130 °C) was the most suitable treatment to improve saccharification. Regarding the production of bacteriocins, the best result was obtained using the enzymatic solution obtained at controlled pH 6.0, supplemented with MRS broth nutrients (except glucose). Thus, the concentrations of bacteriocins obtained in this culture medium (9.21 BU/mL) was 1.22 and 1.98 times higher than those obtained in the nutrient supplemented medium buffered at pH 4.8 (7.56 BU/mL) and in the commercial MRS broth (4.65 BU/mL), respectively. These results highlight the feasibility of the technology developed in this work.

Valorization of chestnut (Castanea sativa) residues: Characterization of different materials and optimization of the acid-hydrolysis of chestnut burrs for the elaboration of culture broths.

Four kinds of waste from the industrial processing of chestnuts (Castanea sativa), namely leaves, pruned material and burrs from chestnut tree plus chestnut shells, were characterized to determine their content in polymers and thus their potential use in biorefinery processes. Results revealed that chestnut burrs have the highest polysaccharide content being the most promising for carrying out the subsequent stages of acid hydrolysis. Treatment with diluted sulfuric acid (prehydrolysis) allowed the solubilization of xylose, glucose and arabinose, but also some toxic compounds such as furan derivatives, aliphatic acids and phenolic constituents. Xylose, the main component released in the hemicellulosic hydrolyzates, was maximized by using a 3**(2-0) full factorial design combined with desirability function. At optimum conditions set at 130 °C and 3% (w/v) H2SO4, this value was 22.6 g L-1 xylose. Three concentrations of activated charcoal (1, 2.5 and 5% w/v) were evaluated to remove certain unwanted byproducts, and it was found that under the highest dosage, 95.27 ±â€¯0.03% of the color was removed with an almost total reduction of furan derivatives, making this liquor an appropriate basis for the development of suitable culture media for lactic acid bacteria. To validate this hypothesis three lactic acid bacteria, namely Lactobacillus plantarum, Lactobacillus pentosus and Lactococcus lactis were positively tested finding lactic acid yields of 0.89, 0.92 and 0.83 g/L·h respectively.

Enzymatic hydrolysis of brewer's spent grain to obtain fermentable sugars.

Lignocellulosic biomass is a feedstock with the potential to be converted into value-added bioproducts. The use of enzymatic hydrolysis allows the cleavage of lignocellulose into their monomeric units, but there are some drawbacks that make its use in industrial biocatalysis unfeasible. In the present study, we describe the hydrolysis of brewer's spent grain (BSG) with an enzymatic cocktail produced by Aspergillus niger CECT 2700 and its comparison with commercial enzymes. In addition, it was determined whether pretreating the BSG (non-pressurized alkaline hydrolysis or treatment with cholinium glycinate ionic liquid) is necessary. Results show that both pretreatments enhanced xylose release (10.55 ±â€¯0.07 g/L and 8.14 ±â€¯0.13 g/L respectively), meanwhile the hydrolysis of raw BSG with the enzymatic cocktail produced solutions containing high levels of glucose (18.45 ±â€¯1.66 g/L) and xylose (6.38 ±â€¯0.26 g/L).

Biorefining brewery spent grain polysaccharides through biotuning of ionic liquids.

Brewery spent grain (BSG), a relevant waste from beer industry mainly composed of polysaccharides and lignin, is experiencing a surge in the production with its associated environmental impact. Thus, this manuscript bets in the application of aqueous solutions of a cholinium-based ionic liquid (IL) containing glycinate as anion ([N1112OH][Gly]) for an efficient delignification pretreatment. The operation at 90 °C yielded drastic lignin reduction (75.89%), greater than the levels attained when a traditional imidazolium-based IL (1-ethyl-3-methylimidazolium acetate, [C2C1im][C1COO]), was used (40.18%). The advantages of this pretreatment positively impacted the subsequent saccharification reaction, as the levels were increased up to about 1.5 times regarding the control (no IL) or the imidazolium-based pretreatment. ATR-FTIR spectrometry and scanning electron microscopy turned out to be useful tools to monitor the structural changes exerted. The results presented in this work make up the basis for a rational design of bio-ILs for delignification of lignocellulosic materials.

Production of a highly concentrated probiotic culture of Lactococcus lactis CECT 539 containing high amounts of nisin.

In this study, probiotic biomass and nisin productions by Lactococcus lactis CECT 539 were followed in two realkalized fed-batch cultures in diluted whey (DW) supplemented with KH2PO4 up to a total phosphorus concentration of 0.459 g/L. Increased biomass (5.12 g/L, 2.18 × 1010 CFU/mL) and nisin (235.23 BU/mL) concentrations were obtained in the culture fed concentrated whey and concentrated mussel processing waste (CMPW) medium supplemented with glucose up to a concentration of 400 g/L (CMPW + G medium) compared with similar fed-batch fermentations in DW medium. In the second fed-batch fermentation, the feeding medium CMPW + G was supplemented with KH2PO4 up to a TP concentration of 3.21 g/L. With this approach, increased production of biomass (5.49 g/L, 2.33 × 1010 CFU/mL) and nisin (257.59 BU/mL) was obtained. Considering the substantial availability of these wastes at very low prices from local dairy and mussel processing plants in Galicia, their use as culture media could offer an attractive alternative for a low-cost production of probiotic biomass and nisin at a high scale.

Fed-batch production of vanillin by Bacillus aryabhattai BA03.

Bacillus aryabhattai BA03, a strain isolated in our laboratory, has interesting properties related to the production of natural aromas and flavors. Specifically, we have found that it was able to produce vanillin from ferulic acid (FA). Furthermore, this strain produces high amounts of 4-vinylguaiacol in only 14h, this being the only intermediate metabolite observed in the process. FA is an inexpensive feedstock for the production of natural value-added compounds when extracted from lignocellulosic wastes. In this study, we optimized the operational conditions (temperature, pH and agitation), medium composition and bioconversion technology (batch or fed-batch) to produce vanillin. In a fed-batch process conducted with just one additional supplementation after 24h, the maximal concentration of vanillin (147.1±0.9mg/L) was observed after 216h (QV=0.681mg/Lh; YV/fFA=0.082mg/mg) after degrading 90.3% FA. In view of our data, we postulate that Bacillus aryabhattai BA03 carries out a decarboxylation of ferulic acid as a metabolic pathway.