Olive mill wastewater anaerobically digested : phenolic compounds with antiradical activity

The recovery of phenolic compounds, present in the olive fruits and its by-products, has been intensively studied by the antioxidant properties. Olive mill wastewater (OMW) is a phenolic-rich industrial effluent that can be advantageously valorized by the anaerobic digestion to the methane and agricultural fertilizer productions. The objective of this work was to evaluate the antiradical activity of OMW after anaerobic digestion in order to maximize the valorization of this type of effluents. The digested flow was obtained from an anaerobic hybrid reactor treating OMW at different organic loading rates (OLR), from 3.3 to 7.1 kg COD m3 d-1. OLR rise was applied by increasing progressively the OMW volume fraction from 8 % to 83 % in the feed mixture. The input and output streams, obtained at different OMW volume fractions, were characterized in terms of antiradical activity and phenolic compounds identification and quantification. Despite of the fraction decrease on total phenolic compounds provided by OMW anaerobic digestion, the antiradical activity is still significantly high (EC50 = 3.24) in the digested effluent. Oleuropein was the main phenolic compound present in the substrate before and after anaerobic digestion (about 15 % of the initial value). Others phenolic compounds present are: gallic acid, hydroxytyrosol, tyrosol, and quercetin. These data confirmed that, after the OMW anaerobic treatment to produce biomethane, the remaining flow yet contain useful compounds with antiradical activity

Aglianico and Fiano wines obtained with an autochthonous non-Saccharomyces yeast

Non-Saccharomyces yeasts are microorganisms that play a prominent role in the fermentation dynamics, composition and flavour of wine. The principal aromatic compounds responsible for varietal aroma in wine are terpenes; of these, the monoterpenes represent the oenologically most important group in terms of volatility and odour, if they are present in a free form. The glycosidically bound forms can be converted into compounds with free odours by hydrolysis with the glycosidases produced by yeasts. We performed a screening of non-Saccharomyces yeasts present in the grapes and must of Aglianico and Fiano cultivars typical of South Italy (Irpinia), which have a high extra-cellular glycosidase activity. Among the species analysed was a strain belonging to Rhodotorula spp. that showed the highest glycosidase activity, an increased free terpene fraction and, simultaneously, little modification of the bouquet. The isolated yeast was subsequently utilized for experimental winemaking processes of Aglianico and Fiano wines. The results demonstrated that the obtained wines had a more intense floral aroma and some sweet and ripened fruit notes

Properties of an alkali-thermo stable xylanase from Geobacillus thermodenitrificans A333 and applicability in xylooligosaccharides generation

An extracellular thermo-alkali-stable and cellulase-free xylanase from Geobacillus thermodenitrificans A333 was purified to homogeneity by ion exchange and size exclusion chromatography. Its molecular mass was 44 kDa as estimated in native and denaturing conditions by gel filtration and SDS-PAGE analysis, respectively. The xylanase (GtXyn) exhibited maximum activity at 70 °C and pH 7.5. It was stable over broad ranges of temperature and pH retaining 88 % of activity at 60 °C and up to 97 % in the pH range 7.5–10.0 after 24 h. Moreover, the enzyme was active up to 3.0 M sodium chloride concentration, exhibiting at that value 70 % residual activity after 1 h. The presence of other metal ions did not affect the activity with the sole exceptions of K+ that showed a stimulating effect, and Fe2+, Co2+ and Hg2+, which inhibited the enzyme. The xylanase was activated by non-ionic surfactants and was stable in organic solvents remaining fully active over 24 h of incubation in 40 % ethanol at 25 °C. Furthermore, the enzyme was resistant to most of the neutral and alkaline proteases tested. The enzyme was active only on xylan, showing no marked preference towards xylans from different origins. The hydrolysis of beechwood xylan and agriculture-based biomass materials yielded xylooligosaccharides with a polymerization degree ranging from 2 to 6 units and xylobiose and xylotriose as main products. These properties indicate G. thermodenitrificans A333 xylanase as a promising candidate for several biotechnological applications, such as xylooligosaccharides preparation

Moving towards Biofuels and High-Value Products through Phytoremediation and Biocatalytic Processes

Phytoremediation is an eco-friendly technology that utilizes plants and plant–microbe interactions to remove a wide spectrum of organic and inorganic pollutants from contaminated environments such as soils, waters and sediments. This low-impact, environmentally sustainable and cost-effective methodology represents a valuable alternative to expensive physical and chemical approaches, characterized by secondary pollution risks, and is gaining increasing attention from researchers and popular acceptance. In this review, the main mechanisms underlying the decontamination activity of plants have been clarified, highlighting the environmental remediation in fertility and soil health. Studies have illustrated the high potential of phytoremediation coupled with green and sustainable biocatalytic processes, which together represent a non-polluting alternative for the conversion of plant biomass into renewable resources. The convenience of this technology also lies in the valorization of the bio-wastes towards biofuels, energy purposes and value-added products, contributing to an effective and sustainable circular approach to phyto-management. The strategy proposed in this work allows, with the use of totally green technologies, the recovery and valorization of contaminated soil and, at the same time, the production of bioenergy with high efficiency, within the framework of international programs for the development of the circular economy and the reduction of greenhouse carbon emissions

Plastid Transformation: New Challenges in the Circular Economy Era

In a circular economy era the transition towards renewable and sustainable materials is very urgent. The development of bio-based solutions, that can ensure technological circularity in many priority areas (e.g., agriculture, biotechnology, ecology, green industry, etc.), is very strategic. The agricultural and fishing industry wastes represent important feedstocks that require the development of sustainable and environmentally-friendly industrial processes to produce and recover biofuels, chemicals and bioactive molecules. In this context, the replacement, in industrial processes, of chemicals with enzyme-based catalysts assures great benefits to humans and the environment. In this review, we describe the potentiality of the plastid transformation technology as a sustainable and cheap platform for the production of recombinant industrial enzymes, summarize the current knowledge on the technology, and display examples of cellulolytic enzymes already produced. Further, we illustrate several types of bacterial auxiliary and chitinases/chitin deacetylases enzymes with high biotechnological value that could be manufactured by plastid transformation

Tobacco Plastid Transformation as Production Platform of Lytic Polysaccharide MonoOxygenase Auxiliary Enzymes

Plant biomass is the most abundant renewable resource in nature. In a circular economy perspective, the implementation of its bioconversion into fermentable sugars is of great relevance. Lytic Polysaccharide MonoOxygenases (LPMOs) are accessory enzymes able to break recalcitrant polysaccharides, boosting biomass conversion and subsequently reducing costs. Among them, auxiliary activity of family 9 (AA9) acts on cellulose in synergism with traditional cellulolytic enzymes. Here, we report for the first time, the production of the AA9 LPMOs from the mesophilic Trichoderma reesei (TrAA9B) and the thermophilic Thermoascus aurantiacus (TaAA9B) microorganisms in tobacco by plastid transformation with the aim to test this technology as cheap and sustainable manufacture platform. In order to optimize recombinant protein accumulation, two different N-terminal regulatory sequences were used: 5′ untranslated region (5′-UTR) from T7g10 gene (DC41 and DC51 plants), and 5′ translation control region (5′-TCR), containing the 5′-UTR and the first 14 amino acids (Downstream Box, DB) of the plastid atpB gene (DC40 and DC50 plants). Protein yields ranged between 0.5 and 5% of total soluble proteins (TSP). The phenotype was unaltered in all transplastomic plants, except for the DC50 line accumulating AA9 LPMO at the highest level, that showed retarded growth and a mild pale green phenotype. Oxidase activity was spectrophotometrically assayed and resulted higher for the recombinant proteins without the N-terminal fusion (DC41 and DC51), with a 3.9- and 3.4-fold increase compared to the fused proteins

The effect of Pleurotus ostreatus arabinofuranosidase and its evolved variant in lignocellulosic biomasses conversion

The fungal arabinofuranosidase from Pleurotus ostreatus PoAbf recombinantly expressed in Pichia pastoris rPoAbf and its evolved variant rPoAbf F435Y/Y446F were tested for their effectiveness to enhance the enzymatic saccharification of three lignocellulosic biomasses, namely Arundo donax, corn cobs and brewer's spent grains (BSG), after chemical or chemical-physical pretreatment. All the raw materials were subjected to an alkaline pretreatment by soaking in aqueous ammonia solution whilst the biomass from A. donax was also pretreated by steam explosion. The capability of the wild-type and mutant rPoAbf to increase the fermentable sugars recovery was assessed by using these enzymes in combination with different (hemi)cellulolytic activities. These enzymatic mixtures were either entirely of commercial origin or contained the cellulase from Streptomyces sp. G12 CelStrep recombinantly expressed in Escherichia coli in substitution to the commercial counterparts. The addition of the arabinofuranosidases from P. ostreatus improved the hydrolytic efficiency of the commercial enzymatic cocktails on all the pretreated biomasses

Optimization of Arundo donax Saccharification by (Hemi)cellulolytic Enzymes from Pleurotus ostreatus

An enzymatic mixture of cellulases and xylanases was produced by Pleurotus ostreatus using microcrystalline cellulose as inducer, partially characterized and tested in the statistical analysis of Arundo donax bioconversion. The Plackett-Burman screening design was applied to identify the most significant parameters for the enzymatic hydrolysis of pretreated A. donax. As the most significant influence during the enzymatic hydrolysis of A. donax was exercised by the temperature (°C), pH, and time, the combined effect of these factors in the bioconversion by P. ostreatus cellulase and xylanase was analyzed by a 33 factorial experimental design. It is worth noting that the best result of 480.10 mg of sugars/gds, obtained at 45°C, pH 3.5, and 96 hours of incubation, was significant also when compared with the results previously reached by process optimization with commercial enzymes