Jorge A. Ferreira

The existing recycling methods of PHA-based material are ineffective in terms of increasing resource efficiency and the production of high value end-of-life products. Therefore, in this study, a novel approach of acidogenic fermentation was proposed to recycle PHB-based composites reinforced with natural fibers such as cellulose, chitin, chitosan, orange waste, sawdust, soy protein, and starch. The inclusion of cellulose, chitosan, and sawdust improved the impact properties of the composites while other fillers had various effects on the mechanical properties. These three composites and neat PHB were subsequently subjected to biological degradation via acidogenic digestion to determine the possibility of converting PHB-based composites into volatile fatty acids (VFAs). Two different pH levels of 6 and 10 were applied to assess the effect of pH on the bioconversion and inhibition of the methanogenesis. The results showed promising PHB degradation, contributing to considerable VFA production of 2.5 g/L at pH 6 after 47 days. At pH 6, the presence of the natural fibers in the biocomposites promoted the degradation rate. On the contrary, pH 10 proved to be more suitable for the degradation of the fibers. The VFA which is produced can be recirculated into PHB production, fitting with the concept of a circulating bioeconomy.

The glucan-rich fraction, hemicellulosic compounds-rich fraction, and a mixture of both fractions obtained from organosolv pretreatment of oil palm empty fruit bunch (OPEFB) were used as substrates to produce volatile fatty acids (VFAs) in acidogenic fermentation. In this study, the effects of medium adjustment (carbon to nitrogen ratio and trace elements supplementation) and methanogenesis inhibition (through the addition of 2-bromoethanesulfonate or by heat shock) to enhance VFAs yield were investigated. The highest VFA yield was 0.50 ± 0.00 g VFAs/g volatile solid (VS), which was obtained when methanogens were inhibited by heat shock and cultivated in a mixture of glucan-rich and hemicellulosic compounds-rich fractions. Under these conditions, the fermentation produced acetic acid as the only VFA. Based on the results, the mass balance of the whole process (from pretreatment and fermentation) showed the possibility to obtain 30.4 kg acetic acid and 20.3 kg lignin with a 70% purity from 100 kg OPEFB.

The rapid growth of aquaculture and the lack of fish meal demand new sustainable ingredients. Although fungal biomass is found to be a promising sustainable fish feed supplementation candidate, the characteristics of this protein-rich source are closely influenced by the quality of the applied growth medium. In this work, the nutritional properties of pure filamentous fungal biomass provided from the cultivation of Aspergillus oryzae, Neurospora intermedia and Rhzopus oryzae were evaluated to assess their potential as alternative novel protein sources in fish feed. In this regard, fungal biomass yields of up to 0.19 ± 0.005 (g dry biomass/g substrate glucose) were obtained during submerged cultivation of fungal strains. The pure fungal biomass acquired could contain significant amounts of protein up to 62.2 ± 1.2% (w/w). The obtained protein had a high quality with notable inclusion of essential amino acids such as lysine, arginine, methionine and threonine with comparable concentrations to those of fish meal. Fungal biomass is mainly considered as protein source, however, entitlement of 6.9 ± 0.5, 4.0 ± 0.7 and 17.2 ± 1.1% (w/w) of lipids and ratio of polyunsaturated fatty acids (PUFA) to saturated fatty acids (SFA) of 1.37:1, 1.74:1 and 1.47:1 in A. oryzae, N. intermedia and R. oryzae, respectively, signal health benefits for the fish. Considering the results, protein-rich pure fungal biomass with amino acid composition is greatly compatible with fish meal, and contains essential nutrients such as fatty acids and minerals. This pure biomass constitutes a promising sustainable alternative supplement to be introduced in fish feed industry.

Filamentous fungi can be used for the valorization of food waste as a value-added product. The goal of this study was the valorization of bread waste through fungal cultivation and the production of value-added products. The fungal cultivation was verified for upscaling from shake flasks to a bench-scale bioreactor (4.5 L) and a pilot-scale bioreactor (26 L). The fungus showed the ability to grow without any additional enzymes or nutrients, and it was able to consume a bread concentration of 4.5% (w/v) over 48 h. The biomass concentration in the shake flasks was 4.1 g/L at a 2.5% bread concentration, which increased to 22.5 g/L at a 15% bread concentration. The biomass concentrations obtained after 48 h of cultivation using a 4.5% bread concentration were 7.2–8.3 and 8.0 g/L in 4.5 and 26 L bioreactors, respectively. Increasing the aeration rate in the 4.5 L bioreactor decreased the amount of ethanol produced and slightly reduced the protein content of the fungal biomass. The initial protein value in the bread was around 13%, while the protein content in the harvested fungal biomass ranged from 27% to 36%. The nutritional value of the biomass produced was evaluated by analyzing the amino acids and fatty acids. This study presents the valorization of bread waste through the production of a protein- and fatty-acid-rich fungal biomass that is simultaneously a source of microfibers.

High production costs still hamper fast expansion of commercial production of polyhydroxyalkanoates (PHAs). This problem is greatly related to the cultivation medium which accounts for up to 50% of the whole process costs. The aim of this research work was to evaluate the potential of using volatile fatty acids (VFAs), derived from acidogenic fermentation of food waste, as inexpensive carbon sources for the production of PHAs through bacterial cultivation. Bacillus megaterium could assimilate glucose, acetic acid, butyric acid, and caproic acid as single carbon sources in synthetic medium with maximum PHAs production yields of 9–11%, on a cell dry weight basis. Single carbon sources were then replaced by a mixture of synthetic VFAs and by a VFAs-rich stream from the acidogenic fermentation of food waste. After 72 h of cultivation, the VFAs were almost fully consumed by the bacterium in both media and PHAs production yields of 9–10%, on cell dry weight basis, were obtained. The usage of VFAs mixture was found to be beneficial for the bacterial growth that tackled the inhibition of propionic acid, iso-butyric acid, and valeric acid when these volatile fatty acids were used as single carbon sources. The extracted PHAs were revealed to be polyhydroxybutyrate (PHB) by characterization methods of Fourier-transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC). The obtained results proved the possibility of using VFAs from acidogenic fermentation of food waste as a cheap substrate to reduce the cost of PHAs production. GRAPHICAL ABSTRACT

Brewer’s spent grain (BSG) is the main solid by-product of the brewing sector. High moisture and nutrient-rich content render BSG easily perishable, leading to waste generation and environmental impacts. BSG has narrow applications in both feed and food sectors due to its composition including high fiber and low protein. Therefore, a processing strategy leading to the nutritional valorization of BSG could widen its applications. In this study, submerged cultivation of edible filamentous fungi (Aspergillus oryzae, Neurospora intermedia, and Rhizopus delemar) was introduced as a strategy to enhance the protein content of BSG. The growth of all strains in BSG increased the protein content of the fermented BSG. The highest increase of protein content (from 22.6% to 34.6%), was obtained by cultivation using A. oryzae and medium supplementation. The protein content increase was followed by a decrease in the content of polysaccharides (up to ca. 50%), namely starch, glucan, xylan, and arabinan. The addition of cellulase resulted in enhanced ethanol production from BSG but led to lower concentration of recovered solids. In conclusion, simple processing of BSG using edible filamentous fungi can lead to quality improvement of BSG, providing potential economic and environmental benefits to the brewing sector.

The feasibility of dry-grind bioethanol plants is extremely dependent on selling prices of ethanol and by-products, known as Dried distillers grains with solubles (DDGS), and sold as animal feed. Increasing the amount and quality of the by-products can widen potential feed and food markets and improve the process economy and robustness to price fluctuations of ethanol and grain. In this study, the techno-economic analysis of a bioethanol plant was investigated. Integration of edible filamentous fungi into the process leading to the conversion of sidestreams into ethanol and protein-rich fungal biomass for food and feed applications was considered, and its impact was investigated. Sensitivity analysis considered variations on process capacity, on the price of grain and ethanol, and on the price of fungal biomass considering its use for various animal feed (e.g., pig and fish) and human food markets. Selling the fungal biomass in the human food market resulted in 5.56 times higher NPV (net present value) than the base case bioethanol plant after 20 years. Integration of a low-performing strain towards ethanol, followed by the usage of the fungal biomass in the food sector, was found to be the most resistant scenario to the low ethanol selling price and increasing grain price. This study showed that the competitiveness of ethanol plants in the fuel market could be reinforced while meeting the increasing demand for protein sources.

Olive oil mills represent an important sector in the Mediterranean Sea Basin but also an environmental hazard due to untreated wastewater. Recovery of nutrients from olive oil mill wastewater (OMWW) as protein-rich microbial biomass can produce novel feed and reduce its chemical oxygen demand; however, low-protein containing products have been reported. New strategies leading to higher protein-containing fungal biomass could renew the research interest on bioconversion for pollution mitigation of OMWW. In this work, through cultivation of edible filamentous fungi (Aspergillus oryzae, Neurospora intermedia, and Rhizopus delemar), a link between the protein content in the originated fungal biomass, and the addition of nitrogen and medium dilution was established. Addition of nitrogen in the form of NaNO₃ reduced the cultivation time from 96 h to 48 h while achieving a similar biomass mass concentration of 8.43 g/L and increased biomass protein content, from w = 15.9% to w = 29.5%. Nitrogen addition and dilution of OMWW, and consequent reduction of suspended solids, led to an increase in the protein content to up to w = 44.9%. To the best of our knowledge, the protein contents achieved are the highest reported to date and can open new research avenues towards bioconversion of OMWW using edible filamentous fungi.