Production of bovine beta-lactoglobulin and hen egg ovalbumin by Trichoderma reesei using precision fermentation technology and testing of their techno-functional properties.

The food protein ingredient market is dominated by dairy and egg proteins. Both milk whey and egg proteins are challenging proteins to replace, e.g. with plant proteins, due to the unique structural features of the animal proteins that render them highly functional. Thus, to provide a non-animal source of these important proteins the fungal host Trichoderma reesei was utilized for the biotechnical production of recombinant hen ovalbumin (TrOVA) and bovine beta lactoglobulin (TrBLG). These food proteins were investigated using two different promoter systems to test the concept of effectively expressing them in a fungal host. Both proteins were successfully produced in 24 well plate and bioreactor scale. The production level of TrBLG and TrOVA were 1 g/L and 2 g/L, respectively. Both proteins were further purified and characterized, and their functional properties were tested. TrBLG and TrOVA secondary structures determined by circular dichroism corresponded to the proteins of bovine and hen. The T. reesei produced proteins were found to be N-glycosylated, mostly with Man 5. TrBLG had emulsification properties matching to corresponding bovine protein. TrOVA showed excellent foaming characteristics and heat-induced gelation, although the strength of the gel was somewhat lower than with hen ovalbumin, possibly due to the partial degradation of TrOVA or presence of other host proteins. Biotechnical production of whey and egg proteins using precision fermentation technology offers an innovative way to increase the sustainability of the conventional food industry, without further reliance on animal farming. Industrial relevance: The food protein ingredient market is dominated by dairy (largely whey proteins) and egg proteins. Whey proteins are valuable and versatile food ingredients due to their functional and nutritional quality. They are largely used in meat and milk products, low fat products, bakery, confectionary, infant formulas and sports nutrition. Similarly, egg white protein ovalbumin is a highly functional protein ingredient that facilitates structure formation and high nutritional quality in most food products. Together they comprise 40-70% of the revenue in the animal protein ingredients market. Both whey and egg proteins are extremely challenging proteins to replace, e.g., by plant proteins due to their unique structural features that render them with high functionality. Biotechnical production of whey and egg proteins using precision fermentation technology offers an innovative way to increase the sustainability of the conventional food industry, without further reliance on animal farming.

Grains - a major source of sustainable protein for health.

Cereal grains are the main dietary source of energy, carbohydrates, and plant proteins world-wide. Currently, only 41% of grains are used for human consumption, and up to 35% are used for animal feed. Cereals have been overlooked as a source of environmentally sustainable and healthy plant proteins and could play a major role in transitioning towards a more sustainable food system for healthy diets. Cereal plant proteins are of good nutritional quality, but lysine is often the limiting amino acid. When consumed as whole grains, cereals provide health-protecting components such as dietary fiber and phytochemicals. Shifting grain use from feed to traditional foods and conceptually new foods and ingredients could improve protein security and alleviate climate change. Rapid development of new grain-based food ingredients and use of grains in new food contexts, such as dairy replacements and meat analogues, could accelerate the transition. This review discusses recent developments and outlines future perspectives for cereal grain use.

Impact of ultra-fine milling and air classification on biochemical and techno-functional characteristics of wheat and rye bran.

Dry milling and air classification were applied to produce three different ingredients from wheat and rye brans. Dried and pin disc-milled brans having particle size medians of 89-131 µm were air classified to produce protein- and soluble dietary fibre-enriched hybrid ingredients (median particle size 7-9 µm) and additionally brans were ultra-finely milled (median particle size 17-19 µm). The samples were characterised in regard to their composition and techno-functional properties. In air classification, protein content increased from 16.4 and 14.7% to 30.9 and 30.7% for wheat and rye brans, which corresponded to protein separation efficiencies of 18.0 and 26.9%, respectively. Concurrently, the ratio between soluble and insoluble dietary fibre increased from 0.22 to 0.85 for wheat and from 0.56 to 1.75 for rye bran. The protein- and soluble dietary fibre-enriched wheat bran fraction showed improved protein solubility at alkaline pH when compared to pin disc- and ultra-finely-milled wheat bran, whereas less difference between the wheat ingredients was observed at native and acidic pH. The protein- and soluble dietary fibre-enriched rye bran fraction exhibited lower solubility than the pin disc- or ultra-finely-milled rye brans at all the studied pH-values. Ultra-fine milling alone decreased protein solubility and increased damaged starch content when compared to the pin disc-milled brans. Both protein enrichment and ultra-fine milling improved colloidal stability in comparison to the pin disc-milled raw materials. The lowest water and oil binding capacities were obtained for the protein-enriched fractions. Ultrasound-assisted emulsification of the protein- and soluble dietary fibre-enriched fractions and the ultra-finely-milled brans revealed no major differences in the visual quality or stability of the emulsions. The results suggest that modification of the techno-functional properties of cereal brans may be acquired via both air classification and ultra-fine milling.

Impact of Ultrasound Treatment and pH-Shifting on Physicochemical Properties of Protein-Enriched Barley Fraction and Barley Protein Isolate.

Ultrasonication alone or in combination with a pH-shifting method could be applied as means for improving the techno-functional properties and performance of barley protein ingredients in liquid food matrix. Ultrasound technology was utilised with and without pH-shifting to 3, 7 and 9 aiming at investigating their impact on primary protein structure, protein solubility, particle size and colloidal stability of an air-classified protein-enriched barley fraction and a barley protein isolate. Shifting the pH of sample dispersion to 9 followed by neutralisation to pH 7 improved protein solubility and colloidal stability of the isolate whereas it had less impact on the protein-enriched fraction. Ultrasound treatment improved both protein solubility and colloidal stability of the protein-enriched fraction at alkaline pH and particle size reduction by ultrasonication was observed at all the studied pH-values. For protein isolate, ultrasonication improved protein solubility at all pH-values and colloidal stability was improved at acidic and neutral pH whereas the sample was inherently stable at alkaline pH. The protein profiles of both ingredients remained unaffected by ultrasound treatment. The results suggest adopting ultrasonication as a promising tool for improving applicability of barley protein ingredients in liquid food systems.

Plant cells as food - A concept taking shape.

Plant cell cultures from cloudberry, lingonberry and stoneberry were studied in terms of their nutritional properties as food. Carbohydrate, lipid and protein composition, in vitro protein digestibility and sensory properties were investigated. Dietary fibre content varied between 21.2 and 36.7%, starch content between 0.3 and 1.3% and free sugar content between 17.6 and 33.6%. Glucose and fructose were the most abundant sugars. High protein contents between 13.7 and 18.9% were recorded and all samples had a balanced amino acid profile. In vitro protein digestion assay showed hydrolysis by digestive enzymes in fresh cells but only limited hydrolysis in freeze-dried samples. The lipid analysis indicated that the berry cells were rich sources of essential, polyunsaturated fatty acids. In sensory evaluation, all fresh berry cells showed fresh odour and flavour. Fresh cell cultures displayed a rather sandy, coarse mouthfeel, whereas freeze-dried cells melted quickly in the mouth. All in all the potential of plant cells as food was confirmed.