1 INTRODUCTION
Canola meal, the by-product of the canola oil extraction process, is gaining tremendous interest in the food industry due to its high protein content (35%-45%) and low cost. The use of canola meals as an alternative protein source can provide more protein options to the plant-based food industry. Canola proteins are dominated by two major proteins, a salt-soluble globulin protein (cruciferin, 12S , MW ~300 kDa) and a smaller water-soluble albumin (napin, 1.7-2S , MW ~14–17 kDa). The amino acid composition of canola meal is well balanced with a relatively high protein efficiency ratio (PER) of 2.64 and therefore can be used for human nutrition (Aider & Barbana, 2011). In recent research, canola proteins have presented high solubility, foaming capacity and stability, and comparable or moderate emulsifying properties compared to other plant proteins such as soybean and pea protein (Chang et al. , 2015; Cheung et al. , 2014; Khattab & Arntfield, 2009; Tan et al. , 2011a; Wu &Muir, 2008). These characteristics make canola protein a potential ingredient for the food industry.
Canola oil can be extracted by hexane extraction (HE) or cold-pressing (CP). The conventional way, HE, is a chemical process involving the use of non-polar solvents, e.g., hexane. During HE, the seeds are ground up and washed with hexane under controlled conditions to release oil molecules stored within. A desolventizing step is followed to remove the hexane by distilling at approximately 100-110oC, after which the solvent is recycled and reused (Cheng et al., 2019). The HE process efficiently produces a high yield of oil, however, the use of chemicals and the application of heat lead to considerate damage to the proteins within the meal, resulting in reduced functionality (e.g., emulsifying properties) (Östbring et al., 2019). The CP technique, on the other hand, solely utilizes mechanical forces (e.g., crushing) without the application of organic solvents or high heat for their subsequent removal. The seeds are pressed at low temperatures (<40oC), leaving nutrients in the remaining meal in a less disturbed form compared to HE, however, the oil yield was lower in HE with a high level of residual oil (6-20%) (Hickling, 2007; Östbring et al., 2019).
Fermentation is the process accomplished by the metabolism of microorganisms that catalyze nutrients, synthesize secondary metabolites, and complete other physiological activities under anaerobic or aerobic conditions. Solid-state fermentation (SSF) is a popular process to modify the functionality and nutritional composition of protein ingredients. As opposed to submerged fermentation (SmF), there’s no free water in SSF, making it suitable for microorganisms that don’t require high moisture or high water activity to grow, for example, fungi. Recently, SSF has been applied to canola meals to improve their quality. Pal Vig and Walia (2001) used Rhizopus oligosporus as the fermenting culture to produce a high-protein product from HE canola meal. The results showed a significant decline in contents of glucosinolates (GLS) (~43%), thiooxazolidones (~31 %), phytic acid (~42%) and fibre (~26%) along with a ~65% increase in crude protein after 10 days. Croat et al. (2016) studied the use of SSF on HE and CP canola meals to modify their nutritional composition. The strains Trichoderma reesei , Aspergillus pullulans , and A. pullulans improved protein content by 22.9, 16.9 and 15.4%, while reducing the total GLS content from 60.6 to 1.0, 3.2 and 10.7 μmol/g, respectively. Significantly higher content of dry matter yield was reported for the HE meal compared to the CP meal, which is mainly due to the high oil residues left by CP. Much work has been done on enhancing the nutritional quality of canola meals, however, the use of SSF to alter the functionality of proteins in HE or CP canola meals has been limited.
Several methods, including alkaline extraction-isoelectric precipitation (AE-IP), salt extraction-dialysis (SE), protein micellation method (PMM), and low pH extraction combined with membrane separation and ultrafiltration (UF), have been widely used for the production of canola protein isolates (Tan et al. , 2011a, 2011b; Wanasundara, 2011). The structural composition and functionality of the canola proteins may vary significantly depending on the extraction method used (Can Karacaet al. , 2011; Hoglund et al. , 1992; Wu & Muir, 2008). In addition, the extraction of canola proteins is especially difficult due to the differences in protein fractions (widely differing isoelectric points of pH 4-11 and molecular weights of 13-320 kDa) and the presence of antinutrients (GLS, phytic acid, and polyphenols), pigments, and fibre in the canola meal (Wu & Muir, 2008). It is expected that pre-treatment, such as fermentation, and specific purification processes may improve protein solubility in the meals and consequently protein extractability.
In our previous study, both CP and HE canola meals had a ~45% degree of protein hydrolysis upon 72 hours of fermentation with Aspergillus niger NRRL 334 andAspergillus oryzae NRRL 5590. In this study, those fermented canola meals were used in the wet fractionation process to produce protein products. Two wet extraction processes, AE-IP and SE, were employed. The resulting protein products were characterized for select functional properties and compared with protein products extracted from meals that were not pre-treated by fermentation. We hypothesize that pre-treatment with SSF has the potential to improve the protein extractability from canola meal because of partial protein hydrolysis and the loosening of protein-carbohydrate interactions while also modifying protein functionality. Moreover, the CP meals are expected to yield protein products with higher functional properties than those processed by HE due to the lack of heat and chemical damage to the proteins. The findings of this work will provide a relatively comprehensive view of the effect of SSF of canola meals on the functionality of extracted proteins with direct comparisons between the test fungal strains, meal type, and protein fractionation techniques.