3.2 Protein solubility
The protein solubility is one of the most important physicochemical properties, as it is generally related to other functional properties, which may affect the rheological, hydrodynamic, and surface activity characteristics (Zayas, 1997). As shown in Table 2, the AE-IP CP control had a significantly higher (p <0.05) solubility at pH 3 where it was 88.9% soluble protein compared to the defatted CP and HE controls (76.2% and 81.4%, respectively). Similar to the results at pH 3, at pH 5 the AE-IP CP control showed higher solubility (53.5%) than the defatted CP (36.9%) and HE (2.0%) controls, whereas at pH 7 the HE control (70.0%) had a higher solubility than both the CP control (27.7%) and the defatted CP control (37.8%). As for the SE products, all of the controls had a higher solubility (≥94%) at pH 3 than the controls produced by AE-IP. At pH 5, a higher solubility was found for the SE HE control (29.4%) compared to AE-IP (2.0%), whereas it was lower than the SE CP control (37.7%) and defatted CP control (37.3%), which had similar solubility values. At pH 7, both the SE CP (51.8%) and defatted CP control (49.3%) had a higher solubility than the respective AE-IP product however, the SE HE control (43.2%) was less soluble than the AE-IP HE control. Overall, the SE products showed higher solubility at pH 3 and 7. Stone et al. (2015) also found a lower solubility of canola protein products obtained by AE-IP compared to the SE process. Similar solubility values were found for both the CP and defatted CP controls at every pH and tended to be higher than the HE control.
The canola protein products prepared from fermented meals showed a lower solubility at each pH compared to the control products. This is hypothesized to be due to partial hydrolysis of protein during fermentation which led to the exposure of previously buried hydrophobic groups. At pH 3, all AE-IP and SE controls showed higher solubility than products from fermented meals. However, the solubility of products fromA. oryzae fermented meals was significantly higher than products from A. niger fermented meals. This indicated that fermentation with A. oryzae was able to maintain the solubility (pH 3) at a relatively higher level than A. niger after SSF. As such, usingA. oryzae may be favorable to obtain a more soluble protein product at pH 3. After fermentation, most extracted samples showed significant decreases (p <0.05) in solubility compared to the controls at pH 5 and had overall low solubility values ranging from 4.5% to 11.8%. However, there was an increase in solubility at pH 5 from 2.0% for the AE-IP HE control to 11.8% and 5.8% when fermented using A. niger and A. oryzae , respectively. According to a previous study, the isoelectric point at pH 6.2 and low solubility at both pH 5 (~37%) and pH 7 (~23%) were reported for a SE canola protein isolate produced from unfermented HE meal (Chang et al. , 2015). Our results indicated that SSF processing decreased the protein solubility at pH 5 which may be due to a change in the isoelectric point (close to pH 5) by partial hydrolysis of the proteins, causing the shift in solubility with pH value.
At pH 7, the solubility of AE-IP canola protein products showed significantly lower (p <0.05) solubility than SE products obtained from CP control meals. The inverse was found for the products extracted from the HE control meal. Unlike at pH 3 and 5, the SSF showed the ability to improve the protein solubility of the extracted products at pH 7. For instance, the AE-IP product from theA. niger fermented CP meal showed a significant (p<0.05) increase in solubility (to 47.5%) compared to both the AE-IP CP controls. In addition, the same level of solubility (70.0%) was found for the AE-IP HE control and the HE product from theA. niger fermented meal. However, there was a decrease in solubility (p <0.05) for protein products extracted (AE-IP) from A. oryzae fermented CP and HE meals compared to the controls, which indicated that A. niger was preferred to maintain or increase the solubility of AE-IP protein products at pH 7. Some opposite results were found for SE protein products, as products fromA. oryzae fermented meals resulted in higher solubility values (90.7% for A. oryzae CP and 55.6% for A. oryzae HE) compared to the controls (p<0.05). An increase in the solubility of the SE product from A. niger fermented HE meal (48.5%) was also observed compared to the HE control (p<0.05). However, for the CP meals a decrease in solubility was reported when using SE on the A. niger fermented meal (37.2%) compared to the CP controls. When extracting proteins via SE, both A. niger and A. oryzae can be acceptable inoculums for meals to maintain or increase the protein solubility at pH 7, whereas A. niger might be a better culture choice for AE-IP products at pH 7.
The results above indicate the differences between AE-IP and SE canola protein (protein fraction, the percentage of napin and cruciferin) and the hydrolysis mechanism of A. niger and A. oryzaeincluding possible different proteinase, length of peptides, structure of hydrolyzed protein, and synthesized metabolism. The protein products extracted using SE showed higher solubility than the AE-IP products mainly due to the low content of non-protein compounds, possible differences in protein fraction, and possible interactions of protein and carbohydrate in the AE-IP products. In addition, the protein products extracted from CP meals showed higher solubility than products from the HE meal. Protein denaturation during the hexane-extraction (heat treatment) could explain the solubility reduction. Heat treatments could result in the exposure of hydrophobic groups, which contribute to the reduction in protein solubility (Khattab & Arntfield, 2009). The partial denaturation of proteins can alternate the balance of protein hydrophobicity/hydrophilicity and further affect solubility (Moureet al. , 2006).