**3. Results and Discussion**

*3.1. Characterization of Mayonnaise Ingredients*

3.1.1. Emulsifying Properties and Protein Content

Two indexes, namely the emulsifying activity (EAI) and emulsifying stability (ESI) were used to characterize the emulsifying properties of aquafaba. The obtained EAI and ESI results and protein content in aquafaba were compared with those values for an egg yolk (Table 2).

**Table 2.** Emulsifying activity index (EAI), emulsion stability index (ESI), and protein content in aquafaba and egg yolk.


*n* = 3; SD—standard deviation; Different letters (a, b) within the same column indicate significant differences between emulsifying parameters (EAI—emulsifying activity index, ESI—emulsifying stability index) and protein content in emulsifiers (Tukey's post hoc test, *p* < 0.05).

The EAI is an oil/water interface area stabilized per unit weight of protein. As can be seen, the EAI of aquafaba (13.75 m2/g) is almost 8-fold higher than the EAI of egg yolk (1.78 m2/g). This calculated EAI result for egg yolk was significantly lower than previously published values ranging between 24.5 and 30.5 m2/g [41,42]. Moreover, Meurer et al. [29] observed a high stability of egg yolk-based emulsion during 4 days (EAI = 100%). In general, differences between the EAI results can be caused by various types and concentrations of proteins, their hydrophobicity, unfolding ability, the treatment and storage conditions of egg yolks, as well as procedures and equipment used in emulsions production. It is probable that the studied egg yolk contained a lower amount of proteins with lower solubility and structural unfolding than the egg yolk proteins investigated by other authors [41,42]. A higher EAI of aquafaba proteins could be attributed to a combination of the less compact structure and higher solubility, which enhanced the ability to form interfacial membranes around the oil droplets.

However, the ESI represents a decrease in turbidity of a diluted emulsion over time. This parameter depends on the resistance of proteins to coalescence, sedimentation, flocculation and creaming over a certain period [43]. It is noteworthy that the ESI of egg yolk-based emulsion was over 110-fold higher than the ESI of vegetable emulsion produced by aquafaba (Table 2). The EAI and ESI results obtained for aquafaba (EAI = 13.75 m2/g and ESI = 20.92 min) were comparable with those reported by other authors (EAI = 12–38.6 m2/g and ESI = 15–25 min) [25,28]. The emulsifying properties of aquafaba were highly dependent on chickpea cultivar, canning process conditions and additives [24,44]. Additionally, amounts of proteins, carbohydrates, saponins, and phenolic compounds, as well as thermal processes, affected the functional properties of aquafaba [45]. It is known that during emulsion generation, proteins reduced the interfacial tension at the oil–water interface due to the presence of hydrophobic and hydrophilic groups. Although the studied aquafaba had a significantly lower protein content (1.26%) than egg yolk (16.12%), aquafaba proteins aggregated at the water–oil interface and formed an intermolecular cohesive film with enough elasticity to stabilize emulsions.

At the same time, polysaccharides can stabilize the emulsion and prevent flocculation and coalescence [46]. Furthermore, proteins and polysaccharides present in aquafaba can be bound by bioactive substances such as saponins and phenolic compounds, causing changes in the emulsion capacity of this natural emulsifier [25]. Therefore, a low ESI value for aquafaba can be related to its high RSA determined by DPPH and ABTS methods and discussed in Section 3.1.3.

For comparison, Günal-Köro ˘glu et al. [47] observed that the EAI (13.9–41.6 m2/g) and ESI (26.1–98.6 min) for emulsions of lentil protein isolate–phenolic solutions were inversely proportional to the concentrations of gallic acid (0.05–0.25 mg/mL) and phenolic extracts

from yellow and red onion skin (0.10–0.50 mg/mL). On the other hand, the amphiphilic structure of saponins generated smaller oil droplets (ODs) during homogenization by lowering interfacial tension [48].

### 3.1.2. Fatty Acid Compositions of Vegetable Oils

The fatty acid compositions of oils used to prepare new vegan mayonnaise samples are presented in Table 3.

**Table 3.** Fatty acid compositions of oils used in making vegan mayonnaises.


*n*= 5; SD—standard deviation; Different letters (a–e) within the same column indicate significant differences between the percentages of fatty acids of vegetable oils: RRO—refined rapeseed oil, CPRO—cold-pressed rapeseed oil, CPSO—cold-pressed sunflower oil, CPLO—cold-pressed linseed oil, CPCO—cold-pressed camelina oil (Tukey's post hoc test, *p* < 0.05); SAFA—saturated fatty acids; MUFA—monounsaturated fatty acids; PUFA— polyunsaturated fatty acid.

Tukey's post hoc test indicated significant differences in the fatty acid percentages of the studied cold-pressed vegetable oils and RRO. Each investigated vegetable oil has a specific fatty acid profile depending on the plant sources. It is noteworthy that all vegetable oils contained a low amount of saturated fatty acids (SAFA = 6.9–10.6%) and a high level of monounsaturated fatty acids (MUFA = 15.7–64.6%) and polyunsaturated fatty acids (PUFA = 27.4–73.8%).

The major SAFA were palmitic acid (C16:0) and stearic acid (C18:0) which were determined in the highest concentrations in CPSO (6.6%) and CPCO (4.1%) samples, respectively. Rapeseed oils (RRO and CPRO) were the richest source of oleic acid (18:1n-9 = 62.2–63.4%), followed by linoleic acid (18:2n-6 = 19.3–19.4%).

On the contrary, linoleic acid was present at the highest level (58.9%) in CPSO, while a moderate content of oleic acid (29%) was found in this oil (Table 3). However, linolenic acid (C18:3n-3 = 30.5–57.5%) was the principal PUFA for two of the cold-pressed oils, CPLO and CPCO. Nevertheless, CPCO had an approximately two times higher C18:3 percentage than CPLO.

As can be seen, CPSO, CPLO and CPCO revealed the highest PUFA content (49.7–73.8%), whereas MUFA accounted for more than 60% of total fatty acids in CPRO and RRO.

It is known that the consumption of vegetable oils with high amounts of unsaturated fatty acids exert a substantial impact on human health, mainly in the prevention of cardiovascular and cancer diseases. On the other hand, vegetable oils rich in PUFA are sensitive to oxidative damage during storage and processing [36].

Our previous studies reported similar results of FAC for rapeseed oils (SAFA = 6.91–7.58%, MUFA = 64.14–66.14%, PUFA = 27.22–30.17%) [36]. However, Symoniuk et al. [6] obtained a higher level of PUFA (64.30–77.42%) in linseed oils.

For comparison, Ratusz et al. [7] studied the quality characteristics of commercial cold-pressed camelina oils and reported somewhat different concentrations of 7.4–10.1% for SAFA, 31.3–36.3% for MUFA, and 55.2–58.4% for PUFA. Specifically, the fatty acid profiles depend on both genetic and environmental factors.

### 3.1.3. Radical Scavenging Activity of Mayonnaise Ingredients

The radical scavenging properties of mayonnaise ingredients were analyzed by the DPPH and ABTS assays, and the obtained RSA results are presented in Table 4.


**Table 4.** Radical scavenging activity of mayonnaise ingredients determined by DPPH and ABTS assays.

*n* = 5; SD—Standard Deviation; Different letters (a–d) within the same column indicate significant differences between radical scavenging activity of samples: RRO—refined rapeseed oil, CPRO—cold-pressed rapeseed oil, CPSO—cold-pressed sunflower oil, CPLO—cold-pressed linseed oil, CPCO—cold-pressed camelina oil (Tukey's post hoc test, *p* < 0.05).

Among the investigated ingredients, nutritional yeas<sup>t</sup> and mustard had the highest DPPH (1687 and 1129 μmol TE/100 g) and ABTS (9192 and 4985 μmol TE/100 g) values. For comparison, Bors et al. [49] reported that the RSA of mustard paste determined using the DPPH method ranged between 23.4–23.7%.

The radical scavenging properties of aquafaba (DPPH = 437 μmol TE/100 g and ABTS = 2097 μmol TE/100 g) were higher than reported by previously published data (0.15–0.38 μmol TE/g) [50]. This variability in total antioxidant content in chickpeas and aquafaba can be explained by the prolonged time of contact of chickpeas with aquafaba in the jar as well as the influence of other factors such as genetic, agronomic, environmental and technological factors. It is known that the ABTS assay is applicable to both hydrophilic and lipophilic antioxidant systems, whereas the DPPH assay is more suited to hydrophobic systems. It can be noted that the ABTS value of aquafaba (2097 μmol TE/100 g) was five times higher than the DPPH value (437 μmol TE/100 g), probably due to water-soluble antioxidants being predominant in the aquafaba containing 95% water (Table 4).

Moreover, the ABTS results (588–738 μmol TE/100 g) of oils used to make vegan mayonnaise samples were higher than DPPH values (272–391 μmol TE/100 g). Insignificant differences for ABTS values of all the studied cold-pressed vegetable oils were observed, whereas RRO, CPRO and CPCO had significantly higher DPPH results than CPSO and CPLO (Tukey's post hoc test, Table 4). Interestingly, rapeseed and camelina cold-pressed oils were found to be richer sources of antioxidants than cold-pressed sunflower and linseed oils. A similar decrease in DPPH values for cold-pressed oils (rapeseed > sunflower > flax and camelina > flaxseed) was reported by Siger et al. [8] and Grajzer et al. [9].

### *3.2. Characterization of Mayonnaise Samples*
