## 3 Results and Discussion
### Nanoemulsion Characterization  
Table 3 presents the characterization of nanoemulsions, including mean droplet size (MDD), polydispersity index (PDI), turbidity, viscosity, and refractive index, prepared using proteins from untreated (control) and HPH-treated (600 bar for 1 pass and 800 bar for 4 passes) biomass. The emulsions were homogenized using either high-speed homogenization (HSH, one-step, 8000 rpm for 10 min) or a combination of HSH followed by high-pressure homogenization (HSH-HPH, two-step, 600 bar for 4 passes), with varying oil content. The main parameter, significantly influencing MDD and PDI, was the homogenization method. Samples homogenized solely with HSH exhibited significantly higher MDD and PDI compared to those processed with the two-step homogenization method (HSH-HPH). This is likely due to the higher energy input of the HSH-HPH emulsification process, which produces smaller droplet sizes and a narrower droplet size distribution by applying more intense shear forces and turbulence [44; 45]. A typical droplet-distribution graph for emulsions stabilized by proteins from untreated biomass and homogenized with HSH (one-step homogenization) and HSH-HPH (two-step homogenization) is presented in Figure 5. Additionally, the turbidity of HSH-HPH samples was significantly lower  
\begin{table}
\begin{tabular}{c c c} \hline \hline
**Interfacial Tension (mN/m)** & **1 Pass** & **4 Passes** \\ \hline Control & 4.25 \(\pm\) 0.19 \({}^{\text{a}}\) & 4.25 \(\pm\) 0.19 \({}^{\text{a}}\) \\
400 bar & 4.62 \(\pm\) 0.33 \({}^{\text{ab}}\) & 5.20 \(\pm\) 0.42 \({}^{\text{bc}}\) \\
600 bar & 5.48 \(\pm\) 0.35 \({}^{\text{c}}\) & 5.27 \(\pm\) 0.39 \({}^{\text{bc}}\) \\
800 bar & 5.65 \(\pm\) 0.41 \({}^{\text{c}}\) & 4.75 \(\pm\) 0.45 \({}^{\text{ab}}\) \\ \hline \hline \end{tabular} Different superscript letters indicate significant differences as calculated by [PERSON]’s multiple range test for a significance level of \(p<\) 0.05. Abbreviations: APPC: _A. perenniosa_ protein concentrate, HPH: high-pressure homogenization.  
\end{table}
Table 2: Interfacial tension between aqueous solutions of APPC from untreated (control) and HPH-treated biomass and sunflower oil.  
Figure 4: Dependence of _A. perenniosa_ protein concentrate (**a**) WHC and (**b**) OHC on HPH treatment of _A. perenniosa_ biomass from 1 bar (untreated) to 800 bar for one- and four-pass treatment. Error bars represent the standard deviation between experimental replications. Different superscript letters indicate significant differences between means \(\pm\) standard deviation as calculated by [PERSON]’s multiple range test for a significance level of \(p<\) 0.05. Abbreviations: WHC: water-holding capacity, OHC: oil-holding capacity, APPC: _A. perenniosa_ protein concentrate, HPH: high-pressure homogenization.  
compared to HSH samples, reflecting the more efficient homogenization and narrow distribution (lower PDI values). These results highlight that the HPH treatment is also effective for preparing nanoemulsions stabilized by proteins from _A. pyrenoidosa_. Furthermore, turbidity increased by up to 335% with the increase in oil content from 2 to 6% \(w/w\). The viscosity and the refractive index of the samples were relatively close to water's respective parameters. Specifically, the viscosity varied between 1.10 and 1.51 cP, and the refractive index between 1.337 and 1.346, while no significant differences between biomass treatment and emulsion homogenization method were observed.