## 3 Characterization of Food-Grade Systems
### Technological Characterization
#### 3.1.1 Morphology and Size of Carriers
Recently, two nanocomplexes consisting of saponin-cholesterol were prepared and characterized by AFM. The study observed that the combination of cholesterol with the saponins escin and dioscin led to the self-assembly of nanoparticle and fibrous structures, respectively. Furthermore, it was observed that the density and roughness of the surfaces varied according to the molar ratios between the two molecules [84]. Although this technique has been introduced more recently compared with electron mi  
Figure 11: SEM analysis of the dissolution/erosion process of microparticles with a matrix consisting of 100% Eudraguard® Biotic and 100% Eudraguard® Control (personal data). croscopy methods, the last ones possess advantages such as the ability to explore larger surfaces and shorter scanning times. Therefore, AFM is generally combined with SEM/TEM analysis to ensure more reliable surface characterization. In addition to morphology, the size and PDI of nanosystems are also critically important parameters that affect properties such as release and encapsulation of the active molecule, absorption, and stability of the formulation. Although morphological analyses also provide information regarding the average diameter of nanosystems, it must be considered that these values are determined on statistically small samples and that effective sample measurements could be modified by the procedures of sample preparation [85]. To overcome these issues and to obtain better statistics, particle average diameters are frequently measured via the dynamic light scattering (DLS) technique. DLS analysis requires a larger number of particles than microscopy techniques and describes also the interactions of the nanosystem with the solvent, providing more detailed dimensional and distributional information. Sometimes, the correlation between measurements obtained from both DLS and electron microscopy techniques is poor, as reported in the case of the above study by Sarma et al. [80]. The authors observed that the sizes evaluated by DLS measurements were much larger (around 900 nm) than those observed with TEM analysis. These differences could be reduced when analyzing spherical particles with homogeneous polydispersion [84]. This phenomenon was confirmed in the study by Doost et al., where NPs of shellac and almond gum were prepared for quercetin delivery. The study investigated how the particle size of the nanosystems was affected by the stirring and dosing speed and the concentrations of almond gum and Tween 80, which was used as a stabilizer. Two types of electron microscopy were performed to evaluate the morphology of the samples, cryogenic scanning (cryo-SEM) and TEM. The size and particle size distribution were measured by DLS analysis. The results of the study showed that the increase in the size of NPs was correlated with a delayed nucleation rate, which was determined by both increasing the agitation speed and the higher concentration of almond gum. Conversely, smaller NPs sizes were obtained when the nucleation rate was faster, which was obtained by increasing the stirring speed and surfactant concentration. The NPs populations were homogeneous (0.252 \(\pm\) 0.01), spherical in shape, and the diameters measured by the three analyses showed consistent results, indicating good agreement between the used methods [86].