## 3 Characterization of Food-Grade Systems
### Technological Characterization  
#### 3.1.1 Morphology and Size of Carriers  
Morphology is one of the parameters that influence the efficacy and biological properties of drug delivery systems. The surface-area-volume ratio determines pharmacokinetic properties such as absorption and biodistribution, permanence in the bloodstream, and toxicity [78]. Microscopy techniques provide information on particle shape, structure, topography, and size. In the case of all-food-grade nanotechnology for colon-targeted delivery, in addition to classical uses, this method has been used for several purposes. The following are some examples. Liu et al. prepared alginate microspheres for colon delivery of quercetin. The authors reported that water evaporation from the microspheres during the freeze-drying process caused shrinkage of the matrix resulting in surface wrinkles. In order to preserve the spherical shape of the microcarriers, inulin was added as a matrix filling material. SEM analysis showed that the addition of inulin provided effective pore filling of the alginate network, improving parameters such as gel strength and encapsulation efficiency. Moreover, comparison of morphological images of the empty and QUE-loaded systems showed that also the QUE contributed to pore filling, leading to the formation of sandy surfaces [79]. In our recent study, SEM analysis was used to evaluate the degradative processes under simulated GIT conditions of food-grade microparticles based on EUGB and EUGC and loaded with resveratrol (RSV). The morphological study showed that matrix degradation was pH- and polymer type-dependent. After 2 h of incubation in SGF, EUGB microparticles maintained unchanged and smooth surfaces, while the first signs of irregularity were detected after incubation in SIF and disaggregation after exposure to SCF, confirming a colonic-targeted release (Figure 11). In contrast, EUGC-based samples showed irregular and porous surfaces already after incubation in SGF and SIF and only partial erosion in SCF, in agreement with the prolonged RSV release profiles. The particle size distribution of the microsystems was investigated using a multidimensional sieve shaker according to the Italian Pharmacopoeia [80]. Figure 10: Characterization steps for food delivery systems. In a study by Sarma et al., core-shell nanoparticles for RSV delivery were prepared with a chitosan core coated with pectin. Different chitosan:pectin ratios (1:4, 1:7 and 1:15) were studied to modify the thickness of the shell. The core-shell morphology, mean core diameter, and shell thickness were investigated using three electron microscopy methods: field emission scanning electron microscopy (FE-SEM), selected area electron diffraction (FE-TEM/SAED), and high-resolution transmission electron microscopy (HR-TEM). FE-SEM images revealed spherical shape NP with a size between 23 and 32 nm. FE-TEM/SAED images were obtained to obtain information on the internal morphology and particle distribution, where FE-TEM analysis revealed the formation of monodisperse core-shell particles with a spherical core of about 13 nm and a shell with an average thickness of about 4 nm; finally, SAED imaging showed circles and bright spots that highlighted the crystalline behavior of the encapsulated RSV. HR-TEM analysis confirmed the crystalline nature and, in relation to the amount of used pectin, showed an increase in the thickness of the coating [81]. One of the limitations of electron microscopy analysis is related to sample preparation. Typically, samples for SEM analysis undergo a drying step before being coated with a metal with high electrical conductivity, while negative dyes or heavy metals are used to confer electronic transmittance in samples for TEM analysis [82]. A microscopic technique with no special sample treatments is atomic force microscopy (AFM), which is used for topographical analysis of nanomedicines. This type of morphological imaging allows to probe surfaces to obtain parameters such as roughness and other mechanical properties [83].