## 3 Characterization of Food-Grade Systems
### Technological Characterization
#### 3.1.4 In Vitro Release Testing
The optimized system had a particle size of 83 \(\pm\) 4 nm and a Zeta potential of 25 \(\pm\) 1 mV in the presence of PEG with an encapsulation efficiency of about 63%. To simulate the gastric environment without enzymes, 1 mL of the system was placed in the dialysis tube and suspended in 20 mL of HCl (0.1 M)-ethanol (in an 80:20 ratio) (pH = 1.2). Next, it was it suspended in 20 mL of phosphate-ethanol buffer (in an 80:20 ratio) (pH = 6.8) to simulate the condition of the small intestine. Finally, the colonic environment was simulated by raising the pH to 7.4 in the presence and absence of 0.6 mg/mL pectinase enzyme. The test was conducted under agitation in an incubator (50 rpm, 37 \({}^{\circ}\)C). The study, in addition, analyzed the release behavior in the pH of grape juice (for potentialuse in fortified food production) by incubating 1 mL of the system inside a dialysis tube in 20 mL of acetate buffer. The pH was adjusted to 3.5 and 4.5 with 0.1 M acetic acid for one month at 4 \({}^{\circ}\)C. In vitro release studies in different simulated gastrointestinal media concluded that 49% and 60% of RSV was released into the simulated colonic environment by the nanoparticles with and without PEG, respectively. Finally, under the conditions simulating grape juice, the-PEG system recorded a higher cumulative release of RSV than the PEG-free form. Therefore, it is more likely that the role of NH\({}_{3}\)\({}^{+}\) groups in swelling at low pH was not as impressive as that of COO\({}^{-}\) groups at pH 6.8 and 7.4 [124]. In another study published in Chinese Medicine, microparticles (Shellac@Cur/MPs) were prepared with the core (zein)-shell (shellac) structure based on coaxial electrospray technology. The particles had an average hydrodynamic size of 2.8 \(\upmu\)m, a zeta potential of \(-\)25.3 mV and a high encapsulation efficiency over 95%. Three types of release media with different pH values were applied to simulate the digestive tract environments, namely artificial colonic fluid (pH 7.8), artificial intestinal fluid (pH 6.8) and SGF (pH 1.2). Both Cur/zein MPs and Shellac@Cur/zein MPs suspended in PBS (equal to 400 \(\upmu\)g of Cur) were introduced into a dialysis tube. The closed tube was placed in a centrifuge tube with 40 mL of the release medium at 150 rpm and 37 \({}^{\circ}\)C for 16 days. At predetermined intervals, an aliquot was taken for HPLC analysis. Both Cur/zein MPs and Shellac@Cur/zein MPs showed very slow-release profiles at pH 1.2, indicating that both systems could prevent burst release of the filler into the acidic stomach microenvironment due to the hydrophobicity of the zein and shellac materials. The intestinal release of CUR from Cur/zein MPs after 11 days had reached 45%. In the presence of shellac, the release was more sustained. CUR release from Shellac@Cur/zein MPs was disruptive at pH 7.8, indicating the pH sensitivity of shellac material [124]. In a study published in Gels, a core-shell bioactive release system was prepared using natural food-grade materials. The beads were fabricated by mixing WPI (where protein isolate) emulsion with pectin using the ionotropic gelling method. The incorporation of high pectin content into the system affected its texture, network structure and thermodynamics. Since the degree of swelling is a crucial factor in the release of quercetin from these systems, the swelling properties of all beads were measured in distilled water after immersion at 25 \({}^{\circ}\)C for 6 h. Beads containing 40% pectin showed the highest swelling ratio after 6 h (48.5%) indicating a porous structure. In contrast, those with 20% had markedly decreased after 3 h, probably indicating the loss of structural integrity following the absorption of water molecules and subsequent dilution of the polymer. The in vitro release test found a maximum release of 10% quercetin for the beads with 20% pectin in SIF. This showed high stability of the systems in SGF and SIF indirectly indicating the potential for use as colon-targeted nutraceutical delivery systems [125]. Liu, S. et al.