## 4 Bionanosystems
### Bionanosystems for RA
#### 4.1.1. Bionanosystems Based on Organic Carriers
demonstrated the efficiency of alginate beads in encapsulating aqueous extracts of _Plectranthus_ plants, having RA and caffe acid as major components [112; 113]. The authors demonstrated that alginate-based bionanosystems retain a high amount of RA and caffe acid, are excellent inhibitors of acetylcholinesteraseinesterase, and have good antioxidant capability. All these features indicate that this strategy could potentiate the action of plant extracts by promoting a protective effect and a sustained release of active constituents and, ultimately, improve their pharmacological effects. Similar results were found by [PERSON] et al. [114] that investigated the encapsulation of thyme aqueous extract containing RA in calcium alginate gel beads prepared by electrostatic droplet generation (electrostatic extrusion). Additionally, in this case, it was demonstrated that the encapsulation preserved the bioactive compounds as the total antioxidant content remained unchanged. Another interesting example of RA immobilization for food and pharmaceutical packaging is that proposed by [PERSON] et al. [115] that prepared gelatin-based edible films containing RA for effective antioxidant and long-term antibacterial activities. In this case, RA was covalently bonded to the carrier. Indeed, the first RA was conjugated onto a gelatin backbone via the carbodiimide (EDC)/N-hydroxysuccinimide (NHS) method (Figure 7) and later the conjugates were plasticized by glycerol and crosslinked by dialdehyde xanthan gum to fabricate active RA-gelatin edible films. The results indicated that the RA-gelatin edible films present good water resistance properties, strong mechanical properties, and excellent ultraviolet light barrier ability. The conjugation of RA conferred gelatin-based edible films with effective antioxidant and antibacterial activities, which was maintained for 3 months. Figure 6: (**A**) Molecular structures of the guest and hosts in RA-CD complexation: (**a**) RA, (**b**) \(\alpha\)-CD and (**c**) \(\beta\)-CD (R = H) and its derivatives. (**B**) Fluorescence spectra of RA in aqueous solution with different CDs at pH 7.0. (**C**) Trolox equivalent antioxidant capacity (TEAC) of RA in free and complex forms in aqueous solutions at pH 7.0 (adapted from [PERSON] et al. [111], copyright 2011 Elsevier). [PERSON] et al. [92] prepared crosslinked poly(RA) particles by an emulsion cross-linking process (Figure 8) and found that poly(RA) particles can release RA in acidic, neutral and basic environments, and that the antioxidant activity of poly(RA) is almost indistinguishable from that of pure RA. In addition, poly(RA) particles were \(\sim\)66% less cytotoxic to mammalian cells than RA monomer and showed a strong inhibition of \(\alpha\)-glycosidase for possible application in the treatment of diabetes. Figure 7: Scheme of preparation of RA-gelatin film. Survival rates of _E. coli_ (**A**) and _S. aureus_ (**B**) after treating with newly prepared (red histogram) and 3 months stored (blue histogram) gelatin-based edible films for 24 h at 37 \({}^{\circ}\)C. (**C**) SEM images of _E. coli_ and _S. aureus_ after treating with gelatin-based edible films for 24 h at 37 \({}^{\circ}\)C (adapted from [PERSON] et al. [115], copyright 2018 Elsevier). [PERSON] et al. [116] described the encapsulation of RA into polylactic-co-glycolic acid (PLGA) and poly-lactic acid (PLA) carriers (with size ranging between 1.5 \(\pm\) 0.5 \(\upmu\)m and 0.3 \(\pm\) 0.1 \(\upmu\)m) in combination with \(\beta\)-carotene (\(\beta\)-CA) andtocopherol (\(\alpha\)-TOC) and using supercritical emulsion extraction technique for the preparation. RA and \(\alpha\)-TOC were added to the formulation as extra protectors of the light-sensitive \(\beta\)-CA thus improving its shelf-life. Results showed that the encapsulation efficiency of RA in both PLA and PLGA was very low; between 1.3 and 3.9%, but it increased by co-encapsulation of RA with \(\beta\)-CA and \(\alpha\)-TOC.