Developing polymer-based delivery systems to improve bioaccessibility and bioefficacy of procyanidins

Procyanidins, also known as condensed tannins, have recently attracted a great deal of attention because of their promising health-promoting effects, including antioxidant, neuroprotective, cardioprotective, anti-inflammatory, anti-obese activity and anti-cancer. They exist in different parts of plants, such as fruits, nuts, flowers and seeds, to defend against biotic and abiotic stressors. The typical structure of procyanidins is monomer procyanidin (also known as catechin or epicatechin) linked by single or multiple interflavan bonds. Nowadays, there is significant amount of procyanidin presenting in food and agriculture waste, taking advantage of which could provide a sustainable source of dietary supplements and functional ingredients. While more research is engaging in fully understanding the extent of their health benefits and incorporating foods rich in procyanidins as part of a balanced diet, studies on how to improve the overall <i>in vivo</i> absorption are rarely reported. Researchers have found that the concentration of orally administrated procyanidins in plasma and organs is very low due to their poor gastrointestinal stability and low bioavailability. In recent years, the polymer-based delivery system composed of protein and polysaccharides has been applied to encapsulate various phytochemicals, because of its good biocompatibility and encapsulation efficiency. However, the majority of reported procyanidins delivery systems were fabricated for monomers and limited formulations have been developed for procyanidins dimers and oligomers. In this study, polymer-based delivery systems were developed to encapsulate procyanidin dimer A₂ (PAC A₂) and B₂ (PAC B₂) to improve their bioaccessibility and bioefficacy. First of all, based on the polarity of the two dimers, gliadin-carboxymethyl dextran (G-CMD) nanoparticles were fabricated through anti-solvent titration method for encapsulating PAC A₂, while whey protein- cellobiose Maillard conjugates (WPI-CE) were developed through Maillard reaction for delivering PAC B₂, with the encapsulation efficiency reaching 84.9% and 88.2%, respectively. The ex vivo permeation study (Franz cells) was performed, and it was found that the encapsulation significantly improved the transmembrane rates of PAC A₂ and B₂ by 1.4-fold and 1.47-fold, respectively. Tested by TNO Gastro-Intestinal model (TIM-1), the bioaccessibility of PAC A₂ and B₂ was found to be increased upon encapsulation by 6-fold and 3.7-fold. Moreover, the cellular uptake study indicated that both encapsulated G-CMD nanoparticles and WPI-CE conjugates could enter Caco-2 cells within 2 hours and were observed surrounding the cell nuclei. Furthermore, G-CMD nanoparticles slightly increased the anti-lipogenic activity of PAC A₂, which reduced lipid accumulation in differentiated 3T3-L1 cells in a dose-dependent manner. PAC B₂ loaded in WPI-CE conjugates exhibited a higher antioxidant ability than the free PAC B₂ at all concentrations (0- 50 µM) on the Hep-G2 cell line. Finally, <i>in vitro</i> microbial fermentation was conducted to investigate the metabolic pathways of the encapsulated and the free procyanidin dimers, and a high-throughput UHPLC-QqQ-MS/MS method combining QuEChERS and (EMR)-lipid technology was successfully developed for sample preparation and analysis. The intermediate metabolites were tentatively identified by UHPLC-QToF MS/MS during colonic fermentation and 34 phenolic compounds were identified and quantified. According to the metabolomic study, different metabolism pathways of PAC A₂ and B₂ were proposed. Compared to the non- encapsulated procyanidin dimers, the encapsulated forms showed a slower release rate and better retention of both parent compounds and metabolites in the fermentation medium, which is very important for procyanidin to continuedly produce local gut and systemic health effects.In conclusion, polymer-based delivery systems were successfully developed for two procyanidin dimers, and the encapsulated procyanidin dimers showed improved bioaccessibility and bio-efficacy compared to their free forms. This work offers a systematic screening framework for assessing the phytochemicals delivery efficiency of oral delivery systems, and it promotes the application of phytochemicals with enhanced bioefficacy in the functional food industry.