ZEIN NANOPARTICLES AS SUSTAINABLE DELIVERY SYSTEM OF ANTHOCYANINS DERIVING FROM PURPLE CORN WASTE

Purple corn (Zea mays L.) has shown preventive effects against several chronic diseases in preclinical and clinical studies. Its health effects have been attributed to the presence of anthocyanins (ACNs), flavonoid pigments highly concentrated in corn seeds and cobs. Nevertheless, the ACN bioavailability is low, mainly due to their poor stability. One of the strategies to limit ACN degradation is their protection through encapsulation in drug delivery systems. Zein (the main storage protein in corn) has already been tested as a suitable and sustainable delivery system in biomedical applications, thanks to its natural abundance, physico-chemical properties, and biodegradability. In this work, ACNs extracted from purple corn cobs, an agro-industrial by-product, were encapsulated in zein nanoparticles (NPs) to assess their potential therapeutic applications. Both loaded and unloaded formulations were tested on enterocytes (CaCo-2 cells) and cardiomyocytes (HL-1 cells). The extraction of ACNs from purple corn cobs was performed by maceration using hydroalcholic solutions acidified with six different acids: hydrochloric, lactic, tartaric, formic, citric and acetic acids. The ACN-enriched extracts were compared in terms of extraction efficiency and selectivity by Liquid Chromatography with Photodiode Array detector (UHPLC-PDA). Acetic, formic and hydrochloric acid presented the best results and the extracts were tested on CaCo-2 cells. Only formic and hydrochloric extracts were not cytotoxic at suitable ACN concentrations (up to 25 μM). These extracts were further characterized by Liquid Chromatography coupled to High Resolution Mass Spectrometry, showing cyanidin 3-O-glucoside as the most abundant anthocyanin followed by cyanidin-3-O-malonyglucoside, peonidin 3-O-glucoside, and peonidin 3-O-malonylglucoside. Formic and hydrochloric extracts were then loaded on zein-based NPs. ACN encapsulation efficiency resulted between 1.8% and 5.5%, as assessed using UHPLC-PDA. The ACN-NPs were also characterized in terms of morphology, size and zeta potential, through Scanning Electron Microscopy and Dynamic Light Scattering: spherical, homogeneous and nanoscaled particles were observed, with a surface potential of +40 mV, confirming the suspension stability. Furthermore, the stability of free and encapsulated ACNs was compared both in storage conditions and simulated biological conditions, highlighting the advantages of encapsulation in zein NPs. The ACN-loaded NPs were tested on CaCo-2 and HL-1 cells. Both formulations were not cytotoxic on CaCo-2 cells. On the contrary, nanoparticles formulated with hydrochloric acid resulted more suitable for applications on cardiomyocytes. All these results highlight the asset of purple corn waste valorisation and the potential of zein nanoparticles as delivery systems for ACNs in therapeutic applications.