Abstract

Cardanol (CA), obtained by distillation of Cashew nut shell liquid, can be considered as the major by-product of the cashew industry having concrete perspectives for the preparation of new fine chemicals and functional hybrid materials.1 Chlorogenic acids (CGA) are natural phenolic compounds which can be extracted from rowanberries or coffee beverages having well known biological effects.2-5 Our attention has been focused on the combination of natural renewable materials which can be used for the environmentally friendly preparation “green nanocarriers”. In this context we proposed the efficient extraction process in water of phenolic compounds from two different berries (Sorbus Americana and Vaccinium sp.) and their encapsulation in a natural cardanol-based vesicular nanodispersion in order to obtain an entirely natural system as carrier loaded with bioactive compounds. Chlorogenic acid derivatives (3-O-caffeoylquinic (3-CQA) and 5-caffeoylquinic acid (5-CQA) were extracted from Sorbus Americana and Vaccinium sp. by vortex vigorous shaking using water as solvent. The identification and quantification of these compounds was achieved by UHPLC-ESI-MS/MS, comparing their retention times, MS/MS fragmentation patterns and by using a calibration curve of commercial standard. Using this methodology, phenolic compound concentrations obtained in S. Americana extract was 21.63 mg/L and 80.51 mg/L of the isomers 3-CQA and 5-CQA, respectively, whereas only the 5-CQA isomer (63.16 mg/L) in Vaccinium sp. extract was detected. These extracts were used to prepare the vesicle nanodispersions (CA-CH-S. Americana, CA-CH-Vaccinium sp. and blank sample with only CA-CH as a comparison) following a procedure similar to that recently reported,6 in which CA can acts as solvent of the phenols and cholesterol due to its amphiphilic properties under alkaline conditions. After exhaustive dialysis in order to remove the non-entrapped (free) phenols in vesicles, the nanodispersion samples were characterized and analyzed to establish stability, size, shape and phenol content. The formation of spherical and regular vesicles was confirmed by transmission electron micrographs (Figure 1). Particle size as hydrodynamic diameters and Zeta–potential were determined by dynamic light scattering measurements. The presence of phenols encapsulated into vesicle nanodispersions has been evidenced by UV–visible measurements. The phenol-loaded vesicles, after their lysis and solubilization in methanol, display a similar absorption to the berries methanolic extract, unlike the blank sample CA-CH that does not exhibit any characteristic absorption throughout the investigated spectral region. The ability of the vesicles to entrap the phenols was studied by UHPLC-ESI-MS/MS analysis performed in Multiple Reaction Monitoring (MRM) mode and expressed as percent of encapsulation efficiency (E%). In the Table 1 was reported a comparison of phenol concentrations in the nanodispersions before and after dialysis process, and the E% of cardanol-based vesicles loaded with phenolic compounds. Table 1.- Phenolic compound concentrations (mg/L) in lysed vesicles measured before (ND) and after (D) dialysis process, and E (%) of cardanol-based vesicles. Samples Phenolic compounds (mg/L) 3-CQA 5-CQA CA – CH – S. Americana ND vesicles 19.96 25.97 D vesicles 1.77 2.53 E (%) 8.88% 7.42% CA – CH – Vaccinium sp. ND vesicles 23.61 25.19 D vesicles 0.32 0.79 E (%) 1.37% 3.14% MRM chromatograms corresponding to the quantified phenolic compounds in S. Americana extract, Vaccinium sp. extract and phenols entrapped by vesicles confirms as reported in table 1, that is when Vacciniun sp. was trapped in the vesicles part of 5-CQA was transformed to 3-CQA. Studies about the influence of pH on 5-CQA thermal stability7 have shown that chlorogenic acid can undergo transformations, such as isomerization in 3-CQA, during the heating of its water solution at different pH. References 1. Mele, G., Vasapollo G. , 2008, Mini-Reviews in Organic Chemistry 5 (3), 243. 2. De Paula A.A., Martins J.B., Dos Santos M.L., Nascente Lde C., Romeiro L.A., Areas T.F., Vieira K.S., Gamboa N.F., Castro N.G., Gargano R., 2009, Eur. J. Med. Chem., 44, 3754. 3. Trevisan M.T., Pfundstein B., Haubner R., Wurtele G., Spiegelhalder B., Bartsch H., Owen R.W., 2006, Food Chem. Toxicol., 44,188. 4. Belay A., Gholap A. V., 2009, African Journal of Pure and Applied Chemistry, 3, 234. 5. Camuesco D., Comalada M., Rodriguez-Cabezas M. E., Nieto A., Lorente M. D., Concha A., Zarzuelo A., Galvez J., 2004, Br. J. Pharmacol., 143, 908. 6. Bloise E., Carbone L., Colafemmina G., D’Accolti L., Mazzetto S. E., Vasapollo G., Mele G., 2012, Molecules, 17, 12252. 7. Dawidowicz A.L., Typek R., 2011, Eur. Food Res. Technol. , 233, 223.