CHEMICAL COMPOSITION, ANTIOXIDANT AND ANTIPROLIFERATIVE ACTIVITY OF HAZELNUT SHELLS

Introduction Hazelnut (Corylus avellana L., Betulaceae family) is one of the most cultivated and consumed nuts in the world, not only as whole fruit (raw or roasted) but also incorporated as an ingredient into manufactured food products. Substantial quantities of by-products, including the green leafy cover, hard shell, and skin, are produced during harvesting and industrial processing of hazelnuts. The hard shells account for a majority of this waste and represent a huge amount of discarded material available at very low cost. Nowadays, the hazelnuts shells are used for burning as a heat source, for mulching, and as a raw material for the production of furfural in the dye industry (Stévigny C., et al. 2007). In the last years, great attention has been paid to the recovery and upgrading of agro-industrial residues as potential sources of bioactive compounds with beneficial effects on human health, and some papers have already showed the potential use of hazelnut wastes as natural antioxidants (Shaidi F., et al. 2007; Contini M., et al. 2008). In this research we report for the first time the chemical composition of a polar extract obtained from hazelnuts hard shells. Moreover, the antiproliferative activity of the extract and its constituents has been evaluated on A375 and SK-Mel-28 (melanoma cancer cell lines) and Hela (cervical carcinoma cell line) cells. Methods Hazelnut (var. Mortarella and San Giovanni in mixture) hard shells were kindly supplied by a local hazelnut processing industry (Hazelnuts South Italy Manufacturing s.r.l., Baiano, AV). The shells were defatted with n-hexane and chloroform and then extracted with methanol. This extract was purified by Sephadex LH-20 followed by RP-HPLC to give twelve compounds. Their structures were elucidated using spectroscopic methods including 1D- and 2D-NMR experiments as well as ESIMS analysis. The polyphenols content of the extract was examined by the Folin-Ciocalteau colorimetric assay (Picerno P., et al. 2011). The concentrations of the major compounds were determined by a RP-HPLC-DAD direct calibration method. The free-radical scavenging activity of both the extract and pure compounds was evaluated by DPPH-test (Picerno P., et al. 2011). The effects of hazelnut shells extract and its constituents on cell proliferation and apoptosis on human cancer cell lines were evaluated by the MTT bioassay and propidium iodide staining, respectively (MencheriniT., er al. 2011)Results The phytochemical investigation of the methanol extract of hazelnut shells led to the isolation and characterization of four neolignans, seven phenolic compounds, and a diarylheptanoid. C-veratroylglycol, lawsonicin and cedrusin resulted the major constituents of the extract (2.93, 1.98, and 1.79%, respectively, HPLC method). The polar extract showed a significant and concentration dependent free-radical scavenging activity (EC50 31.87 ug/ml, with respect to alpha-tocopherol used as positive control EC50 10.10 ug/ml) correlated to its high polyphenols content (193.80 ug/mg), mainly gallic acid and methyl gallate (EC50 1.20 and 1.19ug/ml, respectively). Moreover, the extract exhibited a significant and concentration-dependent inhibitory effect on tested cancer cell growth (IC50 500 μg/mL), and it increase the percentage of hypodiploid cells at the same concentration. The activity seem to be correlated with the presence of pure compounds, balanophonin (a neolignan) and gallic acid, which showed a citotoxic effect on all cell lines (IC50 150 uM). Furthermore, the neolignan cedrusin showed a citotoxic activity only on the A375 and HeLa cells. Conclusion The obtained results showed the potential use of a hazelnut shell extract, contained different classes of compound including neolignans, phenolics and a diarylheptanoid, as antioxidant and anticancer agent.