Avenanthramides (AVNs) are natural polyphenols obtained from oat sprouts and can also be chemically synthetized. The aim of the present study was to assess the anticancer, anti-inflammatory and antioxidant effects of individual synthetized AVNs (s-2c, s-2p, s-2f) and a natural AVN mixture (n-MIX) on CaCo-2 and Hep3B cancer cells. In CaCo-2, the AVN s-2c was found to be the most cytotoxic followed by the n-MIX. In Hep3B cells, a marked cytotoxic effect was found but no significant difference was observed between the synthesized AVNs and the n-MIX. In both CaCo-2 and Hep3B cells, natural and synthetic AVNs activated caspases 8 and 3, and the n-MIX and the AVN s-2c were also able to activate caspase 2. Both synthetic and natural AVNs downregulated pro-survival genes hTERT, COX-2 and MDR1, inhibited the activity of pro-inflammatory COX-2 enzyme and reduced prostaglandin E2 levels, showing the potent chemopreventive effects of these oat-derived phytochemicals. Synthetic AVN s-2c was found to have the highest chemical antioxidant capacity, as indicated by ORAC, DPPH and ABTS values, whereas all AVNs and n-MIX were shown to have similar intracellular antioxidant activity, evaluated by means of the DCFH-DA assay. As AVNs have high bioavailability in humans, results of this study suggest that oat-based foods, fortified with AVNs, could be an alternative to produce functional foods with anticancer, anti-inflammatory and antioxidant effects for health benefits.