Abstract The potential of cantabrian anchovy (Engraulis encrarischolus) viscera as a source of bioactive compounds is of interest for both, pharmaceutical and food industries. Cantabrian anchovy guts and heads were freeze-dried, extracted with methanol and subjected to fractionation by solvent partitioning using hexane, ethyl acetate, and butanol. Fractions were tested for antimutagenic, antioxidant, antifungal, and antibacterial activity using the Ames test; DPPH, ABTS, and FRAP assays; the radial grown inhibition assay; and the microbroth dilution method, respectively. Five fractions were obtained from the anchovy gut methanolic extract, in addition to the hexane- (HF), ethyl acetate- (EAF), and butanol-soluble (BF) fractions, an aqueous-soluble fraction (ALF) and precipitated crystals (ACF) in this were also obtained. HF and EAF resulted to be antimutagenic, HF and ALF showed antifungal activity, BF and ACF showed the highest antioxidant potential, and HF and BF were antibacterial against several strains. Anchovy gut, which to the present study had not been reported for any bioactivity, has antimutagenic, antifungal, antioxidant, and antibacterial compounds, which need to be isolated for full characterization and study.