This study aimed at investigating the use of the water-soluble extract of amaranth seeds for extending the shelf-life of gluten-free and wheat flour breads. The antifungal activity of the amaranth water-soluble extract was shown by agar diffusion, conidia germination and dry biomass assays, using Penicillium roqueforti DPPMAF1 as the indicator fungus. The crude water-soluble extract had minimal inhibitory concentration (MIC) of 5 mg of peptides/ml and showed inhibition towards a large number of fungal species isolated from bakeries. Four novel antifungal peptides, encrypted in amaranth agglutinin sequences, were identified from the water-soluble extract by nano-Liquid Chromatography-Electrospray Ionisation-Mass Spectra/Mass Spectra (nano-LC-ESI-MS/MS). The water-soluble extract of amaranth was used as an ingredient for the manufacture of gluten-free and wheat flour breads and the inhibitory activity was confirmed during long-term shelf-life under pilot plant conditions. The effect of the water-soluble extract on gluten-free bread rheology and sensory properties was also shown.