This study aimed to investigate the effect of Pyracantha fortuneana fruit extract (PFE) as a nitrite substitute on the quality, flavor, contents of biogenic amines and the formation of antioxidant peptides of fermented beef sausages during storage. Compared with the control group, the fermented beef sausages treated with PFE showed elevated L* and a × values, enhanced antioxidant activity, and increased resistance to microbial growth and lipid and protein oxidation. In particular, there were 2.44 ± 0.02 mg MDA/kg thiobarbituric acid reactive substances after 35 d. Interestingly, PFE had a positive effect on the texture, flavor, and antioxidant protection of fermented beef sausages that was superior to that of nitrite. PFE not only improved the microstructure of fermented beef sausage but also delayed the degradation of myofibrillar and sarcoplasmic proteins as shown in SDS-PAGE analysis, which resulted in a significant improvement in the sausage flavor and reduction of biogenic amines (P < 0.05). The LC-MS/MS results showed that PFE-treated fermented sausages significantly promoted peptide formation and had high 2,2-diphenyl-1-picrylhydrazyl radical scavenging and ferric ion reducing antioxidant power (P < 0.05). Therefore, PFE has the potential to be effective and promising substitute for nitrite in industrial production and application in fermented sausages.