To explore the antibacterial activity of lemon peel and pulp fresh juice against seven common foodborne pathogens and its effects on bacteria biofilm formation.Methods The fresh lemon peel and pulp juice were prepared by juice extractor. The total acids of fresh lemon peel and pulp juice were determined. Volatile compounds of lemon peel and pulp fresh juice were extracted by headspace sampling and the extracts were detected by gas chromatography-mass spectroscopy (GC-MS). The antibacterial tests of lemon peel and pulp fresh juice were examined on tryptic soy agar (TSA). The bacteria biofilm formations were tested by tube method and 96-well method. Results The total acids were 98.86 g/kg for pulp fresh juice, and 6.05 g/kg for peel fresh juice, respectively. GC-MS analysis showed limonene, γ-terpinene, α-terpineol and citral were the main volatile compounds in lemon peel and pulp fresh juice. The growths of seven target pathogens on TSA were inhibited distinctly by lemon pulp juice and did not affect by lemon peel juice. The tube and 96-well methods for the bacterial biofilm formation tests showed that the lemon pulp juice had obvious inhibition to several bacteria biofilm formation, and lemon peel juice had obvious promoting effects to most bacteria biofilm formation.Conclusion The effects of lemon juice on common foodborne pathogens and their biofilm formation provide meaningful scientific data for the prevention and control of foodborne pathogens.