Abstract: Purpose: To investigate the inhibitory effects of grapefruit seed extract (GSE) and its nanoemulsion (GNE) on biofilm formation of Escherichia coli and Staphylococcus aureus. Methods: The inhibitory effects of GSE and GNE on the formation of single and mixed-species biofilms were explored by analyzing the changes in the number of adherent cells and the content of extracellular polymeric substances (EPS) as well as microstructure observation, and the effectiveness of GSE and GNE in eliminating biofilms formed by E. coli and S. aureus was compared. Results: The number of adherent cells and EPS production during single-species biofilm formation were inhibited by GSE and GNE at 1/2 minimum inhibitory concentration (MIC), and S. aureus was more strongly inhibited. The changes of biofilm microstructure observed under a scanning electron microscope (SEM) and a confocal laser scanning microscope (CLSM) confirmed the inhibitory effect of GSE and GNE on biofilm formation by the two bacteria. GSE and GNE could also eliminate E. coli and S. aureus biofilms, and were more effective for S. aureus biofilm, but the resistance of the mixed-species biofilms was stronger than that of the single-species biofilms. GNE was more effective than GSE in inhibiting biofilm formation and eliminating biofilms. The amount of single-species biofilms formed by the two bacteria decreased by 0.6 (lg (CFU/cm2)) when treated with GNE at 1/2 MIC for 120 h compared with GSE, and the elimination rate of biofilms by 1.8% GNE increased by 7.7%–27.2% compared with GSE. Conclusion: GSE and GNE have inhibitory effect on on biofilm formation of Escherichia coli and Staphylococcus aureus. This study can provide a scientific basis for the application of GSE and GNE in biofilm control.

Key words: grapefruit seed extract; nanoemulsion; Escherichia coli; Staphylococcus aureus; biofilm