Abstract: Fungal contamination is a concern of food industry. Aspergillus flavus(A. flavus) spores can resist food sterilization and produce aflatoxin which is toxic to animals and humans. Therefore, it is advisable to explore technologies of inactivating spores and toxins without affecting food quality. In this paper, the spores of A. flavus were treated by a low temperature plasma device produced by dielectric barrier discharge for different time. Firstly, the inhibition effect of plasma on A. flavus spores was studied. Compared with the control group, the inhibition effect of plasma on A. flavus spores was confirmed by inverted microscope and microplate analysis. The mycelial growth and of spore germination were inhibited after 30 s treatment. When the treatment time reached 90 s, A. flavus spores aggregated. It was speculated that A. flavus spores were damaged by oxidative stress. This discovery provides a new idea for the subsequent study on the inactivation mechanism of plasma against A. flavus. The scanning electron microscopy chracterization showed that the spore surface was deformed and shrunk after plasma treatment for 30 s, and the degree of deformation was strengthened with the extension of treatment time. When the treatment time was increased to 90 s, the spore morphology of A. flavus changed significantly compared with the control group, such as depression and shrinkage, and the cell surface tended to be smooth, and the number of membrane proteins decreased. These results showed that plasma treatment changed the morphology of A. flavus spores, and this effect was dose-dependent and increased with treatment time. Therefore, it is of great significance to study the effect of plasma on the cell membrane of A. flavus to explore its antifungal mechanism. Cell membrane permeability plays an important role in maintaining cell microenvironment and normal metabolism. After plasma treatment, A. flavus spores showed leakage of cellular constituent, decrease of pH and increase of conductivity, which indicated that the cell membrane of A. flavus spores was damaged and the permeability of cell membrane was increased, which was one of the main reasons for the inhibition of plasma on A. flavus. Through the analysis of fluorescence microscope and fluorescence spectrophotometer, it was confirmed that a large number of reactive oxygen species (ROS) were produced in the treated cells, indicating that after plasma treatment, spores suffered oxidative damage, which might be one reason for the inactivation of A. flavus spores. In conclusion, the fungal spores can be inactivated in a few minutes by dielectric barrier discharge low temperature plasma treatment.