Abstract: Aspergillus flavus poses a serious threat to food safety and human health, making the development of green and efficient antimicrobial compounds crucial. This study therefore focuses on the natural monoterpene compound α-terpineol, preliminarily investigating its inhibitory effect on A. flavus growth and potential mechanisms. The minimum inhibitory concentration (MIC) of α-terpineol against A. flavus was determined using a gradient concentration addition method. Subsequently, Aspergillus flavus was treated with α-terpineol at concentrations of 0, 1/4MIC, 1/2MIC, and MIC. The antibacterial mechanism was analyzed through measurements of mycelial dry weight, observations of cell wall and cell membrane integrity, and assessments of relative electrical conductivity, pH, total lipid content, and malondialdehyde (MDA) levels. Concurrently, in vitro antibacterial tests were conducted using peanuts as the test substrate. Results indicated α-pinene exhibited an MIC of 4 μL/mL against A. flavus. Compared with the control group, treated samples showed significantly reduced mycelial dry weight and impaired cell wall/membrane integrity. Extracellular relative electrical conductivity increased while pH decreased. Total lipid were markedly reduced. At the MIC concentration, A. flavus growth on peanuts was completely inhibited. In summary, α-terpineol effectively inhibits Aspergillus flavus growth by disrupting its cell wall and membrane integrity. This demonstrates its potential application in food mold prevention and provides theoretical support for developing green antimicrobial.