Antifungal Activity of Menthol, Eugenol and Their Combination against Aspergillus ochraceus and Aspergillus niger In Vitro and in Stored Cereals

Aspergillus ochraceus and Aspergillus niger are spoilage and mycotoxin-producing fungi that can contaminate agricultural commodities and derived products. In the present study, menthol, eugenol, and their combination (mix 1:1) were tested to determine their contact and fumigation toxicity against the two fungi. Menthol, eugenol, and their mixture significantly reduced mycelial growth and spore germination at concentrations from 300 to 600 µg/mL, and the inhibitory effects showed clear dose dependence. The minimum inhibitory concentration (MIC) values against A. ochraceus were 500 µg/mL (menthol), 400 µg/mL (eugenol), and 300 µg/mL (mix 1:1), while the MIC values for A. niger were 500 µg/mL (menthol), 600 µg/mL (eugenol), and 400 µg/mL (mix 1:1). Additionally, the analyzed compounds exhibited more than 50% protection against A. ochraceus and A. niger by fumigation of stored cereal grains (maize, barley, and rice) in sealed containers. The binary mixture of menthol and eugenol showed synergistic effects against both fungi in both in vitro direct contact and stored grain fumigation trials. The results of the present study provide a scientific basis for the application of a combination of natural antifungals in food preservation.