Effects of Tropical Citrus Essential Oils on Growth, Aflatoxin Production, and Ultrastructure Alterations of Aspergillus flavus and Aspergillus parasiticus

Ethyl acetate extracts and hydrodistillated essential oils from five cultivars of tropical citrus epicarps were evaluated for their inhibitory activities against Aspergillus fumigatus, Aspergillus niger, Aspergillus flavus, Aspergillus parasiticus, and Penicillium sp. using disk diffusion and broth microdilution assays. Essential oils prepared from kaffir lime (Citrus hystrix DC) and acid lime (Citrus aurantifolia Swingle) epicarps exhibited stronger antifungal activity to all fungi than their ethyl acetate extracts with minimum inhibitory concentration and minimum fungicidal concentration values of 0.56 and 1.13 mg/ml (dry matter), respectively, against aflatoxin-producing A. flavus and A. parasiticus. The dominant components of the essential oil from kaffir lime were limonene, citronellol, linalool, o-cymene, and camphene, whereas limonene and p-cymene were major components of acid lime essential oil. Pure limonene, citronellal, and citronellol were five to six times less fungicidal than the natural essential oils, indicating the synergistic activity of many active compounds present in the oils. Kaffir and acid lime essential oils significantly reduced aflatoxin production of A. flavus and A. parasiticus, particularly lime essential oil, which completely inhibited growth and aflatoxin production of A. flavus at the concentration of 2.25 mg/ml. Target cell damage caused by acid lime essential oil was investigated under transmission electron microscopy. Destructive alterations of plasma and nucleus membrane, loss of cytoplasm, vacuole fusion, and detachment of fibrillar layer were clearly exhibited in essential-oil-treated cells.