A bis-quaternary ammonium compound (bis-QAC), N,N'-hexamethylenebis(4-carbamoyl-1decylpyridinium bromide) (D-38), exhibited a wide antifungal spectrum and a strong activity against sixteen strains of fungi including nine strains isolated from various kinds of food. The antifungal activity was higher than that of N-dodecylpyridinium Iodide, a typical mono-QAC, and that of the commonly used fungicide, 2-(4-thiazolyl) benzimidazole. The activity of D-38, however, was comparatively low when it was measured by the agar dilution method. It is considered that the hydrophobic D-38 molecules, having two long alkyl chains, interacted with the agar medium, and therefore showed lower activity results. On the other hand, the activity of D-38 against Aspergillus niger IFO 6342 was not significantly influenced by temperature, pH and the initial spore concentration. These results indicate that bis-QACs can exhibit a high antifungal activity whether or not the environmental conditions change.