Purification and characterization of antifungal compounds produced by Bacillus subtilis KS1

A bacterial strain, KSl, possessing strong antifungal activity was isolated from soil samples of ginseng fields and identified as Bacillus subtilis. In greenhouse test, the culture filtrate of B. subtilis KS1 showed strong protective effect against several fungal diseases of agricultural plants such as cucumber gray mold and wheat leaf rust. In addition, the crude butanol fraction of the culture filtrate exhibited antagonistic effect against several fungi including plant or human pathogens, such as Botrytis maydis, Chytridium lagenarium and Candida albicans. The antifungal compound, SW1, produced by B. subtilis KS1 was purified through consecutive chromatographic separations on a pep-RPC column and a Bondapak reverse phase column. Temperature and pH showed little effect on the stability of the compound in the ranges and pH 4.0-10.0, respectively. The composition and structural characteristics of SW1 were analysed by HPLC and by , NOESY, COSY-NOESY and HOHAHA NMR spectroscopy, respectively, which revealed that the compound belongs to iturin A, a typical cyclic antifungal compound produced by B. subtilis. In contrast to the previously reported iturin A compounds which have one or no side chain in the hydrophobic hydrocarbon chain of acids, SW1 was shown to have a acid containing 12-carbon skeleton with two side chains.