Bacillus subtilis N1 showed antagonistic effect against Curvularia sp., a causal agent of leaf spot on Siam tulip. The aims of this research were to verify the optimal conditions for production of antifungal substances by B. subtilis N1 and to purify the compound. In order to obtain the optimal conditions to produce the antifungal substances, factors which were types of cultivation medium (NB, LB and TSB), pH of the medium (6, 7, 8 and 9), temperatures of cultivation (30, 37 and 40 degree Celsius) and cultivation times (0 – 24 hours) were investigated. The results showed that the optimal cultivation conditions for B. subtilis N1 were grown in TSB medium, at pH 7, at 37 degree Celsius, for 12 hours. The antifungal substances were non-growth associated product since it was produced during stationary phase of growth. The active compound was thermostable at 20 – 121 degree Celsius for 20 minutes. The activity remained unchanged when exposed to pH ranged from 2 – 10. Separation of protein by 20 - 40 % ammonium sulfate precipitation gave partial purified proteins with specific activity 379.82 AU/mg. SDS-PAGE analysis revealed mixed proteins with major four bands at 61.38, 41.98, 36.14 and 22.13 kDa. MICs of culture filtrate and partial purified protein were 350 µg/ml and 23.43 µg/ml, respectively. The later exhibited stronger activity by 14 folds more than the culture filtrate. Observation under light microscope indicated that the antifungal proteins caused severe alteration in cell morphogenesis. It caused abnormal thickenings, swollen and rounded hyphae. These results revealed that B. subtilis N1 and its antifungal substances might be used as a biocontrol agent against Curvularia sp. to prevent damage from this fungal infection on Siam tulip production.
