Studies On Antifungal Active Substance Of A Bacillus Subtilis Strain

Bacillus subtilis is considered to be a useful rhizobacteria, which can suppressthe growth of plant fungal pathogens and promote the growth of plant. So far, a seriesof lipopeptide complexes produced by Bacillus subtilis have been found haveantifungal and antimicrobial activities, including surfactin, iturin and fengycin. Weisolated two Bacillus sp. RJ-1 and RJ-2. By the methods of traditional strainidentification and 16S ribosomal DNA sequencing, both of the two strains wereidentified as Bacillus subtilis. When cultivated with plant fungal pathogen Fusariumoxysporum f.sp.cucumerinum on PDA plate, RJ-1 did not show any activity tosuppress the growth of F.o.c, while RJ-2 showed strong ability to suppress the growthof F.o.c. Compared the extract of RJ-1 with RJ-2 by HPLC, we found RJ-2 producedcertain substance that RJ-1 did not. Then we used LC-MS to analyze the substanceselectively by the result of the HPLC analysis, and found that the possible molecularweight of the main substance were 1477.0, 1489.1 and 1505.1. By the method ofpeptide mass fingerprinting, we identified them as C_17-18 Fengycin A or C_16-18Fengycin B. In order to confirmed the conjecture, high precision mass spectrumMALDI-TOF MS was used to make a further analysis of the RJ-2 extract at the rangeof 900¹600Da where Bacillus subtilis lipopeptide lay in. Then we found a group ofhomologues whose molecular weight were 1462.8,1476.8, 1490.8,1504.8 and 1518.8.By the method of peptide mass fingerprinting, we identified them as C_14-18 FengycinB, and the content of C₁₇ Fengycin B is the highest of them. We also found anothergroup of homologues whose molecular weight were 1432.8,1446.8,1460.8, 1474.8and 1488.8, which were identified as a isomer of C_14-18 Fengycin A, lacked 2H incarbon chain when compared with fengycin A homologues.