|Budget Amount *help
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1990: ¥200,000 (Direct Cost: ¥200,000)
Fiscal Year 1989: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1988: ¥1,400,000 (Direct Cost: ¥1,400,000)
1. After screening many antibiotics-producing strains, 11 strains (Streptomyces longwoodensis, S. tubercidicus, S. antibioticus, S. phaeofaciens, S. toyocaensis, S. celluloflavus, S. sclerogranulatus, S. clavurigerus, S. griseus and Bacillus brevis) were found to produce VB-like inducing factors. The VB activity of them were 500 units/ml, 500 units/ml, 100 units/ml, 100 units/ml, 5 units/ml, 5 units/ml, 4 units/ml, 1 unit/ml, 0.5 unit/ml and 0.2 unit/ml, respectively.
2. By C_<18> reverse phase HPLC using linear gradient elution from 20% to 80% CH_3CN, natural inducing factors VB-A and VB-B were eluted at 12 min and VB-C at 8 min. Under the same conditions, active compounds from the above 11 strains were separated. Of them, activity of S. lincolnensis, S. clavurigerus and B. brevis was found to be constituted from only one component, that of S. tubercidicus, S. phaeofaciens and S. toyokaensis was from 2 components, that of S. longwoodensis, S. celluloflavus and S. sclerogranulatus was f
rom 3 components, and that of S. antibioticus and S. griseus was from 4 components.
3. To confirm that the activity deected in those organisms was actually due to VB-like compounds, S. antibioticus was selected and screened for higher producing strains and high producer strain NF-18 was obtained. After optimization of carbon and nitrogen sources of cultivation medium, conditions to produce 5000 units/ml were found. This amount of inducing activity was 50-fold higher than that produced by the wild type S. antibioticus. From the 5-liter culture broth of S. antibioticus NF-18, 2.3mg of pure inducing factor was isolated by ethyl acetate extraction, active charcoal column chromatography and C_<18> reverse phase HPLC. By usual spectroscopic analyses (NMR, IR, MS and CD), the compound was identified as virginiae butanolide A which was previously isolated from S. virginiae.
4. Several VB analogs were chemically synthesized and their effectiveness on inducing virginiamycin production were compared leading to the establishment of the essential partial-structure in VB to exert its inducing activity. Less