Identification, Culture Condition Optimization And Bioactive Product Separation Of Actinomycete FRo2from Dongxiang Wild Rice

Endophyte is of affinity and plant growth-promoting ability to the host plant,moreover, its rich metabolites can be used for biological control. So far, about75%of the antibiotics were produced by actinomycetes. Dongxiang wild rice (Oryzarufipogon) is an important germplasm resource, and its unique and rich endophyte is avaluable biological resources. Endophytic actinomycetes screened from Dongxiangwild rice (Oryza rufipogon) can be used as the the agricultural antibiotic developmentof new resources.Four actinomycetes (FRo1, FRo2, FRo3and the FRh1) from Dongxiang wildrice were subjected to preliminary taxonomy, physiological and biochemical tests, andidentification including morphological and culture characteristics and phylogeneticanalysis based on16S rDNA gene sequence analysis. FRo1、FRh1、 FRo2and FRo3were identified as Streptomyces venezuelae, Streptomyces hygroscopicus,Streptomyces rochei and Streptomyces carnosus respectively. The fermentation brothof four actinomycetes has a wide antibacterial spectrum, and strain FRo2is ofantagonistic activity for the mainly seven plant pathogenic fungi and displayspotential of development prospects.The physiological and biochemical tests results of strain FRo2showed thatoptimum growth temperature and pH is30C and pH8respectively. It could grow inISP2liquid medium containing NaCl concentration varied from0to12%(W/V).Strain FRo2could hydrolysis starch, gelatin liquefaction, Milk peptonization but notsolidification, Nitrate not deoxidation, Cellulose hydrolysis, no melanin productionand no H2S formation.It could used D-xylose、D-fructose、rhamnose、D-arabinose andD-glucose but not raffinose. Its chemotaxonomic properties further demonstrated thatthe stains FRo2is Streptomyces rochei.The antimicrobial substance had good stabilityand inhibitory activities under the condition of50~100C, pH2~10and UVirradiation.In addition, it couldn t be degraded by proteases.The fermentation medium and fermentation conditions of the strain FRo2wereoptimized by Suppression of mycelial growth results. The optimal mediumcomposition：corn starch4%, Soybean1%, starch2%, K2HPO40.025%, CaCO30.3%,KNO30.025%and NaCl0.05%。The optimal cultural conditions were mediumvolume50mL/250mL, initial pH8.0, inoculum volume10%, temperature28C, and rotation speed120r/min. The average inhibition rate of fermentation broth on F.graminearumt was81.7%.Through different solvents extract, a variety of column chromatography,bioautography, thin layer chromatography and other ways, an active component wasseparationed and purificationed from strain FRo2. The spectra data of GC-MS,1HNMR and14C NMR were used to identify the compound. The molecular formula ofthe active compound was confirmed as Dibutyl phthalate (C16H22O4).