Strctural Identification Of Secindary Metabolically Active Substance Of Streptomyces Albosponus CT205 And Its Antagonistic Effect Against Fusarium Oxysporum

Antibiotics are the secondary metabolism substances produced by microorganisms.They are widely used in agriculture and medicine.Streptomyces albospinus CT205 is an actinomycete strain screened by our laboratory,which has strong antagonistic activity against many plant pathogenic fungi.Field experiments showed that the biocontrol agent from strain CT205 had better control effect on soil-borne diseases such as strawberry root rot and cucumber Fusarium wilt.In order to explore its prevention and control mechanism,firstly,the active substance from fermentation broth of S.albospinus CT205 were extracted,purified and identified.Secondly,the influence of environmental factors on the production of active substance from strain CT205 was studied.Finally,the synergistic effect of active substance from CT205 liquid fermentation and 2-phenylethanol,the main volatile component produced by CT205 solid fermentation,on the inhibition of Fusarium oxysporum f.sp.cucumber(F.oxysporum)was explored.The results are as follows:S.albospinus CT205 was fermented using the liquid shake flask,and the supernatant of fermentation was centrifuged,extracted with ethyl acetate,concentrated under vacuum,then active substance was dissolved with methanol and collected.Thin layer chromatography and biological development was used to determine Rf value of the active substance.The compound was obtained by the separation of prepared silica gel plate.Ultraviolet scanning showed that the active substance had the maximum absorption peak at ?=215 nm.UPLC was used to separate and determine the absorption peak position of the active substance.HPLC-TOF-HR-MS analysis showed that the molecular weight of the active substance was 282.2.The molecular weight of the active substance was the same as that of actidione.1H NMR and 13C NMR were used to detect the skeleton structure of active substance.FTIR was used to confirm the existence of functional groups(carbonyl,hydroxyl,amino,etc.)of active substance.The substance was identified as actidione,also known as Cycloheximide.A chemical method for rapid determination of the active substance in fermentation broth was established,and the content of actidione in fermentation broth was determined to be 37.95 mg/L.The effects of fermentation conditions on S.albospinus CT205 liquid fermentation were studied.The different initial pH,fermentation temperature and fermentation period affected the content of active substance produced by S.albospinus CT205 fermentation.When the initial pH of liquid fermentation was 8.0,the fermentation temperature was 28 C,and the fermentation period was 96 h,the actidione content in fermentation broth was the highest.The effects of main components of cucumber root exudates(glucose,oxalic acid and glutamic acid)on S.albospinus CT205 liquid fermentation were studied.The results showed that adding appropriate glucose and oxalic acid could increase the yield of actidione,but adding glutamic acid had no significant difference in the yield of actidione.The inhibition effects of actidione and 2-phenylethanol on F.oxysporum were explored.It was found that actidione and 2-phenylethanol could inhibit the growth activity of F.oxysporum when they individually treated F.oxysporum,and the combination of actidione and 2-phenylethanol could synergistically inhibit F.oxysporum.When treated with 40 ?g/mL of actidione and 25 ?L 2-phenylethanol,the relative inhibition rate of mycelial growth of F.oxysporum was 82.49%,and the mycelium did not grow at all;when treated with actidione 8?g/mL and 2-phenylethanol 30 ?L,the inhibition rate of spore germination of F.oxysporum was 100%,and the effect of complete inhibition was achieved.In this study,the secondary metabolism active substance produced by S.albospinus CT205 was identified as actinone,and a chemical detection method for active substance content in fermentation broth was established.It was found that different environmental factors had a great influence on the content of active substance produced by strain CT205 fermentation,and it was confirmed that actidione and 2-phenylethanol had synergistic inhibition effect on F.oxysporum.The results provide a theoretical basis for the preparation of safe and efficient biocontrol agents for the prevention and control of soil-borne wilt.