Screening Of 11 Medicinal Fungi In Nyingchi For Antibacterial Activity And Preliminary Study On The Active Components Of Laetiporus Sulphureus

In order to develop new agricultural antibiotics,this study used 6kinds of plant pathogenic fungi（Rhizoctonia cerealis,Alternaria solani,Bipolaris sorokiniana,Fusarium on potato,Sclerotinia sclerotiorum,and Fusarium graminearum）as target fungi,and compared antibacterial activity of 11 kinds of medicinal fungi fruiting bodies in Tibet Nyingchi area with tablet confrontation and hyphal growth rate method.Among these fruiting bodies,the crude extract of Laetiporus sulphureus fruiting body has better antibacterial effect on Fusarium graminearum and Fusarium on potato.Besides,the alkaloids components represented by betaine were determined as the active material base.Bacteriostatic and virulence tests of alkaloids components in Laetiporus sulphureus fruiting body were carried out with commercial pesticides Carbendazim and Chlorothalonil as controls.This study established a certain foundation for the research and development of the biological antibacterial agent using Laetiporus sulphureus as raw materials.（1）This part of study determined preparation method of crude extract of 11 kinds of medicinal fungi fruiting bodies collected in Nyingchi area,including Trichaptum pargamenum,Calvatia caelata（Bull.ex DC.）Morg.,Daedaleopsis confragosa（Bolton）J.Schr?t.,Lactarius piperatus,Pholiota lubrica（Fr.）Sing.,Coriolus versicolor,Lycoperdon perlatum Pers.,Pholiota terrigena and Craterellus aureus,with antibacterial activity for 6 kinds of pathogenic bacteria as the index.The results showed that the optimal preparation method was as follows:95%ethanol was the solvent;the solid-liquid ratio was 50:1;fruiting body was extracted at 89℃for 7 hours with Soxhlet extraction method,and the extraction solution was filtered and freeze-dried.（2）The preliminary screening test was carried out to measure antibacterial activity of 11 medicinal fungi fruiting bodies,and the results showed that the inhibition rate of Trichaptum pargamenum against Rhizoctonia cerealis was 80.87%;that of Lactarius piperatus against Fusarium graminearum was 83.43%;the inhibition rates of Craterellus aureus against Alternaria solani,Bipolaris sorokiniana and Sclerotinia sclerotiorum were 83.17%,80.51%and 83.16%,respectively;that of Laetiporus sulphureus against Fusarium graminearum and Fusarium on potato were 85.75%and 82.69%respectively when the concentration of crude extract of fruiting body is at 100 mg/m L.（3）Double screening for the antibacterial activity of plant pathogen fungi and toxicity test were carried out,and the results showed that the inhibition rates of crude extract of Laetiporus sulphureus fruiting body against Fusarium graminearum and Fusarium on potato were 86.27%and 85.39%respectively when the concentration of crude extract of fruiting body is at 100 mg/m L;the EC₅₀is 14.50 mg/m L,13.73 mg/m L respectively.At present,there are few studies on the bacteriostatic activity of Laetiporus sulphureus against these two pathogenic fungi,and there is not relevant reports.The Laetiporus sulphureus has been identified as target medicinal fungi for bacteriostatic active component analysis.（4）Through literature and experimental studies,the alkaloids components represented by betaine in the fruiting body of Laetiporus sulphureus were determined as its active components,and their contents were determined by HPLC.The content of betaine in fruiting body of Laetiporus sulphureus was 0.79%.（5）The bacteriostatic rate and active concentration of alkaloids components（containing 0.79%betaine）in the fruiting body of Laetiporus sulphureus were measured with Commercial pesticides Carbendazim and Chlorothalonil as controls.Results showed that inhibition rates of Alkaloids components containing 0.8 mg/m L betaine against Fusarium graminearum and Fusarium on potato were 88.16%and 86.23%respectively,which were consistent with the toxicity test results.The inhibition rates of Laetiporus sulphureus against Fusarium graminearum have not increased when concentration of alkaloidal betaine have increased to 1.6 mg/m L or 3.2 mg/m L,and there was no significant difference between the results of two concentrations（P>0.05）.The above results indicated that the maximum bacteriostatic rate of the extract of Laetiporus sulphureus fruiting body against Fusarium graminearum was88.42%;the inhibition rates of Laetiporus sulphureus against Fusarium on potato have not increased when concentration of alkaloidal betaine have increased to 1.6 mg/m L or 3.2 mg/m L,and there was no significant difference between the results of two concentrations（P>0.05）;the maximum bacteriostatic rate of the extract of Laetiporus sulphureus fruiting body against Fusarium on potato was 86.46%.（6）Optical microscope was used to observe the bacteriostatic effect of Laetiporus sulphureus fruiting body extract on mycelium of pathogenic fungi.The results showed that compared with the control group,the Fusarium graminearum mycelium treated by the extract of Laetiporus sulphureus fruiting body became shallow,thinner,atrophic and deformed.Besides,The mycelium of Fusarium on potato treated became shallow and deformed,and its growth quantity decreased and its morphology and structure changed.（7）The bacteriostatic effect of alkaloids from crude extract of Laetiporus sulphureus fruiting body were studied with betaine as reference substance,the results showed that the inhibition rates of alkaloids components containing 100ug/m L betaine on Fusarium graminearum and Fusarium on potato were 47.27%and 43.48%respectively and the inhibition rates of betaine reference substance on Fusarium graminearum and Fusarium on potato were 34.66%and29.54%respectively when its concentration is 100ug/m L.The above results indicated that betaine is only one of the active substances in the fruiting body of Laetiporus sulphureus,so other active ingredients need to be further studied.