Study On Extraction,separation And Partial Bioactivity Of Total Flavonoids From Pleurotus Eryngii Spent Substrate

With the rapid development of Pleurotus eryngii industry,the Pleurotus eryngii spent substrate is also growing.They are usually treated by discarding,piling up,burning and as renewable resource,and the general treatments cannot effectively solve the problem of waste materials.Therefore,the rational use of Pleurotus eryngii spent substrate has become a challenge that restricted the development of Pleurotus eryngii and the construction of ecological civilization.Some studies have shown that the waste materials contain a large number of nutrients and bioactive components from composts,mycelia,and metabolites of the mycelium.In this study,taking the Pleurotus eryngii spent substrate as raw material and the content of total flavonoids as index,we studied separation and purification of total flavonoids from the Pleurotus eryngii spent substrate and their antioxidation and stability by the ultrasonic assisted method.The results can provide a theoretical basis for the comprehensive utilization of Pleurotus eryngii spent substrate.The main contents and results are the following:(1)According to color reaction and HPLC chromatography test to determine the flavonoids in the extracts of Pleurotus eryngii spent substrate,the single factor experiments were studied respectively.Through it,we made an orthogonal experiment design of 4 factors and 3 levels of the extraction conditions of total flavonoids from Pleurotus eryngii spent substrate.The experiment was optimized to get the optimal conditions for the extraction.The optimum extraction conditions were the ethanol concentration 70%,the ratio of material to liquid 1:35 g/ml,the ultrasonic time 20 min,and the water bath temperature about 70℃.Under these conditions,the total flavonoids of Pleurotus eryngii spent substrate is 2.852±0.015mg/g,RSD=0.38%,and the process is stable.(2)On the basis of inspecting the effects of adsorption and desorption of six kinds of resin,we chose AB-8 macroporous resin to separate and purified total flavonoids from Pleurotus eryngii spent substrate.Based on the single factor experiments,the orthogonal optimization tests for the process of adsorption and desorption were made respectively.The purification process of total flavonoids from Pleurotus eryngii spent substrate was as follows: the flow rate of sample 2 m L/min,p H6,the concentration of the sample solution 0.25 mg/m L,the liquid flow rate 3 m L/min;the eluent concentration 90% and the elution volume 4.5BV.The results showed that the total flavonoids increased from 8.049% to 30.37%,and the purity was increased by 3.77 times.At the same time,through the investigation of repeating use the performance of AB-8 resin,the adsorption rate could reach above 50% after repeating use 7 times,and AB-8 resin is suited to separate the total flavonoids from Pleurotus eryngii spent substrate.(3)Compared with VC,the total reduction ability of purified total flavonoids of Pleurotus eryngii spent substrate and on DPPH,hydroxyl radical,superoxide anion free radical scavenging effect were investigated.When the mass concentration of total flavonoids of the Pleurotus eryngii spent substrate was larger than 1.2 mg/ml,the total flavonoids’ reducing capacity was stronger than VC,in addition,within the scope of choosing the experiment’s mass concentration,the scavenging rate of total flavonoids for DPPH was 68.51%,the scavenging rate for hydroxyl radical was 50.38%,the scavenging rate for superoxide anion was 56.06%,the scavenging activity of total flavonoids of the eryngiibacterial chaff for DPPH and hydroxyl radical was stronger than VC,and weaker than VC for the scavenging activity of superoxide anion,the total flavonoids of Pleurotus eryngii spent substrate had good antioxidant activity.(4)By using the punch method to conduct the anti-bacterial tests on 4 kinds of common bacteria and 3 fungus,we got the minimum inhibitory concentration of the total flavonoids of Pleurotus eryngii spent substrate on 7 pathogenic bacteria: Escherichia coli MIC 40mg/m L,Staphylococcus aureus MIC 20 mg/m L,Salmonella typhi MIC 20 mg/m L,Shigella flexneri MIC 40 mg/m L,Watermelon Rot Pathogen of wheat scab MIC 40 mg/m L,Wheat Scab MIC 40 mg/m L,and Cotton Wilt Fusarium MIC 20 mg/m L,That total flavonoids of Pleurotus eryngii spent substrate had good bacteriostatic activity.(5)After investigating the stability of light,temperature,p H and metal cation for the influence on the total flavonoids of Pleurotus eryngii spent substrate,we obtained the following results that the solution change of the total flavonoids of Pleurotus eryngii spent substrate stored at 4 degrees and 20 degrees for 10 hours was small for the total flavonoids of Pleurotus eryngii spent substrate stored 24 hours under p H6 and stored 7 days with dark environment,the change of K+ and Na+ for the effect of absorbance change was also not big and Cu2+ and Fe3+ for the solution’s absorbance had different effects.