Study On The Mutagenesis Of Grifola Frondosa And The Production Of Mycelium And Polysaccharides From Rice Bran And Wheat Bran By Liquid Fermentation Of The New Strain

Grifola frondosa is a medicinal and edible fungus with multiple chemical components and pharmacological effects.Rice bran and wheat bran are low-value,cheap,renewable,and nutrient-rich agricultural products processing by-products.These valuable by-products can further expand the application.In this study,the valuable mycelia and polysaccharides were produced by transforming the whole material of the rice bran and wheat bran with the liquid fermentation of the new strain after mutagenesis,providing a way for the high-value utilization of the rice bran and wheat bran.The detailed contents and results are as follows:（1）UV mutagenesis and ARTP mutagenesis were used to improve the species of Grifola frondosa.After mutagenesis improvement,UV mutagenesis strain GFU6 and ARTP mutagenesis strain GFA2 were obtained,which had faster growth rate and higher conversion efficiency of rice bran and wheat bran than the original strain.The mycelial growth rate of the original strain,GFU6 and GFA2 strains was 4.918mm/d,5.775mm/d,6.008mm/d.After liquid fermentation,the dry weight of mycelium was1.9980g/100mL,2.1865g/100mL,2.2259g/100mL.Mycelial polysaccharides was244.40mg/100mL,263.05mg/100mL,265.33mg/100mL.Compared with the original strain,the mycelial growth rate of GFU6 and GFA2 strains were increased by 17.24%and 22.16%,the dry weight of mycelium increased by 9.43%and 11.41%,and mycelial polysaccharides increased by 7.63%and 8.56%.It can be found that GFA2strain has the fastest growth rate and fermentation performance among the three strains.（2）Optimization of liquid fermentation medium of Grifola frondosa GFA2strain.The single factor and U₁₁（11³）uniform experiments design of rice bran,wheat bran,potassium dihydrogen phosphate and magnesium sulfate heptahydrate were conducted to analyze the results of the fermentation index mycelial dry weight,mycelial polysaccharides and extracellular polysaccharides and obtain regression model equation,from which the optimal medium composition is obtained.When the dry weight of mycelium or mycelium polysaccharide was used as the fermentation index,the 100mL optimized medium has wheat bran 12g,rice bran,potassium dihydrogen phosphate and magnesium sulfate heptahydrate were all 0g,they all reached the maximum value 2.726g and 590.352mg.when using extracellular polysaccharide as the fermentation index,the 100mL optimized medium has rice bran0g,wheat bran 12g,potassium dihydrogen phosphate 0.15g,magnesium sulfate heptahydrate 0.02g,the maximum value of extracellular polysaccharide was2601.252mg.（3）Polysaccharides from Grifola frondosa mycelium were extracted by ultrasonic synergistic compound enzyme method.Using the yield of mycelium polysaccharide as the evaluation index,the technological conditions for extracting mycelium polysaccharide by compound enzymatic method were studied,and the ratio of material to liquid was 1:30（g/mL）,the pH of enzymolysis was 4.5,the temperature of enzymolysis was 50℃,and the amount of enzyme was 2.0%,the enzymolysis time of 50min is the best condition for the extraction of polysaccharides by complex enzymatic method,and the polysaccharide yield is 15.12%.The ultrasonic conditions were optimized on the basis of compound enzymatic extraction.The single factor analysis of variance showed that the main factors affecting the yield of mycelium polysaccharides were ultrasonic temperature,enzymatic hydrolysis pH and ultrasonic time.After the response surface optimization experiments,the optimal extraction process conditions of mycelium polysaccharides were as follows:ultrasonic temperature 50℃,enzymatic hydrolysis pH4.5,ultrasonic time 40min,material-liquid ratio 1:30（g/mL）,enzyme addition amount 2.0%,ultrasonic frequency was synchronous dual-frequency 28/40kHz,Ultrasonic power 240W.Under this condition,compared with hot water extraction and compound enzymatic extraction of mycelial polysaccharides,polysaccharide yield increased by 8.84%and 3.37%.（4）Detection of heavy metal contaminants in rice bran and wheat bran and Grifola frondosa mycelium.The content of lead,arsenic,mercury,copper,cadmium and chromium in heavy metal pollutants was detected according to the national standard method.The test results showed that the content of lead,arsenic,mercury and cadmium in rice bran and wheat bran were within the limits of the national standard.The content of copper in rice bran is within the standard range,while the content of copper in wheat bran is 15.7mg/kg,which exceeds the limit range.The chromium content in rice bran and wheat bran exceeds the national standard limit range.The heavy metal detection of Grifola frondosa mycelium before and after citric acid treatment found that the contents of heavy metals arsenic,mercury and cadmium before and after treatment were within the limits of the national standard,and the contents of lead,copper and chromium all exceeded the limit.However,compared with before treatment,the contents of lead,copper,and chromium were reduced by24.69%,18.75%,and 9.94%,indicating that citric acid immersion has a certain effect on the removal of heavy metals,but if the content of heavy metals in the mycelium is to be reduced to the limit of the national standard,the method for removing heavy metals needs to be improved.