Study On Technical Optimization Of Increasing Nucleosides Content In Fruiting Bodies Of Cordyceps Chanhua

Cordyceps chanhua,as a traditional and precious Chinese medicinal material,is a kind of entomogenous fungus,which is homology of medicine and food.Its coremiums（fruiting bodies）and host cicadae can be used as compatibility of traditional Chinese medicine or as health care products.The natural Cordyceps chanhua cannot meet the growing demand of the market.As an important medicinal part of Cordyceps chanhua,fruiting bodies have good substitutability for natural Cordyceps chanhua.Therefore,artificial cultivation of Cordyceps chanhua fruiting bodies has become the focus of cordyceps resource research.Cordyceps chanhua has many active ingredients,among which nucleosides have pharmacological functions such as anti-neuroinflammation and antioxidation,which can improve memory and delay neural degeneration.Based on the above,this study conducted a test on the cultivation of Cordyceps chanhua fruiting bodies with high contents of nucleosides.Using high performance liquid chromatography（HPLC）to detect the content of five nucleosides in the fruiting bodies,including uridine,guanosine,adenosine,cordycepin,and N6-（2-hydroxyethyl）adenosine（HEA）,to screen a strain of Cordyceps chanhua with high yield of nucleosides,to optimize the solid culture medium of Cordyceps chanhua,and then this study explore the protective effect of Cordyceps chanhua on degenerative neurological diseases.The main results are as follows:1.A strain of Cordyceps chanhua with high activity and high nucleosides content was screened.Using wheat as a substrate,different strains of Cordyceps chanhua were cultivated,and morphological characteristics such as fruiting body shape and color were observed.Growth indicators such as fruiting body height,budding time,lodging time,growth rate,fresh weight,water content,and biological conversion rate were observed.HPLC was used to detect the content of uridine,guanosine,adenosine,cordycepin,and HEA in the fruiting bodies.Growth indicators such as growth rate,fresh weight,and biological conversion rate of the fruiting body were combined,this study screened and evaluated four strains.The results showed that GZUIFR-3716（abbreviated as 3716）was significantly higher than other Cordyceps chanhua in terms of total nucleosides content（P<0.0001）,fresh fruit weight（P<0.05）,and biological conversion rate（P<0.0001）.2.To optimize the extraction process of nucleosides from Cordyceps chanhua fruiting body with the aim of fully extracting nucleosides.During the preparation of the extract,this study investigated the effects of fruiting body processing methods,extraction methods,and extraction solvents on the content of nucleosides.After optimization,the extraction method was as follows:Adding 0.5%phosphoric acid solution to the Cordyceps chanhua fresh fruiting body,ultrasound extraction（40 k Hz,500 w）for 30 minutes,followed by a 100℃water bath for 1 hour.Compared with the original extraction solution,the optimized extraction solution has a 1.72 fold improvement in uridine content,4.63 fold improvement in guanosine content,1.25fold improvement in adenosine content,0.43 fold improvement in HEA content,and a3.4 fold improvement in total nucleosides content.3.In order to improve the yield of nucleosides in the fruiting body of Cordyceps chanhua,the conditions for the cultivation of Cordyceps chanhua were optimized.A single factor cultivation experiment was conducted using different carbon and nitrogen sources.Based on the results of the single factor experiment,a L9（3⁴）orthogonal cultivation experiment was designed using composite carbon and nitrogen sources as four factors,with three levels set for each factor.The yield of uridine,guanosine,adenosine,cordycepin,and HEA,under different cultivation conditions,was detected by HPLC to determine the optimal ratio of carbon and nitrogen sources.The optimized cultivation nutrient conditions were as follows:The nutrient solution of the culture medium was composed of 12 g sucrose,10 g yeast extract,50 g soybean powder,1 g KH₂PO₄,1 g Mg SO₄,10 tablets of vitamin B1,1 g citric acid,and 1000m L water.The total nucleosides content increased by 38.5%compared to the initial conditions.4.To explore the degenerative neuroprotective effect of Cordyceps chanhua.Using the wild type of Caenorhabditis elegans N2 and the Alzheimer’s disease transgenic Caenorhabditis elegans CL4176 as biological models,the effects of Cordyceps chanhua on delaying senility of N2 and decreasing Aβdeposition of CL4176 were evaluated by observing the swallowing,movement,paralysis,and reproduction of the Caenorhabditis elegans.The results showed that Cordyceps chanhua had the effects of promoting the reproduction of CL4176（P<0.0001）and delaying the decline of movement rate of N2（P<0.05）.