Parameter Optimization Of Culture Processing Of Multigrain Cordyceps Militaris Hyphae Symbionts

ObjectiveTo explore the optimal extraction conditions of main functional components in multigrain cordyceps militaris hyphae symbionts and make the content analysis under different culture conditions, and to optimize the parameters of the culture processing of multigrain cordyceps militaris hyphae symbionts, it could provide a scientific basis for the pilot production process and the substantive examination of invention patent application. MethodsThe optimal extraction and content analysis of cordycepin, adenosine, cordycepic acid, cordyceps polysaccharide and total flavonoids in multigrain cordyceps militaris hyphae symbionts were determined by single factor experiments combined with response surface methodology under different culture conditions of inoculums concentration, solid-liquid ratio, ratio of medium, duration of illumination and milk powder amount, and the predominant influential factors of the above five indicators were determined. The functional based method was used to synthesize the five indicators into a aggregative indicator-comprehensive value Y, taking Y as response value, the three predominant influential factors were optimized and analyzed by Box-Behnken test and the optimal culture process was determined. The functional components were analyzed by high performance liquid chromatography and spectrophotometry. SPSS 17.0 software was applied for statistical analysis, and one-way analysis of variance and SNK test were used to analyze the difference between groups. The values of α=0.05 were regarded as statistically significant. Results1. The optimal extraction conditions of cordycepin in multigrain Cordyceps militaris hyphae symbionts were: extraction time 6.59 h, extraction temperature 81.9 °C, solid-liquid ratio 1:90, the contents of cordycepin and adenosine were 0.5309±0.0114~11.9629±0.0519 mg/g and 0.1597±0.0022~0.5590±0.0064 mg/g with this condition, and were significantly different(P<0.0001) under different culture conditions. Inoculums concentration, duration of illumination and milk powder amount were closely related with the content of cordycepin. Duration of illumination, milk powder amount and solid-liquid ratio were closely related with the content of adenosine.2. The optimal extraction conditions of cordycepic acid in multigrain Cordyceps militaris hyphae symbionts were: solid-liquid ratio 1:26.42, extraction time 3 h, extraction temperature 35 °C, the contents of cordycepic acid were 0.5309±0.0114~11.9629±0.0519 mg/g with this condition, and were significantly different(P<0.0001) under different culture conditions. Inoculums concentration, duration of illumination and milk powder amount were closely related with the content of cordycepin.3. The optimal extraction conditions of cordyceps polysaccharide in multigrain Cordyceps militaris hyphae symbionts were: microwave power 568.63 W, solid-liquid ratio 1:23.18, microwave time 120 s, the contents of cordyceps polysaccharide were 69.7736±3.0390~208.2109±5.5284 mg/g with this condition, and were significantly different(P<0.0001) under different culture conditions. Milk powder amount, solid-liquid ratio and duration of illumination were closely related with the content of cordyceps polysaccharide.4. The contents of total flavonoids were 0.5309±0.0114~11.9629±0.0519 mg/g in multigrain Cordyceps militaris hyphae symbionts and were significantly different(P<0.0001) under different culture conditions. Inoculums concentration, solid-liquid ratio and duration of illumination were closely related with the content of total flavonoids.5. Combined with the results of single factor experiments, duration of illumination, inoculums concentration and milk powder amount were identified as significant influential factors, and the optimal culture process of multigrain Cordyceps militaris hyphae symbionts was that duration of illumination was21.68 d, inoculums concentration was 7.93 mL, milk powder amount was 2.94 g, the regression model was very significant(P<0.0001) and the relative error was 2.46% through verification test.ConclusionsThe fungus of Cordyceps militaris can effectively transform multigrain and get multigrain Cordyceps militaris hyphae symbionts, the main functional components are improved by the optimized extraction. The optimal culture process is determined within the scope of the experiment, and it provides a scientific basis for the pilot production process and the substantive examination of invention patent application.