Method for increasing yield of total flavonoids in Ganoderma lucidum mycelium

A method for improving the yield of flavonoids from Ganoderma lucidum mycelium, comprises the following steps of:

• (1) inoculating Ganoderma lucidum strains into a potato dextrose (PD) liquid fermentation medium comprising 200 g peeled potato, 20 g glucose and 1000 ml water;
• and performing activation culture to obtain a seed liquid;
• preparing an expansin solution by sterilizing a broad bean or cucumber seed with mercury chloride (HgCl2) for 4 to 6 minutes, washing with running water for 5 to 7 hours, culturing in the darkroom for 4 to 6 days at the temperature between 15 and 28° C.;
• precooling seedling hypocotyl apices for 0.5 hours at −20° C., adding a homogenate buffer solution that is pre-cooled to 4° C., filtering with a nylon net having an aperture of 70 μm after homogenate, washing the filter residue with a homogenate buffer solution, adding the filter residue in the homogenate buffer solution, settling for 1 to 3 hours to obtain a settled solution, adding an extracting solution in the settled solution for 44 to 50 hours at 4° C., adding 0.3 to 0.5 g/ml of ammonium sulfate ((NH4)2SO4) in the filtrate while stirring to prevent a partial supersaturation of the (NH4)2SO4, settling for 25 to 30 hours centrifuging for 5 to 10 minutes at 4° C. dissolving the sediment with an acid buffer solution, dialyzing in a dialysis bag with a molecular weight of 3000 Da at 4° C., centrifuging the dialyzate at 20000 g for 10 minutes, and taking the supernatant to obtain the expansin solution;
• (2) inoculating the seed liquid obtained in step (1) into a liquid fermentation medium to perform liquid fermentation culture at the temperature between 25 and 30° C. for 3 to 5 days, then adding an expansin solution further culturing for additional 5 to 7 days, and separating to obtain Ganoderma lucidum mycelium, wherein the liquid fermentation medium has a pH between 5 and 7, and contains lactose, sucrose, soybean powder, peptone, yeast extract, MgSO4. 7H20, KH2P04;
• and
• (3) extracting flavonoids from the Ganoderma lucidum mycelium obtained in step (2), comprising the following steps:
• a) drying Ganoderma lucidum mycelium obtained in step (2) at 65° C. and grinding to power, b) adding ethanol solution up to 70-80% by volume and extracting by ultrasonic at 40 kHz and 200 W for 3 to 5 hours at the temperature between 60 and 70° C., then c) filtrating and separating the filtrate, and repeating 2 to 3 times of steps b) and c) for the precipitates;
• pooling the filtrates to yield the purified flavonoids.