CN103130550A - Culture medium and culture method of male agaric mycelium - Google Patents

Abstract

The invention provides a culture medium and a preparation method of male agaric mycelium, and a method for culturing the male agaric mycelium by using the culture medium. The culture medium comprises rice and a culture solution. As rice is used as the raw material of the culture medium, the raw material is available and is economic, the operation is simple, and the effect is good.

Description

A kind of substratum of phellinus igniarius mycelium and cultural method

Technical field

The present invention relates to edible mushrooms and cultivate the field, relate to specifically a kind of substratum and cultural method of phellinus igniarius mycelium.

Background technology

Bao nurse Phellinus (Phellinus baumii) is commonly called as Phellinus mushroom, mulberry ear, is a kind of perennial medicinal fungi of developing in recent years, records the earliest in the LI Shi-Zhen Compendium of Material Medica.Traditional Chinese Medicine thinks that Phellinus is sweet, flat, and bitter, suffering are returned liver, urinary bladder channel, be used for the treatment of that metrorrhagia, blood are drenched, prolapse of the anus is rushed down blood, under band, amenorrhoea, diarrhea due to hypofunction of the spleen etc.Numerous experimental studies show, the polysaccharide in Phellinus and flavonoid compound have important biological activity, are playing an important role aspect treatment cancer, liver protecting, enhancing immunity, neural reparation.

Free radical is the metabolic product of human body, and generally, human body has the ability to remove the free radical that self produces, and still along with the aging of human body, the ability of removing free radical weakens.If the free radical that produces can not in time be removed, will accelerate body aging, even make human body generation pathology etc.Existing studies show that: the polysaccharide in Medicinal Fungus Phellinus igniarius and Flavonoid substances have good antioxygenation; But present research focuses mostly on aspect the research of Phellinus fruiting body extract and liquid fermenting, and less to the research of solid fermentation phellinus igniarius mycelium and antioxygenation thereof.Wild sporophore mainly is distributed in the forest zones such as Heilungkiang, Shaanxi, and the resource standing stock are rare, cause expensive owing to yielding poorly.The Phellinus sporophore mainly obtains by artificial culture at present, but the artificial culture cycle is long, needs larger manpower financial capacity to drop into.By contrast, fermentation technique has short, the advantage such as controllability good, experimental situation is simple of cycle, is the good approach that obtains the Phellinus secondary metabolite.And liquid fermenting easily pollutes in the process of cultivating, and can not reach the biomass of expection in certain incubation time, bad control artificially in culturing process.

Summary of the invention

One of purpose of the present invention is to provide a kind of substratum of phellinus igniarius mycelium, and this substratum comprises following component:

Rice and nutrient solution, wherein the volume (ml) of the weight of rice (g) and nutrient solution is than being x:(x+(0-30), x 〉=10 wherein;

Wherein nutrient solution contains: 2.5% glucose, 0.3%KH ₂PO ₃And 0.15%MgSO ₄7H ₂O。

Above-mentioned substratum is prepared by the following method:

Configuration nutrient solution (2.5% glucose, 0.3%KH ₂PO ₃, 0.15%MgSO ₄7H ₂O), rice is placed in nutrient solution, autoclaving.

The present invention also provides a kind of method of utilizing above-mentioned culture medium culturing phellinus igniarius mycelium, and the method comprises the steps:

The female kind of Phellinus is inoculated on above-mentioned substratum, is placed in 26 ℃ of biochemical cultivation cases and cultivates, growth cycle 25-30 days;

With cultured mycelium together with substratum as for drying in 30 ℃ of convection oven.

Phellinus igniarius mycelium after above-mentioned rice medium fermentation has oxidation resistant biological action, can be used as foodstuff additive and uses, and improves Nutritive value of food.

Innovation of the present invention:

1, use substratum of the present invention to cultivate, liquid culture has advantages of convenient and easy, save energy, is difficult for polluting relatively.

2, substratum of the present invention, adopt rice as raw material not only material easily get, and very economical, respond well.Very large in the energy consumption of liquid culture medium power, and the present invention cultivates only need standing cultivation at a certain temperature of bacterial classification, save the energy, and growth conditions also is better than liquid culture.Find by anti-oxidant experiment simultaneously, the mycelium of cultivation has stronger antioxidant effect.Nowadays foodstuff additive are subject to extensive concern, and rice is as a kind of food of nutrient safe, and the mycelium of cultivation can be used as additive and improves Nutritive value of food.

Embodiment

(Phellinus baummii Pilat from edible mushrooms branch center, Shanghai, Chinese microorganism strain preservation center, is numbered: Phellinus baumii Pilat3249) the Phellinus bacterial classification.

Embodiment 1-4 prepares substratum

Configure 2.5% glucose, 0.3%KH ₂PO ₃, 0.15%MgSO ₄7H ₂The nutrient solution of O is got respectively 60ml in the 250ml culturing bottle, then takes respectively 30,40,50, the rice of 60g is placed in nutrient solution, soaks 121 ℃ of autoclaving 30min about 6 hours.Rice (profit the village fragrant round-grained rice) is provided by Shang Hairun village agricultural science and technology company limited.

The impact of table 1 rice loading amount on biomass

Annotate: the mycelium after the rate of recovery of biomass refers to finally dry and the ratio of rice loading amount.

The substratum of embodiment 5 use embodiment 3 is cultivated

The Phellinus bacterial classification is longer than on the PDA inclined-plane, and growth cycle is a week.

Get above-mentioned Phellinus bacterial classification and be inoculated on the substratum of embodiment 1, be placed in 26 ℃ of biochemical cultivation cases and cultivate, growth cycle 25-30 days.

Take out phellinus igniarius mycelium, as for drying in the convection oven of 30 ℃.

The mensuration of flavones content in embodiment 6 mycelium alcohol extracts

80% ethanol that the mycelium of oven dry is added 10 times of volumes soaks 24h, filters; Collect filtrate, residue spends the night with 80% alcohol immersion of 10 times of volumes again, repeats aforesaid operations, merges twice filtrate, and is concentrated, obtains the phellinus igniarius mycelium alcohol extract.

Use NaNO ₂-Al (NO ₃) ₃The colorimetric method for determining flavones content, in the mycelium alcohol extract after oven dry, the weight percent content of flavones is 40%.

The mensuration of embodiment 7 phellinus igniarius mycelium anti-oxidant activities

The source of phellinus igniarius mycelium alcohol extract is with in embodiment 6; Measure the anti-oxidant activity of phellinus igniarius mycelium alcohol extract, mainly remove hydrogen peroxide, superoxide anion and DPPH experiment.

Concrete grammar is as follows

One, remove H ₂O ₂The free radical experiment

Alcohol extract is mixed with four concentration with 70% ethanol, is respectively 1 μ g/mL, 10 μ g/mL, 20 μ g/mL, 50 μ g/mL.To H ₂O ₂Removing experiment use luminol,3-aminophthalic acid cyclic hydrazide-H ₂O ₂The luminous reaction system, with 80% ethanol as blank.Get respectively 5 μ L and add 96 orifice plates, pump into 150 μ L luminol,3-aminophthalic acid cyclic hydrazides-CBS solution (pH9.5) by pump 1, pump 2 pumps into 10 μ L6%H ₂O ₂, start luminous reaction at 20 ℃, luminous value of every 0.6s record, continuous recording 30s.Light-emitting procedure is as showing 3-1:

Table 3-1H ₂O ₂The light-emitting procedure of free radical

H ₂O ₂The free radical scavenging activity calculation formula is as follows

Result shows, the phellinus igniarius mycelium alcohol extract of high density is to H ₂O ₂The clearance rate of free radical reaches 80%.

Two, remove the superoxide anion experiment

Alcohol extract is mixed with different concentration with 80% dissolve with ethanol, i.e. 500 μ g/mL, 800 μ g/mL, 1000 μ g/mL, 2000 μ g/mL do blank with 80% ethanol.Adopt the chemoluminescence method of luminol,3-aminophthalic acid cyclic hydrazide-pyrogallol-CBS system.Each concentration sample is got respectively 10 μ L and is added 96 orifice plates.Pump into 6.25 * 10 by pump 1 ^-4Mol/L pyrogallol solution, pump 2 pump into 150 μ L luminol,3-aminophthalic acid cyclic hydrazides-CBS solution (pH10.2).Start luminous reaction at 20 ℃, every 0.6S collects an optical signal, collects continuously 30S.

Light-emitting procedure is as showing 3-2:

Table 3-2 removes the light-emitting procedure of superoxide anion

The clearance rate formula of superoxide anion is with removing H ₂O ₂The free radical scavenging activity formula

Result shows, when lower concentration 0.5mg/mL, has just showed the ability of higher removing superoxide anion, and concentration is when 2mg/mL, and clearance rate is near 100%.

Three, remove the experiment of DPPH free radical

Get 6 * 10 ^-4The storing solution 100 μ L of mol/L DPPH add 900 μ L80% ethanol, are diluted to 6 * 10 ^-5Mol/L DPPH working fluid.With 80% ethanol, mycelium alcohol extract sample ligand is made different concns (1 μ g/mL, 20 μ g/mL, 100 μ g/mL, 200 μ g/mL).The sample of getting respectively 500 μ L different concns adds in 1mL DPPH working fluid, and mixing is measured light absorption value in the 517nm place after standing 20min, with 80% ethanol as blank.The DPPH free radical scavenging activity is calculated as follows:

Result shows, the alcohol extract of high density reaches 40% to the clearance rate of DPPH free radical.

Claims (3)

1. the substratum of a phellinus igniarius mycelium is characterized in that this substratum comprises following component:

Rice and nutrient solution, wherein the volume (ml) of the weight of rice (g) and nutrient solution is than being x:(x+(0-30), x 〉=10 wherein;

Wherein nutrient solution contains: 2.5% glucose, 0.3%KH ₂PO ₃And 0.15%MgSO ₄7H ₂O。

2. method for preparing the described substratum of claim 1 is characterized in that the method is:

The configuration nutrient solution is placed in nutrient solution with rice, autoclaving.

3. method of utilizing the described culture medium culturing phellinus igniarius mycelium of claim 1, the method is:

The female kind of Phellinus is inoculated on substratum, is placed in 26 ℃ of biochemical cultivation cases and cultivates, growth cycle 25-30 days;

With cultured mycelium together with substratum as for drying in 30 ℃ of convection oven.