CN103555786A - Method for producing polysaccharides through liquid state fermentation of rice bran and wheat bran complete material by using ganoderma lucidum mutant strain - Google Patents
Method for producing polysaccharides through liquid state fermentation of rice bran and wheat bran complete material by using ganoderma lucidum mutant strain Download PDFInfo
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Abstract
The invention provides a ganoderma lucidum polysaccharide production method, and relates to the technical field of food microorganism applications. The method comprises: weighing raw materials, wherein rice bran is 1-5 g/100 mL, wheat bran is 1-4.5 g/100 mL, potassium dihydrogen phosphate is 0.1-0.5 g/100 mL, and magnesium sulfate is 0.1-0.5 g/100 mL; adding required water, filling the materials to obtain the nature pH value, sterilizing at a temperature of 121 DEG C, and cooking for 30 min; cooling, and inoculating the obtained ganoderma lucidum CCTCC M2013287 liquid seed, wherein the inoculation amount is 8-10%, the fermentation temperature is 25-30 DEG C, and the fermentation time is 4-7 d; and carrying out deproteinization, alcohol precipitation, centrifugation separation and vacuum drying on the fermentation product centrifugation separation liquid or the centrifugation separation liquid obtained through wall-breaking hot water extraction of mycelium so as to respectively obtain extracellular polysaccharides and mycelium polysaccharides. With the method, the purpose of high efficiency and high value conversion of rice bran and wheat bran into ganoderma lucidum polysaccharides through liquid state fermentation is well achieved.
Description
Technical field
The present invention relates to food microorganisms applied technical field, relate in particular to a kind of method of utilizing high yield ganoderan in the new bacterial strain liquid state fermentation of glossy ganoderma mutagenesis rice bran wheat bran complete feed substratum.
Background technology
Since the people such as Chihara (reference: Chihara G, MaedaY, Hamuro J, et al.Inhibition of mouse sarcoma180by polysaccharides from Lentinus edodes (Berk.) sing[J] .Nature, 1969, 222 (5194): 687-688.) in 1969, on < < Nature > >, publish thesis, since announcement lentinan has anti-tumor activity, have been found that the medicinal fungi with anti-tumor activity reaches kind more than 200, wherein a part or edible fungus, as Grifola frondosa, Hericium erinaceus (Bull. Ex Fr.) Pers., glossy ganoderma etc.In the mycelium of these edible mushroomss, sporophore, sclerotium or spore, can produce the composition such as the various active such as amino acid, protein, VITAMIN, polysaccharide, glycoside, flavonoid and microbiotic or Rich in Trace Element selenium, zinc etc., have improve body immunity, antitumor, strengthen liver function, the multiple efficacies such as anti-oxidant.The existing edible mushroom health-care food in domestic market is if ganoderma lucidum capsule, Grifola frondosa capsule and capsule of Chinese caterpillar fungus etc. are in fast sale.New Zealand, Japan, the U.S. etc. all produce and have similar product in the world.Zelanian " GanoPoly " series product are formed by the polysaccharide composite chitosan of the extractions such as glossy ganoderma, rainbow conk, are mainly used in improving the aspects such as immunizing power, auxiliary antineoplaston.With regard to ganoderan, research for it shows: ganoderan has the curative effects such as good raising immunizing power, auxiliary antineoplaston, the test of pesticide effectiveness of extracting ganoderan such as Huang, Seto etc. from glossy ganoderma thalline shows, ganoderan has immunoregulation effect and antitumor action, and (reference is shown in 1. Huang S.Q., Ning Z.X.Extraction of polysaccharide from Ganoderma lucidum and its immune enhancement activity[J] .Int J Biol Macromol, 2010,47 (3): 336～341; 2. Seto S.W., Lam T.Y, Tam H.L., et al.Novel hypoglycemic effects of Ganoderma lueidum water-extract in obese/diabetic (db/+db) mice[J] .Phytomedicine, 2009, (16): 426～436; 3. Hikino H, Konno C, Mirin Y, et al.Isolation and hypoglycemic activity of ganoderans A and B, glycans of Ganoderma lucidum fruit bodies[J] .Planta Med, 1985, (12): 339～340)
Glossy ganoderma (Ganoderma luccidum) another name sesame grass, auspicious grass, it is Mycophyta, Basidiomycotina, Hymenomycetes, Aphyllophorales, glossy ganoderma Cordycepps, Ganoderma fungi, is mainly distributed in the torrid zone and the subtropical zone in Asia, Australia, Africa and America, and minority is also distributed in Temperate Region in China, only there are 4 kinds of Ganoderma in the Europe that is located in temperate zone, the Northern Hemisphere, and about 5 kinds of North America.China extends across the torrid zone to cool temperature zone, and Ganodermataceae kind is many and one of country that distribution Guang，Shi world Ganoderma fungal species diversity is the abundantest.Traditionally, the obtain manner of glossy ganoderma is mainly artificial culture or field acquisition, with sporophore or spore powder, is used as medicine.But the Ganoderma Lucidum cycle is long, production efficiency is low, labour intensity is large, is subject to again the restrictions such as season, environment, subjects to disease and pest, and quality and output are unstable, and field acquisition glossy ganoderma limited amount is difficult to meet large production requirement.Edible fungus liquid submerged fermentation technology, has obvious advantage than traditional sporophore cultivation.For High-efficient Production ganoderan, researchist attaches great importance to promoting the research of the liquid cultivating method of suitability for industrialized production, has obtained many achievements in research.Hoe Xiaoyan, Zeng Xi and other liquid culture of Ganoderma conditions were studied, the main contents of such research involves the production strain, medium composition and fermentation parameters, will reduce production costs and increase yields of polysaccharide as a research priority.Wherein most process using are if starch, potato, glucose etc. are as medium component, even if use processing of farm products byproduct as rice bran or wheat bran, also be using it as submember, be still that to take grain class raw material, glucose or other raw materials be main Carbon and nitrogen sources.
China is large agricultural country, and agricultural byproducts source is abundant.Rice bran, wheat bran are as the by product of paddy and wheat processing, and not only its nutritive ingredient is abundant, and cheap.The material such as rich in starch, Mierocrystalline cellulose in rice bran, and wheat bran is rich in the materials such as albumen, Mierocrystalline cellulose.Theoretically, rice bran and wheat bran are compound has possessed the glossy ganoderma needed Carbon and nitrogen sources material of growing.Under the effect of glossy ganoderma self cellulase and other enzymes, glossy ganoderma can change into rice bran and wheat bran the nutritive substance of self and grow, and produces ganoderan.Therefore, use the cheap agricultural byproducts such as rice bran, wheat bran, the required nutritive substances of Ganoderma lucidum submerged fermentation such as complete place of glucose, produce the ganodermal drop with auxiliary oncotherapy, high-valued application to low side raw material is significant, has high economic worth.
According to inventor herein Liu Wei people seminar years of researches experience, some edible mushroom strainses, not add on the rice bran of glucose or other grain class raw materials or wheat bran liquid culture medium upgrowth situation very desirable, need to be done special processing if induction mutation of bacterium is to obtain good growth.For example, Liu Weimin instructs several Master degree candidates to carry out deep research to edible mushrooms Grifola frondosa liquid state fermentation rice bran or wheat bran, having obtained some achievements, formed as Publication about Document: (1) Yang Suohua. Grifola frondosa ferment rice bran is prepared polysaccharide [D]. master thesis. Zhenjiang: the .2006 of Jiangsu University; (2) Gu Huimin. Grifola frondosa is liquid research [D] of cultivating product polysaccharide and enrichment organoselenium in rice bran substratum. master thesis. Zhenjiang: the .2009 of Jiangsu University; (3) to open and build. Physical mutagenesis Grifola frondosa liquid fermenting rice bran wheat bran produces the research [D] of polysaccharide. master thesis. Zhenjiang: Jiangsu University, 2010; (4) Guo Chunmei. the induction mutation of bacterium of Grifola frondosa, liquid fermenting rice bran wheat bran produce polysaccharide and rich selenium research [D]. master thesis. Zhenjiang: Jiangsu University, 2011; (5) Li Yongzhuan. transform rice bran and the Grifola frondosa strain mutagenesis of wheat bran high polysaccharide and the fermentation test [D] of existing bacterium. master thesis. Zhenjiang: Jiangsu University, 2012; (6) Liu Weimin, opens and builds, Guo Chunmei, etc. for the Grifola frondosa strain [P] of rice bran and wheat bran compound material production polysaccharide, 10579078.5,2010; (7) Liu Weimin, opens and builds, Guo Chunmei, etc. use rice bran wheat bran compound material and Grifola frondosa mutagenic strain to produce the method [P] of polysaccharide, 1010579048.4,2010; (7) Liu Weimin, Guo Chunmei, opens and builds, etc. for the bacterial strain [P] of ferment rice bran and wheat bran extracting solution production grifolan, 10150888.3,2011 (applications); (8) Li Yanan. the mutagenesis of Grifola frondosa bacterial classification and fermentation and product property research [D]. master thesis. Zhenjiang: Jiangsu University, 2013 (this paper publication date is in June, 2013, submits to the days identical with the application of this patent); (9) Zhao Li. the research [D] of Grifola frondosa liquid state fermentation enrichment manganese in rice bran wheat bran complex medium. master thesis. Zhenjiang: Jiangsu University, 2012; (10) Liu Lili. high value transforms Ganoderma lucidum submerged fermentation and the induction mutation of bacterium [D] of rice bran wheat bran. master thesis. Zhenjiang: Jiangsu University, 2012; (11) Guo Tianlong. the full rice bran wheat bran of the high-valued conversion of Ganderma lucidum strain mutagenesis and liquid state fermentation research [D]. master thesis. Zhenjiang: Jiangsu University, 2013 (this paper publication date is in June, 2013, submits to the days identical with the application of this patent).
From above-mentioned reference, the inventor herein Liu Wei people study around efficient high-valued this special topic of rare edible fungi polysaccharide that is converted into of rice bran wheat bran always, realize a plurality of innovation and creation imaginations, and proposed to carry out rice bran wheat bran complete feed liquid fermenting with glossy ganoderma, the Research Thinking that efficient high-valued conversion rice bran wheat bran is ganoderan, to original strain mutagenic treatment in addition, unsuccessful at Liu Lili mutagenesis testing (is negative mutagenesis, see the analysis of Liu Lili Master's thesis p46 his-and-hers watches 5.2, the Master's thesis of Liu Lili all adopts rice bran wheat bran to cross leaching juice and makes substratum, rice bran wheat bran complete feed liquid culture medium of the present invention) basis on, summing up experience, instruct Guo Tianlong to carry out systematic research, obtained positive mutagenic strain, and mutagenic strain liquid state fermentation rice bran wheat bran complete feed newly proposed, the method that efficient high-valued conversion rice bran wheat bran is ganoderan, the content of another patent of invention that has formed patent of the present invention and declared simultaneously.Master's thesis publication date of Guo Tianlong is in June, 2013, submits to days identical with the application of this patent, does not affect novelty of the present invention.
One of key issue of the present invention is: obtaining on the basis of the new bacterial strain of a kind of glossy ganoderma, the present invention will provide the method that this new bacterial strain is effectively grown on rice bran wheat bran complete feed liquid culture medium and Efficient Conversion rice bran wheat bran complete feed is ganoderan.The foundation of new bacterial strain and new bacterial strain all need innovation to the mode of newly utilizing of rice bran wheat bran, be suitable for that the rice bran of new ganoderma strain capable and wheat bran form the proportioning of liquid nutrient medium and the processing condition of liquid state fermentation all need recent studies on to draw, have not been reported, therefore the present invention has uniqueness, innovation and practicality, possesses the essential characteristic of patent of invention.
Special instruction: rice bran wheat bran complete feed liquid culture medium of the present invention refers in particular to rice bran and wheat bran unique carbon source, nitrogenous source in substratum, do not add other carbon sources, nitrogenous source, and rice bran, wheat bran add the water shape that is in a liquid state, and do not filter, and complete feed is used.
Another patent of invention of simultaneously submitting to the present invention has provided by the walk abreast method of mutagenesis of ultraviolet microwave, take high yield ganoderan as index, directed Efficient Conversion rice bran wheat bran complete feed liquid culture medium is basis, and seed selection obtains the new bacterial strain glossy ganoderma of the glossy ganoderma CCTCC M2013287 (Ganoderma lucidum CCTCC M 2013287) of high yield.The resulting new glossy ganoderma CCTCC M2013287 of another patent of invention that utilization of the present invention and the present invention submit to simultaneously carries out liquid fermenting in rice bran wheat bran complete feed liquid culture medium, produces ganoderan, obtains thus a kind of new ganoderan production method.Mutagenic strain glossy ganoderma CCTCC M2013287 is new bacterial strain, produces sugared characteristic and changes, and output is higher.This new bacterial strain, for rice bran wheat bran complete feed liquid culture medium, is formed to new rice bran wheat bran and utilizes method, realize the target of high yield ganoderan, there are no bibliographical information ganoderan production method of the present invention and level.So, the present invention adopting new mutagenic strain, realize the efficient high-valued trans-utilization of rice bran wheat bran complete feed liquid state fermentation, to the ganoderan manufacture method making new advances (new bacterial strain adds different rice bran wheat bran complete feed liquid culture mediums and forms and zymotechnique), rice bran wheat bran, as the liquid state fermentation gained ganoderan of substratum, (substratum is different, the molecular composition of glossy ganoderma meta-bolites ganoderan can be different) aspect such as output height has uniqueness, novelty and practicality, established the basis of patent of the present invention.
Summary of the invention
The present invention is in order to reduce production costs, efficient high-valued conversion rice bran and wheat bran, high yield has the ganoderan of the effects such as strengthening immunity, auxiliary tumour, by the ultraviolet microwave mutagenesis screening that walks abreast, go out to be applicable to the high yield Ganoderma Varieties By Uv Induced of ferment rice bran wheat bran complete feed liquid culture medium, utilize this novel bacterial to realize the method that efficient high-valued fermentation transforms rice bran wheat bran complete feed liquid culture medium high yield ganoderan.
The technical solution used in the present invention is as follows, on rice bran wheat bran complete feed liquid culture medium, by the method for the ganoderma strain capable fermentative production ganoderan of mutagenesis gained, according to following step, carry out: (1) adds 121 ℃ of sterilizings of water boiling 30min that holds concurrently by rice bran and wheat bran; (2) rice bran and wheat bran complete feed are directly used in to glossy ganoderma CCTCC M2013287 fermention medium as carbon source, nitrogenous source, rice bran usage quantity is 1-5g/100mL, the usage quantity of wheat bran is 1-4.5g/100mL, add potassium primary phosphate 0.1-0.5g/100mL, sal epsom 0.1-0.5g/100mL, pH nature, shaking flask charge amount 40%, fermentor tank charge amount 70-80%, inoculum size 8-10%, culture temperature 25-30 ℃, shaking speed 150-180r/min or fermentation stirring arm rotating speed 50-70r/min, fermentation time 4-7d; (3) by the mycelium containing a small amount of rice bran wheat bran of liquid culture gained after centrifugation, wash with water, after mycelium broken wall with 70 ℃ of hot water lixiviate 3h, mistake leaching clear liquid; (4) fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.
The ganoderma strain capable glossy ganoderma JSU6161314 using in step of the present invention (2) has been deposited in Chinese Typical Representative culture collection center (CCTCC) on June 25th, 2013, bacterial strain deposit number is for being numbered CCTCC M 2013287, name is called glossy ganoderma CCTCC M2013287 (latin name: Ganoderma lucidum CCTCC M2013287), the application for a patent for invention of this bacterial strain is identical with the patent application of the present invention submission date.
Charge amount in step of the present invention (2) during shaking flask is 40% of shaking flask volume, and inoculum size is stocking volume 10%, rotating speed 150-180r/min, and incubation time is 4-7d.
When in step of the present invention (2), upper tank ferments, sample-loading amount is 80% of fermentor tank volume, and during upper tank, ventilation is tinning liquid volume/min, culture temperature 25-30 ℃, stir speed (S.S.) 50-70r/min, upper tank fermentation 4-7d.
Beneficial effect of the present invention
Contriver is set out by existing glossy ganoderma CFCC6043, carry out the parallel mutagenesis of ultraviolet and microwave, with growth velocity, mycelia dry weight and exocellular polysaccharide are that index is screened, the final glossy ganoderma CCTCC M2013287 that obtains, in not adding the rice bran wheat bran complete feed liquid culture medium of other Carbon and nitrogen sources, produce glossy ganoderma and produce polysaccharide, to compare output higher with original strain glossy ganoderma CFCC6043, while being fermented by tank on microwave irradiation bacterial strain obtained strains glossy ganoderma CCTCC M2013287, mycelia dry weight and mycelia polysaccharide productive rate output are 6.3g/100mL substratum and 0.203mg/100mL substratum, starting strain glossy ganoderma CFCC6043 mycelia dry weight and mycelia polysaccharide output are 5.12g/100mL substratum and 0.14mg/100mL substratum, the mycelia dry weight of mutagenic strain glossy ganoderma CCTCC M2013287 fermentation and mycelia dry weight and the mycelia polysaccharide output of mycelia polysaccharide productivity ratio starting strain glossy ganoderma CFCC6043 have increased respectively 23.0% and 45%.The fermentation capacity of obtained strains is strong, and ganoderan output increases significantly, has significant uniqueness and creative feature, has promoted the technical progress in this field, has obtained useful technique effect.
Contriver is with described new bacterial strain glossy ganoderma CCTCC M2013287 liquid state fermentation rice bran wheat bran complete feed substratum, and efficient high-valued conversion rice bran wheat bran is ganoderan, and obtains the ganoderan of high yield.The rice bran wheat bran of low value is changed into the ganoderan with plurality of health care functions of high value, to reducing production costs, the reasonable higher value application of low side resource high-efficiency, protect general health, all there is useful social effect.
The liquid fermentative production polysaccharide of glossy ganoderma before the present invention adopts grain class raw material more, and rice bran and wheat bran are used as auxiliary material, and consumption is few, crosses leaching juice use-pattern and makes the service efficiency of rice bran wheat bran low.The present invention is applied rice bran wheat bran complete feed, and as the only Carbon and nitrogen sources of substratum, obtain higher ganoderan output, illustrate and new bacterial strain described in utilization provided by the present invention carry out the liquid state fermentation of rice bran wheat bran complete feed to produce the method for ganoderan advanced, economic benefit is high, obtains useful economical effectiveness.
In sum, the present invention has possessed uniqueness, creativeness and the practicality of patent of invention, has produced the technology that patent of invention should possess, economy and social beneficial effect.
Accompanying drawing explanation
Fig. 1 is the parallel mutagenic and breeding schema of bacterial strain glossy ganoderma CCTCC M2013287 ultraviolet microwave of the present invention.
Embodiment
The invention provides glossy ganoderma CCTCC M2013287 (apply for a patent separately), can with respect to original ganoderma strain capable, there is high growth rates and high polysaccharide yield not adding on the rice bran wheat bran complete feed liquid culture medium of other Carbon and nitrogen sources simultaneously; This ganoderma strain capable glossy ganoderma CCTCC M2013287 has been deposited in the Chinese Typical Representative culture collection center (CCTCC) in the Wuhan University of Wuhan, China on June 25th, 2013, preservation strain is numbered CCTCC M2013287, and name is called glossy ganoderma CCTCC M2013287 (latin name is Ganoderma lucidum CCTCC M2013287).The invention provides the production method that adopts bacterial strain that mutagenesis obtains to produce ganoderan on Ganoderma lucidum CCTCC M2013287 rice bran wheat bran complete feed liquid culture medium.
In embodiment, rice bran and wheat bran are added to 121 ℃ of sterilizings of water boiling 30min that holds concurrently, as substratum for fermentation.Described mycelia is the mycelia that mycelia mixes a small amount of raw material.
Embodiment mono-
Ganoderma strain capable adopts mutagenic strain glossy ganoderma CCTCC M2013287.Fermentor tank sample-loading amount is 80% of fermentor tank volume, culture temperature is 28 ℃, ventilation 1:0.8v/v/mim, stirring velocity 60r/min, tank gauge pressure 0.05MPa, inoculum size 8%, incubation time 5d, fermention medium is rice bran 5g/100mL, wheat bran 3g/100mL, potassium primary phosphate 0.15g/100mL, sal epsom 0.1g/100mL, pH nature.Add water to required volume, 121 ℃ of sterilizings boiling 30min that holds concurrently, as substratum for fermentation, by the mycelium containing a small amount of rice bran wheat bran of liquid culture gained after centrifugation, wash with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 6.3g/100mL, and exocellular polysaccharide is 436.0mg/100mL nutrient solution, and intracellular polyse is 203.0mg/100mL nutrient solution.
Embodiment bis-
The glossy ganoderma CCTCC M2013287 bacterial strain that ganoderma strain capable adopts mutagenesis to obtain.Rice bran usage quantity is 2.0g/100mL substratum, and the usage quantity of wheat bran is 2.5g/100mL substratum, adds KH
2pO
40.25g/100mL, MgSO
40.35g/100mL, pH nature, adds water to required volume, 121 ℃ of sterilizings boiling 30min that holds concurrently, as substratum, for fermentation, shaking flask sample-loading amount is 40%, inoculum size is 10%, 25 ℃ of culture temperature, rotating speed 150r/min, incubation time 7d.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 3.51g/100mL, and exocellular polysaccharide is 263.3mg/100mL nutrient solution, and intracellular polyse is 92.7mg/100mL nutrient solution.
Embodiment tri-
The glossy ganoderma CCTCC M2013287 bacterial strain that ganoderma strain capable adopts mutagenesis to obtain.Rice bran usage quantity is 2.5g/100mL substratum, and the usage quantity of wheat bran is 3.5g/100mL substratum, adds KH
2pO
40.5g/100mL, MgSO
40.15g/100mL, pH nature, adds water to required volume, 121 ℃ of sterilizings boiling 30min that holds concurrently, as substratum, for fermentation, shaking flask sample-loading amount is 40%, inoculum size is 10%, 28 ℃ of culture temperature, rotating speed 150r/min, incubation time 4d.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 4.92g/100mL, and exocellular polysaccharide is 357.6mg/100mL nutrient solution, and intracellular polyse is 138.3mg/100mL nutrient solution.
Embodiment tetra-
The glossy ganoderma CCTCC M2013287 bacterial strain that ganoderma strain capable adopts mutagenesis to obtain.Rice bran usage quantity is 3.0g/100mL substratum, and the usage quantity of wheat bran is 4.5g/100mL substratum, adds KH
2pO
40.20g/100mL, MgSO
40.5g/100mL, pH nature, adds water to required volume, 121 ℃ of sterilizings boiling 30min that holds concurrently, as substratum, for fermentation, shaking flask sample-loading amount is 40%, inoculum size is 10%, 26 ℃ of culture temperature, rotating speed 150r/min, incubation time 5d.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 5.91g/100mL, and exocellular polysaccharide is 428.7mg/100mL nutrient solution, and intracellular polyse is 173.2mg/100mL nutrient solution.
Embodiment five
The glossy ganoderma CCTCC M2013287 bacterial strain that ganoderma strain capable adopts mutagenesis to obtain.Rice bran usage quantity is 4.5g/100mL substratum, and the usage quantity of wheat bran is 2.0g/100mL substratum, adds KH
2pO
40.40g/100mL, MgSO
40.45g/100mL, pH nature, adds water to required volume, 121 ℃ of sterilizings boiling 30min that holds concurrently, as substratum, for fermentation, shaking flask sample-loading amount is 40%, inoculum size is 10%, 280 ℃ of culture temperature, rotating speed 150r/min, incubation time 7d.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 4.92g/100mL, and exocellular polysaccharide is 324.3mg/100mL nutrient solution, and intracellular polyse is 149.7mg/100mL nutrient solution.
Embodiment six
The glossy ganoderma CCTCC M2013287 bacterial strain that ganoderma strain capable adopts mutagenesis to obtain.Rice bran usage quantity is 5.0g/100mL substratum, and the usage quantity of wheat bran is 3.0g/100mL substratum, adds KH
2pO
40.10g/100mL, MgSO
40.25g/100mL, pH nature, adds water to required volume, 121 ℃ of sterilizings boiling 30min that holds concurrently, as substratum, for fermentation, shaking flask sample-loading amount is 40%, inoculum size is 10%, 30 ℃ of culture temperature, rotating speed 180r/min, incubation time 6d.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 6.0g/100mL, and exocellular polysaccharide is 416.6mg/100mL nutrient solution, and intracellular polyse is 179.5mg/100mL nutrient solution.
Embodiment seven
Ganoderma strain capable adopts mutagenic strain glossy ganoderma CCTCC M2013287.Fermentor tank sample-loading amount is 80% of fermentor tank volume, culture temperature is 28 ℃, ventilation 1:0.8v/v/mim, stirring velocity 60r/min, tank gauge pressure 0.05MPa, inoculum size 8%, incubation time 5d, fermention medium is rice bran 20g/L, wheat bran 25g/L, potassium primary phosphate 2.5g/L, sal epsom 3.5g/L, pH nature.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 3.34g/100mL, and exocellular polysaccharide is 241.3mg/100mL nutrient solution, and intracellular polyse is 123.8mg/100mL nutrient solution.
Embodiment eight
Ganoderma strain capable adopts mutagenic strain glossy ganoderma CCTCC M2013287.Fermentor tank sample-loading amount is 70% of fermentor tank volume, culture temperature is 28 ℃, ventilation 1:0.8v/v/mim, stirring velocity 70r/min, tank gauge pressure 0.05MPa, inoculum size 10%, incubation time 6d, fermention medium is rice bran 30g/L, wheat bran 45g/L, potassium primary phosphate 2.0g/L, sal epsom 5.0g/L, pH nature.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 5.7g/100mL, and exocellular polysaccharide is 400.4mg/100mL nutrient solution, and intracellular polyse is 192.9mg/100mL nutrient solution.
Embodiment nine
Ganoderma strain capable adopts mutagenic strain glossy ganoderma CCTCC M2013287.Fermentor tank sample-loading amount is 80% of fermentor tank volume, and culture temperature is 25 ℃, ventilation 1:0.8v/v/mim, tank gauge pressure 0.05MPa, inoculum size 10%, incubation time 5d, fermention medium is rice bran 45g/L, wheat bran 20g/L, potassium primary phosphate 4.0g/L, sal epsom 4.5g/L, pH nature.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 5.1g/100mL, and exocellular polysaccharide is 364.3mg/100mL nutrient solution, and intracellular polyse is 158.7mg/100mL nutrient solution.
Embodiment ten
Ganoderma strain capable adopts mutagenic strain glossy ganoderma CCTCC M2013287.Fermentor tank sample-loading amount is 70% of fermentor tank volume, and culture temperature is 30 ℃, ventilation 1:0.8v/v/mim, tank gauge pressure 0.05MPa, inoculum size 10%, incubation time 7d, fermention medium is rice bran 50g/L, wheat bran 20g/L, potassium primary phosphate 1.5g/L, sal epsom 1.0g/L, pH nature.The mycelium containing a small amount of rice bran wheat bran of liquid culture gained, after centrifugation, is washed with water, after mycelium broken wall, with 70 ℃ of hot water lixiviate 3h, cross leaching clear liquid.Fermented product centrate or the mycelia centrate after hot water lixiviate, after deproteinated, alcohol precipitation, centrifugation, vacuum-drying, obtain respectively exocellular polysaccharide and mycelia polysaccharide, adopt phenol sulfuric acid process to measure the polysaccharide content in mycelia polysaccharide and exocellular polysaccharide.Result is mycelia dry weight 5.3g/100mL, and exocellular polysaccharide is 377.4mg/100mL nutrient solution, and intracellular polyse is 178.5mg/100mL nutrient solution.
Claims (5)
1. a glossy ganoderma mutagenic fungi liquid state fermentation rice bran wheat bran complete feed is produced the method for polysaccharide, it is characterized in that the ganoderma strain capable that utilizes mutagenesis to obtain, with rice bran wheat bran complete feed liquid culture medium fermentative production ganoderan, according to following step, carry out: (1) takes raw material rice bran is 1-5g/100mL, wheat bran is 1-4.5g/100mL, potassium primary phosphate 0.1-0.5g/100mL, sal epsom 0.1-0.5g/100mL (2) adds after required water, charging, pH nature, 121 ℃ of sterilizings boiling 30min that holds concurrently, the glossy ganoderma CCTCC M2013287 liquid seeds obtaining is invented in cooling rear access simultaneously, carry out liquid fermenting, inoculum size 8-10%, leavening temperature 25-30 ℃, fermentation time 4-7d, (3) by the mycelium of liquid culture gained after centrifugation, water cleans, and obtains mycelia thing, (4) after mycelia broken wall with 70 ℃ of hot water lixiviate 3h, centrifugation, obtains mycelia polysaccharide after getting supernatant liquor deproteinated, alcohol precipitation, centrifugation, vacuum-drying, (5) fermented product filtered liquid obtains exocellular polysaccharide after deproteinated, alcohol precipitation, centrifugation, vacuum-drying.
2. the method that a kind of glossy ganoderma mutagenic fungi liquid state fermentation rice bran wheat bran complete feed according to claim 1 is produced polysaccharide, the bacterial strain glossy ganoderma CCTCC M2013287 (Ganoderma lucidum CCTCC M2013287) that the new mutagenesis that it is characterized in that wherein using obtains, this bacterial strain is deposited in Chinese Typical Representative culture collection center, and preservation strain is numbered CCTCC M2013287.
3. a kind of glossy ganoderma mutagenic fungi liquid state fermentation rice bran wheat bran complete feed described in claim 1 or 2 is produced the method for polysaccharide, it is characterized in that it is that complete feed uses that rice bran and wheat bran complete feed are added to water uses as liquid culture medium.
4. the method that a kind of glossy ganoderma mutagenic fungi liquid state fermentation rice bran wheat bran complete feed according to claim 1 is produced polysaccharide, while it is characterized in that shake flask fermentation, inoculum size 8-10%, sample-loading amount is 40%, leavening temperature 25-30 ℃, shaking speed 150-180r/min, fermentation time 4-7d.
5. the method that a kind of glossy ganoderma mutagenic fungi liquid state fermentation rice bran wheat bran complete feed according to claim 1 is produced polysaccharide, while it is characterized in that tank fermentation, inoculum size 8-10%, sample-loading amount is 70-80%, leavening temperature 25-30 ℃, stirring arm rotating speed 50-70r/min, fermentation time 4-7d.
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| CN105586270A (en) * | 2015-12-25 | 2016-05-18 | 江苏大学 | Production method of selenium-rich ganoderma lucidum mycelium raw material |
| CN105586267A (en) * | 2015-12-25 | 2016-05-18 | 江苏大学 | Ganoderma lucidum mutant strain for production of Ganoderma lucidum mycelia |
| CN105754865A (en) * | 2015-12-25 | 2016-07-13 | 江苏大学 | Method for producing ganoderma lucidum mycelium material by induced mutant strain |
| CN115024159A (en) * | 2022-06-23 | 2022-09-09 | 海南热带海洋学院 | Method for screening heat-resistant ganoderma lucidum variety |
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| CN101317526A (en) * | 2007-06-06 | 2008-12-10 | 福建省农业科学院农业生态研究所 | Production and cultivation of glossy ganoderma with tea branch, tea, stem material |
| CN101591688A (en) * | 2009-06-30 | 2009-12-02 | 华中科技大学 | But a kind of method of obtaining conversion of substrate by the bacterium chaff |
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| CN101317526A (en) * | 2007-06-06 | 2008-12-10 | 福建省农业科学院农业生态研究所 | Production and cultivation of glossy ganoderma with tea branch, tea, stem material |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105586270A (en) * | 2015-12-25 | 2016-05-18 | 江苏大学 | Production method of selenium-rich ganoderma lucidum mycelium raw material |
| CN105586267A (en) * | 2015-12-25 | 2016-05-18 | 江苏大学 | Ganoderma lucidum mutant strain for production of Ganoderma lucidum mycelia |
| CN105754865A (en) * | 2015-12-25 | 2016-07-13 | 江苏大学 | Method for producing ganoderma lucidum mycelium material by induced mutant strain |
| CN105586270B (en) * | 2015-12-25 | 2018-10-09 | 江苏大学 | The production method of Se-rich lucid ganoderma mycelia raw material |
| CN105586267B (en) * | 2015-12-25 | 2018-11-06 | 江苏大学 | Produce the ganoderma lucidum mutagenic strain of ganoderma lucidum mycelium |
| CN105754865B (en) * | 2015-12-25 | 2019-04-30 | 江苏大学 | Utilize the method for mutagenic strain production ganoderma lucidum mycelium raw material |
| CN115024159A (en) * | 2022-06-23 | 2022-09-09 | 海南热带海洋学院 | Method for screening heat-resistant ganoderma lucidum variety |
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