CN1273010C - Technique for production of Tricholoma matsutake and polysaccharide thereof by large scale liquid submerged fermentation - Google Patents
Technique for production of Tricholoma matsutake and polysaccharide thereof by large scale liquid submerged fermentation Download PDFInfo
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- CN1273010C CN1273010C CN 200310112685 CN200310112685A CN1273010C CN 1273010 C CN1273010 C CN 1273010C CN 200310112685 CN200310112685 CN 200310112685 CN 200310112685 A CN200310112685 A CN 200310112685A CN 1273010 C CN1273010 C CN 1273010C
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Abstract
The present invention relates to a technique for producing tricholoma mushroom mycelium and polysaccharidess thereof by large scale liquid submerged fermentation. The labor cultivation of fruiting bodies of unusual edible fungus tricholoma mushroom is not achieved; the report on a large scale liquid submerged fermentation cultivation technology does not exist at present. The present invention has a goal for producing the tricholoma mushroom mycelium by the large scale liquid submerged fermentation; the tricholoma mushroom fungus of a fungus academy of in Jinxiang, Shandong is used as original strain; bevel inclined plane fungus seeds are adopted for liquid bottle shaking cultivation, liquid bottle shaking enlarging cultivation, first stage seed cultivation and fermentation cultivation; finally, the volume of a fermentation tank can reach 1000L to 5000L. The fermentation technology is utilized for fermenting 160h in the fermentation tank with 1500L; the mycelium bodies in the fermentation liquid can reach 2.5% by dry weight. If the polysaccharide needs to be obtained, the temperature of final fermentation liquid can be heated to 60 DEG C to 100 DEG C; a stirring speed is 150 rpm/min to 250 rpm/min; the temperature and the speed are maintained in two to four hours; the fermentation liquor is conveyed to a storage tank by a pump and is pressed and filtered by a plate frame for obtaining the tricholoma mushroom polysaccharides.
Description
Technical field
The present invention relates to a kind of large-scale deep liquid fermentation and produce Trichotoma matsutake and polysaccharide process thereof.
Technical background
Trichotoma matsutake (Tricholoma matsutaka) is called matsutake, pine mushroom, closes gill fungus, the platform bacterium, belongs to Basidiomycotina, Hymenomycetes, Holobasidiomycetidae, Agaricales, Tricholomataceae, Tricholoma, is famous and precious wild edible fungus.One of matsutake starts from China, and is just on the books in the Classic Classified Materia Medica for Emergencies of Song dynasty.In Japan's multi-section ancient books also relevant for the record of its local flavor." national Chinese herbal medicine compilation " record this product can be used for phlegm many, breathe hard, disease such as lumbocrural pain." Yunnan natural resources of Chinese medicinal materials register " records reducing fever and causing diuresis, beneficial stomach, pain relieving, effect such as anticancer.
The bright product culinary art of Trichotoma matsutake, delicious unusual, left a lingering fragrance in one's mouth, liked by the place of production people of various countries.In Japan, the custom of edible matsutake spreads so far always, becomes the first-class delicacies in Japan's food in autumn, owing to cost an arm and a leg, is described as " diamond in the mountain ", have saying of " Herring roes in the sea, the matsutake of land ".In Japan, first-class wild Trichotoma matsutake is by a sale.Trichotoma matsutake is delicious flavour not only, and has very high food therapy value.Trichotoma matsutake removes several amino acids, the Cobastab that contains needed by human body
1, B
2, outside the material such as C, PP and calcium, phosphorus, iron, still contain aromatic substances such as octanol alkene, methyl cinnamate, octanone and blazei polysaccharide, mannitol, matsutakealcohol, isomatsutakeol isoreactivity composition.
Modern scientific research shows, Trichotoma matsutake has treatment diabetes, specific function such as anticancer.Japan scientist's research points out that the hot water extract of Trichotoma matsutake, comes out at the top in basidiomycetes up to 91.8% to the tumour inhibiting rate of murine sarcoma S-180.The main pharmaceutical component of Trichotoma matsutake is a polysaccharide.Its polysaccharide energy enhancing body immunocompetence of report is closely arranged, promote that cAMP (cyclic adenosine monophosphate (cAMP)) improves in tumor-bearing mice blood, spleen and the tumor tissue.
In recent ten years, the price of Trichotoma matsutake was more and more higher, and its international market price rises to the 1275 dollars/kg of nineteen ninety-five from 200 dollars/kg in 1975.Because predatoriness is in recent years excavated wild Trichotoma matsutake resource is reduced rapidly.And the not success as yet of the artificial cultivation of Trichotoma matsutake.In order to satisfy people's needs, the research of Trichotoma matsutake is become the problem that people gaze at always.At present, the artificial cultivation of Trichotoma matsutake fruit body not success as yet.Have not yet to see large-scale deep liquid fermentation and produce the report of Trichotoma matsutake mycelium and polysaccharide thereof.
Summary of the invention
The objective of the invention is to produce Trichotoma matsutake mycelium and polysaccharide is target, invents a kind of Trichotoma matsutake mycelium of Cheap highly effective and the large-scale deep liquid fermentation process of polysaccharide thereof, for from now on commercial application lays the first stone.
Realize technical scheme of the present invention: with the Trichotoma matsutake fungi is starting strain, adopts slant strains to carry out liquid shaking bottle cultivation, liquid shaking bottle enlarged culture, first order seed cultivation and fermented and cultured.This bacterial strain is available from Jinxiang, Shandong fungal studies institute.
Realize that concrete steps of the present invention are as follows:
(1) takes out slant strains and insert in the fresh slant medium of preparing, cultivation temperature 25-33 ℃, cultivated 150-250 hour.
(2) slant strains in the step 1 is transferred the shaking in the bottle of shake-flask culture base is housed, 25-33 ℃ of cultivation, rotating speed 120-180rpm/min cultivates the enlarged culture of shaking bottle after 60-100 hour.
(3) bottle bacterial classification that shakes in the step 2 is transferred and carried out enlarged culture again into the bottle that shakes that shakes bottle enlarged culture base is housed, 25-33 ℃ of cultivation, rotating speed 120-180rpm/min cultivates after 20-50 hour the access first class seed pot and cultivates.
(4) bottle bacterial classification that shakes in the step 3 is transferred one and is equipped with in the first class seed pot of seed culture medium, inoculum concentration is 4-10%, keep warm 25-33 ℃ in jar, tank pressure 0.5-0.9kPa, stir speed (S.S.) 80-150rpm/min, throughput 1: 0.3-0.5v/v.m fermented 30-55 hour, and ferment tank is gone in switching then.
(5) bacterial classification in the first class seed pot is transferred one be equipped with in the fermentation tank of fermentation medium, inoculum concentration is 4-10%, keeps jar warm 25-33 ℃, tank pressure 0.5-0.9kPa, stir speed (S.S.) 80-150rpm/min, throughput 1: 0.3-0.5v/v.m fermented 120-180 hour.When being reduced to the 1.0-2.0% left and right sides, content of reducing sugar puts jar.
(6) obtain its polysaccharide as need, the end of a period liquid temp that then will ferment is heated to 60-100 ℃, and mixing speed is 150-250rpm/min, keeps 1-3 hour, with pump zymotic fluid is transported in the basin, obtains the Trichotoma matsutake polysaccharide through plate compression.
The prescription of slant medium is (g/100ml) among the present invention: potato 10-20, glucose 1-3, agar 1.5-2.5, add water be settled to volume required, pH value nature.
The shake-flask culture base is (g/100ml) with shaking bottle prescription of enlarged culture base among the present invention: glucose 2-4, yeast extract 0.05-0.5, analysis for soybean powder 0.5-2, wheat bran juice 1.0-2.5, KH
2PO
40.1-0.5, MgSO
40.01-0.2, adding water and be settled to volume requiredly, the pH value is 4.5-5.0.
The culture medium prescription of first order seed cultivation and fermented and cultured is (g/100ml) among the present invention: glucose 2-4, yeast extract 0.05-0.5, corn flour 0.5-2, analysis for soybean powder 0.5-2, wheat bran juice 1-3, KH
2PO
40.1-0.5, MgSO
40.01-0.2, adding water and be settled to volume requiredly, the pH value is 4.5-5.0.
The shake-flask culture base is 1: 5 with shaking bottle volumetric ratio in the step 2 of the present invention and 3.
The volume of first class seed pot is 100-300L in the step 4 of the present invention, and the first order seed medium that is equipped with in this first class seed pot should be 50-180L mutually; The volume of fermentation tank is 1000-5000L in the step 5, and the fermentation medium that is equipped with in this fermentation tank should be 500-3500L mutually.
Jar standard of putting of the present invention is that zymotic fluid begins the thickness that becomes, and content of reducing sugar is reduced to about 1.0-2.0%, and microscopy finds that mycelium senesces.
Obtain its polysaccharide as need, then after fermentation of the present invention stopped, the end of a period liquid temp that will ferment was heated to 60-100 ℃, mixing speed is 150-250rpm/min, kept 2-4 hour, and with pump zymotic fluid was transported in the basin, through plate compression, concentrating filter liquor, make the 1/4-1/10 of last volume in stoste, add an amount of ethanol and make concentration of alcohol reach 75%, alcohol was analysed 20-30 hour, get the precipitate with deionized water dissolving, atomized drying obtains the Trichotoma matsutake polysaccharide.
Beneficial effect of the present invention: the present invention is applicable to large-scale deep liquid fermentation production Trichotoma matsutake mycelium and polysaccharide thereof, solve large-scale production and cultivated a difficult problem of amplifying, significant for the development of from now on suitability for industrialized production and relevant application industry.The tunning color that the present invention obtains after fermentation ends is brown, and smell delicate fragrance is pleasant, and the fragrant of pine needle is arranged, and dry mycelium is brown.Mycelium counts 2.5% with dry weight in the zymotic fluid.
Embodiment
The present invention is described in further detail below in conjunction with specific embodiment.
Embodiment 1:
Inoculation Trichotoma matsutake mycelium in slant medium, 28 ℃ of cultivation temperature, incubation time 180 hours.Fresh slant strains access is equipped with in the 500ml triangular flask of 100ml medium, and 28 ℃ of shaking tables were cultivated shaking speed 150rpm/min 80 hours.Then gained is shaken transfer 500ml triangle that the 100ml medium is housed of bottle bacterial classification and shake and carry out enlarged culture in the bottle, 28 ℃ of shaking tables were cultivated shaking speed 150rpm/min 28 hours.During the bacterial classification 2200ml that triangular flask is enlarged transfers the 100L first class seed pot that the 50L seed culture medium is housed then, keep 28 ℃ of jar temperature, tank pressure 0.7kPa, stir speed (S.S.) 100rpm/min, throughput 1: 0.5v/v.m, fermentation time 45 hours.The first class seed pot bacterial classification is transferred in the 1500L fermentation tank that the 800L fermentation medium is housed, keep 28 ℃ of jar temperature, tank pressure 0.7kPa, stir speed (S.S.) 100rpm/min, throughput 1: 0.5v/v.m, fermentation time 160 hours, mycelium counts 2.5% with dry weight in the zymotic fluid, and polyoses content is 2.5%.
To form be (g/100ml) to slant medium in the present embodiment 1: potato 15, and glucose 2, agar 2 adds water and is settled to volume requiredly, and the pH value is a nature.
The shake-flask culture base is (g/100ml) with shaking a bottle enlarged culture base composition in the present embodiment 1: glucose 3, yeast extract 0.2, analysis for soybean powder 1, wheat bran juice 2, KH
2PO
40.2, MgSO
40.1, adding water and be settled to volume requiredly, the pH value is 4.5-5.0.
The medium of 100L first class seed pot composition is (g/100ml) in the present embodiment 1: glucose 3, yeast extract 0.2, corn flour 1.5, analysis for soybean powder 1, wheat bran juice 2, KH
2PO
40.2, MgSO
40.1, adding water and be settled to 50L, the pH value is 4.5-5.0.
The culture medium prescription of 1500L fermentation tank is (g/100ml) in the present embodiment 1: glucose 3, yeast extract 0.2, corn flour 1.5, analysis for soybean powder 1.0, wheat bran juice 2, KH
2PO
40.2, MgSO
40.1, adding water and be settled to 800L, the pH value is 4.5.
The fermentation ends standard is in the present embodiment: zymotic fluid begins the thickness that becomes, and content of reducing sugar is reduced to 1.5%, and microscopy finds that mycelium senesces.
The method of obtaining polysaccharide is: after fermentation of the present invention stops, temperature is heated to 90 ℃, and mixing speed is 220rpm/min, keeps 3 hours, with pump zymotic fluid is transported in the basin, through plate compression, concentrating filter liquor makes last volume in 1/5 of stoste, adding an amount of ethanol makes concentration of alcohol reach 75%, alcohol was analysed 24 hours, got the precipitate with deionized water dissolving, and atomized drying obtains the Trichotoma matsutake polysaccharide.
Claims (4)
1, a kind of Trichotoma matsutake liquid deep layer fermenting production method, it is characterized in that: with the Trichotoma matsutake fungi is starting strain, adopts slant strains to carry out liquid shaking bottle cultivation, liquid shaking bottle enlarged culture, first order seed cultivation and fermented and cultured and gets the Trichotoma matsutake mycelium fermentation broth; Or after the fermentation ends end of a period zymotic fluid is continued to be heated to uniform temperature, and stirring and keep certain hour, zymotic fluid obtains the Trichotoma matsutake polysaccharide through plate compression;
Described slant strains medium is formed in g/100ml: potato 10-20, and glucose 1-3, agar 1.5-2.5 adds water and is settled to volume requiredly, and the pH value is a nature, and 25-33 ℃ of slant strains cultivation temperature cultivated 150-250 hour;
The medium of described shake-flask culture is formed in g/100ml: glucose 2-4, yeast extract 0.05-0.5, analysis for soybean powder 0.5-2, wheat bran juice 1.0-2.5, KH
2PO
40.1-0.5, MgSO
40.01-0.2, adding water and be settled to volume requiredly, the pH value is 4.5-5.0, and condition of culture is: cultivation temperature 25-33 ℃, rotating speed 120-180rpm/min cultivated 60-100 hour, and switching is gone into to shake bottle and is carried out enlarged culture then;
The described medium that shakes bottle enlarged culture is formed in g/100ml: glucose 2-4, yeast extract 0.05-0.5, analysis for soybean powder 0.5-2, wheat bran juice 1.0-2.5, KH
2PO
40.1-0.5, MgSO
40.01-0.2, adding water and be settled to volume requiredly, the pH value is 4.5-5.0, and condition of culture is: cultivation temperature 25-33 ℃, rotating speed 120-180rpm/min cultivated 20-50 hour, and switching is gone into first order seed and is cultivated then;
The medium that described first order seed is cultivated is formed in g/100ml: glucose 2-4, yeast extract 0.05-0.5, corn flour 0.5-2, analysis for soybean powder 0.5-2, wheat bran juice 1-3, KH
2PO
40.1-0.5, MgSO
40.01-0.2, adding water is settled to volume required, the pH value is 4.5-5.0, inoculum concentration is 4%-10%, and condition of culture is: cultivation temperature 25-33 ℃, and tank pressure 0.5-0.9kPa, stir speed (S.S.) 80-150rpm/min, throughput 1: 0.3-0.5v/v.m fermented 30-55 hour, and ferment tank is gone in switching then;
Described fermented and cultured, fermentation medium are formed in g/100ml: glucose 2-4, yeast extract 0.05-0.5, corn flour 0.5-2, analysis for soybean powder 0.5-2, wheat bran juice 1-3, KH
2PO
40.1-0.5, MgSO
40.01-0.2, adding water and be settled to volume requiredly, the pH value is 4.5-5.0, inoculum concentration is 4%-10%, and condition of culture is: cultivation temperature 25-33 ℃, and tank pressure 0.5-0.9kPa, stir speed (S.S.) 80-150rpm/min, throughput 1: 0.3-0.5v/v.m fermented 120-200 hour.
2, Trichotoma matsutake liquid deep layer fermenting production method according to claim 1 is characterized in that: being starting strain available from Jinxiang, Shandong fungal studies institute Trichotoma matsutake fungi.
3, Trichotoma matsutake liquid deep layer fermenting production method according to claim 1 is characterized in that: described fermented and cultured, and can select the fermentation tank of 1000-5000L to carry out fermented and cultured.
4, Trichotoma matsutake liquid deep layer fermenting production method according to claim 1, it is characterized in that: need obtain its polysaccharide, then end of a period zymotic fluid temperature is heated to 60-100 ℃, mixing speed is 150-250rpm/min, kept 2-4 hour, with pump zymotic fluid is transported in the basin, obtains the Trichotoma matsutake polysaccharide through plate compression.
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| CN 200310112685 CN1273010C (en) | 2003-12-16 | 2003-12-16 | Technique for production of Tricholoma matsutake and polysaccharide thereof by large scale liquid submerged fermentation |
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| CN 200310112685 CN1273010C (en) | 2003-12-16 | 2003-12-16 | Technique for production of Tricholoma matsutake and polysaccharide thereof by large scale liquid submerged fermentation |
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| CN1273010C true CN1273010C (en) | 2006-09-06 |
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Families Citing this family (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102191297A (en) * | 2011-04-06 | 2011-09-21 | 黑龙江省轻工科学研究院 | Method for making tricholoma matsutake polysaccharides |
| CN102771772A (en) * | 2012-07-20 | 2012-11-14 | 黄晓青 | Pine mushroom health product prepared by submerged fermentation and preparation method thereof |
| CN103467170A (en) * | 2013-08-12 | 2013-12-25 | 凤台县星展食用菌有限公司 | Tricholoma matsutake cultivation medium and preparation method thereof |
| CN104904498A (en) * | 2015-06-30 | 2015-09-16 | 兴安县铭晖食用菌种植有限公司 | Cultivation method of black fungi |
| CN105557299A (en) * | 2015-12-05 | 2016-05-11 | 刘晓红 | Wild tricholoma matsutake artificial domestication method |
| CN105505786A (en) * | 2015-12-21 | 2016-04-20 | 宋瑞鹏 | Culture method for tricholoma matsutake liquid strain |
| CN109498513B (en) * | 2018-12-28 | 2021-08-24 | 广州艾儿化妆品有限公司 | Natural tricholoma matsutake extract and application thereof in mild shampoo |
| CN109431916B (en) * | 2018-12-28 | 2021-06-01 | 广东汉粹生物药业有限公司 | Mild baby shampoo and preparation method thereof |
| CN110199778A (en) * | 2019-06-04 | 2019-09-06 | 临沂信邦生物科技有限公司 | A kind of production technology using fermentation of corn starch production tricholoma matsutake mycelium |
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