CN102823428B - Method for solid liquid mixed culture of medical hericium erinaceus mycelia - Google Patents
Method for solid liquid mixed culture of medical hericium erinaceus mycelia Download PDFInfo
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Abstract
The invention discloses a method for solid liquid mixed culture of medical hericium erinaceus mycelia. According to the method, special medical white hericium erinaceus strains cultured from wild hericium erinaceus in Zhongtiao mountain primeval forest through optimization for many years are adopted. Compared with the ordinary edible hericium erinaceus, the mycelia produced from the strains have the advantages that polysaccharides and peptide are contained, five kinds of anticancer substances, seven kinds of vitamins, three kinds of active cellulase, 18 kinds of amino acids, human body necessary trace elements and the like are also contained, the contents of cystine, histidine and arginine with great medical values are high, the amino acids are identified for 6 to 7 points, and the protein content is 16.65 percent, and is respectively 3 times and 1.8 times that of other hericium erinaceus.
Description
Technical field
The present invention relates to microorganism and cultivate technical field, the mycelial cultural method of especially medicinal Hericium erinaceus (Bull. Ex Fr.) Pers..
Background technology
Hericium erinaceus (Bull. Ex Fr.) Pers., to gain the name because its sporophore shape exactly likes the head of little monkey, it is a kind of famous and precious edible mushrooms that has edible and pharmaceutical use concurrently, property flat, taste is sweet, energy sharp the five internal organs, aid digestion, nourishing, anticancer, treats neurasthenia, is used for the treatment of disease and the tumour of the Digestive tract such as maldigestion, stomach ulcer, esophagus cancer, cancer of the stomach, duodenal cancer, cardiac cancer.Hericium erinaceus (Bull. Ex Fr.) Pers. has been successfully applied in currently available products at present, and as commercially available HOUTOU JIANWEI JIAONANG, wherein Hericium erinaceus (Bull. Ex Fr.) Pers. is the major ingredient of this product.
The production ways of Hericium erinaceus (Bull. Ex Fr.) Pers. mainly contains three kinds at present: 1, artificial culture, and advantage is simple, cost is low; Shortcoming is that the cultivation cycle is long, approximately 3 months; 2, solid fermentation, cultivates Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium with solid medium, and to obtain secondary metabolite, its advantage is: the simple and wide material sources of substratum, low price; Fermenting process does not generally need strict aseptic technique; Culturing process oxygen supply and temperature can adopt direct force air to ventilate to control; The processing of fermentation residue is simple, directly as feed or fertilizer; Shortcoming is: substratum water-activity is low, the inhomogeneous and difficult stirring of substratum, the growth of thalline, all inhomogeneous to the secretion of the absorption of nutritive substance and meta-bolites, makes the monitoring of fermentation parameter and controls difficulty, thus operate continuously and automatization very difficult; The temperature of the heat that microbial respiratory and metabolism produce is controlled very difficult; Adopt solid-state cultivation mostly based on rule of thumb data and operating experience, easily pollution microbes; 3, liquid submerged fermentation, to obtain optimum operation condition, produce a large amount of mycelium and meta-bolites within a short period of time, advantage is: can carry out industrialization and produce continuously, cultivate raw material sources extensive, by controlled fermentation device, reach optimal culture condition, the cycle, fugitive rate was high; Shortcoming is that mycelial turnout is low, is difficult to form large-scale application.
Summary of the invention
The present invention is directed to the deficiencies in the prior art, propose a kind of method of medicinal Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium solids liq mixed culture, the method can be produced the medicinal Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium of better quality, and method is more perfect.
In order to realize foregoing invention object, the invention provides following technical scheme: the mycelial solid medium cultural method of a kind of medicinal Hericium erinaceus (Bull. Ex Fr.) Pers., comprise the following steps: the percentage ratio of following substances is weight percentage,
(1). preparation solid medium
The water that adds surplus, pH value 6～7.5, mixes, 118～121 ℃ of sterilizings, 25～30 minutes;
(2). the optimization screening of bacterial classification and sporophore are separated:
The sporophore strong, that physically well develop, do not have disease and pest, seed not to be scattered of growing in wild Hericium erinaceus (Bull. Ex Fr.) Pers. in selecting and purchasing Zhongtiao Shan Mountain virgin forest;
At transfer room more than 100 grades in Bechtop, to in the alcohol of hericium erinaceus fruiting body immersion 75%, sterilize 1～2 minute, clean with distilled water flushing after taking-up, with antiseptic gauze, dry again, sporophore is placed on culture dish, with Bell jar or similar apparatus, cover, room temperature remains on 18～20 ℃, sporophore is slowly grown, cross after 2～3 days, seed sheds, fall into the culture dish that step (1) substratum is housed, then with tweezers, dip the seed falling, by method of scoring, be inoculated on the inclined-plane of test tube and cultivate, at 22～25 ℃, cultivating 4～5 days seeds can sprout, when mycelia is covered with inclined-plane substantially, selection look white, substrate mycelium prosperity is sturdy, bacterium bundle is obvious, insert shaking table bottle graft kind standby,
(3). the cultivation of bacterial classification
Before inoculation, inoculation mouthful should first be sterilized by ordinary method, with the rayon balls of 75% alcohol-pickled mistake, wipe examination, rear unlatching ozonizer, make its air outlet all the time facing to inoculation mouthful, in inoculation mouthful, form a local aseptic area around, then with sterile nontoxic rubber pipe box in inoculation the mouth of pipe on, shaking table bottle bacterial classification access through spore separation is equipped with in the tank of step (1) substratum, and inoculum size is 9%～11%, and all processes all carries out under aseptic condition, after inoculation, open stirrer, 200 revs/min of stirring velocitys, 25 ℃ of constant temperature, cultivate 2 days;
(4). deep liquid is cultivated:
4.1 seed tank culture bases are:
The water that adds surplus, 118～121 ℃ of sterilizings 28～32 minutes;
4.2 are seeded to seeding tank by the cultured bacterial classification of step (3), and 25～27 ℃, cultivate 3～4 days, to result be:
1.. microscopy: mycelia is dendroid, without miscellaneous bacteria;
2.. outward appearance: have more white peloid in liquid;
(5). Mycelium culture:
Wheat bran 35～40%, liquorice dregs 15～20%, the Yanhusuo dregs of a decoction 7～10%, Semen Maydis powder 7～10%, cavings 7～10%, corn ear 3～5%, wood chip 3～5%; Adding water mixes, 1% the gypsum that adds again total amount, stir, be divided in polypropylene pouch, 118～121 ℃ of sterilizings 115～120 minutes, in 100 grades of aseptic areas, access the solid spawn of step 4.2, inoculum size is 1/30, front 12 days 26～28 ℃, within latter 28 days, in 23～25 ℃, cultivate, after mycelial growth is full, break 55～65 ℃ of oven dry of piece, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. filament.
Compared with prior art, the present invention adopts the wild Hericium erinaceus (Bull. Ex Fr.) Pers. in Zhongtiao Shan Mountain virgin forest through for many years, to optimize improvement, adopt solid medium in conjunction with the mode of liquid nutrient medium, and the special medicinal white monkey bacterial classification of cultivating, the mycelium of producing with this bacterial classification is compared with general edible Hericium erinaceus (Bull. Ex Fr.) Pers., except containing polysaccharide polypeptide, still also have five kinds different " callose ", the various saccharides combinations such as same glucose, N.F,USP MANNITOL and semi-lactosi, immunocompetence is high, there is certain antitumous effect, contain VitAVitE, vitamins B
1, vitamins B
2, vitamins B
6, vitamins B
12, seven kinds of VITAMIN such as vitamins C, contain cellulase, hemicellulase, amylase, guaiacolase and polyphenoloxidase isoreactivity cellulase, and contain leucine, Isoleucine, α-amino-isovaleric acid, Methionin, Threonine, methionine(Met), tryptophane, phenylalanine, Histidine, arginine, L-Ala, aspartic acid, Gelucystine, L-glutamic acid, proline(Pro), tyrosine, glycine, the trace element of 18 seed amino acids such as Serine and needed by human etc., the larger Gelucystine of pharmaceutical use wherein, Histidine, arginine content is more, amino acid is differentiated 6～7 points, protein content 16.65%, respectively 3 times and 1.8 times of other Hericium erinaceus (Bull. Ex Fr.) Pers..
Embodiment
Below in conjunction with specific embodiment, describe the present invention, the description of this part is only exemplary and explanatory, should not have any restriction to protection scope of the present invention.
Embodiment 1
A method for medicinal Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium solids liq mixed culture, comprises the following steps: the percentage ratio of following substances is weight percentage,
(1). preparation solid medium
The water that adds surplus, pH nature, mixes 121 ℃ of sterilizings, 30 minutes;
(2). the optimization screening of bacterial classification and sporophore are separated:
The sporophore strong, that physically well develop, do not have disease and pest, seed not to be scattered of growing in wild Hericium erinaceus (Bull. Ex Fr.) Pers. in selecting and purchasing Zhongtiao Shan Mountain virgin forest;
At transfer room more than 100 grades in Bechtop, to in the alcohol of hericium erinaceus fruiting body immersion 75%, sterilize 2 minutes, clean with distilled water flushing after taking-up, with antiseptic gauze, dry again, sporophore is placed on culture dish, with Bell jar or similar apparatus, cover, room temperature remains on 20 ℃, sporophore is slowly grown, cross after 3 days, seed sheds, fall into the culture dish that step (1) substratum is housed, then with tweezers, dip the seed falling, by method of scoring, be inoculated on the inclined-plane of test tube and cultivate, at 22 ℃, cultivating 5 days seeds can sprout, when mycelia is covered with inclined-plane substantially, selection look white, substrate mycelium prosperity is sturdy, bacterium bundle is obvious, insert shaking table bottle graft kind standby,
(3). the cultivation of bacterial classification
Before inoculation, inoculation mouthful should first be sterilized by ordinary method, with the rayon balls of 75% alcohol-pickled mistake, wipe examination, rear unlatching ozonizer, make its air outlet all the time facing to inoculation mouthful, in inoculation mouthful, form a local aseptic area around, then with sterile nontoxic rubber pipe box in inoculation the mouth of pipe on, shaking table bottle bacterial classification access through spore separation is equipped with in the tank of step (1) substratum, and inoculum size is 10%, and all processes all carries out under aseptic condition, after inoculation, open stirrer, 200 revs/min of stirring velocitys, 25 ℃ of constant temperature, cultivate 2 days;
(4). deep liquid is cultivated:
4.1 seed tank culture bases are:
The water that adds surplus, 121 ℃ of sterilizings 30 minutes;
4.2 are seeded to seeding tank by the cultured bacterial classification of step (3), and 26 ℃, cultivate 3 days, to result be:
1.. microscopy: mycelia is dendroid, without miscellaneous bacteria;
2.. outward appearance: have more white peloid in liquid;
(5). Mycelium culture:
Wheat bran 40%, liquorice dregs 20%, the Yanhusuo dregs of a decoction 10%, Semen Maydis powder 10%, cavings 10%, corn ear 5%, wood chip 5%; Adding water mixes, 1% the gypsum that adds again total amount, stir, be divided in polypropylene pouch, 121 ℃ of sterilizings 2 hours, in 100 grades of aseptic areas, access the solid spawn of step 4.2, inoculum size is 1/30, front 12 days 27 ℃, within latter 28 days, in 24 ℃, cultivate, after mycelial growth is full, break 60 ℃ of oven dry of piece, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. filament.
Embodiment 2
A method for medicinal Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium solids liq mixed culture, comprises the following steps: the percentage ratio of following substances is weight percentage,
(1). preparation solid medium
The water that adds surplus, pH value 7.5, mixes, 118 ℃ of sterilizings 30 minutes;
(2). the optimization screening of bacterial classification and sporophore are separated:
The sporophore strong, that physically well develop, do not have disease and pest, seed not to be scattered of growing in wild Hericium erinaceus (Bull. Ex Fr.) Pers. in selecting and purchasing Zhongtiao Shan Mountain virgin forest;
At transfer room more than 100 grades in Bechtop, to in the alcohol of hericium erinaceus fruiting body immersion 75%, sterilize 1 minute, clean with distilled water flushing after taking-up, with antiseptic gauze, dry again, sporophore is placed on culture dish, with Bell jar or similar apparatus, cover, room temperature remains on 20 ℃, sporophore is slowly grown, cross after 2 days, seed sheds, fall into the culture dish that step (1) substratum is housed, then with tweezers, dip the seed falling, by method of scoring, be inoculated on the inclined-plane of test tube and cultivate, at 25 ℃, cultivating 4 days seeds can sprout, when mycelia is covered with inclined-plane substantially, selection look white, substrate mycelium prosperity is sturdy, bacterium bundle is obvious, insert shaking table bottle graft kind standby,
(3). the cultivation of bacterial classification
Before inoculation, inoculation mouthful should first be sterilized by ordinary method, with the rayon balls of 75% alcohol-pickled mistake, wipe examination, rear unlatching ozonizer, make its air outlet all the time facing to inoculation mouthful, in inoculation mouthful, form a local aseptic area around, then with sterile nontoxic rubber pipe box in inoculation the mouth of pipe on, shaking table bottle bacterial classification access through spore separation is equipped with in the tank of step (1) substratum, and inoculum size is 11%, and all processes all carries out under aseptic condition, after inoculation, open stirrer, 200 revs/min of stirring velocitys, 25 ℃ of constant temperature, cultivate 2 days;
(4). deep liquid is cultivated:
4.1 seed tank culture bases are:
The water that adds surplus, 118 ℃ of sterilizings 32 minutes;
4.2 are seeded to seeding tank by the cultured bacterial classification of step (3), and 27 ℃, cultivate 3 days, to result be:
1.. microscopy: mycelia is dendroid, without miscellaneous bacteria;
2.. outward appearance: have more white peloid in liquid;
(5). Mycelium culture:
Wheat bran 35%, liquorice dregs 20%, the Yanhusuo dregs of a decoction 7%, Semen Maydis powder 10%, cavings 7%, corn ear 5%, wood chip 3%; Adding water mixes, 1% the gypsum that adds again total amount, stir, be divided in polypropylene pouch, 121 ℃ of sterilizings 115 minutes, in 100 grades of aseptic areas, access the solid spawn of step 4.2, inoculum size is 1/30, front 12 days 28 ℃, within latter 28 days, in 23 ℃, cultivate, after mycelial growth is full, break 65 ℃ of oven dry of piece, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. filament.
Embodiment 3
A method for medicinal Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium solids liq mixed culture, comprises the following steps: the percentage ratio of following substances is weight percentage,
(1). preparation solid medium
The water that adds surplus, pH value 6, mixes, 121 ℃ of sterilizings, 25 minutes;
(2). the optimization screening of bacterial classification and sporophore are separated:
The sporophore strong, that physically well develop, do not have disease and pest, seed not to be scattered of growing in wild Hericium erinaceus (Bull. Ex Fr.) Pers. in selecting and purchasing Zhongtiao Shan Mountain virgin forest;
At transfer room more than 100 grades in Bechtop, to in the alcohol of hericium erinaceus fruiting body immersion 75%, sterilize 2 minutes, clean with distilled water flushing after taking-up, with antiseptic gauze, dry again, sporophore is placed on culture dish, with Bell jar or similar apparatus, cover, room temperature remains on 18 ℃, sporophore is slowly grown, cross after 3 days, seed sheds, fall into the culture dish that step (1) substratum is housed, then with tweezers, dip the seed falling, by method of scoring, be inoculated on the inclined-plane of test tube and cultivate, at 22 ℃, cultivating 5 days seeds can sprout, when mycelia is covered with inclined-plane substantially, selection look white, substrate mycelium prosperity is sturdy, bacterium bundle is obvious, insert shaking table bottle graft kind standby,
(3). the cultivation of bacterial classification
Before inoculation, inoculation mouthful should first be sterilized by ordinary method, with the rayon balls of 75% alcohol-pickled mistake, wipe examination, rear unlatching ozonizer, make its air outlet all the time facing to inoculation mouthful, in inoculation mouthful, form a local aseptic area around, then with sterile nontoxic rubber pipe box in inoculation the mouth of pipe on, shaking table bottle bacterial classification access through spore separation is equipped with in the tank of step (1) substratum, and inoculum size is 9%, and all processes all carries out under aseptic condition, after inoculation, open stirrer, 200 revs/min of stirring velocitys, 25 ℃ of constant temperature, cultivate 2 days;
(4). deep liquid is cultivated:
4.1 seed tank culture bases are:
The water that adds surplus, 121 ℃ of sterilizings 28 minutes;
4.2 are seeded to seeding tank by the cultured bacterial classification of step (3), and 25 ℃, cultivate 4 days, to result be:
1.. microscopy: mycelia is dendroid, without miscellaneous bacteria;
2.. outward appearance: have more white peloid in liquid;
(5). Mycelium culture:
Wheat bran 40%, liquorice dregs 15%, the Yanhusuo dregs of a decoction 10%, Semen Maydis powder 7%, cavings 10%, corn ear 3%, wood chip 5%; Adding water mixes, 1% the gypsum that adds again total amount, stir, be divided in polypropylene pouch, 118 ℃ of sterilizings 120 minutes, in 100 grades of aseptic areas, access the solid spawn of step 4.2, inoculum size is 1/30, front 12 days 26 ℃, within latter 28 days, in 25 ℃, cultivate, after mycelial growth is full, break 55 ℃ of oven dry of piece, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. filament.
Detect above-described embodiment 1 to embodiment 3 resulting Hericium erinaceus (Bull. Ex Fr.) Pers. filament, result is compared with general edible Hericium erinaceus (Bull. Ex Fr.) Pers., except containing polysaccharide polypeptide, still also has the trace element of five kinds of cancer-resisting substances, seven kinds of VITAMIN, three kinds of active cellulases, 18 seed amino acids and needed by human etc., Gelucystine, Histidine, arginine content that wherein pharmaceutical use is larger are more, amino acid is differentiated 6～7 points, protein content 16.65%, is respectively 3 times and 1.8 times of other Hericium erinaceus (Bull. Ex Fr.) Pers..
Below be only the preferred embodiment of the present invention; it should be pointed out that for those skilled in the art, under the premise without departing from the principles of the invention; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (1)
1. a method for medicinal Hericium erinaceus (Bull. Ex Fr.) Pers. mycelium solids liq mixed culture, comprises the following steps: the percentage ratio of following substances is weight percentage,
(1). preparation solid medium
The water that adds surplus, pH value 6～7.5, mixes, 118～121 ℃ of sterilizings, 25～30 minutes;
(2). the optimization screening of bacterial classification and sporophore are separated;
The sporophore strong, that physically well develop, do not have disease and pest, seed not to be scattered of growing in wild Hericium erinaceus (Bull. Ex Fr.) Pers. in selecting and purchasing Zhongtiao Shan Mountain virgin forest;
At transfer room more than 100 grades in Bechtop, to in the alcohol of hericium erinaceus fruiting body immersion 75%, sterilize 1～2 minute, clean with distilled water flushing after taking-up, with antiseptic gauze, dry again, sporophore is placed on culture dish, with Bell jar apparatus, cover, room temperature remains on 18～20 ℃, sporophore is slowly grown, cross after 2～3 days, seed sheds, fall into the culture dish that step (1) substratum is housed, then with tweezers, dip the seed falling, by method of scoring, be inoculated on the inclined-plane of test tube and cultivate, at 22～25 ℃, cultivating 4～5 days seeds can sprout, when mycelia is covered with inclined-plane substantially, selection look white, substrate mycelium prosperity is sturdy, bacterium bundle is obvious, insert shaking table bottle graft kind standby,
(3). the cultivation of bacterial classification
Before inoculation, inoculation mouthful should first be sterilized by ordinary method, with the rayon balls of 75% alcohol-pickled mistake, wipe examination, rear unlatching ozonizer, make its air outlet all the time facing to inoculation mouthful, in inoculation mouthful, form a local aseptic area around, then with sterile nontoxic rubber pipe box in inoculation the mouth of pipe on, shaking table bottle bacterial classification access through spore separation is equipped with in the tank of step (1) substratum, and inoculum size is 9%～11%, and all processes all carries out under aseptic condition, after inoculation, open stirrer, 200 revs/min of stirring velocitys, 25 ℃ of constant temperature, cultivate 2 days;
(4). deep liquid is cultivated:
4.1 seed tank culture bases are:
The water that adds surplus, 118～121 ℃ of sterilizings 28～32 minutes;
4.2 are seeded to seeding tank by the cultured bacterial classification of step (3), and 25～27 ℃, cultivate 3～4 days, to result be:
1.. microscopy: mycelia is dendroid, without miscellaneous bacteria;
2.. outward appearance: have more white peloid in liquid;
(5). Mycelium culture:
Wheat bran 40%, liquorice dregs 20%, the Yanhusuo dregs of a decoction 10%, Semen Maydis powder 10%, cavings 10%, corn ear 5%, wood chip 5%; Adding water mixes, 1% the gypsum that adds again total amount, stir, be divided in polypropylene pouch, 118～121 ℃ of sterilizings 115～120 minutes, in 100 grades of aseptic areas, access the solid spawn of step 4.2, inoculum size is 1/30, front 12 days 26～28 ℃, within latter 28 days, in 23～25 ℃, cultivate, after mycelial growth is full, break 55～65 ℃ of oven dry of piece, obtain Hericium erinaceus (Bull. Ex Fr.) Pers. filament.
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