CN106520569A - White rot fungi medium taking crop straws as main body and preparation method of white rot fungi medium - Google Patents
White rot fungi medium taking crop straws as main body and preparation method of white rot fungi medium Download PDFInfo
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- CN106520569A CN106520569A CN201610976590.0A CN201610976590A CN106520569A CN 106520569 A CN106520569 A CN 106520569A CN 201610976590 A CN201610976590 A CN 201610976590A CN 106520569 A CN106520569 A CN 106520569A
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- culture medium
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
Abstract
The invention provides a white rot fungi medium taking crop straws as a main body and a preparation method of the white rot fungi medium. White rot fungi mainly relate to phanerochaete chrysosporium. The medium is prepared from the following raw material components in parts by weight: 60-73 parts of the crop straws, 23-36 parts of bran, 2.5 parts of glucose, 0.5 part of KH2PO4 and 1 part of (NH4)2SO4, and meanwhile, a solid-to-liquid ratio is at 1 to 4.5-1 to 6.5; the straws that coarse peels are peeled off are crushed by virtue of a pulverizer and the crushed straws are filtered by virtue of a sieve; the components are fully mixed in accordance with the proportions; water at a corresponding quantity is added in accordance with the solid-to-liquid ratio; and pH value is adjusted to be 6.9-7.2. The white rot fungi medium provided by the invention can solve the problems of an existing white rot fungi medium which is complex in process, long in cycle and poor in growth vigor. Meanwhile, in accordance with the nutrition demand of the white rot fungi, the provided medium is capable of guaranteeing balanced nutrition and is conducive to the growth of the fungi; and by taking the crop straws as the main body, the medium achieve resource recycling and provides pre-processing for a biological pulping process of the crop straws.
Description
Technical field
The invention belongs to agricultural straw resource utilizes technical field, more particularly to one kind based on agricultural crop straw
The white-rot fungi culture medium of body and its compound method.
Background technology
China is large agricultural country, the generation for having substantial amounts of crop straw refuse every year, containing a large amount of in these stalks
Can be used as the cellulose and hemicellulose of feed and paper making raw material.In plant tissue, lignin and hemicellulose are with covalent bond
Form is combined, and cellulosic molecule is embedded wherein, forms a kind of firm natural cover for defense, makes general microorganism be difficult degraded profit
With constraining the recycling of agricultural crop straw.
Mycophyta micro- life of the whiterot fungi as most effective so far, topmost degradable lignocellulosic in nature
Thing, can lignocellulose degradation be thoroughly CO2And H20.Ligocellulose degradation fungi can be divided into 3 classes：Whiterot fungi (White rot
Fungi), brown rot fungus (Brown rot fungi) and soft-rot fungi (Soft rot fungi).Makings brown rot fungus can only be attacked and be dropped
Cellulose, hemicellulose fraction in solution cell membrane, and soft-rot fungi acts only on the hemicellulose in cell, only whiterot fungi is
In nature, a unique class has the microorganism of independent lignin degrading ability, and brown rot fungus, soft-rot fungi are generally acknowledged that in lignin
Secondary action is played in degraded only.Eccrine fiber element enzyme, hemicellulase, pectin are gone back while whiterot fungi secretion lignin-degrading enzymes
The enzymes such as enzyme, thus be more suitable for decomposing natural lignocellulosic materials than trichoderma, aspergillus etc..
Cellulose is the main component of plant cell wall, is widely present in nature, is most abundant on the earth, most cheap
Renewable resource, and the main source of the possible energy of human future, food and industrial chemicals.Using cellulase by fiber
Element is converted into available sugar, for the enforcement of the strategy of sustainable development is significant.Therefore lignin degrading is first had to,
Cellulose is utilized to greatest extent could.
Lignocellulosic is the three-dimensional netted high score of poly- phenols being formed by connecting by ehter bond and carbon-carbon bond by benzene oxide unit
Sub- compound.Lignocellulosic is difficult to be degraded by microorganisms due to the complicated of bonding with various bio-stables.It is how high
Using lignocellulosic material, which it is critical only that effectively degraded is wrapped in lignin and half fiber outside cellulose crystals to effect
Element, so as to increase cellulose surface product, makes cellulose be easy to degraded, so as to generate fermentable sugar, and is applied to biological system
During slurry.
The content of the invention
The technical problem to be solved in the present invention is to provide a kind of white-rot fungi culture medium based on agricultural crop straw, and
Its preparation method is provided.
To reach above-mentioned purpose, the technical scheme is that：
A kind of semisolid culturemedium of the present invention, culture medium component by weight include following components：
| Stalk | 56-75 parts |
| Wheat bran | 20-40 parts |
| (NH4)2SO4 | 1 part |
| KH2PO4 | 0.5 part |
| Glucose | 2.5 part |
The summation of said components is 100 parts.
Preferably, the solid-to-liquid ratio of the culture medium is 1: 4.5～1:6.5.(solid-to-liquid ratio is solid matter and water in culture medium
Mass ratio)
Preferably, the pH value of the culture medium is 6.9-7.2.
Preferably, the semisolid culturemedium, culture medium component by weight include following components：
| Stalk | 60-73 parts |
| Wheat bran | 23-36 parts |
| (NH4)2SO4 | 1 part |
| KH2PO4 | 0.5 part |
| Glucose | 2.5 part |
The summation of said components is 100 parts.
Described fungi is white-rot fungi, for degraded using lignin in stalk.
The preparation method of the culture medium is：
(1) pre-process：The crushed stalk for having been peeled off tertia with pulverizer is sieved, and the size of sieve is 80-100 mesh；
(2) said components are sufficiently mixed according to the proportioning；
(3) added after a certain amount of water according to solid-to-liquid ratio, it is that 6.9-7.2 can be used to adjust pH value.
The agricultural crop straw is common stalk：Rice straw, wheat straw waste and maize straw.It is furthermore preferred that the straw
Stalk is wheat straw waste.
The present invention also provides the purposes of the culture medium, for the culture medium of whiterot fungi.
Preferably, the purposes of the culture medium, the culture medium that making is finished, is aseptically inoculated with whiterot fungi,
And be put in homoiotherm incubator, 32～37 DEG C of design temperature, 70～90rpm of rotating speed.It is furthermore preferred that rotating speed is 80rpm.
Whiterot fungi applied in the present invention is：Phanerochaete chrysosporium (Phanerochaete chrysosporium)
The culture medium of whiterot fungi involved in the present invention, can ensure the same of the supply of nutrient needed for whiterot fungi growth course
When optimization culture technique, shorten cultivation cycle, reduce production cost and be conducive to promote whiterot fungi Biological Pretreatment crops
Stalk resource and industrialized utilization.
Beneficial effects of the present invention one：Growth demand of the present invention according to bacterial classification, there is provided a kind of nutrition is balanced, beneficial to bacterial classification
Production, cheap fungi culture medium；
Beneficial effects of the present invention two：The preparation of the culture medium of whiterot fungi involved in the present invention, can ensure whiterot fungi
Optimization culture technique while the supply of nutrient needed for growth course, shortens cultivation cycle, reduces production cost, is conducive to promoting
The industrialized utilization of whiterot fungi Biological Pretreatment agricultural crop straw
Beneficial effects of the present invention three：Culture medium provided by the present invention based on agricultural crop straw, using whiterot fungi
Selection degradability to lignin, improves the recycling value of agricultural crop straw, be straw refuse in feed and
Application in terms of bio-pulping is provided may.
Description of the drawings
Fig. 1 is that (PSD is culture provided by the present invention to two kinds of culture medium degraded 30d Lignin degradation rate comparison diagrams of wheat straw
Base；PSC is traditional liquid culture medium)
Fig. 2 is that (PSD is culture provided by the present invention to two kinds of culture medium degraded 30d mycelial growth situation comparison diagrams of corn
Base；PSC is traditional liquid culture medium)
Specific embodiment
Embodiment 1：
A kind of Phanerochaete chrysosporium containing wheat stalk (Phanerochaete chrysosporium) culture medium
Including the component of following weight portion：
| Wheat straw waste | 73g, |
| (NH4)2SO4 | 1g, |
| Glucose | 2.5g, |
| Wheat bran | 23g, |
| KH2PO4 | 0.5g, |
| Solid-to-liquid ratio | 1:4.5 |
Preparation method is：Wheat straw waste is removed into the tertia of appearance, straw section is deducted with scissors, is crushed to 80-100 mesh, presses
Each material is sufficiently mixed according to said ratio, adjust moisture and pH value is corrected for 7.2, you can use.In incubation, training
Foster base is placed in 32 DEG C of constant incubators, rotating speed 80rpm.
Aseptically, the culture medium 5g through a period of time culture is taken out, vacuum freeze drying 8h both can be according to
Klason methods determine content of lignin in matrix, are contrasted with the front content of culture and be both obtained lignin degradation in incubation
Rate.Experimental result is shown in Fig. 1.
It is demonstrated experimentally that by content of lignin in matrix before and after measure culture it is known that culture provided by the present invention
Base can lignin in high-efficient culture Phanerochaete chrysosporium degradation selectivity mechanism, while mycelium growth vigor is trained than conventional solid
Sturdy many of the growing way of foster base, and the speed of growth of mycelia 3-5 days in advance.
Embodiment 2：
A kind of Phanerochaete chrysosporium containing rice straw (Phanerochaete chrysosporium) Semi-solid cell culture
Base
Including the component of following weight portion：
| Rice straw | 60g |
| (NH4)2SO4 | 1g |
| Glucose | 2.5g |
| Wheat bran | 36g |
| KH2PO4 | 0.5g |
| Solid-to-liquid ratio | 1:5.5 |
Preparation method is：Rice straw is removed into the tertia of appearance, straw section is deducted with scissors, is crushed to 80-100 mesh, presses
Each material is sufficiently mixed according to said ratio, adjust moisture and pH value is corrected for 6.9, you can use.In incubation, training
Foster base is placed in 37 DEG C of constant incubators, rotating speed 80rpm.
Phanerochaete chrysosporium incubation Enzymatic characteristic is determined in incubation.Specific embodiment：
(1) formulation of crude enzyme liquid
Under aseptic condition, by the PSC (traditional liquid culture medium) of the rice straw in shaking table in first ten days incubator,
PSD (semisolid culturemedium provided by the present invention) culture medium respectively takes out one, prepares six small beakers, and six culture dishes will
Each culture medium is divided equally into two halves, and wherein half is transferred in small beaker together with the bacterium on second half surface, adds 40ml
4h is extracted under distilled water, room temperature, 10min is centrifuged with 10000rpm/min after 4 layers of filtered through gauze, supernatant is crude enzyme liquid, puts 4
DEG C refrigerator is standby.Second half is paved in being transferred to culture dish, wraps culture dish with preservative film, pricks some apertures, is put into -20 DEG C
It is standby in refrigerator.
(2) measure of enzymatic activity
Phanerochaete chrysosporium can produce the lignin that extracellular oxidizing ferment comes in degrading straw, and which mainly includes manganese peroxide
Compound enzyme (MnP), lignin peroxidase (LiP), laccase (Lac).Here is illustrated by taking manganese peroxidase as an example.
MnP
Take sodium lactate buffer solution (pH4.5) 3.4mL of 50mmol/L, the MnSO of 1.6mmol/L4Solution 0.1mL, 0.4mL
Crude enzyme liquid be preheated to 37 DEG C addition 1.6mmol/L hydrogenperoxide steam generator 0.1mL start reaction.Determine in initial 3min
Absorbance at 240nm, 1 enzyme activity unit min-1·g-1Crude enzyme liquid increases 0.1OD to represent.
1 Phanerochaete chrysosporium of table Mnp enzymatic activitys activity change (U/mL) on different culture media
Table4 Phanerochaete chrysosporium Mnp activity changes of activity
(U/mL)on different media
Can analyze from table 1 and obtain, during 5-10d after incubation, it is active to start to show Mnp, and in different cultures
The trend for raising-reducing is presented in base, is peaked in the 15d of culture, PSC the and PSD enzyme activity of straw is up to 49.70U/
ML and 51.81U/mL.Hereafter there is downward trend in enzymatic activity.
It is demonstrated experimentally that semi-solid fungi culture medium provided by the present invention in incubation with conventional solid culture medium phase
Than in identical incubation time, the yield of manganese peroxidase is significantly improved, and illustrates which selects degradability more to lignin in matrix
Good, Phanerochaete chrysosporium growth is more vigorous, and biomass more enriches.And mycelium growth vigor is than conventional solid culture medium
Growing way is equally dense, sturdy, and the speed of growth of mycelia is slightly in advance.
Embodiment 3
A kind of Phanerochaete chrysosporium containing maize straw (Phanerochaete chrysosporium) culture medium
Including the component of following weight portion：
| Maize straw | 65g |
| (NH4)2SO4 | 1g |
| Glucose | 2.5g |
| Wheat bran | 31g |
| KH2PO4 | 0.5g |
| Solid-to-liquid ratio | 1:6.5 |
Preparation method is：Wheat straw waste is removed into the tertia of appearance, straw section is deducted with scissors, is crushed to 80-100 mesh, presses
Each material is sufficiently mixed according to said ratio, adjust moisture and pH value is corrected for 7.0, you can use.In incubation, training
Foster base is placed in 32 DEG C of constant incubators, rotating speed 80rpm.
Aseptically, PSC (traditional liquid culture medium), the PSD of the maize straw through a period of time culture are taken out
(culture medium provided by the present invention) culture medium 5g is moved in the bacterium bag culture medium of same component, is placed in 32 DEG C of constant incubators
Continuous culture 30 days.Mycelial growth situation was recorded every 5 days, mycelia covering position was marked using marking pen, with mycelia after 30 days
Cover bacterium bag surface area changing value to count.Experimental result is shown in Fig. 2.
It is demonstrated experimentally that by contrasting PSC with the net speed of growth of two kinds of different culture media mycelia of PSD it is known that the present invention
The net speed of growth of culture medium PSD mycelia for being provided wants fast conventional medium PSC, and reaches maximum growth rate at the 15th day,
Maximum growth area was reached at 30 days.During experiment proves culture medium provided by the present invention, Phanerochaete chrysosporium grows more
Plus it is vigorous, biomass more enriches.And mycelium growth vigor is denser than the growing way of conventional solid culture medium, sturdy, the life of mycelia
Long speed shifts to an earlier date in advance for 3-5 days.
Claims (10)
1. a kind of semisolid culturemedium, culture medium component by weight include following components：
The summation of said components is 100 parts.
2. semisolid culturemedium as claimed in claim 1, it is characterised in that the solid-to-liquid ratio of the culture medium is 1: 4.5～1:
6.5。
3. semisolid culturemedium as claimed in claim 1 or 2, it is characterised in that the pH value of the culture medium is 6.9-7.2.
4. the semisolid culturemedium as described in any one of claims 1 to 3, it is characterised in that culture medium component by weight includes
Following components：
The summation of said components is 100 parts.
5. the semisolid culturemedium as described in any one of Claims 1 to 4, it is characterised in that the stalk is rice straw, wheat
Careless stalk or maize straw.It is furthermore preferred that the stalk is wheat straw waste.
6. the preparation method of the semisolid culturemedium as described in any one of Claims 1 to 5, comprises the following steps：
(1) pre-process：The crushed stalk for having been peeled off tertia with pulverizer is sieved, and the size of sieve is 80-100 mesh；
(2) said components are sufficiently mixed according to the proportioning；
(3) added after a certain amount of water according to solid-to-liquid ratio, it is that 6.9-7.2 can be used to adjust pH value.
7. the purposes of the semisolid culturemedium as described in any one of Claims 1 to 5, for the culture medium of whiterot fungi.
8. the purposes of semisolid culturemedium as claimed in claim 7, it is characterised in that described whiterot fungi is：Yellow archespore hair
Flat lead fungi (Phanerochaete chrysosporium).
9. the purposes of semisolid culturemedium as claimed in claim 7, it is characterised in that the culture medium for finishing will be made, in nothing
Whiterot fungi is inoculated with the conditions of bacterium, and is put in homoiotherm incubator, 32～37 DEG C of design temperature, 70～90rpm of rotating speed.
10. the purposes of semisolid culturemedium as claimed in claim 7, it is characterised in that rotating speed is 80rpm.
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| CN201610976590.0A CN106520569A (en) | 2016-10-28 | 2016-10-28 | White rot fungi medium taking crop straws as main body and preparation method of white rot fungi medium |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108130277A (en) * | 2017-12-01 | 2018-06-08 | 齐鲁工业大学 | A kind of white-rot fungi culture medium and its preparation method based on lignin |
| CN113396777A (en) * | 2021-06-17 | 2021-09-17 | 齐鲁工业大学 | Method for treating agricultural organic solid waste by using ammonia fiber expansion in cooperation with white rot fungi |
| CN114586640A (en) * | 2022-03-24 | 2022-06-07 | 山西农业大学农学院(山西省农业科学院作物科学研究所) | Wheat straw based cultivation mechanism and manufacturing method thereof |
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| CN101045918A (en) * | 2006-01-05 | 2007-10-03 | 安徽大学 | Solid state tramete AH28-2 fermenting process for producing laccase |
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2016
- 2016-10-28 CN CN201610976590.0A patent/CN106520569A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101045918A (en) * | 2006-01-05 | 2007-10-03 | 安徽大学 | Solid state tramete AH28-2 fermenting process for producing laccase |
Non-Patent Citations (2)
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| 武忠伟 等: "白腐真菌固体发酵玉米秸秆产漆酶条件的优化", 《湖北农业科学》 * |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108130277A (en) * | 2017-12-01 | 2018-06-08 | 齐鲁工业大学 | A kind of white-rot fungi culture medium and its preparation method based on lignin |
| CN113396777A (en) * | 2021-06-17 | 2021-09-17 | 齐鲁工业大学 | Method for treating agricultural organic solid waste by using ammonia fiber expansion in cooperation with white rot fungi |
| CN114586640A (en) * | 2022-03-24 | 2022-06-07 | 山西农业大学农学院(山西省农业科学院作物科学研究所) | Wheat straw based cultivation mechanism and manufacturing method thereof |
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Application publication date: 20170322 |
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