CN106071106A - Fermentation cottonseed protein and the method for solid-state aerobic fermentation preparation fermentation cottonseed protein - Google Patents
Fermentation cottonseed protein and the method for solid-state aerobic fermentation preparation fermentation cottonseed protein Download PDFInfo
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Abstract
The invention belongs to technical field of microbial fermentation, be specifically related to a kind of cottonseed protein and method of solid-state aerobic fermentation preparation fermentation cottonseed protein of fermenting.The technical problem to be solved in the present invention is to provide a kind of fermentation cottonseed protein.This fermentation cottonseed protein is formed by the fermenting raw materials of following weight proportion: cottonseed meal powder 97～99 parts, Semen Maydis powder 0～1.5 parts, mixed yeast solid spawn 0.7～1.5 parts, mixotrophism salt 0.1～0.4 part, mixing enzyme preparation 0.03～0.1 part, water；The gross weight of water accounts for 0.6～0.9 times of other five kinds of raw material gross weights.And the solid-state aerobic fermentation preparation method of the cottonseed protein that ferments, then mainly comprise the steps that and take each raw material mix homogeneously, in 30～35 DEG C of bottom fermentations 45～48h under good oxygen condition, be dried.The inventive method can remove the gossypol in cottonseed meal and antinutritional factor effectively, provides a better way for effectively utilizing of cottonseed meal.
Description
Technical field
The invention belongs to prepare technical field of microbial fermentation, be specifically related to a kind of fermentation cottonseed protein and aerobic of solid-state
Ferment preparation fermentation cottonseed protein its preparation method.
Background technology
China's aquaculture heavy dependence corn-soybean meal diet at present, particularly main protein raw material bean cake, the biggest portion
Being divided into import, be limited by international market, often result in aquaculture cost fluctuation, risk increases.It addition, containing turning in import bean cake
Gene element, does not meets the requirement of organic farm products.
Cottonseed meal are the by-product after Semen Gossypii oil expression, rich in nutrition content, and albumen is more than 40%, and aminoacid composition is more flat
Weighing apparatus, arginine content is higher than bean cake.China's annual cottonseed meal yield about 6,000,000 tons, the stock number whole world first, but cottonseed meal contain more
Poisonous gossypol and antinutritional factor, limit its application in animal diets.
Gossypol is primarily present in Cottonseed pigment gland, is a kind of water insoluble and be dissolved in the poly-phenol of yellowish-brown of organic solvent
Pigment.During liquefaction, due to heat effects such as steaming are fried, squeezing, major part gossypol is combined with protein, aminoacid and becomes knot
Closing gossypol, bound gossoypol is not absorbed by animal in animal alimentary canal, therefore toxicity is the least.Another part gossypol is the most in a free form
Existing, relatively big to animal toxicity, especially nonruminant excess ingestion or the picked-up time is longer, may result in growth retardation, reproductive ability
Energy and production performance decline, and even result in death.Brood is lower to the tolerance of gossypol.Undressed cottonseed meal middle reaches
From gossypol content 800～1100mg/kg.Due to the existence of toxicity gossypol, cottonseed meal usage amount in animal and fowl fodder be 3～
6%, young stock fowl does not the most use cottonseed meal.
At present commercialization cottonseed protein product only has the dephenolize cottonseed meal using organic solvent extraction, and free gossypol content can be
Below 400mg/kg, but extraction can cause the loss of part nutrient substance.
And the bioremediation that prior art uses is mainly anaerobe fermentation method, but if to hold concurrently simultaneously
Turn round and look at detoxification and eliminate antinutritional factor, improve digestibility, the effect of the raising aspect such as nutritive value and feeding effect, being then
Extremely difficult, it is extremely difficult to the effect wanted.For solving an above-mentioned difficult problem, existing experimental technique mainly uses the bacterial strain group of complexity
Closing, be separately added into different strains and decompose corresponding material, but these methods often run counter to Microbiological Principle, bacterial strain is the most past
Toward contaminated bacteria each other, influence each other, long potential difference；Or the optimum growing condition difference of these bacterial strains itself is relatively big, in same base
In matter, growth interferes.Additionally, in the fermentation process of prior art, often add more wheat bran and corn as auxiliary materials (account for 10～
20%), and mixing dregs of rice class fermentation is used, the bad factor contents such as cottonseed meal content is usually less than 80% in total raw material, gossypol
Dilute by rushing, difficulty far below single cottonseed meal ferment, can not reach single feed raw material requirement (single component account for 95% with
On).
From microbiology angle, to a kind of stock substrate, the bacterial strain of uniqueness, and corresponding condition of culture need to be selected.Existing
In technology, also using general leaven, technique is simple, for various fermenting raw materials (such as bean cake, cottonseed meal and grain slag etc.), actual
On it is optimal that each raw material all can not be reached；If after industry is amplified, fermentation efficiency is relatively low, it is suitable only for sideline production agricultural raw
Produce.
This area there is presently no develops preferably, can be under the conditions of single-matrix, to the gossypol in cottonseed meal and anti-battalion
Support the factor etc. and can carry out resolution process well, and improve the biological method of albumen solubility.
Summary of the invention
The defect existed for existing chemistry dephenolize method and microbe fermentation method, the present invention provides a kind of fermentation Semen Gossypii egg
Pseudobulbus Bletillae (Rhizoma Bletillae) its preparation method.The method cottonseed meal account for total raw material more than 97%, for the fermentation of single raw material.Use high aerobic fermentation and enzyme
Solve synchronization to carry out processing cottonseed meal, poisonous free gossypol can not only be reduced, can also obvious degradation raw material resist such as phytic acid etc. simultaneously
Trophic factors and increase acid-soluble protein content, thus improve the nutritive value of product.
First technical problem to be solved by this invention is to provide a kind of fermentation cottonseed protein.This fermentation cottonseed protein by
The fermenting raw materials of following weight proportion forms: cottonseed meal powder 97～99 parts, Semen Maydis powder 0～1.5 parts, mixed yeast solid spawn
0.7～1.5 part, mixotrophism salt 0.1～0.4 part, mixing enzyme preparation 0.03～0.1 part, water；The gross weight of water accounts for other five kinds
0.6～0.9 times of raw material gross weight.
Preferably, above-mentioned fermentation cottonseed protein is formed by the fermenting raw materials of following weight proportion: 98 parts of cottonseed meal powder, Semen Maydis powder
0.5 part, mixed yeast solid spawn 1.2 parts, mixotrophism salt 0.25 part, mixing enzyme preparation 0.05 part and water, the gross weight of water
Amount accounts for 0.8 times of other five kinds of raw material gross weights.
Wherein, the cottonseed meal powder described in above-mentioned fermentation cottonseed protein is after the cottonseed meal pulverizer through 2.0mm aperture is pulverized,
Obtain with 20 mesh sieve sub-sieves again.
Further, in above-mentioned fermentation cottonseed protein, mixed yeast solid spawn includes following strain: detain capsule laminating adhesive
Spore yeast (Saccharomycopsis fibuligera), Candida utilis (Candida utilis) and Marx gram
Shandong dimension yeast (Kluyveromyces marxianus).
Prepare saccharomycopsis fibuligera solid spawn and Candida utilis and yeast Kluyveromyces marxianus bacterium the most respectively
Compound barm culture fluid.Again by saccharomycopsis fibuligera solid spawn and Candida utilis and yeast Kluyveromyces marxianus
The mixing liquid spawn combined inoculation of bacterium, in wheat bran corn culture medium, is obtained by culture mixed yeast solid spawn.
Concrete, the preparation method of the saccharomycopsis fibuligera solid spawn described in above-mentioned fermentation cottonseed protein is:
Picking 1 ring from the slant preservation strain of saccharomycopsis fibuligera (Saccharomycopsis fibuligera)
(inoculating loop), accesses in 100mL Rhizoma Solani tuber osi sucrose culture fluid, in 28～32 DEG C of temperature shake-flask culture (150～200rpm) cultivate 2～
3d, obtains saccharomycopsis fibuligera culture fluid.The most by weight, saccharomycopsis fibuligera liquid wheat bran corn culture medium=0.5
～1.5 100, culture fluid is proceeded in wheat bran corn culture medium, cultivate 2～3d in 28～32 DEG C of temperature, capsule laminating adhesive spore must be detained
Yeast solids strain.
Concrete, the preparation method of the compound barm culture fluid described in above-mentioned fermentation cottonseed protein is:
Respectively from Candida utilis (Candida utilis) and yeast Kluyveromyces marxianus bacterium
In the slant preservation strain of (Kluyveromyces marxianus), each picking 1 ring (inoculating loop), is respectively connected to 100mL Rhizoma Solani tuber osi
In sucrose culture fluid, cultivate 2～3d in 28～32 DEG C of temperature shake-flask culture (150～200rpm), respectively obtain Candida utilis ferment
Female bacteria culture fluid and yeast Kluyveromyces marxianus bacteria culture fluid；The most by weight, Candida utilis culture fluid Marx gram
Dimension yeast culture fluid=0.75, Shandong～0.9 0.1～0.25 mixing access in Rhizoma Solani tuber osi sucrose culture fluid, in 28～32 DEG C of temperature
Shake-flask culture (150～200rpm) cultivates 2～3d, i.e. obtains compound barm liquid；Wherein, Candida utilis culture fluid and horse
Gram this kluyveromyces bacteria culture fluid gross weight and Rhizoma Solani tuber osi sucrose culture fluid weight ratio are 1 100；
Concrete, the preparation method of the mixed yeast solid spawn described in above-mentioned fermentation cottonseed protein is:
By weight, saccharomycopsis fibuligera solid spawn: compound barm liquid wheat bran corn culture medium=0.5～
1.0 0.2～0.5:100, mix homogeneously, cultivate 2～3d in 28～32 DEG C of temperature, obtain mixed yeast solid spawn.
Preferably, Candida utilis culture fluid yeast Kluyveromyces marxianus bacterium described in above-mentioned fermentation cottonseed protein
Culture fluid=0.85 0.15.
Concrete, the preparation method of the wheat bran corn culture medium described in above-mentioned fermentation cottonseed protein is: by weight ratio
Meter, wheat bran Semen Maydis water=7～8 2～3 8 by three mix after at 118～126 DEG C sterilizing 25～40min, cool down.
Concrete, the preparation method of the mixotrophism salt described in above-mentioned fermentation cottonseed protein is: counts by weight ratio, takes
1 part of KCl, 0.5 part of MgSO4, 0.05 part of FeSO4With 0.025 part of ZnSO4Mix.
Concrete, the preparation method of the mixing enzyme preparation described in above-mentioned fermentation cottonseed protein is: counts by weight ratio, plants
Acid enzyme-cellulose enzyme xylanase pectase papain=10～20 2～3 0.8～1.5 0.8～1.5 0.8～
1.5 mix five.
Preferably, the mixing enzyme preparation described in above-mentioned fermentation cottonseed protein is to count phytase cellulose by weight ratio
Enzyme xylanase pectase papain=10 2111.
Concrete, described in above-mentioned fermentation cottonseed protein, the enzyme activity of phytase is 5000 units, described cellulase
Enzyme activity is 5000 units, and the enzyme activity of described xylanase is 100,000 units, and the enzyme activity of described pectase is 30,000 units,
The enzyme activity of described papain is 800,000 units.
It is good that second technical problem to be solved by this invention is to provide a kind of solid-state preparing above-mentioned fermentation cottonseed protein
Aerobe fermentation method.The method comprises the following steps:
A, pulverize and sieve: the pulverizer taking cottonseed meal 2.0 sieves is pulverized.Again after the sieve dressing sieve with 20～30 mesh apertures
Cottonseed meal powder；
B, mixing: take cottonseed meal powder, Semen Maydis powder, mixed yeast solid spawn, mixotrophism salt, mixing enzyme preparation and water mixed
Even fermentation material；
C, fermentation: by fermentation material under good oxygen condition in 30～35 DEG C of bottom fermentations 45～50h, the most i.e. obtain fermented cotton
Seed albumen.
Conventional drying mode temperature is natural drying in the environment of not higher than 60 degree.
Preferably, above-mentioned preparation method, in step B, during fermentation, the thickness of fermentation material is no more than 60mm.
Preferably, above-mentioned preparation method, the container used that ferments in step B is any one of stainless steel or bamboo
Kind.Such as, rustless steel fermentation dish or bamboo fermentation dish can be used.
The beneficial effects of the present invention is: the present invention cottonseed meal that ferment compared with untreated cottonseed meal (crude protein about 44%),
Under identical Water Content Conditions, crude protein >=50%, total amino acids >=44%, free gossypol clearance >=60%, phytic acid is degraded
Rate about 30%, pectin degrading rate 42%, acid-soluble protein content increases by more than 100%, Feeding Value is greatly improved total.The present invention
In fermentation raw material, cottonseed meal account for more than 97%, can more substantially effectively process cottonseed meal than prior art.Apply product of the present invention permissible
Reduce aquaculture cost, additional income；Use product of the present invention can improve animal body and intestinal health level, strengthen disease-resistant
Power, can be conventional without antibiotic, is conducive to carrying out nonreactive healthy aquaculture；Through ferment treatment and fermentation, rich in somatomedin and
Peptides etc., can improve the qualities such as meat egg, improve agricultural product quality；Improve breeding environment, be conducive to decontamination of human excreta and money
Source, development cleaning cultivation and ecological circulation.
Detailed description of the invention
Prior art uses chemical-solvent method and anaerobe fermentation method, will take into account detoxification simultaneously and eliminate anti-nutrition
The factor, raising digestibility, raising nutritive value and feeding effect are extremely difficult to the effect wanted.For solving above-mentioned difficulties,
Prior art mainly uses the strain combinations of complexity, adds bacterial strain and decomposes corresponding material, but these methods are often run counter to micro-
Biological Principles, bacterial strain contaminated bacteria the most each other, influence each other, long potential difference；Or the most suitable growth of these bacterial strains itself
Condition difference is relatively big, grows and interfere in same matrix.Therefore it is necessary to reasonably to select bacterial strain and enzyme preparation, ability
Under the conditions of single-matrix, the gossypol in cottonseed meal and antinutritional factor are carried out good processed.
In order to remove poisonous and harmful substance well, select suitable strain, the present invention antibacterial, yeast and silk
Hundreds of strain bacterium of shape fungus three major types microorganism, carry out anaerobic and aerobic cultivation in cottonseed meal culture medium respectively, determine bacterial strain
Increment, gossypol resolution ratio, crude protein content and outward appearance abnormal smells from the patient, eliminate filamentous fungi；Further, mix according to bacterial strain again
Close and cultivate indexs such as measuring the increment of bacterial strain, gossypol resolution ratio, crude protein content and outward appearance abnormal smells from the patient, then consider bacterial strain originally
The safety of body, the complexity of industrialized production, the resultant effect of fermentation etc. finally determines selection saccharomycopsis fibuligera, product
Protein candidiasis and three kinds of bacterial strains of yeast Kluyveromyces marxianus bacterium carry out processing cottonseed meal.
Being found through experiments, in above-mentioned three kinds of bacterial strains, main bacterial strain is saccharomycopsis fibuligera, and Biomass height has fall gossypol energy
Power, secondary bacterial strain Candida utilis has faint fall gossypol ability, yeast Kluyveromyces marxianus Pseudomonas SHENGXIANG yeast, can produce a small amount of
Ester is fragrant, improves fermentation cottonseed protein palatability；This three strains bacterium all can have inhibitory action to intestinal gram negative bacteria, can improve intestinal
Road non-specific immunity, part cell can pass through intestinal, enters feces, is conducive to cleaning cultivation.
It has also been found that, secondary inoculation amount is crossed conference and is reduced the fall gossypol ability of main bacterial strain, accordingly, it is preferred that scheme
It it is the inoculative proportion more accurately determining main and secondary bacterial strain under industrial process conditions.Experimental studies have found that through substantial amounts of, first
Obtain compound barm bacteria culture fluid after Candida utilis and yeast Kluyveromyces marxianus bacterium are mixed amplification culture, control to add
Being added to the ratio in mixed yeast solid spawn, then the ratio with saccharomycopsis fibuligera solid spawn is 0.2～0.5:
Gossypol can be decomposed well when 0.5～1.0.When preparing mixed yeast solid spawn, preferably mixed yeast is cultivated
Liquid 0.4 part, saccharomycopsis fibuligera solid spawn 0.8 part.
Be further discovered that, although above-mentioned three kinds of strain combinations fermentation can decompose gossypol, but to degraded antinutritional factor and
Improve and digest and assimilate limited use.In order to solve the problems referred to above, the present invention initially passes through a large amount of screening, selects phytase producing strains
With Pectinase Producing Strain etc., adding in strain combinations, it is desired to be able to play the effect reducing antinutritional factor, experiment later is sent out
Existing, under conditions of with single cottonseed meal raw material and multiple bacterium, it not the optimal condition of enzyme production of phytase and pectase, more complicated
Strain combinations is difficult to take into account both had decomposed gossypol, reduced again antinutritional factor.
In the face of an above-mentioned difficult problem, further studying, discovery can add compound enzymic preparation to solve the problems referred to above, by reality
The common effect of existing employing cellulase, phytase, pectase and xylanase of issuing after examination and approval can not only be decomposed in rapeseed meal and mainly be resisted
Trophic factors, moreover it is possible to improve the dephenolize ability of microorganism；It has furthermore been found that enzyme compound use has certain synergism, such as cotton
In the dregs of rice, phytic acid content is higher, the phytase decomposition to phytic acid, can reduce the phytic acid combination to nutrient, decomposes from phytic acid and releases
The phosphorus put provides nutrient, beneficially growth of microorganism for growth of microorganism again, and the catabolite of pectin and xylan etc. is also in addition
It is provided that the nutrient of growth of microorganism.
Experiment also finds, under the conditions of same use, papain has Degradation to other several pherons, with
Time lengthening, enzymatic activity declines.Therefore, consumption and ratio to papain have done experimentation again, control Fructus Chaenomelis egg
The addition of white enzyme, can make the activity of other several enzymes retain more than 50% in the front 10h of processing procedure.Raw by industry
Produce experiment find, when phytase cellulase xylanase pectase papain=10～20 2～3 0.8～
1.5 0.8～1.5 0.8～1.5 ratios prepare mixing enzyme preparation, can process the antinutritional factor in cottonseed meal well.
Preferably phytase cellulase xylanase pectase papain=10 2111, it was found that fermentation Semen Gossypii
The acid original cottonseed meal of molten protein ratio of albumen improves about 100%, and gossypol resolution ratio is brought up to about 60%.
Present invention discover that: while adding enzyme preparation, it should control strain inoculum concentration, enzymolysis process is also strain amplification
Process.If inoculum concentration is excessive, cell proliferation is too fast, and fermentation time is too short, is unfavorable for zymolysis；Otherwise inoculum concentration is too small,
Longer fermentation times, easily causes miscellaneous bacteria hypertrophy.
It has also been found that, the water yield of addition is other solid feeds 0.6～when 0.9 times, both can meet enzymolysis well
Needs to water, can reach again preferably to increase bacterium and dephenolization effect.
Inventor studies discovery, the thinnest easy dehydration of fermentation material, and the container needed is the most, and cost is high, if fermentation
Expecting that the thickest meeting becomes anaerobic fermentation, effect is deteriorated, so the thickness of fermentation material is no more than 60mm, it is preferred that ferment during fermentation
The thickness of material is no more than 50mm.I.e. use thin layer solid-state aerobic fermentation, be conducive to improving Biomass.
Preferably, the container that aerobic fermentation uses is any one of stainless steel or bamboo.
It is further preferred that the preparation method of above-mentioned fermentation cottonseed protein, the container that the container used that ferments in step B is
For rustless steel fermentation dish or bamboo fermentation dish can be used.Such as, can use and fail to grow up in 2000mm, wide no more than 1000mm,
The high rustless steel not metal tray of 60mm that is not more than is as fermentation dish.
Generally speaking, relative to the biological method being mainly simple anaerobic fermentation of report in the past.The inventive method is
Based on aerobic fermentation mode, it is simultaneously introduced ferment treatment process, while improving gossypol clearance, effectively reduces anti-nutrition
The factor and raising protein digestibility, improve the Feeding Value of cottonseed meal, serve the effect of complex optimum, and its difficulty realized is non-
Chang great, but extraordinary comprehensive benefit can be obtained.The inventive method compares with traditional fermentation process and is shown in Table 1.
Table 1 the inventive method processes cottonseed meal with traditional fermentation process and compares
The present invention is different from other bioanalysis, for single cottonseed meal raw material, creatively uses high aerobic solid-state fermentation side
Formula, can fast decoupled toxicant and significantly improve Amino acid and protein content, the enzyme hydrolysis effect synchronizing to carry out then can be very
Effectively degrade antinutritional factor.
In following test example and embodiment, the saccharomycopsis fibuligera (Saccharomycopsis used
Fibuligera) strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms on March 17th, 2016
Center, deposit number is CGMCC No.12221.Preservation address: Yard 1, BeiChen xi Road, Chaoyang District, Beijing City 3 Chinese Academy of Sciences
Institute of microbiology.This bacterial strain all has stronger detoxification ability to gossypol in giucosinolate in rapeseed cake and cottonseed meal, and examination is oral
Safety non-toxic.
Candida utilis preserves from Microbiological Lab of Sichuan University, and primary source grinds in Chinese Academy of Sciences's microorganism
Study carefully institute (Beijing), be preserved in China General Microbiological culture presevation administrative center (CGNCC), bacterium numbering CGNCC2.281；
Yeast Kluyveromyces marxianus is preserved in Chinese microorganism strain and preserves administration committee's common micro-organisms center, preservation
Numbering CGMCCNo.9427；
Phytase is purchased from Beijing Smistyle Science & Technology Development Co., Ltd., cellulase, pectase, xylanase and Fructus Chaenomelis egg
White enzyme is purchased from ROTA Bioengineering Co., Ltd..
Embodiment 1 cottonseed meal screening pretreatment condition
Being pulverized by cottonseed meal raw material industry pulverizer, pulverizer sieve aperture is 2.0mm.Cottonseed meal after pulverizing are respectively with 10
The screen cloth sub-sieve of mesh, 20 mesh, 30 mesh, 40 mesh, 60 mesh and 80 mesh, claims sieve upper and the weight of siftage respectively.Wherein on 10 mesh sieves
Thing is entirely down, accounts for raw material gross weight ratio about 2%；20 mesh oversizes are down and a small amount of cotton seed hull impurity, account for raw material
Gross weight ratio about 8%, siftage is visible by naked eyes obvious down；From 30 mesh to 80 mesh sub-sieve results, Cottonseed in oversize
The thinnest and increase with sieve aperture.More than Zong He, choose the pretreatment before 20～30 mesh sieves are allocated as fermentation, reduce fibrous matter etc. miscellaneous
Matter content.Preferably 20 mesh, analysis result: before screening, raw material crude protein is 44%, and free gossypol is 852mg/kg；Thick egg after screening
Bai Weiwei 46.2%, free gossypol is 833mg/kg.
The preparation of embodiment 2 solids manufacture strain
Picking 1 ring from the slant preservation strain of saccharomycopsis fibuligera (Saccharomycopsis fibuligera)
(inoculating loop), accesses in 100mL Rhizoma Solani tuber osi culture fluid, in 30 DEG C of temperature shake-flask culture (180rpm) 52h, obtains detaining capsule laminating adhesive spore ferment
Female culture fluid.
Take saccharomycopsis fibuligera liquid 50g, access in sterilized 5000g wheat bran corn culture medium (in terms of siccative), mix
Close uniformly, cultivate 48h in 30 DEG C of temperature, obtain saccharomycopsis fibuligera solid spawn.
Prepared by Rhizoma Solani tuber osi sucrose culture fluid: be cut into the bulk referring to greatly nurse size after being peeled by fresh Rhizoma Solani tuber osi, weigh 100g, adds
The tap water of 500g, boils 20 minutes, filtered through gauze, and waste takes supernatant, with tap water constant volume to 500g, adds sucrose
10g, after dissolving, subpackage 250ml triangular flask, every bottle of 100g.Sterilizing 120 DEG C maintains 30 minutes.
The preparation of wheat bran corn culture medium: taking wheat bran 750g, Semen Maydis 250g, tap water 800g, after mix is uniform, in 120
DEG C sterilizing 30min, can inoculate after cooling.
Respectively from Candida utilis (Candida utilis) and yeast Kluyveromyces marxianus bacterium
Picking 1 ring (inoculating loop) in the slant preservation strain of (Kluyveromyces marxianus), is respectively connected to 100g Rhizoma Solani tuber osi sucrose
In culture fluid, cultivate 50h in 30 DEG C of temperature, respectively obtain Candida utilis culture fluid and the training of yeast Kluyveromyces marxianus bacterium
Nutrient solution；Candida utilis culture fluid 0.85g yeast Kluyveromyces marxianus bacteria culture fluid 0.15g mixing is accessed 100g soil
In bean sucrose culture fluid, cultivate 48h in 30 DEG C of temperature, i.e. obtain compound barm liquid；
Taking saccharomycopsis fibuligera solid spawn 8g, compound barm liquid 4g again, it is beautiful that mixing accesses sterilized 1000g wheat bran
In rice culture medium (in terms of siccative), mix is uniform, cultivates 52h in 30 DEG C of temperature, obtains mixed yeast solid spawn, for sending out
Ferment produces.Cell counting is 6.45 × 109/ g culture.
The addition of the embodiment 3 nutritive salt impact on result
Cottonseed meal raw material is pulverized by embodiment 1, divides with 20 mesh sieves, obtain cottonseed meal powder.Compound barm is prepared again by embodiment 2
Bacterium solid spawn.
By KH2PO4、KCl、MgSO4、FeSO4、ZnSO4Adding with the packet of phytase according to the form below, mix is uniform, in 30 DEG C of temperature
Degree cultivates 48h, measures the cell number after fermentation and crude protein content.
| 1 | 2 | 3 | 4 | 5 | 6 | 7 | 8 | |
| Cottonseed meal powder (g) | 1000 | 1000 | 1000 | 1000 | 1000 | 1000 | 1000 | 1000 |
| Water (g) | 700 | 700 | 700 | 700 | 700 | 700 | 700 | 700 |
| Strain (g) | 5 | 5 | 5 | 5 | 5 | 5 | 5 | 5 |
| KH<sub>2</sub>PO<sub>4</sub>(g) | —— | 1 | —— | 1 | 1 | 1 | —— | —— |
| KCl(g) | —— | —— | 1 | —— | —— | —— | 1 | 1 |
| MgSO<sub>4</sub>(g) | —— | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 | 0.5 |
| FeSO<sub>4</sub>(g) | —— | —— | —— | 0.05 | —— | 0.05 | 0.05 | 0.05 |
| ZnSO<sub>4</sub>(g) | —— | —— | —— | —— | 0.025 | 0.025 | 0.025 | 0.025 |
| Phytase (g) | —— | —— | —— | —— | —— | —— | —— | 0.2 |
| Cell number (× 10<sup>9</sup>/g) | 1.45 | 1.75 | 1.56 | 1.73 | 1.76 | 1.81 | 1.55 | 1.83 |
| Crude protein (%) | 48.20 | 49.35 | 48.82 | 49.84 | 49.86 | 50.20 | 48.84 | 50.61 |
Choose nutritive salt and be added to the 8th group, it may be assumed that KCl 0.1%, MgSO40.05%, FeSO40.005%,
ZnSO40.0025%, phytase 0.02%.It can be that cell growth provides phosphorus source that phytic acid in cottonseed meal is decomposed by phytase.More than plant
Acid enzyme activity unit is 5000u/g.
Embodiment 4 fermentation process cottonseed meal
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 833mg/kg.Press again
Embodiment 2 prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 100g, MgSO4 50g、FeSO45g and ZnSO42.5g pulverizer mixes
Pulverize, whole mistake 100 mesh.
Preparation mixing enzyme preparation: weigh phytase 100g, cellulase 20g, xylanase 10g, pectase 10g, Fructus Chaenomelis
Protease 10g, mixing and stirring.
Take cottonseed meal powder 980g, Semen Maydis powder 5g, mixed yeast solid spawn 12g, mixotrophism salt 2.5g, mixing enzyme preparation
0.5g and 800g water mix mixes, and in 700 × 350mm stainless steel disc, tiling is uniformly, and thickness 4cm ferments at a temperature of 30 DEG C
48h, after being dried 24h, i.e. obtains the cottonseed protein that ferments at a temperature of 50 DEG C.
Measuring the cell number after fermentation is 2.2 × 109/ g, moisture 12.2%, crude protein 51.7%, free gossypol 276mg/
kg。
Aspartic acid 4.41%, threonine 1.60%, serine 2.06%, glutamic acid 9.22%, glycine 1.97%,
Alanine 2.01%, cystine 0.56%, valine 2.04%, methionine 0.75%, isoleucine 1.53%, leucine
3.05%, tyrosine 1.33%, phenylalanine 2.72%, lysine 2.07%, histidine 1.33%, arginine 5.16%, dried meat
Propylhomoserin 2.03%, total amino acid content 43.83%.Little peptide content 10.3% (comparison raw material 4.4%, moisture 8.08%).
Embodiment 5 fermentation process cottonseed meal
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 833mg/kg.Press again
Embodiment 2 prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 100g, MgSO4 50g、FeSO45g and ZnSO42.5g pulverizer mixes
Pulverize, whole mistake 100 mesh.
Take cottonseed meal powder 974g, Semen Maydis powder 15g, mixed yeast solid spawn 6.7g, mixotrophism salt 4g, mixing enzyme preparation
0.3g and 700g water mix mixes, and in 700 × 350 stainless steel discs, tiling is uniformly, and thickness 6cm ferments at a temperature of 32 DEG C
48h, after being dried 20h, i.e. obtains the cottonseed protein that ferments at a temperature of 50 DEG C.
Measuring the cell number after fermentation is 1.65 × 109/g.Moisture 9.8%, crude protein 51.7%, lactic acid content
1.62%, free gossypol 329mg/kg.The molten protein content of acid 10.01% (comparison raw material is 4.94%), stachyose ＜ 50.0mg/
Kg, cottonseed sugar ＜ 50.0mg/kg.
Embodiment 6 fermentation process cottonseed meal
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 833mg/kg.Press again
Embodiment 2 prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 100g, MgSO4 50g、FeSO45g and ZnSO42.5g pulverizer mixes
Pulverize, whole mistake 100 mesh.
Preparation mixing enzyme preparation: weigh phytase 100g, cellulase 20g, xylanase 10g, pectase 10g, Fructus Chaenomelis
Protease 10g, mixing and stirring.
Take cottonseed meal powder 980g, Semen Maydis powder 3g, mixed yeast solid spawn 15g, mixotrophism salt 1g, mixing enzyme preparation 1g
Mixing with 600g water mix, in 700 × 350mm stainless steel disc, tiling is uniformly, thickness 6cm, and ferment at a temperature of 32 DEG C 50h,
After being dried 20h at a temperature of 50 DEG C, i.e. obtain the cottonseed protein that ferments.
Measuring the cell number after fermentation is 1.8 × 109/ g, free gossypol 318mg/kg, moisture 11.12%, crude protein
50.11%, lactic acid content 1.49%, sour molten protein content 10.8% (non-fermentation raw material is 4.4%, moisture 8.08%), water-soluble
Protein content 12.3%, stachyose ＜ 50.0mg/kg, cottonseed sugar ＜ 50.0mg/kg.
Embodiment 7 scale fermentation process cottonseed meal
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 833mg/kg.Press again
Embodiment 2 prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 2000g, MgSO4 1000g、FeSO4100g and ZnSO450g pulverizer
Co-grinding, whole mistake 100 mesh.
Preparation mixing enzyme preparation: by the every 1000g of phytase join cellulase 200g, xylanase 100g, pectase 100g,
The proportioning of papain 100g weighs the raw material of requirement, mixing and stirring.
Take cottonseed meal powder 970kg, Semen Maydis powder 15kg, mixed yeast solid spawn 11kg, mixotrophism salt 3.2kg, mixing
Enzyme preparation 0.8kg and the mixing of 800kg water mix, in 2000 × 1000 × 60mm rustless steel fermentation dish (bottom is 40 eye mesh screens)
Uniformly, thickness 5cm, often dish charging 20kg (in terms of siccative), fills 50 dishes altogether, and ferment at a temperature of 33 DEG C 48h, 50 DEG C of temperature in tiling
Under be dried 20h after, i.e. obtain ferment cottonseed protein.
Measuring the cell number after fermentation is 2.05 × 109/ g, moisture 12.5%, crude protein 50.8%, free gossypol
322mg/kg。
Aspartic acid 4.52%, threonine 1.64%, serine 2.15%, glutamic acid 9.35%, glycine 1.99%,
Alanine 2.01%, cystine 0.68%, valine 2.06%, methionine 0.76%, isoleucine 1.51%, leucine
3.02%, tyrosine 1.38%, phenylalanine 2.76%, lysine 2.12%, histidine 1.38%, arginine 5.44%, dried meat
Propylhomoserin 1.95%, total amino acid content 44.76%.The molten protein 11 .5% of acid (comparison raw material 4.45%, moisture 8.9%), stachyose
＜ 50.0mg/kg, cottonseed sugar ＜ 50.0mg/kg.
Embodiment 8 scale fermentation process cottonseed meal
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 845mg/kg.Press again
Embodiment 2 method prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 10kg, MgSO4 5kg、FeSO40.5kg and ZnSO40.25kg pulverizes
Machine co-grinding, whole mistake 100 mesh.
Preparation mixing enzyme preparation: weigh phytase 10kg, cellulase 2kg, xylanase 1kg, pectase 1kg, Fructus Chaenomelis
Protease 1kg, mixing and stirring.
Take cottonseed meal powder 990kg, mixed yeast solid spawn 7kg, mixotrophism salt 2.2kg, mixing enzyme preparation 0.8kg and
900kg water mix mixes, and in 2000 × 1000 × 60mm rustless steel fermentation dish (bottom is 40 eye mesh screens), tiling is uniformly, thickness
4cm, often dish charging 20kg (in terms of siccative), fill 50 dishes, fermenting cellar temperature control 33 DEG C altogether, and ferment 48h, is dried 20h at a temperature of 50 DEG C
After, i.e. obtain the cottonseed protein that ferments.
Measuring the cell number after fermentation is 1.9 × 109/g.Free gossypol 325mg/kg, crude protein 52.89%, moisture
9.96%, ash 7.25%, aspartic acid 4.81%, lysine 2.29%, proline 1.84%, leucine 2.98%, phenylpropyl alcohol
Propylhomoserin 2.74%, glycine 2.24%, valine 2.14%, histidine 1.61%, serine 2.44%, isoleucine
1.53%, methionine 0.61%, alanine 2.08%, glutamic acid 9.35%, threonine 1.71%, arginine 5.62%, Guang ammonia
Acid 0.86%, free ammonia 1.43%.The molten protein 11 .5% of acid (comparison raw material 4.41%, moisture 8.5%), stachyose ＜
50.0mg/kg, cottonseed sugar ＜ 50.0mg/kg.
Embodiment 9 scale fermentation process cottonseed meal also carry out growing and fattening pigs culture experiment
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 845mg/kg.Press again
Embodiment 2 method prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 10kg, MgSO4 5kg、FeSO40.5kg and ZnSO40.25kg pulverizes
Machine co-grinding, whole mistake 100 mesh.
Preparation mixing enzyme preparation: weigh phytase 10kg, cellulase 2kg, xylanase 1kg, pectase 1kg, Fructus Chaenomelis
Protease 1kg, mixing and stirring.
Take cottonseed meal powder 4900kg, Semen Maydis powder 30kg, mixed yeast solid spawn 50kg, mixotrophism salt 15kg, mixing
Enzyme preparation 5kg and the mixing of 4000kg water mix, in 2000 × 1000 × 60mm rustless steel fermentation dish (bottom is 40 eye mesh screens)
Tiling is uniform, thickness 5cm, often dish charging 20kg (in terms of siccative), fills 250 dishes, fermenting cellar temperature 34.5 DEG C altogether, and ferment 46h, and 50
After being dried 20h at a temperature of DEG C, i.e. obtain the cottonseed protein that ferments.
Measuring the cell number after fermentation is 1.7 × 109/g.Free gossypol 322mg/kg.Crude protein 53.47%, moisture
10.12%, ash 6.89%, aspartic acid 4.83%, lysine 2.18%, proline 1.77%, leucine 2.95%, phenylpropyl alcohol
Propylhomoserin 2.99%, glycine 2.26%, valine 2.05%, histidine 1.64%, serine 2.45%, isoleucine
1.45%, methionine 0.60%, alanine 2.07%, glutamic acid 9.49%, threonine 1.69%, arginine 5.64%, Guang ammonia
Acid 0.89%, free ammonia 1.68%.The molten protein 11 .8% of acid (comparison raw material 4.47%, moisture 8.65%), stachyose ＜
50.0mg/kg, cottonseed sugar ＜ 50.0mg/kg.
Select Neijiang Pig and Duroc bi-crossbreeding 120, be randomly divided into 2 groups, often group 60, feed traditional raw material for 1 group
Complete feedstuff, another 1 group of biological feedstuff fed added with fermentation cottonseed protein.Conventional feed group average weight 65.4kg, biological feedstuff group
Average weight 64.85kg.
Conventional feed formula: Semen Maydis 69%, bean cake (crude protein 43%) 16%, cottonseed meal (crude protein 44%) 6%, wheat bran
5%, trace regulator 4%.Biological feedstuff formula: Semen Maydis 69%, bean cake 7%, fermentation cottonseed protein (crude protein 53.47%)
13%, wheat bran 7%, trace regulator 4%.
Biological feedstuff is that fermentation cottonseed protein 13% is replaced cottonseed meal and part bean cake in conventional feed group, keep two groups thick
Albumen is consistent, and metabolizable energy is suitable, and differential section is with 2% wheat bran polishing.
Two group of formula unit prices: conventional feed group is higher than biological feedstuff group 36 yuan/ton.
Feed 82 days under equal rearing conditions.Conventional feed group: daily gain 685g, feedstuff-meat ratio 3.572, money meat ratio
7.86；Biological feedstuff group: daily gain 736g, feedstuff-meat ratio 3.410, money meat ratio 7.38, cost reduces by 0.48 yuan/every kg body weight and increases
Weight.
4 parts of above-mentioned trace regulators include following composition: calcium hydrogen phosphate 10g, lysine 2g, copper Cu8.0mg, ferrum
Fe80.0mg, manganese Mn60.0mg, zinc Zn 40.0mg, selenium Se 0.15mg, iodine I 0.35mg, vitamin A 50000IU, vitamin
D310000IU, vitamin E 25IU, vitamin K3 5mg, vitamin B12 10mg, vitamin B12 0mg, vitamin B2 16mg, dimension
Raw element B6 6mg, nicotiamide 35mg, pantothenic acid 25mg, folic acid 0.5mg.IU iu in trace regulator.
Embodiment 10 scale fermentation process cottonseed meal also carry out growing broiler breeding experiment
Being pulverized by embodiment 1 by cottonseed meal raw material, divide with 20 mesh sieves, obtain cottonseed meal powder, free gossypol is 845mg/kg.Press again
Embodiment 2 method prepares saccharomycopsis fibuligera solid spawn and compound barm bacterium solution respectively.
Preparation mixotrophism salt: weigh KCl 10kg, MgSO4 5kg、FeSO40.5kg and ZnSO40.25kg pulverizes
Machine co-grinding, whole mistake 100 mesh.
Preparation mixing enzyme preparation: weigh phytase 10kg, cellulase 2kg, xylanase 1kg, pectase 1kg, Fructus Chaenomelis
Protease 1kg, mixing and stirring.
Take cottonseed meal powder 980kg, Semen Maydis powder 7kg, mixed yeast solid spawn 10kg, mixotrophism salt 2.2kg, mixed enzyme
Preparation 0.8kg and the mixing of 800kg water mix, in 2000 × 1000 rustless steel fermentation dishes (bottom is 40 eye mesh screens), tiling is all
Even, thickness 4cm, often dish charging 20kg (in terms of siccative), fill 50 dishes, fermenting cellar temperature control 34.8 DEG C altogether, ferment 45h, 50 DEG C of temperature
Under be dried 20h after, i.e. obtain ferment cottonseed protein.
Measuring cell number in fermentation cottonseed protein finished product is 2.15 × 109/g.Crude protein 49.4%, moisture 13.3%, trip
From gossypol 317mg/kg.
Aspartic acid 4.52%, threonine 1.64%, serine 2.17%, glutamic acid 9.77%, glycine 2.03%,
Alanine 1.98%, cystine 0.60%, valine 2.02%, methionine 0.76%, isoleucine 1.53%, leucine
3.04%, tyrosine 1.38%, phenylalanine 2.62%, lysine 2.19%, histidine 1.40%, arginine 5.58%, dried meat
Propylhomoserin 2.00%, total amino acid content 45.21%.Phytic acid degradation rate about 30%, pectin degrading rate 42%, acid-soluble protein 11.2%
(comparison raw material 4.43%, moisture 8.5%).
Nursing broiler chicken is tested
Select the blue or green foot Rhizoma Euonymus fiber crops plumage chicken of 600 about 1.6kg, be randomly divided into 2 groups, often group 300, the average initial weight of matched group
Only, average initial weight 1.618kg/ of biological group is only for 1.645kg/.
Matched group formula is: Semen Maydis 68%, bean cake (crude protein 43%) 18%, cottonseed meal (crude protein 44%) 6%, wheat bran
3%, Oleum Glycines 1%, trace regulator 4%；Biological feedstuff group of formula is: Semen Maydis 68%, bean cake (crude protein 46%) 10%, fermentation
Cottonseed protein (conversion of crude protein 51.3%, moisture presses 10%) 13%, wheat bran 4%, Oleum Glycines 1%, trace regulator 4%.Biological
Feedstuff group replaces 8% bean cake and 6% cottonseed meal with 13% fermentation cottonseed protein, and not enough 1% is supplied with wheat bran.Two groups of energy are with thick
Protein level is basically identical.Bioprotein group of formula cost unit price reduces by 30 yuan/ton of feedstuffs.
Feed 31 days under equal rearing conditions.Matched group: total augment weight 110.27kg, feedstuff-meat ratio is 2.72, feed price
2.2 yuan/kg money meat than 5.98:1, biological feedstuff group: total augment weight 120.08kg, feedstuff-meat ratio is 2.59, feed price 2.17 yuan/
Kg, money meat increase weight than for 5.62:1, i.e. per kilogram and reduce cost 0.36 yuan.Biological group dies of illness several 0, and matched group dies of illness several 1.
Biological group feedstuff group does not adds any antibiotic.
Above-mentioned trace regulator is by every kg diet: calcium hydrogen phosphate 10g, choline 1g, sodium bicarbonate 1g, lysine 2g, egg ammonia
Acid 1g, phytase 0.2g, copper Cu8.0mg, ferrum Fe80.0mg, manganese Mn60.0mg, zinc Zn 40.0mg, selenium Se 0.15mg, iodine I
0.35mg, vitamin A 50000IU, vitamin D310000IU, vitamin E 25IU, vitamin K3 5mg, vitamin B1210mg、
Vitamin B120mg, vitamin B216mg, vitamin B66mg, nicotiamide 35mg, pantothenic acid 25mg, folic acid 0.5mg.Matched group is raised
In material trace regulator every kilogram possibly together with virginiamycin 20mg, Salinomycin 60mg.In biological group feedstuff trace regulator
Every kilogram is supplemented more: lysine 0.4g.IU iu in trace regulator.
Claims (8)
1. fermentation cottonseed protein, it is characterised in that: formed by the fermenting raw materials of following weight proportion: cottonseed meal powder 97～99 parts, jade
Rice flour 0～1.5 parts, mixed yeast solid spawn 0.7～1.5 parts, mixotrophism salt 0.1～0.4 part, mixing enzyme preparation 0.03
～0.1 part, water；The gross weight of water accounts for 0.6～0.9 times of other five kinds of raw material gross weights.
Fermentation cottonseed protein the most according to claim 1, it is characterised in that: by the fermenting raw materials of following weight proportion
Become: 98 parts of cottonseed meal powder, Semen Maydis powder 0.5 part, mixed yeast solid spawn 1.2 parts, 0.25 part of nutritive salt, mixing enzyme preparation 0.05
Part and 80 parts of water.
Fermentation cottonseed protein the most according to claim 1 and 2, it is characterised in that: described cottonseed meal powder is that Semen Gossypii is through oil expression
After the dregs of rice, then through pulverizing and screening remove impurity.
4. according to the fermentation cottonseed protein described in any one of claims 1 to 3, it is characterised in that: described mixed yeast is solid
The strain that body strain includes is: saccharomycopsis fibuligera (Saccharomycopsis fibuligera), Candida utilis
(Candida utilis) and yeast Kluyveromyces marxianus bacterium (Kluyveromyces marxianus).
5. according to the fermentation cottonseed protein described in any one of Claims 1 to 4, it is characterised in that: the preparation of described nutritive salt
Method is: count by weight ratio, takes 1 part of KCl, 0.5 part of MgSO4, 0.05 part of FeSO4With 0.025 part of ZnSO4Mix.
6. according to the fermentation cottonseed protein described in any one of Claims 1 to 5, it is characterised in that: count phytase by weight ratio
Cellulase xylanase pectase papain=10 2111, mix five；Wherein, described phytic acid
The enzyme activity of enzyme is 5000 units, and the enzyme activity of described cellulase is 5000 units, and the enzyme activity of described xylanase is 10
Ten thousand units, the enzyme activity of described pectase is 30,000 units, and papain enzyme activity is 800,000 units.
7. the solid-state aerobic fermentation method of the fermentation cottonseed protein described in preparation any one of claim 1～8, it is characterised in that:
Comprise the following steps:
A, pulverize and sieve: the pulverizer taking cottonseed meal 2.0 sieves is pulverized, then with the cotton after the sieve dressing sieve in 20～30 mesh apertures
Dregs of rice powder；
B, mixing: take cottonseed meal powder, Semen Maydis powder, mixed yeast solid spawn, nutritive salt, mixing enzyme preparation and water and mix to obtain fermentation
Material；
C, fermentation: by fermentation material under good oxygen condition in 30～35 DEG C of bottom fermentations 45～50h, the most i.e. obtain ferment Semen Gossypii egg
In vain.
The preparation method of fermentation cottonseed protein the most according to claim 8, it is characterised in that: in step B, good oxygen condition is sent out
During ferment, the thickness of fermentation material is no more than 60mm.
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