CN103343118A - Biological selenium product applied to organic selenium-rich agriculture and preparation method thereof - Google Patents
Biological selenium product applied to organic selenium-rich agriculture and preparation method thereof Download PDFInfo
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- CN103343118A CN103343118A CN2013102700507A CN201310270050A CN103343118A CN 103343118 A CN103343118 A CN 103343118A CN 2013102700507 A CN2013102700507 A CN 2013102700507A CN 201310270050 A CN201310270050 A CN 201310270050A CN 103343118 A CN103343118 A CN 103343118A
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Abstract
The invention belongs to the biotechnology field, and relates to a biological selenium product applied to organic selenium-rich agriculture and a preparation method thereof. According to the method, by-products, such as molasses and bean pulp, from processing of agriculture products are taken as a fermentation substrate; an inorganic selenium compound is added into the fermentation substrate and is transformed into biological selenium though the fermentation by transformed selenium produced muscle candida efficient strains cultivated through induced mutation; the non-poisonous biological selenium product, which is easy to be absorbed by plants, is prepared by enzymolysis of selenium macromolecules. All the selenium derives from the process of biosynthesis and transformation. When the product is applied to the production of selenium-rich organic agricultural products, not only is the safety of the selenium-rich products guaranteed, but also the risk of environment pollution caused by utilizing the inorganic selenium compound in production is effectively avoided.
Description
Technical field
The invention belongs to biological technical field, relate to adding the agricultural-food of inorganic selenium
By product is matrix, has the high Candida utilis fermentation that transforms the selenium ability by screening and transforms inorganic selenium, prepares biological selenium product and the production technology of easily being utilized by plant absorbing thereof through enzymolysis.
Background technology
Selenium is the essential trace element of humans and animals, and the existing humans and animals that studies show that has hundreds of disease to be associated with the selenium intake deficiency, and endemy one Keshan disease that occurs in my north is exactly to lack due to the selenium.Food is the main source that humans and animals is taken in selenium.Because selenium is in the earth's crust
Spend low and skewness, the whole world lacks the selenium area in various degree and accounts for 2/3.Up to now, increase the approach that selenium intake improves selemium nutrition, the one, in food or the healthcare products course of processing, add inorganic selenium and (use Na usually
2SeO) this method is simple and easy to do, with low cost, but inorganic selenium toxicity is big, bioavailability is low; The 2nd, use yeast rich in selenium, nontoxic but tough and tensile cell walls difficulty is arranged by the utilization of humans and animals digestion influence to selenium; The 3rd, use at crop growth period and to contain selenium fertilizer, utilize the assimilation of plant to produce selenium-enriched crop, obtain to contain the food of nontoxic, high bioavailability biological selenium thus.This is to mend the most scientific and effective approach of selenium, also is current extensive employing method and the generally accepted measure of human consumer.But without exception the selenium source of use all is inorganic selenium in rich selenium agriculture production, and this just makes and also do not meet modern Organic farming production requirement by human consumer rich selenium Product Safety is left a question open, also has the risk of contaminate environment.So far for this reason, also there is not a kind of non-mineral compound selenium source product that rich selenium agriculture production is used that is applied to.
Summary of the invention
The present invention is directed to the prior art existing problems, use composite factor mutagenic treatment Candida utilis, carry out anti-selenium, selenium rich ability screening with high seleno culture medium; Be that the substratum of carbon, nitrogenous source carries out the adaptability screening in order to the processing of farm products by product, obtain the biomass height, strong and can be the efficient bacterial strain of substrate fermentation with the processing of farm products by product to the selenium conversion capability, by fermentation the inorganic selenium that adds is converted into biological selenium in matrix.Fermented substrate is carbon, nitrogenous source with the processing of farm products by product, reduced production cost, adopt the initial selenium concentration of reduction in fermented substrate according to the strain growth metabolic rule, and gradation adds the selenium novel process during the fermentation, both ensured the supply of selenium, avoided the disposable too high inhibition phenomenon that bacterial strain is produced of fermentation initial stage selenium concentration that a large amount of selenium cause that adds again, kept bacterial strain from start to finish and breed at a high speed and the high selenium vigor that transforms.The remaining matrix of centrifugal removal after the fermentation ends, through high-pressure homogeneous smudge cells, enzymolysis contains selenium people molecule and obtains the nontoxic good biological selenium product that is easily directly absorbed by plant, and contained selenium all derives from biosynthesizing and transforms.Product application can ensure the rich selenium security of products of producing in the production of Organic farming Se-rich farm products, can effectively avoid the contaminate environment risk of using inorganic selenium to exist again.
Realization flow of the present invention is: the bacterium → mutagenesis of setting out → conversion selenium ability screening → culture medium adaptability is screened → is transformed the efficient bacterial strain of selenium → preparation fermentation seed liquid → seed liquor enlarged culturing → fermentation and transforms the remaining fermentation of selenium → removal
→ broken somatocyte → enzymolysis contains selenium macromole → separation removal of impurities → concentrated or drying → finished product packing.Implementation step is as follows:
The bacterial strain complex mutation: the Candida utilis thalline was handled 2～15 minutes with 50～100Lx UV-irradiation 10～120 seconds and 0.2～2.0% ethyl sulfate in the darkroom, carried out composite factor mutagenesis.
The screening of selenium conversion capability: the half plate substratum of preparation selenium concentration 100～300mg/L, coat the resistance screening culture medium flat plate after will diluting with stroke-physiological saline solution through the thalline that complex mutation is handled and carry out anti-selenium and the screening of selenium conversion capability.
Matrix adaptability screening: it is to cultivate on the substratum of carbon, nitrogenous source to carry out the adaptability screening that the thalline that resistant panel filters out is inoculated in the processing of farm products by product.Filtering out with the molasses dregs of beans is carbon, nitrogenous source, biomass height, the efficient bacterial strain of the Candida utilis that the selenium conversion capability is strong, and its selenium content reaches 800～2000mg/Kg.
Liquid strain preparation: the efficient conversion selenium bacterial strain activation back of depositing of going bail for is inoculated in dress by 5～15% inoculum size and contains the triangular flask that selenite concentration is the molasses dregs of beans substratum of 20～300mg Se/L, and 26～32 ℃ of shaking tables were cultivated 24～48 hours.
The seed liquor enlarged culturing: the liquid bacterial classification that shake-flask culture is gone out inserts to be equipped with by 6～6% inoculum size and contains the seeding tank that selenite concentration is the molasses dregs of beans substratum of 20～300mg Se/L, 26～32 ℃ of fermentation culture 24～48 hours
Fermentation transforms selenium: the seed liquor that enlarged culturing is gone out inserts to be equipped with by 8～20% inoculum size and contains the fermentor tank that selenite concentration is the molasses dregs of beans substratum of 20～300mgSe/L, 26～32 ℃ of fermentations 48 hours, its interbody spacer 12～24 hours are added fermentating liquid volume 1～10 ‰ in the fermentor tank, concentration is 500～1000mg Se/L selenite solution.
Remaining fermented substrate is removed: fermentation ends secondary fermentation thing separates by horizontal spiral centrifuge removes remaining fermented substrate, changes subsequent processing over to.
Somatic cells is broken to be discharged: the fermented product of removing after the remaining matrix carries out the somatic cells Mechanical Crushing through high pressure homogenizer, is conveyed into hydrolytic decomposition pot then.
Degraded contains the selenium macromole: the compound protease liquid that adds 0.1～5.0 ‰ volume in the hydrolytic decomposition pot that broken somatic cells fermented product is housed, stirring at low speed, 30～55 ℃ of insulations 12～24 hours, the biological selenium that matrix inorganic selenium fermentation is transformed generate is by can not being the small molecules that plant easily absorbs by the macromolecules degradation of plant absorbing.
Separate removal of impurities: enzymolysis is finished by the butterfly centrifugal machine and is separated, and removes the impurity that cell wall fragment, starch etc. can not be utilized by plant absorbing.
Concentrate with dry: the hydrolyzed solution behind the removal impurity is sent into the multiple-effect thickener and is concentrated, with bucket, bottle packing; Be transported to perhaps that spray-drying tower spray is dried to become powder, packed.
The present invention has the following advantages compared with the prior art:
1. product is biogenetic derivation, safety non-toxic, easily absorbs, and meets green agriculture, Organic farming code requirement, has overcome thoroughly that the toxicity of using inorganic selenium to exist up to now is big, the risk of contaminate environment etc.
2. fermentative production is made carbon, the nitrogenous source of fermented substrate with the processing of farm products by product, has not only reduced production cost but also widened the application market of processing of farm products by product.
3. use during the fermentation repeatedly to add selenium technology, overcome effectively because of 1 property adding high density selenium in the matrix and improved the thalline biomass and transformed the selenium amount in the early stage drawback that produces retarding effect of fermentation.
Embodiment
Embodiment 1:
Transform the efficient bacterial strain screening process of selenium Candida utilis: the bacterium activation culture of setting out → ultraviolet mutagenesis processing → chemomorphosis processing → resistant panel primary dcreening operation → shake-flask culture sieves → the high selenium ability bacterial strain that transforms again.Implementation step is as follows:
1) gets 24 hours Candida utilis of shake-flask culture cultivation thalline and dilute with stroke-physiological saline solution, in 6 culture dish that are placed in, shone respectively 10～120 seconds with the 50Lx UV-light in the darkroom.
2) thalline after the uviolizing added in 6 test tubes that 0.2% ethyl sulfate solution is housed oscillation treatment respectively 15 minutes, and 2% hypo solution termination reaction is centrifugal, the stroke-physiological saline solution washing.
3) plate culture medium of preparation glucose 3%, peptone 1%, yeast extract paste 0.1%, agar 2%, selenium concentration 100～300mg/L, coat the resistance culture medium flat plate after will diluting with stroke-physiological saline solution through the thalline that complex mutation is handled, cultivated 48 hours about 28 ℃, select eugonic light bacterium colony.
4) preparation contains 6 groups of the high seleno culture mediums of sugared close dregs of beans of industry sodium selenate of sugar 40% molasses 4～25%, dregs of beans 2～15%, 50～300mgSe/L in triangular flask respectively, inoculating 24～36 ℃ of shaking tables of the selected bacterium colony of primary dcreening operation respectively cultivated 48 hours, carrying out with the processing of farm products by product is the fermention medium adaptability screening of carbon, nitrogenous source, and obtaining with the processing of farm products by product is the efficient bacterial strain of Candida utilis that the matrix growth and breeding is fast, the selenium conversion capability is strong.
Be used for the biogenic selenium Production Flow Chart of organic Se-rich agricultural: transform the efficient bacterial strain seed liquor of selenium preparation → seed liquor enlarged culturing → substrate fermentation conversion selenium → remaining matrix removal → somatic cells fragmentation → enzymolysis and contain selenium macromole → separation removal of impurities → concentrated → can.Concrete technology is as follows:
1) preparation seed liquor: add the molasses 4% that contain sugar 40% in the triangular flask, dregs of beans 2%, sal epsom 0.05 ‰, potassium primary phosphate 0.5%, Sodium Selenite 20mgSe/L, sterilization.Inoculum size by 5% after the efficient bacterial strain activation of the conversion selenium of preserving is inoculated in triangular flask, and 26 ℃ of shaking tables were cultivated 48 hours.
2) seed liquor enlarged culturing: add the molasses 10% that contain sugar 40% in the seeding tank, dregs of beans 5%, sal epsom 0.1 ‰, potassium primary phosphate 1.0%, Sodium Selenite 20mgSe/L, sterilization.Inoculum size by 16% inserts seed liquor, and 32 ℃ of shaking tables were cultivated 24 hours.
3) fermentation transforms selenium: add the molasses 8% that contain sugar 40% in the fermentor tank, dregs of beans 4%, sal epsom 0.05 ‰, potassium primary phosphate 0.5%, Sodium Selenite 20mgSe/L, sterilization.Insert the seed liquor that spreads cultivation by 8% inoculum size, 26 ℃ are stirred fermentation 48 hours, do fermentation added 1 ‰ volumes respectively in the jar to 24 and 36 hours 1000mgSe/L sodium selenite solution.
4) remove remaining fermented substrate: pump into the horizontal spiral centrifuge centrifugation after fermentation is finished and remove the fermented substrate residue.
5) high-pressure homogeneous smudge cells: the fermented product of removing behind the remaining fermented substrate is transported to the high pressure homogenizer smudge cells, discharges by the synthetic selenium-containing biological molecule that enters in the born of the same parents that transforms that ferments.
6) enzymolysis contains the selenium macromole: the smudge cells fermented product is transported to hydrolytic decomposition pot, adds the compound protease liquid of 0.1 ‰ volumes, stirring at low speed, and 30 ℃ are incubated 24 hours, make the middle inorganic selenium fermentation of matrix transform the biological selenium that generates
Can not be the small molecules that plant easily absorbs by the macromolecules degradation of plant absorbing.
7) separate removal of impurities: the enzymolysis solution after degraded is finished separates through the butterfly centrifugal machine to be removed
The impurity that fragment, starch etc. can not be utilized by plant absorbing.
8) concentrate can: the hydrolyzed solution behind the removal impurity is sent into the economic benefits and social benefits thickener and is concentrated, and uses plastic tank or the Plastic Bottle packing of good airproof performance as required.
Embodiment 2:
Transform the efficient bacterial strain screening process of selenium Candida utilis: the bacterium activation culture of setting out → chemomorphosis processing → ultraviolet mutagenesis processing → resistant panel primary dcreening operation → shake-flask culture sieves → the high selenium ability bacterial strain that transforms again.Implementation step is as follows:
1) get 24 hours Candida utilis of shake-flask culture and cultivate thalline and added in 6 test tubes that 2.0% ethyl sulfate solution is housed oscillation treatment respectively 5 minutes, 2% sulfo-is dredged the acid sodium solution termination reaction, and is centrifugal, and stroke-physiological saline solution is washed.
2) thalline after ethyl sulfate is handled is changed in the culture dish, the stroke-physiological saline solution dilution was shone respectively 10～120 seconds with the 50Lx UV-light in the darkroom.
3) plate culture medium of preparation glucose 3%, peptone 1%, yeast extract paste 0.1%, agar 2%, selenium concentration 100～300mg/L, coat the resistance culture medium flat plate after will diluting with stroke-physiological saline solution through the thalline that complex mutation is handled, cultivated 48 hours about 28 ℃, select eugonic light bacterium colony.
4) preparation contains 6 groups of the high seleno culture mediums of molasses dregs of beans of Sodium Selenite of sugar 40% molasses 4～25%, dregs of beans 2～15%, 50～300mgSe/L in triangular flask respectively, inoculating 24～36 ℃ of shaking tables of the selected bacterium colony of primary dcreening operation respectively cultivated 48 hours, carrying out with the processing of farm products by product is the fermention medium adaptability screening of carbon, nitrogenous source, and obtaining with the processing of farm products by product is the efficient bacterial strain of Candida utilis that the matrix growth and breeding is fast, the selenium conversion capability is strong.
Be used for the biogenic selenium Production Flow Chart of organic Se-rich agricultural: transform the efficient bacterial strain seed liquor of selenium cultivation → seed liquor enlarged culturing → substrate fermentation and transform selenium → remaining matrix removal → somatic cells fragmentation → enzymolysis and contain selenium macromole → separation removal of impurities → spraying drying → pack.Concrete technology is as follows:
1) preparation seed liquor: add the molasses 20% that contain sugar 40% in the triangular flask, dregs of beans 10%, sal epsom 0.2 ‰, potassium primary phosphate 2.0%, Sodium Selenite 300mgSe/L, sterilization.Inoculum size by 15% after the efficient bacterial strain activation of the conversion selenium of preserving is inoculated in triangular flask, and 32 ℃ of shaking tables were cultivated 24 hours.
2) seed liquor enlarged culturing: add the molasses 10% that contain sugar 40% in the seeding tank, dregs of beans 5%, sal epsom 0.1 ‰, potassium primary phosphate 1.0%, Sodium Selenite 20mgSe/L, sterilization.Inoculum size by 6% inserts seed liquor, and 32 ℃ of shaking tables were cultivated 24 hours.
3) fermentation transforms selenium: add the molasses 20% that contain sugar 40%, dregs of beans in the fermentor tank
Sal epsom 0.5 ‰, potassium primary phosphate 2.0%, Sodium Selenite 300mgSe/L, sterilization.Insert the seed liquor that spreads cultivation by 20% inoculum size, 32 ℃ are stirred fermentation 48 hours, add the 500mgSe/L sodium selenite solution of 2 ‰ volumes respectively in the jar to 24 and 36 hours in fermentation.
4) remove remaining fermented substrate: pump into the horizontal spiral centrifuge centrifugation after fermentation is finished and remove the fermented substrate residue.
5) high-pressure homogeneous smudge cells: the fermented product of removing behind the remaining fermented substrate is transported to the high pressure homogenizer smudge cells, discharges by the synthetic selenium-containing biological molecule that enters in the born of the same parents that transforms that ferments.
6) enzymolysis contains the selenium macromole: the smudge cells fermented product is transported to hydrolytic decomposition pot, adds the compound protease liquid of 50 ‰ volumes.Stirring at low speed, 55 ℃ of insulations 12 hours, the biological selenium that inorganic selenium fermentation on the matrix is transformed generate is by can not being the water molecules that plant easily absorbs by the macromolecules degradation of plant absorbing.
7) separate removal of impurities: the enzymolysis solution after degraded is finished separates through the butterfly centrifugal machine removes the impurity that cell wall fragment, starch etc. can not be utilized by plant absorbing.
8) dry packing: the hydrolyzed solution behind the removal impurity is conveyed into spray-drying tower, and 150 170 ℃ of instantaneous powder that are dried to of spraying are down packed with compound membrane bag.
Embodiment 3:
Transform the efficient bacterial strain screening process of selenium Candida utilis: the bacterium activation culture of setting out → ultraviolet mutagenesis processing → chemomorphosis processing → resistant panel primary dcreening operation → shake-flask culture sieves → the high selenium ability bacterial strain that transforms again.Implementation step is as follows:
1) gets 24 hours Candida utilis of shake-flask culture cultivation thalline and dilute with stroke-physiological saline solution, in 6 culture dish that are placed in, shone respectively 10～120 seconds with the 100Lx UV-light in the darkroom.
2) thalline after the uviolizing added in 6 test tubes that 0.2% ethyl sulfate solution is housed oscillation treatment respectively 15 minutes, and 2% hypo solution termination reaction is centrifugal, the stroke-physiological saline solution washing.
3) plate culture medium of preparation glucose 3%, peptone 1%, yeast extract paste 0.1%, agar 2%, selenium concentration 100～300mg/L, to after thalline that complex mutation is handled is with the stroke-physiological saline solution dilution, coat the resistance culture medium flat plate, cultivate about 28 ℃ 48 hours, select eugonic light bacterium colony.
4) preparation contains 6 groups of the high seleno culture mediums of molasses dregs of beans of Sodium Selenite of sugar 40% molasses 4～25%, dregs of beans 2～15%, 50～300mgSe/L in triangular flask respectively, inoculating 24～36 ℃ of shaking tables of the selected bacterium colony of primary dcreening operation respectively cultivated 48 hours, carrying out with the processing of farm products by product is the fermention medium adaptability screening of carbon, nitrogenous source, and obtaining with the processing of farm products by product is the efficient bacterial strain of Candida utilis that the matrix growth and breeding is fast, the selenium conversion capability is strong.
Be used for the biogenic selenium Production Flow Chart of organic Se-rich agricultural: transform the efficient bacterial strain seed liquor of selenium cultivation → seed liquor enlarged culturing → substrate fermentation conversion selenium → remaining matrix removal → somatic cells fragmentation → enzymolysis and contain selenium macromole → separation removal of impurities → concentrated → can.Concrete technology is as follows:
1) preparation seed liquor: add the molasses 20% that contain sugar 40% in the triangular flask, dregs of beans 10%, sal epsom 0.5 ‰, potassium primary phosphate 2.0%, Sodium Selenite 300mgSe/L, sterilization.Inoculum size by 15% after the efficient bacterial strain activation of the conversion selenium of preserving is inoculated in triangular flask, and 26 ℃ of shaking tables were cultivated 48 hours.
2) seed liquor enlarged culturing: add the molasses 10% that contain sugar 40% in the seeding tank, dregs of beans 5%, sal epsom 0.1 ‰, potassium primary phosphate 1.0%, Sodium Selenite 20mgSe/L, sterilization.Inoculum size by 6% inserts seed liquor, and 32 ℃ Oscillating beds were cultivated 24 hours.
3) fermentation transforms selenium: add the molasses 8% that contain sugar 40% in the fermentor tank, dregs of beans 4%, sal epsom 0.05 ‰, potassium primary phosphate 0.5%, Sodium Selenite 20mgSe/L, sterilization.Insert the seed liquor that spreads cultivation by 8% inoculum size, 26 ℃ are stirred fermentation 48 hours, add the 1000mgSe/L sodium selenite solution of 1 ‰ volumes respectively in the jar to 24 and 36 hours in fermentation.
4) remove remaining fermented substrate: pump into the horizontal spiral centrifuge centrifugation after fermentation is finished and remove the fermented substrate residue.
5) high-pressure homogeneous smudge cells: the fermented product of removing behind the remaining fermented substrate is transported to the high pressure homogenizer smudge cells, discharges by the synthetic selenium-containing biological molecule that enters in the born of the same parents that transforms that ferments.
6) enzymolysis contains the selenium macromole: the smudge cells fermented product is transported to hydrolytic decomposition pot, the compound protease liquid that adds 50 ‰ volumes, stirring at low speed, 30 ℃ of insulations 24 hours, the biological selenium that inorganic selenium fermentation in the matrix is transformed generate is by can not being the small molecules that plant easily absorbs by the macromolecules degradation of plant absorbing.
7) separate removal of impurities: the enzymolysis solution after degraded is finished separates through the butterfly centrifugal machine removes the impurity that cell wall fragment, starch etc. can not be utilized by plant absorbing.
8) concentrate can: the hydrolyzed solution behind the removal impurity is sent into the economic benefits and social benefits thickener and is concentrated, and uses plastic tank or the Plastic Bottle packing of good airproof performance as required.
Embodiment 4:
Transform the efficient bacterial strain screening process of selenium Candida utilis: the bacterium activation culture of setting out → chemomorphosis processing → ultraviolet mutagenesis processing → resistant panel primary dcreening operation → shake-flask culture sieves → the high selenium ability bacterial strain that transforms again.Implementation step is as follows:
1) get 24 hours Candida utilis of shake-flask culture and cultivate thalline and added in 6 test tubes that 2.0% ethyl sulfate solution is housed oscillation treatment respectively 15 minutes, 2% hypo solution termination reaction, centrifugal, the stroke-physiological saline solution washing.
2) thalline after ethyl sulfate is handled is changed in the culture dish, the stroke-physiological saline solution dilution was shone respectively 10～120 seconds with the 100Lx UV-light in the darkroom.
3) plate culture medium of preparation glucose 3%, peptone 1%, yeast extract paste 0.1%, agar 2%, selenium concentration 100-300mg/L, coat the resistance culture medium flat plate after will diluting with stroke-physiological saline solution through the thalline that complex mutation is handled, cultivated 48 hours about 28 ℃.Select eugonic light bacterium colony.
4) preparation contains 6 groups of the high seleno culture mediums of molasses dregs of beans of Sodium Selenite of sugar 40% molasses 4～25%, dregs of beans 2～15%, 50～300mgSe/L in triangular flask respectively.Inoculating 24～36 ℃ of shaking tables of the selected bacterium colony of primary dcreening operation respectively cultivated 48 hours, carrying out with the processing of farm products by product is the fermention medium adaptability screening of carbon, nitrogenous source, and obtaining with the processing of farm products by product is the efficient bacterial strain of Candida utilis that the matrix growth and breeding is fast, the selenium conversion capability is strong.
Be used for the biogenic selenium Production Flow Chart of organic Se-rich agricultural: transform the efficient bacterial strain seed liquor of selenium cultivation → seed liquor enlarged culturing → substrate fermentation and transform selenium → remaining matrix removal → somatic cells fragmentation → enzymolysis and contain selenium macromole → separation removal of impurities → spraying drying → pack.Concrete technology is as follows:
1) preparation seed liquor: add the molasses 4% that contain sugar 40% in the triangular flask, dregs of beans 2%, sal epsom 0.05 ‰, potassium primary phosphate 0.5%, Sodium Selenite 300mgSe/L, sterilization.Inoculum size by 5% after the efficient bacterial strain activation of the conversion selenium of preserving is inoculated in triangular flask, and 32 ℃ of shaking tables were cultivated 48 hours.
2) seed liquor enlarged culturing: add the molasses 40% that contain sugar 40% in the seeding tank, dregs of beans 5%, sal epsom 0.1 ‰, potassium primary phosphate 1.0%, industry sodium selenate 20mgSe/L, sterilization.Inoculum size by 16% inserts seed liquor, and 32 ℃ of shaking tables were cultivated 24 hours.
3) fermentation transforms selenium: add the molasses 20% that contain sugar 40% in the fermentor tank, dregs of beans 10%, sal epsom 0.5 ‰, potassium primary phosphate 2.0%, Sodium Selenite 300mgSe/L, sterilization.Insert the seed liquor that spreads cultivation by 20% inoculum size, 32 ℃ are stirred fermentation 48 hours, add the 500mgSe/L sodium selenite solution of 2 ‰ volumes respectively in the jar to 24 and 36 hours in fermentation.
4) remove remaining fermented substrate: pump into the horizontal spiral centrifuge centrifugation after fermentation is finished and remove the fermented substrate residue.
5) high-pressure homogeneous smudge cells: the fermented product of removing behind the remaining fermented substrate is transported to the high pressure homogenizer smudge cells, discharges by the synthetic selenium-containing biological molecule that enters in the born of the same parents that transforms that ferments.
6) enzymolysis contains the selenium macromole: the smudge cells fermented product is transported to hydrolytic decomposition pot, the compound protease liquid that adds 50 ‰ volumes, stirring at low speed, 55 ℃ of insulations 12 hours, the biological selenium that inorganic selenium fermentation in the matrix is transformed generate is by can not being the small molecules that plant easily absorbs by the macromolecules degradation of plant absorbing.
7) separate removal of impurities: the enzymolysis solution after degraded is finished separates through the butterfly centrifugal machine removes the impurity that cell wall fragment, starch etc. can not be utilized by plant absorbing.
8) dry packing: the hydrolyzed solution behind the removal impurity is conveyed into spray-drying tower, and 150 170 ℃ of instantaneous powder that are dried to of spraying are down packed with compound membrane bag.
Claims (3)
1. a mutagenesis screening transforms the method for selenium Candida utilis Black Liquor with Efficient Bacteria, and the bacterial strain that filters out can be fermented substrate with the processing of farm products by product, inorganic selenium toxicity is big, that bioavailability is low is converted into biological selenium nontoxic, high bioavailability efficiently.Its feature comprises following step:
1) Candida utilis bacterial strain physics+chemically composited factor mutagenesis: the Candida utilis thalline of shake-flask culture is used 50～100Lx UV-irradiation 10～120 seconds respectively in the darkroom; 0.2～2.0% ethyl sulfate was handled respectively 2～15 minutes.To bring out sudden change.
2) selenium conversion capability screening: the plate culture medium of preparation selenium concentration 100～300mg/L, coat the resistance screening culture medium flat plate after will diluting with stroke-physiological saline solution through the thalline that complex mutation is handled and carry out anti-selenium and the screening of selenium conversion capability.
3) matrix adaptability screening: being inoculated in the processing of farm products by product through the thalline that the selenium conversion capability filters out is to cultivate on the substratum of carbon, nitrogenous source to carry out the adaptability screening.Filtering out with the molasses dregs of beans is carbon, nitrogenous source, biomass height, the efficient bacterial strain of the Candida utilis that the selenium conversion capability is strong, and its selenium content reaches 800～2000mg/Kg.
2. the conversion selenium Candida utilis Black Liquor with Efficient Bacteria that obtains according to claim 1 is substrate fermentation with the processing of farm products by product, and the inorganic selenium that adds in the matrix is converted into biological selenium.Its feature comprises:
1) is carbon, nitrogenous source with the processing of farm products by product, adds inorganic selenium salt and make matrix that the conversion selenium Candida utilis Black Liquor with Efficient Bacteria that obtains with claim 1 is that kind of daughter bacteria transforms selenium through the secondary fermentation that spreads cultivation.
When 2) the conversion selenium Candida utilis Black Liquor with Efficient Bacteria that obtains with claim 1 is fermented, seed liquor spreads cultivation and the substratum that ferments is the consumption of carbon, nitrogenous source with molasses and dregs of beans, be 40% molasses 4～20%, dregs of beans 2～15% by mass fraction, selenium 20～300mgSe/L.
3) the conversion selenium Candida utilis Black Liquor with Efficient Bacteria that obtains with claim 1 is carried out in the fermenting process, adds the Sodium Selenite of 500～1000mgSe/L at interval in 8～12 hours in the fermentor tank.
3. the conversion selenium Candida utilis Black Liquor with Efficient Bacteria fermented product that obtains according to claim 2 is applicable to that through a series for the treatment of processess preparations such as removal matrix, smudge cells, complex enzyme hydrolysis, separation concentrate its feature of biological selenium product that rich selenium Organic farming is produced comprises the steps:
1) the remaining matrix of fermentation is removed: the conversion selenium Candida utilis Black Liquor with Efficient Bacteria fermented product that claim 2 fermentation obtains pumps into the horizontal spiral centrifuge centrifugation and removes the fermented substrate residue.
2) high-pressure homogeneous smudge cells: the fermented liquid of removing behind the remaining fermented substrate is transported to the high pressure homogenizer smudge cells, discharges by the synthetic selenium-containing biological molecule that enters in the born of the same parents that transforms that ferments.
3) selenium-containing biological macromolecules degradation: the smudge cells fermented product is transported to hydrolytic decomposition pot, compound protease liquid with 50 ‰ volumes, stirring at low speed, 55 ℃ of insulations 12 hours, the biological selenium that inorganic selenium fermentation on the matrix is transformed generate is by can not being the small molecules that plant easily absorbs by the macromolecules degradation of plant absorbing.
4) separate removal of impurities: the enzymolysis solution after hydrolysis is finished is transported to the butterfly centrifugal machine to be separated and removes the impurity that cell wall fragment, starch etc. can not be utilized by plant absorbing.
5) concentrate or dry: after separating the hydrolyzed solution of removing behind the impurity and sending into the multiple-effect thickener and concentrate
The bottle packing; Or be transported to 150～170 ℃ of instantaneous powder that are dried to of following spraying of spray-drying tower, use composite membrane for packing.
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| CN107094991A (en) * | 2017-04-26 | 2017-08-29 | 中国科学院合肥物质科学研究院 | A kind of animal selenium-enriched health care nutrient solution, preparation method and application |
| CN109182128A (en) * | 2018-10-16 | 2019-01-11 | 广西壮族自治区农业科学院农业资源与环境研究所 | A kind of strain culturing method of Efficient Conversion Selenium in Soil |
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