CN104256086A - Technology for preparing docosahexaenoic acid (DHA)-rich feed additive by grain dreg raw material through fermentation - Google Patents
Technology for preparing docosahexaenoic acid (DHA)-rich feed additive by grain dreg raw material through fermentation Download PDFInfo
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Abstract
The invention discloses a technological method for preparing an animal feed additive which is rich in polyunsaturated fatty acid, namely, docosahexaenoic acid (DHA), protein and xylo-oligosaccharide by taking a grain dreg raw material as a culture medium through a microbiological fermentation method. The method concretely comprises the steps of (1) mixing the grain dreg raw material with water to prepare a material with a certain solid-liquid mass ratio; (2) carrying out pretreatment on the mixed grain dreg raw material by a hydrothermal extraction method; (3) carrying out enzymolysis on the pretreated grain dregs; (4) culturing microorganism capable of producing DHA by the grain dregs treated by enzymolysis; and (5) concentrating or drying fermentation liquor of schizochytrium limacinum to prepare a liquid or solid-state feed. The method fully utilizes the principle that the grain dregs contain rich hemicellulose and cellulose so as to be used for preparing xylo-oligosaccharide and cello-oligosaccharide with a probiotic function, and soluble lignin is used for preparing a polyphenol antioxidant; and the degraded cellulose and starch can be used as a carbon source for microbial growth, and the protein can be used as a nitrogen source for cell growth, so that the costs of the carbon source and the nitrogen source for microbial fermentation can be lowered.
Description
Technical field
The present invention relates to the production technology of feed addictive, be specifically related to a kind of with distillers grains and dregs material for the process of rich DHA feed addictive produced by raw material.
Background technology
DHA (DHA) is a kind of extremely important highly unsaturated fatty acid, is the important composition material of human body cell film and nerve fiber.DHA is at angiocardiopathy preventing, suppression and Therapeutic cancer and ensure the normal development of fetus and promote to have important function in infant development growth.But the DHA that the Chinese common people absorb every day from food is only 40mg, and recommends minimum standard intake to be 160mg), belong to famine state.Patent of invention 201210385417.5 discloses " one is rich in DHA microalgae powder and preparation method thereof ", the zymotic fluid being rich in DHA is directly processed into microalgae powder by this patent, then it can be used as feed addictive for being also effectively enriched in animal body in herding, aquaculture, again by the food delivery route of " being rich in the microalgae powder → animal body → people of DHA ", make the mankind from diet, take in the DHA of q.s.But in prior art, the production cost of DHA algal powder is very high, is difficult to the increase realizing common people DHA intake by this method.Therefore for reducing the cost of material of fermenting and producing DHA algal powder, need to find cheap microorganism and cultivate Carbon and nitrogen sources, and distillers grains and dregs resource is for addressing this problem the approach of providing.Grain slag is that it contains the nutritional labelings such as a large amount of crude fibre, thick protein and crude fat by remaining juicy residual substances after the utilized carbohydrate extraction in raw material.China's distillers grains and dregs material resource enriches, and only brewages, starch, Fruit quality can produce poor slag about 100,000,000 tons every year, are a kind of available valuable regenerated resources.
The poor slag of existing multiple utilization prepares the process of animal and fowl fodder, mainly comprises and will make dry feed after poor slag convection drying and biological fermentation process prepares animal feeding-stuff containing somatic protein.Such as prepare DDGS with maize alcohol stillage, not only cost is high, energy consumption is high for this method, and does not make full use of each nutritional labeling in poor slag.It is the process route that another kind utilizes poor slag material that biological fermentation process prepares animal feeding-stuff containing somatic protein, at present, most based on simple directly cultivation saccharomyces cerevisiae or other microorganisms.Patent of invention 201210276611.X discloses " a kind of method of preparing feedstuff by biological modification of brewery mash ", patent of invention 201010525743.2 discloses " utilizing cassava grain stillage to produce the method for biological feedstuff ", and patent of invention 200510019848.X discloses " a kind of spirit stillage produces the method for thalli feed ".But biological fermentation process can only utilize residual starch in poor slag and monose, and cellulose and hemicellulose are not still utilized further.CN201210420826 discloses a kind of reuse method of vinasse, is specially: after being dried by vinasse, pulverize and add aqueous medium and fully soak, and carry out steam blasting pretreatment, obtain steam blasting pretreatment material; Described steam blasting pretreatment material is made solid medium, adds probio and carry out solid fermentation, obtain fermented feed.Although the method partly can utilize cellulose in vinasse and hemicellulose, its technical problem solved produces the biological feedstuff containing probio, and need first by raw material drying, causes power consumption excessive.
Hydro-thermal extraction process has been widely used in the pretreatment of lignocellulosic, to improve cellulosic enzymolysis efficiency.Hydro-thermal extraction process possesses following advantage: (1), without the need to the participation of the chemical substance such as soda acid or ammonia, can not cause corrosion to consersion unit; Water is environmental friendliness resource, not only reduces cost, and can not cause serious environmental problem; (2) high-pressure water heating can break the structure of lignin, without the need to the crushing material process before pretreatment, reduces energy consumption; (3) with optimal conditions, hemicellulose can be made farthest to dissolve with compound sugar form, and the generation controlling monose generate the fermentation inhibitors such as furfural to prevent it from continuing degraded.Starch in poor slag, hemicellulose and portion of cellulose can be converted into monose (glucose, wood sugar and arabinose) and have wood oligose and the cell-oligosaccharide of prebiotic function by application hydro-thermal method process grain slag; The protein in concentrated poor slag, fat and micro-can be extracted again simultaneously, and starch after enzymolysis in extract and cell-oligosaccharide and protein can as the high-quality of growth of microorganism cheap carbon source and nitrogenous source.
In sum, in conjunction with the high added value advantage of rich DHA product and the low cost feature of distillers grains and dregs raw material, hydrothermal pretreatment is utilized to process poor slag in conjunction with the process of enzymolysis, then the nutriment after application process in extract produces the feed addictive of rich DHA as the Carbon and nitrogen sources producing DHA growth of microorganism, the maximum using of distillers grains and dregs material nutrient component can be realized on the one hand, again reducing the cost that fermentation method produces DHA on the other hand, is therefore a kind of method of feasible efficiency utilization distillers grains and dregs agricultural by-products.
Patent EP 2520608 A1 discloses a kind of continuous hydrothermal method preprocessing lignocellulose of applying to improve the method for cellulosic enzymolysis efficiency and ethanol production.But the method solve only the degradation problem of ligno-cellulosic materials, do not solve the process problem of the distillers grains and dregs material of high protein, high starch and high-moisture.Hydrothermal carbonization method is applied to process and brewages slag by patent CN 103221518 A, but biomaterial there occurs hydrothermal carbonization under its process conditions, and the method will be brewageed slag and finally be converted into high-carbon product (biological coal and humus).
Summary of the invention
Based on the problem of prior art in poor slag utilization and rich DHA Feed Manufacturing, the invention provides one with distillers grains and dregs raw material for culture medium, the process of the feed addictive of employing Production by Microorganism Fermentation rich DHA, protein and wood oligose.
Technical scheme of the present invention: adopt distillers grains and dregs material production to be rich in the technique of the additive for farm animal feed of DHA, comprise the following steps:
1. material pretreatment: pending distillers grains and dregs material is mixed with water and obtains mixed material, the pH value of adjustment mixed material is 3-8; Dry poor slag in described distillers grains and dregs material and the mass ratio of water are 1:3-1:50; Described distillers grains and dregs material is any one or the combination multiple arbitrarily of brewageing in slag, fecula and pomace; Described step 1. in the preferred 1:3-1:10 of mass ratio of dry poor slag in distillers grains and dregs material and water, most preferably 1:3-1:8.
Described slag of brewageing refers to starch yielding plant property material for fermentation raw material, the distillers grains and dregs accessory substance that the production process that the fermentable production being filling auxiliary material with straw and seed shell contains ethanol and acetic acid product produces, such as spirit stillage and vinegar grain.Described starch yielding plant property material comprises the stem tuber of starch-containing material, root, whole grain and their any combination.Described starch-containing material comprises any combination of corn, wheat, jowar, barley, cassava, Ipomoea batatas, potato, rye, pea, soya bean or above-mentioned material.The straw of filling auxiliary material used is preferably barley-straw, wheat-straw and cornstalk; The seed shell of filling auxiliary material used is preferably rice husk.This kind of distillers grains and dregs accessory substance contains: (a) is from starch, the thalline of the fiber of starch material, shell, embryo, oil, albumen and non-complete fermentation; (b) and from filling the lignin of auxiliary material rice husk and stalk, cellulose and hemicellulose.
Described fecula refers to and adopts starch-containing material to prepare the fecula that starch process produces.Described starch-containing material comprises any combination of corn, wheat, jowar, barley, cassava, Ipomoea batatas, potato, rye, pea, soya bean or above-mentioned material.Preferred fecula comprises corn starch residue, batata starch dregs, cassava starch dregs, potato starchy residue.Fecula contains the starch from the fiber of starch material, shell, embryo, oil and protein ingredient and remnants.
Described pomace refers to the accessory substance obtained after fruit is squeezed the juice.Described fruit comprises apple, grape and oranges and tangerines.Described pomace contains from the cellulose of pericarp, fruit stone and carpopodium, hemicellulose, lignin, protein and fat.
2. hydro-thermal method process: mixed material step 1. prepared proceeds in container, is slowly heated to temperature under agitation and reaches T1; Under T1 temperature conditions, be incubated 1-180 minute, then fast cooling is to T2, obtains hydro-thermal grain slag; Described T1 is 120-230 DEG C, and described T2 is 20-80 DEG C.After hydro-thermal method process, starch degradation in grain slag is dextrin, hemicellulose degradation is the xylan of low polymerization degree and wood oligose and wood sugar, and some cellulose dissolves the indigestibility albumen-alcohol soluble protein be degraded in cell-oligosaccharide, poor slag can be changed into the protein of easy enzymolysis.The physicochemical properties of grain slag there occurs great variety, more easily by enzyme hydrolysis.
3. enzyme hydrolysis process: (a) batch enzymolysis: the temperature of hydro-thermal grain slag is adjusted to 20-90 DEG C, and pH value is adjusted to 3-8; Then in hydro-thermal grain slag, add glucoamylase, zytase, cellulase, react after 48-72 hour, then add mmp reaction 2-24 hour, obtain enzymolysis material; Or (b) fed-batch: hydro-thermal grain slag is added in enzyme solutions in batches, described enzyme solutions is the mixed enzyme solution of glucoamylase, zytase and cellulase, react after 48-72 hour, then add mmp reaction 2-24 hour, obtain enzymolysis material.After enzyme hydrolysis, starch and few fibers oligosaccharides are degraded to glucose, and xylan degrading is wood oligose, and protein degradation is small peptide and amino acid.
Described glucoamylase (α-Isosorbide-5-Nitrae-glucose hydrolysis enzyme, EC.3.2.1.3) can use one or more glucoamylases, and addition is the enzyme that every gram of dry poor slag solid adds 0.02-200AGU.Described zytase is inscribe-Isosorbide-5-Nitrae-β zytase (3.2.1.8), the inscribe-Isosorbide-5-Nitrae-β zytase of preferred GH10 or GH11; The addition of described zytase is every 1 × 10
2-1 × 10
5gram dry poor slag adds 1g zytase or total enzyme lives 1 × 10
6the zytase of IU.Described cellulase for there is the enzyme of cellobiohydrolase activity (E.C.3.2.1.91) and there is the enzyme of active and beta glucan glycosides enzyme (E.C.3.2.1.21) activity of endoglucanase (E.C.3.2.1.91), described in there is cellobiohydrolase activity (E.C.3.2.1.91) enzyme comprise cellobiohydrolase I and cellobiohydrolase II; The addition of described cellulase is the enzyme that every gram of dry poor slag solid adds 0.1-100FPU.Described protease is acid protease, and its addition is every gram of distiller's dried grain solid 1 × 10
-4-1 × 10
2the enzyme of LAPU.
4. solid-liquor separation: the solid in enzymolysis material is separated from liquid, described separate mode be centrifugal, compacting or filter; Preferred embodiment comprise plate and frame filter press, belt filter press, screw press and butterfly centrifugal machine.Solid-phase component is cellulose, lignin and mineral matter mainly; Liquid phase ingredient comprises glucose, cell-oligosaccharide, wood oligose, protein, polypeptide, amino acid and aldehydes matter and a small amount of acetic acid.
5. fermentable: the concentration and the ratio that regulate Carbon and nitrogen sources in liquid phase after solid-liquor separation, add required nutritional labeling (yeast extract, vitamin, metal ion, phosphate etc.), then the microorganism into product DHA is inoculated, carry out fermentable, obtain microbial fermentation solution.The microorganism of described product DHA is that schizochytrium limacinum belongs to (Schizochytrium/Aurantiochytrium sp), genus thraustochytrium (Thraustochytrium), hidden dinoflagellate belongs to any one or population mixture multiple arbitrarily in (Crypthecodinium), my Ken Shi Chytridium (Ulkenia) and Mortierella (Mortierella).
6. drying and crushing: the mixed liquor of multiple-microorganism fermentation culture is carried out drying, pulverizing by (a), obtains solid-state feed additive; Or (b) is by mixing after the drying of single microbial fermentation solution, pulverizing, obtains solid-state feed additive.The method of described drying is freeze drying, vacuum drying, heated-air drying or spraying dry.Preferably freeze drying, because carry out the oxidizable and DHA of temperature-sensitive of dry available protecting under low temperature and vacuum condition.The Polyphenols antioxidant that hydro-thermal extracting grain slag lignin produces simultaneously also can play the protective effect to DHA.
Adopt the purposes of additive for farm animal feed prepared by said method, as the feed addictive of livestock and poultry, aquiculture animal and companion's property animal; Described livestock and poultry comprise ox, pig, chicken, duck and goose, and described aquiculture animal comprises shrimp and fish; Described companion's property animal is cat and dog.
Beneficial effect of the present invention:
(1) what method of the present invention can make full use of that poor slag contains enrich hemicellulose and cellulose can be used for preparing the wood oligose and cell-oligosaccharide with prebiotic effect, utilize soluble lignin to prepare Polyphenols antioxidant; And degraded after cellulose and starch can by the carbon source with growth of microorganism, protein can be used as the nitrogenous source of Growth of Cells, thus reduces the Carbon and nitrogen sources cost of fermentable.
(2) the poor slag after hydro-thermal method process is used for the fermentable producing DHA by the present invention, takes full advantage of the feature that material moisture is high, directly processes, reduce the consumption of water when can directly separately not add water or add water less; The acidulous material contained of grain slag material itself can as the self-catalysis agent in hydrothermal treatment consists, without the need to adding external source acidic catalyst;
(3) hot conditions of hydro-thermal method can by starch liquefacation, not only makes that starchy material is easier to be separated with other solid matters, and reduces diastatic consumption in enzymolysis step; Cellulose after hydrothermal treatment consists and dextrin are more easily glucose by enzymolysis, reduce the consumption of cellulase and glucoamylase;
(4), after hydrothermal treatment consists, the hemicellulose major part in poor slag is converted into wood oligose and xylan and soluble in water; The Polyphenols antioxidant that lignocellulosic contains is extracted, and is conducive to stablizing containing DHA algal powder; The enzymatic hydrolyzation of non-digestible protein (mainly alcohol soluble protein) can significantly improve.
Detailed description of the invention
Below in conjunction with embodiment, the present invention is described further.
Embodiment 1:
Spirit stillage is the byproduct of wine-making industry.According to statistics, China 2013 produces white wine 1,000 ten thousand tons per year, and spirit stillage reaches more than 3,000 ten thousand tons.If processed not in time, will be putrid and deteriorated, not only waste valuable resource, also can severe contamination surrounding environment.
Get the fresh spirit stillage 100g of water content 70%, add 20ml water, after fully stirring and evenly mixing, make that solid-liquid mass ratio is 1:3, pH value is the mixed material of 3.Be placed in hydrothermal reactor by material after mixing, mixing speed 200rpm, heats 180 DEG C, keeps this temperature 10 minutes.Process rear stopping heating and be cooled to rapidly 55 DEG C.Adjust ph is 3, adds the glucoamylase that gross activity is 3000AGU, and the zytase of 20000IU, the cellulase of 2000FPU react 72 hours.Add the acid protease of 2000LAPU after 72 hours and act on 9 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, Cobastab 150mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts schizochytrium limacinum Aurantiochy trium sp.SD116 (be preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, its deposit number is CGMCC No:6208).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
Embodiment 2:
Get the fresh vinegar grain 100g of water content 90%, add 410ml water, after fully stirring and evenly mixing, to make solid-liquid mass ratio be 1:50 and pH value is the mixed material of 8.Be placed in hydrothermal reactor by material after mixing, mixing speed 300rpm, is heated to 230 DEG C, keeps this temperature 1 minute.Process rear stopping heating and be cooled to rapidly 80 DEG C.Adjustment temperature is 90 DEG C, pH value is 8, and add the glucoamylase that gross activity is 1AGU, the zytase of 1000IU, the cellulase of 1000FPU react 48 hours.Add the acid protease of 1000LAPU after 48 hours and act on 2 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, Cobastab 150mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts thraustochytriale Thraustochytrium (purchased from American Type Culture Collection, numbering ATCC 28210).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
Embodiment 3:
Get the corn starch residue 100g of water content 80%, add 420ml water, after fully stirring and evenly mixing, make that solid-liquid mass ratio is 1:25, pH value is the mixed material of 5.Be placed in hydrothermal reactor by material after mixing, mixing speed 150rpm, is heated to 120 DEG C, keeps this temperature 180 minutes.Process rear stopping heating and be cooled to rapidly 20 DEG C.Adjust ph is 5, adds the glucoamylase that gross activity is 1000AGU, and the zytase of 2500IU, the cellulase of 2FPU react 68 hours.Add the acid protease of 0.002LAPU after 68 hours and act on 24 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, pangamic acid 0mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts hidden dinoflagellate Crypthecodinium (purchased from American Type Culture Collection, numbering ATCC 30556).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
Embodiment 4:
Get the citrus pulp 100g of water content 70%, add 230ml water, after fully stirring and evenly mixing, make that solid-liquid mass ratio is 1:10, pH value is the mixed material of 4.Be placed in hydrothermal reactor by material after mixing, mixing speed 150rpm, is heated to 140 DEG C, keeps this temperature 120 minutes.Process rear stopping heating and be cooled to rapidly 30 DEG C.Adjust ph is 4, adds the glucoamylase that gross activity is 30AGU, and the zytase of 1000IU, the cellulase of 900FPU react 54 hours.Add the acid protease of 300LAPU after 54 hours and act on 8 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, Cobastab 150mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts my Ken Shi chytrid Ulkenia sp (purchased from American Type Culture Collection, numbering ATCC 28207).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
Embodiment 5:
Get the spirit stillage 100g of water content 80%, add 80ml water, after fully stirring and evenly mixing, make that solid-liquid mass ratio is 1:8, pH value is the mixed material of 6.Be placed in hydrothermal reactor by material after mixing, mixing speed 150rpm, is heated to 180 DEG C, keeps this temperature 90 minutes.Process rear stopping heating and be cooled to rapidly 50 DEG C.Adjust ph is 6, adds the glucoamylase that gross activity is 500AGU, and the zytase of 5000IU, the cellulase of 20FPU react 58 hours.Add the acid protease of 20LAPU after 58 hours and act on 16 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, Cobastab 150mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts schizochytrium limacinum Aurantiochy trium sp.SD116 (be preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, its deposit number is CGMCC No:6208).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
Embodiment 6:
Get the pomace 100g of water content 60%, add 140ml water, after fully stirring and evenly mixing, make that solid-liquid mass ratio is 1:5, pH value is the mixed material of 8.Be placed in hydrothermal reactor by material after mixing, mixing speed 150rpm, is heated to 200 DEG C, keeps this temperature 60 minutes.Process rear stopping heating and be cooled to rapidly 65 DEG C.Adjust ph is 8, adds the glucoamylase that gross activity is 8000AGU, and the zytase of 5000IU, the cellulase of 200FPU react 60 hours.Add the acid protease of 4LAPU after 60 hours and act on 20 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, Cobastab 150mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts Mortierella Mortierella sp (purchased from American Type Culture Collection, numbering ATCC 32222).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
Embodiment 7:
Get the potato starchy residue 100g of water content 50%, add 300ml water, after fully stirring and evenly mixing, make that solid-liquid mass ratio is 1:7, pH value is the mixed material of 5.Be placed in hydrothermal reactor by material after mixing, mixing speed 150rpm, is heated to 220 DEG C, keeps this temperature 10 minutes.Process rear stopping heating and be cooled to rapidly 75 DEG C.Adjust ph is 5, adds the glucoamylase that gross activity is 500AGU, and the zytase of 1000IU, the cellulase of 500FPU react 62 hours.Add the acid protease of 0.5LAPU after 62 hours and act on 5 hours.After 200 order gauzes filter, get filtrate and measure concentration of glucose and ammonia ion concentration.Filtrate concentration of glucose is regulated with glucose and water.Fermentation medium main component is glucose 60g/L, yeast extract 10g/L, potassium dihydrogen phosphate 5g/L, magnesium sulfate 3g/L, natrium citricum 3g/L, Cobastab 150mg/L, vitamin B6 310mg/L, vitamin B12 20mg/L, biotin 20mg/L.Medium pH 6.8, cultivation temperature 25 DEG C.The microorganism producing DHA adopts schizochytrium limacinum Aurantiochy trium sp.SD116 (be preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, its deposit number is CGMCC No:6208).Initial inoculum is often liter of 0.5g (0.1-0.6g) dry weight, and shaking table cultivates 72 hours, rotating speed 200rpm.After cultivation terminates, vacuum freeze drying fermentation culture also pulverizes dry rear solid, obtains rich DHA additive for farm animal feed.
The nutrient component meter (wt%) of the rich DHA feed of table 1 gained of the present invention
| ? | Embodiment 1 | Embodiment 2 | Embodiment 3 | Embodiment 4 | Embodiment 5 | Embodiment 6 | Embodiment 7 |
| Wood oligose | 8 | 7 | 6 | 8 | 8 | 8 | 8 |
| DHA | 30 | 22 | 24 | 21 | 20 | 28 | 25 |
| Protein | 49 | 52 | 51 | 55 | 48 | 40 | 53 |
| Cell-oligosaccharide | 3 | 3 | 2 | 1 | 2 | 5 | 4 |
Claims (10)
1. adopt distillers grains and dregs material production to be rich in the technique of the additive for farm animal feed of DHA, it is characterized in that: comprise the following steps:
1. material pretreatment: pending distillers grains and dregs material is mixed with water and obtains mixed material, the pH value of adjustment mixed material is 3-8; Dry poor slag in described distillers grains and dregs material and the mass ratio of water are 1:3-1:50;
2. hydro-thermal method process: mixed material step 1. prepared proceeds in container, is slowly heated to temperature under agitation and reaches T1; Under T1 temperature conditions, be incubated 1-180 minute, then fast cooling is to T2, obtains hydro-thermal grain slag; Described T1 is 120-230 DEG C, and described T2 is 20-80 DEG C;
3. enzyme hydrolysis process: the temperature of hydro-thermal grain slag is adjusted to 20-90 DEG C, and pH value is adjusted to 3-8; Then in hydro-thermal grain slag, add glucoamylase, zytase, cellulase, react after 48-72 hour, then add protease, reaction 2-24 hour, obtains enzymolysis material;
4. solid-liquor separation: the solid in enzymolysis material is separated from liquid, described separate mode be centrifugal, compacting or filter;
5. DHA fermentation: the concentration and the ratio that regulate Carbon and nitrogen sources in liquid phase after solid-liquor separation, add required nutritional labeling, then inoculate the microorganism into product DHA, carry out fermentable, obtain microbial fermentation solution;
6. drying and crushing: microbial fermentation broth is carried out drying, pulverizing, obtains solid-state feed additive.
2. the technique of the additive for farm animal feed of DHA is rich in employing distillers grains and dregs according to claim 1 material production, it is characterized in that: described step enzyme hydrolysis processing mode is 3. for adding hydro-thermal grain slag in enzyme solutions in batches, described enzyme solutions is glucoamylase, zytase and cellulase mixed enzyme solution, react after 48-72 hour, add protease again, reaction 2-24 hour, obtains enzymolysis material.
3. the technique of the additive for farm animal feed of DHA is rich in employing distillers grains and dregs according to claim 1 material production, it is characterized in that: the microorganism of the product DHA that step is 5. described is schizochytrium limacinum genus, genus thraustochytrium, hidden dinoflagellate belong to any one or population mixture multiple arbitrarily in I Ken Shi Chytridium, genus Shewanella and Mortierella.
4. be rich in the technique of the additive for farm animal feed of DHA according to the employing distillers grains and dregs material production in claim 1-3 described in any one, it is characterized in that: the addition of step 3. described glucoamylase is the enzyme that every gram of dry poor slag solid adds 0.02-200AGU; Described zytase is inscribe-Isosorbide-5-Nitrae-β zytase, and the addition of described zytase is that every gram of dry poor slag adds 10-1 × 10
4the zytase of IU; Described cellulase is have the enzyme of cellobiohydrolase activity and have the enzyme of endoglucanase activity and beta glucan glycosides enzymatic activity, and the addition of described cellulase is the enzyme that every gram of dry poor slag solid adds 0.1-100FPU; The addition of described protease is every gram of distiller's dried grain solid 1 × 10
-4-1 × 10
2the enzyme of LAPU.
5. the technique of the additive for farm animal feed of DHA is rich in employing distillers grains and dregs according to claim 4 material production, it is characterized in that: the 3. described glucoamylase of step is α-Isosorbide-5-Nitrae-glucose hydrolysis enzyme; Described zytase is the inscribe-Isosorbide-5-Nitrae-β zytase of GH10 or GH11; The described enzyme with cellobiohydrolase activity comprises cellobiohydrolase I and cellobiohydrolase II; Described protease is acid protease.
6. the technique of employing distillers grains and dregs material production additive for farm animal feed according to claim 1, is characterized in that: described distillers grains and dregs material is any one or the combination multiple arbitrarily of brewageing in slag, fecula and pomace; Described step 1. in the mass ratio of distillers grains and dregs material and water be 1:3-1:10, preferably 1:3-1:8.
7. the technique of employing distillers grains and dregs material production additive for farm animal feed according to claim 2, is characterized in that: described step 6. in microbial fermentation solution be the mixed liquor of multiple-microorganism zymotic fluid; The method of described drying is freeze drying, vacuum drying, heated-air drying or spraying dry.
8. the technique of employing distillers grains and dregs material production additive for farm animal feed according to claim 2, is characterized in that: described step 6. in by dry for single microbial fermentation solution, pulverize after mixing, obtain solid-state feed additive; The method of described drying is freeze drying, vacuum drying, heated-air drying or spraying dry.
9. the additive for farm animal feed adopting method according to claim 1 to prepare, is characterized in that: by mass fraction, comprises 40-55 part protein, 20-30 part DHA, 8-10 part wood oligose and 1-5 part cell-oligosaccharide.
10. the purposes of additive for farm animal feed adopting method according to claim 1 to prepare, is characterized in that: as the feed addictive of livestock and poultry, aquiculture animal and companion's property animal; Described livestock and poultry comprise ox, pig, chicken, duck and goose, and described aquiculture animal comprises shrimp and fish; Described companion's property animal is cat and dog.
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