CN105028955A - Poultry feed compound enzyme and preparation thereof - Google Patents
Poultry feed compound enzyme and preparation thereof Download PDFInfo
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- CN105028955A CN105028955A CN201510156124.3A CN201510156124A CN105028955A CN 105028955 A CN105028955 A CN 105028955A CN 201510156124 A CN201510156124 A CN 201510156124A CN 105028955 A CN105028955 A CN 105028955A
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Abstract
The invention discloses a poultry feed compound enzyme and preparation thereof and belongs to the field of enzyme preparations. The invention solves a technical problem of providing the poultry feed compound enzyme which is composed of following raw materials, by weight, 10-15 parts of xylanase, 8-15 parts of cellulase, 5-10 parts of [beta]-glucanase, 1-2 parts of galactosidase, 1-2 parts of pectinase, 0.5-1 part of mannase, 3-6 parts of lactobacillus plantarum capsules, 6-10 parts of a plant preparation, 2-4 parts of a fresh ginger extract and 1-2 parts of liquorice. In the product, by means of a synergistic effect of a protective agent with the enzyme preparation, the plant preparation, the fresh ginger extract and the enzyme preparation, the compound enzyme achieves best enzyme activity and enzyme efficacy, and can correspondingly improve utilization rate of a feed and growth rate of animals, can improve appetite and disease resistance of animals, is prolonged in shelf life and can protect environment.
Description
Technical field
The invention belongs to enzymic preparation field, specifically complex enzyme and preparation method thereof.
Background technology
The enzyme preparation applied in feed industry at present mainly contains 4 large classes: be used for degraded cellulose, protein, starch and phytic acid respectively.
A kind of complex enzyme preparation for feeding piglets, application number is 201110026602.0, a kind of complex enzyme preparation for feeding piglets of disclosure of the invention.This complex enzyme preparation for feeding piglets comprises following seven kinds of enzymes: acid protease, amylase, zytase, 1,4 beta-glucanase, cellulase, pectase and phytase; The enzyme activity ratio of described seven kinds of enzymes is followed successively by 1: 1: (8-10): (3.3-4): 1: (0.5-0.55): (0-0.02).Various ANFs in complex enzyme formulation degradable feed of the present invention, reduces enteron aisle chyme viscosity, improves feedstuff metabolisable energy; Improve the digestibility of piglet to protein and starch; Reduce harmful microbe excessive multiplication in enteron aisle, reduce the damage of intestines wall, reconcile intestinal microecology balance; Improve piglet survival rate, premunition, reduce diarrhea rate, improve the overall uniformity; Promote weight gain of piglets, reduce aquaculture cost.
A kind of pig's feed complex enzyme formulation being added with activator, application number: 201210500011.7, invention provides a kind of pig's feed complex enzyme formulation being added with activator, organophosphor in feed can be resolved into the inorganic phosphate that can absorb by the phytase in formula, protein-based nutriment in protease degradable feed, promote that live pig is to the absorption of Dietary phosphorus and nitrogen, also reduces the discharge capacity of phosphorus and nitrogen in live pig fecaluria simultaneously, reduces its pollution on the environment.And activator can improve the reaction rate of enzyme, enzyme preparation is taken effect faster, and each ingredient combination in activator and enzyme preparation use the shearing force of cold cuts also can be made to reduce, and improves pork tenderness.The pig's feed complex enzyme formulation that the present invention obtains can improve the digestibility of pig to nitrogen, phosphorus in feed, thus makes the excretion of phosphorus in live pig fecaluria reduce more than 40%, and the excretion of nitrogen reduces more than 29%; Instant effect, namely has significant effect in 2-3 days; Cold cuts shearing force reduces by more than 50%.
Name is called a kind of pig's feed complex enzyme formulation and preparation method thereof application number: 201410275557.6, the invention provides a kind of pig's feed complex enzyme formulation, belongs to feed additive field.This additive is made up of complex enzyme and reinforcing agent; Complex enzyme is lipase, neutral proteinase, beta amylase, isoamylase, neutral phytase, 'beta '-mannase, zytase, alpha-galactosidase; Reinforcing agent is made up of extract from pine needles, leaf of bamboo Thick many candies, caulis lonicerae powder, galangal powder, radix jurineae powder, Asian puccoon powder, vitamin C, sodium chloride, sodium butyrate, ferrous sulfate and calcium gluconae.This enzyme preparation effectively can improve the utilization rate of feed, reduces feedstuff-meat ratio, promotes the speed of growth of pig; Can also play the effect of the immunity strengthening pig, to the health degree improving pig, reduce the pig incidence of disease and have very large help, market prospects are very wide.
Name is called a kind of efficient pig's feed complex enzyme formulation being added with enzymatic protective reagent, application number is 201210381755.1, invention provides a kind of efficient pig's feed complex enzyme formulation being added with enzymatic protective reagent, the materials such as ANFs and large biological molecule such as the beta glucan in degradable feed, phytic acid, pectin, thus promote that pig is to the absorption of feed Middle nutrition composition, improves efficiency of feed utilization; Protectantly add the inactivation rate that can reduce enzyme, extend effective time, make the effect of this enzyme preparation more significantly and lasting; The high efficiency composition enzyme preparation feed conversion rate that the present invention obtains is high, and feedstuff-meat ratio can be used to decline more than 19%; The pig speed of growth improves more than 23%; The enzymatic activity retention time is long, and in 18 months, enzymatic activity can remain on more than 90%.
Name is called a kind of efficient pig's feed complex enzyme formulation being added with enzymatic protective reagent, application number: 201210360110.X, invention provides a kind of efficient pig's feed complex enzyme formulation being added with enzymatic protective reagent, the materials such as ANFs and large biological molecule such as the beta glucan in degradable feed, phytic acid, pectin, thus promote that pig is to the absorption of feed Middle nutrition composition, improves efficiency of feed utilization; Protectantly add the inactivation rate that can reduce enzyme, extend effective time, make the effect of this enzyme preparation more significantly and lasting; The high efficiency composition enzyme preparation feed conversion rate that the present invention obtains is high, and feed coefficient can be used to decline more than 19%; The pig speed of growth improves more than 23%; The enzymatic activity retention time is long, and in 30 months, enzymatic activity can remain on more than 90%.
Name is called a kind of the feed for piglet complex enzyme formulation and production method thereof and application, application number: 201410150709.X, a kind of the feed for piglet complex enzyme formulation of disclosure of the invention and production method thereof and application.The enzyme of this complex enzyme formulation is lived and is consisted of: glucose oxidase 80-100U/g, alpha-galactosidase 20-25U/g, zytase 3000-4000U/g, 1,4 beta-glucanase 500-600U/g, 'beta '-mannase 500-600U/g, acid protease 2000-3000U/g, neutral proteinase 800-1000U/g, fungal alpha-amylase 400-500U/g, glucoamylase 3000-4000U/g.This complex enzyme formulation adopts cornstarch and potassium dihydrogen phosphate, dipotassium hydrogen phosphate, mixture of calcium sulfate as diluent.This complex enzyme formulation using method is: the addition in newborn piglet perfect compound feed per ton is 500-750 gram.
The bark of eucommia is rich in phenylpropanoids, iridoids isoreactivity composition, has rubbish in purged body, strengthens cellular material metabolism, decomposer inner cholesterol, reduces body fat, broad-spectrum antiseptic, improves white blood cell count, the remarkable efficacy such as develop immunitypty.The result of pure eucommia bark powder in zoopery shows: can promote cholesterol and lipid-metabolism; Add the eel that pure eucommia bark powder is raised, there is the taste of natural eel; Pure eucommia bark powder is added to the quality that can improve broiler chicken in feed, make it that there is tempting fragrance and quality.
Poria cocos has clearing damp and promoting diuresis, strengthening the spleen and reducing phlegm, antitoxic heart-soothing and sedative, relieve internal heat anticancer effect, and can relax gastrointestinal smooth muscle, gastric acid secretion inhibiting, prevent necrosis of liver cells, pachymic acid contained by the effect such as antibacterial has develop immunitypty, antitumor and calm, hypoglycemic etc. effect; Pachymaran can reduce malaber reefcod serum A/G value, improves serum complement C3 content, improves blood leucocyte number and leukocytes phagocytic rate, improves its non-specific immunity.
In sum, the market space that the application of enzyme has it wide and huge economic worth, but the heat endurance of feed enzyme, security, composite comprehensive and action effect give full play to the major issue being still enzyme preparation manufacturer and numerous raisers and jointly paying close attention to, prepare safer, more comprehensively, the better complex enzyme of enzyme action effect is corporation responsibility and the pursuit of industry technical staff.
Summary of the invention
A kind of poultry feed complex enzyme of technical problem solved by the invention, be made up of the raw material of following parts by weight: zytase 10-15, cellulase 8-15,1,4 beta-glucanase 5-10, galactosidase 1-2, pectase 1-2, mannase 0.5-1, Lactobacillus plantarum capsule 3-6, galenical 6-10 part, Ginger P.E 2-4 part, Radix Glycyrrhizae 1-2 part.
Described acidic xylanase, phytase, 1,4 beta-glucanase, cellulase, acid protease are food-grade enzyme preparation;
The preparation method of described galenical is as follows:
Take cassia seed 10-18 part; Bark of eucommia 5-15 part; Poria cocos 10-15 part; Respectively said herbal medicine being crushed to particle diameter is less than 2 millimeters, then Homogeneous phase mixing add the water of 3-6 times of weight in container, add the acidic cellulase of said mixture constituent mass 1-2%, control temperature 45-55 DEG C, pH:3.5-4, keeps 1-2h, adds the mixture of mixed material 2-3 times of w ethanol and propyl alcohol subsequently, control temperature to 60 DEG C-78 DEG C of maintenance 3-4h, filter; Filter vacuum concentrates postlyophilization and obtains galenical.
The mass ratio of described ethanol and propyl alcohol is 1:1-1.5; Concentration of alcohol is 95%; Propanol concentration is 100%;
The preparation method of described Ginger P.E is as follows:
To be crushed to particle diameter is that less than 2 millimeters gingers are placed in container, adds the water of 3-6 times of weight, and control temperature 50 DEG C-60 DEG C keeps 2-3h, adds the mixture of mixed material 2-3 times of w ethanol and methyl alcohol, and control temperature to 30 DEG C-40 DEG C of maintenance 3-8h, filter; Filter vacuum concentrates postlyophilization and obtains Ginger P.E.
The mass ratio of described ethanol and methyl alcohol is 1:2, concentration of alcohol 85-95%.Methanol concentration 100%.
The preparation method of described Lactobacillus plantarum capsule is as follows:
Lactobacillus plantarum capsule product is made up of wall material, Lactobacillus plantarum, freeze drying protectant, stachyose, hickory chick enzymolysis powder.
Described wall material is made up of enzymatic soybean protein isolate, shitosan, xanthans, carragheen, glycerine and trehalose; Concentration when above-mentioned each material is made into mixed solution is respectively: enzymatic soybean protein isolate 7-10%, shitosan 1-2%, xanthans 1-3%, carragheen 0.1-0.5%, glycerine 0.3-1%, trehalose 0.5-2%.
The preparation method of enzymatic soybean protein isolate is: compound concentration is that 10-13% soybean protein isolate solution is heated to 30-45 DEG C, adjustment pH to 3-5, add the acid protease insulation enzymolysis 0.5-1.5 hour of soybean protein isolate weight 0.1-1%, after enzymolysis, solution spray drying obtains enzymatic soybean protein isolate.
Hickory chick enzymolysis powder, preparation method thereof is as follows:
(1) Morchella esculenta (L.) Pers sporophore drying and crushing;
(2) water-soluble homogeneous: the Morchella esculenta (L.) Pers sporophore of pulverizing is added in stainless steel cylinder, add the water of fructification 3-6 times of weight, soak 3-5 hour, then this Morchella esculenta (L.) Pers sporophore liquid is passed through colloid mill, colloid mill operation condition is: adjustment colloid mill stator and the gap of rotor are 0.5-1 micron, colloid mill flow be 0.4-1 ton/hour;
(3) intensification enzymolysis: the Morchella esculenta (L.) Pers sporophore liquid mixed liquor through milling treatment of colloid is transferred in stainless steel enzymatic vessel and is heated to 50-60 DEG C, adjustment pH to 4.5-6.0, add the cellulase of Morchella esculenta (L.) Pers sporophore weight 0.05-0.1%, the 1,4 beta-glucanase of 0.01-0.1%, the protease of 0.01-0.1%, insulation enzymolysis 0.5-1.5 hour, constantly stirs in enzymolysis process.
(4) dry: dry acquisition hickory chick enzymolysis powder after the mash filtrations after enzymolysis.
Described drying can adopt the dried forms such as conventional spraying dry, freeze drying.
Described freeze drying protectant is made up of skimmed milk power, trehalose, maltodextrin.
The preferred CGMCCNO.9405 of the Lactobacillus plantarum that capsule product of the present invention comprises.
Above-mentioned Lactobacillus plantarum capsule product preparation method is as follows:
1) core is prepared: in the Lactobacillus plantarum zymotic fluid cultivated through ferment tank, add the stachyose of zymotic fluid quality 3-5% and the hickory chick enzymolysis powder of 5-8%, then mix with frozen-dried protective agent solution, pre-freeze 0.5h at-50 DEG C, then freeze-drying 10-18h in vacuum freeze drier, grinds after freeze-drying and makes core;
The ratio of described zymotic fluid and frozen-dried protective agent solution is 1:1.4-0.8; Described frozen-dried protective agent solution is made up of skimmed milk power, trehalose, maltodextrin solution; The volume ratio of three kinds of solution is skimmed milk power: trehalose: maltodextrin=3:1:0.5;
The concentration of described skimmed milk power, trehalose, maltodextrin solution is respectively: skimmed milk power 5%, trehalose 1%, maltodextrin 2%.
2) bag quilt: core is suspended in fluid bed, wall material spraying bag quilt, the mixed liquor of shitosan, glycerine and trehalose is sprayed into from a shower nozzle, the mixed liquor of xanthans, carragheen and enzymatic soybean protein isolate is sprayed into from another shower nozzle, spray velocity controls identical, bag between 25-38 DEG C, is namely formed capsule after 30 minutes by temperature in fluid bed in process.This drying means adopts device processes in patent 201120503311.1.
The preparation method of poultry feed complex enzyme of the present invention is as follows:
By described Radix Glycyrrhizae, galenical and Ginger P.E ultramicro grinding respectively, then with zytase, cellulase, 1,4 beta-glucanase, galactosidase, pectase, mannase, packs the poultry feed complex enzyme that gets product after Lactobacillus plantarum capsule Homogeneous phase mixing.
Described enzyme is applicable to feed-processing plant and plant's autogamy feed, should mix, can the present invention be mixed with a small amount of feed in advance, then be mixed in large quantities of feed, Direct-fed during use with other raw material in feed.Advise that complete diet pellet addition per ton is: 100-150g.
Beneficial effect:
Adopt galenical, Ginger P.E, Radix Glycyrrhizae science composite in product of the present invention, effectively slow down the moisture regain of enzyme preparation; Can strengthen complex enzyme simultaneously resistance toly to freeze, heat resistance, keep identical enzyme activity, its heat resisting temperature can improve 10-15 DEG C, resistance to cryogenic temperature can reduce 5-10 DEG C, effectively prevent the loss of complex enzyme enzyme activity in transport, preservation and use procedure, extend the shelf-life of complex enzyme, reach same enzyme activity, the like product shelf-life can extend 3-5 month.
The galenical that product of the present invention adds both can extend the shelf-life of complex enzyme formulation, can improve again the immunity of raising livestock and poultry, effectively prevent the generation of livestock and poultry epidemic disease.
Lactobacillus micro-capsule provided by the invention, modified soybean protein isolate is with the addition of in its wall material, modified soybean protein isolate improves 15-25% than soybean protein isolate emulsifying capacity raising 25%, gelling ability, modified soybean protein isolate xanthans and carragheen and shitosan with the use of, gel strength improves more than 10%, enhances the stomach juice-resistant of capsule;
Capsule of the present invention adopts modified soybean protein isolate, has good enteric solubility, complete disintegration can discharge Lactobacillus plantarum, and propagation becomes dominant microflora rapidly after capsule arrives enteron aisle in 1-1.5 hour, thus reaches effects such as suppressing growth of pathogenic bacteria.
The common synergy of protective agent and enzyme preparation, galenical, Ginger P.E and enzyme preparation in product of the present invention; the enzyme activity of complex enzyme and effect are played to greatest extent; and improve the utilization rate of feed and the growth rate of animal accordingly; enhance appetite and the resistance against diseases of animal, extend the shelf-life of product and protect environment.
Detailed description of the invention
The present invention is described below by specific embodiment.Unless stated otherwise, technological means used in the present invention is method known in those skilled in the art.In addition, embodiment is interpreted as illustrative, but not limits the scope of the invention, and the spirit and scope of the invention only limited by claims.To those skilled in the art, under the prerequisite not deviating from essence of the present invention and scope, the various change carry out the material component in these embodiments and consumption or change also belong to protection scope of the present invention.
Lactobacillus plantarum provided by the present invention (Lactobacillusplantarum) tlj-2014, this bacterial strain is preserved in China Committee for Culture Collection of Microorganisms's common micro-organisms center, deposit number is CGMCCNo.9405, preservation address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode 100101, preservation date on July 2nd, 2014.
Embodiment 1
The Breeding Process of the present invention's Lactobacillus plantarum used CGMCCNo.9405 is as follows:
The original bacterial classification that sets out → test tube activation → dithyl sulfate (DES) mutagenesis → nitrosoguanidine (NTG) mutagenesis → plasma mutagenesis → dull and stereotyped primary dcreening operation → shaking flask sieves → mitotic stability test again.
Starting strain of the present invention is in MRS dextrose culture-medium, and the throughput rate of its lactic acid is 1.5g/L/d, almost stops growing when medium pH is 3.5, is 0.34mg/h/kg Chinese cabbage to the decomposition rate of natrium nitrosum.Starting strain is collected in the greenfeed of Fattening Sheep field, Yanchi county Ningxia by Li Zheng, acquisition time on September 15th, 2013.
In order to improve the decomposition rate of its production of lactic acid speed, acid-fast ability and nitrite, DES and NTG technology is adopted to carry out mutagenesis to this bacterial classification successively, after mutagenesis, bacterial strain adopts MRS calcium carbonate flat board to carry out primary dcreening operation, then 500mL shake flask fermentation is adopted, biosensor analysis instrument carries out multiple sieve to Producing Strain, the lactobacillus plantarum strain that seed selection is excellent, then does passage assays, evaluates its genetic stability.
Lactobacillus plantarum tlj-2014 genetic stability result shows: through continuous passage ten times, property indices is all more stable, and heredity is better, and proterties is not replied, therefore using the object bacterial strain that Lactobacillus plantarum tlj-2014 obtains as seed selection.
Empirical tests finds: the production of lactic acid speed of this mutagenic strain can reach 35g/L/d, and this bacterial strain lactic acid concn after 71 hours fermentation reaches 95g/L; Can survive under pH is the condition of 1.80.Degrading nitrite speed is fast, and capacity of decomposition reaches 9.8mg/h/kg (speed of spontaneous fermentation process nitrite accumulation is approximately 1.1mg/h/kg), can resistance to 1% cholate.
1) DES mutagenic and breeding
A. on super-clean bench, get Lactobacillus plantarum L mono-ring (set out bacterium) on test tube slant, access is equipped with in the 250mL triangular flask of 50mL culture medium MRS (without agar, glucose 20g/L), 200rpm, cultivate about 12h for 37 DEG C, make thalline be in logarithmic growth in earlier stage.
B. get 5mL bacterium liquid, the centrifugal 10min of 5000rpm collects thalline, with brine 2 times.
C. 10 are diluted to pH7.0 phosphate buffer
7individual/mL bacteria suspension.
D. get the kaliumphosphate buffer of 32mLpH7.0,8mL bacteria suspension, 150mL triangular flask that 0.4mLDES to put into rotor in advance fully mix, make DES ultimate density be 1% (v/v).
E. in 37 DEG C of shaking tables, 150rpm reacts 30min, gets 1mL mixed liquor, adds 0.5mL25%Na
2s
2o
3solution stopped reaction.
F. suitably dilute, get last dilution bacterium liquid 0.2mL, coat in calcium carbonate screening and culturing base (the calcium carbonate MRS culture medium containing 100g/L glucose) plate.Cultivate after 2 ~ 3 days at 37 DEG C, adopting photolithography the bacterial strain of this screening flat board to be transferred to pH is on the upper and natrium nitrosum screening and culturing base (single nitrogenous source is the modification MRS screening and culturing base of 2g/L natrium nitrosum) of LPHMRS culture medium (low ph value modification MRS culture medium) of 1.5,1.8 and 2.0.
G. cultivate after 2 ~ 3 days at 37 DEG C, choosing colony is comparatively large, can grow respectively and on LPHMRS culture medium, natrium nitrosum screening and culturing base on calcium carbonate screening and culturing base.Through Preliminary screening, the bacterium colony called after Lactobacillus plantarum L1 that picking goes out.
2) nitrosoguanidine mutagenesis
A. on super-clean bench, get Lactobacillus plantarum L1 mono-ring on test tube slant, access is equipped with in the 250mL triangular flask of 50mL culture medium MRS (without agar) (concentration of glucose is 60g/L), 200rpm, cultivates about 12h for 37 DEG C, makes thalline be in logarithmic growth in earlier stage.
B. get the centrifugal 10min of 5mL bacterium liquid 5000rpm and collect thalline, with brine 2 times.
C. 10 are diluted to pH6.0 phosphate buffer
7individual/mL bacteria suspension.
D. get 10mL bacteria suspension to be transferred in 100mL triangular flask, add the NTG of 10mg, be mixed with the NTG solution that final concentration is 10mg/mL, and add 4-5 and drip acetone, be beneficial to NTG and dissolve.
E. at 37 DEG C, the centrifugal 10min of 200rpm oscillating reactions 30min, 5000rpm collects thalline, with SPSS washing several, and stopped reaction.
F. suitably dilute, get last dilution bacterium liquid 0.2mL, coat in calcium carbonate screening and culturing base (the calcium carbonate MRS culture medium containing 100g/L glucose) plate.Cultivate after 2 ~ 3 days at 37 DEG C, adopting photolithography the bacterial strain of this screening flat board to be transferred to pH is on the upper and natrium nitrosum screening and culturing base (single nitrogenous source is the modification MRS screening and culturing base of 2g/L natrium nitrosum) of LPHMRS culture medium (low ph value modification MRS culture medium) of 1.5,1.8 and 2.0.
G. select bacterial strain method: choosing colony is comparatively large, to grow on LPHMRS culture medium, natrium nitrosum screening and culturing base respectively and on calcium carbonate screening and culturing base.Through Preliminary screening, picking 100 meets the bacterium colony of above condition.
3) shaking flask is sieved again
A. on super-clean bench, get Lactobacillus plantarum one ring on each test tube slant respectively, access is equipped with in the 250mL triangular flask of 50mL culture medium MRS (without agar) (concentration of glucose is 100g/L), 200rpm, cultivates about 15h, makes thalline be in mid log phase for 37 DEG C.
B. get 5mL bacterium liquid respectively, LPHMRS fluid nutrient medium (low ph value modification MRS culture medium) and the natrium nitrosum liquid screening medium (single nitrogenous source is the modification MRS screening and culturing base of 2g/L natrium nitrosum) upper (note: adopt 250mL triangular flask) that 50mL calcium carbonate screens in fluid nutrient medium (the calcium carbonate MRS culture medium containing 250g/L glucose) plate, pH is 1.5,1.8 and 2.0 is equipped with in access.200rpm, cultivates 3-4 days for 37 DEG C, detects the wear rate that Pfansteihl in calcium carbonate screening fluid nutrient medium produces speed, biomass in LPHMRS fluid nutrient medium and natrium nitrosum liquid screening medium nitrite every day respectively.After fermentation ends, compare the wear rate that Pfansteihl in the calcium carbonate screening fluid nutrient medium of 100 strain bacterial classifications produces speed, biomass in LPHMRS fluid nutrient medium and natrium nitrosum liquid screening medium nitrite.
C. select to have concurrently the bacterial strain that high Pfansteihl produces speed, the wear rate of tolerate low pH (this bacterial classification only can grow in the minimum culture medium for pH1.8) and nitrite is high, by its called after L2 bacterium.
4) genetic stability test
L2 bacterium is gone down to posterity for continuous ten times on inclined-plane, and detects the fermentation situation after at every turn going down to posterity by the method that shaking flask is sieved again.Experiment finds, inclined-plane goes down to posterity for continuous ten times, and this bacterial classification proterties does not have significant change, and property indices is all normal, illustrates that the genetic stability of this bacterial classification is stronger.Strain Designation is Lactobacillus plantarum (Lactobacillusplantarum) tlj-2014.
5) 5L fermentation tank test
A. get Lactobacillus plantarum L2 mono-ring on inclined-plane, access is equipped with in the 250mL triangular flask of 50mL culture medium MRS (without agar) (concentration of glucose is 150g/L), 200rpm, cultivates about 12h, makes thalline be in mid log phase for 37 DEG C.
B. the access of the bacterial classification of logarithmic phase is equipped with in the 5L fermentation tank of 3LMRS fluid nutrient medium (initial glucose is 150g/L).Inoculum concentration is that at 10%, 37 DEG C, 100rpm cultivates 8 hours, and logarithm dissolved oxygen in early stage controls 10% (ventilation 0.5L/min), later stage Anaerobic culturel 63 hours.After fermentation ends, the lactic acid production of Lactobacillus plantarum L2 reaches 95g/L.
C. 3LpH being equipped with in the access of the bacterial classification of logarithmic phase is in the 5L fermentation tank of LPHMRS fluid nutrient medium (initial glucose is 50g/L) of 1.8.Inoculum concentration is that at 10%, 37 DEG C, 100rpm cultivates 8 hours, and logarithm dissolved oxygen in early stage controls 10% (ventilation 0.5L/min), and later stage anaerobism, zymotic fluid pH controls 1.8 by the NaOH of whole process 0.5mol/L, and total incubation time is 48 hours.After fermentation ends, the biomass detecting Lactobacillus plantarum L2 is 2.5g/L, illustrates that Lactobacillus plantarum L2 can survive in the environment of pH1.8.
D. the access of the bacterial classification of logarithmic phase is equipped with in the 5L fermentation tank of 3L natrium nitrosum liquid screening medium (single nitrogenous source is the modification MRS screening and culturing base of 2g/L natrium nitrosum).Inoculum concentration is that at 10%, 37 DEG C, 100rpm cultivates 8 hours, and logarithm dissolved oxygen in early stage controls 10% (ventilation 0.5L/min), and later stage anaerobism, sweat adds the sodium nitrite solution of 20g/L according to the wear rate stream of nitrite, cultivates 2-3 days.After fermentation ends, calculate sweat Lactobacillus plantarum L2 to the degradation rate of natrium nitrosum.Found that: under this condition, L2 can reach 563mg/h/L to the degradation rate of natrium nitrosum.
Embodiment 2
Lactobacillus plantarum capsule product is made up of wall material, Lactobacillus plantarum, freeze drying protectant, stachyose, hickory chick enzymolysis powder.
Described wall material is made up of enzymatic soybean protein isolate, shitosan, xanthans, carragheen, glycerine and trehalose; Concentration when above-mentioned each material is made into mixed solution is respectively: enzymatic soybean protein isolate 10%, shitosan 1%, xanthans 3%, carragheen 0.1%, glycerine 0.5%, trehalose 0.5%.
The preparation method of enzymatic soybean protein isolate is: compound concentration is that 10-13% soybean protein isolate solution is heated to 30-33 DEG C, pH is to 3 in adjustment, the acid protease adding soybean protein isolate weight 0.2% is incubated enzymolysis 1.5 hours, and after enzymolysis, solution spray drying obtains enzymatic soybean protein isolate.
Lactobacillus plantarum is deposit number CGMCCNO.9405 bacterial classification.
The preparation method of described hickory chick enzymolysis powder is as follows:
(1) Morchella esculenta (L.) Pers sporophore drying and crushing;
(2) water-soluble homogeneous: the Morchella esculenta (L.) Pers sporophore of pulverizing is added in stainless steel cylinder, add the water of fructification 3 times of weight, soak 5 hours, then this Morchella esculenta (L.) Pers sporophore liquid is passed through colloid mill, colloid mill operation condition is: the gap of adjustment colloid mill stator and rotor is 0.6 ± 1 micron, and colloid mill flow is 0.5 ton/hour;
(3) intensification enzymolysis: the Morchella esculenta (L.) Pers sporophore liquid mixed liquor through milling treatment of colloid is transferred in stainless steel enzymatic vessel and is heated to 50 DEG C, pH is to 6.0 in adjustment, add the cellulase of Morchella esculenta (L.) Pers sporophore weight 0.05%, the 1,4 beta-glucanase of 0.01%, the protease of 0.1%, insulation enzymolysis 1.5 hours, constantly stirs in enzymolysis process.
(4) dry: after the mash filtrations after enzymolysis, spraying dry obtains hickory chick enzymolysis powder.
The preparation process of above-mentioned Lactobacillus plantarum capsule product is as follows:
1) core is prepared: in the Lactobacillus plantarum zymotic fluid cultivated through ferment tank, add the stachyose of zymotic fluid quality 3% and the hickory chick enzymolysis powder of 8%, then mix with frozen-dried protective agent solution, pre-freeze 0.5h at-50 DEG C, then freeze-drying 10h in vacuum freeze drier, grinds after freeze-drying and makes core; The ratio of zymotic fluid and frozen-dried protective agent solution is 1:1.4; Frozen-dried protective agent solution is made up of skimmed milk power, trehalose, maltodextrin solution; The volume ratio of three kinds of solution is skimmed milk power: trehalose: maltodextrin=3:1:0.5;
The concentration of described skimmed milk power, trehalose, maltodextrin solution is respectively: skimmed milk power 5%, trehalose 1%, maltodextrin 2%.
2) bag quilt: core is suspended in fluid bed, wall material spraying bag quilt, the mixed liquor of shitosan, glycerine and trehalose is sprayed into from a shower nozzle, the mixed liquor of xanthans, carragheen and enzymatic soybean protein isolate is sprayed into from another shower nozzle, spray velocity controls identical, bag at 28 DEG C, is namely formed capsule after 30 minutes by temperature in fluid bed in process.Adopt device preparation as described in patent 201120503311.1.
Lactobacillus plantarum capsule product bile tolerance is tested.
Capsule product in embodiment 2 be placed in pig cholate solution (solution concentration 3.0%) insulation of 37 DEG C and constantly stir, take out after 2h, with sterile saline washing, with opening one's purse, liquid dissolves, measure Lactobacillus Survival, and compare with the bacterium liquid do not embedded.Test result is as shown in the table.
| Metamorphosis | Survival rate | |
| Embodiment 1 | Not disintegration | 87.5％ |
| Embodiment 2 | Not disintegration | 89.3％ |
| Do not wrap and contrasted | -- | 0.65％ |
Embodiment 3
A kind of poultry feed complex enzyme of technical problem solved by the invention, be made up of the raw material of following parts by weight: zytase 12, cellulase 10,1,4 beta-glucanase 8, galactosidase 1, pectase 1, mannase 0., 8, Lactobacillus plantarum capsule 5, galenical 8 parts, Ginger P.E 3 parts, 1.5 parts, Radix Glycyrrhizae.
The preparation method of described galenical is as follows:
Take cassia seed 15 parts; The bark of eucommia 10 parts; 12 parts, Poria cocos; Respectively said herbal medicine being crushed to particle diameter is less than 2 millimeters, then Homogeneous phase mixing add the water of 5 times of weight in container, add the acidic cellulase of said mixture constituent mass 2%, control temperature 50 DEG C, pH:3.8, keeps 1h, adds the mixture of mixed material 3 times of w ethanol and propyl alcohol subsequently, control temperature to 70 DEG C keeps 3h, filters; Filter vacuum concentrates postlyophilization and obtains galenical.
The mass ratio of described ethanol and propyl alcohol is 1:1; Concentration of alcohol is 95%; Propanol concentration is 100%;
The preparation method of described Ginger P.E is as follows:
To be crushed to particle diameter is that less than 2 millimeters gingers are placed in container, adds the water of 4 times of weight, and control temperature 55 DEG C keeps 2h, adds the mixture of mixed material 3 times of w ethanol and methyl alcohol, and control temperature to 35 DEG C keeps 5h, filters; Filter vacuum concentrates postlyophilization and obtains Ginger P.E.
The mass ratio of described ethanol and methyl alcohol is 1:2, concentration of alcohol 90%.Methanol concentration 100%.
Embodiment 4 is basic with routine 1-3
A kind of poultry feed complex enzyme of technical problem solved by the invention, be made up of the raw material of following parts by weight: zytase 10, cellulase 15,1,4 beta-glucanase 5, galactosidase 2, pectase 2, mannase 1, Lactobacillus plantarum capsule 3, galenical 10 parts, Ginger P.E 2 parts, 2 parts, Radix Glycyrrhizae.
Described acidic xylanase, phytase, 1,4 beta-glucanase, cellulase, acid protease are food-grade enzyme preparation;
Embodiment 6
A kind of poultry feed complex enzyme of technical problem solved by the invention, be made up of the raw material of following parts by weight: zytase 15, cellulase 8,1,4 beta-glucanase 10, galactosidase 1, pectase 1, mannase 0.5, Lactobacillus plantarum capsule 3, galenical 6 parts, Ginger P.E 2 parts, 1 part, Radix Glycyrrhizae.
The preparation method of poultry feed complex enzyme of the present invention is as follows:
By described Radix Glycyrrhizae, galenical and Ginger P.E ultramicro grinding respectively, then with zytase, cellulase, 1,4 beta-glucanase, amylase, galactosidase, pectase, mannase, packs the poultry feed complex enzyme that gets product after Lactobacillus plantarum capsule Homogeneous phase mixing.
Feeding experiment
Materials and methods
The selection of test pig and grouping
The a certain large-scale regular pig farm in Hunan, chooses the healthy DLY three way cross grower pigs that 36 body weight are (30 ± 2) kg, is divided into 3 groups at random, often organizes 2 hurdles and repeats, often repeat 6, galt and gilt half and half.The prerun of 1 week formal test advance behavior phase, and complete expelling parasite and routine immunization work.Test point two phases of front and back carry out, in earlier stage (30kg-60kg), and the later stage (61kg ~ 90kg).
Feeding and management
Feed dry mash, free choice feeding, is limited cannot not have enough surplusly, and automatic drinking bowl is drunk water.Day in early stage feeds three times, and day in later stage feeds secondary, in units of hurdle, record feed consumption rate.Pig house is brick structure, cement flooring, single-column type, and examination pig is raised respectively in 9 hurdles, and the environmental condition of each column home is consistent, cleans colony house morning and afternoon every day each once, observes the behavior of pig, appetite, ight soil simultaneously.Duration of test record disease and treatment situation.
Feed and formula
Based on full price GB grower pigs feed, feed per ton adds the grower pigs specific enzyme 100g of test group, control group 1 and control group 2;
Test index: before the test at the end of (30kg-60kg), mid-term and later stage (61kg ~ 90kg), point another name individual weight, weighs and carries out on an empty stomach all in the morning.Stage by stage, record its every daily material consumption, incidence of disease in units of hurdle, calculate daily ingestion amount and food utilization efficiency simultaneously, and carry out statistical analysis to above-mentioned data, grower pigs growth performance is in table 2.
Table 2
Claims (8)
1. a poultry feed complex enzyme, be made up of the raw material of following parts by weight: zytase 10-15, cellulase 8-15,1,4 beta-glucanase 5-10, galactosidase 1-2, pectase 1-2, mannase 0.5-1, Lactobacillus plantarum capsule 3-6, galenical 6-10 part, Ginger P.E 2-4 part, Radix Glycyrrhizae 1-2 part.
2. poultry feed complex enzyme according to claim 1, the preparation method of described galenical is as follows:
Take cassia seed 10-18 part; Bark of eucommia 5-15 part; Poria cocos 10-15 part; Respectively said herbal medicine being crushed to particle diameter is less than 2 millimeters, then Homogeneous phase mixing add the water of 3-6 times of weight in container, add the acidic cellulase of said mixture constituent mass 1-2%, control temperature 45-55 DEG C, pH:3.5-4, keeps 1-2h, adds the mixture of mixed material 2-3 times of w ethanol and propyl alcohol subsequently, control temperature to 60 DEG C-78 DEG C of maintenance 3-4h, filter; Filter vacuum concentrates postlyophilization and obtains galenical.
3. poultry feed complex enzyme according to claim 1, the preparation method of described Ginger P.E is as follows:
To be crushed to particle diameter is that less than 2 millimeters gingers are placed in container, adds the water of 3-6 times of weight, and control temperature 50 DEG C-60 DEG C keeps 2-3h, adds the mixture of mixed material 2-3 times of w ethanol and methyl alcohol, and control temperature to 30 DEG C-40 DEG C of maintenance 3-8h, filter; Filter vacuum concentrates postlyophilization and obtains Ginger P.E.
4., according to poultry feed complex enzyme described in claim 1, the preparation method of described Lactobacillus plantarum capsule is as follows:
Lactobacillus plantarum capsule product is made up of wall material, Lactobacillus plantarum, freeze drying protectant, stachyose, hickory chick enzymolysis powder.
Described wall material is made up of enzymatic soybean protein isolate, shitosan, xanthans, carragheen, glycerine and trehalose; Concentration when above-mentioned each material is made into mixed solution is respectively: enzymatic soybean protein isolate 7-10%, shitosan 1-2%, xanthans 1-3%, carragheen 0.1-0.5%, glycerine 0.3-1%, trehalose 0.5-2%; The preparation method of enzymatic soybean protein isolate is: compound concentration is that 10-13% soybean protein isolate solution is heated to 30-45 DEG C, adjustment pH to 3-5, add the acid protease insulation enzymolysis 0.5-1.5 hour of soybean protein isolate weight 0.1-1%, after enzymolysis, solution spray drying obtains enzymatic soybean protein isolate.
5. according to poultry feed complex enzyme described in claim 4, the preferred CGMCCNO.9405 of Lactobacillus plantarum.
6., according to poultry feed complex enzyme described in claim 1-4, preparation method is as follows for Lactobacillus plantarum capsule product:
1) core is prepared: in the Lactobacillus plantarum zymotic fluid cultivated through ferment tank, add the stachyose of zymotic fluid quality 3-5% and the hickory chick enzymolysis powder of 5-8%, then mix with frozen-dried protective agent solution, pre-freeze 0.5h at-50 DEG C, then freeze-drying 10-18h in vacuum freeze drier, grinds after freeze-drying and makes core;
The ratio of described zymotic fluid and frozen-dried protective agent solution is 1:1.4-0.8; Described frozen-dried protective agent solution is made up of skimmed milk power, trehalose, maltodextrin solution; The volume ratio of three kinds of solution is skimmed milk power: trehalose: maltodextrin=3:1:0.5;
The concentration of described skimmed milk power, trehalose, maltodextrin solution is respectively: skimmed milk power 5%, trehalose 1%, maltodextrin 2%.
2) bag quilt: core is suspended in fluid bed, wall material spraying bag quilt, the mixed liquor of shitosan, glycerine and trehalose is sprayed into from a shower nozzle, the mixed liquor of xanthans, carragheen and enzymatic soybean protein isolate is sprayed into from another shower nozzle, spray velocity controls identical, bag between 25-38 DEG C, is namely formed capsule after 30 minutes by temperature in fluid bed in process.
7., according to poultry feed complex enzyme described in claim 1, be made up of the raw material of following parts by weight:
Zytase 12, cellulase 10,1,4 beta-glucanase 8, galactosidase 1, pectase 1, mannase 0., 8, Lactobacillus plantarum capsule 5, galenical 8 parts, Ginger P.E 3 parts, 1.5 parts, Radix Glycyrrhizae;
A kind of poultry feed complex enzyme, is made up of the raw material of following parts by weight:
Zytase 10, cellulase 15,1,4 beta-glucanase 5, galactosidase 2, pectase 2, mannase 1, Lactobacillus plantarum capsule 3, galenical 10 parts, Ginger P.E 2 parts, 2 parts, Radix Glycyrrhizae;
A kind of poultry feed complex enzyme, is made up of the raw material of following parts by weight:
Zytase 15, cellulase 8,1,4 beta-glucanase 10, galactosidase 1, pectase 1, mannase 0.5, Lactobacillus plantarum capsule 3, galenical 6 parts, Ginger P.E 2 parts, 1 part, Radix Glycyrrhizae.
8. according to the preparation method of poultry feed complex enzyme described in claim 1, as follows:
By described Radix Glycyrrhizae, galenical and Ginger P.E ultramicro grinding respectively, then with zytase, cellulase, 1,4 beta-glucanase, amylase, galactosidase, pectase, mannase, packs the poultry feed complex enzyme that gets product after Lactobacillus plantarum capsule Homogeneous phase mixing.
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