CN103859169A

CN103859169A - Feed compound enzyme containing neutral protease and preparation method thereof

Info

Publication number: CN103859169A
Application number: CN201310682190.5A
Authority: CN
Inventors: 李洪兵; 李贵骏; 杨中秋; 孙霞; 王章林; 王永兰
Original assignee: Hunan Hongying Biological Science & Technology Co Ltd
Current assignee: Hunan Hongying Biological Science & Technology Co Ltd
Priority date: 2013-12-13
Filing date: 2013-12-13
Publication date: 2014-06-18
Anticipated expiration: 2033-12-13
Also published as: CN103859169B

Abstract

The invention discloses a feed compound enzyme containing neutral protease and a preparation method thereof, and belongs to the technical field of preparation of an enzyme preparation. The feed compound enzyme containing neutral protease is prepared from concentrated malt enzyme, acid proteinase, neutral protease, acidic xylanase, cellulose, beta-glucanase, pectinase, amylase, seminase, phytase, Chinese herb extracts, a protective agent and an activating agent in a scientific distribution manner. The feed compound enzyme is high in enzyme activity, wide in action condition, strong in stability, and more applicable to feed production. A safe digestive enzyme can be provided for beasts and birds, the digestion burden is relieved, the utilization rate and the growth rate of the raw materials are improved, the environment is protected, joyful natural malt fragrance is brought for the feed, the livestock and poultry appetite is enhanced, meanwhile, the efficacy of the enzyme preparation can be fully played by proper quantity of activating agent under the same condition, the adding amount of the enzyme preparation is saved, the expiration date of the compound enzyme preparation is prolonged by scientific distribution of the Chinese herb extracts, and the immunity of the fed beasts and birds can be improved.

Description

A kind of feed complex enzyme containing neutral proteinase and preparation method thereof

Technical field

The invention belongs to enzyme preparation technical field, specifically a kind of feed complex enzyme containing neutral proteinase and preparation method thereof.

Background technology

In feed, add enzyme preparation and mainly contain following 4 reasons: 1. the ANFs that degraded exists in animal feed.These materials can not be degraded by animal endogenous enzymes, thereby disturb the eubolism of animal, cause animal digestion bad, and production performance declines.2. improve the utilization rate of starch, protein and mineral matter.These materials or surrounded by rich fibrous cell membrane, or can not be had by the version of animal digestion with some that (for example, in plant feed raw material, a large amount of phosphorus exists with the form of phytate phosphorus.）。3. some specific chemical bond of degrading in raw material.These chemical bonds can not be degraded by the enzyme of animal self, add exogenous enzymes can discharge more nutrients later.4. because young animal autodigestion system is also immature, the not enough interpolation of endogenous enzymes exogenous enzymes can improve feed digestibility, prevents indigestion symptom.Except can improving the utilization rate of daily ration, the enzyme-added difference that can also reduce between feedstuff, the accuracy of raising feed formula, the while can also be improved the regularity of growth of animal, reduces management cost, increases economic efficiency.Use all right protection of the environment of enzyme preparation.Because the utilization rate of feed has improved, corresponding ight soil discharge capacity has declined.In the situation that effect is obvious, the discharge capacity of ight soil can reduce by 20% left and right, the discharge of nitrogen 15% left and right that declines in pig manure, and in chicken manure, the discharge of nitrogen declines 20%.For phytate phosphorus, can significantly reduce the pollution of phosphorus to environment.

The enzyme preparation of applying in feed industry at present mainly contains 4 large classes: be used for respectively degraded cellulose, protein, starch and phytic acid.

Fiber degradation enzyme: for nonruminant, the maximum resistance of digestion is the enzyme that can not produce degradation of fibers, in the daily ration that contains the components such as wheat, barley, oat, fiber is araboxylan and beta-glucan greatly.Water miscible fiber can improve the viscosity of small intestine contents, hinders the absorption of nutrient, thereby reduces the growth performance of animal.Simultaneously this situation also with some because the disease that indigestion causes is relevant.As black toe disease and the piggy of the anorexia of pig, fowl have loose bowels.Due to the impact of the factors such as kind, growth place gentle time condition, the altering a great deal of fiber content in barley and wheat, causes the nutritive value of the daily ration that contains these components widely different.Fiber degradation enzyme, zytase and beta-glucan can reduce these differences, improve growth performance and the regularity of animal.Can also reduce some dyspeptic disease simultaneously.

PD enzyme: protein is from all feeds raw material in animal diets, and they finally stockpile in lean meat by the amino acid of degraded.In nonruminant daily ration, add protease (DIFFERENT FEED material protein and quality and utilizability) except fully degrading most of storage protein or Storage protein or for the available small-molecular peptides of animal, can also improve feed nutritive value by degraded anti-nutritional factors.The efficiency variance stockpiling is very large.At plant protein source, as existed some ANFs in beancake powder, as several tannins and trypsin ihhibitor, may cause damage to intestinal absorption surface, affect nutraceutical absorption.In addition, the incomplete digestion system of young animal can not well be utilized the protein in vegetable protein (as beancake powder).

Starch degrading enzyme: many nutritionists think that corn is feedstuff " golden standard ".Large absolutely number nutritionist thinks that corn does not exist lienteric, digestibility exceedes 95%, but Noy and Sklan research show (1994) in the ideal situation recently, in the daily ration of broiler of 4-12 ages in days, the digestibility of starch seldom exceedes 85%, adds amylase and can make starch obtain more degradeds faster at small intestine.In the weaned piglet phase, due to nutrition, environment and immune variation, body weight can decline.In daily ration, add amylase and some other enzyme, can increase the endogenous digestive ferment secretion of animal, and then improve digesting and assimilating of nutrition, improve food conversion ratio and growth of animal rate.

Phytic acid digestive enzyme: for all animals, phosphorus is all vital for mineralising, immunity, breeding, the growth of bone.Pig and poultry nonruminant can only in utilize in plant feed 30-40% phosphorus, all the other phytate phosphorus of 60-70% are unserviceable.In many cases, in feed diet, Phos must be supplemented and meet the needs of growth of animal.Phosphorus over half in feed is along with ight soil is discharged in environment, contaminated environment.Add the phytase phytic acid of can degrading, discharge the phosphorus in phytic acid molecule.Can produce like this 2 benefits: 1. the addition that has reduced Dietary phosphorus.2. having reduced feces of livestock and poultry pollutes the phosphorus of environment.

Apparent: as four large leading roles of feed enzyme, their mechanism of action and pattern determine or promoted animal feed industry to use for the absorption of enzyme preparation technology to a great extent.The ratio for input and output example of adding enzyme at present in broiler chicken material exceedes 2:1.Comparatively speaking, in pig industry field, the service condition of enzyme preparation, with regard to more complicated, seems uncertain.Intensive degree is low, relates to link many, and the result of use of enzyme preparation is difficult to carry out business calculating.Although had the imagination that uses enzyme preparation in the 1950's, until just start to understand strength how to bring into play enzyme in feed industry the eighties in 20th century.Feed grain, as Wheat and barley all contains the unavailable fiber of higher nonruminant.As fruit fiber can be degraded, animal just can utilize nutrients better.In Europe, barley is more cheap, and bird nutritionist and zymologist have dropped into great effort and studied and in the daily ration of broiler that contains barley, add beta-glucan enzyme to reduce the possibility of its negative effect.Its result is proved to be successfully, and has obtained a gold law: barley+beta-glucan enzyme=wheat.Be subject to above-mentioned successful inspiration, wheat is the research object of second.Theory hypothesis is: wheat+zytase=corn.The research of this step has also obtained success.In the mid-90 in 20th century, enzyme has obtained generally accreditation at feed industry.Can not rant out: 1996, in the broiler chicken material (viscosity cereal is energy source) in Europe 80%, contain fiber degradation enzyme.Strengthen thus and accelerate the application of feed industry to new technology.In the world, about 65% the poultry feed that can produce viscosity cereal that contains has added fiber degradation enzyme.And application percentage in pig feed is much lower, approaches 10%.Its main cause is the complicated structure in market, and market is diversification, even cannot calculate.From geographical distribution, use the area of cellulose degrading enzyme mainly to concentrate on the producing region of viscosity cereal as main energy feed, for example: Europe, Canada, Australia and New Zealand.In addition, in the U.S., South America and the Asian-Pacific area, service condition depends on the rate of exchange between corn and wheat.In this sense, Europe is to use the core of degraded cellulose enzyme to segment market.In order to obtain global accreditation, feeding enzyme producer must carry out on a large scale marching taking corn---and bean pulp type daily ration is main North America and the Asian-Pacific area.Corn---bean pulp type daily ration is always counted as " golden standard ", although many nutritionists think that the mobility of these raw materials is more much bigger than the mobility of original imagination.Now, increasing evidence shows that this gold daily ration also can improve its production performance by enzyme, although this class daily ration problem relevant to crude fibre or viscosity is not serious.Past 10 years was expended a lot in research and development Corn-soybean first generation feed enzyme, and started successful Application in 1996, and initial stage application result is multifarious, but industry is just starting to become how more and more understand could be handy, adds zymotechnic and obtains maximum economy return.It is estimated, this feed a part enzyme market share is 2,000 ten thousand dollars, and actual only have 5% for enzyme-added feed at the broiler fodder that uses Corn-soybean daily ration.Within 1999/2000 year one, reduce the feed enzyme market value that viscosity and crude fibre are daily ration and exceed 100,000,000 dollars.At present, phytase has obtained admitting and applying of the whole world.The market share of phytase is approximately annual 5000 ten thousand dollars, approximately has the pig fowl feed of 8.0% left and right to add phytase in the whole world.Except the reason of economic interests, also having a factor is to have reduced the content of phytate phosphorus in excrement to be conducive to protection of the environment.

In sum, the application of feed enzyme has its wide market space and huge economic worth, but the heat endurance of feed enzyme, security, composite comprehensive and giving full play to of action effect are still enzyme preparation manufacturer and the common major issue of paying close attention to of numerous raisers, prepare safer, more comprehensively, enzyme action effect is better, and the feed complex enzyme that can improve raising appetite of livestock and poultry is industry technical staff's corporation responsibility and pursuit.

Summary of the invention

Technical problem solved by the invention is with enzyme system comprehensively, natural safety, there is concentrated maltase and the high enzymatic activity of Fructus Hordei Germinatus fragrance, action condition is wide in range, the neutral proteinase that stability is strong is basis, and the composite Chinese herbal medicine extract of science, protective agent, activator and other food stage feed enzyme, the feed complex enzyme containing neutral proteinase making not only provides safety for raising livestock and poultry, comprehensively digestive ferment, alleviate digestion burden, improve raw material availability and growth rate, effectively protection of the environment, and can bring joyful native malt fragrance to feed, increase the appetite of livestock and poultry, appropriate activator can under equal conditions be given full play to the effect of enzyme preparation simultaneously, make the best use of everything, save enzyme preparation addition, the composite shelf-life that both can extend complex enzyme formulation of science of Chinese herbal medicine extract, can improve again the immunity of raising livestock and poultry, thereby reach the multiplex effect of an enzyme.

In order to achieve the above object, the present invention is by the following technical solutions:

Containing a feed complex enzyme for neutral proteinase, formed by the enzyme preparation of following parts by weight:

Concentrated maltase 30-40 part, acid protease 20-30 part, neutral proteinase 20-30 part; acidic xylanase 20-30 part; cellulase 20-30 part, 1,4 beta-glucanase 15-20 part, pectase 15-20 part; amylase 15-20 part; seminase 10-15 part, phytase 10-15 part, Chinese herbal medicine extract 10-15 part; protective agent 10-15 part, activator 10-15 part.

Described acid protease, neutral proteinase, acidic xylanase, cellulase, 1,4 beta-glucanase, pectase, amylase, seminase, phytase are food-grade enzyme preparation;

The preparation method of described neutral proteinase is as follows:

Bacillus subtilis 1398-2-12 through actication of culture and step by step expand cultivate obtain liquid seeds; Liquid seeds is accessed to fermentation tank, cultivation temperature 30-36 DEG C, mixing speed 200-700r/m, ventilation (V/V) 1:1-3, incubation time 10-15h with 6% inoculum concentration; Then with 1-2 DEG C/h rate of temperature fall slow cooling to 10-15 DEG C, constant temperature culture 15-20h; Continuation to 2-5 DEG C, now, is appended access fermentation tank, constant temperature culture 20-30h by liquid seeds with 4% inoculum concentration with 1-2 DEG C/h rate of temperature fall slow cooling; Finally slowly be warming up to 10-15 DEG C, constant temperature culture 15-20h with 1-2 DEG C/h heating rate; Continue to be slowly warming up to 30-36 DEG C, constant temperature culture 15-20h with 1-2 DEG C/h heating rate; Zymotic fluid after filtration, concentrated, allotment, essence filter, the dry solid neutral proteinase that to obtain; Described allocation process adds concentrated enzyme liquid gross weight 0.5-5% Chinese herbal medicine powder.

Described slant medium consists of: beef extract 3-10g, and sodium chloride 5-12g, peptone 10-20g, glucose 2-5g, (NH) ₂sO ₄3-5g, K ₂hPO ₄6-8g, CaCl ₂1-3g, agar 15-20g, Chinese herbal medicine powder 5-10g, distilled water l000mL, pH value 7.0-7.2,121 DEG C of sterilizing 20min;

The preparation method of described Chinese herbal medicine powder is as follows:

Take Radix Astragali 20-30 part; Radix Codonopsis 10-18 part; Radix bupleuri 10-15 part; Root of large-flowered skullcap 10-15 part; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C-90 DEG C and keep 2-4h, then be cooled to 45-60 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 5.5-6.8, enzymolysis 2-4h, finally add the mixture of mixed material 0.5-3 times of weight ethanol and propyl alcohol, control temperature to 60 DEG C-78 DEG C of maintenance 3-4h, filter; Filtrate Vacuum Concentration postlyophilization obtains Chinese herbal medicine powder.

Described mixed enzyme addition is the 5-10% of mixed material gross weight.

The parts by weight of described mixed enzyme consist of: endo-beta-glucanase 10-20 part, outer 1,4 beta-glucanase 10-20 part, beta-glucosidase 10-15 part, zytase 15-20 part, pentosanase 15-20 part, Pullulanase 20-30 part, beta amylase 10-15 part, neutral proteinase 10-15 part, acid protease 10-15 part, superoxide dismutase 5-10 part, glucose oxidase 5-10 part, acid phosphatase 5-10 part.

The mass ratio of described ethanol and propyl alcohol is 1:1-1.5.

Described seed culture medium percentage by weight consists of:

Dusty yeast 0.3-0.5%, glucose 1-1.5%, peptone 0.3-0.5%, beef extract 0.5-0.8%, dipotassium hydrogen phosphate 0.8-1.5%, Chinese herbal medicine powder 1.5-2%, trehalose 1-3%, calcium sulfate 1g, magnesium chloride 2g, natrium citricum 1-3g, insufficient section pure water is supplied, pH value 7.0-7.2,121-123 DEG C of sterilizing 30-40min.

Described seed tank culture base percentage by weight consists of:

Maltodextrin 5-15%, dusty yeast 0.4-0.8%, Chinese herbal medicine powder 1.5-2%, trehalose 1-3%, peptone 0.1-0.5%, corn steep liquor 0.1-0.5%, dipotassium hydrogen phosphate 0.8-1.5%, magnesium sulfate 0.05-0.1%, natrium citricum 0.1-0.5%, insufficient section pure water is supplied, pH value 7.0-7.2,121-123 DEG C of sterilizing 30-40min.

Described seeding tank zymotic fluid cell concentration is 7.0x10 ⁸-8.0x10 ⁸individual/ml;

Described fermentation medium consists of: maltodextrin 50-150g, corn flour 50-60g, beancake powder 15-25g, Chinese herbal medicine powder 30-50g, trehalose 30-40g, dusty yeast 4-8g, corn steep liquor 1-5g, ammonium sulfate 1-3g, dipotassium hydrogen phosphate 1-2g, potassium dihydrogen phosphate 1-2g, natrium citricum 1-5g, defoamer 0.1-1g, pure water l000mL, pH value 7.0-7.2,121 DEG C of sterilizing 20min;

Described supplemented medium percentage by weight consists of: maltodextrin 20-30%, and corn flour 10-20%, bean powder 15-25%, herbal mediciment powder 5-10%, insufficient section pure water is supplied, pH value 7.0-7.2,121-123 DEG C of sterilizing 30-40min.

The concocting method of described fermentation medium is:

Accurately take in proportion raw material, pure water in raw material, corn flour, beancake powder are dropped in material-compound tank, regulate pH value 7.0-7.2, add middle temperature amylase (3u/g corn flour) and alpha-amylase (30u/g corn flour), DEG C insulation 15-30min of warming while stirring to 70 simultaneously, is then slowly warming up to 90 DEG C of insulation 15-30min and liquefies, finally add other raw material, stir, adjust initial p H7.0-7.2,121-123 DEG C of sterilizing 30-40min is for subsequent use.

Described concentrated maltase percentage by weight consists of: former maltase 30-40%, fragrance maltase 60-70%.

Described former maltase preparation method is as follows:

(1) high-pressure pulse electric (PEF) immersion treatment: by Fructus Hordei Germinatus in pre-immersion trough with 3-6 times of 20-50 DEG C of emerge in worm water 10-20min, make Fructus Hordei Germinatus moisture reach 25-35%, carry out high-pressure pulse electric (PEF) processes simultaneously, electric-field intensity 20-40KV/cm, burst length 150-200 μ S, pulse frequency 200-300Hz;

(2) pulverize: add maltcrusher band pigment broken in the Fructus Hordei Germinatus of handling well in step (1) and immersion water, adjusting roller spacing is 0.5mm, and rotating speed is 250rpm, obtains Fructus Hordei Germinatus slurries;

(3) ultrasonic, PEF extracts: Fructus Hordei Germinatus slurries are placed in to pill tank, and limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 0.6A/25w-1.5A/275w condition, carried out ultrasonic extraction 10-15min; Then carry out high-pressure pulse electric (PEF) and extract 15-20min, electric-field intensity 20-40KV/cm, burst length 400-600 μ S, pulse frequency 200-300Hz;

(4) filter: be 100 order-300 order diatomite in room temperature by 500-800 order filter cloth precoating granularity by extract, adopt flame filter press to filter, operating pressure is 0.14-0.24Mpa;

(5) ultrafiltration concentration: adopt the daltonian milipore filter circulation of molecular cut off <100000 concentrated, until concentrate enzyme content is 10-15 times before ultrafiltration;

(6) dry: to the protective agent of the present invention that adds concentrate weight 3-5% in concentrate, then dry with the special spray dryer of enzyme preparation, EAT 150-160 DEG C; temperature of outgoing air 75-85 DEG C; drying time 5-15s, product moisture < 5%, both former maltase.

Described fragrance maltase preparation method is as follows:

(1) Fructus Hordei Germinatus moisture regain: use malt weight 3-4%, temperature is 40-50 DEG C of warm water even spraying Fructus Hordei Germinatus grain surface, puts the even 3-5min of mixing in mixer.

(2) cure: pack moisture regain Fructus Hordei Germinatus into drum-type and fry stove, in 10-15min, temperature rises to 170-200 DEG C, insulation 10-15min, is then cooled to 100-110 DEG C, and insulation 20-30min, finally takes out spreading for cooling, obtains fragrance Fructus Hordei Germinatus;

(3) pulverize: add maltcrusher to pulverize in fragrance Fructus Hordei Germinatus, adjusting roller spacing is 0.5mm, and rotating speed is 250rpm, obtains fragrance malt flour;

(4) ultrasonic, PEF extracts: fragrance malt flour is placed in to pill tank, adds 3-6 water doubly, limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 0.6A/25w-1.5A/275w condition, carried out ultrasonic extraction 10-15min; Then carry out high-pressure pulse electric (PEF) and extract 15-20min, electric-field intensity 20-40KV/cm, burst length 400-600 μ S, pulse frequency 200-300Hz;

(5) filter: adopt flame filter press to filter in room temperature 500-800 order filter cloth extract, operating pressure is 0.14-0.24Mpa;

(6) ultrafiltration concentration: adopt the daltonian milipore filter circulation of molecular cut off <100000 concentrated, until concentrate enzyme content is 10-15 times before ultrafiltration;

(7) dry: to the protective agent of the present invention that adds concentrate weight 3-5% in concentrate, then dry with the special spray dryer of enzyme preparation, EAT 150-160 DEG C; temperature of outgoing air 75-85 DEG C; drying time 5-15s, product moisture < 5%, both fragrance maltase.

The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C-90 DEG C and keep 2-4h, then be cooled to 45-60 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 5.5-6.8, enzymolysis 2-4h, finally add the mixture of mixed material 0.5-3 times of weight ethanol and propyl alcohol, control temperature to 60 DEG C-78 DEG C of maintenance 3-4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then freeze drying obtains Chinese herbal medicine extract.

The parts by weight of described Chinese herbal medicine consist of: sea-buckthorn 20-30 part, cassia seed 20-30 part, matrimony vine 10-15 part, Chinese yam 10-15 part, radix glehniae 5-10 part, fruit of negundo 5-10 part, radix polygonati officinalis 3-5 part, seed of Job's tears 3-5 part, fructus hordei germinatus 3-5 part, sweet osmanthus 3-5 part, Radix Astragali 3-5 part;

Described mixed enzyme addition is the 5-10% of mixed material gross weight;

The parts by weight of described mixed enzyme consist of: endo-beta-glucanase 10-20 part, outer 1,4 beta-glucanase 10-20 part, beta-glucosidase 10-15 part, zytase 15-20 part, pentosanase 15-20 part, Pullulanase 20-30 part, beta amylase 10-15 part, neutral proteinase 10-15 part, acid protease 10-15 part, superoxide dismutase 5-10 part, glucose oxidase 5-10 part, acid phosphatase 5-10 part;

Described protective agent is made up of the raw material of following parts by weight: trehalose 20-30 part, NaCl20-30 part, (NH ₄) SO ₄10-15 part, cysteine 10-15 part.

Described activator is evenly to be mixed by the inorganic salts of following quality component: zinc chloride 30-40 part, calcium chloride 10-20 part, sodium sulphate 10-20 part, magnesium chloride 5-10 part.

The present invention contains the preparation method of the feed complex enzyme of neutral proteinase:

By described protective agent, Chinese herbal medicine extract, concentrated maltase ultramicro grinding respectively; guarantee that granularity is less than described neutral proteinase and other enzyme preparation; then with neutral proteinase, acid protease, acidic xylanase, cellulase, 1,4 beta-glucanase, pectase, amylase, seminase, phytase; evenly mix; finally add activator, after mixing, pack the feed complex enzyme getting product containing neutral proteinase.

Bacillus subtilis 1398-2-12 provided by the invention is obtained through UV-LiCl-dithyl sulfate Mutation screening by bacillus subtilis (Bacillus subtilis) 1398-2 of a strain product neutral proteinase of laboratory preservation.

The bacterial strain of product heat-flash stability neutral proteinase provided by the invention is specially bacillus subtilis 1398-2-12.This bacterial strain is preserved in Chinese Typical Representative culture collection center on November 3rd, 2013 and (is called for short CCTCC, address: China. Wuhan. Wuhan University, postcode: 430072), preserving number is CCTCC NO:M2013539, Classification And Nomenclature: bacillus subtilis 1398-2-12(Bacillus subtilis1398-2-12).

Described bacillus subtilis (Bacillus subtilis) 1398-2-12 bacterial strain feature is as follows:

Described bacterial strain colony colour on solid plate is milky, and dry tack free is opaque, and neat in edge, for having the aerobic bacteria of motility.Microscopy is elongated rod shape, and Gram's staining is positive.This bacterium can utilize citrate, and nitrate reductase, V-P test into positive.

Bacillus subtilis 1398-2-12 provided by the invention has that produced neutral proteinase tolerable temperature is high, the feature of applicable pH value wide scope, 75 DEG C of enzymes of zymotic fluid crude enzyme liquid complete stability alive, 70 DEG C of optimal reactive temperatures, pH value 4.5-8.5 enzyme is lived stable, optimal reaction pH value 7.2.This bacterial strain the most suitable growth pH value 7.0-7.2, optimum growth temperature 30-36 DEG C, the suitableeest product enzyme temperature 32-35 DEG C.

Press mutagenesis screening scheme, mutant strain step-sizing is eliminated, finally to strain excellent through fermenting property test screen, obtain a strain and produce the bacterial strain bacillus subtilis bacterium 1398-2-12 of heat-flash stability neutral proteinase, the work of zymotic fluid neutral proteinase enzyme can reach 5500-7000U/mL, heat endurance to enzyme is analyzed, and crude enzyme liquid is placed in respectively at 40 DEG C, 45 DEG C, 50 DEG C, 55 DEG C, 60 DEG C, 65 DEG C, 70 DEG C, 75 DEG C, lives every 10 minutes sampling and measuring enzymes.At 40 DEG C, 45 DEG C, 50 DEG C, 55 DEG C, 60 minutes enzymes are lived and are not declined.At 60 DEG C and 65 DEG C, what within 30 minutes, drop to constitutive enzyme work drops to 85% in 95%, 60 minute.At 70 DEG C, what within 30 minutes, drop to constitutive enzyme work drops to 80% in 85%, 60 minute.At 75 DEG C, what within 30 minutes, drop to constitutive enzyme work drops to 70% in 80%, 60 minute.

Beneficial effect:

1. the present invention can reach 5500-7000U/mL containing the neutral proteinase fermentation broth enzyme work in the feed complex enzyme of neutral proteinase, is 2.5 times of starting strain.Between 40-70 DEG C, have higher enzyme and live, optimal reactive temperature is 70 DEG C; Enzyme complete stability alive in the time that pH value is 4.5-8.5, optimal reaction pH value is 7.2; This enzyme still can keep 80% above enzyme to live preserve 1h under 70 DEG C of conditions after, keep 70% above enzyme work after preserving 1h under 75 DEG C of conditions.Stronger than existing neutral protein enzyme heat stability, enzyme activity is higher, enzyme effect optimum pH wide scope, and storage stability is high, is more applicable to raising the interpolation of animal and fowl fodder.

The present invention containing the concentrated maltase in the feed complex enzyme of neutral proteinase taking high-quality import Fructus Hordei Germinatus as raw material, adopting the methods such as ultrasonic wave, high-voltage pulse electric field technology (pulsed electric field is called for short " PEF ") to extract to greatest extent maltase is enzyme source, it is a kind of natural complex enzyme, its enzyme source kind and single enzyme characteristic 100% be from brewers malt, and hydrolysis result is far superior to adopt microorganism the ferment single enzyme and the combination thereof that make; Fragrance maltase wherein not only can provide enzyme source, and most importantly its strong, unique Fructus Hordei Germinatus fragrance can obviously improve the appetite of raising livestock and poultry.

3. the present invention adopts polysaccharide, inorganic salts and amino acid science composite containing the protective agent in the feed complex enzyme of neutral proteinase, has effectively slowed down the moisture regain of fragrance maltase and complex enzyme formulation; Can strengthen complex enzyme simultaneously resistance toly freeze, heat resistance, keep identical enzyme activity, its heat resisting temperature can improve 20-30 DEG C, resistance to cryogenic temperature can reduce 10-15 DEG C, effectively prevent the loss of complex enzyme enzyme activity in transport, preservation and use procedure, extended the shelf-life of complex enzyme, reached same enzyme activity, the like product shelf-life can extend 2-3.

4. the present invention adds inorganic salts as activator containing the feed complex enzyme of neutral proteinase; create the optimum condition of enzyme catalysis; give full play to the vigor of the each enzyme component of complex enzyme; the macromolecular substances such as starch in feed, protein, cellulose, phytic acid are thoroughly effectively decomposed; greatly alleviate the digestion burden of livestock and poultry animal; improve the growth rate of raw material availability and livestock and poultry, effectively prevented the environmental pollution that feces of livestock and poultry causes, protected feeding environment simultaneously.

5. the Chinese herbal medicine extract that the present invention adds containing the feed complex enzyme of neutral proteinase both can extend the shelf-life of complex enzyme formulation, can improve again the immunity of raising livestock and poultry, effectively prevented the generation of livestock and poultry epidemic disease.

6. the present invention is containing the common synergy of protective agent in the feed complex enzyme of neutral proteinase and enzyme preparation, activator and enzyme preparation, Chinese herbal medicine extract and enzyme preparation; enzyme activity and the effect of complex enzyme are brought into play to greatest extent; and the utilization rate of feed and the growth rate of animal are improved accordingly; strengthen appetite and the resistance against diseases of animal, extended the shelf-life of complex enzyme and protected environment.

Detailed description of the invention

Below by specific embodiment narration the present invention.Unless stated otherwise, in the present invention, technological means used 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 are only limited by claims.To those skilled in the art, do not deviating under the prerequisite of essence of the present invention and scope various changes that the material component in these embodiments and consumption are carried out or change and also belong to protection scope of the present invention.

Embodiment 1

35 parts of concentrated maltases, 25 parts of acid proteases, 25 parts of neutral proteinases, 25 parts of acidic xylanases, 25 parts of cellulases; 17 parts of 1,4 beta-glucanases, 17 parts of pectases, 17 parts of amylase, 12 parts of seminases; 13 parts of phytases, 12 parts of Chinese herbal medicine extracts, 13 parts of protective agents, 12 parts of activator.

The preparation method of described neutral proteinase comprises the steps:

(1) slant strains of intact bacillus subtilis 1398-2-12 is inoculated in to slant medium, cultivates 24h for 30 DEG C and carry out actication of culture, so activate 2 times;

Described slant medium consists of: beef extract 3g, and sodium chloride 5g, peptone 10g, glucose 2g, (NH) ₂sO ₄3g, K ₂hPO ₄6g, CaCl ₂1g, agar 15g, Chinese herbal medicine powder 5g, distilled water l000mL, 7.0,121 DEG C of sterilizing 20min of pH value;

Take 20 parts of the Radixs Astragali; 10 parts of Radix Codonopsis; 10 parts of radix bupleuri; 10 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3 times of weight, control temperature 70 C and keep 2h, be then cooled to 45 DEG C, add the mixing enzyme preparation of mixed material gross weight 5% to carry out enzymolysis, with newborn acid for adjusting pH value be 5.5, enzymolysis 2h, finally adds the mixture of 0.5 times of weight ethanol of mixed material and propyl alcohol, and the mass ratio of described ethanol and propyl alcohol is 1:1, control temperature to 60 DEG C and keep 3h, filter; Filtrate Vacuum Concentration postlyophilization obtains Chinese herbal medicine powder.

The parts by weight of described mixed enzyme consist of: 10 parts of endo-beta-glucanases, 10 parts of outer 1,4 beta-glucanases, 10 parts of beta-glucosidases, 15 parts of zytases, 15 parts of pentosanases, 20 parts of Pullulanases, 10 parts of beta amylases, 10 parts of neutral proteinases, 10 parts of acid proteases, 5 parts of superoxide dismutases, 5 parts of glucose oxidases, 5 parts of acid phosphatases.

(2) liquid seeds expands cultivation

1. first order seed is cultivated: slant strains 1 articulating after step (1) activation is entered in 500 ml shake flasks to 100 milliliters of culture medium loading amounts, 180 revs/min of rotary shaking tables, 30 DEG C of cultivation temperature, incubation time 10h;

2. secondary seed is cultivated: by first order seed, according in 500 milliliters of secondary seed shaking flasks of inoculum concentration access of 10%, condition of culture is identical with first order seed;

3. three grades of seed culture: secondary seed is accessed in 5000 milliliters of three grades of seed shaking flasks to 1000 milliliters of culture medium loading amounts, 100 revs/min of rotary shaking tables, 30 DEG C of cultivation temperature, incubation time 10h with 10% inoculum concentration;

4. first class seed pot is cultivated: the first class seed pot by three grades of seeds taking 10% inoculum concentration access total measurement (volume) as 150L, fermentation medium loading amount 100L, 30 DEG C of cultivation temperature, mixing speed 200rpm, ventilation (V/V) 1:1, tank pressure 0.05Mpa, incubation time 10h;

Described one-level, secondary, three grades of seed culture medium percentage by weights consist of:

Dusty yeast 0.3%, glucose 1%, peptone 0.3%, beef extract 0.5%, dipotassium hydrogen phosphate 0.8%, Chinese herbal medicine powder 1.5%, trehalose 1%, calcium sulfate 1g, magnesium chloride 2g, natrium citricum 1g, insufficient section pure water is supplied, 7.0,121 DEG C of sterilizing 30min of pH value.

Described seed tank culture base percentage by weight consists of:

Maltodextrin 5%, dusty yeast 0.4%, Chinese herbal medicine powder 1.5%, trehalose 1%, peptone 0.1%, corn steep liquor 0.1%, dipotassium hydrogen phosphate 0.8%, magnesium sulfate 0.05%, natrium citricum 0.1%, insufficient section pure water is supplied, 7.0,121 DEG C of sterilizing 30min of pH value.

Described seeding tank zymotic fluid cell concentration is 7.0x10 ⁸individual/ml;

(3) ferment tank

First class seed pot zymotic fluid in step (2) is accessed to fermentation tank, 30 DEG C of cultivation temperature, mixing speed 200r/m, ventilation (V/V) 1:1, incubation time 10h with 6% inoculum concentration; Then with 1 DEG C/h rate of temperature fall slow cooling to 10 DEG C, constant temperature culture 15h; Continue with 1 DEG C/h rate of temperature fall slow cooling to 2 DEG C, now, first class seed pot zymotic fluid in step (2) is appended to access fermentation tank, constant temperature culture 20h with 4% inoculum concentration; Finally slowly be warming up to 10 DEG C with 1 DEG C/h heating rate, constant temperature culture 15h; Continue to be slowly warming up to 30 DEG C with 1 DEG C/h heating rate, constant temperature culture 15h;

Dissolved oxygen control: by adjusting speed of agitator and ventilation, control dissolved oxygen 15%;

PH controls: by mending ammoniacal liquor or phosphoric acid,diluted, in controlled fermentation process, pH value remains on 7.0;

Control of additive raw material: add supplemented medium, to maintain zymotic fluid content of reducing sugar as 2-5mg/ml;

Put tank standard: the old and feeble self-dissolving of 60-80% thalline, enzyme activity increasess slowly.

Described fermentation medium consists of: maltodextrin 50g, corn flour 50g, beancake powder 15g, Chinese herbal medicine powder 30g, trehalose 30g, dusty yeast 4g, corn steep liquor 1g, ammonium sulfate 1g, dipotassium hydrogen phosphate 1g, potassium dihydrogen phosphate 1g, natrium citricum 1g, defoamer 0.1g, pure water l000mL, 7.0,121 DEG C of sterilizing 20min of pH value;

Described supplemented medium percentage by weight consists of: maltodextrin 20%, and corn flour 10%, bean powder 15%, herbal mediciment powder 5%, insufficient section pure water is supplied, 7.0,121 DEG C of sterilizing 30min of pH value.

The concocting method of described fermentation medium is:

Accurately take in proportion raw material, pure water in raw material, corn flour, beancake powder are dropped in material-compound tank, regulate pH value 7.0, add middle temperature amylase (3u/g corn flour) and alpha-amylase (30u/g corn flour), DEG C insulation 15min of warming while stirring to 70 simultaneously, is then slowly warming up to 90 DEG C of insulation 15min and liquefies, finally add other raw material, stir, adjust initial p H7.0,121 DEG C of sterilizing 30min are for subsequent use.

(4) zymotic fluid after filtration, concentrated, allotment, essence filter, the dry heat-flash stability neutral proteinase that to obtain, described allocation process adds concentrated enzyme liquid gross weight 0.5% Chinese herbal medicine powder.

Described concentrated maltase percentage by weight consists of: former maltase 35%, fragrance maltase 65%.

Described former maltase preparation method is as follows:

(1) high-pressure pulse electric (PEF) immersion treatment: by Fructus Hordei Germinatus in pre-immersion trough with 5 times of 35 DEG C of emerge in worm water 15min, make Fructus Hordei Germinatus moisture reach 30%, carry out high-pressure pulse electric (PEF) simultaneously and process, electric-field intensity 30KV/cm, burst lengths 175 μ S, pulse frequency 250Hz;

(3) ultrasonic, PEF extracts: Fructus Hordei Germinatus slurries are placed in to pill tank, and limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 1.0A/150w condition, carried out ultrasonic extraction 12min; Then carry out high-pressure pulse electric (PEF) and extract 12min, electric-field intensity 30KV/cm, burst lengths 500 μ S, pulse frequency 250Hz;

(4) filter: be 200 order diatomite in room temperature by 650 order filter cloth precoating granularities by extract, adopt flame filter press to filter, operating pressure is 0.19Mpa;

(5) ultrafiltration concentration: adopt the daltonian milipore filter circulation of molecular cut off <100000 concentrated, until concentrate enzyme content is 12 times before ultrafiltration;

(6) dry: to the protective agent of the present invention that adds concentrate weight 4% in concentrate, then dry with the special spray dryer of enzyme preparation, 155 DEG C of EATs; 80 DEG C of temperature of outgoing airs; drying time 10s, product moisture < 5%, both former maltase.

Described fragrance maltase preparation method is as follows:

(1) Fructus Hordei Germinatus moisture regain: with malt weight 3.5%, temperature is 45 DEG C of warm water even spraying Fructus Hordei Germinatus grain surfaces, puts the even 4min of mixing in mixer.

(2) cure: pack moisture regain Fructus Hordei Germinatus into drum-type and fry stove, in 12min, temperature rises to 185 DEG C, insulation 12min, is then cooled to 105 DEG C, and insulation 25min, finally takes out spreading for cooling, obtains fragrance Fructus Hordei Germinatus;

(4) ultrasonic, PEF extracts: fragrance malt flour is placed in to pill tank, adds the water of 4 times, limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 1.0A/150w condition, carried out ultrasonic extraction 12min; Then carry out high-pressure pulse electric (PEF) and extract 17min, electric-field intensity 30KV/cm, burst lengths 500 μ S, pulse frequency 250Hz;

(5) filter: adopt flame filter press to filter in room temperature 650 order filter clothes extract, operating pressure is 0.19Mpa;

(6) ultrafiltration concentration: adopt the daltonian milipore filter circulation of molecular cut off <100000 concentrated, until concentrate enzyme content is 12 times before ultrafiltration;

(7) dry: to the protective agent of the present invention that adds concentrate weight 4% in concentrate, then dry with the special spray dryer of enzyme preparation, 155 DEG C of EATs; 80 DEG C of temperature of outgoing airs; drying time 10s, product moisture < 5%, both fragrance maltase.

The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 5 times of weight, control 80 DEG C of temperature and keep 3h, then be cooled to 53 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 6.2, enzymolysis 3h, finally add the mixture of 2 times of weight ethanol of mixed material and propyl alcohol, control temperature to 69 DEG C and keep 4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then freeze drying obtains Chinese herbal medicine extract.

The parts by weight of described Chinese herbal medicine consist of: 25 parts of sea-buckthorns, 25 parts of cassia seeds, 17 parts of matrimony vines, 13 parts of Chinese yams, 8 parts of radix glehniaes, 7 parts of fruits of negundo, 4 parts of radix polygonati officinalis, 4 parts of the seeds of Job's tears, 4 parts of fructus hordei germinatus, 4 parts of sweet osmanthus, 4 parts of the Radixs Astragali;

Mixed enzyme addition is the 5-10% of mixed material gross weight;

The parts by weight of described mixed enzyme consist of: 15 parts of endo-beta-glucanases, 15 parts of outer 1,4 beta-glucanases, 13 parts of beta-glucosidases, 13 parts of zytases, 13 parts of pentosanases, 25 parts of Pullulanases, 12 parts of beta amylases, 13 parts of neutral proteinases, 13 parts of acid proteases, 7 parts of superoxide dismutases, 7 parts of glucose oxidases, 7 parts of acid phosphatases;

Described protective agent is made up of the raw material of following parts by weight: 25 parts of trehaloses, NaCl25 part, (NH ₄) SO ₄13 parts, 12 parts of cysteines.

Described activator is evenly to be mixed by the inorganic salts of following quality component: 35 parts of zinc chloride, 15 parts, calcium chloride, 15 parts, sodium sulphate, 7 parts, magnesium chloride.

Contain the preparation method of the feed complex enzyme of neutral proteinase:

Embodiment 2:

30 parts of concentrated maltases, 20 parts of acid proteases, 20 parts of neutral proteinases, 20 parts of acidic xylanases, 20 parts of cellulases; 15 parts of 1,4 beta-glucanases, 15 parts of pectases, 15 parts of amylase, 10 parts of seminases; 10 parts of phytases, 10 parts of Chinese herbal medicine extracts, 10 parts of protective agents, 10 parts of activator.

The preparation method of described neutral proteinase comprises the steps:

(1) actication of culture

The slant strains of intact bacillus subtilis 1398-2-12 is inoculated in to slant medium, cultivates 30h for 33 DEG C and carry out actication of culture, so activate 2 times;

Described slant medium consists of: beef extract 6g, and sodium chloride 8g, peptone 15g, glucose 4g, (NH) ₂sO ₄4g, K ₂hPO ₄7g, CaCl ₂2g, agar 18g, Chinese herbal medicine powder 8g, distilled water l000mL, 7.2,121 DEG C of sterilizing 20min of pH value;

Take 25 parts of the Radixs Astragali; 16 parts of Radix Codonopsis; 12 parts of radix bupleuri; 12 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 5 times of weight, control 80 DEG C of temperature and keep 3h, be then cooled to 50 DEG C, add the mixing enzyme preparation of mixed material gross weight 8% to carry out enzymolysis, with newborn acid for adjusting pH value be 6.0, enzymolysis 3h, finally adds the mixture of 2 times of weight ethanol of mixed material and propyl alcohol, and the mass ratio of described ethanol and propyl alcohol is 1:1.2, control temperature to 70 DEG C and keep 4h, filter; Filtrate Vacuum Concentration postlyophilization obtains Chinese herbal medicine powder.

The parts by weight of described mixed enzyme consist of: 15 parts of endo-beta-glucanases, 15 parts of outer 1,4 beta-glucanases, 12 parts of beta-glucosidases, 18 parts of zytases, 18 parts of pentosanases, 25 parts of Pullulanases, 12 parts of beta amylases, 12 parts of neutral proteinases, 12 parts of acid proteases, 8 parts of superoxide dismutases, 8 parts of glucose oxidases, 8 parts of acid phosphatases.

(2) liquid seeds expands cultivation

1. first order seed is cultivated: slant strains 2 articulatings after step (1) activation are entered in 500 ml shake flasks to 100 milliliters of culture medium loading amounts, 180 revs/min of rotary shaking tables, 33 DEG C of cultivation temperature, incubation time 12h;

3. three grades of seed culture: secondary seed is accessed in 5000 milliliters of three grades of seed shaking flasks to 1000 milliliters of culture medium loading amounts, 100 revs/min of rotary shaking tables, 33 DEG C of cultivation temperature, incubation time 12h with 10% inoculum concentration;

4. first class seed pot is cultivated: the first class seed pot by three grades of seeds taking 10% inoculum concentration access total measurement (volume) as 150L, fermentation medium loading amount 100L, 33 DEG C of cultivation temperature, mixing speed 300rpm, ventilation (V/V) 1:1.5, tank pressure 0.05Mpa, incubation time 15h;

Dusty yeast 0.4%, glucose 1.2%, peptone 0.4%, beef extract 0.6%, dipotassium hydrogen phosphate 1.0%, Chinese herbal medicine powder 1.8%, trehalose 2%, calcium sulfate 1g, magnesium chloride 2g, natrium citricum 2g, insufficient section pure water is supplied, 7.2,121 DEG C of sterilizing 30min of pH value.

Described seed tank culture base percentage by weight consists of:

Maltodextrin 10%, dusty yeast 0.5%, Chinese herbal medicine powder 1.8%, trehalose 2%, peptone 0.2%, corn steep liquor 0.3%, dipotassium hydrogen phosphate 1.0%, magnesium sulfate 0.08%, natrium citricum 0.3%, insufficient section pure water is supplied, 7.2,121 DEG C of sterilizing 30min of pH value.

Described seeding tank zymotic fluid cell concentration is 7.5x10 ⁸individual/ml;

(3) ferment tank

First class seed pot zymotic fluid in step (2) is accessed to fermentation tank, 35 DEG C of cultivation temperature, mixing speed 400r/m, ventilation (V/V) 1:2, incubation time 12h with 6% inoculum concentration; Then with 2 DEG C/h rate of temperature fall slow cooling to 12 DEG C, constant temperature culture 18h; Continue with 2 DEG C/h rate of temperature fall slow cooling to 4 DEG C, now, first class seed pot zymotic fluid in step (2) is appended to access fermentation tank, constant temperature culture 25h with 4% inoculum concentration; Finally slowly be warming up to 12 DEG C with 2 DEG C/h heating rate, constant temperature culture 18h; Continue to be slowly warming up to 33 DEG C with 2 DEG C/h heating rate, constant temperature culture 18h;

Dissolved oxygen control: by adjusting speed of agitator and ventilation, control dissolved oxygen 20%;

PH controls: by mending ammoniacal liquor or phosphoric acid,diluted, in controlled fermentation process, pH value remains on 7.2;

Described fermentation medium consists of: maltodextrin 100g, corn flour 55g, beancake powder 20g, Chinese herbal medicine powder 40g, trehalose 35g, dusty yeast 6g, corn steep liquor 3g, ammonium sulfate 2g, dipotassium hydrogen phosphate 2g, potassium dihydrogen phosphate 2g, natrium citricum 3g, defoamer 0.5g, pure water l000mL, 7.2,121 DEG C of sterilizing 20min of pH value;

Described supplemented medium percentage by weight consists of: maltodextrin 25%, and corn flour 15%, bean powder 20%, herbal mediciment powder 8%, insufficient section pure water is supplied, 7.2,121 DEG C of sterilizing 30min of pH value.

The concocting method of described fermentation medium is:

Accurately take in proportion raw material, pure water in raw material, corn flour, beancake powder are dropped in material-compound tank, regulate pH value 7.2, add middle temperature amylase (3u/g corn flour) and alpha-amylase (30u/g corn flour), DEG C insulation 20min of warming while stirring to 70 simultaneously, is then slowly warming up to 90 DEG C of insulation 20min and liquefies, finally add other raw material, stir, adjust initial p H7.2,121 DEG C of sterilizing 30min are for subsequent use.

(4) zymotic fluid after filtration, concentrated, allotment, essence filter, the dry heat-flash stability neutral proteinase that to obtain, described allocation process adds concentrated enzyme liquid gross weight 3% Chinese herbal medicine powder.

Described concentrated maltase percentage by weight consists of: former maltase 30%, fragrance maltase 70%.

Described former maltase preparation method is as follows:

(1) high-pressure pulse electric (PEF) immersion treatment: by Fructus Hordei Germinatus in pre-immersion trough with 3 times of 20 DEG C of emerge in worm water 10min, make Fructus Hordei Germinatus moisture reach 25%, carry out high-pressure pulse electric (PEF) simultaneously and process, electric-field intensity 20KV/cm, burst lengths 150 μ S, pulse frequency 200Hz;

(3) ultrasonic, PEF extracts: Fructus Hordei Germinatus slurries are placed in to pill tank, and limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 0.6A/25w condition, carried out ultrasonic extraction 10min; Then carry out high-pressure pulse electric (PEF) and extract 15min, electric-field intensity 20KV/cm, burst lengths 400 μ S, pulse frequency 200Hz;

(4) filter: be 100 order diatomite in room temperature by 500 order filter cloth precoating granularities by extract, adopt flame filter press to filter, operating pressure is 0.14Mpa;

(5) ultrafiltration concentration: adopt the daltonian milipore filter circulation of molecular cut off <100000 concentrated, until concentrate enzyme content is 10 times before ultrafiltration;

(6) dry: to the protective agent of the present invention that adds concentrate weight 3% in concentrate, then dry with the special spray dryer of enzyme preparation, 150 DEG C of EATs; 75 DEG C of temperature of outgoing airs; drying time 5s, product moisture < 5%, both former maltase.

Described fragrance maltase preparation method is as follows:

(1) Fructus Hordei Germinatus moisture regain: with malt weight 3%, temperature is 40 DEG C of warm water even spraying Fructus Hordei Germinatus grain surfaces, puts the even 3min of mixing in mixer.

(2) cure: pack moisture regain Fructus Hordei Germinatus into drum-type and fry stove, in 105min, temperature rises to 170 DEG C, insulation 10min, is then cooled to 100 DEG C, and insulation 20min, finally takes out spreading for cooling, obtains fragrance Fructus Hordei Germinatus;

(4) ultrasonic, PEF extracts: fragrance malt flour is placed in to pill tank, adds the water of 3 times, limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 0.6A/25w condition, carried out ultrasonic extraction 10min; Then carry out high-pressure pulse electric (PEF) and extract 15min, electric-field intensity 20KV/cm, burst lengths 400 μ S, pulse frequency 200Hz;

(5) filter: adopt flame filter press to filter in room temperature 500 order filter clothes extract, operating pressure is 0.14Mpa;

(7) dry: to the protective agent of the present invention that adds concentrate weight 3% in concentrate, then dry with the special spray dryer of enzyme preparation, 150 DEG C of EATs; 75 DEG C of temperature of outgoing airs; drying time 5s, product moisture < 5%, both fragrance maltase.

The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3 times of weight, control temperature 70 C and keep 2h, then be cooled to 45 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 5.5, enzymolysis 2h, finally add the mixture of 0.5 times of weight ethanol of mixed material and propyl alcohol, control temperature to 60 DEG C and keep 3h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then freeze drying obtains Chinese herbal medicine extract.

The parts by weight of described Chinese herbal medicine consist of: 20 parts of sea-buckthorns, 20 parts of cassia seeds, 10 parts of matrimony vines, 10 parts of Chinese yams, 5 parts of radix glehniaes, 5 parts of fruits of negundo, 3 parts of radix polygonati officinalis, 3 parts of the seeds of Job's tears, 3 parts of fructus hordei germinatus, 3 parts of sweet osmanthus, 3 parts of the Radixs Astragali;

Mixed enzyme addition is 5% of mixed material gross weight;

The parts by weight of described mixed enzyme consist of: 10 parts of endo-beta-glucanases, 10 parts of outer 1,4 beta-glucanases, 10 parts of beta-glucosidases, 15 parts of zytases, 15 parts of pentosanases, 20 parts of Pullulanases, 10 parts of beta amylases, 10 parts of neutral proteinases, 10 parts of acid proteases, 5 parts of superoxide dismutases, 5 parts of glucose oxidases, 5 parts of acid phosphatases;

Described protective agent is made up of the raw material of following parts by weight: 20 parts of trehaloses, NaCl20 part, (NH ₄) SO ₄10 parts, 10 parts of cysteines.

Described activator is evenly to be mixed by the inorganic salts of following quality component: 30 parts of zinc chloride, 10 parts, calcium chloride, 10 parts, sodium sulphate, 5 parts, magnesium chloride.

Containing the preparation method of the feed complex enzyme of neutral proteinase with embodiment 1.

Embodiment 3:

40 parts of concentrated maltases, 30 parts of acid proteases, 30 parts of neutral proteinases, 30 parts of acidic xylanases, 30 parts of cellulases; 20 parts of 1,4 beta-glucanases, 20 parts of pectases, 0 part of amylase 2,15 parts of seminases; 15 parts of phytases, 15 parts of Chinese herbal medicine extracts, 15 parts of protective agents, 15 parts of activator.

The preparation method of described neutral proteinase comprises the steps:

(1) actication of culture

The slant strains of intact bacillus subtilis 1398-2-12 is inoculated in to slant medium, cultivates 36h for 36 DEG C and carry out actication of culture, so activate 3 times;

Described slant medium consists of: beef extract 10g, and sodium chloride 12g, peptone 20g, glucose 5g, (NH) ₂sO ₄5g, K ₂hPO ₄8g, CaCl ₂3g, agar 20g, Chinese herbal medicine powder 10g, distilled water l000mL, 7.2,121 DEG C of sterilizing 20min of pH value;

Take 30 parts of the Radixs Astragali; 18 parts of Radix Codonopsis; 15 parts of radix bupleuri; 15 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 6 times of weight, control 90 DEG C of temperature and keep 4h, be then cooled to 60 DEG C, add the mixing enzyme preparation of mixed material gross weight 10% to carry out enzymolysis, with newborn acid for adjusting pH value be 6.8, enzymolysis 4h, finally adds the mixture of 3 times of weight ethanol of mixed material and propyl alcohol, and the mass ratio of described ethanol and propyl alcohol is 1:1.5, control temperature to 78 DEG C and keep 4h, filter; Filtrate Vacuum Concentration postlyophilization obtains Chinese herbal medicine powder.

The parts by weight of described mixed enzyme consist of: 20 parts of endo-beta-glucanases, 20 parts of outer 1,4 beta-glucanases, 15 parts of beta-glucosidases, 20 parts of zytases, 20 parts of pentosanases, 30 parts of Pullulanases, 15 parts of beta amylases, 15 parts of neutral proteinases, 15 parts of acid proteases, 10 parts of superoxide dismutases, 10 parts of glucose oxidases, 10 parts of acid phosphatases.

(2) liquid seeds expands cultivation

1. first order seed is cultivated: slant strains 2 articulatings after step (1) activation are entered in 500 ml shake flasks to 100 milliliters of culture medium loading amounts, 180 revs/min of rotary shaking tables, 36 DEG C of cultivation temperature, incubation time 15h;

3. three grades of seed culture: secondary seed is accessed in 5000 milliliters of three grades of seed shaking flasks to 1000 milliliters of culture medium loading amounts, 100 revs/min of rotary shaking tables, 36 DEG C of cultivation temperature, incubation time 15h with 10% inoculum concentration;

4. first class seed pot is cultivated: the first class seed pot by three grades of seeds taking 10% inoculum concentration access total measurement (volume) as 150L, fermentation medium loading amount 100L, 36 DEG C of cultivation temperature, mixing speed 400rpm, ventilation (V/V) 1:2, tank pressure 0.05Mpa, incubation time 20h;

Dusty yeast 0.5%, glucose 1.5%, peptone 0.5%, beef extract 0.8%, dipotassium hydrogen phosphate 1.5%, Chinese herbal medicine powder 2%, trehalose 3%, calcium sulfate 1g, magnesium chloride 2g, natrium citricum 3g, insufficient section pure water is supplied, 7.2,123 DEG C of sterilizing 40min of pH value.

Described seed tank culture base percentage by weight consists of:

Maltodextrin 15%, yeast 0.8%, Chinese herbal medicine powder 2%, trehalose 3%, peptone 0.5%, corn steep liquor 0.5%, dipotassium hydrogen phosphate 1.5%, magnesium sulfate 0.1%, natrium citricum 0.5%, insufficient section pure water is supplied, 7.2,123 DEG C of sterilizing 40min of pH value.

Described seeding tank zymotic fluid cell concentration is 8.0x10 ⁸individual/ml;

(3) ferment tank

First class seed pot zymotic fluid in step (2) is accessed to fermentation tank, 36 DEG C of cultivation temperature, mixing speed 700r/m, ventilation (V/V) 1:3, incubation time 15h with 6% inoculum concentration; Then with 2 DEG C/h rate of temperature fall slow cooling to 15 DEG C, constant temperature culture 20h; Continue with 2 DEG C/h rate of temperature fall slow cooling to 5 DEG C, now, first class seed pot zymotic fluid in step (2) is appended to access fermentation tank, constant temperature culture 30h with 4% inoculum concentration; Finally slowly be warming up to 15 DEG C with 2 DEG C/h heating rate, constant temperature culture 20h; Continue to be slowly warming up to 36 DEG C with 2 DEG C/h heating rate, constant temperature culture 20h;

Dissolved oxygen control: by adjusting speed of agitator and ventilation, control dissolved oxygen 30%;

Described fermentation medium consists of: maltodextrin 150g, corn flour 60g, beancake powder 25g, Chinese herbal medicine powder 50g, trehalose 40g, dusty yeast 8g, corn steep liquor 5g, ammonium sulfate 3g, dipotassium hydrogen phosphate 2g, potassium dihydrogen phosphate 2g, natrium citricum 5g, defoamer 1g, pure water l000mL, 7.2,121 DEG C of sterilizing 20min of pH value;

Described supplemented medium percentage by weight consists of: maltodextrin 30%, and corn flour 20%, bean powder 25%, herbal mediciment powder 10%, insufficient section pure water is supplied, 7.2,123 DEG C of sterilizing 40min of pH value.

The concocting method of described fermentation medium is:

Accurately take in proportion raw material, pure water in raw material, corn flour, beancake powder are dropped in material-compound tank, regulate pH value 7.2, add middle temperature amylase (3u/g corn flour) and alpha-amylase (30u/g corn flour), DEG C insulation 30min of warming while stirring to 70 simultaneously, is then slowly warming up to 90 DEG C of insulation 30min and liquefies, finally add other raw material, stir, adjust initial p H7.2,123 DEG C of sterilizing 40min are for subsequent use.

(4) zymotic fluid after filtration, concentrated, allotment, essence filter, the dry heat-flash stability neutral proteinase that to obtain, described allocation process adds concentrated enzyme liquid gross weight 5% Chinese herbal medicine powder.

Described concentrated maltase percentage by weight consists of: former maltase 40%, fragrance maltase 60%.

Described former maltase preparation method is as follows:

(1) high-pressure pulse electric (PEF) immersion treatment: by Fructus Hordei Germinatus in pre-immersion trough with 6 times of 50 DEG C of emerge in worm water 20min, make Fructus Hordei Germinatus moisture reach 35%, carry out high-pressure pulse electric (PEF) simultaneously and process, electric-field intensity 40KV/cm, burst lengths 200 μ S, pulse frequency 300Hz;

(3) ultrasonic, PEF extracts: Fructus Hordei Germinatus slurries are placed in to pill tank, and limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 1.5A/275w condition, carried out ultrasonic extraction 15min; Then carry out high-pressure pulse electric (PEF) and extract 20min, electric-field intensity 40KV/cm, burst lengths 600 μ S, pulse frequency 300Hz;

(4) filter: be 300 order diatomite in room temperature by 800 order filter cloth precoating granularities by extract, adopt flame filter press to filter, operating pressure is 0.24Mpa;

(5) ultrafiltration concentration: adopt the daltonian milipore filter circulation of molecular cut off <100000 concentrated, until concentrate enzyme content is 15 times before ultrafiltration;

(6) dry: to the protective agent of the present invention that adds concentrate weight 5% in concentrate, then dry with the special spray dryer of enzyme preparation, 160 DEG C of EATs; 85 DEG C of temperature of outgoing airs; drying time 15s, product moisture < 5%, both former maltase.

Described fragrance maltase preparation method is as follows:

(1) Fructus Hordei Germinatus moisture regain: with malt weight 4%, temperature is 50 DEG C of warm water even spraying Fructus Hordei Germinatus grain surfaces, puts the even 5min of mixing in mixer.

(2) cure: pack moisture regain Fructus Hordei Germinatus into drum-type and fry stove, in 15min, temperature rises to 200 DEG C, insulation 15min, is then cooled to 110 DEG C, and insulation 30min, finally takes out spreading for cooling, obtains fragrance Fructus Hordei Germinatus;

(4) ultrasonic, PEF extracts: fragrance malt flour is placed in to pill tank, adds the water of 6 times, limit is slowly stirred limit probe type ultrasonic extraction apparatus and under current strength 1.5A/275w condition, carried out ultrasonic extraction 15min; Then carry out high-pressure pulse electric (PEF) and extract 20min, electric-field intensity 40KV/cm, burst lengths 600 μ S, pulse frequency 300Hz;

(5) filter: adopt flame filter press to filter in room temperature 800 order filter clothes extract, operating pressure is 0.14-0.24Mpa;

(7) dry: to the protective agent of the present invention that adds concentrate weight 5% in concentrate, then dry with the special spray dryer of enzyme preparation, 160 DEG C of EATs; 85 DEG C of temperature of outgoing airs; drying time 15s, product moisture < 5%, both fragrance maltase.

The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 6 times of weight, control 90 DEG C of temperature and keep 4h, then be cooled to 60 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 6.8, enzymolysis 4h, finally add the mixture of 3 times of weight ethanol of mixed material and propyl alcohol, control temperature to 78 DEG C and keep 4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then freeze drying obtains Chinese herbal medicine extract.

The parts by weight of described Chinese herbal medicine consist of: 30 parts of sea-buckthorns, 30 parts of cassia seeds, 15 parts of matrimony vines, 15 parts of Chinese yams, 10 parts of radix glehniaes, 10 parts of fruits of negundo, 5 parts of radix polygonati officinalis, 5 parts of the seeds of Job's tears, 5 parts of fructus hordei germinatus, 5 parts of sweet osmanthus, 5 parts of the Radixs Astragali;

Mixed enzyme addition is 10% of mixed material gross weight;

The parts by weight of described mixed enzyme consist of: 20 parts of endo-beta-glucanases, 20 parts of outer 1,4 beta-glucanases, 15 parts of beta-glucosidases, 20 parts of zytases, 20 parts of pentosanases, 30 parts of Pullulanases, 15 parts of beta amylases, 15 parts of neutral proteinases, 15 parts of acid proteases, 10 parts of superoxide dismutases, 10 parts of glucose oxidases, 10 parts of acid phosphatases;

Described protective agent is made up of the raw material of following parts by weight: 30 parts of trehaloses, NaCl30 part, (NH ₄) SO ₄15 parts, 15 parts of cysteines.

Described activator is evenly to be mixed by the inorganic salts of following quality component: 40 parts of zinc chloride, 20 parts, calcium chloride, 20 parts, sodium sulphate, 10 parts, magnesium chloride.

Claims

1. containing a feed complex enzyme for neutral proteinase, formed by the enzyme preparation of following parts by weight: concentrated maltase 30-40 part, acid protease 20-30 part, neutral proteinase 20-30 part, acidic xylanase 20-30 part, cellulase 20-30 part, 1,4 beta-glucanase 15-20 part, pectase 15-20 part, amylase 15-20 part, seminase 10-15 part, phytase 10-15 part, Chinese herbal medicine extract 10-15 part, protective agent 10-15 part, activator 10-15 part;

The preparation method of described neutral proteinase is as follows: bacillus subtilis 1398-2-12 through actication of culture and step by step expand cultivate obtain liquid seeds; Liquid seeds is accessed to fermentation tank, cultivation temperature 30-36 DEG C, mixing speed 200-700r/m, ventilation (V/V) 1:1-3, incubation time 10-15h with 6% inoculum concentration; Then with 1-2 DEG C/h rate of temperature fall slow cooling to 10-15 DEG C, constant temperature culture 15-20h; Continuation to 2-5 DEG C, now, is appended access fermentation tank, constant temperature culture 20-30h by liquid seeds with 4% inoculum concentration with 1-2 DEG C/h rate of temperature fall slow cooling; Finally slowly be warming up to 10-15 DEG C, constant temperature culture 15-20h with 1-2 DEG C/h heating rate; Continue to be slowly warming up to 30-36 DEG C, constant temperature culture 15-20h with 1-2 DEG C/h heating rate; Zymotic fluid after filtration, concentrated, allotment, essence filter, the dry solid neutral proteinase that to obtain; Described allocation process adds concentrated enzyme liquid gross weight 0.5-5% Chinese herbal medicine powder.

2. a kind of feed complex enzyme containing neutral proteinase as claimed in claim 1, is characterized in that, the slant medium of preparing neutral proteinase consists of: beef extract 3-10g, and sodium chloride 5-12g, peptone 10-20g, glucose 2-5g, (NH) ₂sO ₄3-5g, K ₂hPO ₄6-8g, CaCl ₂1-3g, agar 15-20g, Chinese herbal medicine powder 5-10g, distilled water l000mL, pH value 7.0-7.2.

3. a kind of feed complex enzyme containing neutral proteinase as claimed in claim 1, is characterized in that, the seed culture medium percentage by weight of preparing neutral proteinase consists of: dusty yeast 0.3-0.5%, glucose 1-1.5%, peptone 0.3-0.5%, beef extract 0.5-0.8%, dipotassium hydrogen phosphate 0.8-1.5%, Chinese herbal medicine powder 1.5-2%, trehalose 1-3%, calcium sulfate 1g, magnesium chloride 2g, natrium citricum 1-3g, insufficient section pure water is supplied, pH value 7.0-7.2.

4. a kind of feed complex enzyme containing neutral proteinase as claimed in claim 1, is characterized in that, the seed tank culture base of preparing neutral proteinase consists of: maltodextrin 5-15%, dusty yeast 0.4-0.8%, Chinese herbal medicine powder 1.5-2%, trehalose 1-3%, peptone 0.1-0.5%, corn steep liquor 0.1-0.5%, dipotassium hydrogen phosphate 0.8-1.5%, magnesium sulfate 0.05-0.1%, natrium citricum 0.1-0.5%, insufficient section pure water is supplied, pH value 7.0-7.2.

5. a kind of feed complex enzyme containing neutral proteinase as claimed in claim 1, is characterized in that, while preparing neutral proteinase, seeding tank zymotic fluid cell concentration is 7.0x10 ⁸-8.0x10 ⁸individual/ml.

6. a kind of feed complex enzyme containing neutral proteinase as claimed in claim 1, is characterized in that, the fermentation medium of preparing neutral proteinase consists of: maltodextrin 50-150g, corn flour 50-60g, beancake powder 15-25g, Chinese herbal medicine powder 30-50g, trehalose 30-40g, dusty yeast 4-8g, corn steep liquor 1-5g, ammonium sulfate 1-3g, dipotassium hydrogen phosphate 1-2g, potassium dihydrogen phosphate 1-2g, natrium citricum 1-5g, defoamer 0.1-1g, pure water l000mL, pH value 7.0-7.2.

7. the preparation method containing the feed complex enzyme of neutral proteinase; it is characterized in that; by described protective agent, Chinese herbal medicine extract, concentrated maltase ultramicro grinding respectively; guarantee that granularity is less than described neutral proteinase and other enzyme preparation; then with neutral proteinase, acid protease, acidic xylanase, cellulase, 1,4 beta-glucanase, pectase, amylase, seminase, phytase; evenly mix; finally add activator, after mixing, pack the feed complex enzyme getting product containing neutral proteinase.

8. the preparation method of a kind of feed complex enzyme containing neutral proteinase as claimed in claim 7, it is characterized in that, the described feed complex enzyme containing neutral proteinase is made up of the enzyme preparation of following parts by weight: concentrated maltase 30-40 part, acid protease 20-30 part, neutral proteinase 20-30 part, acidic xylanase 20-30 part, cellulase 20-30 part, 1,4 beta-glucanase 15-20 part, pectase 15-20 part, amylase 15-20 part, seminase 10-15 part, phytase 10-15 part, Chinese herbal medicine extract 10-15 part, protective agent 10-15 part, activator 10-15 part,

9. the preparation method of a kind of feed complex enzyme containing neutral proteinase as claimed in claim 8, it is characterized in that, the preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3-6 times of weight, control temperature 70 C-90 DEG C and keep 2-4h, then be cooled to 45-60 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 5.5-6.8, enzymolysis 2-4h, finally add the mixture of mixed material 0.5-3 times of weight ethanol and propyl alcohol, control temperature to 60 DEG C-78 DEG C of maintenance 3-4h, filter, it is more than 20% that filtrate decompression is concentrated into solid content, and then freeze drying obtains Chinese herbal medicine extract,

Described mixed enzyme addition is the 5-10% of mixed material gross weight;

10. the preparation method of a kind of feed complex enzyme containing neutral proteinase as claimed in claim 8, it is characterized in that, the described feed complex enzyme containing neutral proteinase, enzyme preparation by following parts by weight forms: 35 parts of concentrated maltases, 25 parts of acid proteases, 25 parts of neutral proteinases, 25 parts of acidic xylanases, 25 parts of cellulases, 17 parts of 1,4 beta-glucanases, 17 parts of pectases, 17 parts of amylase, 12 parts of seminases, 13 parts of phytases, 12 parts of Chinese herbal medicine extracts, 13 parts of protective agents, 12 parts of activator;

The preparation method of described neutral proteinase comprises the steps:

Take 20 parts of the Radixs Astragali; 10 parts of Radix Codonopsis; 10 parts of radix bupleuri; 10 parts of the roots of large-flowered skullcap; Respectively said herbal medicine being crushed to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 3 times of weight, control temperature 70 C and keep 2h, be then cooled to 45 DEG C, add the mixing enzyme preparation of mixed material gross weight 5% to carry out enzymolysis, with newborn acid for adjusting pH value be 5.5, enzymolysis 2h, finally adds the mixture of 0.5 times of weight ethanol of mixed material and propyl alcohol, and the mass ratio of described ethanol and propyl alcohol is 1:1, control temperature to 60 DEG C and keep 3h, filter; Filtrate Vacuum Concentration postlyophilization obtains Chinese herbal medicine powder;

(2) liquid seeds expands cultivation

Dusty yeast 0.3%, glucose 1%, peptone 0.3%, beef extract 0.5%, dipotassium hydrogen phosphate 0.8%, Chinese herbal medicine powder 1.5%, trehalose 1%, calcium sulfate 1g, magnesium chloride 2g, natrium citricum 1g, insufficient section pure water is supplied, 7.0,121 DEG C of sterilizing 30min of pH value;

Described seed tank culture base percentage by weight consists of:

Maltodextrin 5%, dusty yeast 0.4%, Chinese herbal medicine powder 1.5%, trehalose 1%, peptone 0.1%, corn steep liquor 0.1%, dipotassium hydrogen phosphate 0.8%, magnesium sulfate 0.05%, natrium citricum 0.1%, insufficient section pure water is supplied, 7.0,121 DEG C of sterilizing 30min of pH value;

(3) ferment tank

Described supplemented medium percentage by weight consists of: maltodextrin 20%, and corn flour 10%, bean powder 15%, herbal mediciment powder 5%, insufficient section pure water is supplied, 7.0,121 DEG C of sterilizing 30min of pH value;

The concocting method of described fermentation medium is:

Accurately take in proportion raw material, pure water in raw material, corn flour, beancake powder are dropped in material-compound tank, regulate pH value 7.0, add middle temperature amylase (3u/g corn flour) and alpha-amylase (30u/g corn flour), DEG C insulation 15min of warming while stirring to 70 simultaneously, is then slowly warming up to 90 DEG C of insulation 15min and liquefies, finally add other raw material, stir, adjust initial p H7.0,121 DEG C of sterilizing 30min are for subsequent use;

(4) zymotic fluid after filtration, concentrated, allotment, essence filter, the dry heat-flash stability neutral proteinase that to obtain, described allocation process adds concentrated enzyme liquid gross weight 0.5% Chinese herbal medicine powder;

Described concentrated maltase percentage by weight consists of: former maltase 35%, fragrance maltase 65%;

Described former maltase preparation method is as follows:

(6) dry: to the protective agent of the present invention that adds concentrate weight 4% in concentrate, then dry with the special spray dryer of enzyme preparation, 155 DEG C of EATs, 80 DEG C of temperature of outgoing airs, drying time 10s, product moisture < 5%, both former maltase;

Described fragrance maltase preparation method is as follows:

(1) Fructus Hordei Germinatus moisture regain: with malt weight 3.5%, temperature is 45 DEG C of warm water even spraying Fructus Hordei Germinatus grain surfaces, puts the even 4min of mixing in mixer;

(7) dry: to the protective agent of the present invention that adds concentrate weight 4% in concentrate, then dry with the special spray dryer of enzyme preparation, 155 DEG C of EATs, 80 DEG C of temperature of outgoing airs, drying time 10s, product moisture < 5%, both fragrance maltase;

The preparation method of described Chinese herbal medicine extract is: respectively Chinese herbal medicine powder being broken to particle diameter is below 2 millimeters, then in container, evenly mix and add the water of 5 times of weight, control 80 DEG C of temperature and keep 3h, then be cooled to 53 DEG C, add mixing enzyme preparation to carry out enzymolysis, with newborn acid for adjusting pH value be 6.2, enzymolysis 3h, finally add the mixture of 2 times of weight ethanol of mixed material and propyl alcohol, control temperature to 69 DEG C and keep 4h, filter; It is more than 20% that filtrate decompression is concentrated into solid content, and then freeze drying obtains Chinese herbal medicine extract;

Mixed enzyme addition is the 5-10% of mixed material gross weight;

Described protective agent is made up of the raw material of following parts by weight: 25 parts of trehaloses, NaCl25 part, (NH ₄) SO ₄13 parts, 12 parts of cysteines;

Described activator is evenly to be mixed by the inorganic salts of following quality component: 35 parts of zinc chloride, 15 parts, calcium chloride, 15 parts, sodium sulphate, 7 parts, magnesium chloride;