CN103911325B - Bacillus coagulans, lactobacterium casei, plant lactobacillus probiotic agent and preparation - Google Patents
Bacillus coagulans, lactobacterium casei, plant lactobacillus probiotic agent and preparation Download PDFInfo
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Abstract
A kind of solid fermentation probiotic agent, belongs to feed microbe technical field.Relate to one with dregs of beans, wheat bran, Semen Maydis powder for matrix, the utilisation technology of solid state fermentation is carried out, wherein Bacillus licheniformis (Bacillus with Bacillus coagulans HM-09, Bacillus licheniformis BL-09, lactobacterium casei HM-09 and plant lactobacillus HM-10? does is licheniformis) BL-09 deposit number CGMCC? No.5686.The present invention adds Bacillus licheniformis BL-09 continuation fermentation 20-28h after passing through that Bacillus coagulans, lactobacterium casei and plant lactobacillus are accessed solid medium mixed fermentation 18-28h after, the product of gained has special sour fragrance, and the viable count of dregs of beans, wheat bran and fermentation of corn starch product is high, the product that the degradation rate of dregs of beans protein is high.This preparation method's production cost is low, can increase economic efficiency and simple to operate.
Description
Technical field
The invention belongs to feed microbe technical field, relate to one with dregs of beans, wheat bran, Semen Maydis powder for matrix, carry out the utilisation technology of solid state fermentation with probiotic bacterium Bacillus coagulans (Bacilluscoagulans) HM-09, Bacillus licheniformis (Bacilluslicheniformis) BL-09, lactobacterium casei (Lactobacilluscasei) HM-09 and plant lactobacillus (Lactobacillusplantarum) HM-10, this technology can be widely used in the aspects such as animal probiotics micro-ecological.
Background technology
At present, probiotic composition many employings probiotic bacterium directly with liquid cow's milk and soya-bean milk fermentation, obtain functional yogurt milk or soybean milk yoghurt, all the other then adopt liquid state fermentation, and directly collection probiotic bacterium thalline, is made into probiotic bacterium pulvis, tablet or capsule.Adopt liquid state fermentation can produce a large amount of waste water, environmental pollution is comparatively large, and aftertreatment is more difficult, machinery equipment complex operation.Therefore the application adopts Bacillus coagulans (Bacilluscoagulans) HM-09, Bacillus licheniformis (Bacilluslicheniformis) BL-09, lactobacterium casei (Lactobacilluscasei) HM-09 and plant lactobacillus (Lactobacillusplantarum) HM-10 mixed solid fermentation dregs of beans, wheat bran and Semen Maydis powder, adopt solid state fermentation, its production cost is low, the fund of investment is less, convenient in downstream processing, pollute little, device structure is simple, easy to operate.
Employing Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 carry out mixed solid fermentation dregs of beans, can produce a kind of fermented product with sour fragrance taste, improve the palatability of animal-feed.Its protein is broken down into little peptide and total free aminoacids, is conducive to animal intestine peptic digestion and absorbs.Such as the patent of invention that application number is " 20041000612.5 " discloses probiotics of a kind of stability and safety and its production and use, be used for the treatment of or the pre-intestinal tract disease such as anti-diarrhea, constipation; The patent of invention that application number is " 201210070157.1 " discloses probiotics with probiotic bacterium and organoselenium dual function and preparation method thereof, the probiotic bacterium live body selenium probiotics with probiotic bacterium and organoselenium dual function can be widely used in livestock and poultry cultivation, culture fishery, plant husbandry, industry of planting forest or fruit tress, Edible Fungi, herbal medicine cultivation, produces organic Se-rich functional foodstuff; Also such as the patent of invention that application number is " 200510045358.7 " discloses multifunctional probiotic preparation and preparation method thereof, have and suppress enteric pathogenic bacteria, enhancing body immunizing power, improve the function of efficiency of feed utilization.
Existing probiotic composition many employings probiotic bacterium directly with liquid cow's milk and soya-bean milk fermentation, obtain functional yogurt milk or soybean milk yoghurt, or after being adopted liquid state fermentation, directly collection probiotic bacterium thalline, is made into probiotic bacterium pulvis, tablet or capsule.When adopting liquid state fermentation, can produce a large amount of contaminated wastewater environment, aftertreatment is more difficult, and machinery equipment operation is also comparatively loaded down with trivial details.Because probiotic bacterium can improve host's microecological balance, improve active bacteria formulation and the meta-bolites of host health level and state of health, therefore, now the research of probiotic bacterium is goed deep into detailed-oriented very much, and be widely used in each fields such as industry, agricultural, environmental protection, family.
At present, probiotic composition is applied to animal microorganism preparation, be added in animal-feed also comparatively common; Such as, publication number is the patent of invention " a kind of effective prevention and improve the compound lactobacillus probiotics of garget " of CN103202391A, disclose a kind of by preserving number be the lactobacterium casei HM-09 of CGMCCNo.6736, preserving number is the compound micro-ecological preparation that the plant lactobacillus HM-10 of CGMCCNo.6741 and zeolite powder combine by mass fraction, can effectively reduce somatic number, prevention reduces recessive garget sickness rate; Publication number is the patent of invention " strain is for improving excellent Bacillus coagulans and the application thereof of the cultivation of inferior piglet " of CN103243043A, disclose the Bacillus coagulans that a kind of preserving number is CGMCCNo.6729, by applying this bacterial strain, effectively can improve the healthy state of piglet, improving the production performance of inferior piglet cultivation.But simultaneously when utilizing liquid state fermentation to make fermented feed, also there is the defects such as above-mentioned as high in cost, environmental pollution is high, aftertreatment is difficult, machinery equipment complex operation, and the fermented product aromatising flavour produced is not enough, palatability is poor.
Summary of the invention
The object of the invention is to adopt Bacillus coagulans HM-09, Bacillus licheniformis BL-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 to carry out mixed solid fermentation dregs of beans, wheat bran and Semen Maydis powder, there is provided a kind of with dregs of beans, wheat bran and Semen Maydis powder for matrix can be widely used in the probiotic solid fermentation technology in Tiny ecosystem field, this method can be produced has special sour fragrance, and the viable count of dregs of beans, wheat bran and fermentation of corn starch product is high, the product that the degradation rate of dregs of beans protein is high.This preparation method's production cost is low, can increase economic efficiency and simple to operate.
The present invention is achieved through the following technical solutions:
A kind of with dregs of beans, wheat bran and Semen Maydis powder for matrix can be widely used in the probiotic solid fermentation technology in Tiny ecosystem field, the cultivation and the solid state fermentation thereof that comprise Bacillus coagulans HM-09, Bacillus licheniformis BL-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 are cultivated, and obtain the solid fermentation products of probiotic bacterium.
Described Bacillus coagulans HM-09, Bacillus licheniformis BL-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 seed culture medium are: peptone 20-50g, glucose 20-50g, adds water to 1L, pH7.0-7.2.
Seed culture condition: Bacillus coagulans HM-09 and Bacillus licheniformis BL-09 32-37 DEG C, 150-180 in thermostat container turn shaking table and cultivate 20-24h; Lactobacterium casei HM-09 and plant lactobacillus HM-10 is 32-37 DEG C of cultivation 20-24h in thermostat container.
Solid-state fermentation culture medium: dregs of beans: wheat bran: Semen Maydis powder=10:8-10:0.5-2, moisture content 40-70%.
Solid state fermentation culture condition: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation temperature 32-37 DEG C, humidity 40-60%, add Bacillus licheniformis BL-09 after mixed fermentation 18-28h and continue fermentation 20-28h, temperature 34-37 DEG C, humidity 40-60%; Its free ammonical nitrogen content of fermentation ends is: 700.5umol/g; Bacillus coagulans HM-09 viable count 2.90 × 10 in fermented product
9cFU/g, lactobacterium casei HM-09 viable count 3.48 × 10
9cFU/g, plant lactobacillus HM-10 viable count 2.87 × 10
9cFU/g, Bacillus licheniformis BL-096.87 × 10
9cFU/g; Fermentation ends drying prepares microbial inoculum.
Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 4-6%(v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively 1-3%(v/w), Bacillus licheniformis BL-09 is 5-7%(v/w).
Described Bacillus licheniformis BL-09 is Bacillus licheniformis (Bacilluslicheniformis) BL-09, China Committee for Culture Collection of Microorganisms's common micro-organisms center is preserved on December 31st, 2011, preserving number is CGMCCNo.5686, and preservation address is: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City.
Bacillus licheniformis BL-09 is separated genus bacillus in the healthy ox intestinal samples in area, Hohhot City ,Inner Mongolia Autonomous Region, belongs to amphimicrobian, Gram-positive genus bacillus; Thalline is bar shape, is good at shaft-like once in a while, raw in gemma, ovalize; Optimum growth temperature is 37 ± 1 DEG C, and the most suitable growth pH is 7.0-7.4, and solid state fermentation dries product normal temperature storage 6 months, and gemma rate of loss is lower than 5%; Nutrient agar plate cultivates about 12h, bacterium colony is irregular, edge is rough, bacterium colony central authorities have obvious protruding peak, be cultured to 24-36h, bacterium colony fold obviously and in fold material be liquid, transfering loop picking has wire drawing situation, and microscopy gemma comes off substantially, Bacillus licheniformis is defined as, called after Bacillus licheniformis BL-09 through uncle's Jie Shi bacterium handbook (the 9th edition) physiological and biochemical test and bacterium, colonial morphology.Experiment in vitro confirms, this bacterial strain has the biochemical characteristic Bacillus licheniformis BL-09 such as good high temperature resistant, acidproof, bile tolerance and also has the several physiological active substances such as the multiple enzyme such as high proteinase yield, amylase, lipase and amino acid, VITAMIN, antimicrobial substance, solid medium viscosity after fermentation is very large, wire drawing is serious, bottom culture vessel, visible a small amount of liquid substance, proves that the moisture-retaining capacity in its solid state fermentation is relatively strong.
Acid resistance test:
Bacillus licheniformis BL-09 is in meat soup seed liquor, 37 DEG C, 180r/min cultivates 24h, in 4 DEG C, 4000r/min centrifugal 10min collection thalline, after 0.85% stroke-physiological saline solution rinsing once, be suspended in pH and be respectively in the buffered soln of 2.0,3.0 and 4.0, cultivate 1 ~ 3h for 37 DEG C, detection viable count changes, and result display Bacillus licheniformis BL-09 has good acid resistance.
Oven test:
Bacillus licheniformis BL-09 is in meat soup seed liquor, 37 DEG C, 180r/min cultivates 24h, in 4 DEG C, 4000r/min centrifugal 10min collection thalline, after 0.85% stroke-physiological saline solution rinsing once, the thalline of acquisition with after appropriate stroke-physiological saline solution dilution in 80 DEG C of heating in water bath process 10min to obtain gemma, then process 5 ~ 30min respectively at 80 DEG C, 90 DEG C and 100 DEG C, detection viable count changes, and result display Bacillus licheniformis BL-09 has good thermotolerance.
Bile tolerance is tested:
Bacillus licheniformis BL-09 is in meat soup seed liquor, 37 DEG C, 180r/min cultivates 24h, in 4 DEG C, 4000r/min centrifugal 10min collection thalline, after 0.85% stroke-physiological saline solution rinsing once, be suspended in and be respectively in the physiological saline of 0.03%, 0.10%, 0.30% containing gallbladder salinity, cultivate 1 ~ 6h for 37 DEG C, detection viable count changes, and result display Bacillus licheniformis BL-09 has good resistance to light salt.
Beneficial effect:
Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 etc. is adopted to carry out mixed solid fermentation dregs of beans, wheat bran and Semen Maydis powder in the present invention.The present invention adopts solid state fermentation production cost low, and the fund of investment is less, and convenient in downstream processing, pollute little, device structure is simple, easy to operate, can be widely used in the probiotic solid fermentation technology in Tiny ecosystem field; Adopt mixed fermentation, can produce the tart flavour such as lactic acid and acetic acid small-molecule substance, these small-molecule substances give improvement and the variation of the organoleptic features such as the distinctive sour fragrance of product, color and luster and mouthfeel, and leavened prod improves animal palatability; Adopt Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09, plant lactobacillus HM-10 mixed fermentation dregs of beans, wheat bran and Semen Maydis powder, the protein in dregs of beans and Semen Maydis powder can be made to be broken down into little peptide and total free aminoacids, to be beneficial to digesting and assimilating of animal stomach.Bacillus licheniformis BL-09 has this good bacterial strain and has the biochemical characteristics such as good high temperature resistant, acidproof, bile tolerance and produce the features such as multiple enzyme such as proteolytic enzyme, amylase, lipase, when making to add use in product of the present invention in feed, when this series products is used for Cow-feeding, effectively can improve the utilising efficiency 15-28% of feed.The addition of product of the present invention in feed is 20-50 gram/kg.
After product of the present invention uses, the conception rate of milk cow improves 14%, can improve the year milk yield more than 310 kilograms of milk cow as calculated.
Embodiment
Below in conjunction with example, the present invention is further described, and following example is illustrative, is not determinate, can not limit protection scope of the present invention with following example.
Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10, specific name is respectively BacilluscoagulansHM-09, LactobacilluscaseiHM-09, BacilluslicheniformisBL-09 and LactobacillusplantarumHM-10.Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 on October 29th, 2012 in the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, preserving number is respectively CGMCCNo.6729, CGMCCNo.6736 and CGMCCNo.6741, Bacillus licheniformis BL-09 is on December 31st, 2012 in the center preservation of China Committee for Culture Collection of Microorganisms's common micro-organisms, and preserving number is CGMCCNo.5686.Preservation address is: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City.
(1) the seed liquor preparation of thalline
Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 receive and are equipped with in the substratum of liquid seeds from the glycerine pipe of cold storage, described Bacillus coagulans HM-09 and Bacillus licheniformis BL-09 seed culture condition: be positioned on shaking table, rotating speed is 160-180r/min, temperature is 33-37 DEG C, cultivates 20-24h; Lactobacterium casei HM-09 and plant lactobacillus HM-10 is 30-37 DEG C of quiescent culture 20-24h in thermostat container.
Described Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 seed culture medium are (g/L): peptone 10-50, glucose 10-50, adds water to 1L, pH7.0-7.2.
(2) solid state fermentation
Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation temperature 32-37 DEG C, humidity 40-60%, add Bacillus licheniformis BL-09 after mixed fermentation 18-28h and continue fermentation 20-28h, temperature 34-37 DEG C, humidity 40-60%; Fermentation ends drying obtains.
Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 4-6%(v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively 1-3%(v/w), Bacillus licheniformis BL-09 is 5-7%(v/w).
Solid-state fermentation culture medium: dregs of beans: wheat bran: Semen Maydis powder=10:8-10:0.5-2, moisture content 40-70%.
(3) mensuration of viable count
General employing colony counting method, operation steps is: take the bean pulp fermentation goods 10g prepared, be placed in the stroke-physiological saline solution of 90ml, be placed on shaking table and shake 30min, shaking up the bacterium liquid that rear liquid-transfering gun gets 1ml adds in the stroke-physiological saline solution of another 9ml, dilute, according to national standard method, take turns doing ten times of gradient dilutions.After having diluted, get suitable extent of dilution 1ml respectively with liquid-transfering gun and throw flat board into, pour into substratum mixing, take turns doing three parallel, be placed in incubator at 37 DEG C of quiescent culture 48h.
(4) mensuration of pH value
Take the rear sample of 10g fermentation, put into triangular flask, add the distilled water of 90ml.Use magnetic stirrer 30min, after leaving standstill 10min, determine its pH value with Accurate pH measurement.
(5) mensuration of amino-acid nitrogen content
Get the rear sample of 10g fermentation, broken with homogenizer, mix with the distilled water of 90ml afterwards, place 1 day at 4 DEG C of refrigerators.The centrifugal 7000r/min of sample, centrifugal 15min.Supernatant liquor 0.8ml is placed in test tube, then adds 3.2ml distilled water, 2.0ml developer, and mixing, puts into boiling water bath and heat 15min, make blank simultaneously.Then cold water cooling, adds the mixing of 5.0ml40% ethanolic soln, uses 1cm cuvette after placing 15min, and returning to zero in 570nm place with blank tube measures A value.
Below the determination of some parameters of the production method to the probiotics fermention product taking dregs of beans as matrix, and the optimization of some culture condition.
Embodiment 1: probiotics preparation containing Bacillus coagulans, lactobacterium casei, plant lactobacillus and preparation method thereof, its preparation method comprises spawn culture and solid state fermentation is cultivated.
Described seed culture condition: Bacillus coagulans HM-09 and Bacillus licheniformis BL-09 seed culture condition: be positioned on shaking table, rotating speed is 180r/min, and temperature is 37 DEG C, cultivates 24h; Lactobacterium casei HM-09 and plant lactobacillus HM-10 is 37 DEG C of quiescent culture 24h in thermostat container.
Described Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 seed culture medium are (g/L): peptone 20, glucose 50, adds water to 1L, pH7.0-7.2.
Described solid state fermentation cultural method is: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation temperature 37 DEG C, humidity 60%, add Bacillus licheniformis BL-09 after mixed fermentation 18h and continue fermentation 28h, temperature 34 DEG C, humidity 50%.
Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 5%(v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively 1%(v/w), Bacillus licheniformis BL-09 is 5%(v/w).
Described Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 seed culture medium are (g/L): peptone 20, glucose 50, adds water to 1L, pH7.0-7.2.
Described solid-state fermentation culture medium mass ratio consists of: dregs of beans: wheat bran: Semen Maydis powder=10:9.5:2, water content controls 55%.
Record Bacillus coagulans HM-09 viable count 2.69 × 10
9cFU/g, lactobacterium casei HM-09 viable count 3.58 × 10
9cFU/g, plant lactobacillus HM-10 viable count 1.86 × 10
9cFU/g, Bacillus licheniformis BL-094.42 × 10
9cFU/g, after fermentation ends, its free ammonical nitrogen content is: 671.3umol/g.
Embodiment 2: probiotics preparation containing Bacillus coagulans, lactobacterium casei, plant lactobacillus and preparation method thereof, its preparation method comprises spawn culture and solid state fermentation is cultivated.
Described seed culture condition: Bacillus coagulans HM-09 and Bacillus licheniformis BL-09 seed culture condition: be positioned on shaking table, rotating speed is 180r/min, and temperature is 37 DEG C, cultivates 24h; Lactobacterium casei HM-09 and plant lactobacillus HM-10 is 37 DEG C of quiescent culture 24h in thermostat container.
Described solid state fermentation cultural method is: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation temperature 33 DEG C, humidity 50%, add Bacillus licheniformis BL-09 after mixed fermentation 24h and continue fermentation 24h, temperature 37 DEG C, humidity 55%.After fermentation ends, fluidised bed drying prepares microbial inoculum.
Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 8%(v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively 3%(v/w), Bacillus licheniformis BL-09 is 5%(v/w).
Described Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 seed culture medium are (g/L): peptone 20, glucose 50, adds water to 1L, pH7.0-7.2.
Described solid-state fermentation culture medium mass ratio consists of: dregs of beans: wheat bran: Semen Maydis powder=10:9:1, water content controls 60%.
Record Bacillus coagulans HM-09 viable count 2.18 × 10
9cFU/g, lactobacterium casei HM-09 viable count 2.57 × 10
9cFU/g, plant lactobacillus HM-10 viable count 3.66 × 10
9cFU/g, Bacillus licheniformis BL-094.15 × 10
9cFU/g, after fermentation ends, its free ammonical nitrogen content is: 659.2umol/g.
Embodiment 3: probiotics preparation containing Bacillus coagulans, lactobacterium casei, plant lactobacillus and preparation method thereof, its preparation method comprises spawn culture and solid state fermentation is cultivated.
Described seed culture condition: Bacillus coagulans HM-09 and Bacillus licheniformis BL-09 seed culture condition: be positioned on shaking table, rotating speed is 180r/min, and temperature is 37 DEG C, cultivates 24h; Lactobacterium casei HM-09 and plant lactobacillus HM-10 is 37 DEG C of quiescent culture 24h in thermostat container.
Described solid state fermentation cultural method is: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation temperature 33 DEG C, humidity 50%, add Bacillus licheniformis BL-09 after mixed fermentation 20h and continue fermentation 28h, temperature 33 DEG C, humidity 55%.
Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 5%(v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively 3%(v/w), Bacillus licheniformis BL-09 is 7%(v/w).
Described Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 seed culture medium are (g/L): peptone 20, glucose 50, adds water to 1L, pH7.0-7.2.
Described solid-state fermentation culture medium mass ratio consists of: dregs of beans: wheat bran: Semen Maydis powder=10:10:1, water content controls 50%.
Record Bacillus coagulans HM-09 viable count 2.84 × 10
9cFU/g, lactobacterium casei HM-09 viable count 3.19 × 10
9cFU/g, plant lactobacillus HM-10 viable count 3.45 × 10
9cFU/g, Bacillus licheniformis BL-095.53 × 10
9cFU/g, after fermentation ends, its free ammonical nitrogen content is: 681.4umol/g.
Embodiment 4(most preferred embodiment): probiotics preparation containing Bacillus coagulans, lactobacterium casei, plant lactobacillus and preparation method thereof, its preparation method comprises spawn culture and solid state fermentation is cultivated.
Described seed culture condition: Bacillus coagulans HM-09 and Bacillus licheniformis BL-09 seed culture condition: be positioned on shaking table, rotating speed is 180r/min, and temperature is 37 DEG C, cultivates 24h; Lactobacterium casei HM-09 and plant lactobacillus HM-10 is 37 DEG C of quiescent culture 24h in thermostat container.
Described solid state fermentation cultural method is: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation temperature 37 DEG C, humidity 60%, add Bacillus licheniformis BL-09 after mixed fermentation 28h and continue fermentation 20h, temperature 37 DEG C, humidity 60%.
Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 5%(v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively 1%(v/w), Bacillus licheniformis BL-09 is 5%(v/w).
Described Bacillus coagulans HM-09, lactobacterium casei HM-09, Bacillus licheniformis BL-09 and plant lactobacillus HM-10 seed culture medium are (g/L): peptone 20, glucose 50, adds water to 1L, pH7.0-7.2.
Described solid-state fermentation culture medium mass ratio consists of: dregs of beans: wheat bran: Semen Maydis powder=10:10:0.5, water content controls 60%.
Record Bacillus coagulans HM-09 viable count 2.90 × 10
9cFU/g, lactobacterium casei HM-09 viable count 3.48 × 10
9cFU/g, plant lactobacillus HM-10 viable count 2.87 × 10
9cFU/g, Bacillus licheniformis BL-096.87 × 10
9cFU/g, after fermentation ends, its free ammonical nitrogen content is: 700.5umol/g.
Embodiment 5: fodder additives (most preferred embodiment group) result of use is tested
Milk cow is selected from Beijing Milk Cow Center's seed multiplication farm, and select age, parity, lactation amount close, the china holstein cows 100 of just childbirth, is divided into 2 groups at random, often organizes 50.It is identical that test group, control group respectively organize basal diet.Test group daily ration is daily daily ration+15% product of the present invention, and control group is daily daily ration.
Test site and time: this test is carried out at milk cow center seed multiplication farm; Trial period (on August 1st, 2013 was to September 30).
Test method and feeding and management are in process of the test, and control group dairy cow diet is undertaken by original formulation.Its forage component, ratio and scale of feeding are consistent with before test; Test group daily ration from childbirth day adds product of the present invention on original basis.All the other are consistent with control group.Every day 3 times, timing feeding, freely drinks water, and first feeds roughage, feeds fine fodder afterwards, milks every day 3 times.
The appetite of mensuration project and record viewing test milk cow every day and healthy state, record bovine lactation amount every day every.Result illustrates:
Search for food and health condition observation: in trial period, cow feeding amount, health condition normally, do not occur unusual phenomenon.Experimental group milk cow is more smooth by hair, and the growth of livestock is better.
Impact on Lactation of Dairy Cow amount: test group August 15, September 8, September 30 lactation amount compared with control group, in table 1.
The impact that table 1, invention product change lactating cows lactation amount
| Group | Lactation amount (kg/d) |
| August 15 | September 8 | September 30 | |
| Experimental group | 38.57 | 34.02 | 32.34 |
| Control group | 30.83 | 25.55 | 24.42 |
Claims (8)
1. a solid state fermentation probiotic agent, with fermented bean dregs, wheat bran and Semen Maydis powder for solid-state fermentation substrate, it is characterized in that: adopt Bacillus coagulans (Bacilluscoagulans) HM-09, Bacillus licheniformis (Bacilluslicheniformis) BL-09, lactobacterium casei (Lactobacilluscasei) HM-09 and plant lactobacillus (Lactobacillusplantarum) HM-10 mixed solid fermentation to obtain;
Described Bacillus licheniformis (Bacilluslicheniformis) BL-09, deposit number is CGMCCNo.5686.
2. a kind of solid state fermentation probiotic agent as claimed in claim 1, is characterized in that: the substratum parts by weight of described solid state fermentation consist of: dregs of beans 29%, wheat bran 29%, Semen Maydis powder 2%, water 40%.
3. a kind of solid state fermentation probiotic agent as claimed in claim 1, is characterized in that: the method for described solid state fermentation is as follows:
Solid state fermentation culture condition: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation, temperature 32-37 DEG C, humidity 40-60%, add Bacillus licheniformis BL-09 after mixed fermentation 18-28h and continue fermentation 20-28h, temperature 34-37 DEG C, humidity 40-60%; After fermentation ends, drying prepares microbial inoculum.
4. a kind of solid state fermentation probiotic agent as described in claim 1 or 3, it is characterized in that: the inoculum size of described Bacillus coagulans HM-09 is 4-6%, the inoculum size of lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively the inoculum size of 1-3%, Bacillus licheniformis BL-09 is 5-7%.
5. a kind of solid state fermentation probiotic agent as described in claim 1 or 3, is characterized in that: described solid state fermentation terminates rear free ammonical nitrogen content and is: 700.5umol/g; In fermented product, Bacillus coagulans HM-09 viable count is 2.90 × 10
9cFU/g, lactobacterium casei HM-09 viable count is 3.48 × 10
9cFU/g, plant lactobacillus HM-10 viable count is 2.87 × 10
9cFU/g, Bacillus licheniformis BL-09 viable count is 6.87 × 10
9cFU/g.
6. a preparation method for solid state fermentation probiotic agent, comprises the steps:
Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium, mixed fermentation temperature 32-37 DEG C, humidity 40-60%, add Bacillus licheniformis BL-09 after mixed fermentation 18-28h and continue fermentation 20-28h, temperature 34-37 DEG C, humidity 40-60%, fermentation ends is dry;
Described Bacillus licheniformis (Bacilluslicheniformis) BL-09, deposit number is CGMCCNo.5686;
The parts by weight of described solid medium consist of: dregs of beans 29%, wheat bran 29%, Semen Maydis powder 2%, water 40%;
The inoculum size of described Bacillus coagulans HM-09 is 4-6%, and the inoculum size of lactobacterium casei HM-09 and plant lactobacillus HM-10 is respectively the inoculum size of 1-3%, Bacillus licheniformis BL-09 is 5-7%;
Described solid state fermentation terminates rear free ammonical nitrogen content: 700.5umol/g; In fermented product, Bacillus coagulans HM-09 viable count is 2.90 × 10
9cFU/g, lactobacterium casei HM-09 viable count is 3.48 × 10
9cFU/g, plant lactobacillus HM-10 viable count is 2.87 × 10
9cFU/g, Bacillus licheniformis BL-09 viable count is 6.87 × 10
9cFU/g.
7. the preparation method of a solid state fermentation probiotic agent, comprise the steps: that the method for described solid state fermentation is: Bacillus coagulans HM-09, lactobacterium casei HM-09 and plant lactobacillus HM-10 access solid medium mixed fermentation, temperature 33 DEG C, humidity 50%, Bacillus licheniformis BL-09 is added after mixed fermentation 20h, deposit number is that CGMCCNo.5686 continues fermentation 28h, temperature 33 DEG C, humidity 55%; Solid state fermentation inoculum size is: Bacillus coagulans HM-09 is 5% (v/w), lactobacterium casei HM-09 and plant lactobacillus HM-10 be respectively 3% (v/w), Bacillus licheniformis BL-09 is 7% (v/w).
8. the application of a kind of solid state fermentation probiotic agent as claimed in claim 1 in detoxification.
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