CN102174421B - Glycerol-producing saccharomyces cerevisiae NAU-ZH-GY1 and application thereof - Google Patents
Glycerol-producing saccharomyces cerevisiae NAU-ZH-GY1 and application thereof Download PDFInfo
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- CN102174421B CN102174421B CN201110057770A CN201110057770A CN102174421B CN 102174421 B CN102174421 B CN 102174421B CN 201110057770 A CN201110057770 A CN 201110057770A CN 201110057770 A CN201110057770 A CN 201110057770A CN 102174421 B CN102174421 B CN 102174421B
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- saccharomyces cerevisiae
- glycerine
- nau
- fermentation
- glycerol
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- 235000015099 wheat brans Nutrition 0.000 description 1
Abstract
The invention provides glycerol-producing saccharomyces cerevisiae NAU-ZH-GY1 and application thereof, belonging to the field of microbial fermentation. The fungus is separated from unprocessed honey and is identified as saccharomyces cerevisiae with a collection number of CGMCC (China General Microbiological Culture Collection Center) No. 4551. The strain is obtained through liquid fermentation; fermentative culture comprises abundant glycerol, a large amount of living saccharomyces and rich microbial metabolism substances such as amino acid, vitamins and the like; the saccharomyces cerevisiae is applied to producing a feedstuff additive at the perinatal period of a ruminant; the glycerol can be used as an energy supplement to relieve negative energy balance; by means of the saccharomyces cerevisiae and the microbial metabolism substances such as amino acid, vitamins and the like, animal gastrointestinal tract flora can be adjusted, pathogenic microorganisms can be antagonized, immunity of organisms can be improved, conversion and absorption of nutritive materials can be promoted, and production performance can be improved; and the glycerol-producing saccharomyces cerevisiae NAU-ZH-GY1 is a novel environmentally-friendly, nutritive and safe feedstuff additive with high cost performance.
Description
Technical field
The present invention relates to a kind of product glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 and application thereof, belong to the microbial fermentation field, is to utilize the method for microbial fermentation to produce glycerine, is applicable to the production of ruminant feed supplement.
Background technology
Be meant this stage at later pregnancy (antenatal 2～3 weeks) to lactation initial stage (2～3 weeks of postpartum) perinatal period of ruminating animal; Also be called the transitional period; Because the later pregnancy food consumption descends, dry-matter is taken in and is reduced, and has satisfied not embryo growth and development and the metabolic needs of self-energy; And peak (8～10 weeks of postpartum) is taken in prior to dry-matter in postpartum milk secretion peak (4～6 weeks of postpartum), makes body be in a kind of like this nutrition stress situation of negative energy balance.Research shows; The glucose that the later pregnancy fetus consumes can account for parent institute glucogenic 46%; The glucose that peak of lactation consumes is then up to 85%; Therefore Chang Yi causes energy metabolism impairment, causes a series of perinatal period diseases such as ketoacidosis, fatty liver, milk fever, has a strong impact on the development of ruminating animal aquacultures such as ox, sheep.
Since introduction, the cultivation of high-yield variety, the exploitation of high energy diet, utilization, the reasons such as raising of large-scale breeding degree, perinatal period, the sickness rate of energy metabolism impairment disease was high always for a long time.The sickness rate of China's ketosis of dairy cows accounts for 15.0%～30.0% of cow in milk, and India is 14.7%, and the U.S. is 5.0%, and Japan is up to 43.1%.The sickness rate of China's milk cow fatty liver surpasses 30.0%, and the U.S. is 35.0%～66.0%.According to external, 1 the ketoacidosis milk cow only loss that caused of medicine and milk yield decline promptly reaches 151～312 dollars.In addition, ketosis of dairy cows, fatty liver are also usually brought out other disease perinatal periods such as abomasum displacement, retention of placenta and parturient paresis because of it, bring serious economy loss to dairy.Therefore, prevention and control high yield cow in perinatal period energy metabolism impairment disease is to guarantee one of vital task that the milk cow highly effective and safe is produced.Although the research of this respect report is a lot, do not find the ideal solution all the time.
Most of glucose is to obtain through the glyconeogenesis approach in the ruminating animal body, and glyconeogenesis is meant that various living sugar precursors change the process of glycogen or glucose into, and is significant to perinatal ruminating animal, is the important channel of regulating energy metabolism impairment.Voltaile fatty acids such as microbial fermentation generation acetate, propionic acid, butyric acid provide energy in the cud because ruminating animal mainly leans on, and glucose or other glucide are difficult to directly get into blood or glyconeogenesis approach.Glycerine is the sugared precursor of a kind of life, and it has the glyconeogenesis approach that is superior to other living sugared precursors, in the glyconeogenesis process, is converted into glucose through glycerol kinase, and does not rely on rate-limiting enzyme pyruvate carboxylase or phosphoric acid enol pyruvic acid carboxylase.Existing people adds glycerine in the daily ration of milk cow perinatal period to as giving birth to sugar precursor, and research shows that glycerine can improve the blood sugar of milk cow perinatal period, improves milk performance, regulates the blood metabolism, reduces ketoboidies content, plays the effect of alleviating negative energy balance.
Mainly synthetic as the glycerine of milk cow energy supplement perinatal period thing at present from the by product or the industry of production of biodiesel process, can introduce industrial objectionable impurities in its course of processing, milk cow appetite is produced detrimentally affect or body is produced destructive stimulus; High purity glycerine then cost is high, has limited promoting the use of glycerine.
In recent years; Along with each side Progress in technique such as biotechnology, microbial strains, zymotechnique and extraction separation; Make ferment glycerin production become and have possibility; It is to utilize raw materials such as microbial fermentation starchy material (corn, Ipomoea batatas etc.) or molasses to produce glycerine that fermentation method produces glycerine, is a kind of natural glycerine, has overcome the shortcoming of industrial production glycerine.Mikrobe commonly used has pichia farinose, produces glycerin candida etc. in the industry.But also not seeing at present has the report that the glycerine of fermentative Production is used as fodder additives, and the product glycerol yeast that tradition is used not is a probiotic bacterium.
Probiotic bacterium is meant that dropping into the back brings into play beneficial effect through improving host's intestinal microflora eubiosis; Reach the active bacteria formulation and the meta-bolites thereof that improve host's (humans and animals) health level and healthy good attitude, mainly comprise milk-acid bacteria, bifidus bacillus, actinomycetes, yeast.In fodder industry, the probiotic bacterium additive is utilize microbe additive that biotechnology produces a kind of, and yeast class probiotic bacterium commonly used has yeast saccharomyces cerevisiae.In the U.S., the yeast additive is widely used as a kind of forage component of routine.Its mechanism of action it has been generally acknowledged that: (1) yeast is as viable bacteria; Breed and the vigor enhancing after getting into gi tract, promote microbial reproduction and enhanced activity, can effectively suppress the breeding of pathogenic micro-organism; Regulate the micro ecology of gastrointestinal tract balance; Participate in the struggle for life of pathogenic micro-organism flora, the repulsion pathogenic bacteria adheres to gastrointestinal tract mucous surface, assists body to eliminate toxin and meta-bolites thereof.(2) improve the animal intestinal digestive enzyme activity.Through improving gastrointestinal tract environment and flora structure, Regulation of Gastric intestinal fermentation.Reduce Lactated generation; Improve pH value stabilization property, promote the raising of profitable strain breeding and vigor, increase the effective concentration of probiotics; Promote stomach and intestine to the decomposition of nutritive substance, synthetic, digestion, absorb and utilize; Thereby the increase food consumption is improved the utilization ratio of animal to feed, improves production performance; Improve the digestibility of animal to Mierocrystalline cellulose and mineral substance; (3) enhancing body immunizing power and disease resistance prevent the absorption of toxin and refuse, and animal digestive system is played a role in health care; But growing animal is then grown in the stimulating gastrointestinal road, ensure animal health; (4) the multiple digestive ferment and the UGF of yeast secretary; Nutritive ingredients such as VITAMINs, SOD, bioactive peptide, coenzyme have growth promoting function; Excretory proteolytic enzyme, glycase and lipase activity are participated in degraded, the digestion of feed, improve the utilization ratio of animal to nutritive substance; And the enzyme of complex carbohydrates (like pectin, VISOSE, Mierocrystalline cellulose etc.) in the feed of degrading in addition, wherein much be the enzyme that animal itself does not have.Some researchs think that the effect of yeast class feed is based on it nutritional factor important, that can stimulate microorganism active is provided.
In sum; With specific probiotic bacterium is bacterial classification; Utilize various starchy materials or molasses to produce glycerine as fermenting substrate; The fermenting culture of probiotic bacterium as ruminant feed supplement perinatal period, can be brought into play the effect of glycerine and probiotic bacterium simultaneously, and glycerine can be used as the energy supplement thing and alleviates negative energy balance; Microbial metabolism such as yeast and amino acid, VITAMINs material, adjustable animal gastrointestinal tract flora, the antagonism pathogenic micro-organism, enhance immunity power, and promote nutritive substance to transform absorption, improve production performance.
Summary of the invention
The objective of the invention is to deficiency to above-mentioned prior art; A kind of yeast saccharomyces cerevisiae NAU-ZH-GY1 that can produce glycerine and other useful microbe metabolic substds is provided, another object of the present invention is to provide this yeast saccharomyces cerevisiae NAU-ZH-GY1 to produce the method for glycerine in fermentation.Another purpose of the present invention is to provide this product glycerine yeast saccharomyces cerevisiae to produce the application in the glycerine in fermentation.Another purpose of the present invention is to provide the application of this product glycerine yeast saccharomyces cerevisiae in producing ruminant feed supplement.Form green, nutrition, safety, novel ruminant feed supplement that cost performance is high through industrial fermentation production.
The object of the invention can be realized through following technical scheme:
The present invention provides a kind of product glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1; This strain classification called after: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae); Be stored in China Committee for Culture Collection of Microorganisms common micro-organisms center on January 13rd, 2011, culture presevation numbering CGMCC No.4551.
Main biological characteristics: microscopically is observed, and strain cell is a subcircular, and size (4.1-8.2) micron * (5.2-9.5) micron does not produce the mould film; Carry out vegetative propagation with the multiterminal budding; No pseudohypha has thecaspore to produce; On the agar plate 30 ℃ cultivate 24-36h after, the bacterium colony oyster white, swell, show smooth moistening, glossy, quality evenly, the edge is more neat.
30 ℃ of optimum growth temperatures, pH5.0.
Can glucose fermentation, SANMALT-S, sucrose, semi-lactosi, unfermentable lactose, cellobiose, synanthrin; Can not assimilate saltpetre, Sodium Nitrite.
The 26S rDNA D1/D2 zone of this bacterial strain has the base sequence shown in the SEQ ID NO.1, all reaches 99% with existing yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) the same area base sequence homology of Genbank.
Produce the method that glycerine is produced in glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 fermentation, its process is following:
1) substratum:
Seed culture medium (g/L): glucose 200, urea 2, yeast extract paste 4, uncomfortable pH, 115 ℃ of sterilization 20min.
Fermention medium (g/L): glucose 200, urea 4, Dried Corn Steep Liquor Powder 5, sodium-chlor 20, sal epsom 0.5, calcium chloride 0.1 is regulated 5.0,115 ℃ of sterilizations of pH 20min.
2) culture condition:
First order seed is cultivated: be shake flask fermentation, with the bottled seed culture medium 100ml of 500ml triangle, will produce glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 and insert in the seed culture medium that 30 ℃, the 150rpm shaking table was cultivated 24 hours.
Secondary seed is cultivated: the canned seed culture medium 6L of 10L seed fermentation, and 5%, 30 ℃ of inoculum size, the 150rpm shaking table was cultivated 24 hours.
The full-automatic stirred fermentor dress of fermentor cultivation: 100L fermention medium 60L adds inoculation by seeding tank stream, and 32 ℃, inoculum size 10%, pH 5.0,300rpm, air flow 0.5vvm cultivates 48h.
Above-mentioned product glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 produces the application in the glycerine in fermentation.
The application of above-mentioned product glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 in producing ruminant feed supplement.
Beneficial effect of the present invention:
The invention provides and produce glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1.The present invention also provides the fermentation process that produces glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1, and the glycerine that produces reaches 35.6g/L, and number of viable reaches 3 * 10
9CFU/mL.
The yeast that relates among the present invention is a yeast saccharomyces cerevisiae, is the probiotic bacterium that the Ministry of Agriculture feeds can directly being used to of allowing, its fermentation the glycerine that produces natural, have no side effect, safe and reliable.This yeast is applied to the production of ruminating animal fodder additives perinatal period, need not to extract glycerine after the fermentation, and production technique is simple, environmentally safe, the glycerine that produces can replenish animal energy, alleviate the animal energy negative balance, correct energy metabolism impairment; A large amount of yeast saccharomyces cerevisiae viable bacterias in the culture; And useful microbe physiological metabolism material such as rich in amino acid, VITAMINs, adjustable animal gastrointestinal tract flora, antagonism pathogenic micro-organism; Enhance immunity power and disease resistance; Promoting the conversion and the absorption of nutritive substance, improve production performance, will be a kind of green, nutrition, safety, novel fodder additive that cost performance is high.
Embodiment
Embodiment 1: the separation of producing the glycerine bacterial strain
Weighing sample (unprocessed honey) 10g puts into the Erlenmeyer flask that fills the 90mL SPSS, and vibration 20min processes bacteria suspension, draws the 0.1mL bacterial suspension inoculation in the wort agar substratum, and 28 ℃ of incubators are cultivated 24h.
Pick out yeast, produce glycerol fermentation respectively and cultivate, shaking table 150rpm, 28 ℃ of temperature are cultivated 48h.Culture medium prescription (g/L): glucose 200, urea 2, KH
2PO
40.4, uncomfortable pH, 115 ℃ of sterilization 20min.
Screening obtains the higher bacterium of a strain glycerine output; Called after NAU-ZH-GY1; Through being accredited as yeast saccharomyces cerevisiae (Saccharomyces cerevisiae), be stored in China Committee for Culture Collection of Microorganisms common micro-organisms center on January 13rd, 2011, culture presevation numbering CGMCC No.4551.
Main biological characteristics: microscopically is observed, and strain cell is a subcircular, and size (4.1-8.2) micron * (5.2-9.5) micron does not produce the mould film; Carry out vegetative propagation with the multiterminal budding; No pseudohypha has thecaspore to produce; On the agar plate 30 ℃ cultivate 24-36h after, the bacterium colony oyster white, swell, show smooth moistening, glossy, quality evenly, the edge is more neat;
30 ℃ of optimum growth temperatures, pH5.0;
Can glucose fermentation, SANMALT-S, sucrose, semi-lactosi, unfermentable lactose, cellobiose, synanthrin; Can not assimilate saltpetre, Sodium Nitrite.
The 26S rDNA D1/D2 zone of this bacterial strain has the base sequence shown in the sequence 1, all reaches 99% with existing yeast saccharomyces cerevisiae (Saccharomyces cerevisiae) the same area base sequence homology of Genbank.
Embodiment 2: utilize and produce glycerine bacterial strain NAU-ZH-GY1 fermentation product glycerine
1) substratum:
Seed culture medium (g/L): glucose 200, urea 2, yeast extract paste 4, uncomfortable pH, 115 ℃ of sterilization 20min.
Fermention medium (g/L): glucose 200, urea 4, Dried Corn Steep Liquor Powder 5, sodium-chlor 20, sal epsom 0.5, calcium chloride 0.1 is regulated 5.0,115 ℃ of sterilizations of pH 20min.
2) culture condition:
First order seed is cultivated: be shake flask fermentation, with the bottled seed culture medium 100ml of 500ml triangle, will produce glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 and insert in the seed culture medium that 30 ℃, the 150rpm shaking table was cultivated 24 hours.
Secondary seed is cultivated: the canned seed culture medium 6L of 10L seed fermentation, and 5%, 30 ℃ of inoculum size, the 150rpm shaking table was cultivated 24 hours.
The full-automatic stirred fermentor dress of fermentor cultivation: 100L fermention medium 60L adds inoculation by seeding tank stream, and 32 ℃, inoculum size 10%, pH 5.0,300rpm, air flow 0.5vvm cultivates 48h.
3) fermentation result: glycerine 35.6g/L, number of viable 3 * 10
9CFU/mL.
Animal experiment
1. experimental animal and feeding and management
2～3 years old healthy goat of 8 local bulls (castration), mean body weight 18.76 ± 0.94kg is numbered 1～8.The permanence lymphoma stomach fistulization pipe is installed in operation.The duration of test experimental animal adopts the drylot feeding of single hurdle; Basal diet consists of fine fodder and clover and does (basis smart slightly than be 1: 2); The fine fodder 90g/ only (fine fodder is that corn, dregs of beans and wheat bran mix by mass ratio at 65: 25: 10) that at every turn feeds; 180g/ of clover grass, every day, 8:00 and 17:00 respectively fed once, guaranteed sufficient drinking-water.The staple of alfalfa hay (%, mass percent): dry-matter 90.54, crude protein 17.52, crude fat 2.62, robust fibre 29.05, calcium 1.64, phosphorus 0.36.
2. test design
4 * 4 complete Latin square test design are adopted in test, and 8 fistula sheep are divided into 4 groups at random.Test was divided into for 4 phases, each issue trial test 7 days, official test 15 days.The control group C basal diet of feeding; The low dose group L basal diet+yeast culture of feeding (contains glycerine 30g and yeast 2.07 * 10
9CFU)/previous day; In the dose groups M basal diet+yeast culture of feeding (contain glycerine 60g and yeast 4.04 * 10
9CFU)/previous day; The high dose group H basal diet+yeast culture of feeding (contains glycerine 90g and yeast 6.07 * 10
9CFU)/previous day.Every day, 8:00 directly added yeast culture in the cud to through rumen fistula.
All animals are gathered rumen fluid sample detection microbial proteinous, ammonia nitrogen and voltaile fatty acid in empty stomach in the 16th day early morning of official test, gather the jugular vein blood sample and detect plasma glucose.
3. sample detection
3.1 rumen microorganism albumen
Rumen microorganism albumen (MCP) carries out The pretreatment with reference to Makkar substep centrifuging, Xylene Brilliant Cyanine G (G250) colorimetric method for determining protein content, and the standard protein that uses is bovine serum albumin (BSA), content is 96%～99%Albumin (Sigma).
(1) reagent preparation:
The preparation of Coomassie brilliant blue solution: take by weighing 100mg Xylene Brilliant Cyanine G G-250, be dissolved in the 50mL90% ethanol (guaranteeing fully dissolving), add the phosphoric acid solution 100mL of 85% (W/V), at last with the zero(ppm) water constant volume to 1000mL.After the stirred overnight, filter and use, one week of quality guaranteed period.
(2) preparation of typical curve
With the 1mgmL for preparing
-1Bovine serum albumin (BSA) mother liquor is with ten concentration point production standards of distilled water stepwise dilution curve.
(3) sample analysis
Rumen fluid takes out and thaws with vortice vortex vibration 45s～1min (mikrobe is separated with chyme) at 1000rmin
-1Centrifugal 5min under the rotating speed.Draw supernatant 1.5mL in the 2mL centrifuge tube in the freezing whizzer with 12000rmin
-1The centrifugal 30min of rotating speed.Take out sample and abandon supernatant, in substrate, add 0.8mL 0.25molL
-1NaOH, behind the mixing in 100 ℃ of water-baths water-bath 20min.React completely the back in 12000rmin
-1The centrifugal 30min of low temperature (4 ℃) draws supernatant 100 μ L then, adds the Xylene Brilliant Cyanine G solution that 5mL prepares, behind the mixing in wavelength 595nm place with 721 spectrophotometer colorimetrics.
3.2 ammonia nitrogen
With ammonium sulfate is standard substance, adopts phenol-hypochlorous acid colourimetry.Concrete steps are following:
(1) reagent preparation:
A reagent: accurately take by weighing the 0.0126g sodium nitroprusside, add 50mL ddH
2(wearing gloves) adds 2.5058g phenol (in 50～60 ℃ of water-baths, melt, use the small beaker weighing again) then in the volumetric flask of O dissolving back immigration 250mL, dissolves to 250mL surely with distilled water behind the mixing, shakes up 4 ℃ of preservations of brown bottle.
B reagent: accurately take by weighing 1.2505g sodium hydroxide and add the 100mL distilled water, add the Na of 12.6645g after the dissolving
2HPO
412H
2The warm stirring and dissolving of O is cooled to and adds 12.5mL 5.25% Youxiaolin mixing after the room temperature, adds distilled water then and to the 250mL constant volume after, uses filter paper filtering, filtrates to be placed in the Plastic Bottle 4 ℃ and to keep in Dark Place.
(2) preparation of typical curve:
Standard reserving solution is 5mmol/L (NH
4)
2SO
4Solution: configuration is preceding with (NH
4)
2SO
480 ℃ of oven dry 2h.Accurately take by weighing (the NH of 0.066g oven dry
4)
2SO
4Be dissolved in 100mL 0.1molL
-1Promptly get standard reserving solution in the hydrochloric acid (0.56mL concentrated hydrochloric acid constant volume is to 100mL).Be diluted to 1mmolL to above-mentioned standard reserving solution with distilled water
-1As mother liquor, stepwise dilution is ten concentration point production standard curves then.
(3) sample analysis:
The rumen fluid taking-up is thawed, press 3000rmin
-1The centrifugal 10min of rotating speed so that remove the chyme particle.0.2mL upper strata liquid (should suitably dilute in case of necessity) is drawn to the 10mL plastics tubing in centrifugal back, adds 1.5mL A liquid earlier, adds 1.5mL B liquid again, reacts 15min with being placed in 75～85 ℃ of water-baths behind the vortex mixed appearance mixing.Question response is cooled to room temperature with frozen water fully, with spectrophotometer colorimetric under 630nm.
3.3 voltaile fatty acid
With Ba Dousuan is that interior mark is used the gas chromatography determination voltaile fatty acid
With the Ba Dousuan of 5mmolL-1 just as interior calibration, with acetate (Acetate, A), propionic acid (Propionate, P), butyric acid (Butyrate, come qualitative, comes quantitative with the area and the concentration regression beeline equation of three kinds of acid by the RT separately of standard substance B).Go up appearance then, set up the area and the concentration regression beeline equation of three kinds of acid.
(1) foundation of reagent preparation and typical curve: set up straight line with 5 grades of gradient chromatogram standards mixing.
The Ba Dousuan of accurately drawing 344 μ L acetate, 373 μ L propionic acid, 184 μ L butanic acids and 43.6mg with liquid-transfering gun places the volumetric flask of 100mL, uses the chloroform constant volume, promptly gets hybrid standard liquid.Its concentration is respectively acetate 60mmolL
-1, propionic acid 50mmolL
-1, butanic acid 20mmolL
-1, Ba Dousuan 5mmolL
-1With above-mentioned hybrid standard liquid doubling dilution successively, be mixed with the standard mixed solution of 5 grades of gradients.
The preparation of acid sorbent material: accurately took by weighing SODIUM SULPHATE ANHYDROUS 99PCT, 50% sulfuric acid (V/V) and zeyssatite mixing by weight 30: 1: 20 and form.Earlier SODIUM SULPHATE ANHYDROUS 99PCT is ground during preparation, add 50% sulfuric acid then, add zeyssatite at last and grind mixing.The sour sorbent material for preparing will seal preservation, prevents that suction from becoming tide, influences measuring result.
(2) chromatographiccondition is specially: chromatographic column is Agilent19091F-413HP-FFAP capillary column (30.0m * 320 μ m * 0.25 μ m); The post case: temperature programming, post oven temperature, degree 140 ℃ (4min) be to 240 ℃ (2min), 50 ℃ (1min).Splitting ratio is 30: 1, and sample size is 1 μ L.Detector FID temperature is 250 ℃.
(3) rumen fluid pre-treatment:
The rumen fluid taking-up is thawed, at 1000rmin
-1Centrifugal 5min under the rotating speed.The rumen fluid supernatant 1mL that gets after centrifugal has in the Glass tubing of plug to the 10mL that is added with 3mL 0.5mM% Ba Dousuan chloroformic solution and 2.4g acidity sorbent material in advance.Jump a queue immediately, draw the 0.5mL supernatant behind the mixing, prepare to measure to Agilent GC bottle.
3.4 plasma glucose: adopt the complete white moving biochemical instruments analysis of BECKMAN.
4. test-results
By the table 1 glycerinated yeast culture of various dose of can feeding, H group propionic acid content is significantly higher than control group (P＜0.05), and H group acetate propionic acid ratio extremely significantly is lower than control group (P＜0.01); H group ammonia nitrogen concentration significantly is lower than control group (P＜0.05); Experimental group microbial proteinous content all is significantly higher than control group (P＜0.05); H group plasma glucose concentration is significantly higher than control group (P＜0.05).
Table 1 influence of glycerinated yeast culture of feeding perinatal period to goat rumen zymosis and plasma glucose
Annotate: * representes and the control group significant difference
5. discuss
Feeding of yeast culture improved the propionic acid content of H group, possibly be the growth that glycerine has promoted to produce in the cud bacterium acidi propionici, also possibly be saccharomycetic effect in the culture.
Have and report that if the more competent situation that glucide and soluble nitrogen provide, viable yeast can promote microbial growth, reduce the loss of nitrogen.This test is behind the feeding yeast culture, and H group ammonia nitrogen concentration descends, and microbial proteinous concentration increases; Also might be that glycerine one side glycerine has suppressed the growth of cud protein decomposing bacteria or reduced proteolytic activity; By-pass protein is increased, and on the other hand, glycerine is that mikrobe provides growth institute energy requirement; Promote that mikrobe utilizes the ammonia nitrogen synthetic animalcule albumen, make the proteinic synthetic increase of rumen microorganism.
The raising of goat blood sugar concentration; Explaining feeds produces the glycerol yeast culture has effect to blood glucose increasing; Possibly be through promoting the output of propionic acid in the cud; Perhaps glycerine directly absorbs the result who gets into blood, and this provides foundation for producing the application of glycerol yeast culture on ruminant feed supplement.
Conclusion (of pressure testing): the glycerinated yeast saccharomyces cerevisiae culture of feeding perinatal period, can effectively improve the goat energy metabolism and regulate gastrointestinal bacterial flora.
Advantage: this yeast is applied to the production of ruminating animal fodder additives perinatal period, need not to extract glycerine after the fermentation, and production technique is simple, and environmentally safe, the glycerine that produces have supplementing energy and alleviate the animal energy negative balance, correct the effect of energy metabolism impairment; A large amount of yeast saccharomyces cerevisiae viable bacterias in the culture; And useful microbe physiological metabolism material such as rich in amino acid, VITAMINs, adjustable animal gastrointestinal tract flora, antagonism pathogenic micro-organism; Enhance immunity power and disease resistance; Promoting the conversion and the absorption of nutritive substance, improve production performance, will be a kind of green, nutrition, safety, novel fodder additive that cost performance is high.
Claims (4)
1. one kind is produced glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1; This strain classification called after: yeast saccharomyces cerevisiae (Saccharomyces cerevisiae); Be stored in China Committee for Culture Collection of Microorganisms common micro-organisms center on January 13rd, 2011, culture presevation numbering CGMCC No.4551.
2. the method for utilizing the described product glycerine of claim 1 yeast saccharomyces cerevisiae NAU-ZH-GY1 fermentation to produce glycerine is characterized in that process is following:
1) substratum:
Seed culture medium: glucose 200g/L, urea 2g/L, yeast extract paste 4g/L, uncomfortable pH, 115 ℃ of sterilization 20min;
Fermention medium: glucose 200g/L, urea 4g/L, Dried Corn Steep Liquor Powder 5g/L, sodium-chlor 20g/L, sal epsom 0.5g/L, calcium chloride 0.1g/L regulates 5.0,115 ℃ of sterilizations of pH 20min;
2) culture condition:
First order seed is cultivated: be shake flask fermentation, with the bottled seed culture medium 100ml of 500ml triangle, will produce glycerine yeast saccharomyces cerevisiae NAU-ZH-GY1 and insert in the seed culture medium that 30 ℃, the 150rpm shaking table was cultivated 24 hours;
Secondary seed is cultivated: the canned seed culture medium 6L of 10L seed fermentation, and 5%, 30 ℃ of inoculum size, the 150rpm shaking table was cultivated 24 hours;
The full-automatic stirred fermentor dress of fermentor cultivation: 100L fermention medium 60L adds inoculation by seeding tank stream, and 32 ℃, inoculum size 10%, pH 5.0,300rpm, air flow 0.5vvm cultivates 48h.
3. the described product glycerine of claim 1 yeast saccharomyces cerevisiae produces the application in the glycerine in fermentation.
4. the application of the described product glycerine of claim 1 yeast saccharomyces cerevisiae in producing ruminant feed supplement.
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| CN103734497B (en) * | 2013-11-29 | 2015-12-09 | 南京农业大学 | A kind of feed addictive of anti-cow heat stress |
| CN104017740B (en) * | 2014-05-28 | 2017-04-26 | 烟台张裕集团有限公司 | Glycerin high-yielding saccharomyces cerevisiae strain and application thereof to dry white wine |
| CN104212730A (en) * | 2014-09-18 | 2014-12-17 | 广东一线天酒业有限公司 | Honey fermented wine |
| CN104855689A (en) * | 2015-05-28 | 2015-08-26 | 河南双成生物科技有限公司 | Method for preparing fatty acid-rich protein feed raw material from amino acid fermentation residual liquor |
| CN113186116B (en) * | 2021-02-08 | 2022-09-23 | 茅台学院 | Non-decarboxylation lechlenibacter NJ22 with lactic acid as carbon source and application thereof |
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|---|---|---|---|---|
| WO2010015122A1 (en) * | 2008-08-06 | 2010-02-11 | Tianjin University | Yeast strains and methods for making and using such yeast strains |
| CN101658243A (en) * | 2009-09-17 | 2010-03-03 | 南京农业大学 | Microbiological feed additive for adjusting energy metabolism impairment of flocks and herds during perinatal period |
-
2011
- 2011-03-10 CN CN201110057770A patent/CN102174421B/en not_active Expired - Fee Related
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010015122A1 (en) * | 2008-08-06 | 2010-02-11 | Tianjin University | Yeast strains and methods for making and using such yeast strains |
| CN101658243A (en) * | 2009-09-17 | 2010-03-03 | 南京农业大学 | Microbiological feed additive for adjusting energy metabolism impairment of flocks and herds during perinatal period |
Non-Patent Citations (2)
| Title |
|---|
| 张彦.利用遗传工程方法构建高产甘油酿酒酵母菌株.《中国优秀硕士学位论文全文数据库基础科学辑》.2009,(第4期),A006-57. * |
| 龙凌.酵母产品及其用途.《中国饲料》.2001,28-30. * |
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| CN102174421A (en) | 2011-09-07 |
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