CN111670998A - Composite microbial inoculum suitable for silage corns in Ningxia region, preparation method and application - Google Patents
Composite microbial inoculum suitable for silage corns in Ningxia region, preparation method and application Download PDFInfo
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- CN111670998A CN111670998A CN202010060472.1A CN202010060472A CN111670998A CN 111670998 A CN111670998 A CN 111670998A CN 202010060472 A CN202010060472 A CN 202010060472A CN 111670998 A CN111670998 A CN 111670998A
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/12—Animal feeding-stuffs obtained by microbiological or biochemical processes by fermentation of natural products, e.g. of vegetable material, animal waste material or biomass
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K30/00—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs
- A23K30/10—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder
- A23K30/15—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder using chemicals or microorganisms for ensilaging
- A23K30/18—Processes specially adapted for preservation of materials in order to produce animal feeding-stuffs of green fodder using chemicals or microorganisms for ensilaging using microorganisms or enzymes
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/20—Bacteria; Culture media therefor
Abstract
The invention relates to the technical field of animal husbandry, and particularly discloses a compound microbial inoculum suitable for silage corns in Ningxia regions, a preparation method and application. The composite microbial inoculum suitable for silage corns in Ningxia areas comprises lactic acid bacteria and saccharomycetes or fermentation products of the lactic acid bacteria and the saccharomycetes. The composite microbial inoculum can obviously reduce the content of neutral detergent fiber, acidic detergent fiber and coarse ash in the silage corn feed, increase the content of coarse fat in the silage corn feed and improve the total energy of the silage corn feed. Meanwhile, the palatability of the silage corn feed is improved, and the silage corn feed is guaranteed to have higher fermentation quality and higher nutritional value.
Description
Technical Field
The invention relates to the technical field of animal husbandry, in particular to a compound microbial inoculum suitable for silage corns in Ningxia regions, a preparation method and application.
Background
Corn is one of three major food crops in Ningxia and is also the main feed crop in Ningxia. Corn is used as a high-quality feed source of ruminants, has the advantages of high nutritional value, good palatability, high digestibility, long storage life and the like after being made into silage, becomes a main feed source of the ruminants in Ningxia in winter, and the demand is continuously increased, but the problems of nutritional loss, aerobic decay and the like in the production process of the corn silage are urgently solved.
The use of silage additives is one of the important measures to improve the quality of silage. The microbial inoculant is a novel silage additive which is widely used and mainly has the functions of regulating a microbial system in silage and regulating a silage fermentation process. The market has more commercial silage microbial inoculants, but the microbial inoculants mainly comprise single strains of lactic acid bacteria, the strains aiming at the climate environment of Ningxia regions are fewer, the microbial inoculants cannot be well adapted to the drought and cold climate environments in Ningxia and winter in the silage process, the activity of the lactic acid bacteria is lower after the microbial inoculants are added, and the silage effect is poor; meanwhile, the problem of higher cost of adding the microbial inoculum exists.
Therefore, it is necessary to provide a new compound microbial inoculum suitable for silage corns in Ningxia regions, a preparation method and application to solve the problems.
Disclosure of Invention
In order to solve the problems in the prior art, the invention aims to provide the compound microbial inoculum which can ensure that the silage corn feed has higher fermentation quality and nutritive value and is suitable for the silage corn in Ningxia regions.
In order to achieve the purpose of the invention, the technical scheme of the invention is as follows:
a composite bacterial preparation suitable for ensiling corn in Ningxia region is prepared from lactobacillus and yeast or the fermented product of said lactobacillus and said yeast.
The preparation of the silage is a very complicated microbial fermentation process, and microorganisms in the silage mainly comprise lactic acid bacteria, saccharomycetes, mould and other putrefying bacteria. According to the invention, researches show that the lactic acid bacteria and the saccharomycetes have a promoting effect on fermentation of the silage, the lactic acid bacteria can reduce the respiration effect and the enzyme activity of plants in the silage process, inhibit the reproduction of harmful accessory microorganisms, reduce the oxidation of the silage raw materials to sugar and the hydrolysis of protein due to the respiration effect of the plants, keep the nutritive value of the silage raw materials, and simultaneously, the lactic acid bacteria generate esterase in the fermentation process and increase the degradation of neutral detergent fibers of the silage raw materials in the silage process. The yeast can increase the content of microbial protein in the feed, and simultaneously generate substances such as ethanol and the like, so that the silage has special fragrance, and the nutritional value and the palatability of the silage are improved.
The invention preferably separates lactobacillus and saccharomycete strains which can adapt to the Ningxia climate environment, have good acid production performance and strong growth and reproduction capacity and can promote the fermentation of the silage corns from high-quality silage corn feed naturally fermented in Ningxia Hui autonomous regions so as to solve the problems of more nutrient loss, easy mildew and the like of the silage corns in Ningxia regions in the silage process.
The invention uses a selective medium to selectively culture microorganisms in silage corn feed of Ningxia Hui nationality autonomous region, separates lactobacillus and saccharomycete strains which promote the fermentation of the silage corn feed, measures the performance of the lactobacillus and saccharomycete strains, selects strains with good acid production performance and high propagation speed, and prepares the composite silage bacterial agent.
In the composite microbial inoculum, the ratio of the viable count of the lactic acid bacteria to the viable count of the yeast is (1-5): (1-5).
Preferably, the ratio of the number of live bacteria of the lactic acid bacteria to the number of live bacteria of the yeast is 1:1, so that the cost is saved on the basis of ensuring the performance of the bacteria.
In the composite microbial inoculum of the invention, the viable count of the composite microbial inoculum is 108-109CFU/g。
Preference is given toThe viable count of the composite microbial inoculum is 109CFU/g to ensure ensiling effect through higher bacterial activity.
The invention also provides a method for preparing the composite microbial inoculum, which comprises the step of respectively preparing freeze-dried powder from the lactic acid bacteria and the product obtained by fermenting the yeast and then mixing.
The invention further provides an application of the compound microbial inoculum or the method in preparation of silage corn feed.
The invention provides application of the compound microbial inoculum or the method in reducing the content of neutral washing fiber, acidic washing fiber and/or coarse ash in silage corn feed.
The invention provides application of the compound microbial inoculum or the method in increasing the crude fat content of the silage corn feed and/or improving the total energy of the silage corn feed.
The invention has the beneficial effects that:
compared with commercial microbial inoculum in the market, the compound microbial inoculum can obviously reduce the content of neutral detergent fiber, acidic detergent fiber and crude ash in the silage corn feed, increase the crude fat content of the silage corn feed and improve the total energy of the silage corn feed. Meanwhile, the palatability of the silage corn feed is improved, and the silage corn feed is guaranteed to have higher fermentation quality and higher nutritional value.
Drawings
FIG. 1 is a lactic acid bacteria acid production curve according to the present invention;
FIG. 2 is a graph showing the growth of lactic acid bacteria according to the present invention;
FIG. 3 is a yeast growth curve of the present invention.
Detailed Description
Preferred embodiments of the present invention will be described in detail with reference to the following examples. It is to be understood that the following examples are given for illustrative purposes only and are not intended to limit the scope of the present invention. Various modifications and alterations of this invention will become apparent to those skilled in the art without departing from the spirit and scope of this invention.
The experimental procedures used in the following examples are all conventional procedures unless otherwise specified. Materials, reagents and the like used in the following examples are commercially available unless otherwise specified.
Example 1 isolation and identification of lactic acid bacteria and yeasts
Taking 50g of silage corn samples from a clean bench, putting the silage corn samples into a sterilized beaker, adding 200mL of sterile normal saline, sealing the beaker, putting the beaker on a shaker at 120r/min, and slowly shaking the beaker for 2 h. Filtering the grass residue with a layer of sterile gauze, centrifuging the filtrate at 12000r/min for 30min, resuspending the precipitate with a little sterile physiological saline, selecting 10-7、10-8、10-9And (3) three dilution gradients, wherein 0.1mL of liquid is sucked in each dilution, and the liquid is respectively coated on an MRS solid culture medium and an YPD culture medium and cultured for 24-72 h at 37 ℃.
After the bacteria are formed on the MRS solid culture medium and the YPD culture medium, the shape of the bacteria is observed in a clean bench, and the lactic acid bacteria are separated and identified according to the method introduced in the literature, "lactic acid bacteria classification identification and experimental method". The colonies of YPD medium were identified by isolation of yeast according to the handbook of characterization and identification of Yeast.
EXAMPLE 2 determination of Strain Performance
The lactic acid bacteria and yeast separated in example 1 were subjected to a performance test to finally determine the lactic acid bacteria and yeast used in the present invention.
The performance test method comprises the following steps:
1. determination of acid production rate of lactic acid bacteria
Inoculating 24h lactobacillus culture solution into MRS liquid culture medium at 3%, and culturing at 37 deg.C in incubator. And (3) measuring the pH value of the lactobacillus fermentation liquor every 2h from the inoculation time of 0h, and drawing an acid production rate curve by taking different fermentation time (h) as a horizontal coordinate and the pH value of the fermentation liquor as a vertical coordinate. The measuring time is 30 h. The test results are shown in FIG. 1.
2. Measurement of growth Performance of lactic acid bacteria
Inoculating 24h culture solution of lactic acid bacteria into MRS liquid culture medium with the inoculation amount of 3%, culturing in an incubator at 37 ℃, taking samples every 2h from inoculation 0h, taking the MRS culture medium as a blank control, measuring the light absorption value of the samples at the wavelength of 600nm, and drawing a growth curve by taking the culture time (h) as an abscissa and the corresponding light absorption value (OD) as an ordinate. The measuring time is 30 h. The test results are shown in FIG. 2.
3. Determination of growth Performance of Yeast
Inoculating yeast 24h culture solution into YPD liquid culture medium at an inoculation amount of 3%, culturing in an incubator at 28 deg.C, taking samples every 2h from inoculation 0h, taking YPD liquid culture medium as blank control, measuring light absorption value of the samples at 660nm wavelength, and drawing growth curve with culture time (h) as abscissa and corresponding light absorption value (OD) as ordinate. The measuring time is 30 h. The test results are shown in FIG. 3.
According to the performance test results, the lactic acid bacteria and the yeast used by the invention are screened out. The test results of the lactic acid bacteria used in the present invention are shown in fig. 1 to 2, and the test results of the yeast are shown in fig. 3. As can be seen from FIGS. 1 to 2, the lactic acid bacteria used in the present invention have good acid productivity and high propagation rate. As can be seen from FIG. 3, the yeast used in the present invention has a high propagation rate.
Example 3 preparation of Complex microbial inoculum
The lactobacillus and yeast strains screened in the embodiment 2 are fermented in a microbial fermentation tank to prepare freeze-dried powder, the freeze-dried powder is compounded into a composite microbial inoculum, and the ratio of the viable count of the lactobacillus to the viable count of the yeast is 1:1, the viable bacteria content of the composite microbial inoculum is about 109CFU/g。
Example 4
In this example, the compound microbial inoculum prepared in example 3, a control group, and a commercial microbial inoculum were used to prepare silage corn feeds, and the prepared silage corn feeds were evaluated.
In the embodiment, fresh whole corn growing to the wax ripeness stage in Ningxia is selected, the corn is cut into straws with the length of about 5cm after being harvested, distilled water is sprayed and added into a control group by a sprayer, a commercial microbial inoculum and a compound microbial inoculum are respectively sprayed and added into a test group by a sprayer, silage corn raw materials sprayed with the microbial inoculum are quickly placed into a silage silo, the silage silo is coated by a plastic film after being compacted, and the silage corn raw materials are fermented in the silage silo in a sealing mode for 60 days. And then, opening the cellar for sensory evaluation, sampling and determining the nutritional ingredients of the silage corn feed.
Specifically, the silage corn feed is prepared in the following manner:
the method comprises the following steps: respectively adding the composite microbial inoculum and the commercial microbial inoculum into distilled water, and activating the microbial inoculum according to the mass ratio of 1: 20.
Step two: the whole fresh corn plant growing to the wax ripeness stage is harvested and crushed into short stalks with the length of 5cm, and the moisture content of the silage raw material is 66.33%.
Step three: the silage test was divided into four groups, which were a control group (group a), a commercial microbial inoculum group (group B), a commercial microbial inoculum group (group C), and a complex microbial inoculum group (group D).
Group A: no additive is added, and only distilled water is adopted; group B: adding 2g/t of a first commercial microbial inoculum; group C: adding 3g/t of a commercial microbial inoculum II; group D: 2.5g/t of the composite microbial inoculum of example 3 is added.
Step four: and (4) respectively spraying the activated commercial microbial inoculum and the activated complex microbial inoculum obtained in the step one onto the silage corn raw materials by using a sprayer.
Step five: compacting the ensiled corn raw material, coating the ensiled corn raw material by using a plastic film, and sealing and fermenting the ensiled corn raw material in an ensiling cellar for 60 days.
Step six: and (5) opening the silage pit for silage fermentation for 60d, and performing sensory evaluation and nutrient component determination on the silage corn feed.
Sensory evaluation criteria are seen in table 1.
TABLE 1 German Agri-Association of agriculture (DLG) silage quality sensory score standards
The sensory evaluation results are shown in table 2.
TABLE 2 silage sensory score
| Group of | Color and luster | Smell(s) | Texture of | Scoring |
| Group A | Yellow colour | Almost has no sour taste | With mild mold | 9 |
| Group B | Light brown | Has weak odor of butyric acid | The stem and leaf structure is well preserved | 15 |
| Group C | Light yellow | Faint fragrance | The stem and leaf parts are well maintained | 15 |
| Group D | Light yellow | Has no odor of butyric acid and aromatic sour taste | The stem and leaf structure is well maintained | 19 |
As can be seen from Table 2, the silage of group A is yellow in color, has slight mildewing phenomenon, has special sharp odor and musty taste, has a total score of 9, and belongs to grade 3 and the like. The group B silage is light brown in color, has weak butyric acid odor, has good storage of the stem leaf structure of the silage, has the overall score of 15, and belongs to the level 2. The group C silage is light yellow and slightly aromatic, the stem leaf structure of the silage is well preserved, the overall score is 15, and the silage belongs to grade 2. The D group silage has special aromatic sour taste, similar color to the raw materials, good stem and leaf structure, 19 points of overall score and good grade 1.
The nutrient content results are shown in table 3.
TABLE 3 silage corn feed nutrient content
Note: the data in the same row shows significant difference (P < 0.05) in different lowercase letters in the shoulder marks, and shows insignificant difference (P > 0.05) in the same or no letters.
Studies have shown that neutral detergent fiber represents a structural carbohydrate in feed, and that the level of neutral detergent fiber in the ration affects ruminant feed intake, in relation to feed palatability. The lower the neutral detergent fiber content of the silage, the higher the feed intake of the livestock. The acidic detergent fiber influences the digestion capability of ruminants on feed, and researches show that the lower the content of the acidic detergent fiber in the silage, the higher the digestibility of the feed. The content of the crude fat is one of the conventional analysis items of the feed, the level of the crude fat is an important index for measuring the quality and the nutritive value of the feed, and the crude fat is used as a high-energy substance and has quite important effects on the aspects of growth, development, fertility and the like of animals. The coarse ash in the feed is mainly minerals such as oxides and salts, and is not easily absorbed and utilized by animals, so the smaller the content of the coarse ash in the feed, the better.
As can be seen from Table 3, compared with the group A, after the commercial microbial inoculum and the complex microbial inoculum are added, the crude protein content of B, C group is remarkably reduced (P < 0.05), the crude protein content of the silage corn feed of group D is slightly reduced compared with the group A, but the difference is not remarkable (P > 0.05), the crude protein content difference between B, C groups is not remarkable (P > 0.05), but the crude protein content of group D is remarkably higher than that of B, C group (P < 0.05), and the action effect of the complex microbial inoculum on inhibiting the decomposition of the crude protein of the silage is better than that of the commercial microbial inoculum. Compared with the group A, the crude fat content of the group B silage corn feed EE is kept unchanged, along with the increase of the addition amount of the commercial microbial inoculum, the crude fat content of the group C silage corn feed is remarkably reduced (P < 0.05), the crude fat content of the group D silage corn feed is remarkably higher than that of the group A, B, C (P < 0.05), and the situation that the compound microbial inoculum can improve the crude fat content of the silage corn feed EE is proved, and the general commercial microbial inoculum has the possibility of reducing the crude fat content of the silage corn feed EE.
Compared with the group A, the content change of the acid washing fiber of the group B is not obvious (P is more than 0.05), the content of the acid washing fiber of the silage corn feed of the group C, D is obviously reduced compared with the group A (P is less than 0.05), and simultaneously, the content of the acid washing fiber of the group D is also obviously reduced compared with the group B, C (P is less than 0.05). B. The content of neutral detergent fibers in the feed of the group C is remarkably increased (P < 0.05) compared with that of the group A, and the content of neutral detergent fibers in the feed of the group D is remarkably reduced (P < 0.05) compared with that of the group A, B, C, so that the effect of the composite microbial inoculum on reducing acidic detergent fibers and neutral detergent fibers in the silage corn feed is better than that of the commercial microbial inoculum. B. C, D groups all significantly reduced the Ash content of silage corn feed (P < 0.05). B. C, D groups all significantly improved the total energy of silage (P < 0.05).
In conclusion, the invention reduces the contents of neutral detergent fiber, acidic detergent fiber and crude ash in the silage corn feed, improves the crude fat content of the silage corn feed, and simultaneously increases the silage corn feed all the time, thereby showing that the invention can effectively improve the nutritive value and the fermentation quality of the silage corn feed.
Although the invention has been described in detail hereinabove with respect to a general description and specific embodiments thereof, it will be apparent to those skilled in the art that modifications or improvements may be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.
Claims (9)
1. The composite microbial inoculum suitable for silage corns in Ningxia regions is characterized by comprising lactic acid bacteria and saccharomycetes or fermentation products of the lactic acid bacteria and the saccharomycetes.
2. The composite microbial inoculum according to claim 1, wherein the ratio of viable count of the lactic acid bacteria to the yeast is (1-5): (1-5).
3. The composite microbial inoculum according to claim 2, wherein the ratio of the viable count of the lactic acid bacteria to the viable count of the yeast is 1: 1.
4. the complex microbial inoculant according to any one of claims 1 to 3, wherein the viable count of the complex microbial inoculant is 108-109CFU/g。
5. The complex microbial inoculant according to claim 4, wherein the viable count of the complex microbial inoculant is 109CFU/g。
6. A method for preparing the composite microbial inoculum of any one of claims 1 to 5, which comprises the step of respectively preparing the lactic acid bacteria and the yeast fermentation product into freeze-dried powder and then mixing.
7. Use of the complex microbial agent of any one of claims 1 to 5 or the method of claim 6 in the preparation of silage corn feed.
8. Use of the complex inoculant of any one of claims 1-5 or the method of claim 6 to reduce the neutral washed fiber, acid washed fiber and/or coarse ash content of silage corn feed.
9. Use of the complex microbial inoculant of any one of claims 1 to 5 or the method of claim 6 for increasing the crude fat content and/or total energy of silage corn feed.
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Application publication date: 20200918 |