Corn silage based on fermentation of propionibacterium freudenreichii subsp. Scheri and phytobacterium plantarum and preparation method of corn silage

Abstract

The invention discloses corn silage based on fermentation of propionibacterium freudenreichii subsp. Scheri and phytobacterium plantarum and a preparation method of the corn silage. The preparation method comprises the following steps: firstly, selecting whole corn or corn straws from the late milk-ripening stage to the early dough-ripening stage, and chopping to obtain silage crops; then uniformly scattering the plant lactobacillus powder and the propionibacterium freudenreichii subsp. Shermannii powder into the silage crops; filling and compacting, exhausting air and sealing; the using amount of the plant lactobacillus powder is controlled to be 20-50 mg / kg silage crops, and the using amount of the propionibacterium freudenreichii subsp. Shermannii powder is controlled to be 10-20 mg / kg silage crops; the prepared sealed crop is subjected to anaerobic dark fermentation at 30-37 DEG C for 45-90 days, and the corn silage is obtained. According to the present invention, the milk cow rumen carbon emission can be significantly inhibited, the greenhouse effect caused by the animal carbon emission can be easily relieved, the immune level of the milk cow and the quality of the fresh milk can be improved, and the animal feeding efficiency can be improved.

Description

Technical Field

[0001] The present invention relates to corn silage, in particular to corn silage for regulating carbon emission of ruminants based on Propionibacterium freudenreichii fermentation and a preparation method thereof. Background Art

[0002] Probiotics, used as silage starter cultures, can significantly improve silage quality and enhance animal nutrition and health. Probiotics are an effective means of enhancing silage quality. Selecting the right probiotic strain, combined with appropriate silage management practices, can significantly enhance the nutritional value of silage, thereby improving animal performance and health.

[0003] The development of the ruminant industry requires maintaining a balance between human needs, resource conservation, and environmental protection. Reducing methane emissions is particularly crucial. Dairy cows produce 30% of the world's total methane from livestock, and methane production also results in a 6% to 13% loss of total feed energy. Therefore, reducing methane emissions from ruminants, such as dairy cows, has become a research hotspot in animal nutrition.

[0004] The American Dairy Association reported a strong negative correlation between methane emissions from dairy cows and feeding efficiency. Reducing methane production by 2.5g per kilogram of standard milk increases standard milk production by 300mL per kilogram of feed. Therefore, reducing gastrointestinal methane emissions from dairy cows will help mitigate the greenhouse effect and improve feeding efficiency.

[0005] Probiotics enhance animal immunity through multiple mechanisms and are a promising alternative to antibiotics. They improve animal productivity and immunity by producing antimicrobial compounds and competing with pathogens for nutrients.

[0006] The development of new probiotic silage can provide ruminants with a nutritious, healthy feed option. Probiotics can also influence the fat and protein content and composition of milk, improving its nutritional profile and flavor, and meeting consumer demand for high-quality dairy products.

[0007] Chinese patent invention CN106721079B discloses a method for producing corn straw fiber feed, comprising the following steps: laying chopped grass on the bottom of the silo, chopping the corn straw into small segments, spreading a mixed inoculum of lactic acid cocci, Lactobacillus acidophilus, and actinomycetes on the chopped corn straw, and then adding ammonium tetraformate to mix thoroughly. During filling, the moisture content is controlled at 65% to 75%. After every 50-70 cm of filling, the corn straw is compacted and sealed, covered with plastic sheeting, and then filled with soil. When the raw material sinks and cracks appear on the roof and edges of the silo, they should be promptly filled with wet soil. The silo is fermented for 30-40 days. The use of silage in this invention can effectively improve the fermentation quality of corn straw fiber feed, resulting in a high success rate for fiber feed production and increased corn straw utilization. However, the fermentation products of lactic acid cocci, Lactobacillus acidophilus, and actinomycetes in this technology are mostly lactic acid, acetic acid, and other peptides. These bacteria or products do not reduce carbon emissions or improve milk quality.

[0008] Chinese invention patent CN112586442B discloses a method for raising dairy cows to increase the content of conjugated linoleic acid in milk, comprising the following steps: feeding each dairy cow with a total bacterial count greater than 10 11 The probiotics are Lactobacillus reuteri, Lactobacillus acidophilus, Propionibacterium freudenreichii subsp. schefflera, and Propionibacterium freudenreichii subsp. freudenreichii, and the unsaturated fatty acids constitute 5% or more of the total dry matter weight of the diet. This technology not only produces milk rich in conjugated linoleic acid (CLA), but also significantly increases feed intake, milk production, milk protein content, and antioxidant capacity. However, after transiting through the mouth and rumen, the viable bacteria ingested by cows using this technology do not remain in the rumen in sufficient numbers to affect the rumen's microbial composition, preventing the propionibacteria from exerting their carbon-reducing properties.

[0009] Chinese invention patent CN108570421B discloses a strain of Lactobacillus plantarum and its silage starter, the Lactobacillus plantarum strain, its deposit number is CGMCC No.12849. This strain is a homofermentative lactic acid bacterium that can produce a large amount of lactic acid, quickly reduce pH, is acid-resistant, has strong antibacterial activity against Staphylococcus aureus, Bacillus cereus, Salmonella, Escherichia coli and yeast, and has a high-efficiency adsorption effect on aflatoxin AFB1, which has obvious advantages over other strains. Using this strain as a silage starter can significantly improve the fermentation quality of silage, and after the silage is opened, it has an inhibitory effect on the production of aflatoxin AFB1 in the feed, and can be used as a silage starter in livestock and poultry farming. Although Lactobacillus plantarum can metabolize a variety of carbohydrates, produce organic acids such as lactic acid and acetic acid, and inhibit the growth of other microorganisms, it cannot compete with other methanogenic archaea for H by producing propionic acid. +To reduce carbon emissions from dairy cows. Summary of the Invention

[0010] In response to the above-mentioned problems existing in the prior art, the purpose of the present invention is to effectively reduce methane emissions of dairy cows by 70%-84% and carbon dioxide emissions by 44%-74%, and increase the nutritional content of milk (such as increasing lactose by more than 8%, milk fat by more than 31%, milk protein by more than 10%, etc.), and to provide a low-cost corn silage fermented by Propionibacterium freudenreichii subsp. shenrietta and Lactobacillus plantarum and a preparation method thereof.

[0011] The purpose of the present invention is achieved through the following technical solutions:

[0012] The preparation method of corn silage based on fermentation of Propionibacterium freudenreichii subsp. shenkelii and Lactobacillus plantarum comprises the following steps:

[0013] 1) Select whole corn plants or corn stalks from the late milky to early waxy stages and chop them short to obtain silage crops;

[0014] 2) Evenly spreading Lactobacillus plantarum powder and Propionibacterium freudenreichii subsp. schneiderii powder on the silage crop; filling and compacting, exhausting air and sealing; controlling the dosage of Lactobacillus plantarum powder to 20-50 mg / kg silage crop and the dosage of Propionibacterium freudenreichii subsp. schneiderii powder to 10-20 mg / kg silage crop; the viable cell count of Propionibacterium freudenreichii subsp. schneiderii powder is 1×10 9 CFU / g or above, the deposit number of Propionibacterium freudenreichii subsp. cheleriana is CGMCC1.2231;

[0015] 3) Fermenting the sealed crop prepared in step 2) anaerobically at 30-37° C. in the dark for 45-90 days to obtain corn silage.

[0016] To further achieve the purpose of the present invention, preferably, the moisture content of the whole corn plant or corn stalk from the late milky stage to the early waxy stage is 65wt%-70wt%, and the dry matter content remaining after removing the moisture is above 30wt%.

[0017] Preferably, the length of the chopped piece is 1-3 cm.

[0018] Preferably, the plant lactobacillus powder is a commercial starter, and the activity of the powder is 1×0 10 CFU / g or above;

[0019] The Propionibacterium freudenreichii subsp. schernobacterium powder is activated by Propionibacterium freudenreichii subsp. schernobacterium and expanded by MRS culture medium until the viable cell count is 1×10 9 CFU / mL or more, and store the obtained product after drying. The number of viable bacteria of Propionibacterium freudenreichii subspecies schizontella powder after drying is 1×10 9CFU / g or above;

[0020] Preferably, the drying is freeze drying, vacuum drying, low temperature spray drying, high temperature spray drying, vacuum foam drying or spray freeze drying.

[0021] Preferably, the anaerobic fermentation in the dark lasts for 60-90 days.

[0022] Preferably, the temperature of the anaerobic fermentation in the dark is 35-37°C.

[0023] Preferably, the dosage of the Lactobacillus plantarum powder is 30-40 mg / kg of silage crops.

[0024] Preferably, the added amount of the Propionibacterium freudenreichii subsp. schreiberi powder is 15-20 mg / kg of silage crops.

[0025] A corn silage fermented by Propionibacterium freudenreichii subsp. shermanii and Lactobacillus plantarum is prepared by the above-mentioned preparation method.

[0026] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0027] 1) The present invention utilizes Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231 and the commercial starter Lactobacillus plantarum to co-ferment whole-plant corn silage, which effectively improves silage quality. Compared to using Lactobacillus plantarum alone, the acid detergent-insoluble protein content and neutral detergent-insoluble protein content of whole-plant corn silage co-fermented with the present invention decrease, and the proportion of rumen-degradable protein to total protein increases. Moreover, the content of acid detergent fiber, neutral detergent fiber, deashed neutral detergent fiber, and lignin all decreases, while the content of starch, non-fibrous carbohydrates, and water-soluble carbohydrates increases, significantly improving silage quality.

[0028] 2) The whole-plant corn silage fermented by the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231 in the present invention can effectively reduce the emissions of methane and carbon dioxide from the rumen of dairy cows, help alleviate the greenhouse effect caused by carbon emissions from ruminants and improve the feeding efficiency of ruminants.

[0029] 3) Whole-plant corn silage fermented with the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schefflerae CGMCC1.2231 effectively improves the quality of fresh milk and the immune system of dairy cows, reducing the somatic cell count in cow serum. The contents of milk fat, lactose, milk protein, and non-fat milk solids in the fresh milk are all increased, while the proportions of unsaturated fatty acids and long unsaturated fatty acids in the fresh milk are significantly increased, significantly improving the fatty acid composition of the fresh milk. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 The figure shows the Kindom heat map of corn silage fermented with different probiotics in Examples and Comparative Examples.

[0031] Figure 2 1 is a genus-level microbial composition diagram of the products obtained in Examples and Comparative Examples.

[0032] Figure 3 Species-level microbial composition diagram of the products obtained in Examples and Comparative Examples. DETAILED DESCRIPTION

[0033] For a better understanding of the present invention, the present invention is further described below with reference to the accompanying drawings and specific embodiments, but the embodiments of the present invention are not limited thereto. The embodiments described are part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0034] The Propionibacterium freudenreichii subsp. Shermanii of the present invention refers to Propionibacterium freudenreichii subsp. Shermanii CGMCC1.2231, i.e., Propionibacterium freudenreichii subsp. Shermanii, with the accession number CGMCC1.2231 and the original number DSM490, originating directly from Germany. It was isolated, identified, and deposited by Zhou Yuguang on July 14, 1998. Propionibacterium freudenreichii subsp. Shermanii (CGMCC1.2231) is widely used in the production of Swiss-style cheese and is crucial to the formation of flavor and perforations. Propionic acid fermentation is the key to forming the unique flavor and perforations in Swiss cheese production.

[0035] The plant lactobacillus of the present invention is a commercial fermentation agent, a commercially available fermentation agent, purchased from Guangzhou Hua Microbiology Technology Co., Ltd., produced by Youchubao, the bacterial powder activity of 1×0 10 CFU / g or above; identified as containing only Lactobacillus plantarum and the detected bacterial activity reached 1×10 10 The address of the production unit is No. 1 Jiantashan Road, Luogang District, Guangzhou.

[0036] Existing feed technology already involves Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231, as demonstrated in Chinese invention patent CN112586442B. This technology produces milk rich in conjugated linoleic acid and also improves feed intake, milk production, milk protein content, and antioxidant capacity in lactating cows to a certain extent. Given that dairy cows produce 30% of the world's total methane from livestock, and methane production causes 6% to 13% of the total energy loss of feed, the present invention has discovered that fermenting corn silage with Propionibacterium freudenreichii subsp. cheleriani and Lactobacillus plantarum can effectively reduce methane emissions by 70%-84% and carbon dioxide emissions by 44%-74%, while also increasing the nutritional content of milk (e.g., increasing lactose by 8% or more, milk fat by 31% or more, and milk protein by 10% or more), while offering significant cost advantages.

[0037] On the one hand, Propionibacterium freudenreichii subsp. shenkelii can convert lactic acid, glucose, etc. into propionic acid, acetic acid, carbon dioxide and water; and Propionibacterium freudenreichii subsp. shenkelii CGMCC1.2231 has a strong ability to convert conjugated linoleic acid (CLA), which can affect the activity of signaling molecules such as mTOR and AMPK, as well as transcription factors such as THRSP, PPARG and SREBP, thereby regulating the expression of lipid synthesis genes such as acetyl CoA carboxylase, fatty acid synthase and stearoyl CoA desaturase 1, affecting the production of milk fat; at the same time, Propionibacterium freudenreichii subsp. shenkelii CGMCC1.2231 can also metabolize certain amino acids, such as aspartic acid and alanine, to produce propionic acid, acetic acid and carbon dioxide; Propionibacterium freudenreichii subsp. shenkelii CGMCC1.2231 also has immunomodulatory properties. By fermenting corn silage, these strains can be introduced into the diet of ruminants, thereby potentially improving their immune function; Propionibacterium freudenreichii subsp. shenkelii CGMCC1.2231 can also affect rumen fermentation in ruminants, indirectly affecting methane production by reducing cellulose content or changing the ratio of volatile fatty acids to unsaturated fatty acids.

[0038] On the other hand, inoculation with Lactobacillus plantarum can optimize the composition of the microbial community during the ensiling process, promote the growth of the dominant bacterial community required by the present invention, improve fermentation efficiency, and promote the function of the subspecies CGMCC1.2231 of Schefflera, thereby improving the quality of silage. The metabolites produced also have the effect of inhibiting mold growth, which can extend the shelf life of silage.

[0039] Specifically, the synergistic effect of Propionibacterium freudenreichii subsp. schreibungii and Lactobacillus plantarum is also reflected in:

[0040] Co-fermentation of corn silage with the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231 effectively inhibits methane and CO2 production, thereby reducing rumen carbon emissions, by reducing silage fiber or increasing silage unsaturated fatty acid concentrations. This dual-bacteria fermentation produces significant amounts of propionate. Approximately 82% of methane in the rumen is synthesized via hydrogenotrophic pathways. Propionate production consumes 4H, leading to reduced methane production. Unsaturated fatty acids are toxic to rumen microorganisms, modulating the activity of bacteria involved in biohydrogenation and suppressing methane production by sinking additional H.

[0041] Corn silage fermentation with the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231 improved the fatty acid composition of fresh milk from dairy cows. The unsaturated fatty acid content in fresh milk from cows fed corn silage enriched with unsaturated fatty acids was significantly increased. This is because rumen microorganisms biohydrogenate unsaturated fatty acids in the feed, but some unsaturated fatty acids escape rumen biohydrogenation and enter the small intestine for absorption, ultimately incorporating into milk fat. Unsaturated fatty acids also regulate milk fat synthesis through complex molecular mechanisms. Among them, miRNAs and circular RNAs play a key role in regulating unsaturated fatty acid synthesis in bovine mammary epithelial cells. For example, the miR-33a / LPPR4 pathway regulates unsaturated fatty acid synthesis in bovine mammary epithelial cells by affecting the expression of genes involved in fatty acid metabolism. circ09863 regulates unsaturated fatty acid metabolism in bovine mammary epithelial cells by recruiting miR-27a-3p. The circRNA-11228 / miR-103 / INSIG1 pathway is also involved in regulating milk fat synthesis in bovine mammary epithelial cells. Transcription factors such as SREBP1 and PPARγ are also crucial in regulating milk fat synthesis. Fatty acid binding protein 3 (FABP3), a target gene of SREBP1 and PPARγ, plays a role in the transport and metabolism of long-chain fatty acids, thereby affecting milk fat synthesis.

[0042] Corn silage fermented with the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schefflerae CGMCC1.2231 has demonstrated excellent immunomodulatory properties in dairy cows. This is attributed to the high concentration of unsaturated fatty acids imparted by the combined fermentation. Saturated fatty acids can modulate the immune system in animals by regulating the function of innate and adaptive immune cells and influencing the activation, differentiation, and function of immune cells such as macrophages and T cells. Some studies have shown that unsaturated fatty acids have antiviral activity, inactivating enveloped viruses. Cows fed corn silage fermented with Lactobacillus plantarum showed a significant decrease in the abundance of Bifidobacteria in their gut microbiota. However, consuming corn silage fermented with the combined fermentation helped maintain the abundance of beneficial intestinal bacteria, such as Bifidobacteria, in the cows' gut. Bifidobacteria, as one of the dominant intestinal bacteria, can influence the immune system through interactions with other microorganisms. For example, Bifidobacteria produce short-chain fatty acids, metabolites of which bind to receptors on immune cells, modulating signaling pathways such as NF-κB, thereby influencing inflammatory responses.

[0043] Based on the above findings, the present invention provides a method for preparing corn silage fermented with Propionibacterium freudenreichii subsp. schneideri and Lactobacillus plantarum, comprising the following steps:

[0044] 1) Select whole corn plants or corn stalks from the late milky to early waxy stages and chop them short to obtain silage crops;

[0045] 2) Evenly spreading Lactobacillus plantarum powder and Propionibacterium freudenreichii subsp. schneiderii powder on the silage crop; filling and compacting, exhausting air and sealing; controlling the dosage of Lactobacillus plantarum powder to 20-50 mg / kg silage crop and the dosage of Propionibacterium freudenreichii subsp. schneiderii powder to 10-20 mg / kg silage crop; the viable cell count of Propionibacterium freudenreichii subsp. schneiderii powder is 1×10 9 CFU / g or above, the deposit number of Propionibacterium freudenreichii subsp. cheleriana is CGMCC1.2231;

[0046] 3) Fermenting the sealed crop prepared in step 2) anaerobically at 30-37° C. in the dark for 45-90 days to obtain corn silage.

[0047] In the above method of the present invention, the moisture content of the whole corn plant or corn stover from the late milky to early waxy stage in step 1) is generally 65wt%-70wt%, and the dry matter content remaining after water removal is above 30wt%. The length of the short cut is preferably 1-3cm, which is also a common practice in the art.

[0048] As a further preferred step 2), the plant lactobacillus powder is a commercial starter, and the activity of the powder is 1×0 10CFU / g or above; Propionibacterium freudenreichii subsp. scheroni powder is activated by Propionibacterium freudenreichii subsp. scheroni and expanded by MRS medium until the viable cell count is 1×10 9 CFU / mL, the obtained product was dried and stored, and the number of viable bacteria of Propionibacterium freudenreichii subspecies shermanii powder after drying was 1×10 9 CFU / g or above; drying methods can be freeze drying, vacuum drying, low-temperature spray drying, high-temperature spray drying, vacuum foam drying, or spray freeze drying. The dosage of Lactobacillus plantarum powder is 30-40 mg / kg of silage. The dosage of Propionibacterium freudenreichii subsp. schefflerae powder is 15-20 mg / kg of silage.

[0049] Preferably, the anaerobic fermentation in the dark is carried out for 60-90 days; and the temperature of the anaerobic fermentation in the dark is 35-37°C.

[0050] In the following examples, the analysis and detection methods of the main indicators are as follows:

[0051] The feed post-feeding test of the present invention comprises the following steps:

[0052] 1) Rumen fluid was collected from three dairy cows and mixed with 0.1 M sodium phosphate buffer (pH 6.8) at a ratio of 1:1-10 to prepare rumen buffer. Then, under anaerobic conditions, 5-10 g of the prepared silage sample (packaged in a non-woven bag) and 200-400 ml of rumen buffer were added to each fermentation vessel. The contents of the fermentation vessel were continuously stirred at 25-50 rpm using a central propeller driven by a magnet. The temperature of the fermentation vessel was maintained at 37-39°C using a circulating hot water bath.

[0053] 2) After 36-48 h of anaerobic fermentation, the gas was sampled and the gas composition was detected by GC-MS.

[0054] 3) Whole-plant corn silage formula feed fermented with the obtained silage starter was fed to dairy cows for 23 days, and changes in serum immunomodulatory factors and milk components of the dairy cows were detected after the feeding period.

[0055] The rumen fluid can be from ruminants such as dairy cows, beef cattle, goats, and sheep, and the quantity can be one or more portions. The mixing ratio of rumen fluid and 0.1M sodium phosphate buffer (pH 6.8) is 1:1-10. In the in vitro simulated rumen fermentation experiment, 5-10g of the prepared silage sample (packaged in a non-woven bag) and 200-400ml of rumen mixed buffer solution are added to each fermentation container. The contents of the fermentation container are continuously stirred at a speed of 25-50r / min by a central propeller device driven by a magnet, and the temperature of the fermentation container is maintained at 37-39°C by a circulating hot water bath. The gas collection methods in the in vitro simulated rumen fermentation experiment include the gas collection bag method, the drainage gas collection method, and the like.

[0056] Main chemical components: The corn silage prepared in the present invention was delivered to Hangzhou Aike Testing Technology Co., Ltd. for chemical composition analysis of the sample using a near-infrared spectrometer according to the method of Wei Yong et al. (2019) (Wei Yong, Wang Dezhi, He Hong, et al. Application of near-infrared detection analyzer in corn silage quality detection [J]. China Animal Husbandry and Poultry Breeding, 2019, 15(01): 4-6.), including acidic detergent fiber, neutral detergent fiber, lignin, starch, non-fibrous carbohydrates, water-soluble carbohydrates, crude protein, etc.

[0057] Volatile fatty acid analysis: According to the method of Zhao Meng et al. (2025) (Zhao Meng, Ma Bo, Bai Haitao, et al. Effects of lactic acid bacteria exopolysaccharides on the physiological functions and gastrointestinal microorganisms of lambs [J]. Journal of Animal Nutrition, 2025, 37(04): 2498-2521.), 0.5 g of whole corn silage was added to 4 mL of ether and fully extracted by ultrasonication for 10 min. The extraction was repeated once, and the upper organic phases were combined and filtered through a 0.22 μm filter. The volatile fatty acid content of whole corn silage was analyzed by GC-MS.

[0058] Fatty acid content analysis: According to the method of Jinyue Zheng et al. (2021) (Jinyue Z, Di S, Xingxia L, et al. The effect of fatty acid chain length and saturation on the emulsification properties of pork myofibrillar proteins [J]. LWT, 2021, 139, 110242.), 0.5 g whole corn silage or 2 mL of milk was added to 4 mL of benzene-petroleum ether (1:1) and 4 mL of 0.4 M KOH-methanol. The mixture was thoroughly shaken and incubated in a 60°C water bath for 30 min. Then, 2 mL of n-hexane and 1 mL of saturated NaCl were added, and the mixture was centrifuged at 25°C × 850 g for 5 min. The extract was filtered through a 0.22 μm filter, and the fatty acid content of whole corn silage was analyzed by GC-MS using methyl hexadecanoate as an internal standard.

[0059] Gas composition analysis: According to the method of Yelim Choi et al. (2017) (Yelim C, Seongwoo L, Shindong K, et al. Determination of VOCs and Sulfuric Compounds generated from Biowaste Digester by using GC / MS with Gas Sampling Bag[J]. Proceedings of the Research Conference of the Society for Waste Resources Recycling, 2017, 28(0):550-551.), the rumen fluid contents in the simulated rumen container were sampled at different time points, and the gas production rate of each rumen container was monitored. After 48 hours of anaerobic fermentation, the gas was sampled and the gas composition was detected by GC-MS.

[0060] Milk composition analysis: The milk collected from the animal experiments in this invention was handed over to Guangdong Yantang Dairy Co., Ltd. to analyze the milk components, including milk fat, milk protein, lactose, non-fat solids, somatic cell count, etc., using a milk analyzer according to the method of Shen Lina et al. (2021) (Shen Lina, Wang Haitong, Luo Xuelu, et al. Analysis of factors affecting the content of fatty acids and conventional nutrients in milk [J]. Chinese Dairy, 2021, (02): 45-51.).

[0061] Serum cytokine analysis: Before and after the animal experiment, the present invention collects plasma from the experimental cows by puncturing the tail vein and stores it in a collection tube without anticoagulant. The supernatant is serum and centrifuged at 2000-3000 rpm for 5 minutes. According to the method of Zhao Meng et al. (2025) (Zhao Meng, Ma Bo, Bai Haitao, et al. Effects of lactic acid bacteria exopolysaccharides on the physiological functions and gastrointestinal microorganisms of lambs [J]. Journal of Animal Nutrition, 2025, 37(04): 2498-2521.), the concentration changes of cytokines in serum are analyzed using IL-1, IL-2, IL-6, IL-8, IgA, IgG, and IgM reagent analysis kits.

[0062] Whole corn plants were provided by Hubei Agricultural Base and chopped into pieces of 1-3 cm in length.

[0063] Example 1:

[0064] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g or more. Activate Propionibacterium freudenreichii subsp. Shermanii and expand the culture in MRS medium until the viable count reaches 1×10 9 CFU / mL or above, dry and store, the number of viable bacteria in the powder after drying should reach 1×10 9 CFU / g or above.

[0065] 2) Select corn stalks at the end of milky maturity to the beginning of waxy maturity, with an average moisture content of 65wt%-70wt%. Harvest corn stalks with a dry matter content of more than 30wt% after removing moisture, and chop them into pieces of 1-3 cm.

[0066] 3) Evenly sprinkle the Lactobacillus plantarum (powder preparation dosage of 20 mg / kg silage) and Propionibacterium freudenreichii subsp. shermanii (powder preparation dosage of 12 mg / kg silage) prepared in step 1) into the corn stalks prepared in step 2), fill and compact, expel air, and seal.

[0067] 4) fermenting the crops to be ensiled prepared in step 3) anaerobically at 37° C. in the dark for 45 days.

[0068] After the silage process was completed, the chemical composition changes and microbial composition differences of corn straw silage were analyzed (see Appendix Figure 1Analysis revealed that corn silage fermented with Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schneideri CGMCC1.2231 had a crude protein content of 9.15%, an acid insoluble protein content of 0.45%, a neutral insoluble protein content of 1.35%, a soluble protein content of 58.00% of total protein, a rumen-degradable protein content of 70.00%, an acid detergent fiber content of 16.00%, a neutral detergent fiber content of 30.40%, an ash-free neutral detergent fiber content of 29.45%, a lignin content of 1.60%, a non-fiber carbohydrate content of 53.60%, a starch content of 46.70%, and a water-soluble carbohydrate content of 3.00%. Furthermore, the bacterial abundance of corn silage fermented with Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schneideri CGMCC1.2231 was 99.20%, the fungal abundance was 0.02%, and the viral abundance was 0.76%.

[0069] Example 2

[0070] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g or more. Activate Propionibacterium freudenreichii subsp. Shermanii and expand the culture in MRS medium until the viable count reaches 1×10 9 CFU / mL, dried and stored, the number of viable bacteria in the powder after drying reaches 1×10 9 CFU / g and above.

[0071] 2) Whole corn plants at the late milky to early waxy stages are selected, with an average moisture content of 65 wt% to 70 wt%. Corn stalks with a dry matter content of more than 30 wt% after water removal are harvested and cut into pieces of 1 to 3 cm.

[0072] 3) Evenly sprinkle the Lactobacillus plantarum (powder preparation at a dosage of 20 mg / kg silage) and Propionibacterium freudenreichii subsp. shermanii (powder preparation at a dosage of 12 mg / kg silage) prepared in step 1) on the whole corn prepared in step 2), and then fill and compact them using a bag wrapping machine. Each bag weighs approximately 900 kg and is sealed and placed in the field in the dark for anaerobic fermentation for 60 days.

[0073] After the silage process was completed, the chemical composition changes and microbial composition differences of the whole corn silage were analyzed ( Figure 2 、 Figure 3The results showed that the crude protein content of the whole corn silage fermented by Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231 was 7.50%, the acid insoluble protein content was 0.70%, the neutral insoluble protein content was 1.03%, the acid detergent insoluble protein accounted for 7.40% of the total protein, the soluble protein accounted for 61.33% of the total protein, the rumen degradable protein accounted for 74.33% of the total protein, the acid detergent fiber content was 25.20%, the neutral detergent fiber content was 46.20%, the ash-free neutral detergent fiber content was 44.57%, the lignin content was 3.03%, the non-fiber carbohydrate content was 38.03%, the starch content was 25.03%, the water-soluble carbohydrate content was 3.37%, the acetic acid content was 12.00 μmol / g, and the propionic acid content was 17.62 μmol / g. In addition, the saturated fatty acid content of whole corn silage fermented by Lactobacillus plantarum was 38.19%, the unsaturated fatty acid content was 61.81%, the medium-chain fatty acid content was 7.67%, the long-chain fatty acid content was 92.33%, the long unsaturated fatty acid content was 56.80%, and the proportion of long unsaturated fatty acids to unsaturated fatty acids was 91.92%. Figure 2 As shown in the figure, the abundance of Lactobacillus at the genus level, the abundance of Lactobacillus at the genus level, the abundance of Lactobacillus at the genus level, the abundance of Lactobacillus at the genus level, and the abundance of Klebsiella at the genus level were 92.21%, 3.25%, and 0.76%, respectively. Figure 3 ), the abundance of Lactobacillus acidophilus was 79.83%, and the abundance of Lactobacillus amyloliquefaciens was 4.77%.

[0074] After the ensilage process was completed, rumen fluid from three cows was collected and mixed with 0.1M sodium phosphate buffer (pH 6.8) in a 1:1 ratio to prepare rumen buffer. Then, under anaerobic conditions, 10g of silage sample (packaged in a non-woven bag) and 400ml of rumen mixed buffer were added to each fermentation container. The contents of the fermentation container were continuously stirred at a speed of 25-50r / min by a central propeller device driven by a magnet, and the temperature of the fermentation container was maintained at 39°C by a circulating hot water bath. After 48h of anaerobic fermentation, the gas was sampled and the gas composition was detected by GC-MS. Analysis showed that after 48h of in vitro simulated rumen digestion of whole corn silage fermented with Lactobacillus plantarum and Propionibacterium freudenreichii subspecies CGMCC1.2231, the methane content in the gas was 0.39% and the carbon dioxide content was 12.87%, which was significantly reduced compared to the methane and carbon dioxide emissions in Comparative Example 2.

[0075] After ensilage was completed, the whole corn silage was mixed with a complete diet and fed to 80 randomly selected healthy dairy cows. After 23 days of feeding, serum and milk were collected from the experimental cows to analyze the effects of whole corn silage fermented with the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schneideri CGMCC1.2231 on the cows' immune system and milk quality. As shown in Table 2, compared to pre-experimental levels, serum levels of interleukin (IL)-1 increased by 31.06%, IL-2 increased by 51.20%, IL-6 decreased by 7.94%, IL-8 decreased by 18.76%, IgA decreased by 16.61%, IgG increased by 77.54%, and IgM increased by 13.23%. In addition, the milk produced by dairy cows has a milk fat content of 7.13%, a milk protein content of 3.94%, a lactose content of 4.91%, a non-fat milk solids content of 9.57%, and a somatic cell count of 9.16×10,000 / mL. The milk also contains 46.89% saturated fatty acids, 53.11% unsaturated fatty acids, 14.24% medium-chain fatty acids, 85.14% long-chain fatty acids, and 53.11% long unsaturated fatty acids, with the proportion of long unsaturated fatty acids to long-chain fatty acids being 62.39%.

[0076] Example 3

[0077] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g or more. Activate Propionibacterium freudenreichii subsp. Shermanii and expand the culture in MRS medium until the viable count reaches 1×10 9 CFU / mL, dried and stored, the number of viable bacteria in the powder after drying reaches 1×10 9 CFU / g and above.

[0078] 2) Whole corn plants at the late milky to early waxy stages are selected, with an average moisture content of 65 wt% to 70 wt%. Corn stalks with a dry matter content of more than 30 wt% after water removal are harvested and cut into pieces of 1 to 3 cm.

[0079] 3) evenly spreading the Lactobacillus plantarum (powder preparation at a dosage of 20 mg / kg silage) and Propionibacterium freudenreichii subsp. shermanii (powder preparation at a dosage of 10 mg / kg silage) prepared in step 1) on the whole corn prepared in step 2), followed by filling and compacting the whole corn using a bag wrapping machine. Each bag weighs approximately 1000 kg and is sealed and placed in the field in the dark for anaerobic fermentation for 80 days.

[0080] After the ensilage process was completed, rumen fluid from three cows was collected and mixed with 0.1M sodium phosphate buffer (pH 6.8) in a 1:1 ratio to prepare rumen buffer. Then, under anaerobic conditions, 10g of silage sample (packaged in a non-woven bag) and 400ml of rumen mixed buffer were added to each fermentation container. The contents of the fermentation container were continuously stirred at a speed of 25-50r / min by a central propeller device driven by a magnet, and the temperature of the fermentation container was maintained at 39°C by a circulating hot water bath. After 48h of anaerobic fermentation, the gas was sampled and the gas composition was detected by GC-MS. Analysis showed that after 48h of in vitro simulated rumen digestion of whole corn silage fermented with Lactobacillus plantarum and Propionibacterium freudenreichii subspecies CGMCC1.2231, the methane content in the gas was 0.74% and the carbon dioxide content was 5.80%, which was significantly reduced compared to the methane and carbon dioxide emissions in Comparative Example 2.

[0081] Example 4

[0082] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g or more. Activate Propionibacterium freudenreichii subsp. Shermanii and expand the culture in MRS medium until the viable count reaches 1×10 9 CFU / mL, dried and stored, the number of viable bacteria in the powder after drying reaches 1×10 9 CFU / h or above.

[0083] 2) Whole corn plants at the late milky to early waxy stages are selected, with an average moisture content of 65 wt% to 70 wt%. Corn stalks with a dry matter content of more than 30 wt% after water removal are harvested and cut into pieces of 1 to 3 cm.

[0084] 3) evenly spreading the Lactobacillus plantarum (powder preparation at a dosage of 30 mg / kg silage) and Propionibacterium freudenreichii subsp. shermanii (powder preparation at a dosage of 12 mg / kg silage) prepared in step 1) on the whole corn prepared in step 2), followed by filling and compacting the whole corn using a bag wrapping machine. Each bag weighs approximately 1000 kg and is sealed and placed in the field in the dark for anaerobic fermentation for 80 days.

[0085] Example 5

[0086] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g and above.

[0087] 2) Whole corn plants at the late milky to early waxy stages are selected, with an average moisture content of 65 wt% to 70 wt%. Corn stalks with a dry matter content of more than 30 wt% after water removal are harvested and cut into pieces of 1 to 3 cm.

[0088] 3) Evenly sprinkle the Lactobacillus plantarum (powder preparation at a dosage of 50 mg / kg silage) and Propionibacterium freudenreichii subsp. shermanii (powder preparation at a dosage of 20 mg / kg silage) prepared in step 1) on the whole corn prepared in step 2), and then fill and compact them using a bag wrapping machine. Each bag weighs about 1000 kg and is placed in a sealed, dark-proof field for anaerobic fermentation for 60 days.

[0089] Comparative Example 1: Corn straw silage fermented with commercial fermentation agent Lactobacillus plantarum

[0090] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g and above.

[0091] 2) Select corn stalks at the end of milky maturity to the beginning of waxy maturity, with an average moisture content of 65wt%-70wt%. Harvest corn stalks with a dry matter content of more than 30wt% after removing moisture, and chop them into pieces of 1-3 cm.

[0092] 3) The Lactobacillus plantarum prepared in step 1) (the dosage of the powder preparation is 20 mg / kg silage) is evenly spread on the corn stalks prepared in step 2), filled and compacted, and the air is exhausted and sealed.

[0093] 4) fermenting the crops to be ensiled prepared in step 3) anaerobically at 37° C. in the dark for 45 days.

[0094] After the silage process was completed, the chemical composition changes and microbial composition differences of corn straw silage were analyzed (see Appendix Figure 1Tests showed that the crude protein content of corn silage fermented by Lactobacillus plantarum was 8.45%, the acid insoluble protein content was 0.50%, the neutral insoluble protein content was 1.00%, the soluble protein accounted for 56.00% of the total protein, the rumen-degradable protein accounted for 66.50% of the total protein, the acid detergent fiber content was 17.75%, the neutral detergent fiber content was 29.85%, the ash-free neutral detergent fiber content was 30.15%, the lignin content was 2.25%, the non-fiber carbohydrate content was 52.80%, the starch content was 44.50%, and the water-soluble carbohydrate content was 2.55%. In addition, the bacterial abundance of corn silage fermented by Lactobacillus plantarum was 98.28%, the fungal abundance was 0.12%, and the viral abundance was 1.60%.

[0095] Comparative Example 2: Whole-plant corn silage fermented with commercial fermentation agent Lactobacillus plantarum

[0096] 1) Preparation of silage starter: The commercial starter Lactobacillus plantarum was purchased from Youchubao, a manufacturer of Guangzhou Huawei Microbiology Technology Co., Ltd. It was identified as containing only Lactobacillus plantarum with a bacterial activity of 1×10 10 CFU / g and above.

[0097] 2) Whole corn plants at the late milky to early waxy stages are selected, with an average moisture content of 65 wt% to 70 wt%. Corn stalks with a dry matter content of more than 30% after water removal are harvested and cut into pieces of 1 to 3 cm.

[0098] 3) Evenly spreading the Lactobacillus plantarum prepared in step 1) (the amount of the powder preparation is 20 mg / kg silage) on the whole corn prepared in step 2), and then filling and compacting it with a bag wrapper. Each bag weighs about 900 kg and is placed in a sealed, dark-proof field for anaerobic fermentation for 60 days.

[0099] After the silage process was completed, the chemical composition changes (see Table 1) and microbial composition differences (see Appendix Figure 2 、 Figure 3). Tests showed that the crude protein content of whole-plant corn silage fermented by Lactobacillus plantarum was 8.50%, the acid insoluble protein content was 0.73%, the neutral insoluble protein content was 1.23%, the acid detergent insoluble protein accounted for 8.57% of the total protein, the ratio of soluble protein to total protein was 60.67%, the rumen-degradable protein accounted for 73.67% of the total protein, the acid detergent fiber content was 29.03%, the neutral detergent fiber content was 49.43%, the ash-free neutral detergent fiber content was 48.67%, the lignin content was 3.40%, the non-fiber carbohydrate content was 36.00%, the starch content was 23.10%, the water-soluble carbohydrate content was 3.20%, the acetic acid content was 7.38 μmol / g, and the propionic acid content was 7.19 μmol / g. In addition, the saturated fatty acid content of whole corn silage fermented by Lactobacillus plantarum in the pilot test was 42.47%, the unsaturated fatty acid content was 57.53%, the medium-chain fatty acid content was 5.18%, the long-chain fatty acid content was 94.82%, the long unsaturated fatty acid content was 52.04%, and the proportion of long unsaturated fatty acids to unsaturated fatty acids was 90.45%. Figure 2 As shown in Figure 2, the abundance of Lactobacillus genus, Lactobacillus mucilaginosus genus and Klebsiella genus in whole corn silage fermented by Lactobacillus plantarum was 83.23% at the genus level, 6.45% at the genus level and 2.10% at the species level (see Appendix). Figure 3 ), the abundance of Lactobacillus acidophilus was 65.25%, and the abundance of Lactobacillus amyloliquefaciens was 10.03%.

[0100] After the ensiling process was completed, rumen fluid from three cows was collected and mixed with 0.1M sodium phosphate buffer (pH 6.8) in a 1:1 ratio to prepare rumen buffer. Then, under anaerobic conditions, 10g of silage sample (packaged in a non-woven bag) and 400ml of rumen mixing buffer were added to each fermentation container. The contents of the fermentation container were continuously stirred at a speed of 25-50r / min by a central propeller device driven by a magnet, and the temperature of the fermentation container was maintained at 39°C by a circulating hot water bath. After 48 hours of anaerobic fermentation, the gas was sampled and the gas composition was detected by GC-MS. Analysis showed that after 48 hours of simulated rumen digestion in vitro of whole-plant corn silage fermented by Lactobacillus plantarum, the methane content in the gas was 2.46% and the carbon dioxide content was 22.93%.

[0101] After ensilage, the whole corn silage was mixed with a complete diet and fed to 80 randomly selected healthy dairy cows. After 23 days of feeding, serum and milk were collected from the experimental cows to analyze the effects of whole corn silage fermented with the commercial starter Lactobacillus plantarum on the cows' immune system and milk quality. As shown in Table 2, compared to pre-experimental levels, serum IL-1 levels increased by 24.25%, IL-2 levels increased by 60.57%, IL-6 levels decreased by 11.70%, IL-8 levels decreased by 7.67%, IgA levels decreased by 19.95%, IgG levels increased by 43.72%, and IgM levels increased by 13.66%. Furthermore, the milk produced by the cows had a milk fat content of 5.42%, milk protein content of 3.56%, lactose content of 4.54%, non-fat milk solids content of 8.71%, and a somatic cell count of 103.27 × 10,000 / mL. The content of saturated fatty acids in fresh milk is 56.67%, the content of unsaturated fatty acids is 42.42%, the content of medium-chain fatty acids is 8.39%, the content of long-chain fatty acids is 91.36%, the content of long unsaturated fatty acids is 31.72%, and the proportion of long unsaturated fatty acids to long-chain fatty acids is 34.72%.

[0102] Table 1 Chemical component contents of whole-plant corn silage fermented by Lactobacillus plantarum and Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231

[0103]

[0104]

[0105] Note: In Table 1, LP: whole-plant corn silage fermented by Lactobacillus plantarum; L.P+1.2231: whole-plant corn silage fermented jointly by Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schneideri CGMCC1.2231.

[0106] In summary, as shown in Table 2, after the commercial fermentation agent Lactobacillus plantarum and Propionibacterium freudenreichii subspecies CGMCC1.2231 were used to co-ferment corn silage in Example 2, the acid washing insoluble protein content of the whole plant corn silage decreased by 4.5%, the neutral washing insoluble protein content decreased by 16.2%, the proportion of rumen degradable protein in the total protein increased by 0.9%, the acid detergent fiber content decreased by 10.9%, the neutral detergent fiber content decreased by 5.2%, the ash-free neutral detergent fiber content decreased by 8.4%, the lignin content decreased by 10.8%, the starch content increased by 8.4%, the non-fibrous carbohydrates increased by 2.9%, and the water-soluble carbohydrate content increased by 5.2%. In addition, after the commercial fermentation agent Lactobacillus plantarum and Propionibacterium freudenreichii subspecies CGMCC1.2231 was used to co-ferment, the acetic acid and propionic acid contents of the whole plant corn silage increased significantly, the saturated fatty acid and long-chain fatty acid contents decreased, and the unsaturated fatty acid and medium-chain fatty acid contents increased. Acetic acid is a strong antibacterial agent that can inhibit the growth of mold, yeast and certain anaerobic bacteria, thereby improving the storage stability of silage. Propionic acid has a strong antifungal effect and can effectively prevent silage from becoming moldy during storage. Propionic acid can also improve the aerobic stability of silage and prevent silage from rapidly deteriorating after being exposed to air. The combined use of acetic acid and propionic acid can play a synergistic role and further improve the fermentation quality and storage stability of silage. In addition, in a small-scale silage test (see Appendix Figure 1 ) and wrapping pilot (attached Figure 2 、 Figure 3 ) clearly showed that the combined fermentation of corn silage with the commercial starter Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schreibungii CGMCC1.2231 significantly reduced the abundance of viruses and fungi in the silage. Simultaneously, the abundance of Klebsiella, a genus known to cause inflammation, decreased significantly, while the abundance of beneficial silage bacteria such as Lactobacillus acidophilus and Lactobacillus plantarum increased. Meanwhile, the abundance of Acetobacter batrix and Lactobacillus mucilaginosus, which can synthesize cellulose but are detrimental to cellulose degradation, decreased. This phenomenon is likely attributed to the powerful acid production during the combined fermentation process, which is consistent with the results obtained in Examples 1 and 2.

[0107] In vitro simulated rumen fermentation tests showed that whole-plant corn silage fermented with Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schreibensis CGMCC1.2231 effectively reduced methane and carbon dioxide emissions compared to commercial starter cultures. However, both whole-plant corn silage fermented with Lactobacillus plantarum and supplemented with P. freudenreichii subsp. schreibensis CGMCC1.2231 increased methane and carbon dioxide levels to varying degrees in the in vitro simulated rumen fermentation experiments, indicating that P. freudenreichii subsp. schreibensis CGMCC1.2231 does not directly reduce methane and carbon dioxide emissions from rumen fluid. Approximately 82% of methane in the rumen is synthesized via the hydrogenotrophic pathway. Fermentation of whole-plant corn silage with the commercial starter cultures Lactobacillus plantarum and P. freudenreichii subsp. schreibensis CGMCC1.2231 produces significant amounts of acetic and propionic acids. The production of propionate consumes 4[H], resulting in reduced methane production. At the same time, the co-fermentation of whole-plant corn silage with the commercial fermentation agents Lactobacillus plantarum and Propionibacterium freudenreichii subsp. shenkelii CGMCC1.2231 significantly increased the content of unsaturated fatty acids in the silage. Unsaturated fatty acids are toxic to rumen microorganisms, can regulate the activity of bacteria involved in the biohydrogenation process, and inhibit methane production by sinking additional [H].

[0108] As shown in Table 2, compared with the diet containing whole-plant corn silage fermented by Lactobacillus plantarum, the concentrations of cytokines such as IL-1 and IgG in the serum of dairy cows fed a mixed complete diet containing whole-plant corn silage fermented by both Lactobacillus plantarum and Propionibacterium freudenreichii subsp. chelerianii CGMCC1.2231 increased, while the concentration of the pro-inflammatory factor IL-8 decreased (compared with before feeding), indicating that whole-plant corn silage fermented by both bacteria can also improve the immune level of dairy cows.

[0109] Table 2 Changes in serum immunomodulatory factor concentrations in dairy cows before and after ingestion of a complete diet containing whole corn silage fermented with Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schreibungii CGMCC1.2231

[0110]

[0111] Note: In Table 2, LP: whole-plant corn silage fermented by Lactobacillus plantarum; L.P+1.2231: whole-plant corn silage fermented jointly by Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani GMCC1.2231.

[0112] In addition, compared with the diet containing whole-plant corn silage fermented by Lactobacillus plantarum, the fresh milk of dairy cows fed a mixed complete diet containing whole-plant corn silage fermented by both Lactobacillus plantarum and Propionibacterium freudenreichii subsp. schreibungii CGMCC1.2231 had increased unsaturated fatty acid and long-unsaturated fatty acid contents, and decreased saturated fatty acid and long-chain fatty acid contents. At the same time, the contents of milk fat, lactose, milk protein, and non-fat milk solids in the fresh milk all increased to varying degrees.

[0113] In general, the use of commercial fermentation agents Lactobacillus plantarum and Propionibacterium freudenreichii subsp. cheleriani CGMCC1.2231 to jointly ferment corn silage can not only effectively reduce the fiber content of silage, increase the content of carbohydrates such as starch, and effectively improve the quality of corn silage, but also significantly reduce the rumen carbon emissions of dairy cows, while improving the immune level of dairy cows and the quality of fresh milk produced.

[0114] It should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the above embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the above embodiments, or replace some or all of the technical features therein with equivalents. However, these modifications or replacements do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subspecies shermanii and Lactobacillus plantarum, characterized in that The steps include: 1) Select whole corn plants or corn stalks from the late milky to early waxy stages and chop them short to obtain silage crops; 2) Evenly spreading Lactobacillus plantarum powder and Propionibacterium freudenreichii subsp. schneiderii powder on the silage crop; filling and compacting, exhausting air and sealing; controlling the dosage of Lactobacillus plantarum powder to 20-50 mg / kg silage crop and the dosage of Propionibacterium freudenreichii subsp. schneiderii powder to 10-20 mg / kg silage crop; the viable cell count of Propionibacterium freudenreichii subsp. schneiderii powder is 1×10 9 CFU / g or above, the deposit number of Propionibacterium freudenreichii subsp. cheleriana is CGMCC1.2231; 3) Fermenting the sealed crop prepared in step 2) anaerobically at 30-37° C. in the dark for 45-90 days to obtain corn silage.

2. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii and Lactobacillus plantarum according to claim 1, characterized in that: The moisture content of the whole corn plant or corn stalk from the late milky stage to the early waxy stage is 65wt%-70wt%.

3. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shermanii and Lactobacillus plantarum according to claim 1, characterized in that, The short length of the guillotine is 1-3 cm.

4. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shermanii and Lactobacillus plantarum according to claim 1, characterized in that: The plant lactobacillus powder is a commercial starter, and the activity of the powder is 1×0 10 CFU / g or above; The Propionibacterium freudenreichii subsp. schernobacterium powder is activated by Propionibacterium freudenreichii subsp. schernobacterium and expanded by MRS culture medium until the viable cell count is 1×10 9 CFU / mL or more, and store the obtained content after drying.

5. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shenkelii and Lactobacillus plantarum according to claim 4, characterized in that, The drying is freeze drying, vacuum drying, low temperature spray drying, high temperature spray drying, vacuum foam drying or spray freeze drying.

6. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shenkelii and Lactobacillus plantarum according to claim 1, characterized in that: The time of the anaerobic light-proof fermentation is 60-90 days.

7. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shenkelii and Lactobacillus plantarum according to claim 1, characterized in that: The temperature of the anaerobic light-proof fermentation is 35-37°C.

8. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shenkelii and Lactobacillus plantarum according to claim 1, characterized in that: The dosage of the plant lactobacillus powder is 30-40 mg / kg of silage crops.

9. The method for preparing corn silage based on fermentation of Propionibacterium freudenreichii subsp. shermanii and Lactobacillus plantarum according to claim 1, characterized in that: The added amount of the Propionibacterium freudenreichii subspecies shermanii bacterial powder is 15-20 mg / kg of silage crops.

10. A corn silage fermented by Propionibacterium freudenreichii subsp. shermanii and Lactobacillus plantarum, characterized in that: The invention discloses a novel novel polyol according to claim 1 .

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