Lactobacillus plantarum microbial preparation as well as preparation method and application thereof

Abstract

The invention provides a lactobacillus plantarum microbial preparation as well as a preparation method and application thereof, and relates to the technical field of microbial preparations. The lactobacillus plantarum microbial preparation is prepared from lactobacillus plantarum fermentation liquor, bacillus subtilis fermentation liquor, candida utilis fermentation liquor, potassium nitrate, dipotassium phosphate, honey, fructo-oligosaccharide, pectin, whey protein, glycerin, lecithin and clear water. According to the invention, through combined use, mutual cooperation and synergistic interaction of a plurality of microbial strains, the feed additive contains rich antibacterial substances capable of preventing and treating plant diseases, can inhibit germination of harmful pathogen spores, has the effects of killing fungi, bacteria, viruses and mycoplasmas, preventing diseases and insect pests, promoting growth of beef cattle, and improving the quality of beef cattle. The nutrient apparent digestibility, the blood antioxidant level and the rumen protease activity can be improved, the antioxidant level can also be improved, the activity of part of rumen digestive enzymes can be improved, and the digestion and absorption of nutrient substances can be improved.

Description

Technical Field

[0001] The present invention relates to the technical field of microbial preparations, in particular to a Lactobacillus plantarum microbial preparation and a preparation method and application thereof. Background Art

[0002] Lactobacillus plantarum is a Gram-positive bacterium belonging to the genus Lactobacillus. It is widely found in fermented plant foods, animal intestines, and the natural environment. Microbial preparations developed with Lactobacillus plantarum are widely used in food, feed, agriculture, and medicine due to their safety and significant probiotic properties.

[0003] The development of efficient and precision agriculture is an inevitable trend in Chinese agriculture, and the area of ​​greenhouse vegetable cultivation has grown rapidly across the country. While greenhouse cultivation has led to increased vegetable yields, the frequent occurrence of pests and diseases is a key factor limiting farmers' income growth. This has led to the extensive use of chemical pesticides for pest control, increasing labor input and reducing vegetable quality.

[0004] In recent years, the concepts and practices of biological control, using insects to control insects, and using bacteria to inhibit bacteria have been accepted by farmers. However, the common biological control methods on the market are still limited to the purchase of biological pesticides and commercial natural enemy insects. They are relatively passive to use, time-lagged, slow to take effect, and require high investment.

[0005] Therefore, those skilled in the art provide a Lactobacillus plantarum microbial preparation and a preparation method and application thereof to solve the problems raised in the above background technology. Summary of the Invention

[0006] In view of the deficiencies in the prior art, the present invention provides a plant lactobacillus microbial preparation and its preparation method and application, which, through the combined use of multiple microbial strains, cooperates with each other, synergizes and enhances efficiency, so that plant lactobacillus, bacillus subtilis, and Candida utilis are effectively propagated, and contains rich antimicrobial substances with plant disease prevention and treatment, including lipopeptides, peptides, phospholipids, amino acids, nucleic acids, lipopeptide antibiotics, surfactants and polyene substances and other compounds, which can inhibit the germination of harmful pathogenic bacteria spores, kill fungi, bacteria, viruses and mycoplasmas, and can achieve the effect of preventing diseases and insect pests. It solves the problem that common biological control methods on the market still stay on the purchase and investment of biological pesticides and commercial natural enemy insects, are relatively passive in use, have a time lag, are slow to take effect, and have a high investment.

[0007] To achieve the above objectives, the present invention is implemented through the following technical solutions: A lactobacillus plantarum microbial preparation comprises raw material components including lactobacillus plantarum fermentation liquid, bacillus subtilis fermentation liquid, Candida utilis fermentation liquid, potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin and clean water.

[0008] In one embodiment of the present invention, the weight ratio of the raw materials of the microbial preparation is: 70-90 parts of Lactobacillus plantarum fermentation broth, 70-90 parts of Bacillus subtilis fermentation broth, 90-110 parts of Candida utilis fermentation broth, 8-12 parts of potassium nitrate, 18-22 parts of dipotassium hydrogen phosphate, 4-6 parts of honey, 70-90 parts of oligofructose, 40-60 parts of pectin, 70-90 parts of whey protein, 80-100 parts of glycerol, 40-60 parts of lecithin, and 600-700 parts of water.

[0009] In one embodiment of the present invention, the weight ratio of the raw materials of the microbial preparation is preferably: 80 parts of Lactobacillus plantarum fermentation broth, 80 parts of Bacillus subtilis fermentation broth, 100 parts of Candida utilis fermentation broth, 10 parts of potassium nitrate, 20 parts of dipotassium hydrogen phosphate, 5 parts of honey, 80 parts of fructooligosaccharides, 50 parts of pectin, 80 parts of whey protein, 90 parts of glycerol, 50 parts of lecithin, and 650 parts of water.

[0010] In one embodiment of the present invention, the Lactobacillus plantarum fermentation broth is made from Lactobacillus plantarum, and the Bacillus subtilis fermentation broth is made from Bacillus subtilis.

[0011] In one embodiment of the present invention, the Candida utilis fermentation broth is prepared from Candida utilis.

[0012] In one embodiment of the present invention, the method for preparing the Lactobacillus plantarum microbial preparation comprises the following steps: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2 using liquid submerged fermentation, the culture medium is mainly glucose, peptone, yeast extract, control the temperature, pH and ventilation, promote bacterial proliferation, obtain Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor in a weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 80 to 150 r / min speed for 5 to 10 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

[0013] In one embodiment of the present invention, the method for obtaining the plant lactobacillus fermentation broth is: 1) Inoculate freeze-dried Lactobacillus plantarum powder into MRS medium and culture at 37°C for 24 hours; 2) Then, take 1% of the inoculum and inoculate it into the fermentation medium. Ferment and culture at 37°C for 16-18 hours. 3) During the entire fermentation period, glucose solution is intermittently added to the fermentation system to maintain the glucose concentration in the fermentation system at 5-15 g / L to obtain Lactobacillus plantarum fermentation broth; The method for obtaining the Bacillus subtilis fermentation broth is: 1) Streak Bacillus subtilis onto LB solid medium and incubate at 37°C for 48 hours. 2) Then, use an inoculation loop to pick up the bacteria and inoculate them into liquid LB medium. Incubate at 180 rpm and 37°C for 24 hours. 3) inoculating a corn flour and soybean meal culture medium with a 1% inoculum size for fermentation at the following conditions: temperature 37°C, tank pressure 0.1 MPa, ventilation 0.8-1 vvm, rotation speed 180 rpm, and culture time 2-3 days to obtain a Bacillus subtilis fermentation liquid; The method for obtaining the Candida utilis fermentation broth is as follows: The freeze-dried Lactobacillus plantarum powder was inoculated into MRS culture medium and cultured at 37°C for 24 hours; 2) Then, 1% of the inoculum amount was inoculated into the fermentation medium, and fermented at 37°C for 20 hours to obtain the Candida utilis fermentation broth.

[0014] The present invention provides the Lactobacillus plantarum microbial preparation and application of the Lactobacillus plantarum microbial preparation prepared by the method in preventing and controlling greenhouse vegetable diseases and insect pests.

[0015] In one embodiment of the present invention, the method for using the Lactobacillus plantarum microbial preparation is to dilute the Lactobacillus plantarum microbial preparation and irrigate the roots or spray the entire vegetable leaves during the growth period.

[0016] The present invention provides a Lactobacillus plantarum microbial preparation, a preparation method thereof, and an application thereof. The preparation has the following beneficial effects: 1. The present invention provides a Lactobacillus plantarum microbial preparation, a preparation method thereof, and an application thereof. The preparation method effectively reproduces Lactobacillus plantarum, Bacillus subtilis, and Candida utilis through the combined use of multiple microbial strains, which cooperate with each other and enhance synergy. The preparation method contains rich antibacterial substances with plant disease prevention and control functions, including lipopeptides, peptides, phospholipids, amino acids, nucleic acids, lipopeptide antibiotics, surfactants, polyenes, and other compounds. These can inhibit the germination of harmful pathogenic bacteria spores, kill fungi, bacteria, viruses, and mycoplasmas, and can prevent diseases and insect pests. Compared with similar commercially available strains, the preparation method has a more excellent fermentation effect.

[0017] 2. The present invention provides a Lactobacillus plantarum microbial preparation, a preparation method thereof, and an application thereof. The Lactobacillus plantarum microbial preparation has the effect of promoting the growth of beef cattle, improving the apparent digestibility of nutrients, blood antioxidant levels, and rumen protease activity, and can also improve antioxidant levels, increase the activity of some rumen digestive enzymes, and improve the digestion and absorption of nutrients. DETAILED DESCRIPTION

[0018] The following is a clear and complete description of the technical solutions in the embodiments of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts are within the scope of protection of the present invention.

[0019] Example 1

[0020] Obtaining Lactobacillus plantarum fermentation broth 1) Inoculate freeze-dried Lactobacillus plantarum powder into MRS medium and culture at 37°C for 24 hours; 2) Then, take 1% of the inoculum and inoculate it into the fermentation medium. Ferment and culture at 37°C for 16-18 hours. 3) During the entire fermentation period, glucose solution needs to be intermittently added to the fermentation system to maintain the glucose concentration in the fermentation system at 5-15 g / L to obtain Lactobacillus plantarum fermentation liquid.

[0021] Example 2

[0022] Obtaining Bacillus subtilis fermentation broth 1) Streak Bacillus subtilis onto LB solid medium and incubate at 37°C for 48 hours. 2) Then, use an inoculation loop to pick up the bacteria and inoculate them into liquid LB medium. Incubate at 180 rpm and 37°C for 24 hours. 3) Inoculating a corn flour and soybean meal culture medium with a 1% inoculum size for fermentation under the following conditions: temperature 37°C, tank pressure 0.1 MPa, ventilation 0.8-1 vvm, rotation speed 180 rpm, and culture time 2-3 days to obtain a Bacillus subtilis fermentation broth.

[0023] Example 3

[0024] Obtaining Candida utilis fermentation broth 1) Inoculate freeze-dried Lactobacillus plantarum powder into MRS medium and culture at 37°C for 24 hours; 2) Then, 1% of the inoculum amount was inoculated into the fermentation medium, and fermented at 37°C for 20 hours to obtain the Candida utilis fermentation broth.

[0025] Example 4

[0026] A Lactobacillus plantarum microbial preparation comprises the following raw materials in the following weight ratios: 80 parts of Lactobacillus plantarum fermentation liquid, 80 parts of Bacillus subtilis fermentation liquid, 100 parts of Candida utilis fermentation liquid, 10 parts of potassium nitrate, 20 parts of dipotassium hydrogen phosphate, 5 parts of honey, 80 parts of fructooligosaccharides, 50 parts of pectin, 80 parts of whey protein, 90 parts of glycerol, 50 parts of lecithin, and 650 parts of clean water.

[0027] The number of viable bacteria in the Lactobacillus plantarum fermentation liquid is 4.5 billion / gram, the number of viable bacteria in the Bacillus subtilis fermentation liquid is 2.5 billion / gram, and the number of viable bacteria in the Candida utilis fermentation liquid is 900 million / gram. The Lactobacillus plantarum and Candida utilis strains are both isolated from healthy egg-laying red wasps, and the Bacillus subtilis is a preserved Bacillus subtilis strain.

[0028] The preparation method of the Lactobacillus plantarum microbial preparation comprises the following steps: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2 using liquid submerged fermentation, the culture medium is mainly glucose, peptone, yeast extract, control the temperature, pH and ventilation, promote bacterial proliferation, obtain Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor according to weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 120 r / min speed for 8 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

[0029] Example 5

[0030] A Lactobacillus plantarum microbial preparation comprises the following raw materials in the following weight ratios: 70 parts of Lactobacillus plantarum fermentation liquid, 70 parts of Bacillus subtilis fermentation liquid, 90 parts of Candida utilis fermentation liquid, 8 parts of potassium nitrate, 18 parts of dipotassium hydrogen phosphate, 4 parts of honey, 70 parts of oligofructose, 40 parts of pectin, 70 parts of whey protein, 80 parts of glycerol, 40 parts of lecithin, and 600 parts of clean water.

[0031] The number of viable bacteria in the Lactobacillus plantarum fermentation liquid is 4.5 billion / gram, the number of viable bacteria in the Bacillus subtilis fermentation liquid is 2.5 billion / gram, and the number of viable bacteria in the Candida utilis fermentation liquid is 900 million / gram. The Lactobacillus plantarum and Candida utilis strains are both isolated from healthy egg-laying red wasps, and the Bacillus subtilis is a preserved Bacillus subtilis strain.

[0032] The preparation method of the Lactobacillus plantarum microbial preparation comprises the following steps: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2 using liquid submerged fermentation, the culture medium is mainly glucose, peptone, yeast extract, control the temperature, pH and ventilation, promote bacterial proliferation, obtain Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor according to weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 120 r / min speed for 8 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

[0033] Example 6

[0034] A Lactobacillus plantarum microbial preparation comprises the following raw materials in the following weight ratios: 90 parts of Lactobacillus plantarum fermentation liquid, 90 parts of Bacillus subtilis fermentation liquid, 110 parts of Candida utilis fermentation liquid, 12 parts of potassium nitrate, 22 parts of dipotassium hydrogen phosphate, 6 parts of honey, 90 parts of fructooligosaccharides, 60 parts of pectin, 90 parts of whey protein, 100 parts of glycerol, 60 parts of lecithin, and 700 parts of clean water.

[0035] The number of viable bacteria in the Lactobacillus plantarum fermentation liquid is 4.5 billion / gram, the number of viable bacteria in the Bacillus subtilis fermentation liquid is 2.5 billion / gram, and the number of viable bacteria in the Candida utilis fermentation liquid is 900 million / gram. The Lactobacillus plantarum and Candida utilis strains are both isolated from healthy egg-laying red wasps, and the Bacillus subtilis is a preserved Bacillus subtilis strain.

[0036] The preparation method of the Lactobacillus plantarum microbial preparation comprises the following steps: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2 using liquid submerged fermentation, the culture medium is mainly glucose, peptone, yeast extract, control the temperature, pH and ventilation, promote bacterial proliferation, obtain Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor according to weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 120 r / min speed for 8 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

[0037] Example 7

[0038] A Lactobacillus plantarum microbial preparation comprises the following raw materials in the following weight ratios: 75 parts of Lactobacillus plantarum fermentation liquid, 75 parts of Bacillus subtilis fermentation liquid, 95 parts of Candida utilis fermentation liquid, 9 parts of potassium nitrate, 19 parts of dipotassium hydrogen phosphate, 4.5 parts of honey, 75 parts of oligofructose, 45 parts of pectin, 75 parts of whey protein, 85 parts of glycerol, 45 parts of lecithin, and 625 parts of clean water.

[0039] The number of viable bacteria in the Lactobacillus plantarum fermentation liquid is 4.5 billion / gram, the number of viable bacteria in the Bacillus subtilis fermentation liquid is 2.5 billion / gram, and the number of viable bacteria in the Candida utilis fermentation liquid is 900 million / gram. The Lactobacillus plantarum and Candida utilis strains are both isolated from healthy egg-laying red wasps, and the Bacillus subtilis is a preserved Bacillus subtilis strain.

[0040] The preparation method of the Lactobacillus plantarum microbial preparation comprises the following steps: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2 using liquid submerged fermentation, the culture medium is mainly glucose, peptone, yeast extract, control the temperature, pH and ventilation, promote bacterial proliferation, obtain Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor according to weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 120 r / min speed for 8 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

[0041] Example 8

[0042] A Lactobacillus plantarum microbial preparation comprises the following raw materials in the following weight ratios: 85 parts of Lactobacillus plantarum fermentation liquid, 85 parts of Bacillus subtilis fermentation liquid, 105 parts of Candida utilis fermentation liquid, 11 parts of potassium nitrate, 21 parts of dipotassium hydrogen phosphate, 5.5 parts of honey, 85 parts of oligofructose, 55 parts of pectin, 85 parts of whey protein, 95 parts of glycerol, 55 parts of lecithin, and 675 parts of clean water.

[0043] The number of viable bacteria in the Lactobacillus plantarum fermentation liquid is 4.5 billion / gram, the number of viable bacteria in the Bacillus subtilis fermentation liquid is 2.5 billion / gram, and the number of viable bacteria in the Candida utilis fermentation liquid is 900 million / gram. The Lactobacillus plantarum and Candida utilis strains are both isolated from healthy egg-laying red wasps, and the Bacillus subtilis is a preserved Bacillus subtilis strain.

[0044] The preparation method of the Lactobacillus plantarum microbial preparation comprises the following steps: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2 using liquid submerged fermentation, the culture medium is mainly glucose, peptone, yeast extract, control the temperature, pH and ventilation, promote bacterial proliferation, obtain Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor according to weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 120 r / min speed for 8 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

[0045] Experimental example: Effect of Lactobacillus plantarum microbial preparation feed on the growth of beef cattle 1. Test methods Forty beef cattle weighing (500±20) kg were randomly divided into two treatment groups, the experimental group and the control group.

[0046] Experimental grouping method: The site was disinfected and the beef cattle were dewormed before the experiment; During the feeding process, the feed was fed in the form of a complete mixed diet, once in the morning and once in the evening, with free access to water; The pre-feeding period was 7 days, and the experimental group was fed 80g / day of the probiotic preparation per cow, while the control group did not add any additives. The main trial period was 55 days, and the feeding method and the use of additives were the same as those in the pre-feeding period; At the beginning and end of the experiment, the fasting body weight of the experimental animals was measured, and at the end of the experiment, venous blood was collected for testing of relevant blood indicators.

[0047] Measurement indicators 1) Determination of production performance indicators The amount of feed and the remaining amount of the diet were accurately weighed at each feeding, and the fasting body weight was measured at the beginning and end of the experiment to calculate the dry matter intake (DMI), average daily gain (ADG) and feed-to-gain ratio (F / G). The results are shown in Table 1 below: Table 1 Effects of Lactobacillus plantarum microbial preparations on the production performance of beef cattle As shown in Table 1, the dry matter intake of the experimental group was 2.74% higher than that of the control group (p < 0.05), indicating that the probiotics promoted the fermentation of rumen and played a certain regulatory role, thereby improving its feed intake level; the average daily weight gain was 27.51% higher than that of the control group (p < 0.05); the feed-to-weight ratio of the experimental group was reduced by 19.38%, which did not reach a significant level, but there was a significant improvement trend.

[0048] 2) Determination of nutrient apparent digestibility The dry matter, crude protein, neutral detergent fiber, and acid fiber content in feed and fecal samples were determined according to the methods in "Feed Analysis and Feed Quality Testing Technology." The apparent digestibility of nutrients was determined using the acid-insoluble ash method, and the calculation formula is as follows: Apparent digestibility of a nutrient (%) = (1-bc / ad) × 100% Where: a is the content of a certain nutrient in the feed; b is the content of a certain nutrient in the feces; c is the content of AIA in the feed; d is the content of AIA in the feces.

[0049] The measurement results are shown in Table 2 below: Table 2 Effects of probiotics on the apparent digestibility of nutrients in beef cattle (%) As shown in Table 2, the digestibility of dry matter in the experimental group did not increase significantly (p>0.05); however, the digestibility of crude protein in the experimental group was 1.59% higher than that in the control group (p<0.05); although the digestibility of neutral detergent fiber and acid detergent fiber was higher than that in the control group, neither reached a significant level (p>0.05).

[0050] 3) Determination of blood antioxidant index At the end of the experimental period, blood was collected from the jugular vein on an empty stomach and centrifuged at 3000 rpm for 10 minutes. The serum was stored at -20°C until further use. Serum levels of superoxide dismutase (SOD), catalase (CAT), malondialdehyde (MDA), and total antioxidant capacity (T-AOC) were determined using kits. The results are shown in Table 3 below: Table 3 Effects of probiotics on antioxidant indexes in beef cattle serum As shown in Table 3, the malondialdehyde content in the blood of the experimental group was 29.70% lower than that of the control group (p < 0.01); the superoxide dismutase activity was 10.03% higher than that of the control group (p < 0.01); and the catalase activity was 16.93% lower than that of the control group (p < 0.01). Although the total antioxidant capacity of the experimental group showed a trend of improvement compared with the control group, no significant difference was found (p > 0.05).

[0051] 4) Determination of digestive enzyme activity in rumen fluid At the end of the trial period, rumen fluid was collected and the activities of rumen fluid digestive enzymes (protease, lipase, amylase, xylanase, pectinase, and filter paper enzyme) were determined using the 3-dinitrosalicylic acid colorimetric method and the kit method. The results are shown in Table 4 below: Table 4 Effects of probiotics on rumen digestive enzyme activities in beef cattle As can be seen from Table 4, the protease activity in the rumen fluid of the experimental group was 7.79% higher than that of the control group (p < 0.01); the amylase activity was 10.23% higher than that of the control group (p < 0.01); however, the lipase activity in the rumen fluid of the two groups did not change significantly (p > 0.05); the pectinase activity in the rumen fluid of the experimental group was 12.80% higher than that of the control group (p < 0.01); the xylanase and filter paper enzyme activities were not significantly different from those of the control group (p > 0.05).

[0052] The Lactobacillus plantarum microbial preparation prepared by the present invention is simple to prepare, and its combination with other microorganisms creates a synergistic effect. It promotes dry matter intake and weight gain in beef cattle, and can improve feed-to-weight ratio to a certain extent. Furthermore, the microecological preparation can increase antioxidant levels, enhance the activity of some rumen digestive enzymes, and improve the digestion and absorption of nutrients.

[0053] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A Lactobacillus plantarum microbial preparation, characterized in that The raw material components of the microbial preparation include lactobacillus plantarum fermentation liquid, bacillus subtilis fermentation liquid, candida utilis fermentation liquid, potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin, and clean water.

2. The plant lactobacillus microbial preparation according to claim 1, wherein The weight ratio of the raw materials of the microbial preparation is: 70-90 parts of Lactobacillus plantarum fermentation liquid, 70-90 parts of Bacillus subtilis fermentation liquid, 90-110 parts of Candida utilis fermentation liquid, 8-12 parts of potassium nitrate, 18-22 parts of dipotassium hydrogen phosphate, 4-6 parts of honey, 70-90 parts of oligofructose, 40-60 parts of pectin, 70-90 parts of whey protein, 80-100 parts of glycerol, 40-60 parts of lecithin, and 600-700 parts of clean water.

3. The plant lactobacillus microbial preparation according to claim 1, wherein The weight ratio of the raw materials of the microbial preparation is preferably: 80 parts of Lactobacillus plantarum fermentation broth, 80 parts of Bacillus subtilis fermentation broth, 100 parts of Candida utilis fermentation broth, 10 parts of potassium nitrate, 20 parts of dipotassium hydrogen phosphate, 5 parts of honey, 80 parts of fructooligosaccharides, 50 parts of pectin, 80 parts of whey protein, 90 parts of glycerol, 50 parts of lecithin, and 650 parts of water.

4. The plant lactobacillus microbial preparation according to claim 1, wherein The Lactobacillus plantarum fermentation liquid is prepared from Lactobacillus plantarum, and the Bacillus subtilis fermentation liquid is prepared from Bacillus subtilis.

5. The plant lactobacillus microbial preparation according to claim 1, wherein The Candida utilis fermentation broth is prepared from Candida utilis.

6. A method for preparing a Lactobacillus plantarum microbial preparation, characterized in that: The following steps are involved: S1. Isolate highly active strains from natural samples and screen for excellent Lactobacillus plantarum, Bacillus subtilis, and Candida utilis strains that are acid-resistant, bile-resistant, and have strong adhesion. S2. Using liquid submerged fermentation, the culture medium is mainly composed of glucose, peptone, and yeast extract. The temperature, pH value, and ventilation volume are controlled to promote bacterial proliferation, and fermentation broths of Lactobacillus plantarum, Bacillus subtilis, and Candida utilis are obtained; S3. Potassium nitrate, dipotassium hydrogen phosphate, honey, oligofructose, pectin, whey protein, glycerol, lecithin were placed in a stainless steel reactor in a weight ratio, and the corresponding amount of water was added, heated to 80 ° C, stirred at 80 to 150 r / min speed for 5 to 10 min, and mixed evenly; S4. The mixture heated in step S3 is cooled to 35°C, and then Lactobacillus plantarum fermentation broth, Bacillus subtilis fermentation broth and Candida utilis fermentation broth are added, and stirred to obtain a finished microbial preparation.

7. The method for preparing a Lactobacillus plantarum microbial preparation according to claim 6, wherein The method for obtaining the plant lactobacillus fermentation liquid is: 1) Inoculate freeze-dried Lactobacillus plantarum powder into MRS medium and culture at 37°C for 24 hours; 2) Then, take 1% of the inoculum and inoculate it into the fermentation medium. Ferment and culture at 37°C for 16-18 hours. 3) During the entire fermentation period, glucose solution is intermittently added to the fermentation system to maintain the glucose concentration in the fermentation system at 5-15 g / L to obtain Lactobacillus plantarum fermentation broth; The method for obtaining the Bacillus subtilis fermentation broth is: 1) Streak Bacillus subtilis onto LB solid medium and incubate at 37°C for 48 hours. 2) Then, use an inoculation loop to pick up the bacteria and inoculate them into liquid LB medium. Incubate at 180 rpm and 37°C for 24 hours. 3) inoculating a corn flour and soybean meal culture medium with a 1% inoculum size for fermentation at the following conditions: temperature 37°C, tank pressure 0.1 MPa, ventilation 0.8-1 vvm, rotation speed 180 rpm, and culture time 2-3 days to obtain a Bacillus subtilis fermentation liquid; The method for obtaining the Candida utilis fermentation broth is as follows: 1) Inoculate freeze-dried Lactobacillus plantarum powder into MRS medium and culture at 37°C for 24 hours; 2) Then, 1% of the inoculum amount was inoculated into the fermentation medium, and fermented at 37°C for 20 hours to obtain the Candida utilis fermentation broth.

8. Use of the Lactobacillus plantarum microbial preparation according to any one of claims 1 to 5 and the Lactobacillus plantarum microbial preparation prepared according to claim 6 in promoting the growth of beef cattle and improving apparent nutrient digestibility, blood antioxidant level and rumen protease activity.

9. Use of the plant lactobacillus microbial preparation according to any one of claims 1 to 5 and the plant lactobacillus microbial preparation prepared according to claim 6 in preventing and controlling plant diseases and insect pests in greenhouse vegetables.

10. A method for using a Lactobacillus plantarum microbial preparation, characterized in that: The Lactobacillus plantarum microbial preparation is diluted and used for root irrigation or sprayed on the leaves of vegetables during the growth period.

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