Lipopeptide feed for promoting growth of weaned piglets and reducing diarrhea and preparation method thereof

Abstract

According to the special feed for the weaned piglets, 60 g of lipopeptide is added into 1 t of feed, so that the growth speed of the weaned piglets can be remarkably increased, the piglets under LPS challenge stimulation are remarkably protected, the effects of protecting liver and resisting inflammation and remarkably promoting growth are achieved, the occurrence rate of diarrhea of the weaned piglets is reduced, and the weaned piglets are prevented from suffering from diarrhea. The intestinal health of the weaned piglets can be remarkably improved, and the breeding efficiency is improved. The invention further provides a preparation method of the feed, the preparation process of the lipopeptide is improved by optimizing a fermentation culture medium and fermentation parameters, adopting a physical defoaming device and the like, the yield of the lipopeptide is effectively increased, and the fermentation yield reaches 1.69 g / L. The method is simple in preparation, the production cost of the feed is greatly reduced, and the method has industrial economy and is worthy of popularization and application. And large-scale preparation and industrial application of the feed are facilitated.

Description

Technical Field

[0001] The invention belongs to the technical field of feed, and in particular relates to a lipopeptide-containing feed capable of promoting growth of weaned piglets and reducing diarrhea, and a preparation method thereof. Background Art

[0002] The long-term addition of antibiotics to feed has led to a series of problems such as bacterial resistance, antibiotic residues in animal food, and environmental pollution, which seriously threaten food safety and public health. To meet this challenge, the Ministry of Agriculture and Rural Affairs has completely banned the addition of antibiotics to feed since 2020, marking the official start of the era of antibiotic-free feeding in my country. However, after the implementation of the ban on antibiotics, pig production faces problems such as decreased production performance, increased diarrhea rates and mortality rates. How to develop green and safe alternative products with equivalent effects to antibiotics has become a key issue that the current pig farming industry needs to solve urgently. Its significance lies not only in alleviating the negative impact of the ban on antibiotics in feed, but also in improving the efficiency of pig farming, promoting the high-quality development of the pig farming industry, and effectively guaranteeing the supply of residents' "pork plates".

[0003] Lipopeptides are a class of low molecular weight biosurfactants produced by microorganisms. They have a unique amphiphilic molecular structure. Their molecules are composed of polar hydrophilic peptide chains formed by the combination of amino acids and non-polar hydrophobic groups composed of fatty acid hydrocarbon chains. This structure gives lipopeptides a variety of biological activities, such as antibacterial, anti-inflammatory and environmental remediation. Compared with traditional antibiotics, lipopeptides have outstanding advantages such as wide sources, broad antibacterial spectrum, low resistance and no residue. However, the complex biosynthetic pathways and metabolic networks of lipopeptides and the coexistence of multiple homologues restrict the microbial development of lipopeptides. In recent years, through the optimization of fermentation conditions, genetic engineering, metabolic engineering methods and biosynthetic strategies have been used to increase the yield of lipopeptide biosynthesis. Although significant progress has been made in the application and production strategies of antimicrobial lipopeptides, they are still not industrially produced. Therefore, further research on the application and yield improvement strategies of antimicrobial lipopeptides is of great significance for improving production efficiency and solving practical application problems. Summary of the invention

[0004] In view of the deficiencies of the above-mentioned prior art, the present invention aims to provide a suckling pig feed that improves the growth performance of weaned piglets, regulates the microbial structure of piglet feces, and reduces the diarrhea rate. Through the method of the present invention, the abundance of beneficial bacteria in the intestinal tract of piglets can be increased, the intestinal health can be improved, and the diarrhea rate of piglets can be reduced.

[0005] The purpose of the first aspect of the present invention is to provide a special feed for weaned piglets, including 56.174% corn, 8.000% puffed corn, 17.500% soybean meal (46%), 4.000% puffed soybean powder, 5.500% fermented soybean meal, 2.000% yeast culture, 1.000% glucose, 1.000% fish meal, 1.000% high-fat oil powder, 0.800% calcium formate, 0.600% calcium hydrogen phosphate, 0.350% salt, 0.800% acidulant, 0.400% lysine, 0.100% choline chloride, 0.100% DL-methionine, 0.080% threonine, 0.070% complex enzyme, 0.520% premix, and 0.006% lipopeptide, calculated in mass percentage.

[0006] Furthermore, the nutritional level of the feed is 3.10 MJ / kg of metabolizable energy, and the contents of the main nutrients are: crude protein 18.60%, calcium 0.53%, total phosphorus 0.48%, available phosphorus 0.30%, lysine 1.25%, methionine 0.40%, methionine + cystine 0.69%, threonine 0.76%, and tryptophan 0.20%, calculated as mass percentage.

[0007] Furthermore, the premix in the above feed provides, per kilogram of feed, the following: Cu 5.12 mg, I 0.15 mg, Fe 87.59 mg, Mn 3.69 mg, Se 0.30 mg, Zn 84.43 mg, VA 9 000 IU, VD 3 000 IU, VE 24 IU, VK 3 mg, thiamine 3 mg, riboflavin 7.5 mg, pantothenic acid 15 mg, niacin 30 mg, pyridoxine 3.60 mg, biotin 0.15 mg, folic acid 1.50 mg, and VB 12 0.036 mg.

[0008] Furthermore, the structural formula of the above lipopeptide is

[0009] Furthermore, the above-mentioned lipopeptide is produced by the Paenibacillus elgii Z121 strain, the preservation number of the strain is CGMCC No.19830, the preservation date is May 15, 2020, the preservation classification name is Paenibacillus elgii, and the preservation unit is the General Microbiology Center of the China Microbiological Culture Collection Administration.

[0010] The second aspect of the present invention is to provide a method for preparing the above feed, comprising the following steps: Step 1, strain activation: the seed culture of Paenibacillus aegypti Z121 (CGMCC No. 19830) was inoculated with 1% of the inoculum and cultured in LB medium. The culture conditions were: 37 °C, initial pH: 7.0, rotation speed 200 rpm, and culture time 12-16 h. Step 2, strain fermentation: the strain was fermented in a fermentation medium at an inoculation rate of 3%, and the fermentation conditions were: temperature 35°C, initial pH 6.5, rotation speed 180 rpm, and fermentation period 26 h; Step 3, first, the fermentation liquid in the fermenter is transported to a butterfly centrifuge through a closed pipe and centrifuged at 8000 rpm to remove bacteria and impurities and collect lipopeptides; secondly, nanofiltration concentration is performed at 40°C, pH 3.0, and inlet and outlet membrane pressures of 2.0 / 1.2 MPa; then, the fermentation concentrate is spray-dried at an inlet and outlet temperature of 170 / 80°C to obtain a lipopeptide solid powder; Step 4, feed preparation: according to the above-mentioned component ratio, puffed corn, puffed soybean powder, yeast culture, glucose, fish meal, high-fat oil powder, calcium formate, calcium hydrogen phosphate, salt, acidulant, lysine, choline chloride, DL-methionine, threonine, compound enzyme, and premix are weighed and premixed for 120 s. After cooling, mixture 1 is obtained; corn, soybean meal (46%), and fermented soybean meal are weighed, high-speed hammer crushing is performed, the screen is 1.0 mm, and they are put into a mixer, the temperature is adjusted to 75-85 ° C, mixed for 180 s, cooled to room temperature, and then the lipopeptide solid powder obtained in step 3 is added to obtain mixture 2; mixture 1 and mixture 2 are mixed together for 180 s, and special feed for weaned piglets is obtained after cooling.

[0011] Furthermore, the fermentation medium contains 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, 0.05% magnesium sulfate, and 1 g / L aspartic acid or arginine, calculated by mass percentage.

[0012] Furthermore, in the fermentation culture stage of step 2, a physical negative pressure defoaming device is used, which starts to operate 6-8 hours after fermentation and continues until the end of fermentation. Beneficial Effects

[0013] The present invention provides a feed with added lipopeptide. Adding 60 g of lipopeptide to 1 t of feed can increase the growth rate of weaned piglets, increase the average daily weight gain (ADG) in the third week by 10.63%, and reduce the feed conversion efficiency (F / G) by 8.56%. Under the stimulation of LPS attack, the immunosuppressive reaction caused by LPS can be significantly alleviated, and the diversity and uniformity of intestinal flora can be maintained, the secretion of short-chain fatty acids (such as butyrate) can be increased, and the intestinal barrier function can be repaired. The feed has a significant protective effect on piglets, has liver protection and anti-inflammatory effects and significant growth-promoting effects, reduces the incidence of diarrhea in weaned piglets, can significantly improve the intestinal health and production performance of weaned piglets, and has a good market application prospect.

[0014] The present invention also provides a method for preparing lipopeptide feed. The preparation process of lipopeptide is improved by optimizing fermentation medium and fermentation parameters, adopting a physical defoaming device and the like, thereby greatly increasing the yield of lipopeptide, so that the fermentation yield reaches 1.69 g / L, reducing the production cost, and facilitating the large-scale preparation of feed and its application in the pig farming industry. Instruction Manual

[0015] Figure 1 Species accumulation curve (left) and Venn diagram (right) (CON: basal diet group without lipopeptide addition; H: 60 g / t lipopeptide addition group) Figure 2 Effects of dietary peptide addition on the diversity of intestinal microbial flora in piglets (the higher the sobs, chao and ace indexes, the higher the species richness, the higher the shannon index, the higher the species uniformity, and the lower the simpson index, the higher the species uniformity) (CON: basic diet group without lipopeptide addition; H: 60 g / t lipopeptide addition group) Figure 3 Effects of dietary peptide supplementation on the overall structure of the intestinal flora of piglets (the significance and explanatory power obtained by PERMANOVA analysis are shown in the figure p and R2 represent) (CON: basal diet group without lipopeptide addition; H: 60 g / t lipopeptide addition group) Figure 4 Analysis of species differences in the intestinal flora of piglets when lipopeptides were added to the diet (CON: basic diet group without lipopeptides; H: 60 g / t lipopeptide addition group) Figure 5 Effects of lipopolysaccharide injection on the diversity of intestinal microbial flora in piglets fed diets without lipopeptide (Con and LPS groups) and with 60 g / t lipopeptide (H and HLPS groups) (**: P <0.01, ns: P >0.05) Figure 6Effects of lipopolysaccharide injection on the overall structure of intestinal flora in piglets fed diets without lipopeptide (con and lps groups) and with 60 g / t lipopeptide (h and hlps groups) (the significance and explanation of the PERMANOVA analysis are shown in the figure) p and R2 represents) Figure 7 LPS injection significantly changed the abundance of key species in the intestinal microbiota of piglets fed diets without lipopeptides (Con and LPS groups) and with lipopeptides (H and HLPS groups) Figure 8 Effects of different carbon sources on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig. 9 Effects of different nitrogen sources on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.10 Effects of amino acids on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.11 Effects of inorganic salts on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.12 Effects of trace elements on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.13 Effect of fermentation pH on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P<0.005;***: P <0.001; ****: P <0.0001) Fig.14 Effect of fermentation temperature on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.15 Effect of inoculum size on lipopeptide expression (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.16 Effect of fermentation speed on lipopeptide expression (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.17 Effect of culture cycle on the expression of lipopeptides (ns: no difference; *: P <0.05; **: P <0.005;***: P <0.001; ****: P <0.0001) Fig.18 The addition of different concentrations of defoamers had no significant effect on the expression of lipopeptides Fig.19 Effect of physical defoaming on the expression of lipopeptides in strain Z121 (defoaming device: air compressor 1, air amplifier 2, ventilation pipe 3, defoaming pipe 4, vacuum shell 5, upper cover 501, lower cover 502, support rib 1 503, support rib 2 504, connecting sleeve 505, defoaming hole 506, groove 507, suction hole 6, fixing frame 7, U-shaped tube 8, solenoid valve 9, controller 10, foam sensor 11, peristaltic pump 12, defoaming agent storage 13, defoaming agent input pipe 14, fermentation tank 15, agitator 16, top cover 17, stirring head 18) Fig. 20 Effects of different centrifuge speeds on the recovery rate of lipopeptide fermentation broth Fig.21 Effect of different pH values ​​on the recovery rate of lipopeptide fermentation broth during nanofiltration Fig. 22 Effect of different temperatures on the recovery rate of lipopeptide fermentation broth during nanofiltration DETAILED DESCRIPTION

[0016] In order to make the purpose, technical scheme and advantages of the embodiments of the present invention clearer, the technical scheme in the embodiments of the present invention will be described clearly and completely below. If the specific conditions are not specified in the embodiments, they are carried out according to conventional conditions or conditions recommended by the manufacturer. If the manufacturer of the reagents or instruments used is not specified, they are all conventional products that can be purchased commercially.

[0017] The features and performance of the present invention are further described in detail below in conjunction with the embodiments.

[0018] Example 1 A special feed for weaned piglets, the specific composition and nutritional level of which are shown in Table 1.

[0019]

[0020] *Premix provides per kg feed: Cu 5.12 mg, I 0.15 mg, Fe 87.59 mg, Mn 3.69 mg, Se 0.30 mg, Zn 84.43 mg, VA 9 000 IU, VD 3 000 IU, VE 24 IU, VK 3 mg, thiamine 3 mg, riboflavin 7.5 mg, pantothenic acid 15 mg, niacin 30 mg, pyridoxine 3.60 mg, biotin 0.15 mg, folic acid 1.50 mg, VB 120.036 mg.

[0021] Example 2 A lipopeptide feed specifically for weaned piglets can improve the growth performance of weaned piglets

[0022] Experimental design and animal husbandry 1 Experimental design and animal husbandry The animal feeding experiment was divided into two stages. In the first stage, a single-factor randomized block design was used. 64 LY weaned piglets aged 21 days and weighing about 6.0 kg were selected. The body weight was weighed after one week of pre-feeding. There was no significant difference in the body weight among the treatment groups. They were randomly divided into two groups: control group (CON) and lipopeptide 60 g / t addition group (H) (Table 2). Each group had 8 replicate pens, and each pen had 4 replicates. The experiment lasted for 21 days. The second stage adopted a double-factor randomized block design. After the first stage of the experiment, 2 pigs with a body weight close to the average body weight of the pen were selected from each of the 8 pens in the CON group and the H group. One pig was injected with lipopolysaccharide (100 μg / kg) and the other was injected with an equal volume of sterile saline. The four treatment groups (con, lps, h and hlps) are shown in Table 3. The feed intake data were repeated per pen (n=4), and the rest of the data were repeated per head (n=32). During the entire experiment, the piglets were fed and drank water freely, and were fed and managed according to the daily management of the pig farm.

[0023]

[0024]

[0025] 2. Determination indicators and methods

[0026] 2.1 Growth performance All experimental pigs were weighed and recorded on an empty stomach (no feeding) at night on the 0th, 7th, 21st and 28th days of the experiment, and at 8:00 in the morning on the 1st, 8th, 15th, 22nd and 29th days. At the same time, feed was fed at 8:00, 12:00, 16:00 and 20:00 every day, and the feed weight and loss during the experiment were accurately recorded. The average daily weight gain (ADG), average daily feed intake (ADFI) and feed-to-meat ratio (F / G) were calculated using the following formula: ADG (g / d) = total weight gain / test days ADFI (g / d) = feed consumption / test days F / G (g / g) = ADFI / ADG 2.2 Diarrhea rate and diarrhea index During the experiment, the diarrhea and fecal index of all experimental pigs were recorded at 09:00 (after feeding at 8:00) and 17:00 (after feeding at 16:00) every day. One diarrhea episode was recorded as one pig diarrhea in one day. The fecal scoring standard is shown in Table 4.

[0027] Diarrhea rate (%) = 100 × total number of diarrhea episodes / (total number of test pigs × number of test days) Diarrhea index = 100 × sum of diarrhea scores / (total number of test pigs × number of test days)

[0028] 2.3 Blood indicators On the morning of the 22nd day of the experiment, two pigs with body weights close to the mean body weight of the circle were selected from each circle of the CON group and the lipopeptide group, and one pig was injected with LPS (100 μg / kg), and the other pig was injected with an equal volume of sterile saline. 10 mL of blood was collected from the jugular vein 4 hours after LPS stimulation and placed in a procoagulant blood collection tube. 10 mL of blood was also collected from a pig with body weights close to the mean body weight of the circle in the antibiotic group and the other two lipopeptide groups, and placed in a procoagulant blood collection tube. After standing at room temperature for 30 min, the blood was centrifuged at 3000 rpm for 10 min, and the supernatant was divided into 0.5 mL centrifuge tubes and stored at -20 ℃ or -80 ℃ for later use.

[0029] The levels of total protein (TP), albumin (ALB), globulin (GLB), urea nitrogen (BUN), triglycerides (TG), total cholesterol (TC), total bile acid (TBA), total bilirubin (TBIL) in serum and the activities of aspartate aminotransferase (AST), alanine aminotransferase (ALT), and glutamyl transpeptidase (GGT) were determined using an automatic biochemical analyzer.

[0030] The levels of serum IgG, IgM, and IgA, and the levels of serum cytokines IL-6 and TNF-α were determined by ELISA. The kits used were from Nanjing Jiancheng Bioengineering Research Institute Co., Ltd., and the detection methods were operated and calculated in accordance with the instructions.

[0031] 2.4 Determination and analysis of fecal microbial 16S rRNA On the morning of the 20th and 27th days of the experiment, fresh feces of pigs were collected in pens. The feces were broken open with a sterile spoon, and the central part that did not touch the ground was taken and placed in a sterile sealed bag. Half of the bag was collected and mixed, and part was divided into EP tubes and stored at -80 °C for fecal colony sequencing. The samples were sent to a sequencing company for 16S rRNA amplicon sequencing and diversity and abundance analysis. GC-MS was used to determine the contents of acetic acid, propionic acid, butyric acid, valeric acid, and isovaleric acid in feces and cecal contents.

[0032] 2.5 Data Statistical Analysis Excel was used for data sorting, and growth performance was statistically analyzed in replicates. Blood, feces, and intestinal flora indicators were statistically analyzed in heads. The chi-square test was used to compare the differences in diarrhea rate, and the growth performance, blood, and feces indicators were statistically analyzed using T test and Two Way ANOVA (LPS challenge test). If the difference was significant, Tukey's multiple comparisons were performed. P The difference was significant when the value was less than 0.05, and the linear quadratic effect was analyzed by orthogonal polynomial analysis.

[0033] 3 Experimental results and analysis 3.1 Effects of lipopeptides on growth performance of weaned piglets As shown in Table 5, there was no significant difference in the growth performance of piglets in the H group compared with the CON group ( P >0.05), but the ADG in the third week increased by 10.63% and the F / G decreased by 8.56%.

[0034]

[0035] CON: basal diet group; H: basal diet + 60 g / t pure lipopeptide group After the piglets were fed different diets for 21 days, 2 piglets were selected from each of the 8 pens in the CON and H groups, one of which was injected with saline and the other with LPS. All piglets were fed the basal diet for 1 week, and the feed intake and body weight were recorded. The results are shown in Table 6. The addition of lipopeptides significantly increased the final weight and ADG of piglets and reduced F / G ( P< 0.01), under LPS challenge, feeding the feed supplemented with 60g / t lipopeptide had a significant protective effect on piglets.

[0036]

[0037] CON: basal diet group; H: basal diet + 60 g / t pure lipopeptide group 3.2 Effect of lipopeptides on diarrhea rate in piglets3.2 Effect of lipopeptides on diarrhea rate in piglets As shown in Table 7, the pigs in this experiment were in good health, and there were no significant differences in diarrhea index and diarrhea rate among the treatment groups at different periods ( P> 0.01). However, in the whole period, the diarrhea rate in the H group was 19.02% lower than that in the CON group.

[0038] As shown in Table 7, the pigs in this experiment were in good health, and there were no significant differences in diarrhea index and diarrhea rate among the treatment groups at different periods ( P> 0.01). However, in the whole period, the diarrhea rate in the H group was 19.02% lower than that in the CON group.

[0039]

[0040] CON: basal diet group; H: basal diet + 60 g / t pure lipopeptide group. The diarrhea rate and diarrhea index were compared using the chi-square test.

[0041] 3.3 Effects of lipopeptides on blood routine in piglets challenged with LPS As shown in Table 8, LPS challenge significantly reduced the number of white blood cells, lymphocytes, monocytes, and eosinophils (P <0.05), significantly increased the percentage of basophils and the number of neutrophils ( P <0.05). Lipopeptide (H) significantly reduced the number of lymphocytes ( P <0.05). H and LPS had significant interactions on lymphocyte percentage, lymphocyte number, monocyte number, and hematocrit ( P <0.05).

[0042]

[0043] con: pigs in CON group were injected with normal saline; lps: pigs in CON group were injected with 100 μg / kg lipopolysaccharide; h: pigs in H group were injected with normal saline; hlps: pigs in H group were injected with 100 μg / kg lipopolysaccharide.

[0044] 3.4 Effects of lipopeptides on blood biochemistry in LPS-challenged piglets As shown in Table 9, LPS challenge significantly increased serum levels of aspartate aminotransferase, alkaline phosphatase, aspartate / glutamic acid, urea, and triglycerides ( P <0.05), indicating that LPS challenge can significantly damage liver, heart, and gallbladder-related functions. LPS and lipopeptide have significant interactions on alkaline phosphatase, triglycerides, and glucose ( P <0.05). Under LPS challenge, alkaline phosphatase and triglyceride increased significantly, and glucose decreased significantly, while lipopeptides could significantly alleviate this phenomenon and protect liver function. The addition of lipopeptides significantly reduced the white-globulin ratio, indicating that lipopeptides can enhance the immune ability of piglets under LPS challenge.

[0045]

[0046] con: pigs in CON group were injected with normal saline; lps: pigs in CON group were injected with 100 μg / kg lipopolysaccharide; h: pigs in H group were injected with normal saline; hlps: pigs in H group were injected with 100 μg / kg lipopolysaccharide.

[0047] 3.5 Effects of lipopeptides on serum immune parameters in piglets challenged with LPS As shown in Table 10, LPS challenge treatment significantly increased the level of lipopolysaccharide endotoxin in piglet serum ( P <0.05); LPS and lipopeptide both significantly increased the IgM content in piglet serum ( P <0.01); LPS significantly reduced the IgG content in piglet serum ( P <0.05), lipopeptide significantly increased the IgG content in piglet serum ( P <0.05); Lipopeptide significantly increased the levels of IL6 and IgA in piglet serum ( P<0.01); LPS and lipopeptide had significant interactive effects on the IgA content in piglet serum ( P <0.05).

[0048]

[0049] con: pigs in CON group were injected with normal saline; lps: pigs in CON group were injected with 100 μg / kg lipopolysaccharide; h: pigs in H group were injected with normal saline; hlps: pigs in H group were injected with 100 μg / kg lipopolysaccharide.

[0050] 3.6 Effects of lipopeptides on volatile fatty acids in piglet feces As shown in Table 11, there were no significant differences in acetic acid, propionic acid, valeric acid, and isovaleric acid in the feces of the H group compared with the CON group, and the butyric acid content was lower than that of the CON group ( P <0.05).

[0051]

[0052] CON: basal diet group; H: basal diet + 60 g / t pure lipopeptide group The effect of 60 g / t lipopeptide on volatile fatty acids in piglet feces under LPS challenge model is shown in Table 12. LPS significantly increased the content of propionic acid in piglet feces ( P <0.01), significantly reduced the valeric acid content in piglet feces ( P <0.01, while lipopeptides significantly increased the contents of fecal acetic acid and butyric acid under LPS challenge ( P <0.01). LPS and lipopeptides had highly significant interactions on acetate, butyrate and valeric acid ( P <0.01); there was a significant interaction with isovaleric acid ( P <0.05). LPS can increase propionic acid and reduce valeric acid in feces, while lipopeptides can increase acetic acid and butyric acid in feces after LPS treatment. Butyric acid has the effect of strengthening the intestinal barrier, indicating that feeding feed supplemented with 60 g / t lipopeptides can effectively maintain intestinal health and slow down the intestinal function damage caused by LPS.

[0053]

[0054] con: CON pigs were injected with normal saline; lps: CON pigs were injected with 100 μg / kg lipopolysaccharide; h: H group pigs were injected with normal saline; hlps: H group pigs were injected with 100 μg / kg lipopolysaccharide.

[0055] 3.7 Effects of lipopeptides on the intestinal flora of piglets 3.7.1 Effects of lipopeptides on the diversity of intestinal flora in piglets Species accumulation curve ( Figure 1 Left), as the sample size increases, the detected OTUs curve tends to be flat, proving that the sample size of this experiment can fully analyze the community richness. Venn diagram ( Figure 1 (right), the control group and the experimental group each contained 843 OTUs, of which 54 and 95 were unique to each group.

[0056] The three diversity indices of sobs, chao and ace were used to measure the species richness of the intestinal flora of piglets, and the two diversity indices of shannon and simpson were used to measure the species uniformity of the intestinal flora of piglets. The results showed that although there was no statistical difference among the groups ( Figure 2 , P >0.05), but the addition of 60g / t lipopeptide in the diet slightly increased the species richness and species uniformity of the intestinal flora of piglets, suggesting that the addition of lipopeptide can affect the diversity of the intestinal flora of piglets.

[0057] 3.7.2 Effects of lipopeptides on the intestinal flora structure of piglets PCoA analysis based on unweighted unifrac distance showed that the addition of 60g / t lipopeptide in the diet significantly changed the overall structure of the intestinal flora of piglets. PERMANOVA analysis showed that the explanations for the changes in the overall structure of the intestinal flora of piglets were 19.55% and 15.75%, respectively ( Figure 3 ).

[0058] The difference in species between each peptide addition group and the control group was analyzed at the species level. The FDR<0.05 was used as the screening threshold. The results showed that there were 11 species with significant changes in abundance when 60 g / t lipopeptide was added, among which the most obvious change was Fusobacteriia and Betaproteobacteria ( P <0.05) ( Figure 4 ). This indicates that 60 g / t lipopeptide in the diet may affect its regulatory pattern on the intestinal flora of piglets.

[0059] 3.7.3 Effect of lipopeptides on the diversity of intestinal flora in piglets challenged with LPS LPS injection significantly reduced the species richness and uniformity of the intestinal flora of piglets fed with a normal diet, but it did not significantly affect the species richness of the intestinal flora of piglets fed with a diet supplemented with 60 g / t lipopeptides. At the same time, the species uniformity of the piglets fed with the 60 g / t lipopeptide diet was significantly improved under LPS stimulation ( Figure 5 The above results indicate that the addition of 60 g / t lipopeptides in the diet will help the intestinal flora of piglets maintain good species richness and uniformity when responding to intestinal damage.

[0060] LPS injection also significantly changed the overall structure of the intestinal flora of piglets fed with a normal diet and a diet supplemented with 60 g / t lipopeptide ( Figure 6 ), indicating that when lipopeptides are not added to the diet, the intestinal flora of piglets is more likely to become disordered when encountering intestinal damage, that is, the addition of lipopeptides can maintain the homeostasis of the intestinal flora structure of piglets to a certain extent.

[0061] The different species in each experimental group compared with the control group were analyzed at the species level, with FDR < 0.05 as the screening threshold. Figure 7 The results show that the relative abundance of the key differential species in each group was significantly changed by LPS injection without the addition of lipopeptides to the diet. Faecalibacterium prausnitzii , its abundance increased by 367% compared with the control group ( Figure 7 ). In published studies, this strain has been shown to be one of the important producers of butyrate in the human intestine, which plays a role in maintaining the integrity of the intestinal lining. Faecalibacterium prausnitzii The significant increase in abundance is most likely a self-repair mechanism adopted by piglets to cope with the intestinal damage caused by lipopolysaccharide injection. Faecalibacterium prausnitzii The abundance was only 63.6%, a decrease of about 300%. On the one hand, this further confirms the conclusion that the addition of lipopeptides can maintain the homeostasis of the intestinal flora structure of piglets to a certain extent. On the other hand, it shows that lipopolysaccharide injection may not cause serious damage to the intestines of the piglets in the treatment group.

[0062] In summary, the addition of 60 g / t lipopeptide to the diet can improve the growth performance of weaned piglets, significantly change the microbial structure of piglet feces, and reduce the diarrhea rate by more than 19%. Lipopolysaccharide challenge has a negative impact on the growth performance, blood routine, blood biochemistry, volatile fatty acids and intestinal microorganisms of piglets, while the addition of 60 g / t lipopeptide to the diet can increase the levels of immunoglobulins and IL6 in serum, enhance the body's immunity, reduce liver damage, maintain blood triglycerides and glucose at normal levels, improve the secretion of volatile fatty acids in feces, and help restore intestinal health.

[0063] Example 3 Optimization of Z121 strain fermentation process and post-treatment process 1. Bacterial strains and culture media The fermentation strain is Paenibacillus aegypti Z121 (the deposit number is CGMCC No. 19830). The seed culture medium is LB culture medium. The fermentation basal culture medium is 1% glucose, 1% yeast powder, and 0.5% sodium chloride.

[0064] 2. Investigate the effect of carbon source on product content in shake flask fermentation The fermentation medium: 1% glucose, 1% yeast powder, 0.5% sodium chloride was used as the control group. Five different carbon sources, glucose, sucrose, lactose, corn flour, and soybean meal, were selected at a dosage of 1% for single-factor shake flask experiment. The fermentation shake flask parameters were as follows: temperature 37 ℃, initial pH 7.0, inoculation size 1%, and rotation speed 200 rpm for 24 h. The product content in the fermentation broth was detected. After repeated verification for 3 times, the optimal carbon source was selected according to the average content, and then different contents (1%, 2%, 3%, and 4%) were selected for the same experimental verification.

[0065] Figure 8 The effects of five carbon sources (glucose, sucrose, lactose, corn sugar, and glycerol) on lipopeptide expression are shown. The lipopeptide expression levels of sucrose and glucose were significantly higher than those of the control group, and the difference in sucrose was more significant ( P <0.005). Figure 8 The right side shows that the sucrose content of 2% has the greatest effect on increasing the expression of lipopeptides, which is 0.55 g / L.

[0066] 3. Effect of nitrogen source on product content in shake flask fermentation The fermentation medium: 2% sucrose, 1% yeast powder, 0.5% sodium chloride was used as the control group. Five different nitrogen sources, peptone, yeast powder, corn syrup powder, soybean meal powder, and biological nutrients were selected at a dosage of 1% for single-factor shake flask experiment. The fermentation shake flask parameters were temperature 37 ℃, initial pH 7.0, inoculation size 1%, and rotation speed 200 rpm for 24 h. The product content in the fermentation broth was detected. After repeated verification for 3 times, the optimal nitrogen source was selected according to the average content, and then the same test verification was carried out with different contents (1%, 2%, 3%, and 4%).

[0067] Fig. 9 The effect of different nitrogen sources (yeast powder, peptone, corn syrup powder, soybean meal powder, and bionutrients) on the expression of lipopeptides in the Z121 strain is shown. The left figure shows that when yeast powder is used as a nitrogen source, the expression of lipopeptides in the Z121 strain is the highest. The right figure shows that when the yeast powder content is 2%, the expression of lipopeptides is the highest, at 0.58 g / L.

[0068] 4. Investigate the effect of amino acids on product content in shake flask fermentation The culture medium (10 g / L ammonium sulfate, 5 g / L potassium dihydrogen phosphate, 2 g / L sucrose, 5 g / L yeast powder + 0.2 g / L magnesium sulfate) was used as the basic culture medium, and different 1 g / L amino acids (L-tyrosine, L-leucine, L-histidine, L-glutamine, L-proline, L-isoleucine, L-aspartic acid, L-threonine, L-arginine, L-lysine, L-methionine, L-cysteine, L-alanine, L-serine, L-valine, L-tryptophan, L-phenylalanine) were added to optimize the amino acids. The culture medium numbers were 1-17.

[0069] Fig.10 The results show that different amino acids have different effects on the expression of active substances, among which aspartic acid (7) and arginine (9) can most significantly increase the expression of active substances.

[0070] 5. Effect of inorganic salts on product content in shake flask fermentation The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% sodium chloride was used as the control group. Four different inorganic salts, dipotassium hydrogen phosphate, potassium dihydrogen phosphate, sodium chloride and potassium chloride, were selected at a dosage of 0.5% for single-factor shake flask test. The fermentation shake flask parameters were temperature 37 ℃, initial pH 7.0, inoculation size 1%, and rotation speed 200 rpm for 24 h. The product content in the fermentation broth was detected. After repeated verification for 3 times, the optimal inorganic salt was selected according to the average content, and then the same test verification was carried out with different contents (0.3%, 0.5%, 0.8%, 1%).

[0071] Fig.11 The effect of different inorganic salts (sodium chloride, dipotassium hydrogen phosphate, potassium dihydrogen phosphate, potassium chloride) on the expression of lipopeptides in the Z121 strain is shown. The left figure shows that the addition of potassium dihydrogen phosphate has the highest expression of lipopeptides in the Z121 strain. The right figure shows that when the content of potassium dihydrogen phosphate is 0.5%, the expression of lipopeptides is the highest, which is 0.64 g / L.

[0072] 6. Effect of trace elements on product content in shake flask fermentation The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate was used as the control group, and 5 different trace elements, magnesium sulfate, ferrous sulfate, zinc chloride, copper sulfate, and calcium chloride, were selected at a dosage of 0.1% for single-factor shake flask experiment. The fermentation shake flask parameters were temperature 37 ℃, initial pH 7.0, inoculation size 1%, and rotation speed 200 rpm. After selecting the optimal trace element, the same experiment was carried out with different contents (0.05%, 0.1%, 0.2%, and 0.3%) for verification.

[0073] Fig.12The effect of different trace elements (magnesium sulfate, zinc chloride, ferrous sulfate, copper sulfate, and calcium chloride) on the expression of lipopeptides in the Z121 strain is shown. The left figure shows that the addition of magnesium sulfate has the highest expression of lipopeptides in the Z121 strain. The right figure shows that the highest expression of lipopeptides is 0.89 g / L when the magnesium sulfate content is 0.05%.

[0074] 7. Effect of initial pH on product content in shake flask fermentation The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, and 0.05% magnesium sulfate were used as fermentation raw materials. On this basis, an initial pH verification test was carried out. The initial pH was adjusted to 6, 6.5, 7 (control group), 7.5, and 8. The culture was carried out at a temperature of 37°C, an inoculation size of 1%, and a rotation speed of 200 rpm for 24 h. The product content in the fermentation broth was detected and the parallel experiment was repeated 3 times.

[0075] Fig.13 The effects of different pH values ​​of 6, 6.5, 7 (control group), 7.5, and 8 on the expression of lipopeptides in the Z121 strain were shown. The results showed that the expression of lipopeptides was the highest at a pH value of 6.5, which was 0.94 g / L.

[0076] 8. Effect of shake flask fermentation temperature on product content The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, and 0.05% magnesium sulfate were used as fermentation raw materials. On this basis, a temperature verification test was carried out. The culture temperature was 30 ℃, 33 ℃, 35 ℃, 37 ℃ (control group), and 39 ℃. The inoculation size was 1%, the optimal initial pH was 6.5, and the rotation speed was 200 rpm. The product content in the fermentation broth was detected and the parallel experiment was repeated 3 times.

[0077] Fig.14 The effect of different temperatures (30 ℃, 33 ℃, 35 ℃, 37 ℃ (control group), 39 ℃) on the expression of lipopeptides in Z121 strain was shown. The results showed that the expression of lipopeptides was the highest at 35 ℃, which was 0.97 g / L.

[0078] 9. Effect of shake flask fermentation inoculum size on product content The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, and 0.05% magnesium sulfate were used as fermentation raw materials. On this basis, an inoculation verification test was carried out. The inoculation amounts were 0.5%, 1% (control group), 2%, 3%, and 4%. The culture was carried out for 24 h at the optimal temperature of 35 °C, initial pH 6.5, and rotation speed of 200 rpm. The product content in the fermentation broth was detected and the parallel experiment was repeated 3 times.

[0079] Fig.15The effect of different inoculation amounts (0.5%, 1% (control group), 2%, 3%, 4%) on the expression of lipopeptides in Z121 strain was shown. The results showed that the expression of lipopeptides was the highest when the inoculation amount was 3%, which was 1.13 g / L.

[0080] 10. Effect of shake flask fermentation speed on product content The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, and 0.05% magnesium sulfate were used as fermentation raw materials. On this basis, a speed verification test was carried out. The speeds were 150 rpm, 180 rpm, 200 rpm (control group), and 220 rpm. The culture was carried out for 24 h at the optimal temperature of 35 °C, initial pH 6.5, and inoculation size of 3%. The product content in the fermentation broth was detected, and the parallel experiment was repeated 3 times.

[0081] Fig.16 The effect of different fermentation speeds (150 rpm, 180 rpm, 200 rpm (control group), 220 rpm) on the expression of lipopeptides in Z121 strain was shown. The results showed that the expression of lipopeptides was the highest at 180 rpm, which was 1.18 g / L.

[0082] 11. Effect of shake flask fermentation culture cycle on product content The fermentation medium: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, and 0.05% magnesium sulfate were used as fermentation raw materials. On this basis, a culture cycle verification test was carried out with cycles of 20 h, 22 h, 24 h (control group), 26 h, 28 h, and 30 h. The culture was carried out under the optimal temperature of 35 °C, initial pH 6.5, inoculation size of 3%, and rotation speed of 180 rpm. The product content in the fermentation broth was detected and the parallel experiment was repeated 3 times.

[0083] Fig.17 The effects of different fermentation culture cycles (20 h, 22 h, 24 h (control group), 26 h, 28 h, and 30 h) on the expression of lipopeptides in the Z121 strain were shown. The results showed that the expression of lipopeptides was the highest at cycle 26, which was 1.24 g / L.

[0084] 12. Investigate the effect of defoamers on product content in shake flask fermentation The culture medium (2 g / L sucrose + 5 g / L yeast extract + 8 g / L ammonium sulfate + 0.2 g / L magnesium sulfate heptahydrate, pH 7.0) was used as the control culture medium. Defoamers of different concentrations (1 / 1000, 1 / 100, and 5 / 100) were added to the basic culture medium formula of the control group, and the effects of defoamers on the growth performance of the strain and the yield of active substances were compared.

[0085] Fig.18The effect of adding different concentrations of defoamers on the expression of lipopeptides in the Z121 strain is shown. It can be seen that the defoamer has no significant effect on the expression of active substances.

[0086] 13. Effect of physical defoaming on product content A physical defoaming device was installed in the experiment, including a vent pipe and a defoaming tube. The defoaming tube was provided with an air suction hole. The defoaming tube was connected to an air amplifier. The air amplifier was connected to an air compressor through the vent pipe. When the air compressor ventilated the air amplifier, a local negative pressure was formed near the defoaming tube based on high-speed air, which played a role of fermentation foam, thereby avoiding the inhibitory effect of chemical defoaming on fermentation and significantly increasing the product expression amount, which could reach up to 1.69 g / L ( Fig.19 ).

[0087] 14. Effect of different centrifuge speeds on product content The fermentation liquid in the fermenter is transported to a butterfly centrifuge through a closed pipe and centrifuged at different centrifugal speeds. The supernatant is collected and the volume and product content are measured, and the recovery rate of the product is calculated.

[0088] from Fig. 20 It can be seen that with the increase of centrifugal separation speed, the recovery rate of the clear liquid after centrifugation gradually increases. When the speed exceeds 8000 rpm, the recovery rate no longer changes, indicating that when the centrifuge reaches a speed of more than 8000 rpm, impurities such as bacteria in the fermentation broth have been basically separated out.

[0089] 15. Effect of different pH values ​​on product content during nanofiltration The pH value of lipopeptide fermentation broth can change the dispersion of solutes in the solution by changing the charge on the surface of the solute, thus affecting the membrane filtration process. Sedimentation can occur under certain pH conditions. The pH value of the fermentation broth was adjusted with 6 mol / L NaOH and HCl, and then the nanofiltration membrane was directly used for filtration. The membrane flux and product recovery rate were investigated within the range of inlet and outlet pressure and temperature.

[0090] from Fig.21 It can be seen that when the pH of the lipopeptide fermentation broth is between 1 and 3, the recovery rate increases with the increase of pH, and when the pH of the lipopeptide fermentation broth is between 3 and 10, the recovery rate decreases with the increase of pH. Therefore, the recovery rate is highest when the pH of the lipopeptide fermentation broth is controlled at 3 during the nanofiltration process.

[0091] 16. Effect of nanofiltration membrane inlet and outlet pressure on product content The upper liquid obtained from the centrifuge was pumped into a liquid storage tank, and the temperature of the lipopeptide fermentation liquid was controlled at 35°C when the membrane flux was stable. The nanofiltration process was performed under different pressures and pressure differences.

[0092] It can be seen from Table 13 that the influence of the inlet and outlet pressure and the pressure difference on the recovery rate is limited, but it has a greater impact on the membrane flux. When the pressure and the pressure difference are very low, the membrane flux decreases, the filtration time is greatly increased, and the filtration efficiency decreases. Therefore, operating condition 1 is selected for subsequent research.

[0093]

[0094] 17. Effect of temperature on lipopeptide fermentation broth during nanofiltration Changes in filtration temperature will lead to changes in the viscosity and diffusion coefficient of the fermentation broth, resulting in different recovery rates of the lipopeptide fermentation broth under the same membrane pressure and inlet and outlet pressure differential conditions. The effect of temperature on the nanofiltration membrane was investigated by controlling different temperatures in a shell-and-tube heat exchanger.

[0095] Depend on Fig. 22 It can be seen that within the allowable operating temperature range (≤40 ℃), the recovery rate increases with increasing temperature. When the temperature exceeds 40 ℃, the recovery rate decreases with increasing temperature. In addition, too high a temperature will also cause the chromaticity of the filtrate to deepen. Therefore, 40 ℃ is selected as the operating temperature of the nanofiltration membrane.

[0096] 18. Effect of spray drying material inlet and outlet temperature on product content After the nanofiltration concentrate is obtained, it is pumped into a spray drying tower through a gear pump, and different spray drying inlet and outlet temperatures are designed to verify their effects on the recovery rate of the lipopeptide fermentation broth.

[0097] It can be seen from Table 5-2 that under 9 different spray drying inlet and outlet temperature conditions, the product recovery rate reaches the highest value when the inlet temperature is 170 ℃ / outlet temperature is 80 ℃.

[0098]

[0099] 19. Data processing and statistical analysis The above experimental data were sorted using Excel and analyzed using One Way ANOVA. If the difference was significant, multiple comparisons between the experimental treatment group and the control group were performed using Tukey. P The difference was significant when the value was less than 0.05.

[0100] Through a series of verification experiments, the optimal fermentation process of strain Z121 was obtained as follows: 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, 0.05% magnesium sulfate, 1 g / L aspartic acid or arginine, culture temperature 35 ℃, initial pH 6.5, inoculation size 3%, shaker speed 180 rpm, fermentation cycle 26 h, centrifuge speed 8000 rpm in the post-treatment process, lipopeptide fermentation liquid pH 3.0, inlet and outlet pressure 2.0 / 1.2 MPa, lipopeptide fermentation liquid temperature 40 ℃ during nanofiltration, spray drying tower material inlet and outlet temperature of 170 / 80 ℃, use of a physical negative pressure defoaming device, and the final fermentation content reached 1.69 g / L.

[0101] Example 4 Preparation method of feed containing lipopeptide Preparation of lipopeptides Strain activation Paenibacillus aegypti Z121 with a preservation number of CGMCC No. 19830 was inoculated into LB slant medium for culture activation. After culturing at 37°C for 12 h, it was inoculated into a sterilized LB medium shake flask at an inoculum rate of 1%, and cultured at 37°C, initial pH 7.0, and rotation speed of 200 rpm for 12-16 h to obtain a seed solution.

[0102] Strain fermentation The seed liquid was inoculated into the optimized fermentation medium (the fermentation medium contained 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, 0.05% magnesium sulfate, and 1g / L aspartic acid or arginine, calculated as mass percentage) at a volume ratio of 3%. The fermentation temperature was 35°C, the initial pH was 6.5, and the rotation speed was 180 rpm. The culture was carried out for 6-8 h using a physical negative pressure defoaming device, which was continuously operated throughout the fermentation cycle until the fermentation was completed to ensure that the foam was effectively controlled to avoid affecting oxygen transfer and bacterial growth. The fermentation cycle was 26 h.

[0103] 1.3 Concentration and purification of lipopeptides (post-treatment process): First, the fermentation liquid in the fermenter was transported to a butterfly centrifuge through a closed pipe and centrifuged at 8000 rpm to remove bacteria and impurities and collect lipopeptides; secondly, nanofiltration concentration was performed at 40 °C, pH 3.0, and inlet and outlet membrane pressures of 2.0 / 1.2 MPa; then, the fermentation concentrate was spray-dried at an inlet and outlet temperature of 170 / 80 °C to obtain lipopeptide solid powder; 1.4 Detection of lipopeptide product content and recovery rate: The Z121 fermentation broth was extracted with acetonitrile and then prepared, separated and purified by high performance liquid chromatography. The yield of lipopeptide obtained by analytical liquid chromatography reached 1.69 g / L, with a maximum purity of 97.019%.

[0104] 2. Feed preparation: According to the formula of Example 1, a special feed for weaned piglets was prepared. Taking one ton of feed as an example, according to the above-mentioned component ratio, puffed corn, puffed soybean powder, yeast culture, glucose, fish meal, high-fat oil powder, calcium formate, calcium hydrogen phosphate, salt, acidulant, lysine, choline chloride, DL-methionine, threonine, compound enzyme, and premix were weighed and premixed respectively. The mixing time was 120 s, and mixture 1 was obtained after cooling; corn, soybean meal (46%), and fermented soybean meal were weighed, high-speed hammer crushed, the screen was 1.0 mm, put into a mixer, the temperature was adjusted to 75-85 ° C, mixed for 180 s, cooled to room temperature, and the lipopeptide solid powder obtained in step 3 was added to obtain mixture 2; mixture 1 and mixture 2 were mixed together for 180 s, and after cooling, a special feed for weaned piglets was obtained.

Claims

1. A special feed for weaned piglets, comprising 56.174% corn, 8.000% puffed corn, 17.500% soybean meal (46%), 4.000% puffed soybean powder, 5.500% fermented soybean meal, 2.000% yeast culture, 1.000% glucose, 1.000% fish meal, 1.000% high-fat oil powder, 0.800% calcium formate, 0.600% calcium hydrogen phosphate, 0.350% salt, 0.800% acidulant, 0.400% lysine, 0.100% choline chloride, 0.100% DL-methionine, 0.080% threonine, 0.070% complex enzyme, 0.520% premix, and 0.006% lipopeptide, calculated in percentage by mass.

2. The feed according to claim 1, wherein the nutritional level of the feed is 3.10 MJ / kg of metabolizable energy, and the contents of the main nutrients are: crude protein 18.60%, calcium 0.53%, total phosphorus 0.48%, available phosphorus 0.30%, lysine 1.25%, methionine 0.40%, methionine + cystine 0.69%, threonine 0.76%, and tryptophan 0.20%, calculated by mass percentage.

3. The feed according to claim 1 or 2, wherein the premix provides, per kilogram of feed, the following: Cu 5.12 mg, I 0.15 mg, Fe 87.59 mg, Mn 3.69 mg, Se 0.30 mg, Zn 84.43 mg, VA 9 000 IU, VD 3 000 IU, VE 24IU, VK 3 mg, thiamine 3 mg, riboflavin 7.5 mg, pantothenic acid 15 mg, niacin 30 mg, pyridoxine 3.60 mg, biotin 0.15 mg, folic acid 1.50 mg, and VB 12 0.036 mg.

4. The feed according to claims 1-3, characterized in that: The structural formula of the lipopeptide is shown in structural formula (I): 。 5. The feed according to claim 4, characterized in that: The lipopeptide is produced by Paenibacillus aegypti Z121 (CGMCC No. 19830).

6. A method for preparing the feed according to claims 1-5, comprising the following steps: Step 1, activation of the strain capable of producing the lipopeptide according to claim 5: the seed solution is inoculated at a rate of 1%, and the strain is activated and cultured in LB medium under the following culture conditions: 37°C, initial pH 7.0, rotation speed 200 rpm, and culture time 12-16 h; Step 2, strain fermentation: the strain was fermented in a fermentation medium at an inoculation rate of 3%, and the fermentation conditions were: temperature 35 °C, initial pH 6.5, rotation speed 180 rpm, and fermentation period 26 h; Step 3, post-treatment process: first, the fermentation liquid in the fermenter is transported to a butterfly centrifuge through a closed pipe and centrifuged at 8000 rpm to remove bacteria and impurities and collect lipopeptides; secondly, nanofiltration concentration is performed at 40°C, pH 3.0, and inlet and outlet membrane pressures of 2.0 / 1.2 MPa; then, the fermentation concentrate is spray-dried at an inlet and outlet temperature of 170 / 80°C to obtain lipopeptide solid powder; Step 4, feed preparation: according to the component ratio of claim 1, puffed corn, puffed soybean powder, yeast culture, glucose, fish meal, high-fat oil powder, calcium formate, calcium hydrogen phosphate, salt, acidulant, lysine, choline chloride, DL-methionine, threonine, complex enzyme, and premix are weighed respectively for premixing, the mixing time is 120 s, and mixture 1 is obtained after cooling; corn, soybean meal (46%), and fermented soybean meal are weighed, high-speed hammer crushing, the screen is 1.0 mm, put into a mixer, the temperature is adjusted to 75-85°C, mixed for 180 s, cooled to room temperature, and the lipopeptide solid powder obtained in step 3 is added to obtain mixture 2; mixture 1 and mixture 2 are mixed together for 180 s, and special feed for weaned piglets is obtained after cooling.

7. The method according to claim 6, characterized in that: The fermentation medium in step 2 contains 2% sucrose, 2% yeast powder, 0.5% potassium dihydrogen phosphate, 0.05% magnesium sulfate, and 1 g / L aspartic acid or arginine, calculated by mass percentage.

8. The method according to claim 6 or 7, characterized in that: A physical negative pressure defoaming device is used during the fermentation and cultivation stage. It starts running 6-8 hours after fermentation and continues until the end of fermentation.

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