CN112263586B - Hesperidin-rosemary anti-diarrhea preparation as well as preparation method and application thereof - Google Patents
Hesperidin-rosemary anti-diarrhea preparation as well as preparation method and application thereof Download PDFInfo
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Abstract
The invention discloses a hesperidin-rosemary anti-diarrhea preparation, a preparation method and application thereof, wherein the preparation comprises the following components in a concentration ratio of 10-20: 1, a hesperidin preparation and a rosmarinic acid preparation, the preparation method comprises the following steps: (1) Extracting pericarpium Citri Tangerinae by hot solvent method, and preparing hesperidin preparation; (2) Extracting rosemary leaves by a solvent extraction method to prepare a rosmarinic acid preparation; (3) Mixing the two preparations, then whirling for 30-60 s, and then placing in a shaking table for 2-3 h to prepare the hesperidin-rosemary anti-diarrhea preparation; the preparation can be used as feed additive in feed. The preparation is nontoxic and non-drug-resistant, reduces the pollution of livestock and poultry feces to the environment, has strong bactericidal capacity, stronger bacteriostatic capacity, obviously enhanced in-vitro free radical scavenging capacity, improved in-vitro fermentation parameters, promotes microbial fermentation, can effectively improve intestinal micro-ecological balance and improve the body resistance, and therefore can be added into feed.
Description
Technical Field
The invention relates to an anti-diarrhea preparation, a preparation method and application thereof, and in particular relates to a hesperidin-rosemary anti-diarrhea preparation, a preparation method and application thereof.
Background
Because a stable micro-ecosystem is not established in the intestinal tract of the piglet, the piglet has low self-resistance, is sensitive to external stimulation and is easily affected by invasion of various pathogenic microorganisms and various stress factors, infectious diarrhea is common for suckling piglets, but diarrhea caused by non-infectious factors such as daily ration antigen allergy, weaning, sudden feed replacement, cold, environmental stress and the like is mainly used for nursing piglets, sick pigs and pigs with viruses are main infection sources, viruses are discharged from excrement to pollute the environment and disseminate and infect, pigs fed in late autumn and early winter with high humidity, low temperature and high density are more easily infected with the disease, the pigs in the whole field are attacked within a few days, and huge economic loss is brought.
Disclosure of Invention
The purpose of the invention is as follows: the invention aims to provide a hesperidin-rosemary anti-diarrhea preparation which has small side effect, does not cause adverse reaction of organisms and does not generate drug resistance, and also aims to provide a preparation method and application of the preparation.
The technical scheme is as follows: the hesperidin-rosemary anti-diarrhea preparation comprises the components with the concentration ratio of 10-20: 1 hesperidin preparation and rosmarinic acid preparation.
Wherein the concentration ratio of the hesperidin preparation to the rosmarinic acid preparation is 10-20: 1.
the preparation method of the hesperidin-rosemary anti-diarrhea preparation comprises the following steps:
(1) Extracting pericarpium Citri Tangerinae by hot solvent method, and preparing hesperidin preparation;
(2) Extracting rosemary leaves by a solvent extraction method to prepare a rosmarinic acid preparation;
(3) Mixing the two preparations, then whirling for 30-60 s, and then placing in a shaking table for 2-3 h to prepare the hesperidin-rosemary anti-diarrhea preparation.
Wherein, step 1 includes the following steps:
(11) Crushing and sieving dried orange peel, and putting the dried orange peel into a reaction kettle;
(12) Adding an ethanol solution into a reaction kettle;
(13) Placing the reaction kettle at 100-150 ℃ for hot solvent extraction, and performing rotary evaporation to obtain hesperidin;
(14) Dissolving hesperidin in dimethyl sulfoxide to obtain hesperidin preparation.
Wherein, the dried orange peel is crushed in the step 11 and then is sieved by a 200-250 mesh sieve, the concentration of the ethanol solution in the step 12 is 60-75%, and the mass ratio of the volume of the added ethanol solution to the dried orange peel powder is 70-80 mL:1g of a compound; in the step 14, hesperidin is dissolved in dimethyl sulfoxide and then filtered by using a filter membrane of 0.22-0.28 μm.
Wherein, step 2 includes the following steps:
(21) Pulverizing herba Rosmarini officinalis leaf, and sieving;
(22) Adding an ethanol solution into the sieved rosemary powder for leaching to obtain an extracting solution, wherein the leaching time is 6-8 hours;
(23) Filtering the extract with a filter membrane, adding n-butanol, extracting, and recovering n-butanol;
(24) Adjusting the pH value of the recovered n-butanol to 2-2.5, adding ethyl acetate for extraction, eluting the extract liquor by using an ethanol solution, collecting filtrate by using X-5 macroporous resin, crystallizing at low temperature, filtering and drying to obtain rosmarinic acid;
(25) Dissolving rosmarinic acid in dimethyl sulfoxide to obtain rosmarinic acid preparation.
Wherein, the rosemary leaves in the step 21 are smashed and then pass through a 20-30 mesh sieve, the concentration of the ethanol solution in the step 22 is 50-60%, and the mass ratio of the volume of the added ethanol solution to the rosemary powder is 15-20 mL:1g, and leaching for 2-3 times; adjusting the pH of the extracting solution to 2-3 before adding n-butyl alcohol for extraction in the step 23, wherein the added n-butyl alcohol solution is 1.5-2 times of the volume of the extracting solution, the extraction time is 20-30 min, and the extraction times are 2-4; in the step 24, the ethyl acetate extraction time is 20-30 min, the extraction times are 3-5 times, the usage amount of the ethyl acetate is 1.5-2 times of the volume of the recycled n-butanol, and in the step 25, the rosmarinic acid is dissolved in the dimethyl sulfoxide and then filtered by a filter membrane of 0.22-0.28 μm.
Wherein, in the step 3, the rotating speed of the shaking table is 150-200 rpm, and the temperature is 30-40 ℃.
The hesperidin-rosemary anti-diarrhea preparation provided by the invention is applied to feed as a feed additive.
The working principle is as follows: the hesperidin has a dioxyflavone oxyglycoside structure, and the contained phenolic hydroxyl structure enables the hesperidin to have higher oxidation resistance. The hesperidin has bacteriostasis mainly by changing the permeability of phenols and terpenoid derivatives to bacterial cell membranes, and the derivatives have weak acidity and can denature or coagulate bacterial proteins. Researches show that hesperidin has antibacterial, antiinflammatory, and antioxidant effects. It has obvious inhibiting effect on colibacillus, bacillus subtilis, staphylococcus aureus, etc. in low pH condition. Rosmarinic acid is a water-soluble polyphenol hydroxy acid, and the strong effects of resisting oxidation and scavenging free radicals are probably related to the hydroxyl group of the catechol. The weak acidic and phenolic derivatives can destroy bacterial cell membranes and change the permeability of the cell membranes. The two components are used in combination to increase the inhibition effect on bacterial growth, and the hesperidin and the rosmarinic acid derivative have weak acidity, so that the damage of low pH to bacterial cell membranes and proteins can be further enhanced, and the antibacterial effect is stronger. In addition, hesperidin has strong antioxidant capacity, the clearance rate of free radicals is second to that of vitamin C, the clearance rate of free radicals of rosmarinic acid is weaker than that of hesperidin, and the antioxidant level of a compatible preparation can be enhanced by using the hesperidin and the rosmarinic acid in a matching way. Our research finds that hesperidin acts better at a higher ratio (10-20). The subsequent test results also show that the combination of the two can enhance the capability of inhibiting pathogenic bacteria, can also improve the functions of the organism such as oxidation resistance, inflammation inhibition and the like, promote the growth and development of animals, increase the average daily gain of rats and improve the structure of intestinal microbial flora. In addition, the fermentation parameters in vitro can be improved, the content of Short Chain Fatty Acids (SCFAs) in a fermentation system is improved or remarkably improved, and the microbial fermentation promoting agent has the effect of promoting microbial fermentation.
Has the advantages that: compared with the prior art, the invention has the remarkable advantages that: 1. the additive is non-toxic and non-drug resistant, and reduces the pollution of the livestock and poultry manure to the environment; 2. the microbial fermentation liquid has strong sterilization capability and bacteriostasis capability, remarkably enhances the in vitro free radical scavenging capability, improves in vitro fermentation parameters and promotes microbial fermentation; 3. can improve intestinal microecological balance and improve body resistance, and can be used as additive in feed.
Drawings
FIG. 1 is a graph of the change in body weight of rats: (a) a rat weight percent change curve; (b) histogram of daily weight gain of rats;
FIG. 2 is a graph comparing the diarrhea rate of rats;
FIG. 3 is a graph showing the effects of control group, hesperidin preparation, rosmarinic acid preparation and hesperidin-rosemary anti-diarrhea preparation on the content of intestinal inflammatory factors in LPS-challenged rats: (a) the MPO content of the ileum, caecum and colon; (b) ileal IL-6, IL-10 and TNF- α levels; (c) the amount of cecal IL-6, IL-10 and TNF- α; (d) colonic IL-6, IL-10 and TNF- α levels.
Detailed Description
Preparing a hesperidin preparation:
crushing dried orange peel, sieving with a 250-mesh sieve, weighing 0.2g of sieved dried orange peel powder in a 25mL stainless steel reaction kettle with a polytetrafluoroethylene lining, adding 60% ethanol solution according to a material-to-liquid ratio of 70 (mL: g), performing hot solvent extraction at 120 ℃ for 45min, centrifuging at 4000r/min for 5min after extraction is finished, immediately performing rotary evaporation to remove ethanol to obtain hesperidin, weighing 4.00g of hesperidin in a centrifuge tube, adding 10mL of dimethyl sulfoxide to dissolve and mix uniformly, placing on a 37 ℃,150rpm shaking table for 2h, taking out and mixing uniformly for 30s every 30min, taking out and using sterile water to 10mL, filtering through a 0.22 mu m filter membrane to prepare a 400mg/mL hesperidin preparation, and storing in a 4 ℃ refrigerator for constant volume standby.
Preparation of rosmarinic acid preparation:
the preparation method comprises the following steps of crushing rosemary leaves, sieving the rosemary leaves by a 20-mesh sieve, weighing 100.0g of sieved rosemary leaf powder, sealing the rosemary leaf powder in a polyethylene bag, adding an ethanol solution with the volume concentration of 15 times and 50%, leaching twice, wherein the leaching time is 6 hours, collecting a leaching solution, pretreating the extracting solution by using a microfiltration membrane, filtering for three times, removing suspended particles in the extracting solution, adjusting the pH of filtrate to 3 by using hydrochloric acid, extracting by using 1.5 times of volume of n-butyl alcohol, and extracting under the conditions that: extracting twice, each time being 20min, recovering n-butanol, adding high pure water into the recovered n-butanol, recording the volume, adjusting the pH to 2.0-2.5 by hydrochloric acid, extracting for 3 times by ethyl acetate with the volume ratio of 10. Weighing 4.00g of rosmarinic acid into a centrifuge tube, respectively adding 10mL of dimethyl sulfoxide to dissolve and mix uniformly, placing the centrifuge tube in a shaking table with 37 ℃ and 150rpm for 2h, taking out the mixture at intervals of 30min, mixing uniformly for 30s, taking out, metering the volume to 10mL by using sterile water, filtering the mixture by using a 0.22 mu m filter membrane to prepare a rosmarinic acid preparation with the volume of 400mg/mL, and storing the rosmarinic acid preparation in a refrigerator with the temperature of 4 ℃ for later use.
Preparing hesperidin-rosemary anti-diarrhea preparation:
diluting the rosmarinic acid preparation and the hesperidin preparation to 40mg/mL, mixing the two combined preparations according to the concentration of 1.
The results of in vitro bacteriostatic experiments on the hesperidin preparation, the rosmarinic acid preparation and the hesperidin-rosemary anti-diarrhea preparation are shown in table 1, and it can be seen from table 1 that the hesperidin-rosemary anti-diarrhea preparation has strong killing capacity on enterotoxigenic escherichia coli, staphylococcus aureus and salmonella; the minimum bacteriostatic concentrations measured by an MIC method test are respectively 0.5mg/mL;0.5mg/mL;0.5mg/mL. :
TABLE 1 inhibitory Effect (mg/mL) of hesperidin preparation, rosmarinic acid preparation and hesperidin-Rosemary diarrhea-suppressing agent against pathogenic bacteria
Note: "_" indicates that no bacteriostatic action was detected in the range of 0 to 400mg/mL
The influence of the hesperidin preparation, the rosmarinic acid preparation and the hesperidin-rosemary anti-diarrhea preparation on the in-vitro fermentation level of the pig colon microorganisms is simulated by using an in-vitro fermentation method, and the results are shown in table 2:
TABLE 2 results of simulated fermentation experiments of hesperidin preparation, rosmarinic acid preparation and hesperidin-rosemary preparation for anti-diarrhea
As shown in Table 2, the experiment proves that the hesperidin, rosmarinic acid and hesperidin-rosemary anti-diarrhea preparation as plant extracts have great improvement on in vitro fermentation parameters, and the improvement of the combined preparation on the in vitro fermentation parameters is better than or obviously better than that of a single plant extract preparation.
The animal tests of the hesperidin preparation, the rosmarinic acid preparation and the hesperidin-rosemary anti-diarrhea preparation are carried out in an animal test center of Nanjing agriculture university, and the animal tests are carried out for 12 months to 2020 months in 2019. The test was carried out on 40 weaned SD rats of 21 days old and weighing about 50 + -2.0 g, divided into 4 treatments, i.e., a control group (Con), a hesperidin group (Hes), a rosmarinic acid + hesperidin group (Ros-A + Hes), and a rosmarinic acid group (Ros-A), wherein 10 rats of similar body weight and age were treated by half of male and female. The gavage amount of each rat is shown in table 3, and the rats were weighed daily and gavaged once. During the test period, the food and the water are taken freely, the pre-test period is 7 days, the formal test period is 14 days, the total period is 21 days, and the food and the water are taken freely during the test process. On the 21 st day of the experiment, all rats were intraperitoneally injected with 1mg/kg LPS, and 8 hours after the injection of lipopolysaccharide, all rats were sacrificed by decapitation and sampled.
TABLE 3 addition amounts of rosmarinic acid preparation, hesperidin preparation and hesperidin-rosemary anti-diarrhea preparation
During the test period, the food intake is recorded every day, and the food intake is weighed and recorded at 8 points in the morning on the 1 st, 4 th, 7 th, 10 th, 11 th, 12 th, 13 th and 14 th days of the test respectively; after a rat is slaughtered, weighing small intestines (including duodenum, jejunum, ileum and chyme thereof), taking jejunum middle sections, ileum middle sections and caecum of each group, carrying out HE (high intensity staining) dyeing, measuring the height of villus and the depth of crypts, and calculating the ratio of the height of the villus to the depth of the crypts, wherein the ratio is used for evaluating the protection effect of the hesperidin-rosemary anti-diarrhea preparation on the morphology of intestinal tracts after the toxicity of LPS is attacked; the kit is adopted to measure the contents of SOD, MDA, T-AOC and GSH-px in the serum of each group of rats, and is used for evaluating the protection efficacy of the hesperidin-rosemary anti-diarrhea preparation and the intestinal morphological organism antioxidant system after the LPS is attacked; the kit is adopted to measure the contents of IL-6, IL-8, IL-10 and TNF-alpha in intestinal mucosa of rats of each group, so as to evaluate the protective effect of the hesperidin-rosemary anti-diarrhea preparation on an organism inflammation system after LPS challenge.
The effects of the control group, the rosmarinic acid preparation, the hesperidin preparation and the hesperidin-rosemary anti-diarrhea preparation on the weight gain of the rat are shown in fig. 1 (a) and fig. 1 (b), and it can be seen from the figure that the rat gavage hesperidin, rosmarinic acid and the hesperidin-rosemary anti-diarrhea preparation can improve the growth performance of the rat. After 5 days of gastric perfusion, the weight of the rats in the hesperidin-rosmarinic acid anti-diarrhea preparation group is obviously separated from the control group, the hesperidin preparation group and the rosmarinic acid preparation group, and the average daily gain of the rats in the hesperidin-rosmarinic acid anti-diarrhea preparation group is obviously higher than that of the control group and a single plant extract group (P is less than 0.05). The effect of the control group, the rosmarinic acid preparation, the hesperidin preparation and the hesperidin-rosemary anti-diarrhea preparation on the diarrhea rate of the rats is shown in figure 2, and it can be seen from the figure that the diarrhea rate of the weaned rats of the gavage plant extract is reduced compared with the control group, wherein the diarrhea rate of the rats of the hesperidin-rosemary anti-diarrhea preparation group is obviously lower than that of the control group (P < 0.05). Furthermore, the diarrhea rate of the hesperidin-rosemary anti-diarrhea preparation group rats is lower than that of the single plant extract group, but the significant level (P > 0.05) is not achieved. The effects of the control group, the rosmarinic acid preparation, the hesperidin preparation and the hesperidin-rosemary anti-diarrhea preparation on the intestinal morphological structure of LPS challenge rats are shown in table 4: compared with the control group, the gavage plant extract can obviously improve the morphological structure of jejunum, ileum and cecum of rats. Wherein, the gavage of the hesperidin-rosemary anti-diarrhea preparation can obviously improve the villus height of the jejunum of a rat and obviously reduce the crypt depth (P < 0.05), and the villus cryptic ratio of the jejunum of the rat in the hesperidin-rosemary anti-diarrhea preparation group is also obviously higher than that of a control group and a single plant extract group (P < 0.05). The ileum villus height of the rats in the gavage hesperidin-rosemary antidiarrheal preparation group is also obviously higher than that of the control group (P < 0.05), but does not reach the obvious level compared with a single plant extract; the depth of the crypts is slightly higher than that of the control group, and does not reach a significant level; however, the ratio of the hesperidin to the rosemary anti-diarrhea preparation in the rat villus hiding ratio is obviously higher than that in the control group and the hesperidin preparation group (P < 0.05). In addition, the hesperidin-rosemary anti-diarrhea preparation group rats had significantly higher cecal villus height and mucosal thickness than the control group (P < 0.05), but did not significantly differ from the single plant extract group (P > 0.05).
TABLE 4 influence of control group, rosmarinic acid preparation, hesperidin preparation and hesperidin-rosemary anti-diarrhea preparation on intestinal morphological structure of LPS-challenged rat
The effects of the control group, the rosmarinic acid preparation, the hesperidin preparation and the hesperidin-rosemary anti-diarrhea preparation on the antioxidant index of the LPS challenge rat are shown in Table 5, and it can be seen from Table 5 that the lavage plant extracts of SD rat and the combination thereof can improve the antioxidant level of the liver and the intestinal tract. Compared with the control group, the hesperidin-rosemary antidiarrheal preparation group can obviously improve the T-AOC and GSH-PX levels of ileum, caecum and liver (P < 0.05), and obviously reduce the MDA content of caecum, colon, jejunum and liver (P < 0.05). The total analysis shows that the antioxidant level of the intestinal tract and the liver of the rats in the hesperidin-rosemary anti-diarrhea preparation group is slightly higher or remarkably higher than that of the single plant extract group.
TABLE 5 influence of control group, rosmarinic acid preparation, hesperidin preparation and hesperidin-rosemary anti-diarrhea preparation on antioxidant index of LPS challenge rat
As can be seen from fig. 3 (a), 3 (b), 3 (c), and 3 (d), the rosmarinic acid formulation, hesperidin formulation, and hesperidin-rosemary antidiarrheal formulation all reduced the MPO levels in the ileum, cecum, and colon of LPS challenged rats compared to the control group, wherein the hesperidin-rosemary antidiarrheal formulation group reached significant levels (P < 0.05). The content of IL-10 in ileum mucosa, caecum and colon mucosa of the hesperidin-rosemary anti-diarrhea preparation group is obviously higher than that of a control group (P is less than 0.05), and the content of IL-10 in the ileum mucosa of the hesperidin-rosemary anti-diarrhea preparation group is also obviously higher than that of a rosmarinic acid preparation group and slightly higher than that of the hesperidin preparation group, but the difference is not obvious. The content of IL-6 in ileum, caecum and colon mucosa of the hesperidin-rosemary anti-diarrhea preparation group is obviously lower than that of a control group and a single plant extract group (P < 0.05). The content of IL-6 in the mucous membranes of the three intestinal sections of the rosmarinic acid preparation group and the hesperidin preparation group is slightly lower than that of the control group, but does not reach the significant level.
The total bacterial count of the intestinal tract reflects the steady state of the intestinal tract, and the increase of the total bacterial count indicates that the intestinal tract has an increased microbial steady state. In the experiment, the total bacterial count in the four intestinal sections of empty, ilex, blindness and colon is obviously higher than that of a control group and higher than or obviously higher than that of a single plant extract preparation group, which shows that the hesperidin-rosemary anti-diarrhea preparation can improve the steady state of the intestinal flora of LPS-challenged rats. Escherichia coli is a conditional pathogen with a large number in intestinal tracts, and the increase of the number of Escherichia coli or Enterobacter flora can cause diseases such as diarrhea and the like. The lactobacillus is beneficial bacteria in intestinal tracts, the lactic acid produced by the metabolism of the lactobacillus can be utilized by organisms, the pH value of the intestinal tracts can be reduced, and the growth of harmful bacteria is inhibited, and the influence of a control group, a rosmarinic acid preparation, a hesperidin preparation and a hesperidin-rosemary anti-diarrhea preparation on the number of intestinal flora of LPS challenge rats is shown in Table 6:
TABLE 6 influence of control group, rosmarinic acid preparation, hesperidin preparation and hesperidin-rosemary anti-diarrhea preparation on the number of intestinal flora in LPS-challenged rats
As can be seen from table 6, compared to the control group, the hesperidin-rosemary anti-diarrhea preparation group can significantly increase the number of lactobacillus in each intestinal segment and significantly reduce the number of escherichia coli (P < 0.05), and compared to the single plant extract group, the hesperidin-rosemary anti-diarrhea preparation group has reduced or significantly reduced number of escherichia coli and increased or significantly increased number of lactobacillus. The hesperidin-rosemary anti-diarrhea preparation is shown to be capable of increasing the number of beneficial bacteria in intestinal tracts, reducing the number of conditional pathogenic bacteria escherichia coli, adjusting the flora structure of the intestinal tracts and improving the health level of the intestinal tracts.
Claims (9)
1. A hesperidin-rosemary preparation for resisting diarrhea is characterized by being prepared from a hesperidin preparation and a rosmarinic acid preparation, wherein the concentration ratio of the hesperidin preparation to the rosmarinic acid preparation is 15.
2. A process for the preparation of a hesperidin-rosemary anti-diarrhea preparation according to claim 1, comprising the steps of:
(1) Extracting pericarpium Citri Tangerinae by hot solvent method, and preparing hesperidin preparation;
(2) Extracting rosemary leaves by a solvent extraction method to prepare a rosmarinic acid preparation;
(3) Mixing the two preparations, then swirling for 30 to 60 seconds, and then placing in a shaking table for 2 to 3 hours to prepare the hesperidin-rosemary anti-diarrhea preparation.
3. The process for the preparation of hesperidin-rosemary anti-diarrhea preparation according to claim 2, wherein the step 1 comprises the following steps:
(11) Crushing and sieving dried orange peel, and putting the dried orange peel into a reaction kettle;
(12) Adding an ethanol solution into a reaction kettle;
(13) Placing the reaction kettle at 100-150 ℃ for hot solvent extraction, and performing rotary evaporation to obtain hesperidin;
(14) Dissolving hesperidin in dimethyl sulfoxide to obtain hesperidin preparation.
4. The preparation method of the hesperidin-rosemary anti-diarrhea preparation according to claim 3, wherein the dried orange peel is pulverized in the step 11 and then sieved through a 200-250 mesh sieve, the concentration of the ethanol solution in the step 12 is 60-75%, and the mass ratio of the volume of the added ethanol solution to the dried orange peel powder is 70-80 mL:1g; in the step 14, the hesperidin is dissolved in dimethyl sulfoxide and filtered by a filter membrane of 0.22 to 0.28 mu m.
5. The process for the preparation of a hesperidin-rosemary preparation for anti-diarrhea according to claim 2, wherein the step 2 comprises the steps of:
(21) Pulverizing herba Rosmarini officinalis leaf, and sieving;
(22) Adding an ethanol solution into the sieved rosemary powder, and leaching for 6 to 8 hours to obtain an extracting solution;
(23) Filtering the extract with a filter membrane, adding n-butanol, extracting, and recovering n-butanol;
(24) Adjusting the pH value of the recycled n-butanol to 2-2.5, adding ethyl acetate for extraction, eluting the extract by using an ethanol solution, collecting filtrate by using X-5 macroporous resin, crystallizing at low temperature, filtering and drying to obtain rosmarinic acid;
(25) Dissolving rosmarinic acid in dimethyl sulfoxide to obtain rosmarinic acid preparation.
6. The preparation method of the hesperidin-rosemary anti-diarrhea preparation according to claim 5, wherein rosemary leaves are ground in the step 21 and then screened through a sieve of 20 to 30 meshes, the concentration of the ethanol solution in the step 22 is 50 to 60 percent, and the mass ratio of the volume of the added ethanol solution to the rosemary powder is 15 to 20mL:1g, the extraction frequency is 2~3, the pH of the extracting solution is adjusted to 2~3 before adding n-butanol in the step 23 for extraction, the added n-butanol solution is 1.5 to 2 times of the volume of the extracting solution, the extraction time is 20 to 30min, and the extraction frequency is 2~4.
7. The method for preparing the hesperidin-rosemary preparation for resisting diarrhea according to claim 5, wherein the ethyl acetate extraction time in the step 24 is 20 to 30min, the extraction times are 3~5, the usage amount of the ethyl acetate is 1.5 to 2 times of the volume of the recovered n-butanol, and the rosmarinic acid is dissolved in dimethyl sulfoxide and then filtered by a 0.22 to 0.28 μm filter membrane in the step 25.
8. The preparation method of the hesperidin-rosemary anti-diarrhea preparation according to claim 2, wherein the shaking table rotation speed in the step 3 is 150 to 200rpm, and the temperature is 30 to 40 ℃.
9. Use of the hesperidin-rosemary anti-diarrhea preparation of claim 1 in the preparation of a feed additive.
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