Preparation method and application of feed additive containing ampelopsis grossedentata extract

Abstract

The invention discloses a preparation method and application of a feed additive containing ampelopsis grossedentata extract. A supercritical fluid extraction technology and a molecular distillation method are combined in front-stage production, main chemical components such as dihydromyricetin and quercetin of ampelopsis grossedentata flavone are extracted, meanwhile, in order to screen out chemical components with a powerful antibacterial function, a creative high-concentration membrane separation esterification technology is adopted, and the yield of ampelopsis grossedentata flavone is improved. Dihydromyricetin and quercetin are subjected to secondary extraction, so that the concentrations of dihydromyricetin and quercetin in the raw materials are kept to a golden proportion for exerting the physiological effect of cells. And the production of the rear section is mainly a mixing process. The natural plant extract is used for replacing antibiotics, so that the drug resistance caused by antibiotic abuse is avoided, and the safety of the livestock feed is improved; meanwhile, gallic acid, a yeast extract and yeast immune polysaccharide are adopted for compatibility modification, so that the synergistic effect of various effective components is better exerted.

Description

Technical Field

[0001] The invention belongs to the technical field of feed additives and relates to a preparation method of a feed additive containing a rattan tea extract and application thereof. Background Art

[0002] Feed additives are substances added in small or trace amounts during feed production, processing, and consumption. While their use in small quantities can have significant effects, modern intensive farming practices currently involve large numbers of animals living in relatively densely packed spaces. This environment facilitates the rapid spread of infectious diseases (such as respiratory and intestinal diseases). Once an outbreak occurs, the losses can be devastating. To prevent and control disease outbreaks, antibiotics are often added to livestock and poultry feed. However, long-term antibiotic use can lead to risks such as antibiotic resistance, antibiotic residues, and environmental pollution. Therefore, the reduction and elimination of antibiotics is an inevitable trend in the development of China's livestock and poultry industry. Currently, the use of natural plant extracts as feed additives, replacing antibiotics, has significant market potential in recent years. Since the reduction and elimination of antibiotics is an inevitable trend in the development of China's livestock and poultry industry, the elimination of antibiotics in feed will comprehensively improve the quality of my country's livestock and poultry products.

[0003] Ampelopsis grossedentata, also known as Ampelopsis grossedentata, is a plant of the genus Ampelopsis in the Vitaceae family. It has been used as a health tea and traditional Chinese medicine for centuries. This plant is primarily found in areas south of the Yangtze River, including Fujian, Yunnan, Guangdong, Guangxi, Guizhou, and Hunan. It is a plant that can be used as both a medicine and a food. The total flavonoids in Ampelopsis grossedentata exhibit a variety of pharmacological activities, including anti-inflammatory, antibacterial, hypoglycemic, hypolipidemic, hypotensive, antioxidant, anti-atherosclerotic, hepatoprotective, and anti-tumor properties. Flavonoids have potent inhibitory effects against Escherichia coli, Salmonella, and Staphylococcus aureus in the intestines. They also possess powerful free radical scavenging properties, increasing the antioxidant capacity of intestinal tissue and liver and reducing inflammatory responses. Therefore, the application of Ampelopsis grossedentata extract as a feed additive in livestock and poultry farming to improve production performance, enhance disease resistance and immunity, and reduce the incidence of gastrointestinal inflammation has become an urgent technical challenge in this field. Summary of the Invention

[0004] In view of this, the purpose of the present invention is to address the deficiencies of the prior art and provide a method for preparing and using a feed additive containing a rattan tea extract. This feed additive avoids drug resistance caused by the abuse of antibiotics, improves the safety of poultry and livestock feed, enhances disease resistance and immunity, and reduces and minimizes the occurrence of digestive tract inflammation.

[0005] The technical solution adopted by the present invention is: a method for preparing a feed additive containing rattan tea extract, comprising the following steps: S1. Crude extraction of Tengcha extract: Supercritical fluid extraction technology combined with molecular distillation was used to extract the main chemical components of Tengcha flavonoids: dihydromyricetin and quercetin; S2. Purification of rattan tea extract: Using high-concentration membrane separation and esterification technology, the crude extract in S1 was subjected to secondary extraction of dihydromyricetin and quercetin, so that the ratio of dihydromyricetin to quercetin in the rattan tea extract was (25-80):1; S3, weighing 15-20 parts of the vine tea extract prepared in S2, 25-30 parts of gallic acid, 30-40 parts of yeast extract, and 20-30 parts of yeast immune polysaccharide by weight, and stirring and mixing them uniformly to form a premix; S4. Weigh 1-5 parts of the premix prepared in S3, add 10-50 parts of an acidifier, 0.5-5 parts of an enzyme preparation, and 1-10 parts of probiotics, and mix well. S5. Dry and store Tengcha flavonoids are extracts from Tengcha, primarily containing dihydromyricetin (DMY), myricetin, quercetin, ampelopsis, and plant polysaccharides. The extremely high content of dihydromyricetin is the most unique and important active ingredient in Tengcha flavonoids. Together with quercetin, it forms a powerful biologically active component, inhibiting harmful intestinal bacteria, eliminating inflammation, and enhancing immunity in animals, making it a viable alternative to antibiotics in animal production. In the aforementioned preparation method, the initial production phase utilizes supercritical fluid extraction combined with molecular distillation to extract the main chemical components of Tengcha flavonoids, including dihydromyricetin and quercetin. To identify chemical components with potent antibacterial properties, a unique high-concentration membrane separation and esterification technique is then employed for secondary extraction of dihydromyricetin and quercetin, maintaining their concentrations in the raw material at the optimal ratio for achieving cellular physiological effects. The subsequent production phase primarily involves a mixing process. Gallic acid, yeast extract, and yeast immune polysaccharide are also used for compatibility modification to maximize the synergistic effects of the various active ingredients.

[0006] Preferably, in step S1, the dried leaves of the Ampelopsis ulmoides L. are first crushed and then dried at low temperature; then loaded into an extraction kettle, and an entrainer ethanol-water is injected into the CO2 flow through a high-pressure pump for supercritical CO2 extraction; then dihydromyricetin and quercetin are purified by molecular distillation, and quercetin is further enriched in the distillation residue phase by polyamide column chromatography (eluent: ethanol-water gradient).

[0007] Preferably, the dried leaves of Tengcha are crushed to 40-60 mesh. Too fine mesh will clog the equipment, while too large mesh will lead to insufficient extraction.

[0008] Preferably, the low-temperature drying temperature is controlled below 40° C. to reduce the moisture content to less than 5% to avoid agglomeration.

[0009] Preferably, the entrainer has an ethanol:water ratio of 8:2, and the entrainment capacity is 15-25% of the CO2 flow rate. Since dihydromyricetin and quercetin are both polar molecules, their solubility in pure CO2 is low, so an entrainer is added to enhance the solubility of polar flavonoids.

[0010] Preferably, in step S1, the extraction pressure is controlled at 30-45 MPa, the temperature is within the range of 50-60°C, the CO2 flow rate is 20-30 L / h, and the extraction time is 1.5-2.5 h. High extraction pressure can increase density and improve solubility.

[0011] Preferably, in step S1, the evaporation temperature of dihydromyricetin is 140-160° C., the evaporation temperature of quercetin is 180-220° C., and the pressure is 0.1-1 Pa. The present invention separates dihydromyricetin and quercetin by using the difference in boiling points.

[0012] Preferably, in step S2, dihydromyricetin and quercetin are first esterified using an esterifying agent, and then separated by a high-concentration membrane to obtain a dihydromyricetin ester-enriched liquid quercetin and a quercetin ester-enriched liquid, and then the dihydromyricetin ester-enriched liquid quercetin and the quercetin ester-enriched liquid are deesterified by a deesterifying agent to obtain dihydromyricetin and quercetin. The present invention increases the molecular weight difference between dihydromyricetin / quercetin and impurities through the esterification reaction, thereby improving the membrane separation selectivity.

[0013] Preferably, the esterification agent of dihydromyricetin is acetic anhydride, so that dihydromyricetin is converted into dihydromyricetin triacetate, and the molecular weight increases from 320.25 to 446.39 Da; the deesterification agent is methanol, so that dihydromyricetin triacetate is deesterified to dihydromyricetin.

[0014] Preferably, the esterifying agent of quercetin is propionic anhydride, so that quercetin is converted into quercetin-tripropionate, and the molecular weight increases from 302.23 to 470.45 Da; the deesterifying agent is ethanol, so that quercetin-tripropionate is deesterified to quercetin.

[0015] Preferably, the high-concentration membrane is a molecular sieve membrane, which can accurately separate dihydromyricetin triacetate and quercetin tripropionate by utilizing the difference in molecular weight after esterification.

[0016] Preferably, the acidifier is a composite solid organic acid or a coated slow-release acidifier. Acidifiers can acidify the digestive tract environment and inhibit the growth of pathogens. Coated slow-release acidifiers can more effectively reach the posterior intestinal tract.

[0017] Preferably, the enzyme preparation is one or more of phytase, protease or amylase. Enzyme preparation can improve feed digestibility and reduce intestinal undigested matter that may nourish harmful bacteria.

[0018] Preferably, the probiotic is a Lactobacillus / Bifidobacterium or Bacillus. Probiotics can regulate the balance of intestinal flora, inhibit pathogens, and enhance intestinal health.

[0019] The use of the feed additive according to claim 1 in poultry feed, wherein the total mass of the feed additive is 0.1 to 0.3‰ of the total mass of the poultry feed.

[0020] The use of the feed additive according to claim 1 in pig feed, wherein the total mass of the feed additive is 0.3 to 0.6‰ of the total mass of the pig feed.

[0021] Compared with the prior art, the present invention has the following advantages: 1. The present invention focuses on screening out dihydromyricetin and quercetin, chemical components with powerful antibacterial functions, from rattan tea, and uses natural plant extracts to replace antibiotics, thereby avoiding drug resistance caused by the abuse of antibiotics and improving the safety of poultry and livestock feed.

[0022] 2. The innovative compounding scheme of the present invention is compounded with multiple functional components such as gallic acid, yeast extract, yeast immune polysaccharide, acidifier, enzyme preparation and probiotics, so that the feed additive has the effects of improving livestock and poultry production performance, enhancing disease resistance and immunity, reducing and reducing the occurrence of digestive tract inflammation, and improving meat quality. It has a wide range of applications and is safe.

[0023] 3. The invention adds yeast extract, which can act as an appetite inducer. The invention develops an innovative flavor improvement solution to enhance the acceptance of natural extracts of rattan tea by poultry, livestock and aquatic products. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 This is the effect of the experiment on the jejunum morphology of piglets 36 days after weaning. DETAILED DESCRIPTION

[0025] To deepen the understanding of the present invention, the present invention will be further described in detail below with reference to implementation cases and drawings, but the content of the present invention is not limited to the following embodiments. Example

[0026] The method for preparing a feed additive containing a rattan tea extract of the present invention comprises the following steps: S1. Crude extraction of Tengcha extract: Supercritical fluid extraction technology combined with molecular distillation was used to extract the main chemical components of Tengcha flavonoids: dihydromyricetin and quercetin; the details are as follows: First, the dried leaves of Camellia tengcha were crushed to 40-60 mesh, and then the temperature was controlled below 40°C and low-temperature dried to a moisture content of <5%; then loaded into the extraction kettle, and the entrainer ethanol-water was injected into the CO2 flow through a high-pressure pump for supercritical CO2 extraction, wherein the entrainer ethanol: water = 8:2, the entrainment amount was 15-25% of the CO2 flow rate, the extraction pressure was set to 40 MPa, the temperature was 55°C, the CO2 flow rate was 25L / h, and the extraction time was 2 h. After the solute-containing fluid entered the separation kettle, the pressure was reduced to 5-10 MPa, the temperature was controlled at 35-40°C, and the solute was precipitated and collected; then dihydromyricetin and quercetin were purified by molecular distillation, and quercetin was further enriched in the distillation residue by polyamide column chromatography (eluent: ethanol-water gradient). The main purpose of molecular distillation purification is to separate dihydromyricetin and quercetin by boiling point difference. The specific distillation purification parameters are as follows Table 1: parameter Dihydromyricetin fraction Quercetin-enriched fraction Evaporation temperature 140-160℃ 180-220℃ Operating pressure 0.1-1 Pa 0.1-1 Pa Feed rate 1-2 mL / min 1-2 mL / min Scraping speed 200-300 rpm 200-300 rpm Condensation temperature 40-50℃ 60-70℃ Table 1 S2. Purification of the Tengcha Extract: Using high-concentration membrane separation and esterification technology, the crude extract from S1 is subjected to secondary extraction of dihydromyricetin and quercetin, resulting in a dihydromyricetin:quercetin ratio of (25-80):1 in the Tengcha Extract. The high-concentration membrane separation and esterification technology described in the present invention primarily includes esterification modification, high-concentration membrane separation into an enriched solution, and deesterification, as follows: 1. The esterification modification scheme is shown in Table 2: Table 2 During high-concentration membrane separation, the temperature is controlled at 25-35°C and the pH is controlled between 6.5-7.0.

[0027] The deesterification scheme is shown in Table 3. After deesterification, neutralization is performed first, then the solvent is removed by rotary evaporation, and finally, pure dihydromyricetin and quercetin are obtained by freeze-drying. The purity of the final product is: dihydromyricetin ≥98%, quercetin ≥95%. Table 3 S3, weighing 18 parts of the vine tea extract prepared in S2, 30 parts of gallic acid, 35 parts of yeast extract, and 25 parts of yeast immune polysaccharide by weight, and stirring and mixing them uniformly to form a premix; S4. Weigh 4 parts of the premix prepared in S3, add 30 parts of an acidifier (composite solid organic acid), 4 parts of an enzyme preparation (protease), and 5 parts of probiotics (Lactobacillus / Bifidobacterium), and mix well. S5. Drying and preserving to obtain the feed additive containing the rattan tea extract.

[0028] Examples 2-5 The preparation methods of Examples 2-5 are the same as steps S1-S3 of Example 1, except that the raw material components of S4 are different from those of Example 1. The raw material components of Examples 1-5 in parts by weight are shown in Table 4: Components Example 1 Example 2 Example 3 Example 4 Example 5 Tengcha Extract 18 18 18 18 18 Gallic acid 30 30 30 30 30 yeast extract 35 35 35 35 35 Yeast immune polysaccharide 25 25 25 25 25 S3 premix 3 2 3 5 1 acidifier Composite solid organic acid 30 Coated sustained-release acidifier 10 Composite solid organic acid 30 Composite solid organic acid 50 Coated sustained-release acidifier 20 enzyme preparations Protease 4 Phytase 4 Phytase 3 Protease 5 Protease 2 Probiotics (Lactobacillus / Bifidobacterium) 5 3 5 10 1 Table 4 The above Examples 1-5 were added to feed, and their uses and total feed mass addition amounts are shown in Table 5 below: Example 1 Example 2 Example 3 Example 4 Example 5 usefulness Conservation materials Medium and large pig feed Pig feed creep material poultry feed Total feed mass addition amount (‰) 0.5 0.3 0.4 0.6 0.2 Table 5 This embodiment 1 was used for feeding weaned piglets to conduct an experiment, and the experimental design was as follows: 1. Experimental pigs: 120 weaned piglets, half male and half female, were evenly distributed into three groups, namely, a blank control group of 40, an experimental group of 40, and an antibiotic group of 40, with 4 replicates in each group and 10 pigs in each replicate.

[0029] 2. Test steps Feed selection: The blank control group was given ordinary piglet feed without additives or antibiotics; The test group was given piglet feed supplemented with the feed additive of Example 1; The antibiotic group was given piglet feed containing 40 ppm of antifungal, 60 ppm of bacitracin, 70 ppm of chlortetracycline and 2000 ppm of zinc oxide.

[0030] Feeding method: The experimental pigs were fed three times a day and had free access to water. The experimental period was 45 days.

[0031] 3. The results of the above test on the diarrhea rate of piglets aged 1-14 days after weaning are shown in Table 6 below: Blank control group Test agent group Antibiotic group Diarrhea rate, % 8.65 4.52 4.76 Stool scoring 0.15 0.12 0.13 Table 6 The results of the above test on the morphological analysis of the jejunum of 36-day-old piglets after weaning are shown in the attached manual. Figure 1 shown.

[0032] The following conclusions can be drawn from the above test results: 3.1. The antibiotic replacement effect of the test group was comparable to that of the antibiotic group, indicating that the feed additive of the present invention can improve intestinal health, reduce intestinal inflammation and nutritional diarrhea, reduce antibiotic resistance, and reduce or replace the amount of antibiotics added to the diet.

[0033] 3.2. Morphological analysis of the jejunum of 36-day-old piglets revealed intact intestinal villi in the experimental and antibiotic groups, while the blank control group showed poorer intestinal villi integrity. The order of intestinal villi integrity was: experimental group ≈ antibiotic group > blank control group. These results demonstrate a high correlation between diarrhea rate and the degree of damage to jejunal villi morphology, further confirming the protective effects of the present invention on the intestine and demonstrating favorable nutritional and physiological characteristics.

[0034] This Example 5 was used for feeding white-feathered broiler chickens and the experimental design was as follows: 1. Experimental chickens: 200 white-feathered broiler chickens from the same batch of vaccine were selected, including 100 each of the experimental agent group and the antibiotic control group.

[0035] 2. Test steps The test group was given poultry feed to which the feed additive of Example 5 was added at an amount of 0.2‰; The antibiotic control group was given poultry feed with 0.1‰ avilamycin added.

[0036] Feeding method: The chickens were raised according to the normal feeding and management procedures of white-feathered broilers, with free access to food and water. The experimental period was 30 days.

[0037] 3. The results of the above test on the growth performance of white-feathered broiler chickens are shown in Table 7 below: project Weight (g) Feed intake (g) F / G Abdominal fat (%) Antibiotic control group 2672 4426 1.688 1.00 Test agent group 2688 4413 1.673 0.92 Table 7 From the above test results, it can be concluded that, compared with the antibiotic control group, the addition of the present invention to broilers has no effect on production performance indices such as weight gain, feed intake and feed-to-meat ratio, further indicating that the present invention can replace antibiotics in poultry feed.

[0038] Finally, it should be noted that the above is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the scope of protection of the present invention.

Claims

1. A method for preparing a feed additive containing a rattan tea extract, characterized in that: The following steps are involved: S1. Crude extraction of Tengcha extract: Supercritical fluid extraction technology combined with molecular distillation was used to extract the main chemical components of Tengcha flavonoids: dihydromyricetin and quercetin; S2. Purification of the extract of Ampelopsis ulmoides: Using high-concentration membrane separation and esterification technology, the crude extract of S1 was subjected to secondary extraction of dihydromyricetin and quercetin, so that the ratio of dihydromyricetin to quercetin in the Ampelopsis ulmoides extract was (25-80):1; S3, weighing 15-20 parts of the vine tea extract prepared in S2, 25-30 parts of gallic acid, 30-40 parts of yeast extract, and 20-30 parts of yeast immune polysaccharide by weight, and stirring and mixing them uniformly to form a premix; S4. Weigh 1-5 parts of the premix prepared in S3, add 10-50 parts of an acidifier, 0.5-5 parts of an enzyme preparation, and 1-10 parts of probiotics, and mix well. S5. Dry and store.

2. The method for preparing a feed additive containing rattan tea extract according to claim 1, characterized in that: In step S1, the dried leaves of the Ampelopsis ulmoides L. are first crushed and then dried at low temperature; then they are loaded into an extraction kettle, and an entrainer ethanol-water is injected into a CO2 stream through a high-pressure pump for supercritical CO2 extraction; then dihydromyricetin and quercetin are purified by molecular distillation, and quercetin is further enriched in the distillation residue phase by polyamide column chromatography (eluent: ethanol-water gradient).

3. The method for preparing a feed additive containing rattan tea extract according to claim 2, characterized in that: The entrainer has an ethanol:water ratio of 8:2, and the entrainment amount is 15-25% of the CO2 flow rate.

4. The method for preparing a feed additive containing rattan tea extract according to claim 3, characterized in that: In step S1, the extraction pressure is controlled at 30-45 MPa, the temperature is in the range of 50-60°C, the CO2 flow rate is 20-30 L / h, and the extraction time is 1.5-2.5 h.

5. The method for preparing a feed additive containing rattan tea extract according to claim 4, characterized in that: In step S1, the evaporation temperature of dihydromyricetin is 140-160° C., the evaporation temperature of quercetin is 180-220° C., and the pressure is 0.1-1 Pa.

6. The method for preparing a feed additive containing rattan tea extract according to claim 1, characterized in that: In step S2, dihydromyricetin and quercetin are first esterified by an esterifying agent, and then separated by a high-concentration membrane to obtain dihydromyricetin ester-enriched liquid quercetin and quercetin ester-enriched liquid, and then the dihydromyricetin ester-enriched liquid quercetin and quercetin ester-enriched liquid are deesterified by a deesterifying agent to obtain dihydromyricetin and quercetin.

7. The method for preparing a feed additive containing rattan tea extract according to claim 6, characterized in that: The esterifying agent of dihydromyricetin is acetic anhydride, and the deesterifying agent is methanol; the esterifying agent of quercetin is propionic anhydride, and the deesterifying agent is ethanol; and the high-concentration membrane is a molecular sieve membrane.

8. The method for preparing a feed additive containing rattan tea extract according to claim 1, characterized in that: The acidulant is a composite solid organic acid or a coated slow-release acidulant, the enzyme preparation is one or more of phytase, protease or amylase, and the probiotic is a lactobacillus / bifidobacterium or bacillus.

9. Use of the feed additive according to claim 1 in poultry feed, characterized in that: The total mass of the feed additive is 0.1 to 0.3‰ of the total mass of the poultry feed.

10. Use of the feed additive according to claim 1 in pig feed, characterized in that: The total mass of the feed additive is 0.3-0.6‰ of the total mass of the pig feed.

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