Chinese herbal medicine additive for delaying senescence in later egg laying period of laying hens

Abstract

The invention relates to the technical field of breeding, in particular to a senescence delaying type Chinese herbal medicine additive for laying hens in the later egg laying period. The matrix dispersed nano-micelle included traditional Chinese medicine compound extract is prepared from the following raw materials in parts by weight: extracts of eight traditional Chinese medicine raw materials and sodium cholate-chitosan quaternary ammonium salt ionic liquid, and the eight traditional Chinese medicine raw materials are as follows: motherwort, glossy privet fruit, astragalus membranaceus, angelica sinensis, dandelion, fructus lycii, poria cocos and hawthorn. According to the invention, an adjusting network formed by compatibility of the traditional Chinese medicine components realizes accurate release and synergistic effect in the body of the laying hen through efficient delivery of the ionic liquid nano-micelles: leonurine and astragaloside act together, so that the ovarian function decline speed is slowed down, and the laying rate is maintained at 82% or above; polysaccharide and flavonoid components synergistically enhance the oxidation resistance, the serum SOD activity is improved to 240 U / mL or above, the MDA content is reduced to 5.2 nmol / mL or below, and the aging process of an organism is delayed.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of breeding, in particular to a Chinese herbal medicine additive for delaying the decline of egg-laying hens in the later egg-laying period. BACKGROUND

[0002] In the breeding of egg-laying hens, the maintenance of production performance in the later egg-laying period (age ≥ 50 weeks) is a key link that determines the breeding benefit. With the increase of age, the ovarian function gradually declines, and the antioxidant capacity of the body decreases, and problems such as rapid decline of egg production rate (monthly decline of 5%-8%), deterioration of egg quality (thin and easy-to-break eggshell, light yellow egg yolk, and increased egg white fluidity) often occur.

[0003] To solve this problem, various technical solutions have been tried in the industry. Although western medicine additives can short-term improve egg production rate, there is a risk of drug residue. Chinese herbal medicine additives have become a research hotspot due to their natural and residue-free advantages, but there are two major problems: first, the extraction of effective components is incomplete, and traditional single solvent extraction can only obtain a small amount of components, missing key active substances such as polysaccharides, resulting in limited decline effect; second, the bioavailability is low, and traditional Chinese medicine extracts are easily degraded in the digestive tract of egg-laying hens and are difficult to be absorbed through the intestinal barrier, so a large dose needs to be added to see the effect in actual application, increasing the breeding cost.

[0004] Therefore, it is necessary to develop a decline-type additive that can realize complete extraction of effective components of traditional Chinese medicine, efficient delivery, and significant synergistic effect, which has become a problem that needs to be solved in the industry. SUMMARY

[0005] The present application aims to overcome the shortcomings of the prior art and provide a Chinese herbal medicine additive for delaying the decline of egg-laying hens in the later egg-laying period.

[0006] The specific technical solution is as follows: a Chinese herbal medicine additive for delaying the decline of egg-laying hens in the later egg-laying period, the additive is a matrix dispersed nano-micelle inclusion traditional Chinese medicine composite extract;

[0007] The preparation raw materials of the matrix dispersed nano-micelle inclusion traditional Chinese medicine composite extract include extracts of 8 kinds of traditional Chinese medicine raw materials and sodium cholate-chitosan quaternary ammonium salt ionic liquid, and the 8 kinds of traditional Chinese medicine raw materials are 35-50 parts of motherwort, 20-40 parts of privet fruit, 18-30 parts of astragalus, 10-20 parts of angelica, 10-24 parts of dandelion, 16-25 parts of medlar, 10-20 parts of poria cocos, and 7-15 parts of hawthorn.

[0008] The matrix dispersed nano-micelle inclusion traditional Chinese medicine composite extract is a nano-micelle with a particle size of 50-200 nm, which is prepared by spray drying after molecular self-assembly of the extracts of the 8 kinds of traditional Chinese medicine raw materials and sodium cholate-chitosan quaternary ammonium salt ionic liquid.

[0009] As a further technical solution, the preparation method of the matrix dispersed nanomicelle inclusion traditional Chinese medicine composite extract comprises the following steps:

[0010] Step one: preparation of the extract of eight traditional Chinese medicine raw materials: take eight traditional Chinese medicine raw materials by weight, ultrafine grinding, and prepare the traditional Chinese medicine composite extract concentrate by pH gradient extraction method:

[0011] Step two: nanomicelle inclusion:

[0012] The traditional Chinese medicine composite extract concentrate obtained in step one is subjected to microwave pretreatment of 300-400W for 6-10min; through the thermal effect and non-thermal effect of the microwave pretreatment, the intermolecular hydrogen bond combination of flavonoids, saponins and polysaccharide components in the traditional Chinese medicine composite extract with sodium cholate-chitosan quaternary ammonium salt ionic liquid is promoted, and compared with the process without microwave pretreatment, the inclusion rate of the final product is increased to more than 95%;

[0013] Then add sodium cholate-chitosan quaternary ammonium salt ionic liquid, and perform molecular self-assembly under the condition of 45℃, 200W ultrasonic and constant temperature stirring at 200-300r / min, and the assembly time is 1-2h;

[0014] Step three: molding:

[0015] The self-assembly liquid of step two is subjected to spray drying, and the inlet air temperature of spray drying is 180-200℃, the outlet air temperature is 80-90℃, and the feeding rate is 15-20mL / min, to obtain the matrix dispersed nanomicelle inclusion traditional Chinese medicine composite extract.

[0016] As a further technical solution, the eight traditional Chinese medicine raw materials are subjected to ultrafine grinding treatment, and the particle size of the ground material is ≤20μm.

[0017] As a further technical solution, the preparation method of the traditional Chinese medicine composite extract concentrate in step one is: mixing with 60% ethanol by volume fraction as solvent according to the solid-liquid ratio of 1:8-10, adjusting pH=4, and extracting twice at 60-65℃, each time for 1.5h, and combining to obtain flavonoids and saponins extract;

[0018] After extracting flavonoids and saponins, the residue is mixed with deionized water as solvent according to the solid-liquid ratio of 1:10-12, the pH is adjusted to 7, and the extraction is carried out twice at 80-85℃, each time for 2h, and the combined polysaccharide extract is obtained;

[0019] Combine the flavonoids and saponins extract and the polysaccharide extract, concentrate under the conditions of vacuum degree-0.08 to-0.09MPa and 55-65℃ to a relative density of 1.10-1.20 to obtain the traditional Chinese medicine composite extract concentrate.

[0020] As a further technical solution, the parameters of microwave pretreatment in step two are 350W and 8min, and the stirring speed of ultrasonic self-assembly is 250r / min, and the assembly time is 1.8h; in step one, the solid-liquid ratio of flavonoids and saponin extraction is 1:9, and the solid-liquid ratio of polysaccharide extraction is 1:11.

[0021] As a further technical solution, the preparation method of the sodium cholate-chitosan quaternary ammonium salt ionic liquid comprises the following steps:

[0022] S1: preparing chitosan quaternary ammonium salt: mixing chitosan and glycidyltrimethylammonium chloride at a mass ratio of 1:2-3, then adding deionized water at a solid-liquid ratio of 1:20, and stirring at 60℃ and 300r / min for 6h; after the reaction is completed, 3 times the volume of 95% ethanol solution is added, and the mixture is precipitated at 25℃ for 2h, and the precipitation is repeated for 3 times to remove the unreacted glycidyltrimethylammonium chloride, and the purified chitosan quaternary ammonium salt is obtained;

[0023] S2: preparing ionic liquid complex: adding the purified chitosan quaternary ammonium salt obtained in step S1 and sodium cholate at a molar ratio of 1:3 into 1-butyl-3-methylimidazolium tetrafluoroborate ionic liquid, and stirring at 400-500r / min for 2-3h;

[0024] S3: purification: the reaction liquid obtained in step S2 is purified by gradient pH dialysis method: a dialysis bag is selected, the initial pH of the dialysis liquid is 5.0, the pH of the dialysis liquid is increased by 0.5 units every 2 hours until the pH is 7.0, and the total dialysis time is 12h; after dialysis, the dialysis liquid is freeze-dried at-50 to-40℃ and a vacuum degree of ≤10Pa to obtain sodium cholate-chitosan quaternary ammonium salt ionic liquid.

[0025] As a further technical solution, the molecular weight cut-off of the dialysis bag in step S3 is 3500Da.

[0026] As a further technical solution, the dialysis liquid in step S3 is 0.01mol / L phosphate buffer.

[0027] As a further technical solution, the dialysis liquid in step S3 is replaced in full every 4 hours.

[0028] As a further technical solution, the use method of the additive is that the additive is mixed into the feed at an addition amount of 1-1.2% of the basic daily diet of laying hens; the late egg-laying stage is the stage when the laying hens are ≥50 weeks old.

[0029] Compared with the prior art, the application has the following beneficial effects:

[0030] The present application adopts eight kinds of traditional Chinese medicines such as Leonurus japonicus, and Ligustrum lucidum, and the components form a synergistic regulation system. The leonurus japonicus contains leonurus japonicus alkaline which can directly act on the granulosa cells of the hen ovary, promote cell proliferation and inhibit apoptosis, and improve the problem of reduced ovulation caused by ovarian aging; the astragalus saponins in astragalus can activate the body's antioxidant pathway, up-regulate the synthesis of superoxide dismutase (SOD), and the polysaccharide components of ligustrum lucidum and medlar can enhance the activity of immune cells, and the two can synergistically relieve the decline of immune function in the later stage of hens. The ferulic acid of angelica and the organic acid of hawthorn can promote intestinal peristalsis and improve the absorption efficiency of nutrients, providing sufficient nutritional support for the reproductive system of hens; the grifola frondosa polysaccharide can regulate the balance of intestinal flora, and the flavonoid components of dandelion can play an anti-inflammatory role and reduce the interference of chronic inflammation on egg production function. This multi-component combination avoids the problem of single component action limitation, and builds a complete body anti-aging network from four dimensions of ovary protection, antioxidant, immune regulation and nutrient absorption.

[0031] The pH gradient extraction method in the preparation process of the components of the present application solves the problem of low extraction rate of traditional extraction by two-step extraction strategy, first obtains flavonoids and saponin liposoluble components with acid ethanol, and then obtains polysaccharide water-soluble components with neutral water extraction, so that the utilization rate of active substances in traditional Chinese medicine is obviously improved. Microwave pretreatment destroys the polymerization structure of polysaccharide through energy input, avoids the agglomeration of extract, and lays a foundation for subsequent inclusion. The sodium cholate-chitosan quaternary ammonium salt ionic liquid carrier can self-assemble into 50-200nm nanomicelles with amphiphilic structure, the hydrophobic core wraps the liposoluble components, the hydrophilic shell improves the water solubility, and the biocompatibility of the ionic liquid itself enhances the intestinal mucosa permeability, so that the absorption rate of effective components is higher than that of ordinary chitosan carrier.

[0032] The regulation network formed by the combination of each traditional Chinese medicine component in the present application realizes precise release and synergistic effect in the body of hens through the efficient delivery of ionic liquid nanomicelles: the combined action of leonurus japonicus alkaline and astragalus saponins slows down the speed of ovarian function decline, and the egg production rate is maintained above 82%; the synergistic effect of polysaccharides and flavonoids enhances the antioxidant capacity, the serum SOD activity is increased to more than 240U / mL, and the MDA content is reduced to less than 5.2nmol / mL, which delays the aging process of the body. At the same time, the improvement of intestinal absorption efficiency reduces the daily ration. BRIEF DESCRIPTION OF DRAWINGS

[0033] Figure 1 The preparation method flow chart of the present application is shown in the figure. DETAILED DESCRIPTION

[0034] The technical solutions in the embodiments of the present application will be clearly and completely described below. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the protection scope of the present application.

[0035] The present application provides a Chinese herbal medicine additive for prolonging the egg-laying period of laying hens in the later stage of egg production. The additive is a substrate-dispersed nanomicelle-encapsulated traditional Chinese medicine (TCM) composite extract. The core is formed by gradient extraction of specific eight TCM raw materials, nanoself-assembly mediated by sodium cholate-chitosan quaternary ammonium salt ionic liquid, and spray drying, so as to realize efficient encapsulation and stable release of TCM effective components, and finally achieve the effects of prolonging the egg-laying period of laying hens in the later stage (≥50 weeks of age) and improving egg quality.

[0036] Raw materials and preparation include:

[0037] The TCM raw materials: Motherwort, Ligustrum lucidum, Astragalus, Angelica sinensis, Taraxacum mongolicum, Medlar, Poria cocos, and Crataegus pinnatifida are selected from dry medicinal materials meeting the standards of Chinese Pharmacopoeia (2020 Edition Part One), which can be purchased on the market and treated by removing impurities, washing, and drying before use.

[0038] Sodium cholate-chitosan quaternary ammonium salt ionic liquid: Its preparation needs to go through three steps of chitosan quaternization, ionic liquid compounding, and dialysis purification.

[0039] Extraction and molding auxiliary materials: 60% ethanol (analytical pure, China National Pharmaceutical Group), deionized water (laboratory self-made), 1-butyl-3-methylimidazolium tetrafluoroborate ionic liquid (purity ≥98%, Shanghai Maikelin), and phosphate buffer (0.01 mol / L, pH 5.0-7.0, laboratory preparation).

[0040] The equipment used includes: ultrafine grinder (XDW-600), constant temperature stirring extraction tank (TJG-50), microwave processor (MW-3000), ultrasonic stirrer (SCIENTZ-IID), spray dryer (SD-1000), and freeze dryer (FD-1A-50).

[0041] The following are specific embodiments:

[0042] Embodiment 1:

[0043] Preparation of sodium cholate-chitosan quaternary ammonium salt ionic liquid:

[0044] S1: Preparation of chitosan quaternary ammonium salt: 100 g of chitosan was mixed with 200 g of glycidyltrimethylammonium chloride (mass ratio 1:2), 2000 mL of deionized water was added according to the solid-liquid ratio of 1:20, and placed in a constant temperature stirring tank, stirred at 60°C and 300 r / min for 6 h. After the reaction was completed, 6000 mL of 95% ethanol solution was added, and the mixture was allowed to stand at 25°C for 2 h to precipitate, and the precipitation was repeated 3 times to remove the unreacted glycidyltrimethylammonium chloride. After vacuum drying, the purified chitosan quaternary ammonium salt was obtained.

[0045] S2: Preparation of ionic liquid complex: 50 g of purified chitosan quaternary ammonium salt was mixed with 82.5 g of sodium cholate (molar ratio 1:3), and 500 mL of 1-butyl-3-methylimidazolium tetrafluoroborate ionic liquid was added, and stirred at 400 r / min for 2 h until the system was completely homogeneous.

[0046] S3: Purification: A dialysis bag with a molecular weight cut-off of 3500 Da was selected, and the above reaction solution was loaded into the bag. 0.01 mol / L phosphate buffer was used as the dialysis solution (initial pH=5.0), and the pH of the dialysis solution was adjusted by 0.5 units every 2 hours (to 5.5, 6.0, 6.5, and 7.0, respectively). The total dialysis time was 12 h, and the dialysis solution was replaced every 4 h. After dialysis, the dialysis solution was dried in a freeze dryer at -50°C and a vacuum degree of ≤10 Pa to obtain sodium cholate-chitosan quaternary ammonium salt ionic liquid.

[0047] Preparation of extracts of 8 traditional Chinese medicine raw materials:

[0048] The traditional Chinese medicine raw materials were weighed according to the weight parts: Motherwort 40 parts, Ligustrum lucidum 30 parts, Astragalus 24 parts, Angelica 15 parts, Dandelion 17 parts, Wolfberry 20 parts, Poria cocos 15 parts, and Hawthorn 11 parts, with a total mass of 500 g.

[0049] The above traditional Chinese medicine raw materials were put into a supermicro grinder and ground to a particle size of ≤20 μm to obtain traditional Chinese medicine supermicro powder.

[0050] Flavonoids and saponins extraction: 4500 mL of 60% ethanol was added to the traditional Chinese medicine supermicro powder (solid-liquid ratio 1:9), the pH was adjusted to 4, and the mixture was stirred at 60°C for 2 times, each for 1.5 h. The two extraction solutions were combined and filtered to obtain the flavonoids and saponins extract.

[0051] Polysaccharide extraction: The filtered traditional Chinese medicine residue was taken and 5500 mL of deionized water was added (solid-liquid ratio 1:11), the pH was adjusted to 7, and the mixture was stirred at 85°C for 2 times, each for 2 h. The two extraction solutions were combined and filtered to obtain the polysaccharide extract.

[0052] Concentration: combine the flavonoids and saponin extract, polysaccharide extract, and place them in a vacuum concentration tank, concentrate under the condition of vacuum degree -0.08 MPa, 55℃ to relative density 1.15 (25℃), to obtain the concentrated extract of traditional Chinese medicine composite extract.

[0053] Nano-micelle inclusion:

[0054] Microwave pretreatment: place the concentrated extract of traditional Chinese medicine composite extract in a microwave processor, 350W for 8min, and cool to room temperature after treatment.

[0055] Molecular self-assembly: add the prepared sodium cholate-chitosan quaternary ammonium salt ionic liquid (addition amount is 5% of the mass of the concentrated extract) to the pretreated concentrated extract, and transfer it to an ultrasonic stirrer, under the condition of 45℃, 200W ultrasonic, constant temperature stirring at 250r / min for 1.8h, complete self-assembly, to obtain the self-assembly liquid.

[0056] Shaping:

[0057] The self-assembly liquid is sent into a spray dryer, the inlet air temperature is set to 190℃, the outlet air temperature is set to 85℃, and the feeding rate is set to 18mL / min, after spray drying, the matrix dispersed nano-micelle inclusion traditional Chinese medicine composite extract (particle size detection is 80-150nm) is obtained, which is the post-lay hen egg production period delaying type Chinese herbal medicine additive.

[0058] Method of use: the additive is mixed into the feed at an addition amount of 1% of the basal daily ration of the laying hen, and is fed to 50-week-old Roman brown-shelled laying hens.

[0059] Example 2:

[0060] Preparation of sodium cholate-chitosan quaternary ammonium salt ionic liquid:

[0061] S1: weigh 100g of chitosan and 300g of glycidyltrimethylammonium chloride (mass ratio 1:3), add 2000mL of deionized water according to the solid-liquid ratio of 1:20, stir at 60℃, 300r / min for 6h, and the subsequent ethanol precipitation and drying steps are the same as in Example 1.

[0062] S2: weigh 50g of purified chitosan quaternary ammonium salt and 82.5g of sodium cholate (molar ratio 1:3), add 500mL of 1-butyl-3-methylimidazolium tetrafluoroborate ionic liquid, stir at 500r / min for 3h, and the rest is the same as in Example 1.

[0063] S3: the dialysis parameters are the same as in Example 1, the freeze-drying conditions are -45℃, vacuum degree ≤10Pa, and the rest is the same as in Example 1.

[0064] Preparation of extracts of 8 kinds of traditional Chinese medicine raw materials:

[0065] Traditional Chinese medicine weight parts: motherwort 35 parts, female privet fruit 20 parts, radix astragali 18 parts, angelica 10 parts, dandelion 10 parts, medlar 16 parts, tuckahoe 10 parts, hawthorn 7 parts, total mass 300g.

[0066] Ultrafine grinding to particle size ≤20 μm, flavonoids and saponins extraction: add 2400 mL of 60% ethanol (solid-liquid ratio 1:8), pH=4, extract 2 times (1.5h each time) at 65℃; polysaccharide extraction: add 3000 mL of deionized water to the residue (solid-liquid ratio 1:10), pH=7, extract 2 times (2h each time) at 80℃.

[0067] Concentration: vacuum degree -0.09 MPa, 65℃, concentrate to relative density 1.10 (measured at 25℃), the rest is the same as example 1.

[0068] Nano-micelle inclusion:

[0069] Microwave pretreatment: 300W for 10min; self-assembly: 45℃, 200W ultrasonic, 200r / min stirring for 2h, the rest is the same as example 1.

[0070] Molding:

[0071] Spray drying parameters: inlet air temperature 180℃, outlet air temperature 80℃, feed rate 15mL / min, obtain nano-micelles (particle size 50-120nm), additive amount is 1% of the basic diet, feed 50-week-old laying hens.

[0072] Example 3:

[0073] Preparation of sodium cholate-chitosan quaternary ammonium salt ionic liquid: the steps are the same as example 1, only S2 stirring speed is 450r / min, stirring time is 2.5h, S3 freeze-drying temperature is -40℃.

[0074] Preparation of extracts of 8 traditional Chinese medicine raw materials:

[0075] Traditional Chinese medicine weight parts: motherwort 50 parts, female privet fruit 40 parts, radix astragali 30 parts, angelica 20 parts, dandelion 24 parts, medlar 25 parts, tuckahoe 20 parts, hawthorn 15 parts, total mass 264g.

[0076] Ultrafine grinding to particle size ≤20 μm, flavonoids and saponins extraction: add 2640 mL of 60% ethanol (solid-liquid ratio 1:10), pH=4, extract 2 times (1.5h each time) at 62℃; polysaccharide extraction: add 3168 mL of deionized water to the residue (solid-liquid ratio 1:12), pH=7, extract 2 times (2h each time) at 83℃.

[0077] Concentration: vacuum degree -0.085 MPa, 60℃, concentrate to relative density 1.20 (measured at 25℃), the rest is the same as example 1.

[0078] Nano-micelle inclusion:

[0079] Microwave pretreatment: 400W for 6min; self-assembly: 45℃, 200W ultrasonic, 300r / min stirring for 1h, the rest is the same as example 1.

[0080] Molding:

[0081] Spray drying parameters: inlet temperature 200℃, outlet temperature 90℃, feed rate 20mL / min, to obtain nano-micelles (particle size 120-200nm), the additive amount is 1.2% of the basic diet, and the 50-week-old laying hens are fed.

[0082] Comparative example 1: the difference from example 1 is that in step 3 "nano-micelle inclusion", no sodium cholate-chitosan quaternary ammonium salt ionic liquid is added, but an equal amount of ordinary chitosan (molecular weight 100,000 Da) is used instead, and the rest of the steps are exactly the same as example 1.

[0083] Comparative example 2: the difference from example 1 is that in step 2 "traditional Chinese medicine raw material weighing", motherwort is removed, and the rest of the 7 traditional Chinese medicines are adjusted according to the original proportion (jujube 35.3 parts, radix astragali 28.2 parts, angelica 17.6 parts, dandelion 20 parts, medlar 23.5 parts, tuckahoe 17.6 parts, haw 12.9 parts), the total mass is still 500g, and the rest of the steps are exactly the same as example 1.

[0084] Comparative example 3: the difference from example 1 is that in step 2 "preparation of traditional Chinese medicine extract", no pH gradient extraction is used, only 4500mL of 60% ethanol (solid-liquid ratio 1:9) is used for 2 times of extraction (60℃, 1.5h / time) on the traditional Chinese medicine ultra-fine powder, no polysaccharide is extracted, and the combined extract is concentrated to a relative density of 1.15, and the rest of the steps are exactly the same as example 1.

[0085] Test:

[0086] In order to verify the effect of the additive, tests were carried out, all 50-week-old healthy Roman brown shell laying hens (no significant difference in initial egg laying rate, P>0.05) were randomly divided into 7 groups (examples 1-3, comparative examples 1-3, blank control group), 30 in each group, the feeding period was 4 weeks, the blank control group was only fed with the basic diet, and the rest of the groups were fed with the same basic diet + corresponding additives (example 3 added amount 1.2%, the rest added amount 1%).

[0087] Test 1: determination of egg laying rate (core effect index of extension and decline);

[0088] Refer to NY / T3872-2021, record the total number of eggs of each group of laying hens (remove broken eggs, soft-shelled eggs) every day, calculate the average egg laying rate of each group every week, calculate the average egg laying rate for 4 weeks after the end of the test, formula: average egg laying rate (%) = total number of eggs / (number of chickens x feeding days) x 100%, the results are as follows:

[0089] Table 1

[0090] Group 4-week average egg laying rate (%) Example 1 85.2 Example 2 82.8 Example 3 86.5 Comparative Example 1 70.3 Comparative Example 2 75.6 Comparative Example 3 68.5 Blank control group 65.2

[0091] As can be seen from Table 1, the average egg laying rate of Examples 1-3 is significantly higher than that of Comparative Examples 1-3 and the blank control group (P<0.05), which shows that the additive of the present application can effectively prolong the egg laying period of laying hens in the post-egg laying period.

[0092] Test 2: Egg quality determination (eggshell strength, yolk color, albumen height);

[0093] Refer to GB / T34270-2017, at the end of the 4th week of the test, 20 fresh eggs (within 24h after laying) were randomly taken from each group, and the following methods were used for determination:

[0094] Eggshell strength: The eggshell strength tester (ETG-1063, Japan FHK) was used to determine the compression resistance of the longitudinal axis direction of the eggshell, with the unit of kg / cm²;

[0095] Yolk color: The Lohmann color fan (DSM, Netherlands) was used to compare under natural light, and the color fan grade (1-15 grade, the higher the grade, the darker the color) was recorded;

[0096] Albumen height: The albumen height tester (NZH-3000, Nanjing Mingao) was used to determine the distance between the midpoint of the yolk edge and the thick albumen edge, with the unit of mm, and the results are as follows:

[0097] Table 2

[0098] Group Eggshell strength (kg / cm 2 ])]] Egg yolk color (Roche grade) Egg white height (mm) Example 1 3.52 8.2 6.8 Example 2 3.35 7.8 6.5 Example 3 3.61 8.5 7.0 Comparative Example 1 2.83 6.3 5.2 Comparative Example 2 3.01 7.1 5.8 Comparative Example 3 2.75 6.0 5.0 Blank control group 2.58 5.8 4.8

[0099] As can be seen from Table 2, the egg quality indicators (eggshell strength, yolk color, albumen height) of Examples 1-3 are significantly better than those of other groups, which shows that the additive of the present application not only can delay the decline, but also can improve the egg quality, the additive can regulate the calcium metabolism of laying hens (enhance the formation of eggshell), promote the absorption of carotenoids (improve the yolk color), and increase the content of ovalbumin in egg white (increase the albumen height).

[0100] Test 3: Serum antioxidant index determination (SOD activity, MDA content);

[0101] Serum collection: At the end of the 4th week of the test, 10 laying hens were randomly taken from each group, 5mL of blood was collected from the wing vein, centrifuged at 3000r / min for 10min, and the upper serum was taken for standby;

[0102] SOD activity determination: according to NY / T1332-2007, the SOD activity (unit: U / mL) was calculated by measuring the absorbance at 420 nm with a UV spectrophotometer (UV-2600, Shimadzu);

[0103] MDA content determination: according to GB / T5009.181-2017, the MDA content (unit: nmol / mL) was calculated by measuring the absorbance at 532 nm with the thiobarbituric acid (TBA) method, and the results were as follows:

[0104] Table 3

[0105] Group Serum SOD activity (U / mL) Serum MDA content (nmol / mL) Example 1 252.3 4.8 Example 2 241.5 5.2 Example 3 258.7 4.5 Comparative Example 1 201.8 7.9 Comparative Example 2 218.6 6.8 Comparative Example 3 195.2 8.5 Blank control group 182.5 9.2

[0106] As can be seen from Table 3, the serum SOD activity of Examples 1-3 was significantly higher than that of other groups, and the MDA content was significantly lower than that of other groups, indicating that the additive of the application can realize the delay of aging by enhancing the antioxidant capacity of laying hens, SOD is a key antioxidant enzyme and can remove free radicals; MDA is a lipid oxidation product, and the lower the content, the lighter the oxidative damage.

[0107] The preferred embodiments of the application disclosed above are only used to help illustrate the application. The preferred embodiments do not describe all the details, nor limit the application to the specific embodiments described. Obviously, many modifications and variations can be made according to the content of the present application.

Claims

1. A Chinese herbal medicine additive for prolonging the laying period of hens, characterized in that, The additive is a matrix dispersed nanomicelle-encapsulated traditional Chinese medicine composite extract; The preparation raw material of the matrix dispersed nanomicelle-encapsulated traditional Chinese medicine composite extract comprises extracts of eight traditional Chinese medicine raw materials and sodium cholate-chitosan quaternary ammonium salt ionic liquid, and the eight traditional Chinese medicine raw materials are as follows in terms of weight parts: motherwort 35-50 parts, privet fruit 20-40 parts, radix astragali 18-30 parts, angelica 10-20 parts, dandelion 10-24 parts, medlar 16-25 parts, poria 10-20 parts, and hawthorn 7-15 parts. The matrix dispersed nanomicelle-encapsulated traditional Chinese medicine composite extract is a nanomicelle with a particle size of 50-200 nm, which is prepared by spray drying of the extracts of the eight traditional Chinese medicine raw materials and sodium cholate-chitosan quaternary ammonium salt ionic liquid after molecular self-assembly.

2. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 1, characterized in that, The preparation method of the matrix dispersed nanomicelle-encapsulated traditional Chinese medicine composite extract comprises the following steps: Step one: preparation of extracts of eight traditional Chinese medicine raw materials: take eight traditional Chinese medicine raw materials in terms of weight parts, perform ultrafine grinding, and prepare traditional Chinese medicine composite extract concentrate by using pH gradient extraction method: Step two: nanomicelle encapsulation: perform microwave pretreatment on the traditional Chinese medicine composite extract concentrate obtained in step one at 300-400 W for 6-10 min; then add sodium cholate-chitosan quaternary ammonium salt ionic liquid, perform molecular self-assembly under the condition of ultrasonic at 45°C and 200 W, and perform constant temperature stirring at 200-300 r / min, and the assembly time is 1-2 h; Step three: molding: perform spray drying on the self-assembly liquid of step two, the inlet air temperature of spray drying is 180-200°C, the outlet air temperature is 80-90°C, and the feeding rate is 15-20 mL / min, to obtain the matrix dispersed nanomicelle-encapsulated traditional Chinese medicine composite extract.

3. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 2, characterized in that, The eight traditional Chinese medicine raw materials are subjected to ultrafine grinding treatment, and the particle size of the ground material is ≤20 μm.

4. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 2, characterized in that, The preparation method of the traditional Chinese medicine composite extract concentrate in step one is as follows: mix with 60% ethanol by volume fraction as a solvent according to a solid-liquid ratio of 1:8-10, adjust pH to 4, and extract at 60-65°C for 2 times, each for 1.5 h, to obtain flavone and saponin extract liquid; after the extraction of flavone and saponin, mix the residue with deionized water as a solvent according to a solid-liquid ratio of 1:10-12, adjust pH to 7, and extract at 80-85°C for 2 times, each for 2 h, to obtain polysaccharide extract liquid; combine the flavone and saponin extract liquid and the polysaccharide extract liquid, concentrate under the condition of a vacuum degree of-0.08 to-0.09 MPa and 55-65°C to a relative density of 1.10-1.20, to obtain the traditional Chinese medicine composite extract concentrate.

5. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 2, characterized in that, The parameters of microwave pretreatment in step two are 350 W and 8 min, and the stirring speed of ultrasonic self-assembly is 250 r / min, and the assembly time is 1.8 h; the solid-liquid ratio of flavone and saponin extraction in step one is 1:9, and the solid-liquid ratio of polysaccharide extraction is 1:

11.

6. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 2, characterized in that, The preparation method of the sodium cholate-chitosan quaternary ammonium salt ionic liquid is characterized by comprising the following steps: S1: preparing chitosan quaternary ammonium salt: chitosan and glycidyltrimethylammonium chloride are mixed at a mass ratio of 1:2-3, then deionized water is added at a solid-liquid ratio of 1:20, and the mixture is stirred at 300 r / min at 60 DEG C for 6 h; after the reaction is completed, 3 times the volume of 95% ethanol solution is added, and the mixture is precipitated at 25 DEG C for 2 h, and the precipitation is repeated 3 times to remove unreacted glycidyltrimethylammonium chloride, and the purified chitosan quaternary ammonium salt is obtained; S2: preparing ionic liquid complex: the purified chitosan quaternary ammonium salt obtained in step S1 is mixed with sodium cholate at a molar ratio of 1:3, and then 1-butyl-3-methylimidazolium tetrafluoroborate ionic liquid is added, and the mixture is stirred at 400-500 r / min for 2-3 h; S3: purification: the reaction solution obtained in step S2 is purified by gradient pH dialysis: a dialysis bag is selected, the initial pH of the dialysis solution is 5.0, the pH of the dialysis solution is increased by 0.5 units every 2 h until the pH is 7.0, and the total dialysis time is 12 h; after dialysis, the dialysis solution is freeze-dried at -50 to -40 DEG C under a vacuum degree of less than or equal to 10 Pa to obtain sodium cholate-chitosan quaternary ammonium salt ionic liquid.

7. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 6, characterized in that, The molecular weight cut-off of the dialysis bag in step S3 is 3500 Da.

8. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 6, characterized in that, The dialysis solution in step S3 is 0.01 mol / L phosphate buffer.

9. The Chinese herbal additive for prolonging the laying period of laying hens according to claim 6, characterized in that, The dialysis solution in step S3 is replaced with a full amount every 4 h.

10. The Chinese herbal additive for the late stage of laying hens according to claim 6, characterized in that, The use method of the additive is: the additive is mixed into the feed at an addition amount of 1-1.2% of the basic daily diet of laying hens; the late egg production stage of laying hens is the stage when the laying hens are greater than or equal to 50 weeks old.

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