Traditional Chinese medicine feed additive for improving animal immunity and preparation method thereof

Abstract

The invention provides a traditional Chinese medicine feed additive for improving animal immunity and a preparation method thereof, and relates to the technical field of traditional Chinese medicine feed additives. The traditional Chinese medicine feed additive is prepared from twelve traditional Chinese medicines including astragalus membranaceus, immature bitter orange, fructus forsythiae, black beans, semen cuscutae, radix paeoniae alba, rosemary, thyme, litsea cubeba, mangosteen peel, cinnamon and origanum vulgare. The traditional Chinese medicine feed additive disclosed by the invention can be used for improving the immune organ index of an immunosuppressed mouse induced by cyclophosphamide, regulating the level of immune protein and inflammation-related factors and enhancing the phagocytic ability of macrophages of the mouse; the antibody titer of the Newcastle disease virus can be improved, the immune organ index of the chicken can be improved, and the secretion of IL-4, TNF-alpha and IFN-gamma in the chicken body can be promoted; mouse and broiler chicken tests show that the traditional Chinese medicine feed additive disclosed by the invention can improve the immune response capability of animals and enhance the immunity of the animals, and has a good application prospect in animal breeding.

Description

Technical Field

[0001] The invention relates to the technical field of Chinese medicinal feed additives, and in particular to a Chinese medicinal feed additive for improving animal immunity and a preparation method thereof. Background Art

[0002] In recent years, it is common in animal breeding that infectious diseases still occur after vaccination, and the dosage of drugs is gradually increased but the ideal treatment effect is not achieved. This is caused by farmers' long-term abuse of chemical drugs and unreasonable vaccination. To effectively solve this problem, the fundamental solution is to activate the immune system of the animal body, increase the formation rate of immune cells, the conversion rate of peripheral blood lymphocytes and the phagocytic ability of macrophages, so that animals can obtain a higher level of response after vaccination.

[0003] At present, there are many kinds of substances that can improve animal immunity and have immune-enhancing effects, and the commonly used ones are propolis, Freund's adjuvant, vitamin E, levamisole, transfer factor and interferon, etc. Although these immunoenhancers have played a certain role in the development of animal husbandry, due to their complex preparation, high price, and some even have side effects on the body, breeders are committed to seeking new alternative products.

[0004] Chinese herbal medicines are derived from nature, are rich in resources and inexpensive, and contain a variety of active substances and nutrients. They have antibacterial, anti-inflammatory, and antiviral effects. Unlike Western medicines, Chinese herbal medicines have little residue after their effects in the body, are less likely to cause pathogens to develop drug resistance, and have fewer toxic side effects on the body. Currently, many studies have shown that Chinese herbal medicines can mobilize the body's anti-stress ability by adjusting the body's immune system, thereby improving the body's immune function and defense function. Therefore, the development and utilization of Chinese herbal medicines to improve animal immunity has broad prospects. Summary of the invention

[0005] 1. Technical issues to be resolved

[0006] In view of the deficiencies in the prior art, the present invention provides a Chinese medicinal feed additive for improving animal immunity and a preparation method thereof.

[0007] (II) Technical solution

[0008] To achieve the above objectives, the present invention is implemented through the following technical solutions:

[0009] Firstly, the present invention provides a traditional Chinese medicine feed additive for improving animal immunity. The traditional Chinese medicine feed additive is composed of the following raw materials in parts by weight: 10-30 parts of astragalus, 10-30 parts of immature bitter orange, 10-30 parts of forsythia, 5-20 parts of black beans, 5-20 parts of dodder seeds, 5-20 parts of white peony roots, 5-20 parts of rosemary, 5-20 parts of thyme, 5-20 parts of litsea cubeba, 5-20 parts of mangosteen peel, 5-20 parts of cinnamon, and 5-20 parts of oregano.

[0010] Specifically, the Chinese medicinal feed additive is composed of the following raw materials in parts by weight: 15-25 parts of astragalus, 15-25 parts of immature bitter orange, 10-20 parts of forsythia, 5-15 parts of black beans, 10-20 parts of dodder seeds, 5-15 parts of white peony roots, 5-15 parts of rosemary, 5-15 parts of thyme, 10-20 parts of litsea cubeba, 10-20 parts of mangosteen peel, 5-15 parts of cinnamon, and 5-15 parts of oregano.

[0011] Specifically, the Chinese medicinal feed additive is composed of the following raw materials in parts by weight: 22 parts of astragalus, 20 parts of immature bitter orange, 16 parts of forsythia, 12 parts of black beans, 16 parts of dodder seeds, 10 parts of white peony, 8 parts of rosemary, 8 parts of thyme, 10 parts of litsea cubeba, 14 parts of mangosteen peel, 10 parts of cinnamon, and 6 parts of oregano.

[0012] On the other hand, the present invention also provides a method for preparing a Chinese medicinal feed additive for improving animal immunity. The Chinese medicinal feed additive is prepared by weighing each raw material according to the above-mentioned portion, washing, drying, and then ultra-finely breaking the wall to obtain the Chinese medicinal feed additive.

[0013] The pharmacological effects of each Chinese medicinal ingredient in the Chinese medicinal feed additive of the present invention are as follows:

[0014] Astragalus: sweet in taste, slightly warm in nature, enters the spleen and lung meridians; has the effects of replenishing qi and strengthening the exterior, expelling toxins and pus, promoting diuresis, and promoting tissue regeneration; used for symptoms such as qi deficiency and fatigue, long-term diarrhea and prolapse of the anus, spontaneous sweating, edema, uterine prolapse, proteinuria of chronic nephritis, diabetes, and non-healing wounds; Astragalus contains effective ingredients such as carbohydrates, multiple amino acids, proteins, choline, betaine, folic acid, vitamin P, amylase, and multiple trace elements, which have the effects of enhancing immunity and anti-virus.

[0015] Citrus aurantium: tastes bitter, pungent, and sour, and is slightly cold in nature; it enters the spleen and stomach meridians; it has the effects of breaking up qi and eliminating accumulation, resolving phlegm and dispersing abdominal distension; it is pungent in nature and bitter in nature, and is strong in nature, and is good at moving the qi of the middle burner; modern research has shown that it has the functions of regulating gastrointestinal smooth muscle, anti-ulcer, bile-promoting, raising blood pressure, strengthening the heart, relieving pain, and regulating uterine contraction.

[0016] Forsythia: bitter in taste, slightly cold in nature, enters the lung, heart, and small intestine meridians; has the effects of clearing away heat and detoxifying, reducing swelling and dispersing nodules; used to treat wind-heat colds, the initial stage of febrile diseases, high fever, thirst, coma and rash; Forsythia contains chemical components such as lignans, flavonoids, volatile components, phenylacetylene, ethylcyclohexanols, triterpenes and coumarins. Modern pharmacological studies have shown that Forsythia has antioxidant, anti-inflammatory, antiviral, phosphodiesterase inhibitory effects, and elastase activity inhibitory effects.

[0017] Black beans: sweet in taste, neutral in nature, enter the spleen and kidney meridians; have the effects of improving essence and eyesight, nourishing blood and dispelling wind, promoting diuresis and detoxification; can be used for yin deficiency and thirst, dizziness, physical weakness and sweating, kidney deficiency and low back pain, edema and oliguria; black beans contain polyphenols such as anthocyanins and isoflavones, which have multiple pharmacological effects such as anti-oxidation, regulating hormone levels, beauty and hair care, cancer prevention and anti-cancer.

[0018] Cuscuta: It tastes spicy and sweet, is neutral in nature, and enters the liver, kidney, and spleen meridians; it has the effects of nourishing the kidney and replenishing essence, nourishing the liver and improving eyesight; it is used to treat kidney deficiency and low back pain, impotence, premature ejaculation, liver and kidney deficiency and other symptoms; Cuscuta mainly contains chemical components such as flavonoids, polysaccharides, alkaloids, volatile oils, amino acids, trace elements, etc., which have pharmacological effects on the reproductive system, bone metabolism, anti-aging, immune system and many other aspects.

[0019] White peony root: bitter and sour in taste, slightly cold in nature, enters the liver and spleen meridians; has the effects of calming the liver and relieving pain, nourishing blood and regulating menstruation, astringing yin and stopping sweating; used for symptoms such as headache and dizziness, chest and flank pain, spontaneous sweating, night sweats, yin deficiency and fever; the chemical components in white peony root are mainly volatile oils, monoterpenes, triterpenes and flavonoid compounds. Monoterpenes represented by paeoniflorin and paeoniflorin and their glycosides are recognized as the active substances in white peony root, and its pharmacological effects are extensive, with anti-inflammatory and analgesic, liver protection, improvement of osteoarthritis and other effects.

[0020] Rosemary: It has the effects of sweating, strengthening the spleen, calming the nerves and relieving pain. It is used to treat headaches, prevent hair loss and other symptoms. Modern pharmacological research shows that rosemary has antioxidant, anti-tumor, antibacterial, anti-inflammatory and liver protection effects.

[0021] Thyme: spicy, slightly warm in nature; has the effects of dispelling wind and relieving exterior symptoms, promoting qi and relieving pain, relieving cough, and lowering blood pressure; used for colds, coughs, headaches, toothaches, indigestion, acute gastroenteritis, hypertension, etc.; modern pharmacological studies have shown that thyme has antioxidant, anti-thrombotic, antibacterial, and antidepressant effects.

[0022] Litsea cubeba: pungent, bitter, warm in nature; has the effects of dispelling wind and cold, regulating qi and relieving pain; root: used for stomach cold and vomiting, cold pain in the abdomen, abdominal pain due to cold hernia, cold and dampness stagnation, turbid urine; leaves: used externally to treat carbuncle, swelling and pain, mastitis, insect and snake bites, and prevent mosquito bites; seeds: used for colds, headaches, indigestion, and stomachaches.

[0023] Mangosteen peel: It tastes bitter and astringent, and is cold in nature. It has the effect of clearing away heat and detoxifying. The peel contains tannic acid, which can precipitate with microbial protein, causing the protein to coagulate and help the body kill bacteria. Mangosteen peel has antioxidant, antibacterial, anti-inflammatory, anti-tumor and immune-enhancing pharmacological effects.

[0024] Cinnamon: It tastes spicy and sweet, is very hot in nature, and enters the kidney, spleen, heart, and liver meridians; it has the effects of warming the middle and dispelling cold, regulating qi and relieving pain; it is used for cold pain in the abdomen, vomiting and diarrhea, soreness and coldness of the waist and knees, damp-cold arthralgia, blood stasis and dysmenorrhea, etc.; cinnamon mainly contains volatile oils, polyphenols, flavonoids, lignans, polysaccharides and other ingredients, which have anti-tumor, antibacterial, anti-inflammatory, antioxidant, anti-allergic, hypoglycemic and blood lipid lowering and other pharmacological effects.

[0025] Oregano: spicy, slightly bitter, cool in nature; has anti-microbial, antispasmodic, immune-enhancing, sedative, antioxidant, and diuretic effects.

[0026] (III) Beneficial effects

[0027] The invention provides a Chinese medicinal feed additive for improving animal immunity and a preparation method thereof. The Chinese medicinal feed additive is prepared from astragalus, immature bitter orange, forsythia, black bean, dodder seed, white peony root, rosemary, thyme, litsea cubeba, mangosteen peel, cinnamon bark and oregano as raw materials. In the formula, astragalus and immature bitter orange are used as monarch drugs, mainly exerting the effects of invigorating qi and strengthening the exterior, and enhancing immunity; forsythia, black bean and dodder seed are used as assistant drugs, assisting the monarch drugs in playing the effects of clearing away heat and detoxifying, invigorating the kidney and replenishing yin, and strengthening physical fitness; white peony root, rosemary, thyme, litsea cubeba and mangosteen peel are used as adjuvant drugs, harmonizing the medicinal properties, dispelling wind and dampness, promoting blood circulation and relieving pain, and clearing away heat and detoxifying; cinnamon bark and oregano are used as guiding drugs, guiding the medicinal effects, enhancing the effects of warming the middle and dispersing cold, clearing away heat and detoxifying, and harmonizing the drugs to achieve synergistic enhancement.

[0028] The Chinese medicinal feed additive of the present invention has the following advantages:

[0029] (1) The Chinese medicinal feed additive of the present invention can improve the immune organ index of cyclophosphamide-induced immunosuppressive mice, regulate the levels of immune proteins and inflammatory related factors, enhance the ability of mice to clear carbon particles, and enhance the phagocytic ability of macrophages.

[0030] (2) The Chinese medicinal feed additive of the present invention can increase the antibody titer of Newcastle disease virus in chickens, improve the immune organ index of chickens, and promote the secretion of IL-4, TNF-α and IFN-γ in chickens.

[0031] (3) The Chinese medicinal feed additive of the present invention can improve the immune response ability of animals and enhance the immunity of animals. The effect is better than that of the positive drug levamisole, and it can reduce the residual amount of the drug in the animal body. It is green and healthy and has good application prospects in animal breeding. DETAILED DESCRIPTION

[0032] In order to make the purpose, technical solution and advantages of the embodiments of the present invention clearer, the technical solution in the embodiments of the present invention will be clearly and completely described below in combination with the embodiments of the present invention. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0033] Example 1

[0034] A Chinese medicinal feed additive for improving animal immunity is prepared by weighing the following raw materials in parts by weight: 10 parts of astragalus, 10 parts of immature bitter orange, 10 parts of forsythia, 5 parts of black beans, 5 parts of dodder seeds, 5 parts of white peony, 5 parts of rosemary, 5 parts of thyme, 5 parts of litsea cubeba, 5 parts of mangosteen peel, 5 parts of cinnamon bark and 5 parts of oregano; washing and drying the raw materials and then performing ultrafine wall breaking and pulverization to obtain the Chinese medicinal feed additive.

[0035] Example 2

[0036] A Chinese medicinal feed additive for improving animal immunity comprises the following raw materials weighed according to the following weight proportions: 30 parts of astragalus, 30 parts of immature bitter orange, 30 parts of forsythia, 20 parts of black bean, 20 parts of dodder seed, 20 parts of white peony root, 20 parts of rosemary, 20 parts of thyme, 20 parts of litsea cubeba, 20 parts of mangosteen peel, 20 parts of cinnamon bark and 20 parts of oregano; the raw materials are washed and dried, and then subjected to ultrafine wall-breaking and pulverization to obtain the Chinese medicinal feed additive.

[0037] Example 3

[0038] A Chinese medicinal feed additive for improving animal immunity is prepared by weighing the following raw materials in parts by weight: 14 parts of astragalus, 12 parts of immature bitter orange, 14 parts of forsythia, 8 parts of black beans, 7 parts of dodder seeds, 6 parts of white peony, 5 parts of rosemary, 70 parts of thyme, 8 parts of litsea cubeba, 10 parts of mangosteen peel, 8 parts of cinnamon bark and 6 parts of oregano; washing, drying and then performing ultra-micro wall-breaking and pulverizing to obtain the Chinese medicinal feed additive.

[0039] Example 4

[0040] A Chinese medicinal feed additive for improving animal immunity comprises the following raw materials weighed according to the following weight proportions: 28 parts of astragalus, 26 parts of immature bitter orange, 25 parts of forsythia, 16 parts of black bean, 18 parts of dodder seed, 13 parts of white peony root, 11 parts of rosemary, 12 parts of thyme, 18 parts of litsea cubeba, 16 parts of mangosteen peel, 15 parts of cinnamon bark and 14 parts of oregano; the raw materials are washed, dried and then subjected to ultrafine wall-breaking and pulverization to obtain the Chinese medicinal feed additive.

[0041] Example 5

[0042] A Chinese medicinal feed additive for improving animal immunity is prepared by weighing the following raw materials in parts by weight: 15 parts of astragalus, 15 parts of immature bitter orange, 10 parts of forsythia, 5 parts of black beans, 10 parts of dodder seeds, 5 parts of white peony, 5 parts of rosemary, 5 parts of thyme, 10 parts of litsea cubeba, 10 parts of mangosteen peel, 5 parts of cinnamon bark and 5 parts of oregano; washing and drying the raw materials and then performing ultrafine wall breaking and pulverization to obtain the Chinese medicinal feed additive.

[0043] Example 6

[0044] A Chinese medicinal feed additive for improving animal immunity is prepared by weighing the following raw materials in parts by weight: 20 parts of astragalus, 20 parts of immature bitter orange, 14 parts of forsythia, 10 parts of black beans, 16 parts of dodder seeds, 12 parts of white peony, 8 parts of rosemary, 10 parts of thyme, 16 parts of litsea cubeba, 14 parts of mangosteen peel, 8 parts of cinnamon bark and 12 parts of oregano; washing and drying the raw materials and then performing ultrafine wall-breaking and pulverizing to obtain the Chinese medicinal feed additive.

[0045] Example 7

[0046] A Chinese medicinal feed additive for improving animal immunity comprises the following raw materials weighed according to the following weight proportions: 25 parts of astragalus, 25 parts of immature bitter orange, 20 parts of forsythia, 15 parts of black bean, 20 parts of dodder seed, 15 parts of white peony root, 15 parts of rosemary, 15 parts of thyme, 20 parts of litsea cubeba, 20 parts of mangosteen peel, 15 parts of cinnamon bark and 15 parts of oregano; the raw materials are washed and dried, and then subjected to ultrafine wall-breaking and pulverization to obtain the Chinese medicinal feed additive.

[0047] Example 8

[0048] A Chinese medicinal feed additive for improving animal immunity comprises the following raw materials weighed according to the following weight proportions: 18 parts of astragalus, 20 parts of immature bitter orange, 14 parts of forsythia, 9 parts of black beans, 12 parts of dodder seeds, 8 parts of white peony, 6 parts of rosemary, 5 parts of thyme, 12 parts of litsea cubeba, 13 parts of mangosteen peel, 6 parts of cinnamon bark and 7 parts of oregano; the raw materials are washed and dried, and then subjected to ultra-micro wall-breaking and pulverization to obtain the Chinese medicinal feed additive.

[0049] Example 9

[0050] A Chinese medicinal feed additive for improving animal immunity is prepared by weighing the following raw materials in parts by weight: 22 parts of astragalus, 20 parts of immature bitter orange, 16 parts of forsythia, 12 parts of black beans, 16 parts of dodder seeds, 10 parts of white peony, 8 parts of rosemary, 8 parts of thyme, 10 parts of litsea cubeba, 14 parts of mangosteen peel, 10 parts of cinnamon bark and 6 parts of oregano; washing and drying the raw materials and then performing ultrafine wall-breaking and pulverizing to obtain the Chinese medicinal feed additive.

[0051] Test Example 1

[0052] Immunoenhancing effect of the Chinese medicinal feed additive of the present invention on mice

[0053] 1 Experimental animals

[0054] Healthy adult SPF female Kunming mice with a body weight of (20±2) g were selected and kept in a sterile animal room (temperature: 23±2°C, humidity: 50%±5%, 12h light-dark cycle). All animals were free to take SPF drinking water and feed.

[0055] 2 Test substances

[0056] (1) 22 parts of Astragalus, 20 parts of Citrus aurantium, 16 parts of Forsythia, 12 parts of Black Bean, 16 parts of Cuscuta, 10 parts of White Peony, 8 parts of Rosemary, 8 parts of Thyme, 10 parts of Litsea cubeba, 14 parts of Mangosteen, 10 parts of Cinnamon, and 6 parts of Oregano;

[0057] (2) 20 parts of Citrus aurantium, 16 parts of Fructus Forsythiae, 12 parts of Black Bean, 16 parts of Cuscuta australis, 10 parts of White Peony Root, 8 parts of Rosemary, 8 parts of Thyme, 10 parts of Cinnamon Bark, and 6 parts of Oregano;

[0058] (3) 22 parts of Astragalus, 16 parts of Forsythia, 12 parts of Black Bean, 16 parts of Cuscuta, 10 parts of White Peony, 10 parts of Litsea cubeba, 14 parts of Mangosteen, 10 parts of Cinnamon, and 6 parts of Oregano;

[0059] (4) 22 parts of Astragalus, 20 parts of Citrus aurantium, 16 parts of Cuscuta australis, 10 parts of White Peony Root, 8 parts of Rosemary, 8 parts of Thyme, 10 parts of Litsea cubeba, 14 parts of Mangosteen Peel, and 10 parts of Cinnamon Bark;

[0060] (5) 22 parts of Astragalus, 20 parts of Citrus aurantium, 16 parts of Forsythia, 12 parts of Black Bean, 8 parts of Rosemary, 8 parts of Thyme, 10 parts of Litsea cubeba, 14 parts of Mangosteen Peel, and 6 parts of Oregano;

[0061] Weigh the raw materials according to the weight proportions in the above-mentioned formulas, wash and dry them, and then perform ultrafine wall-breaking pulverization to obtain test substances 1 to 5. Prepare test substances 1 to 5 into 1 g / mL solutions with 100°C deionized water, cool them to below room temperature, and use them immediately after preparation. Stir them evenly before administration.

[0062] 3 Animal grouping and drug treatment

[0063] After 140 mice were adaptively fed for 7 days, 20 healthy mice were randomly selected as the blank control group and continued to be fed. The remaining 120 mice were injected with cyclophosphamide (60 mg / kg) for 3 consecutive days, and the food and water intake were maintained at normal levels to establish an immune-impaired model. Mice with symptoms such as body weight loss, large hair loss, and lethargy were used as the standard for successful modeling. After the modeling was successful, the 120 mice were randomly divided into a model control group and test groups 1-5, with 20 mice in each group. Test groups 1-5 were gavaged with solutions of test substances 1-5, respectively, with a dose of 0.2 mL / kg; the blank control group and the model control group were gavaged with an equal volume of normal saline; each group was gavaged once a day for 30 consecutive days.

[0064] 4 Observation indicators

[0065] 4.1 Body weight and immune organ index

[0066] At the end of the experiment, the mice in each group were weighed, and the mice were killed by cervical dislocation. The thymus and spleen were weighed and the immune index was calculated.

[0067] Immune index = thymus or spleen weight (mg) / body weight (g)

[0068] 4.2 Determination of immune proteins and inflammatory factors

[0069] Blood was collected from the eye sockets of mice in each group, left at room temperature until the upper serum was precipitated, and centrifuged at 4°C and 3000 r / min for 10 min to obtain serum. According to the operating procedures in the kit manual, the levels of immune proteins IgA, IgG, IgM and inflammatory factors IL-4, IFN-γ, and TNF-α were detected.

[0070] 4.3 Carbon clearance experiment in mice

[0071] 24h after the last administration of each group of mice, 0.1mL / 10g India ink (diluted 4 times with saline) was injected into the tail vein of the mice, and the time was immediately started. 20μL of blood was collected from the medial angular venous plexus at 2min (t1) and 10min (t2), respectively, and mixed with 2mL of 0.1% sodium carbonate solution by mass fraction, and the OD value was measured at a wavelength of 600nm. 2mL of 0.1% sodium carbonate solution by mass fraction was taken as blank. After blood collection, the mice were killed by cervical dislocation, and the liver and spleen of the mice were dissected and weighed, and the phagocytic index α was calculated.

[0072]

[0073] Where K represents the phagocytic rate; W0 represents the weight of the mouse; W L W represents mouse liver weight; S represents the mouse spleen weight.

[0074] 4.4 Phagocytosis of chicken red blood cells by mouse peritoneal macrophages

[0075] After the last administration, mice were intraperitoneally injected with 0.2 mL of 2% packed sheep red blood cells to activate macrophages. Four days later, mice were killed by cervical dislocation, and 4 mL of Hank's solution plus calf serum was intraperitoneally injected. The abdomen was gently massaged, the abdominal wall was cut, 0.5 mL of peritoneal wash fluid was drawn and added to a test tube containing 0.5 mL of 1% chicken red blood cell suspension, and mixed. 0.5 mL of the mixed solution was drawn and added to the agar circle of the slide, and placed in an incubator at 37 ° C for 20 minutes. After the end, it was quickly rinsed with saline and dried, fixed in methanol solution, stained with Giemsa stain, rinsed with distilled water, observed under a microscope, and the phagocytic rate and phagocytic index were calculated.

[0076] P=N1 / N0Equation 3I=N2 / N0Equation 4

[0077] Wherein, P represents the phagocytic rate, %; N1 represents the number of macrophages that phagocytize chicken blood red blood cells; N0 represents the total number of macrophages counted; I represents the phagocytic index; N2 represents the total number of chicken blood red blood cells phagocytized.

[0078] 5 Statistical analysis

[0079] Statistical analysis was performed using GraphPad Prism 8.0. ± standard deviation (s), one-way analysis of variance was used for comparison among the groups, and the results were considered to be significantly different when P < 0.05.

[0080] 6 Results

[0081] 6.1 Effects of each experimental group on mouse body weight and immune organ indexes

[0082] As shown in Table 1, compared with the model control group, there was no significant difference in the weight gain of mice (P>0.05), indicating that the Chinese medicine feed additive of the present invention had no significant effect on the weight of mice. As shown in Table 2, compared with the blank control group, the thymus index and spleen index of the mice in the model control group were significantly decreased (P<0.01); compared with the model control group, the thymus index and spleen index of the mice in the test group 1 and the test groups 4-5 were significantly increased (P<0.05), while the thymus index and spleen index of the mice in the test groups 2-3 were increased but without significant difference, indicating that the Chinese medicine feed additive of the present invention can promote the development of immune organs and enhance immune function.

[0083] Table 1. Changes in body weight of mice in each experimental group

[0084] Grouping Initial weight / g Final weight / g Weight gain / g Blank control group 20.09±0.48 28.73±0.29 8.43±0.78 Model control group 20.34±0.19 28.37±0.37 8.06±0.45 Test Group 1 20.43±0.27 28.24±0.58 7.81±0.68 Test Group 2 20.22±0.49 27.98±0.42 7.76±0.63 Experiment 3 groups 20.77±0.57 28.71±0.59 7.94±0.30 Test 4 groups 20.31±0.91 28.43±0.54 8.12±0.62 Test 5 groups 20.41±0.67 28.93±0.59 8.52±0.74

[0085] Table 2 Effects of each test group on mouse immune organ index

[0086] Grouping Thymus index (mg / g) Spleen index (mg / g) Blank control group 2.76±0.19 2.71±0.07 Model control group <![CDATA[1.84±0.11 ## ]]> <![CDATA[1.80±0.06 ## ]]> Test Group 1 <![CDATA[2.60±0.06 ** ]]> <![CDATA[2.56±0.04 ** ]]> Test Group 2 1.96±0.05 1.86±0.06 Experiment 3 groups 2.07±0.09 1.94±0.04 Test 4 groups <![CDATA[2.20±0.13 * ]]> <![CDATA[2.15±0.13 ** ]]> Test 5 groups <![CDATA[2.43±0.12 ** ]]> <![CDATA[2.40±0.06 ** ]]>

[0087] Compared with the blank control group, # P<0.05, ## P<0.01. Compared with the model control group, * P<0.05, ** P<0.01.

[0088] 6.2 Effects of each experimental group on mouse immune proteins and inflammatory-related factors

[0089] As shown in Table 3, compared with the blank control group, the levels of immune proteins IgG, IgA and IgM in mice after cyclophosphamide modeling were significantly decreased (P<0.01), and the levels of inflammatory factors IL-4, TNF-α and IFN-γ were significantly decreased (P<0.01); compared with the model control group, the levels of immune proteins IgG, IgA and IgM and inflammatory factors IL-4, TNF-α and IFN-γ in mice in test group 1 and test groups 4-5 were increased to varying degrees, and the difference was significant (P<0.01). Although the levels of immune proteins and inflammatory factors in test groups 2-3 were increased, the difference was not significant (P>0.05), indicating that the Chinese medicine feed additive of the present invention can increase the amount of immune proteins in mouse serum, promote the secretion of inflammatory factors, induce the production and expression of interleukins, interferons and tumor necrosis factors, and enhance the body's immune function.

[0090] Table 3 Effects of each test group on mouse immune protein

[0091] Grouping IgG (pg / mL) Ig A (pg / mL) Ig M (pg / mL) Blank control group 125.53±0.78 150.30±1.99 212.62±2.53 Model control group <![CDATA[96.59±1.59 ## ]]> <![CDATA[120.81±2.71 ## ]]> <![CDATA[165.12±2.40 ## ]]> Test Group 1 <![CDATA[120.78±0.34 ** ]]> <![CDATA[147.70±2.34 ** ]]> <![CDATA[205.88±2.09 ** ]]> Test Group 2 97.96±0.62 122.51±2.54 167.87±2.02 Experiment 3 groups <![CDATA[99.70±2.37 * ]]> 125.85±1.67 169.46±1.09 Test 4 groups <![CDATA[105.70±1.28 ** ]]> <![CDATA[137.93±2.58 ** ]]> <![CDATA[188.75±3.41 ** ]]> Test 5 groups <![CDATA[115.83±0.88 ** ]]> <![CDATA[145.79±2.34 ** ]]> <![CDATA[197.94±2.13 ** ]]>

[0092] Compared with the blank control group, # P<0.05, ## P<0.01. Compared with the model control group, * P<0.05, ** P<0.01.

[0093] Table 4 Effects of each experimental group on inflammatory-related factors in mice

[0094] Grouping IL-4 (pg / mL) TNF-α(pg / mL) IFN-γ (pg / mL) Blank control group 223.88±2.77 174.17±1.58 118.15±2.67 Model control group <![CDATA[152.79±1.53 ## ]]> <![CDATA[142.17±1.72 ## ]]> <![CDATA[76.84±1.65 ## ]]> Test Group 1 <![CDATA[206.44±3.61 ** ]]> <![CDATA[165.96±2.05 ** ]]> <![CDATA[107.94±2.78 ** ]]> Test Group 2 157.86±1.20 145.00±2.45 79.03±1.23 Experiment 3 groups <![CDATA[160.76±1.02 ** ]]> 146.17±1.05 80.79±1.50 Test 4 groups <![CDATA[177.19±1.44 ** ]]> <![CDATA[150.67±1.45 ** ]]> <![CDATA[90.03±1.60 ** ]]> Test 5 groups <![CDATA[198.48±2.31 ** ]]> <![CDATA[160.68±1.52 ** ]]> <![CDATA[102.35±2.76 ** ]]>

[0095] Compared with the blank control group, # P<0.05, ## P<0.01. Compared with the model control group, * P<0.05, ** P<0.01.

[0096] 6.3 Effects of each experimental group on the carbon clearance function of mice

[0097] As shown in Table 5, compared with the blank control group, the phagocytic index of the model control group was significantly reduced (P<0.01), indicating that the clearance ability of the model group mice was significantly reduced; compared with the model control group, the phagocytic index of the test group 1 and the test groups 3-5 was significantly increased (P<0.05), and the phagocytic index of the test group 2 was increased but the difference was not significant (P>0.05), indicating that the Chinese medicine feed additive of the present invention can enhance the ability of mice to clear carbon particles.

[0098] Table 5 Effects of each test group on carbon clearance function of mice

[0099] Grouping K α Blank control group 0.022±0.003 5.36±0.12 Model control group 0.005±0.001 <![CDATA[3.48±0.11 ## ]]> Test Group 1 0.043±0.002 <![CDATA[6.75±0.17 ** ]]> Test Group 2 0.006±0.001 3.59±0.05 Experiment 3 groups 0.007±0.001 <![CDATA[3.77±0.09 * <!-- 7 -->]]> Test 4 groups 0.011±0.001 <![CDATA[4.51±0.13 ** ]]> Test 5 groups 0.022±0.003 <![CDATA[5.45±0.09 ** ]]>

[0100] Compared with the blank control group, # P<0.05, ## P<0.01. Compared with the model control group, * P<0.05, ** P<0.01.

[0101] 6.4 Effects of each test group on the phagocytosis of chicken red blood cells by mouse peritoneal macrophages

[0102] As shown in Table 6, compared with the blank control group, the phagocytic index and phagocytic rate of the model control group were significantly reduced (P<0.01), compared with the model group, the phagocytic index and phagocytic rate of test group 1 and test groups 4-5 were significantly increased (P<0.05), and although the phagocytic index and phagocytic rate of test groups 2-3 showed an increasing trend, the difference was not significant (P>0.05), indicating that the Chinese medicinal feed additive of the present invention can enhance the phagocytic ability of macrophages, thereby improving the clearance ability of the immune system.

[0103] Table 6 Effects of each test group on the test of mouse peritoneal macrophage phagocytosis of chicken red blood cells

[0104] Grouping I P Blank control group 0.35±0.02 50.75±1.31 Model control group <![CDATA[0.23±0.02 ## ]]> <![CDATA[32.08±1.45 ## ]]> Test Group 1 <![CDATA[0.36±0.02 ** ]]> <![CDATA[52.08±2.14 ** ]]> Test Group 2 0.24±0.03 33.01±1.53 Experiment 3 groups 0.26±0.02 35.37±1.07 Test 4 groups <![CDATA[0.28±0.02 * ]]> <![CDATA[40.79±1.57 ** ]]> Test 5 groups <![CDATA[0.31±0.02 ** ]]> <![CDATA[45.27±0.96 ** ]]>

[0105] Compared with the blank control group, # P<0.05, ## P<0.01. Compared with the model control group, * P<0.05, ** P<0.01.

[0106] 7 Discussions

[0107] The Chinese medicinal feed additive of the present invention can improve the immune organ index of cyclophosphamide-induced immunosuppressed mice, promote the development of immune organs, regulate the levels of immune proteins and inflammation-related factors, enhance the ability to remove carbon particles, enhance the phagocytic ability of macrophages, thereby improving the removal ability of the immune system and enhancing the immune function of mice; and the Chinese medicinal feed additive of the present invention is obtained by rationally combining according to the formula theory after dialectical analysis of the cause and determination of the treatment method, and the Chinese medicinal components in the formula are synergistic and indispensable. When a certain component is missing, a good effect of enhancing immunity cannot be achieved.

[0108] Test Example 2

[0109] Immunoenhancing effect of the Chinese medicinal feed additive of the present invention on broiler chickens

[0110] 1 Materials

[0111] 1.1 Experimental animals

[0112] 240 healthy broiler chickens of one day old were selected, male and female mixed, and kept in three-layer cages with infrared lamps for heat preservation. During the feeding period, all animals had free access to food and unlimited drinking water.

[0113] 1.2 Trial Drugs

[0114] 22 parts of astragalus, 20 parts of immature bitter orange, 16 parts of forsythia, 12 parts of black beans, 16 parts of dodder seeds, 10 parts of white peony root, 8 parts of rosemary, 8 parts of thyme, 10 parts of litsea cubeba, 14 parts of mangosteen peel, 10 parts of cinnamon bark and 6 parts of oregano were respectively weighed, washed, dried and then ultra-micro-wall-broken to obtain the traditional Chinese medicine feed additive.

[0115] Levamisole was provided by Qingdao Servier Biopharmaceutical Co., Ltd.

[0116] 2 Methods

[0117] 2.1 Experimental Animal Grouping and Dosing

[0118] 240 broiler chickens were randomly divided into a blank control group, a model control group, a Chinese medicine feed low-dose, medium-dose and high-dose groups, and a levamisole group, with 40 chickens in each group, and adaptive feeding was performed for 7 days; each group of experimental chickens was immunized with a Newcastle disease-infectious bronchitis dual live vaccine by intranasal drops at 8 days of age, and a second immunization was performed 2 weeks later, and the blank control group was intranasally dropped with an equal amount of normal saline; the control group was fed with a complete powder, and the Chinese medicine feed low-dose, medium-dose and high-dose groups were respectively added with 5g / kg, 10g / kg and 15g / kg of the Chinese medicine feed additive of the present invention in the complete powder based on the original drug amount; the levamisole group was added with 2.4g / kg of levamisole powder in the complete feed, and the mixture was evenly mixed and then fed, and the test period was 49 days.

[0119] 2.2 Determination of antibody titer

[0120] On the 7th, 21st and 28th day after the second immunization, 5 chickens in each group were randomly selected for heart blood sampling, and the serum was centrifuged for 2 hours to separate the serum. The Newcastle disease virus antibody titer was determined by the microplate method.

[0121] 2.3 Determination of immune organ index

[0122] On the 7th, 21st and 28th day after the second vaccination, 5 chickens were randomly selected from each group and weighed. Then they were killed and their spleens and bursae of Fabricius were taken out. The excess tissues were stripped off and absorbed with filter paper and then weighed. The spleen index and bursa index were calculated.

[0123] Spleen index = spleen weight (mg) / body weight (g);

[0124] Bursal index = bursal weight (mg) / body weight (g).

[0125] 2.4 Determination of inflammation-related factors

[0126] On the 28th day after the second immunization, 5 chicken hearts were randomly selected in each group for blood collection, and the blood was allowed to stand for 2 hours and centrifuged at 4000 rpm for 10 minutes to separate the serum. The levels of serum cytokines IL-4, TNF-α, and IFN-γ were determined by ELISA.

[0127] 2.5 Statistical methods

[0128] Statistical analysis was performed using GraphPad Prism 8.0. ± standard deviation (s), one-way analysis of variance was used for comparison among groups, and the results were considered to be significantly different when P < 0.05.

[0129] 3 Results

[0130] 3.1 Effects of each group on Newcastle disease virus antibody levels

[0131] As shown in Table 7, compared with the blank control group, the antibody levels of broilers in the model control group were significantly reduced at 7, 21, and 28 days after the second vaccination (P<0.05); compared with the model control group, the antibody levels of broilers in the medium and high dose groups of Chinese medicinal feed were significantly increased at 7, 21, and 28 days after the second vaccination (P<0.05), while the antibody level of the low dose group of Chinese medicinal feed increased but the difference was not significant (P>0.05), and the antibody level of the high dose group of Chinese medicinal feed was significantly higher than that of the positive drug levamisole group (P<0.05), indicating that the Chinese medicinal feed additive of the present invention can promote the formation of Newcastle disease virus antibodies, improve the body's immune response ability, and has a dose-dependent effect on the level of Newcastle disease virus antibodies in chickens.

[0132] Table 7 Effect of each group on Newcastle disease virus antibody level

[0133] Grouping 7d 21d 28d Blank control group <![CDATA[3.66±0.01 c ]]> <![CDATA[5.58±0.02 c ]]> <![CDATA[6.01±0.03 c ]]> Model control group <![CDATA[3.57±0.02 d ]]> <![CDATA[5.51±0.02 d ]]> <![CDATA[5.91±0.02 d ]]> Chinese medicine feed low dose group <![CDATA[3.60±0.01 d ]]> <![CDATA[5.56±0.02 c ]]> <![CDATA[5.98±0.03 c ]]> Chinese medicine feed medium dose group <![CDATA[3.70±0.03 c ]]> <![CDATA[5.62±0.02 b ]]> <![CDATA[6.07±0.01 bc ]]> Chinese medicine feed high dose group <![CDATA[3.84±0.02 a ]]> <![CDATA[5.69±0.02 a ]]> <![CDATA[6.15±0.04 a ]]> Levamisole group <![CDATA[3.77±0.02 b ]]> <![CDATA[5.64±0.02 b ]]> <![CDATA[6.09±0.01 b ]]>

[0134] Data in the same column with different lowercase letters indicate significant differences (P<0.05), and data with the same lowercase letters indicate no significant differences (P>0.05).

[0135] 3.2 Effects of each group on immune organ indexes

[0136] As shown in Table 8, compared with the blank control group, the spleen index of broilers in the model control group decreased significantly at 7, 21, and 28 days after the second vaccination (P<0.05). Compared with the model control group, the spleen index of broilers in the medium and high dose groups of Chinese medicine feed increased significantly (P<0.05), while the spleen index of the low dose group increased but the difference was not significant (P>0.05), and the spleen index of the high dose group of Chinese medicine feed was significantly higher than that of the positive drug levamisole group (P<0.05). As shown in Table 9, compared with the blank control group, the spleen index of broilers in the model control group decreased significantly at 7, 21, and 28 days after the second vaccination (P<0.05). The bursal index of 8d decreased significantly (P<0.05). Compared with the model control group, the bursal index of broiler chickens in the medium and high dose groups of Chinese herbal feed increased significantly (P<0.05), while the bursal index of the low dose group increased but the difference was not significant (P>0.05). The bursal index of the high dose group of Chinese herbal feed was significantly higher than that of the positive drug levamisole group (P<0.05), and the change trend of the bursal index was consistent with the spleen index; this shows that the Chinese herbal feed additive of the present invention can promote the development of immune organs such as spleen and bursa of broiler chickens and enhance nonspecific immune function.

[0137] Table 8 Effect of each group on spleen index

[0138]

[0139]

[0140] Data in the same column with different lowercase letters indicate significant differences (P<0.05), and data with the same lowercase letters indicate no significant differences (P>0.05).

[0141] Table 9 Effect of each group on bursal index

[0142] Grouping 7d(mg / g) 21d(mg / g) 28d(mg / g) Blank control group <![CDATA[3.64±0.01 b ]]> <![CDATA[4.35±0.01 c ]]> <![CDATA[4.42±0.01 b ]]> Model control group <![CDATA[3.55±0.03 c ]]> <![CDATA[4.23±0.02 d ]]> <![CDATA[4.33±0.02 c ]]> Low dose group of Chinese medicine feed <![CDATA[3.58±0.01 c ]]> <![CDATA[4.32±0.01 c ]]> <![CDATA[4.35±0.01 c ]]> Chinese medicine feed medium dose group <![CDATA[3.65±0.03 b ]]> <![CDATA[4.38±0.01 bc ]]> <![CDATA[4.43±0.02 b ]]> High-dose Chinese medicine feed group <![CDATA[3.77±0.02 a ]]> <![CDATA[4.48±0.02 a ]]> <![CDATA[4.55±0.03 a ]]> Levamisole group <![CDATA[3.67±0.01 b ]]> <![CDATA[4.39±0.02 b ]]> <![CDATA[4.45±0.03 b ]]>

[0143] Data in the same column with different lowercase letters indicate significant differences (P<0.05), and data with the same lowercase letters indicate no significant differences (P>0.05).

[0144] 3.3 Effects of each group on serum inflammation-related factors

[0145] As shown in Table 10, compared with the blank control group, the concentrations of IL-4, TNF-α and IFN-γ in the serum of the model control group were significantly decreased (P<0.05); compared with the model control group, the concentrations of IL-4, TNF-α and IFN-γ in the serum of the medium and high dose groups of the Chinese medicine feed were significantly increased (P<0.05), while the concentrations of IL-4, TNF-α and IFN-γ in the serum of the low dose group of the Chinese medicine feed increased but the difference was not significant (P>0.05), and the concentrations of IL-4, TNF-α and IFN-γ in the high dose group of the Chinese medicine feed additive of the present invention were significantly higher than those in the positive drug levamisole group, indicating that the Chinese medicine feed additive of the present invention can stimulate the secretion of IL-4, TNF-α and IFN-γ in chickens and enhance the immune function.

[0146] Table 10 Effects of each group on serum inflammation-related factors

[0147] Grouping IL-4 (pg / mL) TNF-α(pg / mL) IFN-γ (pg / mL) Blank control group <![CDATA[178.79±2.25 a ]]> <![CDATA[135.03±1.46 a ]]> <![CDATA[135.45±3.16 a ]]> Model control group <![CDATA[138.56±2.08 e ]]> <![CDATA[90.33±2.19 e ]]> <![CDATA[82.67±1.86 e ]]> Low dose group of Chinese medicine feed <![CDATA[142.74±2.27 e ]]> <![CDATA[95.11±1.50 e ]]> <![CDATA[90.20±1.84 e ]]> Chinese medicine feed medium dose group <![CDATA[157.76±1.74 d ]]> <![CDATA[110.81±2.44 d ]]> <![CDATA[105.16±3.84 d ]]> Chinese medicine feed high dose group <![CDATA[173.05±1.48 b ]]> <![CDATA[128.48±2.83 b ]]> <![CDATA[125.83±2.86 b ]]> Levamisole group <![CDATA[163.90±1.95 c ]]> <![CDATA[118.50±2.77 c ]]> <![CDATA[115.74±2.99 c ]]>

[0148] Data in the same column with different lowercase letters indicate significant differences (P<0.05), and data with the same lowercase letters indicate no significant differences (P>0.05).

[0149] 4 Discussion

[0150] The Chinese medicinal feed additive of the present invention can improve the antibody titer of Newcastle disease virus in chickens and improve the immune protection of Newcastle disease vaccine in chickens; improve the immune organ index of chickens and promote the development of immune organs; stimulate the expression of interleukin, tumor necrosis factor and interferon in chickens, promote the secretion of inflammation-related factors IL-4, TNF-α and IFN-γ in chickens, and improve the immune response ability of chickens. The Chinese medicinal feed additive of the present invention can achieve a good effect of enhancing immunity at a dosage of 1.5 g / kg for mixing with feed.

[0151] The present invention is a technical solution, rather than a limitation thereto; although the present invention has been described in detail with reference to the aforementioned embodiments, a person skilled in the art should understand that the technical solutions described in the aforementioned embodiments may still be modified, or some of the technical features thereof may be replaced by equivalents; and these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A Chinese medicinal feed additive for improving animal immunity, characterized in that: The Chinese medicinal feed additive is composed of the following raw materials in parts by weight: 10-30 parts of astragalus, 10-30 parts of immature bitter orange, 10-30 parts of forsythia, 5-20 parts of black beans, 5-20 parts of dodder seeds, 5-20 parts of white peony roots, 5-20 parts of rosemary, 5-20 parts of thyme, 5-20 parts of litsea cubeba, 5-20 parts of mangosteen peel, 5-20 parts of cinnamon, and 5-20 parts of oregano.

2. A Chinese medicinal feed additive for improving animal immunity according to claim 1, characterized in that: The Chinese medicinal feed additive is composed of the following raw materials in parts by weight: 15-25 parts of astragalus, 15-25 parts of immature bitter orange, 10-20 parts of forsythia, 5-15 parts of black beans, 10-20 parts of dodder seeds, 5-15 parts of white peony, 5-15 parts of rosemary, 5-15 parts of thyme, 10-20 parts of litsea cubeba, 10-20 parts of mangosteen peel, 5-15 parts of cinnamon, and 5-15 parts of oregano.

3. A Chinese medicinal feed additive for improving animal immunity according to claim 1, characterized in that: The Chinese medicinal feed additive is composed of the following raw materials in parts by weight: 22 parts of astragalus, 20 parts of immature bitter orange, 16 parts of forsythia, 12 parts of black beans, 16 parts of dodder seeds, 10 parts of white peony root, 8 parts of rosemary, 8 parts of thyme, 10 parts of litsea cubeba, 14 parts of mangosteen peel, 10 parts of cinnamon, and 6 parts of oregano.

4. A method for preparing a Chinese medicinal feed additive for improving animal immunity according to any one of claims 1 to 3, characterized in that: The raw materials are weighed according to the above-mentioned portions, washed, dried and then ultra-finely crushed to obtain the Chinese medicinal feed additive.

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