Compound traditional Chinese medicine preparation for resisting porcine reproductive and respiratory syndrome virus as well as preparation method and application of compound traditional Chinese medicine preparation

Abstract

The invention provides an anti-porcine reproductive and respiratory syndrome virus compound traditional Chinese medicine preparation and a preparation method and application thereof, and belongs to the technical field of traditional Chinese medicine preparations. The preparation method comprises the following steps: mixing and crushing 9-11 parts by weight of radix isatidis, 5-6 parts by weight of radix scutellariae and 1.5-2 parts by weight of pericarpium citri reticulatae, adding 26-30 parts by weight of papaya pulp, uniformly mixing, carrying out heat preservation treatment at 45 + / -2 DEG C for 2-3 hours, adding into an ethanol solution of which the mass is 8-10 times that of the papaya pulp, and carrying out ultrasonic reflux and filtration to obtain an extracting solution a; mixing 6-8 parts of honeysuckle, 4-5 parts of astragalus membranaceus, 2.5-3 parts of fructus forsythiae, 1-1.5 parts of polygonum cuspidatum, 1-1.5 parts of puffball and 1-1.5 parts of burdock, crushing, adding into an ethanol solution of which the mass is 8-10 times that of the mixture, soaking, heating, refluxing and filtering, adding 0.3-0.5 part of beta-cyclodextrin into filtrate, and uniformly mixing to obtain an extracting solution b; respectively carrying out vacuum rotary evaporation on the extracting solution a and the extracting solution b until the relative density is 1.12-1.16, mixing, and carrying out spray drying, so as to obtain the traditional Chinese medicine composition. The compound traditional Chinese medicine preparation disclosed by the invention has the advantages of high effect taking speed and obvious curative effect on the porcine reproductive and respiratory syndrome.

Description

Technical Field

[0001] The present invention belongs to the technical field of traditional Chinese medicine preparations, and particularly relates to a compound traditional Chinese medicine preparation for resisting porcine reproductive and respiratory syndrome virus, a preparation method thereof, and an application thereof. Background Art

[0002] Porcine reproductive and respiratory syndrome (PRRS), also known as swine blue ear disease, is a highly contagious disease of pigs caused by porcine reproductive and respiratory syndrome virus (PRRSV). Pigs of different ages, breeds, and genders can be infected, but pregnant sows and piglets within 1 month old are the most susceptible; the disease is mainly characterized by sow abortion, stillbirth, weak fetus, mummified fetus, as well as piglet dyspnea, septicemia, high mortality, etc. It has the characteristics of fast transmission speed and wide incidence, and has caused huge economic losses to the global pig industry.

[0003] Currently, traditional Chinese medicine products for porcine reproductive and respiratory syndrome mainly act through three mechanisms: directly antiviral (preventing adsorption / inhibiting replication), immunomodulatory (enhancing immunity), and relieving symptoms (clearing heat and detoxifying). In terms of specific components: Isatis root contains isatis root polysaccharide and adenosine. The polysaccharide blocks the adsorption of the virus to host cells; adenosine directly inhibits the replication of porcine reproductive and respiratory syndrome virus; it enhances non-specific immunity and increases the proportion of CD3+ / CD4+ lymphocytes. Scutellaria baicalensis contains baicalin, which inhibits the replication of porcine reproductive and respiratory syndrome virus genome and protein synthesis; it relieves respiratory tract inflammation and improves clinical symptoms. Flos Lonicerae contains chlorogenic acid, which interferes with the virus replication process and reduces the virus load; it synergistically enhances the antiviral effect with baicalin.

[0004] Combining different traditional Chinese medicine components can produce different combined action mechanisms / effects. For example, when isatis root and flos lonicerae are combined, the core mechanism is to block adsorption and inhibit replication, achieving the effect of directly antiviral. When scutellaria baicalensis and forsythia suspensa are combined, the core mechanism is anti-inflammatory and neutralizing endotoxin, achieving the effect of relieving symptoms (clearing heat and detoxifying).

[0005] In order to better treat porcine reproductive and respiratory syndrome, it is necessary to further optimize the formula of the traditional Chinese medicine compound preparation for resisting porcine reproductive and respiratory syndrome virus (PRRSV) by combining the pharmacological synergistic mechanisms of existing core traditional Chinese medicine components. However, in the existing preparation process, some drugs are mutually exclusive, resulting in obstruction of the extraction of relevant pharmacodynamic components, and further having a negative impact on the final efficacy. Summary of the Invention

[0006] To solve the problems in the background art, the present invention provides a compound traditional Chinese medicine preparation for resisting porcine reproductive and respiratory syndrome virus, a preparation method thereof, and an application thereof. Through the compatibility of various traditional Chinese medicines and the preparation process of blocking mutual exclusion reactions, it is ensured that the compound traditional Chinese medicine preparation prepared has a fast onset speed and remarkable curative effect against porcine reproductive and respiratory syndrome.

[0007] To achieve the above object, the present invention provides the following technical solutions:

[0008] A preparation method of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus includes the following steps:

[0009] Step 1: Peel and seed green papayas and make them into pulp to obtain papaya pulp; by weight, mix 9-11 parts of isatis root, 5-6 parts of skullcap root, and 1.5-2 parts of dried tangerine peel, crush (40-60 mesh), add 26-30 parts of papaya pulp, mix evenly, keep warm at 45±2°C for 2-3 hours, then add it to an ethanol solution with a mass 8-10 times that of the mixture, carry out ultrasonic reflux, and filter (filter with double-layer 300-mesh filter cloth) to obtain extract a;

[0010] Step 2: By weight, mix 6-8 parts of honeysuckle flower, 4-5 parts of astragalus root, 2.5-3 parts of forsythia fruit, 1-1.5 parts of polygonum cuspidatum, 1-1.5 parts of lasiosphaera fenzlii, and 1-1.5 parts of great burdock achene, crush (30-40 mesh), then add it to an ethanol solution with a mass 8-10 times that of the mixture, soak, heat and reflux, filter (filter with double-layer 300-mesh filter cloth), add 0.3-0.5 part of β-cyclodextrin to the filtrate, mix evenly to obtain extract b;

[0011] Step 3: Vacuum rotary evaporate the extract a obtained in Step 1 and the extract b obtained in Step 2 respectively to a relative density of 1.12-1.16 (measured at 60°C), then mix the two, and carry out spray drying to obtain the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus.

[0012] Preferably, in Step 1, the volume fraction of the ethanol solution is 60%-70%.

[0013] Preferably, in Step 1, the specific operation of the ultrasonic reflux is: carry out ultrasonic treatment at 40-45 kHz, accelerate the release through the cavitation effect, and at the same time, carry out reflux treatment at 80±2°C for efficient dissolution; the duration of the ultrasonic treatment is 30-35 minutes, and the duration of the reflux treatment is 2.5-3.5 hours.

[0014] Preferably, in Step 2, the volume fraction of the ethanol solution is 45%-50%.

[0015] Preferably, in Step 2, the soaking time is 8-12 hours, and the heat reflux is: heat to 60-65°C and reflux for 2-3 hours.

[0016] Preferably, in Step 3, the vacuum rotary evaporation is: carry out rotary evaporation at 60°C under a vacuum degree of -0.08 MPa.

[0017] Preferably, in Step 3, the spray drying is: the inlet air temperature for spray drying is 110-125°C, and the outlet air temperature is 60-70°C.

[0018] A compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus is prepared by using the above preparation method.

[0019] Application of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus (PRRSV). The compound traditional Chinese medicine preparation against PRRSV is prepared by the above-mentioned preparation method; the compound traditional Chinese medicine preparation against PRRSV is mixed into a basal diet to prepare a feed against PRRSV.

[0020] The related effects of each raw material component are as follows:

[0021] Isatis root: contains isatis root polysaccharide and adenosine. The polysaccharide blocks the adsorption of the virus to host cells; adenosine directly inhibits the replication of PRRSV; enhances non-specific immunity and increases the proportion of CD3+ / CD4+ lymphocytes.

[0022] Scutellaria baicalensis: contains baicalin, which inhibits the replication of the PRRSV genome and protein synthesis; relieves respiratory tract inflammation and improves clinical symptoms.

[0023] Tangerine peel: rich in pectinase, which can degrade the pectin colloid (the main source of viscosity) in isatis root polysaccharide, reduce the entanglement between polysaccharide molecules, directly reduce the viscosity, improve the fluidity, and enhance the release efficiency of polysaccharide.

[0024] Green papaya: rich in proteases (such as papain), which can efficiently decompose the heteroprotein components in isatis root, reduce the protein-polysaccharide complex in isatis root polysaccharide, thereby reducing the fluid resistance and increasing the polysaccharide dissolution rate at the same time.

[0025] Honeysuckle flower: contains chlorogenic acid, which interferes with the virus replication process and reduces the virus load; synergizes with baicalin to enhance the antiviral effect.

[0026] Astragalus membranaceus: contains astragalus polysaccharide and saponins, regulates the expression of cytokines, blocks the pathogenic metabolic pathway of the virus; enhances humoral immunity and cellular immunity (such as antibody level, macrophage activity).

[0027] Forsythia suspensa: contains effective components such as phillyrin, volatile oil, luteolin, etc., has a broad-spectrum antiviral effect and has a significant inhibitory effect on PRRSV, etc.

[0028] Reynoutria japonica: contains effective components such as emodin, polydatin, resveratrol, etc., has antiviral and immunomodulatory effects, interferes with the replication of PRRSV, and enhances macrophage activity.

[0029] Lasiosphaera fenzlii: contains effective components such as lasiospaeranin, ergosterol, amino acids, etc., has the effects of hemostasis, anti-inflammatory, detoxifying and relieving sore throat, and antibacterial.

[0030] Arctium lappa: contains arctiin, arctigenol, lignans, etc., has the effects of antiviral and immune enhancement, specifically inhibits parainfluenza virus, and enhances the immunity of respiratory mucosa.

[0031] The present application has the following beneficial effects:

[0032] 1. In the preparation of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus of the present invention, in step one, papaya pulp (green papaya) and dried tangerine peel are introduced into the mixture of isatis root and skullcap root, and after heat preservation treatment at 45 ± 2°C for 2 - 3 h, high-temperature (80 ± 2°C) reflux extraction is carried out to obtain an extract.

[0033] Proteases in green papaya (such as papain) can efficiently decompose the heteroprotein components in isatis root, reduce the protein-polysaccharide complexes in isatis root polysaccharide, thereby reducing the fluid resistance and simultaneously increasing the polysaccharide dissolution rate; dried tangerine peel is rich in pectinase, which can degrade the pectin colloid (the main source of viscosity) in isatis root polysaccharide, reduce the entanglement between polysaccharide molecules, directly reduce the viscosity, improve the fluidity, and enhance the release efficiency of polysaccharide; in this way, papain focuses on protein clearance and pectinase acts on colloid decomposition, avoiding the insufficient effect of a single enzyme, improving the extraction rate, reducing the viscosity, increasing the polysaccharide purity, optimizing the bioavailability of polysaccharide, improving the intestinal absorption rate, enhancing the immune regulation function (such as activating macrophages and natural killer cells), and thus improving the onset speed and curative effect.

[0034] 2. In step two of the present invention, β-cyclodextrin is introduced into the mixed extract of honeysuckle flower, astragalus root, forsythia fruit, polygonum cuspidatum, calvatia gigantea, and great burdock achene, and then it is mixed with the extract obtained in step one and spray-dried.

[0035] The hydrophobic cavity of β-cyclodextrin (β-CD) can form a "molecular isolation chamber" to block the direct contact between chlorogenic acid (honeysuckle flower) and baicalin (skullcap root), avoiding thermal-sensitive mutual damage caused by high-temperature mutual aggregation; the β-CD cavity shields oxidation-sensitive groups (such as the phenolic hydroxyl group of chlorogenic acid), improving the thermal stability after inclusion; when the inlet air temperature of the inclusion complex is 110 - 125°C, the β-CD shell preferentially dehydrates to form a protective layer, shortening the heating time of the internal drug. When the outlet temperature is 60 - 70°C, the glass transition temperature (Tg) of the inclusion complex is higher than that of the free drug, avoiding melting and adhesion; effectively avoiding the decline of the combined pharmacodynamic activity and ensuring a fast onset speed and significant curative effect. Description of the Drawings

[0036] Figure 1 It is a comparison chart of the number of pigs with body temperature recovery when the compound traditional Chinese medicine preparations prepared in Examples 1 - 3 and Comparative Examples 1 - 4 in the test examples of the present invention are used to treat porcine reproductive and respiratory syndrome after 3 days and 7 days of administration (infected piglets);

[0037] Figure 2 It is a comparison chart of the body temperature recovery rate when the compound traditional Chinese medicine preparations prepared in Examples 1 - 3 and Comparative Examples 1 - 4 in the test examples of the present invention are used to treat porcine reproductive and respiratory syndrome after 3 days and 7 days of administration (infected piglets);

[0038] Figure 3In the test examples of the present invention, it is a comparison chart of the number of deaths and cured cases of the compound traditional Chinese medicine preparation prepared in Examples 1-3 and Comparative Examples 1-4 for treating porcine reproductive and respiratory syndrome (PRRS) after 14 days of drug administration (infected piglets).

[0039] Figure 4 In the test examples of the present invention, it is a comparison chart of the mortality rate and cure rate of the compound traditional Chinese medicine preparation prepared in Examples 1-3 and Comparative Examples 1-4 for treating porcine reproductive and respiratory syndrome (PRRS) after 14 days of drug administration (infected piglets). Detailed implementation mode

[0040] The following further elaborates on the present application in conjunction with examples. The raw materials of the examples and comparative examples of the present application are all ordinary commercially available products unless otherwise specified.

[0041] Example 1

[0042] A preparation method of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus (PRRSV) includes the following steps:

[0043] Step 1: Peel and seed green papayas and make them into pulp to obtain papaya pulp.

[0044] By weight, mix 10 parts of Isatis root, 5.5 parts of Scutellaria baicalensis, and 1.8 parts of dried tangerine peel, crush them to 40-60 mesh, add 28 parts of papaya pulp, mix well, after heat preservation treatment at 45 °C for 2.5 h, add them to an ethanol solution with 9 times the mass, and the concentration (volume fraction) of this ethanol solution is 65%.

[0045] Then carry out ultrasonic reflux. The specific operation is: ultrasonic treatment at 42 kHz, accelerating release through cavitation effect. At the same time, carry out reflux treatment at about 80 °C for efficient dissolution; the duration of ultrasonic treatment is 32 min, and the duration of reflux treatment is 3 h. Then use a double-layer 300-mesh nylon filter cloth (moistened with pure water before use and sterilized at 121 °C under high pressure for 20 min) for filtration to obtain a filtrate, which is the extract a.

[0046] Step 2: By weight, mix 7 parts of honeysuckle, 4.5 parts of astragalus membranaceus, 2.8 parts of forsythia suspensa, 1.2 parts of polygonum cuspidatum, 1.2 parts of lasiosphaera fenzlii, and 1.2 parts of arctium lappa, crush them to 30-40 mesh, and then add them to an ethanol solution with 9 times the mass, and the concentration (volume fraction) of this ethanol solution is 48%. After soaking for 10 h, carry out heating reflux, specifically heating to 62 °C and refluxing for 2.5 h. Then use a double-layer 300-mesh nylon filter cloth (moistened with pure water before use and sterilized at 121 °C under high pressure for 20 min) for filtration to obtain a filtrate, and add 0.4 part of β-cyclodextrin to the filtrate and mix well to obtain the extract b.

[0047] Step 3: Separately vacuum rotary evaporate the extract a obtained in Step 1 and the extract b obtained in Step 2 to a relative density of 1.14 (thermally measured at 60°C). The specific operation of vacuum rotary evaporation is as follows: under the condition of a vacuum degree of -0.08 MPa, rotary evaporate at 60°C. Then mix the two, and perform spray drying. Specifically, use a centrifugal atomization disk with a rotation speed of 17,000 rpm and a particle size D50 = 90 - 120 μm. The inlet air temperature for spray drying is about 115°C and the outlet air temperature is about 65°C, thus obtaining the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus.

[0048] Example 2

[0049] A preparation method of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus, comprising the following steps:

[0050] Step 1: Peel and seed green papayas and make them into pulp to obtain papaya pulp.

[0051] By weight, mix 9 parts of isatis root, 5 parts of scutellaria baicalensis, and 1.5 parts of dried tangerine peel, crush them to 40 - 60 mesh, add 26 parts of papaya pulp, mix well, after heat preservation treatment at 45°C for 2 h, add them to an ethanol solution with 8 times the mass, and the concentration (volume fraction) of this ethanol solution is 70%.

[0052] Then perform ultrasonic reflux. The specific operation is as follows: perform ultrasonic treatment at 45 kHz, accelerate the release through the cavitation effect, and at the same time, perform reflux treatment at 80°C for efficient dissolution; the duration of ultrasonic treatment is 30 min, and the duration of reflux treatment is 2.5 h. Then use a double-layer 300-mesh nylon filter cloth (moistened with pure water before use and sterilized at 121°C under high pressure for 20 min) for filtration to obtain a filtrate, which is the extract a.

[0053] Step 2: By weight, mix 6 parts of honeysuckle, 4 parts of astragalus membranaceus, 2.5 parts of forsythia suspensa, 1 part of polygonum cuspidatum, 1 part of lasiosphaera fenzlii, and 1 part of arctium lappa, crush them to 30 - 40 mesh, and then add them to an ethanol solution with 8 times the mass, and the concentration (volume fraction) of this ethanol solution is 50%. After soaking for 8 h, perform heating reflux, specifically heat to 60°C and reflux for 3 h. Then use a double-layer 300-mesh nylon filter cloth (moistened with pure water before use and sterilized at 121°C under high pressure for 20 min) for filtration to obtain a filtrate, and add 0.3 part of β-cyclodextrin to the filtrate and mix well to obtain the extract b.

[0054] Step 3: Separately vacuum rotary evaporate the extract a obtained in Step 1 and the extract b obtained in Step 2 to a relative density of 1.12 (thermally measured at 60°C). The specific operation of vacuum rotary evaporation is as follows: under the condition of a vacuum degree of -0.08 MPa, perform rotary evaporation at 60°C. Then mix the two, and perform spray drying. Specifically, use a centrifugal atomization disk with a rotation speed of 17,000 rpm, a particle size D50 = 90 - 120 μm, a spray drying inlet air temperature of about 115°C, and an outlet air temperature of about 65°C to obtain the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus.

[0055] Example 3

[0056] A preparation method of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus, comprising the following steps:

[0057] Step 1: Peel and seed green papayas and make them into pulp to obtain papaya pulp.

[0058] By weight, mix 11 parts of Isatis root, 6 parts of Scutellaria baicalensis, and 2 parts of dried tangerine peel, crush them to 40 - 60 mesh, add 30 parts of papaya pulp, mix well, after heat preservation treatment at 45°C for 3 h, add them to an ethanol solution with a mass 10 times that of the above mixture, and the concentration (volume fraction) of this ethanol solution is 60%.

[0059] Then perform ultrasonic reflux. The specific operation is as follows: perform ultrasonic treatment at 40 kHz, accelerate the release through the cavitation effect, and at the same time, perform reflux treatment at 80°C for efficient dissolution; the duration of ultrasonic treatment is 35 min, and the duration of reflux treatment is 3.5 h. Then use a double-layer 300-mesh nylon filter cloth (moisten it with pure water before use and perform high-pressure sterilization treatment at 121°C for 20 min) for filtration to obtain a filtrate, which is the extract a.

[0060] Step 2: By weight, mix 8 parts of honeysuckle, 5 parts of astragalus membranaceus, 3 parts of forsythia suspensa, 1.5 parts of polygonum cuspidatum, 1.5 parts of lasiosphaera fenzlii, and 1.5 parts of arctium lappa, crush them to 30 - 40 mesh, and then add them to an ethanol solution with a mass 10 times that of the above mixture, and the concentration (volume fraction) of this ethanol solution is 45%. After soaking for 12 h, perform heating reflux, specifically heat to 65°C and reflux for 2 h. Then use a double-layer 300-mesh nylon filter cloth (moisten it with pure water before use and perform high-pressure sterilization treatment at 121°C for 20 min) for filtration to obtain a filtrate, and add 0.5 part of β-cyclodextrin to the filtrate and mix well to obtain the extract b.

[0061] Step 3: Separately vacuum-rotate the extract a obtained in Step 1 and the extract b obtained in Step 2 until the relative density is 1.16 (thermally measured at 60 °C). The specific operation of vacuum rotation is as follows: under the condition of a vacuum degree of -0.08 MPa, rotate and evaporate at 60 °C. Then mix the two and perform spray drying. Specifically, use a centrifugal atomization disk with a rotation speed of 17,000 rpm and a particle size D50 = 90 - 120 μm. The inlet air temperature for spray drying is about 115 °C and the outlet air temperature is about 65 °C, thus obtaining the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus.

[0062] Comparative Example 1

[0063] The difference between this comparative example and Example 1 is only that: in Step 1, tangerine peel and papaya pulp are not added; and the 45 °C heat preservation treatment is not carried out.

[0064] Specifically as follows: Step 1: Mix 9 parts of isatis root and 5 parts of scutellaria baicalensis by weight, pulverize to 40 - 60 mesh, and add them to an ethanol solution with 8 times the mass. The concentration (volume fraction) of this ethanol solution is 70%.

[0065] Then perform ultrasonic reflux. The specific operation is as follows: perform ultrasonic treatment at 45 kHz, accelerate the release through the cavitation effect, and at the same time, perform reflux treatment at 80 °C for efficient dissolution; the duration of ultrasonic treatment is 30 min, and the duration of reflux treatment is 2.5 h. Then use a double-layer 300-mesh nylon filter cloth (moistened with pure water before use and autoclaved at 121 °C for 20 min) for filtration to obtain a filtrate, which is the extract a.

[0066] Comparative Example 2

[0067] The difference between this comparative example and Example 1 is only that: in Step 1, the 45 °C heat preservation treatment is not carried out.

[0068] Specifically as follows: Step 1: Peel and seed the green papaya and make it into pulp to obtain papaya pulp.

[0069] Mix 10 parts of isatis root, 5.5 parts of scutellaria baicalensis, and 1.8 parts of tangerine peel by weight, pulverize to 40 - 60 mesh, add 28 parts of papaya pulp, mix well, and add them to an ethanol solution with 9 times the mass. The concentration (volume fraction) of this ethanol solution is 65%.

[0070] Then perform ultrasonic reflux. The specific operation is as follows: perform ultrasonic treatment at 42 kHz, accelerate the release through the cavitation effect, and at the same time, perform reflux treatment at about 80 °C for efficient dissolution; the duration of ultrasonic treatment is 32 min, and the duration of reflux treatment is 3 h. Then use a double-layer 300-mesh nylon filter cloth (moistened with pure water before use and autoclaved at 121 °C for 20 min) for filtration to obtain a filtrate, which is the extract a.

[0071] Comparative Example 3

[0072] The difference between this comparative example and Example 1 is only that: honeysuckle and skullcap root are refluxed and extracted together at about 80°C in Step 1.

[0073] Specifically as follows: Step 1, peel and seed green papaya and then make it into pulp to obtain papaya pulp.

[0074] By weight, mix 10 parts of isatis root, 5.5 parts of skullcap root, 7 parts of honeysuckle, and 1.8 parts of dried tangerine peel, crush them to 40 - 60 mesh, add 28 parts of papaya pulp, mix well, after heat preservation treatment at 45°C for 2.5 h, add them into an ethanol solution with 9 times the mass, and the concentration (volume fraction) of this ethanol solution is 65%.

[0075] Then carry out ultrasonic reflux, and the specific operation is: ultrasonic treatment at 42 kHz, accelerating release through cavitation effect. At the same time, carry out reflux treatment at about 80°C for efficient dissolution; the duration of ultrasonic treatment is 32 min, and the duration of reflux treatment is 3 h. Then use a double - layer 300 - mesh nylon filter cloth (moistened with pure water before use and sterilized at 121°C under high pressure for 20 min) for filtration to obtain a filtrate, which is the extract a.

[0076] Step 2, by weight, mix 4.5 parts of astragalus root, 2.8 parts of forsythia fruit, 1.2 parts of polygonum cuspidatum, 1.2 parts of lasiosphaera seu calvatia, and 1.2 parts of great burdock achene, crush them to 30 - 40 mesh, and then add them into an ethanol solution with 9 times the mass, and the concentration (volume fraction) of this ethanol solution is 48%. After soaking for 10 h, carry out heating reflux, specifically heat to 62°C and reflux for 2.5 h. Then use a double - layer 300 - mesh nylon filter cloth (moistened with pure water before use and sterilized at 121°C under high pressure for 20 min) for filtration to obtain a filtrate, add 0.4 part of β - cyclodextrin to the filtrate, and mix well to obtain the extract b.

[0077] Comparative Example 4

[0078] The difference between this comparative example and Example 1 is only that: in Step 2, β - cyclodextrin is not added.

[0079] Specifically as follows: Step 2, by weight, mix 7 parts of honeysuckle, 4.5 parts of astragalus root, 2.8 parts of forsythia fruit, 1.2 parts of polygonum cuspidatum, 1.2 parts of lasiosphaera seu calvatia, and 1.2 parts of great burdock achene, crush them to 30 - 40 mesh, and then add them into an ethanol solution with 9 times the mass, and the concentration (volume fraction) of this ethanol solution is 48%. After soaking for 10 h, carry out heating reflux, specifically heat to 62°C and reflux for 2.5 h. Then use a double - layer 300 - mesh nylon filter cloth (moistened with pure water before use and sterilized at 121°C under high pressure for 20 min) for filtration to obtain a filtrate, which is the extract b.

[0080] Test Example

[0081] 1. Experimental animals and grouping: 408 fattening pigs with positive PRRSV antigen (confirmed by RT-PCR) and typical clinical symptoms (body weight 20 ± 2 kg) were selected and randomly divided into 8 groups (51 pigs in each group, raised in independent pig pens). One group was the blank group, without medication after infection; the other 7 groups were experimental groups (Example 1 group - Example 3 groups and Comparative Example 1 group - Comparative Example 4 groups), and corresponding medications were given respectively (compound traditional Chinese medicine preparations prepared in Example 1 - Example 3 and Comparative Example 1 - Comparative Example 4).

[0082] 2. Viral challenge and medication regimen: Intramuscular injection of NADC30 strain (1×10 5 TCID 50 / mL, 2 mL / head) was carried out uniformly. Medication started after the appearance of fever (>40.5 °C) and respiratory symptoms, for 14 consecutive days. Medication was given by mixing with feed (the mass ratio of compound traditional Chinese medicine preparation to feed was 1%), fed 4 times a day, the dosage was 10 g / day, and the daily feed intake was 1.0 kg / day.

[0083] 3. Test items and results: 3-1. Speed of effect: On the 3rd and 7th days after infection, the body temperature was measured 2 times a day. <40 °C was regarded as returning to normal, and the body temperature recovery rate was calculated. Calculation formula: Body temperature recovery rate (%) = Number of recovered body temperatures / 51 × 100%; the larger the value of the body temperature recovery rate, the faster the speed of effect of the compound traditional Chinese medicine preparation prepared in the corresponding group. The test results are shown in Table 1.

[0084] 3-2. Efficacy: ① On the 14th day after infection, the number of deaths was recorded and the mortality rate was calculated. Calculation formula: Mortality rate (%) = Number of deaths / 51 × 100%; the smaller the value of the mortality rate, the better the efficacy of the compound traditional Chinese medicine preparation prepared in the corresponding group. ② On the 14th day after infection, the number of cured pigs (symptoms completely disappeared and the virus turned negative) was recorded and the cure rate was calculated. Calculation formula: Cure rate (%) = Number of cured pigs / 51 × 100%; the larger the value of the cure rate, the better the efficacy of the compound traditional Chinese medicine preparation prepared in the corresponding group. The test results are shown in Table 2.

[0085] Table 1. Statistical data of speed of effect - body temperature recovery rate

[0086]

[0087] Table 2. Statistical data of efficacy - mortality rate / cure rate

[0088]

[0089] Result analysis: Combining the data in Table 1 - Table 2 and Figures 1-4, By analyzing Examples 1 - 3, it can be seen that the compound traditional Chinese medicine preparation prepared by the present invention (Examples 1 - 3) is used to treat porcine reproductive and respiratory syndrome. The body temperature recovery rate reaches over 58.8% after 3 days of administration, and over 96.1% after 7 days of administration; and the cure rate reaches over 84.3% after 14 days of administration. This indicates that the compound traditional Chinese medicine preparation prepared by the present invention has a fast onset speed and significant curative effect against porcine reproductive and respiratory syndrome.

[0090] Combined with the data in Tables 1 - 2 and Figures 1-4 , By analyzing Example 1 and Comparative Examples 1 - 4, specifically by comparing Comparative Example 1 and Comparative Example 2, it can be seen that compared with Comparative Example 1, in Comparative Example 2, papaya pulp and tangerine peel were added together for extraction. As a result, the body temperature recovery rates after 3 days and 7 days of administration both decreased, the mortality rate increased after 14 days of administration, and the cure rate decreased. This indicates that adding papaya pulp and tangerine peel here will instead lead to a slower onset speed and reduced curative effect of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome.

[0091] The main reason is that when green papaya and tangerine peel are introduced alone without heat preservation enzymatic hydrolysis treatment, during the extraction process, the ethanol reflux temperature of 80°C far exceeds the tolerance ranges of papain (optimum 40 - 50°C) and pectinase (optimum 45 - 55°C), resulting in the instantaneous inactivation of the enzyme protein and the loss of enzymatic hydrolysis ability. And the high - concentration ethanol solution (65%) causes irreversible denaturation of the enzyme's spatial structure. The denatured enzyme protein can bind to polysaccharides, leading to an increase in molecular aggregates and viscosity, thereby hindering subsequent absorption (reducing intestinal absorption rate), having a negative impact on the immune activation effect, and further resulting in a slower onset speed and reduced curative effect of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome.

[0092] By comparing with Example 1 again, it can be seen that adding papaya pulp and tangerine peel and performing heat preservation enzymatic hydrolysis treatment at 45 ± 2°C for 2 - 3 h can accelerate the onset speed and improve the curative effect of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome.

[0093] By comparing Example 1 and Comparative Example 3, it can be seen that compared with Example 1, in Comparative Example 3, honeysuckle and scutellaria baicalensis were reflux - extracted together at about 80°C in Step 1. As a result, the body temperature recovery rates after 3 days and 7 days of administration both decreased, the mortality rate increased after 14 days of administration, and the cure rate decreased. This indicates that reflux - extracting honeysuckle and scutellaria baicalensis together at about 80°C in Step 1 will lead to a slower onset speed and reduced curative effect of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome.

[0094] The main reason is that chlorogenic acid (honeysuckle) and baicalin (scutellaria baicalensis) have thermal - sensitive mutual damage: when co - extracted at high temperature of 80°C, the two are prone to form molecular aggregates, resulting in precipitation and decreased activity, affecting the exertion of the corresponding drug effects.

[0095] By comparing Example 1 with Comparative Example 4, it can be seen that, compared with Example 1, β-cyclodextrin is not added in Comparative Example 4, and as a result, the body temperature recovery rates on the 3rd and 7th days after administration both decrease, the mortality rate increases on the 14th day after administration, and the cure rate decreases. This shows that deleting β-cyclodextrin will lead to a slower onset speed and reduced efficacy of the prepared compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome.

[0096] This is mainly because after the inclusion blocking effect brought by β-cyclodextrin is lacking, spray drying will, to a certain extent, cause chlorogenic acid (honeysuckle) and baicalin (scutellaria baicalensis) to polymerize with each other at high temperature, resulting in a decrease in application activity and affecting the exertion of the corresponding drug efficacy.

[0097] In addition, it should be noted that among the various specific technical features described in the above specific embodiments, without conflict, they can be combined in any appropriate manner. To avoid unnecessary repetition, the present invention will not separately describe various possible combination methods.

[0098] In addition, any combination can be made among various different embodiments of the present invention, as long as it does not violate the idea of the present invention, it should also be regarded as the content disclosed by the present invention.

Claims

1. A preparation method of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus, characterized in that It includes the following steps: Step 1: Peel and seed the green papaya and make it into pulp to obtain papaya pulp; by weight, mix 9 - 11 parts of Isatis indigotica Fort., 5 - 6 parts of Scutellaria baicalensis Georgi, and 1.5 - 2 parts of Pericarpium Citri Reticulatae, pulverize them, add 26 - 30 parts of papaya pulp, mix well, keep it at 45 ± 2 °C for heat treatment for 2 - 3 h, then add it into an ethanol solution with a mass 8 - 10 times that of the above mixture, perform ultrasonic reflux, filter, and obtain extract a; Step 2: By weight, mix 6 - 8 parts of Lonicera japonica Thunb., 4 - 5 parts of Astragalus membranaceus (Fisch.) Bunge, 2.5 - 3 parts of Forsythia suspensa (Thunb.) Vahl, 1 - 1.5 parts of Polygonum cuspidatum Sieb. et Zucc., 1 - 1.5 parts of Lasiosphaera fenzlii Reichb., and 1 - 1.5 parts of Arctium lappa L., pulverize them, then add them into an ethanol solution with a mass 8 - 10 times that of the above mixture, soak, heat and reflux, filter, add 0.3 - 0.5 part of β - cyclodextrin to the filtrate, mix well, and obtain extract b; Step 3: Vacuum - rotate evaporate the extract a obtained in Step 1 and the extract b obtained in Step 2 respectively until the relative density is 1.12 - 1.16, then mix the two, and perform spray - drying to obtain the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus.

2. The preparation method of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus according to claim 1, characterized in that, In Step 1, the volume fraction of the ethanol solution is 60% - 70%.

3. The preparation method of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus according to claim 1, characterized in that, In Step 1, the specific operation of ultrasonic reflux is: perform ultrasonic treatment at 40 - 45 kHz, accelerate the release through cavitation effect, and at the same time, perform reflux treatment at 80 ± 2 °C for efficient dissolution; the duration of ultrasonic treatment is 30 - 35 min, and the duration of reflux treatment is 2.5 - 3.5 h.

4. The preparation method of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus according to claim 1, characterized in that, In Step 2, the volume fraction of the ethanol solution is 45% - 50%.

5. The preparation method of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus according to claim 1, characterized in that, In Step 2, the soaking time is 8 - 12 h, and the heat - reflux is: heat to 60 - 65 °C and reflux for 2 - 3 h.

6. The preparation method of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus according to claim 1, characterized in that, In Step 3, the vacuum - rotation evaporation is: perform rotation evaporation at 60 °C under the condition of a vacuum degree of - 0.08 MPa.

7. The preparation method of the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus according to claim 1, characterized in that, In Step 3, the spray - drying is: the inlet air temperature for spray - drying is 110 - 125 °C, and the outlet air temperature is 60 - 70 °C.

8. A compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus, characterized in that, Prepared by the preparation method described in any one of claims 1 - 7.

9. Use of a compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus, characterized in that, The compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus is prepared by the preparation method described in any one of claims 1 - 7; mix the compound traditional Chinese medicine preparation against porcine reproductive and respiratory syndrome virus into the basal feed to prepare the feed against porcine reproductive and respiratory syndrome virus.

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