Extraction method of cassia twig and radix puerariae soup

Abstract

The invention provides an extraction method of cassia twig and radix puerariae decoction, and belongs to the technical field of traditional Chinese medicine extraction. In the extraction method, volatile oil components in the cassia twig and the ginger are firstly extracted by adopting ethanol, and then the cassia twig and ginger residues and effective components in the remaining four medicines are extracted by adopting a water extraction mode, so that the loss of the volatile oil components is avoided, and the medicinal effect of the cassia twig and radix puerariae decoction is reduced. Besides, cyclodextrin is added into the alcohol extract of cassia twig and ginger, and the volatile oil component dissolved in ethanol is adsorbed into cyclodextrin, so that the stability of the volatile oil component is effectively improved through cyclodextrin, the volatile oil component cannot be lost along with precipitation of ethanol in the vacuum concentration process, and the content of the volatile oil component in the alcohol extract concentrated solution is improved. According to the extraction method provided by the invention, effective components in the cassia twig and radix puerariae decoction can be extracted to the greatest extent, and the extraction process is simple and easy to implement.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of traditional Chinese medicine extraction, and relates to an extraction method of Guizhiga Gegen decoction. BACKGROUND

[0002] Guizhiga Gegen decoction is a traditional Chinese medicine decoction composed of six traditional Chinese medicines, namely, Gegen, Guizhi, Baishao, Zhigancao, Shengjiang and Dazao, and has the effects of relieving superficies and dispelling cold and tonifying middle qi, and is mainly used for treating exogenous fever and intestinal diseases.

[0003] At present, the extraction process of Guizhiga Gegen decoction mainly uses water as a solvent, but Guizhi and Shengjiang in the prescription contain volatile oil components. Modern research shows that volatile components have pharmacological effects such as anti-inflammatory, antiviral and antipyretic, and can be used as one of the effective material bases for resisting influenza. However, due to the poor solubility of volatile oil components in water, they are lost with water vapor in the traditional decoction process, which leads to the neglect of the role of volatile components in the previous administration mode. In addition, the processing of traditional Chinese medicine processing may also destroy the active ingredients in the volatile oil that play a pharmacological role, resulting in a decrease in the curative effect of volatile oil.

[0004] Regarding the extraction of volatile oil in Guizhiga Gegen decoction, the traditional steam distillation method has the problems of destruction of heat-labile components in the extraction process and poor oil-water separation effect, which easily leads to low oil yield, and thus the pharmacological effects of Guizhiga Gegen decoction cannot be effectively exerted. SUMMARY

[0005] The purpose of the present application is to provide an extraction method of Guizhiga Gegen decoction to solve the problem that the existing extraction method cannot completely extract the effective components in Guizhiga Gegen decoction.

[0006] To achieve the above-mentioned purpose, the present application adopts the following technical solutions: The present application provides an extraction method of Guizhiga Gegen decoction, which comprises the following steps: Soaking Guizhi and Shengjiang in ethanol and heating to reflux extraction to obtain an alcohol extraction liquid and a residue; Adding cyclodextrin to the alcohol extraction liquid and reducing pressure to concentrate to obtain an alcohol extraction concentrate; Soaking the residue, Gegen, Baishao, Zhigancao and Dazao in water and heating to reflux extraction three times, combining and concentrating the filtrate, and then mixing the alcohol extraction concentrate to obtain Guizhiga Gegen decoction extract.

[0007] The present application has the following beneficial effects: (1) In the present application, ethanol is used to extract volatile oil components in Guizhi and Shengjiang first, and then water extraction is used to extract effective components in Guizhi and Shengjiang residue and the remaining four medicines, which avoids the loss of volatile oil components and reduces the pharmacological effects of Guizhiga Gegen decoction.

[0008] (2) In the present application, cyclodextrin is added to the alcohol extract of Ramulus Cinnamomi and raw ginger, and the volatile oil components dissolved in ethanol are adsorbed into cyclodextrin, so as to effectively improve the stability of the volatile oil components by cyclodextrin, and then the volatile oil components are not lost with the precipitation of ethanol in the process of vacuum concentration, and the content of the volatile oil components in the alcohol extraction concentrate is improved.

[0009] (3) The extraction method provided by the present application can extract the effective components in Guizhiga Gegen Decoction to the maximum extent, and the extraction process is simple and easy to implement. BRIEF DESCRIPTION OF DRAWINGS

[0010] Figure 1 is an ethanol volume concentration influence diagram; Figure 2 is an alcohol extraction time influence diagram; Figure 3 is an ethanol multiple influence diagram; Figure 4 is a cyclodextrin addition amount influence diagram; Figure 5 is a concentration temperature influence diagram on the retention of alcohol extract components in the concentration process; Figure 6 is a comparison diagram of the body weight of each group of mice; Figure 7 is a comparison diagram of the lung index of each group of mice; Figure 8 is a comparison diagram of the thymus index of each group of mice; Figure 9 is a comparison diagram of the spleen index of each group of mice; Figure 10 is a comparison diagram of the colon length of each group of mice; Figure 11 is a comparison diagram of the lung viral load of each group of mice; Figure 12 is a comparison diagram of the relative expression amount of the inflammatory factor IL-1β gene of each group of mice; Figure 13 is a lung staining diagram of each group of mice, wherein a-e are the staining diagrams of the normal group, the model group, the positive drug group, the comparison diagram and the experimental group, respectively. DETAILED DESCRIPTION

[0011] The present application provides an extraction method of Guizhiga Gegen Decoction, which comprises: S01: Ramulus Cinnamomi and raw ginger are added to 6-8 times of ethanol with a volume concentration of 70%-90% and soaked for 30-50 min, and heated to reflux extraction at 80-100℃ for 90-120 min, so as to extract the volatile oil components in Ramulus Cinnamomi and raw ginger by ethanol, and obtain alcohol extract and residue.

[0012] S02: The cyclodextrin and the alcohol extract are mixed in a mass-volume ratio of 1:45, and then concentrated under reduced pressure at a temperature of 45-60℃ to obtain an alcohol extract concentrate. When cyclodextrin is added to the alcohol extract, the volatile oil components dissolved in ethanol are adsorbed into the cyclodextrin, so as to effectively improve the stability of the volatile oil components by the cyclodextrin, and then the volatile oil components are not lost with the ethanol during the concentration under reduced pressure, thereby improving the content of the volatile oil components in the alcohol extract concentrate.

[0013] S03: The residue, together with Puerariae radix, Radix Paeoniae Alba, Glycyrrhizae radix praeparata and Jujube, is soaked in 8-10 times the amount of water for 30-50 min, and then heated to reflux extraction at 80-100℃ for 1.5h to obtain an extract and a residue a; 8 times the amount of water is added to the residue a, and then heated to reflux at 80-100℃ for 1h to obtain a filtrate a and a residue b; 8 times the amount of water is added to the residue b, and then heated to reflux at 80-100℃ for 1h to obtain a filtrate b and a residue; the extract, the filtrate a and the filtrate b are combined and concentrated under reduced pressure to obtain a concentrate, which is mixed with the alcohol extract concentrate to obtain a Gui Zhi Jia Gegen Decoction extract.

[0014] To determine the optimal process parameters in the extraction method of Gui Zhi Jia Gegen Decoction, the content of cinnamic acid, cinnamyl aldehyde and 6-gingerol is taken as a reference in this application, and single factor test and orthogonal test are carried out respectively to determine the influence of the volume concentration of ethanol, alcohol extraction time, ethanol multiple, cyclodextrin addition amount and concentration temperature on the retention of alcohol extract components during the concentration process. The specific content is as follows: I. Single factor test 1. Volume concentration of ethanol According to the prescription of Gui Zhi Jia Gegen Decoction, 12g of Cinnamomum cassia and 18g of Zingiber officinale are weighed, and 5 portions are obtained. 50%, 60%, 70%, 80% and 90% of ethanol are added to the 5 portions of Cinnamomum cassia and Zingiber officinale respectively, and the solid-liquid ratio is 1:10. The mixture is heated to reflux extraction for 1h to obtain an extract. The content of volatile oil components, cinnamic acid, cinnamyl aldehyde and 6-gingerol in the extract is determined by high performance liquid chromatography, and the comprehensive score is calculated by AHP weighted scoring method to obtain Figure 1 . The chromatographic conditions are as follows: chromatographic column: Thermo Syncronis-C 18 (250 mm×4.6 mm, 5μm); mobile phase: acetonitrile (A)-0.1% formic acid aqueous solution (B); flow rate: 1.0 mL / min; detection wavelength: 235 nm, 280 nm, 250 nm. Gradient elution: 0-20 min, 12-16% A; 20-24 min, 16-20% A; 24-32 min, 20-25% A; 32-53 min, 25-40% A; 53-65 min, 40-42% A; 65-75 min, 42-100% A; column temperature: 28℃; sample size: 25μL.

[0015] From the attached Figure 1 It can be seen that with the increase of the volume concentration of ethanol, the comprehensive score first rises and then falls, so it is known that when extracting the effective components in cassia twig and ginger, the optimal volume fraction of ethanol is 80%.

[0016] 2, alcohol extraction time According to the prescription of Guizhijia Gegen decoction 2 times, 12g of cassia twig and 18g of ginger were weighed, and 5 portions were obtained. To the 5 portions of cassia twig and ginger, 80% volume concentration of ethanol was added, the solid-liquid ratio was 1:10, and the heating reflux extraction time was 30, 60, 90, 120 and 150 min respectively, and the extraction liquid was obtained. The contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol in the extraction liquid were determined by high performance liquid chromatograph according to the chromatographic conditions in "1, volume concentration of ethanol", and the comprehensive score was calculated by AHP weighted score method, and the results were obtained Figure 2 .

[0017] From the attached Figure 2 It can be seen that with the extension of alcohol extraction time, the comprehensive score first rises and then falls, so it is known that when extracting the effective components in cassia twig and ginger, the optimal alcohol extraction time is 2h.

[0018] 3, ethanol multiple According to the prescription of Guizhijia Gegen decoction 2 times, 12g of cassia twig and 18g of ginger were weighed, and 5 portions were obtained. To the 5 portions of cassia twig and ginger, 80% volume concentration of ethanol was added, the solid-liquid ratio was 1:4, 1:6, 1:8, 1:10 and 1:12, and the extraction time was 1h, and the extraction liquid was obtained. The contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol in the extraction liquid were determined by high performance liquid chromatograph according to the chromatographic conditions in "1, volume concentration of ethanol", and the comprehensive score was calculated by AHP weighted score method, and the results were obtained Figure 3 .

[0019] From the attached Figure 3 It can be seen that with the increase of ethanol multiple, the comprehensive score first rises and then falls, so it is known that when extracting the effective components in cassia twig and ginger, the optimal ethanol multiple is 8 times.

[0020] 4, cyclodextrin addition amount According to the Guizhiga Gegen Decoction 2 times prescription ratio of 12 g of cassia twig and 18 g of ginger, 5 parts. The cassia twig and ginger were added to 180 mL of 80% volume concentration of ethanol, soaked for 30 min, and then heated to reflux. The obtained extract was filtered through 4 layers of gauze to obtain an alcohol extract. To the alcohol extract, 80% volume concentration of ethanol was added to make up to 180 mL. According to the volume-mass ratio of the alcohol extract to cyclodextrin of 35:1, 40:1, 45:1, 50:1 and 55:1, 5.14, 4.50, 4.00, 3.60, 3.27 g of cyclodextrin was weighed, mixed, vacuum concentrated, and made up to 50 mL. The contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol were determined by high performance liquid chromatography according to the chromatographic conditions in "1, Volume concentration of ethanol", and the comprehensive score was calculated by AHP weighted scoring method to obtain the following table: Figure 4 .

[0021] From the table Figure 4 It can be seen that with the increase of the amount of cyclodextrin, the comprehensive score first increases and then decreases, so it is known that when extracting the effective components of cassia twig and ginger, the volume-mass ratio of alcohol extract to cyclodextrin is optimal at 45:1.

[0022] 5, Effect of concentration temperature on the retention of alcohol extract components during concentration According to the Guizhiga Gegen Decoction 2 times prescription ratio of 12 g of cassia twig and 18 g of ginger, 5 parts. The cassia twig and ginger were added to 180 mL of 80% volume concentration of ethanol, soaked for 30 min, and then heated to reflux. The obtained extract was filtered through 4 layers of gauze to obtain an alcohol extract. To the alcohol extract, 80% volume concentration of ethanol was added to make up to 180 mL. To the alcohol extract, 4 g of cyclodextrin was added, and vacuum concentration was carried out at 30, 40, 50, 60 and 70°C, respectively, and made up to 50 mL. The contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol were determined by high performance liquid chromatography according to the chromatographic conditions in "1, Volume concentration of ethanol", and the comprehensive score was calculated by AHP weighted scoring method to obtain the following table: Figure 5 .

[0023] From the table Figure 5 It can be seen that with the increase of the amount of cyclodextrin, the comprehensive score first increases and then decreases, so it is known that when extracting the effective components of cassia twig and ginger, the volume-mass ratio of alcohol extract to cyclodextrin is optimal at 45:1.

[0024] II. Orthogonal test 1. Orthogonal test of the optimal condition of alcohol extraction process According to the Guizhiga Gegen Decoction 2 times prescription ratio of 12 g of cassia twig and 18 g of ginger, 9 parts. According to L9(34) orthogonal test table, 9 parts of cassia twig and ginger were added to 270 mL of 80% volume concentration of ethanol, soaked for 30 min, and then heated to reflux. The obtained extract was filtered through 4 layers of gauze to obtain an alcohol extract. To the alcohol extract, 80% volume concentration of ethanol was added to make up to 270 mL. According to the volume-mass ratio of the alcohol extract to cyclodextrin of 35:1, 40:1, 45:1, 50:1 and 55:1, 5.14, 4.50, 4.00, 3.60, 3.27 g of cyclodextrin was weighed, mixed, vacuum concentrated, and made up to 50 mL. The contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol were determined by high performance liquid chromatography according to the chromatographic conditions in "1, Volume concentration of ethanol", and the comprehensive score was calculated by AHP weighted scoring method to obtain the following table: 4) orthogonal test table to heat reflux extraction, filter, put cold, concentrated filtrate to 50ml, get extract. 5ml extract using 50% methanol solution and transfer to 10ml capacity bottle, constant volume, using 0.45 μm microporous filter membrane filter, get test solution. With test solution in volatile oil components cinnamyl aldehyde, cinnamic acid, 6-gingerol content and dry extract yield as evaluation index, solvent multiple (A), extraction time (B), extraction times (C), alcohol volume (D) as influencing factors, set the weighted coefficient of volatile oil components cinnamyl aldehyde, cinnamic acid, 6-gingerol content is 1, the weighted coefficient of dry extract yield is 1 to determine the optimum extraction process of guizhiga gen root decoction, get table 1, 2. Among them, comprehensive score = (cinnamyl aldehyde content / maximum value + 6-gingerol content / maximum value + cinnamic acid content / maximum value) × 100%.

[0025] Table 1: orthogonal test results Table 2: variance analysis Note: F test critical value: P0.01 (2, 2) = 99.00; P0.05 (2, 2) = 19.00.

[0026] From table 1, the influence degree of each factor is B > A > D > C, that is, extraction time > solvent multiple > alcohol volume > extraction times. From table 2, the influence of ABCD four factors has no statistical significance (P>0.05). Considering the extraction efficiency and energy saving factors, combined with single factor investigation results, the optimal conditions of guizhiga gen root decoction extraction method are finally determined as A1B1C1D2, that is, solvent multiple is 6 times, extraction time is 90 min, extraction 1 times, ethanol volume concentration is 80%.

[0027] To verify the above optimal conditions, the present application embodiment according to the prescription respectively takes 3 batches of medicinal materials, extracts guizhiga gen root decoction according to the above optimal process, and determines the content of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol in alcohol extract by high performance liquid chromatograph according to the chromatographic conditions in "1, ethanol volume concentration". The comprehensive score is calculated by AHP weighted scoring method, and table 3 is obtained.

[0028] Table 3: extraction process verification results (n=3) From table 3, it can be seen that the contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol extracted by each test group under the above optimal process conditions are relatively small, which shows that the process is stable and feasible.

[0029] 2, verification of vacuum concentration process conditions To verify the optimal process conditions of the reduced pressure concentration, three batches of medicinal materials were respectively weighed according to the prescription in the embodiment of the application, and alcohol extraction concentrated liquid was prepared according to the optimal process conditions of the reduced pressure concentration. The contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol in the alcohol extraction concentrated liquid were determined by high performance liquid chromatography according to the chromatographic conditions in “1, volume concentration of ethanol”, and the comprehensive score was calculated by AHP weighted scoring method, and Table 4 was obtained.

[0030] Table 4: verification results of the reduced pressure concentration process (n = 3) As can be seen from Table 4, the contents of volatile oil components cinnamic acid, cinnamyl aldehyde and 6-gingerol extracted by each test group under the above optimal process conditions are relatively small, which indicates that the process is stable and feasible.

[0031] The technical solutions of the application will be further explained and described below through specific embodiments.

[0032] Embodiment 1 The embodiment of the application provides an extraction method of Guizhijia Gegen Decoction, which comprises the following steps: S101: 30g of Ramulus Cinnamomi and 45g of raw ginger are added into 6 times of ethanol with a volume concentration of 80% for soaking for 30min, and heated to reflux for 90min to obtain alcohol extraction liquid and residue.

[0033] S102: cyclodextrin and the alcohol extraction liquid are mixed uniformly according to a mass-volume ratio of 1:45, and then reduced pressure concentration is performed under the condition of a temperature of 60℃ to obtain alcohol extraction concentrated liquid.

[0034] S103: the residue, 60g of Radix Puerariae, 30g of Radix Paeoniae Alba, 30g of Radix Glycyrrhizae Preparata and 35g of Fructus Jujubae are added into 10 times of water for soaking for 30min, and heated to reflux for 1.5h to obtain extraction liquid and residue a; 8 times of water is added into the residue a, and heated to reflux for 1h to obtain filtrate a and residue b; 8 times of water is added into the residue b, and heated to reflux for 1h to obtain filtrate b and residue; the extraction liquid, the filtrate a and the filtrate b are combined, and then reduced pressure concentration is performed to obtain concentrated liquid; and the concentrated liquid is mixed uniformly with the alcohol extraction concentrated liquid to obtain Guizhijia Gegen Decoction extract.

[0035] Embodiment 2 The embodiment of the application provides an extraction method of Guizhijia Gegen Decoction, which comprises the following steps: S201: 30g of Ramulus Cinnamomi and 45g of raw ginger are added into 6 times of ethanol with a volume concentration of 90% for soaking for 30min, and heated to reflux for 120min to obtain alcohol extraction liquid and residue.

[0036] S202: cyclodextrin and the alcohol extraction liquid are mixed uniformly according to a mass-volume ratio of 1:45, and then reduced pressure concentration is performed under the condition of a temperature of 60℃ to obtain alcohol extraction concentrated liquid.

[0037] S203: Soak the residue with 60g of kudzu root, 30g of white peony root, 30g of prepared licorice root, and 35g of jujube in 10 times the amount of water for 30 minutes, then heat and reflux for 1.5 hours to obtain the extract and residue a; add 8 times the amount of water to residue a, heat and reflux for 1 hour to obtain filtrate a and residue b; add 8 times the amount of water to residue b, heat and reflux for 1 hour to obtain filtrate b and residue; combine the extract, filtrate a, and filtrate b, concentrate under reduced pressure, and mix the concentrate with the alcohol extract concentrate to obtain Guizhi Jia Gegen Tang extract.

[0038] Example 3 This application provides an extraction method for Guizhi Jia Gegen Tang (Cinnamon Twig and Kudzu Root Decoction), the method comprising: S301: Add 30g of cinnamon twigs and 45g of ginger to 6 times the volume of 70% ethanol and soak for 50 minutes. Then heat and reflux to extract for 90 minutes to obtain the ethanol extract and residue.

[0039] S302: Cyclodextrin and alcohol extract are mixed at a mass-to-volume ratio of 1:45 and then concentrated under reduced pressure at a temperature of 60℃ to obtain concentrated alcohol extract.

[0040] S303: Soak the residue with 60g of kudzu root, 30g of white peony root, 30g of prepared licorice root, and 35g of jujube in 10 times the amount of water for 30 minutes, then heat and reflux for 1.5 hours to obtain the extract and residue a; add 8 times the amount of water to residue a, heat and reflux for 1 hour to obtain filtrate a and residue b; add 8 times the amount of water to residue b, heat and reflux for 1 hour to obtain filtrate b and residue; combine the extract, filtrate a, and filtrate b, concentrate under reduced pressure, and mix the concentrate with the alcohol extract concentrate to obtain Guizhi Jia Gegen Tang extract.

[0041] Comparative Example 1 This application provides a comparative example of an extraction method for Guizhi Jia Gegen Tang (Cinnamon Twig and Kudzu Root Decoction), the method comprising: Soak 60g of kudzu root, 30g of cinnamon twig, 30g of white peony root, 30g of prepared licorice root, 35g of jujube, and 45g of ginger in 10 times the amount of water for 30 minutes, then heat and reflux for 1.5 hours to obtain the extract and residue a. Add 8 times the amount of water to residue a and heat and reflux for 1 hour to obtain filtrate a and residue b. Add 8 times the amount of water to residue b and heat and reflux for 1 hour to obtain filtrate b and residue. Combine the extract, filtrate a, and filtrate b and concentrate under reduced pressure to obtain Guizhi Jia Gegen Tang extract.

[0042] The contents of cinnamic acid, cinnamaldehyde, and 6-gingerol in the Guizhi Jia Gegen Tang extract prepared in Example 1 and Comparative Example 1 were determined by high performance liquid chromatography according to the chromatographic conditions in "1. Volume concentration of ethanol". The comprehensive score was calculated by AHP weighted scoring method and the results are shown in Table 5.

[0043] Table 5: Content of volatile oil components in Example 1 and Comparative Example 1 As shown in Table 5, compared with Comparative Example 1, the content of cinnamic acid, a volatile oil component, in the extract of Guizhi Jia Gegen Tang extracted in Example 1 of this application is relatively similar, the content of cinnamaldehyde is significantly increased, and the content of 6-gingerol is slightly decreased. However, the average comprehensive score is greater than that of Comparative Example 1, which indicates that the effective components in the extract of Guizhi Jia Gegen Tang extracted in this application are significantly increased.

[0044] In addition, the embodiments of this application also use the Guizhi Jia Gegen Tang extract prepared in Example 1 and Comparative Example 1 to conduct a comparative efficacy experiment on mice with pneumonia-enteritis model infected with H1N1 influenza virus, as detailed below: 1. Preparation of an influenza mouse model Balb / c mice were randomly divided into four groups (n=6 per group) according to body weight and rectal temperature: normal group, model group, positive drug group, experimental group, and control group. Mice in the model group, positive drug group, experimental group, and control group received 20 µL of intranasal instillation and TCID50=1×10⁻⁶ dextrose instillation, respectively. 6 / mL of influenza A H1N1 / PR8 virus was administered to normal mice via intranasal inoculation with an equal volume of 0.9% sodium chloride solution.

[0045] 2. Drug administration intervention Equal masses of the Guizhi Jia Gegen Tang extract prepared in Example 1 and Comparative Example 1 were taken and added to water to prepare a Guizhi Jia Gegen Tang extract solution with a concentration of 1.1464 mg / mL. 28.66 mg of the contents of oseltamivir phosphate capsules were weighed, dissolved in water, and diluted to 25 mL to form a 1.1464 mg / mL oral administration drug.

[0046] Mice were weighed daily, and their water and food intake, fur condition, and mental state were observed. When the model group mice lost weight, the normal and model groups were administered 0.02 mL of distilled water daily by gavage, the positive control group was administered an equal volume of the drug daily by gavage, and the experimental and control groups were administered an equal volume of Guizhi Jia Gegen Tang extract solution prepared in Example 1 and Comparative Example 1 daily by gavage, based on the mice's weight. Administration was continued for 6 consecutive days, once daily.

[0047] 3. Mouse body weight, organ indices, lung tissue pathological morphology, viral load in the lungs, and relative gene expression of IL-1β and M genes. On day 7 of drug administration, the weight of the mice was recorded, and the following data were obtained. Figure 6 Mice were euthanized, and lung tissue, thymus, spleen, and colon were harvested. The mass of each organ was accurately weighed, and the organ index was calculated. The formula for calculating the organ index is: Index = Organ Mass (mg) / Body Mass (g). Figures 7-10In addition, RT-qPCR was used to detect viral load and the relative expression level of the inflammatory factor IL-1β gene in lung tissue. Viral load was visualized using the M gene. Figure 11 , 12 Simultaneously, H&E staining analysis was performed on lung tissue to compare the pathological damage of lung tissue in different groups of mice, obtaining the results. Figure 13 .

[0048] From the appendix Figure 6 As can be seen, compared with the model group, the mice in the normal group, positive drug group, experimental group, and control group all had higher body weights, and the body weight difference between the positive drug group and the experimental group was small. This indicates that the Guizhi Jia Gegen Tang extract extracted in Example 1 of this application can effectively maintain the body weight of mice infected with H1N1 influenza virus.

[0049] From the appendix Figure 7 As can be seen, compared with the model group, the lung index of mice in the normal group, positive drug group, experimental group, and control group was significantly reduced. The lung index of mice in the positive drug group and experimental group was similar and higher than that of the normal group. This indicates that the Guizhi Jia Gegen Tang extract extracted in Example 1 of this application can effectively reduce the lung index of mice infected with H1N1 influenza virus.

[0050] From the appendix Figure 8 As can be seen, compared with the model group, the thymus index of mice in the normal group, positive drug group, experimental group, and control group was significantly increased, and the thymus index of mice in the experimental group was higher than that of mice in the control group. This indicates that the Guizhi Jia Gegen Tang extract extracted in Example 1 of this application can effectively increase the thymus index of mice infected with H1N1 influenza virus.

[0051] From the appendix Figure 9 , 10 As can be seen, there were no significant differences in spleen index and colon length among the normal group, model group, positive drug group, experimental group, and control group mice. This indicates that H1N1 influenza virus infection in mice did not affect the spleen and colon.

[0052] From the appendix Figure 11 , 12 As can be seen, compared with the model group, the relative expression levels of the M gene and the inflammatory cytokine IL-1β gene were decreased in the positive drug group, experimental group, and control group mice. Furthermore, the relative expression levels of the M gene and IL-1β gene in the experimental group were significantly lower than those in the control group. This indicates that the Guizhi Jia Gegen Tang extract extracted in Example 1 of this application can effectively increase the relative expression levels of the M gene and the inflammatory cytokine IL-1β gene in H1N1 influenza virus-infected mice.

[0053] From the appendix Figure 13As can be seen, the lung tissue of the normal group mice showed normal morphology, with no degeneration, necrosis and shedding of the mucosal epithelium, no inflammatory cell infiltration in the pulmonary artery wall and surrounding tissue, and no thickening of the alveolar septa. In the model group mice, necrotic cells and a large amount of exudate were observed in the lung tissue, the surrounding lung tissue structure was unclear, inflammatory cell infiltration was present, and the alveolar walls were thickened. The inflammatory cell infiltration in the lung tissue of the positive control group, experimental group, and control group mice was improved, the alveolar septa returned to normal, and the overall degree of lesion was lower than that of the model group.

[0054] As can be seen from the above, the extract of Guizhi Jia Gegen Tang obtained by the extraction method of Guizhi Jia Gegen Tang provided in this application embodiment can effectively maintain the body weight of mice infected with H1N1 influenza virus, reduce the lung index of mice infected with H1N1 influenza virus, and increase the thymus index of mice infected with H1N1 influenza virus.

[0055] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. A method for extracting Guizhi Jia Gegen Tang (Cinnamon Twig and Kudzu Root Decoction), characterized in that, include: Cinnamon twigs and ginger were soaked in ethanol and extracted by heating and reflux to obtain an ethanol extract and residue. Cyclodextrin was added to the alcohol extract, and the mixture was concentrated under reduced pressure to obtain a concentrated alcohol extract. The residue, along with kudzu root, white peony root, prepared licorice root, and jujube, was soaked in water and extracted three times by heating and reflux. The filtrates were combined, concentrated, and then mixed with the alcohol extract concentrate to obtain Guizhi Jia Gegen Tang extract.

2. The extraction method of Guizhi Jia Gegen Tang according to claim 1, characterized in that, The volume concentration of ethanol is 70-90%, and the mass-to-volume ratio of the total mass of cinnamon twigs and ginger to ethanol is 1:6-8.

3. The extraction method of Guizhi Jia Gegen Tang according to claim 1, characterized in that, The ethanol soaking time is 30-50 min, the reflux temperature is 80-100℃, the reflux time is 90-120 min, and the reflux method is total reflux.

4. The extraction method of Guizhi Jia Gegen Tang according to claim 1, characterized in that, The mass-to-volume ratio of the cyclodextrin to the alcohol extract is 1:

45.

5. The extraction method of Guizhi Jia Gegen Tang according to claim 1, characterized in that, The temperature for vacuum concentration is 45-60℃.

6. The extraction method of Guizhi Jia Gegen Tang according to claim 1, characterized in that, The residue, along with kudzu root, white peony root, prepared licorice root, and jujube, was soaked in water and extracted three times by reflux. The filtrates were combined, concentrated, and then mixed with the concentrated liquid to obtain Guizhi Jia Gegen Tang extract, which includes: The residue, along with kudzu root, white peony root, prepared licorice root, and jujube, is soaked in 8-10 times the amount of water for 30-50 minutes, then heated under reflux for 1.5 hours to obtain the extract and residue a. The dregs a were added to 8 times the amount of water and heated under reflux for 1 hour to obtain filtrate a and dregs b. The dregs b were added to 8 times the amount of water and heated under reflux for 1 hour to obtain filtrate b and dregs. The extract, filtrate a, and filtrate b are combined and concentrated under reduced pressure. The resulting concentrate is then mixed with the alcohol extract concentrate to obtain Guizhi Jia Gegen Tang extract.

7. The extraction method of Guizhi Jia Gegen Tang according to claim 6, characterized in that, The heating reflux temperature is 80-100℃, and the reflux method is full reflux.

8. The extraction method of Guizhi Jia Gegen Tang according to claim 1, characterized in that, The ingredients for preparing Guizhi Jia Gegen Tang (Cinnamon Twig and Kudzu Root Decoction) include, by weight, 60g of kudzu root, 30g of cinnamon twig, 30g of white peony root, 30g of prepared licorice root, 35g of jujube, and 45g of ginger.

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