A pueraria and perilla composite extract, a preparation method and applications thereof

Abstract

The present application relates to the technical field of extraction processing of pueraria and perilla, and specifically discloses a pueraria and perilla compound extract, a preparation method and application thereof. The preparation method is as follows: after pueraria is sliced, the pueraria and perilla are cleaned and dried together, the water content is adjusted by adding water, and a fermenting agent is added to ferment the mixture to a specific pH value to obtain a fermented product. Then, the fermented product is extracted by using a low eutectic solvent to obtain a low eutectic solvent extract. The extract is mixed with ethanol and then extracted by supercritical carbon dioxide to obtain a supercritical carbon dioxide extract. Then, the extract is treated by water extraction and alcohol precipitation to obtain a preliminarily purified extract. Finally, the preliminarily purified extract is separated by using a macroporous resin to obtain the final pueraria and perilla compound extract. The method significantly improves the extraction rate of flavonoids and other bioactive substances in the compound extract, and the compound extract has excellent effects in reducing blood pressure, blood sugar and blood lipid, and can be used as a daily dietary supplement and widely applied in the fields of food, medicine and health care products.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of extraction and processing of pueraria and perilla, and specifically relates to a pueraria and perilla composite extract, a preparation method and applications thereof. BACKGROUND

[0002] In recent years, with the continuous improvement of consumers' health awareness, functional foods, as a category of foods that can provide specific health benefits, have gradually become a market hotspot. Functional foods not only meet people's basic nutritional needs, but also help prevent chronic diseases and regulate body functions by adding ingredients with specific physiological functions, such as plant extracts. For example, in daily baked foods such as biscuits, bread, and other foods, consumers are increasingly inclined to choose functional products with added natural plant extracts. These natural products not only retain the deliciousness of traditional foods, but also have high safety and additional health values, such as regulating blood lipids, blood sugar, and blood pressure.

[0003] Hypertension, hyperglycemia, and hyperlipidemia are common metabolic diseases in modern society, posing a serious threat to people's health. Pueraria and perilla are two traditional Chinese medicinal materials commonly used in traditional Chinese medicine in China, which have certain effects of lowering blood pressure, blood sugar, and blood lipids. Currently, there have been some studies on the extraction methods of pueraria and perilla, but how to improve the extraction efficiency and the efficacy of the composite extract is still a problem to be solved.

[0004] The existing extraction technology of pueraria and perilla mainly includes traditional solvent extraction, ultrasonic extraction, microwave extraction, etc. These methods can extract effective components from pueraria and perilla to some extent, but have the following defects:

[0005] (1) The traditional extraction method is not complete in extracting effective components, resulting in low content of effective components in the extract, which affects its effects of lowering blood pressure, blood sugar, and blood lipids.

[0006] (2) Some extraction methods require high temperature and pressure, which may cause the structure of effective components to be damaged, reducing their activity.

[0007] (3) Some extraction methods use organic solvents, which may pollute the environment and have certain toxicity to the human body.

[0008] Therefore, a pueraria and perilla composite extract, a preparation method and applications thereof are proposed to solve the above problems. By optimizing the extraction process, the effective components in pueraria and perilla are fully extracted, and a composite extract with high efficiency in lowering blood pressure, blood sugar, and blood lipids is prepared, providing new ideas and solutions for the development of the functional food industry. SUMMARY

[0009] The present invention aims to address the problem that the existing extraction methods often fail to fully extract the effective components from kudzu root and perilla, resulting in low content of effective components in the extract, unstable purity and content, and unstable product efficacy, which affects its effects on lowering blood pressure, blood sugar, and blood lipids. Traditional extraction and purification methods often involve cumbersome steps and high production costs.

[0010] To achieve the above objectives, the present invention provides the following technical solution:

[0011] A method for preparing a compound extract of kudzu root and perilla leaves includes the following steps:

[0012] S1. Wash and slice kudzu root and perilla separately, dry them until the moisture content is less than 10wt%, put them into a fermentation tank, add water to adjust the moisture content to 45wt%-55wt% to obtain the fermentation substrate, add fermentation agent, ferment at 40-45℃ until the pH is 4-4.5, take it out, dehydrate and concentrate to obtain the fermented product.

[0013] The starter culture includes Lactobacillus bulgaricus, Candida lipolyticis, Streptococcus thermophilus, and Lactobacillus casei; the strain number of Lactobacillus bulgaricus is CICC 25031; the strain number of Candida lipolyticis is CICC 32245; the strain number of Streptococcus thermophilus is CICC 6038; and the strain number of Lactobacillus casei is CICC 20241.

[0014] S2. Extract the fermentation product by reflux with a eutectic solvent 2-3 times, 1-2 hours each time, filter, combine the extracts, separate the eutectic solvent, and concentrate to obtain the eutectic solvent extract.

[0015] S3. Mix the eutectic solvent extract with a 90%-95% ethanol solution, stir well, and then put it into a supercritical carbon dioxide extractor for supercritical extraction. Recover the ethanol, collect the extract and concentrate it to obtain the supercritical carbon dioxide extract.

[0016] S4. The supercritical carbon dioxide extract is subjected to water extraction and alcohol precipitation to obtain a preliminarily purified extract.

[0017] S5. The preliminarily purified extract is separated using macroporous resin to obtain a compound extract of kudzu root and perilla.

[0018] As a preferred option, the dosage of Lactobacillus bulgaricus is 10. 6 -10 7 CFU / g fermentation substrate; the amount of *Candida lipolyticis* used was 10. 8 -10 9 CFU / g fermentation substrate; the amount of thermophilic streptococci used was 10. 7 -10 8CFU / g fermentation substrate; Lactobacillus casei dosage was 10 5 -10 6 CFU / g fermentation substrate.

[0019] As a preferred option, the mass ratio of kudzu root to perilla is (5-7):(3-5).

[0020] Preferably, the mass ratio of fermentation product to eutectic solvent is 1:(30-40), and the water content of eutectic solvent is 25wt%-35wt%.

[0021] Preferably, the components of the eutectic solvent include betaine, glycerol, lactic acid, urea, citric acid, and water.

[0022] As a preferred option, the molar ratio of betaine, glycerol, lactic acid, urea, and citric acid is 1:(1-3):(0.5-2):(0.8-1.2):(0.5-2).

[0023] Preferably, the mass ratio of the eutectic solvent extract to the 90%-95% ethanol solution is 1:(30-40), and the method for separating the eutectic solvent is bipolar electrodialysis membrane separation.

[0024] As a preferred method, the conditions for supercritical extraction are: carbon dioxide dosage is 2-4 times that of 90%-95% ethanol solution, pressure is 30-40 MPa, temperature is 35-45℃, and time is 1-2 h.

[0025] As a preferred method, the specific steps for water extraction and alcohol precipitation are as follows:

[0026] Add the supercritical carbon dioxide extract to 8-12 times the volume of water, heat to 80-90℃, extract for 1-2 hours, filter, and collect the extract;

[0027] Cool the extract to room temperature, add 95%-98% ethanol to a concentration of 60%-70%, stir well, let stand for 12-24 hours, centrifuge, collect the precipitate, wash the precipitate successively with water, 50% ethanol solution, and 95% ethanol solution until neutral, evaporate the ethanol solution, and dry at 55-65℃ to obtain the preliminarily purified extract.

[0028] As a preferred method, the specific steps for macroporous resin separation are as follows: the preliminarily purified extract is dissolved in water and passed through macroporous resin at a flow rate of 2.5-3.5 BV / h, and eluted sequentially with 3-5 BV of water and 2-4 BV of 45%-55% ethanol solution. The eluent is collected, concentrated and dried under reduced pressure to obtain the kudzu root and perilla compound extract.

[0029] Preferably, the mass ratio of the pre-purified extract to water is 1:(10-30).

[0030] Preferably, the macroporous resin is a styrene-type nonpolar macroporous resin with an average pore size of 12-16 nm, a porosity of 45%-50%, and a specific surface area of ​​450-500 m² / g. The macroporous resin was purchased from Tianjin Haoju Resin Technology Co., Ltd., model ADS-8.

[0031] This application also provides a kudzu root and perilla compound extract, which is prepared by the above-described method for preparing the kudzu root and perilla compound extract.

[0032] This application also provides an application of the above-mentioned kudzu root and perilla compound extract in the preparation of functional products for regulating blood pressure, blood sugar and blood lipids.

[0033] Before extracting kudzu root and perilla, fermentation with specific fermenting agents can improve the extraction efficiency of active ingredients, thereby enhancing the efficacy of the compound extract. This is because these fermenting agents work synergistically. *Lactobacillus bulgaricus* and *Streptococcus thermophilus* produce lactic acid, lowering the pH of the fermentation broth, inhibiting the growth of harmful microorganisms, protecting flavonoids from oxidative damage, and promoting the dissolution and stability of flavonoids. *Candida lipolytica* and *Lactobacillus casei* produce various enzymes, such as β-glucosidase, protease, cellulase, and amylase. These enzymes can break down cell walls and macromolecules, releasing more active ingredients, such as flavonoids and polysaccharides from perilla and kudzu root. Furthermore, these fermenting bacteria also have antioxidant effects, protecting the stability of flavonoids, while producing organic acids and alcohols, improving the flavor and aroma of the raw materials, and enhancing the sensory quality of the final product. Through these synergistic effects, the extraction rate and purity of active ingredients can be significantly improved, thus enhancing the efficacy of the compound extract.

[0034] A compound extract from kudzu root and perilla leaves, when used in functional foods, not only helps control hypertension, hyperglycemia, and hyperlipidemia, but also has a positive impact on improving cardiovascular health and can be used as a daily dietary supplement. This is because kudzu root and perilla are rich in flavonoids (such as puerarin, daidzein, apigenin, luteolin, etc.), isoflavones, polysaccharides, saponins, rosmarinic acid, perillaldehyde, and other components, which work synergistically.

[0035] First, puerarin has a natural β-receptor antagonistic effect, which can block the stimulation of adenylate cyclase by adrenaline, thereby counteracting the pressor effect and achieving a blood pressure lowering effect. Simultaneously, puerarin and daidzein can also block α-receptors, lowering blood pressure. Meanwhile, the flavonoids in perilla indirectly affect blood pressure levels by regulating the contraction and relaxation of vascular smooth muscle and exerting antioxidant and anti-inflammatory effects, maintaining vascular health and promoting blood pressure stability. Both work synergistically in the blood pressure regulation system to enhance the blood pressure lowering effect. Second, puerarin can counteract the hyperglycemic effect of adrenaline, enhance insulin sensitivity, and correct hyperinsulinemia, thereby lowering blood sugar. The flavonoids in perilla indirectly affect blood sugar levels by regulating insulin secretion and utilization. Both work synergistically on the insulin signaling pathway, enhancing insulin sensitivity, lowering blood sugar levels, and reducing the risk of metabolic diseases such as diabetes.

[0036] Furthermore, puerarin can lower total cholesterol and LDL cholesterol levels, reducing fat deposition on arterial walls, while the flavonoids and other active ingredients in perilla have lipid-lowering effects. Together, they act on the lipid metabolism pathway, promoting fat breakdown and utilization, lowering blood lipid levels, and further reducing the risk of arteriosclerosis. Finally, isoflavones in pueraria help dilate coronary arteries, improve cardiovascular circulation, and prevent cardiovascular disease. The active ingredients in perilla help reduce inflammation and oxidative stress in blood vessel walls, maintaining vascular health. Both work together on the cardiovascular system, improving cardiovascular health and reducing the risk of cardiovascular disease through mechanisms such as vasodilation, inhibition of oxidative stress, and inflammation. Therefore, the higher the concentration of active ingredients in the pueraria and perilla compound extract, the more significant its effects in lowering blood pressure, blood sugar, and blood lipids.

[0037] Using betaine, glycerol, lactic acid, urea, and citric acid as a eutectic solvent to extract active substances from perilla and kudzu root can improve the selectivity and efficiency of extraction. This is because the flavonoids and other active substances in both raw materials typically possess a certain degree of polarity and hydrophobicity. In the eutectic solvent composed of betaine, glycerol, lactic acid, urea, and citric acid, betaine and urea stabilize the solvent structure by forming a strong hydrogen bond network, while glycerol and lactic acid optimize extraction conditions by dissolving and adjusting the pH. The polycarboxyl groups of citric acid can interact with flavonoids and other active substances, improving the selectivity and efficiency of extraction. Simultaneously, the strong hydrogen bond interaction between urea and betaine binds the urea molecules, making it difficult for them to diffuse into the lipase molecules, thus reducing the impact on the activity of the active substances and the extraction efficiency. Compared with traditional ethanol extraction, the eutectic solvent described in this invention has higher solubility, better stability, and higher selectivity for effective substances. Furthermore, the bipolar electrodialysis membrane separation technology removes the eutectic solvent while recovering its effective components, greatly improving extraction efficiency and product purity, thereby enhancing its effects in lowering blood pressure, blood sugar, and blood lipids.

[0038] Using a specific macroporous resin for secondary separation and purification of the initially purified extract can further improve the purity and content of active ingredients in the composite extract. This is likely because ADS-8, as a styrene-type nonpolar adsorption resin, has certain ester groups on its surface, making it hydrophobic. Therefore, it can effectively adsorb flavonoids, isoflavones, and other active ingredients in kudzu root and perilla. The polar groups on its surface can also interact with the polar parts of molecules such as rosmarinic acid. Simultaneously, it does not adsorb sugars, inorganic acids, alkalis, salts, or small-molecule hydrophilic organic compounds, achieving selective separation and thus increasing the content of active substances in the composite extract. By controlling the ethanol concentration during elution, flavonoids and other active substances can be eluted from the resin, avoiding the structural damage caused by excessively high ethanol concentrations and the poor elution effect caused by excessively low concentrations, ensuring the purity and yield of active substances in the composite extract.

[0039] Compared with the prior art, the technical effects and advantages of the present invention are:

[0040] (1) This invention provides a kudzu root and perilla compound extract and its preparation method. Through the optimized combination of fermentation, low eutectic solvent extraction, supercritical carbon dioxide extraction, water extraction and alcohol precipitation and macroporous resin separation and purification, the extraction rate of flavonoids and other bioactive substances in the compound extract is significantly improved. The prepared compound extract not only helps to control hypertension, hyperglycemia and hyperlipidemia, but also has a positive effect on improving cardiovascular health. The raw materials are derived from plants and are non-toxic and harmless, and can be used as a daily dietary supplement.

[0041] (2) Before extracting kudzu root and perilla, the present invention first uses a specific fermentation agent to ferment them, which can improve the extraction efficiency of the effective components in kudzu root and perilla, thereby improving the efficacy of the compound extract.

[0042] (3) The present invention uses betaine, glycerol, lactic acid, urea and citric acid as eutectic solvents to extract the effective substances in perilla and kudzu root, which can improve the selectivity and extraction efficiency of the extraction.

[0043] (4) The present invention uses a specific macroporous resin to perform secondary separation and purification on the preliminarily purified extract, which can further improve the purity and content of the effective components in the composite extract. Detailed Implementation

[0044] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0045] Example 1

[0046] This embodiment provides a compound extract of kudzu root and perilla, the preparation method of which includes the following steps:

[0047] S1. After washing and slicing kudzu root and perilla, dry them to a moisture content of 10 wt%. Place them in a fermentation tank, add water to adjust the total moisture content of the material to 50 wt% to obtain the fermentation substrate, add the fermentation agent, ferment at 44℃ until the pH reaches 4, take it out, dehydrate and concentrate it to obtain the fermented product.

[0048] The starter culture includes *Lactobacillus bulgaricus*, *Candida lipolyticis*, *Streptococcus thermophilus*, and *Lactobacillus casei*. All starter cultures are commercially available. The strain number for *Lactobacillus bulgaricus* is CICC 25031; the strain number for *Candida lipolyticis* is CICC32245; the strain number for *Streptococcus thermophilus* is CICC 6038; and the strain number for *Lactobacillus casei* is CICC 20241.

[0049] S2. The fermentation product was refluxed with a eutectic solvent three times, each time for 1.5 hours. The extracts were filtered, combined, and the eutectic solvent was separated and concentrated to obtain the eutectic solvent extract.

[0050] S3. Mix the eutectic solvent extract and 90% ethanol solution, stir well, and then put it into a supercritical carbon dioxide extractor for supercritical extraction. Recover the ethanol, collect the extract and concentrate it to obtain the supercritical carbon dioxide extract.

[0051] S4. The supercritical carbon dioxide extract is subjected to water extraction and alcohol precipitation to obtain a preliminarily purified extract.

[0052] S5. The preliminarily purified extract is separated using macroporous resin to obtain a compound extract of kudzu root and perilla.

[0053] The mass ratio of kudzu root to perilla is 3:2.

[0054] The starter culture consists of Lactobacillus bulgaricus, Candida lipolytica, Streptococcus thermophilus, and Lactobacillus casei.

[0055] The dosage of Lactobacillus bulgaricus is 10. 6 CFU / g fermentation substrate; the amount of *Candida lipolyticis* used was 10. 8 CFU / g fermentation substrate; the amount of thermophilic streptococci used was 10. 7 CFU / g fermentation substrate; Lactobacillus casei dosage was 10 5 CFU / g fermentation substrate.

[0056] The mass ratio of fermentation product to eutectic solvent is 1:35.

[0057] The components of the eutectic solvent are betaine, glycerol, lactic acid, urea, citric acid, and water.

[0058] The eutectic solvent has a water content of 30%.

[0059] The molar ratio of betaine, glycerol, lactic acid, urea, and citric acid is 1:2:1:1:1.

[0060] The mass ratio of the eutectic solvent extract to the 90% ethanol solution is 1:35.

[0061] The method for separating eutectic solvents is bipolar electrodialysis membrane separation, and the specific implementation method is described in Example 1 of the patent authorization announcement number "CN115531490B".

[0062] The conditions for supercritical extraction were: carbon dioxide dosage was 3 times that of 90% ethanol solution, pressure was 35 MPa, temperature was 40℃, and time was 1.5 h.

[0063] The specific steps for water extraction and alcohol precipitation are as follows:

[0064] Add the supercritical carbon dioxide extract to 10 times the volume of water, heat to 85°C, extract for 1.5 hours, filter, and collect the extract;

[0065] Cool the extract to room temperature, add 95% ethanol to a concentration of 65%, stir well, let stand for 20 hours, centrifuge, collect the precipitate, wash the precipitate successively with water, 50% ethanol solution, and 95% ethanol solution until neutral, evaporate the ethanol solution, and dry at 60°C to obtain the preliminarily purified extract.

[0066] The specific steps for macroporous resin separation are as follows: the preliminarily purified extract is dissolved in water and passed through macroporous resin at a flow rate of 3 BV / h. It is then eluted sequentially with 4 BV of water and 3 BV of 50% ethanol solution. The eluent is collected, concentrated and dried under reduced pressure to obtain the kudzu root and perilla compound extract.

[0067] The mass ratio of the preliminarily purified extract to water was 1:20.

[0068] The macroporous resin is a styrene-type non-polar macroporous resin with an average pore size of 12-16 nm, a porosity of 45%-50%, and a specific surface area of ​​450-500 m² / g. It was purchased from Tianjin Haoju Resin Technology Co., Ltd., ADS-8.

[0069] Example 2

[0070] Based on Example 1, the difference between this example and Example 1 is that the dosage of Lactobacillus bulgaricus is 10. 7 CFU / g fermentation substrate; the amount of *Candida lipolyticis* used was 10. 9 CFU / g fermentation substrate; the amount of thermophilic streptococci used was 10. 8 CFU / g fermentation substrate; Lactobacillus casei dosage was 10 6 CFU / g fermentation substrate.

[0071] Example 3

[0072] Based on Example 1, the difference between this example and Example 1 is that the molar ratio of betaine, glycerol, lactic acid, urea, and citric acid is 1:1:2:1:2.

[0073] Comparative Example 1

[0074] The difference between this comparative example and Example 1 is that the starter culture is Lactobacillus bulgaricus, Candida lipolyticis, and Streptococcus thermophilus.

[0075] The dosage of Lactobacillus bulgaricus is 10. 6 CFU / g fermentation substrate; the amount of *Candida lipolyticis* used was 10. 8 CFU / g fermentation substrate; the amount of thermophilic streptococci used was 10. 7 CFU / g fermentation substrate.

[0076] Comparative Example 2

[0077] The difference between this comparative example and Example 1 is that the amount of Lactobacillus bulgaricus used is 10. 5 CFU / g fermentation substrate; the amount of *Candida lipolyticis* used was 10. 6 CFU / g fermentation substrate; the amount of thermophilic streptococci used was 10. 6CFU / g fermentation substrate; Lactobacillus casei dosage was 10 4 CFU / g fermentation substrate.

[0078] Comparative Example 3

[0079] The difference between this comparative example and Example 1 is as follows: S2, the fermentation product was refluxed with 70% ethanol aqueous solution three times, 1.5h each time, filtered, the extracts were combined and concentrated to obtain the extract.

[0080] Comparative Example 4

[0081] The difference between this comparative example and Example 1 is that the components of the eutectic solvent are glycerol, lactic acid, urea, citric acid, and water.

[0082] The molar ratio of glycerol, lactic acid, urea, and citric acid is 2:1:1:1.

[0083] Comparative Example 5

[0084] The difference between this comparative example and Example 1 is that the macroporous resin is a polar macroporous resin with an average pore size of 1.3-1.4 nm and a specific surface area of ​​480-520 m² / g.

[0085] The macroporous resin was purchased from Bengbu Sanyi Technology Co., Ltd., model A-8.

[0086] Comparative Example 6

[0087] The difference between this comparative example and Example 1 is as follows: The specific steps of macroporous resin separation are as follows: the preliminarily purified extract is dissolved in water and passed through macroporous resin at a flow rate of 3 BV / h. It is then eluted sequentially with 4 BV of water and 3 BV of 80% ethanol solution. The eluent is collected, concentrated and dried under reduced pressure to obtain the kudzu root and perilla compound extract.

[0088] Comparative Example 7

[0089] The difference between this comparative example and Example 1 is that the preparation method of the kudzu root and perilla compound extract is as follows:

[0090] S1. After washing and slicing kudzu root and perilla, dry them until the moisture content is less than 10 wt%. Put them into a fermentation tank, add water to adjust the moisture content to 50 wt% to obtain the fermentation substrate, add fermentation agent, ferment at 44℃ until the pH is 4, take it out, dehydrate and concentrate to obtain the fermented product.

[0091] S2. The fermentation product was refluxed with a eutectic solvent three times, each time for 1.5 hours. The extracts were filtered, combined, and the eutectic solvent was separated and concentrated to obtain the eutectic solvent extract.

[0092] S3. The eutectic solvent extract is subjected to water extraction and alcohol precipitation to obtain a preliminarily purified extract.

[0093] S4. The preliminarily purified extract is separated using macroporous resin to obtain a compound extract of kudzu root and perilla.

[0094] Performance testing

[0095] Referring to the method in patent CN114306544B, a mouse experiment was conducted to test the hypoglycemic effect of the tea bag. Mice were fed a high-fat, high-sugar, and high-salt diet for 10 weeks to ensure that the fasting blood glucose level in all mice increased by more than 4 times. The mice were then administered the drug by gavage at a daily oral gavage dose of 1 g / kg·Bw. The changes in fasting blood glucose and cholesterol levels in the mice were tested after 30 days.

[0096] Blood glucose reduction rate = (pre-test blood glucose - post-test blood glucose) / pre-test blood glucose × 100%;

[0097] Cholesterol reduction rate = (pre-test cholesterol - post-test cholesterol) / pre-test cholesterol × 100%.

[0098] Referring to the method in the paper "The antihypertensive effect of kudzu root fermentation liquid on essential hypertensive rats" published by Su Lei et al., essential hypertensive rats were selected, and mice were administered the drug by gavage at a daily oral gavage dose of 1 g / kg·Bw. After 30 days, the diastolic and systolic blood pressure of the mice were tested.

[0099] Diastolic blood pressure decrease rate = (Pre-test diastolic blood pressure - Post-test diastolic blood pressure) / Pre-test diastolic blood pressure × 100%;

[0100] Systolic blood pressure drop rate = (pre-test systolic blood pressure - post-test systolic blood pressure) / pre-test systolic blood pressure × 100%.

[0101] The results are shown in Table 1.

[0102] Table 1 Measurement Results

[0103]

[0104] According to statistics, the kudzu root and perilla compound extracts prepared in Examples 1-3 of this invention showed high reduction rates in fasting blood glucose, cholesterol, diastolic blood pressure, and systolic blood pressure, indicating excellent effects in lowering blood sugar, blood lipids, and blood pressure. Comparative Example 1 did not add Lactobacillus casei; Comparative Example 2 used low amounts of each fermentation agent; Comparative Example 3 did not use a eutectic solvent for extraction; Comparative Example 4 did not add betaine; Comparative Example 5 did not use a specific macroporous resin; Comparative Example 6 had excessively high ethanol concentration during elution; and Comparative Example 7 did not undergo supercritical extraction. The kudzu root and perilla compound extracts prepared in these examples showed poor reduction rates in fasting blood glucose, cholesterol, diastolic blood pressure, and systolic blood pressure. Therefore, the kudzu root and perilla compound extracts prepared using the method of this application help control hypertension, hyperglycemia, and hyperlipidemia, and have a positive impact on improving cardiovascular health. The raw materials are derived from plants, are non-toxic and harmless, and can be used as a daily dietary supplement.

[0105] This invention significantly improves the extraction rate of flavonoids and other bioactive substances in the compound extract through an optimized combination of fermentation, eutectic solvent extraction, supercritical carbon dioxide extraction, water extraction and alcohol precipitation, and macroporous resin separation and purification. This means that more active ingredients can be obtained from kudzu root and perilla, thereby enhancing the product's efficacy. Fermentation treatment can improve the extraction efficiency of active ingredients from kudzu root and perilla, thus enhancing the efficacy of the compound extract. The fermentation agent not only protects the active ingredients but may also generate new beneficial components, making the compound extract more effective in controlling hypertension, hyperglycemia, and hyperlipidemia.

[0106] The use of a eutectic solvent composed of betaine, glycerol, lactic acid, urea, and citric acid improves the selectivity and efficiency of extraction. This solvent better dissolves and stabilizes the target components while reducing the extraction of unnecessary components, thus increasing the purity of the final product.

[0107] The use of eutectic solvents reduces reliance on traditional organic solvents, thus lowering environmental impact. Simultaneously, the extremely low water content of the bipolar solvents makes separation more efficient, reducing energy consumption and costs.

[0108] Secondary separation and purification using a specific macroporous resin further improved the purity and content of the active ingredients in the composite extract. This selective adsorption not only removed impurities but also ensured that the structure of the active ingredients was not damaged, thereby improving the product yield and stability.

[0109] The compound extract is rich in various bioactive components, such as puerarin and flavonoids, which have a positive effect on improving cardiovascular health, helping to dilate blood vessels, lower blood pressure, and regulate blood sugar and blood lipids. The raw materials are derived from plants, are non-toxic and harmless, and are suitable for use as a daily dietary supplement, increasing consumer confidence and acceptance.

[0110] The kudzu root and perilla compound extract provided by this invention is suitable for functional foods and other fields, and has broad application prospects.

[0111] In summary, this invention has significant technical effects and advantages in improving extraction efficiency, enhancing product efficacy, promoting environmentally friendly production, and increasing product purity and yield.

[0112] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.

Claims

1. A method for preparing a compound extract of kudzu root and perilla, characterized in that, Includes the following steps: S1. Wash and slice kudzu root and perilla separately, dry them until the moisture content is less than 10 wt%, put them into a fermentation tank, add water to adjust the moisture content to 45%-55 wt% to obtain the fermentation substrate, add fermentation agent, ferment at 40-45℃ until the pH is 4-4.5, take it out, dehydrate and concentrate to obtain the fermented product. The starter culture includes Lactobacillus bulgaricus, Candida lipolyticis, Streptococcus thermophilus, and Lactobacillus casei; the strain number of Lactobacillus bulgaricus is CICC25031; the strain number of Candida lipolyticis is CICC32245; the strain number of Streptococcus thermophilus is CICC6038; and the strain number of Lactobacillus casei is CICC20241. S2. Extract the fermentation product by reflux with a eutectic solvent 2-3 times, 1-2 hours each time, filter, combine the extracts, separate the eutectic solvent, and concentrate to obtain the eutectic solvent extract. S3. Mix the eutectic solvent extract with a 90%-95% ethanol solution, stir well, and then put it into a supercritical carbon dioxide extractor for supercritical extraction. Recover the ethanol, collect the extract and concentrate it to obtain the supercritical carbon dioxide extract. The conditions for supercritical extraction are: carbon dioxide dosage is 2-4 times that of 90%-95% ethanol solution, pressure is 30-40 MPa, temperature is 35-45℃, and time is 1-2 h. S4. The supercritical carbon dioxide extract is subjected to water extraction and alcohol precipitation to obtain a preliminarily purified extract. The specific steps of the water extraction and alcohol precipitation are as follows: Add the supercritical carbon dioxide extract to 8-12 times the volume of water, heat to 80-90℃, extract for 1-2 hours, filter, and collect the extract; Cool the extract to room temperature, add 95%-98% ethanol to the ethanol concentration of 60%-70%, stir well, let stand for 12-24 hours, centrifuge, collect the precipitate, wash the precipitate with water, 50% ethanol solution and 95% ethanol solution in sequence until neutral, evaporate the ethanol solution, dry at 55-65℃ to obtain the preliminarily purified extract. S5. The preliminarily purified extract is separated using macroporous resin to obtain a compound extract of kudzu root and perilla. The specific steps of macroporous resin separation are as follows: the preliminarily purified extract is dissolved in water and passed through macroporous resin at a flow rate of 2.5-3.5 BV / h. It is then eluted sequentially with 3-5 BV of water and 2-4 BV of 45%-55% ethanol solution. The eluent is collected, concentrated and dried under reduced pressure to obtain the kudzu root and perilla compound extract. The macroporous resin is a styrene-type nonpolar macroporous resin with an average pore size of 12-16 nm, a porosity of 45%-50%, and a specific surface area of ​​450-500 m² / g. The dosage of Lactobacillus bulgaricus is 10. 6 -10 7 CFU / g fermentation substrate; the amount of *Candida lipolyticis* used was 10. 8 -10 9 CFU / g fermentation substrate; the amount of thermophilic streptococci used was 10. 7 -10 8 CFU / g fermentation substrate; Lactobacillus casei dosage was 10 5 -10 6 CFU / g fermentation substrate; The components of the eutectic solvent include betaine, glycerol, lactic acid, urea, citric acid, and water; The molar ratio of betaine, glycerol, lactic acid, urea, and citric acid is 1:(1-3):(0.5-2):(0.8-1.2):(0.5-2).

2. A compound extract of kudzu root and perilla, characterized in that, The kudzu root and perilla compound extract was prepared by the method described in claim 1.

3. The application of the kudzu root and perilla compound extract according to claim 2 in the preparation of functional foods that help lower blood pressure, blood sugar and blood lipids.

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