Houttuynia cordata extract and preparation method thereof

Abstract

The preparation method comprises the following steps: S1, crushing cordate houttuynia rhizomes, adding the crushed cordate houttuynia rhizomes into a mixed acid aqueous solution, and carrying out immersion treatment to obtain a cordate houttuynia mixed solution; s2, adjusting the pH value of the herba houttuyniae mixed solution to 4-5, adding pectinase, carrying out primary enzymolysis and enzyme deactivation, adjusting the pH value to 6-7, adding compound enzyme, carrying out secondary enzymolysis and enzyme deactivation, so as to obtain an enzymatic hydrolysate; s3, menthol is added into the enzymatic hydrolysate, water extraction is conducted, and water extract and filter residues are obtained; s4, the filter residues are added into ethyl alcohol, then salicylic acid and diacetyl tartaric acid ester of mono (di) glycerides are added, alcohol extraction and filtering are conducted, and an alcohol extraction solution is obtained; s5, mixing the water extract with the alcohol extract, and performing reduced pressure distillation to obtain the herba houttuyniae extract. According to the method, the extraction rate of polysaccharide and flavone in houttuynia cordata can be effectively increased, and meanwhile, the prepared houttuynia cordata extract has better oxidation resistance and has a good application prospect.

Description

Technical Field

[0001] The present invention belongs to the technical field of plant extracts, and particularly relates to a houttuynia cordata extract and a preparation method thereof. Background Art

[0002] Houttuynia cordata, also known as houttuynia, cenchrus grass, heartleaf houttuynia herb, and stinking houttuynia, etc., is the whole herb of the houttuynia plant of Saururaceae, and is a perennial herb. Houttuynia cordata is mainly distributed in the central, southeastern and southwestern provinces south of the Yangtze River in China, especially in Sichuan, Hubei, Zhejiang, Fujian, Guangxi, Guizhou and other places.

[0003] Houttuynia cordata is a plant resource with great development potential that "is both a medicine and a food". The chemical constituents of houttuynia cordata are complex, and the main compounds include flavonoid compounds, alkaloids, volatile oils, polysaccharides, terpenoids and sterols, etc. Modern pharmacological studies have shown that houttuynia cordata extract has various biological activities such as anti-inflammatory, antiviral, anti-allergic, antibacterial, antioxidant, anti-tumor and immunomodulatory effects. Houttuynia cordata is mainly used in clinical treatment of dysentery, cold and fever, epidemic parotitis, acute icteric hepatitis and other diseases, and compound houttuynia cordata tablets, houttuynia cordata eye drops, etc. have been widely used in clinical practice.

[0004] However, other bioactive components in houttuynia cordata, such as flavonoids and water-soluble polysaccharides, have rarely attracted people's attention. Flavonoids mainly refer to compounds with 2-phenylchromone as the nuclear group, and usually exist in many plants in the form of free glycosides or glycosides combined with sugars. Flavonoid compounds are good scavengers of active oxygen free radicals. In today's emerging natural health foods, a large part is related to anti-aging and scavenging excessive free radicals in the body, so flavonoids may be the bioactive factors that play a role in them. Polysaccharides are widely present in animal, plant and microbial tissues, and are a class of natural macromolecular compounds composed of various monosaccharides. Various types of cellulose, pectin, mucilage, etc. have the functions of structural polysaccharides. Polysaccharide substances are organic compounds with a broad-spectrum chemical structure and biological functions existing in nature, widely present in plants, animals and microorganisms, and have effects such as regulating immunity, anti-tumor, antiviral, reducing blood sugar and blood lipids, and anti-aging.

[0005] Chinese patent application CN108530548A discloses a method for preparing a polysaccharide extract of Houttuynia cordata, comprising the following steps: (1) crushing fresh Houttuynia cordata, adding purified water at a solid-liquid ratio of 1:60-65 g / mL, extracting for 120-160 min at a pH of 7-7.5 and a temperature of 85-90°C, and filtering to obtain a filtrate a and a Houttuynia cordata filter residue; (2) taking the Houttuynia cordata filter residue, refluxing with ether for 160-180 min, centrifuging, drying the filter residue at 40°C, refluxing with ethanol for 220-250 min, centrifuging, drying the filter residue at 40°C, adding 40 times of deionized water, extracting in a water bath at 85-90°C for 10 h, repeating 4 times, and collecting by centrifugation to obtain a filtrate b; (3) mixing the filtrate a and the filtrate b, concentrating under reduced pressure, and drying to obtain a Houttuynia cordata polysaccharide extract. The Houttuynia cordata polysaccharide of the invention has a high extraction rate and is relatively pure. Chinese patent application CN104906238A discloses a method for preparing total flavonoids of Houttuynia cordata, and the processing steps include: drying Houttuynia cordata in the shade, crushing, adding 50-60% ethanol of 1-2 times the raw material, placing in a homogenizer for mixing and homogenization; after homogenization, placing the material in an ultrasonic device, adding 50-60% ethanol solution of 3-5 times the raw material, ultrasonic extraction for 90-120 minutes, decompressing and concentrating, recovering ethanol, and obtaining a concentrated solution; applying the obtained concentrated solution to a column and eluting through a 80-100 mesh polyamide resin, collecting the eluate, concentrating, and drying to obtain total flavonoids of Houttuynia cordata. The invention has the advantages of rapidity, high efficiency, low solvent usage, good reproducibility, and the resin can be reused.

[0006] However, the above patents only extract a single active ingredient from Houttuynia cordata, and are unable to simultaneously extract polysaccharides and flavonoids from Houttuynia cordata. Therefore, it is of great significance to develop a new method for extracting Houttuynia cordata extracts and to improve the extraction rates of polysaccharides and flavonoids from Houttuynia cordata. Summary of the invention

[0007] In view of the deficiencies in the prior art, the object of the present invention is to provide a Houttuynia cordata extract and a preparation method thereof, which can effectively improve the extraction rate of polysaccharides and flavonoids in Houttuynia cordata. At the same time, the prepared Houttuynia cordata extract has better antioxidant properties and good application prospects.

[0008] To achieve the above object, the present invention provides the following technical solutions: A method for preparing a houttuynia cordata extract comprises the following steps: S1. Pretreatment of Houttuynia cordata: washing and drying the rhizomes of Houttuynia cordata, crushing them and adding them into a mixed acid aqueous solution for immersion treatment, and obtaining a Houttuynia cordata mixed solution after the immersion is completed; S2. Enzymatic hydrolysis of the complex enzyme: Adjust the pH of the Houttuynia cordata mixture in step S1 to 4 - 5, then add pectinase for the first enzymatic hydrolysis. After the enzymatic hydrolysis is completed, inactivate the enzyme, adjust the pH to 6 - 7, add the complex enzyme for the second enzymatic hydrolysis, and inactivate the enzyme after the enzymatic hydrolysis is completed to obtain the enzymatic hydrolysate; S3. Preparation of the aqueous extract: Add menthol to the enzymatic hydrolysate in step S2 for aqueous extraction. After the aqueous extraction is completed, filter to obtain the aqueous extract and the filter residue. The filter residue is dried for later use; S4. Preparation of the ethanol extract: Add the filter residue in step S3 to ethanol, then add salicylic acid and diacetyl tartaric acid monoglyceride, and perform ethanol extraction under ultrasonic conditions. After the ethanol extraction is completed, filter to obtain the ethanol extract; S5. Preparation of the Houttuynia cordata extract: Mix the aqueous extract in step S3 and the ethanol extract in step S4 evenly, and then perform vacuum distillation to remove the solvent to obtain the Houttuynia cordata extract.

[0009] Preferably, the mesh number of the pulverized Houttuynia cordata powder in step S1 is 80 - 120 meshes, the temperature of the impregnation treatment is 30 - 40 °C, and the time is 2 - 3 h.

[0010] Preferably, the mixed acid aqueous solution in step S1 is composed of malic acid, tartaric acid and water. The concentration of malic acid is 7 - 10 g / L, and the concentration of tartaric acid is 3 - 5 g / L; the material - liquid ratio of the Houttuynia cordata rhizome to the mixed acid aqueous solution is 1:10 - 15.

[0011] In the present invention, by treating Houttuynia cordata with a mixed acid, malic acid in the mixed acid can partially hydrolyze components such as cellulose, hemicellulose and pectin in the plant cell wall, making the cell wall structure loose, which is beneficial to subsequent enzymatic hydrolysis and solvent penetration. Tartaric acid has certain antioxidant and chelating effects. The antioxidant effect can protect easily oxidized components such as flavonoids in Houttuynia cordata and reduce their loss during the extraction process; compared with other inorganic acids and organic acids, the action of the mixed acid on Houttuynia cordata is milder, reducing the damage to polysaccharide and flavonoid components and increasing the content of polysaccharide and flavonoid in the subsequent extract.

[0012] Preferably, the addition amount of the pectinase in step S2 is 0.5 - 0.8% of the mass of the Houttuynia cordata rhizome, the temperature of the first enzymatic hydrolysis is 45 - 50 °C, and the time is 1 - 2 h.

[0013] Preferably, the complex enzyme in step S2 is composed of cellulase and xylanase mixed in a mass ratio of 5 - 7:3 - 5. The addition amount of the complex enzyme is 1 - 1.2% of the mass of the Houttuynia cordata rhizome, the temperature of the second enzymatic hydrolysis is 40 - 45 °C, and the time is 30 - 50 min.

[0014] In the present invention, through two-step enzymatic hydrolysis, different components of the cell wall are more thoroughly degraded, improving the extraction efficiency. Using pectinase for preliminary enzymatic hydrolysis can reduce the integrity of the cell wall and decrease the resistance to the dissolution of intracellular substances; subsequently, adding a complex enzyme, cellulase can hydrolyze cellulose in the cell wall, and xylanase can hydrolyze hemicellulose in the cell wall. Through the combined action of the two enzymes, the cell wall of Houttuynia cordata is damaged to the greatest extent, intracellular substances are fully released, and the extraction rate of polysaccharides is increased.

[0015] Preferably, the addition amount of menthol in step S3 is 0.07 - 0.1% of the mass of the enzymatic hydrolysate, the temperature of water extraction is 80 - 90 °C, and the time is 1 - 2 h.

[0016] In the present invention, by adding menthol to the enzymatic hydrolysate and then performing water extraction, after the cell wall is enzymatically hydrolyzed by the complex enzyme, menthol can accelerate the penetration of the solvent into the cells, accelerate the dissolution of intracellular substances, shorten the extraction time and increase the yield.

[0017] Preferably, the solid-liquid ratio of the filter residue to ethanol in step S4 is 1:8 - 12, the addition amount of salicylic acid is 0.2 - 0.3% of the mass of the filter residue, and the addition amount of diacetyl tartaric acid ester of mono- and diglycerides is 0.3 - 0.5% of the mass of the filter residue.

[0018] In the present invention, salicylic acid and diacetyl tartaric acid ester of mono- and diglycerides are added during the ethanol extraction process. Salicylic acid has certain antioxidant and anti-inflammatory effects and may form complexes with certain flavonoid compounds, making the flavonoid substances easier to be extracted; diacetyl tartaric acid ester of mono- and diglycerides has certain cell membrane penetration ability, which helps ethanol enter the cell interior and increases the extraction rate of flavonoid substances.

[0019] Preferably, the power of ultrasonic in step S4 is 60 - 70 W, the frequency is 30 - 40 kHz, the temperature of ethanol extraction is 60 - 65 °C, and the time is 1 - 2 h.

[0020] Preferably, the pressure of vacuum distillation in step S5 is 5 - 10 kPa.

[0021] The present invention also protects a Houttuynia cordata extract prepared by the method as described above.

[0022] Compared with the prior art, the present invention has the following beneficial effects: (1)The preparation method of the houttuynia cordata extract provided by the present invention realizes the efficient synchronous extraction of polysaccharides and flavonoids through the combined pretreatment of mixed acids and multi-stage enzymatic hydrolysis technology. A mixed acid system composed of a compound of malic acid and tartaric acid is used to effectively loosen the plant cell wall structure under mild conditions, and through the synergistic effect of a composite enzyme system of pectinase - cellulase - xylanase, the cellulose, hemicellulose and pectin in the cell wall are directionally decomposed, significantly improving the dissolution rate of intracellular substances. Then, combined with the stepwise water extraction and ultrasonic-assisted ethanol extraction processes, water-soluble polysaccharides and ethanol-soluble flavonoids are efficiently extracted respectively. Compared with the traditional single extraction method, the present invention can simultaneously obtain high yields of polysaccharides and flavonoids, avoiding waste of resources and improving the comprehensive utilization value of houttuynia cordata.

[0023] (2)The preparation method of the houttuynia cordata extract provided by the present invention innovatively uses a mixed acid solution of malic acid and tartaric acid in the pretreatment stage. Malic acid can more gently hydrolyze the cell wall of houttuynia cordata, loosen the structure, and create favorable conditions for subsequent enzymatic hydrolysis and solvent penetration. Tartaric acid has antioxidant and chelating effects, which can protect polysaccharides and flavonoid compounds from being damaged, thereby improving the yield and quality of the target products. Subsequently, a two-step enzymatic hydrolysis process with the synergistic action of pectinase and a composite enzyme (cellulase, xylanase) is adopted in the enzymatic hydrolysis stage. Pectinase can effectively degrade the pectin component in the cell wall and reduce the resistance to the dissolution of intracellular substances. The composite enzyme further acts on cellulose and hemicellulose to more thoroughly destroy the cell wall structure and promote the release of polysaccharides and flavonoid compounds. This enzymatic hydrolysis combination fully considers the composition characteristics of the houttuynia cordata cell wall, has strong pertinence and high enzymatic hydrolysis efficiency.

[0024] (3)The preparation method of the houttuynia cordata extract provided by the present invention introduces menthol in the water extraction stage. Menthol can accelerate the penetration of the solvent into the cell interior and shorten the extraction time. In the ethanol extraction stage, salicylic acid and diacetyl tartaric acid esters of mono- and diglycerides are respectively introduced as co-extraction agents. Their synergistic effect can effectively break the hydrogen bond binding between flavonoids and lignin, promote the permeability of the cell membrane, further improve the extraction rate of flavonoid compounds, and at the same time inhibit the activity of polyphenol oxidase to avoid the oxidative loss of flavonoid components. At the same time, the cavitation effect of ultrasonic waves also helps cell rupture and the dissolution of target components. Through the comprehensive application of these co-extraction means, the extraction process is more efficient and thorough, thereby effectively improving the extraction rates of polysaccharides and flavonoids in houttuynia cordata, and the prepared houttuynia cordata extract has better antioxidant properties. Detailed implementation mode

[0025] The technical solution of the present invention will be clearly and completely described below in conjunction with embodiments. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all embodiments. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts belong to the scope of protection of the present invention.

[0026] Example 1 A preparation method of houttuynia cordata extract, comprising the following steps: S1. Pretreatment of houttuynia cordata: Wash and dry the rhizome of houttuynia cordata, crush it to 100 meshes, and then add it to a mixed acid aqueous solution (the concentration of malic acid is 9 g / L, and the concentration of tartaric acid is 4 g / L). The material-liquid ratio of the rhizome of houttuynia cordata to the mixed acid aqueous solution is 1:13, and soak it at 35 °C for 2.5 h. After soaking, obtain a houttuynia cordata mixed solution; S2. Enzymatic hydrolysis with composite enzyme: Adjust the pH of the houttuynia cordata mixed solution in step S1 to 4.5 by adding 0.1 mol / L sodium hydroxide solution, then add pectinase (0.7% of the mass of the rhizome of houttuynia cordata), and carry out enzymatic hydrolysis at 48 °C for 1.5 h. After enzymatic hydrolysis, inactivate the enzyme in boiling water for 10 min, add 0.1 mol / L sodium hydroxide solution to adjust the pH to 6.5, add a composite enzyme (cellulase and xylanase with a mass ratio of 6:4), and the addition amount of the composite enzyme is 1.1% of the mass of the rhizome of houttuynia cordata. Carry out enzymatic hydrolysis at 43 °C for 40 min, and inactivate the enzyme in boiling water for 10 min after enzymatic hydrolysis to obtain an enzymatic hydrolysate; S3. Preparation of water extract: Add menthol (0.09% of the mass of the enzymatic hydrolysate) to the enzymatic hydrolysate in step S2, carry out water extraction at 85 °C for 1.5 h, filter after water extraction is completed to obtain a water extract and filter residue, and dry the filter residue for later use; S4. Preparation of alcohol extract: Add the filter residue in step S3 to ethanol, the material-liquid ratio of the filter residue to ethanol is 1:10, then add salicylic acid (0.25% of the mass of the filter residue) and diacetyl tartaric acid monoglyceride (0.4% of the mass of the filter residue), and carry out alcohol extraction under ultrasonic conditions. The power of ultrasonic is 65 W, the frequency is 35 kHz, the temperature of alcohol extraction is 63 °C, and the time is 1.5 h. Filter after alcohol extraction is completed to obtain an alcohol extract; S5. Preparation of houttuynia cordata extract: Mix the water extract in step S3 and the alcohol extract in step S4 evenly, and then carry out vacuum distillation to remove the solvent. The pressure of vacuum distillation is 7 kPa to obtain houttuynia cordata extract.

[0027] Example 2 A preparation method of houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash and dry the rhizomes of Houttuynia cordata, crush them to 100 mesh, and then add them to an aqueous mixed acid solution (the concentration of malic acid is 8 g / L, and the concentration of tartaric acid is 4 g / L). The material-liquid ratio of the rhizomes of Houttuynia cordata to the aqueous mixed acid solution is 1:12, and soak at 35 °C for 2.5 h. After soaking, obtain the Houttuynia cordata mixed solution; S2. Enzymatic hydrolysis with composite enzyme: Adjust the pH of the Houttuynia cordata mixed solution in step S1 to 4.5 by adding 0.1 mol / L sodium hydroxide solution, then add pectinase (0.6% of the mass of the rhizomes of Houttuynia cordata), and carry out enzymatic hydrolysis at 47 °C for 1.5 h. After enzymatic hydrolysis, inactivate the enzyme in boiling water for 10 min, add 0.1 mol / L sodium hydroxide solution to adjust the pH to 6.5, add composite enzyme (cellulase and xylanase with a mass ratio of 7:3), and the addition amount of the composite enzyme is 1.1% of the mass of the rhizomes of Houttuynia cordata, and carry out enzymatic hydrolysis at 42 °C for 40 min. After enzymatic hydrolysis, inactivate the enzyme in boiling water for 10 min to obtain the enzymatic hydrolysate; S3. Preparation of water extract: Add menthol (0.08% of the mass of the enzymatic hydrolysate) to the enzymatic hydrolysate in step S2, and carry out water extraction at 85 °C for 1.5 h. After water extraction, filter to obtain the water extract and the filter residue. The filter residue is dried for later use; S4. Preparation of ethanol extract: Add the filter residue in step S3 to ethanol, and the material-liquid ratio of the filter residue to ethanol is 1:10. Then add salicylic acid (0.25% of the mass of the filter residue) and diacetyl tartaric acid monoglyceride (0.4% of the mass of the filter residue), and carry out ethanol extraction under ultrasonic conditions. The power of ultrasonic is 65 W, the frequency is 35 kHz, the temperature of ethanol extraction is 62 °C, and the time is 1.5 h. After ethanol extraction, filter to obtain the ethanol extract; S5. Preparation of Houttuynia cordata extract: Mix the water extract in step S3 and the ethanol extract in step S4 evenly, and then carry out vacuum distillation to remove the solvent. The pressure of vacuum distillation is 8 kPa to obtain the Houttuynia cordata extract.

[0028] Example 3 A method for preparing Houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash and dry the rhizomes of Houttuynia cordata, crush them to 80 mesh, and then add them to an aqueous mixed acid solution (the concentration of malic acid is 7 g / L, and the concentration of tartaric acid is 3 g / L). The material-liquid ratio of the rhizomes of Houttuynia cordata to the aqueous mixed acid solution is 1:10, and soak at 30 °C for 3 h. After soaking, obtain the Houttuynia cordata mixed solution; S2. Enzymatic hydrolysis of the complex enzyme: Add the Houttuynia cordata mixture in step S1 to 0.1 mol / L sodium hydroxide solution to adjust the pH to 4. Then add pectinase (0.5% of the mass of Houttuynia cordata rhizomes), and carry out enzymatic hydrolysis at 45 °C for 2 h. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min. Add 0.1 mol / L sodium hydroxide solution to adjust the pH to 6. Add the complex enzyme (cellulase and xylanase with a mass ratio of 5:5), and the addition amount of the complex enzyme is 1% of the mass of Houttuynia cordata rhizomes. Carry out enzymatic hydrolysis at 40 °C for 50 min. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min to obtain the enzymatic hydrolysate; S3. Preparation of the water extract: Add menthol (0.07% of the mass of the enzymatic hydrolysate) to the enzymatic hydrolysate in step S2, and carry out water extraction at 80 °C for 2 h. After the water extraction is completed, filter to obtain the water extract and the filter residue. The filter residue is dried for later use; S4. Preparation of the ethanol extract: Add the filter residue in step S3 to ethanol, and the material-liquid ratio of the filter residue to ethanol is 1:8. Then add salicylic acid (0.2% of the mass of the filter residue) and diacetyl tartaric acid monoglyceride (0.3% of the mass of the filter residue), and carry out ethanol extraction under ultrasonic conditions. The power of ultrasonic is 60 W, the frequency is 40 kHz, the temperature of ethanol extraction is 60 °C, and the time is 2 h. After the ethanol extraction is completed, filter to obtain the ethanol extract; S5. Preparation of the Houttuynia cordata extract: Mix the water extract in step S3 and the ethanol extract in step S4 evenly, and then carry out vacuum distillation to remove the solvent. The pressure of vacuum distillation is 10 kPa to obtain the Houttuynia cordata extract.

[0029] Example 4 A method for preparing a Houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash and dry the Houttuynia cordata rhizomes, crush them to 120 mesh, and then add them to the mixed acid aqueous solution (the concentration of malic acid is 10 g / L, and the concentration of tartaric acid is 5 g / L). The material-liquid ratio of the Houttuynia cordata rhizomes to the mixed acid aqueous solution is 1:15, and carry out impregnation treatment at 40 °C for 2 h. After the impregnation is completed, obtain the Houttuynia cordata mixture; S2. Enzymatic hydrolysis of the complex enzyme: Add the Houttuynia cordata mixture in step S1 to 0.1 mol / L sodium hydroxide solution to adjust the pH to 5. Then add pectinase (0.8% of the mass of Houttuynia cordata rhizomes), and carry out enzymatic hydrolysis at 50 °C for 1 h. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min. Add 0.1 mol / L sodium hydroxide solution to adjust the pH to 7. Add the complex enzyme (cellulase and xylanase with a mass ratio of 7:3), and the addition amount of the complex enzyme is 1.2% of the mass of Houttuynia cordata rhizomes. Carry out enzymatic hydrolysis at 45 °C for 30 min. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min to obtain the enzymatic hydrolysate; S3. Preparation of aqueous extract: Add menthol (0.1% of the mass of the enzymolysis solution) to the enzymolysis solution in step S2, perform aqueous extraction at 90 °C for 1 h, filter after the aqueous extraction is completed to obtain the aqueous extract and the filter residue, and dry the filter residue for later use; S4. Preparation of ethanol extract: Add the filter residue in step S3 to ethanol, with the solid-liquid ratio of the filter residue to ethanol being 1:12, then add salicylic acid (0.3% of the mass of the filter residue) and diacetyl tartaric acid esters of mono- and diglycerides (0.5% of the mass of the filter residue), perform ethanol extraction under ultrasonic conditions, with the ultrasonic power being 70 W, the frequency being 30 kHz, the temperature of ethanol extraction being 65 °C, and the time being 1 h, filter after the ethanol extraction is completed to obtain the ethanol extract; S5. Preparation of Houttuynia cordata extract: Mix the aqueous extract in step S3 and the ethanol extract in step S4 evenly, then perform vacuum distillation to remove the solvent, with the pressure of vacuum distillation being 5 kPa, to obtain the Houttuynia cordata extract.

[0030] Comparative Example 1 A method for preparing Houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash and dry the Houttuynia cordata rhizome, crush it to 100 meshes, then add it to a citric acid aqueous solution (concentration: 13 g / L), with the solid-liquid ratio of the Houttuynia cordata rhizome to the citric acid aqueous solution being 1:13, perform impregnation treatment at 35 °C for 2.5 h, and after the impregnation is completed, obtain the Houttuynia cordata mixture; S2. Enzymolysis with composite enzyme: Adjust the pH of the Houttuynia cordata mixture in step S1 to 4.5 by adding 0.1 mol / L sodium hydroxide solution, then add pectinase (0.7% of the mass of the Houttuynia cordata rhizome), perform enzymolysis at 48 °C for 1.5 h, inactivate the enzyme in boiling water for 10 min after the enzymolysis is completed, adjust the pH to 6.5 by adding 0.1 mol / L sodium hydroxide solution, add a composite enzyme (cellulase and xylanase with a mass ratio of 6:4), with the addition amount of the composite enzyme being 1.1% of the mass of the Houttuynia cordata rhizome, perform enzymolysis at 43 °C for 40 min, inactivate the enzyme in boiling water for 10 min after the enzymolysis is completed, to obtain the enzymolysis solution; S3. Preparation of aqueous extract: Add menthol (0.09% of the mass of the enzymolysis solution) to the enzymolysis solution in step S2, perform aqueous extraction at 85 °C for 1.5 h, filter after the aqueous extraction is completed to obtain the aqueous extract and the filter residue, and dry the filter residue for later use; S4. Preparation of ethanol extract: Add the filter residue in step S3 to ethanol, with the solid-liquid ratio of the filter residue to ethanol being 1:10, then add salicylic acid (0.25% of the mass of the filter residue) and diacetyl tartaric acid esters of mono- and diglycerides (0.4% of the mass of the filter residue), perform ethanol extraction under ultrasonic conditions, with the ultrasonic power being 65 W, the frequency being 35 kHz, the temperature of ethanol extraction being 63 °C, and the time being 1.5 h, filter after the ethanol extraction is completed to obtain the ethanol extract; S5. Preparation of Houttuynia cordata extract: Mix the water extract in step S3 and the ethanol extract in step S4 evenly, then perform vacuum distillation to remove the solvent at a pressure of 7 kPa to obtain the Houttuynia cordata extract.

[0031] Compared with Example 1, the present invention replaces the mixed acid with citric acid.

[0032] Comparative Example 2 A method for preparing Houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash and dry the Houttuynia cordata rhizomes, crush them to 100 meshes, then add them to an aqueous solution of mixed acid (the concentration of malic acid is 9 g / L, and the concentration of tartaric acid is 4 g / L). The material-liquid ratio of the Houttuynia cordata rhizomes to the aqueous solution of mixed acid is 1:13, and soak at 35 °C for 2.5 h. After soaking, obtain the Houttuynia cordata mixture; S2. Enzymatic hydrolysis with composite enzyme: Adjust the pH of the Houttuynia cordata mixture in step S1 to 4.5 with 0.1 mol / L sodium hydroxide solution, then add pectinase (0.7% of the mass of the Houttuynia cordata rhizomes), and perform enzymatic hydrolysis at 48 °C for 1.5 h. After enzymatic hydrolysis, inactivate the enzyme in boiling water for 10 min, adjust the pH to 6.5 with 0.1 mol / L sodium hydroxide solution, add composite enzyme (cellulase and xylanase with a mass ratio of 6:4), the addition amount of the composite enzyme is 1.1% of the mass of the Houttuynia cordata rhizomes, and perform enzymatic hydrolysis at 43 °C for 40 min. After enzymatic hydrolysis, inactivate the enzyme in boiling water for 10 min to obtain the enzymatic hydrolysate; S3. Preparation of water extract: Subject the enzymatic hydrolysate in step S2 to water extraction at 85 °C for 1.5 h. After water extraction, filter to obtain the water extract and the filter residue. The filter residue is dried for later use; S4. Preparation of ethanol extract: Add the filter residue in step S3 to ethanol. The material-liquid ratio of the filter residue to ethanol is 1:10, then add salicylic acid (0.25% of the mass of the filter residue) and diacetyl tartaric acid ester of mono- and diglycerides (0.4% of the mass of the filter residue), and perform ethanol extraction under ultrasonic conditions. The power of ultrasonic is 65 W, the frequency is 35 kHz, the temperature of ethanol extraction is 63 °C, and the time is 1.5 h. After ethanol extraction, filter to obtain the ethanol extract; S5. Preparation of Houttuynia cordata extract: Mix the water extract in step S3 and the ethanol extract in step S4 evenly, then perform vacuum distillation to remove the solvent at a pressure of 7 kPa to obtain the Houttuynia cordata extract.

[0033] Compared with Example 1, menthol was not added during the water extraction process in this comparative example.

[0034] Comparative Example 3 A method for preparing Houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash the rhizomes of Houttuynia cordata, dry them, pulverize them to 100 mesh, and then add them to an aqueous mixed acid solution (the concentration of malic acid is 9 g / L, and the concentration of tartaric acid is 4 g / L). The material-liquid ratio of the rhizomes of Houttuynia cordata to the aqueous mixed acid solution is 1:13. Immerse them at 35 °C for 2.5 h. After the immersion is completed, obtain the Houttuynia cordata mixed solution; S2. Enzymatic hydrolysis with composite enzyme: Adjust the pH of the Houttuynia cordata mixed solution in step S1 to 4.5 by adding 0.1 mol / L sodium hydroxide solution. Then add pectinase (0.7% of the mass of the rhizomes of Houttuynia cordata), and carry out enzymatic hydrolysis at 48 °C for 1.5 h. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min. Add 0.1 mol / L sodium hydroxide solution to adjust the pH to 6.5, add composite enzyme (cellulase and xylanase with a mass ratio of 6:4), and the addition amount of the composite enzyme is 1.1% of the mass of the rhizomes of Houttuynia cordata. Carry out enzymatic hydrolysis at 43 °C for 40 min. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min to obtain the enzymatic hydrolysate; S3. Preparation of water extract: Add menthol (0.09% of the mass of the enzymatic hydrolysate) to the enzymatic hydrolysate in step S2, and carry out water extraction at 85 °C for 1.5 h. After the water extraction is completed, filter to obtain the water extract and the filter residue. Dry the filter residue for later use; S4. Preparation of alcohol extract: Add the filter residue in step S3 to ethanol, and the material-liquid ratio of the filter residue to ethanol is 1:10. Then add salicylic acid (0.65% of the mass of the filter residue), and carry out alcohol extraction under ultrasonic conditions. The power of ultrasonic is 65 W, the frequency is 35 kHz, the temperature of alcohol extraction is 63 °C, and the time is 1.5 h. After the alcohol extraction is completed, filter to obtain the alcohol extract; S5. Preparation of Houttuynia cordata extract: Mix the water extract in step S3 and the alcohol extract in step S4 evenly, and then carry out vacuum distillation to remove the solvent. The pressure of vacuum distillation is 7 kPa to obtain the Houttuynia cordata extract.

[0035] Compared with Example 1, in this comparative example, diacetyl tartaric acid ester of mono- and diglycerides was not added during the alcohol extraction process.

[0036] Comparative Example 4 A preparation method of Houttuynia cordata extract, comprising the following steps: S1. Pretreatment of Houttuynia cordata: Wash the rhizomes of Houttuynia cordata, dry them, pulverize them to 100 mesh, and then add them to an aqueous mixed acid solution (the concentration of malic acid is 9 g / L, and the concentration of tartaric acid is 4 g / L). The material-liquid ratio of the rhizomes of Houttuynia cordata to the aqueous mixed acid solution is 1:13. Immerse them at 35 °C for 2.5 h. After the immersion is completed, obtain the Houttuynia cordata mixed solution; S2. Enzymatic hydrolysis of the complex enzyme: Adjust the pH of the houttuynia cordata mixture in step S1 to 4.5 by adding 0.1 mol / L sodium hydroxide solution. Then add pectinase (0.7% of the mass of houttuynia cordata rhizomes), and perform enzymatic hydrolysis at 48 °C for 1.5 h. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min. Add 0.1 mol / L sodium hydroxide solution to adjust the pH to 6.5, add the complex enzyme (cellulase and xylanase with a mass ratio of 6:4), and the addition amount of the complex enzyme is 1.1% of the mass of houttuynia cordata rhizomes. Perform enzymatic hydrolysis at 43 °C for 40 min. After the enzymatic hydrolysis is completed, inactivate the enzyme in boiling water for 10 min to obtain the enzymatic hydrolysate; S3. Preparation of the water extract: Add menthol (0.09% of the mass of the enzymatic hydrolysate) to the enzymatic hydrolysate in step S2, and perform water extraction at 85 °C for 1.5 h. After the water extraction is completed, filter to obtain the water extract and the filter residue. The filter residue is dried for later use; S4. Preparation of the ethanol extract: Add the filter residue in step S3 to ethanol, and the material-liquid ratio of the filter residue to ethanol is 1:10. Then add diacetyl tartaric acid ester of mono- and diglycerides (0.65% of the mass of the filter residue), and perform ethanol extraction under ultrasonic conditions. The power of ultrasonic is 65 W, the frequency is 35 kHz, the temperature of ethanol extraction is 63 °C, and the time is 1.5 h. After the ethanol extraction is completed, filter to obtain the ethanol extract; S5. Preparation of the houttuynia cordata extract: Mix the water extract in step S3 and the ethanol extract in step S4 evenly, and then perform vacuum distillation to remove the solvent. The pressure of vacuum distillation is 7 kPa to obtain the houttuynia cordata extract.

[0037] Compared with Example 1, salicylic acid was not added during the ethanol extraction process in this comparative example.

[0038] For the preparation methods of Examples 1-4 and Comparative Examples 1-4, calculate the polysaccharide yield and total flavonoid yield in the houttuynia cordata extract. The determination method of polysaccharide content adopts the phenol-sulfuric acid colorimetric method, and the polysaccharide yield / % = the mass of polysaccharide in the extract / the mass of houttuynia cordata used × 100; the determination of flavonoids adopts the ultraviolet spectrophotometry, and the flavonoid yield / % = the mass of flavonoids in the extract / the mass of houttuynia cordata used × 100. At the same time, test the antioxidant activity of the houttuynia cordata extract. The specific test method is as follows: Accurately pipette 2.0 mL of 0.2 mmol·L -1 DPPH·ethanol solution into a 10 mL test tube, and then add 2 mL of the houttuynia cordata extract solution of Examples 1-4 and Comparative Examples 1-4, with a concentration of 5 mg·mL -1 , quickly mix evenly, and place it statically in the dark under the condition of a 37 °C constant temperature water bath. React for 30 min, and measure the absorbance A 1 . Take 2 mL of ethanol and mix it evenly with 2 mL of DPPH·ethanol (0.2 mmol·L -1 ), and measure the absorbance A 0, simultaneously measure the absorbance A after mixing 2 mL of 95% ethanol and 2 mL of the sample solution 2 ; The DPPH· scavenging rate is calculated according to the following formula: DPPH· scavenging rate = , and the test results are shown in Table 1

[0039] Table 1 As can be seen from Table 1 above, the preparation method of the houttuynia cordata extract provided by the present invention effectively improves the yields of houttuynia cordata polysaccharide and total flavonoids. At the same time, the prepared houttuynia cordata extract has better antioxidant activity and has good application prospects

[0040] Although the embodiments of the present invention have been shown and described, for those of ordinary skill in the art, it can be understood that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principle and spirit of the present invention. The scope of the present invention is defined by the appended claims and their equivalents

Claims

1. A method for preparing a houttuynia cordata extract, characterized in that: The following steps are involved: S1, crushing the rhizome of Houttuynia cordata and adding it into a mixed acid aqueous solution, and immersing it to obtain a Houttuynia cordata mixed solution; S2, adjusting the pH of the mixed solution of Houttuynia cordata to 4-5, adding pectinase, performing enzymolysis once, inactivating the enzyme, adjusting the pH to 6-7, adding complex enzyme, performing enzymolysis twice, inactivating the enzyme, and obtaining an enzymolysis solution; S3, adding menthol to the enzymatic hydrolyzate, extracting with water, and obtaining a water extract and a filter residue; S4, adding the filter residue to ethanol, then adding salicylic acid and diacetyl tartaric acid mono- and diglycerides, extracting with alcohol, filtering, and obtaining an alcohol extract; S5. Mix the water extract and the alcohol extract, and perform vacuum distillation to obtain a Houttuynia cordata extract.

2. The preparation method according to claim 1, characterized in that: The mesh size of the crushed Houttuynia cordata powder in step S1 is 80-120 meshes, and the temperature of the immersion treatment is 30-40° C. and the time is 2-3 hours.

3. The preparation method according to claim 1, characterized in that: The mixed acid aqueous solution in step S1 is composed of malic acid, tartaric acid and water, the concentration of malic acid is 7-10 g / L, and the concentration of tartaric acid is 3-5 g / L; the solid-liquid ratio of the Houttuynia cordata rhizome to the mixed acid aqueous solution is 1:10-15.

4. The preparation method according to claim 1, characterized in that: In step S2, the amount of pectinase added is 0.5-0.8% of the mass of Houttuynia cordata rhizome, the temperature of the primary enzymolysis is 45-50° C., and the time is 1-2 h.

5. The preparation method according to claim 1, characterized in that: The complex enzyme in step S2 is a mixture of cellulase and xylanase in a mass ratio of 5-7:3-5. The amount of the complex enzyme added is 1-1.2% of the mass of the rhizome of Houttuynia cordata. The temperature of the secondary enzymolysis is 40-45° C. and the time is 30-50 min.

6. The preparation method according to claim 1, characterized in that: In step S3, the amount of menthol added is 0.07-0.1% of the mass of the enzymatic hydrolysate, the water extraction temperature is 80-90° C., and the time is 1-2 h.

7. The preparation method according to claim 1, characterized in that: In step S4, the solid-liquid ratio of the filter residue to ethanol is 1:8-12, the amount of salicylic acid added is 0.2-0.3% of the mass of the filter residue, and the amount of diacetyl tartaric acid mono- and diglycerides added is 0.3-0.5% of the mass of the filter residue.

8. The preparation method according to claim 1, characterized in that: In step S4, the power of the ultrasound is 60-70W, the frequency is 30-40kHz, the temperature of the alcohol extraction is 60-65°C, and the time is 1-2h.

9. The preparation method according to claim 1, characterized in that: The pressure of the reduced pressure distillation in step S5 is 5-10 kPa.

10. A Houttuynia cordata extract prepared by the method according to any one of claims 1 to 9.

Images