High-activity diaphragma juglandis aqueous extract as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of diaphragma juglandis extraction, and particularly relates to a high-activity diaphragma juglandis aqueous extract as well as a preparation method and application thereof. The preparation method of the high-activity diaphragma juglandis aqueous extract comprises the step of crushing diaphragma juglandis to obtain diaphragma juglandis powder. And mixing the diaphragma juglandis powder and distilled water according to a mass ratio of 1: (10-30), carrying out subcritical extraction at 120-180 DEG C for 30-60 minutes, collecting extract liquor, and filtering to obtain the diaphragma juglandis aqueous extract. Wherein after the subcritical extraction is finished, stirring is kept, condensate water is introduced, the temperature of the extraction liquid is reduced to the room temperature, and then the extraction liquid is collected. The preparation method of the high-activity diaphragma juglandis aqueous extract provided by the invention has the advantages of no solvent residue, short extraction time and high biological activity of the extract.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of walnut split wood extraction, and particularly relates to a high-activity walnut split wood water extract as well as a preparation method and application thereof. BACKGROUND

[0002] Walnut split wood, also known as walnut septum or walnut husk, is the lignified septum inside the walnut fruit. Walnut split wood has a long application history in traditional Chinese medicine. According to the theory of traditional Chinese medicine, walnut split wood tastes bitter and astringent, and is neutral in nature, and is attributed to the spleen and kidney channels, and has the effects of tonifying kidney and astringency, soothing the nerves and helping sleep, and astringing and stopping bleeding, and is often used for the treatment of symptoms such as spermatorrhea, enuresis, insomnia, frequent urination and the like. Walnut split wood is rich in bioactive components such as flavonoids, phenolic acids, polysaccharides and saponins, and these substances endow it with various pharmacological effects such as antioxidant, anti-inflammatory, hypoglycemic and hypolipidemic effects. Literature reports that walnut split wood extract can significantly improve the glucose metabolism disorder of diabetic model mice, and reduce the blood glucose and blood lipid levels, suggesting that it has potential application value in the management of metabolic diseases.

[0003] In the prior art, common walnut split wood extraction technologies include ultrasonic-assisted enzyme method, organic solvent extraction method, supercritical carbon dioxide extraction, microwave-assisted extraction, spray drying to prepare instant powder, and water extraction method. These methods have the following advantages and disadvantages. The ultrasonic-assisted enzyme extraction has high extraction efficiency and is environmentally friendly, but has high cost and long extraction time. The cavitation effect of ultrasonic waves can accelerate the contact between the solvent and the walnut split wood. The ultrasonic-assisted enzyme extraction uses biological enzymes such as cellulase to avoid the use of large amounts of organic solvents and reduce environmental pollution. The ultrasonic-assisted enzyme extraction can be carried out at a relatively low temperature to avoid the destruction of active ingredients by high temperature. The cost of enzymes is relatively high, and the addition amount of enzymes and reaction conditions need to be accurately controlled. The traditional organic solvent extraction method is mature and widely applicable, but has high energy consumption and solvent residue problems. The traditional organic solvent extraction method is the most commonly used method at present, which is mature and relatively simple to operate. It can extract various types of active ingredients, including polyphenols, flavonoids, etc. However, the use of organic solvents may cause solvent residue, affecting product quality and safety. Moreover, high temperature and long time are required to complete the extraction process, which has high energy consumption. The supercritical carbon dioxide extraction equipment has high cost and high technical requirements. The supercritical carbon dioxide extraction requires high-pressure equipment, which has high purchase and maintenance costs. Moreover, the control of temperature, pressure and other parameters during operation is strict, which has high technical difficulty. The microwave-assisted extraction has equipment limitations and temperature control problems. The microwave-assisted extraction requires special microwave extraction equipment, which has high equipment cost. The temperature rises quickly during microwave extraction, and the temperature needs to be accurately controlled to avoid the destruction of active ingredients. The spray drying equipment is complex and has high cost. Spray drying requires complex equipment and accurate parameter control. The equipment investment and operation cost are high, which limits its large-scale application. The water extraction method has low extraction efficiency and many impurities. The water extraction method has low extraction efficiency for some non-polar or weakly polar components, and many impurities may be brought in during the extraction process, which needs to be further purified.

[0004] Therefore, the existing extraction technology of walnut split wood has the problems of solvent residue and destruction of active ingredients. SUMMARY

[0005] In order to solve the above technical problems, the present application provides a high-activity walnut split wood water extract and a preparation method and application thereof.

[0006] The object of the present application is to provide a preparation method of high-activity walnut split wood water extract, which comprises the following steps: The walnut split wood is crushed to obtain split wood powder. The split wood powder and water are mixed in a mass ratio of 1:10-30, and then subcritical extraction is carried out at 120-180 DEG C for 30-60 min. The extraction liquid is collected, filtered, and dried to obtain the walnut split wood water extract. The subcritical water extraction of walnut split wood uses water as the extraction agent. Under this condition, the solubility of polar compounds is enhanced, and the high temperature and pressure conditions promote the dissolution of phenolic acid and flavonoid glycosides, which can effectively solve the problems of solvent residue and long extraction time in traditional extraction.

[0007] After the extraction is completed, the stirring is maintained, condensed water is passed to reduce the temperature of the extraction liquid to room temperature, and then the extraction liquid is collected again. Rapid cooling after the extraction is completed helps to reduce the problem of damage to active ingredients.

[0008] The water is distilled water or deionized water.

[0009] Preferably, the particle size of the split wood powder is 170-200 µm.

[0010] Preferably, the mass ratio of the split wood powder to distilled water is 1:20.

[0011] Preferably, the temperature of the subcritical extraction is 140 DEG C.

[0012] Preferably, the time of the subcritical extraction is 30 min.

[0013] Preferably, the rotation speed of the subcritical extraction is 120-160 rpm.

[0014] Preferably, the rotation speed of the subcritical extraction is 140 rpm.

[0015] Another object of the present application is to provide the use of the high-activity walnut split wood water extract in the preparation of instant tea, solid beverage and antioxidant products. The walnut split wood water extract is prepared into a solid powder by spray drying, which is convenient for quantitative use, and is also beneficial for storage and transportation.

[0016] Compared with the prior art, the present application has the following beneficial effects: 1. The preparation method of the high-activity walnut split wood water extract comprises the following steps: The walnut split wood is crushed to obtain split wood powder. The split wood powder is added to water in a mass ratio of 1:10-30, and subcritical extraction is carried out at 120-180 DEG C for 30-60 min. The extraction liquid is collected, filtered, and dried to obtain the walnut split wood water extract. After the subcritical extraction is completed, the stirring is maintained, the temperature of the extraction liquid is reduced to room temperature, and then the extraction liquid is collected again.

[0017] The walnut split wood is crushed to obtain split wood powder. The walnut split wood is crushed into powder, which increases the specific surface area of the walnut split wood, and is beneficial to the full contact of water and the walnut split wood to make extraction more sufficient. The split wood powder and distilled water are mixed according to a mass ratio of 1:10-30, and are extracted at an extraction temperature of 120-180 DEG C and a rotating speed of 120-160 rpm for 30-60 min. The extraction liquid is collected, filtered, and the extraction liquid is obtained. Under the condition, the active substances in the split wood powder can be retained, the bioactivity of the phenolic acid substances and the flavonoid glycosides is retained, and the active substances in the split wood powder can be fully extracted. The subcritical water extraction method uses water as the extraction agent, the dielectric constant of water is low, the solubility of polar compounds is enhanced, and the dissolution of the phenolic acid substances and the flavonoid glycosides is promoted under the condition of high temperature and high pressure. These substances neutralize free radicals by providing hydrogen atoms or electrons, and the scavenging mechanism involves the chelation of the adjacent hydroxyl groups on the benzene ring and the free radicals. After the extraction is completed, the stirring is maintained, the condensation water is passed, the temperature of the extraction liquid is reduced to room temperature, and then the extraction liquid is collected. In addition, water is used as the extraction agent, and the composition is single, and there is no residual problem of other solvent components.

[0018] The preparation process of the high-activity walnut split wood water extract of the present application has the highest antioxidant property when the subcritical extraction temperature is 140 DEG C, the extraction time is 30 min, the stirring speed is 140 rpm, and the raw material liquid ratio is 1:20. The ABTS free radical scavenging rate reaches 86.5%, the DPPH free radical scavenging rate reaches 67.0%, and the hydroxyl radical scavenging rate reaches 59.9%. Compared with the traditional water extraction method, the ABTS free radical scavenging is increased by 24.1%, the DPPH free radical scavenging rate is increased by 21.2%, and the hydroxyl radical scavenging rate is increased by 21.3%. Therefore, the preparation method of the high-activity walnut split wood water extract of the present application has no solvent residue problem, has good extraction effect, and has high bioactivity of the walnut split wood water extract.

[0019] 2. Compared with the traditional organic solvent extraction method, the subcritical extraction technology of the preparation process of the high-activity walnut split wood water extract of the present application can achieve a higher extraction efficiency in only 30 min, while the traditional organic solvent extraction usually needs several hours, and the extraction time is shortened by 50%-70%. The present application aims at the high-value utilization of walnut split wood, and innovatively uses the subcritical extraction technology to solve the technical bottlenecks of solvent residue and low extraction efficiency in the traditional extraction method, and provides a scientific basis for the development of related functional foods and health products. The preparation process of the high-activity walnut split wood water extract of the present application not only helps to improve the comprehensive economic benefit of the walnut industry, but also promotes the utilization of agricultural waste resources. BRIEF DESCRIPTION OF DRAWINGS

[0020] Figure 1Figure for the antioxidant activity of the Juglans mandshurica water extract of Example 1 to Example 4 and Comparative Example 1 of the present application.

[0021] Figure 2 Figure for the antioxidant activity of the Juglans mandshurica water extract of Example 5 to Example 8 and Comparative Example 2 of the present application.

[0022] Figure 3 Figure for the antioxidant activity of the Juglans mandshurica water extract of Example 9 to Example 12 and Comparative Example 3 of the present application.

[0023] Figure 4 Figure for the antioxidant activity of the Juglans mandshurica water extract of Example 13 to Example 15 and Comparative Example 4 to Comparative Example 5 of the present application. DETAILED DESCRIPTION

[0024] In order for those skilled in the art to better understand the technical solutions of the present application and to implement them, the technical solutions in the embodiments of the present application are described clearly and completely below in combination with the preferred embodiments and the accompanying drawings. Obviously, the described embodiments are only some of the embodiments of the present application, but not all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor fall within the scope of protection of the present application.

[0025] It should be noted that all the professional terms used in the present application are only for the purpose of describing the specific embodiments and are not intended to limit the scope of protection of the present application. Unless otherwise specified, all the materials, reagents, instruments and equipment used in the following embodiments of the present application can be purchased from the market or prepared by the existing methods.

[0026] I. Materials and equipment 1. Experimental materials The materials used in the present application mainly include Juglans mandshurica, 1,1-diphenyl-2-trinitrophenylhydrazine, 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid and ascorbic acid standard.

[0027] Among them, Juglans mandshurica is purchased from Shaoisheng Health Industry Co., Ltd. in Huaiyuan County. 1,1-diphenyl-2-trinitrophenylhydrazine, abbreviated as DPPH, has a CAS number of 1898-66-4 and is purchased from Tianjin Huasheng Chemical Reagent Co., Ltd. 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid, abbreviated as ABTS, has a CAS number of 30931-67-0 and is purchased from Tianjin Huasheng Chemical Reagent Co., Ltd. Ascorbic acid standard has a purity of more than 98% and a CAS number of 50-81-7 and is purchased from National Pharmaceutical Group Chemical Reagent Co., Ltd.

[0028] 2. Experimental equipment The main equipment used in the present application is a subcritical water extraction device, model FCFSK-0.5, purchased from Henan Subcritical Extraction Technology Research Institute.

[0029] II. Experimental method The antioxidant activity of the walnut Fagussieboldii Hance water extract of the present application is detected by the following method.

[0030] 1. Determination of ABTS free radical scavenging rate Mix 7.4 mmol / L of ABTS solution with 2.6 mmol / L of potassium persulfate solution in equal volume, and react for 12 h to 16 h at room temperature in the dark to generate ABTS stock solution. Dilute the ABTS stock solution to an absorbance of 0.70±0.02 at 734 nm before use to obtain ABTS working solution. Mix 100 μL of the sample solution with 3.9 mL of the ABTS working solution, and immediately measure the absorbance value at 734 nm after reacting for 6 min. Use distilled water instead of the sample as a blank control. The formula (1) is used to calculate the sample scavenging rate:

[0031] Formula (1): Wherein, A0 is the absorbance of water; A is the absorbance of the sample. s

[0032] 2. Determination of DPPH free radical scavenging rate Mix 1 mL of the sample solution with 4 mL of 0.1 mmol / L DPPH ethanol solution, and measure the absorbance at 517 nm after reacting for 30 min in the dark. Use ethanol instead of the sample as a blank control. The scavenging rate is calculated by formula (2):

[0033] Formula (2): Wherein, A0 is the absorbance of water; A1 is the absorbance of the sample solution.

[0034] 3. Determination of hydroxyl radical scavenging rate Mix 1.0 mL of the sample solution with 1.0 mL of 9 mmol / L FeSO4 solution, add 1.0 mL of 9 mmol / L salicylic acid-ethanol solution, and then add 1.0 mL of 8.8 mmol / L H2O2 to start the reaction. Measure the absorbance at 510 nm after reacting for 30 min in a 37℃ water bath. The absorbance value of the reaction system with distilled water instead of H2O2 solution is A2; the absorbance value of the reaction system with solvent instead of the sample solution is A0. The formula (3) is used to calculate the scavenging rate:

[0035] Formula (3): ​Note: A0 is the absorbance of water; A1 is the absorbance of the test solution; A2 is the absorbance of the background of the test solution.

[0036] Example 1 A preparation method of a high-activity Juglans regia L. extract, comprising the following steps: crushing Juglans regia L. into a powder with a particle size of 170 µm to obtain Juglans regia L. powder. The Juglans regia L. powder and distilled water are mixed at a mass ratio of 1:20, and the Juglans regia L. powder is added to the distilled water. Subcritical extraction is performed at 120 ℃ and a rotation speed of 140 rpm for 30 min. After the extraction is completed, stirring is maintained, and condensed water is passed to reduce the temperature of the extract to room temperature. The extract is collected and filtered through filter paper with a pore size of 20 µm to obtain the Juglans regia L. extract. The ABTS free radical scavenging rate of the Juglans regia L. extract reaches 84.6%, the DPPH free radical scavenging rate reaches 48.2%, and the hydroxyl free radical scavenging rate reaches 29.3%.

[0037] Example 2 A preparation method of a high-activity Juglans regia L. extract, comprising the following steps: adjusting the temperature of the subcritical extraction of Example 1 to 140 ℃, and keeping other conditions consistent with Example 1 to obtain the Juglans regia L. extract. The ABTS free radical scavenging rate of the Juglans regia L. extract reaches 87.1%, the DPPH free radical scavenging rate reaches 66.3%, and the hydroxyl free radical scavenging rate reaches 34.8%.

[0038] Example 3 A preparation method of a high-activity Juglans regia L. extract, comprising the following steps: adjusting the temperature of the subcritical extraction of Example 1 to 160 ℃, and keeping other conditions consistent with Example 1 to obtain the Juglans regia L. extract. The ABTS free radical scavenging rate of the Juglans regia L. extract reaches 86.6%, the DPPH free radical scavenging rate reaches 67.0%, and the hydroxyl free radical scavenging rate reaches 44.6%.

[0039] Example 4 A preparation method of a high-activity Juglans regia L. extract, comprising the following steps: adjusting the temperature of the subcritical extraction of Example 1 to 180 ℃, and keeping other conditions consistent with Example 1 to obtain the Juglans regia L. extract. The ABTS free radical scavenging rate of the Juglans regia L. extract reaches 84.7%, the DPPH free radical scavenging rate reaches 64.1%, and the hydroxyl free radical scavenging rate reaches 38.7%.

[0040] Comparative Example 1 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: adjusting the temperature of the subcritical extraction of Example 1 to 200 DEG C, and keeping other conditions consistent with Example 1 to prepare the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 82.1%, the DPPH free radical scavenging rate reaches 61.4%, and the hydroxyl free radical scavenging rate reaches 31.7%.

[0041] The antioxidant activities of the Juglans regia L. water extracts prepared in Examples 1-4 and Comparative Example 1 are shown in Table 1. Figure 1 The results show that the temperature of subcritical extraction has a significant influence on the antioxidant activity of the Juglans regia L. water extract. When the temperature is 120-200 DEG C, the ABTS free radical scavenging rate, the DPPH scavenging rate and the hydroxyl free radical scavenging rate all show a trend of first increasing and then decreasing with the increase of temperature. And when the temperature is 140-160 DEG C, the ABTS free radical scavenging rate and the DPPH scavenging rate are obviously higher than those of other experimental groups, and the highest values reach 87.1% and 67.0% respectively. When the temperature exceeds 140 DEG C, the DPPH scavenging rate does not change significantly. When the temperature reaches 160 DEG C, the hydroxyl free radical scavenging rate reaches the highest value of 44.6%. But when the temperature is greater than 160 DEG C, with the continuous increase of temperature, some heat-labile components begin to degrade, resulting in the decrease of the hydroxyl free radical scavenging rate.

[0042] Example 5 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: crushing Juglans regia L. into powder with a particle size of 180 µm to obtain Juglans regia L. powder. The Juglans regia L. powder and distilled water are mixed at a mass ratio of 1:15, and the Juglans regia L. powder is added into the distilled water. Subcritical extraction is carried out at 140 DEG C and a rotation speed of 140 rpm for 30 min. After the extraction is completed, stirring is kept, condensed water is passed to reduce the temperature of the extract to room temperature, the extract is collected, filtered through a filter paper with a pore size of 20 µm, and the Juglans regia L. water extract is obtained. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 85.3%, the DPPH free radical scavenging rate reaches 62.1%, and the hydroxyl free radical scavenging rate reaches 40.8%.

[0043] Example 6 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: mixing the Juglans regia L. powder of Example 1 and distilled water at a mass ratio of 1:20, and keeping other conditions consistent with Example 5 to prepare the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 87.1%, the DPPH free radical scavenging rate reaches 66.3%, and the hydroxyl free radical scavenging rate reaches 55.2%.

[0044] Example 7 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: preparing the Juglans regia L. water extract by mixing the Juglans regia L. powder of Example 1 with distilled water at a mass ratio of 1:25, and keeping other conditions consistent with those of Example 5. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 86.6%, the DPPH free radical scavenging rate reaches 65.7%, and the hydroxyl free radical scavenging rate reaches 58.4%.

[0045] Example 8 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: preparing the Juglans regia L. water extract by mixing the Juglans regia L. powder of Example 1 with distilled water at a mass ratio of 1:30, and keeping other conditions consistent with those of Example 5. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 85.9%, the DPPH free radical scavenging rate reaches 60.3%, and the hydroxyl free radical scavenging rate reaches 55.3%.

[0046] Example 9 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: preparing the Juglans regia L. water extract by mixing the Juglans regia L. powder of Example 1 with distilled water at a mass ratio of 1:10, and keeping other conditions consistent with those of Example 5. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 84.7%, the DPPH free radical scavenging rate reaches 49.1%, and the hydroxyl free radical scavenging rate reaches 34.8%.

[0047] The antioxidant activities of the Juglans regia L. water extracts of Examples 5-9 are shown in Table 1. Figure 2 The results show that the raw material-to-liquid ratio has an effect on the antioxidant activity of the subcritical extraction Juglans regia L. water extract. The effect on the ABTS free radical scavenging rate is not significant, but the effect on the DPPH free radical and hydroxyl free radical scavenging rates is significant. When the raw material-to-liquid ratio is 1:10-30, both the DPPH free radical and hydroxyl free radical scavenging rates first increase and then slowly decrease with the decrease of the raw material-to-liquid ratio, and both reach the highest value when the raw material-to-liquid ratio is 1:20. This may be because the increase of the solvent amount improves the mass transfer efficiency, so that the active ingredients are more fully dissolved. However, when the solvent continues to increase, the dissolved substances may be diluted due to the further increase of the solvent amount, resulting in the decrease of the antioxidant activity of the water extract per unit mass. When the raw material-to-liquid ratio is large, moderately increasing the solvent volume has a positive effect on improving the antioxidant activity of the subcritical extraction Juglans regia L. water extract.

[0048] Example 10 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: crushing Juglans regia L. into a powder with a particle size of 200 µm to obtain Juglans regia L. powder; adding the Juglans regia L. powder into distilled water at a mass ratio of 1:20. Subcritical extraction is performed at 140 ℃ and a rotation speed of 140 rpm for 30 min. After the extraction is completed, stirring is maintained, and condensed water is passed to reduce the temperature of the extract to room temperature. The extract is collected and filtered through a filter paper with a pore size of 20 µm to obtain the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 87.1%, the DPPH free radical scavenging rate reaches 66.3%, and the hydroxyl free radical scavenging rate reaches 37.8%.

[0049] Example 11 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: adjusting the subcritical extraction time of Example 1 to 40 min, and keeping other conditions consistent with Example 9 to obtain the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 83.8%, the DPPH free radical scavenging rate reaches 62.2%, and the hydroxyl free radical scavenging rate reaches 38.3%.

[0050] Example 12 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: adjusting the subcritical extraction time of Example 1 to 50 min, and keeping other conditions consistent with Example 9 to obtain the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 82.5%, the DPPH free radical scavenging rate reaches 60.1%, and the hydroxyl free radical scavenging rate reaches 45.1%.

[0051] Example 13 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: adjusting the subcritical extraction time of Example 1 to 60 min, and keeping other conditions consistent with Example 9 to obtain the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 82.1%, the DPPH free radical scavenging rate reaches 61.2%, and the hydroxyl free radical scavenging rate reaches 52.5%.

[0052] Comparative Example 2 A preparation method of high-activity Juglans regia L. water extract, comprising the following steps: adjusting the subcritical extraction time of Example 1 to 20 min, and keeping other conditions consistent with Example 9 to obtain the Juglans regia L. water extract. The ABTS free radical scavenging rate of the Juglans regia L. water extract reaches 85.1%, the DPPH free radical scavenging rate reaches 50.2%, and the hydroxyl free radical scavenging rate reaches 35.6%.

[0053] The antioxidant activities of the walnut Juglans mandshurica water extracts of Examples 10-13 and Comparative Example 2 were determined as shown in Table 1. Figure 3 As shown in Table 1, the extraction time had some effect on the antioxidant activity of the subcritical extraction walnut Juglans mandshurica water extract. When the extraction time was 20-60 min, the ABTS radical scavenging rate and the DPPH radical scavenging rate both increased first and then decreased with increasing extraction time, and both reached a maximum when the extraction time was 30 min, i.e., 87.1% and 66.3%, respectively. However, the hydroxyl radical scavenging rate increased with increasing extraction time in the test time range. This phenomenon can be attributed to two reasons. On the one hand, a shorter extraction time cannot fully destroy the plant cell wall structure, resulting in incomplete release of the active ingredients. On the other hand, a longer extraction time can cause degradation or oxidation of some heat-sensitive ingredients. In addition, the active ingredients with different molecular weights respond differently to the extraction time. The maximum extraction amount of the small-molecule phenolic substances is reached at 20 min, while the polysaccharides need 40 min to be fully dissolved.

[0054] Example 14 A method for preparing a high-activity walnut Juglans mandshurica water extract comprises the following steps: crushing walnut Juglans mandshurica into a powder with a particle size of 190 µm to obtain Juglans mandshurica powder. The Juglans mandshurica powder and distilled water are mixed at a mass ratio of 1:20, and the Juglans mandshurica powder is added to the distilled water. Subcritical extraction is performed at 140°C and a rotation speed of 120 rpm for 30 min. After the extraction is completed, stirring is maintained, and condensed water is passed to reduce the temperature of the extract to room temperature. The extract is collected and filtered through a filter paper with a pore size of 20 µm to obtain the walnut Juglans mandshurica water extract. The ABTS radical scavenging rate of the walnut Juglans mandshurica water extract is 86.4%, the DPPH radical scavenging rate is 66.1%, and the hydroxyl radical scavenging rate is 54.8%.

[0055] Example 15 A method for preparing a high-activity walnut Juglans mandshurica water extract comprises the following steps: adjusting the rotation speed of the subcritical extraction of Example 1 to 140 rpm, and keeping the other conditions the same as in Example 13 to obtain the walnut Juglans mandshurica water extract. The ABTS radical scavenging rate of the walnut Juglans mandshurica water extract is 87.1%, the DPPH radical scavenging rate is 66.3%, and the hydroxyl radical scavenging rate is 55.2%.

[0056] Example 16 A method for preparing a high-activity walnut Juglans mandshurica water extract comprises the following steps: adjusting the rotation speed of the subcritical extraction of Example 1 to 160 rpm, and keeping the other conditions the same as in Example 13 to obtain the walnut Juglans mandshurica water extract. The ABTS radical scavenging rate of the walnut Juglans mandshurica water extract is 86.2%, the DPPH radical scavenging rate is 55.4%, and the hydroxyl radical scavenging rate is 56.1%.

[0057] Comparative Example 3 A method for preparing a high-activity Juglans regia L. bark water extract, comprising the following steps: adjusting the rotation speed of the subcritical extraction of Example 1 to 180 rpm, and keeping other conditions consistent with Example 13, to obtain a Juglans regia L. bark water extract. The ABTS free radical scavenging rate of the Juglans regia L. bark water extract reaches 82.7%, the DPPH free radical scavenging rate reaches 48.3%, and the hydroxyl free radical scavenging rate reaches 53.1%.

[0058] Comparative Example 4 A method for preparing a high-activity Juglans regia L. bark water extract, comprising the following steps: adjusting the rotation speed of the subcritical extraction of Example 1 to 200 rpm, and keeping other conditions consistent with Example 13, to obtain a Juglans regia L. bark water extract. The ABTS free radical scavenging rate of the Juglans regia L. bark water extract reaches 81.3%, the DPPH free radical scavenging rate reaches 41.7%, and the hydroxyl free radical scavenging rate reaches 50.4%.

[0059] The antioxidant activity of the Juglans regia L. bark water extract of Example 14-Example 16 and Comparative Example 3-Comparative Example 4 is shown in Table 2. Figure 3 The results show that the rotation speed of the subcritical extraction has an effect on the antioxidant activity of the Juglans regia L. bark water extract. The rotation speed of the subcritical extraction has a smaller effect on the ABTS free radical and hydroxyl free radical scavenging rates, but has a significant effect on the DPPH free radical scavenging rate, and when the stirring rotation speed is 120 rpm-140 rpm, there is no overall change. When the stirring rotation speed is 140 rpm-200 rpm, there is a downward trend overall. This is related to the degradation of heat-sensitive phenolic substances caused by high-speed shearing force, and the increase in temperature fluctuations in the system caused by excessively high stirring speed, and the possible exacerbation of the oxidative decomposition of active ingredients due to thermodynamic instability. The cell structure of the raw material treated at 140 rpm presents a uniform honeycomb-like rupture, while the fiber rupture and charring phenomenon are obvious in the group with excessively high rotation speed.

[0060] Comparative Example 6 A method for preparing a traditional Juglans regia L. bark water extract, comprising the following steps: adding Juglans regia L. bark powder and distilled water at a mass ratio of 1:20, and adding the bark powder into the distilled water. Extracting at 100℃ for 30 min to obtain a Juglans regia L. bark water extract. The ABTS free radical scavenging rate of the Juglans regia L. bark water extract is 62.4%, the DPPH free radical scavenging rate is 45.8%, and the hydroxyl free radical scavenging rate is 38.6%.

[0061] Compared with Example 1-Example 15, it can be seen that the antioxidant activity of the Juglans regia L. bark subcritical water extract prepared by the method of the present application is significantly higher than that of the water extract prepared by the traditional method.

[0062] In order to illustrate the influence of the three conditions of extraction temperature, material liquid ratio and extraction time on the antioxidant activity of the walnut Fissistigma glomeratum water extract, the following experiments are further made.

[0063] The temperature and time of the subcritical extraction and the material liquid ratio are selected, and the ABTS free radical clearance rate, the DPPH free radical clearance rate and the hydroxyl radical clearance rate are taken as indexes for comprehensive evaluation. The comprehensive evaluation standard is ABTS free radical clearance rate: DPPH free radical clearance rate: hydroxyl radical clearance rate = 1:1:1. The temperature of the subcritical extraction is denoted as A, the time is denoted as B, and the material liquid ratio is denoted as C, and the conditions are shown in Table 1.

[0064] Table 1 Extraction conditions of walnut Fissistigma glomeratum water extract The influence of the three conditions of the subcritical extraction temperature and extraction time and the material liquid ratio on the antioxidant activity of the walnut Fissistigma glomeratum water extract is shown in Table 2. The results show that from the data in Table 2, RA>RC>RB, and the order of the factors affecting the antioxidant activity of the subcritical extraction walnut Fissistigma glomeratum water extract is A>C>B. It can be known from the comparison of K1, K2 and K3 that when the process condition is A1B2C2, that is, A is 140℃, B is 1:20 g / mL, and C is 30 min, the antioxidant activity of the extracted walnut Fissistigma glomeratum water extract is the highest.

[0065] Table 2 Condition score of walnut Fissistigma glomeratum water extract In the table, "-" represents that there is no such item.

[0066] The antioxidant activity of the walnut Fissistigma glomeratum water extract prepared at the extraction temperature of 140℃, the extraction time of 30 min, the stirring speed of 140 rpm and the material liquid ratio of 1:20 g / mL is shown in Table 3. The results show that under the process condition, the ABTS free radical clearance rate of the walnut Fissistigma glomeratum water extract prepared is 86.5±0.3%, the DPPH free radical clearance rate is 67.7±0.4%, the hydroxyl radical clearance rate is 59.9±0.4%, and the comprehensive antioxidant activity index is significantly higher than that of other test groups.

[0067] Table 3 Antioxidant activity of walnut Fissistigma glomeratum water extract under the optimal extraction process condition IV. Result discussion The test results show that the extraction temperature, extraction time and raw material liquid ratio of the application have a significant influence on the antioxidant activity of the subcritical extraction walnut split wood water extract, and the stirring speed has no significant influence on the antioxidant activity of the subcritical extraction walnut split wood water extract. Under the conditions of extraction temperature 140 DEG C, extraction time 30 min, stirring speed 140 rpm and liquid ratio 1:20, the ABTS free radical scavenging rate of the walnut split wood water extract reaches 86.5+ / -0.3%, the DPPH free radical scavenging rate reaches 67.0+ / -0.4%, and the hydroxyl radical scavenging rate reaches 59.9+ / -0.4%. Compared with the traditional water extraction method, the ABTS free radical scavenging is increased by 24.1+ / -0.3%, the DPPH free radical scavenging rate is increased by 21.2+ / -0.4%, and the hydroxyl radical scavenging rate is increased by 21.2+ / -0.4%. This shows that the dielectric constant of the subcritical water extraction method of the application is reduced, the solubility of polar compounds is enhanced, and the high temperature and high pressure conditions promote the dissolution of phenolic acid and flavonoid glycosides. These substances neutralize free radicals by providing hydrogen atoms or electrons, and the scavenging mechanism involves the chelation of the adjacent hydroxyl group on the benzene ring with free radicals. Compared with the literature reported, the scavenging rate obtained in this study is higher than that of ADELA et al. in 2018, which is 52.4% higher than that of the conventional solvent extraction of walnut septum polyphenols, and the difference is related to the distribution of antioxidant components in the raw material and the extraction parameters.

[0068] In addition, the subcritical water extraction technology can realize efficient extraction in a short time, and significantly improve the production efficiency. In the application, only 30 min is needed to achieve high extraction efficiency, while the traditional organic solvent extraction usually needs several hours. However, the high temperature and high pressure conditions may cause degradation of some heat-sensitive components, and the temperature and time of extraction need to be carefully controlled during the extraction process.

[0069] It should be noted that when numerical ranges are involved in the application, both endpoints of each numerical range and any number between the two endpoints can be selected. Since the same steps and examples are used, the preferred embodiments of the application are described to prevent redundancy. Although the preferred embodiments of the application have been described, those skilled in the art can make additional changes and modifications to these embodiments once they understand the inventive concept of the application, and these changes and modifications all fall within the scope of the application.

[0070] Obviously, those skilled in the art can make various modifications and variations to the application without departing from the spirit and scope of the application. If these modifications and variations of the application fall within the scope of the equivalent technology of the application, the application also intends to include these modifications and variations.

Claims

1. A method for preparing a highly active aqueous extract of walnut septum, characterized in that, Includes the following steps: The walnut septum is crushed to obtain septum powder; the septum powder is added to water at a mass ratio of 1:10~30, and subcritical extraction is performed at 120℃~180℃ for 30min~60min. The extract is collected, filtered, and the walnut septum aqueous extract is obtained. After the subcritical extraction is completed, stirring is maintained, the temperature of the extract is lowered to room temperature, and then the extract is collected.

2. The method for preparing a highly active walnut septum aqueous extract according to claim 1, characterized in that, The particle size of the septum wood powder is 170µm~200µm.

3. The method for preparing a highly active walnut septum aqueous extract according to claim 1, characterized in that, The mass ratio of the wood septum powder to distilled water is 1:

20.

4. The method for preparing a highly active walnut septum aqueous extract according to claim 1, characterized in that, The subcritical extraction temperature is 140°C.

5. The method for preparing a highly active walnut septum aqueous extract according to claim 1, characterized in that, The subcritical extraction time is 30 min.

6. The method for preparing a highly active walnut septum aqueous extract according to claim 1, characterized in that, The subcritical extraction speed is 120 rpm to 160 rpm.

7. The method for preparing a highly active walnut septum aqueous extract according to claim 6, characterized in that, The subcritical extraction speed was 140 rpm.

8. Aqueous extract of walnut septum prepared by the preparation method according to any one of claims 1 to 7.

9. The application of the highly active walnut septum water extract according to claim 8 in the preparation of instant tea, solid beverages, antioxidant health products or antioxidant drugs.

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