Preparation method of Elsholtzia extract and its product and application

Abstract

The present invention relates to a preparation method of an Elsholtzia extract, and a product and application thereof. The preparation method comprises: mixing Elsholtzia with water, extracting, and obtaining an Elsholtzia extract; decompressing and concentrating the Elsholtzia extract, mixing with yeast, and fermenting, and obtaining an Elsholtzia fermented liquid; filtering and drying the Elsholtzia fermented liquid. The present invention uses high-pressure hot water extraction to increase the dissolution rate of effective components in Elsholtzia, and then uses fermentation technology to enhance the biological activity of effective components in Elsholtzia; the preparation process of the present invention does not use any organic solvent or toxic and harmful chemical substances, is harmless to the human body, is green and environmentally friendly, and can be applied to the fields of functional foods, health products, medicines, or functional cosmetics to achieve industrial production.

Description

Technical Field

[0001] The invention belongs to the technical field of material extraction, and relates to a preparation method of an Elsholtzia ciliata extract, a product and an application thereof. Background Art

[0002] In the millennium Chinese herbal medicine culture, there are records of the application of plants, from human food to medicine for treating diseases, and then to beauty and hair products. Elsholtzia ciliata is a plant of the genus Elsholtzia in the family Lamiaceae. The whole plant can be used as medicine, and it is a plant with both medicinal and edible properties. Elsholtzia ciliata is pungent and slightly warm, and has the effects of sweating, dispelling dampness and harmonizing the middle. The Compendium of Materia Medica records that "Elsholtzia ciliata is a medicine for dispelling exterior pathogens in summer, just like ephedra in winter. People with qi deficiency should not take too much", so Elsholtzia ciliata is known as "June ephedra" or "summer ephedra". Modern pharmacological studies have shown that the material basis of the action of Elsholtzia ciliata is mainly volatile oils and flavonoids, showing anti-inflammatory, antibacterial, antioxidant, and antiviral effects.

[0003] At present, the extraction of Elsholtzia ciliata extract is mostly done by organic solvent extraction to obtain flavonoid active ingredients, which is time-consuming, has low resource utilization, and is only used in cosmetics for external use, and cannot be used in oral beauty products. The biological activity of Elsholtzia ciliata extract obtained by water extraction is low. Therefore, based on the current technical problems, the development of a safe, green, and effective Elsholtzia ciliata extract and its application in functional foods and functional cosmetics is the research focus in this field. Summary of the invention

[0004] In view of the deficiencies of the prior art, the object of the present invention is to provide a preparation method of Elsholtzia ciliata extract and its product and application.

[0005] In order to achieve the purpose of the invention, the present invention adopts the following technical solutions:

[0006] In a first aspect, the present invention provides a method for preparing an Elsholtzia ciliata extract, the preparation method comprising:

[0007] (1) mixing Elsholtzia ciliata with water, and extracting the mixture to obtain an Elsholtzia ciliata extract;

[0008] (2) concentrating the Elsholtzia extract under reduced pressure, mixing with yeast, and fermenting to obtain the Elsholtzia fermented liquid;

[0009] (3) filtering the Elsholtzia fermented liquid and drying it to obtain the product.

[0010] The total flavonoid content of the Elsholtzia ciliata extract prepared by the present invention is not less than 20%. The content of the effective ingredient rosmarinic acid is greater than 1%, which can remove DPPH free radicals, ABTS +The invention uses high-pressure hot water extraction to improve the dissolution rate of the effective ingredients in Elsholtzia, and then uses fermentation technology to improve the biological activity of the effective ingredients in Elsholtzia; the preparation process of the invention does not use any organic solvents or toxic and harmful chemicals, is harmless to the human body, is green and environmentally friendly, and can be applied to the fields of functional foods, health products, medicines or functional cosmetics to achieve industrial production.

[0011] Preferably, the mass ratio of the Elsholtzia ciliata to water is 1:(5-30), wherein the specific point values ​​in (5-30) can be selected from 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, etc., and other specific point values ​​within the above numerical range can be selected, which will not be repeated here.

[0012] Preferably, the extraction temperature is 90-121° C. and the extraction time is 20-60 min.

[0013] The present invention finds that temperature affects the extraction effect, and further affects the content of flavonoids in the extract.

[0014] The temperature can be selected from 90°C, 92°C, 94°C, 96°C, 98°C, 100°C, 102°C, 104°C, 106°C, 108°C, 110°C, 112°C, 114°C, 116°C, 118°C, 120°C, 121°C, etc., and the time can be selected from 20min, 25min, 30min, 35min, 40min, 45min, 50min, 55min, 60min, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be elaborated here one by one.

[0015] Preferably, the yeast comprises any one of high temperature resistant yeast, high sugar resistant yeast or aroma producing yeast, or a combination of at least two thereof.

[0016] Preferably, the fermentation temperature is 25-35°C and the fermentation time is 24-72h.

[0017] The temperature can be selected as 25°C, 26°C, 27°C, 28°C, 29°C, 30°C, 31°C, 32°C, 33°C, 34°C, 35°C, etc., and the time can be selected as 24h, 28h, 32h, 36h, 40h, 44h, 48h, 52h, 56h, 60h, 64h, 68h, 72h, etc. Other specific point values ​​within the above numerical range can be selected, so they will not be described one by one here.

[0018] Preferably, the mass of the yeast is 0.1-2% of the mass of the extract, for example 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, 1.1%, 1.2%, 1.3%, 1.4%, 1.5%, 1.6%, 1.7%, 1.8%, 1.9%, 2%, etc. Other specific values ​​within the above numerical range can be selected, which will not be repeated here.

[0019] Preferably, the fermentation is carried out under stirring, and the stirring speed is 100-200rpm, for example, 100rpm, 110rpm, 120rpm, 130rpm, 140rpm, 150rpm, 160rpm, 170rpm, 180rpm, 190rpm, 200rpm, etc. Other specific point values ​​within the above numerical range can be selected, which will not be repeated here.

[0020] Preferably, the extraction further includes filtering.

[0021] Preferably, the filtration screen aperture is 400-800 mesh, for example, 400 mesh, 450 mesh, 500 mesh, 550 mesh, 600 mesh, 650 mesh, 700 mesh, 750 mesh, 800 mesh, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be described one by one here.

[0022] Preferably, the temperature of the reduced pressure concentration is 50-70° C. and the pressure is 90-120 Mbar.

[0023] The temperature can be selected from 50°C, 52°C, 55°C, 58°C, 60°C, 62°C, 65°C, 68°C, 70°C, etc., and the pressure can be selected from 90Mbar, 92Mbar, 95Mbar, 98Mbar, 100Mbar, 102Mbar, 105Mbar, 108Mbar, 110Mbar, 112Mbar, 115Mbar, 118Mbar, 120Mbar, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be elaborated here one by one.

[0024] Preferably, the mass percentage of solids in the concentrated Elsholtzia liquid is 1-5%, for example 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, etc. Other specific points within the above numerical range can be selected and will not be described here one by one.

[0025] Preferably, the fermentation further includes filtering and sterilization.

[0026] Preferably, the sterilization temperature is 80-90°C and the time is 25-35 minutes.

[0027] The present invention adopts pasteurization to prevent the fermentation liquid from undergoing secondary fermentation and affecting the biological activity.

[0028] The temperature can be selected as 80°C, 81°C, 82°C, 83°C, 84°C, 85°C, 86°C, 87°C, 88°C, 89°C, 90°C, etc., and the time can be selected as 25min, 26min, 27min, 28min, 29min, 30min, 31min, 32min, 33min, 34min, 35min, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be elaborated here one by one.

[0029] Preferably, the filtration includes primary filtration and secondary filtration.

[0030] Preferably, the pore size of the filter for the primary filtration is 150-400 mesh, for example, 150 mesh, 200 mesh, 250 mesh, 300 mesh, 350 mesh, 400 mesh, etc. Other specific point values ​​within the above numerical range can be selected, and they will not be described one by one here.

[0031] Preferably, the secondary filtration comprises vacuum filtration.

[0032] Preferably, the adsorbent material for suction filtration includes diatomaceous earth.

[0033] Preferably, the drying method in step (4) includes spray drying.

[0034] The product is prepared by spray drying method, and the obtained Elsholtzia ciliata extract has fine powder, uniform particle quality, high biological activity of effective ingredients, and wide application.

[0035] Preferably, the skeleton support agent used in the spray drying comprises any one of mannitol, sorbitol, maltodextrin or maltitol, or a combination of at least two thereof.

[0036] Preferably, the added amount of the skeleton support agent is 10-50% of the solid mass in the Elsholtzia fermented liquid, for example, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, etc. Other specific point values ​​within the above numerical range can be selected and will not be described one by one here.

[0037] Preferably, the drying temperature is 100-150°C, for example, 100°C, 105°C, 110°C, 115°C, 120°C, 125°C, 130°C, 135°C, 140°C, 145°C, 150°C, etc. Other specific point values ​​within the above numerical range can be selected, which will not be described here one by one.

[0038] In a second aspect, the present invention provides an Elsholtzia ciliata extract prepared according to the method for preparing the Elsholtzia ciliata extract according to the first aspect.

[0039] The mass percentage of total flavonoids in the Elsholtzia extract is ≥20%, and rosmarinic acid is greater than 1%. The DPPH free radical scavenging rate IC50 value of the Elsholtzia extract prepared by the invention is 38.75 μg / mL, and the ABTS + The free radical inhibition rate IC50 value is 0.85 mg / mL, and the hyaluronidase activity inhibition rate IC50 value is 187 μg / mL. It has high biological activity and good antioxidant and moisturizing effects.

[0040] In a third aspect, the present invention provides a use of the Elsholtzia ciliata extract according to the second aspect in the preparation of foods, health products, medicines or functional cosmetics with antioxidant and moisturizing effects.

[0041] Compared with the prior art, the present invention has the following beneficial effects:

[0042] (1) Novel preparation technology:

[0043] Traditional extraction techniques such as heating reflux extraction start from the surface of the medicinal material. It is difficult to extract the active ingredients to the maximum extent within a limited temperature, and too much residue is discarded, resulting in excessive loss of active ingredients. Using pure water as the solvent, high-pressure hot water extraction is used to extract Elsholtzia ciliata. Under high pressure conditions, the tissue cells and subcellular structures of the extract are disintegrated, allowing the intracellular components to fully contact the solvent, thereby increasing the dissolution rate of the active ingredients. At the same time, with the assistance of fermentation technology, under the action of microorganisms, the active ingredients can not only be fully extracted to improve biological activity, but also decompose factors that are adverse to the human body, thereby increasing the utilization rate of plants.

[0044] (2) Expansion of application scope:

[0045] The Elsholtzia ciliata extract provided by the invention does not use an organic solvent, and the extracted effective ingredients can be used not only in external cosmetics, but also in the fields of functional foods, health products, etc.

[0046] (3) Low cost:

[0047] The extraction technology adopts high-pressure hot water extraction and microbial fermentation. The preparation process is simple, green and environmentally friendly. The extract has a wide market application range and low cost. BRIEF DESCRIPTION OF THE DRAWINGS

[0048] Figure 1 UPLC test results of rosmarinic acid standard and Elsholtzia ciliata extract prepared in Example 1 are shown in FIG.

[0049] Figure 2 This is a graph showing the DPPH radical scavenging rate of the Elsholtzia ciliata extract and vitamin C prepared in Example 1 at different concentrations.

[0050] Figure 3 ABTS of the Elsholtzia extract and vitamin C prepared in Example 1 at different concentrations + Free radical inhibition rate graph.

[0051] Figure 4 This is a graph showing the hyaluronidase inhibition rate of the Elsholtzia ciliata extract and vitamin C prepared in Example 1 at different concentrations. DETAILED DESCRIPTION

[0052] The technical solution of the present invention is further described below by specific implementation methods. It should be understood by those skilled in the art that the embodiments are only to help understand the present invention and should not be regarded as specific limitations of the present invention.

[0053] The sources of the functional ingredients contained in the products involved in the following embodiments and comparative examples are as follows (only the functional ingredients are reflected, and the necessary auxiliary ingredients contained in other commercially available raw materials are not repeated):

[0054] High temperature resistant and highly active yeast for brewing: Angel Yeast Co., Ltd.

[0055] Aroma active dry yeast: Angel Yeast Co., Ltd.

[0056] High sugar tolerant and highly active yeast: Angel Yeast Co., Ltd.

[0057] Example 1

[0058] This embodiment provides an Elsholtzia extract, which is prepared by the following preparation method:

[0059] (1) Powdering the raw medicinal material of Elsholtzia ciliata, mixing it with 10 times the amount of water, extracting it under high pressure at 121° C. for 40 min, and passing it through a 400-mesh sieve to obtain an Elsholtzia ciliata extract;

[0060] (2) concentrating the Elsholtzia extract under reduced pressure at 60° C. and 110 Mbar to a solid content of 2.5% to obtain a concentrated solution; inoculating the concentrated solution with 0.1% of a high-temperature-resistant and highly active yeast for brewing, stirring at a speed of 150 rpm, and fermenting at 30° C. for 48 h; and sterilizing at 85° C. for 30 min after the fermentation is completed to obtain an Elsholtzia fermented solution;

[0061] (3) The fermented liquid of Elsholtzia ciliata is filtered through a 400-mesh sieve and then vacuum-filtered using diatomaceous earth. The mixture is then mixed with maltodextrin having a solid content of 30% by weight and spray-dried at 150° C. to obtain the product.

[0062] Example 2

[0063] This embodiment provides an Elsholtzia extract, which is prepared by the following preparation method:

[0064] (1) Powdering the raw medicinal material of Elsholtzia ciliata, mixing it with 15 times the amount of water, extracting it under high pressure at 110° C. for 30 min, and passing it through a 600-mesh sieve to obtain an Elsholtzia ciliata extract;

[0065] (2) concentrating the Elsholtzia extract under reduced pressure at 60° C. and 110 Mbar to a solid content of 3% to obtain a concentrated solution; inoculating 0.3% of brewing thermostable and highly active yeast into the concentrated solution, stirring at a speed of 180 rpm, fermenting at 35° C. for 24 h, and sterilizing at 85° C. for 30 min after the fermentation is completed to obtain the Elsholtzia fermented solution;

[0066] (3) The fermented liquid of Elsholtzia ciliata is filtered through a 400-mesh sieve and then vacuum-filtered using diatomaceous earth. The mixture is then mixed with maltodextrin having a solid content of 20%, and spray-dried at 150° C. to obtain the product.

[0067] Example 3

[0068] This embodiment provides an Elsholtzia extract, which is prepared by the following preparation method:

[0069] (1) Powdering the raw medicinal material of Elsholtzia ciliata, mixing it with 20 times the amount of water, extracting it under high pressure at 90° C. for 60 min, and passing it through an 800-mesh sieve to obtain an Elsholtzia ciliata extract;

[0070] (2) concentrating the Elsholtzia extract under reduced pressure at 60° C. and 110 Mbar to a solid content of 1.5% to obtain a concentrated solution; inoculating 0.5% of brewing high-temperature resistant and highly active yeast into the concentrated solution, stirring at a speed of 150 rpm, fermenting at 25° C. for 72 h, and sterilizing at 85° C. for 30 min after the fermentation is completed to obtain the Elsholtzia fermented solution;

[0071] (3) The fermented liquid of Elsholtzia ciliata is filtered through a 800-mesh sieve and then vacuum-filtered using diatomaceous earth. The mixture is then mixed with maltodextrin having a solid content of 25%, and spray-dried at 150° C. to obtain the product.

[0072] Example 4

[0073] This embodiment provides an Elsholtzia ciliata extract, which differs from Embodiment 1 only in that in step (2), an equal amount of high-temperature-resistant and highly active yeast for brewing is replaced with high-sugar-resistant and highly active yeast, and other operations remain unchanged.

[0074] Example 5

[0075] This embodiment provides an Elsholtzia ciliata extract, which differs from Embodiment 1 only in that in step (2), an equal amount of the high temperature resistant and highly active yeast for brewing is replaced by an active dry yeast for aroma production, and other operations remain unchanged.

[0076] Example 6

[0077] This embodiment provides an Elsholtzia ciliata extract, which differs from Embodiment 1 only in that step (1) is "grinding the raw Elsholtzia ciliata medicinal material into powder, mixing it with 10 times the amount of water, extracting it under high pressure at 80° C. for 40 min, and passing it through a 400-mesh sieve to obtain an Elsholtzia ciliata extract", and other operations remain unchanged.

[0078] Example 7

[0079] This embodiment provides an Elsholtzia ciliata extract, which differs from Embodiment 1 only in that step (1) is "grinding the raw Elsholtzia ciliata medicinal material into powder, mixing it with 10 times the amount of water, extracting it under high pressure at 130° C. for 40 min, and passing it through a 400-mesh sieve to obtain an Elsholtzia ciliata extract", and other operations remain unchanged.

[0080] Comparative Example 1

[0081] This comparative example provides an Elsholtzia ciliata extract, which differs from Example 1 only in that step (1) is "grinding the raw Elsholtzia ciliata medicinal material into powder, mixing it with 10 times the amount of water, passing it through a 400-mesh sieve, and heating and refluxing to obtain an Elsholtzia ciliata extract", and other operations remain unchanged.

[0082] Comparative Example 2

[0083] This comparative example provides a herba elsholtzia extract, which is prepared by the following preparation method:

[0084] (1) Powdering the raw medicinal material of Elsholtzia ciliata, mixing it with 10 times the amount of water, extracting it under high pressure at 121° C. for 40 min, and passing it through a 400-mesh sieve to obtain an Elsholtzia ciliata extract;

[0085] (2) concentrating the Elsholtzia extract under reduced pressure at 60° C. and 110 Mbar to a solid content of 2.5% to obtain a concentrated solution, and sterilizing at 85° C. for 30 min to obtain an Elsholtzia concentrate;

[0086] (3) The Elsholtzia ciliata concentrate is filtered through a 400-mesh sieve and then vacuum-filtered using diatomaceous earth. The mixture is then mixed with 30% maltodextrin by solid content and spray-dried at 150° C. to obtain a product.

[0087] Comparative Example 3

[0088] This comparative example provides a herba elsholtzia extract, which is prepared by the following preparation method:

[0089] (1) grinding the raw medicinal material of Elsholtzia ciliata into powder, mixing it with 10 times the amount of water, passing it through a 400-mesh sieve, and heating and refluxing to obtain an Elsholtzia ciliata extract;

[0090] (2) concentrating the Elsholtzia extract under reduced pressure at 60° C. and 110 Mbar to a solid content of 2.5% to obtain a concentrated solution, and sterilizing at 85° C. for 30 min to obtain an Elsholtzia concentrate;

[0091] (3) The Elsholtzia ciliata concentrate is filtered through a 400-mesh sieve and then vacuum-filtered using diatomaceous earth. The mixture is then mixed with 30% maltodextrin by solid content and spray-dried at 150° C. to obtain a product.

[0092] Test Example 1

[0093] Determination of total flavonoids content:

[0094] Flavonoid compounds refer to a series of compounds with the basic core structure of 2-phenylchromone in the molecular structure of the substance. The flavonoid core contains basic oxygen atoms, and half of them have phenolic hydroxyl groups. Under neutral or weak alkaline conditions and the presence of sodium nitrite, flavonoid compounds and aluminum salts form chelates. After adding sodium hydroxide to adjust the alkalinity, they appear red-orange, with maximum absorption at around 500nm, and the absorption value is proportional to the content of flavonoid compounds. Rutin is used as a reference substance, and aluminum nitrate is used as a color developer for flavonoid determination. The absorbance is linearly related to the concentration of rutin.

[0095] Test samples: Examples 1-7, Comparative Examples 1-3

[0096] Test method: Precisely measure 1.0 mL of each of the Elsholtzia extract solution (2 mg / mL) and the reference solution (1, 0.8, 0.6, 0.4, 0.2 mg / mL) in a 10 mL volumetric flask, accurately add 0.5 mL of 5% sodium nitrite solution, mix, and place for 6 minutes, then accurately add 0.5 mL of 10% aluminum nitrate solution, mix, and place for 6 minutes, then add 4.0 mL of 4% sodium hydroxide solution, mix, and dilute to the scale with 95% ethanol, mix, and place at room temperature for 15 minutes. After the reaction is completed, 200uL of each sample is drawn into a 96-well plate, and the reagent is used as a blank. The absorbance is measured at a wavelength of 500±2nm. A standard curve is drawn according to the absorbance value and the concentration of the rutin reference solution. The total flavonoid content is calculated according to the standard curve. The results are shown in Table 1.

[0097] Total flavonoid content (%) = total flavonoid mass / extract mass × 100%

[0098] Table 1

[0099] Group Total flavonoids content (%) Example 1 35.26 Example 2 30.7 Example 3 30.5 Example 4 32.1 Example 5 32.5 Example 6 28.6 Example 7 30 Comparative Example 1 30.29 Comparative Example 2 30.5 Comparative Example 3 27.5

[0100] The Elsholtzia extracting method provided by the invention can increase the extraction rate of effective components, has a good extraction effect on flavonoids, and the extraction temperature is very important to the extraction effect.

[0101] Test Example 2

[0102] Determination of rosmarinic acid content:

[0103] The samples were tested by ultra-high performance liquid chromatography, and the chromatographic conditions were:

[0104] Chromatographic column: Poroshell 120SB-C18 column (100mm×3.0mm, 2.7μm); column temperature: 35℃; UV detector wavelength: 326nm; injection volume: 5μL; flow rate: 0.8mL / min; mobile phase: 0.1% formic acid water (A), acetonitrile (B), elution program: 0~5min, 15% B; 5~10min, 20% B; 10~30min, 25% B; 30.00~35min, 30% B; 35~40min, 15% B.

[0105] The test results of the samples described in Example 1 are as follows Figure 1 The results of determination of rosmarinic acid content by the above chromatography are shown in Table 2.

[0106] Table 2

[0107] Group Rosmarinic acid content (%) Example 1 1.53 Example 2 1.41 Example 3 1.36 Example 4 1.45 Example 5 1.43 Example 6 1.28 Example 7 1.33 Comparative Example 1 1.33 Comparative Example 2 1.39 Comparative Example 3 1.05

[0108] Test Example 3

[0109] DPPH free radical scavenging experiment

[0110] DPPH (1,1-diphenyl-2-trinitrophenylhydrazine, 2,2-diphenyl-1-picrylhydrazyl) is a very stable nitrogen-centered free radical with a single electron. In the presence of a free radical scavenger, the single electron of DPPH is captured, causing its color to become lighter and the absorbance at the maximum light absorption wavelength to decrease. The decrease in the absorbance level indicates an increase in antioxidant activity, thereby evaluating the antioxidant capacity of the test sample.

[0111] Test method: DPPH solution was dissolved in anhydrous ethanol to a concentration of 0.1 mg / mL. A 0.5 mg / mL vitamin C standard solution was prepared at the same time. The concentrations of the Elsholtzia extract prepared in Examples 1-7 and Comparative Examples 1-3 were dissolved in different proportions of aqueous solution (1-0.03125 mg / mL). 150 μL of DPPH ethanol solution and 150 μL of Elsholtzia extract solution were mixed evenly. A blank control group was set up. The reaction was carried out at room temperature in the dark for 30 minutes. The reaction was shaken and the absorbance was measured at 517 nm. The scavenging rate of the Elsholtzia sample for DPPH free radicals was calculated according to the following formula:

[0112]

[0113] Vitamin C was used as a positive control in this experiment. A scatter plot with smooth lines and data markers was prepared with the concentration of the sample to be tested as the X-axis and the absorbance as the Y-axis. The upper, middle and lower three points with a clearance rate of about 50% were selected as a straight line to obtain the straight line equation, and the DPPH free radical scavenging IC50 value and clearance rate of the sample and vitamin C were calculated. The DPPH free radical scavenging IC50 value results are shown in Table 3, and the DPPH free radical scavenging rate results of the Elsholtzia extract prepared in Example 1 and vitamin C are shown in Table 3. Figure 2 shown.

[0114] Table 3

[0115]

[0116]

[0117] The Elsholtzia ciliata extract provided by the present invention has good antioxidant effect, and the selection of yeast has a great influence on the antioxidant effect.

[0118] Test Example 4

[0119] ABTS + Free radical scavenging experiment

[0120] ABTS is oxidized to green ABTS by appropriate oxidants. + , in the presence of antioxidants, ABTS + The production of ABTS is inhibited and measured at 405 nm. + The total antioxidant capacity of the sample can be determined and calculated by the absorbance.

[0121] ABTS + The kit includes: reagent one, detection buffer; reagent two, ABTS solution; reagent three, concentrated substrate solution, prepared into application solution with distilled water at a ratio of 1:39; ABTS working solution preparation: prepared into ABTS working solution at a ratio of reagent one: reagent two: reagent three application solution = 76:5:4; reagent four, enzyme solution, prepared into reagent four application solution at a ratio of enzyme solution: reagent one = 1:9. At the same time, prepare 1 mg / mL vitamin C standard solution, and the concentration of Elsholtzia ciliata extract prepared by Examples 1-7 and Comparative Examples 1-3 dissolved in different proportions of aqueous solution (1-0.03125 mg / mL). Mix 20 μL of reagent four application solution, 170 μL of ABTS working solution and 10 μL of sample solution evenly, set up a blank control group, react at room temperature in the dark for 6 minutes, shake well, measure the absorbance at 405 nm, and calculate the Elsholtzia ciliata sample to ABTS according to the following formula + Free radical scavenging rate IR:

[0122]

[0123] Ai represents the absorbance of the solution after the test solution and ABTS are mixed;

[0124] Aj represents the absorbance of the solution after the test solution and the solvent are mixed;

[0125] A0 represents the absorbance of the solution after ABTS and solvent are mixed.

[0126] Vitamin C was used as a positive control in this experiment. A scatter plot with smooth lines and data markers was made with the concentration of the sample to be tested as the X-axis and the absorbance as the Y-axis. A straight line was drawn from the top, middle and bottom three points where the clearance rate was about 50%, and the equation of the straight line was obtained to calculate the ABTS of the sample and vitamin C. + Free radical scavenging IC50 values ​​and scavenging rates. ABTS of samples and vitamin C + The free radical scavenging IC50 values ​​are shown in Table 4. The ABTS of the Elsholtzia extract and vitamin C prepared in Example 1 + Free radical scavenging rate Figure 3 shown.

[0127] Table 4

[0128] <![CDATA[ABTS + Free radical scavenging (IC50 value mg / mL)]]> Example 1 0.85 Example 2 0.91 Example 3 0.95 Example 4 1.09 Example 5 1.13 Example 6 1.37 Example 7 1.25 Comparative Example 1 0.9 Comparative Example 2 1.3 Comparative Example 3 1.42

[0129] The Elsholtzia ciliata extract provided by the present invention has good antioxidant effect, and the selection of yeast has a great influence on the antioxidant effect.

[0130] Test Example 5

[0131] Hyaluronidase inhibition assay

[0132] Hyaluronidase (HAase) is an enzyme that mainly hydrolyzes hyaluronic acid (HA). Inhibiting the activity of HAase can ensure the normal content and function of HA. HAase can hydrolyze HA to produce β-N-acetylglucosamine, which condenses with p-dimethylaminobenzaldehyde under alkaline conditions to produce the chromogen 2-methyl-3-diacetylpyrrole derivative (purple). The lower the degree of HA hydrolysis, the better the moisturizing effect of the test sample.

[0133] Test method:

[0134] Weigh 2.5-10 mg, dissolve in 10 mL of 0.1 mM acetate buffer and vortex to prepare 0.25-1 mg / mL HAS solution; weigh 10 mg, dissolve in 10 mL of 0.1 mM acetate buffer and vortex to prepare 1 mg / mL HA solution. Use water as solvent to prepare 1 mg / mL tannic acid solution and the Elsholtzia extract prepared in Example 1-7 and Comparative Example 1-3 (concentration range: 0.125-1 mg / mL). The operation is shown in the following table:

[0135] Table 5

[0136]

[0137]

[0138] A—Absorbance value of sample solution

[0139] B—Absorbance value of sample blank

[0140] C—absorbance value of control solution

[0141] D—Absorbance value of control blank

[0142] Tannic acid was used as a positive control in this experiment. A scatter plot with smooth lines and data markers was prepared with the concentration of the sample to be tested as the X-axis and the absorbance as the Y-axis. The upper, middle and lower three points with an inhibition rate of about 50% were selected as a straight line to obtain the straight line equation, and the hyaluronidase inhibition IC50 value and inhibition rate of the sample and tannic acid were calculated. The hyaluronidase activity inhibition rates of the Elsholtzia extract prepared in Example 1 and the positive control at different concentrations are shown in Figure 1. Figure 4 The IC50 values ​​of hyaluronidase inhibition of Elsholtzia ciliata extract are shown in Table 6.

[0143] Table 6

[0144] Hyaluronidase inhibition (IC50 value μg / mL) Example 1 187 Example 2 326 Example 3 376 Example 4 407 Example 5 423 Example 6 459 Example 7 399 Comparative Example 1 414 Comparative Example 2 521 Comparative Example 3 552

[0145] It can be seen from the data in Table 6 that the Elsholtzia ciliata extract provided by the present invention has good moisturizing effect, and the selection of yeast has a greater impact on the moisturizing effect.

[0146] In summary, when the temperature is too high or too low during the high-pressure extraction of Elsholtzia, the total flavonoids content in the Elsholtzia extract will be reduced, and the DPPH free radical scavenging rate, ABTS+ free radical inhibition rate and hyaluronidase inhibition activity will be reduced; when the high-pressure hot water extraction process of Elsholtzia is replaced by a heating reflux process, the total flavonoids content of the Elsholtzia extract will also be reduced, and the antioxidant and moisturizing activities will be reduced; when the Elsholtzia extract is directly filtered without undergoing a fermentation step, the obtained Elsholtzia extract will be significantly reduced in antioxidant and moisturizing effects compared with the Elsholtzia extract that has undergone a fermentation process; and the use of a high-pressure extraction process to extract the raw medicinal material of Elsholtzia can increase the dissolution rate of the effective ingredients in Elsholtzia, and then supplemented by a microbial fermentation process to greatly improve the biological activity of the extract, and the addition of a skeleton support agent allows the Elsholtzia extract to be used as a food raw material in functional foods, health products and other fields.

[0147] The applicant declares that the present invention uses the above-mentioned embodiments to illustrate the preparation method of the Elsholtzia extract of the present invention, its product and application, but the present invention is not limited to the above-mentioned embodiments, that is, it does not mean that the present invention must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement of the present invention, equivalent replacement of the raw materials of the product of the present invention, addition of auxiliary components, selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

[0148] The preferred embodiments of the present invention are described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the technical concept of the present invention, a variety of simple modifications can be made to the technical solution of the present invention, and these simple modifications all belong to the protection scope of the present invention.

[0149] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not further describe various possible combinations.

Claims

1. A method for preparing an Elsholtzia extract, characterized in that: The preparation method comprises: (1) mixing Elsholtzia ciliata with water, and extracting at 90-121° C. for 20-60 min to obtain an Elsholtzia ciliata extract; (2) concentrating the Elsholtzia extract under reduced pressure, mixing with a thermostable yeast, and fermenting to obtain an Elsholtzia fermented liquid; (3) filtering the Elsholtzia fermented liquid and drying it to obtain the product.

2. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The mass ratio of the Elsholtzia ciliata to water is 1:(5-30).

3. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The fermentation temperature is 25-35°C and the fermentation time is 24-72h.

4. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The mass of the yeast is 0.1-2% of the mass of the extract.

5. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The fermentation is carried out under stirring at a rotation speed of 100-200 rpm.

6. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The method further comprises filtering after the leaching.

7. The method for preparing the Elsholtzia ciliata extract according to claim 6, characterized in that: The filtration screen has an aperture of 400-800 meshes.

8. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The fermentation process also includes filtering and sterilization.

9. The method for preparing the Elsholtzia ciliata extract according to claim 8, characterized in that: The sterilization temperature is 80-90°C and the time is 25-35 minutes.

10. The method for preparing the Elsholtzia ciliata extract according to claim 1, characterized in that: The drying method in step (3) includes spray drying.

11. The Elsholtzia ciliata extract prepared by the method for preparing the Elsholtzia ciliata extract according to any one of claims 1 to 10.

12. Use of the Elsholtzia ciliata extract according to claim 11 in the preparation of food, health products, medicines or cosmetics with antioxidant effects.

13. Use of the Elsholtzia ciliata extract according to claim 11 in preparing cosmetics with moisturizing effect.

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