Gooseberry extract as well as extraction method and application thereof

Abstract

The invention relates to an India gooseberry extract as well as an extraction method and application thereof, and the India gooseberry extract is extracted by utilizing an improved natural deep eutectic solvent (NaDES), so that the yield of active ingredients in the India gooseberry extract is improved.

Description

Technical Field

[0001] The present invention relates to an Indian gooseberry extract and an extraction method thereof, in particular to an extraction method using a modified natural deep eutectic solvent (NaDES) as an extraction solvent to increase the yield of active ingredients in the Indian gooseberry extract, and the Indian gooseberry extract obtained by the extraction method. Background Art

[0002] In recent years, due to the natural disasters caused by the intensification of climate change, the world has begun to realize the importance of sustainable development of the environment and consciously implement environmental protection actions. The beauty industry has also set off a similar topic boom, from "environmental protection" and "purity" to "plastic reduction". Now, "carbon neutrality" has become a trending term related to ecology. However, plant extracts often used in cosmetics are often extracted with organic solvents in the production process, but organic solvents are not biodegradable and cause environmental damage, and the concentration and purification process consumes a lot of energy and increases carbon emissions. If we can develop and use green solutions that are safe and environmentally friendly based on the principles of green chemistry, it is necessary to balance environmental issues and economic development.

[0003] Recently, a large number of green solvents have been studied, including bio-solvents, supercritical fluids, and ionic liquids. The properties of ionic liquids include low vapor pressure, thermal stability and electrochemical stability, non-flammability, and reusability. Studying their application as solvents in industrial reactions, extraction separations, and material synthesis is very helpful. In the past, the composition of ionic liquids was mostly based on imidazolium ions, which had problems such as high toxicity and poor biodegradability.

[0004] Deep eutectic solvents (DESs) have many similarities with ionic liquids, but their composition is different from that of ionic liquids. Because deep eutectic solvents are composed of hydrogen-bond acceptors and hydrogen-bond donors, the melting point of the homogeneous mixture formed will be significantly lower than the melting point of the original composition, and the preparation of such mixtures is relatively simple, as long as they are heated and stirred to form a homogeneous liquid; Natural deep eutectic solvents (NaDES) are composed of natural compounds, and the raw materials are from natural sources, using a specific ratio of choline chloride and urea. Later, other scholars proposed that raw materials that can be obtained in nature can be used for reactions, usually using choline derivatives, sugars, organic acids, amino acids, and alcohols. Natural deep eutectic solvents have the characteristics of biodegradability and low toxicity, which is in line with the trend of modern green chemistry.

[0005] However, in cosmetics, the addition of choline chloride is expressly prohibited due to the principle of safety risk assessment; therefore, it is necessary to find raw materials that are mild and low in irritation to the skin without using choline chloride and other banned ingredients in cosmetics, to prepare natural deep eutectic solvents, and to use them to extract various natural plant extracts.

[0006] On the other hand, Indian gooseberry (Phyllanthus emblica, also known as Phyllanthus emblica or emblica oleracea) can be used in traditional medicine to treat fever, cough, skin diseases, liver diseases and other diseases. Recent pharmacological studies have shown that it has anti-cancer, anti-inflammatory, anti-lipid, antioxidant, anti-aging, liver protection, anti-diarrhea, immunomodulatory and antibacterial effects. Other studies have shown that Indian gooseberry fruit is rich in phenolic compounds, flavonoids, tannins and other active ingredients. Among them, β-glucogallin is known to inhibit aldose reductase to reduce lipopolysaccharide-induced inflammatory response; ellagic acid has anti-inflammatory and antioxidant functions and has potential preventive effects on cardiovascular diseases and diabetes; gallic acid has antioxidant and anti-allergic effects; rutin has the effect of reducing hypertension, vascular hemorrhage and other related diseases and microvascular repair, and has good health care effects.

[0007] However, currently common extraction methods are unable to fully and effectively extract the active ingredients contained in Indian gooseberry. Therefore, it is necessary to find an efficient, safe and environmentally friendly extraction method that can not only effectively extract the active ingredients in Indian gooseberry, but also conform to the trend of modern green chemistry. Summary of the invention

[0008] In view of the above-mentioned defects, the inventors have made unremitting efforts in research and development and experiments, and finally developed an extraction method that can improve the yield of active ingredients in Indian gooseberry extract and the Indian gooseberry extract obtained by the extraction method. In addition to significantly increasing the content of active ingredients in the Indian gooseberry extract, the extraction method uses a non-toxic extraction solvent and does not cause pollution to the environment, which is an environmentally friendly extraction method.

[0009] The first object of the present invention is to provide a method for extracting active ingredients from Indian gooseberry extract to increase the yield, comprising the following steps:

[0010] Step S1: dissolving at least two natural deep eutectic solvents at 50° C. and then mixing them to obtain a mixed solution of natural deep eutectic solvents;

[0011] Step S2: adding at least one emulsifier to the mixed solution of the natural deep eutectic solvent, stirring evenly at 50° C. to obtain an emulsified mixed solution;

[0012] Step S3: extracting Indian gooseberry with the emulsified mixed solution to obtain Indian gooseberry extract.

[0013] In step S1, the natural deep eutectic solvent is selected from glycerin, urea, 1,3-butanediol, fructose, betaine, sorbitol and / or lactic acid.

[0014] In step S2, the emulsifier is selected from sorbitan monolaurate (Sorbitan Laurate, Span 20), sorbitan monooleate (Sorbitan Monooleate, Span 80) and / or polysorbate 80 (Polysorbate 80, Tween 80).

[0015] In step S3, the extraction method is to use ultrasound to extract Indian gooseberry with the emulsified mixed solution at 30-60° C. for 10-30 minutes, and the obtained extract is centrifuged to obtain the liquid part, and further filtered with filter paper to obtain the clear filtrate, which is the Indian gooseberry extract.

[0016] The Indian gooseberry is a powdered Indian gooseberry obtained by drying, freeze-drying, crushing and sieving.

[0017] The second object of the present invention is to provide an Indian gooseberry extract obtained by the aforementioned extraction method; wherein the yield of the active ingredients in the Indian gooseberry extract is significantly improved compared to the Indian gooseberry extract obtained by extraction using a general traditional organic solvent, thereby significantly improving the antioxidant, tyrosinase inhibition, IL-1β inhibition, UV light protection and / or melanin inhibition capabilities of the Indian gooseberry extract.

[0018] Through the extraction method provided by the present invention, the content of active ingredients in the Indian gooseberry extract can be significantly increased, and the extraction solvent used is a non-toxic natural deep eutectic solvent. The natural deep eutectic solvent has the characteristics of biodegradability and low toxicity, and can reduce pollution to the environment, which is in line with the trend of modern green chemistry.

[0019] Hereinafter, the present invention will be described in detail by means of specific embodiments in combination with the accompanying drawings, so that it will be easier to understand the purpose, technical content, characteristics and technical effects of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The present invention will be further described below in conjunction with the accompanying drawings:

[0021] Figure 1 A flow chart of the extraction method for improving the yield of Indian gooseberry extract according to the present invention; Figure 2 Schematic diagram of DPPH free radical scavenging ability; Figure 3 It is a schematic diagram of the in vitro tyrosinase inhibition rate; Figure 4 Schematic diagram of IL-1β inhibitory ability; Figure 5 is a schematic diagram of UV light protection capability; Figure 6 Schematic diagram of the ability to inhibit melanin production. DETAILED DESCRIPTION

[0022] In order to facilitate understanding of the technical features, contents and advantages of the present invention and make the technical effects that can be achieved more obvious, the present invention will now be described in detail in the form of embodiments in conjunction with the accompanying drawings. The drawings used therein are only for illustration and auxiliary explanation, and may not be the actual proportions and precise configurations after the implementation of the present invention. Therefore, the proportions and configuration relationships of the attached drawings should not be interpreted or limited to the scope of rights of the present invention in actual implementation.

[0023] Unless otherwise defined, all terms (including technical and scientific terms) used herein have the meanings commonly understood by those of ordinary skill in the art to which the invention belongs. It will be further understood that those terms, such as those defined in commonly used dictionaries, should be interpreted as having a meaning consistent with their meanings in the context of the relevant technology and the present invention, and will not be interpreted as idealized or overly formal meanings, unless explicitly defined as such herein.

[0024] Example 1 Preparation of Indian Gooseberry Extract

[0025] See also Figure 1 , which is a flow chart of the extraction method for improving the yield of Indian gooseberry extract of the present invention, the extraction method comprises the following steps:

[0026] Step S1: dissolving at least two natural deep eutectic solvents at 50° C. and then mixing them to obtain a mixed solution of natural deep eutectic solvents;

[0027] Step S2: adding at least one emulsifier to the mixed solution of the natural deep eutectic solvent, stirring evenly at 50° C. to obtain an emulsified mixed solution;

[0028] Step S3: extracting Indian gooseberry with the emulsified mixed solution to obtain Indian gooseberry extract.

[0029] In step S1, the natural deep eutectic solvent is selected from glycerin, urea, 1,3-butanediol, fructose, betaine, sorbitol and / or lactic acid; preferably, the mixed solution of the natural deep eutectic solvent is selected from glycerin and urea (represented by the code GlyU), 1,3-butanediol and fructose (represented by the code BGF), betaine and sorbitol (represented by the code BetSor) or glycerin and lactic acid (represented by the code GlyLa); wherein any two natural deep eutectic solvents are mixed in a molar ratio of 1:1; more preferably, the mixed solution of the natural deep eutectic solvent is obtained by stirring and mixing at 50°C for 10 minutes.

[0030] In step S2, the emulsifier is selected from sorbitan monolaurate (Sorbitan Laurate, Span 20), sorbitan monooleate (Sorbitan Monooleate, Span 80) and / or polysorbate 80 (Polysorbate 80, Tween 80); preferably, the two natural deep eutectic solvents in the mixed solution of natural deep eutectic solvents are mixed with the emulsifier in a molar ratio of 1:1:1 to obtain the emulsified mixed solution; more preferably, the emulsifier consists of sorbitan oleate and polysorbate 80 (represented by code ST); more preferably, the emulsifier consists of 60wt% sorbitan oleate and 40wt% polysorbate 80.

[0031] In step S3, the extraction method is to use ultrasound to extract Indian gooseberry with the emulsified mixed solution at 30-60° C. for 10-30 minutes, and the obtained extract is centrifuged to obtain the liquid portion, and further filtered with filter paper to obtain the clear filtrate, which is the Indian gooseberry extract; preferably, the emulsified mixed solution and Indian gooseberry are mixed and extracted in a weight ratio of 4:1; more preferably, the extraction method is to use ultrasound to extract at 30° C. for 30 minutes.

[0032] Wherein, the Indian gooseberry is a powdered Indian gooseberry obtained by drying, freeze-drying, crushing and sieving; preferably, the powdered Indian gooseberry is obtained by filtering through a 200-mesh sieve, and the moisture content after drying is less than 5wt%.

[0033] Comparative Example 1 Comparison of extraction differences before and after adding emulsifier

[0034] In order to find out whether adding an emulsifier to the mixture of the natural deep eutectic solvent is helpful for increasing the active ingredients in the Indian gooseberry extract, a mixture of the natural deep eutectic solvent without adding an emulsifier and a mixture of the natural deep eutectic solvent with an emulsifier added were compared; the mixture of the natural deep eutectic solvent was selected from glycerol and urea (represented by the code GlyU), 1,3-butylene glycol and fructose (represented by the code BGF), betaine and sorbitol (represented by the code BetSor) or glycerol and lactic acid (represented by the code GlyLa), as a group without adding an emulsifier; and a single emulsifier was added to the mixture of the natural deep eutectic solvent, that is, sorbitan monolaurate (Span 20), sorbitan monooleate (Span80) or polysorbate-80 (Tween80) was added to the mixture of the natural deep eutectic solvent, as a group with adding an emulsifier.

[0035] Next, the above-mentioned groups without and with emulsifiers were used to extract Indian gooseberry, respectively. The extraction conditions were the same as those described in Example 1 (the group without emulsifiers skipped step S2). The Indian gooseberry extracts obtained from each group were quantitatively analyzed by HPLC, and the gallic acid content of the Indian gooseberry extracts was used as a comparison basis. The results are shown in Table 1. For the sake of simplicity, each component is expressed in English.

[0036] Table 1

[0037] From the results in Table 1 above, it can be seen that under the same extraction conditions, the gallic acid content of the group with emulsifier added is higher than that of the group without emulsifier added; in addition, it can be seen from the group with emulsifier added that the addition of sorbitan monolaurate (Span 20), sorbitan monooleate (Span 80) and polysorbate-80 (Tween80) can increase the gallic acid content, and sorbitan monolaurate (Span 20) has the best effect, followed by polysorbate-80 (Tween80) and sorbitan monooleate (Span 80), indicating that different HLB values ​​will affect their extraction efficiency.

[0038] From the above results, it can be found that the hydrophilic-lipophilic balance (HLB) value of the emulsifier has a significant effect on the extraction efficiency of gallic acid. The HLB value of sorbitan monolaurate (Span 20) is 8.6, the HLB value of sorbitan monooleate (Span 80) is 4.3, and the HLB value of polysorbate 80 (Tween 80) is 15.0. It can be seen that gallic acid is more easily extracted by an HLB value of 8.6.

[0039] Comparative Example 2 Extraction Effect of Adding Two Emulsifiers

[0040] From the results of the aforementioned comparative example 1, it can be seen that the HLB value of the emulsifier has a significant effect on the extraction efficiency of gallic acid, and when the HLB value is around 8.6, the extraction efficiency of gallic acid is better. Therefore, the HLB values ​​of the two compounded emulsifiers need to be around 8.6; sorbitan oleate (Span 80, HLB value is 4.3) and polysorbate-80 (Tween80, HLB value is 15.0) are used for compounding. After repeated adjustment of the ratio, when the composition of the compounded emulsifier is 60% sorbitan oleate and 40% polysorbate-80, its HLB value is 8.6, which is just around the HLB value of 8.6.

[0041] Next, the composite emulsifier was added to the mixed solution of each group of deep eutectic solvents, i.e., glycerol and urea (represented by code GlyU), 1,3-butanediol and fructose (represented by code BGF), betaine and sorbitol (represented by code BetSor) or glycerol and lactic acid (represented by code GlyLa), and Indian gooseberry was extracted with them respectively. The extraction conditions were as described in Example 1. The Indian gooseberry extracts obtained from each group were quantitatively analyzed by HPLC, and the gallic acid content of the Indian gooseberry extracts was used as a comparison basis. The results are shown in Table 2. For the sake of simplicity, each component is also represented in English.

[0042] Table 2

[0043] From the results in Table 2 above, it can be seen that the compound emulsifier with two emulsifiers added, especially the compound emulsifier with a low HLB value (<6) and a high HLB value (>10) selected to compound to the required HLB value, compared with the use of a single emulsifier with an HLB value within the required range, the Indian gooseberry extract obtained by the extraction has a significantly increased gallic acid content, and the concentration thereof is greater than 500 μg / mL. In particular, the natural deep eutectic solvent mixture composed of betaine and sorbitol and the compound emulsifier (i.e., the BetSorST group) has the best extraction efficiency, reaching 1249 μg / mL.

[0044] Example 2 Comparison of extraction effects between emulsified mixed solution and general organic solvent

[0045] From the results of the aforementioned comparative example 2, it can be seen that the emulsified mixed solution with the best extraction efficiency is the BetSorST group, so this group is used to compare the extraction effect with general organic solvents with lower environmental pollution; wherein the general organic solvents with lower environmental pollution are selected from corn-derived plant propylene glycol (Zemea) and biological fermentation-derived ethanol (Ethanol).

[0046] The emulsified mixed solution, propylene glycol and ethanol of the BetSorST group were used to extract Indian gooseberry, respectively, and the extraction conditions were the same as those described in Example 1 (i.e., step S3). The Indian gooseberry extracts obtained from each group were quantitatively analyzed by HPLC, and the gallic acid content of the Indian gooseberry extracts was used as a comparison basis. The results are shown in Table 3.

[0047] Table 3 Group Gallic acid (μg / mL) BetSorST 1249 Zemea 280 Ethanol 300

[0048] From the results in Table 3 above, it can be seen that when the natural deep eutectic solvent mixture and the compound emulsifier (i.e., the BetSorST group) are used as the extraction solvent, the gallic acid content of the Indian gooseberry extract is significantly increased compared to the general traditional organic solvents ethanol or propylene glycol.

[0049] Example 3 Comparison of Antioxidant Capacity

[0050] The Indian gooseberry extracts obtained from the groups with better extraction rates for gallic acid content of the Indian gooseberry extracts described in Comparative Example 2, i.e., GlyUST, BGFST, BetSorST, GlyLaST, etc., and the Indian gooseberry extracts obtained by extraction using ethanol and propylene glycol described in Example 2 were subjected to DPPH free radical scavenging ability test analysis, and the analysis method is as follows:

[0051] First, 50 μL of the prepared positive control group (Vit.C), GlyUST, BGFST, BetSorST, GlyLaST and other groups of Indian gooseberry extracts, and the Indian gooseberry extracts extracted with ethanol and propylene glycol were transferred to different wells of the 96-well cell analysis plate, and then 150 μL of 0.2 mM DPPH ethanol solution was added, mixed evenly, and allowed to react at room temperature in the dark for 30 minutes, and then the OD was measured by a microplate reader (SPECTROstar Nano). 517nm The absorbance value under the condition of 1:1 was measured, and each group was tested three times. The final concentration of the Indian gooseberry extract obtained by the extraction method of the present invention was 0.1wt%, and ascorbic acid (Vit.C) was used as the positive control group in this test. The calculation method of DPPH free radical scavenging rate was as follows: DPPH free radical scavenging rate (%) = (1-A / B) × 100% Where A is the sample group at OD 517nm B is the absorbance value of the control group at OD 517nm The absorbance value below.

[0052] See also Figure 2 , which is a schematic diagram of DPPH free radical scavenging ability; it can be seen from the figure that when the concentration of the Indian gooseberry extract obtained by the extraction method of the present invention is 0.1wt%, its DPPH free radical scavenging ability is much higher than that of the general traditional organic solvents ethanol or propylene glycol, and the BetSorST group has the best DPPH free radical scavenging ability, with an inhibition rate of 75%. Therefore, the extraction solvent composed of the natural deep eutectic solvent mixture and the compound emulsifier can indeed improve the antioxidant capacity of the Indian gooseberry extract compared with the general traditional organic solvent.

[0053] Example 4 In vitro tyrosinase inhibition assay

[0054] Similar to Example 3, the Indian gooseberry extracts obtained from the GlyUST, BGFST, BetSorST, GlyLaST groups, and the Indian gooseberry extracts obtained by extraction with ethanol and propylene glycol were subjected to in vitro tyrosinase inhibition analysis, and the analysis method is as follows:

[0055] First, 50 μL of the prepared positive control group (kojic acid), GlyUST, BGFST, BetSorST, GlyLaST and other groups of Indian gooseberry extracts, and the Indian gooseberry extracts extracted with ethanol and propylene glycol were transferred to different wells of the 96-well cell analysis plate, and then 100 μL of L-Dopa solution and 50 μL of Tyrosinase solution were added, mixed evenly, and allowed to react at 37°C in the dark for 20 minutes. The OD values ​​were measured at 0 and 20 minutes using a microplate reader. 475nm The absorbance value under the condition of 1:1 was measured, and each group was tested three times. The final concentration of the Indian gooseberry extract obtained by the extraction method of the present invention was 5wt%, and kojic acid was used as the positive control group in this test. The calculation method of tyrosinase inhibition rate was as follows: Tyrosinase inhibition rate (%) = ((A 20min -A 0min )-(B 20min -B 0min )) / (A 20min -A 0min )×100% where A is the control group at OD 475nm The absorbance value of the sample group at OD 475nm The absorbance value below.

[0056] See also Figure 3 , which is a schematic diagram of in vitro tyrosinase inhibition rate; as can be seen from the figure, when the concentration of the Indian gooseberry extract obtained by the extraction method of the present invention is 5wt%, its in vitro tyrosinase inhibition ability is much higher than that of the general traditional organic solvents ethanol or propylene glycol, and the GlyLaST group has the best in vitro tyrosinase inhibition ability, with an inhibition rate of 100% (because lactic acid itself has a slight inhibitory effect on tyrosinase activity, it has a multiplying effect), and the BetSorST group is second, with an inhibition rate of 42.15%. Therefore, the extraction solvent composed of the natural deep eutectic solvent mixture and the compound emulsifier can indeed improve the tyrosinase inhibition ability of the Indian gooseberry extract compared with the general traditional organic solvent.

[0057] Example 5 Analysis of the inhibitory ability of pro-inflammatory factor interleukin-1β (IL-1β)

[0058] Similar to Example 4, the Indian gooseberry extracts obtained from the BGFST, BetSorST, GlyLaST and other groups (because the experimental data of the GlyUST group was significantly worse than the above groups, the subsequent experiments did not provide its experimental data for comparison), and the Indian gooseberry extract obtained by extraction with propylene glycol (because ethanol is cytotoxic to RAW264.7 cells, the Indian gooseberry extract obtained was not used for the test) were analyzed for IL-1β inhibition ability, and the analysis method is as follows:

[0059] Mouse macrophages (RAW264.7) were cultured at a cell density (2×10 5 cell / well) in a 24-well plate with a total volume of 500 μL and cultured at 37°C and 5% CO2 for at least 12 hours; the next day, the old culture medium was removed, the plate was washed with PBS and replaced with DMEM medium (200 μL / well) containing the Indian gooseberry extract obtained by the extraction method of the present invention for 1 hour, and then 5 μL LPS (1 μg / mL) was added to each well to induce inflammation, and cultured at 37°C and 5% CO2 for 24 hours. The cell culture supernatant was collected from the 24-well plate the next day for analysis by RayBio Mouse IL-1β ELISA Kit (Catalog#: ELM-1L 1b).

[0060] See also Figure 4 , which is a schematic diagram of the IL-1β inhibitory ability; it can be seen from the figure that the content of IL-1β produced by RAW264.7 cells after LPS induction is 94.45pg / mL. The lower the IL-1β content, the more anti-inflammatory it is. When the concentration of the Indian gooseberry extract obtained by the extraction method of the present invention is 0.1wt%, it has the ability to significantly inhibit the production of IL-1β, indicating that the Indian gooseberry extract has an anti-inflammatory effect; among them, the BetSorST group has the best inhibitory effect. However, the Indian gooseberry extract obtained by extraction with the general traditional organic solvent propylene glycol does not reduce the expression of the pro-inflammatory protein IL-1β, but instead promotes inflammation.

[0061] Example 6 UV light protection test

[0062] Similar to Example 5, the Indian gooseberry extracts obtained by the BGFST, BetSorST, GlyLaST groups (the experimental data of the GlyUST group was significantly worse than the above groups, so the subsequent experiments did not provide its experimental data for comparison), and the Indian gooseberry extracts obtained by extraction with ethanol and propylene glycol were subjected to UV light protection analysis, and the analysis method was as follows:

[0063] Keratinocytes (HaCaT) were cultured at a cell density (3×10 5The cells were seeded into 24-well plates with a total volume of 500 μL, divided into 2 plates, and cultured at 37°C, 5% CO2 for at least 12 hours; the next day, the medium was replaced with serum-free DMEM medium containing the Indian gooseberry extract obtained by the extraction method of the present invention, and then cultured at 37°C, 5% CO2 for 24 hours. The 24-well plates were washed with PBS the next day, and one plate of cells was placed in a nucleic acid fixation instrument with the cover opened and irradiated with UVB 2.5 J / cm 2 After irradiation, PBS was removed and replaced with 500 μL serum-free DMEM medium. After culturing for 1 hour, 50 μL MTT reagent was added to each cell and reacted in the incubator for 30 minutes. After removing the MTT culture medium, 500 μL DMSO was added to dissolve the blue-purple crystals and the OD was calculated. 570nm The absorbance value was measured. The final concentration of the Indian gooseberry extract obtained by the extraction method of the present invention was 0.1 wt %, and Silymarin was used as a positive control group in this experiment.

[0064] See also Figure 5 , which is a schematic diagram of UV light protection ability; it can be seen from the figure that HaCaT cells caused light damage after UV light irradiation, the photoprotection ability of the blank group was 0%, and the photoprotection ability of the positive control group (Silymarin) was 37%. The Indian gooseberry extract obtained by the extraction method of the present invention has a significant photoprotection ability when the concentration is 0.1wt%; among them, the photoprotection ability of the GlyLaST group is the best, reaching 54.53%, and the photoprotection ability of the Indian gooseberry extract of the present invention is significantly higher than that of the general traditional organic solvents ethanol and propylene glycol, and its protection ability is even higher than that of silymarin as the positive control group.

[0065] Example 7 Whitening Test

[0066] Similar to Example 5, the Indian gooseberry extracts obtained from the BGFST, BetSorST, GlyLaST groups (the experimental data of the GlyUST group was significantly worse than the above groups, so the subsequent experiments did not provide its experimental data for comparison), and the Indian gooseberry extracts obtained by extraction with ethanol and propylene glycol were subjected to whitening analysis, and the analysis method is as follows:

[0067] Mouse melanocytes (B16F10) were cultured at a cell density (1×10 5Cells were seeded into 24-well plates with a total volume of 500 μL and cultured at 37°C and 5% CO2 for at least 12 hours. The next day, the culture medium was replaced with DMEM medium containing the Indian gooseberry extract obtained by the extraction method of the present invention and 500 nM inducer α-MSH, and then cultured at 37°C and 5% CO2 for 48 hours. The supernatant was removed and 2N NaOH solution was added. The cells were shaken for 3 to 5 minutes until the cells were dissolved and transferred to a microcentrifuge tube (Eppendorf). After heating at 80°C for 60 minutes, the OD was measured using an enzyme immunoassay analyzer. 405nm Absorbance value. The final concentration of the Indian gooseberry extract obtained by the extraction method of the present invention is 0.2wt%, and in this experiment, oxidized resveratrol is used as the positive control group, and the calculation method of the melanin inhibition rate is as follows: Melanin inhibition rate (%) = (AB) / A×100% Where A is α-MSH in OD 405nm The absorbance value of the sample group at OD 405nm The absorbance value below.

[0068] See also Figure 6 , which is a schematic diagram of the ability to inhibit melanin production; it can be seen from the figure that after the addition of melanin stimulating hormone ɑ-MSH induction, the amount of melanin production increased to 144.26%. When the concentration of the Indian gooseberry extract obtained by the extraction method of the present invention is 0.2wt%, it has the ability to significantly inhibit melanin production; among them, the melanin content of the GlyLaST group is the lowest, at 61.76%. Compared with the group of melanin stimulating hormone ɑ-MSH, its melanin inhibition rate is as high as 82.50%, and the ability of the Indian gooseberry extract of the present invention to inhibit melanin production is significantly higher than that of the general traditional organic solvents ethanol and propylene glycol.

[0069] In summary, the Indian gooseberry extract and the extraction method thereof provided by the present invention can significantly increase the active ingredients in the Indian gooseberry extract by using a mixture of a natural deep eutectic solvent and a compound emulsifier as an extraction solvent, compared with extraction using a conventional organic solvent. In addition, the Indian gooseberry extract obtained by the extraction method of the present invention has significantly improved antioxidant, anti-inflammatory, whitening and photoprotective effects compared with the Indian gooseberry extract obtained by extraction using a conventional organic solvent.

[0070] Those skilled in the art to which the present invention belongs can understand from the foregoing that the present invention can be illustrated by other specific embodiments without changing the technical concepts or essential features of the present invention. In this regard, the embodiments disclosed herein are only for illustrative purposes and should not be interpreted as limiting the scope of the present invention. On the contrary, the present invention tends to cover not only the above-mentioned embodiments, but also various changes, modifications, equivalents, and other embodiments within the spirit and scope of the present invention as defined in the appended patent claims.

Claims

1. An Indian gooseberry extract, characterized in that The Indian gooseberry extract is extracted by a mixture of natural deep eutectic solvents (NaDES), wherein The gallic acid concentration of the Indian gooseberry extract is greater than 500 μg / mL.

2. The Indian gooseberry extract according to claim 1, characterized in that The mixed solution of the natural deep eutectic solvents comprises at least two natural deep eutectic solvents, and the natural deep eutectic solvents are selected from glycerol, urea, 1,3-butanediol, fructose, betaine, sorbitol and / or lactic acid.

3. The Indian gooseberry extract according to claim 2, characterized in that The mixed solution of the natural deep eutectic solvent comprises a combination of the following natural deep eutectic solvents: Glycerol and urea, 1,3-butylene glycol and fructose, betaine and sorbitol or glycerol and lactic acid, wherein In the combination of natural deep eutectic solvents, the natural deep eutectic solvents are mixed in a molar ratio of 1:

1.

4. The Indian gooseberry extract according to any one of claims 1 to 3, characterized in that The mixed solution of the natural deep eutectic solvent is emulsified by at least one emulsifier, and the emulsifier is selected from sorbitan monolaurate, sorbitan oleate and / or polysorbate-80.

5. The Indian gooseberry extract according to claim 4, characterized in that The mixed liquid of the natural deep eutectic solvent is emulsified by a compound emulsifier composed of sorbitan oleate and polysorbate-80, and in the compound emulsifier, sorbitan oleate accounts for 60% and polysorbate-80 accounts for 40%.

6. The Indian gooseberry extract according to claim 5, characterized in that In the mixed solution of the natural deep eutectic solvent, each of the natural deep eutectic solvents and the compound emulsifier are mixed in a molar ratio of 1:1:

1.

7. The Indian gooseberry extract according to claim 5, characterized in that The hydrophile-lipophile balance (HLB) value of the compound emulsifier is between 8 and 10.

8. A method for extracting Indian gooseberry extract according to any one of claims 1 to 7, comprising the following steps: Step S1: dissolving at least two natural deep eutectic solvents at 50° C. and then mixing them to obtain a mixed solution of natural deep eutectic solvents; Step S2: adding at least one emulsifier to the mixed solution of the natural deep eutectic solvent, stirring evenly at 50° C. to obtain an emulsified mixed solution; and Step S3: extracting Indian gooseberry with the emulsified mixed solution to obtain Indian gooseberry extract.

9. The extraction method according to claim 8, characterized in that In step S3, extraction is performed by ultrasonic method at 30 to 60°C for 10 to 30 minutes.

10. The extraction method according to claim 8, characterized in that In step S3, the weight ratio of the emulsified mixed solution to the Indian gooseberry is 4:

1.

11. Use of the Indian gooseberry extract according to any one of claims 1 to 7, wherein the use is selected from anti-oxidation, inhibition of tyrosinase production, inhibition of IL-1β production, UV light protection and / or inhibition of melanin production.

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