Deep eutectic solvents and their applications, extracts containing berberine and their applications

Abstract

The application relates to the technical field of traditional Chinese medicine raw material extraction, in particular to a eutectic solvent and application thereof, and an extract containing berberine and application thereof. The eutectic solvent is composed of choline hydroxide and maleic acid, wherein the molar ratio of the choline hydroxide to the maleic acid is (2-4):1. The eutectic solvent can effectively extract berberine in traditional Chinese medicine raw materials, improve the yield of the extract containing berberine, and improve the content of berberine in the extract.

Description

Technical Field

[0001] This invention relates to the field of traditional Chinese medicine raw material extraction technology, and more specifically, to eutectic solvents and their applications, and extracts containing berberine and their applications. Background Technology

[0002] Preservatives are a crucial component of cosmetic safety, ensuring that cosmetics are not easily contaminated or spoiled by microorganisms during production, storage, transportation, and use. Currently, chemically synthesized preservatives, such as parabens and phenoxyethanol, are frequently used in cosmetics. These preservatives have significant antibacterial effects. However, research has found that some chemically synthesized preservatives are significant contributors to skin irritation, inflammation, allergies, and even cancer, posing certain safety risks. Therefore, safe, effective, and naturally derived preservatives better align with people's pursuit of health and have become a hot topic in cosmetic research. With increasing demands for natural, safe, and gentle products, the demand for natural and plant-based raw materials and preservatives will grow, leading to more in-depth research. Furthermore, many plants possess medicinal properties, allowing for further exploration of their applications.

[0003] Berberine can be used not only as a preservative but also for skin care, and can be used in skin care products. Currently, the commonly used methods for extracting berberine or its salts in industry are as follows: (1) Acid water method, but because this method uses acid water, it has the problems of corroding equipment and causing pollution. (2) Extraction with ethanol as solvent, which overcomes the disadvantages of the acid water method and has the characteristics of solvent recycling and reuse without corroding equipment. However, under the condition of similar extraction rate, the purity of the product obtained by ethanol leaching is low, and the same result is obtained even with ultrasonic-assisted extraction, thus limiting the application of organic solvent extraction methods.

[0004] In view of this, the present invention is proposed. Summary of the Invention

[0005] The purpose of this invention is to provide a eutectic solvent and its application, as well as an extract containing berberine and its application. An embodiment of this invention provides a eutectic solvent that exhibits good solubility for berberine, effectively extracting it from traditional Chinese medicine raw materials, thereby improving the extraction yield and purity of berberine.

[0006] This invention is implemented as follows:

[0007] In a first aspect, the present invention provides a eutectic solvent for preparing berberine, its salts, or extracts containing berberine, which is composed of choline hydroxide and maleic acid, wherein the molar ratio of choline hydroxide to maleic acid is (2-4):1.

[0008] In an optional embodiment, the molar ratio of choline hydroxide to maleic acid is (2-3.5):1.

[0009] Secondly, the present invention provides the use of the eutectic solvent described in the foregoing embodiments in the preparation of berberine, its salts, or extracts containing berberine.

[0010] In an optional embodiment, the preparation includes extracting the traditional Chinese medicine raw material containing berberine or its salt using the eutectic solvent;

[0011] Preferably, the herbal raw material is selected from Coptis chinensis.

[0012] Thirdly, the present invention provides a method for preparing berberine, its salts, or extracts containing berberine, comprising: extracting a traditional Chinese medicine raw material containing berberine or its salts using the eutectic solvent described in the foregoing embodiments.

[0013] In an optional embodiment, the eutectic solvent is mixed with the traditional Chinese medicine raw material and extracted at 40-60°C for 3.5-6.5 hours;

[0014] Preferably, post-processing is performed after extraction;

[0015] Preferably, the post-processing includes: filtration of the extraction mixture, adjustment of the pH of the filtrate to neutral, filtration again, washing of the filter residue to neutral, and drying.

[0016] Preferably, each gram of the traditional Chinese medicine raw material corresponds to 10-30 ml of the eutectic solvent;

[0017] Preferably, the herbal raw material is selected from Coptis chinensis.

[0018] Fourthly, the present invention provides berberine, its salt, or an extract containing berberine, which is prepared by the preparation method of berberine, its salt, or an extract containing berberine described in the foregoing embodiments.

[0019] Fifthly, the present invention provides a nanoemulsion whose raw materials include water, emulsifier, co-emulsifier, stabilizer, emollient, penetration enhancer, and berberine, its salt, or extract containing berberine as described in the foregoing embodiments.

[0020] In an optional embodiment, the raw materials, by weight percentage, comprise 7-15% emulsifier, 3-8% co-emulsifier, 6-10% stabilizer, 6.5-9.5% emollient, 0.5-1.5% penetration enhancer, 1-2.5% berberine, its salt or extract containing berberine, with the remainder being water.

[0021] Preferably, by weight percentage, the raw materials comprise 7-15% castor oil, 3-8% sorbitan oleate, 6-10% propylene glycol, 6.5-9.5% caprylic / capric triglycerides, 0.5-1.5% squalane, 1-2.5% berberine, its salts or extracts containing berberine, with the remainder being water.

[0022] In a sixth aspect, the present invention provides a preservative whose raw materials include berberine, its salt, or an extract containing berberine as described in the foregoing embodiments, or the nanoemulsion described in the foregoing embodiments.

[0023] The present invention has the following beneficial effects: The embodiments of the present invention form a new eutectic solvent by specifically selecting choline hydroxide and maleic acid and limiting their ratio. This eutectic solvent can effectively extract berberine from Chinese herbal raw materials, improve the yield of extracts containing berberine, and increase the content of berberine in the extracts. Attached Figure Description

[0024] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the embodiments will be briefly introduced below. It should be understood that the following drawings only show some embodiments of the present invention and should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can be obtained based on these drawings without creative effort.

[0025] Figure 1 This is an HPLC result chromatogram of the extract provided in Example 5 of the present invention;

[0026] Figure 2 This is an HPLC result chromatogram of the extract provided in Example 6 of the present invention;

[0027] Figure 3 This is an HPLC result chromatogram of the extract provided in Example 7 of the present invention;

[0028] Figure 4 This is an HPLC result chromatogram of the extract provided in Example 8 of the present invention;

[0029] Figure 5 The HPLC results of the extract provided in Comparative Example 1 of this invention are shown in the figure.

[0030] Figure 6 The image shows the HPLC results of the extract provided in Comparative Example 2 of this invention. Detailed Implementation

[0031] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer shall apply. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased commercially.

[0032] This invention provides a eutectic solvent that can be used to extract berberine from traditional Chinese medicine raw materials, thereby obtaining berberine or its salt, or an extract containing berberine. Specifically, the eutectic solvent is composed of choline hydroxide and maleic acid, wherein the molar ratio of choline hydroxide to maleic acid is (2-4):1, for example, any value between 2:1, 2.5:1, 3:1, 3.5:1, and 4:1, or any range between any two values, preferably (2-3.5):1.

[0033] The preparation of the aforementioned eutectic solvent involves mixing and stirring the materials to obtain a clear and homogeneous liquid. The stirring temperature can be either room temperature or 60°C.

[0034] In this embodiment of the invention, the choline hydroxide and maleic acid formed by the above-mentioned raw materials can effectively extract berberine from the Chinese herbal raw materials, thereby increasing the extraction rate and the content of berberine in the extract.

[0035] Therefore, this invention provides the application of the eutectic solvent described in the foregoing embodiments in the preparation of berberine, its salts, or extracts containing berberine. The preparation includes extracting a traditional Chinese medicine raw material containing berberine or its salt using the eutectic solvent; for example, the traditional Chinese medicine raw material includes, but is not limited to, Coptis chinensis. It is understood that other traditional Chinese medicine raw materials containing berberine can also be selected.

[0036] Specifically, the present invention provides a method for preparing berberine, its salts, or extracts containing berberine, comprising:

[0037] The above-mentioned eutectic solvent is mixed with the Chinese herbal raw materials, wherein each gram of the Chinese herbal raw materials corresponds to 10-30 ml of the eutectic solvent; for example, 1 gram of Chinese herbal raw materials corresponds to any value between 10-30 ml or any range between two values, such as 10 ml, 15 ml, 20 ml, 25 ml, and 30 ml.

[0038] Then, extract for 3.5-6.5 hours at 40-60℃. For example, temperatures can be any value between 40-60℃ or any range between any two values, such as 40℃, 43℃, 45℃, 47℃, 50℃, 51℃, 54℃, 55℃, 58℃, and 60℃. The extraction time can be any value between 3.5-6.5 hours, such as 3.5h, 4h, 4.5h, 5h, 6h, and 6.5h.

[0039] The mixture is then filtered to remove the herbal raw materials. The pH of the filtrate is then adjusted to neutral, 6.5-7.5. The residue is then collected by filtration, washed until neutral, and then dried to obtain the desired extract.

[0040] To obtain pure berberine, the extract can be further purified. The purification method can be any known purification method in the existing technology, such as column chromatography, crystallization, extraction, etc.

[0041] This invention also provides a nanoemulsion, the raw materials of which include berberine, its salt, or extracts containing berberine prepared as described above. Specifically, the raw materials for forming the nanoemulsion also include water, emulsifiers, co-emulsifiers, emollients, stabilizers, and penetration enhancers.

[0042] More specifically, by weight percentage, the raw materials comprise 7-15% emulsifier, 3-8% co-emulsifier, 6-10% stabilizer, 6.5-9.5% emollient, 0.5-1.5% penetration enhancer, 1-2.5% berberine, its salt or extract containing berberine, with the remainder being water; for example, the raw materials comprise 7-15% castor oil, 3-8% sorbitan oleate, 6-10% propylene glycol, 6.5-9.5% caprylic / capric triglycerides, 0.5-1.5% squalane, 1-2.5% berberine, its salt or extract containing berberine, with the remainder being water.

[0043] Castor oil includes, but is not limited to, PEG-40 castor oil.

[0044] The preparation of the above-mentioned nanoemulsions is a known method for preparing nanoemulsions, therefore, the embodiments of the present invention will not be described in detail.

[0045] Furthermore, the present invention provides a preservative whose raw materials include berberine, its salt, or an extract containing berberine as described in the foregoing embodiments, or the nanoemulsion described in the foregoing embodiments.

[0046] The features and performance of the present invention will be further described in detail below with reference to embodiments.

[0047] Example 1

[0048] This embodiment provides a eutectic solvent composed of choline hydroxide and maleic acid, wherein the molar ratio of choline hydroxide to maleic acid is 2:1.

[0049] The preparation method of this eutectic solvent includes:

[0050] Add the above materials to a beaker according to the specified ratio, and stir with a magnetic stirrer at 60°C until a clear and homogeneous liquid is obtained, thus obtaining the prepared eutectic solvent.

[0051] Examples 2-4 and Comparative Examples 1-15

[0052] Examples 2-4 and Comparative Examples 1-15 each provide a eutectic solvent, the only difference being the raw materials or their proportions, as shown in Table 1. Their preparation methods are the same as those in Example 1.

[0053] Table 1 Formulations of eutectic solvents

[0054]

[0055]

[0056] Detection Example 1

[0057] The eutectic solvents prepared in Examples 1-4 and Comparative Examples 1-15 were tested. The states of the eutectic solvents were observed and recorded after being placed at room temperature (25°C) for 24 hours, at 0°C for 24 hours, and at 40-50°C for 60 minutes by rotary evaporation. See Table 2 for details.

[0058] Table 2. Detection results of eutectic solvents

[0059]

[0060]

[0061] As can be seen, the eutectic solvent provided in the embodiments of the present invention remained a yellow and transparent liquid after being placed at room temperature (25°C) for 24 hours, at 0°C for 24 hours, and by rotary evaporation at 40-50°C for 60 minutes. Other comparative examples either showed solid analysis at room temperature, precipitated solids at low temperatures, or precipitated solids immediately upon rotary evaporation. These comparative examples' eutectic solvents all failed and cannot be used as extractants for berberine. However, a eutectic solvent can be prepared when the molar ratio of maleic acid to choline hydroxide is between 1:2 and 1:4. When the molar ratio of maleic acid to choline hydroxide is 1:3, the prepared eutectic solvent exhibits the best fluidity and a homogeneous and transparent liquid, which is beneficial for achieving a higher yield of berberine hydrochloride prepared from this eutectic solvent.

[0062] Example 5

[0063] This embodiment provides a method for preparing an extract containing berberine, comprising:

[0064] 1) After cleaning the Coptis chinensis, crush it to obtain Coptis chinensis powder.

[0065] 2) Add 20 mL of the eutectic solvent from Example 1 and 1.0 g of Coptis chinensis powder to a flat-bottomed flask (100 mL). Then, place the sample into the reactor and react and extract at 50 °C for 4-6 h. After the reaction is complete, filter the sample, adjust the pH of the filtrate, wash the extraction residue with anhydrous ethanol and distilled water until neutral, dry it in a 40 °C oven, and collect it for component analysis.

[0066] Examples 6-8

[0067] Examples 6-8 provide a method for preparing an extract containing berberine, which is the same as the method provided in Example 5, except that the eutectic solvent in Example 1 is changed to the eutectic solvent in Examples 2-4.

[0068] Comparative Example 1: This comparative example provides a method for preparing an extract containing berberine: 10g of finely ground Coptis chinensis was weighed into a 250mL round-bottom flask, 100mL of ethanol was added, and the flask was placed in an ultrasonic cleaner for ultrasonic extraction at 20kHz power and constant temperature of 60℃ for 30min. (Hereinafter referred to as alcohol extraction).

[0069] Comparative Example 2: This comparative example provides a method for preparing an extract containing berberine: 20g of Coptis chinensis was added to 200ml of water and decocted for 20 minutes. The mixture was filtered, allowed to stand and cool, and no precipitate formed. 4% NaCl was added to the filtrate, and a yellow precipitate formed. After filtration, the precipitate turned cherry red in bleaching water, confirming it as berberine hydrochloride. The precipitate was purified by recrystallization in water and dried to obtain high-quality berberine hydrochloride (hereinafter also referred to as water extraction).

[0070] Detection Example 2

[0071] The berberine content of the extracts prepared in Examples 5-8 and Comparative Examples 1-2 was determined by HPLC, as follows:

[0072] Chromatographic column: Hypersil BDS C18 (250 mm × 4.6 mm, 5 μm); Mobile phase: Acetonitrile: 0.02 mol·L⁻¹ -1 KH₂PO₄ = 30:70, pH = 3.0; Flow rate: 1.0 mL / min -1Detection wavelength: 350nm; column temperature: 30℃; injection volume: 10μL; theoretical plate number calculated based on berberine hydrochloride is greater than 3000. Under these conditions, the resolution of berberine hydrochloride in the sample from adjacent peaks is greater than 1.5.

[0073] Preparation of the reference solution: Accurately weigh 1.25 mg of berberine hydrochloride reference standard dried to constant weight, place it in a 10 mL volumetric flask, and dilute to the mark with methanol to obtain a concentration of 0.125 mg / mL. -1 The reference solution was shaken well and filtered through a 0.45 μm filter membrane to obtain the final product.

[0074] The results are shown in Table 3 and... Figure 1-6 .

[0075] Table 3 Results of extracting Coptis chinensis using different methods

[0076] Example Extraction solvent or method Berberine hydrochloride content Example 5 Example 1 3.4 mg / g Example 6 Example 2 5.21 mg / g Example 7 Example 3 4.13 mg / g Example 8 Example 4 2.27 mg / g Comparative Example 1 Alcohol extraction 0.051 mg / g Comparative Example 2 Water lifting 0.013mg / g

[0077] It should be noted that the berberine hydrochloride content mentioned above refers to the berberine hydrochloride content per gram of extract.

[0078] Detection Example 3

[0079] The antibacterial effects of the extracts prepared in Examples 6 and 7, and Comparative Examples 1-2, as well as the positive control substance, were tested. Details are as follows:

[0080] (1) Oxford cup test for Staphylococcus aureus

[0081] Take fresh culture from the activated third-generation Staphylococcus aureus slant and add 5 mL of sterile physiological saline to prepare a 10 6 -10 7 CFU / mL bacterial suspension, use a pipette to transfer 0.3 mL of the test bacterial suspension to a concentration of 10. 5 -10 6 Add CFU / mL to the surface of the above culture medium plate and spread evenly with a disposable spreader. Cover the plate and let it dry on a horizontal surface for 5 minutes. Using sterile forceps, gently place the sterilized and dried Oxford cups onto the prepared culture medium plate. Gently press the Oxford cups with the forceps to ensure better contact with the culture medium. Place four Oxford cups evenly in the plate, cover, and let stand on a horizontal work surface for 5 minutes. Use a micropipette to pipette 200 μL of the test sample into the Oxford cups and cover. Incubate upright in a 30°C biochemical incubator for 24 hours and observe the results. Measure and record the diameter of the inhibition zone using calipers. Repeat the experiment three times.

[0082] The experimental samples were prepared by diluting the above-mentioned extract and 0.5g of the positive control into 100mL of purified water, resulting in a concentration of 5mg / mL. The positive control drug was phenoxyethanol, with an aqueous solution concentration of 0.5%.

[0083] The results are shown in Table 4.

[0084] Table 4. Results of Staphylococcus aureus inhibition.

[0085] sample Diameter of the inhibition zone (mm) Water lifting 27.96、26.54、27.11 Alcohol extraction 32.49、30.05、30.12 Example 6 36.87、35.22、35.08 Example 7 28.63、29.19、27.16 Positive (phenoxyethanol) 20.54、22.90、20.40

[0086] As can be seen, the antibacterial effect is as follows: Example 6 > Example 7 > alcohol extraction > water extraction > positive result, indicating that the extracts of the present invention have excellent antibacterial effect.

[0087] (2) Malassezia Oxford cup test

[0088] Take fresh culture from the activated third-generation Malassezia furfur slant and add 5 mL of sterile physiological saline to prepare a 10... 6 -10 7 CFU / mL bacterial suspension, use a pipette to transfer 0.3 mL of the test bacterial suspension to a concentration of 10. 5 -10 6 Add CFU / mL to the surface of the above culture medium plate and spread evenly with a disposable spreader. Cover the plate and let it dry on a horizontal surface for 5 minutes. Using sterile forceps, gently place the sterilized and dried Oxford cups onto the prepared culture medium plate. Gently press the Oxford cups with the forceps to ensure better adhesion. Place four Oxford cups evenly in the plate, cover, and let stand on a horizontal work surface for 5 minutes. Use a micropipette to pipette 200 μL of the test sample into the Oxford cups and cover. Incubate upright in a 30°C biochemical incubator for 48 hours and observe the results. Measure and record the diameter of the inhibition zone using calipers. Repeat the experiment three times.

[0089] The experimental samples were the above-mentioned extract and 0.5g of the positive control, each diluted to 100mL of purified water to obtain a concentration of 5mg / mL. The positive control drug was OCT.

[0090] The results are shown in Table 5.

[0091] Table 5. Results of Malassezia's antibacterial activity

[0092] sample Diameter of the inhibition zone (mm) Water lifting 28.75、28.44、28.29 Alcohol extraction 35.93、37.02、36.93 Example 6 41.73、43.95、44.96 Example 7 40.91、41.00、39.89 Positive (OCT) 32.66、31.05、31.66

[0093] As can be seen, the antibacterial effect is as follows: Example 6 > Example 7 > alcohol extraction > positive result > water extraction, indicating that the extracts of the present invention have excellent antibacterial effect.

[0094] (3) Oxford cup test for Candida albicans

[0095] Take fresh culture from the activated third-generation Candida albicans slant culture medium and add 5 mL of sterile physiological saline to prepare a 10 6 -10 7 CFU / mL bacterial suspension, use a pipette to transfer 0.3 mL of the test bacterial suspension to a concentration of 10. 5 -10 6 Add CFU / mL to the surface of the above culture medium plate and spread evenly with a disposable spreader. Cover the plate and let it dry on a horizontal surface for 5 minutes. Using sterile forceps, gently place the sterilized and dried Oxford cups onto the prepared culture medium plate. Gently press the Oxford cups with the forceps to ensure better adhesion. Place four Oxford cups evenly in the plate, cover, and let stand on a horizontal work surface for 5 minutes. Use a micropipette to pipette 200 μL of the test sample into the Oxford cups and cover. Incubate upright in a 30°C biochemical incubator for 48 hours and observe the results. Measure and record the diameter of the inhibition zone using calipers. Repeat the experiment three times.

[0096] The experimental samples were the above-mentioned extract and 0.5g of the positive control, each diluted to 100mL of purified water to obtain a concentration of 5mg / mL. The positive control was OCT.

[0097] The results are shown in Table 6.

[0098] Table 6. Antibacterial results of Candida albicans

[0099] sample Diameter of the inhibition zone (mm) Water lifting 22.82、20.89、21.95 Alcohol extraction 22.86、21.47、23.65 Example 6 41.12、40.69、40.66 Example 7 37.21、36.22、36.13 OCT 31.88、32.06、32.01

[0100] As can be seen, the antibacterial effect is as follows: Example 6 > Example 7 > Positive result > Alcohol extraction > Water extraction, indicating that the extracts of the present invention have excellent antibacterial effects.

[0101] Examples 9-11

[0102] Examples 9-11 each provide a nanoemulsion with the following composition: 9.5% of the oil phase consists of caprylic / capric triglycerides and squalane; 15% is a mixed surfactant consisting of sorbitan oleate and PEG-40 castor oil; 8% is the co-surfactant propylene glycol (1.5% for the examples or comparative samples); and the remainder is the aqueous phase. See Table 7 for details.

[0103] Table 7. Composition of Nanoemulsions

[0104]

[0105]

[0106] The preparation method of the nanoemulsion is as follows: At room temperature, PEG-40 castor oil and sorbitan oleate are mixed as a compound emulsifier, propylene glycol as a co-emulsifier is added and mixed, and then caprylic / capric triglyceride and squalane are added to the oil phase. The mixture is placed on a magnetic stirrer to prepare three portions. After stirring the three portions at a speed of 400 r / min until homogeneous, purified water containing the same amount of preservative (Example 1, Example 2 and Example 4) is slowly added dropwise. After stirring for 30 min, three clear and transparent nanoemulsion solutions are obtained.

[0107] It should be noted that the nanoemulsion of Example 9 corresponds to the extract of Example 1, the nanoemulsion of Example 10 corresponds to the extract of Example 2, and the nanoemulsion of Example 11 corresponds to the extract of Example 4.

[0108] Detection Example 4

[0109] The nanoemulsions prepared in Examples 9-11 above were subjected to corrosion resistance tests, as detailed below:

[0110] 1. Bacteria: Staphylococcus aureus (ATCC 6538 recommended), Escherichia coli (ATCC 8739 recommended), and Pseudomonas aeruginosa (ATCC 9027 recommended). In addition to these three bacteria, depending on the characteristics of the product and raw materials, Burkholderia cepacia (ATCC 25416 recommended) or bacteria isolated from contaminated samples may be added as indicator strains, and the submitter should specify this requirement before testing.

[0111] 2. Fungi: Candida albicans (Candida albicans is recommended) (ATCC 10231), Aspergillus niger (ATCC 16404 is recommended).

[0112] 3. Generation number of strains: Freeze-dried strains obtained from the strain preservation center are the 0th generation, and the original freeze-dried strains after subculturing are the 1st generation. The 3rd to 5th generation strains should be selected as the working strains for the preservation challenge test.

[0113] 4. Inoculation method: Bacteria are inoculated as mixed bacteria (if the characteristics of the sample require single inoculation, the submitting party shall specify the requirements before the test); fungi are inoculated as single bacteria, that is, take two samples and inoculate Candida albicans and Aspergillus niger respectively.

[0114] 5. Inoculum concentration: For bacteria, whether in single-strain or mixed-strain tests, the inoculum concentration is 1.0-5.0 × 10⁻⁶. 6 CFU / mL (or CFU / g); for fungi, the inoculation concentration for Candida albicans is 1.0–5.0 × 10⁻⁶. 5 The inoculation concentration of Aspergillus niger is 1.0-5.0 × 10⁻⁶ CFU / mL (or CFU / g).4 CFU / mL (or CFU / g).

[0115] 6. Viable bacteria count: After adding the test bacterial solution to the sample to be tested, samples were taken at 0 hours, day 7, day 4, day 21 and day 28 for plate viable bacteria count.

[0116] 7. Inoculation and Sample Detection: Take the weighed cosmetic product (30g) and add 0.3mL of mixed bacterial suspension and 0.3mL of mixed fungal suspension respectively. Mix thoroughly using a vortex mixer. Store the sample at room temperature (22.5±2.5℃). Detect the colony count in the cosmetic product at 0, 1, 7, 14, 21, and 28 days. Specifically, add 1g or 1mL of the sample to 9mL of modified LETHEEN broth, mix thoroughly, and dilute to an appropriate concentration. Detect the colony count in the sample. Perform two parallel tests for each test.

[0117] See Table 8 for testing standards.

[0118] Table 8 Testing Standards

[0119]

[0120] The test results are shown in Tables 9-11.

[0121] Table 9. Detection results of nanoemulsions in Example 9

[0122]

[0123] Mixed bacteria, B standard; Candida albicans, A standard; Aspergillus niger, A standard.

[0124] Table 10 Detection results of nanoemulsion in Example 10

[0125]

[0126] Mixed bacteria, A standard; Candida albicans, A standard; Aspergillus niger, A standard.

[0127] Table 11 Detection results of nanoemulsion in Example 11

[0128]

[0129] Mixed bacteria, C standard; Candida albicans, A standard; Aspergillus niger, A standard.

[0130] This demonstrates that the nanoemulsion provided in the embodiments of the present invention has a good anti-corrosion effect.

[0131] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. The use of a eutectic solvent in the preparation of berberine, its salts, or extracts containing berberine, characterized in that, The eutectic solvent is composed of choline hydroxide and maleic acid, wherein the molar ratio of choline hydroxide to maleic acid is (2-4):1; The preparation includes extracting a traditional Chinese medicine raw material containing berberine or its salt using the eutectic solvent, wherein the raw material is selected from Coptis chinensis.

2. The application according to claim 1, characterized in that, The molar ratio of choline hydroxide to maleic acid is (2-3.5):

1.

3. A method for preparing berberine, its salt, or an extract containing berberine, characterized in that, include: A traditional Chinese medicine raw material containing berberine or its salt is extracted using a eutectic solvent, wherein the eutectic solvent is composed of choline hydroxide and maleic acid, wherein the molar ratio of choline hydroxide to maleic acid is (2-4):1, and the traditional Chinese medicine raw material is selected from Coptis chinensis.

4. The preparation method according to claim 3, characterized in that, The eutectic solvent is mixed with the Chinese herbal raw material and extracted at 40-60℃ for 3.5-6.5 hours.

5. The preparation method according to claim 4, characterized in that, Post-processing is performed after extraction.

6. The preparation method according to claim 5, characterized in that, Post-processing includes: The extract mixture was filtered, the pH of the filtrate was adjusted to neutral, then filtered again, and the filter residue was washed to neutral and then dried.

7. The preparation method according to claim 3, characterized in that, Each gram of the aforementioned Chinese herbal raw material corresponds to 10-30 ml of the aforementioned eutectic solvent.

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