Preparation method of modified ganoderma lucidum extract with whitening effect

Abstract

The invention belongs to the technical field of deep processing of lucid ganoderma, and particularly relates to a preparation method of a modified lucid ganoderma extract with a whitening effect, which comprises the following steps: I, modifying carboxymethyl chitosan with hyaluronic acid to obtain modified carboxymethyl chitosan; iI, the modified carboxymethyl chitosan obtained in the step I is chlorinated with thionyl chloride, and chlorinated modified carboxymethyl chitosan is obtained; and III, under the protection of inert gas, dissolving the chlorinated modified carboxymethyl chitosan, the lucid ganoderma extract and pyridine in dichloromethane, stirring at 25-35 DEG C for 35-37 hours, filtering, washing and drying to obtain the modified lucid ganoderma extract. The modified ganoderma lucidum extract provided by the invention has a relatively high inhibition rate on tyrosinase, is good in skin absorbability and has a whitening effect.

Description

Technical Field

[0001] The invention belongs to the technical field of deep processing of ganoderma lucidum, and in particular relates to a method for preparing a modified ganoderma lucidum extract with whitening effect. Background Art

[0002] Ganoderma lucidum is a traditional Chinese medicinal and edible fungus with a history of more than 2,000 years. It has extremely high pharmacological activity. Triterpenes and polysaccharides are the main active ingredients. Polysaccharides can improve immunity, promote metabolism, effectively prevent the formation of melanin, and make the skin white and tender. Triterpenes have the effect of inhibiting tyrosinase activity, reducing the formation of melanin, and can effectively lighten dark spots and freckles. They are widely used in the preparation of whitening cosmetics. Ganoderma lucidum exists in three forms: Ganoderma lucidum fruiting body, Ganoderma lucidum spore powder, and Ganoderma lucidum mycelium. Ganoderma lucidum fruiting body is usually used. The mature Ganoderma lucidum fruiting body is highly lignified, so it cannot be eaten directly. The outer wall of the Ganoderma lucidum fruiting body cell is mainly composed of cellulose, hemicellulose, chitin and lignin, etc. The structure is tight, which causes the extraction of active ingredients such as Ganoderma lucidum polysaccharides and triterpenes to have the problem of long extraction time and incomplete extraction. Therefore, how to efficiently destroy the cell wall to improve the extraction rate of active ingredients in Ganoderma lucidum has become a hot issue in current research.

[0003] Patent CN113425751B discloses a method for preparing a ganoderma extract: using dried ganoderma fruiting bodies as raw materials, the fruiting bodies are firstly crushed by a coarse crusher, and then ultrafinely broken by a freezing crusher at low temperature, and directed enzymatic hydrolysis by biological enzymes, and then heated for extraction, centrifugation, reduced pressure concentration, vacuum drying, semi-finished product crushing, sieving and other processes to prepare the ganoderma extract. Patent CN109893547B discloses a method for extracting ganoderma triterpenic acid and ganoderma polysaccharide from ganoderma, (1) raw material pretreatment: putting the crushed ganoderma fruiting bodies into an aqueous solution of sodium hydroxide for extraction, filtering after ultrasonic treatment, and obtaining a filtrate and an extract; (2) adjusting pH; (3) dynamic reflux adsorption: (4) segmented elution; the invention effectively improves the extraction rate of ganoderma triterpenic acid and ganoderma polysaccharide and the purity of ganoderma triterpene. Although the above patents extract the active ingredients in ganoderma, the skin absorbs them slowly and the whitening effect is not significant.

[0004] Therefore, there is an urgent need for a method for preparing a modified Ganoderma lucidum extract with whitening effect. Summary of the invention

[0005] In order to overcome the deficiencies of the above-mentioned prior art, the present invention provides a method for preparing a modified Ganoderma lucidum extract with whitening effect, wherein the cell walls of Ganoderma lucidum are broken by adding a wall-breaking extraction liquid, thereby reducing the loss of active ingredients in Ganoderma lucidum and improving the extraction rate, and then the Ganoderma lucidum extract is modified by using modified carboxymethyl chitosan so that the ingredients in the Ganoderma lucidum extract are more easily absorbed by the skin, thereby increasing the whitening effect.

[0006] In order to achieve the above object, the technical solution adopted by the present invention is:

[0007] The first aspect of the present invention provides a method for preparing a modified Ganoderma lucidum extract having whitening effect, characterized in that the method comprises the following steps:

[0008] I. adding hyaluronic acid, carboxymethyl chitosan and sodium hydroxide into ethylene dichloride, reacting at 70-80° C. for 7-8 hours, filtering, washing and drying to obtain modified carboxymethyl chitosan;

[0009] II. The modified carboxymethyl chitosan obtained in step I is added to N, N-dimethylformamide, and thionyl chloride is added while stirring, and the reaction is carried out at 0-5°C for 3-5h, filtered, washed, and dried to obtain chlorinated modified carboxymethyl chitosan;

[0010] III. Under the protection of inert gas, the chlorinated modified carboxymethyl chitosan, Ganoderma lucidum extract and pyridine were dissolved in dichloromethane, stirred at 25-35° C. for 35-37 hours, filtered, washed and dried to obtain the modified Ganoderma lucidum extract.

[0011] Preferably, the mass ratio of the modified carboxymethyl chitosan to thionyl chloride in step II is 1:(0.5-0.7).

[0012] The present invention firstly utilizes hyaluronic acid to modify carboxymethyl chitosan, and grafts hyaluronic acid onto the carboxymethyl chitosan to enhance the water retention of the skin, while promoting the proliferation and differentiation of skin cells to repair damaged skin. The modified carboxymethyl chitosan is then chlorinated and reacted with the ganoderma lucidum extract. The two are connected by an ether bond to increase the affinity with the skin, which is beneficial to the absorption of the effective ingredients in the ganoderma lucidum extract.

[0013] In some embodiments, the mass ratio of hyaluronic acid to carboxymethyl chitosan in step I is (1-2):1.

[0014] In some embodiments, the mass ratio of the chlorinated modified carboxymethyl chitosan to the Ganoderma lucidum extract in step III is (0.3-0.5):1.

[0015] The present invention limits the mass ratio of hyaluronic acid to carboxymethyl chitosan so that the carboxyl groups of carboxymethyl chitosan are fully reacted to reduce irritation to the skin, and further limits the mass ratio of chlorinated modified carboxymethyl chitosan to ganoderma lucidum extract so that the chlorine atoms in the chlorinated modified chitosan are fully reacted, thereby enhancing the affinity of the ganoderma lucidum extract to the skin.

[0016] In some embodiments, the extraction process of the Ganoderma lucidum extract comprises the following steps:

[0017] (1) Wall breaking: The crushed Ganoderma lucidum is placed in a wall breaking extraction solution to fully dissolve the active ingredients, thereby obtaining a solid-liquid mixture, and then centrifuged to separate the solid phase and the liquid phase;

[0018] (2) Separation and purification: The liquid phase obtained in step (1) is separated, and the aqueous phase is extracted, purified, and dried to obtain the Ganoderma lucidum extract.

[0019] Preferably, the Ganoderma lucidum in step (1) is a Ganoderma lucidum fruiting body.

[0020] In some embodiments, the cell wall breaking step in step (1) comprises: adding the crushed Ganoderma lucidum to the cell wall breaking extract, ultrasonicating for 25-35 minutes with an ultrasonic power of 80-100W, filtering and repeating the above operation 2-4 times on the residue, and combining the filtrate.

[0021] In some embodiments, the raw materials of the cell wall-breaking extract in step (1) include: deionized water and 1-carboxyethyl-3-methylimidazolium tetrafluoroborate.

[0022] In some embodiments, the mass ratio of deionized water to 1-carboxyethyl-3-methylimidazolium tetrafluoroborate is 1:(0.11-0.16).

[0023] In some embodiments, the method for preparing 1-carboxyethyl-3-methylimidazolium tetrafluoroborate comprises the following steps:

[0024] S1. Dissolve 1-(allyl)-3-methylimidazolium chloride in dichloromethane, and react at -77°C to -79°C with O3 for 1.5-2h, then add triphenylphosphine, stir at room temperature for 25-35min, quench the reaction, purify, and dry to obtain an intermediate;

[0025] S2, dissolving the intermediate obtained in step S1 in dichloromethane, passing O2, adding a catalyst, reacting at 60-80° C. for 1-2 h, then separating, purifying, and drying to obtain 1-carboxyethyl-3-methylimidazolium chloride;

[0026] S3. Add the 1-carboxyethyl-3-methylimidazolium chloride and sodium tetrafluoroborate in step S2 to ethyl acetate, stir for 6-8 hours, filter, wash and dry to obtain 1-carboxyethyl-3-methylimidazolium tetrafluoroborate.

[0027] In some embodiments, the molar ratio of 1-(allyl)-3-methylimidazolium chloride to triphenylphosphine in step S1 is 1:(2-2.2).

[0028] In some embodiments, the molar ratio of 1-carboxyethyl-3-methylimidazolium chloride to sodium tetrafluoroborate in step S3 is 1:(1-1.2).

[0029] Currently, the commonly used methods for breaking the cell wall of Ganoderma lucidum are enzymatic hydrolysis and external force-assisted method. The enzymatic hydrolysis method uses the specific decomposition effect of enzymes to rupture the cell wall, but during the extraction process, the enzyme may denature, resulting in incomplete dissolution of the active ingredients and a low extraction rate. The external force-assisted method usually uses microwaves, ultrasound and other equipment to generate rapid vibrations to generate a large amount of heat energy, temperature and high pressure to rupture the cell membrane and cell wall and promote the precipitation of active ingredients. This method is pollution-free and not easy to denature, but the equipment cost is relatively high.

[0030] The applicant has found that the active ingredients in Ganoderma lucidum can be fully dissolved by using a cell wall breaking extracting solution to break the cell wall of plant cells, and with the assistance of ultrasound. The present invention uses deionized water and 1-carboxyethyl-3-methylimidazolium tetrafluoroborate as the cell wall breaking extracting solution, and limits the mass ratio of the two so that the cell wall can be quickly broken and the active ingredients can be fully dissolved. 1-carboxyethyl-3-methylimidazolium tetrafluoroborate is environmentally friendly, pollution-free, easy to separate from the product, easy to recycle and easy to use, avoiding serious environmental, health, safety and equipment corrosion problems, and is suitable for the requirements of the currently advocated clean technology and sustainable development. At the same time, when the cell wall is broken, the carboxyl group can chelate with the metal ions in the cell, and the holes on the cell wall surface are enlarged, so that the active ingredients are dissolved more fully, and at the same time, it is conducive to the removal of metal ions and increases the extraction rate.

[0031] In some embodiments, the method for purifying the aqueous phase in step (2) is an alcohol precipitation method.

[0032] Preferably, the alcohol precipitation method comprises the following steps: adding 95% ethanol in an amount 3-4 times the volume of the aqueous phase to the aqueous phase, stirring evenly and standing for 24 hours, then centrifuging to remove the supernatant, and drying.

[0033] In some embodiments, the extraction rate of Ganoderma lucidum polysaccharide in the Ganoderma lucidum extract is ≥ 91%.

[0034] Compared with the prior art, the present invention has the following beneficial effects:

[0035] 1. The present invention obtains a modified Ganoderma lucidum extract by extracting Ganoderma lucidum and modifying the extract, which has high affinity with the skin and has a good whitening effect.

[0036] 2. The present invention firstly utilizes hyaluronic acid to modify carboxymethyl chitosan, and grafts hyaluronic acid onto carboxymethyl chitosan to enhance the water retention of the skin, while promoting the proliferation and differentiation of skin cells to repair damaged skin. The modified carboxymethyl chitosan is then chlorinated and reacted with the ganoderma lucidum extract. The two are connected by an ether bond to increase the affinity with the skin, which is beneficial to the absorption of the effective ingredients in the ganoderma lucidum extract.

[0037] 3. The present invention uses deionized water and 1-carboxyethyl-3-methylimidazolium tetrafluoroborate as cell wall-breaking extracting solutions, and limits the mass ratio of the two so that the cell wall can be quickly broken and the active ingredients are fully dissolved. 1-carboxyethyl-3-methylimidazolium tetrafluoroborate is environmentally friendly, pollution-free, easy to separate from the product, easy to recycle and easy to use. At the same time, when the cell wall is broken, the carboxyl group can chelate with the metal ions in the cell, and the holes on the cell wall surface are enlarged, so that the active ingredients are more fully dissolved, and at the same time, it is beneficial to the removal of metal ions and the extraction rate is increased. DETAILED DESCRIPTION

[0038] The present invention will be described below in conjunction with specific embodiments. It should be noted that the following examples and comparative examples are examples of the present invention and are only used to illustrate the present invention, but not to limit the present invention. Other combinations and various modifications within the concept of the present invention may be performed without departing from the spirit or scope of the present invention.

[0039] In order to facilitate those skilled in the art to implement the present invention, some raw materials and manufacturers of the embodiments and comparative examples are described as follows: zebrafish embryo culture medium was purchased from Shanghai Feixi Biotechnology Co., Ltd.; Ganoderma lucidum fruiting bodies were planted by our company.

[0040] Preparation Example 1

[0041] The preparation method of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1 comprises the following steps:

[0042] S1, dissolving 16g of 1-(allyl)-3-methylimidazolium chloride in dichloromethane, reacting at -78°C with O3 for 2h, then adding 52.4g of triphenylphosphine, stirring at room temperature for 30min, quenching the reaction, purifying, and drying to obtain an intermediate;

[0043] S2, dissolving 15 g of the intermediate obtained in step S1 in 100 ml of dichloromethane, passing O2, adding 0.15 g of manganese acetate, reacting at 70° C. for 1.5 h, then purifying by column chromatography and drying to obtain 1-carboxyethyl-3-methylimidazolium chloride;

[0044] S3. Add 17.7 g of 1-carboxyethyl-3-methylimidazolium chloride obtained in step S2 and 12.1 g of sodium tetrafluoroborate to 200 ml of ethyl acetate, stir at room temperature for 7 h, filter, wash and dry to obtain 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1.

[0045] Preparation Example 2

[0046] The preparation method of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-2 is the same as that of Preparation Example 1, except that the amount of triphenylphosphine added is 25 g.

[0047] Preparation Example 3

[0048] The extraction process of Ganoderma lucidum extract-1 includes the following steps:

[0049] (1) Wall breaking: 10 g of crushed Ganoderma lucidum was added to 1000 g of deionized water and 140 g of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1, and ultrasonicated for 30 min. The solid phase and the liquid phase were separated by centrifugation. The solid phase was then subjected to the above operation for 3 times, and the liquid phase was combined and the solid phase was stored for later use.

[0050] (2) Separation and purification: The liquid phase obtained in step (1) is separated, and the aqueous phase is extracted and purified: After the aqueous phase is extracted, a 95 wt% ethanol solution is added in an amount 3 times the volume of the aqueous phase, the mixture is stirred evenly and allowed to stand for 24 hours, then the supernatant is removed by centrifugation, and the mixture is dried at 35° C. for 6 hours to obtain Ganoderma lucidum extract-1.

[0051] Preparation Example 4

[0052] The specific implementation method of the extraction process of Ganoderma lucidum extract-2 is the same as that of Preparation Example 3, except that in step (1), 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1 is replaced by an equal amount of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-2.

[0053] Preparation Example 5

[0054] The specific implementation method of the extraction process of Ganoderma lucidum extract-3 is the same as that of Example 1, except that the amount of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1 added in step (1) is 100 g.

[0055] Preparation Example 6

[0056] The specific implementation method of the extraction process of Ganoderma lucidum extract-4 is the same as that of Example 1, except that the amount of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1 added in step (1) is 170 g.

[0057] Preparation Example 7

[0058] The extraction process of Ganoderma lucidum extract-5 comprises the following steps:

[0059] (1) Cell wall breaking: 10 g of crushed Ganoderma lucidum was added to 1000 g of deionized water and 0.05 g of cellulase, and ultrasonicated for 25 min at a power of 90 W at 95° C. to inactivate the enzyme, and centrifuged to separate the solid phase and the liquid phase. The solid phase was then subjected to the above operation twice, and the liquid phase was combined and the solid phase was stored for later use;

[0060] (2) Separation and purification: The liquid phase obtained in step (1) is separated, and the aqueous phase is extracted and purified: After the aqueous phase is extracted, a 95 wt% ethanol solution is added in an amount 3 times the volume of the aqueous phase, the mixture is stirred evenly and allowed to stand for 24 hours, and then the supernatant is removed by centrifugation, and the mixture is dried at 35° C. for 6 hours to obtain a Ganoderma lucidum extract.

[0061] Example 1

[0062] A method for preparing a modified ganoderma lucidum extract having whitening effect comprises the following steps:

[0063] I. Add 15g hyaluronic acid, 10g carboxymethyl chitosan and 1g sodium hydroxide into 100ml ethylene dichloride, react at 75°C for 7.5h, filter, wash and dry to obtain modified carboxymethyl chitosan;

[0064] II. The 10 g modified carboxymethyl chitosan obtained in step I was added to 100 ml N, N-dimethylformamide, 6 g thionyl chloride was added while stirring, and the reaction was carried out at 3 ° C for 4 h, filtered, washed, and dried to obtain chlorinated modified carboxymethyl chitosan;

[0065] III. Under nitrogen protection, 4 g of chlorinated modified carboxymethyl chitosan, 10 g of Ganoderma lucidum extract-1 and 1 g of pyridine were dissolved in 50 ml of dichloromethane, stirred at 30°C for 36 h, filtered, washed and dried to obtain the modified Ganoderma lucidum extract.

[0066] Example 2

[0067] A method for preparing a modified ganoderma lucidum extract having whitening effect comprises the following steps:

[0068] I. Add 10 g hyaluronic acid, 10 g carboxymethyl chitosan and 1 g sodium hydroxide into 100 ml ethylene dichloride, react at 70° C. for 8 h, filter, wash and dry to obtain modified carboxymethyl chitosan;

[0069] II. The 3g modified carboxymethyl chitosan obtained in step I was added to 100ml N,N-dimethylformamide, 5g of thionyl chloride was added while stirring, and the reaction was carried out at 0°C for 5h, filtered, washed, and dried to obtain chlorinated modified carboxymethyl chitosan;

[0070] III. Under nitrogen protection, 3 g of chlorinated modified carboxymethyl chitosan, 10 g of Ganoderma lucidum extract-2 and 1 g of pyridine were dissolved in 50 ml of dichloromethane, stirred at 25°C for 37 h, filtered, washed and dried to obtain the modified Ganoderma lucidum extract.

[0071] Example 3

[0072] A method for preparing a modified ganoderma lucidum extract having whitening effect comprises the following steps:

[0073] I. Add 20g hyaluronic acid, 10g carboxymethyl chitosan and 1g sodium hydroxide into 100ml ethylene dichloride, react at 80°C for 7h, filter, wash and dry to obtain modified carboxymethyl chitosan;

[0074] II. The 10 g modified carboxymethyl chitosan obtained in step I was added to 100 ml N, N-dimethylformamide, 7 g thionyl chloride was added while stirring, and the reaction was carried out at 5 ° C for 3 h, filtered, washed, and dried to obtain chlorinated modified carboxymethyl chitosan;

[0075] III. Under nitrogen protection, 5 g of chlorinated modified carboxymethyl chitosan, 10 g of Ganoderma lucidum extract-3 and 1 g of pyridine were dissolved in 50 ml of dichloromethane, stirred at 35 ° C for 35 h, filtered, washed and dried to obtain the modified Ganoderma lucidum extract.

[0076] Example 4

[0077] A method for preparing a modified Ganoderma lucidum extract with whitening effect, the specific implementation method is the same as Example 1, the difference is that the Ganoderma lucidum extract-1 is replaced by Ganoderma lucidum extract-2 in equal amounts.

[0078] Example 5

[0079] A method for preparing a modified Ganoderma lucidum extract with whitening effect, the specific implementation method is the same as Example 1, the difference is that the Ganoderma lucidum extract-1 is replaced by Ganoderma lucidum extract-3 in equal amounts.

[0080] Example 6

[0081] A method for preparing a modified Ganoderma lucidum extract with whitening effect, the specific implementation method is the same as Example 1, the difference is that the Ganoderma lucidum extract-1 is replaced by Ganoderma lucidum extract-4 in equal amounts.

[0082] Example 7

[0083] A method for preparing a modified Ganoderma lucidum extract with whitening effect, the specific implementation method is the same as Example 1, the difference is that the Ganoderma lucidum extract-1 is replaced by Ganoderma lucidum extract-5 in equal amounts.

[0084] Example 8

[0085] A method for preparing a modified Ganoderma lucidum extract with whitening effect, the specific implementation method is the same as Example 1, except that the added amount of hyaluronic acid is 8g.

[0086] Example 9

[0087] A method for preparing a modified Ganoderma lucidum extract with whitening effect, the specific implementation method is the same as Example 1, the difference is that the added amount of chlorinated modified carboxymethyl chitosan is 6g.

[0088] Performance Testing

[0089] 1. Tyrosinase inhibition test

[0090] DMSO and deionized water were mixed as solvents in a volume ratio of 1:19, and the modified Ganoderma lucidum extract obtained in the examples and comparative examples was configured into a 10 mg / mL solution with a solvent as a test solution. In a 96-well plate, 130 μL of phosphate buffer solution (50 mmol / L, pH=6.8), 10 μL of tyrosinase solution (final concentration 33.3 U / mL) and 10 μL of the test solution were added to each well and mixed evenly, 50 μL of L-Dopa (final concentration 0.5 mmol / L) was added and mixed evenly, reacted for 10 min, and the OD value was measured at a wavelength of 490 nm. 10 μL of solvent was added to the control group instead of the test solution. Each group was tested 3 times in parallel, and the average value was calculated. The inhibition rate of the test solution on tyrosinase was calculated according to the formula: inhibition rate (%) = (OD control-OD test solution) / OD control × 100%.

[0091] 2. Animal Experiments

[0092] Healthy zebrafish embryos developed to 24 hpf were placed in a 24-well plate at a rate of 10 embryos / well. The modified Ganoderma lucidum extract obtained in each embodiment and comparative example was dissolved in a solvent to prepare a 50 μg / ml solution as a test sample. The solvent was DMSO and zebrafish embryo culture medium, and the volume ratio of the two was 1:1000. 2 ml of the test sample was added to each well. The experiment was set up in 9 groups, and 4 parallels were set in each group. The culture was continued for 48 hours. Five zebrafish with normal phenotype and behavior in each group were randomly selected, anesthetized, and fixed on a slide with a 1wt% methylcellulose aqueous solution. The zebrafish melanin signal intensity was expressed by optical density value (OD) using Image j 64 image analysis software. The smaller the OD value, the better its whitening activity.

[0093] The test results are shown in Table 1.

[0094] Table 1

[0095]

[0096]

[0097] From the comparison of Examples 1-3 in the experimental data of Table 1, it can be seen that the modified Ganoderma lucidum extract has a higher tyrosinase inhibition rate and whitening effect; from the comparison of Example 1 and Example 4, it can be seen that 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-1 is replaced by an equal amount of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate-2, resulting in a decrease in the content of Ganoderma lucidum polysaccharide, and a decrease in the tyrosinase inhibition rate and whitening effect; from the comparison of Example 1 and Examples 5 and 6, it can be seen that too low a concentration of the broken wall extract will lead to incomplete wall breaking and incomplete active component precipitation, and too high a concentration of the broken wall extract will cause the extract viscosity to be too high, affecting the active components. The precipitation of the active ingredients leads to the decrease of the active ingredient content and the extraction rate, and the decrease of the tyrosinase inhibition rate and the whitening effect; from the comparison between Example 1 and Example 7, it can be seen that in the process of using cellulase to break the cell wall and extract the active ingredients, the enzyme may be denatured, resulting in incomplete dissolution of the active ingredients of the Ganoderma lucidum extract, and the decrease of the tyrosinase inhibition rate and the whitening effect; from the comparison between Example 1 and Example 8, it can be seen that the change of the ratio of hyaluronic acid and carboxymethyl chitosan may lead to the decrease of the repairability of the skin and the decrease of the whitening effect; from the comparison between Example 1 and Example 9, it can be seen that the change of the ratio of chlorinated modified carboxymethyl chitosan and Ganoderma lucidum extract leads to the decrease of the whitening effect.

[0098] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Although the present invention has been disclosed as a preferred embodiment as above, it is not used to limit the present invention. Any technician familiar with this profession can make some changes or modifications to equivalent embodiments of equivalent changes using the technical contents disclosed above without departing from the scope of the technical solution of the present invention. However, any simple modification, equivalent change and modification made to the above embodiments based on the technical essence of the present invention without departing from the content of the technical solution of the present invention still fall within the scope of the technical solution of the present invention.

Claims

1. A method for preparing a modified Ganoderma lucidum extract having whitening effect, characterized in that: The following steps are involved: I. adding hyaluronic acid, carboxymethyl chitosan and sodium hydroxide into ethylene dichloride, reacting at 70-80° C. for 7-8 hours, filtering, washing and drying to obtain modified carboxymethyl chitosan; II. The modified carboxymethyl chitosan obtained in step I is added to N, N-dimethylformamide, and thionyl chloride is added while stirring, and the reaction is carried out at 0-5°C for 3-5h, filtered, washed, and dried to obtain chlorinated modified carboxymethyl chitosan; III. Under the protection of inert gas, the chlorinated modified carboxymethyl chitosan, Ganoderma lucidum extract and pyridine were dissolved in dichloromethane, stirred at 25-35° C. for 35-37 hours, filtered, washed and dried to obtain the modified Ganoderma lucidum extract.

2. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 1, characterized in that: The mass ratio of hyaluronic acid to carboxymethyl chitosan in step I is (1-2):

1.

3. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 1, characterized in that: The mass ratio of the chlorinated modified carboxymethyl chitosan to the Ganoderma lucidum extract in step III is (0.3-0.5):

1.

4. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 1, characterized in that: The extraction process of the ganoderma lucidum extract comprises the following steps: (1) Wall breaking: The crushed Ganoderma lucidum is placed in a wall breaking extraction solution to fully dissolve the active ingredients, thereby obtaining a solid-liquid mixture, and then centrifuged to separate the solid phase and the liquid phase; (2) Separation and purification: The liquid phase obtained in step (1) is separated, and the aqueous phase is extracted, purified, and dried to obtain the Ganoderma lucidum extract.

5. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 4, characterized in that: The cell wall breaking step in step (1) comprises: adding the crushed Ganoderma lucidum to the cell wall breaking extract, ultrasonicating for 25-35 minutes with an ultrasonic power of 80-100W, and after filtering, repeating the above operation 2-4 times on the residue, and combining the filtrate.

6. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 4, characterized in that: The raw materials of the cell wall-breaking extract in step (1) include: deionized water and 1-carboxyethyl-3-methylimidazolium tetrafluoroborate.

7. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 6, characterized in that: The preparation method of 1-carboxyethyl-3-methylimidazolium tetrafluoroborate comprises the following steps: S1, dissolving 1-(allyl)-3-methylimidazolium chloride in dichloromethane, reacting with O3 at -77°C to -79°C for 1.5-2h, then adding triphenylphosphine, stirring for 25-35min, quenching the reaction, purifying, and drying to obtain an intermediate; S2, dissolving the intermediate obtained in step S1 in dichloromethane, passing O2, adding a catalyst, reacting at 60-80° C. for 1-2 h, then separating, purifying, and drying to obtain 1-carboxyethyl-3-methylimidazolium chloride; S3. Add the 1-carboxyethyl-3-methylimidazolium chloride and sodium tetrafluoroborate in step S2 to ethyl acetate, stir for 6-8 hours, filter, wash and dry to obtain 1-carboxyethyl-3-methylimidazolium tetrafluoroborate.

8. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 7, characterized in that: In the step S1, the molar ratio of 1-(allyl)-3-methylimidazolium chloride to triphenylphosphine is 1:(2-2.2).

9. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to claim 4, characterized in that: The method for purifying the aqueous phase in step (2) is an alcohol precipitation method.

10. The method for preparing the modified Ganoderma lucidum extract with whitening effect according to any one of claims 4 to 9, characterized in that: The extraction rate of ganoderma lucidum polysaccharide in the ganoderma lucidum extract is ≥91%.

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