A freckle-reducing and whitening essence and its preparation method

Abstract

The present invention provides a freckle-removing and whitening essence and a preparation method thereof, belonging to the technical field of cosmetics. It is prepared from the following raw materials in parts by weight: 10-20 parts of plant extract, 3-5 parts of hirudin / phloretin / ellagic acid chelate complex, 10-20 parts of emollient, 5-10 parts of humectant, 0.3-1 part of thickener, 0.5-3 parts of emulsifier, and 40-60 parts of deionized water. The freckle-removing and whitening essence prepared by the present invention has good effects of whitening, freckle-removing, firming the skin, antioxidant, anti-inflammatory, moisturizing, improving skin elasticity and delaying aging, and has wide sources of raw materials and simple preparation method, and has broad application prospects.

Description

Technical Field

[0001] The present invention relates to the technical field of cosmetics, and particularly relates to a freckle-removing and whitening essence and a preparation method thereof. Background Art

[0002] With the improvement of people's living standards and aesthetic standards, fair and clean skin is increasingly favored by most women. All along, whitening and freckle removal have been hot topics of concern. People hope to obtain fair, smooth, bright and translucent skin through the use of whitening care products, and some use functional whitening cosmetics to reduce the deposition of skin spots. Environmental factors such as ultraviolet rays, polluted air, and reactive oxygen species will accelerate the generation of melanin in the skin, accelerate skin aging, cause skin relaxation, skin spot deposition, dull yellow skin color, lack of luster and various fine lines and wrinkles.

[0003] There are many whitening active ingredients on the market, but they all have certain disadvantages. First, some whitening active ingredients will cause skin irritation, allergies and other phenomena; second, most whitening active ingredients are unstable, easily oxidized, deteriorated and discolored, thus losing their whitening effect; third, the epidermal permeability of some whitening active ingredients is poor, and it is difficult to penetrate the stratum corneum and enter the basal layer of the skin to play a role. Fourth, to reduce the generation of melanin by inhibiting the activity of tyrosinase, or to decompose, metabolize and reduce the already generated melanin to achieve the effect of whitening and freckle removal, the main active ingredients used are synthetic chemical preparations such as arbutin, sodium ascorbyl phosphate (vitamin C), kojic acid palmitate, etc., and the effect is limited and easy to rebound. Summary of the Invention

[0004] The purpose of the present invention is to provide a freckle-removing and whitening essence and a preparation method thereof, which have good effects of whitening, freckle removal, firming skin, antioxidant, anti-inflammatory, moisturizing, improving skin elasticity and delaying aging, and have a wide range of raw material sources and a simple preparation method, and have broad application prospects.

[0005] The technical solution of the present invention is realized as follows:

[0006] The present invention provides a freckle-removing and whitening essence, which is prepared from the following raw materials by weight: 10-20 parts of plant extract, 3-5 parts of hirudin / phloretin / ellagic acid chelate complex, 10-20 parts of emollient, 5-10 parts of humectant, 0.3-1 part of thickener, 0.5-3 parts of emulsifier, and 40-60 parts of deionized water.

[0007] As a further improvement of the present invention, the preparation method of the plant extract includes the following steps:

[0008] S1. Enzymolysis in deep eutectic solvent: Wash, dry, and pulverize birch bark, roots of Peristrophe baphica, European plum, peony root, and angelica root respectively to obtain a mixed powder. Add it to an aqueous solution containing a deep eutectic solvent, add a composite enzyme, carry out enzymolysis, inactivate the enzyme, and obtain an enzymolysis product;

[0009] S2. Fermentation: Sterilize the enzymolysis product in step S1, inoculate the seed solutions of Bifidobacterium longum and Bifidobacterium infantis, carry out fermentation culture, filter, wash the solid, collect the eluate and concentrate to obtain a bacterial solution, and keep the filtrate for use;

[0010] S3. Preparation of lysate of Saccharomyces boulardii: Freeze the bacterial solution obtained in step S2 in liquid nitrogen, dissolve it at room temperature, centrifuge, collect the supernatant, and freeze-dry to obtain the lysate of Saccharomyces boulardii;

[0011] S4. Preparation of polysaccharide composition: Add ethanol to precipitate the filtrate obtained in step S2, centrifuge, collect the solid, recover ethanol from the supernatant, keep the obtained solution for use, wash and dry the solid to obtain a polysaccharide composition;

[0012] S5. Separation and purification: Add an organic solvent to the solution in step S3, extract, collect the organic phase, and separate and purify with macroporous resin to obtain a flavonoid / saponin complex;

[0013] S6. Preparation of preservative: Mix vitamin C and vitamin E evenly to obtain a preservative;

[0014] S7. Preparation of plant extract: Mix evenly the lysate of Saccharomyces boulardii obtained in step S3, the polysaccharide composition obtained in step S4, the flavonoid / saponin complex obtained in step S5, and the preservative obtained in step S6 to obtain a plant extract.

[0015] As a further improvement of the present invention, in step S1, the mass ratio of the birch bark, roots of Peristrophe baphica, European plum, peony root, and angelica root is 3 - 5:1 - 3:7 - 12:3 - 5:1 - 2, the mass ratio of the mixed powder to the aqueous solution containing the deep eutectic solvent is 1:5 - 7, the composite enzyme is composed of a mixture of cellulase and pectinase, and the mass ratio is 10 - 15:3 - 5. The addition amount of the composite enzyme is 2 - 3 wt% of the total mass of the system. The water content in the aqueous solution containing the deep eutectic solvent is 40 - 60 wt%. The deep eutectic solvent is prepared by mixing choline chloride and glucose according to a molar ratio of 1:1. The temperature of the enzymolysis is 40 - 50 °C, and the time is 1 - 3 h; in step S2, the inoculation amounts of the seed solutions of Bifidobacterium longum and Bifidobacterium infantis are 1 - 2 v / v% and 2 - 4 v / v% respectively, and the bacterial content of the seed solution of the strain is 10 8 -10 9cfu / mL, and the conditions for the fermentation culture are anaerobic conditions, 36 - 39 °C, 150 - 200 r / min, and the fermentation culture is carried out for 24 - 48 h.

[0016] As a further improvement of the present invention, in step S4, ethanol is added to the system so that the ethanol content is 60 - 80 wt%, and the precipitation time is 3 - 5 h; in step S5, the mass ratio of the organic solvent to the solution is 1 - 3:2 - 4, the organic solvent is a mixed solvent of dichloromethane and tetrahydrofuran with a volume ratio of 3 - 5:1, and the macroporous resin is selected from at least one of XDA - 2, NKA - 9, SP - 825, D101, HPD600, X - 5, LSA - 10, NKI - 9, and AB - 8.

[0017] As a further improvement of the present invention, in step S6, the mass ratio of vitamin C to vitamin E is 10 - 15:3 - 7; in step S7, the mass ratio of the bifida ferment lysate, the polysaccharide composition, the flavonoid / saponin complex, and the preservative is 10 - 12:15 - 22:17 - 20:2 - 4.

[0018] As a further improvement of the present invention, the preparation method of the hirudin / phloretin / ellagic acid chelate complex is as follows:

[0019] T1. Add hirudin, phloretin, and ellagic acid to water, add tris(hydroxymethyl)aminomethane and tannic acid, adjust the pH value of the solution, and heat and stir to react to obtain a complex;

[0020] T2. Add the complex obtained in step T1 to water, add silver nitrate, stir to react, dialyze, and dry to obtain the hirudin / phloretin / ellagic acid chelate complex.

[0021] As a further improvement of the present invention, in step T1, the mass ratio of hirudin, phloretin, ellagic acid, tris(hydroxymethyl)aminomethane, and tannic acid is 12 - 15:3 - 5:2 - 4:2 - 4:5 - 7, the adjustment of the solution pH value is 8.2 - 8.7, the temperature of the heat and stir reaction is 40 - 50 °C, and the time is 1 - 3 h; in step T2, the mass ratio of the complex, water, and silver nitrate is 10 - 15:100:0.2 - 0.5, and the time of the stir reaction is 20 - 30 min.

[0022] As a further improvement of the present invention, the emollient is selected from at least one of triglyceride (ethylhexanoate), isononyl isononanoate, polydimethylsiloxane, caprylic / capric triglyceride, shea butter (BUTYROSPERMUM PARKII), and Limnanthes Alba seed oil; the humectant is selected from at least one of glycerol, butanediol, propylene glycol, hexanediol, sodium hyaluronate, betaine, trehalose, and 1,2-pentanediol; the emulsifier is selected from at least one of glyceryl stearate, PEG-100 stearate, cetearyl olivate, sorbitan olivate, C12-20 alkyl glucoside, and polyglyceryl-2 stearate; the thickener is selected from at least one of carbomer, acrylate / C10-30 alkyl acrylate cross-linked polymer, sodium polyacryloyldimethyl taurate, polyacrylate cross-linked polymer-6, and xanthan gum.

[0023] The present invention further provides a method for preparing the above-mentioned freckle-removing and whitening essence, comprising the following steps:

[0024] (1) Add the humectant and thickener to deionized water, heat to 70 - 80 °C, stir at 300 - 500 r / min for 10 - 20 min to obtain Phase A;

[0025] (2) Mix the emollient, emulsifier, and hirudin / phloretin / ellagic acid chelate complex, heat to 70 - 80 °C, stir at 300 - 500 r / min for 10 - 20 min to obtain Phase B;

[0026] (3) Add Phase B to Phase A, stir at 300 - 500 r / min for 5 - 15 min, emulsify at 7000 - 10000 r / min for 10 - 20 min, keep warm for 10 - 20 min, and then cool down;

[0027] (4) After cooling down to 35 - 40 °C, add the plant extract, stir at 300 - 500 r / min for 5 - 15 min, and filter to obtain the freckle-removing and whitening essence.

[0028] The present invention further provides an application of the above-mentioned freckle-removing and whitening essence in skin whitening, freckle removal, and moisturizing.

[0029] The present invention has the following beneficial effects:

[0030] The plant extract of the present invention uses birch bark, the root of Cyanotis arachnoidea C. B. Clarke, European plum, peony root, and angelica root as raw materials, and uses enzymes as catalysts to hydrolyze plant cell walls, promoting the diffusion of active ingredients from inside the cells into the extraction medium, solving the disadvantages of traditional extraction processes such as complex procedures, low efficiency, and poor extraction quality. However, the extraction efficiency of bioenzymes for water-insoluble components is poor. The present invention uses deep eutectic solvents as reaction media, greatly improving the extraction of compounds that are poorly soluble in water, and at the same time having many advantages such as low toxicity, greenness, dynamic tunability, non-flammability, and strong solubility for compounds. Glucose is used as one of the raw materials in the deep eutectic solvent of the present invention, providing a carbon source for subsequent fermentation.

[0031] The present invention ferments with Bifidobacterium longum and Bifidobacterium infantis. The fermentation products contain rich vitamins, amino acids, and minerals that nourish cells, accelerating skin metabolism and repairing damaged skin; at the same time, it can also improve the dissolution of active components in the raw materials, making the fermentation products contain rich substances such as flavonoids, organic acids, terpene alcohols, polysaccharides, and saponins. The fermentation of the two strains also has a synergistic effect.

[0032] The polysaccharide composition obtained by alcohol precipitation of the fermentation product has excellent moisturizing, whitening, and antioxidant properties. After separation and purification by macroporous resin, the obtained active mixture contains rich flavonoid / saponin complexes.

[0033] Among them, the birch bark extract contains rich components such as volatile oils and triterpenol alcohols, having antibacterial and anti-inflammatory effects; the extract of the root of Cyanotis arachnoidea C. B. Clarke has good exfoliating and whitening effects, and can be used in combination with the antibacterial and anti-inflammatory birch bark extract, having good repair effects on facial chloasma, freckles, melanin deposition, etc., and also having obvious effects on acne. The European plum extract contains traditional moisturizing and skin care ingredients, and its seed extract has repair and soothing effects, which can help the skin repair dull skin color and deeply moisturize the skin. The peony root extract contains antioxidant components, inhibits metalloproteinase and elastase, has the effect of delaying skin aging, contains collagen fiber gel, has a contracting effect, can tighten the skin, remove fine lines, wrinkles, and eye bags. The angelica root extract can promote the production of hemoglobin and red blood cells, and ferulic acid contained therein can improve peripheral circulation, having antioxidant and free radical scavenging effects, indicating a certain anti-aging effect.

[0034] On the one hand, the fermentation process produces rich fermentation by-products, and at the same time promotes the large proliferation of Bifidobacterium. After liquid nitrogen freezing-room temperature dissolution treatment, the cell walls of Bifidobacterium rupture and the contents dissolve out. The prepared bifida ferment lysate can regulate the expression of the sirtuin-6 gene, helping to extend cell lifespan and delay aging, and having the effects of whitening, antioxidant, and anti-aging.

[0035] The hirudin / phloretin / ellagic acid chelate complex prepared by the present invention forms a complex with a composite network multi-molecular structure under the polymerization of tannic acid for hirudin, phloretin, and ellagic acid. The phloretin structure has good antioxidant, anti-inflammatory, and moisturizing effects. The hirudin part accelerates cell metabolism, dredges capillaries, improves facial microcirculation, inhibits and reduces the growth of melanin, and has the effects of anti-inflammatory, detumescence, and elimination of facial pigmentation. The ellagic acid structure can block the activity of tyrosinase, fight free radicals, help the skin play an anti-aging role, prevent the occurrence of skin problems such as fine lines and wrinkles, prevent the accumulation of skin melanin, accelerate the decomposition of melanin, and prevent the generation of melanin from the source, helping the skin to whiten, brighten the skin tone, and fade freckles. Further, by forming a complex with silver ions, the participation of silver ions can inhibit the activity and function of melanocytes, reduce the production of melanin, and achieve a whitening effect. At the same time, it can also inhibit the activity of tyrosinase, thereby reducing the synthesis of melanin.

[0036] The freckle-removing and whitening essence prepared by the present invention has good effects of whitening, freckle removal, skin tightening, antioxidant, anti-inflammatory, moisturizing, improving skin elasticity, and delaying aging. Moreover, the raw materials are widely sourced and the preparation method is simple, having broad application prospects. Specific Embodiments

[0037] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0038] Cellulase, 11,000 U / g, pectinase, 60,000 U / g, purchased from Xiasheng (Beijing) Biotechnology Development Co., Ltd.

[0039] Bifidobacterium longum, 10 billion cfu / g, purchased from Guangdong Hongyou Biotechnology Co., Ltd., Bifidobacterium infantis, 10 billion cfu / g, purchased from Hubei Mingtuo Biotechnology Co., Ltd.

[0040] Preparation method of the strain seed liquid: Inoculate the strain into the Gao's medium, culture at 37 °C and 100 r / min for 24 h to obtain a strain seed liquid with a bacterial content of 10 8 -10 9 cfu / mL.

[0041] The preparation method of the plant extract in Preparation Example 1 includes the following steps:

[0042] S1. Enzymolysis in deep eutectic solvent: Wash 3 parts by weight of Betula platyphylla bark, 1 part by weight of Peristrophe baphica roots, 7 parts by weight of Prunus domestica, 3 parts by weight of Paeonia suffruticosa roots, and 1 part by weight of Angelica sinensis roots respectively, dry them, pulverize them to obtain a mixed powder. Add 10 parts by weight of the mixed powder to 50 parts by weight of an aqueous solution containing a deep eutectic solvent, add a composite enzyme, and the addition amount of the composite enzyme is 2 wt% of the total mass of the system. Enzymolyze at 40 °C for 1 h, inactivate the enzyme to obtain an enzymolysis product;

[0043] The composite enzyme is composed of a mixture of cellulase and pectinase, and the mass ratio is 10:3;

[0044] The water content in the aqueous solution containing the deep eutectic solvent is 40 wt%, and the deep eutectic solvent is prepared by mixing choline chloride and glucose according to a molar ratio of 1:1;

[0045] S2. Fermentation: Sterilize the enzymolysis product in step S1, inoculate the seed solutions of Bifidobacterium longum and Bifidobacterium infantis, and the inoculation amounts of the seed solutions of Bifidobacterium longum and Bifidobacterium infantis are 1 v / v% and 2 v / v% respectively. Under anaerobic conditions, at 36 °C, 150 r / min, ferment and culture for 24 h, filter, wash the solid, collect the eluate and concentrate to obtain a bacterial solution, and keep the filtrate for use;

[0046] S3. Preparation of bifidobacterium lysate: Freeze the bacterial solution obtained in step S2 in liquid nitrogen, dissolve it at room temperature, centrifuge, collect the supernatant, and freeze-dry it to obtain a bifidobacterium lysate;

[0047] S4. Preparation of polysaccharide composition: Add ethanol to the filtrate obtained in step S2 until the ethanol content in the system is 60 wt%, precipitate for 3 h, centrifuge, collect the solid, recover ethanol from the supernatant, keep the obtained solution for use, wash the solid, dry it to obtain a polysaccharide composition;

[0048] S5. Separation and purification: Add 10 parts by weight of an organic solvent to 20 parts by weight of the solution in step S3, extract, collect the organic phase, remove the solvent from the organic phase under reduced pressure, dilute it to 1 mg / mL with 70 wt% ethanol for loading, wet-pack the column to obtain an AB-8 macroporous resin chromatography column. The chromatography column specifications are 15 mm * 500 mm, the column volume is 50 mL, collect the eluate with a 70 wt% ethanol concentration, remove ethanol under reduced pressure, dry it to obtain a flavonoid / saponin complex;

[0049] The organic solvent is a mixed solvent of dichloromethane and tetrahydrofuran, and the volume ratio is 3:1;

[0050] S6. Preparation of preservative: Stir and mix 10 parts by weight of vitamin C and 3 parts by weight of vitamin E for 10 min to obtain a preservative;

[0051] S7. Preparation of plant extract: Stir and mix 10 parts by weight of the lysate of Saccharomyces boulardii prepared in step S3, 15 parts by weight of the polysaccharide composition prepared in step S4, 17 parts by weight of the flavonoid / saponin complex prepared in step S5, and 2 parts by weight of the preservative prepared in step S6 for 20 min to obtain the plant extract.

[0052] The preparation method of the plant extract in Preparation Example 2 includes the following steps:

[0053] S1. Enzymolysis in deep eutectic solvent: Wash, dry, and pulverize 5 parts by weight of white birch bark, 3 parts by weight of the root of Peristrophe baphica, 12 parts by weight of European plum, 5 parts by weight of peony root, and 2 parts by weight of angelica root respectively to obtain a mixed powder. Add 10 parts by weight of the mixed powder to 70 parts by weight of an aqueous solution containing a deep eutectic solvent, add a composite enzyme, and the addition amount of the composite enzyme is 3 wt% of the total mass of the system. Carry out enzymolysis at 50 °C for 3 h, inactivate the enzyme, and obtain an enzymolysis product;

[0054] The composite enzyme is composed of a mixture of cellulase and pectinase, and the mass ratio is 15:5;

[0055] The water content in the aqueous solution containing the deep eutectic solvent is 60 wt%, and the deep eutectic solvent is prepared by mixing choline chloride and glucose in a molar ratio of 1:1;

[0056] S2. Fermentation: Sterilize the enzymolysis product in step S1, inoculate the seed solutions of Bifidobacterium longum and Bifidobacterium infantis, and the inoculation amounts of the seed solutions of Bifidobacterium longum and Bifidobacterium infantis are 2 v / v% and 4 v / v% respectively. Under anaerobic conditions, at 39 °C and 200 r / min, ferment and culture for 48 h, filter, wash the solid, collect the eluate and concentrate to obtain a bacterial solution, and keep the filtrate for use;

[0057] S3. Preparation of lysate of Saccharomyces boulardii: Freeze the bacterial solution prepared in step S2 with liquid nitrogen, dissolve it at room temperature, centrifuge, collect the supernatant, and freeze-dry to obtain the lysate of Saccharomyces boulardii;

[0058] S4. Preparation of polysaccharide composition: Add ethanol to the filtrate prepared in step S2 until the ethanol content in the system is 80 wt%, precipitate for 5 h, centrifuge, collect the solid, recover ethanol from the supernatant, keep the obtained solution for use, wash the solid, and dry to obtain the polysaccharide composition;

[0059] S5. Separation and purification: Add 30 parts by weight of an organic solvent to 40 parts by weight of the solution in step S3, extract, collect the organic phase, remove the solvent from the organic phase under reduced pressure, dilute it with 80 wt% ethanol to 1.5 mg / mL for loading, wet-pack the column to obtain an AB-8 macroporous resin chromatography column. The specifications of the chromatography column are 15 mm * 500 mm, the column volume is 50 mL, collect the eluate with an ethanol concentration of 80 wt%, remove ethanol under reduced pressure, and dry to obtain the flavonoid / saponin complex;

[0060] The organic solvent is a mixed solvent of dichloromethane and tetrahydrofuran with a volume ratio of 5:1;

[0061] S6. Preparation of the preservative: Stir and mix 15 parts by weight of vitamin C and 7 parts by weight of vitamin E for 10 min to obtain the preservative;

[0062] S7. Preparation of the plant extract: Stir and mix 12 parts by weight of the lysate of Saccharomyces boulardii prepared in step S3, 22 parts by weight of the polysaccharide composition prepared in step S4, 20 parts by weight of the flavonoid / saponin complex prepared in step S5, and 4 parts by weight of the preservative prepared in step S6 for 20 min to obtain the plant extract.

[0063] The preparation method of the plant extract in Preparation Example 3 includes the following steps:

[0064] S1. Enzymolysis in the deep eutectic solvent: Wash, dry, and pulverize 4 parts by weight of Betula platyphylla Suk. bark, 2 parts by weight of Peristrophe baphica (Spreng.) Bremek. root, 10 parts by weight of Prunus domestica L., 4 parts by weight of Paeonia suffruticosa Andr. root, and 1.5 parts by weight of Angelica sinensis (Oliv.) Diels root respectively to obtain a mixed powder. Add 10 parts by weight of the mixed powder to 60 parts by weight of an aqueous solution containing the deep eutectic solvent, add a composite enzyme, the addition amount of the composite enzyme is 2.5 wt% of the total mass of the system, carry out enzymolysis at 46 °C for 2 h, inactivate the enzyme, and obtain an enzymolysis product;

[0065] The composite enzyme is composed of a mixture of cellulase and pectinase with a mass ratio of 12:4;

[0066] The water content in the aqueous solution containing the deep eutectic solvent is 50 wt%, and the deep eutectic solvent is prepared by mixing choline chloride and glucose according to a molar ratio of 1:1;

[0067] S2. Fermentation: Sterilize the enzymolysis product in step S1, inoculate the seed solutions of Bifidobacterium longum and Bifidobacterium infantis, the inoculation amounts of the seed solutions of Bifidobacterium longum and Bifidobacterium infantis are 1.2 v / v% and 3 v / v% respectively, under anaerobic conditions, at 37 °C, 170 r / min, ferment and culture for 36 h, filter, wash the solid, collect the eluate and concentrate to obtain a bacterial solution, and keep the filtrate for use;

[0068] S3. Preparation of the lysate of Saccharomyces boulardii: Freeze the bacterial solution prepared in step S2 with liquid nitrogen, dissolve it at room temperature, centrifuge, collect the supernatant, and freeze-dry to obtain the lysate of Saccharomyces boulardii;

[0069] S4. Preparation of the polysaccharide composition: Add ethanol to the filtrate prepared in step S2 until the ethanol content in the system is 70 wt%, precipitate for 4 h, centrifuge, collect the solid, recover ethanol from the supernatant, keep the obtained solution for use, wash the solid, and dry to obtain the polysaccharide composition;

[0070] S5. Separation and purification: Add 20 parts by weight of an organic solvent to 30 parts by weight of the solution in step S3, extract, collect the organic phase, remove the solvent from the organic phase under reduced pressure, dilute it with 75 wt% ethanol to 1.5 mg / mL for loading, and obtain an AB-8 macroporous resin chromatography column by wet packing. The specifications of the chromatography column are 15 mm * 500 mm, the column volume is 50 mL, collect the eluent with a 75 wt% ethanol concentration, remove ethanol under reduced pressure, and dry to obtain a flavonoid / saponin complex;

[0071] The organic solvent is a mixed solvent of dichloromethane and tetrahydrofuran with a volume ratio of 4:1;

[0072] S6. Preparation of preservative: Stir and mix 12 parts by weight of vitamin C and 5 parts by weight of vitamin E for 10 min to obtain a preservative;

[0073] S7. Preparation of plant extract: Stir and mix 11 parts by weight of the lysate of Saccharomyces boulardii prepared in step S3, 20 parts by weight of the polysaccharide composition prepared in step S4, 18 parts by weight of the flavonoid / saponin complex prepared in step S5, and 3 parts by weight of the preservative prepared in step S6 for 20 min to obtain a plant extract.

[0074] Preparation Example 4

[0075] Compared with Preparation Example 3, the difference is that the complex enzyme is a single cellulase.

[0076] Preparation Example 5

[0077] Compared with Preparation Example 3, the difference is that the complex enzyme is a single pectinase.

[0078] Comparative Preparation Example 1

[0079] Compared with Preparation Example 3, the difference is that no complex enzyme was added in step S1.

[0080] Comparative Preparation Example 2

[0081] Compared with Preparation Example 3, the difference is that no seed solution of Bifidobacterium longum was inoculated in step S2.

[0082] Comparative Preparation Example 3

[0083] Compared with Preparation Example 3, the difference is that no seed solution of Bifidobacterium infantis was inoculated in step S2.

[0084] Comparative Preparation Example 4

[0085] Compared with Preparation Example 3, the difference is that step S2 was not carried out.

[0086] Comparative Preparation Example 5

[0087] Compared with Preparation Example 3, the difference lies in that no bifidobacterium lysate was added in step S7.

[0088] Comparative Preparation Example 6

[0089] Compared with Preparation Example 3, the difference lies in that no polysaccharide composition was added in step S7.

[0090] Comparative Preparation Example 7

[0091] Compared with Preparation Example 3, the difference lies in that no flavonoid / saponin complex was added in step S7.

[0092] Preparation of hirudin / phloretin / ellagic acid chelate complex in Preparation Example 6

[0093] The method is as follows:

[0094] T1. Add 12 parts by weight of hirudin, 3 parts by weight of phloretin, and 2 parts by weight of ellagic acid to 200 parts by weight of water, add 2 parts by weight of tris(hydroxymethyl)aminomethane and 5 parts by weight of tannic acid, adjust the pH value of the solution to 8.2, heat to 40 °C, and stir and react for 1 h to obtain a complex;

[0095] T2. Add 10 parts by weight of the complex obtained in step T1 to 100 parts by weight of water, add 0.2 parts by weight of silver nitrate, stir and react for 20 min, dialyze, and dry to obtain the hirudin / phloretin / ellagic acid chelate complex.

[0096] Preparation of hirudin / phloretin / ellagic acid chelate complex in Preparation Example 7

[0097] The method is as follows:

[0098] T1. Add 15 parts by weight of hirudin, 5 parts by weight of phloretin, and 4 parts by weight of ellagic acid to 200 parts by weight of water, add 4 parts by weight of tris(hydroxymethyl)aminomethane and 7 parts by weight of tannic acid, adjust the pH value of the solution to 8.7, heat to 50 °C, and stir and react for 3 h to obtain a complex;

[0099] T2. Add 15 parts by weight of the complex obtained in step T1 to 100 parts by weight of water, add 0.5 parts by weight of silver nitrate, stir and react for 30 min, dialyze, and dry to obtain the hirudin / phloretin / ellagic acid chelate complex.

[0100] Preparation of hirudin / phloretin / ellagic acid chelate complex in Preparation Example 8

[0101] The method is as follows:

[0102] T1. Add 13 parts by weight of hirudin, 4 parts by weight of phloretin, and 3 parts by weight of ellagic acid to 200 parts by weight of water. Add 3 parts by weight of tris(hydroxymethyl)aminomethane and 6 parts by weight of tannic acid, adjust the pH value of the solution to 8.5, heat to 45 °C, and stir and react for 2 h to obtain a complex;

[0103] T2. Add 12 parts by weight of the complex prepared in step T1 to 100 parts by weight of water, add 0.3 parts by weight of silver nitrate, stir and react for 25 min, dialyze, and dry to obtain a hirudin / phloretin / ellagic acid chelate complex.

[0104] Comparative Preparation Example 8

[0105] Compared with Preparation Example 8, the difference lies in that phloretin was not added in step T1.

[0106] Comparative Preparation Example 9

[0107] Compared with Preparation Example 8, the difference lies in that ellagic acid was not added in step T1.

[0108] Comparative Preparation Example 10

[0109] Compared with Preparation Example 8, the difference lies in that hirudin was not added in step T1.

[0110] Comparative Preparation Example 11

[0111] Compared with Preparation Example 8, the difference lies in that tannic acid was not added in step T1.

[0112] Comparative Preparation Example 12

[0113] Compared with Preparation Example 8, the difference lies in that step T2 was not carried out.

[0114] Example 1

[0115] This example provides a freckle-removing and whitening essence, and the preparation method includes the following steps:

[0116] (1) Add 5 parts by weight of glycerol and 0.3 parts by weight of xanthan gum to 40 parts by weight of deionized water, heat to 70 °C, and stir at 300 r / min for 10 min to obtain phase A;

[0117] (2) Mix 10 parts by weight of tri(ethylhexanoate) glycerol, 0.5 parts by weight of sorbitan oleate, and 3 parts by weight of the hirudin / phloretin / ellagic acid chelate complex prepared in Preparation Example 6, heat to 70 °C, and stir at 300 r / min for 10 min to obtain phase B;

[0118] (3) Add phase B to phase A, stir at 300 r / min for 5 min, emulsify at 7000 r / min for 10 - 20 min, keep warm for 10 - 20 min, and then cool down;

[0119] (4) After cooling to 35°C, add 10 parts by weight of the plant extract prepared in Preparation Example 1, stir at 300 r / min for 5 min, and filter to obtain the freckle-reducing and whitening essence.

[0120] Example 2

[0121] This example provides a freckle-reducing and whitening essence, and the preparation method includes the following steps:

[0122] (1) Add 10 parts by weight of hexylene glycol and 1 part by weight of polyacrylate cross-linked polymer-6 to 60 parts by weight of deionized water, heat to 80°C, and stir at 500 r / min for 20 min to obtain Phase A;

[0123] (2) Mix 20 parts by weight of polydimethylsiloxane, 3 parts by weight of cetearyl olivate, and 5 parts by weight of the hirudin / phloretin / ellagic acid chelate complex prepared in Preparation Example 7, heat to 80°C, and stir at 500 r / min for 20 min to obtain Phase B;

[0124] (3) Add Phase B to Phase A, stir at 500 r / min for 15 min, emulsify at 10000 r / min for 20 min, keep warm for 20 min, and then cool down;

[0125] (4) After cooling to 40°C, add 20 parts by weight of the plant extract prepared in Preparation Example 2, stir at 500 r / min for 15 min, and filter to obtain the freckle-reducing and whitening essence.

[0126] Example 3

[0127] This example provides a freckle-reducing and whitening essence, and the preparation method includes the following steps:

[0128] (1) Add 7 parts by weight of sodium hyaluronate and 0.7 part by weight of carbomer to 50 parts by weight of deionized water, heat to 75°C, and stir at 400 r / min for 15 min to obtain Phase A;

[0129] (2) Mix 15 parts by weight of glyceryl tricaprylate, 1.5 parts by weight of glyceryl stearate, and 4 parts by weight of the hirudin / phloretin / ellagic acid chelate complex prepared in Preparation Example 8, heat to 75°C, and stir at 400 r / min for 15 min to obtain Phase B;

[0130] (3) Add Phase B to Phase A, stir at 400 r / min for 10 min, emulsify at 8000 r / min for 15 min, keep warm for 15 min, and then cool down;

[0131] (4) After cooling to 37°C, add 15 parts by weight of the plant extract prepared in Preparation Example 3, stir at 400 r / min for 10 min, and filter to obtain the freckle-reducing and whitening essence.

[0132] Example 4

[0133] Compared with Example 3, the difference lies in that the plant extract is prepared from Preparation Example 4.

[0134] Example 5

[0135] Compared with Example 3, the difference lies in that the plant extract is prepared from Preparation Example 5.

[0136] Comparative Example 1

[0137] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 1.

[0138] Comparative Example 2

[0139] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 2.

[0140] Comparative Example 3

[0141] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 3.

[0142] Comparative Example 4

[0143] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 4.

[0144] Comparative Example 5

[0145] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 5.

[0146] Comparative Example 6

[0147] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 6.

[0148] Comparative Example 7

[0149] Compared with Example 3, the difference lies in that the plant extract is prepared from Comparative Preparation Example 7.

[0150] Comparative Example 8

[0151] Compared with Example 3, the difference lies in that the hirudin / phloretin / ellagic acid chelate complex is prepared from Comparative Preparation Example 8.

[0152] Comparative Example 9

[0153] Compared with Example 3, the difference lies in that the hirudin / phloretin / ellagic acid chelate complex is prepared from Comparative Preparation Example 9.

[0154] Comparative Example 10

[0155] Compared with Example 3, the difference lies in that the hirudin / phloretin / ellagic acid chelate complex is prepared from Comparative Preparation Example 10.

[0156] Comparative Example 11

[0157] Compared with Example 3, the difference lies in that the hirudin / phloretin / ellagic acid chelate complex is prepared from Comparative Preparation Example 11.

[0158] Comparative Example 12

[0159] Compared with Example 3, the difference lies in that the hirudin / phloretin / ellagic acid chelate complex is prepared from Comparative Preparation Example 12.

[0160] Comparative Example 13

[0161] Compared with Example 3, the difference lies in that no plant extract is added.

[0162] Comparative Example 14

[0163] Compared with Example 3, the difference lies in that no hirudin / phloretin / ellagic acid chelate complex is added.

[0164] Test Example 1 Cytotoxicity Study

[0165] The cytotoxicity of Zhenyao White was determined by the CCK-8 method. HaCaT cells in the logarithmic growth phase were collected and seeded in 96-well plates at a density of 2×10 5 cells / mL, 100 μL per well, and cultured for 24 h under the conditions of 5% CO2 and 37 °C. 100 μL of DMEM complete medium containing the lightening and whitening essence prepared in Examples 1-5 or Comparative Examples 1-14 (the concentration of the lightening and whitening essence was 200 μg / mL) was added to each well, and the control group was only added with 100 μL of DMEM complete medium, with 3 replicates in each group. After continuous culture for 24 h, the cell survival rate was measured by the CCK-8 method. The results are shown in Table 1.

[0166] Table 1

[0167]

[0168]

[0169] As can be seen from the above table, the lightening and whitening essence prepared by the present invention is non-toxic to HaCaT cells.

[0170] Test Example 2 Study on Cell Tyrosinase Activity and Melanin Content

[0171] Take B16F10 cells at 7×10 4Inoculate at 1×10⁴ cells / mL into 24-well and 6-well plates respectively. Add 500 μL to the 24-well plate and 2 mL to the 6-well plate. Incubate for 24 h, and then add 100 nmol / L α-melanocyte-stimulating hormone (α-MSH) for induction according to the experimental groups (normal control group, model group, Example 1-5 groups, Comparative Example 1-14 groups) (except for the normal control group) to construct an α-MSH-induced high melanin expression model. Then add the lightening and whitening essence prepared in Example 1-5 or Comparative Example 1-14 (the concentration of the lightening and whitening essence is 200 μg / mL). Set 3 replicate wells for each group and incubate for 48 h. The results are shown in Table 2.

[0172] Discard the supernatant of the 24-well cells, wash 3 times with PBS, lyse the cells, take 60 μL of the supernatant into a 96-well plate, add 140 μL of 0.1% L-dopamine, incubate at 37 °C for 1 h, and measure the absorbance (A) of each well at a wavelength of 490 nm to determine the tyrosinase activity.

[0173] After washing the 6-well plate cells 3 times with PBS, add 300 μL of 1.0 mmol / L NaOH solution (containing 10% DMSO) to each well, fully lyse the cells at 80 °C for 1 h, and measure the absorbance (A) of each well at a wavelength of 405 nm to determine the cellular melanin content.

[0174] Table 2

[0175]

[0176]

[0177] As can be seen from the above table, the lightening and whitening essence prepared in Examples 1-3 of the present invention has good inhibitory effects on tyrosinase activity and cellular melanin production.

[0178] Test Example 3 Scavenging rate of ·OH

[0179] Dilute the lightening and whitening essence prepared in Example 1-5 or Comparative Example 1-14 with deionized water to 1 mg / mL as the sample solution.

[0180] Take 2 mL of 1.00×10 -2 mol / L salicylic acid-ethanol solution, 2 mL of 9.00×10 -3 mol / L FeSO₄ solution and the sample solution and add them into a test tube, then add 2 mL of 8.8×10 -2 mol / L H₂O₂ solution, shake well and place in a 37 °C water bath for reaction for 30 min, and measure the absorbance A at 510 nm i ; Measure the absorbance A with 2 mL of pure water instead of the salicylic acid solution j; Use 2 mL of pure water instead of the sample solution to measure the absorbance A0. Calculate the scavenging rate (%) of ·OH according to the formula. Use vitamin C as the control group. The results are shown in Table 3.

[0181] Scavenging rate of ·OH (%) = [1 - (A i - A j ) / A0] × 100%

[0182] Table 3

[0183]

[0184]

[0185] As can be seen from the above table, the freckle-reducing and whitening essence prepared in Examples 1-3 of the present invention has good antioxidant activity.

[0186] Test Example 4

[0187] Select volunteers aged 32-45 years who meet the volunteer inclusion criteria, and randomly divide them into groups of Examples 1-5, Comparative Examples 1-14, and blank group, with 10 people in each group.

[0188] Experimental samples:

[0189] The experimental group uses the freckle-reducing and whitening essence prepared in Examples 1-5 or Comparative Examples 1-14. The blank group uses an equal amount of water.

[0190] Test conditions: On the day of the test, the subjects clean their faces with the designated cleansing sample and sit quietly in a laboratory at a temperature of (25 ± 1) °C and a humidity of 45% ± 5% RH for 30 minutes.

[0191] Experimental method: Before the subjects apply the sample, they first wash the test area, dry it, and then apply the sample. An area of 5 × 5 cm is used as the test area; the subjects use the sample twice a day, morning and evening, in the test area. During the experiment, the subjects cannot apply any other cosmetics to the test area. The subjects are tested once at the 0th and 4th weeks after continuous use of the sample. At the same time, the applied area is washed, and the melanin content of the applied area is measured by the Mexameter MX18 tester and the MicroSkin II multi-functional dermoscope image analysis system. Each point is measured five times, and the average value is taken. The results are shown in Table 4.

[0192] Melanin reduction rate (%) = (last melanin content - first melanin content) / first melanin content × 100%

[0193] Relative melanin reduction rate (%) = melanin reduction rate of the sample group (%) - melanin reduction rate of the blank group (%) Table 4

[0194] Group Melanin reduction rate (%) Relative black reduction rate (%) Blank group -1.4 / Example 1 41.2 42.6 Example 2 40.9 42.3 Example 3 42.3 43.7 Example 4 38.5 39.9 Example 5 37.9 39.3 Comparative example 1 35.2 36.6 Comparative example 2 34.9 36.3 Comparative example 3 35.5 36.9 Comparative example 4 32.1 33.5 Comparative example 5 30.8 32.2 Comparative example 6 36.2 37.6 Comparative example 7 34.8 36.2 Comparative example 8 37.5 38.9 Comparative example 9 36.9 38.3 Comparative example 10 32.0 33.4 Comparative example 11 33.9 35.3 Comparative example 12 34.5 35.9 Comparative example 13 26.1 27.5 Comparative example 14 28.9 30.3

[0195] As can be seen from the above table, the lightening and whitening essence prepared in Examples 1-3 of the present invention has a good whitening effect.

[0196] The above are only the preferred embodiments of the present invention, and are not intended to limit the present invention. Any modifications, equivalent replacements, improvements, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A spot-lightening and whitening essence, characterized in that: The invention is prepared from the following raw materials by weight: 10-20 parts of plant extract, 3-5 parts of hirudin / phloretin / ellagic acid chelate complex, 10-20 parts of emollient, 5-10 parts of humectant, 0.3-1 parts of thickener, 0.5-3 parts of emulsifier and 40-60 parts of deionized water; The preparation method of the plant extract comprises the following steps: S1. Enzymatic hydrolysis in a deep eutectic solvent: wash, dry, and crush birch bark, spider silk blue ear grass root, European plum, peony root, and angelica root to obtain a mixed powder, add it to an aqueous solution containing a deep eutectic solvent, add a composite enzyme, perform enzymolysis, inactivate the enzyme, and obtain an enzymatic hydrolysis product; the composite enzyme is composed of a mixture of cellulase and pectinase, with a mass ratio of 10-15:3-5; S2 fermentation: the enzymatic product of step S1 is sterilized, inoculated with Bifidobacterium longum, Bifidobacterium infantis seed solution, fermented, filtered, washed with solids, the eluate is collected and concentrated to obtain a bacterial solution, and the filtrate is retained; S3. Preparation of bifid yeast lysate: freezing the bacterial solution obtained in step S2 with liquid nitrogen, thawing it at room temperature, centrifuging it, collecting the supernatant, and freeze-drying it to obtain bifid yeast lysate; S4. Preparation of a polysaccharide composition: adding ethanol to the filtrate obtained in step S2 for precipitation, centrifuging, collecting the solid, recovering ethanol from the supernatant, retaining the obtained solution, washing the solid, and drying to obtain a polysaccharide composition; S5. Separation and purification: adding an organic solvent to the solution in step S3, extracting, collecting the organic phase, separating and purifying with a macroporous resin to obtain a flavonoid / saponin complex; S6. Preparation of preservative: Mixing vitamin C and vitamin E to obtain a preservative; S7. Preparation of plant extract: The bifid yeast lysate obtained in step S3, the polysaccharide composition obtained in step S4, the flavonoid / saponin complex obtained in step S5, and the preservative obtained in step S6 are uniformly mixed to obtain a plant extract; The preparation method of the hirudin / phloretin / ellagic acid chelate complex is as follows: T1. Add hirudin, phloretin, and ellagic acid to water, add tris(hydroxymethyl)aminomethane and tannic acid, adjust the pH value of the solution, heat and stir the reaction to obtain a complex; T2. Add the complex prepared in step T1 into water, add silver nitrate, stir to react, dialyze, and dry to obtain a hirudin / phloretin / ellagic acid chelate complex.

2. The spot-lightening and whitening essence according to claim 1, characterized in that: The mass ratio of birch bark, spider silk blue ear grass root, European plum, peony root, and angelica root in step S1 is 3-5:1-3:7-12:3-5:1-2, the mass ratio of the mixed powder and the aqueous solution containing a deep eutectic solvent is 1:5-7, the amount of the complex enzyme added is 2-3wt% of the total mass of the system, the water content in the aqueous solution containing a deep eutectic solvent is 40-60wt%, the deep eutectic solvent is prepared by mixing choline chloride and glucose in a molar ratio of 1:1, the temperature of the enzymolysis is 40-50°C, and the time is 1-3h; the inoculation amounts of the long bifidobacterium and infant bifidobacterium seed solutions in step S2 are 1-2v / v% and 2-4v / v%, respectively, and the bacterial content of the seed solution is 10 8 -10 9 cfu / mL, and the fermentation culture conditions are anaerobic conditions, 36-39°C, 150-200r / min, and the fermentation culture is 24-48h.

3. The spot-lightening and whitening essence according to claim 1, characterized in that: In step S4, ethanol is added to the system to obtain an ethanol content of 60-80wt%, and the precipitation time is 3-5h; in step S5, the mass ratio of the organic solvent to the solution is 1-3:2-4, the organic solvent is a mixed solvent of dichloromethane and tetrahydrofuran, and the volume ratio is 3-5:1, and the macroporous resin is selected from at least one of XDA-2, NKA-9, SP-825, D101, HPD600, X-5, LSA-10, NKI-9 and AB-8.

4. The spot-lightening and whitening essence according to claim 1, characterized in that: The mass ratio of vitamin C to vitamin E in step S6 is 10-15:3-7; the mass ratio of bifida yeast lysate, polysaccharide composition, flavonoid / saponin complex and preservative in step S7 is 10-12:15-22:17-20:2-4.

5. The spot-lightening and whitening essence according to claim 1, characterized in that: In step T1, the mass ratio of hirudin, phloretin, ellagic acid, tris(hydroxymethyl)aminomethane) and tannic acid is 12-15:3-5:2-4:2-4:5-7, the pH value of the adjusted solution is 8.2-8.7, the temperature of the heated stirring reaction is 40-50°C, and the time is 1-3h; in step T2, the mass ratio of the complex, water and silver nitrate is 10-15:100:0.2-0.5, and the stirring reaction time is 20-30min.

6. The spot-lightening and whitening essence according to claim 1, characterized in that: The emollient is selected from at least one of glyceryl ethylhexanoate, isononyl isononanoate, polydimethylsiloxane, caprylic / capric triglyceride, butyrospermum parkii fruit fat, and liminanthes alba seed oil; the moisturizer is selected from at least one of glycerol, butylene glycol, propylene glycol, hexylene glycol, sodium hyaluronate, betaine, trehalose, and 1,2-pentanediol; the emulsifier is selected from at least one of glyceryl stearate, PEG-100 stearate, cetearyl olivate, sorbitan olivate, C12-20 alkyl glucoside, and polyglyceryl-2 stearate; the thickener is selected from at least one of carbomer, sodium polyacryloyl dimethyl taurate, polyacrylate crosspolymer-6, and xanthan gum.

7. A method for preparing the spot-lightening and whitening essence according to any one of claims 1 to 6, characterized in that: The following steps are involved: (1) Add a moisturizer and a thickener into deionized water, heat to 70-80° C., and stir at 300-500 r / min for 10-20 min to prepare phase A; (2) mixing the emollient, emulsifier, and hirudin / phloretin / ellagic acid chelate complex, heating to 70-80° C., and stirring at 300-500 r / min for 10-20 min to prepare phase B; (3) Add phase B to phase A and stir at 300-500r / min for 5-15min. 7000-10000r / min emulsification for 10-20min, heat preservation for 10-20min, cooling; (4) After cooling to 35-40° C., add plant extract, stir at 300-500 r / min for 5-15 min, filter, and obtain a spot-lightening and whitening essence.

8. Use of the spot-lightening and whitening essence according to any one of claims 1 to 6 in the preparation of skin whitening, spot-lightening and moisturizing products.

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