CN118286096A - Freckle-removing and whitening essence and preparation method thereof - Google Patents

Abstract

The invention provides a spot-lightening and whitening essence and a preparation method thereof, and belongs to the technical field of cosmetics. The composite material 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 spot-lightening and whitening essence prepared by the invention has good effects of whitening, spot-lightening, tightening skin, resisting oxidation, resisting inflammation, moisturizing, improving skin elasticity and delaying aging, and has wide raw material sources, simple preparation method and wide application prospect.

Description

A kind of spot-lightening and whitening essence and preparation method thereof

Technical Field

The invention relates to the technical field of cosmetics, and in particular to a spot-lightening and whitening essence and a preparation method thereof.

Background technique

With the improvement of people's living standards and aesthetic standards, fair and clean skin is increasingly admired by most women. Whitening and lightening of spots have always been a topic of keen concern. People hope to get fair, smooth and translucent skin through the use of whitening care products, and some use functional whitening cosmetics to reduce the deposition of spots. Environmental factors such as ultraviolet rays, polluted air, and active oxygen will accelerate the production of melanin in the skin, accelerate skin aging, loose skin, pigmentation, dark yellow skin color, lack of luster, and various fine lines and wrinkles.

There are many whitening active ingredients on the market, but they all have certain disadvantages. First, some whitening active ingredients can cause skin irritation, allergies and other phenomena; second, most whitening active ingredients are unstable and easily oxidized, deteriorated, discolored and lose their whitening effects; third, some whitening active ingredients have poor epidermal permeability and are difficult to penetrate the stratum corneum into the basal layer of the skin to exert their effects. Fourth, the whitening and spot-removing effects are achieved by inhibiting the activity of tyrosinase to reduce the production of melanin, or by decomposing and reducing the melanin that has been produced. The active ingredients used are mainly synthetic chemical preparations such as arbutin, sodium ascorbyl phosphate (vitamin C), and kojic acid palmitate, which have limited effects and are prone to rebound.

Summary of the invention

The purpose of the present invention is to provide a whitening and freckle removal essence and a preparation method thereof, which has good whitening, freckle removal, skin tightening, anti-oxidation, anti-inflammatory, moisturizing, improving skin elasticity, and anti-aging effects, and has a wide source of raw materials, a simple preparation method, and broad application prospects.

The technical solution of the present invention is achieved in this way:

The invention provides a spot-lightening 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 parts of thickener, 0.5-3 parts of emulsifier and 40-60 parts of deionized water.

As a further improvement of the present invention, the method for preparing the plant extract comprises the following steps:

S1. Enzymatic hydrolysis in a deep eutectic solvent: washing, drying, and crushing birch bark, root of blue ear grass, European plum, peony root, and angelica root respectively to obtain a mixed powder, adding the mixed powder to an aqueous solution containing a deep eutectic solvent, adding a composite enzyme, performing enzymatic hydrolysis, and inactivating the enzyme to obtain an enzymatic hydrolysis product;

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: The bacterial solution obtained in step S2 was frozen in liquid nitrogen, dissolved at room temperature, centrifuged, the supernatant was collected, and freeze-dried 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: uniformly mix 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 to obtain a plant extract.

As a further improvement of the present invention, the mass ratio of birch bark, pearl 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 composite enzyme is composed of a mixture of cellulase and pectinase, the mass ratio is 10-15:3-5, the addition amount of the composite enzyme 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.

As a further improvement of the present invention, the ethanol content in the system added in step S4 is 60-80wt%, and the precipitation time is 3-5h; the mass ratio of the organic solvent and the solution in step S5 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.

As a further improvement of the present invention, the mass ratio of vitamin C and 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.

As a further improvement of the present invention, 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.

As a further improvement of the present invention, the mass ratio of hirudin, phloretin, ellagic acid, tris(hydroxymethyl)aminomethane) and tannic acid in step T1 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 and stirred reaction is 40-50°C, and the time is 1-3h; the mass ratio of the complex, water and silver nitrate in step T2 is 10-15:100:0.2-0.5, and the stirring reaction time is 20-30min.

As a further improvement of the present invention, the emollient is selected from at least one of tri(ethylhexanoin)glyceryl, 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 olive oil ester, sorbitan olive oil ester, C12-20 alkyl glucoside, and polyglyceryl-2 stearate; the thickener is selected from at least one of carbomer, acrylic acid (ester)/C10-30 alkyl acrylate crosspolymer, sodium polyacryloyldimethyl taurate, polyacrylate crosspolymer-6, and xanthan gum.

The present invention further provides a method for preparing the above-mentioned spot-lightening and whitening essence, comprising the following steps:

(1) Add a moisturizer and a thickener to 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, 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 cool down;

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

The present invention further provides a use of the above-mentioned spot-lightening and whitening essence in skin whitening, spot-lightening and moisturizing.

The present invention has the following beneficial effects:

The plant extract of the present invention uses birch bark, pearl silk blue ear grass root, European plum, peony root, and angelica root as raw materials, uses enzymes as catalysts to hydrolyze plant cell walls, promotes the diffusion of effective ingredients from the cell to the extraction medium, and solves the shortcomings of traditional extraction processes such as complex procedures, low efficiency, and poor extraction quality. However, the extraction efficiency of biological enzymes for water-insoluble components is poor. The present invention uses deep eutectic solvents as reaction media, greatly improving the extraction of water-insoluble compounds, and has many advantages such as low toxicity, green, dynamically adjustable, non-flammable, and strong solubility for compounds. Glucose is used as one of the raw materials in the deep eutectic solvent of the present invention, which also provides a carbon source for subsequent fermentation.

The present invention uses Bifidobacterium longum and Bifidobacterium infantis for fermentation, and the fermented product contains rich vitamins, amino acids and minerals that nourish cells, etc., which accelerates skin metabolism and repairs damaged skin; at the same time, it can also improve the dissolution of active components in the raw materials, so that the fermented product contains rich flavonoids, organic acids, terpenoids, polysaccharides, saponins and other substances. The fermentation of the two strains also has a synergistic effect.

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 flavonoids/saponin complexes.

Among them, birch bark extract contains rich volatile oils and triterpenoid alcohols, which have antibacterial and anti-inflammatory effects; the root extract of the blue ear grass has good exfoliating, spot removal and whitening effects, and can be used in combination with the antibacterial and anti-inflammatory birch bark extract. It has a good repair effect on facial chloasma, freckles, melanin deposition, etc., and also has a significant effect on acne. European plum extract contains traditional moisturizing skin care ingredients, and its seed extract has a repairing and soothing effect, which can help the skin repair the dull and dull skin color and deeply moisturize the skin. Peony root extract contains antioxidant ingredients, which have an inhibitory effect on metalloproteinases and elastases, and has the effect of delaying skin aging. It contains collagen fiber gel, which has a contraction effect, can tighten the skin, and remove fine lines, wrinkles and eye bags. Angelica root extract can promote hemoglobin and erythrocyte production. The ferulic acid it contains can improve peripheral circulation, has antioxidant and free radical scavenging effects, indicating that it has a certain anti-aging effect.

On the one hand, the fermentation process produces abundant fermentation by-products, and on the other hand, it promotes the massive proliferation of bifidobacteria. After liquid nitrogen freezing-room temperature thawing treatment, the bifidobacterium bacterial wall is broken and the contents are dissolved. The obtained bifid yeast lysate can regulate the expression of the deacetylase-6 gene, help to prolong cell life, delay aging, and has whitening, anti-oxidation and anti-aging effects.

The hirudin/phloretin/ellagic acid chelate complex prepared by the present invention forms a complex of a composite network multi-molecular structure by hirudin, phloretin, and ellagic acid under the polymerization of tannic acid, and the phloretin structure has good anti-oxidation, anti-inflammatory, and moisturizing effects, and 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 disappearance of facial pigmentation; the ellagic acid structure can block the activity of tyrosinase, resist free radicals, help the skin to play an anti-aging effect, prevent the skin problems such as fine lines and wrinkles from occurring, prevent the accumulation of skin melanin, accelerate the decomposition of melanin, and prevent the generation of melanin from the root, help the skin to play a whitening effect, brighten the skin color, and dilute spots. Further by forming a complex with silver ions, the participation of silver ions can inhibit the activity and function of melanocytes, reduce melanin production, and achieve a whitening effect. At the same time, the activity of tyrosinase can also be inhibited, thereby reducing the synthesis of melanin.

The spot-lightening and whitening essence prepared by the invention has good whitening, spot-lightening, skin-tightening, anti-oxidation, anti-inflammatory, moisturizing, skin elasticity-enhancing and anti-aging effects, and has a wide source of raw materials, a simple preparation method and broad application prospects.

Detailed ways

The technical solutions in the embodiments of the present invention are described clearly and completely below. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

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

Bifidobacterium longum, 10 billion cfu/g, was purchased from Guangdong Hongyou Biotechnology Co., Ltd., and Bifidobacterium infantis, 10 billion cfu/g, was purchased from Hubei Mingtuo Biotechnology Co., Ltd.

Preparation method of bacterial seed solution: inoculate the bacterial strain into Gould's medium, activate and culture at 37°C, 100 r/min for 24 hours, and obtain bacterial seed solution with a bacterial content of 10 ⁸ -10 ⁹ cfu/mL.

Preparation Example 1 The preparation method of the plant extract comprises the following steps:

S1. Enzymatic hydrolysis in a deep eutectic solvent: 3 parts by weight of birch bark, 1 part by weight of blue ear grass root, 7 parts by weight of European plum, 3 parts by weight of peony root, and 1 part by weight of angelica root were washed, dried, and crushed to obtain a mixed powder, 10 parts by weight of the mixed powder were added to 50 parts by weight of an aqueous solution containing a deep eutectic solvent, and a composite enzyme was added, wherein the amount of the composite enzyme added was 2 wt % of the total mass of the system, and the enzymatic hydrolysis was performed at 40° C. for 1 h, and the enzyme was inactivated to obtain an enzymatic hydrolysis product;

The complex enzyme is composed of a mixture of cellulase and pectinase, with a mass ratio of 10:3;

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 in a molar ratio of 1:1;

S2. Fermentation: The enzymatic hydrolysate in step S1 was sterilized and inoculated with Bifidobacterium longum and Bifidobacterium infantis seed solution, wherein the inoculation amount of Bifidobacterium longum and Bifidobacterium infantis seed solution was 1v/v% and 2v/v%, respectively, under anaerobic conditions, at 36°C, 150r/min, and the fermentation culture was carried out for 24h, filtered, the solid was washed, the eluate was collected and concentrated to obtain a bacterial solution, and the filtrate was retained;

S3. Preparation of bifid yeast lysate: The bacterial solution obtained in step S2 was frozen in liquid nitrogen, dissolved at room temperature, centrifuged, the supernatant was collected, and freeze-dried to obtain bifid yeast lysate;

S4. Preparation of a polysaccharide composition: adding ethanol to the filtrate obtained in step S2 until the ethanol content in the system is 60 wt %, precipitating for 3 h, 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 10 parts by weight of an organic solvent to 20 parts by weight of the solution in step S3, extracting, collecting the organic phase, removing the solvent from the organic phase under reduced pressure, diluting it with 70 wt % ethanol to 1 mg / mL for loading, wet packing to obtain an AB-8 macroporous resin chromatography column, the chromatography column specification is 15 mm * 500 mm, the column volume is 50 mL, collecting the eluate with a concentration of 70 wt % ethanol, removing the ethanol under reduced pressure, and drying to obtain a flavonoid / saponin complex;

The organic solvent is a mixed solvent of dichloromethane and tetrahydrofuran, with a volume ratio of 3:1;

S6. Preparation of preservative: 10 parts by weight of vitamin C and 3 parts by weight of vitamin E were stirred and mixed for 10 min to obtain a preservative;

S7. Preparation of plant extract: 10 parts by weight of the bifida yeast lysate obtained in step S3, 15 parts by weight of the polysaccharide composition obtained in step S4, 17 parts by weight of the flavonoid/saponin complex obtained in step S5, and 2 parts by weight of the preservative obtained in step S6 were stirred and mixed for 20 minutes to obtain a plant extract.

Preparation Example 2 The preparation method of the plant extract comprises the following steps:

S1. Enzymatic hydrolysis in a deep eutectic solvent: 5 parts by weight of birch bark, 3 parts by weight of blue ear grass root, 12 parts by weight of European plum, 5 parts by weight of peony root, and 2 parts by weight of angelica root were washed, dried, and crushed to obtain a mixed powder, 10 parts by weight of the mixed powder were added to 70 parts by weight of an aqueous solution containing a deep eutectic solvent, and a composite enzyme was added, wherein the amount of the composite enzyme added was 3wt% of the total mass of the system, and the enzymatic hydrolysis was performed at 50°C for 3h, and the enzyme was inactivated to obtain an enzymatic hydrolysis product;

The complex enzyme is composed of a mixture of cellulase and pectinase, with a mass ratio of 15:5;

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;

S2. Fermentation: The enzymatic hydrolysate in step S1 was sterilized and inoculated with Bifidobacterium longum and Bifidobacterium infantis seed solution, wherein the inoculation amount of Bifidobacterium longum and Bifidobacterium infantis seed solution was 2 v / v% and 4 v / v%, respectively, under anaerobic conditions, 39 ° C, 200 r / min, and the fermentation culture was carried out for 48 h, filtered, the solid was washed, and the eluate was collected and concentrated to obtain a bacterial solution, and the filtrate was retained;

S3. Preparation of bifid yeast lysate: The bacterial solution obtained in step S2 was frozen in liquid nitrogen, dissolved at room temperature, centrifuged, the supernatant was collected, and freeze-dried to obtain bifid yeast lysate;

S4. Preparation of a polysaccharide composition: adding ethanol to the filtrate obtained in step S2 until the ethanol content in the system is 80 wt %, precipitating for 5 h, centrifuging, collecting the solid, recovering ethanol from the supernatant, retaining the resulting solution, washing the solid, and drying to obtain a polysaccharide composition;

S5. Separation and purification: adding 30 parts by weight of an organic solvent to 40 parts by weight of the solution in step S3, extracting, collecting the organic phase, removing the solvent from the organic phase under reduced pressure, diluting it with 80 wt % ethanol to 1.5 mg / mL for loading, wet packing to obtain an AB-8 macroporous resin chromatography column, the chromatography column specification is 15 mm * 500 mm, the column volume is 50 mL, collecting the eluate with an ethanol concentration of 80 wt %, removing the ethanol under reduced pressure, and drying to obtain a flavonoid / saponin complex;

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

S6. Preparation of preservative: 15 parts by weight of vitamin C and 7 parts by weight of vitamin E were stirred and mixed for 10 min to obtain a preservative;

S7. Preparation of plant extract: 12 parts by weight of the bifida yeast lysate obtained in step S3, 22 parts by weight of the polysaccharide composition obtained in step S4, 20 parts by weight of the flavonoid/saponin complex obtained in step S5, and 4 parts by weight of the preservative obtained in step S6 were stirred and mixed for 20 minutes to obtain a plant extract.

Preparation Example 3 The preparation method of the plant extract comprises the following steps:

S1. Enzymatic hydrolysis in a deep eutectic solvent: 4 parts by weight of birch bark, 2 parts by weight of blue ear grass root, 10 parts by weight of European plum, 4 parts by weight of peony root, and 1.5 parts by weight of angelica root were washed, dried, and crushed to obtain a mixed powder, 10 parts by weight of the mixed powder were added to 60 parts by weight of an aqueous solution containing a deep eutectic solvent, and a composite enzyme was added, wherein the amount of the composite enzyme added was 2.5 wt% of the total mass of the system, and the enzymatic hydrolysis was performed at 46° C. for 2 h, and the enzyme was inactivated to obtain an enzymatic hydrolysis product;

The complex enzyme is composed of a mixture of cellulase and pectinase, with a mass ratio of 12:4;

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 in a molar ratio of 1:1;

S2. Fermentation: The enzymatic hydrolysate in step S1 was sterilized and inoculated with Bifidobacterium longum and Bifidobacterium infantis seed solution, wherein the inoculation amount of Bifidobacterium longum and Bifidobacterium infantis seed solution was 1.2 v / v% and 3 v / v%, respectively, under anaerobic conditions, 37 ° C, 170 r / min, and the fermentation culture was carried out for 36 h, filtered, the solid was washed, and the eluate was collected and concentrated to obtain a bacterial solution, and the filtrate was retained;

S3. Preparation of bifid yeast lysate: The bacterial solution obtained in step S2 was frozen in liquid nitrogen, dissolved at room temperature, centrifuged, the supernatant was collected, and freeze-dried to obtain bifid yeast lysate;

S4. Preparation of a polysaccharide composition: adding ethanol to the filtrate obtained in step S2 until the ethanol content in the system is 70 wt %, precipitating for 4 h, 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 20 parts by weight of an organic solvent to 30 parts by weight of the solution in step S3, extracting, collecting the organic phase, removing the solvent from the organic phase under reduced pressure, diluting with 75 wt% ethanol to 1.5 mg/mL for loading, wet packing to obtain an AB-8 macroporous resin chromatography column, the chromatography column having a specification of 15 mm*500 mm, a column volume of 50 mL, collecting an eluate having an ethanol concentration of 75 wt%, removing ethanol under reduced pressure, and drying to obtain a flavonoid/saponin complex;

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

S6. Preparation of preservative: 12 parts by weight of vitamin C and 5 parts by weight of vitamin E were stirred and mixed for 10 min to obtain a preservative;

S7. Preparation of plant extract: 11 parts by weight of the bifida yeast lysate obtained in step S3, 20 parts by weight of the polysaccharide composition obtained in step S4, 18 parts by weight of the flavonoid/saponin complex obtained in step S5, and 3 parts by weight of the preservative obtained in step S6 were stirred and mixed for 20 minutes to obtain a plant extract.

Preparation Example 4

Compared with Preparation Example 3, the difference is that the composite enzyme is a single cellulase.

Preparation Example 5

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

Comparative Preparation Example 1

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

Comparative Preparation Example 2

Compared with Preparation Example 3, the difference is that the seed liquid of Bifidobacterium longum is not inoculated in step S2.

Comparative Preparation Example 3

Compared with Preparation Example 3, the difference is that the seed liquid of Bifidobacterium infantis is not inoculated in step S2.

Comparative Preparation Example 4

Compared with Preparation Example 3, the difference is that step S2 is not performed.

Comparative Preparation Example 5

Compared with Preparation Example 3, the difference is that no bifida yeast lysate is added in step S7.

Comparative Preparation Example 6

Compared with Preparation Example 3, the difference is that no polysaccharide composition is added in step S7.

Comparative Preparation Example 7

Compared with Preparation Example 3, the difference is that no flavonoid/saponin complex is added in step S7.

Preparation Example 6 Preparation of hirudin/phloretin/ellagic acid chelate complex

Methods as below:

T1. 12 parts by weight of hirudin, 3 parts by weight of phloretin, and 2 parts by weight of ellagic acid were added to 200 parts by weight of water, 2 parts by weight of tris(hydroxymethyl)aminomethane and 5 parts by weight of tannic acid were added, the pH value of the solution was adjusted to 8.2, heated to 40 ° C, and stirred for 1 h to obtain a complex;

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

Preparation Example 7 Preparation of hirudin/phloretin/ellagic acid chelate complex

Methods as below:

T1. 15 parts by weight of hirudin, 5 parts by weight of phloretin, and 4 parts by weight of ellagic acid were added to 200 parts by weight of water, 4 parts by weight of tris(hydroxymethyl)aminomethane and 7 parts by weight of tannic acid were added, the pH value of the solution was adjusted to 8.7, heated to 50 ° C, and stirred for 3 hours to obtain a complex;

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

Preparation Example 8 Preparation of hirudin/phloretin/ellagic acid chelate complex

Methods as below:

T1. 13 parts by weight of hirudin, 4 parts by weight of phloretin, and 3 parts by weight of ellagic acid were added to 200 parts by weight of water, 3 parts by weight of tris(hydroxymethyl)aminomethane and 6 parts by weight of tannic acid were added, the pH value of the solution was adjusted to 8.5, heated to 45 ° C, and stirred for 2 hours to obtain a complex;

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 minutes, dialyze, and dry to obtain a hirudin/phloretin/ellagic acid chelate complex.

Comparative Preparation Example 8

Compared with Preparation Example 8, the difference is that phloretin is not added in step T1.

Comparative Preparation Example 9

Compared with Preparation Example 8, the difference is that ellagic acid is not added in step T1.

Comparative Preparation Example 10

Compared with Preparation Example 8, the difference is that hirudin is not added in step T1.

Comparative Preparation Example 11

Compared with Preparation Example 8, the difference is that tannic acid is not added in step T1.

Comparative Preparation Example 12

Compared with Preparation Example 8, the difference is that step T2 is not performed.

Example 1

This embodiment provides a whitening and lightening essence, the preparation method of which comprises the following steps:

(1) 5 parts by weight of glycerol and 0.3 parts by weight of xanthan gum were added to 40 parts by weight of deionized water, heated to 70° C., and stirred at 300 r/min for 10 min to prepare phase A;

(2) 10 parts by weight of glyceryl tri(ethylhexanoate), 0.5 parts by weight of sorbitan olivate, and 3 parts by weight of the hirudin/phloretin/ellagic acid chelate complex prepared in Preparation Example 6 were mixed, heated to 70° C., and stirred at 300 r/min for 10 min to prepare phase B;

(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 cool down;

(4) After cooling to 35° C., 10 parts by weight of the plant extract prepared in Preparation Example 1 was added, stirred at 300 r/min for 5 min, and filtered to prepare a spot-lightening and whitening essence.

Example 2

This embodiment provides a whitening and lightening essence, the preparation method of which comprises the following steps:

(1) adding 10 parts by weight of hexanediol and 1 part by weight of polyacrylate cross-linked polymer-6 to 60 parts by weight of deionized water, heating to 80° C., and stirring at 500 r/min for 20 min to prepare phase A;

(2) 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 were mixed, heated to 80° C., and stirred at 500 r/min for 20 min to prepare phase B;

(3) Add phase B to phase A, stir at 500 r/min for 15 min, emulsify at 10,000 r/min for 20 min, keep warm for 20 min, and cool down;

(4) After cooling to 40° C., 20 parts by weight of the plant extract prepared in Preparation Example 2 was added, stirred at 500 r/min for 15 min, and filtered to prepare a spot-lightening and whitening essence.

Example 3

This embodiment provides a whitening and lightening essence, the preparation method of which comprises the following steps:

(1) 7 parts by weight of sodium hyaluronate and 0.7 parts by weight of carbomer were added to 50 parts by weight of deionized water, heated to 75° C., and stirred at 400 r/min for 15 min to prepare phase A;

(2) 15 parts by weight of capric triglyceride, 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 were mixed, heated to 75° C., and stirred at 400 r/min for 15 min to prepare phase B;

(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 cool down;

(4) After cooling to 37° C., 15 parts by weight of the plant extract prepared in Preparation Example 3 was added, stirred at 400 r/min for 10 min, and filtered to prepare a spot-lightening and whitening essence.

Example 4

Compared with Example 3, the difference is that the plant extract is prepared by Preparation Example 4.

Example 5

Compared with Example 3, the difference is that the plant extract is prepared by Preparation Example 5.

Comparative Example 1

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 1.

Comparative Example 2

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 2.

Comparative Example 3

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 3.

Comparative Example 4

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 4.

Comparative Example 5

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 5.

Comparative Example 6

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 6.

Comparative Example 7

Compared with Example 3, the difference is that the plant extract is prepared by Comparative Preparation Example 7.

Comparative Example 8

Compared with Example 3, the difference is that the hirudin/phloretin/ellagic acid chelate complex is prepared by Comparative Preparation Example 8.

Comparative Example 9

Compared with Example 3, the difference is that the hirudin/phloretin/ellagic acid chelate complex is prepared by Comparative Preparation Example 9.

Comparative Example 10

Compared with Example 3, the difference is that the hirudin/phloretin/ellagic acid chelate complex is prepared by Comparative Preparation Example 10.

Comparative Example 11

Compared with Example 3, the difference is that the hirudin/phloretin/ellagic acid chelate complex is prepared by Comparative Preparation Example 11.

Comparative Example 12

Compared with Example 3, the difference is that the hirudin/phloretin/ellagic acid chelate complex is prepared by Comparative Preparation Example 12.

Comparative Example 13

Compared with Example 3, the difference is that no plant extract is added.

Comparative Example 14

Compared with Example 3, the difference is that the hirudin/phloretin/ellagic acid chelate complex is not added.

Test Example 1 Cytotoxicity Study

The cytotoxicity of Zhenyaobai was determined by CCK-8 method. HaCaT cells in logarithmic growth phase were collected and inoculated into 96-well plates at a density of 2×10 ⁵ cells/mL, 100 μL per well, 5% CO ₂ , and cultured at 37°C for 24 hours. 100 μL of DMEM complete medium containing the whitening essence prepared in Example 1-5 or Comparative Example 1-14 was added to each well (the concentration of the whitening essence was 200 μg/mL), and only 100 μL of DMEM complete medium was added to the control group, with 3 replicates in each group. After continuing to culture for 24 hours, the cell survival rate was measured by CCK-8 method. The results are shown in Table 1.

Table 1

It can be seen from the above table that the spot-lightening and whitening essence prepared by the present invention has no toxicity to HaCaT cells.

Test Example 2 Study on Cellular Tyrosinase Activity and Melanin Content

B16F10 cells were inoculated at 7×10 ⁴ cells/mL in 24-well and 6-well plates, 500 μL was added to the 24-well plate, and 2 mL was added to the 6-well plate. After 24 hours of culture, 100 nmol/L melanocyte stimulating hormone (α-MSH) was added according to the experimental groups (normal control group, model group, Example 1-5 group, Comparative Example 1-14 group) for induction (except the normal control group), and an α-MSH-induced melanin high expression model was constructed, and then the spot-lightening and whitening essence prepared in Example 1-5 or Comparative Example 1-14 was added (the concentration of the spot-lightening and whitening essence was 200 μg/mL), and 3 replicates were set for each group, and cultured for 48 hours. The results are shown in Table 2.

The supernatant of 24-well cells was discarded, the plates were washed three times with PBS, the cells were lysed, 60 μL of the supernatant was placed in a 96-well plate, 140 μL of 0.1% L-dopamine was added, and the plates were incubated at 37°C for 1 h. The absorbance of each well was measured at a wavelength of 490 nm (A) to determine the tyrosinase activity.

After washing the cells in the 6-well plate three 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, measure the absorbance (A) of each well at a wavelength of 405 nm, and determine the cellular melanin content.

Table 2

It can be seen from the above table that the spot-lightening and whitening essences prepared in Examples 1-3 of the present invention have good effects of inhibiting tyrosinase activity and inhibiting cellular melanin production.

Test Example 3: Removal rate of OH

The spot-lightening and whitening essence prepared in Example 1-5 or Comparative Example 1-14 was diluted with deionized water to 1 mg/mL as a sample solution.

Take 2mL 1.00×10 ^-2 mol/L salicylic acid-ethanol solution, 2mL 9.00×10 ^-3 mol/L FeSO ₄ solution and sample solution into a test tube, then add 2mL 8.8×10 ^-2 mol/L H ₂ O ₂ solution, shake well and place in a 37℃ water bath to react for 30min, measure the absorbance A _i at 510nm; use 2mL pure water instead of salicylic acid solution to measure the absorbance A _j ; use 2mL pure water instead of sample solution to measure the absorbance A _0. Calculate the OH clearance rate (%) according to the formula. Vitamin C was used as the control group. The results are shown in Table 3.

OH removal rate (%) = [1-(A _i -A _j )/A ₀ ] × 100%

table 3

It can be seen from the above table that the spot-lightening and whitening essences prepared in Examples 1-3 of the present invention have good antioxidant activity.

Test Example 4

Volunteers aged 32-45 years old who met the voluntary inclusion criteria were selected and randomly divided into Example 1-5, Comparative Example 1-14 group and blank group, with 10 people in each group.

Experimental samples:

The experimental group used the spot-lightening and whitening essence prepared in Examples 1-5 or Comparative Examples 1-14, and the blank group used an equal amount of water.

Test conditions: On the test day, the subjects cleaned their faces with the designated cleansing samples and sat quietly in a laboratory with a temperature of (25±1)°C and a humidity of 45%±5% RH for 30 minutes.

Experimental method: Before applying the sample, the subjects should wash the test area and apply the sample after drying. An area of 5×5cm was used as the test area; the subjects used the sample twice a day in the morning and evening on the test area. During the experiment, the subjects could not apply any other cosmetics on the test area. The subjects were tested once in the 0th and 4th weeks after continuous use of the sample. At the same time, the application area was washed and the melanin content of the application area was measured by the Mexameter MX18 tester and the MicroSkin II multifunctional dermatoscope image analysis system. Each point was measured five times and the average value was taken. The results are shown in Table 4.

Melanin reduction rate (%) (last melanin content - first melanin content) / first melanin content × 100%

Relative melanin reduction rate (%) = sample group melanin reduction rate (%) - blank group melanin reduction rate (%) Table 4

Group Melanin reduction rate (%) Black relative 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

It can be seen from the above table that the spot-lightening and whitening essences prepared in Examples 1-3 of the present invention have good whitening effects.

The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principle of the present invention should be included in the protection scope of the present invention.

Claims (10)

1. A spot-lightening and whitening essence, characterized in that it 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. 2. The spot-lightening and whitening essence according to claim 1, characterized in that the preparation method of the plant extract comprises the following steps: S1. Enzymatic hydrolysis in a deep eutectic solvent: washing, drying, and crushing birch bark, root of blue ear grass, European plum, peony root, and angelica root respectively to obtain a mixed powder, adding the mixed powder to an aqueous solution containing a deep eutectic solvent, adding a composite enzyme, performing enzymatic hydrolysis, and inactivating the enzyme to obtain an enzymatic hydrolysis product; 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: The bacterial solution obtained in step S2 was frozen in liquid nitrogen, dissolved at room temperature, centrifuged, the supernatant was collected, and freeze-dried 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: uniformly mix 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 to obtain a plant extract. 3. The spot-lightening and whitening essence according to claim 2, characterized in that the mass ratio of the birch bark, the root of the blue ear grass, the European plum, the peony root, and the angelica root in step S1 is 3-5:1-3:7-12:3-5:1-2, the mass ratio of the mixed powder to the aqueous solution containing a deep eutectic solvent is 1:5-7, the complex enzyme is composed of a mixture of cellulase and pectinase, and the mass ratio is 10-15:3-5, the addition amount of the complex enzyme is 2-3wt% of the total mass of the system, the water content in the aqueous solution containing the 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. 4. The spot-lightening and whitening essence according to claim 2 is characterized in that the ethanol content in the system added in step S4 is 60-80wt%, and the precipitation time is 3-5h; the mass ratio of the organic solvent and the solution in step S5 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. 5. The spot-lightening and whitening essence according to claim 2, characterized in that the mass ratio of vitamin C and 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. 6. The spot-lightening and whitening essence according to claim 1, characterized in that 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. 7. The whitening and freckle removal essence according to claim 6, characterized in that the mass ratio of hirudin, phloretin, ellagic acid, tris(hydroxymethyl)aminomethane) and tannic acid in step T1 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 and stirred reaction is 40-50°C, and the time is 1-3h; the mass ratio of the complex, water and silver nitrate in step T2 is 10-15:100:0.2-0.5, and the stirring reaction time is 20-30min. 8. The spot-lightening and whitening essence according to claim 1, characterized in that the emollient is selected from at least one of triethylhexanoin, isononyl isononanoate, dimethicone, caprylic/capric triglyceride, butyrospermumparkii fruit fat, and limnanthes alba seed oil; the humectant 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 olive oil ester, sorbitan olive oil ester, C12-20 alkyl glucoside, and polyglyceryl-2 stearate; the thickener is selected from at least one of carbomer, acrylic acid (ester)/C10-30 alkyl acrylate crosspolymer, sodium polyacryloyldimethyl taurate, polyacrylate crosspolymer-6, and xanthan gum. 9. A method for preparing the spot-lightening and whitening essence according to any one of claims 1 to 8, characterized in that it comprises the following steps: (1) Add a moisturizer and a thickener to 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, 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 cool down; (4) After cooling to 35-40°C, add the plant extract, stir at 300-500 r/min for 5-15 min, filter, and obtain the spot-lightening and whitening essence. 10. Use of the spot-lightening and whitening essence according to any one of claims 1 to 8 in skin whitening, spot-lightening and moisturizing.