A skin-whitening and spot-fading skincare composition and its preparation method

Abstract

This invention provides a skin-whitening and spot-fading composition and its preparation method, belonging to the field of skin-whitening technology. The composition includes modified glycyrrhizin, niacinamide-tranexamic acid co-coated compound, camel thorn flower extract, and artemisia seed extract. Modified glycyrrhizin is obtained by modifying glycyrrhizin with tocopheryl succinate, polyethylene glycol, and artemisia seed gum. The niacinamide-tranexamic acid co-coated compound is obtained by coating a mixture of niacinamide and tranexamic acid with sodium alginate and chitosan hydrochloride. Camel thorn flower extract is obtained by enzymatic hydrolysis of dried camel thorn flower powder with a compound enzyme and glucosidase, followed by purification using a mixed packing adsorption column. Artemisia seed extract is obtained by fermenting artemisia seed powder with Aspergillus oryzae, removing impurities with ADS-7 resin, and purifying using a polyamide resin column. This invention uses special preparation and purification methods to prepare each effective ingredient, which are used in specific ratios to achieve whitening effects through multi-effect synergy, including inhibiting tyrosinase, blocking melanin production, and scavenging free radicals.

Description

Technical Field

[0001] This invention pertains to the field of skin whitening and skincare technology, specifically relating to a skin whitening and spot-fading composition and its preparation method. Background Technology

[0002] Facial pigmentation is primarily caused by the overactivation of melanocytes, triggered by factors such as UV radiation, endocrine disorders, genetic factors, and post-inflammatory hyperpigmentation. Symptoms include age spots, uneven skin tone, and dullness, affecting appearance. Early whitening and spot-fading products were limited to the surface effects of single ingredients, relying heavily on potent components like hydroquinone and mercury compounds to inhibit tyrosinase activity. While these ingredients could quickly fade spots, they posed serious safety risks, easily causing skin irritation, allergies, thinning of the stratum corneum, and even permanent pigmentation, and their use has been restricted or banned. Subsequently, the industry shifted towards exploring gentler ingredients, with vitamin C and its derivatives, arbutin, and kojic acid becoming mainstream. These work by acting as antioxidants and blocking melanin synthesis pathways. However, single-ingredient products have limited target areas and are easily impeded by the skin barrier, resulting in poor penetration and unsatisfactory whitening and spot-fading effects, failing to meet consumers' demand for long-lasting improvement.

[0003] With in-depth research in skin physiology and molecular biology, the entire mechanism of melanin production, transport, and metabolism has become clearer, propelling skin whitening and spot-fading technology into a "multi-target synergistic" stage. Modern skincare composition development no longer relies on single ingredients but rather on the synergistic effect of multiple active ingredients: first, by inhibiting the activity of tyrosinase and tyrosine-related proteins through ingredients such as glycyrrhizin and niacinamide, melanin synthesis is reduced at its source; second, tranexamic acid and tranexamic acid are used to block the transport of melanosomes to keratinocytes; third, gentle exfoliating ingredients such as fruit acids and salicylic acid are added to accelerate stratum corneum metabolism and promote the shedding of pigmented keratinocytes; and fourth, antioxidant ingredients such as vitamin E and glutathione are combined to scavenge free radicals and reduce the activation of melanocytes by ultraviolet radiation and inflammation. Simultaneously, breakthroughs in carrier technology have further improved the effectiveness and safety of these compositions. Liposomes, nanoemulsions, and microencapsulation technologies can precisely deliver active ingredients to the dermis or around melanocytes, preventing inactivation on the surface and reducing skin irritation.

[0004] However, the market is currently flooded with whitening and spot-fading products, with varying degrees of effectiveness. Many products suffer from poor results, irritation, or allergic reactions. There is a need to continuously develop and improve the safety and efficacy of ingredients, and to balance "efficacy" and "safety" by utilizing safe multi-target ingredients to meet market demands. Summary of the Invention

[0005] To address the issues of poor efficacy, irritation, or allergic reactions associated with existing whitening and spot-fading ingredients, a good balance between "high efficacy" and "safety" is difficult to achieve. This invention provides a skin-whitening and spot-fading composition and its preparation method. It employs special preparation and purification methods to prepare the active ingredients, achieving whitening effects through multi-effect synergy, including inhibiting tyrosinase, blocking melanin production, and scavenging free radicals. Furthermore, processes such as "modification, encapsulation, and purification" reduce the irritation of the ingredients, avoiding the shortcomings of traditional products that are either "highly effective but toxic" or "mild but ineffective." The specific technical solution is as follows:

[0006] A skin care composition for whitening and fading spots, wherein the composition comprises modified glycyrrhizin, nicotinamide-tranexamic acid co-encapsulated compound, camel thorn flower extract and sand wormwood seed extract in a mass ratio of (0.8-1.2):(4-5):(3-5):(1-2);

[0007] The preparation of the modified glycyrrhizin includes: a glycyrrhizin-tocopherol succinate-anhydrous ethanol organic phase and a polyethylene glycol-Artemisia argyi seed gum-phosphate buffer solution aqueous phase, which are mixed, sheared and emulsified, and spray-dried to obtain the product.

[0008] The preparation of the nicotinamide-tranexamic acid co-coated compound includes: mixing and shearing the nicotinamide-tranexamic acid-deionized water aqueous phase and sodium alginate-Span 80-liquid paraffin oil phase to obtain a primary emulsion, which is then treated with calcium chloride-isopropanol solution and chitosan hydrochloride-acetic acid aqueous solution, allowed to stand and separate into layers, washed, and spray-dried to obtain the product.

[0009] The preparation of the camel thorn flower extract includes: camel thorn flower powder is dissolved in a citrate-disodium hydrogen phosphate buffer solution, enzymatically hydrolyzed by a complex enzyme and a glucosidase, then extracted with anhydrous ethanol. The supernatant after centrifugation is concentrated under reduced pressure, loaded onto a mixed-packing adsorption column for purification, concentrated, and freeze-dried to obtain the product. The complex enzyme includes cellulase, pectinase, and β-glucanase. The mixed-packing adsorption column is packed with HPD-100 macroporous adsorption resin and polyamide.

[0010] The preparation of the Artemisia argyi seed extract includes: Artemisia argyi seed powder, wheat bran powder, and zinc sulfate are mixed to prepare a fermentation medium, Aspergillus oryzae is inoculated for fermentation and tyrosine and phenylalanine are added, the fermentation broth is sterilized and centrifuged, the supernatant is concentrated, impurities are removed by ADS-7 resin, the sample is loaded onto a polyamide resin column for purification, concentrated, and freeze-dried to obtain the product.

[0011] The modified glycyrrhizin in the above composition is prepared by: preparing an organic phase at a mass ratio of glycyrrhizin:tocopherol succinate:anhydrous ethanol = 1:(0.3-0.5):(12-15) under a water bath at 60℃-65℃; preparing an aqueous phase at a mass ratio of polyethylene glycol:Artemisia argyi seed gum:phosphate buffer solution = 1:(0.08-0.12):(12-15); adding the organic phase to the aqueous phase at a volume ratio of organic phase:aqueous phase = 1:(4-6) under stirring conditions, shearing and emulsifying to obtain an emulsion, and spray drying to obtain modified glycyrrhizin.

[0012] In the above-mentioned method for preparing modified glycyrrhizin, the polyethylene glycol is PEG-1000; the pH value of the phosphate buffer solution is 5.8-6.3; the stirring speed is 300 rpm-500 rpm; and the shear emulsification is performed at 6000 rpm-8000 rpm for 5 min-8 min.

[0013] The preparation method of the nicotinamide-tranexamic acid co-coated compound in the above composition includes: preparing an inner aqueous phase according to the mass ratio of sodium alginate:nicotinamide:tranexamic acid:deionized water = (2-3)(4-5):(1-1.5):(30-40); preparing an oil phase according to the mass ratio of Span 80:liquid paraffin = (1-1.5):(90-100); and adding the inner aqueous phase to the oil phase under stirring according to the volume ratio of inner aqueous phase:oil phase = 1:(2-3), and shearing emulsion. The mixture was stirred to obtain a primary emulsion. While stirring, an isopropanol solution containing 2 wt%–2.5 wt% calcium chloride was added at 1.5–2 times the volume of the primary emulsion. The mixture was stirred again, and then an aqueous solution containing 1.0 wt%–1.5 wt% chitosan hydrochloride and 0.8 wt%–1.0 wt% acetic acid was added at 2–3 times the volume of the primary emulsion. The mixture was stirred, allowed to stand for separation, and the oil phase was discarded to obtain a microsphere suspension. This suspension was washed alternately with n-hexane and deionized water, and then spray-dried to obtain a nicotinamide-tranexamic acid co-coated product.

[0014] In the preparation method of the above-mentioned nicotinamide-tranexamic acid co-coated compound, sodium alginate, nicotinamide and tranexamic acid are dissolved in deionized water at 55℃~60℃, and the pH is adjusted to 7.2~7.5 to prepare an inner aqueous phase; the pH of the inner aqueous phase is adjusted by 0.8M~1.2M sodium hydroxide aqueous solution; the shear emulsification is performed at 4000rpm~6000rpm for 10min~15min; the washing is performed alternately with n-hexane and deionized water for a total of 4 to 6 times.

[0015] The preparation method of the camel thorn flower extract in the above composition includes: drying and pulverizing camel thorn flowers into dry powder; adding the dry powder to a citrate-disodium hydrogen phosphate buffer solution at a material-to-liquid mass ratio of 1:(15-20), then adding a compound enzyme and glucosidase, enzymatically hydrolyzing at 45℃-50℃ for 2-3 hours, inactivating the enzyme in a water bath, and cooling to room temperature to obtain an enzymatic hydrolysate; adding anhydrous ethanol to the enzymatic hydrolysate, extracting by stirring at 60℃-65℃, centrifuging, taking the supernatant, concentrating under reduced pressure to remove ethanol, and obtaining a concentrated solution; loading the concentrated solution onto a mixed packing adsorption column, first eluting with a 5%-10% volume concentration ethanol aqueous solution for 2-3 column volumes to remove impurities, then eluting with a 30%-35% volume concentration ethanol aqueous solution for 1-2 column volumes to remove impurities, and finally eluting with a 60%-70% volume concentration ethanol aqueous solution for 3-4 column volumes, collecting the eluent, concentrating under reduced pressure to remove ethanol, and freeze-drying to obtain the camel thorn flower extract.

[0016] In the above method for preparing camel thorn flower extract, the particle size of the dried powder is sieved through a 40-60 mesh sieve; the pH of the citrate-disodium hydrogen phosphate buffer is pH 4.5-5.0; the amount of the compound enzyme added is 1.5%-2.5% of the mass of the dried powder; the mass ratio of the components of the compound enzyme is cellulase: pectinase: β-glucanase = (2-2.5):(1-1.5):(0.8-1.2); the amount of glucosidase added is 0.2%-0.4% of the mass of the dried powder; the enzyme inactivation in the water bath is performed at 90℃-95℃ for 10 min-1 5 min; the amount of anhydrous ethanol added is 60% to 65% of the volume of the enzymatic hydrolysate; the stirring extraction is carried out at 200 rpm to 300 rpm for 1 to 2 hours; the centrifugation is carried out at 6000 rpm to 8000 rpm for 10 to 15 minutes; the mixed packing adsorption column is prepared by mixing HPD-100 macroporous adsorption resin and polyamide at a mass ratio of (4 to 5): (1 to 1.5), and pre-equilibrated with 90% to 95% volume concentration ethanol aqueous solution and distilled water in sequence; the temperature of the vacuum concentration is 50℃ to 55℃.

[0017] The preparation method of the Artemisia argyi seed extract in the above composition includes: crushing and breaking the shells of Artemisia argyi seeds to obtain Artemisia argyi seed powder; mixing Artemisia argyi seed powder: wheat bran powder: zinc sulfate = 10:(2-3):(1-1.5) by mass to obtain a substrate; mixing the substrate with water at a material-to-liquid mass ratio of 1:(12-15), adjusting the pH to 6.0-6.5 to obtain a fermentation medium, and sterilizing it; aseptically inoculating Aspergillus oryzae into the fermentation medium, and aerobic fermenting at 28℃-30℃ and 150rpm-200rpm for 48h-52h, and supplementing with 0.5%-1.0% of the substrate mass of tyrosine and... The substrate contains 0.3%–0.6% phenylalanine. After fermentation, the mixture is boiled for sterilization, centrifuged, and the supernatant is collected and concentrated under reduced pressure to obtain a concentrate. ADS-7 resin is added, the mixture is mixed, and the ADS-7 resin is removed by filtration. The filtrate is collected and the pH is adjusted to 5.0–5.5. The concentrate is then loaded onto a polyamide resin column. The column is first rinsed with deionized water for 2–3 column volumes to remove impurities, then rinsed with 10%–20% (v / v) ethanol aqueous solution for 1–2 column volumes to remove impurities, and finally eluted with 60%–70% (v / v) ethanol aqueous solution for 3–4 column volumes. The eluent is collected, concentrated under reduced pressure to remove ethanol, and then freeze-dried to obtain the Artemisia argyi seed extract.

[0018] In the above-mentioned method for preparing Artemisia argyi seed extract, the sterilization is performed by steam sterilization at 90℃~100℃ for 30min~45min; the inoculation amount of Aspergillus oryzae is 5%~8% of the substrate mass based on dry cell count; the boiling sterilization is performed by boiling for 10min~15min; the centrifugation is performed at 4℃~6℃ and 8000rpm~10000rpm for 15min~20min; the supernatant is concentrated under reduced pressure to 30%~40% of its volume; the amount of ADS-7 resin added is 2%~5% of the mass of the concentrate; the mixing time is 20min~30min; and the temperature for reduced pressure concentration is 50℃~55℃.

[0019] The preparation method of the above-mentioned skin whitening and spot-fading skin care composition includes the following steps:

[0020] The modified glycyrrhizin, nicotinamide-tranexamic acid co-coated compound, camel thorn flower extract and sand wormwood seed extract were uniformly mixed according to the mass ratio to obtain the composition.

[0021] The above-mentioned skin whitening and spot-fading composition is combined with pharmaceutically or cosmetically available ingredients to prepare a product for whitening and fading spots.

[0022] This invention provides a skin-whitening and spot-fading composition and its preparation method, the beneficial effects of which include:

[0023] I. This invention employs a special preparation and purification method to prepare the effective ingredients. Through multi-effect synergy, it achieves whitening effects by inhibiting tyrosinase, blocking melanin production, and scavenging free radicals. Furthermore, processes such as "modification, encapsulation, and purification" reduce the irritation of the ingredients, avoiding the shortcomings of traditional products that are either "highly effective but toxic" or "mild but ineffective." Breaking through traditional simple mixing or single-component encapsulation techniques, it utilizes self-assembly and ionic cross-linking processes to overcome the application bottlenecks of active ingredients (such as the poor water solubility of glycyrrhizin, the irritation caused by niacinamide, and the easy degradation of tranexamic acid), ensuring that the ingredients do not lose their effectiveness during storage and can penetrate the skin to fully exert their efficacy.

[0024] 2. Glycyrrhizin and tocopherol succinate form an organic phase, while polyethylene glycol 1000 (a hydrophilic surfactant) and Artemisia argyi seed gum (a natural colloid) form an aqueous phase. Self-assembly of nanomicelles is achieved through specific temperature and shear speed. 60-65℃ ensures ethanol solubility and micelle stability, the phase volume ratio avoids micelle aggregation, and the high shear speed ensures uniform micelle particle size. Ultimately, this improves the water solubility and skin permeability of glycyrrhizin. At the same time, the pre-antioxidant effect of tocopherol succinate extends the shelf life of the ingredients.

[0025] III. In the preparation of nicotinamide-tranexamic acid co-coated compounds, sodium alginate and chitosan hydrochloride encapsulate nicotinamide and tranexamic acid; pH 7.2-7.5 ensures that nicotinamide and tranexamic acid dissolve and become charged, adapting to the wall material for adsorption; a specific shear speed forms a stable primary emulsion; the amount of calcium chloride and chitosan controls the microsphere wall thickness, achieving sustained release of the components, reducing the irritation of nicotinamide directly contacting the skin, while protecting tranexamic acid from enzymatic degradation and prolonging the action time.

[0026] IV. In the preparation of camel thorn flower extract, a complex enzyme (cellulase + pectinase + β-glucanase) degrades the cell wall of camel thorn flower, glucosidase converts glycosides into free flavonoids, and HPD-100 macroporous resin and polyamide are used for synergistic purification to remove impurities, enrich highly active ingredients, and enhance antioxidant and melanin-inhibiting abilities.

[0027] V. In the preparation of Artemisia argyi seed extract, Aspergillus oryzae converts the macromolecules (proteins and polysaccharides) of Artemisia argyi seeds into active peptides (barrier repair) and fatty acids (absorption promotion). Tyrosine and phenylalanine are added to provide substrates for Aspergillus oryzae and increase the yield of active peptides. ADS-7 resin adsorbs fermentation byproducts and removes impurities to reduce the stimulation of cells. At the same time, polyamide resin purification further enriches active peptides, enhancing barrier repair and synergistic effects of components.

[0028] VI. The formulation of this compound exhibits excellent synergistic effects. Modified glycyrrhizin, niacinamide-tranexamic acid co-encapsulated compound, camel thorn flower extract, and artemisia seed extract work synergistically across the entire process of "generation-transport-inducing-repair," resulting in more thorough melanin inhibition than single-ingredient formulations. The active peptides in artemisia seed extract repair the skin barrier, reducing potential irritation from modified glycyrrhizin and niacinamide; the sustained-release design of the niacinamide-tranexamic acid co-encapsulated compound reduces direct irritation from niacinamide; the antioxidant components of camel thorn flower extract scavenge free radicals, mitigating oxidative stress from the metabolism of other components. This multi-layered safety profile reduces overall irritation. The tocopheryl succinate of modified glycyrrhizin synergistically combats oxidation with the polyphenols / flavonoids of camel thorn flower extract, preventing other components in the formulation from being oxidized and rendered ineffective; artemisia seed gum (modified glycyrrhizin) and sodium alginate (co-encapsulated compound) are both natural colloids that synergistically stabilize the formulation system, preventing component stratification or aggregation and improving product storage stability. Detailed Implementation

[0029] The present invention will be further described below with reference to specific implementation examples, but the present invention is not limited to these embodiments.

[0030] Example 1

[0031] A skin-whitening and spot-fading skincare composition comprising modified glycyrrhizin, niacinamide-tranexamic acid co-encapsulated compound, camel thorn flower extract, and Artemisia argyi seed extract in a mass ratio of 1:4.5:4:1.5.

[0032] The preparation method of modified glycyrrhizin includes: preparing an organic phase at a mass ratio of glycyrrhizin:tocopherol succinate:anhydrous ethanol = 1:0.4:13 under a 62℃ water bath; preparing an aqueous phase at a mass ratio of polyethylene glycol (PEG-1000):Artemisia argyi seed gum:phosphate buffer solution (pH 6.0) = 1:0.1:13; adding the organic phase to the aqueous phase at a volume ratio of organic phase:aqueous phase = 1:5 under stirring at 400 rpm, and emulsifying at 7000 rpm for 6 min to obtain an emulsion, which is then spray-dried to obtain modified glycyrrhizin.

[0033] The preparation method of the nicotinamide-tranexamic acid co-coated compound includes: dissolving sodium alginate, nicotinamide, and tranexamic acid in deionized water at 58℃ according to a mass ratio of sodium alginate:nicotinamide:tranexamic acid:deionized water = 2.5:4.5:1.3:35, adjusting the pH to 7.4 with 1M sodium hydroxide aqueous solution to obtain an inner aqueous phase; mixing Span 80:liquid paraffin according to a mass ratio of 1.3:95 to obtain an oil phase; adding the inner aqueous phase to the oil phase at a volume ratio of inner aqueous phase:oil phase = 1:2.5 under stirring at 400 rpm, and shearing emulsifying at 5000 rpm for 12 min to obtain a primary emulsion; stirring at 250 rpm... Add 1.8 times the volume of the initial emulsion to an isopropanol solution containing 2.2 wt% calcium chloride, and stir at 250 rpm for 35 min. Then, while stirring at 250 rpm, add 2.5 times the volume of the initial emulsion to an aqueous solution containing 1.2 wt% chitosan hydrochloride and 0.9 wt% acetic acid, stir at 250 rpm for 35 min, allow to stand and separate into layers, discard the oil phase, and obtain a microsphere suspension. Wash the microsphere suspension alternately with n-hexane and deionized water (each time adding 1.2 times the volume of the microsphere suspension as washing solution, stirring for 2 min, allowing to stand and separate into layers, discarding the washing solution, and obtaining a cleaned microsphere suspension) 4 times, and spray dry to obtain the nicotinamide-tranexamic acid co-coated material.

[0034] The preparation method of camel thorn flower extract includes: drying and pulverizing camel thorn flowers, passing them through a 50-mesh sieve to obtain dry powder; adding the dry powder to a citrate-disodium hydrogen phosphate buffer solution at a material-liquid mass ratio of 1:18, then adding 2% by weight of a compound enzyme (of which cellulase:pectinase:β-glucanase = 2.3:1.2:1) and 0.3% by weight of glucosidase, hydrolyzing at 48℃ for 2.5 h, inactivating the enzyme in a 92℃ water bath for 12 min, and cooling to room temperature to obtain an enzymatic hydrolysate; adding 63% by volume of anhydrous ethanol to the enzymatic hydrolysate, and extracting by stirring at 62℃ and 250 rpm for 1.5 h, followed by centrifugation at 7000 rpm for 12 minutes. After 1 minute, the supernatant was collected and concentrated under reduced pressure at 52°C to remove ethanol, yielding a concentrated solution. The concentrated solution was then loaded onto a mixed-packing adsorption column (the mixed-packing adsorption column was prepared by packing HPD-100 macroporous adsorption resin and polyamide at a mass ratio of 4.5:1.2, and pre-equilibrated with 92% ethanol aqueous solution and distilled water sequentially). The column was first eluted with 8% ethanol aqueous solution for 2.5 column volumes to remove impurities, then eluted with 32% ethanol aqueous solution for 1.5 column volumes to remove impurities, and finally eluted with 65% ethanol aqueous solution for 3.5 column volumes. The eluent was collected, concentrated under reduced pressure at 52°C to remove ethanol, and then freeze-dried to obtain camel thorn flower extract.

[0035] The preparation method of Artemisia annua seed extract includes:

[0036] (1) Preparation and sterilization of fermentation medium: The seeds of Artemisia argyi were crushed and shelled, and passed through a 120-mesh sieve to obtain Artemisia argyi seed powder; the substrate was obtained by mixing Artemisia argyi seed powder, wheat bran powder and zinc sulfate in a mass ratio of 10:2.5:1.3. The substrate was mixed with water in a mass ratio of 1:13, the pH was adjusted to 6.3, and the fermentation medium was obtained. The medium was then sterilized by steaming at 95°C for 40 min and cooled to room temperature for later use.

[0037] (2) Preparation of Aspergillus oryzae seed culture (activation and expansion): Aspergillus oryzae strain was inoculated onto PDA slant medium and cultured at 29℃ for 4 days until the spores matured. The spores were washed off with sterile water to prepare a spore suspension. The spore suspension was inoculated into sterilized seed culture medium (wheat bran extract, natural pH) at an inoculation rate of 8% (v / v) and cultured on a shaker at 29℃ and 150 rpm for 30 h to obtain Aspergillus oryzae seed culture.

[0038] (3) Fermentation and extraction: Aseptically inoculate Aspergillus oryzae into the fermentation medium at an inoculation amount of 6.5% of the substrate mass (based on dry cell count, i.e., the required mass of Aspergillus oryzae seed culture for inoculation is calculated based on the bacterial concentration). Ferment at 29℃ and 200 rpm on a shaker (aerobic) for 50 h. During this period, 0.8% tyrosine and 0.5% phenylalanine by substrate mass are added at 26 h of fermentation. After fermentation, the fermentation broth is boiled for 12 min to terminate the reaction and sterilize, and then centrifuged at 5℃ and 9000 rpm for 18 min to collect the supernatant. The supernatant is concentrated under reduced pressure at 53℃ to 35% of its volume to obtain the concentrate. Add 3% ADS-7 resin by mass of the concentrate to the concentrate, mix for 25 min, and then filter to remove the ADS-7 resin. The filtrate was collected and the pH was adjusted to 5.2. The sample was loaded onto a polyamide resin column and first rinsed with deionized water for 2.5 column volumes to remove impurities. Then, it was rinsed with 15% ethanol aqueous solution for 1.5 column volumes to remove impurities. Finally, it was eluted with 65% ethanol aqueous solution for 3.5 column volumes. The eluent was collected, concentrated under reduced pressure at 53°C to remove ethanol, and then freeze-dried to obtain Artemisia argyi seed extract.

[0039] Example 2

[0040] A skin-whitening and spot-fading skincare composition comprising modified glycyrrhizin, niacinamide-tranexamic acid co-encapsulated compound, camel thorn flower extract, and Artemisia argyi seed extract in a mass ratio of 0.8:5:3:2.

[0041] The preparation method of modified glycyrrhizin includes: preparing an organic phase at a mass ratio of glycyrrhizin:tocopherol succinate:anhydrous ethanol = 1:0.5:12 under a 60℃ water bath; preparing an aqueous phase at a mass ratio of polyethylene glycol (PEG-1000):Artemisia argyi seed gum:phosphate buffer solution (pH 6.3) = 1:0.08:15; adding the organic phase to the aqueous phase at a volume ratio of organic phase:aqueous phase = 1:4 under stirring at 500 rpm, and emulsifying at 6000 rpm for 8 min to obtain an emulsion, which is then spray-dried to obtain modified glycyrrhizin.

[0042] The preparation method of the nicotinamide-tranexamic acid co-coated compound includes: dissolving sodium alginate, nicotinamide, and tranexamic acid in deionized water at 60℃ according to a mass ratio of sodium alginate:nicotinamide:tranexamic acid:deionized water = 2:4:1.5:30, adjusting the pH to 7.5 with 0.8M sodium hydroxide aqueous solution to obtain an inner aqueous phase; mixing Span 80:liquid paraffin at a mass ratio of 1.5:90 to obtain an oil phase; adding the inner aqueous phase to the oil phase at a volume ratio of inner aqueous phase:oil phase = 1:3 under stirring at 300 rpm, and shearing emulsifying at 6000 rpm for 10 min to obtain a primary emulsion; and further emulsifying at 200 rpm... Under stirring, add an isopropanol solution containing 2 wt% calcium chloride, twice the volume of the initial emulsion, and stir at 200 rpm for 40 min. Then, under stirring at 200 rpm, add an aqueous solution containing 1.5 wt% chitosan hydrochloride and 0.8 wt% acetic acid, twice the volume of the initial emulsion, and stir at 200 rpm for 40 min. After standing and separating, discard the oil phase to obtain a microsphere suspension. Wash the microsphere suspension alternately with n-hexane and deionized water (each time add a washing solution equal to the volume of the microsphere suspension, stir for 3 min, let stand and separate, discard the washing solution, and obtain a clean microsphere suspension) 4 times. Spray dry to obtain nicotinamide-tranexamic acid co-coated material.

[0043] The preparation method of camel thorn flower extract includes: drying and pulverizing camel thorn flowers, passing them through a 60-mesh sieve to obtain dry powder; adding the dry powder to a pH 5.0 citrate-disodium hydrogen phosphate buffer solution at a material-liquid mass ratio of 1:15, then adding 1.5% by weight of a compound enzyme (of which cellulase:pectinase:β-glucanase = 2.5:1:1.2) and 0.2% by weight of a glucosidase, enzymatically hydrolyzing at 50℃ for 2 hours, inactivating the enzyme in a 95℃ water bath for 10 minutes, and cooling to room temperature to obtain an enzymatic hydrolysate; adding 65% by volume of anhydrous ethanol to the enzymatic hydrolysate, and extracting by stirring at 60℃ and 300 rpm for 1 hour, centrifuging at 8000 rpm. After 10 min, the supernatant was collected and concentrated under reduced pressure at 55°C to remove ethanol, yielding a concentrated solution. The concentrated solution was then loaded onto a mixed-packing adsorption column (the mixed-packing adsorption column was prepared by packing HPD-100 macroporous adsorption resin and polyamide at a mass ratio of 4:1.5 and pre-equilibrated with 90% ethanol aqueous solution and distilled water sequentially). The column was first eluted with 10% ethanol aqueous solution for 2 column volumes to remove impurities, then eluted with 35% ethanol aqueous solution for 1 column volume to remove impurities, and finally eluted with 70% ethanol aqueous solution for 3 column volumes. The eluent was collected, concentrated under reduced pressure at 55°C to remove ethanol, and then freeze-dried to obtain camel thorn flower extract.

[0044] The preparation method of Artemisia annua seed extract includes:

[0045] (1) Preparation and sterilization of fermentation medium: The seeds of Artemisia argyi were crushed and shelled, and passed through a 100-mesh sieve to obtain Artemisia argyi seed powder; the substrate was obtained by mixing Artemisia argyi seed powder, wheat bran powder and zinc sulfate in a mass ratio of 10:3:1. The substrate was mixed with water in a mass ratio of 1:15, the pH was adjusted to 6.0, and the fermentation medium was obtained. The medium was then sterilized by steaming at 100℃ for 30 min and cooled to room temperature for later use.

[0046] (2) Preparation of Aspergillus oryzae seed culture (activation and expansion): Aspergillus oryzae strain was inoculated onto PDA slant medium and cultured at 30℃ for 3 days until the spores matured. The spores were washed off with sterile water to prepare a spore suspension. The spore suspension was inoculated into sterilized seed culture medium (wheat bran extract, natural pH) at an inoculation rate of 10% (v / v) and cultured on a shaker at 28℃ and 200 rpm for 24 h to obtain Aspergillus oryzae seed culture.

[0047] (3) Fermentation and extraction: Aseptically inoculate *Aspergillus oryzae* into the fermentation medium at an inoculation amount equal to 8% of the substrate mass (i.e., the required *Aspergillus oryzae* seed culture mass is calculated based on the bacterial concentration). Ferment at 28℃ and 200 rpm on a shaker (aerobic) for 48 h. During this period, at the 30th h of fermentation, 0.5% tyrosine and 0.6% phenylalanine by substrate mass are added. After fermentation, the fermentation broth is boiled for 10 min to terminate the reaction and sterilize, and then centrifuged at 6℃ and 8000 rpm for 20 min, and the supernatant is collected. The supernatant is concentrated under reduced pressure at 50℃ to 40% of its volume to obtain a concentrated solution. Add 2% ADS-7 resin by mass of the concentrated solution to the concentrated solution, mix for 30 min, and then filter to remove the ADS-7 resin. The filtrate was collected and the pH was adjusted to 5.0. The sample was loaded onto a polyamide resin column. First, it was rinsed with deionized water for 3 column volumes to remove impurities. Then, it was rinsed with 10% ethanol aqueous solution for 2 column volumes to remove impurities. Finally, it was eluted with 60% ethanol aqueous solution for 4 column volumes. The eluent was collected, concentrated under reduced pressure at 50°C to remove ethanol, and then freeze-dried to obtain Artemisia argyi seed extract.

[0048] Example 3

[0049] A skin-whitening and spot-fading skincare composition comprising modified glycyrrhizin, niacinamide-tranexamic acid co-encapsulated compound, camel thorn flower extract, and Artemisia argyi seed extract in a mass ratio of 1.2:4:5:1.

[0050] The preparation method of modified glycyrrhizin includes: preparing an organic phase at a mass ratio of glycyrrhizin: tocopherol succinate: anhydrous ethanol = 1:0.3:15 under a 65℃ water bath; preparing an aqueous phase at a mass ratio of polyethylene glycol (PEG-1000): Artemisia argyi seed gum: phosphate buffer solution (pH 5.8) = 1:0.12:12; adding the organic phase to the aqueous phase at a volume ratio of organic phase: aqueous phase = 1:6 under stirring at 300 rpm, and emulsifying at 8000 rpm for 5 min to obtain an emulsion, which is then spray-dried to obtain modified glycyrrhizin.

[0051] The preparation method of the nicotinamide-tranexamic acid co-coated compound includes: dissolving sodium alginate, nicotinamide, and tranexamic acid in deionized water at 55℃ according to the mass ratio of sodium alginate:nicotinamide:tranexamic acid:deionized water = 3:5:1:40, adjusting the pH to 7.2 with 1.2M sodium hydroxide aqueous solution to obtain the inner aqueous phase; mixing with Span 80:liquid paraffin at a mass ratio of 1:100 to obtain the oil phase; adding the inner aqueous phase to the oil phase at a volume ratio of inner aqueous phase:oil phase = 1:2 under stirring at 500 rpm, and shearing emulsifying at 4000 rpm for 15 min to obtain the primary emulsion; and stirring at 300 rpm... Add 1.5 times the volume of the primary emulsion to an isopropanol solution containing 2.5 wt% calcium chloride, and stir at 300 rpm for 30 min. Then, while stirring at 300 rpm, add 3 times the volume of the primary emulsion to an aqueous solution containing 1.0 wt% chitosan hydrochloride and 1.0 wt% acetic acid, and stir at 300 rpm for 30 min. Allow to stand and separate into layers, discard the oil phase, and obtain a microsphere suspension. Wash the microsphere suspension 6 times with hexane and deionized water alternately (add 1.5 times the volume of the washing solution to the microsphere suspension each time, stir for 2 min, allow to stand and separate into layers, discard the washing solution, and obtain a clean microsphere suspension). Spray dry to obtain nicotinamide-tranexamic acid co-coated material.

[0052] The preparation method of camel thorn flower extract includes: drying and pulverizing camel thorn flowers, passing them through a 40-mesh sieve to obtain dry powder; adding the dry powder to a citrate-disodium hydrogen phosphate buffer solution at a material-liquid mass ratio of 1:20, then adding 2.5% by weight of a compound enzyme (of which cellulase:pectinase:β-glucanase = 2:1.5:0.8) and 0.4% by weight of a glucosidase, enzymatically hydrolyzing at 45℃ for 3 hours, inactivating the enzyme in a 90℃ water bath for 15 minutes, and cooling to room temperature to obtain an enzymatic hydrolysate; adding 60% by volume of anhydrous ethanol to the enzymatic hydrolysate, and extracting at 65℃ and 200 rpm for 2 hours, followed by stirring at 6000 rpm. Centrifuge for 15 min, collect the supernatant, and concentrate under reduced pressure at 50℃ to remove ethanol, obtaining a concentrated solution. Load the concentrated solution onto a mixed-packing adsorption column (the mixed-packing adsorption column is prepared by packing HPD-100 macroporous adsorption resin and polyamide in a mass ratio of 5:1, and pre-equilibrated with 95% ethanol aqueous solution and distilled water sequentially). First, wash with 5% ethanol aqueous solution for 3 column volumes to remove impurities, then wash with 30% ethanol aqueous solution for 2 column volumes to remove impurities, and finally elute with 60% ethanol aqueous solution for 4 column volumes. Collect the eluent, concentrate under reduced pressure at 50℃ to remove ethanol, and freeze-dry to obtain camel thorn flower extract.

[0053] The preparation method of Artemisia annua seed extract includes:

[0054] (1) Preparation and sterilization of fermentation medium: The seeds of Artemisia argyi were crushed and shelled, and passed through a 150-mesh sieve to obtain Artemisia argyi seed powder; the substrate was obtained by mixing Artemisia argyi seed powder, wheat bran powder and zinc sulfate in a mass ratio of 10:2:1.5. The substrate was mixed with water in a mass ratio of 1:12, the pH was adjusted to 6.5, and the fermentation medium was obtained. The medium was then sterilized by steaming at 90°C for 45 min and cooled to room temperature for later use.

[0055] (2) Preparation of Aspergillus oryzae seed culture (activation and expansion): Aspergillus oryzae strain was inoculated onto PDA slant medium and cultured at 28℃ for 5 days until the spores matured. The spores were washed off with sterile water to prepare a spore suspension. The spore suspension was inoculated into sterilized seed culture medium (wheat bran extract, natural pH) at an inoculation rate of 5% (v / v) and cultured on a shaker at 30℃ and 150 rpm for 36 h to obtain Aspergillus oryzae seed culture.

[0056] (3) Fermentation and extraction: Aseptically inoculate Aspergillus oryzae into the fermentation medium at a rate of 5% of the substrate mass (based on dry cell count, i.e., the required mass of Aspergillus oryzae seed culture is calculated based on the bacterial concentration). Ferment at 30℃ and 150 rpm on a shaker (aerobic) for 52 h. During this period, 1.0% tyrosine and 0.3% phenylalanine by substrate mass are added at 24 h of fermentation. After fermentation, the fermentation broth is boiled for 15 min to terminate the reaction and sterilize it, and then centrifuged at 4℃ and 10000 rpm for 15 min to collect the supernatant. The supernatant is concentrated under reduced pressure at 55℃ to 30% of its volume to obtain the concentrate. Add 5% ADS-7 resin by mass of the concentrate to the concentrate, mix for 20 min, and then filter to remove the ADS-7 resin. The filtrate was collected and the pH was adjusted to 5.5. The sample was loaded onto a polyamide resin column. The column was first rinsed with deionized water for 2 column volumes to remove impurities, then rinsed with 20% ethanol aqueous solution for 1 column volume to remove impurities, and finally eluted with 70% ethanol aqueous solution for 3 column volumes. The eluent was collected, concentrated under reduced pressure at 55°C to remove ethanol, and then freeze-dried to obtain Artemisia argyi seed extract.

[0057] A method for preparing a skin-whitening and spot-fading skincare composition according to the above embodiments includes the following steps:

[0058] The modified glycyrrhizin, nicotinamide-tranexamic acid co-coated compound, camel thorn flower extract and sand wormwood seed extract were uniformly mixed according to the mass ratios in each embodiment to obtain the composition.

[0059] The skin care composition for whitening and fading spots in the above embodiments is compounded with pharmaceutically or cosmetically available ingredients to prepare a product for whitening and fading spots.

[0060] Comparative Example 1

[0061] The composition comprises modified glycyrrhizin, nicotinamide-tranexamic acid co-coated compound, camel thorn flower extract, and Artemisia argyi seed extract in a mass ratio of 1:4.5:1.5:4. Other methods and parameters are the same as in Example 1.

[0062] Comparative Example 2

[0063] The composition comprises modified glycyrrhizin, nicotinamide-tranexamic acid co-coated compound, camel thorn flower extract, and Artemisia argyi seed extract in a mass ratio of 1:4.5:5.3:0.2. Other methods and parameters are the same as in Example 1.

[0064] Comparative Example 3

[0065] Modified glycyrrhizin was replaced with glycyrrhizin without modification. Other parameters and methods were the same as in Example 1.

[0066] Comparative Example 4

[0067] In the preparation of modified glycyrrhizin, no Artemisia argyi seed gum was added. Other parameters and methods were the same as in Example 1.

[0068] Comparative Example 5

[0069] The nicotinamide-tranexamic acid co-coating was directly replaced by a mixture of nicotinamide and tranexamic acid in appropriate proportions. Other parameters and methods were the same as in Example 1.

[0070] Comparative Example 6

[0071] In the preparation of camel thorn flower extract, no β-glucanase or glucosidase was added. Other parameters and methods were the same as in Example 1.

[0072] Comparative Example 7

[0073] In the preparation of camel thorn flower extract, the mixed-packed adsorption column was filled only with HPD-100 macroporous adsorption resin. Other parameters and methods were the same as in Example 1.

[0074] Comparative Example 8

[0075] In the preparation of camel thorn flower extract, the mixed packing adsorption column was filled only with polyamide. Other parameters and methods were the same as in Example 1.

[0076] Comparative Example 9

[0077] In the preparation of camel thorn flower extract, HPD-100 macroporous adsorption resin was replaced with D101 macroporous adsorption resin. Other parameters and methods were the same as in Example 1.

[0078] Comparative Example 10

[0079] In the preparation of camel thorn flower extract, AB-8 macroporous adsorption resin was used instead of HPD-100 macroporous adsorption resin. Other parameters and methods were the same as in Example 1.

[0080] Comparative Example 11

[0081] Artemisia annua seed powder was extracted by soaking in a 70% (v / v) ethanol aqueous solution. After removing the ethanol from the extract, it was freeze-dried to obtain Artemisia annua seed extract. Other parameters and methods were the same as in Example 1.

[0082] Comparative Example 12

[0083] In the preparation of Artemisia annua seed extract, Aspergillus oryzae was replaced by Bacillus subtilis. Other parameters and methods were the same as in Example 1.

[0084] Comparative Example 13

[0085] In the preparation of Artemisia annua seed extract, ADS-7 resin adsorption was not added; the concentrate was directly loaded onto a polyamide resin column. Other parameters and methods were the same as in Example 1.

[0086] Comparative Example 14

[0087] In the preparation of Artemisia annua seed extract, polyamide was replaced with D101 macroporous adsorption resin. Other parameters and methods were the same as in Example 1.

[0088] The raw materials used in the above embodiments and comparative examples are as follows: Glycyrrhizin was sourced from Bloomage Biotechnology (Shanxi) Co., Ltd., with a purity of 98%. Tocopherol succinate was sourced from Shaanxi Xinyao Biotechnology Co., Ltd., D-α-tocopherol succinate, model 1210IU. Polyethylene glycol, with an average molecular weight of 1000, was sourced from Shanghai Minke New Materials Co., Ltd., model PEG-1000. Artemisia annua seed gum was sourced from Wuhan Bojuxin Biotechnology Co., Ltd. Nicotinamide was sourced from Xi'an Qinshengyuan Biotechnology Co., Ltd., with a purity of 99%. Tranexamic acid was sourced from Shenzhen Yibohui Biotechnology Co., Ltd., with a purity of 99%. Sodium alginate was sourced from Fujian Rongsen Biotechnology Co., Ltd., 200 mesh, with a purity of 99%. Span 80 was sourced from Guangzhou Zhongjie Chemical Technology Co., Ltd. Liquid paraffin was sourced from Henan Yefeng Chemical Products Co., Ltd., food grade. Isopropanol was sourced from Shanghai Yaokan Chemical Co., Ltd. Chitosan hydrochloride was sourced from Qingdao Boyite Biomaterials Co., Ltd. n-Hexane was sourced from Shanghai Yaokan Chemical Co., Ltd. Cellulase was sourced from Sichuan Huatang Jurui Biotechnology Co., Ltd., with an enzyme activity of 100,000 U / g. Pectinase was sourced from Sichuan Huatang Jurui Biotechnology Co., Ltd., with an enzyme activity of 30,000 U / g. β-glucanase was sourced from Weifang Ruichen Biotechnology Co., Ltd., with an enzyme activity of 50,000 U / g. Glucosidase was sourced from Shanghai Mairui Biochemical Technology Co., Ltd., derived from almonds, with an enzyme activity of 30,000 U / g. HPD-100 macroporous adsorption resin was sourced from Langfang Fucai Chemical Co., Ltd. Polyamide was sourced from Shanghai Yuanye Biotechnology Co., Ltd., 60-100 mesh, model S14144. Aspergillus oryzae strain was sourced from Shanghai Fuxiang Biotechnology Co., Ltd., ATCC 42149. Tyrosine was sourced from Anhui Weimao Biotechnology Co., Ltd., L-tyrosine. Phenylalanine was sourced from Shandong Changxiao Bioengineering Co., Ltd., L-phenylalanine. ADS-7 resin was sourced from Shanghai Yuanye Biotechnology Co., Ltd., ADS-7 macroporous adsorption resin, model S26813. Polyamide resin was sourced from Shanghai Yuanye Biotechnology Co., Ltd., 100-200 mesh, model S14146. D101 macroporous adsorption resin is sourced from Shanghai Yuanye Biotechnology Co., Ltd., model S14161. AB-8 macroporous adsorption resin is sourced from Shanghai Yuanye Biotechnology Co., Ltd., model S30931. Bacillus subtilis is sourced from Shanghai Fuxiang Biotechnology Co., Ltd., ACCC 10627.

[0089] I. Cytotoxicity test (MTT method):

[0090] Sample preparation: The composition powder was dissolved in DMSO to prepare a 100 mg / mL stock solution (sterilized by a 0.22 μm filter membrane), and then diluted with DMEM to 50 μg / mL and 100 μg / mL. The final concentration of DMSO was ≤0.1%.

[0091] The detection method includes: HaCaT cells at 1×104 Cells / wells were seeded into 96-well plates and cultured at 37°C and 5% CO2 for 24 hours to allow adhesion. The medium was then replaced with the sample-containing medium (6 replicates per group), and a negative control containing only the medium (without the composition) was included. The plates were cultured for 24 hours. 20 μL of MTT prepared with 5 mg / mL PBS was added to each well, and the plates were cultured for 4 hours. The medium was then discarded, and 150 μL of LDMSO was added (vortexed at 200 rpm for 10 minutes). The OD value at 570 nm was measured using a microplate reader (630 nm reference correction).

[0092] Evaluation index: Cell viability (%) = (OD of sample group / OD of negative control group) × 100%.

[0093] Table 1. Cytotoxicity test results (average value)

[0094]

[0095] II. Tyrosinase Inhibitory Activity Test:

[0096] Sample preparation: The composition powder was dissolved in DMSO to prepare a 10 mg / mL stock solution, and then diluted with PBS at pH 6.8 (measured at 25°C) to 50 μg / mL and 100 μg / mL. The final concentration of DMSO was ≤1%.

[0097] The detection method included adding 50 μL of sample, 50 μL of 100 U / mL mushroom tyrosinase (prepared in PBS), and 50 μL of L-DOPA prepared in 2.5 mM fresh PBS sequentially to a 96-well plate; a PBS blank control (without enzyme and substrate) and a negative control (containing enzyme and substrate, but without the composite) were set up; the plates were incubated at 37°C for 30 minutes, and the OD value at 475 nm was measured using a microplate reader. Each group had 3 replicates.

[0098] Evaluation index: Inhibition rate (%) = [1 - (OD of sample group - OD of blank group) / (OD of negative control group - OD of blank group)] × 100%.

[0099] Table 2. Results of tyrosinase inhibitory activity assay (average value)

[0100]

[0101] III. Antioxidant Capacity Test:

[0102] DPPH working solution: Prepare a 0.1 mM DPPH solution with anhydrous ethanol, protect from light throughout the process, and store in a sealed, light-protected, and refrigerated container. Stock solution: Prepare a 10 mg / mL stock solution of the test composition with anhydrous ethanol, and filter through a 0.22 μm filter for sterilization. Test solution: Dilute the stock solution with anhydrous ethanol, setting up concentration gradients of 50 μg / mL and 100 μg / mL. Solvent blank control: Pure anhydrous ethanol, used to subtract background absorbance.

[0103] The detection method included: using 96-well plates, with 3 replicates per group. The experimental group (100 μL of the test solution + 100 μL of DPPH working solution), the background group (100 μL of the test solution + 100 μL of anhydrous ethanol), the DPPH blank group (100 μL of anhydrous ethanol + 100 μL of DPPH working solution), and the solvent blank group (100 μL of anhydrous ethanol + 100 μL of anhydrous ethanol) were incubated at 25°C for 30 minutes in the dark. The absorbance was measured at 517 nm using a microplate reader.

[0104] Evaluation index: DPPH removal rate (%) = [1(OD experimental group - OD sample background group) / (OD DPPH blank group - OD solvent blank group)] × 100%.

[0105] Table 3. DPPH free radical scavenging detection results (average value)

[0106]

[0107] IV. Melanin Synthesis Inhibition Test:

[0108] Sample preparation: The composition powder was dissolved in DMSO to prepare a 10 mg / mL stock solution, and then diluted to 50 μg / mL and 100 μg / mL with DMEM medium containing 10% fetal bovine serum and 1% penicillin-dextrose antibodies, respectively. The final concentration of DMSO was ≤0.1%.

[0109] The detection method includes: B16F10 cells at 5×10⁻⁶ 4 Cells / wells were seeded into 6-well plates and cultured at 37°C and 5% CO2 for 24 hours. The medium was then replaced with medium containing 100 nM α-MSH (containing the composition), with a negative control containing α-MSH (without the composition) also included. Cells were cultured for 72 hours. Cells were digested with 0.25% EDTA-containing trypsin, washed three times with PBS, and incubated in 1 mL of 1 mol / L NaOH containing 10% DMSO at 80°C for 1 hour. After centrifugation at 12000 rpm for 10 minutes, the supernatant was collected and the OD value at 405 nm was measured. Melanin content was calculated using a 0-200 μg / mL levodopa standard curve. Three replicates were performed for each group.

[0110] Evaluation index: Melanin synthesis inhibition rate (%) = [1 - (Melanin content of sample group / Melanin content of negative control group)] × 100%.

[0111] Table 4. Results of melanin synthesis inhibition detection (average value)

[0112]

[0113] The results above show that Examples 1 to 3 performed well in all tests. The core reason is that their technical solutions achieved the dual advantages of synergistic component composition and optimized process. With a reasonable ratio, each substance synergistically inhibits melanin synthesis, blocks transport, enhances antioxidant activity, and repairs the skin barrier without any imbalance. Modified glycyrrhizin utilizes a self-assembly technology of "nonionic surfactant-natural colloid-lipid antioxidant" to improve stability and permeability. The nicotinamide-tranexamic acid co-encapsulation compound achieves sustained release and reduced irritation through "pH-driven ion cross-linking." Camelthorn flower extract is purified and enriched with active ingredients using compound enzymatic hydrolysis and mixed filler. Artemisia seed extract undergoes Aspergillus oryzae fermentation and ADS-7 resin impurity removal to enhance activity and purity, thereby improving efficacy and safety.

[0114] Comparative Example 1 (Insufficient Camel Thorn Flower Extract, Excessive Artemisia Seed Extract): This disrupted the synergistic effect of the two natural extracts. Camel Thorn Flower Extract contains polyphenols, flavonoids, and other components, which can help inhibit melanocyte activation and enhance antioxidant capacity. Artemisia Seed Extract, after fermentation with Aspergillus oryzae, produces specific active peptides, whose main function is to repair the cell barrier and reduce the irritation of other components. When Artemisia Seed Extract is excessive and Camel Thorn Flower Extract is insufficient, on the one hand, the antioxidant and melanin-inhibiting active ingredients decrease, resulting in a decline in the efficacy of tyrosinase inhibition, DPPH scavenging, and melanin synthesis inhibition; on the other hand, when Artemisia Seed Extract is excessive, its large molecular components may increase the cell burden, thereby affecting the relative activity of tissues and cell survival rate at high concentrations, thus disrupting the synergistic cycle of "melanin inhibition-antioxidation-barrier repair".

[0115] Comparative Example 2 (Excessive Camel Thorn Flower Extract, Insufficient Artemisia Seed Extract): Although Camel Thorn Flower Extract contains polyphenols and flavonoids, which are antioxidants and help inhibit melanin production, they need to be within a reasonable ratio range to be effective. When the ratio is increased to 5.3, the effect will not be enhanced. Excessive polyphenols will compete with modified glycyrrhizin for tyrosinase active sites, thus reducing the enzyme inhibition efficiency of modified glycyrrhizin. At the same time, excessive flavonoids will compete with tranexamic acid for melanin transport blocking targets, weakening the overall melanin inhibition effect. Excessive Camel Thorn Flower Extract also contains incompletely purified impurities, affecting cell metabolism and stimulating cells to produce a small amount of inflammatory factors; and insufficient Artemisia seed extract results in the loss of barrier protection and synergistic effects of components, leading to a decrease in its safety and efficacy.

[0116] Comparative Example 3 (Unmodified glycyrrhizin as a substitute for modified glycyrrhizin): Modified glycyrrhizin was prepared through a specific organic phase and emulsified with an aqueous phase containing Artemisia selengensis seeds. Firstly, tocopheryl succinate enhances the lipid solubility and permeability of glycyrrhizin. Secondly, the emulsion system formed by Artemisia selengensis seeds encapsulates glycyrrhizin, reducing oxidation and direct stimulation of cells. In contrast, the direct use of unmodified glycyrrhizin results in poor water solubility, insufficient lipid solubility, and poor biocompatibility, leading to decreased inhibitory efficacy. Furthermore, the lack of emulsion encapsulation protection makes it prone to irritation upon direct contact with cells, disrupting cell membrane integrity and increasing susceptibility to free radical oxidation, thus reducing antioxidant capacity and safety.

[0117] Comparative Example 4 (Preparation of modified glycyrrhizin without Artemisia argyi seed gum): Artemisia argyi seed gum plays both the role of "emulsifying stabilizer" and "mild carrier" in the preparation of modified glycyrrhizin. Its high molecular weight polysaccharide structure can synergistically reduce interfacial tension with polyethylene glycol, forming a stable emulsion, preventing glycyrrhizin aggregation, and it has good biocompatibility, forming a protective film on the surface of glycyrrhizin. After removing Artemisia argyi seed gum, the uniformity and stability of the modification are poor, the stability of the emulsion system decreases, and the dispersibility is poor. On the one hand, the local concentration is too high when in contact with cells, which stimulates the cell membrane; on the other hand, the overall synergy of the composition is poor, and the safety, inhibitory efficacy and antioxidant capacity are also reduced.

[0118] Comparative Example 5 (Niacinamide-Tranexamic Acid Mixture as a Substitute for Co-coating): In this example, the nicotinamide-tranexamic acid co-coating was encapsulated using "sodium alginate-chitosan bilayer microspheres," which controlled the release rate of the components, prolonged the duration of action, reduced the irritation of nicotinamide, and enhanced the stability of tranexamic acid. Direct use of the mixture resulted in easy degradation and a rapid decrease in the concentration of the active ingredient; unencapsulated nicotinamide directly contacted skin cells, easily activating TRPV1 channels and causing cell damage; it could not form a stable synergistic effect, and after degradation, its ability to assist in scavenging free radicals decreased, leading to a decline in related inhibitory efficacy, antioxidant capacity, and safety.

[0119] Comparative Example 6 (Camelthorn Flower Extract Preparation Without β-glucanase and Glucosidase): The β-glucanase added in the example can degrade β-glucan in the cell wall of Camelthorn Flower, and Glucosidase can hydrolyze the glycosides in the extract into free flavonoids (aglycones). The antioxidant and melanin-inhibiting activities of free flavonoids are much higher than those of the glycoside form. After removing the two enzymes, on the one hand, the lack of β-glucanase leads to insufficient degradation of the cell wall, making it difficult to release the active ingredients within the cells; on the other hand, the lack of Glucosidase prevents the conversion of glycosides into free aglycones, weakening the overall activity of the extract. At the same time, the insufficiently degraded cell wall residue may increase the cell burden, resulting in a comprehensive reduction in related efficacy and safety.

[0120] Comparative Examples 7 and 8 (using one type of resin for adsorption): HPD-100 macroporous resin and polyamide synergistically achieve efficient impurity removal and active ingredient enrichment. Using only HPD-100 resin, the variety of active ingredients in the purified extract decreased, while the impurity content increased. On the one hand, the reduction in active ingredients leads to a decrease in related efficacy; on the other hand, residual impurities may trigger cellular immune responses, stimulate the production of inflammatory factors, and reduce safety.

[0121] Comparative Examples 9 and 10 (Camel Thorn Flower Extract Purification Using D101 or AB-8 Resin Instead of HPD-100 Resin): The pore size of HPD-100 resin is better suited for the extraction and purification of active ingredients from Camel Thorn Flower. Replacing HPD-100 resin with D101 or AB-8 resin resulted in different pore sizes and polarities, altering the types of active ingredients in the extract, increasing impurities, and negatively impacting various performance characteristics.

[0122] Comparative Example 11 (Ethanol soaking instead of Aspergillus oryzae fermentation for Artemisia annua seed extract): Aspergillus oryzae can secrete enzymes that convert macromolecular components in Artemisia annua seeds into small-molecule bioactive peptides and fatty acids. These conversion products have stronger cell barrier repair capabilities and can also help enhance the melanin-inhibiting activity of other components. Ethanol soaking extraction can only extract water-soluble components and cannot obtain the bioactive peptides and fatty acids produced during fermentation. On the one hand, the lack of bioactive peptides' barrier repair function reduces the cell's resistance to stimuli; on the other hand, the lack of fatty acids' absorption-promoting effect on other components reduces bioavailability. Furthermore, the polysaccharides extracted by ethanol have weak antioxidant activity, leading to a decline in related properties.

[0123] Comparative Example 12 (Fermentation of Artemisia argyi seed extract with Bacillus subtilis instead of Aspergillus oryzae): The enzyme system secreted by Aspergillus oryzae is mainly composed of "protease + saccharifying enzyme", which can efficiently convert the protein in Artemisia argyi seeds into bioactive peptides. After Bacillus subtilis was used as a substitute, the conversion efficiency of protein in Artemisia argyi seeds decreased, and the yield of effective bioactive peptides decreased. On the one hand, the lack of bioactive peptides led to a decrease in the proliferation rate of keratinocytes and a decrease in the integrity of the cell barrier. On the other hand, bioactive peptides can inhibit melanin production through activation pathways. The reduction in their yield led to insufficient activation of this pathway. In addition, the metabolites of Bacillus subtilis lack the fatty acids unique to Aspergillus oryzae, which cannot help enhance the efficacy of other components.

[0124] Comparative Example 13 (purification of Artemisia annua seed extract without ADS-7 resin adsorption): ADS-7 resin can adsorb Aspergillus oryzae metabolic byproducts in the Artemisia annua seed fermentation broth through hydrophobic interactions, avoiding their stimulation of cells or interference with the activity of active ingredients. After removing ADS-7 resin, more harmful impurities such as organic acids remain in the fermentation broth, stimulating inflammatory responses in cells; or they may bind to tyrosinase, affecting the binding efficiency of active ingredients, and the residual organic acids may oxidize active ingredients, reducing antioxidant capacity and leading to a decline in related properties.

[0125] Comparative Example 14 (Purification of Artemisia annua seed extract using D101 resin instead of polyamide resin): Polyamide resin specifically adsorbs the active ingredients in Artemisia annua seed extract via hydrogen bonding, allowing for precise elution; D101 resin is a non-polar macroporous resin that mainly relies on hydrophobic interactions for adsorption, exhibiting low selectivity and easily adsorbing impurities simultaneously. Replacing the resin resulted in a decrease in the recovery rate of active ingredients; impurities increased the cellular burden, affecting related efficacy and safety.

Claims

1. A skin care composition for whitening and fading spots, characterized by, The modified glabridin, the nicotinamide-co-coated with citric acid, the extract of the camel flower and the extract of the sahu seed are in a mass ratio of (0.8-1.2):(4-5):(3-5):(1-2); The preparation of the modified glabridin comprises: an organic phase of glabridin-tocopherol succinate-anhydrous ethanol and an aqueous phase of polyethylene glycol-sahu seed gum-phosphate buffer solution, mixing and shearing emulsification, and spray drying to obtain the product. The preparation of the nicotinamide-co-coated with citric acid comprises: an inner aqueous phase of nicotinamide-co-coated with citric acid-deionized water and an oil phase of sodium alginate-Span 80-liquid paraffin, mixing and shearing emulsification to obtain a primary emulsion, and then treating with calcium chloride-isopropyl alcohol solution and chitosan hydrochloride-acetic acid aqueous solution, washing after static layering, and spray drying to obtain the product. The preparation of the camel flower extract comprises: dry powder of the camel flower in a citric acid-disodium hydrogen phosphate buffer solution, co-enzymolysis by composite enzymes and glucosidase, and then adding anhydrous ethanol for extraction, and then centrifuging, concentrating under reduced pressure, and purifying by loading into a mixed filler adsorption column, concentrating, and freeze-drying to obtain the product; the composite enzymes comprise cellulase, pectinase and β-glucanase; and the filler of the mixed filler adsorption column comprises HPD-100 macroporous adsorption resin and polyamide. The preparation of the sahu seed extract comprises: mixing sahu seed powder, bran powder and zinc sulfate to prepare a fermentation medium, inoculating Aspergillus oryzae for fermentation and supplementing tyrosine and phenylalanine, sterilizing and centrifuging the fermentation liquor, concentrating the supernatant, removing impurities by ADS-7 resin, purifying by loading into a polyamide resin column, concentrating, and freeze-drying to obtain the product.

2. The skin care composition for whitening and fading freckles according to claim 1, wherein The preparation method of the modified glabridin comprises: under a water bath at 60-65°C, preparing an organic phase according to a mass ratio of glabridin:tocopherol succinate:anhydrous ethanol=1:(0.3-0.5):(12-15); preparing an aqueous phase according to a mass ratio of polyethylene glycol:sahu seed gum:phosphate buffer solution=1:(0.08-0.12):(12-15); and under stirring, adding the organic phase into the aqueous phase, shearing emulsification to obtain an emulsion, and spray drying to obtain the modified glabridin.

3. The skin care composition for whitening and freckle removal according to claim 2, wherein The polyethylene glycol is PEG-1000; the pH value of the phosphate buffer solution is 5.8-6.3; the stirring speed is 300-500 rpm; and the shearing emulsification is shearing emulsification at 6000-8000 rpm for 5-8 min.

4. The skin care composition for whitening and freckle removal according to claim 1, wherein The preparation method of the nicotinamide-tartaric acid co-coating includes: preparing an inner water phase according to the mass ratio of sodium alginate: nicotinamide: tartaric acid: deionized water = (2-3) (4-5) : (1-1.5) : (30-40); preparing an oil phase according to the mass ratio of Span 80: liquid paraffin = (1-1.5) : (90-100); adding the inner water phase into the oil phase under stirring to shear emulsify and obtain a primary emulsion according to the volume ratio of the inner water phase: oil phase = 1: (2-3); adding an isopropanol solution containing 2wt%-2.5wt% calcium chloride in 1.5-2 times the volume of the primary emulsion under stirring, stirring, adding an aqueous solution containing 1.0wt%-1.5wt% chitosan hydrochloride and 0.8wt%-1.0wt% acetic acid in 2-3 times the volume of the primary emulsion, stirring, and standing to separate layers to discard the oil phase and obtain a microsphere suspension, which is washed alternately with n-hexane and deionized water, spray dried to obtain the nicotinamide-tartaric acid co-coating.

5. The skin care composition for whitening and freckle removal according to claim 4, wherein The sodium alginate, nicotinamide and tartaric acid are dissolved in deionized water at 55-60°C, the pH is adjusted to 7.2-7.5 to prepare the inner water phase; the pH of the inner water phase is adjusted by using a 0.8M-1.2M sodium hydroxide aqueous solution; the shear emulsification is 4000rpm-6000rpm shear emulsification for 10-15min; the n-hexane and deionized water are alternately washed for 4-6 times.

6. The skin care composition for whitening and freckle removal according to claim 1, wherein The preparation method of the camel thorn flower extract includes: drying and crushing camel thorn flower into dry powder; adding the dry powder into a citric acid-sodium phosphate buffer solution according to a solid-liquid mass ratio of 1: (15-20), then adding a compound enzyme and a glucosidase, and performing enzymolysis at 45-50°C for 2-3h, water bath enzyme inactivation, and cooling to room temperature to obtain an enzymolysis liquid; adding anhydrous ethanol to the enzymolysis liquid, stirring and extracting at 60-65°C, centrifuging, taking the supernatant, and removing ethanol under reduced pressure to obtain a concentrated liquid; loading the concentrated liquid into a mixed filler adsorption column, first washing impurities with 5%-10% volume concentration ethanol aqueous solution for 2-3 column volumes, then washing impurities with 30%-35% volume concentration ethanol aqueous solution for 1-2 column volumes, finally eluting with 60%-70% volume concentration ethanol aqueous solution for 3-4 column volumes, collecting the eluate, removing ethanol under reduced pressure, and freeze-drying to obtain the camel thorn flower extract.

7. The skin care composition for whitening and freckle removal according to claim 6, wherein The particle size of the dry powder is 40-60 mesh; the pH of the citric acid-disodium hydrogen phosphate buffer is 4.5-5.0; the addition amount of the complex enzyme is 1.5-2.5% of the dry powder; the mass ratio of the components of the complex enzyme is cellulase: pectinase: beta-glucanase = (2-2.5): (1-1.5): (0.8-1.2); the addition amount of the glucosidase is 0.2-0.4% of the dry powder; the water bath enzyme inactivation is 90-95°C water bath enzyme inactivation for 10-15 min; the addition amount of the anhydrous ethanol is 60-65% of the enzyme hydrolysis solution; the stirring extraction is 200-300 rpm stirring extraction for 1-2 h; the centrifugation is 6000-8000 rpm centrifugation for 10-15 min; the mixed filler adsorption column is filled with HPD-100 macroporous adsorption resin and polyamide at a mass ratio of (4-5):(1-1.5), and is pre-equilibrated with 90-95% volume concentration ethanol aqueous solution and distilled water in sequence; and the temperature of the reduced pressure concentration is 50-55°C.

8. The skin care composition for whitening and freckle removal according to claim 1, wherein The preparation method of the Artemisia sphaerocephala Krasn. seed extract comprises the following steps: crushing and breaking the shells of Artemisia sphaerocephala Krasn. seeds to obtain Artemisia sphaerocephala Krasn. seed powder; mixing the Artemisia sphaerocephala Krasn. seed powder, bran powder and zinc sulfate according to a mass ratio of 10:(2-3):(1-1.5) to obtain a substrate; mixing the substrate and water according to a material-liquid mass ratio of 1:(12-15), adjusting the pH to 6.0-6.5 to obtain a fermentation medium, and sterilizing; inoculating Aspergillus oryzae into the fermentation medium through aseptic operation, and carrying out aerobic fermentation at 28-30°C and 150-200 rpm for 48-52 h; adding tyrosine and phenylalanine at 0.5-1.0% and 0.3-0.6% of the mass of the substrate at the 24th-30th hour of fermentation; after the fermentation is completed, boiling and sterilizing, centrifuging, taking the supernatant, reducing pressure and concentrating to obtain a concentrated solution, adding ADS-7 resin, mixing, filtering to remove the ADS-7 resin, collecting the filtrate, adjusting the pH to 5.0-5.5, and loading the filtrate into a polyamide resin column; first, washing the column with deionized water for 2-3 column volumes to remove impurities, then washing the column with 10-20% volume concentration ethanol aqueous solution for 1-2 column volumes to remove impurities, finally, eluting the column with 60-70% volume concentration ethanol aqueous solution for 3-4 column volumes, collecting the eluate, removing ethanol through reduced pressure concentration, and freeze-drying to obtain the Artemisia sphaerocephala Krasn. seed extract.

9. The skin care composition for whitening and freckle removal according to claim 8, wherein The sterilization is 90-100°C steam sterilization for 30-45 min; the inoculation amount of Aspergillus oryzae is 5-8% of the mass of the substrate; the boiling sterilization is boiling for 10-15 min; the centrifugation is centrifugation at 4-6°C and 8000-10000 rpm for 15-20 min; the supernatant is reduced pressure concentrated to 30-40% of the volume; the addition amount of the ADS-7 resin is 2-5% of the mass of the concentrated solution; the mixing time is 20-30 min; and the temperature of the reduced pressure concentration is 50-55°C.

10. A method of preparing a skin care composition for whitening and freckle improvement according to claim 1, characterized by, The method comprises the following steps: The modified glabridin, the nicotinamide-aminic acid co-coating, the extract of K. camelorum and the extract of S. potaninii are uniformly mixed in a mass ratio to obtain a composition.

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