An anti-inflammatory whitening composition, and a method of preparing and using the same

Abstract

The application belongs to the technical field of traditional Chinese medicines, and relates to an anti-inflammatory and whitening composition as well as a preparation method and application thereof. The composition comprises radix adiantidis, herba lepidagathis and rhizoma atractylodis. The radix adiantidis, herba lepidagathis and rhizoma atractylodis are selected as raw materials, and the three raw materials are scientifically compounded to achieve synergistic effect, so that the activity of tyrosinase can be efficiently inhibited, and the anti-inflammatory and whitening effects can be achieved. Through strict formula and process research, the raw materials in the composition are stable and compatible, the proportion is clear, and the safety is high, so that the problems of complex composition and large toxic and side effects of the composition are avoided.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine technology, and relates to an anti-inflammatory and whitening composition, its preparation method and application. Background Technology

[0002] The main process of skin pigmentation is as follows: Melanin is synthesized by mature melanosomes within melanocytes. Melanosomes then migrate from the perinuclear region to the distal dendrites, are transferred to adjacent keratinocytes, and are subsequently redistributed and degraded within the keratinocytes. During this process, increased melanin granule synthesis, decreased transport of melanin granules from melanosomes to adjacent keratinocytes, and a decline in the metabolic level of keratinocytes can all lead to an increase in melanin granules in the skin. The currently accepted pathway for melanin formation is as follows: In melanocytes, tyrosine is oxidized under the catalysis of tyrosinase to produce dopa and dopaquinone. Dopaquinone is further oxidized to dopachrome, which is then rearranged and polymerized through dihydroxyindole to form melanin. Melanin then combines with proteins to form melanin protein.

[0003] Increased melanin synthesis and accumulation can lead to various diseases in the human body, such as acanthosis nigricans, atopic dermatitis of the neck, melasma, periorbital hyperpigmentation, freckle-like nevi, and other skin diseases, as well as the two neurodegenerative diseases Parkinson's disease and Alzheimer's disease.

[0004] In the process of melanin formation, tyrosinase is a major rate-limiting enzyme in the conversion of tyrosine to melanin in melanocytes, playing a crucial role in the formation of melanosomes and the transformation of melanin. Therefore, inhibiting tyrosinase is one of the important ways to reduce melanin formation.

[0005] Glycyrrhizin, an active ingredient extracted from licorice root, is hailed as "whitening gold" due to its powerful skin-whitening effects. Glycyrrhizin inhibits tyrosinase activity and has excellent sun protection properties. Peptido, a patented skin-whitening ingredient developed by Beiersdorf, is a tyrosinase inhibitor used to treat post-inflammatory hyperpigmentation. However, both glycyrrhizin and Peptido are expensive and exhibit some cytotoxicity. In recent years, naturally derived tyrosinase inhibitors have attracted widespread interest.

[0006]

[0007]

[0008] Chinese invention patent CN103202928A discloses a traditional Chinese medicine composition with anti-aging function, which is composed of wolfberry extract, jasmine flower extract, lotus seed heart extract, Polygonum multiflorum extract, Ligusticum chuanxiong extract, Forsythia suspensa extract, and Eriocaulon buergerianum extract. This traditional Chinese medicine composition can be used in anti-aging and skin whitening formulas, and has the functions of inhibiting tyrosinase activity, scavenging free radicals, and whitening the skin.

[0009] Chinese invention patent CN111419747A discloses a skin-whitening and brightening herbal composition, including Cimicifuga rhizome, Angelica pubescens root, Saposhnikovia root, Glycyrrhiza root, Morus root bark, Atractylodes rhizome, Astragalus root, Ginseng root, and Pueraria root. This composition can inhibit tyrosinase activity and has a good scavenging effect on DPPH free radicals. However, this herbal composition, with its complex combination of multiple herbs and complex mechanism of action, has the potential to produce side effects.

[0010] Therefore, developing new, safe, and efficient tyrosinase inhibitors is of great significance. Summary of the Invention

[0011] In view of the technical problems of low inhibitory activity and certain side effects of existing tyrosinase inhibitors, the purpose of this invention is to provide an anti-inflammatory and whitening composition, its preparation method and application.

[0012] To achieve the above objectives, the present invention adopts the following technical solution:

[0013] The present invention provides an anti-inflammatory and skin-whitening composition comprising North American ginseng, Eriocaulon buergerianum and Atractylodes lancea.

[0014] Furthermore, the North American ginseng is North American ginseng slices, North American ginseng water extract, or North American ginseng alcohol extract.

[0015] Furthermore, the *Eriocaulon buergerianum* is *Eriocaulon buergerianum* tablets, *Eriocaulon buergerianum* water extract, or *Eriocaulon buergerianum* alcohol extract.

[0016] Furthermore, the Atractylodes lancea mentioned above is Atractylodes lancea slices, Atractylodes lancea water extract, or Atractylodes lancea alcohol extract.

[0017] Furthermore, the composition comprises, by weight, 10-50 parts of North American ginseng, 10-30 parts of Eriocaulon buergerianum and 10-20 parts of Atractylodes lancea.

[0018] Furthermore, the composition comprises, by weight, 10-40 parts of North American ginseng, 10-25 parts of Eriocaulon buergerianum and 10-20 parts of Atractylodes lancea.

[0019] Furthermore, in the composition, the mass ratio of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea is 0.5-3:0.5-2.5:1.

[0020] Furthermore, in the composition, the mass ratio of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea is 1:1:1.

[0021] Glehnia littoralis F.Schmidt ex Miq., also known as coral vegetable, is a plant belonging to the Apiaceae family, and its root is used medicinally. The main components of Glehnia littoralis are coumarins, lignans, polyacetylenes, flavonoids, organic acids, terpenes, and steroids. Various ancient texts record more than 50 traditional Chinese medicine prescriptions related to Glehnia littoralis, which can be used to clear lung heat, relieve cough and moisten the lungs, and treat chronic bronchitis. It is also used in folk cuisine for making soups and porridges. Clinically, Glehnia littoralis is combined with other Chinese herbs to create prescriptions for treating gastrointestinal diseases, cardiovascular and cerebrovascular diseases, and liver dysfunction, enhancing treatment efficacy and ensuring treatment safety.

[0022] Eriocaulon buergerianum It is a herbaceous plant belonging to the genus *Eriocaulon* in the family Eriocaulonceae. According to the *Compendium of Materia Medica*, *Eriocaulon* treats headaches, blindness, corneal opacity, corneal opacity after smallpox, and stops bleeding. It is pungent and sweet in taste, neutral in nature, and enters the liver and lung meridians.

[0023] Atractylodes lancea (Thunb.) DC. is a perennial herb belonging to the genus Atractylodes in the family Asteraceae. Its rhizome is used medicinally. Atractylodes lancea is warm in nature and pungent in taste. It is often used to treat abdominal distension, nausea, diarrhea, wind-cold-dampness arthralgia, and swelling and pain in the feet and knees caused by dampness obstructing the middle jiao.

[0024] The present invention also provides a method for preparing the composition, comprising the following steps:

[0025] S1. Crush and grind North American ginseng, Eriocaulon buergerianum and Atractylodes lancea respectively to obtain North American ginseng powder, Eriocaulon buergerianum powder and Atractylodes lancea powder;

[0026] S2. Add ethanol to the powder of North American ginseng, the powder of Eriocaulon buergerianum, or the powder of Atractylodes lancea in step S1, heat and reflux to extract, filter, and obtain North American ginseng extract, Eriocaulon buergerianum extract and Atractylodes lancea extract respectively.

[0027] S3. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S2 are concentrated and dried to obtain alcohol extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea.

[0028] S4. Mix the extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S3 according to the formula to obtain the composition.

[0029] Furthermore, the particle size of the North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder mentioned in step S1 is 60-120 mesh.

[0030] Furthermore, the mass concentration of ethanol in step S2 is 60%-80%.

[0031] Furthermore, the ethanol concentration in step S2 is 75%.

[0032] Further, in step S2, the ratio of North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder to ethanol is 1:30-50g / 100mL; the temperature of the heating and reflux extraction in step S2 is 70-85℃, and the heating and reflux extraction time is 1.5-4h.

[0033] Furthermore, in step S2, the ratio of North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder to ethanol is 1:40g / 100mL; the temperature for the heating and reflux extraction in step S2 is 75℃, and the heating and reflux extraction time is 3h.

[0034] Furthermore, the concentration described in step S3 is to concentrate the extract to 1 / 3 to 1 / 5 of its original volume.

[0035] Furthermore, the drying process described in step S3 is freeze drying.

[0036] The present invention also provides the use of the composition in the preparation of anti-inflammatory and whitening pharmaceuticals and cosmetics.

[0037] The present invention also provides an anti-inflammatory and whitening product comprising the aforementioned composition.

[0038] Furthermore, the aforementioned anti-inflammatory and whitening products also contain medically acceptable excipients.

[0039] Furthermore, the products mentioned are pharmaceuticals and cosmetics.

[0040] Furthermore, the dosage form of the product is tablets, capsules, oral liquids, decoctions, pills, granules, powders, sprays, ointments, or emulsions.

[0041] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0042] This invention selects North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea as raw materials. Through the scientific compounding of these three raw materials, they exert a synergistic effect, which can effectively inhibit the activity of tyrosinase and has anti-inflammatory and whitening effects.

[0043] This invention has undergone rigorous formulation and process research. The raw materials in the composition are stable and compatible, with a clear proportion, and high safety, avoiding the problems of complex composition and high toxicity. Attached Figure Description

[0044] Figure 1To investigate the effect of different concentrations of Example 1 on the proliferation activity of B16 cells, compared with the control group, ***, P < 0.001; ****, P < 0.0001;

[0045] Figure 2 To investigate the effect of different concentrations of Comparative Example 6 on the proliferation activity of B16 cells, compared with the control group, **, P < 0.01, ****, P < 0.0001;

[0046] Figure 3 The effect of different concentrations of Comparative Example 7 on the proliferation activity of B16 cells was compared with that of the control group. The results showed that the effect was statistically significant (P < 0.0001).

[0047] Figure 4 The effects of different concentrations of Example 1 and Comparative Examples 6-7 on the tyrosinase activity of B16 cells were compared with the control group. The results showed that the effect was statistically significant (P < 0.0001).

[0048] Figure 5 The effects of different concentrations of Example 1 on zebrafish;

[0049] Figure 6 The effect of different concentrations of Example 1 on zebrafish embryo survival;

[0050] Figure 7 The effect of different concentrations of Example 1 on the number of melanin patches in the head of zebrafish was compared with the control group. The results showed that the effect was ****, and P < 0.0001.

[0051] Figure 8 The NO content in the supernatant of RAW264.7 cells after treatment with different concentrations of Example 1 and Comparative Examples 6-7 was detected. Compared with the +LPS group, ****, P < 0.0001; ***, P < 0.001. Detailed Implementation

[0052] The following specific examples illustrate the implementation of the present invention. Those skilled in the art can easily understand other advantages and effects of the present invention from the content disclosed in this specification. The present invention can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of the present invention.

[0053] Before further describing specific embodiments of the present invention, it should be understood that the scope of protection of the present invention is not limited to the specific embodiments described below; it should also be understood that the terminology used in the embodiments of the present invention is for describing specific embodiments and not for limiting the scope of protection of the present invention.

[0054] When numerical ranges are given in the embodiments, it should be understood that, unless otherwise stated in the invention, both endpoints of each numerical range and any value between the two endpoints may be selected. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

[0055] Unless otherwise specified, all raw materials used in this invention are commercially available products commonly used in this technical field.

[0056] Example

[0057] The formulations of the compositions in Examples 1-3 and Comparative Examples 1-4 are shown in Table 1.

[0058] Table 1

[0059]

[0060]

[0061] The preparation methods of the compositions of Examples 1-3 and Comparative Examples 1-4 include the following steps:

[0062] S1. Dry the medicinal materials of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea at 40℃, crush them, and grind them into powder to obtain North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder.

[0063] S2. Add 70wt% ethanol to the powders of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea in step S1 at a ratio of 1:40g / 100mL, heat to 75℃, reflux for 3h, cool to 25℃ and filter to remove impurities to obtain North American ginseng extract, Eriocaulon buergerianum extract and Atractylodes lancea extract.

[0064] S3. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S2 are concentrated to 1 / 5 of their original volume, and then freeze-dried for 48 hours to obtain the alcohol extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea.

[0065] S4. Mix the extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S3 according to the formula to obtain the composition.

[0066] The particle size of the North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder mentioned in step S1 is 100 mesh.

[0067] Example 4

[0068] The difference between this embodiment and Example 1 lies in the preparation method of the composition, as detailed below:

[0069] The method for preparing the composition includes the following steps:

[0070] S1. Dry the medicinal materials of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea at 40℃, crush them, and grind them into powder to obtain North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder.

[0071] S2. Add 60wt% ethanol to the powders of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea in step S1 at a ratio of 1:30g / 100mL, heat to 70℃, reflux for 1.5h, cool to 25℃, and filter to remove impurities to obtain North American ginseng extract, Eriocaulon buergerianum extract, and Atractylodes lancea extract.

[0072] S3. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S2 are concentrated to 1 / 4 of their original volume, and then freeze-dried for 48 hours to obtain the alcohol extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea.

[0073] S4. Mix the extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S3 according to the formula to obtain the composition.

[0074] The particle size of the North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder mentioned in step S1 is 60 mesh.

[0075] Example 5

[0076] The difference between this embodiment and Example 1 lies in the preparation method of the composition, as detailed below:

[0077] The method for preparing the composition includes the following steps:

[0078] S1. Dry the medicinal materials of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea at 40℃, crush them, and grind them into powder to obtain North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder.

[0079] S2. Add 80wt% ethanol to the powders of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea in step S1 at a ratio of 1:50g / 100mL, heat to 85℃, reflux for 4h, cool to 25℃ and filter to remove impurities to obtain North American ginseng extract, Eriocaulon buergerianum extract and Atractylodes lancea extract.

[0080] S3. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S2 are concentrated to 1 / 5 of their original volume, and then freeze-dried for 48 hours to obtain the alcohol extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea.

[0081] S4. Mix the extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S3 according to the formula to obtain the composition.

[0082] The particle size of the North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder mentioned in step S1 is 120 mesh.

[0083] Comparative Example 5

[0084] The difference between this comparative example and Example 1 lies in the preparation method of the composition, as detailed below:

[0085] The method for preparing the composition includes the following steps:

[0086] S1. Dry the medicinal materials of North American ginseng, Eriocaulon buergerianum and Atractylodes lancea at 40℃, crush them, and grind them into powder to obtain North American ginseng powder, Eriocaulon buergerianum powder and Atractylodes lancea powder.

[0087] S2. Add 50wt% methanol to the powdered Glehnia littoralis, Eriocaulon buergerianum, and Atractylodes lancea in step S1 at a ratio of 1:20g / 100mL, heat to 75℃, reflux for 3h, cool to 25℃ and filter to remove impurities to obtain Glehnia littoralis extract, Eriocaulon buergerianum extract and Atractylodes lancea extract.

[0088] S3. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S2 are concentrated to 1 / 5 of their original volume, and then freeze-dried for 48 hours to obtain the alcohol extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea.

[0089] S4. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S3 are mixed evenly according to the formula amount in Example 1 to obtain the composition.

[0090] The particle size of the North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder mentioned in step S1 is 100 mesh.

[0091] Comparative Example 6

[0092] Glycyrrhizin, CAS: 59870-68-7, purity: ≥98%, purchased from Shanghai Xige Biotechnology Co., Ltd.

[0093] Comparative Example 7

[0094] Peptide Anmido, CAS: 1428450-95-6, purity: 99%, purchased from Wuhan Pushida Biotechnology Co., Ltd.

[0095] Performance test

[0096] I. Detection of toxicity to B16 cells

[0097] 1. Test Methods

[0098] B16 cells cultured to the logarithmic growth phase were placed in 24-well plates and incubated overnight at 37°C with 5% (v / v) CO2. The cell concentration was adjusted to 2.0 × 10⁶ cells / well.5 Cells were added at a concentration of 0.4 mL per well, resulting in a total cell count of approximately 80,000. After 24 hours of culture, different concentrations of solutions from Example 1 and Comparative Examples 6-7 (dissolved in DMSO, 0.1 mL per well) were added, while the control group received culture medium. Culture was continued for another 24 hours after drug addition. After 24 hours, 50 μL of CCK-8 reagent (Cell Counting Kit-8) was added to each well, and the cells were cultured for 1 hour. The supernatant was collected and transferred to a 96-well plate (100 μL per well), with three replicates per group. The OD value was measured at 450 nm using a microplate reader.

[0099]

[0100] 2. Test Results

[0101] The effects of different concentrations of Example 1 and Comparative Examples 6-7 on the proliferation activity of B16 cells were investigated, and the results are as follows: Figure 1-3 In Example 1, the concentrations were set at 3, 10, 30, 50, 75, 100, 150, and 200 μg / mL. (From...) Figure 1 As can be seen, after 24 hours of treatment, the survival rate of B16 cells was higher than 90% at concentrations of 3-30 μg / mL with the composition of Example 1. As the concentration of the composition of Example 1 increased, the growth of B16 cells was affected to some extent. At 50 μg / mL, the cell survival rate was 80%. When the concentration of the composition of Example 1 reached 75 μg / mL and higher, it began to produce a very significant inhibitory effect.

[0102] The effect of glycyrrhizin at concentrations of 1, 3, 10, and 20 μg / mL on the proliferation activity of B16 cells was investigated in Comparative Example 6. The results are as follows: Figure 2 At concentrations >10 μg / mL, glycyrrhizin exhibits strong toxicity to B16 cell growth.

[0103] The effect of the comparative 7-peptide amidato at concentrations of 0.3, 1, 3, and 10 μg / mL on the proliferation activity of B16 cells was investigated, and the results are as follows: Figure 3 Peptido did not inhibit the growth of B16 cells at a concentration of ≤3 μg / mL.

[0104] II. Effects on tyrosinase activity in B16 cells

[0105] 1. Test Methods

[0106] B16 cells cultured to the logarithmic growth phase were placed in 24-well plates and incubated overnight at 37°C with 5% (v / v) CO2. The cell concentration was adjusted to 2.0 × 10⁻⁶ cells / well. 5Cells were cultured at a concentration of 0.4 mL / well, with a total cell count of approximately 80,000 per well. After 24 hours of culture, different concentrations of the drugs from Example 1 and Comparative Examples 6-7 (0.1 mL / well) were added, while the control group received only culture medium. After drug administration (Examples 1 and Comparative Examples 6-7), cells were cultured for another 24 hours, then the culture medium was discarded, and the cells were washed twice with 0.25 mL of PBS. 0.25 mL of 1 wt% Triton X-100 solution was added to each well, and the cells were gently shaken at 400 rpm for 5 minutes. The cells were then frozen at -80°C for 1 hour. Cells were thawed at room temperature, and after lysis, they were preheated at 37°C for 10 minutes. After preheating, 100 μL of 0.1 wt% levodopa (PBS) solution was added to each well, and the cells were incubated at 37°C for 2 hours. Cells were then transferred to 96-well plates (100 μL / well, three replicates per well) and the OD was measured using a microplate reader. 490nm value.

[0107]

[0108] 2. Test Results

[0109] The inhibitory effects of different concentrations of Example 1 and Comparative Examples 6-7 on the activity of tyrosinase in B16 cells were shown in the following results. Figure 4 .

[0110] Example 1 showed a concentration-dependent inhibitory effect on tyrosinase activity in B16 cells at concentrations of 3, 10, and 30 μg / mL. The 10 μg / mL composition of Example 1 showed comparable inhibitory effects on tyrosinase activity in B16 cells to the 10 μg / mL comparative example 6-glycyrrhizin. However, comparative example 6-glycyrrhizin at concentrations >10 μg / mL exhibited significant toxicity to B16 cell growth. The 30 μg / mL composition of Example 1 achieved a 97% inhibition rate on tyrosinase activity in B16 cells, significantly superior to the inhibitory effect of peptidylcholine in comparative example 7.

[0111] III. Effects on melanin synthesis in zebrafish models

[0112] 1. Test Methods

[0113] Wild-type AB zebrafish with normal growth and development at 6 days post-flood (dpf) were randomly selected and enclosed in 6-well plates, with 15 zebrafish per well and 3 wells duplicated. Two experimental groups were set up with different concentrations of the composition from Example 1: 1 and 2 μg / mL, with each well containing 3 mL. A normal control group without the composition from Example 1 was also included. After incubation at 28°C in the dark for 45 hours, images were taken under a stereomicroscope, and the number of melanin patches on the zebrafish heads was analyzed using advanced image analysis software.

[0114] 2. Test Results

[0115] like Figure 5 , 6As shown, no significant toxicity was observed in zebrafish growth at concentrations of 1 and 2 μg / mL of the composition of Example 1. The survival rate of zebrafish reached 92% at a concentration of 1 μg / mL and 83% at a concentration of 2 μg / mL. Compared with the control group, the number of melanin patches on the head of zebrafish was significantly reduced at concentrations of 1 and 2 μg / mL of the composition of Example 1, as shown in the figure. Figure 7 In Example 1, when the concentration of the composition was 1 μg / mL, the number of melanin patches in the head of zebrafish decreased by 27%, and when the concentration of the composition was 2 μg / mL, the number of melanin patches in the head decreased by 36%.

[0116] IV. Detection of NO content in RAW264.7 cell supernatant

[0117] 1. Test Methods

[0118] RAW264.7 cells were seeded in 24-well plates (cell concentration adjusted to 7.0 × 10⁶ cells / well). 5 Cells were added at a concentration of 300 μL / mL (approximately 210,000 cells per well) and cultured overnight at 37°C with 5% (v / v) CO2. After 24 h, different concentrations of the composition from Example 1, comparative example 6 (glycyrrhizin), and comparative example 7 (peptide-amidoxuron) solutions were added, 100 μL per well. The blank group received drug-free culture medium. After 1 h, 100 μL of LPS (lipopolysaccharide) dilution buffer (final concentration 1 μg / mL) was added to each well. The normal group received LPS-free culture medium. NaNO2 was prepared to create a standard curve with concentrations of 100 μM, 50 μM, 25 μM, 12.5 μM, 6.25 μM, 3.125 μM, 1.56 μM, and 0 μM, with 75 μL of standard solution added to each well. The supernatant from the 24-well plate was transferred to a 96-well plate, 75 μL per well, with two replicates per group. The remaining samples were stored at -80°C. Add 75 μL of Griess reagent to each well, vortex for 1 min to mix, and incubate at room temperature in the dark for 30 min. After 30 min, detect the NO content using a microplate reader at a wavelength of 548 nm. Calculate the NO content using the formula derived from the standard curve.

[0119] 2. Test Results

[0120] Using an LPS-induced RAW264.7 cell inflammation model, the anti-inflammatory effects of different concentrations of Example 1 and Comparative Examples 6-7 were compared. The results are as follows: Figure 8As shown. Compared with the LPS-induced control group, the addition of 10 and 30 μg / mL of the composition of Example 1 significantly reduced the NO content produced by RAW264.7 cells. 10 μg / mL of the composition of Example 1 reduced NO content by 57%, and 30 μg / mL of the composition of Example 1 reduced NO content by 91%. 10 μg / mL of Comparative Example 6, glycyrrhizin, reduced cellular NO content by 68%, and 3 μg / mL of Comparative Example 7, amidophos, reduced cellular NO content by 58%.

[0121] in conclusion

[0122] This invention discloses an anti-inflammatory and skin-whitening composition. Using B16 mouse melanoma cell models, RAW264.7 cell inflammation models, and zebrafish melanin models, the composition's simultaneous anti-inflammatory and skin-whitening effects were verified at both cellular and animal levels. Compared to glycyrrhizin and peptidoxuridine, this invention has a higher safe usage concentration. In cellular level experiments, the highest safe usage concentration reached 30 μg / mL, while the highest safe usage concentration for glycyrrhizin was 10 μg / mL, and for peptidoxuridine, it was 3 μg / mL. At the highest safe usage concentration, the composition of this invention significantly inhibited tyrosinase activity in B16 cells, melanin synthesis in zebrafish, and NO production in RAW264.7 cells, with effects superior to glycyrrhizin and peptidoxuridine.

[0123] The above description, in conjunction with specific embodiments, further illustrates the present invention. However, these embodiments are merely exemplary and do not constitute any limitation on the scope of the present invention. Those skilled in the art should understand that modifications or substitutions to the details and form of the technical solutions of the present invention can be made without departing from the spirit and scope of the invention, and all such modifications and substitutions fall within the protection scope of the present invention.

Claims

1. An anti-inflammatory and whitening composition, characterized in that, The composition, by weight, is made of 10 parts North American ginseng, 10 parts Eriocaulon buergerianum and 10 parts Atractylodes lancea; or 30 parts North American ginseng, 25 parts Eriocaulon buergerianum and 10 parts Atractylodes lancea; or 10 parts North American ginseng, 10 parts Eriocaulon buergerianum and 20 parts Atractylodes lancea.

2. The composition according to claim 1, characterized in that, The aforementioned North American ginseng is an alcoholic extract of North American ginseng; The aforementioned *Eriocaulon buergerianum* is an alcoholic extract of *Eriocaulon buergerianum The Atractylodes lancea mentioned above is an alcoholic extract of Atractylodes lancea.

3. A method for preparing the composition according to any one of claims 1-2, characterized in that, Includes the following steps: S1. Crush and grind North American ginseng, Eriocaulon buergerianum and Atractylodes lancea respectively to obtain North American ginseng powder, Eriocaulon buergerianum powder and Atractylodes lancea powder; S2. Add ethanol to the powder of North American ginseng, the powder of Eriocaulon buergerianum, or the powder of Atractylodes lancea in step S1, heat and reflux to extract, filter, and obtain North American ginseng extract, Eriocaulon buergerianum extract and Atractylodes lancea extract respectively. S3. The extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea from step S2 are concentrated and dried to obtain alcohol extracts of North American ginseng, Eriocaulon buergerianum, and Atractylodes lancea. S4. Mix the North American ginseng extract, Eriocaulon buergerianum extract and Atractylodes lancea extract obtained in step S3 according to the formula to obtain the composition. The ethanol concentration in step S2 is 70%.

4. The preparation method according to claim 3, characterized in that, The particle size of the North American ginseng powder, Eriocaulon buergerianum powder, and Atractylodes lancea powder mentioned in step S1 is 60-120 mesh.

5. The preparation method according to claim 3, characterized in that, The ratio of the powdered North American ginseng, the powdered Eriocaulon buergerianum, and the powdered Atractylodes lancea to ethanol in step S2 is 1:30-50 g / 100mL. The temperature of the heating and reflux extraction in step S2 is 70-85℃, and the heating and reflux extraction time is 1.5-4h.

6. The preparation method according to claim 3, characterized in that, The drying process described in step S3 is freeze drying.

7. The use of the composition according to any one of claims 1-2 or the composition prepared by the preparation method according to any one of claims 3-6 in the preparation of anti-inflammatory and whitening pharmaceuticals and cosmetics.

8. An anti-inflammatory and whitening product, characterized in that, Made from the composition according to any one of claims 1-2.

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