Chamomile extract based on supramolecular solvents, process for its preparation and use

Abstract

The application relates to the technical field of natural product extraction, in particular to a chamomile extract based on a supramolecular solvent as well as a preparation method and application thereof. The preparation method comprises the following steps: drying chamomile, crushing the chamomile to prepare chamomile powder; adding a supramolecular solvent to the chamomile powder, mixing, ultrasonically treating the obtained mixture at a first temperature to prepare a chamomile extract solution; the composition of the supramolecular solvent comprises a supramolecular acceptor, a supramolecular substrate and water; the supramolecular acceptor comprises an organic acid; the supramolecular substrate comprises at least one of betaine and choline chloride; the first temperature is 40 DEG C-80 DEG C; performing solid-liquid separation on the chamomile extract solution, collecting the liquid to prepare the chamomile extract. The chamomile extract has better anti-inflammatory soothing effect.

Description

Technical Field

[0001] This application relates to the field of natural product extraction technology, and in particular to chamomile extract based on supramolecular solvents, its preparation method and application. Background Technology

[0002] Chamomile, a traditional plant used in both medicine and food, is rich in active ingredients such as flavonoids, volatile oils, and phenolic acids. It possesses anti-inflammatory, antifungal, and antispasmodic properties and is widely used in medicine, food, and cosmetics. Currently, the main methods for extracting active ingredients from chamomile include organic solvent extraction, steam distillation, and ultrasound-assisted extraction. Organic solvent extraction is widely used due to its simplicity and low cost, but it has the following drawbacks: the use of organic solvents such as methanol and ethanol can easily cause environmental pollution and poses a risk of residue; the active ingredients are easily oxidized and degraded during extraction, resulting in a low extraction rate; and solvent recovery is difficult, which is inconsistent with the concept of green development.

[0003] Supramolecular solvents, as novel green extractants, form homogeneous systems through non-covalent self-assembly via hydrogen bonds, van der Waals forces, and other means. They have advantages such as low toxicity, environmental friendliness, recyclability, and good compatibility with active ingredients, and have been gradually applied in the field of natural plant extraction. However, no chamomile extraction method based on supramolecular solvents has yet been discovered.

[0004] Furthermore, traditional extraction methods are usually optimized from the perspective of increasing extraction yield and efficiency, but lack the combined application of research on the efficacy mechanism of chamomile. Summary of the Invention

[0005] Based on this, one or more embodiments of this application provide a chamomile extract based on a supramolecular solvent, a method for its preparation, and its application, wherein the chamomile extract has better anti-inflammatory and soothing effects.

[0006] The technical solution of this application includes the following:

[0007] In a first aspect, this application provides a method for preparing chamomile extract based on supramolecular solvents, comprising the following steps:

[0008] Chamomile is dried and pulverized to obtain chamomile powder;

[0009] A supramolecular solvent is added to the chamomile powder and mixed. The resulting mixture is then sonicated at a first temperature to obtain a chamomile extract. The supramolecular solvent comprises a supramolecular acceptor and a supramolecular substrate. The supramolecular acceptor comprises an organic acid. The supramolecular substrate comprises at least one of betaine and choline chloride. The first temperature is 40°C-80°C.

[0010] The chamomile extract was subjected to solid-liquid separation, and the liquid was collected to prepare the chamomile extract.

[0011] Furthermore, after collecting the liquid, the obtained liquid is concentrated to obtain a concentrated solution;

[0012] The concentrated liquid was freeze-dried to obtain the chamomile extract.

[0013] Furthermore, the method for preparing the supramolecular solvent includes the following steps:

[0014] The supramolecular acceptor and the supramolecular substrate are mixed, and the resulting mixture is stirred at 50℃-80℃ for 2h-12h. Then, the water is added and mixed to obtain the supramolecular solvent.

[0015] Optionally, the water accounts for 20%-40% of the volume in the supramolecular solvent.

[0016] Further, the molar ratio of the supramolecular acceptor to the supramolecular substrate is 1:(0.5-5); and / or,

[0017] The mass-to-volume ratio of the chamomile powder to the supramolecular solvent is 1g:10-50mL.

[0018] In some embodiments, the organic acid includes at least one selected from succinic acid, mandelic acid, salicylic acid, and malic acid.

[0019] In some embodiments, the step of solid-liquid separation of the chamomile extract includes: centrifuging the chamomile extract and then filtering the resulting supernatant; optionally, the centrifugation speed is 8000rpm-10000rpm.

[0020] In some embodiments, the preparation method satisfies at least one of the following conditions:

[0021] (1) The ultrasound time is 20 min-60 min, and the first temperature is 40℃-60℃;

[0022] (2) The freeze-drying temperature is -70℃ to -60℃.

[0023] Secondly, this application provides a chamomile extract based on a supramolecular solvent, which is prepared using the preparation method described above.

[0024] Furthermore, the chamomile extract based on supramolecular solvents has the effect of targeting and inhibiting the secretion and release of IL-33, a key inflammatory mediator; and / or,

[0025] The supramolecular solvent-based chamomile extract includes at least one of the following active ingredients: apigenin, luteolin, quercetin, rutin, and naringenin.

[0026] Thirdly, this application provides the application of the supramolecular solvent-based chamomile extract described above in the preparation of cosmetics with anti-inflammatory and soothing effects.

[0027] Compared with the prior art, the preparation method of this application has at least the following beneficial effects:

[0028] The supramolecular solvents used in the preparation method of this application are all cosmetic-grade raw materials and are recyclable. The extraction process has no volatile organic pollutant emissions and fully complies with cosmetic safety standards. The prepared extract can be directly used in the research and development of skin care products such as anti-inflammatory and soothing masks and moisturizing lotions. It has a wide range of applications and broad industrialization prospects.

[0029] The chamomile extract of this application has excellent anti-inflammatory and soothing effects compared with existing chamomile extracts, which solves the problems of low industrialization efficiency and unclear efficacy caused by the traditional natural product research that "emphasizes extraction and neglects mechanism". Attached Figure Description

[0030] To more clearly illustrate the technical solutions in the specific embodiments of this application or the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of this application. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0031] Figure 1 The figure shows the effect of different extraction conditions on the extraction rate of chamomile polyphenol extract.

[0032] Figure 2 The figure shows the DPPH free radical scavenging rate of betaine-mandelic acid-chamomile extract (BMMS).

[0033] Figure 3 The figure shows the results of ABTS free radical scavenging determination of betaine-mandelic acid-chamomile extract (BMMS).

[0034] Figure 4 The figure shows the effect of different concentrations of BMMS extract on the viability of RAW264.7 macrophages.

[0035] Figure 5 The graph shows the results of the downregulation test of the inflammatory factor IL-6 by BMMS extract.

[0036] Figure 6The graph shows the results of the test on the downregulation rate of the inflammatory factor IL-33 by BMMS extract.

[0037] Figure 7 The Venn diagram for the chamomile active ingredient target and the sensitive skin-related target in Example 1 shows the intersection of the two sets of targets.

[0038] Figure 8 This is a network diagram of protein interactions (PPIs) between chamomile active ingredients and target sites on sensitive skin.

[0039] Figure 9 This is a topology analysis diagram of the PPI network obtained using Cytoscape software, showing the order of the core targets.

[0040] Figure 10 Figure showing the results of KEGG pathway enrichment analysis for the active components of chamomile.

[0041] Figure 11 This is a network diagram illustrating the "component-target" interaction between chamomile active ingredients and core targets in sensitive skin.

[0042] Figure 12 This is a ranking diagram of the node degrees of nine active ingredients in chamomile in the "ingredient-target" network. Detailed Implementation

[0043] The present application is further described below with reference to embodiments and examples. It should be understood that these embodiments are for illustrative purposes only and are not intended to limit the scope of the application. Furthermore, it should be understood that after reading the teachings of this application, those skilled in the art can make various alterations or modifications to this application, and these equivalent forms also fall within the protection scope of the appended claims.

[0044] 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 application belongs. The terminology used herein is for the purpose of describing particular embodiments only and is not intended to be limiting of the application.

[0045] In a first aspect, this application provides a method for preparing chamomile extract based on supramolecular solvents, comprising the following steps:

[0046] Chamomile is dried and pulverized to obtain chamomile powder;

[0047] A supramolecular solvent is added to the chamomile powder and mixed. The resulting mixture is then sonicated at a first temperature to obtain a chamomile extract. The supramolecular solvent comprises a supramolecular acceptor, a supramolecular substrate, and water. The supramolecular acceptor comprises an organic acid. The supramolecular substrate comprises at least one of betaine and choline chloride. The first temperature is 40°C-80°C.

[0048] The chamomile extract was subjected to solid-liquid separation, and the resulting liquid was concentrated to obtain a concentrated solution.

[0049] The concentrated liquid was freeze-dried to obtain the chamomile extract.

[0050] In some preferred embodiments, the first temperature is 50°C.

[0051] Furthermore, the molar ratio of the supramolecular acceptor to the supramolecular substrate is 1:(0.5-5), for example 1:0.5, 1:1, 1:1.5, 1:2, 1:2.5, 1:3, 1:3.5, 1:4, 1:4.5, 1:5, etc.

[0052] In some preferred embodiments, the molar ratio of the supramolecular acceptor to the supramolecular substrate is 1:2.

[0053] Further, the supramolecular acceptor and the supramolecular substrate are mixed, and the resulting mixture is stirred at 50℃-80℃ (e.g., 50℃, 60℃, 70℃, 80℃, etc.) for 2h-12h (e.g., 2h, 4h, 6h, 8h, 10h, 12h, etc.) to form a deep eutectic solvent (DES). The water is then added and mixed to obtain the supramolecular solvent.

[0054] Optionally, the water in the supramolecular solvent accounts for 20%-40% by volume, for example, 20%, 25%, 30%, 35%, 40%, etc.

[0055] Furthermore, the mass-to-volume ratio of the chamomile powder to the supramolecular solvent is 1g:10-50mL, for example, 1g:10mL, 1g:20mL, 1g:30mL, 1g:40mL, 1g:50mL, etc.

[0056] In some embodiments, the organic acid includes at least one selected from succinic acid, mandelic acid, salicylic acid, and malic acid.

[0057] In some embodiments, the step of solid-liquid separation of the chamomile extract includes: centrifuging the chamomile extract and then filtering the resulting supernatant; optionally, the centrifugation speed is 8000rpm-10000rpm;

[0058] In some embodiments, the ultrasound duration is 20-60 minutes, and the first temperature is 40°C-60°C.

[0059] In some embodiments, the solid content of the concentrate is 10%-15%.

[0060] In some embodiments, the freeze-drying temperature is -70°C to -60°C.

[0061] In some preferred embodiments, the supramolecular acceptor is mandelic acid; the supramolecular substrate is betaine, with a volume ratio of 1:2.

[0062] In some preferred embodiments, the mass-to-volume ratio of the chamomile powder to the supramolecular solvent is 1 g: 40 mL.

[0063] In some preferred embodiments, the ultrasonic power is 200W and the duration is 30min.

[0064] Secondly, this application provides a chamomile extract based on a supramolecular solvent, which is prepared using the preparation method described above.

[0065] Furthermore, the chamomile extract based on supramolecular solvents has the effect of targeting and inhibiting the secretion and release of IL-33, a key inflammatory mediator, and can block the activation and cascade amplification of local pro-inflammatory signaling pathways in the skin, thereby effectively downregulating the intensity of the inflammatory response and indirectly improving the skin sensitivity from the pathological mechanism level, demonstrating excellent anti-inflammatory and soothing bioactivity.

[0066] Furthermore, the supramolecular solvent-based chamomile extract includes at least one of the following active ingredients: apigenin, luteolin, quercetin, rutin, and naringenin. These ingredients work synergistically to exhibit excellent anti-inflammatory, antioxidant, and soothing effects. Their corresponding core targets include TNF, AKT1, EGFR, STAT3, and CASP3, and key pathways include the NF-κB signaling pathway, the JAK-STAT signaling pathway, and the PI3K-Akt signaling pathway.

[0067] Furthermore, the dominant active ingredients are apigenin, luteolin, and quercetin.

[0068] In some embodiments, network pharmacology analysis was performed on the above-mentioned supramolecular solvent-based chamomile extract:

[0069] The potential targets of active ingredients are predicted by the SwissTarget Prediction platform, and disease targets related to the target efficacy are retrieved by the GeneCards database. The intersection of the two is taken to obtain the core target.

[0070] We constructed a core target protein-protein interaction (PPI) network using the String database and performed KEGG pathway analysis using the Metascape database to build a "component-target-pathway" network.

[0071] Based on the above analysis results, it was found that the chamomile extract based on supramolecular solvents has excellent anti-inflammatory, antioxidant, and sedative effects.

[0072] Thirdly, this application provides the application of the supramolecular solvent-based chamomile extract described above in the preparation of cosmetics. In some embodiments, chamomile extract is combined with rose extract for the preparation of cosmetics. The combination system with rose fermentation broth can further enhance the anti-inflammatory efficacy, providing valuable application potential and scientific basis for the research and development of anti-inflammatory and soothing skincare products for sensitive skin.

[0073] In some embodiments, the cosmetic has at least one of anti-inflammatory, antioxidant, and soothing effects.

[0074] Cosmetic formulations include, but are not limited to, aqueous products, oil-based products, emulsions, surfactant-solvent-based products, gels, and creams. The following are some specific examples.

[0075] For experimental parameters not specified in the following specific embodiments, please refer to the guidelines given in this application document first, or refer to experimental manuals or other experimental methods known in the art, or refer to the experimental conditions recommended by the manufacturer.

[0076] The raw materials and reagents involved in the following specific embodiments can be obtained commercially or prepared by those skilled in the art using known methods.

[0077] I. Preparation of chamomile extract based on supramolecular solvent.

[0078] Example 1

[0079] This embodiment provides a method for preparing chamomile extract based on supramolecular solvents, comprising the following steps:

[0080] S1. Raw material pretreatment: Dry the chamomile flower heads, crush them, and pass them through a 50-mesh sieve to obtain chamomile powder;

[0081] S2. Extraction: Betaine and mandelic acid were mixed at a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water was added and mixed to obtain a supramolecular solvent. 400 mL of supramolecular solvent (mass-volume ratio 1 g: 40 mL) was added to 10 g of chamomile powder and mixed. The resulting mixture was placed in an ultrasonic extractor for extraction. The extraction temperature was set at 50°C, the extraction power at 200 W, and the extraction time at 30 min to obtain chamomile extract.

[0082] S3. Separation and concentration: The chamomile extract was centrifuged at 10,000 r / min for 15 min, and then the supernatant was vacuum filtered at 0.05 MPa for 4 min. The resulting liquid was concentrated by rotary evaporation to obtain a concentrated solution with a solid content of 5%.

[0083] S4. Vacuum drying: The concentrate is freeze-dried at -60℃ to -70℃ to obtain chamomile extract.

[0084] Example 2

[0085] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: betaine and succinic acid are mixed at a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0086] Example 3

[0087] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: betaine and malic acid are mixed in a molar ratio of 1:2, heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES), and then 30% deionized water is added and mixed.

[0088] Example 4

[0089] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: betaine and salicylic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0090] Example 5

[0091] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: choline chloride and mandelic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0092] Example 6

[0093] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: choline chloride and succinic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0094] Example 7

[0095] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: choline chloride and malic acid are mixed in a molar ratio of 1:2, heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES), and then 30% deionized water is added and mixed.

[0096] Example 8

[0097] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the preparation method of supramolecular solvent is as follows: choline chloride and salicylic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0098] Example 9

[0099] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the supramolecular solvent is prepared as follows: betaine and mandelic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 20% deionized water is then added and mixed.

[0100] Example 10

[0101] This embodiment provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the supramolecular solvent is prepared as follows: betaine and mandelic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 40% deionized water is then added and mixed.

[0102] Example 11

[0103] This embodiment provides a method for preparing chamomile extract based on supramolecular solvents, which is basically the same as the method in Example 1. The difference is that in the step of extracting the obtained mixture in an ultrasonic extractor, the extraction temperature is set to 40°C, the extraction power to 200W, and the extraction time to 60min.

[0104] Example 12

[0105] This embodiment provides a method for preparing chamomile extract based on supramolecular solvents, which is basically the same as the method in Example 1. The difference is that in the step of extracting the obtained mixture in an ultrasonic extractor, the extraction temperature is set to 80°C, the extraction power is 200W, and the extraction time is 20min.

[0106] Comparative Example 1

[0107] This comparative example provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1, except that water is used instead of supramolecular solvent in step S2 and the extraction step.

[0108] Comparative Example 2

[0109] This comparative example provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1, except that ethanol is used instead of supramolecular solvent in step S2 and the extraction step.

[0110] Comparative Example 3

[0111] This comparative example provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the supramolecular solvent is prepared as follows: betaine and lactic acid are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0112] Comparative Example 4

[0113] This comparative example provides a method for preparing chamomile extract based on supramolecular solvent, which adopts a method that is basically the same as that in Example 1. The difference is that the supramolecular solvent is prepared as follows: betaine and 1,3-propanediol are mixed in a molar ratio of 1:2 and heated and stirred at 80°C for 8 hours to form a deep eutectic solvent (DES). 30% deionized water is then added and mixed.

[0114] The polyphenol extraction rate and hyaluronidase were used as screening indicators.

[0115] The extraction rate was determined and calculated as follows:

[0116] Preparation of gallic acid standard solution: Accurately weigh 10 mg of gallic acid analytical standard (AR≥98%), place it in a 50 mL volumetric flask, and dilute to volume with distilled water to prepare a 0.2 mg / mL gallic acid standard solution. Store in a refrigerator at 4℃ protected from light. Preparation of 10% (V / V) Folin-Ciocalteu reagent: Accurately measure 5 mL of Folin-Ciocalteu reagent, dilute to volume with distilled water in a 100 mL volumetric flask, and shake well to obtain a 10% Folin-Ciocalteu reagent solution. Preparation of NaCO3 solution (7.5%): Accurately weigh 7.5 g of NaCO3 solid, dissolve it in distilled water, transfer it to a 100 mL volumetric flask, dilute to volume with distilled water, and shake well to obtain a 7.5% NaCO3 solution. The composition of the above gallic acid standard solutions is shown in Table 1.

[0117] Table 1 Composition of gallic acid standard solution

[0118]

[0119] The absorbance was measured at 765 nm, and three replicates were performed for each group.

[0120] The absorbance of the test solution was determined using the method described above. Three replicates were taken for each group, with the absorbance value measured at 760 nm. The total phenol content in the crude extract was obtained from the standard curve. The absorbance value was substituted into the regression equation to calculate the total phenol content in the sample. The result is expressed as the equivalent grams of gallic acid per gram of sample, in mg / g.

[0121] The standard curve for gallic acid was obtained as follows: Y = 0.009510X + 0.154R 2 =0.998; Y is absorbance, and X is polyphenol extraction yield.

[0122] Table 2 Comparison of extraction rates of chamomile extract with different extraction solvents

[0123]

[0124] The results are shown in Table 2. Betaine-mandelic acid had the highest extraction rate, followed by ethanol. Most polyphenols are moderately or weakly polar molecules; excessively high polarity makes it difficult to effectively dissolve moderately polar polyphenols, resulting in the lowest extraction rates, for example, water's extraction rate was only 8.54%. The betaine-organic acid system forms a deep eutectic solvent (DES), which breaks down plant cell walls through hydrogen bonds and van der Waals forces, releasing polyphenols. The type of acid affects the physicochemical properties of the DES (polarity, viscosity, pH), thus affecting the extraction efficiency. The high extraction rate of mandelic acid is due to its benzene ring structure, which is highly hydrophobic and may generate π-π stacking with the aromatic rings of polyphenols, enhancing solubility. In summary, the extraction scheme of Example 1 improves the extraction efficiency of the target component while meeting the requirements of green and environmentally friendly technology. Differences in the combination of hydrogen bond donors (HBD) and hydrogen bond acceptors (HBA) lead to different types of interaction forces between the deep eutectic solvents (DESs) and the target component during extraction, thus affecting the solubility of the target component and the solvent's ability to dissolve it. Based on the principle of "like dissolves like," the intermolecular forces formed in Example 1 are better suited to the solubility characteristics of polyphenolic compounds, thus exhibiting superior extraction efficiency compared to traditional ethanol solvents. Although the high viscosity of the DES system may limit mass transfer efficiency, mandelic acid, with its strong hydrophobicity, can effectively reduce system viscosity and improve solvent permeability. Furthermore, compared to the toxicity and high volatility of ethanol, DESs offer superior environmental compatibility and safety, making them more suitable for research fields with high requirements for raw material safety, such as cosmetics, food, and pharmaceuticals.

[0125] Example 13

[0126] This embodiment is a response surface methodology for optimizing the preparation method of chamomile extract based on supramolecular solvents.

[0127] Using the test results in Table 2, the yield of chamomile polyphenols was optimized using Design-Expert 8.0.6.1 software. Three factors significantly affecting the yield of chamomile polyphenols were used as influencing factors: ultrasonic extraction time (A), ultrasonic temperature (B), and moisture content (C), as shown in Table 3. A three-factor, three-level response surface methodology was conducted. The Box-Behnken test was used to perform response surface analysis, and the results are shown in Table 4. Figure 1 As shown.

[0128] Table 3 Influence factors of response surface methodology for extraction process optimization

[0129]

[0130] Table 4 Orthogonal experimental design for extraction process optimization

[0131]

[0132] The experimental data were fitted using Design-Expert 8.0.6.1 software through multiple regression analysis, yielding a quadratic polynomial regression equation between the yield of chamomile polyphenol extract (Y) and each experimental factor: Chamomile polyphenol extraction rate = -88.12 + 1.89A + 3.09B + 0.59C + 0.0080AB + 0.0055AC + 0.0061BC - 0.028A 2 -0.043B 2 -0.019C 2 .

[0133] The response surface methodology (RSM) results show that the optimal extraction conditions for chamomile polyphenols using supramolecular solvents are: ultrasonic extraction temperature of 41.76℃, ultrasonic extraction time of 39.58 min, and water content of 33.70%. Under these optimal conditions, the theoretical yield of chamomile polyphenols is 25.13 mg / g. Considering the need for simplicity and convenience in practical operation, the process was modified by comprehensively considering various factors that may affect the process. The optimal conditions are: ultrasonic extraction temperature of 42℃, ultrasonic extraction time of 40 min, and water content of 33%, resulting in a chamomile polyphenol yield of 26.41 mg / g.

[0134] Antioxidant activity assay:

[0135] 1.1 Determination of DPPH free radical scavenging rate.

[0136] a. Prepare a DPPH solution (2.3 mg: 50 mL anhydrous ethanol), store it protected from light, and measure its absorbance at 517 nm. The absorbance should be between 1.2 and 1.3. It is ready for use.

[0137] b. Take a centrifuge tube, add 2 mL of betaine-mandelic acid-chamomile extract, then add 2 mL of DPPH solution, and label it A1.

[0138] c. Take a centrifuge tube, add 2 mL of betaine-mandelic acid-chamomile extract, and then add 2 mL of deionized water solution. This is recorded as A2.

[0139] d. Take a centrifuge tube, add 2 mL of deionized water, and then add 2 mL of DPPH solution. Record this as A3.

[0140] e. Calculate the DPPH free radical scavenging capacity of its betaine-mandelic acid-chamomile extract according to the formula (Ⅰ) below:

[0141] (I);

[0142] In formula (Ⅰ), A1 is the absorbance of DPPH and sample after mixing; A2 is the absorbance of sample solution and deionized water after mixing; A3 is the absorbance of DPPH and deionized water after mixing; and A0 is the absorbance of deionized water (one set of tests is sufficient).

[0143] The results are as follows Figure 2 As shown, using vitamin C (VC) as a positive control, the DPPH free radical scavenging rate (Y-axis, %) of different concentrations of betaine-mandelic acid-chamomile extract (BMMS) (x-axis, mg / mL) was determined. Within the concentration range of 0.1–1.0 mg / mL, the DPPH free radical scavenging rate of the BMMS extract showed a gradient increasing trend with increasing concentration, reaching 97.95% at a concentration of 1.0 mg / mL. Compared with the positive control VC, the DPPH free radical scavenging ability of the BMMS extract was basically the same, indicating that it has a strong free radical scavenging ability and the potential to be developed into a natural antioxidant.

[0144] 1.2 Determination of ABTS free radical scavenging rate.

[0145] a. Mix 5 mL of 7.4 mmol / L ABTS stock solution with 88 μL of 2.6 mmol / L K₂S₂O₈ and let stand for 12–16 hours to prepare the working solution of ABTS. Take 0.4 mL of ABTS solution and dilute with PBS. An absorbance of 734 nm is required at room temperature.

[0146] b. Take a test tube, add 0.5 mL of BMMS extraction solution, then add 2 mL of ABTS solution, and record it as A1.

[0147] c. Take a test tube, add 0.5 mL of BMMS extraction solution, then add 2 mL of PBS solution, and record it as A2.

[0148] d. Take a test tube, add 0.5 mL of PBS solution, then add 2 mL of ABTS solution, and record it as A3.

[0149] e. Calculate the ABTS free radical scavenging capacity of its BMMS solution according to formula (II):

[0150] (II);

[0151] In formula (II), A1 is the absorbance of the sample solution and ABTS; A2 is the absorbance of the sample solution and distilled water; A3 is the absorbance of ABTS without sample solution; and A0 is the absorbance of distilled water.

[0152] The results are as follows Figure 3As shown, using vitamin C (VC) as a positive control, the ABTS free radical scavenging rate (Y-axis, %) of different concentrations of betaine-mandelic acid-chamomile extract (BMMS) (x-axis, mg / mL) was investigated. Within the concentration range of 0.1–1.0 mg / mL, the ABTS free radical scavenging rate of the BMMS extract gradually increased with increasing concentration, reaching a high of 99.86% at a concentration of 1.0 mg / mL. Compared with the positive control VC, the ABTS free radical scavenging ability of the BMMS extract was similar, confirming that the extract possesses good antioxidant activity.

[0153] 1.3 Macrophage RAW264.7 viability assay.

[0154] a. RAW264.7 macrophages were seeded in DMEM medium in 96-well plates and incubated in a CO2 cell culture incubator for 24 hours.

[0155] b. Discard the supernatant in the 96-well plate, serially dilute the BMMS sample with DMEM, add the diluted sample to the 96-well plate, and incubate in a CO2 cell culture incubator for 24 hours.

[0156] c. Discard the supernatant from the 96-well plate and use a CCK-8 assay kit to detect the viability of RAW264.7 macrophages. (OD values ​​of experimental wells are recorded as follows: RAW264.7 macrophages + diluted BMMS sample + CCK-8; OD values ​​of blank wells are recorded as follows: RAW264.7 macrophages + CCK-8; OD values ​​of control wells are recorded as follows: RAW264.7 macrophages + DMEM medium + CCK-8).

[0157] d. Calculate the activity of BMMS solution against RAW264.7 macrophages using the following formula:

[0158] Cell viability = (OD value of experimental wells - OD value of blank wells) / (OD value of control wells - OD value of blank wells) × 100%;

[0159] The results are as follows Figure 4As shown, BMMS extract was administered at different concentrations (x-axis, mg / mL) to determine the viability of RAW264.7 macrophages (y-axis, %). Within the BMMS extract concentration range of 50–100 μg / mL, there was no significant difference in cell viability compared to the concentration without BMMS extract; cell viability remained above 100% and promoted cell proliferation. When the BMMS extract concentration reached 200 μg / mL, a significant difference was observed compared to the concentration without BMMS extract, with cell viability reaching 87.88%. When the BMMS extract concentration reached 400 μg / mL, a significant difference was observed compared to the concentration without BMMS extract, with cell viability reaching 70.50%, indicating an inhibitory effect on cell growth. When the BMMS extract concentration reached 800 μg / mL, a highly significant difference was observed compared to the concentration without BMMS extract, with cell viability reaching 57.00%, indicating a significant inhibitory effect on cell growth. Therefore, 200 μg / mL was selected as the maximum concentration of BMMS extract for a series of cell experiments.

[0160] 1.4 Inflammatory factor (IL)-6 downregulation rate test.

[0161] IL-6 assay: RAW264.7 cell suspension was adjusted to a non-cytotoxic concentration and added to 96-well cell plates. Cells were incubated overnight at 37°C with 5% CO2 for detection. The test samples were diluted with LPS-containing cell culture medium to a non-cytotoxic concentration before addition. LPS-containing medium served as the negative control (NC), and LPS-free cell culture medium served as the blank control (BC). A positive control (PC, dexamethasone sodium phosphate) diluted with LPS-containing cell culture medium was used as the positive control. Cells were cultured for 20–24 hours after addition. After the assay, the supernatant was collected, and the results were analyzed according to the IL-6 ELISA kit procedure.

[0162] Result calculation and judgment:

[0163] The downregulation rate of the inflammatory factor IL-6 was calculated according to formula (III):

[0164] Downward adjustment rate = (1-T / NC) × 100% (Ⅲ);

[0165] In formula (III), T - the average value of inflammatory factor IL-6 in the experimental group; NC - the average value of inflammatory factor IL-6 in the negative control group.

[0166] Perform statistical and significance analysis on the data, calculate the P-value, P < 0.05 indicates a significant difference, otherwise there is no statistical difference.

[0167] When the IL-6 level in the negative control group was significantly higher than that in the blank control group (P < 0.05), the experiment was considered successful.

[0168] When the level of the inflammatory factor IL-6 in the sample group was lower than that in the negative control group, and the difference between the two was statistically significant (P < 0.05), the sample could be considered to have a soothing effect.

[0169] The results are as follows Figure 5 As shown, using dexamethasone sodium phosphate (DSP) as a positive control, its inhibition rate against the key inflammatory mediator IL-6 reached 91.34%. The results showed that BMMS exhibited a significant dose-dependent effect in inhibiting IL-6 release: its anti-inflammatory efficacy continuously increased with increasing BMMS concentration gradient. When the concentration reached 200 μg / mL, the IL-6 inhibition rate was as high as 9.39%, which was highly similar to the anti-inflammatory activity of the positive control DSP. Furthermore, when BMMS was combined with rose fermentation broth (RFL), the synergistic anti-inflammatory effect was significantly enhanced, and the IL-6 inhibition rate further increased to 84.50%, demonstrating even superior anti-inflammatory activity. These results fully demonstrate that BMMS can effectively reduce inflammatory cell infiltration and tissue damage by targeting and inhibiting the expression and release of IL-6, a core effector of the inflammatory pathway, thereby blocking the initiation and amplification of the skin inflammatory cascade from upstream. This improves skin sensitivity from the pathological mechanism level and exhibits excellent anti-inflammatory and soothing bioactivity. Furthermore, its compound system with rose ferment filtrate can further enhance anti-inflammatory efficacy, providing valuable application potential and scientific basis for the research and development of anti-inflammatory and soothing skincare products for sensitive skin.

[0170] 1.5 IL-33 downregulation rate test.

[0171] IL-33 (Interleukin-33), as a potent pro-inflammatory cytokine and important immunomodulatory cytokine in type 2 immune responses, plays a key pro-inflammatory role not only in allergic diseases (such as asthma and atopic dermatitis), autoimmune diseases (such as rheumatoid arthritis), and chronic inflammation, but its association with the pathophysiological process of pruritus has also received widespread attention. Studies have confirmed that its specific receptor (IL-33R) is functionally expressed in dorsal root ganglion (DRG) sensory neurons. IL-33 can activate downstream signaling pathways by directly binding to this receptor, thereby activating sensory neurons and participating in the transmission and amplification of pruritus signals. Based on this, the IL-33 inflammatory factor was measured in the sample to evaluate its inhibitory effect and soothing effect on IL-33-mediated excessive inflammation (such as allergic diseases and chronic inflammation).

[0172] RAW 264.7 cells were used to determine IL-33 levels. Cell suspensions were adjusted to an appropriate concentration and seeded in 96-well plates, incubated overnight at 37°C with 5% CO2. The experiment included a sample group (containing LPS and chamomile extract), a negative control (containing LPS), a positive control (containing tranilast and LPS), and a blank control (without LPS). After 24 hours of treatment, the culture medium was slowly washed with PBS, and repeated freeze-thaw cycles were performed with the addition of an appropriate amount of PBS. The supernatant was then aspirated, centrifuged, and the IL-33 level was detected using a double-antibody sandwich ELISA method: standards / samples, biotinylated detection antibody, HRP-labeled streptavidin, and TMB substrate were added sequentially. After color development, the absorbance at 450 nm was measured. The IL-33 inhibition rate was calculated using the formula: IL-33 inhibition rate = 1 - (sample - blank) / (stimulus - blank) × 100%. All experiments were performed in triplicate.

[0173] The results are as follows Figure 6 As shown, the model validation results indicated that after repeated freeze-thaw stimulation with LPS, the IL-33 secretion level in the cell supernatant remained stable at 117.20 pg / mL, clearly confirming the successful construction of the IL-33-mediated inflammatory cell model. The positive control drug tranilast (TLT) showed an inhibition rate of 92.45% against IL-33, further validating the model's good drug response sensitivity and experimental reliability, laying a solid foundation for subsequent activity evaluation. In vitro experimental data showed that within the safe concentration range, BMMS exhibited a significant dose-dependent effect in inhibiting IL-33 release, with the 200 μg / mL concentration group showing the best inhibitory activity, reaching an IL-33 inhibition rate as high as 82.89%. When BMMS was combined with rose fermentation broth (RFL), the synergistic anti-inflammatory effect was significantly enhanced, further increasing the IL-33 inhibition rate to 85.00%, which is closer to the inhibitory efficacy of the positive control drug. The above results fully demonstrate that BMMS can effectively downregulate the intensity of inflammatory response by targeting and inhibiting the secretion and release of IL-33, a key inflammatory mediator, thereby blocking the activation and cascade amplification of local pro-inflammatory signaling pathways in the skin and indirectly improving the skin's sensitive state from the pathological mechanism level, exhibiting excellent anti-inflammatory and soothing bioactivity. Furthermore, its compound system with rose fermented liquid can further enhance the anti-inflammatory efficacy, providing valuable application potential and scientific basis for the research and development of anti-inflammatory and soothing skincare products for sensitive skin.

[0174] II. Application of Chamomile Extract.

[0175] To further investigate the mechanism of action of chamomile in relieving skin sensitivity, a network pharmacology analysis was conducted.

[0176] 1. Network pharmacology method of active chamomile extract.

[0177] (1) Screening of components and prediction of active targets of chamomile extract.

[0178] Nine relevant potential active ingredients were identified through database and literature searches, as shown in Table 5 below.

[0179] Table 5 Active ingredients of chamomile extract

[0180]

[0181] After obtaining the structural information of the active ingredients in chamomile, their compound structure diagrams were converted into a format supported by the SwissTargetPrediction platform (http: / / swisstargetprediction.ch / ) and uploaded to the platform for predictive analysis of potential targets of the active ingredients. In this database operation, the species parameter was explicitly set to "human" to ensure the correlation between the predicted targets and human physiological and pathological processes, and to avoid interference from non-target species targets in subsequent analyses. The obtained target data were integrated, and duplicate target entries were removed through deduplication, ultimately yielding the core target set of chamomile for its active ingredients, laying the foundation for subsequent target-related analyses.

[0182] (2) Prediction of active ingredients of chamomile and related targets of sensitive skin.

[0183] To screen for target genes related to sensitive skin diseases, the comprehensive human genome database GeneCards (https: / / www.genecards.org / ) was used to search for and screen for disease-related targets. During the search, "Sensitive skin" was used as the core keyword, and the search species was set to "human" to ensure the correlation between the targets and human pathological processes. Ultimately, a database of targets related to sensitive skin diseases was constructed, laying the foundation for subsequent intersection analysis between disease targets and active ingredient targets.

[0184] To screen for common targets of action between chamomile active ingredients and sensitive skin diseases, Venn diagram analysis was conducted using the Venny online analysis platform (http: / / www.liuxiaoyuyuan.cn / ), which visually presents the overlap relationships between different datasets. The visualization results confirmed the intersection targets between chamomile active ingredients and sensitive skin diseases. These intersection targets can be preliminarily identified as potential core targets for chamomile's intervention in sensitive skin, providing key research objects for subsequent functional enrichment analysis and mechanism elucidation.

[0185] (3) Construction of PPI for chamomile active ingredients in sensitive skin, screening of core ingredient targets, and analysis of signaling pathways.

[0186] To analyze the interactions between chamomile active ingredients and overlapping targets in sensitive skin diseases, a protein-protein interaction (PPI) network analysis of these overlapping targets was conducted using the String database (https: / / cn.string-db.org / ), with the species set to "human". The results were then uploaded to Cytoscape 3.9.1 software for data visualization and in-depth analysis.

[0187] To systematically analyze the biological functions and signaling pathway associations of potential targets, the Metascape database (https: / / metascape.org / gp / index.html# / main / step1) was used to conduct Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis on the screened potential targets.

[0188] To visually demonstrate the relationship between chamomile active ingredients and key targets in sensitive skin, the information on key targets in sensitive skin identified in the initial screening was integrated with that of chamomile active ingredients and imported into Cytoscape 3.9.1 software. This generated a visual network diagram of "potential ingredients - key disease targets" for sensitive skin and skin inflammation.

[0189] 2. Analysis of network pharmacology results of chamomile extract.

[0190] (1) Prediction of active targets of chamomile extract and targets of sensitive skin diseases.

[0191] Using the SwissTarget Prediction platform, 270 potential targets for chamomile active ingredients were identified through screening and integration. Simultaneously, 1814 targets related to sensitive skin were retrieved and screened from the GeneCards database. Venn diagram analysis was used to compare the two groups of targets; the comparison results are shown below. Figure 7 The study identified 130 common targets between chamomile active ingredient targets and sensitive skin-related targets. These intersecting targets were then imported into the STRING database, limited to the species "Homo sapiens," and free protein nodes (non-interacting proteins) were removed. The screened protein-protein interaction network data was exported and processed using Cytoscape 3.9.1 software for network topology analysis and visualization. The results are shown below. Figure 8 The software calculates topological parameters such as node degree and betweenness centrality to rank the importance of target nodes in the network. Figure 9This is a ranking diagram of core targets obtained after topological analysis of the PPI network using Cytoscape software. The top-ranked core targets are AKT1, TNF, EGFR, BCL-2, and STAT3. These targets can be preliminarily identified as key targets for the active ingredients of chamomile in regulating sensitive skin.

[0192] (2) Screening of core components and signaling pathway analysis of chamomile active ingredients in sensitive skin.

[0193] Sensitive skin is a complex pathological state involving multiple biological processes such as inflammatory response, cell signal transduction, and metabolic regulation. Its clinical manifestations include both distinct subjective feelings and objective signs: subjectively, it is characterized by a tightness, stinging sensation, transient burning sensation, and persistent itching; objectively, it is accompanied by typical symptoms such as telangiectasia, flushing, erythema, desquamation, and dryness. The occurrence and development of this pathological state are jointly regulated by internal and external factors. It is not only easily aggravated by external stimuli such as temperature fluctuations, ultraviolet radiation, and environmental dust, but can also be induced by the interaction of internal factors such as abnormal activation of inflammatory responses, accumulated psychological stress, and imbalance of the skin microbiota. Furthermore, the synergistic effect of these factors often leads to a vicious cycle of repeated exacerbations of sensitive skin. To clarify the potential mechanism of action of chamomile in soothing sensitive skin, enrichment analysis was performed using the KEGG pathway. The results are as follows: Figure 10 This suggests that chamomile may exert its effects by regulating key signaling pathways through multiple targets. A total of 185 significantly enriched signaling pathways were identified (P<0.05), including core pathways closely related to inflammation regulation and cellular function, such as the PI3K-AKT signaling pathway, NF-κB signaling pathway, and JAK-STAT signaling pathway. This provides important molecular biological evidence for further elucidating the soothing mechanism of chamomile.

[0194] Using Cytoscape 3.9.1 software, a component-target interaction network related to sensitive skin was constructed, vividly demonstrating the complex interactions between various components and their corresponding targets, thus forming a complete network structure. By constructing the compound-target interaction network, the interaction relationships between nine active ingredients and core targets related to sensitive skin were analyzed. The results are shown below. Figure 11 In this visualized network, blue nodes represent active compounds, green nodes label key target proteins, and connecting lines visually demonstrate the specific binding relationships between compounds and targets. Network topology analysis reveals that these phytochemicals may regulate sensitive skin responses through multi-target synergistic effects. The degree of each compound node in the network was calculated using Cytoscape 3.9.1 software. A higher degree indicates a greater number of targets that the compound can act on, and a more crucial regulatory role in the network. Figure 12This is a network diagram showing the "component-target" interaction between chamomile active ingredients and core targets in sensitive skin. Based on the degree ranking, the nine active ingredients are, in order: 7-methoxycoumarin, apigenin, luteolin, quercetin, naringenin, caffeic acid, apigenin-7-O-glucoside, rutin, and coumarin. Among these, 7-methoxycoumarin, apigenin, luteolin, and quercetin have the highest node degrees, indicating they can act on the most sensitive skin-related targets; therefore, they can be identified as the core active ingredients for regulating sensitive skin.

[0195] Based on the above results, it can be concluded that chamomile's ability to alleviate skin sensitivity may be due to polyphenolic compounds.

[0196] The technical features of the above-described embodiments and examples can be combined in any suitable manner. For the sake of brevity, not all possible combinations of the technical features in the above-described embodiments and examples are described. However, as long as there is no contradiction in the combination of these technical features, they should be considered to be within the scope of this specification.

[0197] The embodiments described above merely illustrate several implementation methods of this application and should not be construed as limiting the scope of the patent application. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of this application, and these all fall within the protection scope of this application. Furthermore, it should be understood that after reading the above teachings, those skilled in the art can make various alterations or modifications to this application, and the equivalent forms obtained also fall within the protection scope of this application. It should also be understood that technical solutions obtained by those skilled in the art based on the technical solutions provided in this application through logical analysis, reasoning, or limited experimentation are all within the protection scope of the appended claims. Therefore, the protection scope of this patent application should be determined by the appended claims, and the specification can be used to interpret the content of the claims.

Claims

1. A method for preparing chamomile extract based on supramolecular solvents, characterized in that, Includes the following steps: Chamomile is dried and pulverized to obtain chamomile powder; A chamomile extract is prepared by mixing a supramolecular solvent with chamomile powder and then sonicating the resulting mixture at a first temperature. The supramolecular solvent comprises a supramolecular acceptor, a supramolecular substrate, and water. The supramolecular acceptor includes an organic acid, specifically mandelic acid. The supramolecular substrate is betaine. The mass-to-volume ratio of the chamomile powder to the supramolecular solvent is 1 g:10-50 mL. The molar ratio of the supramolecular acceptor to the supramolecular substrate is 1:(0.5-5). The sonication time is 20-60 min, and the first temperature is 40-60℃. The preparation method of the supramolecular solvent includes the following steps: mixing the supramolecular acceptor and the supramolecular substrate, stirring the resulting mixture at 50-80℃ for 2-12 h, and then adding water to obtain the supramolecular solvent. The volume percentage of water in the supramolecular solvent is 25%-35%. The chamomile extract was subjected to solid-liquid separation, the liquid was collected, and the resulting liquid was concentrated to obtain a concentrated solution. The concentrated liquid was freeze-dried to prepare the chamomile extract; the freeze-drying temperature was -70℃ to -60℃.

2. The preparation method according to claim 1, characterized in that, The water content in the supramolecular solvent is 30% by volume.

3. The preparation method according to any one of claims 1-2, characterized in that, The steps for solid-liquid separation of the chamomile extract include: centrifuging the chamomile extract and filtering the resulting supernatant; wherein the centrifugation speed is 8000rpm-10000rpm.

4. A chamomile extract based on a supramolecular solvent, characterized in that, It is prepared by the preparation method according to any one of claims 1-3.

5. The chamomile extract based on a supramolecular solvent according to claim 4, characterized in that, The chamomile extract based on supramolecular solvents has the effect of targeting and inhibiting the secretion and release of IL-33, a key inflammatory mediator; and / or, The supramolecular solvent-based chamomile extract includes at least one of the following active ingredients: apigenin, luteolin, quercetin, rutin, and naringenin.

6. The use of the supramolecular solvent-based chamomile extract as described in claim 4 or 5 in the preparation of cosmetics with anti-inflammatory and soothing effects.

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