A plant composition with soothing effect and its preparation method and application

Abstract

The application discloses a plant composition with soothing effect and a preparation method and application thereof, and the composition comprises saponins extracted from radix cynanchi rossii and radix anemarrhenae, and polysaccharides extracted from the radix anemarrhenae, and the mass ratio of the saponins to the polysaccharides is (1-5):(1-3). The technical scheme of the application is as follows: the radix cynanchi rossii and the radix anemarrhenae are mixed in a certain proportion, crushed, extracted by an alcohol solution, and then saponins are obtained through composite membrane separation; the remaining residues are extracted by water, impurities are removed by a clarifying agent, and the radix anemarrhena polysaccharides are obtained through ultrafiltration; and the saponins and the polysaccharides are compounded in a certain proportion, supplemented by polyhydric alcohol to be dissolved, so that the composition with the soothing effect is obtained, and the composition can be applied to the preparation of skin care cosmetics, especially soothing and anti-irritation products.

Description

Technical Field

[0001] This invention relates to the field of A61K8 / 9789, and more specifically to a plant composition with soothing effects, its preparation method, and its application. Background Technology

[0002] Sensitive skin is an overreaction of the skin, a sensory reaction to external environmental factors, including cosmetics, that causes the skin to react allergically. It manifests as a stress response to even mild external stimuli, causing discomfort such as stinging, itching, burning, redness, and tightness. In a global consumer survey, 52% of consumers considered themselves to have sensitive skin, making it a leading skin problem. Saponins and polysaccharides found in plants have excellent effects on sensitive skin.

[0003] CN1546151A discloses a method for preparing lyophilized powder for injection of Codonopsis pilosula and Astragalus membranaceus. The method involves precipitating Codonopsis pilosula and Astragalus membranaceus polysaccharides with alcohol. After recovering the ethanol from the supernatant, the total saponins of Codonopsis pilosula and Astragalus membranaceus are obtained by extraction with n-butanol and isoamyl alcohol. However, the organic solvents used in the extraction are highly toxic, posing a safety risk. CN106880654B discloses a method for extracting total saponins from Codonopsis pilosula using an HP-400 macroporous resin column. A certain proportion of ethanol is added to the Codonopsis pilosula powder for soaking, followed by reflux extraction and filtration. After reducing pressure to recover the ethanol, a solution of a certain concentration is prepared. The solution is then eluted sequentially with water and ethanol to obtain an ethanol eluent. The ethanol eluent is then freeze-dried to obtain the total saponins of Codonopsis pilosula. However, the resin packing material recovery cost is high. CN108586560B discloses a one-step aqueous two-phase extraction method for bitter melon saponins and polysaccharides with hypoglycemic activity. The extraction method uses an ultrasonic-microwave assisted aqueous two-phase system. However, the preparation of crude extracts of bitter melon saponins and polysaccharides involves the extensive use of isopropanol, which results in high preparation costs and environmental pollution, making it unsuitable for large-scale production.

[0004] It is evident that existing technologies for extracting saponins and polysaccharides present problems such as the potential for organic solvents to cause harmful effects on operators, environmental pollution, and high costs associated with the recycling of resin fillers. Summary of the Invention

[0005] To address the aforementioned problems, this invention discloses a plant composition with soothing effects, the plant composition comprising saponins extracted from Cynanchum paniculatum and Anemarrhena asphodeloides, and polysaccharides extracted from Anemarrhena asphodeloides.

[0006] Preferably, the mass ratio of saponin to polysaccharide is (1-5):(1-3).

[0007] More preferably, the mass ratio of saponin to polysaccharide is 5:2.

[0008] Bai Wei

[0009] According to the 2020 edition of the Chinese Pharmacopoeia (Part I), *Cynanchum paniculatum* (Bai Wei) is cold in nature and bitter and salty in taste. It possesses various effects such as clearing heat and cooling blood, promoting diuresis and relieving strangury, and detoxifying and healing sores. It can be used to treat diseases such as fever due to febrile diseases, fever due to yin deficiency, bone steaming fever, postpartum blood deficiency fever, strangury, hematuria, carbuncles, and boils. Currently, C21 steroidal saponins, acetophenones, volatile oils, and alkaloids have been identified from *Cynanchum paniculatum*. Among them, C21 steroidal saponins are the main chemical components and pharmacologically active ingredients, possessing numerous pharmacological effects such as antibacterial, anti-inflammatory, antitumor, memory-improving, immunosuppressive, whitening, and liver-damaging effects. Acetophenone derivatives have pharmacological effects including whitening and antioxidant properties. Alkaloids have antibacterial and antiviral pharmacological effects.

[0010] Anemarrhena asphodeloides

[0011] The dried rhizome of Anemarrhena asphodeloides is bitter and sweet in taste, cold in nature, and enters the lung, stomach, and kidney meridians. It has the effects of clearing heat and purging fire, nourishing yin and moistening dryness. Components isolated from Anemarrhena asphodeloides include saponins, bisbenzopyrones, polysaccharides, and alkaloids, among which saponins are the main active ingredients, accounting for approximately 6%. Modern pharmacological studies have shown that Anemarrhena asphodeloides saponins have in vitro anti-inflammatory activity, inhibiting the production of inflammatory factors such as IL-1β, TNF-α, and IL-6 at both the mRNA and protein expression levels; total polysaccharides from Anemarrhena asphodeloides can reduce PGE2 in inflamed tissues by inhibiting the synthesis or release of PGE2.

[0012] Current research on the extraction of effective components from *Cynanchum paniculatum* and *Anemarrhena asphodeloides* mainly focuses on single-type component extraction. Existing saponin extraction methods suffer from drawbacks such as high toxicity of organic solvents, the need for regeneration of macroporous resins, and inconvenient operation. Polysaccharide extraction primarily involves water extraction and alcohol precipitation, which requires a relatively high amount of ethanol. Although ethanol consumption can be reduced through recycling, it still presents problems of high cost and time consumption. Furthermore, various components in traditional Chinese medicine often exhibit synergistic effects, and the synergistic combination of components plays an indispensable role in the overall efficacy. In addition, extracting only single-type components also leads to a waste of medicinal resources. Therefore, this invention utilizes alcohol extraction and composite membrane separation technology for the combined extraction and utilization of saponins and polysaccharides from *Cynanchum paniculatum* and *Anemarrhena asphodeloides*.

[0013] A second aspect of this invention discloses a method for preparing the plant composition, comprising the following steps:

[0014] (1) Raw material crushing;

[0015] (2) Saponin extraction;

[0016] (3) Polysaccharide extraction;

[0017] (4) Compound preparation.

[0018] In one embodiment, step (1) includes: mixing the root of Baiwei and the root of Zhimu, crushing them, and passing them through a 200-mesh sieve.

[0019] Preferably, the mass ratio of the root of *Cynanchum paniculatum* to the root of *Anemarrhena asphodeloides* is 1:(1-3).

[0020] More preferably, the mass ratio of the root of *Cynanchum paniculatum* to *Anemarrhena asphodeloides* is 1:2.

[0021] In one implementation, step (2) includes the following steps:

[0022] A. Extraction;

[0023] B. Filtration and concentration;

[0024] C. Composite membrane separation;

[0025] In one embodiment, step A includes: mixing the pulverized and sieved raw material powder with a certain proportion of alcohol solution, stirring and extracting to obtain liquid 1 and residue.

[0026] Preferably, the alcohol solution is an ethanol solution with a mass concentration of 30%-80%, and the mass ratio of the raw material powder to the alcohol solution is 1:(10-30).

[0027] More preferably, the alcohol solution is a 70% ethanol solution, and the mass ratio of the raw material powder to the alcohol solution is 1:20.

[0028] Preferably, the extraction time in step A is 2-4 hours, the extraction speed is 300-600 rpm, and the extraction temperature is room temperature.

[0029] More preferably, the extraction time in step A is 3 hours, the extraction speed is 500 rpm, and the extraction temperature is room temperature.

[0030] In one embodiment, step B includes: filtering liquid 1 through a membrane and concentrating it under reduced pressure to obtain liquid 2.

[0031] Preferably, the temperature for vacuum concentration in step B is 50-70°C, and the mass ratio for vacuum concentration is (2-5):1.

[0032] More preferably, the vacuum concentration temperature in step B is 60°C, and the vacuum concentration mass ratio is 3:1.

[0033] In one embodiment, step C includes: ultrafiltration of liquid 2 using a #1 ultrafiltration membrane to obtain liquid 3; and nanofiltration of liquid 3 using a #2 nanofiltration membrane to obtain liquid 4, namely, white peony root and anemarrhena saponins.

[0034] Preferably, the molecular weight cutoff of the #1 ultrafiltration membrane is 1000-1200 Da.

[0035] More preferably, the molecular weight cutoff of the #2 nanofiltration membrane is 300-500 DaL.

[0036] Flavonoids are plant secondary metabolites with free radical scavenging and antioxidant capabilities. Alkaloids in traditional Chinese medicine are an important class of natural organic compounds with antibacterial and anti-inflammatory effects. As those skilled in the art know, combining some functional components can make the composition possess the effects of various substances. However, the applicant found in experiments that if only one ultrafiltration is performed, the solution 3 contains not only saponins but also flavonoids and alkaloids. After nanofiltration, saponins can be separated from flavonoids and alkaloids to obtain purer saponins. The effect of combining solution 3 with polysaccharides is worse than that of combining solution 4 with polysaccharides. The possible reason is that the three functional components of saponins, flavonoids and alkaloids compete with each other. For people with sensitive skin, the efficacy will not be enhanced by the combination of functional components. However, due to the complexity of the components, the skin's tolerance is poor and the efficacy cannot be achieved. Furthermore, nanofiltration not only retains flavonoids and alkaloids but also retains many small molecule impurities, thus ensuring good results and no side effects for people with sensitive skin.

[0037] In one implementation, step (3) includes the following steps:

[0038] D. Polysaccharide extraction;

[0039] E. Clarifying agent for impurity removal;

[0040] F. Polysaccharide isolation.

[0041] In one embodiment, step D includes: mixing the residue from step A with a certain proportion of deionized water, heating and stirring to extract liquid 5.

[0042] Preferably, the mass ratio of the medicinal residue to deionized water is 1:(8-15).

[0043] More preferably, the mass ratio of the medicinal residue to deionized water is 1:10.

[0044] Preferably, the extraction time in step D is 1-3 hours, and the extraction temperature is 70-90℃.

[0045] More preferably, the extraction time in step D is 2 hours and the extraction temperature is 80°C.

[0046] In one embodiment, step E includes: adjusting the pH of feed solution 5 to 5-5.5, using chitosan clarifying agent to adsorb and precipitate impurities, and then filtering through a membrane to obtain feed solution 6; adjusting the pH of feed solution 6 to ≥ 6.5 to precipitate chitosan clarifying agent, and then filtering through a membrane to obtain feed solution 7.

[0047] Preferably, in step E, the concentration of the chitosan clarifying agent is 0.5%-2%.

[0048] More preferably, the concentration of the chitosan clarifying agent is 1%.

[0049] Preferably, the amount of chitosan clarifying agent used is 0.02%-0.12% of the mass of the liquid 5.

[0050] More preferably, the amount of chitosan clarifying agent used is 0.06% of the mass of liquid 5.

[0051] In one embodiment, step F includes: using a 18,000-20,000 molecular weight ultrafiltration membrane to separate polysaccharides from feed solution 7, resulting in feed solution 8, i.e., Anemarrhena polysaccharide.

[0052] In one embodiment, step (4) includes: mixing liquid 4 and liquid 8 in a certain proportion and dissolving them with a polyol to obtain a composition with soothing effects.

[0053] The applicant discovered that using a combination of saponins and polysaccharides in a controlled mass ratio of (1-5):(1-3) resulted in a stronger inhibitory effect on the expression of inflammatory factors NO and PGE2. Experiments showed that the inhibition rates were 73.72% and 77.65% when the saponin-polysaccharide mass ratio was 1:1 and 3:1, respectively. The highest inhibition rate, reaching 92.72%, was observed when the ratio was 5:2. In contrast, the inhibition rates were only 24.02% and 12.08% when saponins and polysaccharides were used alone. This is because saponins have the effect of scavenging oxygen free radicals, maintaining a dynamic balance in intracellular redox reactions, while polysaccharides have in vitro antioxidant effects and protect erythrocytes. The two components have a synergistic effect, and this synergistic combination enhances the overall efficacy, thereby improving the product's soothing effect.

[0054] Preferably, the polyol is selected from polyols that comply with regulations for use in cosmetics, such as propylene glycol, butylene glycol, and dipropylene glycol.

[0055] More preferably, the polyol is 50 wt% butanediol.

[0056] This invention utilizes alcohol extraction and composite membrane separation technology to obtain saponins from *Cynanchum paniculatum* and *Anemarrhena asphodeloides*, solving problems such as the harmful effects of organic solvents on operators, environmental pollution, and high costs of resin and filler recovery. During alcohol extraction, a 30%-80% ethanol solution is used, controlling the mass ratio of raw material powder to alcohol solution to 1:(10-30). In particular, using a 70% ethanol solution with a mass ratio of 1:20 ensures that *Cynanchum paniculatum* and *Anemarrhena asphodeloides* saponins dissolve in the alcohol solution without dissolving the polysaccharide components, leaving the polysaccharide components in the residue for subsequent water extraction.

[0057] In the extraction process of Anemarrhena asphodeloides polysaccharides, the principle that chitosan clarifiers better exert their adsorption and clarification effects under acidic conditions is utilized. Impurities such as proteins in the aqueous extract solution are removed by adjusting the pH; then, the pH is adjusted to the isoelectric point of chitosan to remove chitosan from the solution. No exogenous substances are introduced during the impurity removal process, reducing potential instability factors caused by exogenous substances. This method is environmentally friendly, low-cost, and results in high product stability.

[0058] The soothing composition of the present invention is used in skin care cosmetics, including but not limited to creams, lotions, lotions, gels, masks, and ointments.

[0059] Beneficial effects:

[0060] 1. This invention uses alcohol extraction and composite membrane separation technology to obtain saponins from Baiwei and Zhimu, which solves the problems of organic solvents causing harmful effects on operators, environmental pollution, and high cost of resin filler recycling.

[0061] 2. This invention extracts polysaccharides from medicinal residues, and combines white peony root and anemarrhena saponins with anemarrhena polysaccharides in a certain proportion. The resulting plant composition has the effect of relieving the production of inflammatory factors induced by adverse external factors and repairing the skin barrier. It can be added to cosmetics as a soothing ingredient.

[0062] 3. The extraction and separation method disclosed in this invention does not involve organic solvents, resin fillers, etc., and has the characteristics of being green, safe and environmentally friendly.

[0063] 4. The plant composition obtained in this invention can inhibit the expression of inflammatory factors NO and PGE2, reduce transdermal water loss and erythema value increase caused by external stimuli, and alleviate the stimulation of sensitive skin by adverse external factors. Attached Figure Description

[0064] Figure 1 The figure shows the test results of the plant composition prepared in this invention inhibiting NO release in RAW264.7 cells.

[0065] Figure 2The figure shows the test results of the plant composition prepared in this invention inhibiting PGE2 release in RAW264.7 cells.

[0066] Figure 3 The graph shows the ΔTEWL (transcutaneous water loss change) test results of the plant composition prepared in this invention in order to alleviate capsaicin irritation.

[0067] Figure 4 The graph shows the ΔErythema (change in erythema value) test results of the plant composition prepared in this invention in alleviating capsaicin stimulation. Detailed Implementation

[0068] Example 1

[0069] This embodiment 1 discloses a plant composition with soothing effects. The plant composition is saponins extracted from Cynanchum paniculatum and Anemarrhena asphodeloides, and polysaccharides extracted from Anemarrhena asphodeloides, wherein the mass ratio of saponins to polysaccharides is 5:2.

[0070] This embodiment 1 discloses another aspect of the preparation method of the plant composition, the steps of which are as follows:

[0071] (1) Raw material crushing: Mix the roots of Baiwei root and Zhimu root in a mass ratio of 1:2, crush them, and pass them through a 200-mesh sieve;

[0072] (2) Extraction of saponins from *Cynanchum paniculatum* and *Anemarrhena asphodeloides*:

[0073] A. Extraction: The pulverized and sieved raw material is mixed with 70wt% alcohol solution at a mass ratio of 1:20 and stirred at 500rpm for 3h at room temperature to obtain liquid 1 and residue;

[0074] B. Filtration and concentration: Filter liquid 1 through a membrane and concentrate under reduced pressure at a temperature of 60℃. The mass ratio of the concentrated liquid under reduced pressure is 3:1, thus obtaining liquid 2.

[0075] C. Composite membrane separation: Saponins, flavonoids and alkaloids are separated from feed liquid 2 using a 1000-molecular-weight ultrafiltration membrane, which is feed liquid 3; the saponins, flavonoids and alkaloids of feed liquid 3 are separated from feed liquid 3 using a 500-molecular-weight nanofiltration membrane, which is feed liquid 4, namely the saponins of white peony and anemarrhena.

[0076] (3) Polysaccharide extraction:

[0077] D. Extraction: Mix the residue with deionized water at a mass ratio of 1:10, and extract at 80℃ with stirring for 2 hours to obtain 5 liters of liquid.

[0078] E. Impurity removal: Adjust the pH of feed solution 5 to 5.3±0.1, use chitosan clarifying agent to adsorb and precipitate impurities, and filter through a membrane to obtain feed solution 6; adjust the pH of feed solution 6 to 6.7±0.1, precipitate chitosan clarifying agent, and filter through a membrane to obtain feed solution 7.

[0079] The concentration of the chitosan clarifying agent is 1%.

[0080] The amount of chitosan clarifying agent used is 0.06% of the mass of liquid 5.

[0081] F. Polysaccharide separation: Polysaccharides were separated from feed solution 7 using a 20,000 molecular weight ultrafiltration membrane to obtain feed solution 8;

[0082] (4) Compounding: Mix liquid 4 and liquid 8 in a mass ratio of 5:2 and dissolve them with 50wt% butanediol to obtain a composition with soothing effect.

[0083] Example 2

[0084] Liquid 4 and liquid 8 are mixed in a 1:1 ratio, and the rest is the same as in Example 1.

[0085] Example 3

[0086] Liquid 4 and liquid 8 are mixed in a 3:1 ratio, and the rest is the same as in Example 1.

[0087] Comparative Example 1

[0088] The extraction steps do not include the polysaccharide extraction part; the components only include Cynanchum paniculatum and Anemarrhena asphodeloides, and the rest is the same as in Example 1.

[0089] Comparative Example 2

[0090] The extraction steps do not include the extraction of white peony root and anemarrhena saponins; the components only include anemarrhena polysaccharide, and the rest are the same as in Example 1.

[0091] Performance testing:

[0092] 1. Detection of NO and PGE2 secretion in RAW264.7 cell supernatant

[0093] Prior to testing, cell viability was determined using the MTT assay to establish safe concentrations of the extracts from each example and comparative example on RAW264.7 cells.

[0094] RAW264.7 cells were cultured in DMEM medium (containing 10% FBS and 1% penicillin antibiotics) at 37°C and 5% CO2 using standard methods. Cells in the logarithmic growth phase were collected and a cell suspension was prepared. Cells were then cultured at 3 × 10⁻⁶ cells / mL. 5 Cells were seeded at a density of 500 μL per 24-well plate. After 24 h of cell culture, positive control drug (dexamethasone, Dex), 0.1% of the example, 0.1% of the comparative example, and LPS were added and co-treated for 24 h. The supernatant was collected and centrifuged at 12000 rpm for 3 min at 4 °C. The NO and PGE2 secretion levels in the culture supernatant were measured according to the Griess method or the instructions of the enzyme-linked immunosorbent assay (ELISA) kit.

[0095] Experimental results:

[0096] Depend on Figure 1 It can be seen that, compared with the blank control group, the NO content in the LPS group (model group) increased significantly, indicating that the stimulation conditions in this experiment were effective.

[0097] Compared with the LPS group, the NO content in the positive control group decreased significantly, indicating that the positive control test was effective. The NO content in Examples 1-3 decreased significantly, with inhibition rates of 92.72%, 73.72%, and 77.65%, respectively. However, the NO content in Comparative Examples 1-2 did not decrease significantly, with inhibition rates of only 24.02% and 12.08%, respectively.

[0098] Test results show that the extracts from the examples effectively inhibited the increase in LPS-induced NO production, and the overall effect was better than that of the comparative examples.

[0099] Depend on Figure 2 It can be seen that, compared with the blank control group, the PGE2 content in the LPS group (model group) increased significantly, indicating that the stimulation conditions in this experiment were effective.

[0100] Compared with the LPS group, the PGE2 content in the positive control group decreased significantly, indicating that the positive control test was effective. The PGE2 content in Examples 1-3 decreased significantly, with inhibition rates of 59.70%, 43.46%, and 65.79%, respectively. However, the PGE2 content in Comparative Examples 1-2 did not decrease significantly, with inhibition rates of only 11.60% and 1.39%, respectively.

[0101] Test results show that the extracts from the examples effectively inhibited the increase in LPS-induced PGE2 production, and the overall effect was better than that of the comparative examples.

[0102] 2. Irritant Clinical Repair Test

[0103] Fifteen healthy volunteers, aged 25-45 years, were selected. A fixed area was selected on the relatively delicate skin of the flexor side of the forearm of the subjects. The test sites were treated with control group (blank), model group (0.075% capsaicin, CAP), and sample group (2% of the example or comparative example + 0.075% capsaicin) for 30 minutes. TEWL and erythema values ​​were measured to evaluate the repair status.

[0104] Experimental results:

[0105] Depend on Figure 3 , 4 It is understood that the plant extract composition of the embodiment can effectively alleviate the increase in transdermal water loss (TEWL) and erythema (ERYTHEMA) caused by capsaicin stimulation, while the comparative composition has limited alleviating effect on TEWL and ERYTHEMA.

Claims

1. A method for preparing a plant composition having soothing efficacy, characterized in that, The plant composition comprises saponins extracted from Cynanchum paniculatum and Anemarrhena asphodeloides, and polysaccharides extracted from Anemarrhena asphodeloides, wherein the mass ratio of saponins to polysaccharides is 5:

2. The preparation method of the plant composition is as follows: (1) Raw material crushing: Mix the roots of Baiwei root and Zhimu root in a mass ratio of 1:2, crush them, and pass them through a 200-mesh sieve; (2) Extraction of saponins from *Cynanchum paniculatum* and *Anemarrhena asphodeloides A. Extraction: The pulverized and sieved raw material is mixed with 70wt% alcohol solution at a mass ratio of 1:20 and stirred at 500rpm for 3h at room temperature to obtain liquid 1 and residue; B. Filtration and concentration: Filter liquid 1 through a membrane and concentrate under reduced pressure at a temperature of 60℃. The mass ratio of the concentrated liquid under reduced pressure is 3:1, thus obtaining liquid 2. C. Composite membrane separation: Saponins, flavonoids and alkaloids are separated from feed solution 2 by ultrafiltration using an ultrafiltration membrane with a molecular weight cutoff of 1000 Da, which is feed solution 3; feed solution 3 is separated from saponins, flavonoids and alkaloids by nanofiltration using a nanofiltration membrane with a molecular weight cutoff of 500 Da, to obtain feed solution 4, which is saponins of white peony and anemarrhena. (3) Polysaccharide extraction: D. Extraction: Mix the residue with deionized water at a mass ratio of 1:10, and extract at 80℃ with stirring for 2 hours to obtain 5 liters of liquid. E. Impurity removal: Adjust the pH of feed solution 5 to 5.3±0.1, and use chitosan clarifying agent to adsorb and precipitate impurities, then filter through a membrane to obtain feed solution 6; adjust the pH of feed solution 6 to 6.7±0.1 to precipitate chitosan clarifying agent, then filter through a membrane to obtain feed solution 7; the concentration of the chitosan clarifying agent is 1%; The amount of chitosan clarifying agent used is 0.06% of the mass of liquid 5; F. Polysaccharide separation: Polysaccharides were separated from feed solution 7 using an ultrafiltration membrane with a molecular weight cutoff of 20,000 Dal, resulting in feed solution 8; (4) Compounding: Mix liquid 4 and liquid 8 in a mass ratio of 5:2 and dissolve them with 50wt% butanediol to obtain a composition with soothing effect.

2. The application of a plant composition obtained by the preparation method of the plant composition with soothing effect according to claim 1, for use in the preparation of skin care cosmetics.

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