Abelmoschus manihot polysaccharide skin cream and preparation method thereof

Abstract

The invention discloses abelmoschus manihot polysaccharide skin cream and a preparation method thereof, and belongs to the technical field of skin care products. The abelmoschus manihot polysaccharide skin cream comprises the following components in parts by mass: 60-85 parts of water, 3-15 parts of a humectant, 5-15 parts of an emollient, 6-10 parts of an emulsifier, 0.1-0.5 part of a preservative and 1-3 parts of an abelmoschus manihot polysaccharide extract, and the pH of a system is adjusted to 4.5-6.5 by utilizing a pH regulator. On the basis of the water-based skin cream, the active ingredient hibiscus manihot polysaccharide extract is added, the hibiscus manihot polysaccharide extract has the characteristics of inflammation resistance, oxidation resistance, allergy resistance, viscoelasticity, film-forming property and the like, and the addition amount is controlled, so that the prepared hibiscus manihot polysaccharide skin cream not only retains the characteristics of refreshing skin feeling and quick absorption of the water-based skin cream, but also can be used for preparing the skin-care cream. The long-acting water locking and barrier repairing functions are also realized.

Description

Technical Field

[0001] The invention belongs to the technical field of skin care products, and particularly relates to a sunflower polysaccharide skin care cream and a preparation method thereof. Background Art

[0002] Skin creams, as common topical preparations, are usually divided into two categories: water-based systems and oil-based systems based on the characteristics of the matrix.

[0003] 1. Water-based skin care cream (O / W emulsion)

[0004] With water as the continuous phase and oil phase dispersed in it, its core advantages are:

[0005] (1) Light feel on the skin: easy to spread, quickly absorbed, no sticky feeling after use, suitable for oily skin or humid and hot environment;

[0006] (2) Compatible with hydrophilic active ingredients: such as vitamin C derivatives, sodium hyaluronate, etc., which can enhance moisturizing and antioxidant effects;

[0007] (3) Good breathability: reduces the risk of pore clogging.

[0008] Its inherent defects include:

[0009] (1) Limited moisturizing ability: Water is easily volatile and the ability to lock moisture in for a long time is insufficient, especially in dry environments where the moisturizing durability is significantly reduced;

[0010] (2) Weak barrier repair: Lack of sufficient lipid components makes it difficult to effectively repair the skin barrier, and has limited effect on dry / damaged skin;

[0011] (3) Stability challenge: It relies on emulsifiers / preservatives to maintain system stability, which may increase the complexity of the formula and the risk of irritation.

[0012] 2. Oily skin cream (W / O type or pure ointment)

[0013] With oil as the continuous phase, its notable features are:

[0014] (1) Strong water-locking and sealing: The oil film can significantly reduce transepidermal water loss (TEWL), and is suitable for extremely dry environments or skin with damaged skin barriers;

[0015] (2) High permeability: promotes the delivery of fat-soluble active ingredients (such as retinol and vitamin E);

[0016] (3) High formula stability: not easy to breed microorganisms, and low demand for preservatives.

[0017] But its main disadvantages are:

[0018] (1) Thick skin feel: poor ductility, easy to produce greasy feeling, may cause acne or discomfort, and poor user experience;

[0019] (2) Limited application scenarios: Oily / combination skin may become more shiny after use, and its applicability is low in hot climates;

[0020] (3) Narrow ingredient compatibility: It is difficult to effectively carry high concentrations of hydrophilic active ingredients.

[0021] Relatively speaking, water-based skin care creams are more popular among consumers because of their easy spreading, rapid absorption, and non-sticky feeling after use. However, their inherent defects limit their effectiveness to a certain extent. How to overcome their defects as much as possible while retaining the advantages of water-based skin care creams, so that water-based skin care creams can retain the refreshing skin feeling and fast absorption characteristics while also having long-term water-locking and barrier repair functions, is the current research direction of technical personnel in this field. Summary of the Invention

[0022] The purpose of the present invention is to provide a sunflower polysaccharide skin care cream and a preparation method thereof. By adding an active ingredient, sunflower polysaccharide extract, to a water-based skin care cream as a basis, and utilizing the anti-inflammatory, antioxidant, anti-allergic, viscoelastic and film-forming properties of the sunflower polysaccharide extract, and by controlling the amount of addition, the sunflower polysaccharide skin care cream prepared not only retains the refreshing skin feel and fast absorption properties of the water-based skin care cream, but also has long-lasting water-locking and barrier repair functions.

[0023] To achieve the above object, the present invention provides the following technical solutions:

[0024] One of the technical solutions of the present invention is to provide a sunflower polysaccharide skin care cream, which comprises, by weight, 60 to 85 parts of water, 3 to 15 parts of moisturizer, 5 to 15 parts of emollient, 6 to 10 parts of emulsifier, 0.1 to 0.5 parts of preservative and 1 to 3 parts of sunflower polysaccharide extract, and the pH of the system is adjusted to 4.5 to 6.5 using a pH regulator.

[0025] The anti-inflammatory effect of the Helianthus annuus polysaccharide extract in the present invention can reduce the inflammatory response of the skin, soothe the secondary damage to the skin caused by external stimuli, and help repair the skin barrier; its antioxidant effect can resist the oxidative damage to the skin caused by free radicals, delay aging and improve the skin condition; the small amount of flavonoid components contained in its extract also contributes to anti-inflammatory, anti-oxidation and anti-allergic effects; in addition, the viscoelasticity and film-forming properties of Helianthus annuus polysaccharide can replace conventional thickeners, while improving the smearing feel, increasing the viscosity and stability of the product, preventing stratification, and can effectively reduce transepidermal water loss, and enhance the water-locking function of the product. In addition, skin care products containing thickeners may cause problems such as acne and blackheads on the skin. The thickener forms a thick layer of "glue" on the skin surface, which not only affects the makeup effect, but also may increase the burden on the skin, may clog pores, destroy the skin's natural barrier, accelerate skin aging, and cause skin sensitivity and other problems. The Helianthus annuus polysaccharide extract as a thickener substitute in the present invention does not have the above problems. The macromolecular Helianthus annuus polysaccharide extract can lock in skin moisture while maintaining good air permeability, and is an excellent component for replacing thickeners.

[0026] Preferably, the moisturizing agent comprises one or more of glycerin, sodium hyaluronate, propylene glycol, butylene glycol, amino acid moisturizing agent, sorbitol and trehalose.

[0027] Optionally, the amino acid moisturizing agent includes one or more of arginine, serine and glycine.

[0028] Preferably, the emollient comprises one or more of light synthetic esters, silicone oil derivatives, natural oils and fatty alcohols.

[0029] Optionally, the light synthetic ester includes one or more of caprylic triglyceride, capric triglyceride, ethylhexyl palmitate and dicaprylyl carbonate; the silicone oil derivative includes one or more of cyclopentasiloxane, polydimethylsiloxane and cyclohexasiloxane; the natural oil includes one or more of squalane (plant source), jojoba oil and almond oil; the fatty alcohol includes one or more of cetyl alcohol, stearyl alcohol and cetearyl alcohol.

[0030] Preferably, the emulsifier includes one or more of polysorbate emulsifiers, lecithin, glyceryl stearate and acrylic emulsifiers.

[0031] Optionally, the polysorbate emulsifier includes one or more of polysorbate-20, polysorbate-60 and polysorbate-80; and the acrylic emulsifier includes one or more of sodium lauryl sulfate, sodium lauryl sulfonate and sodium dodecylbenzene sulfonate.

[0032] Preferably, the preservative comprises one or more of phenoxyethanol, sodium benzoate, potassium sorbate, a combination of p-hydroxyacetophenone and caprylhydroxamic acid, a polyol preservative, an isothiazolinone preservative and a paraben preservative.

[0033] Optionally, the polyol preservative includes pentylene glycol and / or hexylene glycol; the isothiazolinone preservative includes methylisothiazolinone and / or methylchloroisothiazolinone; and the paraben preservative includes methylparaben and / or propylparaben.

[0034] Preferably, the extracting solution used when extracting the Helianthus annuus polysaccharide extract is a weakly acidic aqueous solution with a pH of 4.5 to 6.5.

[0035] The main reason why the present invention adopts an aqueous solution with a specific pH value as the extracting solution is that the pH value is in the acidic range of the skin surface. The Helianthus annuus polysaccharide extract obtained by extracting with the extracting solution in this pH range has better anti-inflammatory and antioxidant effects.

[0036] Preferably, the pH adjuster includes: citric acid, lactic acid, triethanolamine, sodium hydroxide or potassium hydroxide.

[0037] The second technical solution of the present invention is to provide a method for preparing the above-mentioned sunflower polysaccharide skin care cream, comprising the following steps:

[0038] Moisturizer, emollient and sunflower polysaccharide extract are added to water accounting for 50 to 70% of the total water usage, mixed evenly and homogenized once, then an emulsifier is added, mixed evenly and homogenized twice, then a preservative and the remaining water are added, mixed evenly and the pH value is adjusted to 4.5 to 6.5 using a pH regulator, and homogenized three times to obtain the sunflower polysaccharide skin care cream.

[0039] Preferably, the conditions for the first homogenization, the second homogenization and the third homogenization are all 2500-3500 rpm for 5-10 min.

[0040] The beneficial technical effects of the present invention are as follows:

[0041] The present invention takes water-based skin care cream as a basis, adds the active ingredient Helianthus annuus polysaccharide extract, utilizes the anti-inflammatory, antioxidant, anti-allergic, viscoelastic and film-forming properties of the Helianthus annuus polysaccharide extract, and controls the added amount so that the prepared Helianthus annuus polysaccharide skin care cream not only retains the refreshing skin feel and fast absorption properties of the water-based skin care cream, but also has long-lasting water-locking and barrier repair functions. DETAILED DESCRIPTION

[0042] Hibiscus manihot L., known as the "Giant Panda of the Plant World," is a gem of the Malvaceae family. Not only are its flowers vibrant, but its polysaccharides, found in its roots, stems, leaves, and flowers, are a focus of ongoing exploration in modern life sciences. Hibiscus manihot L. polysaccharides (HMLP), one of its core active ingredients, demonstrates remarkable potential for applications in health maintenance, disease prevention, and even cutting-edge biomaterials, thanks to its complex structural characteristics and wide-ranging physiological activities.

[0043] Sunflower is an annual herb with tall, upright stems, deeply palmate leaves, large, bright yellow flowers with a deep red center, and a unique shape. Its capsule is tower-shaped and its seeds are kidney-shaped.

[0044] Sunflower is not a new discovery of modern science. In traditional Chinese medicine, its roots, leaves, flowers, and seeds are all used as medicine. TCM believes it has a sweet and cold nature, with benefits such as clearing heat and detoxifying, promoting diuresis and relieving stranguria, cooling blood and stopping bleeding, and moistening and lubricating the intestines. It is commonly used to treat sore throats, dysentery, stranguria, sores, swelling, constipation, and bleeding from trauma. Its young leaves, pods, and petals are also edible as a nutritious vegetable, making it a classic example of a plant with both medicinal and edible properties. Modern research is further exploring the material basis for its traditional benefits.

[0045] Heliotropin polysaccharides are a type of high molecular weight heteropolysaccharide mixture extracted from different parts of Heliotropinus chinensis (especially flowers and leaves). Its chemical nature and properties are the basis for understanding its biological activity.

[0046] The mainstream extraction method remains hot water extraction, which exploits the water solubility of polysaccharides. To improve efficiency, it is often supplemented with ultrasound (using the cavitation effect to disrupt cells), microwaves (using rapid molecular motion to generate heat), or enzymatic hydrolysis (using enzymes such as cellulase and pectinase to selectively destroy cell walls). This is then followed by impurity removal and purification to obtain relatively pure polysaccharide fractions. Ethanol graded precipitation is the most commonly used method, utilizing the differences in solubility of polysaccharides of different molecular weights or structures at different ethanol concentrations to separate them.

[0047] Sunflower polysaccharide has multidimensional biological activity:

[0048] (1) “Intelligent Regulator” of the Immune System:

[0049] Activating Immune Cells: HMLP can significantly activate macrophages, enhancing their ability to phagocytose foreign matter (phagocytic activity) and promoting their secretion of a variety of key immune signaling molecules (cytokines), such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and interferons (IFNs) with antiviral activity. These cytokines form a complex signaling network that recruits and activates more immune cells (such as neutrophils and natural killer (NK) cells), initiating inflammatory defense responses. HMLP also promotes the proliferation and differentiation of T and B lymphocytes, enhancing both cellular immunity (such as killing infected cells) and humoral immunity (producing more antibodies).

[0050] (2) “Free radical scavenger” on the anti-oxidation front:

[0051] Neutralize Reactive Oxygen Species (ROS): Human metabolism and environmental stress continuously generate free radicals (ROS). Excessive ROS attack lipids, proteins, and DNA, leading to cell damage, aging, and various chronic diseases (arteriosclerosis, neurodegenerative diseases, cancer, etc.). HMLP can effectively remove superoxide anions (O 2·- ), hydroxyl radicals (·OH), hydrogen peroxide (H2O2) and other ROS.

[0052] (3) Potential interveners of glucose metabolism disorders:

[0053] Improving insulin resistance and lowering blood sugar: Animal models and some cell experiments have shown that HMLP can improve insulin sensitivity, reduce insulin resistance, and thus help lower blood sugar levels.

[0054] (4) “Ecological guardian” and “physical barrier worker” of intestinal health:

[0055] Prebiotic Effect: As a type of dietary fiber, HMLP is not easily broken down by enzymes in the upper digestive tract and reaches the colon intact. There, it becomes the exclusive "food" (prebiotic) for beneficial intestinal bacteria (such as Bifidobacterium and Lactobacillus). These beneficial bacteria ferment HMLP to produce short-chain fatty acids (SCFAs, such as acetate, propionate, and butyrate). SCFAs are the primary energy source for colonocytes and can lower intestinal pH (inhibiting the growth of harmful bacteria), enhance intestinal barrier function, modulate host immunity, and have systemic anti-inflammatory and metabolic regulatory effects.

[0056] Physical barrier protection: HMLP absorbs water and expands to form a gel-like substance, increasing stool volume and water content, promoting intestinal motility, and relieving constipation. Simultaneously, this mucus-like gel layer forms a physical protective film on the surface of the intestinal mucosa, reducing direct irritation and damage to the intestinal wall from harmful substances and helping to maintain the integrity of the intestinal barrier.

[0057] (5) “Multi-faceted collaborative fighters” in the anti-tumor battlefield:

[0058] Indirect immune mediation: HMLP's powerful immune-enhancing effect is an important basis for its anti-tumor potential. By activating macrophages, NK cells, T cells, etc., it enhances the immune system's ability to recognize and eliminate tumor cells.

[0059] (6) The “loyal guardian” of liver health:

[0060] Protection against chemical liver injury: HMLP has demonstrated significant protective effects in animal models of liver injury induced by toxins such as alcohol, carbon tetrachloride (CCl4), and acetaminophen (APAP). It can reduce serum transaminase (ALT, AST, markers of hepatocellular damage) and bilirubin levels, and improve pathological changes in liver tissue (such as steatosis, necrosis, and inflammatory infiltration).

[0061] Based on the excellent biological activity and high safety of sunflower polysaccharide, sunflower polysaccharide shows attractive application prospects in many fields, such as functional food and health food, medicine, etc.

[0062] Sunflower has been approved by China's National Health Commission as a new resource food (new food raw material), which has laid a regulatory foundation for its application in the food field.

[0063] At present, sunflower polysaccharide is rarely used in the field of skin care products. How to fully utilize its characteristics and develop skin care products that are more easily accepted by consumers is a new research direction in this field.

[0064] The present invention effectively overcomes the adverse effects of thickeners on skin care products by replacing the thickeners in skin care products with Helianthus annuus polysaccharide extract. The skin care products prepared with Helianthus annuus polysaccharide extract as the active ingredient can not only retain the refreshing skin feel and fast absorption characteristics of water-based skin creams, but also have long-lasting water-locking and barrier repair functions.

[0065] In addition, the inventors also found that the golden sunflower polysaccharide extract extracted with a specific extract has more prominent anti-inflammatory and antioxidant effects in skin care products.

[0066] The components of the sunflower polysaccharide skin care cream provided by the present invention include: water, moisturizing agent, emollient, emulsifier, preservative, sunflower polysaccharide extract and pH regulator.

[0067] In addition to the Heliotropium polysaccharide extract, the functions of the other ingredients in the present invention are as follows:

[0068] Water: Mainly acts as a solvent, dissolving other water-soluble ingredients and forming the base texture of the product. It can also provide instant hydration and cooling sensation.

[0069] Moisturizer: absorbs moisture from deep layers of the skin and the environment, and adsorbs it into the stratum corneum to increase the skin's moisture content.

[0070] Emollients: fill the spaces between keratinocytes, smooth the skin texture, provide a soft touch, and reduce transepidermal water loss. Emollients in water-based creams are usually lighter in texture.

[0071] Emulsifier: Stably mixes the water phase and oil phase in the product to form a uniform cream texture. Without emulsifier, water and oil will separate into layers.

[0072] Preservatives: Prevent the growth of microorganisms (bacteria, fungi), ensure product safety and extend shelf life. Water-based products are particularly in need of preservatives.

[0073] Various exemplary embodiments of the present invention are now described in detail. This detailed description should not be considered as a limitation of the present invention, but should be understood as a more detailed description of certain aspects, features, and embodiments of the present invention. It should be understood that the terms used in the present invention are only for describing specific embodiments and are not intended to limit the present invention.

[0074] It should be pointed out that the matters not described in detail in the present invention are conventional operating means in this field and are not the focus of the present invention.

[0075] In addition, for numerical ranges in the present invention, it is understood that each intervening value between the upper and lower limits of the range is also specifically disclosed. Each smaller range between any stated value or stated range, and any other stated value or intervening value in the stated range, is also included in the present invention. The upper and lower limits of these smaller ranges may independently be included or excluded in the range.

[0076] 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 the invention pertains. Although preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the present invention.

[0077] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0078] Example 1

[0079] Preparation of sunflower polysaccharide skin care cream:

[0080] (1) The dried sunflower petals were crushed and then soaked in a 10 wt.% citric acid anhydrous ethanol solution for 20 min for degreasing, filtered and washed with water, and the filter residue was added to a deionized water solution adjusted to pH 5.0 with citric acid at a solid-liquid ratio of 1:20, refluxed at 90°C for 1 hour, and then filtered. After the filtrate was cooled, anhydrous ethanol was added to the system until the alcohol concentration was 70 wt.%, at which time the sunflower polysaccharide was precipitated, filtered, and freeze-dried to obtain the crude sunflower polysaccharide.

[0081] (2) The obtained crude Helianthus annuus polysaccharide was loaded onto a column (DEAE-Sepharose FF column) and connected to a fraction collector and a peristaltic pump; elution was performed with deionized water, and the eluate was collected in one tube every 5 minutes; the absorbance value of the obtained eluate at 490 nm was measured in real time by the phenol-sulfuric acid method, and the eluate before and after the absorbance peak and the eluate within 5 minutes was collected, concentrated, dialyzed with a 3500Da dialysis bag, and freeze-dried to obtain a purified Helianthus annuus polysaccharide extract with a polysaccharide purity of 98.1%.

[0082] (3) Prepare the raw materials according to the components in Table 1, add the moisturizer, emollient and purified sunflower polysaccharide extract to 50 parts of deionized water, stir at 800 rpm for 10 minutes to mix the materials evenly, and then homogenize at a rate of 2800 rpm for 10 minutes; continue to add emulsifier, stir at 800 rpm for 10 minutes to mix the materials evenly, and then homogenize at a rate of 2800 rpm for 10 minutes; then add preservatives and the remaining 25 parts of deionized water, stir at 800 rpm for 10 minutes to mix the materials evenly, use citric acid to adjust the pH value of the system to 5.0, and then homogenize at a rate of 2800 rpm for 10 minutes to prepare sunflower polysaccharide skin care cream.

[0083] Table 1 Dosage of each component

[0084]

[0085] Example 2

[0086] Preparation of sunflower polysaccharide skin care cream:

[0087] Compared with Example 1, the only difference is that in the step of extracting crude Heliotrope polysaccharide, the extracting solution used is deionized water, and the polysaccharide purity of the purified Heliotrope polysaccharide extract is 97.9%.

[0088] Comparative Example 1

[0089] Preparation of conventional skin care cream:

[0090] Compared with Example 1, the difference is that 2.5 parts of purified Heliotrope polysaccharide extract are replaced by 1.5 parts of carbomer and 1 part of xanthan gum.

[0091] The irritation of each group of skin care creams prepared in Examples 1 to 2 and Comparative Example 1 to the skin was examined: the test was based on 2.3.3 Skin Irritation Test in the Technical Specifications for Disinfection (2002 Edition).

[0092] Animal Preparation: New Zealand rabbits were pre-housed in a laboratory animal room for three days to acclimate to the environment. 24 hours before applying the skincare product, the hair on both sides of the rabbit's back spine was removed using electric clippers, avoiding any damage to the skin. The area to be removed should be no less than 5 cm x 5 cm on each side.

[0093] Skincare product application method: Using the left and right side self-comparison method, apply 0.5g of skincare cream evenly to a 3cm x 3cm area of ​​intact, hairless skin on the left side. Cover with a layer of non-irritating cellophane and secure with non-irritating adhesive tape. Apply water to the right side of the skin in the same manner as a negative control.

[0094] Symptom Observation: Observe local skin reactions 1 hour, 24 hours, and 48 hours after removing the test substance. Score according to the scoring criteria for skin irritation in Table 2-11 in 2.3.3 of the Technical Specifications for Disinfection (2002 edition) and the skin irritation intensity rating in Table 2-12. Specific scoring results are shown in Table 2.

[0095] Table 2 Scoring results of a complete skin irritation test

[0096]

[0097] The results in Table 2 show that the skin care products prepared in Examples 1 and 2 of the present invention did not cause skin irritation; the skin care product prepared in Comparative Example 1 showed barely visible erythema after 24 hours, indicating that using the Helianthus annuus polysaccharide extract provided by the present invention instead of conventional thickeners can avoid the irritation caused by the thickeners.

[0098] The moisturizing effects of the skin care creams prepared in Examples 1 to 2 and Comparative Example 1 were investigated:

[0099] Detection method:

[0100] The subjects cleaned their faces with a facial cleanser and dried their skin with dry tissue paper, and then sat in a laboratory at a temperature of 21±1°C and 50±10% RH for 30 minutes.

[0101] One side of the subject's cheek was randomly selected as the skin cream 1 area, and the other side was selected as the skin cream 2 area.

[0102] The researchers used a Tewameter TM300 skin water loss probe to measure transepidermal water loss (TEWL) at 0, 2, and 6 hours after applying the cream. The results are shown in Table 3.

[0103] Table 3 Moisturizing efficacy test results

[0104]

[0105] It can be seen from Table 3 that the sunflower polysaccharide skin care creams prepared in Examples 1 and 2 of the present invention can lock in moisture for a long time, while the moisture locking ability of ordinary water-soluble skin care creams is average.

[0106] The purified Heliotrope polysaccharide extracts obtained in Examples 1 and 2 were subjected to an inhibition test on the expression level of the inflammatory factor IL-6:

[0107] Using conventional ELISA method:

[0108] The experiment was conducted using HDF-a cells in the logarithmic growth phase. The cells were cultured at a density of 2×10 5 / well were inoculated in a 12-well plate and cultured in an environment of 37°C and 5% CO2 until the cells grew to 70% and then set aside. The groups were set up as follows: blank stimulation control group, lipopolysaccharide (LPS) stimulation group and sunflower polysaccharide group (final concentrations were 20, 40, 60, 100, 150, and 200 μM, respectively). The culture medium was carefully removed, and fresh complete culture medium containing different concentrations of sunflower polysaccharide extracts was added to the sunflower polysaccharide group, while equal volumes of water were added to the blank stimulation control group and the LPS stimulation group. After 1 hour, 1 μg / mL LPS was added to all groups except the blank stimulation control group for 4 hours to induce cell inflammation. The well plate was removed and the ELISA experiment was performed according to the instructions of the kit.

[0109] Collect the culture supernatant: collect the supernatant into a 1.5 mL EP tube, centrifuge at 1000 rpm for 10 minutes, and use the supernatant for ELISA detection of cytokines.

[0110] IL-6 secretion in the culture supernatant was detected using an enzyme-linked immunosorbent assay (ELISA) kit (Biyuntian, Cat. No.: IL-6:PI236). The procedure was as follows:

[0111] (1) Reagent preparation

[0112] ① Remove from the refrigerator and allow to equilibrate at room temperature for 20 minutes. ② Dilute the wash buffer (20×) to 1× with double-distilled water to prepare the required wash buffer. ③ Add the standard diluent to one bottle of standard according to the volume indicated on the standard label and incubate at room temperature for 15 minutes. ④ Prepare five clean 1.5 mL centrifuge tubes and pre-add 250 μL of the standard diluent to each tube. Perform serial dilutions of the standard to obtain six standard concentrations of 1000, 500, 250, 125, 62.5, and 31.25 pg / mL. Finally, add the diluted standards to the wells of the pre-coated plate in sequence. Add the standard diluent directly to the wells of the pre-coated plate to achieve a concentration of 0 pg / mL, for a total of seven standard concentrations. ⑤ Add 300 μL to each well and wait approximately 15-30 seconds before washing again. Wash the plate five times in total and pat dry on paper.

[0113] (2) Operation steps

[0114] ① Calculate the number of pre-coated strips required for one experiment, take out the required strips and place them in the 96-well frame.

[0115] ② Add samples or standards of different concentrations into the corresponding wells at 100 μL / well, seal the reaction wells with a transparent sealing film, and incubate at room temperature for 120 minutes.

[0116] ③ Wash the plate 5 times and pat dry on thick absorbent paper for the last wash.

[0117] ④ Add 100 μL / well of biotinylated antibody, seal the reaction wells with a transparent sealing film, and incubate at room temperature for 60 minutes.

[0118] ⑤ Wash the plate 5 times and pat dry on thick absorbent paper for the last wash.

[0119] ⑥ Add 100 μL / well of horseradish peroxidase-labeled streptavidin. Seal the wells with white sealing film and incubate at room temperature in the dark for 20 minutes.

[0120] ⑦ Wash the plate 5 times and pat dry on thick absorbent paper for the last wash.

[0121] ⑧Add 100 μL / well of TMB solution, seal the reaction wells with a white sealing film, and incubate at room temperature in the dark for 20 minutes.

[0122] ⑨Add 50 μL / well of stop solution, mix well and immediately measure the expression level of IL-6.

[0123] The test results showed that compared with the LPS group, the purified Heliotrope polysaccharide extracts in Examples 1 and 2 could downregulate the secretion level of inflammatory factor IL-6. The purified Heliotrope polysaccharide extract IC in Example 1 50 =75.62 μM, IC of the polysaccharide extract of Heliotropium truncatum purified in Example 250 =135.62μM, and all showed a dose-dependent decrease. Compared with conventional water-extracted polysaccharides, the Helianthus annuus polysaccharide extract obtained by the present invention using a weak acid solution with pH=5.0 has better anti-inflammatory activity on skin cells.

[0124] The antioxidant capacity of the purified Heliotrope polysaccharide extracts obtained from Examples 1 and 2 was investigated:

[0125] DPPH free radical (DPPH·) scavenging ability determination: 0.5 mg of purified Helianthus annuus polysaccharide extract of each group was dissolved in 0.5 mL of water, and 0.5 mmol·L -1 3.5 mL of DPPH solution was mixed and allowed to stand at room temperature in the dark for 30 min; the solution was adjusted to zero with anhydrous ethanol and the absorbance was measured at 517 nm, which was recorded as A 测定 ; Distilled water replaces the extract, recorded as A 空白 ; Anhydrous ethanol replaces DPPH, recorded as A 对照 ; Calculate the DPPH clearance rate according to formula (1).

[0126] Determination of hydroxyl radical (·OH) scavenging ability: 0.5 mg of purified Helianthus annuus polysaccharide extract from each group was dissolved in 0.5 mL of water, and 6 mmol·L -1 Ferrous sulfate 1mL, 6mmol·L -1 2 mL of hydrogen peroxide, 6 mmol·L -1 1 mL of salicylic acid was mixed and placed in a 37°C water bath for 30 min; the solution was adjusted to zero with distilled water and the absorbance was measured at 510 nm, which was recorded as A. 测定 ; Distilled water replaces the extract, recorded as A 空白 ; Distilled water instead of hydrogen peroxide is recorded as A 对照 ; Calculate the OH clearance rate according to formula (1).

[0127] Clearance (%) = [A 空白 -(A 测定 -A 对照 )] / A 空白 ×100%(1)

[0128] The calculated DPPH·scavenging rates and ·OH·scavenging rates are shown in Table 4.

[0129] Table 4 DPPH· and ·OH scavenging rates

[0130] DPPH clearance rate (%) OH removal rate (%) Example 1 <![CDATA[89.5 a ]]> <![CDATA[96.4 a ]]> Example 2 <![CDATA[76.9 b ]]> <![CDATA[87.5 b ]]>

[0131] Note: In Table 4, lowercase letters with different headers in the same column indicate significant differences.

[0132] It can be seen from Table 4 that, compared with conventional water-extracted polysaccharides, the Helianthus annuus polysaccharide extract obtained by extraction with a weak acid solution at pH=5.0 in the present invention has better antioxidant activity.

[0133] The embodiments described above are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Without departing from the spirit of the present invention, various modifications and improvements made to the technical solutions of the present invention by persons skilled in the art should fall within the scope of protection defined by the claims of the present invention.

Claims

1. A sunflower polysaccharide skin care cream, characterized in that: The components include, by mass, 60 to 85 parts of water, 3 to 15 parts of moisturizer, 5 to 15 parts of emollient, 6 to 10 parts of emulsifier, 0.1 to 0.5 parts of preservative and 1 to 3 parts of sunflower polysaccharide extract, and the pH of the system is adjusted to 4.5 to 6.5 using a pH regulator.

2. The sunflower polysaccharide skin care cream according to claim 1, characterized in that The moisturizing agent includes one or more of glycerin, sodium hyaluronate, propylene glycol, butylene glycol, amino acid moisturizing agent, sorbitol and trehalose.

3. The sunflower polysaccharide skin care cream according to claim 1, characterized in that The emollient comprises one or more of light synthetic esters, silicone oil derivatives, natural oils and fatty alcohols.

4. The sunflower polysaccharide skin care cream according to claim 1, characterized in that The emulsifier includes one or more of polysorbate emulsifiers, lecithin, glyceryl stearate and acrylic acid emulsifiers.

5. The sunflower polysaccharide skin care cream according to claim 1, characterized in that The preservatives include: one or more of phenoxyethanol, sodium benzoate, potassium sorbate, a combination of p-hydroxyacetophenone and caprylhydroxamic acid, polyol preservatives, isothiazolinone preservatives and paraben preservatives.

6. The sunflower polysaccharide skin care cream according to claim 1, characterized in that The extracting solution used when extracting the Helianthus annuus polysaccharide extract is a weakly acidic aqueous solution with a pH of 4.5 to 6.

5.

7. The sunflower polysaccharide skin care cream according to claim 1, characterized in that The pH regulator includes: citric acid, lactic acid, triethanolamine, sodium hydroxide or potassium hydroxide.

8. A method for preparing the sunflower polysaccharide skin care cream according to any one of claims 1 to 7, characterized in that: The following steps are involved: Moisturizer, emollient and sunflower polysaccharide extract are added to water accounting for 50 to 70% of the total water usage, mixed evenly and homogenized once, then an emulsifier is added, mixed evenly and homogenized twice, then a preservative and the remaining water are added, mixed evenly and the pH value is adjusted to 4.5 to 6.5 using a pH regulator, and homogenized three times to obtain the sunflower polysaccharide skin care cream.

9. The method for preparing the sunflower polysaccharide skin care cream according to claim 8, characterized in that: The conditions for the first homogenization, the second homogenization and the third homogenization are all 2500-3500 rpm homogenization for 5-10 minutes.

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