Preparation of a plant polysaccharide extract and its use in skin care

Abstract

The present application belongs to the technical field of polysaccharide extraction and application, and relates to preparation of a plant polysaccharide extract and application thereof in skin care. The present application uses coix seed as raw material, and obtains coix seed polysaccharide through the steps of pretreatment, defatting + enzyme treatment, ultrasonic-alkali solution-arginine synergistic extraction, preliminary purification, purification and collection. Compared with the prior art, the present application significantly improves the extraction rate and purity of coix seed polysaccharide through the synergistic effect of each process, and the prepared coix seed polysaccharide has good application prospect in the field of skin care, and can be used for preparing skin care products of antioxidant, moisturizing and soothing repair types.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of polysaccharide extraction and application, and particularly relates to a preparation method of a plant polysaccharide extract and application thereof in skin care. BACKGROUND

[0002] Plant polysaccharides are a class of natural high-molecular compounds widely existing in plants, and have various biological activities and unique advantages in skin care. Studies have shown that plant polysaccharides can improve skin condition by replenishing skin moisture, enhancing skin barrier function, scavenging free radicals, and inhibiting inflammatory response. For example, some plant polysaccharides have excellent moisturizing properties and can reduce skin moisture loss; some plant polysaccharides have antioxidant activity and can delay skin aging; and some plant polysaccharides can promote the proliferation and repair of skin cells and accelerate the healing of damaged skin.

[0003] Coix seed is a plant with medicinal and edible properties, and is rich in protein, oil, soluble polysaccharides and other components. Chinese Patent Publication No. CN105367676A discloses an extraction method of coix seed polysaccharides. The coix seed kernels are dried and crushed, then extracted with high-pressure water, concentrated under reduced pressure, treated with trifluoroacetic acid and centrifuged, precipitated with ethanol, and finally freeze-dried to obtain coix seed polysaccharide product. However, the yield of coix seed polysaccharides is low, only 5%-6%. Chinese Patent Publication No. CN105037576A discloses an extraction and separation method of multiple active components of coix seed. The coix seed is first crushed and granulated, then defatted by supercritical CO2 extraction, and then extracted with water and enzymatically treated, combined with membrane separation technology, to finally obtain a coix seed extract containing polysaccharides and other components, realizing the separation and extraction of oil, protein and polysaccharides and other components in coix seed. However, the yield of polysaccharides is only about 0.53%, and the purity of polysaccharides is only about 55.02%.

[0004] However, the current preparation method of plant polysaccharide extract still has some problems. Traditional extraction methods such as hot water extraction and alcohol precipitation have the defects of low extraction efficiency and low purity. Therefore, it is of great significance to develop an efficient preparation method of plant polysaccharide extract, improve the extraction rate and purity of polysaccharides, and further study the application mechanism in skin care, for promoting the development of natural skin care products. SUMMARY

[0005] To overcome the shortcomings of the prior art, the present application provides a preparation method of a plant polysaccharide extract, which uses coix seed as raw material and optimizes the extraction process to improve the extraction rate and purity of polysaccharides. The specific steps are as follows:

[0006] (1) Pretreatment: After sieving and removing impurities, rinse the coix seed with flowing distilled water for 3-4 times, drain the surface moisture, dry in a 55-65℃ drying oven until the moisture content is ≤6%, then crush with a high-speed universal crusher, pass through a 120-150 mesh sieve, and collect the undersize powder.

[0007] (2) Defatting + enzyme treatment: Add petroleum ether to the crushed coix seed powder at a solid-liquid ratio of 1:9-11 (g:mL), defat at 45-55℃ water bath for 1.2-1.8 hours, filter, and wash the residue with distilled water until there is no ether smell, and dry at 55-65℃; add distilled water to the dried residue at a solid-liquid ratio of 1:14-16, adjust the pH to 5.8-6.2 with one of the following buffers: citric acid-sodium citrate buffer, disodium hydrogen phosphate-sodium dihydrogen phosphate buffer, or acetic acid-sodium acetate buffer, add 0.8%-1.2% papain based on the mass of the residue, and enzymatically hydrolyze at 48-52℃ water bath shaker for 0.8-1.2 hours, then inactivate the enzyme by boiling in a water bath for 8-12 minutes, and filter after cooling.

[0008] (3) Ultrasonic-alkali-arginine synergistic extraction: The residue after enzymatic hydrolysis is extracted by three-stage ultrasonic, 0.3%-0.7% Na2CO3 solution and 0.1%-0.5% arginine are added at a solid-liquid ratio of 1:20-30 (g:mL); the parameters of three-stage ultrasonic are as follows: the first stage, power 320-340W, temperature 60-70℃, extraction time 10-20 minutes; the second stage, power 360-380W, temperature 50-60℃, extraction time 10-20 minutes; the third stage, power 360-380W, temperature 65-75℃, extraction time 10-20 minutes; a total of 2-3 times of extraction, centrifugation after each extraction, and combination of supernatants.

[0009] (4) Preliminary purification: Concentrate the supernatant under reduced pressure at 53-57℃ to 1 / 6-1 / 5 of the original volume; first add anhydrous ethanol to a concentration of 38%-42%, and stand at 3-5℃ for 1.8-2.2 hours, then centrifuge and discard the precipitate; take the supernatant and continue to add ethanol to a concentration of 73%-77%, and stand at 3-5℃ for 9-11 hours, then centrifuge to collect the precipitate, and wash with 78%-82% ethanol for 2-4 times, and dry under vacuum at 55-65℃.

[0010] (5) Purification and collection: AB-8 macroporous resin is used to purify the crude polysaccharide obtained by preliminary purification, the macroporous resin is pretreated by soaking with 95% ethanol for 22-26 hours, and then rinsing with distilled water until there is no alcohol smell; the concentration of the crude polysaccharide solution is 2.8-3.2 mg / mL, and the flow rate of column loading is 0.7-0.9 mL / min; during elution, first wash with distilled water until the effluent is colorless, then elute with 53%-57% ethanol solution, and collect the eluate; combine the eluate, concentrate under reduced pressure, and freeze-dry to obtain coix seed polysaccharide.

[0011] Compared with the prior art, the technical effect of the present application is that:

[0012] (1) The extraction rate and purity are significantly improved: The present application uses a three-stage ultrasonic process of ultrasonic-alkali dissolution-arginine synergistic extraction, combined with optimized parameters such as solid-liquid ratio and reagent concentration, so that the yield of coix seed polysaccharide can reach 7.9%-9.2%, and the purity can reach 80.7%-84.5%, which is much higher than the yield of 5%-6% and the purity of about 55.02% in the prior art.

[0013] (2) Good purification effect: The present application uses a combined process of fractional precipitation and macroporous resin purification, which effectively removes impurities and improves the purity of polysaccharide, providing high-quality raw materials for subsequent application in skin care.

[0014] (3) High application value: The coix seed polysaccharide prepared by the present application has excellent antioxidant and moisturizing properties, and has a wide application prospect in the field of skin care, such as antioxidant essence which can significantly reduce facial fine lines and improve skin gloss, moisturizing cream and soothing repair mask which also show good skin care effect. BRIEF DESCRIPTION OF DRAWINGS

[0015] Figure 1 : The yield and purity of coix seed polysaccharide in each group. DETAILED DESCRIPTION

[0016] In order to make the purpose, technical scheme of the present application more clear, the following examples are used to further illustrate the present application, but the protection scope of the present application is not limited to these examples, and the examples are only used to explain the present application. Those skilled in the art should understand that any changes or equivalent replacements without departing from the concept of the present application are included in the protection scope of the present application.

[0017] Example 1 Preparation of coix seed polysaccharide

[0018] (1) Pretreatment:

[0019] Select raw materials, take small particles of coix seed, remove impurities, rinse with flowing distilled water for 3-4 times, take out and drain the surface water; evenly spread on a drying tray, set the drying temperature to 65℃ in the drying box, dry to the moisture content of coix seed ≤6%, put into a high-speed universal pulverizer, and the coix seed powder is passed through a 130 mesh sieve, and the powder under the sieve is collected.

[0020] (2) Double treatment of degreasing and enzyme hydrolysis:

[0021] ① Degreasing: Add petroleum ether to the pulverized coix seed powder at a solid-liquid ratio of 1:10 (g:mL), stir for 1.5 hours in a 50℃ water bath, filter the residue, wash with distilled water until there is no ether smell, and dry at 60℃.

[0022] ② Enzymatic deproteinization: to the dried residue, distilled water was added at a ratio of 1:15, and the pH was adjusted to 6.0 with 0.1 mol / L citric acid-sodium citrate buffer. Papain was added at a concentration of 1.0% of the residue mass, and enzymatic hydrolysis was performed at 50°C for 1 hour on a water bath shaker. The enzyme was inactivated by boiling in a water bath for 10 minutes. After cooling, the mixture was filtered.

[0023] (3) Ultrasonic-alkali-arginine synergistic extraction:

[0024] The residue after enzymatic hydrolysis was added with 0.5% Na2CO3 solution and 0.3% arginine at a ratio of 1:25 (g:mL) and stirred uniformly. It was placed in an ultrasonic extraction instrument for three-stage ultrasonic extraction:

[0025] First stage: power 330W, temperature 65°C, extraction time 15 minutes;

[0026] Second stage: power 370W, temperature 55°C, extraction time 15 minutes;

[0027] Third stage: power 370W, temperature 70°C, extraction time 15 minutes.

[0028] A total of three extractions were performed. After each extraction, centrifugation was performed, and the supernatants from multiple extractions were combined.

[0029] (4) Preliminary purification:

[0030] ① Concentration: the supernatant was concentrated at 55°C under reduced pressure to 1 / 6 of the original volume.

[0031] ② Fractionated precipitation: first, add anhydrous ethanol to a concentration of 40%, and let it stand at 4°C for 2 hours. Centrifuge and discard the precipitate. Take the supernatant and continue to add ethanol to a concentration of 75%. Let it stand at 4°C for 10 hours. Centrifuge to collect the precipitate. Wash with 80% ethanol for 3 times. Dry at 60°C under vacuum.

[0032] (5) Purification and collection of coix seed polysaccharides

[0033] ① Pretreatment of macroporous resin: soak macroporous resin AB-8 type with 95% ethanol for 24 hours, then rinse with distilled water until there is no alcohol smell.

[0034] ② Sample loading and elution: increase the concentration of the crude polysaccharide solution to 3.0 mg / mL, and reduce the flow rate to 0.8 mL / min. When eluting, first wash with distilled water until the effluent is colorless, then elute with 55% ethanol solution, and collect the eluate.

[0035] (6) Collection of coix seed polysaccharides:

[0036] ① Combine the eluate and place it in a rotary evaporator under the following conditions: 52°C, vacuum degree 0.07 MPa, and reduce pressure to concentrate the eluate to 1 / 7 of the original volume without obvious ethanol odor.

[0037] 2. Transfer the concentrated solution to a clean culture dish, and place it in a vacuum freeze dryer. Set the freezing temperature to -37°C, the vacuum degree to 15 Pa, and the drying time to 26 hours until the solution is completely freeze-dried into loose powder to obtain the coix seed polysaccharide.

[0038] Example 2. Preparation of coix seed polysaccharide

[0039] (1) Pretreatment:

[0040] Select raw materials, take small-grained coix seed, remove impurities, rinse with flowing distilled water for 3-4 times, take out, and drain the surface water. Uniformly spread on a drying tray, place in a drying oven, set the drying temperature to 60°C, dry until the moisture content of the coix seed is ≤6%, place in a high-speed universal pulverizer, and pulverize the coix seed powder. Collect the powder under the sieve.

[0041] (2) Double treatment of defatting and enzymatic hydrolysis:

[0042] ① Defatting: add petroleum ether to the pulverized coix seed powder at a solid-liquid ratio of 1:9 (g:mL), and defat at 45-55°C for 1.2 hours. After filtration, wash the residue with distilled water until there is no ether smell, and dry at 55°C.

[0043] ② Enzymatic hydrolysis for protein removal: add distilled water to the dried residue at a solid-liquid ratio of 1:14, adjust the pH to 5.8 with 0.1 mol / L disodium hydrogen phosphate-sodium dihydrogen phosphate buffer, add 0.8% papain based on the mass of the residue, and perform enzymatic hydrolysis at 48°C for 0.8 hours. Boil for 8 minutes to inactivate the enzyme, and then filter after cooling.

[0044] (3) Ultrasonic-alkali-arginine synergistic extraction:

[0045] Take the enzymatically hydrolyzed residue, add 0.3% Na2CO3 solution and 0.1% arginine at a solid-liquid ratio of 1:20 (g:mL), and stir uniformly. Place it in an ultrasonic extractor for three-stage ultrasonic extraction:

[0046] First stage: power 320W, temperature 60°C, extraction time 10 minutes;

[0047] Second stage: power 360W, temperature 50°C, extraction time 10 minutes;

[0048] Third stage: power 360W, temperature 65°C, extraction time 10 minutes;

[0049] Extract for a total of 3 times. Centrifuge after each extraction, and combine the supernatants.

[0050] (4) Preliminary purification:

[0051] ① Concentration: The supernatant was concentrated at 53°C under reduced pressure to 1 / 6 of the original volume.

[0052] ② Fractionated precipitation: First, add anhydrous ethanol to a concentration of 38%, and stand at 3°C for 1.8 hours. Centrifuge and discard the precipitate. Take the supernatant and continue to add ethanol to a concentration of 73%, and stand at 3°C for 9 hours. Centrifuge to collect the precipitate, wash twice with 78% ethanol, and dry at 55°C under vacuum.

[0053] (5) Purification and collection of coix seed polysaccharides

[0054] ① Macroporous resin pretreatment: Soak the AB-8 type macroporous resin in 95% ethanol for 22 hours, and then rinse directly with distilled water until there is no alcohol smell.

[0055] ② Sample loading and elution: Increase the concentration of the crude polysaccharide solution to 2.8 mg / mL, and reduce the flow rate to 0.7 mL / min. When eluting, first wash with distilled water until the effluent is colorless, and then elute with a 53% ethanol solution. Collect the eluate.

[0056] (6) Collection of coix seed polysaccharides:

[0057] ① Combine the eluate and place it in a rotary evaporator under the following conditions: 50°C, vacuum degree 0.06 MPa, and reduced pressure. Concentrate until the eluate is concentrated to 1 / 8 of the original volume and there is no obvious ethanol smell.

[0058] ② Transfer the concentrated solution to a clean culture dish, and place it in a vacuum freeze-drying machine. Set the freezing temperature to -40°C, the vacuum degree to 10 Pa, and the drying time to 24 hours. Dry until the solution is completely freeze-dried into a loose powder, and obtain coix seed polysaccharides.

[0059] Example 3 Preparation of coix seed polysaccharides

[0060] (1) Pretreatment:

[0061] Select raw materials, take small coix seed, remove impurities, rinse 3-4 times with flowing distilled water, pick out, and drain the surface water. Uniformly spread on a drying tray, place in a drying oven, set the drying temperature to 70°C, dry until the moisture content of coix seed is ≤6%, place in a high-speed universal pulverizer, pulverize, and collect the powder under the sieve.

[0062] (2) Double treatment of degreasing and enzyme hydrolysis:

[0063] ① Degreasing: Add petroleum ether to the pulverized coix seed powder at a solid-liquid ratio of 1:11 (g:mL), and stir in a 55°C water bath for 1.8 hours. After filtration, wash the residue with distilled water until there is no ether smell, and dry at 65°C.

[0064] ② Enzymatic deproteinization: To the dried residue, distilled water was added at a ratio of 1:16, and the pH was adjusted to 6.2 with 0.1 mol / L acetic acid-sodium acetate buffer. Papain was added at a concentration of 1.2% of the residue mass, and the mixture was incubated in a 52°C water bath for 1.2 hours. The enzyme was inactivated by boiling in a water bath for 12 minutes. After cooling, the mixture was filtered.

[0065] (3) Ultrasonic-alkali-arginine synergistic extraction:

[0066] The residue after enzymatic hydrolysis was added with 0.7% Na2CO3 solution and 0.5% arginine at a ratio of 1:30 (g:mL) and stirred uniformly. The mixture was subjected to three-stage ultrasonic extraction in an ultrasonic extraction instrument:

[0067] First stage: power 340 W, temperature 70°C, extraction time 20 minutes;

[0068] Second stage: power 380 W, temperature 60°C, extraction time 20 minutes;

[0069] Third stage: power 380 W, temperature 75°C, extraction time 20 minutes;

[0070] The extraction was performed twice. After each extraction, the mixture was centrifuged, and the supernatants were combined.

[0071] (4) Preliminary purification:

[0072] ① Concentration: The supernatant was concentrated at 57°C under reduced pressure to 1 / 5 of the original volume.

[0073] ② Fractionated precipitation: First, add anhydrous ethanol to a concentration of 42%, and let it stand at 5°C for 2.2 hours. Centrifuge and discard the precipitate. Take the supernatant and continue to add ethanol to a concentration of 77%. Let it stand at 5°C for 11 hours. Centrifuge to collect the precipitate. Wash the precipitate with 82% ethanol 4 times, and dry it at 65°C under vacuum.

[0074] (5) Purification and collection of coix seed polysaccharides

[0075] ① Pretreatment of macroporous resin: Soak macroporous resin AB-8 type with 95% ethanol for 26 hours, then rinse with distilled water until there is no alcohol smell.

[0076] ② Sample loading and elution: Increase the concentration of the crude polysaccharide solution to 3.2 mg / mL, and reduce the flow rate to 0.9 mL / min. First, wash with distilled water until the effluent is colorless, then elute with 57% ethanol solution, and collect the eluate.

[0077] (6) Collection of coix seed polysaccharides:

[0078] ① Combine the eluate, and concentrate it in a rotary evaporator at 55°C and a vacuum degree of 0.08 MPa until the eluate is concentrated to 1 / 6 of the original volume and there is no obvious ethanol smell.

[0079] 2. Transfer the concentrated solution to a clean culture dish, and place it in a vacuum freeze dryer. Set the freezing temperature to -35°C, the vacuum degree to 20 Pa, and the drying time to 30 hours. Dry the solution until it is completely freeze-dried into a loose powder, and obtain the polysaccharide from the seeds of Job's tears.

[0080] Experimental design of the effect of ultrasonic-alkali-arginine synergistic extraction on the yield and purity of polysaccharide from the seeds of Job's tears

[0081] Set up the following 10 groups of experiments. Each group of experiments is subjected to the pre-treatment of (1) pre-treatment and (2) double treatment of degreasing and enzymolysis in Example 1. Starting from the step of (3) ultrasonic-alkali-arginine synergistic extraction, the experimental conditions are changed, and the subsequent steps of (4) preliminary purification, (5) purification, and (6) collection of polysaccharide from the seeds of Job's tears are performed according to the operations in Example 1.

[0082] Example 1 group: Perform the ultrasonic-alkali-arginine synergistic extraction according to the conditions in (3) of Example 1, that is, add 0.5% Na2CO3 solution and 0.3% arginine, and use three-stage ultrasonic extraction (first stage: power 330 W, temperature 65°C, extraction time 15 minutes; second stage: power 370 W, temperature 55°C, extraction time 15 minutes; third stage: power 370 W, temperature 70°C, extraction time 15 minutes), and extract for 3 times.

[0083] Example 2 group: Perform the ultrasonic-alkali-arginine synergistic extraction according to the conditions in (3) of Example 2.

[0084] Example 3 group: Perform the ultrasonic-alkali-arginine synergistic extraction according to the conditions in (3) of Example 3.

[0085] Control 1 group (Na2CO3 absence group): Take the residue after enzymolysis, add distilled water according to the solid-liquid ratio of 1:25 (g:mL), and add 0.3% arginine. Stir uniformly. The ultrasonic extraction conditions are the same as those in Example 1 group, and extract for 3 times. After each extraction, centrifuge and combine the supernatants.

[0086] Control 2 group (arginine absence group): Take the residue after enzymolysis, add 0.5% Na2CO3 solution according to the solid-liquid ratio of 1:25 (g:mL), and stir uniformly. The ultrasonic extraction conditions are the same as those in Example 1 group, and extract for 3 times. After each extraction, centrifuge and combine the supernatants.

[0087] Control 3 group (sodium carbonate and arginine absence group): Take the residue after enzymolysis, add distilled water according to the solid-liquid ratio of 1:25 (g:mL), and stir uniformly. The ultrasonic extraction conditions are the same as those in Example 1 group, and extract for 3 times. After each extraction, centrifuge and combine the supernatants.

[0088] Control group 4 (histidine group): take the residue after enzymatic hydrolysis, add 0.5% Na2CO3 solution and 0.3% histidine according to the solid-liquid ratio of 1:25 (g:mL), and stir uniformly. Ultrasonic conditions are the same as the three-stage ultrasonic of example 1 group, and a total of 3 times of extraction. After each extraction, centrifugation is performed, and the supernatants of multiple times are combined.

[0089] Control group 5 (single-stage ultrasonic group): take the residue after enzymatic hydrolysis, add 0.5% Na2CO3 solution and 0.3% arginine according to the solid-liquid ratio of 1:25 (g:mL), and stir uniformly. Place in the ultrasonic extraction instrument for single-stage ultrasonic: power 370W, temperature 65℃, extraction time 45 minutes. A total of 3 times of extraction. After each extraction, centrifugation is performed, and the supernatants of multiple times are combined.

[0090] Control group 6 (two-stage ultrasonic group): take the residue after enzymatic hydrolysis, add 0.5% Na2CO3 solution and 0.3% arginine according to the solid-liquid ratio of 1:25 (g:mL), and stir uniformly. Place in the ultrasonic extraction instrument for two-stage ultrasonic: first stage: power 330W, temperature 65℃, extraction time 25 minutes; second stage: power 370W, temperature 60℃, extraction time 25 minutes. A total of 3 times of extraction. After each extraction, centrifugation is performed, and the supernatants of multiple times are combined.

[0091] Control group 7 (three-stage gradient temperature group): take the residue after enzymatic hydrolysis, add 0.5% Na2CO3 solution and 0.3% arginine according to the solid-liquid ratio of 1:25 (g:mL), and stir uniformly. Place in the ultrasonic extraction instrument for three-stage ultrasonic: first stage: power 330W, temperature 50℃, extraction time 15 minutes; second stage: power 370W, temperature 60℃, extraction time 15 minutes; third stage: power 370W, temperature 70℃, extraction time 15 minutes. A total of 3 times of extraction. After each extraction, centrifugation is performed, and the supernatants of multiple times are combined.

[0092] Specific experimental steps:

[0093] a. According to the pretreatment step (1) in example 1, the small grain coix seed is treated to obtain the powder screened.

[0094] b. According to the double treatment step of defatting and enzymatic hydrolysis (2) in example 1, the powder is treated to obtain the residue after enzymatic hydrolysis.

[0095] According to the different conditions of the above 10 groups of experiments, the ultrasonic-alkali solution-arginine synergistic extraction step (3) is carried out.

[0096] c. All groups are operated according to the preliminary purification (4) and purification (5) and coix seed polysaccharide collection steps in example 1.

[0097] d. The coix seed polysaccharides obtained in each group are collected.

[0098] Detection index and method

[0099] Yield determination

[0100] Yield = (mass of coix seed polysaccharide / mass of coix seed powder after pretreatment) x 100%. The mass of coix seed polysaccharide was determined by gravimetric method.

[0101] Purity determination

[0102] The purity of coix seed polysaccharide was determined by phenol-sulfuric acid method, using glucose as standard, and the absorbance of the sample at 490 nm was determined to calculate the purity of polysaccharide.

[0103] Table 1: Yield and purity of coix seed polysaccharide in each group

[0104]

[0105] Table 1 shows that the optimal extraction conditions are as follows: adding 0.5% Na2CO3 and 0.3% arginine according to the solid-liquid ratio of 1:25, using three-stage ultrasonic (330W / 65℃→370W / 55℃→370W / 70℃), and extracting for 3 times, which can achieve a yield of 9.2% and a purity of 84.5%, which is significantly improved compared with the prior art. Through the ultrasonic-alkali-arginine synergistic system and the dual mechanisms of "high-efficiency dissolution + structure protection", the problems of low extraction rate and insufficient purity in traditional methods are solved, and high-quality raw material guarantee is provided for the application of coix seed polysaccharide in skin care field.

[0106] Application 1: Coix seed polysaccharide antioxidant essence

[0107] Formulation composition (by weight percentage): coix seed polysaccharide 2.5% of example 1, vitamin C derivative 1.0%, glutathione 0.5%, propylene glycol 4.0%, ethanol 5.0%, xanthan gum 0.1%, disodium EDTA 0.05%, deionized water balance.

[0108] Preparation process: dissolve coix seed polysaccharide, propylene glycol and disodium EDTA in deionized water, stir until completely dissolved; dissolve vitamin C derivative and glutathione in ethanol, slowly add it to the above aqueous solution, stir uniformly; finally add xanthan gum, high-speed stirring to form a uniform transparent essence.

[0109] Efficacy verification:

[0110] DPPH radical scavenging rate determination method: accurately weigh DPPH reagent, prepare DPPH solution with a concentration of 0.1 mmol / L with anhydrous ethanol, and store in the dark. Take 1 mL of essence from the experimental group and the control group respectively, add 4 mL of DPPH solution, mix thoroughly, and then place in the dark at room temperature for 30 minutes. Measure the absorbance at 517 nm wavelength (recorded as A sample) by ultraviolet-visible spectrophotometer. At the same time, set up a blank group: take 1 mL of anhydrous ethanol and add 4 mL of DPPH solution, and measure the absorbance (recorded as A blank); control group: take 1 mL of essence matrix (without active ingredient coix seed polysaccharide) and add 4 mL of DPPH solution, and measure the absorbance (recorded as A control). Calculate the DPPH radical scavenging rate according to the formula: scavenging rate (%) = [1- (A sample-A control) / A blank] x 100%, repeat each experiment 3 times, and take the average value.

[0111] Table 2 DPPH radical scavenging rate

[0112]

[0113] The antioxidant capacity of the essence was determined by DPPH radical scavenging experiment, and the results in Table 2 showed that the DPPH radical scavenging rate of coix seed polysaccharide antioxidant essence could reach 82.5%, which was significantly higher than that of the control group without coix seed polysaccharide (scavenging rate 46.7%).

[0114] Volunteer use effect

[0115] Volunteer information: 15 healthy volunteers aged 25-40 years old with fine lines on the face were selected, including 3 males and 12 females, all without skin diseases and allergic history, and no other skin care products or drugs affecting skin condition were used during the experiment.

[0116] Method of use: the volunteers evenly applied an appropriate amount of experimental group essence on the face after cleansing every night, and continuously used for 4 weeks. Detection was carried out before use (day 0), 2 weeks after use, and 4 weeks after use, respectively.

[0117] Detection index and data:

[0118] Number of facial fine lines: the same area of the volunteers' face was photographed and analyzed by skin image analyzer, and the number of fine lines was counted, and the specific data were as follows:

[0119] Table 3 Number of facial fine lines and reduction rate of 15 volunteers

[0120]

[0121] 4Skin gloss: the skin gloss of the volunteers' face was determined by skin image analyzer (expressed as relative numerical value, the higher the value, the better the gloss), and the specific data were as follows:

[0122] Table 4 Facial skin glossiness of 15 volunteers

[0123]

[0124] Table 3 and Table 4 show that 15 volunteers with facial fine lines were selected, and the serum was applied every night, and used continuously for 4 weeks. Through the skin image analyzer, the number of facial fine lines of the volunteers was reduced by an average of 20.1%, and the skin glossiness was increased by 15.0%.

[0125] Application 2: Coixan Moisturizing Emulsion

[0126] Formulation composition (by weight percentage): Example 2 coixan 3.0%, glycerol 5.0%, sodium hyaluronate 0.1%, caprylic / capric triglyceride 3.0%, stearic acid 1.5%, lanolin 2.0%, triethanolamine 0.5%, nipagin ethyl ester 0.1%, deionized water balance.

[0127] Preparation process: Dissolve glycerol, sodium hyaluronate, and coixan in deionized water, heat to 80°C as the water phase; mix caprylic / capric triglyceride, stearic acid, and lanolin, heat to 80°C as the oil phase; slowly add the oil phase to the water phase under stirring, keep 80°C stirring for 20 minutes, add triethanolamine and nipagin ethyl ester when the temperature is lowered to 50°C, continue stirring to room temperature, and the moisturizing emulsion is obtained.

[0128] Efficacy verification: 20 volunteers with skin water content less than 30% were selected, and the emulsion was applied to the face every morning and evening, and used continuously for 28 days. The skin water content of the face was measured before and after use by using the skin moisture tester, and the results showed that the skin water content was increased to about 45.2% after use, indicating that the emulsion has excellent moisturizing effect.

Claims

1. A method for preparing a plant polysaccharide extract, characterized in that, The preparation method includes the following steps: (1) Pretreatment: The Job's tears are screened, washed, dried and crushed; (2) Degreasing + enzymatic hydrolysis: The crushed coix seed powder is degreased, and the residue after degreasing is enzymatically hydrolyzed to remove protein. (3) Ultrasonic-alkali-arginine synergistic extraction: Three-stage ultrasonic extraction is used to extract the residue after enzymatic hydrolysis. 0.3%-0.7% Na2CO3 solution and 0.1%-0.5% arginine are added at a material-liquid ratio of 1g:20-30mL. (4) Preliminary purification: The supernatant obtained from the extraction is concentrated and fractionated for precipitation; (5) Purification and collection: The crude polysaccharide obtained from the preliminary purification was purified by macroporous resin, and the coix seed polysaccharide was collected; The parameters for the three-stage ultrasound are as follows: Stage 1: Power 320-340W, Temperature 60-70℃, Extraction time 10-20 minutes; Stage 2: Power 360-380W, Temperature 50-60℃, Extraction time 10-20 minutes; Stage 3: Power 360-380W, Temperature 65-75℃, Extraction time 10-20 minutes; Extraction is performed 2-3 times in total, centrifuged after each extraction, and the supernatants from multiple extractions are combined. The preliminary purification steps include: concentration: concentrating the supernatant under reduced pressure at 53-57℃ to 1 / 6-1 / 5 of its original volume; fractionation precipitation: first adding anhydrous ethanol to a concentration of 38%-42%, letting it stand at 3-5℃ for 1.8-2.2 hours, centrifuging to discard the precipitate; taking the supernatant and adding more ethanol to a concentration of 73%-77%, letting it stand at 3-5℃ for 9-11 hours, centrifuging to collect the precipitate, washing it 2-4 times with 78%-82% ethanol, and vacuum drying at 55-65℃; The purification and collection steps include: sample loading and elution: the concentration of crude polysaccharide solution is 2.8-3.2 mg / mL, and the column loading flow rate is 0.7-0.9 mL / min; during elution, first wash with distilled water until the eluent is colorless, then elute with 53%-57% ethanol solution, and collect the eluent; coix seed polysaccharide collection: combine the eluents, concentrate under reduced pressure, and freeze-dry to obtain coix seed polysaccharide.

2. The preparation method according to claim 1, characterized in that, In the ultrasound-alkali dissolution-arginine synergistic extraction step: 0.5% Na2CO3 solution and 0.3% arginine are added at a material-liquid ratio of 1 g: 20-30 mL.

3. The preparation method according to claim 1, characterized in that, The parameters of the three-segment ultrasound are as follows: First stage: Power 330W, temperature 65℃, extraction time 15 minutes; Second stage: Power 370W, temperature 55℃, extraction time 15 minutes; Third stage: Power 370W, temperature 70℃, extraction time 15 minutes.

4. The preparation method according to claim 1, characterized in that, The defatting + enzymatic hydrolysis dual treatment step includes: Degreasing: Add petroleum ether to the pulverized coix seed powder at a material-to-liquid ratio of 1 g: 9-11 mL, stir and degrease in a water bath at 45-55℃ for 1.2-1.8 hours, filter and wash the residue with distilled water until there is no ether odor, and dry at 55-65℃. Enzymatic deproteinization: Add distilled water to the dried residue at a material-to-liquid ratio of 1:14-16, adjust the pH to 5.8-6.2, add 0.8%-1.2% papain by weight of the residue, and enzymatically hydrolyze in a water bath shaker at 48-52℃ for 0.8-1.2 hours. Inactivate the enzyme in a boiling water bath for 8-12 minutes, cool, and then filter.

5. The preparation method according to claim 4, characterized in that, The reagent used to adjust the pH is one of the following: citrate-sodium citrate buffer, disodium hydrogen phosphate-sodium dihydrogen phosphate buffer, or acetic acid-sodium acetate buffer.

6. The preparation method according to claim 1, characterized in that, In the purification and collection steps, the macroporous resin is AB-8 type, and the pretreatment method is to soak it in 95% ethanol for 22-26 hours, and then rinse it with distilled water until there is no alcohol odor.

7. The application of a plant polysaccharide extract in skin care, characterized in that, The plant polysaccharide extract is obtained by the preparation method described in claim 1.

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