An olea europaea polysaccharide extract, and a preparation method and application thereof

Abstract

The application relates to the field of biotechnology, in particular to an Olea europaea L. polysaccharide extract and a preparation method and application thereof; the monosaccharide composition of the Olea europaea L. polysaccharide extract is as follows: mannose is greater than or equal to 5%, rhamnose is greater than or equal to 6%, glucuronic acid is greater than or equal to 3%, galacturonic acid is greater than or equal to 33%, glucose is greater than or equal to 6%, galactose is greater than or equal to 20% and arabinose is greater than or equal to 20% in terms of mass fraction. The Olea europaea L. polysaccharide extract can be combined with the tyrosinase inhibiting effect of a liquorice extract based on the antioxidation and anti-inflammatory effects, can more effectively reduce melanin deposition, improve skin spots, and thus achieve a whitening effect.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biotechnology, in particular to an Olea europaea L. polysaccharide extract and a preparation method and application thereof. BACKGROUND

[0002] Gel refers to a thick liquid or semi-solid preparation of a solution, suspension or emulsion type prepared from a drug and a gelling adjuvant. Generally, a gel contains a gel-type matrix, and the gel-type matrix belongs to a single-phase dispersion system, which is divided into aqueous and oily types. At present, the aqueous gel is more commonly used in clinical application. The aqueous gel has the characteristics of easy spreading and washing, and in addition, the aqueous gel has no greasy feeling and can effectively absorb tissue exudate, so the normal physiological activities of the skin are generally not hindered by the aqueous gel. These characteristics enable the drug contained in the aqueous gel to be quickly released, but the lubricating effect of the aqueous gel is poor, it is easy to lose water and cause mold, so it is often necessary to add stabilizers and preservatives; the commonly used preservative is nepal gold methine, and the commonly used stabilizer is disodium ethylenediaminetetraacetate. Although the amount of these stabilizers and preservatives is small, they are all chemically synthesized drugs, and large amounts of use can cause damage to the human body.

[0003] In the typical application field of gels, commonly used whitening drugs include tyrosine kinase inhibitors, hydroquinone substances, vitamin C, flavones and polyphenol substances, etc. However, tyrosine kinase inhibitors and hydroquinone substances have strong irritability to the skin of the human body, and vitamin C, flavones and polyphenol substances are easily oxidized in the air environment and are difficult to store. Although most of the plant extracts harmless to the human body are currently used in gels, the components of these plant extracts are mostly difficult to identify and difficult to store for a long time. SUMMARY

[0004] The present application provides an Olea europaea L. polysaccharide extract and a preparation method and application thereof, to solve the technical problem of how to improve the whitening ability of the gel in the case of being harmless to the human body.

[0005] In a first aspect, the present application provides an Olea europaea L. polysaccharide extract, wherein the monosaccharide composition of the Olea europaea L. polysaccharide extract is, in terms of mass fraction: mannose ≥ 5%, rhamnose ≥ 6%, glucuronic acid ≥ 3%, galacturonic acid ≥ 33%, glucose ≥ 6%, galactose ≥ 20% and arabinose ≥ 20%.

[0006] Optionally, the monosaccharide composition of the Olea europaea L. polysaccharide extract is, in terms of mass fraction: mannose: 5% to 7%, rhamnose: 6% to 7%, glucuronic acid: 3% to 4%, galacturonic acid: 33% to 35%, glucose: 6% to 8%, galactose: 20% to 24% and arabinose: 20% to 24%.

[0007] Optionally, the oil olive polysaccharide extract has a weight average molecular weight of 1100000±100000.

[0008] In a second aspect, the application provides a method for preparing the oil olive polysaccharide extract of the first aspect, the method comprising:

[0009] adsorbing impurities from the remaining liquid phase after oil extraction of the oil olive to obtain an adsorbed liquid phase;

[0010] filtering the adsorbed liquid phase to obtain a filtrate;

[0011] mixing the filtrate and anhydrous ethanol to extract polysaccharide components of the filtrate to obtain a mixed liquid;

[0012] performing first centrifugation on the mixed liquid, then resolubilizing and freeze-drying the centrifugal precipitate to obtain crude oil olive polysaccharide;

[0013] mixing a trichloroacetic acid solution and the crude oil olive polysaccharide and adjusting the pH to remove impurities to obtain an extracted liquid;

[0014] performing second centrifugation on the extracted liquid to obtain an extracted supernatant;

[0015] sequentially dialyzing and rotary-evaporating concentrating the extracted supernatant to obtain the oil olive polysaccharide extract.

[0016] Optionally, the first centrifugation is performed at a speed of 7800rpm-8200rpm for 5min-10min.

[0017] Optionally, the second centrifugation is performed at a speed of 9000rpm-11000rpm for 5min-10min.

[0018] Optionally, the crude oil olive polysaccharide has a mass concentration of ≥10mg / mL, the trichloroacetic acid solute in the trichloroacetic acid solution has a weight greater than or equal to 5% of the weight of the trichloroacetic acid solution; and / or

[0019] the volume V1 of the crude oil olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship V1:V2≥1:1.

[0020] In a third aspect, the application provides a gel having whitening and moisturizing functions, the gel comprising the oil olive polysaccharide extract of the first aspect and a liquorice extract.

[0021] Optionally, the gel has the following composition by mass fraction:

[0022] The oil olive polysaccharide extract is 0.2%-1%, the butanediol glycerol mixed solution is 20%, the triethanolamine is 1%, the carbomer is 1%, the propylene glycol is 10%, the liquorice extract is 20%, the citric acid is 1%, the rest is distilled water and inevitable impurities; wherein, the preparation method of the liquorice extract comprises:

[0023] Mixing ethanol and ammonia water to obtain a mixed solution containing ethanol and ammonia water;

[0024] Ultrasonic extraction of liquorice and the mixed solution to obtain an extract;

[0025] Rotary evaporation and freeze-drying of the extract to obtain a liquorice extract.

[0026] In a fourth aspect, the application provides a method for preparing the gel of the third aspect, the method comprising:

[0027] Mixing the oil olive polysaccharide extract and distilled water to obtain a polysaccharide solution;

[0028] Mixing the carbomer and the polysaccharide solution to make the carbomer adsorb the polysaccharide solution to obtain a mixture;

[0029] Mixing the propylene glycol, the butanediol glycerol mixed solution and the mixture to obtain a gel base solution;

[0030] Mixing the liquorice extract, the citric acid and distilled water to obtain a liquorice acid solution;

[0031] Mixing the triethanolamine, the gel base solution and the liquorice acid solution and adjusting the pH value to obtain the gel with the whitening and moisturizing functions.

[0032] The above technical solution provided by the embodiments of the application has the following advantages compared with the prior art:

[0033] The oil olive polysaccharide extract provided by the embodiment of the present application contains a monosaccharide structure composed of complex mannose, rhamnose, glucuronic acid, galacturonic acid, glucose, galactose and arabinose, which can be combined with licorice extract, based on the emulsification, stabilization and preservative effect of the licorice extract, the monosaccharide structure has good antioxidant and anti-inflammatory effects, can scavenge free radicals and reduce oxidative stress, so as to protect skin cells from damage; in addition, the monosaccharide structure and the licorice extract can both inhibit the activity of tyrosinase, so as to effectively reduce the generation of melanin, and at the same time, the monosaccharide structure and the licorice extract are harmless to the human body, so that the whitening ability of the gel can be improved without harm to the human body; therefore, the oil olive polysaccharide extract can more effectively reduce melanin deposition and improve skin spots based on the combination of the antioxidant and anti-inflammatory effects and the tyrosinase inhibition effect of the licorice extract, so as to achieve the effect of whitening. BRIEF DESCRIPTION OF DRAWINGS

[0034] The drawings incorporated into the specification and constituting a part of the specification show embodiments consistent with the present application and, together with the specification, serve to explain the principles of the present application.

[0035] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed to be used in the embodiments or the prior art description will be briefly introduced as follows, and obviously, other drawings can also be obtained by those skilled in the art without creative labor under the premise of not paying the creative labor.

[0036] Figure 1 A high-performance liquid chromatogram of an oil olive polysaccharide extract provided in Embodiment 2 of the present application is provided;

[0037] Figure 2 A process schematic diagram of a method for preparing an oil olive polysaccharide extract provided in the present application is provided;

[0038] Figure 3 A process schematic diagram of a preparation method of licorice extract provided in the present application is provided;

[0039] Figure 4 A process schematic diagram of a method for preparing the gel provided in the present application is provided;

[0040] Figure 5 A differential refractive index detection result diagram of the oil olive polysaccharide extract provided in Embodiment 2 of the present application is provided;

[0041] Figure 6 A GPC software calculation result diagram of the oil olive polysaccharide extract provided in Embodiment 2 of the present application is provided;

[0042] Figure 7The infrared detection result chart of the oil olive polysaccharide extract provided for the embodiment 2 of the present application is shown. DETAILED DESCRIPTION

[0043] In order to make the objects, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be described clearly and completely below with reference to the drawings in the embodiments of the present application. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments of the present application. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the protection scope of the present application.

[0044] Various embodiments of the present application can exist in the form of a range; it should be understood that the description in the form of a range is merely for the convenience and brevity, and should not be understood as a hard limit on the scope of the present application; therefore, it should be considered that the described range has disclosed all possible sub-ranges and single values in the range; for example, it should be considered that the range description from 1 to 6 has disclosed sub-ranges, such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6, etc., and single numbers in the range, such as 1, 2, 3, 4, 5 and 6, which is applicable regardless of the range; in addition, whenever a numerical range is indicated in the present application, it refers to any cited number (fraction or integer) in the indicated range.

[0045] In this document, the terms "comprises", "comprising", "includes", "including" or "contains", "containing" means "including, but not limited to". The terms "first", "second", "third", etc., are used only to distinguish one entity or action from another, and do not necessarily require or imply any actual relationship or order between or among such entities or actions. The term "and / or" describes the associated relationship of the associated objects, which means that there can be three kinds of relationships, for example, A and / or B, which can represent: the existence of A alone, the existence of A and B together, and the existence of B alone; wherein A and B can be singular or plural. "At least one" means one or more, "multiple" means two or more; "at least one" or "at least one of the following" or the like means any combination of the items, including single item or multiple items; for example, "at least one of a, b, or c", or "at least one of a, b, and c", can represent a, b, c, a-b (i.e. a and b), a-c, b-c, or a-b-c, wherein a, b, and c can be single or multiple. The "parts ratio" such as weight parts, mass parts, etc. represents the proportional relationship between components. In the proportional relationship described in this document, the parameters that need to be described by the order of description should be understood as the front item of the proportional formula, and the proportional number should be understood as the rear item of the proportional formula, for example, the mass ratio of substance A, substance B and substance C is 1:2:3, then substance A, substance B and substance C should be one-to-one corresponding in the proportional formula according to the description order, that is, the mass of substance A: the mass of substance B: the mass of substance C = 1:2:3.

[0046] Unless otherwise specifically stated, the various raw materials, reagents, instruments and equipment used in this document can be purchased from the market or can be prepared by existing methods.

[0047] Figure 1 Exemplarily, the application embodiment 2 provides a high performance liquid chromatography result graph of the olive polysaccharide extract;

[0048] As Figure 1 shown, the application embodiment provides an olive polysaccharide extract, the monosaccharide composition of the olive polysaccharide extract is: mannose ≥ 5%, rhamnose ≥ 6%, glucuronic acid ≥ 3%, galacturonic acid ≥ 33%, glucose ≥ 6%, galactose ≥ 20% and arabinose ≥ 20% by mass fraction.

[0049] In some optional embodiments, the monosaccharide composition of the olive polysaccharide extract is: mannose: 5%~7%, rhamnose: 6%~7%, glucuronic acid: 3%~4%, galacturonic acid: 33%~35%, glucose: 6%~8%, galactose: 20%~24% and arabinose: 20%~24% by mass fraction;

[0050] In these embodiments, the monosaccharide composition of the olive polysaccharide extract can be: mannose: 5% to 7%, rhamnose: 6% to 7%, glucuronic acid: 3% to 4%, galacturonic acid: 33% to 35%, glucose: 6% to 8%, galactose: 20% to 24%, and arabinose: 20% to 24%, prompting the olive polysaccharide extract to have a complex monosaccharide structure to improve the whitening ability of the gel in a way that is harmless to the human body.

[0051] The mass fraction of the mannose can be 5.0%, 5.5%, 6.0%, 6.5%, or 7.0%.

[0052] The mass fraction of the rhamnose can be 6.0%, 6.1%, 6.2%, 6.3%, 6.4%, 6.5%, 6.6%, 6.7%, 6.8%, 6.9%, or 7.0%.

[0053] The mass fraction of the glucuronic acid can be 3.0%, 3.1%, 3.2%, 3.3%, 3.4%, 3.5%, 3.6%, 3.7%, 3.8%, 3.9%, or 4.0%.

[0054] The mass fraction of the galacturonic acid can be 33.0%, 33.5%, 34.0%, 34.5%, or 35%.

[0055] The mass fraction of the glucose can be 6%, 6.5%, 7%, 7.5%, or 8%.

[0056] The mass fraction of the galactose can be 20.0%, 20.5%, 21.0%, 21.5%, 22.0%, 22.5%, 23.0%, 23.5%, or 24.0%.

[0057] The mass fraction of the arabinose can be 20.0%, 20.5%, 21.0%, 21.5%, 22.0%, 22.5%, 23.0%, 23.5%, or 24.0%.

[0058] In some alternative embodiments, the weight average molecular weight of the olive polysaccharide extract is 1100000 ± 100000;

[0059] In these embodiments, the weight average molecular weight of the olive polysaccharide extract can be 1100000 ± 100000, which indicates that the olive polysaccharide extract has a sufficient number of monosaccharide structures, and the synergistic effect between the sufficient number of monosaccharide structures and the licorice extract can improve the whitening ability of the gel in a way that is harmless to the human body.

[0060] Figure 2An exemplary flowchart of a method for preparing an olive polysaccharide extract is shown.

[0061] Based on a general inventive concept, as Figure 2 As shown, the method for preparing the olive polysaccharide extract comprises the following steps:

[0062] S1. Impurity adsorption is performed on the residual liquid phase after olive oil extraction to obtain an adsorbed liquid phase;

[0063] S2. Filtration is performed on the adsorbed liquid phase to obtain a filtrate;

[0064] S3. The filtrate and anhydrous ethanol are mixed to extract polysaccharide components of the filtrate, thereby obtaining a mixed liquid;

[0065] S4. First centrifugation is performed on the mixed liquid, and the centrifugal precipitate is then redissolved and freeze-dried to obtain crude olive polysaccharide;

[0066] S5. Trichloroacetic acid solution and the crude olive polysaccharide are mixed and adjusted in pH to remove impurities, thereby obtaining an extract;

[0067] S6. Second centrifugation is performed on the extract to obtain an extract supernatant;

[0068] S7. The extract supernatant is sequentially subjected to dialysis and rotary evaporation concentration to obtain an olive polysaccharide extract.

[0069] The method is directed to the above-described method for preparing an olive polysaccharide extract. The specific composition of the olive polysaccharide extract can refer to the above-described embodiments. Since the method uses part or all of the technical solutions of the above-described embodiments, it at least has all the beneficial effects brought by the technical solutions of the above-described embodiments, which will not be repeated here.

[0070] In some optional embodiments, the rotation speed of the first centrifugation is 7800 rpm to 8200 rpm, and the time of the first centrifugation is 5 min to 10 min.

[0071] In these embodiments, the rotation speed of the first centrifugation can be 7800 rpm to 8200 rpm, and the time of the first centrifugation can be 5 min to 10 min, so that the polysaccharide components of the mixed liquid can be fully separated from the liquid phase components, facilitating the subsequent redissolution and freeze-drying to obtain crude olive polysaccharide.

[0072] The rotation speed of the first centrifugation can be 7800 rpm, 7850 rpm, 7900 rpm, 7950 rpm, 8000 rpm, 8050 rpm, 8100 rpm, 8150 rpm, or 8200 rpm.

[0073] The time of the first centrifugation can be 5 min, 6 min, 7 min, 8 min, 9 min, or 10 min.

[0074] In some alternative embodiments, the speed of the second centrifugation is 9000 rpm to 11000 rpm, and the time of the second centrifugation is 5 min to 10 min.

[0075] In these embodiments, the speed of the second centrifugation can be 9000 rpm to 11000 rpm, and the time of the second centrifugation can be 5 min to 10 min, so that the liquid phase and the solid phase impurities of the extraction solution can be separated out sufficiently through the second centrifugation, thereby facilitating subsequent concentration by rotary evaporation to obtain the high-purity olive polysaccharide extract.

[0076] The speed of the second centrifugation can be 9000 rpm, 9100 rpm, 9200 rpm, 9300 rpm, 9400 rpm, 9500 rpm, 9600 rpm, 9700 rpm, 9800 rpm, 9900 rpm, 10000 rpm, 10100 rpm, 10200 rpm, 10300 rpm, 10400 rpm, 10500 rpm, 10600 rpm, 10700 rpm, 10800 rpm, 10900 rpm, or 11000 rpm.

[0077] The time of the second centrifugation can be 5 min, 6 min, 7 min, 8 min, 9 min, or 10 min.

[0078] In some alternative embodiments, the mass concentration of the crude olive polysaccharide is ≥10 mg / mL, the weight of the trichloroacetic acid solute of the trichloroacetic acid solution is greater than or equal to 5% of the weight of the trichloroacetic acid solution; and / or

[0079] The volume V1 of the crude olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship: V1:V2≥1:1.

[0080] In these embodiments, the mass concentration of the crude olive polysaccharide is ≥10 mg / mL, and the weight of the trichloroacetic acid solute of the trichloroacetic acid solution is greater than or equal to 5% of the weight of the trichloroacetic acid solution, in addition, the volume V1 of the crude olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship: V1:V2≥1:1, so that the crude olive polysaccharide has sufficient trichloroacetic acid, and the sufficient trichloroacetic acid can effectively remove the impurities of the crude olive polysaccharide, thereby facilitating subsequent second centrifugation to obtain the supernatant containing the pure olive polysaccharide extract.

[0081] Based on a general inventive concept, the embodiments of the present application provide a gel with whitening and moisturizing functions, which has the olive polysaccharide extract and the licorice extract.

[0082] The gel is realized based on the olive polysaccharide extract, and the specific composition of the olive polysaccharide extract can refer to the above embodiments. Since the gel adopts part or all of the technical solutions of the above embodiments, it at least has all the beneficial effects brought by the technical solutions of the above embodiments, which will not be repeated here.

[0083] Figure 3 An exemplary preparation method flowchart of the licorice extract provided by the embodiments of the present application is shown;

[0084] In some optional embodiments, the composition of the gel is as follows in terms of mass fraction:

[0085] Olive polysaccharide extract: 0.2% to 1%, butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, licorice extract: 20%, citric acid: 1%, and the rest is distilled water and unavoidable impurities; wherein, as shown in the table, the preparation method of the licorice extract includes: Figure 3

[0086] S1. mixing ethanol and ammonia water to obtain a mixed solution containing ethanol and ammonia water;

[0087] S2. ultrasonic extraction of licorice and the mixed solution to obtain an extract;

[0088] S3. rotary evaporation and freeze-drying of the extract to obtain a licorice extract;

[0089] In these embodiments, the gel uses the olive polysaccharide extract and the licorice extract as raw materials, and based on the antioxidant and anti-inflammatory effects of the olive polysaccharide extract, the combination of these effects and the tyrosinase inhibitory effect of the licorice extract can more effectively reduce the deposition of melanin to improve skin stains, thereby achieving a better whitening effect. In addition, the licorice extract also has flavonoids, so that the gel formed by the olive polysaccharide extract and the licorice extract has good sunscreen performance, so that the gel can effectively absorb ultraviolet rays to reduce the damage of ultraviolet rays to the skin, thereby improving the whitening ability of the gel.

[0090] ​It should be noted that the licorice extract can act as an emulsifier of the gel: the surface active ingredients in the licorice extract can reduce the interfacial tension of the oil-water interface, so that the oil phase and the water phase in the oleuropein polysaccharide extract can be better mixed to form a uniform gel system; this emulsification not only improves the uniformity of the gel, but also enhances its stability and sensory performance.

[0091] It should be noted that the licorice extract can act as a stabilizer of the gel: the licorice extract can stabilize the colloidal particles of the gel through hydrogen bonding and hydrophobic interaction, preventing their aggregation and precipitation; in addition, the antioxidant properties of the licorice extract can inhibit the oxidation reaction of the gel, further extending the shelf life of the gel.

[0092] It should be noted that the licorice extract can act as a preservative of the gel: the licorice extract contains various antibacterial ingredients, such as glycyrrhizic acid, flavonoids, etc., which can inhibit the growth and reproduction of microorganisms, thereby preventing the mold and spoilage of the gel.

[0093] It should be noted that the carbomer can be carbomer 941.

[0094] It should be noted that the weight ratio of butanediol and glycerol in the butanediol glycerol mixed solution is 1:4.

[0095] Figure 4 An exemplary flowchart of a method for preparing the gel provided by the embodiments of the present application is shown;

[0096] Based on a general inventive concept, as Figure 4 shown, the embodiments of the present application provide a method for preparing the gel, which comprises:

[0097] S1. mixing oleuropein polysaccharide extract and distilled water to obtain a polysaccharide solution;

[0098] S2. mixing carbomer and the polysaccharide solution so that the carbomer adsorbs the polysaccharide solution to obtain a mixture;

[0099] S3. mixing propylene glycol, butanediol glycerol mixed solution and the mixture to obtain a gel base solution;

[0100] S4. mixing licorice extract, citric acid and distilled water to obtain a glycyrrhizic acid solution;

[0101] S5. mixing triethanolamine, the gel base solution and the glycyrrhizic acid solution and adjusting the pH value to obtain a gel with whitening and moisturizing functions.

[0102] The method is for the preparation method of the gelatinous agent described above, and the specific composition of the gelatinous agent can refer to the above examples. Since the method adopts part or all of the technical solutions of the above examples, it at least has all the beneficial effects brought by the technical solutions of the above examples, which will not be repeated here.

[0103] It should be noted that there are hydrogen bonds and hydrophobic interactions between the polysaccharide molecules of the olive polysaccharide extract. During the dissolution process, these polysaccharide molecules will unfold and fully hydrate, and during the subsequent mixing process, these polysaccharide molecules will rearrange and connect with each other through hydrogen bonds to form a stable three-dimensional network structure, thereby imparting the gelatinous agent with unique elasticity and viscosity.

[0104] The application will be further described below in conjunction with specific examples. The experimental methods in the following examples are not specified, and the experimental methods are generally determined according to national standards / industry standards; if there is no corresponding national standard / industry standard, the general international standard, conventional conditions or the conditions suggested by the manufacturer are used. Example 1

[0105] As shown in Figure 2 , the application provides a method for preparing an olive polysaccharide extract, comprising:

[0106] S1. 500 mL of residual liquid phase after olive oil extraction is subjected to impurity adsorption using 500 g of DA201 macroporous adsorption resin to obtain an adsorption liquid phase; the impurity adsorption site is a conical flask, the impurity adsorption temperature is 30°C; the impurity adsorption is carried out under shaking conditions, the shaking speed is 50 r / min, and the shaking time is 6 h;

[0107] S2. The adsorption liquid phase is filtered to obtain a filtrate;

[0108] S3. The filtrate and anhydrous ethanol are mixed to prepare an ethanol solution with a mass concentration of 80%, and then the mixture is placed at 4°C overnight to extract the polysaccharide components of the filtrate, thereby obtaining a mixed solution;

[0109] S4. The mixed solution is subjected to first centrifugation at 4°C, and the centrifugal precipitate is then redissolved and freeze-dried to obtain a crude olive polysaccharide;

[0110] S5. A trichloroacetic acid solution and the crude olive polysaccharide are mixed, and then the mixture is placed at 4°C overnight, and then the pH of the mixed solution is adjusted to neutral using a sodium hydroxide solution with a molar concentration of 6 mol / L to remove impurities, thereby obtaining an extraction liquid;

[0111] S6. The extraction liquid is subjected to second centrifugation to obtain an extraction supernatant;

[0112] S7. The extraction supernatant was sequentially subjected to dialysis (dialysis for 48 h using a 3500 Da dialysis bag) and rotary evaporation concentration to obtain the olive polysaccharide extract.

[0113] The first centrifugation speed was 8000 rpm, and the first centrifugation time was 10 min.

[0114] The second centrifugation speed was 10000 rpm, and the second centrifugation time was 10 min.

[0115] The mass concentration of the crude olive polysaccharide was 10 mg / mL, and the weight of the trichloroacetic acid solute of the trichloroacetic acid solution was 5% of the weight of the trichloroacetic acid solution.

[0116] The volume V1 of the crude olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfied the relationship V1:V2=1:1. Example 2

[0117] On the basis of the olive polysaccharide extract disclosed in Example 1, the olive polysaccharide extract was detected:

[0118] I. Molecular weight detection of the olive polysaccharide extract:

[0119] 1. Chromatographic conditions: chromatographic column: Shodex OHpak SB-806HQ; mobile phase: water; flow rate: 0.7 mL / min; detector: RID detector; column oven: 35°C; run time: 25 min;

[0120] 2. Sample preparation: the olive polysaccharide extract was prepared into a solution with a mass concentration of 5 mg / mL using water, and then the prepared solution was filtered through a filter membrane with a pore size of 0.45 μm.

[0121] The sample was detected using the above chromatography, and the results are shown in Figure 5 and Table 1.

[0122] Table 1: Measurement results of detector A channel

[0123] ,

[0124] The detected results were automatically calculated by GPC software, and the results are shown in Figure 6 and Table 2, and the weight average molecular weight Mw is 1100000±100000.

[0125] Table 2: Calculation results of GPC software

[0126] ,

[0127] Based on the results of this test, a standard curve for the test is plotted, and its formula is: f(x) = -0.0315597 * x 3 +0.949904*x-9.465083*x+37.30616 (R 2 =0.9805728).

[0128] II. Determination of Monosaccharide Composition:

[0129] 1. Hydrolysis of crude olive polysaccharide extract:

[0130] The crude olive polysaccharide extract was prepared into a sample solution with a mass concentration of 5 mg / mL using a 2 mol / L trifluoroacetic acid solution. The sample solution was then placed into ampoules in groups of 4 mL, purged with nitrogen, and sealed. The solution was then hydrolyzed at 105 °C for 6 h. The pH was then adjusted to neutral using a 6 mol / L sodium hydroxide solution. 0.5 mL of the neutral hydrolysate was then taken and 0.5 mL of a 0.25 mol / L sodium hydroxide solution and 0.5 mL of a 0.4 mol / L PMP-methanol solution were added sequentially and mixed thoroughly.

[0131] The reaction was carried out in an oven at 70°C for 90 min. Then, the dried material was removed and placed in an ice water bath for 10 min. After that, the dried material was allowed to return to room temperature. Then, 0.5 mL of 0.3 mol / L hydrochloric acid solution was added and mixed well. Finally, PMP was extracted three times with dichloromethane. The first extraction was carried out overnight, and the interval between the next two extractions was 2 h to 3 h.

[0132] 2. High Performance Liquid Chromatography (HPLC) Detection Procedure:

[0133] Column: Diamonsil C18, 250×4.6mm, 5μm; Mobile phase: 0.05mol / L potassium dihydrogen phosphate (potassium dihydrogen phosphate and acetonitrile volume ratio 20:80); Flow rate: 1mL / min; Detector: DAD UV detection wavelength 250nm; Column oven: 35℃; Run time: 80min.

[0134] Test results as follows Figure 1 As shown, the ultraviolet detection results are as follows: Figure 7 As shown.

[0135] according to Figure 1 and Figure 7 Calculations show that the monosaccharide composition of the olive polysaccharide extract is as follows: mannose: 5%–7%, rhamnose: 6%–7%, glucuronic acid: 3%–4%, galacturonic acid: 33%–35%, glucose: 6%–8%, galactose: 20%–24%, and arabinose: 20%–24%. Example 3

[0136] On the basis of the specific monosaccharide composition of the olive polysaccharide extract determined in Example 2, further preparation of the gel is carried out:

[0137] The composition of the gel is, by mass fraction: olive polysaccharide extract: 5%, butanediol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, liquorice extract: 20%, citric acid: 1%, and the rest is distilled water and unavoidable impurities; wherein, as shown in the following formula (I), the preparation method of the liquorice extract comprises: Figure 3

[0138] S1. mixing ethanol and ammonia water to obtain a mixed solution containing ethanol and ammonia water; wherein, the mass ratio of ethanol and ammonia water is 0.75:0.03.

[0139] S2. ultrasonic extraction of liquorice and the mixed solution for 25 min to obtain an extraction liquid;

[0140] S3. rotary evaporation and freeze-drying of the extraction liquid to obtain a liquorice extract.

[0141] As shown in the following formula (II), a method for preparing a gel comprises: Figure 4

[0142] S1. mixing olive polysaccharide extract and distilled water to obtain a polysaccharide solution;

[0143] S2. mixing carbomer and the polysaccharide solution, and then slowly stirring at 70°C to make the carbomer adsorb the polysaccharide solution and swell, to obtain a mixture;

[0144] S3. mixing propylene glycol, butanediol glycerol mixed solution and the mixture at 70°C to obtain a gel base solution;

[0145] S4. mixing liquorice extract, citric acid and distilled water at 55°C to obtain a liquorice acid solution;

[0146] S5. first cooling the gel base solution to 55°C, and then mixing triethanolamine, the gel base solution and the liquorice acid solution and adjusting the pH to 6-8 to obtain a gel with whitening and moisturizing functions. Comparative Example 1

[0147] On the basis of the disclosure in Example 3, the following modifications are further carried out:

[0148] ​​Without using the polysaccharide extract of Olea europaea L. and the extract of Glycyrrhiza uralensis Fisch., only using the stabilizer instead, a gel is obtained (its components are, in mass fraction: disodium ethylenediaminetetraacetate: 0.4%, butanediol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, citric acid: 1%, the rest being distilled water and unavoidable impurities). Comparative Example 2

[0149] On the basis of the disclosure of Example 3, the following modifications are further made:

[0150] Without using the polysaccharide extract of Olea europaea L., a gel is obtained, and its components are: butanediol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, extract of Glycyrrhiza uralensis Fisch.: 20%, citric acid: 1%, the rest being distilled water and unavoidable impurities. Comparative Example 3

[0151] On the basis of the disclosure of Example 3, the following modifications are further made:

[0152] Without using the extract of Glycyrrhiza uralensis Fisch., a gel is obtained, and its components are: polysaccharide extract of Olea europaea L.: 1%, butanediol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, extract of Glycyrrhiza uralensis Fisch.: 20%, citric acid: 1%, the rest being distilled water and unavoidable impurities.

[0153] Related experiments and effect data:

[0154] 1. Stability test of the gel:

[0155] All examples and comparative examples are subjected to stability test, and the test method is:

[0156] (1) Centrifugal stability: 2.0 g of the sample to be tested is weighed into a centrifuge tube, and then centrifuged at 4000 r / min, 8000 r / min and 12000 r / min, respectively, and the state of the gel after centrifugation is observed.

[0157] (2) Thermal stability: 2.0 g of the sample to be tested is weighed into a centrifuge tube, and then incubated at 40℃ for 24 h, and after taking out, whether turbidity, stratification and other phenomena occur is observed.

[0158] (3) Cold stability: 2.0 g of the sample to be tested is weighed into a centrifuge tube, and then incubated at -20℃ for 24 h, and after taking out, whether turbidity, stratification and other phenomena occur is observed.

[0159] (4) Cold and hot cycle stability: 2.0 g of the sample to be tested was weighed into a centrifuge tube, and was kept at -20°C for 12 h, then was kept at 40°C for 12 h. The above operation was repeated twice, and whether turbidity, delamination or other phenomena occurred was observed.

[0160] (5) Sample observation: 2.0 g of the sample was weighed into a centrifuge tube, and was kept at room temperature for one month, and whether turbidity, delamination or other phenomena occurred was observed.

[0161] 2. Moisture retention capacity test of the gel:

[0162] The moisture retention capacity of the obtained gel was determined by using a weighing method: 5 g of each of the moisturizing gels of the experimental examples was weighed, then pure water was added in a weight ratio of 1:1 to shake and mix uniformly, and then the mixture was loaded into a weighing tank which had been dried to a constant weight in advance. Under room temperature conditions, it was placed into a desiccator containing saturated ammonium sulfate solution (mass concentration of 81%), saturated potassium carbonate solution (mass concentration of 43%) and dry silica gel to simulate different humidity environments, and the mass was weighed again after 1 h, 2 h, 4 h, 6 h, 8 h, 12 h and 24 h. Three parallel samples were determined under the same conditions, and the average value was taken, and the formula was as follows:

[0163] ,

[0164] In the formula, M1 is the mass of the sample when it is loaded; and M2 is the mass of the sample when it is taken out.

[0165] 3. Whitening capacity test of the gel:

[0166] (1) Preparation of detection reagent:

[0167] Preparation of tyrosinase solution with a mass concentration of 100 μg / mL: 10.34 mg of tyrosinase was accurately weighed, and was dissolved in a 10 mL centrifuge tube using PBS buffer with a pH of 6.8. The dissolved solution was transferred to a 100 mL volumetric flask, and was diluted to the calibration line with PBS buffer, to obtain a tyrosinase solution with a mass concentration of 100 μg / mL, which was prepared for use.

[0168] Preparation of L-tyrosine solution with a molar concentration of 2.5 mmol / L: 45.3 mg of tyrosine was accurately weighed, and was dissolved in a 10 mL centrifuge tube using PBS buffer with a pH of 6.8. The dissolved solution was transferred to a 100 mL volumetric flask, and was diluted to the calibration line with PBS buffer, to obtain an L-tyrosine solution with a molar concentration of 2.5 mmol / L, which was prepared for use.

[0169] (2) Test method:

[0170] To centrifuge tube, respectively, add 1 mL of the gel prepared in Example 3 and each of the comparative examples, then add 1 mL of a tyrosinase solution having a mass concentration of 100 μg / mL, 1.8 mL of a PBS solution, and mix the added solutions, and then incubate in an incubator at 37°C for 10 min. Then add 1.2 mL of an L-tyrosine solution having a molar concentration of 2.5 mmol / L, and again incubate in an incubator at 37°C for 10 min. Immediately after the incubation, take samples and add to a 96-well plate, and measure the absorbance at 475 nm three times in parallel for each sample, and then calculate the inhibition rate according to the following formula:

[0171]

[0172] In the formula, A1 is the absorbance of the sample well; A2 is the absorbance with PBS instead of tyrosinase; A3 is the absorbance with PBS instead of the sample; and A4 is the absorbance with PBS instead of the sample and tyrosinase.

[0173] 4 Test results are as follows:

[0174] (1) Stability test results

[0175] Centrifugal stability: None of Examples 3 and Comparative Examples 1 to 3 showed turbidity, stratification, flocculation, and the like under three centrifugal conditions, indicating that the product has good stability under accelerated conditions.

[0176] Thermal stability: None of Examples 3 and Comparative Examples 1 to 3 showed turbidity, stratification, flocculation, and the like under high-temperature conditions, indicating that the product has good stability under high-temperature conditions.

[0177] Cold stability: None of Examples 3 and Comparative Examples 1 to 3 showed turbidity, stratification, crystallization, and the like under low-temperature conditions, indicating that the product has good stability under low-temperature conditions.

[0178] Cold-heat cycle stability: None of Examples 3 and Comparative Examples 1 to 3 showed special phenomena after cold-heat cycling, indicating that the product has good stability under cold-heat cycling conditions.

[0179] Sample observation: Examples 3 and Comparative Examples 1 to 3 were observed again after standing for one month, and none showed turbidity, stratification, deterioration, dryness, or solidification, indicating that the product can be stored for a long time at room temperature.

[0180] (2) Moisturizing ability test results

[0181] The water loss rate (%) under a relative humidity of 43% is shown in Table 3.

[0182] Table 3: Results of water loss rate of each example and comparative example under a relative humidity of 43% ​

[0183] ,

[0184] The water loss rate (%) under the condition of relative humidity 81% is shown in Table 4.

[0185] Table 4 Water loss rate result table of each example and comparative example under the condition of relative humidity 81%

[0186] ,

[0187] The whitening ability test results are shown in Table 5.

[0188] Table 5 Whitening ability result table of each example and comparative example

[0189] ,

[0190] In summary, the olive polysaccharide extract provided by the embodiments of the present application can effectively reduce melanin deposition and improve skin spots by combining the antioxidant and anti-inflammatory effects of the olive polysaccharide extract with the tyrosinase inhibiting effect of the licorice extract, thereby achieving the effect of whitening.

[0191] In addition, the gel provided by the embodiments of the present application has the functions of whitening and moisturizing. The main components of the olive polysaccharide extract and the licorice extract used in the gel are derived from natural extracts, and have no irritation to the skin, high safety, and no adverse reactions. In addition, the interaction between the olive polysaccharide extract and the licorice extract can not only improve the whitening effect of the gel, but also improve the water-locking and moisturizing effect of the gel. In addition, the olive polysaccharide extract and the licorice extract can also synergize with traditional moisturizing agents to further improve the water-locking and moisturizing effect of the gel.

[0192] In addition, the gel provided by the embodiments of the present application has the functions of whitening and moisturizing. The licorice extract used in the gel can also act as an emulsifier, a stabilizer and a preservative of the gel system, and can simultaneously enhance the stability and the anti-mildew ability of the gel. In addition, the gel has antioxidant and anti-inflammatory auxiliary effects based on the antioxidant and anti-inflammatory effects of the olive polysaccharide extract, and has development advantages when used as a whitening and moisturizing cosmetic.

[0193] In addition, the method for preparing the gel provided by the embodiments of the present application is simple to operate, and the preparation conditions are mild, so that the gel product with the best efficacy can be obtained.

[0194] The foregoing detailed description of the application has been presented for purposes of illustration and description. Various modifications to the description will be readily apparent to those skilled in the art, and the generic principles defined herein can be applied to other implementations without departing from the spirit or scope of the application. Accordingly, the application is not intended to be limited to the implementations described herein but is to be accorded the widest scope consistent with the principles and novel features to the application.

Claims

1. A gel having a whitening and moisturizing function, the gel having an Olea europaea L. polysaccharide extract and a Glycyrrhiza extract. The preparation method of the oil olive polysaccharide comprises the following steps: After the oil olive is pressed, the remaining liquid phase is subjected to impurity adsorption to obtain an adsorbed liquid phase; The adsorbed liquid phase is filtered to obtain a filtrate; The filtrate and anhydrous ethanol are mixed to extract polysaccharide components of the filtrate, and a mixed liquid is obtained; The mixed liquid is subjected to first centrifugation, and the centrifugal precipitate is redissolved and freeze-dried to obtain crude oil olive polysaccharide; Trichloroacetic acid solution and the crude oil olive polysaccharide are mixed and adjusted in pH to remove impurities, and an extraction liquid is obtained; The extraction liquid is subjected to second centrifugation to obtain an extraction supernatant; The extraction supernatant is subjected to dialysis and rotary evaporation concentration in sequence to obtain oil olive polysaccharide extract; The preparation method of the liquorice extract comprises the following steps: Ethanol and ammonia water are mixed to obtain a mixed solution containing ethanol and ammonia water; Liquorice and the mixed solution are subjected to ultrasonic extraction to obtain an extraction liquid; The extraction liquid is subjected to rotary evaporation and freeze-drying to obtain liquorice extract.

2. The gel of claim 1, wherein, The monosaccharide composition of the oil olive polysaccharide extract comprises: mannose ≥ 5%, rhamnose ≥ 6%, glucuronic acid ≥ 3%, galacturonic acid ≥ 33%, glucose ≥ 6%, galactose ≥ 20% and arabinose ≥ 20%.

3. The gel of claim 1, wherein, The monosaccharide composition of the oil olive polysaccharide extract comprises, in terms of mass fraction: mannose: 5% to 7%, rhamnose: 6% to 7%, glucuronic acid: 3% to 4%, galacturonic acid: 33% to 35%, glucose: 6% to 8%, galactose: 20% to 24% and arabinose: 20% to 24%.

4. The gel of claim 1, wherein, The weight average molecular weight of the oil olive polysaccharide extract is 1100000 ± 100000.

5. The gel of claim 1, wherein, The rotation speed of the first centrifugation is 7800 rpm to 8200 rpm, and the time of the first centrifugation is 5 min to 10 min.

6. The gel of claim 1, wherein, The rotation speed of the second centrifugation is 9000 rpm to 11000 rpm, and the time of the second centrifugation is 5 min to 10 min.

7. The gel of claim 1, wherein, The mass concentration of the crude oil olive polysaccharide is ≥ 10 mg / mL, the weight of trichloroacetic acid solute in the trichloroacetic acid solution is greater than or equal to 5% of the weight of the trichloroacetic acid solution; and / or The volume V1 of the crude oil olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship: V1:V2 ≥ 1:

1.

8. The gel of claim 1, wherein, The composition of the gel, in terms of mass fraction, comprises: Oil olive polysaccharide extract: 0.2% to 1%, butanediol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, liquorice extract: 20%, citric acid: 1%, and the rest is distilled water and unavoidable impurities.

9. A method for preparing the gel of any one of claims 1 to 8, the method comprising: Oil olive polysaccharide extract and distilled water are mixed to obtain a polysaccharide solution; Carbomer and the polysaccharide solution are mixed to enable the carbomer to adsorb the polysaccharide solution, and a mixture is obtained; Propylene glycol, butanediol glycerol mixed solution and the mixture are mixed to obtain a gel base solution; Liquorice extract, citric acid and distilled water are mixed to obtain liquorice acid solution; Mixing triethanolamine, the gel base solution and the glycyrrhizic acid solution and adjusting the pH value to obtain a gel with whitening and moisturizing functions.

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