Olive polysaccharide extract as well as preparation method and application thereof

Abstract

The invention relates to the technical field of biology, in particular to an olive polysaccharide extract as well as a preparation method and application thereof. The monosaccharide of the olea europaea polysaccharide extract comprises the following components in percentage by mass: more than or equal to 5% of mannose, more than or equal to 6% of rhamnose, more than or equal to 3% of glucuronic acid, more than or equal to 33% of galacturonic acid, more than or equal to 6% of glucose, more than or equal to 20% of galactose and more than or equal to 20% of arabinose. The olive polysaccharide extract is combined with the tyrosinase inhibiting effect of the licorice extract based on the antioxidant and anti-inflammatory effects, melanin deposition can be more effectively reduced, skin stains are improved, and therefore the whitening effect is achieved.

Description

Technical Field

[0001] The present application relates to the field of biotechnology, and in particular to an olive polysaccharide extract and a preparation method and application thereof. Background Art

[0002] Gels refer to thick liquid or semi-solid preparations made of drugs and excipients that can form gels, such as solutions, suspensions or emulsions. Generally, gels contain gel-type matrices, which belong to single-phase dispersion systems. There are aqueous and oily types. At present, aqueous gels are more commonly used in clinical practice. Aqueous gels have the characteristics of easy spreading and washing. In addition, aqueous gels are non-greasy and can effectively absorb tissue exudates. Therefore, aqueous gels generally do not hinder the normal physiological activities of the skin. These characteristics allow the drugs contained in aqueous gels to be released quickly, but aqueous gels have poor lubrication and are prone to losing moisture and mildew. Therefore, stabilizers and preservatives are often added. At present, commonly used preservatives are Nepal gold methyl ester, and commonly used stabilizers are disodium ethylenediaminetetraacetic acid. Although the usage of these stabilizers and preservatives is small, they are all chemically synthesized drugs, and large-scale use still has certain damage to the human body.

[0003] In the typical application field of gels, cosmetics, commonly used whitening drugs include tyrosine kinase inhibitors, hydroquinones, vitamin C, flavonoids and polyphenols, etc. However, tyrosinase inhibitors and hydroquinones are highly irritating to human skin, while vitamin C, flavonoids and polyphenols are easily oxidized in the air environment and difficult to preserve; although most plant extracts that are harmless to the human body are currently used in gels, the components of most of these plant extracts are difficult to identify and difficult to preserve for a long time. Summary of the invention

[0004] The present application provides an olive polysaccharide extract and a preparation method and application thereof to solve the following technical problem: how to improve the whitening ability of the gel without causing harm to the human body.

[0005] In a first aspect, the present application provides an olive polysaccharide extract, wherein the monosaccharide composition of the olive polysaccharide extract is, by 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 olive polysaccharide extract is, by 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 weight average molecular weight of the olive polysaccharide extract is 1100000±100000.

[0008] In a second aspect, the present application provides a method for preparing the olive polysaccharide extract according to the first aspect, the method comprising: The remaining liquid phase after olive oil extraction is subjected to impurity adsorption to obtain an adsorbed liquid phase; Filtering the adsorbed liquid phase to obtain a filtrate; Mixing the filtrate with anhydrous ethanol to extract the polysaccharide components of the filtrate to obtain a mixed solution; The mixed solution is first centrifuged, and then the centrifugal precipitate is redissolved and freeze-dried to obtain crude olive polysaccharide; The trichloroacetic acid solution and the crude olive polysaccharide are mixed and the pH is adjusted to remove impurities to obtain an extract; The extract is subjected to a second centrifugation to obtain an extraction supernatant; The extraction supernatant is dialyzed and concentrated by rotary evaporation in sequence to obtain an olive polysaccharide extract.

[0009] Optionally, 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.

[0010] Optionally, 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.

[0011] Optionally, the mass concentration of the crude olive polysaccharide is ≥10 mg / mL, and the weight of the 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 olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship: V1:V2≥1:1.

[0012] In a third aspect, the present application provides a gel with whitening and moisturizing functions, wherein the gel comprises the olive polysaccharide extract and licorice extract described in the first aspect.

[0013] Optionally, the composition of the gelling agent is, by mass fraction: Olive polysaccharide extract: 0.2%-1%, butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, licorice extract: 20%, citric acid: 1%, and the rest are distilled water and inevitable impurities; wherein the preparation method of the licorice extract comprises: Mixing ethanol and aqueous ammonia to obtain a mixed solution containing ethanol and aqueous ammonia; subjecting licorice and the mixed solution to ultrasonic extraction to obtain an extract; The extract is subjected to rotary evaporation and freeze drying to obtain a liquorice extract.

[0014] In a fourth aspect, the present application provides a method for preparing the gel according to the third aspect, the method comprising: Mixing the olive polysaccharide extract and distilled water to obtain a polysaccharide solution; Mixing carbomer and the polysaccharide solution so that the carbomer absorbs the polysaccharide solution to obtain a mixture; Mixing a mixed solution of propylene glycol and butylene glycol and glycerol with the mixture to obtain a gel matrix solution; Mixing licorice extract, citric acid and distilled water to obtain glycyrrhizic acid solution; The triethanolamine, the gel matrix solution and the glycyrrhizic acid solution are mixed and the pH is adjusted to obtain a gel with whitening and moisturizing functions.

[0015] The above technical solution provided by the embodiment of the present application has the following advantages compared with the prior art: The present application provides an olive polysaccharide extract, which contains a complex monosaccharide structure composed of mannose, rhamnose, glucuronic acid, galacturonic acid, glucose, galactose and arabinose. The monosaccharide structure can be combined with a licorice extract. Based on the emulsification, stabilization and preservative effects of the licorice extract, the monosaccharide structure has good antioxidant and anti-inflammatory effects, can scavenge free radicals and reduce oxidative stress to protect skin cells from damage; in addition, both the monosaccharide structure and the licorice extract can inhibit the activity of tyrosinase to effectively reduce the formation of melanin, and at the same time, the monosaccharide structure and the licorice extract are harmless to the human body, thereby improving the whitening ability of the gel without harming the human body; therefore, the olive polysaccharide extract can more effectively reduce melanin deposition and improve skin spots based on the antioxidant and anti-inflammatory effects combined with the tyrosinase inhibitory effect of the licorice extract, thereby achieving a whitening effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] The accompanying drawings, which are incorporated in and constitute a part of this specification, illustrate embodiments consistent with the present application and, together with the description, serve to explain the principles of the present application.

[0017] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative labor.

[0018] Figure 1 A high performance liquid chromatography result diagram of an olive polysaccharide extract is provided for Example 2 of the present application; Figure 2 A schematic flow chart of a method for preparing an olive polysaccharide extract provided in an embodiment of the present application; Figure 3 A schematic diagram of the preparation method of the licorice extract provided in the embodiment of the present application; Figure 4 A schematic flow chart of a method for preparing the gel provided in an embodiment of the present application; Figure 5 This is a graph showing the results of differential refraction detection of the olive polysaccharide extract provided in Example 2 of the present application; Figure 6 This is a graph showing the calculation results of the GPC software for the olive polysaccharide extract provided in Example 2 of the present application; Figure 7 This is a graph showing the infrared detection results of the olive polysaccharide extract provided in Example 2 of the present application. DETAILED DESCRIPTION

[0019] In order to make the purpose, technical solution and advantages of the embodiments of the present application clearer, the technical solution in the embodiments of the present application will be clearly and completely described below in conjunction with the drawings in the embodiments of the present application. Obviously, the described embodiments are part of the embodiments of the present application, not all of the embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of this application.

[0020] Various embodiments of the present application may be presented in the form of a range; it should be understood that the description in the form of a range is only for convenience and brevity and should not be understood as a rigid limitation on the scope of the present application; therefore, it should be considered that the range description has specifically disclosed all possible sub-ranges and single numerical values ​​within the range; for example, it should be considered that the range description from 1 to 6 has specifically 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., as well as single numbers within the range, such as 1, 2, 3, 4, 5 and 6, which applies regardless of the range; in addition, whenever a numerical range is indicated in this document, it is meant to include any cited numbers (fractions or integers) within the indicated range.

[0021] In this document, the terms including "including" and "including" mean "including but not limited to". Relational terms such as "first" and "second" are used only to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. "And / or" describes the association relationship of associated objects, indicating that there can be three relationships. For example, A and / or B can mean: A exists alone, A and B exist at the same time, and B exists alone; where A and B can be singular or plural. "At least one" means one or more, "plurality" means two or more; "at least one", "at least one of the following" or similar expressions refer to any combination of these items, including any combination of single or plural items; for example, "at least one of a, b, or c", or "at least one of a, b, and c" can all mean: a, b, c, ab (i.e. a and b), ac, bc, or abc, where a, b, and c can be single or plural, respectively. "Parts" such as parts by weight and parts by mass indicate the proportional relationship between the components. In the proportional relationship involved in this article, the parameters that need to be described by proportion should be understood as the first term of the proportional formula in the order of description, and the proportional numbers should be understood as the second term of the proportional formula. For example, if the mass ratio of substance A, substance B and substance C is 1:2:3, then substance A, substance B and substance C should correspond to the proportional numbers in the proportional formula in the order of description, that is, the mass of substance A: the mass of substance B: the mass of substance C = 1:2:3.

[0022] Unless otherwise specified, various raw materials, reagents, instruments and equipment used in this article can be purchased from the market or prepared by existing methods.

[0023] Figure 1 The high performance liquid chromatography result diagram of an olive polysaccharide extract provided by Example 2 of the present application is exemplarily shown; like Figure 1 As shown, the embodiment of the present application provides an olive polysaccharide extract, and the monosaccharide composition of the olive polysaccharide extract is, by mass fraction, mannose ≥5%, rhamnose ≥6%, glucuronic acid ≥3%, galacturonic acid ≥33%, glucose ≥6%, galactose ≥20% and arabinose ≥20%.

[0024] In some optional embodiments, the monosaccharide composition of the olive polysaccharide extract is, by 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%; 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%, so that the olive polysaccharide extract has a complex monosaccharide structure to improve the whitening ability of the gel without being harmful to the human body.

[0025] The mass fraction of mannose may be 5.0%, 5.5%, 6.0%, 6.5% or 7.0%.

[0026] The mass fraction of rhamnose may be 6.0%, 6.1%, 6.2%, 6.3%, 6.4%, 6.5%, 6.6%, 6.7%, 6.8%, 6.9% or 7.0%.

[0027] The mass fraction of 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%.

[0028] The mass fraction of galacturonic acid can be 33.0%, 33.5%, 34.0%, 34.5% or 35%. The mass fraction of glucose can be 6%, 6.5%, 7%, 7.5% or 8%.

[0029] The mass fraction of galactose may be 20.0%, 20.5%, 21.0%, 21.5%, 22.0%, 22.5%, 23.0%, 23.5% or 24.0%.

[0030] The mass fraction of arabinose may be 20.0%, 20.5%, 21.0%, 21.5%, 22.0%, 22.5%, 23.0%, 23.5% or 24.0%.

[0031] In some optional embodiments, the weight average molecular weight of the olive polysaccharide extract is 1100000±100000; 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 enough monosaccharide structures, and the synergistic effect between the sufficient monosaccharide structures and the licorice extract can improve the whitening ability of the gel without causing harm to the human body.

[0032] Figure 2 A schematic flow chart of a method for preparing an olive polysaccharide extract provided in an embodiment of the present application is exemplarily shown; Based on a general inventive concept, such as Figure 2As shown, the present application embodiment provides a method for preparing the olive polysaccharide extract, the method comprising: S1. The remaining liquid phase after olive oil extraction is subjected to impurity adsorption to obtain an adsorbed liquid phase; S2. filtering the adsorbed liquid phase to obtain a filtrate; S3. mixing the filtrate and anhydrous ethanol to extract the polysaccharide component of the filtrate to obtain a mixed solution; S4. The mixed solution is first centrifuged, and then the centrifugal precipitate is redissolved and freeze-dried to obtain crude olive polysaccharide; S5. The trichloroacetic acid solution and the crude olive polysaccharide are mixed and the pH is adjusted to remove impurities to obtain an extract; S6. The extract is subjected to a second centrifugation to obtain an extract supernatant; S7. The extracted supernatant is dialyzed and concentrated by rotary evaporation in sequence to obtain an olive polysaccharide extract.

[0033] This method is a method for preparing the above-mentioned olive polysaccharide extract. The specific composition of the olive polysaccharide extract can refer to the above-mentioned embodiment. Since this method adopts part or all of the technical solutions of the above-mentioned embodiment, it at least has all the beneficial effects brought by the technical solutions of the above-mentioned embodiment, which will not be repeated here one by one.

[0034] In some optional embodiments, the first centrifugation speed is 7800 rpm to 8200 rpm, and the first centrifugation time is 5 min to 10 min; In these embodiments, the speed of the first centrifugation can be 7800rpm~8200rpm, and the time of the first centrifugation can be 5min~10min, so that the polysaccharide components of the mixed solution can be fully separated from the liquid phase components, facilitating the subsequent re-dissolution and freeze-drying to obtain crude olive polysaccharide.

[0035] 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.

[0036] The first centrifugation time can be 5 min, 6 min, 7 min, 8 min, 9 min or 10 min.

[0037] In some optional embodiments, the second centrifugation speed is 9000 rpm to 11000 rpm, and the second centrifugation time is 5 min to 10 min; In these embodiments, the rotation speed of the second centrifugation can be 9000rpm~11000rpm, and the time of the second centrifugation can be 5min~10min. The liquid phase and solid phase impurities of the extract can be fully separated by the second centrifugation, thereby facilitating the subsequent rotary evaporation concentration to obtain a high-purity olive polysaccharide extract.

[0038] The rotation speed of the second centrifugation can be 9000rpm, 9100rpm, 9200rpm, 9300rpm, 9400rpm, 9500rpm, 9600rpm, 9700rpm, 9800rpm, 9900rpm, 10000rpm, 10100rpm, 10200rpm, 10300rpm, 10400rpm, 10500rpm, 10600rpm, 10700rpm, 10800rpm, 10900rpm or 11000rpm.

[0039] The second centrifugation time can be 5 min, 6 min, 7 min, 8 min, 9 min or 10 min.

[0040] In some optional embodiments, the mass concentration of the crude olive polysaccharide is ≥10 mg / mL, and the weight of the 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 olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship: V1:V2≥1:1.

[0041] In these embodiments, the mass concentration of crude olive polysaccharide is ≥10 mg / mL, and 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. 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 a sufficient amount of trichloroacetic acid, and the sufficient amount of trichloroacetic acid can effectively remove impurities from the crude olive polysaccharide, thereby facilitating the subsequent second centrifugation to obtain a pure supernatant containing an olive polysaccharide extract.

[0042] Based on a general inventive concept, an embodiment of the present application provides a gel with whitening and moisturizing functions, wherein the gel has the olive polysaccharide extract and the licorice extract.

[0043] The gel is realized based on the above-mentioned olive polysaccharide extract. The specific composition of the olive polysaccharide extract can be referred to the above-mentioned embodiment. Since the gel adopts part or all of the technical solutions of the above-mentioned embodiments, it has at least all the beneficial effects brought by the technical solutions of the above-mentioned embodiments, which will not be described one by one here.

[0044] Figure 3 The schematic diagram of the preparation method of the licorice extract provided in the embodiment of the present application is exemplarily shown; In some optional embodiments, the composition of the gel is, by mass fraction: Olive polysaccharide extract: 0.2%-1%, butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, licorice extract: 20%, citric acid: 1%, the rest is distilled water and inevitable impurities; among which, Figure 3 As shown, the preparation method of the licorice extract comprises: S1. mixing ethanol and aqueous ammonia to obtain a mixed solution containing ethanol and aqueous ammonia; S2. subjecting licorice and the mixed solution to ultrasonic extraction to obtain an extract; S3. The extract was subjected to rotary evaporation and freeze-drying to obtain a licorice extract; In these embodiments, the gel uses olive polysaccharide extract and licorice extract as raw materials. Based on the antioxidant and anti-inflammatory effects of the olive polysaccharide extract, through the combination of these effects and the inhibitory effect of the licorice extract on tyrosinase, the deposition of melanin can be more effectively reduced to improve skin pigmentation, thereby achieving a better whitening effect; in addition, the licorice extract also has flavonoid compounds, so that the gel formed by the olive polysaccharide extract and the licorice extract has a good sunscreen effect, 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.

[0045] It should be noted that the licorice extract can be used as an emulsifier for the gel: the surfactant in the licorice extract can reduce the tension of the oil-water interface, so that the oil phase and the water phase in the olive polysaccharide extract can be better mixed to form a uniform gel system; this emulsification effect not only improves the uniformity of the gel, but also enhances its stability and sensory properties.

[0046] It should be noted that the licorice extract can be used as a stabilizer for the gel: the licorice extract can stabilize the colloidal particles of the gel through hydrogen bonds and hydrophobic interactions to prevent their aggregation and precipitation; in addition, the antioxidant properties of the licorice extract can inhibit the oxidation reaction of the gel to further extend the shelf life of the gel.

[0047] It should be noted that the licorice extract can be used as a preservative for the gel: the licorice extract contains a variety of antibacterial ingredients, such as glycyrrhizic acid, flavonoids, etc. These ingredients can inhibit the growth and reproduction of microorganisms, thereby preventing the gel from mildewing and corruption.

[0048] It should be noted that the carbomer may be carbomer 941.

[0049] It should be noted that the weight ratio of butanediol to glycerol in the butanediol-glycerol mixed solution is 1:4.

[0050] Figure 4 A schematic flow chart of a method for preparing the gel provided in an embodiment of the present application is shown exemplarily; Based on a general inventive concept, such as Figure 4 As shown, the present application embodiment provides a method for preparing the gel, the method comprising: S1. mixing the olive polysaccharide extract and distilled water to obtain a polysaccharide solution; S2. mixing the carbomer and the polysaccharide solution so that the carbomer adsorbs the polysaccharide solution to obtain a mixture; S3. The propylene glycol, butylene glycol glycerol mixed solution and the mixture are mixed to obtain a gel matrix solution; S4. The licorice extract, citric acid and distilled water are mixed to obtain a glycyrrhizic acid solution; S5. Mixing triethanolamine, the gel matrix solution and the glycyrrhizic acid solution and adjusting the pH to obtain a gel with whitening and moisturizing functions.

[0051] This method is a method for preparing the above-mentioned gelling agent. The specific composition of the gelling agent can refer to the above-mentioned embodiment. Since this method adopts part or all of the technical solutions of the above-mentioned embodiment, it has at least all the beneficial effects brought by the technical solutions of the above-mentioned embodiment, which will not be described one by one here.

[0052] 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. In 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 giving the gel a unique elasticity and viscosity.

[0053] The present application is further described below in conjunction with specific examples. The experimental methods in the following examples where specific conditions are not specified are usually measured in accordance with national standards / industry standards; if there are no corresponding national standards / industry standards, they are measured in accordance with common international standards, conventional conditions or conditions recommended by the manufacturer. Example 1

[0054] like Figure 2 As shown, the present application embodiment provides a method for preparing olive polysaccharide extract, comprising: S1. The remaining 500 mL of the olive oil after the liquid phase was extracted was adsorbed with 500 g of DA201 macroporous adsorption resin to obtain an adsorbed liquid phase; the impurity adsorption site was a conical flask, and the impurity adsorption temperature was 30 ° C; the impurity adsorption was carried out under shaking conditions, the shaking speed was 50 r / min, and the shaking time was 6h; S2. filtering the adsorbed liquid phase to obtain a filtrate; S3. The filtrate and anhydrous ethanol are mixed to prepare an ethanol solution with a mass concentration of 80%, and then allowed to stand overnight at 4°C to extract the polysaccharide components of the filtrate to obtain a mixed solution; S4. The mixed solution is first centrifuged at 4°C, and the centrifugal precipitate is then redissolved and freeze-dried to obtain crude olive polysaccharide; S5. The trichloroacetic acid solution and the crude olive polysaccharide are mixed, and then allowed to stand overnight at 4° C., and then the pH of the mixture is adjusted to neutral using sodium hydroxide having a molar concentration of 6 mol / L to remove impurities to obtain an extract; S6. The extract is subjected to a second centrifugation to obtain an extract supernatant; S7. The extracted supernatant was dialyzed (using a 3500Da dialysis bag for 48 hours) and concentrated by rotary evaporation to obtain an olive polysaccharide extract.

[0055] The rotation speed of the first centrifugation is 8000 rpm, and the time of the first centrifugation is 10 min.

[0056] The rotation speed of the second centrifugation was 10000 rpm, and the time of the second centrifugation was 10 min.

[0057] The mass concentration of crude olive polysaccharide was 10 mg / mL, and the weight of trichloroacetic acid solute in the trichloroacetic acid solution was 5% of the weight of the trichloroacetic acid solution; The volume V1 of crude olive polysaccharide and the volume V2 of trichloroacetic acid solution satisfy the relationship: V1:V2=1:1. Example 2

[0058] Based on the olive polysaccharide extract disclosed in Example 1, the olive polysaccharide extract was tested: 1. Molecular weight detection of olive polysaccharide extract: 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; running time: 25 min; 2. Sample preparation: Use water to prepare the olive polysaccharide extract into a solution with a mass concentration of 5 mg / mL, and then filter the prepared solution through a filter membrane with a pore size of 0.45 μm.

[0059] The sample was tested using the above chromatogram, and the results were as follows Figure 5 And as shown in Table 1.

[0060] Table 1 Detection results of channel A of detector , The test results are automatically calculated by GPC software. Figure 6 As shown in Table 2, the weight average molecular weight Mw was 1,100,000±100,000.

[0061] Table 2 Calculation results of GPC software , The standard curve of the test is drawn according to the test results, and its formula is: f(x)==-0.0315597*x 3 +0.949904*x-9.465083*x+37.30616 (R 2 =0.9805728).

[0062] 2. Determination of monosaccharide composition: 1. Hydrolysis of crude olive polysaccharide extract: The crude olive polysaccharide extract was prepared into a sample solution with a mass concentration of 5 mg / mL using a trifluoroacetic acid solution with a molar concentration of 2 mol / L. The sample solution was filled into an ampoule bottle with 4 mL as a group, filled with nitrogen and sealed; it was then hydrolyzed at 105°C for 6 hours, and then the pH was adjusted to neutral using a sodium hydroxide solution with a molar concentration of 6 mol / L; then 0.5 mL of the neutral hydrolyzate was taken, and 0.5 mL of sodium hydroxide with a molar concentration of 0.25 mol / L and 0.5 mL of PMP-methanol solution with a molar concentration of 0.4 mol / L were added in sequence and mixed.

[0063] React in an oven at 70°C for 90 minutes, then take out the dried product and place it in an ice water bath for 10 minutes, then leave it until the dried product returns to room temperature, then add 0.5mL of 0.3mol / L hydrochloric acid solution to mix well, and finally use dichloromethane to extract PMP three times. The first extraction method is overnight extraction, and the interval time between the adjacent extractions of the last two extractions is 2h to 3h.

[0064] 2. High performance liquid chromatography detection process: Chromatographic column: Diamonsil C18, 250×4.6mm, 5μm; mobile phase: potassium dihydrogen phosphate with a molar concentration of 0.05mol / L (the volume ratio of potassium dihydrogen phosphate and acetonitrile is 20:80); flow rate: 1mL / min; detector: DAD ultraviolet detection wavelength 250nm; column oven: 35℃; running time: 80min.

[0065] Test results such as Figure 1 As shown, the UV detection results are as follows Figure 7 shown.

[0066] according to Figure 1 and Figure 7 Calculation shows that 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%. Example 3

[0067] Under the condition of the specific monosaccharide composition of the olive polysaccharide extract determined in Example 2, the gel was further prepared: The composition of the gel is as follows by mass fraction: olive polysaccharide extract: 5%, 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 inevitable impurities; wherein, Figure 3 As shown, the preparation method of licorice extract comprises: S1. Mix ethanol and ammonia water to obtain a mixed solution containing ethanol and ammonia water; wherein the mass ratio of ethanol to ammonia water is 0.75:0.03.

[0068] S2. subjecting the licorice and the mixed solution to ultrasonic extraction for 25 min to obtain an extract; S3. The extract is subjected to rotary evaporation and freeze-drying to obtain a licorice extract.

[0069] like Figure 4 As shown, a method for preparing a gel comprises: S1. mixing the olive polysaccharide extract and distilled water to obtain a polysaccharide solution; S2. The carbomer and the polysaccharide solution are mixed, and then slowly stirred at 70° C. to allow the carbomer to adsorb the polysaccharide solution and swell to obtain a mixture; S3. Mixing the propylene glycol, butylene glycol and glycerol mixed solution and the mixture at 70° C. to obtain a gel matrix solution; S4. The licorice extract, citric acid and distilled water were mixed at 55°C to obtain a glycyrrhizic acid solution; S5. First, cool the gel matrix solution to 55° C., then mix triethanolamine, the gel matrix solution and glycyrrhizic acid solution and adjust the pH to 6 to 8 to obtain a gel with whitening and moisturizing functions. Comparative Example 1

[0070] Based on the contents disclosed in Example 3, the following modifications are further made: Without using olive polysaccharide extract and licorice extract, only using stabilizer as a substitute, a gel is obtained (the components, by mass fraction, are: disodium oxalate tetraacetate: 0.4%, butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, citric acid: 1%, and the rest are distilled water and inevitable impurities.) Comparative Example 2

[0071] Based on the contents disclosed in Example 3, the following modifications are further made: Without using olive polysaccharide extract, a gel is obtained, the components of which are: butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, licorice extract: 20%, citric acid: 1%, and the rest are distilled water and inevitable impurities. Comparative Example 3

[0072] Based on the contents disclosed in Example 3, the following modifications are further made: Without using licorice extract, a gel is obtained, whose components are: olive polysaccharide extract: 1%, butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, licorice extract: 20%, citric acid: 1%, and the rest are distilled water and inevitable impurities.

[0073] Related experiments and effect data: 1. Gel stability test: All embodiments and comparative examples were subjected to stability tests using the following method: (1) Centrifugal stability: Weigh 2.0 g of the sample to be tested into a centrifuge tube, and then centrifuge at 4000 r / min, 8000 r / min, and 12000 r / min, respectively, and observe the state of the gel after centrifugation.

[0074] (2) Thermal stability: Weigh 2.0 g of the sample to be tested into a centrifuge tube and keep it at 40°C for 24 h. Take it out and check whether it is turbid or stratified.

[0075] (3) Cold stability: Weigh 2.0 g of the sample to be tested into a centrifuge tube and keep it at -20°C for 24 h. Take it out and check whether it is turbid or stratified.

[0076] (4) Thermal cycling stability: Weigh 2.0 g of the sample to be tested into a centrifuge tube and keep it at -20°C for 12 h. After taking it out, keep it at 40°C for 12 h. Repeat the above operation twice and observe whether turbidity, stratification, etc. occur.

[0077] (5) Sample retention for observation: Weigh 2.0 g of the sample into a centrifuge tube and leave it at room temperature for one month to observe whether turbidity, stratification, etc. appear.

[0078] 2. Moisturizing ability test of gel: The moisturizing properties of the obtained gel were measured by weighing method: 5g of each moisturizing gel of each experimental example was weighed, and then pure water was added in a weight ratio of 1:1 to shake evenly, and then the mixture was placed in a weighing tank that had been dried to a constant weight in advance. At room temperature, they were placed in a dryer filled with 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 after being placed for 1h, 2h, 4h, 6h, 8h, 12h and 24h. Three groups of parallel samples were measured under the same conditions, and the average value was taken. The formula is as follows: , Where M1 is the mass of the sample when it is put in; M2 is the mass of the sample when it is taken out.

[0079] 3. Whitening ability test of gel: (1) Configuration of detection reagents: Preparation of 100 μg / mL tyrosinase solution: Accurately weigh 10.34 mg of tyrosinase and dissolve it in a 10 mL centrifuge tube using PBS buffer with a pH of 6.8. Transfer the dissolved solution to a 100 mL volumetric flask and dilute to the mark with PBS buffer to obtain a 100 μg / mL tyrosinase solution for later use.

[0080] L-tyrosine solution with a molar concentration of 2.5mmol / L: Accurately weigh 45.3mg of tyrosine and dissolve it in a 10mL centrifuge tube using PBS buffer with a pH of 6.8. Transfer the dissolved solution to a 100mL volumetric flask and dilute to the mark with PBS buffer to obtain an L-tyrosine solution with a molar concentration of 2.5mmol / L for later use.

[0081] (2) Test method: Add 1 mL of the gel prepared in Example 3 and each comparative example to the centrifuge tube, then add 1 mL of tyrosinase solution with a mass concentration of 100 μg / mL and 1.8 mL of PBS solution, mix the added solutions evenly, and then incubate in an incubator at 37°C for 10 min. Then add 1.2 mL of L-tyrosine solution with a molar concentration of 2.5 mmol / L, and incubate in an incubator at 37°C for 10 min again. After the incubation, take samples immediately after the incubation and add them to a 96-well plate. Each sample is measured three times at 475 nm, and the average value is taken. Then the inhibition rate is calculated according to the following formula: , Wherein, A1 is the absorbance of the sample well; A2 is the absorbance when PBS replaces tyrosinase; A3 is the absorbance when PBS replaces the sample; and A4 is the absorbance when PBS replaces the sample and tyrosinase.

[0082] 4The test results are: (1) Stability test results Centrifugal stability: Under the three centrifugal conditions, Example 3 and Comparative Examples 1 to 3 did not show turbidity, stratification, flocculation and the like, indicating that the product has good stability under accelerated conditions.

[0083] Thermal stability: Under high temperature conditions, Example 3 and Comparative Examples 1 to 3 did not show turbidity, stratification, flocculation and the like, indicating that the product has good stability under high temperature conditions.

[0084] Cold stability: Under low temperature conditions, Example 3 and Comparative Examples 1 to 3 did not show turbidity, stratification, crystallization and the like, indicating that the product has good stability under low temperature conditions.

[0085] Thermal cycling stability: After thermal cycling, no special phenomenon occurred in Example 3 and Comparative Examples 1 to 3, indicating that the product has good stability under thermal cycling conditions.

[0086] Sample observation: Example 3 and Comparative Examples 1 to 3 were left to stand for one month and then re-observed, and no turbidity, stratification, deterioration, drying or solidification occurred, indicating that the product can be stored for a long time at room temperature.

[0087] (2) Moisturizing ability test results The water loss rate (%) under the condition of relative humidity of 43% is shown in Table 3.

[0088] Table 3 Water loss rate results of various embodiments and comparative examples under relative humidity of 43% , The water loss rate (%) under the condition of relative humidity of 81% is shown in Table 4.

[0089] Table 4 Water loss rate results of various embodiments and comparative examples under relative humidity of 81% , Whitening ability test, the results are shown in Table 5.

[0090] Table 5 Whitening ability results of various embodiments and comparative examples , In summary, the olive polysaccharide extract provided in the embodiments of the present application, based on the antioxidant and anti-inflammatory effects combined with the tyrosinase inhibitory effect of licorice extract, can more effectively reduce melanin deposition, improve skin spots, and thus achieve a whitening effect.

[0091] In addition, the embodiment of the present application provides a gel with whitening and moisturizing functions, in which the main ingredients of the olive polysaccharide extract and the licorice extract used in the gel are derived from natural extracts, are non-irritating to the skin, have high safety, and have 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 cooperate with traditional moisturizers to further improve the water-locking and moisturizing effect of the gel.

[0092] In addition, the embodiment of the present application provides a gel with whitening and moisturizing functions. The licorice extract used in the gel can also be used as an emulsifier, stabilizer and preservative of the gel system, which can simultaneously enhance the stability and anti-mildew ability of the gel; in addition, the gel is based on the antioxidant and anti-inflammatory effects of olive polysaccharide extract, so that the gel also has auxiliary effects such as antioxidant and anti-inflammatory, and has more development advantages when used as a whitening and moisturizing cosmetic.

[0093] In addition, the present application provides a method for preparing a gel, which is simple to operate and has mild preparation conditions, and can obtain a gel product with optimal efficacy.

[0094] The above description is only a specific implementation of the present application, so that those skilled in the art can understand or implement the present application. It will be apparent to those skilled in the art that various modifications to these embodiments are possible, and the general principles defined in the present application can be implemented in other embodiments without departing from the spirit or scope of the present application. Therefore, the present application will not be limited to the embodiments shown in the present application, but will conform to the widest range consistent with the principles and novel features applied for by the present application.

Claims

1. An olive polysaccharide extract, wherein the monosaccharide composition of the olive polysaccharide extract is, by mass fraction, mannose ≥5%, rhamnose ≥6%, glucuronic acid ≥3%, galacturonic acid ≥33%, glucose ≥6%, galactose ≥20% and arabinose ≥20%.

2. The olive polysaccharide extract according to claim 1, wherein the monosaccharide composition of the olive polysaccharide extract is, by 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%.

3. The olive polysaccharide extract according to claim 1, wherein the weight average molecular weight of the olive polysaccharide extract is 1100000±100000.

4. A method for preparing the olive polysaccharide extract according to any one of claims 1 to 3, the method comprising: The remaining liquid phase after olive oil extraction is subjected to impurity adsorption to obtain an adsorbed liquid phase; Filtering the adsorbed liquid phase to obtain a filtrate; Mixing the filtrate with anhydrous ethanol to extract the polysaccharide components of the filtrate to obtain a mixed solution; The mixed solution is first centrifuged, and then the centrifugal precipitate is redissolved and freeze-dried to obtain crude olive polysaccharide; The trichloroacetic acid solution and the crude olive polysaccharide are mixed and the pH is adjusted to remove impurities to obtain an extract; The extract is subjected to a second centrifugation to obtain an extraction supernatant; The extraction supernatant is dialyzed and concentrated by rotary evaporation in sequence to obtain an olive polysaccharide extract. 5 . The method according to claim 4 , 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 method according to claim 4, 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 method according to claim 4, wherein the mass concentration of the crude olive polysaccharide is ≥ 10 mg / mL, and the weight of the 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 olive polysaccharide and the volume V2 of the trichloroacetic acid solution satisfy the relationship: V1:V2≥1:

1.

8. A gel with whitening and moisturizing functions, comprising the olive polysaccharide extract according to any one of claims 1 to 3 and a liquorice extract.

9. The gelling agent according to claim 8, wherein the composition of the gelling agent is, by mass fraction: Olive polysaccharide extract: 0.2%-1%, butylene glycol glycerol mixed solution: 20%, triethanolamine: 1%, carbomer: 1%, propylene glycol: 10%, licorice extract: 20%, citric acid: 1%, the rest is distilled water and inevitable impurities; among them, The preparation method of the licorice extract comprises: Mixing ethanol and aqueous ammonia to obtain a mixed solution containing ethanol and aqueous ammonia; subjecting licorice and the mixed solution to ultrasonic extraction to obtain an extract; The extract is subjected to rotary evaporation and freeze drying to obtain a liquorice extract.

10. A method for preparing the gel according to claim 8 or 9, the method comprising: Mixing the olive polysaccharide extract and distilled water to obtain a polysaccharide solution; Mixing carbomer and the polysaccharide solution so that the carbomer absorbs the polysaccharide solution to obtain a mixture; Mixing a mixed solution of propylene glycol and butylene glycol and glycerol with the mixture to obtain a gel matrix solution; Mixing licorice extract, citric acid and distilled water to obtain glycyrrhizic acid solution; The triethanolamine, the gel matrix solution and the glycyrrhizic acid solution are mixed and the pH is adjusted to obtain a gel with whitening and moisturizing functions.

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