Raspberry leaf extract as well as preparation method and cosmetic thereof

Abstract

The invention relates to the field of cosmetics, in particular to a raspberry leaf extract, a preparation method thereof and cosmetics. The invention provides a preparation method of a raspberry leaf extract, which comprises the following steps: preparing raspberry leaves and water into homogenate; mixing the homogenate with a deep-eutectic solvent, and extracting; wherein the eutectic solvent is prepared from the following components in percentage by mass: 55 to 70 percent of L-carnitine, 12 to 30 percent of gallic acid and the balance of water. According to the technical scheme, the raspberry leaves are extracted by adopting the eutectic solvent, so that active substances can be effectively extracted; moreover, the extraction rates of the ellagic acid and the kaempferol-3-O-rutinoside are higher. The raspberry leaf extract extracted by the method has the characteristics of good solubility and remarkable oxidation resistance. The contents of ellagic acid, kaempferol-3-O-rutinoside and other active ingredients in the raspberry leaf extract extracted by the method are obviously increased.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of cosmetics, in particular, a raspberry leaf extract and a preparation method thereof, and a cosmetic. BACKGROUND

[0002] At present, in the field of cosmetics, the development and utilization of raspberries mainly focuses on the development and utilization of raspberry fruits; for example, the extraction of raspberry fruits, and the application of raspberry fruit extracts in cosmetics.

[0003] However, this is limited for the development and utilization of raspberries. SUMMARY

[0004] The purpose of the embodiments of the present application is to provide a raspberry leaf extract and a preparation method thereof, and a cosmetic.

[0005] In a first aspect, the present application provides a preparation method of a raspberry leaf extract, comprising: homogenizing raspberry leaves with water; mixing the homogenate with a deep eutectic solvent for extraction; wherein the deep eutectic solvent consists of the following components in mass percentage: 55%~70% L-carnitine, 12%~30% gallic acid, and the balance water.

[0006] In the above technical solution, the raspberry leaves are extracted by using a deep eutectic solvent consisting of 55 wt%~70 wt% L-carnitine, 12 wt%~30 wt% gallic acid, and the balance water, which can effectively extract active substances, and the extraction rates of ellagic acid and kaempferol-3-O-rutinoside are higher. The raspberry leaf extract obtained by the method has good solubility and significant antioxidant performance. The content of active ingredients such as ellagic acid and kaempferol-3-O-rutinoside in the raspberry leaf extract obtained by the method is significantly increased.

[0007] In other embodiments of the present application, the mixing of the homogenate with the deep eutectic solvent comprises: mixing the homogenate with the deep eutectic solvent according to a volume ratio of the deep eutectic solvent to the mass of the raspberry leaves of (1:10)~(1:25).

[0008] In other embodiments of the present application, the concentration of the deep eutectic solvent is 25%-70% in mass percentage.

[0009] In other embodiments of the present application, the deep eutectic solvent consists of the following components in mass percentage: 60%~65% L-carnitine, 19%~23% gallic acid, and the balance water.

[0010] In other embodiments of the present application, the homogenate is mixed with a deep eutectic solvent for extraction, including: The extraction is performed under high-voltage pulsed electric field conditions.

[0011] In other embodiments of the present application, the extraction is performed under high-voltage pulsed electric field conditions, including: The extraction is performed under a field strength of 2.5 kV / cm ~ 5.0 kV / cm.

[0012] In other embodiments of the present application, the extraction is performed under high-voltage pulsed electric field conditions, including: The extraction is performed under a pulse number of 25-75 times.

[0013] In a second aspect, the present application provides a raspberry leaf extract, which is prepared by the preparation method of the raspberry leaf extract according to any one of the first aspect.

[0014] In other embodiments of the present application, the active ingredients of the raspberry leaf extract include, in terms of mass percentage: 231-500 µg / mL ellagic acid, 4.6-50 µg / mL kaempferol-3-O-rutinoside.

[0015] In a third aspect, the present application provides a cosmetic product, which includes the raspberry leaf extract prepared by the preparation method of the raspberry leaf extract according to any one of the first aspect; or the cosmetic product includes the raspberry leaf extract according to any one of the second aspect. BRIEF DESCRIPTION OF DRAWINGS

[0016] In order to more clearly illustrate the technical solutions of the embodiments of the present application, the following will briefly introduce the drawings needed to be used in the embodiments. It should be understood that the following drawings only show some embodiments of the present application, and therefore should not be regarded as a limitation on the scope. For those skilled in the art, other related drawings can also be obtained without creative labor.

[0017] Figure 1 ABTS experimental standard curve; Figure 2 DPPH experimental standard curve. DETAILED DESCRIPTION

[0018] In order to make the purpose, technical solutions and advantages of the embodiments of the present application more clear, the technical solutions in the embodiments of the present application will be described clearly and completely below. Obviously, the described embodiments are some of the embodiments of the present application, but not all the embodiments.

[0019] Therefore, the following detailed description of the embodiments of this application is not intended to limit the scope of the claimed application, but merely to illustrate selected embodiments of the application. All other embodiments obtained by those skilled in the art based on the embodiments of this application without inventive effort are within the scope of protection of this application.

[0020] The inventors discovered that raspberry leaves are rich in active substances that can be used in cosmetics to prepare antioxidant, whitening, and other cosmetic products.

[0021] Further research revealed that conventional extraction techniques, such as water extraction and alcohol extraction, yielded too low a concentration of active substances from raspberry leaves, and the amount of active substances that could be extracted, such as ellagic acid and kaempferol-3-O-rutin, was limited; thus, they had almost no value for industrial application.

[0022] Further research revealed that by using specific eutectic solvents (eutectic mixtures formed by two or more components through intermolecular forces such as hydrogen bonds, whose melting points are significantly lower than those of any single pure component), the active substances in raspberry leaves can be effectively extracted, and the extraction rates of the active ingredients ellagic acid and kaempferol-3-O-rutin can be increased.

[0023] Based on the above research, this application provides a method for preparing raspberry leaf extract, comprising: Mix raspberry leaves with water to form a paste; The homogenate was mixed with a eutectic solvent for extraction; The eutectic solvent is composed of the following components: L-carnitine 55%~70%, gallic acid 12%~30%, and the balance water.

[0024] For example, in some embodiments of this application, the eutectic solvent described above, by mass percentage, consists of the following components: L-carnitine 55%, 58%, 61%, 64%, 67%, 70% or any two of the aforementioned values; gallic acid 12%, 15%, 18%, 21%, 24%, 27%, 30% or any two of the aforementioned values, and the balance being water.

[0025] In the above technical solution, the extraction of raspberry leaves using a eutectic solvent composed of 55 wt%~70 wt% L-carnitine, 12 wt%~30 wt% gallic acid (3,4,5-trihydroxybenzoic acid, also known as gallic acid), and the balance water not only effectively extracts active substances but also achieves higher extraction rates of ellagic acid and kaempferol-3-O-rutin. The raspberry leaf extract obtained using this method exhibits good solubility and significant antioxidant properties. The content of active ingredients such as ellagic acid and kaempferol-3-O-rutin in the raspberry leaf extract obtained using this method is significantly increased.

[0026] Further, optionally, in some embodiments of this application, the eutectic solvent, by mass percentage, consists of the following components: L-carnitine 60%~65%, gallic acid 19%~23%, and the balance water.

[0027] Furthermore, in some embodiments of this application, the method for preparing raspberry leaf extract includes the following steps: Step S1: Prepare homogenate.

[0028] Furthermore, in some embodiments of this application, the step of preparing the homogenate includes: Mix raspberry leaves with water to make a paste.

[0029] Furthermore, in some embodiments of this application, the raspberry leaves described above are dried raspberry leaves.

[0030] For example, in some embodiments of this application, fresh raspberry leaves can be dried by drying.

[0031] Further, optionally, in some embodiments of this application, preparing a homogenate includes: Take dried raspberry leaves, wash them clean, dry them, and then crush or grind them into powder. Then add water to the powder to prepare a raspberry leaf homogenate.

[0032] Step S2: Mix the homogenate with a eutectic solvent and extract.

[0033] In some embodiments of this application, the eutectic solvent, by mass percentage, consists of the following components: 60% to 65% L-carnitine, 19% to 23% gallic acid, and the balance water.

[0034] For example, in some embodiments of this application, the eutectic solvent is composed of the following components by mass percentage: 60% L-carnitine, 20% gallic acid, and 20% water.

[0035] Further, alternatively, in some embodiments of this application, the concentration of the eutectic solvent is 25%-70% by mass percentage.

[0036] For example, the concentration of the eutectic solvent, by mass percentage, is 25%, 26%, 28%, 30%, 35%, 38%, 40%, 45%, 50%, 55%, 58%, 60%, 65%, 68%, 70%, or any two of the aforementioned values.

[0037] Further, optionally, in some embodiments of this application, the homogenate is mixed with a eutectic solvent for extraction, including: Extraction was performed under high-voltage pulsed electric field conditions.

[0038] Further, optionally, in some embodiments of this application, extraction is performed under high-voltage pulsed electric field conditions, including: Extraction was performed under electric field strength conditions of 2.5 kV / cm to 5.0 kV / cm.

[0039] Exemplary examples, in some embodiments of this application, extraction is performed under high-voltage pulsed electric field conditions, including: Extraction is performed under the condition that the electric field strength is 2.5 kV / cm, 2.6 kV / cm, 2.7 kV / cm, 2.8 kV / cm, 2.9 kV / cm, 3.0 kV / cm, 3.1 kV / cm, 3.2 kV / cm, 3.5 kV / cm, 3.8 kV / cm, 4.0 kV / cm, 4.2 kV / cm, 4.3 kV / cm, 4.5 kV / cm, 4.6 kV / cm, 4.7 kV / cm, 4.9 kV / cm, 5.0 kV / cm, or any two of the aforementioned values.

[0040] Further, optionally, in some embodiments of this application, extraction is performed under high-voltage pulsed electric field conditions, including: Extraction was performed under conditions of 25-75 pulses.

[0041] Exemplary examples, in some embodiments of this application, extraction is performed under high-voltage pulsed electric field conditions, including: Extraction is performed when the number of pulses is 25, 26, 28, 30, 32, 35, 38, 40, 42, 45, 48, 50, 52, 55, 58, 60, 65, 68, 70, 72, 75, or any two of the aforementioned values.

[0042] Some embodiments of this application provide a raspberry leaf extract, which is prepared by the preparation method of raspberry leaf extract provided in any of the foregoing embodiments.

[0043] Furthermore, in some embodiments of this application, the active ingredients of the raspberry leaf extract, by mass percentage, include: 231~500 µg / mL ellagic acid and 4.6~50 µg / mL kaempferol-3-O-rutin.

[0044] Exemplary examples, in some embodiments of this application, the active ingredients of the above-mentioned raspberry leaf extract, by mass percentage, include: 231 µg / mL, 232 µg / mL, 235 µg / mL, 250 µg / mL, 255 µg / mL, 260 µg / mL, 265 µg / mL, 270 µg / mL, 275 µg / mL, 280 µg / mL, 290 µg / mL, 300 µg / mL, 320 µg / mL, 350 µg / mL, 380 µg / mL, 400 µg / mL, 420 µg / mL, and 450 µg / mL. Ellagic acid in amounts of 480 µg / mL, 500 µg / mL, or any two of the aforementioned values; kaempferol-3-O-rutin in amounts of 4.6 µg / mL, 5 µg / mL, 6 µg / mL, 7 µg / mL, 8 µg / mL, 9 µg / mL, 10 µg / mL, 11 µg / mL, 12 µg / mL, 13 µg / mL, 14 µg / mL, 15 µg / mL, 16 µg / mL, 18 µg / mL, 20 µg / mL, 22 µg / mL, 25 µg / mL, 28 µg / mL, 30 µg / mL, 35 µg / mL, 40 µg / mL, 45 µg / mL, 46 µg / mL, 50 µg / mL, or any two of the aforementioned values.

[0045] Further optionally, in some embodiments of this application, the active ingredients of the raspberry leaf extract, by mass percentage, include: 359.296 µg / mL ~ 463.981 µg / mL; ellagic acid; 5.107~30 µg / mL kaempferol-3-O-rutin.

[0046] Further optionally, in some embodiments of this application, the active ingredients of the raspberry leaf extract, by mass percentage, include: 359.296 µg / mL ~ 463.981 µg / mL; ellagic acid; 5.107~30 µg / mL kaempferol-3-O-rutin.

[0047] Further optionally, in some embodiments of this application, the active ingredients of the raspberry leaf extract, by mass percentage, include: 359.296 µg / mL ~ 463.981 µg / mL; ellagic acid; 5.107~30 µg / mL kaempferol-3-O-rutin.

[0048] Further optionally, in some embodiments of this application, the active ingredients of the raspberry leaf extract, by mass percentage, include: 413.652 µg / mL ~ 463.981 µg / mL; ellagic acid; 15.327~30 µg / mL kaempferol-3-O-rutin.

[0049] Some embodiments of this application provide a cosmetic product comprising raspberry leaf extract prepared by the method for preparing raspberry leaf extract provided in any of the foregoing embodiments; or the cosmetic product comprises raspberry leaf extract provided in any of the foregoing embodiments.

[0050] The features and performance of this application will be further described in detail below with reference to embodiments: Example 1 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 50% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:10.

[0051] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 2.5 kV / cm and the number of pulses at 25. Combine the extracted mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0052] Example 2 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 63% L-carnitine, 21% gallic acid, and the remainder water by mass percentage. The concentration of the eutectic solvent is 50% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:17.

[0053] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. The field strength is controlled at 3.7 kV / cm, and the number of pulses is 50. After filtration, the extracted mixture is combined and the supernatant is collected to obtain the sample solution.

[0054] Example 3 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 65% L-carnitine, 19% gallic acid, and the remainder water by mass percentage. The concentration of the eutectic solvent is 50% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:25.

[0055] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 5.0 kV / cm and the number of pulses at 75. Combine the extracted mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0056] Example 4 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 25% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:10.

[0057] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. The field strength is controlled at 3.7 kV / cm, and the number of pulses is 75. After filtration, the extracted mixture is combined and the supernatant is collected to obtain the sample solution.

[0058] Example 5 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 70% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:17.

[0059] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 5.0 kV / cm and the number of pulses at 25. Combine the extracted mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0060] Example 6 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 60% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:25.

[0061] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 2.5 kV / cm and the number of pulses at 50. Combine the extracted mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0062] Example 7 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 70% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:10.

[0063] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 5.0 kV / cm and the number of pulses at 50. Combine the extracted mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0064] Example 8 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 70% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:17.

[0065] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 2.5 kV / cm and the number of pulses at 75. Combine the extracted extract mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0066] Example 9 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 60% L-carnitine, 20% gallic acid, and 20% water by mass percentage. The concentration of the eutectic solvent is 70% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:25.

[0067] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 3.7 kV / cm and the number of pulses at 25. Combine the extracted extract mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0068] Comparative Example 1 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a slurry. Mix the slurry with water to obtain a mixture. In the mixture, the volume ratio of water to the mass ratio of raspberry leaves is 1:25.

[0069] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 5.0 kV / cm and the number of pulses at 75. Combine the extracted extract mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0070] Comparative Example 2 A raspberry fruit extract is provided, prepared according to the following steps: Step (1): Take dried raspberry fruits, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 63% L-carnitine, 21% gallic acid, and 16% water by mass percentage. The concentration of the eutectic solvent is 50% by mass percentage. In the mixture, the volume ratio of the ethanol solution to the mass of the raspberry leaves is 1:17.

[0071] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 5.0 kV / cm and the number of pulses at 75. Combine the extracted extract mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0072] Comparative Example 3 A raspberry leaf extract is provided, prepared according to the following steps: Step (1): Take dried raspberry leaves, wash them clean, and then dry them. Crush them into powder, add water to the powder, and make a homogenate. Mix the homogenate with a eutectic solvent to obtain a mixture. The eutectic solvent consists of 20% betaine, 75% glycerol, and 5% propylene glycol by mass percentage. The concentration of the eutectic solvent is 50% by mass percentage. The volume ratio of the eutectic solvent to the mass of the raspberry leaves in the mixture is 1:10.

[0073] Step (2): Extract the mixture obtained in step (1) using high-voltage pulsed electric field technology. Control the field strength at 5.0 kV / cm and the number of pulses at 75. Combine the extracted mixtures and filter them, collecting the supernatant filtrate to obtain the sample solution.

[0074] The performance of the sample solutions prepared in Examples 1-9 and Comparative Examples 1-3 was tested.

[0075] 1. The content of active ingredients such as ellagic acid and kaempferol-3-O-rutin in each sample solution was used as the extraction rate indicator.

[0076] The contents of ellagic acid and kaempferol-3-O-rutin in each sample solution were determined by high performance liquid chromatography.

[0077] (1) Sample pretreatment: Centrifuge the sample solutions provided in each embodiment or comparative example, take the supernatant and filter it through a 0.22 μm filter membrane, and take the filtrate as the test solution for later use.

[0078] (2) Chromatographic conditions Chromatographic column: Eclipse XDB C18 column (250mm × 4.6mm, 5μm); Mobile phase: Phase A is acetonitrile; Phase B is 0.2 wt% aqueous phosphoric acid solution. Gradient elution conditions are shown in Table 1. Table 1

[0079] Flow rate: 1.0 mL / min; Injection volume: 20 μL; Column temperature: 25℃; Measurement wavelength: 344nm.

[0080] (3) Plotting the standard curve Ellagic acid and kaempferol-3-O-rutinoside standards were dissolved in methanol to prepare standard stock solutions (ellagic acid concentration approximately 0.2 mg / mL, kaempferol-3-O-rutinoside concentration approximately 0.5 mg / mL). 1 mL of each ellagic acid and kaempferol-3-O-rutinoside standard stock solution was accurately pipetted to prepare a mixed standard intermediate solution. This mixed standard intermediate solution was serially diluted to the corresponding concentrations to obtain a series of mixed standard working solutions of ellagic acid and kaempferol-3-O-rutinoside. The ellagic acid concentrations in this standard series were approximately 2 µg / mL, 4 µg / mL, 8 µg / mL, 16 µg / mL, and 24 µg / mL, respectively, and the kaempferol-3-O-rutinoside concentrations were approximately 5 µg / mL, 10 µg / mL, 20 µg / mL, 40 µg / mL, and 60 µg / mL, respectively.

[0081] (4) Sample testing Each example or comparative sample was injected into a liquid chromatograph under the same chromatographic conditions, and the chromatograms were recorded. The contents of ellagic acid and kaempferol-3-O-rutinoside in each example or comparative sample could be calculated using the standard curves of ellagic acid and kaempferol-3-O-rutinoside.

[0082] The contents of ellagic acid and kaempferol-3-O-rutin in the sample solutions of each embodiment and comparative example obtained by testing are shown in Table 2.

[0083] Table 2

[0084] The data in the table above shows that: The contents of ellagic acid and kaempferol-3-O-rutin in the sample solutions of raspberry leaf extracts prepared in each embodiment were higher than those in the sample solutions of raspberry leaf extracts prepared in each comparative example; this shows that the scheme of the embodiments of this application can effectively improve the extraction rate of active ingredients in raspberry leaves.

[0085] Furthermore, a comparison between Comparative Example 2 and Example 2 shows that the raspberry leaf extract contains higher levels of ellagic acid and kaempferol-3-O-rutin compared to the raspberry fruit extract.

[0086] Furthermore, comparing Comparative Example 1 with Example 3, it can be seen that, compared with the traditional water extraction process, the solution of this application embodiment can effectively improve the extraction rate of active ingredients in raspberry leaves.

[0087] Furthermore, compared with Example 3, it can be seen that when other eutectic solvents (not within the scope of this application) are used, the extraction rate of active ingredients in raspberry leaves is significantly reduced.

[0088] 2. The antioxidant properties of samples prepared in some examples and comparative examples were tested.

[0089] (1) ABTS·scavenging experiment to test antioxidant activity ABTS (2,2′-hydrazine-bis(3-ethylbenzothiazoline-6-sulfonic acid) diamine salt) generates a stable blue-green cationic radical ABTS· in the presence of K₂S₂O₈, which exhibits maximum absorbance at 734 nm. The generation of ABTS· is inhibited when antioxidants are added to the reaction system; therefore, the antioxidant activity of the sample can be evaluated by measuring the absorbance at 734 nm.

[0090] The testing steps are as follows: Prepare a 2.45 mmol / L K₂S₂O₈ solution; prepare a 7 mmol / L ABTS stock solution; prepare a 0.2 mol / L PBS solution (pH=7.4); prepare the test sample solution using the PBS solution; use VC as a positive control sample to construct a standard curve. See the instruction manual appendix. Figure 1 Before the experiment, the ABTS stock solution was diluted with PBS solution to adjust its OD value. 734 A concentration of 0.6-0.8 is considered the working solution for ABTS.

[0091] The sample solutions of each example and comparative example were diluted 1000 times to prepare the test solutions. 10 μL of the test solution from each example or comparative example and 190 μL of ABTS working solution were mixed, incubated at room temperature for 6 min, and the absorbance value (A1) at 734 nm was measured. Similarly, 10 μL of the sample solution from each example or comparative example and 190 μL of PBS solution were mixed, incubated at room temperature for 6 min, and the absorbance value (A2) at 734 nm was measured. Finally, 10 μL of PBS solution and 190 μL of ABTS working solution were mixed, incubated at room temperature for 6 min, and the absorbance value (A0) at 734 nm was measured. Three replicates were prepared for each sample solution, and the ABTS clearance rate was calculated.

[0092]

[0093] The test results are shown in Table 3.

[0094] (2) DPPH· scavenging experiment to test antioxidant activity.

[0095] DPPH radicals (1,1-diphenyl-2-trinitrophenylhydrazine radicals) appear purple in anhydrous ethanol solution, exhibiting a characteristic absorption peak at 517 nm, and their concentration shows a linear relationship with absorbance. When antioxidants are added to the reaction system, they neutralize DPPH radicals through electron or hydrogen atom transfer mechanisms, leading to a decrease in absorbance and decolorization of the solution. Therefore, the antioxidant activity of the sample can be evaluated by measuring the absorbance at 517 nm.

[0096] The testing method is as follows: Prepare a 0.2 mmol / L DPPH·anhydrous ethanol solution; prepare a 200 mg / mL solution of the test sample; use VC as a positive control sample to construct a standard curve. See the instruction manual appendix. Figure 2 .

[0097] Take 100 μL of the test solution from each example or comparative example and 100 μL of DPPH solution, mix well, and react at room temperature in the dark for 30 min. Measure the absorbance at 517 nm (A1). Take 100 μL of the sample solution and 100 μL of ethanol, mix well, and react in the dark for 30 min. Measure the absorbance at 517 nm (A2). Take 100 μL of DPPH solution and 100 μL of ethanol, mix well, and react in the dark for 30 min. Measure the absorbance at 517 nm (A0). Perform three replicates for each sample and calculate the DPPH scavenging rate.

[0098]

[0099] The test results are shown in Table 3.

[0100] Table 3

[0101] The data in the table above shows that: The ABTS and DPPH scavenging rates of the sample solutions of raspberry leaf extract prepared in each embodiment were higher than those of the sample solutions of raspberry leaf extract prepared in each comparative example; this demonstrates that the solution of the present application embodiment can effectively improve the ABTS and DPPH scavenging rates of raspberry leaf extract.

[0102] Furthermore, a comparison between Comparative Example 2 and Example 2 shows that the ABTS and DPPH scavenging rates of the raspberry leaf extract are higher than those of the raspberry fruit extract.

[0103] Furthermore, comparing Comparative Example 1 with Example 3, it can be seen that, compared with the traditional water extraction process, the solution of this application embodiment can effectively improve the ABTS· scavenging rate and DPPH· scavenging rate of raspberry leaf extract.

[0104] Furthermore, compared with Example 3, it can be seen that when other eutectic solvents (not within the scope of this application) are used, the ABTS· scavenging rate and DPPH· scavenging rate of raspberry leaf extract are significantly reduced.

[0105] 3. The whitening efficacy of samples prepared in some of the embodiments and comparative examples was tested.

[0106] The whitening effect was evaluated using a tyrosinase inhibition assay. Tyrosinase, a key rate-limiting enzyme in the melanin synthesis pathway, directly regulates the rate of melanin formation by catalyzing the hydroxylation of tyrosine to dopa and further oxidizing dopa to dopaquinone. When the activity of this enzyme is inhibited, the accumulation of melanin precursors is hindered, and the pigment synthesis chain is effectively blocked. Therefore, the ability of a sample to interfere with melanin production can be evaluated by measuring its inhibitory activity on tyrosinase, providing a scientific basis for screening skin-whitening ingredients.

[0107] The testing steps are as follows: Prepare the following reagents: ① 1.5 mmol / L L-tyrosine solution; ② 0.2 mmol / L PBS phosphate buffer (pH=6.8); ③ Tyrosinase solution with an enzyme activity of 500 U / mL; ④ 1 mg / mL kojic acid stock solution as a positive control; ⑤ the test solution provided in each example or comparative example.

[0108] Add 80 μL of phosphate buffer, 40 μL of test solution or kojic acid at different concentration gradients, and 40 μL of enzyme solution to a 96-well plate sequentially. Finally, add 80 μL of L-tyrosine and react for 20 min. Measure the absorbance at 475 nm. Perform three replicates for each sample and calculate the tyrosinase inhibition rate.

[0109]

[0110] The test results are shown in Table 4.

[0111] Table 4

[0112] The experimental data in the table above shows that: The tyrosinase inhibition rates of the sample solutions of raspberry leaf extracts prepared in each embodiment were higher than those of the sample solutions of raspberry leaf extracts prepared in each comparative example; this demonstrates that the solution of the present application embodiment can effectively improve the tyrosinase inhibition rate of raspberry leaf extracts.

[0113] Furthermore, a comparison between Comparative Example 2 and Example 2 shows that the raspberry leaf extract exhibits a higher tyrosinase inhibition rate compared to the raspberry fruit extract.

[0114] Furthermore, comparing Comparative Example 1 with Example 3, it can be seen that, compared with the traditional water extraction process, the solution of this application embodiment can effectively improve the tyrosinase inhibition rate of raspberry leaf extract.

[0115] Furthermore, compared with Example 3, it can be seen that when other eutectic solvents (not within the scope of this application) are used, the tyrosinase inhibition rate of raspberry leaf extract is significantly reduced.

[0116] 4. Cytotoxicity tests were conducted on samples prepared in some of the embodiments and comparative examples.

[0117] Cytotoxicity was determined using the CCK8 assay. The main component of CCK8 is WST-8 (2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl)-5-(2,4-disulfonic acid benzene)-2H-tetrazole monosodium salt), which can be reduced by dehydrogenases in the mitochondria of living cells to produce orange-yellow formazan, with a characteristic absorption wavelength of 450 nm. Therefore, cytotoxicity of samples can be evaluated by measuring their survival rate against CCK8.

[0118] The testing steps are as follows: Cells in the logarithmic growth phase were seeded into 96-well plates at a seeding density of 4 x 10⁻⁶ cells / well. 5Cells were cultured per well for 24 hours. Samples from each example or comparative study were diluted five times with serum-free medium and added to 96-well plates. Sample concentrations were set at 10 wt% with four replicates. A blank control (2% FBS) and a positive control (10 wt% FBS) were also included. Cells were cultured for another 24 hours, and cell morphology and number were observed periodically under a microscope. After 48 hours, CCK-8 was diluted 10-fold with 2 wt% FBS medium, and 100 μL of the CCK-8 dilution was added to each well. The plates were then incubated at 37°C for 1.5 hours. Cell absorbance was measured using a microplate reader at 450 nm, and cell viability was calculated.

[0119] Survival rate = OD 样品组 / OD 空白组 ×100%.

[0120] The test results are shown in Table 5.

[0121] Table 5

[0122] The experimental data in the table above shows that: The sample solutions of raspberry leaf extract prepared in each embodiment showed good survival rates (survival rate above 80% with no significant cytotoxicity). They can be well applied in cosmetics.

[0123] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. A method for preparing raspberry leaf extract, characterized in that, include: Mix raspberry leaves with water to form a paste; The homogenate is mixed with a eutectic solvent for extraction; The eutectic solvent, by mass percentage, comprises the following components: L-carnitine 55%~70%, gallic acid 12%~30%, and the balance water.

2. The method for preparing raspberry leaf extract according to claim 1, characterized in that, The mixing of the homogenate with a eutectic solvent includes: The homogenate is mixed with the eutectic solvent at a ratio of (1:10) to (1:25) between the volume of the eutectic solvent and the mass of the raspberry leaves.

3. The method for preparing raspberry leaf extract according to claim 1, characterized in that, The concentration of the eutectic solvent is 25%-70% by mass percentage.

4. The method for preparing raspberry leaf extract according to claim 1, characterized in that, The eutectic solvent, by mass percentage, consists of the following components: L-carnitine 60%~65%, gallic acid 19%~23%, and the balance water.

5. The method for preparing raspberry leaf extract according to any one of claims 1-4, characterized in that, The homogenate is mixed with a eutectic solvent for extraction, including: Extraction was performed under high-voltage pulsed electric field conditions.

6. The method for preparing raspberry leaf extract according to claim 5, characterized in that, The extraction under high-voltage pulsed electric field conditions includes: Extraction was performed under electric field strength conditions of 2.5 kV / cm to 5.0 kV / cm.

7. The method for preparing raspberry leaf extract according to claim 5, characterized in that, The extraction under high-voltage pulsed electric field conditions includes: Extraction was performed under conditions of 25-75 pulses.

8. A raspberry leaf extract, characterized in that, The raspberry leaf extract was prepared using the method described in any one of claims 1-7.

9. The raspberry leaf extract according to claim 8, characterized in that, The active ingredients of the raspberry leaf extract, by weight percentage, include: 231~500 µg / mL ellagic acid and 4.6~50 µg / mL kaempferol-3-O-rutin.

10. A cosmetic product, characterized in that, The raspberry leaf extract includes the raspberry leaf extract prepared by the method described in any one of claims 1-7; or the cosmetic product includes the raspberry leaf extract described in claim 8 or 9.

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