Method for extracting squalane from camellia, product and use thereof in skincare

Abstract

The application provides a method for extracting squalane from camellia as raw material, a product and an application thereof in skin care products, and belongs to the technical field of squalane extraction and purification. The method comprises the following steps: preparing camellia seed oil raw liquid through microwave pretreatment, microwave-ultrasonic synergistic extraction and magnetization extraction multiple operations; the camellia seed oil raw liquid is subjected to saponification and methyl esterification reaction with alkali-alcohol system, and saponified substances and unsaponified substances are extracted and separated; silver salt eutectic solvent is used as an extractant to synergistically extract squalene in the unsaponified substances through negative pressure cavitation, and high-purity squalene is prepared; and the high-purity squalene is hydrogenated to prepare squalane, wherein the silver salt eutectic solvent is prepared from silver nitrate and polyacrylamide. The squalane is used as a main bioactive ingredient of infant skin care products, and is used for soothing and / or repairing the skin in an irritating state. The camellia seed is synergistically treated, and the silver salt eutectic solvent is coupled with negative pressure cavitation, so that a process for improving the oil yield of camellia and the purity of squalene is provided.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of extraction and purification of squalane, and particularly relates to a method for extracting squalane from Camellia, a product and an application of the product in skin care products. BACKGROUND

[0002] Squalane is obtained by hydrogenation of squalene, and has excellent antioxidant, thermal stability and chemical stability, good affinity to skin, no allergy and irritation, and can accelerate the penetration of other active ingredients in the formula into the skin. Squalane has low polarity and moderate spreading, is pure, colorless and odorless, and can inhibit the growth of mold, so it is widely used in skin care products, especially baby care products.

[0003] Squalane is a hydrocarbon oil with excellent performance, which is obtained by hydrogenation of squalene extracted from the liver of deep-sea sharks, so it is also called deep-sea shark liver oil. Based on the protection of marine animals, squalene extraction from animals gradually changes to plant extraction. Camellia seed is rich in unsaturated fatty acids, squalene and other active ingredients, so Camellia is commonly used as a raw material for extracting squalene.

[0004] Currently, squalene is mainly obtained from Camellia oil using Camellia as a raw material, but there are the following defects:

[0005] (1) Conventional pressing or microwave / ultrasound assisted pressing is used to obtain Camellia oil from Camellia seeds, and the oil yield is low, about 20% to 25%, because the tea oil cake contains a lot of residual oil;

[0006] (2) Since the main component of Camellia oil is unsaturated fatty acid, in order to extract squalene therefrom, the existing technology generally separates most of the unsaturated fatty acid by saponification first, and then extracts squalene by silver ion complexation, but the conventional silver salt complexing extractant has low selectivity for long-chain olefins, resulting in low purity of squalene in the product, generally about 70% to 80%. SUMMARY

[0007] In view of this, the present application aims to provide a method for extracting squalane from Camellia, a product and an application of the product in skin care products, and to solve at least one technical problem in the background art.

[0008] The present application is implemented as follows:

[0009] The present application provides a method for extracting squalane from Camellia, which comprises the following steps:

[0010] The camellia seed is pre-treated by microwave, and then pressed to obtain camellia crude oil and oil tea cake; the oil tea cake is treated by microwave-ultrasonic synergistic extraction to obtain filtrate one and residue; the residue is treated by magnetization extraction to obtain filtrate two; the camellia crude oil, the filtrate one and the filtrate two are combined to form camellia seed oil original liquid;

[0011] The camellia seed oil original liquid is subjected to saponification and methyl esterification reaction with alkali-alcohol system, and the saponified matter and non-saponified matter are extracted and separated;

[0012] The non-saponified matter is mixed with an extractant to synergistically extract squalene in the non-saponified matter by negative pressure cavitation, so as to obtain high-purity squalene, wherein the extractant is a gel-type silver salt eutectic solvent prepared by taking silver nitrate and polyacrylamide as raw materials;

[0013] The high-purity squalene is hydrogenated to obtain squalane.

[0014] Further, the preparation step of the camellia seed oil original liquid specifically comprises

[0015] The shelled fresh camellia seed is pre-treated by microwave, and then pressed to obtain camellia crude oil and oil tea cake, wherein the power of the microwave pre-treatment is 200W-500W, and the time is 4min-10min;

[0016] The oil tea cake is crushed to obtain powder with a size of 50-100 meshes, the powder is mixed with an extraction solvent, and then placed in a microwave-ultrasonic extraction instrument for treatment for 10min-15min, so as to obtain filtrate one and residue after solid-liquid separation;

[0017] The residue is mixed with an organic solvent in a non-magnetic container, a magnetic field is applied to the non-magnetic container, and a stirrer is used to stir the mixture at low temperature for at least 50min, wherein the magnetic field strength is 0.2T-0.5T, and the temperature is 5℃-20℃; solid-liquid separation is performed to obtain filtrate two and residue;

[0018] The camellia crude oil, the filtrate one and the filtrate two are combined and mixed, and then centrifuged to separate liquid phase, and the solvent is removed by distillation to obtain camellia seed oil original liquid.

[0019] Further, in the microwave-ultrasonic synergistic extraction process, the ultrasonic operation is continuous, and the ultrasonic power is 50W-100W; the microwave operation is intermittent, and the microwave power is 50W-100W; the interval time between two microwave operations is 60s-120s, and the single microwave time is 30s-60s.

[0020] Further, the extraction solvent is a composite extractant of at least one polar solvent and at least one non-polar solvent;

[0021] The polar solvent adopts at least one of methanol, ethanol, isopropanol and n-butanol; the non-polar solvent adopts at least one of cyclohexane, petroleum ether, hexane and pentane; the volume ratio of the polar solvent to the non-polar solvent in the composite extractant is 1.5:1-5:1.

[0022] The organic solvent adopts a short-chain alcohol solvent.

[0023] Further, the unsaponifiable matter is mixed with the extractant, and squalene in the unsaponifiable matter is extracted by synergistic negative pressure cavitation to obtain high-purity squalene, and the method specifically comprises the following steps.

[0024] Silver nitrate, polyacrylamide and water are mixed to form a gel-type silver salt eutectic solvent.

[0025] The silver salt eutectic solvent and the unsaponifiable matter are stirred and mixed in a negative pressure cavitation extraction tank, then are fully reacted at 5-15 DEG C for 0.5-2 hours, and then are statically placed, and the liquid phase is separated, and the lower extraction phase is collected.

[0026] The non-polar solvent is added to the extraction phase, and after heating and reflux, the silver salt eutectic solvent is dissociated, the liquid phase is separated, and the organic phase containing high-purity squalene is collected.

[0027] After the organic phase is repeatedly cleaned, the non-polar solvent is removed by reduced pressure distillation, and then the organic phase is cleaned and dried to obtain high-purity squalene.

[0028] Further, the molar ratio of silver nitrate to polyacrylamide in the silver salt eutectic solvent is 1:1-1.5, and the molar ratio of silver nitrate to squalene in the unsaponifiable matter in the silver salt eutectic solvent is 1:0.5-0.8.

[0029] The application also provides squalane obtained by the method for extracting squalane from camellia seeds.

[0030] The application also provides the use of squalane in skin care products.

[0031] Further, the main biological active ingredient of the skin care product is squalane, and the infant skin care product is used for soothing and / or repairing the skin in an irritating state.

[0032] Further, the skin care product is a cream or a paste, and the skin care product is an infant skin care product.

[0033] Compared with the prior art, the application has the following beneficial effects:

[0034] 1. The application provides a process for improving the oil yield of camellia seeds and the purity of squalene by synergistically treating the camellia seeds and coupling negative pressure cavitation with a silver salt eutectic solvent.

[0035] 2. This invention utilizes multiple operations, including microwave pretreatment, microwave-ultrasound synergistic extraction, and magnetization extraction, to improve the oil yield of camellia and effectively extract residual oil from camellia seed residue.

[0036] 3. In this invention, silver nitrate and polyacrylamide are used as hydrogen bond acceptors and hydrogen bond donors, respectively, to prepare a silver salt eutectic solvent, thereby improving its complexation efficiency with squalene and thus improving the purity of squalene.

[0037] 4. The high-purity squalane obtained by this invention can be used to make infant skin care products that soothe and / or repair irritated skin, with fast and significant soothing and repairing effects. Attached Figure Description

[0038] Figure 1 This is a process flow diagram of the present invention;

[0039] Figure 2 This is a flowchart of step S1 of the present invention;

[0040] Figure 3 This is a graph showing the changes in skin hemoglobin content after using the skincare product prepared in Example 5 of the present invention;

[0041] Figure 4 This is a graph showing the change in the percentage of red area on the skin after using the skincare product prepared in Example 5 of this invention;

[0042] Figure 5 This is an application effect diagram of the skin care product prepared in Example 5 of the present invention after 5 minutes;

[0043] Figure 6 This is an image showing the application effect of the skincare product prepared in Example 5 of the present invention after 30 minutes. Detailed Implementation

[0044] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative and not intended to limit the invention.

[0045] The process flow for extracting squalane from camellia is as follows: Figure 1 As shown, the method includes the following steps:

[0046] S1. Camellia seed oil concentrate was prepared by multiple processes including microwave pretreatment, microwave-ultrasound synergistic extraction, and magnetization extraction. The specific process is as follows: Figure 2 As shown;

[0047] (1) The shelled fresh camellia seeds are microwave pretreated and then pressed to extract crude camellia oil and camellia cake.

[0048] This step involves microwave pretreatment of camellia seeds, which reduces their internal water content and disrupts the internal cell membrane, cell wall structure, and intracellular liposomes, thereby increasing the oil yield of the camellia seeds. In addition, pretreatment of whole camellia seeds does not reduce their plasticity or cause oxidation of active ingredients.

[0049] The power of microwave pretreatment is 200W to 500W, for example, it can be 200W, 300W, 400W, or 500W, but is not limited to the above values. Other unlisted values ​​are also within the scope of protection.

[0050] The microwave pretreatment time is 4 min to 10 min, for example, it can be 4 min, 6 min, 8 min, or 10 min, but it is not limited to the above values. Other unlisted values ​​are also within the protection scope.

[0051] Furthermore, this invention uses freshly picked camellia seeds because the content of bioactive components, especially squalene, in camellia seeds decreases over time.

[0052] (2) Pulverize the camellia oil cake to obtain 50-100 mesh powder. Mix the powder with the extraction solvent and place it in a microwave-ultrasonic extractor for 10-15 minutes. After solid-liquid separation, obtain filtrate 1 and filter residue.

[0053] The extraction solvent is a composite extractant consisting of at least one polar solvent and at least one non-polar solvent. The polar solvent is at least one selected from methanol, ethanol, isopropanol, and n-butanol; the non-polar solvent is at least one selected from cyclohexane, petroleum ether, hexane, and pentane. Ethanol + cyclohexane and isopropanol + cyclohexane are preferred, but not limited to the above combinations. Other combinations not listed are also within the scope of protection. The volume ratio of the polar solvent to the non-polar solvent in the composite extractant is 1.5:1 to 5:1, such as 1.5:1, 3:1, 4:1, or 5:1, but not limited to the above values. Other values ​​not listed are also within the scope of protection.

[0054] In the microwave-ultrasound synergistic extraction process, ultrasound is operated continuously with a power of 50W–100W; microwave is operated intermittently with a power of 50W–100W, with an interval of 60s–120s between microwave operations and a single microwave operation time of 30s–60s. During the microwave-ultrasound synergistic process, both microwaves and ultrasound will deform and vibrate biomolecules, further disrupting cell membrane and cell wall structures and promoting the release of active components. In addition, since microwaves also produce a thermal effect, appropriate heating can increase the thermal motion of oil molecules, promoting their diffusion into the solvent and improving extraction efficiency. However, to avoid excessively high temperatures from continuous microwave treatment that could cause thermal decomposition and thermal oxidation of oils, microwave operation is used intermittently during the synergistic extraction process.

[0055] (3) The filter residue and organic solvent are mixed in a non-magnetic container, a magnetic field is applied to the non-magnetic container, and the mixture is stirred at low temperature for at least 50 minutes using a stirrer. The magnetic field strength is 0.2T to 0.5T and the temperature is 5℃ to 20℃. The solid and liquid are separated to obtain filtrate II and residue.

[0056] Non-magnetic containers can be made of glass or other materials. The reaction apparatus containing the reactants is placed in a permanent magnetic field, and magnetic fields of different intensities are applied. A stirrer is used to magnetize the materials while stirring. During this process, the active ingredients remaining in the filter residue dissolve in the organic solvent, thus separating them.

[0057] The magnetic field strength can be 0.2T, 0.3T, 0.4T, or 0.5T, but is not limited to the above values; other unlisted values ​​are also within the protection range. The temperature can be 5℃, 10℃, 15℃, or 20℃, but is not limited to the above values; other unlisted values ​​are also within the protection range.

[0058] In practice, short-chain alcohol solvents are used as organic solvents, such as methanol, ethanol, and isopropanol.

[0059] (4) The crude camellia oil, filtrate one and filtrate two are combined and mixed, then centrifuged and separated. The liquid phase is collected and distilled to remove the solvent, thus obtaining the original camellia seed oil solution.

[0060] The liquid phases obtained in steps (1) to (3) are combined and the small particulate impurities in the mixed liquid phase are separated by low-speed centrifugation at less than 1000 rpm to obtain a pure liquid phase.

[0061] The solvent (compound extractant, organic solvent) in the liquid phase is then removed by vacuum distillation to obtain the original camellia seed oil solution.

[0062] S2. Camellia seed oil concentrate undergoes saponification and methyl esterification reactions with an alkali-alcohol system to extract and separate saponified and unsaponified products.

[0063] In this invention, the saponification and methyl esterification reactions adopt the operating steps permitted in the art, such as mixing camellia seed oil concentrate with sodium hydroxide + ethanol solution in a certain volume ratio, stirring and heating under reflux reaction, and adding n-hexane for extraction after the reaction is completed, wherein the extract phase is a non-saponifiable matter; other saponification and methyl esterification reaction processes may also be used in this invention, which are not specifically limited here.

[0064] S3. Using silver salt eutectic solvent as the extractant, squalene is extracted from unsaponifiables by synergistic negative pressure cavitation to obtain high-purity squalene. The specific steps include:

[0065] (1) Mix silver nitrate, polyacrylamide and water to form a gel-type silver salt eutectic solvent;

[0066] In the process of preparing camellia oil concentrate, multiple operations not only increase the oil yield and the amount of each component extracted, but also make the composition of camellia oil more diversified. This makes it more difficult to extract squalene from non-saponifiable matter. Since conventional silver salts or silver salt eutectic solvents dissolve olefins (especially short-chain olefins) and alkanes during extraction, the purity of squalene in the extracted product is not high. Therefore, this invention uses a gel-type silver salt eutectic solvent composed of silver nitrate and polyacrylamide to more effectively complex squalene.

[0067] In silver salt eutectic solvents, the molar ratio of silver nitrate to polyacrylamide is 1:1 to 1.5; compared with existing silver salt complexing agents, silver salt eutectic solvents have a better effect on squalene (C 30 H 50 The extraction efficiency is higher; compared with existing silver salt eutectic solvents (such as silver nitrate + acetamide), the silver salt eutectic solvent of the present invention does not require heating reaction (such as oil bath at 60℃~70℃), the preparation is simpler, and it will not form products with excessive viscosity, thus avoiding the adverse effects on subsequent complexation reactions.

[0068] (2) The silver salt eutectic solvent and the unsaponifiables are placed in a negative pressure cavitation extraction tank and stirred and mixed. Then, the mixture is allowed to react fully at 5℃~15℃ for 0.5h~2h and then allowed to stand. The liquid phases are separated and the lower extract phase is collected.

[0069] The molar ratio of silver nitrate in the silver salt eutectic solvent to squalene in the unsaponifiables is 1:0.5-0.8. Before this step, samples can be taken and the content of squalene in the unsaponifiables can be analyzed and measured using methods such as gas chromatography-mass spectrometry.

[0070] This invention combines complexation extraction with negative pressure cavitation to enhance the complexation effect of anions on squalene double bonds in silver salt eutectic solvents, thereby increasing the solubility of squalene in silver salt eutectic solvents and separating squalene from unsaponifiables.

[0071] (3) Add a non-polar solvent to the extraction phase, heat to reflux and let stand to dissociate the silver salt eutectic solvent, separate the liquid phase and collect the organic phase, which contains high-purity squalene; the non-polar solvent is at least one of cyclohexane, petroleum ether, hexane and pentane.

[0072] (4) After repeatedly washing the organic phase, the nonpolar solvent was removed by vacuum distillation, and then washed and dried to obtain high-purity squalene.

[0073] S4. Squalane is prepared by hydrogenation of high-purity squalene.

[0074] The hydrogenation reaction of squalene can be carried out using any operation permitted in the art, such as adding a hydrogenation catalyst such as Pd / C or Pt / C to an organic solvent such as toluene or n-hexane, heating to 100°C to 200°C, and adding hydrogen under high pressure, so that squalene reacts with hydrogen to produce squalane. Other hydrogenation reaction processes can also be used, and no specific limitation is made here.

[0075] The squalane obtained by the above method has high purity and can be used as a raw material for skin care products, especially for infant skin care products, such as infant skin care lotions or creams, specifically for soothing and / or repairing irritated skin.

[0076] Commonly used excipients in this field, such as solvents, moisturizers, antioxidants, thickeners, emulsifiers, pH adjusters, preservatives, fragrances, or oils (preferably camellia seed oil), may be added to infant skin care products. The emulsion is prepared using common skin care product preparation processes, and no specific limitations are made here.

[0077] Example 1

[0078] This embodiment describes a method for extracting squalane from camellia, including the following steps:

[0079] S1. Preparation of Camellia Seed Oil Stock Solution

[0080] The shelled fresh camellia seeds are pretreated in a microwave at 200W for 10 minutes, and then pressed to extract crude camellia oil and camellia cake.

[0081] The camellia oil cake was crushed to obtain 50-mesh powder. The powder was mixed with the extraction solvent and then placed in a microwave-ultrasonic extractor for 10 minutes. The extraction solvent was a mixed solvent of ethanol and cyclohexane = 1.5:1. During the microwave-ultrasonic synergistic extraction process, the ultrasonic was continuously operated at 100W. After the ultrasonic was started for 60 seconds, the microwave was started at 100W for 30 seconds. The microwave was stopped for 60 seconds and then started again. This operation was repeated until the extraction was completed. After solid-liquid separation, filtrate 1 and filter residue were obtained.

[0082] The filter residue was mixed with methanol at a liquid-to-solid ratio of 6:1 in a non-magnetic container. A magnetic field of 0.2T was applied to the non-magnetic container, and the mixture was stirred at a low temperature of 5℃ for 60 minutes. Solid-liquid separation was performed to obtain filtrate 2 and residue. The above-mentioned camellia crude oil, filtrate 1 and filtrate 2 were combined and centrifuged. The liquid phase was collected and the solvent was removed by distillation to obtain camellia seed oil concentrate. The oil yield was measured, and the results are shown in Table 1.

[0083] S2. Camellia seed oil concentrate was mixed with 0.5 mol / L sodium hydroxide + 0.9 mol / L ethanol solution at a volume ratio of 1:10. The mixture was stirred and heated to 30°C under reflux for about 1 hour. After the reaction was completed, hexane was added for extraction. The extract phase was unsaponifiable matter. The content of squalene in the unsaponifiable matter was analyzed.

[0084] S3. Silver nitrate, polyacrylamide, and water were mixed in a 1:1 molar ratio to form a gel-type silver salt eutectic solvent. The silver salt eutectic solvent and the unsaponifiables were placed in a negative pressure cavitation extraction tank and stirred. The molar ratio of silver nitrate in the silver salt eutectic solvent to squalene in the unsaponifiables was 1:0.8. The mixture was then reacted at 5°C for 2 hours and allowed to stand. The liquid phases separated, and the lower extract phase was collected. Cyclohexane was added to the extract phase, and the mixture was heated under reflux and allowed to stand to dissociate the silver salt eutectic solvent. The liquid phases separated, and the organic phase containing high-purity squalene was collected. After repeatedly washing the organic phase, the cyclohexane was removed by vacuum distillation, followed by washing and drying to obtain high-purity squalene. The purity of the product was tested, and the results are shown in Table 1.

[0085] S4. Mix high-purity squalene with toluene, add Pd / C as a catalyst, heat to 150℃, and hydrogenate under a high pressure of 6 MPa for 3 hours to react squalene with hydrogen to produce squalane.

[0086] Example 2

[0087] This embodiment describes a method for extracting squalane from camellia, including the following steps:

[0088] S1. Preparation of Camellia Seed Oil Stock Solution

[0089] The shelled fresh camellia seeds were pretreated in a microwave at 500W for 4 minutes, and then pressed to extract crude camellia oil and camellia cake.

[0090] The camellia oil cake was crushed to obtain 100-mesh powder. The powder was mixed with the extraction solvent and then placed in a microwave-ultrasonic extractor for 15 minutes. The extraction solvent was a mixture of isopropanol and cyclohexane in a ratio of 5:1. During the microwave-ultrasonic synergistic extraction process, the ultrasonic was continuously operated at 50W. After the ultrasonic was started for 60 seconds, the microwave was started at 100W for 60 seconds. The microwave was stopped for 60 seconds and then started again. This operation was repeated until the extraction was completed. After solid-liquid separation, filtrate 1 and filter residue were obtained.

[0091] The filter residue was mixed with methanol at a liquid-solid ratio of 6:1 in a non-magnetic container. A magnetic field of 0.5T was applied to the non-magnetic container, and the mixture was stirred at a low temperature of 10℃ for 70 minutes. Solid-liquid separation was performed to obtain filtrate two and residue. The above-mentioned camellia crude oil, filtrate one and filtrate two were combined and centrifuged. The liquid phase was collected and the solvent was removed by distillation to obtain camellia seed oil concentrate. The oil yield was measured, and the results are shown in Table 1.

[0092] S2. Camellia seed oil concentrate was mixed with 0.5 mol / L sodium hydroxide + 0.9 mol / L ethanol solution at a volume ratio of 1:10. The mixture was stirred and heated to 30°C under reflux for about 1 hour. After the reaction was completed, hexane was added for extraction. The extract phase was unsaponifiable matter. The content of squalene in the unsaponifiable matter was analyzed.

[0093] S3. Silver nitrate, polyacrylamide, and water were mixed in a molar ratio of 1:1.2 to form a gel-type silver salt eutectic solvent. The silver salt eutectic solvent and the unsaponifiables were placed in a negative pressure cavitation extraction tank and stirred. The molar ratio of silver nitrate in the silver salt eutectic solvent to squalene in the unsaponifiables was 1:0.6. The mixture was then reacted at 10°C for 1 hour and allowed to stand. The liquid phases separated, and the lower extract phase was collected. Cyclohexane was added to the extract phase, and the mixture was heated to reflux and allowed to stand to dissociate the silver salt eutectic solvent. The liquid phases separated, and the organic phase containing high-purity squalene was collected. After repeatedly washing the organic phase, the cyclohexane was removed by vacuum distillation, followed by washing and drying to obtain high-purity squalene. The purity of the product was tested, and the results are shown in Table 1.

[0094] S4. Mix high-purity squalene with toluene, add Pd / C as a catalyst, heat to 150℃, and hydrogenate under a high pressure of 6 MPa for 3 hours to react squalene with hydrogen to produce squalane.

[0095] Example 3

[0096] This embodiment describes a method for extracting squalane from camellia, including the following steps:

[0097] S1. Preparation of Camellia Seed Oil Stock Solution

[0098] The shelled fresh camellia seeds were pretreated in a microwave at 300W for 6 minutes, and then pressed to extract crude camellia oil and camellia cake.

[0099] The camellia oil cake was crushed to obtain 60-mesh powder. The powder was mixed with the extraction solvent and then placed in a microwave-ultrasonic extraction instrument for 15 minutes. The extraction solvent was a mixture of isopropanol and cyclohexane in a ratio of 3:1. During the microwave-ultrasonic synergistic extraction process, the ultrasonic treatment was continuously operated at 70W. After the ultrasonic treatment started for 120 seconds, the microwave treatment was started at 70W for 30 seconds. The microwave treatment was stopped for 120 seconds and then started again. This operation was repeated until the extraction was completed. After solid-liquid separation, filtrate 1 and filter residue were obtained.

[0100] The filter residue was mixed with methanol at a liquid-solid ratio of 6:1 in a non-magnetic container. A magnetic field of 0.3T was applied to the non-magnetic container, and the mixture was stirred at a low temperature of 20°C for 60 minutes. Solid-liquid separation was performed to obtain filtrate two and residue. The above-mentioned camellia crude oil, filtrate one and filtrate two were combined and centrifuged. The liquid phase was collected and the solvent was removed by distillation to obtain camellia seed oil concentrate. The oil yield was measured, and the results are shown in Table 1.

[0101] S2. Camellia seed oil concentrate was mixed with 0.5 mol / L sodium hydroxide + 0.9 mol / L ethanol solution at a volume ratio of 1:10. The mixture was stirred and heated to 30°C under reflux for about 1 hour. After the reaction was completed, hexane was added for extraction. The extract phase was unsaponifiable matter. The content of squalene in the unsaponifiable matter was analyzed.

[0102] S3. Silver nitrate, polyacrylamide, and water were mixed in a molar ratio of 1:1.5 to form a gel-type silver salt eutectic solvent. The silver salt eutectic solvent and the unsaponifiables were placed in a negative pressure cavitation extraction tank and stirred. The molar ratio of silver nitrate in the silver salt eutectic solvent to squalene in the unsaponifiables was 1:0.5. The mixture was then reacted at 15°C for 0.5 h and allowed to stand. The liquid phases separated, and the lower extract phase was collected. Cyclohexane was added to the extract phase, and the mixture was heated to reflux and allowed to stand to dissociate the silver salt eutectic solvent. The liquid phases separated, and the organic phase containing high-purity squalene was collected. After repeatedly washing the organic phase, the cyclohexane was removed by vacuum distillation, followed by washing and drying to obtain high-purity squalene. The purity of the product was tested, and the results are shown in Table 1.

[0103] S4. Mix high-purity squalene with toluene, add Pd / C as a catalyst, heat to 150℃, and hydrogenate under a high pressure of 6 MPa for 3 hours to react squalene with hydrogen to produce squalane.

[0104] Example 4

[0105] This embodiment describes a method for extracting squalane from camellia, including the following steps:

[0106] S1. Preparation of Camellia Seed Oil Stock Solution

[0107] The shelled fresh camellia seeds were pretreated in a microwave at 400W for 8 minutes, and then pressed to extract crude camellia oil and camellia cake.

[0108] The camellia oil cake was crushed to obtain 70-mesh powder. The powder was mixed with the extraction solvent and then placed in a microwave-ultrasonic extraction instrument for 14 min. The extraction solvent was a mixture of isopropanol and cyclohexane in a ratio of 4:1. During the microwave-ultrasonic synergistic extraction process, the ultrasonic treatment was continuously operated at 80W. After the ultrasonic treatment started for 100 s, the microwave treatment was started at 80W for 45 s. The microwave treatment was stopped for 100 s and then started again. This operation was repeated until the extraction was completed. After solid-liquid separation, filtrate 1 and filter residue were obtained.

[0109] The filter residue was mixed with methanol at a liquid-solid ratio of 6:1 in a non-magnetic container. A magnetic field of 0.4T was applied to the non-magnetic container, and the mixture was stirred at a low temperature of 15℃ for 60 minutes. Solid-liquid separation was performed to obtain filtrate two and residue. The above-mentioned camellia crude oil, filtrate one and filtrate two were combined and centrifuged. The liquid phase was collected and the solvent was removed by distillation to obtain camellia seed oil concentrate. The oil yield was measured, and the results are shown in Table 1.

[0110] S2. Camellia seed oil concentrate was mixed with 0.5 mol / L sodium hydroxide + 0.9 mol / L ethanol solution at a volume ratio of 1:10. The mixture was stirred and heated to 30°C under reflux for about 1 hour. After the reaction was completed, hexane was added for extraction. The extract phase was unsaponifiable matter. The content of squalene in the unsaponifiable matter was analyzed.

[0111] S3. Silver nitrate, polyacrylamide, and water were mixed in a molar ratio of 1:1.4 to form a gel-type silver salt eutectic solvent. The silver salt eutectic solvent and the unsaponifiables were placed in a negative pressure cavitation extraction tank and stirred. The molar ratio of silver nitrate in the silver salt eutectic solvent to squalene in the unsaponifiables was 1:0.7. The mixture was then reacted at 15°C for 0.5 h and allowed to stand. The liquid phases separated, and the lower extract phase was collected. Cyclohexane was added to the extract phase, and the mixture was heated to reflux and allowed to stand to dissociate the silver salt eutectic solvent. The liquid phases separated, and the organic phase containing high-purity squalene was collected. After repeatedly washing the organic phase, the cyclohexane was removed by vacuum distillation, followed by washing and drying to obtain high-purity squalene. The purity of the product was tested, and the results are shown in Table 1.

[0112] S4. Mix high-purity squalene with toluene, add Pd / C as a catalyst, heat to 150℃, and hydrogenate under a high pressure of 6 MPa for 3 hours to react squalene with hydrogen to produce squalane.

[0113] Comparative Example 1

[0114] The method for extracting squalane from camellia in this comparative example differs from that in Example 1 in step S3. Other steps and parameters are the same as in Example 1. In step S3 of this comparative example, the complexation reaction between the silver salt eutectic solvent and the unsaponifiables is carried out in a conventional environment, rather than by negative pressure cavitation. The oil yield and squalene purity were measured, and the results are shown in Table 1.

[0115] Comparative Example 2

[0116] The method for extracting squalane using camellia as raw material in this comparative example differs from that in Example 1 in step S3. The other steps and parameters are the same as in Example 1. In step S3 of this comparative example, the raw materials for the silver salt eutectic solvent are silver nitrate and acetamide. The synthesis steps are as follows: silver nitrate and acetamide are mixed in a 1:1 molar ratio and added to a three-necked flask. After stirring thoroughly for 1.5 hours in an oil bath at 65°C in the dark, a pale yellow viscous liquid is formed, which is the silver salt eutectic solvent. The oil yield and squalene purity are measured, and the results are shown in Table 1.

[0117] Comparative Example 3

[0118] The method for extracting squalane from camellia in this comparative example differs from that in Example 1 in step S1. All other steps and parameters are the same as in Example 1. Specifically, step S1 in this comparative example is as follows:

[0119] Fresh camellia seeds after shelling were pretreated in a microwave at 200W for 10 minutes, and then pressed to obtain crude camellia oil and camellia cake. The crude camellia oil was centrifuged to obtain camellia seed oil concentrate. The oil yield and squalene purity were measured, and the results are shown in Table 1.

[0120] Comparative Example 4

[0121] The method for extracting squalane from camellia in this comparative example differs from that in Example 1 in step S1. Other steps and parameters are the same as in Example 1. Step S1 in this comparative example only includes microwave pretreatment of fresh camellia seeds and microwave-ultrasonic extraction synergistic treatment, and the magnetization treatment step of the filter residue is deleted.

[0122] The above-mentioned crude camellia oil and filtrate were combined and centrifuged. The liquid phase was collected and distilled to remove the extraction solvent, thus obtaining the original camellia seed oil. The oil yield and squalene purity were measured, and the results are shown in Table 1.

[0123] Comparative Example 5

[0124] The method for extracting squalane from camellia in this comparative example differs from that in Example 1 in step S1. The other steps and parameters are the same as in Example 1. In step S1 of this comparative example, the microwave-ultrasonic extraction instrument is operated with simultaneous and continuous microwave and ultrasonic operation to measure the oil yield and squalene purity. The results are shown in Table 1.

[0125] Comparative Example 6

[0126] The method for extracting squalane from camellia in this comparative example differs from that in Example 1 in step S1. The other steps and parameters are the same as in Example 1. In step S1 of this comparative example, the filter residue is dissolved in methanol in a normal way, rather than being magnetized. The oil yield and squalene purity were measured, and the results are shown in Table 1.

[0127] Table 1

[0128] Test subject Oil out rate / % Squalene purity % Example 1 29.1 97.0 Example 2 29.4 97.3 Example 3 29.7 97.2 Example 4 29.5 97.0 Comparative Example 1 29.1 83.6 Comparative Example 2 29.1 81.4 Comparative Example 3 25.2 97.1 Comparative Example 4 27.1 97.0 Comparative Example 5 26.9 96.2 Comparative Example 6 27.0 97.0

[0129] Oil yield (%) = weight of raw camellia seed oil / weight of camellia seeds * 100.

[0130] As shown in Table 1, the embodiments of the present invention can significantly improve the oil yield of camellia seed oil and the extracted squalene has high purity.

[0131] Comparing Comparative Example 1 and Example 1, it can be seen that when silver salt eutectic solvent is used to extract squalene, the negative pressure cavitation assisted by Example 1 can significantly improve the complexation efficiency between silver salt eutectic solvent and squalene, and the purity of the squalene obtained is higher than that of Comparative Example 1.

[0132] Comparing Comparative Example 2 with Example 1, it can be seen that Example 1 uses polyacrylamide as a hydrogen bond donor, while Comparative Example 2 uses acetamide as a hydrogen bond donor. Example 1 can significantly improve the complexation efficiency of silver salt eutectic solvent and squalene, and the purity of the squalene obtained is higher than that of Comparative Example 2.

[0133] Data from Comparative Examples 3 to 5 show that microwave pretreatment, microwave-ultrasound synergistic extraction, and magnetization extraction can all improve the oil yield. The oil yield of the products in Comparative Examples 3 to 5 is lower than that in Example 1. Furthermore, in Comparative Example 5, simultaneous microwave-ultrasound extraction led to the thermal oxidation of fatty acids, which indirectly resulted in a slight decrease in product purity.

[0134] Example 5

[0135] I. Skin irritation test

[0136] Using the squalane obtained in Example 1 as the main active ingredient, an emulsion was prepared with excipients. The emulsion contained 5-30% squalane. In this example, an emulsion containing 10% squalane was selected as the test object.

[0137] Rabbits were used as test animals, and 5 groups were set up, with 2 male rabbits and 2 female rabbits in each group, weighing 2.5kg to 2.7kg. The safety of the emulsion was tested according to the requirements of the "Cosmetic Safety Technical Specifications" (2015 edition), and the results are shown in Table 2.

[0138] Table 2

[0139]

[0140] The data in Table 2 show that the emulsion of this invention showed no irritation in multiple skin irritation tests.

[0141] II. Security Testing

[0142] Subjects aged 19 to 36 years (half male and half female) were selected, and the safety of the samples was evaluated in accordance with Chapter 7 of the "Cosmetic Safety Technical Specifications (2015 Edition)" regarding human skin patch testing.

[0143] Using qualified patch testing equipment, and employing the closed patch test method, 0.020g to 0.025g of squalane prepared in Example 1 was placed in the patch testing device as the test group and applied to the subject's forearm when bent forward. Water was used as the control group. The test substance was removed after 24 hours, and skin reactions were observed at 0.5, 24, and 48 hours after removal. The results were recorded according to the skin reaction grading standards in the currently effective technical specifications, as shown in Table 3.

[0144] Table 3

[0145]

[0146] Table 3 shows that the human skin patch test showed no adverse reactions and was safe.

[0147] III. Moisturizing Efficacy Test

[0148] The moisturizing efficacy of the squalane prepared in Example 1 was tested according to QB / T4256-2011 "Guidelines for Evaluation of Moisturizing Efficacy of Cosmetics".

[0149] 1. Subjects: 24 subjects, aged 18-60 years, with no statistically significant differences in age, gender, etc., and meeting the inclusion criteria. Subjects waited at least 20 minutes in the experimental environment, and a test area of ​​3cm*3cm was marked. After adaptation, the moisture content of the stratum corneum of their forearm skin was measured. Based on the screening results, a sufficient number of subjects with skin stratum corneum moisture content meeting the set subject number requirements were finally selected. Subjects whose baseline values ​​of the forearm test area using the capacitive skin moisture meter were between 15 and 45 (Corneometer Units, CU) were considered to meet the criteria.

[0150] 2. Test samples: Squalane prepared in Example 1 was used as the product group sample, and olive squalane was used as the positive control group sample.

[0151] 3. Testing environment: The test results should be observed in an environment with a temperature of (21±1)℃ and a relative humidity of (50±10)%RH, and real-time dynamic monitoring should be carried out. The testing conditions should be kept consistent throughout the testing process, such as the tester, the location, and the instruments. All subjects should adapt to this environment for at least 20 minutes before the test can be conducted.

[0152] 4. Test method: Before the eligible subjects included in the group use the product, wipe the arm with a dry tissue paper, wait for more than 20 minutes in the environmental test chamber, do not drink water or beverages, expose the forearm, place it in a test state, and keep relaxed; after adaptation, measure the water content of the stratum corneum of the skin in the test sample application area and the control area respectively, measure 3 times in parallel within each area, and the measurement results are expressed as the average value of the 3 measurements, which is used as the initial value; the sample is applied once at a dosage of (2.0 ± 0.1) mg / cm 2 by quantitative sampling through a syringe or equivalent, and use a latex finger cot to evenly apply the sample within the specified area and record the actual application amount; after the sample is applied, test the water content of the stratum corneum at 2h, 4h, and 8h set, and the test results are shown in Table 4, and the significant comparison results of the test results are shown in Table 5.

[0153] Table 4

[0154]

[0155] Table 5

[0156]

[0157] Note: n.s indicates no statistically significant difference or no significant difference, at this time P > 0.05; if P ≤ 0.05, it indicates a statistically significant difference or a significant difference. * indicates 0.01 < P ≤ 0.05; ** indicates 0.001 < P ≤ 0.01; *** indicates P ≤ 0.001. When the percentage change in the average value is positive, the direction of the difference is positive, otherwise it is negative.

[0158] After the moisturizing efficacy test, the results show that: (1) There are no cases of adverse reactions in the subjects of this test, and the test product has high safety; (2) The test of the water content of the stratum corneum shows that after the subjects use the product, the water content of the stratum corneum in the product test area at the follow-up points (2h, 4h, 8h) has an upward trend compared with the initial value, and there is no significant difference in the change of the water content of the stratum corneum in the positive control group area of olive squalane, and there is a significant difference in the change of the water content of the stratum corneum in the blank control group area.

[0159] To sum up, using the camellia squalane product for 8 hours can increase the water content of the stratum corneum of the skin and achieve the effect of moisturizing, indicating that the product has a moisturizing effect, and there is no obvious difference between camellia squalane and olive squalane in terms of moisturizing performance.

[0160] IV. Soothing and repairing efficacy test

[0161] Conduct the soothing and repairing efficacy test on the test product in accordance with the "Operating Instruction for the Evaluation of the Soothing Human Body Efficacy of Cosmetics" and the "Operating Instruction for the Evaluation of the Special Declared Efficacy of Cosmetics".

[0162] 1. Subjects: 30 subjects aged 18-55 years, with no statistically significant differences in age, gender, etc.; all subjects had at least one symptom of impaired skin barrier function, such as dryness, peeling, or redness, and met the inclusion criteria.

[0163] 2. Using the squalane obtained in Example 1 as the main active ingredient, an emulsion was prepared with excipients. The emulsion contained 5-30% squalane. In this example, an emulsion containing 10% squalane was selected as the test product.

[0164] 3. Testing environment: The test results should be observed in an environment with a temperature of (21±1)℃ and a relative humidity of (50±10)%RH, and real-time dynamic monitoring should be carried out. The testing conditions should be kept consistent throughout the testing process, such as the tester, the location, and the instruments. All subjects should adapt to this environment for at least 20 minutes before the test can be conducted.

[0165] 4. Testing Method: Before using the product, qualified subjects should wipe their arms with a dry paper towel and wait in the environmental testing room for at least 20 minutes. They should not drink water or beverages, and should remain relaxed with their faces exposed, avoiding contact with the skin. Lactic acid stinging tests should be performed on both cheeks to induce facial immune erythema. After adaptation, baseline assessments should be conducted, and the skin hemoglobin content on the face should be measured as the initial value. Assessments and tests should be performed 5 minutes and 30 minutes after product use. Changes in skin hemoglobin content should be recorded. Figure 3 As shown; using VISIA7, the red areas of the subject's face (front, left, and right sides) were photographed, and the proportion of the red area was calculated. The results of this proportion change are shown below. Figure 4 As shown.

[0166] Before and after effects are as follows: Figure 5 and Figure 6 As shown; Figure 5 (a) shows the test area 5 minutes after applying the emulsion of this embodiment. Figure 5 (b) is the blank control area. Figure 6 (a) shows the test area 30 minutes after applying the emulsion of this embodiment. Figure 6 (b) is the blank control area.

[0167] Depend on Figures 3 to 6 As can be seen, after 5 minutes and 30 minutes of use, the skin hemoglobin content and the proportion of red area were significantly reduced. Therefore, the lotion in this embodiment has the effect of relieving skin irritation such as immune inflammation, and has a soothing and repairing effect. Moreover, the effect is enhanced with the increase of use time.

[0168] The embodiments described above are merely illustrative of several implementations of the present invention, and while the descriptions are specific and detailed, they should not be construed as limiting the scope of the present invention. It should be noted that those skilled in the art can make various modifications and improvements without departing from the concept of the present invention, and these modifications and improvements all fall within the scope of protection of the present invention. Therefore, the scope of protection of this patent should be determined by the appended claims.

Claims

1. A method for extracting squalane from camellia, characterized in that, The method includes the following steps: S1. First, microwave pre-treat whole camellia seeds, then press to obtain crude camellia oil and camellia cake. Next, microwave-ultrasonic synergistic extraction is performed on the camellia cake to obtain filtrate one and residue. Then, magnetization extraction is used to treat the residue to obtain filtrate two. The crude camellia oil, filtrate one, and filtrate two are combined to form camellia seed oil concentrate. Specifically, this includes: The shelled fresh camellia seeds are microwave pretreated and then pressed to extract crude camellia oil and camellia cake. The microwave pretreatment power is 200W~500W and the time is 4min~10min. Camellia oleifera cake is pulverized to obtain a powder of 50-100 mesh. The powder is mixed with an extraction solvent and then treated in a microwave-ultrasonic extraction instrument for 10-15 minutes. After solid-liquid separation, filtrate and filter residue are obtained. The extraction solvent is a composite extractant consisting of at least one polar solvent and at least one non-polar solvent. The polar solvent is at least one of methanol, ethanol, isopropanol, and n-butanol. The non-polar solvent is at least one of cyclohexane, petroleum ether, hexane, and pentane. The volume ratio of the polar solvent to the non-polar solvent in the composite extractant is 1.5:1 to 5:

1. The filter residue is mixed with an organic solvent in a non-magnetic container. A magnetic field is applied to the non-magnetic container, and the mixture is stirred at a low temperature for at least 50 minutes using a stirrer. The magnetic field strength is 0.2T~0.5T, and the temperature is 5℃~20℃. Solid-liquid separation is performed to obtain filtrate and residue. The organic solvent used is a short-chain alcohol. The crude camellia oil, filtrate one and filtrate two were combined and mixed, then centrifuged to separate them. The liquid phase was collected and distilled to remove the solvent, thus obtaining the original camellia seed oil solution. S2. Camellia seed oil concentrate undergoes saponification and methyl esterification reactions with an alkali-alcohol system to extract and separate saponified and unsaponified products. S3. The unsaponifiables are mixed with the extractant, and squalene in the unsaponifiables is extracted by synergistic negative pressure cavitation to obtain high-purity squalene. The extractant is a gel-type silver salt eutectic solvent prepared from silver nitrate and polyacrylamide. Specifically, it includes: Silver nitrate, polyacrylamide, and water are mixed to form a gel-type silver salt eutectic solvent; The silver salt eutectic solvent and the unsaponifiables are placed in a negative pressure cavitation extraction tank and stirred. The mixture is then allowed to react fully at 5℃~15℃ for 0.5h~2h, and then allowed to stand. The liquid phases separate, and the lower extract phase is collected. A nonpolar solvent was added to the extraction phase, and the mixture was heated to reflux and allowed to stand. The silver salt eutectic solvent was dissociated, the liquid phase was separated, and the organic phase was collected. The organic phase contained high-purity squalene. After repeatedly washing the organic phase, the nonpolar solvent was removed by vacuum distillation, followed by washing and drying to obtain high-purity squalene. S4. Squalane is prepared by hydrogenation of high-purity squalene.

2. The method for extracting squalane from camellia as a raw material according to claim 1, characterized in that, In the microwave-ultrasound synergistic extraction process, the ultrasound is operated continuously with an ultrasound power of 50W~100W; the microwave is operated intermittently with a microwave power of 50W~100W, the interval between two microwave operations is 60s~120s, and the duration of a single microwave operation is 30s~60s.

3. The method for extracting squalane from camellia as a raw material according to claim 1, characterized in that, In the silver salt eutectic solvent, the molar ratio of silver nitrate to polyacrylamide is 1:1 to 1.5; the molar ratio of silver nitrate to squalene in the unsaponifiables is 1:0.5 to 0.8.

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