A preparation method of a fat-soluble rosemary extract

Abstract

The present invention discloses a preparation method of a fat-soluble rosemary extract, which relates to the technical field of plant extracts. The preparation method includes: extracting rosemary with an acidic ethanol aqueous solution at 80-100°C, separating the first solid phase after liquid-phase de-alcoholization; mixing the first solid phase with extraction reagent A in a mass-to-volume ratio of 1 g: 10-22 L for extraction, removing the organic solvent in the extraction solution, and separating the oil phase and the second solid phase; extraction reagent A is a vegetable oil and a composite low-polar organic solvent with a volume ratio of 1:20-40; mixing the second solid phase with extraction reagent B in a mass-to-volume ratio of 1 g: 2-6 L, dissolving at 50-60°C, and performing hot solid-liquid separation, the third solid phase is the carnosol product; the liquid phase is crystallized at low temperature to obtain the rosmarinic acid product; extraction reagent B is ethyl acetate and a second low-polar organic solvent with a volume ratio of 1:2-8. Thus, high-purity carnosol and rosmarinic acid can be prepared.

Description

Technical Field

[0001] The present invention relates to the technical field of plant extracts, and in particular to a preparation method of a fat-soluble rosemary extract. Background Art

[0002] Rosemary (Rosmarinus officinalis L.) is a plant of the genus Rosmarinus in the Lamiaceae family. Its antioxidant active ingredients are divided into two categories: water-soluble components and fat-soluble components. The water-soluble components include rosmarinic acid, etc. The fat-soluble components include carnosic acid, carnosol, etc., all of which belong to diterpenoid compounds. They have good research and application prospects in aspects such as food antioxidant, anti-corrosion, improving sleep, and protecting the liver.

[0003] In related technologies, the processes for preparing fat-soluble rosemary extracts mostly rely on treatments such as column chromatography for separation, with complex procedures, low production efficiency, and insufficient utilization of resources, which easily lead to high production costs and resource waste. For example, a preparation method for separating carnosic acid from rosemary leaf extract (2011100010175) includes: dissolving the rosemary leaf extract in ethyl acetate, adding phosphoric acid-silica gel stationary phase, fully stirring and then recovering ethyl acetate to obtain the stationary phase adsorbed with the rosemary leaf extract, adding the stationary phase adsorbed with the rosemary leaf extract to a phosphoric acid-silica gel chromatography column, eluting, collecting the fractions and drying them under vacuum to obtain carnosic acid. This process has high and complex requirements for process conditions, requires self-preparation of phosphoric acid-silica gel stationary phase, and requires high-purity nitrogen protection during the process. In addition, the production efficiency is relatively low when using silica gel column chromatography for separation.

[0004] A preparation method for high-purity carnosol (2013103050811) includes: extracting and concentrating the dried raw material containing carnosol with a lipophilic organic solvent, dissolving it with a 10% - 90% hydrophilic organic solvent, and performing macroporous adsorption resin refining treatment; then, dissolving and precipitating it with a mixed lipophilic organic solvent to obtain a crude carnosol product; and then crystallizing it with a hydrophilic organic solvent to obtain high-purity carnosol crystals with a content detected by HPLC above 95%. This process requires macroporous resin treatment, which increases the complexity of the process, and the process does not consider the separation and collection of carnosic acid, resulting in resource waste.

[0005] A preparation method for simultaneously preparing high-purity carnosol and rosmarinic acid from rosemary extract (2018100928155) includes: dissolving the rosemary extract in ethyl acetate saturated with water, then mixing the sample with normal-phase silica gel and loading the column by the dry method; concentrating the eluent under reduced pressure to obtain crude carnosol with a purity of about 85% and crude rosmarinic acid with a purity of about 90%; and then separately performing Sephadex LH-20 gel column chromatography purification to obtain carnosol and rosmarinic acid with a purity above 98%. This process uses a separation and preparation method combining silica gel column chromatography and gel column chromatography, with relatively complex process conditions and low production efficiency.

[0006] Therefore, it is very important to provide a preparation method that can effectively separate carnosol and rosmarinic acid without relying on column chromatography. Summary of the Invention

[0007] The present invention aims to solve at least one of the technical problems existing in the prior art. For this purpose, the present invention provides a preparation method for a fat-soluble rosemary extract, which can effectively separate high-purity carnosol and rosmarinic acid from rosemary.

[0008] The present invention also discloses a fat-soluble rosemary extract prepared by the above preparation method.

[0009] The present invention also discloses the application of the above fat-soluble rosemary extract in the preparation of foods, health products or drugs.

[0010] A preparation method for a fat-soluble rosemary extract according to an embodiment of the first aspect of the present invention includes the following steps:

[0011] S1. Extract rosemary with an acidic ethanol aqueous solution at 80°C to 100°C to obtain an ethanol extract, take the liquid phase, remove alcohol, and perform first solid-liquid separation to obtain a first solid phase;

[0012] S2. Extract the first solid phase with extraction reagent A, perform second solid-liquid separation to obtain an extract; remove the organic solvent, and separate to obtain an oil phase and a second solid phase, and the oil phase is the oily product;

[0013] The extraction reagent A includes vegetable oil and a composite low-polarity organic solvent with a volume ratio of 1:20 to 40;

[0014] The composite low-polarity organic solvent includes ethyl acetate and a first low-polarity organic solvent with a volume ratio of 1:10 to 25;

[0015] The mass-volume ratio of the first solid phase to the extraction reagent A is 1 kg:10 L to 22 L;

[0016] S3, mixing the extraction reagent B with the second solid phase, heating at 50°C to 60°C to dissolve, performing a third solid-liquid separation while hot to obtain a third solid phase, which is the carnosin product; performing low-temperature crystallization of the liquid phase, and the precipitated crystals are the carnosic acid product;

[0017] The extraction reagent B includes ethyl acetate and a second low-polarity organic solvent;

[0018] The volume ratio of the ethyl acetate to the second low-polarity organic solvent is 1:2-8;

[0019] The mass volume ratio of the second solid phase to the extraction reagent B is 1kg:2L-6L.

[0020] The preparation method according to the embodiment of the present invention has at least the following beneficial effects:

[0021] The preparation method of the embodiment has a simple process and high production efficiency. The extraction and separation only use solvent treatment, and there is no need to use a column chromatography separation process. Moreover, the used solvents can be recycled. The production process is environmentally friendly and suitable for industrial production.

[0022] The preparation method of the embodiment can realize the full utilization of resources, and can not only obtain three specifications of fat-soluble rosemary extract products (including carnosic acid products, carnosic acid products and oily products) at the same time. Among them, the purity of carnosic acid products and carnosic acid products can reach more than 90%; the carnosic acid content of the oily product is more than 10%. In addition, step S2 can obtain a crude ursolic acid product through the second solid-liquid separation, which contains about 20% of ursolic acid and can be used as a raw material for further purification of ursolic acid.

[0023] Based on the principle of like dissolves like, a composite low-polarity organic solvent (a combination of ethyl acetate and other low-polarity organic solvents) is used to extract carnosic acid and carnosol from the fat-soluble rosemary extract, and reduce the extraction of impurities (such as ursolic acid). Adding vegetable oil can further decolorize and remove impurities, and at the same time help dissolve more impurities.

[0024] In step S3, the carnosic acid component is dissolved as much as possible by heating, which is not only conducive to the preparation of a high-purity carnosic acid product, but also conducive to improving the purity of the subsequent carnosin product. If the temperature is too low, the carnosic acid cannot be fully dissolved; if the temperature is too high, the solvent is easily volatilized and the extraction efficiency is reduced. A better separation and purification effect can be achieved by hot dissolution and cold analysis.

[0025] According to some embodiments of the present invention, the rosemary in step S1 includes rosemary leaves.

[0026] According to some embodiments of the present invention, the pH of the acidic ethanol aqueous solution is 3 to 6. Thus, the stability of carnosic acid can be improved. The pH of the acidic ethanol aqueous solution is adjusted by an acidifying agent. The acidifying agent includes at least one of hydrochloric acid, citric acid, malic acid, and L-ascorbic acid.

[0027] According to some embodiments of the present invention, in the acidic ethanol aqueous solution, the volume percentage of ethanol is 50% to 90%.

[0028] According to some embodiments of the present invention, the extraction method is the reflux extraction method.

[0029] According to some embodiments of the present invention, the extraction time is 50 min to 140 min. By performing the extraction under the conditions of 80°C to 100°C, the active ingredients can be fully extracted.

[0030] In step S1, if the ethanol in the liquid phase is not removed, it will affect the subsequent separation effect and cause waste of the solvent.

[0031] According to some embodiments of the present invention, step S1 further includes performing a decolorization treatment on the ethanol extract. The decolorization treatment time is 40 min to 80 min.

[0032] According to some embodiments of the present invention, the decolorizing agent for the decolorization treatment includes but is not limited to activated carbon, diatomaceous earth, or a combination thereof. The addition amount of the decolorizing agent is 1 g / L to 2.5 g / L.

[0033] According to some embodiments of the present invention, the mass-volume ratio of rosemary to the acidic ethanol aqueous solution is 1 kg: 6 L to 15 L. Preferably, it is 1 kg: 8 L to 12 L.

[0034] According to some embodiments of the present invention, in step S1, the number of repetitions of the extraction treatment is 2 to 3 times.

[0035] According to some embodiments of the present invention, in step S1, the method for removing alcohol includes but is not limited to vacuum concentration. The temperature for removing alcohol is 60°C to 70°C.

[0036] According to some embodiments of the present invention, step S1 further includes performing a drying treatment on the first solid phase. The drying treatment temperature is 60°C to 70°C. Thus, the influence of moisture on the extraction and separation effect can be reduced, and it is avoided that the separated rosemary oil-like product contains more moisture later, which is difficult to remove.

[0037] According to some embodiments of the present invention, in step S2, the particle size of the first solid phase is 40 mesh to 80 mesh.

[0038] According to some embodiments of the present invention, in step S2, the extraction time is 40 min to 80 min.

[0039] According to some embodiments of the present invention, the vegetable oil includes at least one of sunflower oil, pumpkin seed oil, rapeseed oil, and peanut oil.

[0040] According to some embodiments of the present invention, the first low-polarity organic solvent includes at least one of petroleum ether, n-hexane, cyclohexane, pentane, and octane.

[0041] In some preferred embodiments of the present invention, the first low-polarity organic solvent is selected from petroleum ether and octane. The volume ratio of the ethyl acetate to the first low-polarity organic solvent is 1:24 to 25. The volume ratio of the petroleum ether to the octane is 3:1 to 3. Further preferably 3:1.5 to 2.5.

[0042] In some preferred embodiments of the present invention, the first low-polarity organic solvent is selected from n-hexane and pentane. The volume ratio of the ethyl acetate to the first low-polarity organic solvent is 1:9 to 11. Further preferably 1:9.5 to 10.5. The volume ratio of the n-hexane to the pentane is 3:1 to 3. Further preferably 3:1.5 to 2.5.

[0043] In some preferred embodiments of the present invention, the first low-polarity organic solvent is selected from cyclohexane. The volume ratio of the ethyl acetate to the first low-polarity organic solvent is 1:13 to 17. Further preferably 1:14 to 16.

[0044] In some preferred embodiments of the present invention, the first low-polarity organic solvent is selected from petroleum ether. The volume ratio of the ethyl acetate to the first low-polarity organic solvent is 1:15 to 19. Further preferably 1:16 to 18.

[0045] According to some embodiments of the present invention, step S2 further includes drying the second solid phase.

[0046] According to some embodiments of the present invention, the second low-polarity organic solvent includes at least one of petroleum ether, n-hexane, cyclohexane, pentane, and octane.

[0047] In some preferred embodiments of the present invention, the second low-polarity organic solvent is selected from petroleum ether. The volume ratio of the ethyl acetate to the second low-polarity organic solvent is 1:2 to 3. Further preferably 1:2 to 2.5.

[0048] In some preferred embodiments of the present invention, the second low-polarity organic solvent is selected from cyclohexane and pentane. The volume ratio of ethyl acetate to the second low-polarity organic solvent is 1:5 to 7. More preferably 1:5.5 to 6.5. The volume ratio of cyclohexane to pentane is 1:0.5 to 2. More preferably 1:0.5 to 1.5.

[0049] In some preferred embodiments of the present invention, the second low-polarity organic solvent is selected from n-hexane and octane. The volume ratio of ethyl acetate to the second low-polarity organic solvent is 1:3 to 5. More preferably 1:3.5 to 4.5. The volume ratio of n-hexane to octane is 1:0.5 to 2. More preferably 1:0.5 to 1.5.

[0050] In some preferred embodiments of the present invention, the second low-polarity organic solvent is selected from petroleum ether and pentane. The volume ratio of ethyl acetate to the second low-polarity organic solvent is 1:7 to 8. More preferably 1:7.5 to 8. The volume ratio of petroleum ether to pentane is 3:4 to 6. More preferably 3:4.5 to 5.5.

[0051] According to some embodiments of the present invention, in step S3, the temperature of the low-temperature crystallization is not higher than 4°C.

[0052] According to some embodiments of the present invention, in step S3, the temperature of the low-temperature crystallization is 2°C to 4°C.

[0053] According to some embodiments of the present invention, in step S3, the time of the low-temperature crystallization is 12h to 24h.

[0054] According to some embodiments of the present invention, each drying treatment is independently selected from hot air drying or vacuum drying.

[0055] According to some embodiments of the present invention, the first solid-liquid separation, the second solid-liquid separation, and the third solid-liquid separation are each independently selected from centrifugation or suction filtration.

[0056] A fat-soluble rosemary extract prepared by the above preparation method according to the second aspect embodiments of the present invention.

[0057] According to some embodiments of the present invention, the fat-soluble rosemary extract includes at least one of a fat-soluble rosemary extract, a carnosol product, and an oily product.

[0058] According to some embodiments of the present invention, the content of carnosic acid in the carnosic acid product is not less than 90%. Preferably not less than 92%. More preferably not less than 94%.

[0059] According to some embodiments of the present invention, the content of carnosol in the carnosol product is not less than 90%.

[0060] According to some embodiments of the present invention, the content of carnosic acid in the oily product is 10% to 20%.

[0061] Use of the above-mentioned fat-soluble rosemary extract according to the third aspect embodiment of the present invention in the preparation of food, health products or drugs.

[0062] Other features and advantages of the present invention will be described in the following description, and some will be obvious from the description, or understood by implementing the present invention. Brief Description of the Drawings

[0063] The above and / or additional aspects and advantages of the present invention will become apparent and easy to understand from the description of the embodiments in conjunction with the following drawings, wherein:

[0064] Figure 1 is the liquid chromatogram of the carnosic acid product of Example 4 of the present invention;

[0065] Figure 2 is the liquid chromatogram of the carnosol product of Example 4 of the present invention;

[0066] Figure 3 is the liquid chromatogram of the rosemary oily product of Example 4 of the present invention. Detailed Embodiments

[0067] The following will clearly and completely describe the concept and technical effects generated by the present invention in conjunction with the embodiments to fully understand the purpose, features and effects of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Other embodiments obtained by those skilled in the art based on the embodiments of the present invention without creative efforts belong to the scope of protection of the present invention.

[0068] For those not specified in the embodiments, they are carried out under conventional conditions or conditions recommended by the manufacturer. Those reagents or instruments not specified by the manufacturer can be obtained as conventional products through commercial purchase.

[0069] In the description of the present invention, if the first, second, third, etc. are described only for the purpose of distinguishing technical features, they should not be construed as indicating or implying relative importance or implicitly indicating the quantity of the indicated technical features or implicitly indicating the sequence of the indicated technical features.

[0070] In the description of the present invention, the terms "include" and "have" and any variations thereof are intended to cover non-exclusive inclusion. For example, a process, method, system, product or device that includes a series of steps or units need not be limited to those steps or units clearly listed, but may include other steps or units not clearly listed or inherent to these processes, methods, products or devices.

[0071] Unless otherwise specified, "not less than" in the present invention means greater than and greater than or equal to.

[0072] Unless otherwise specified, "about" in the present invention means allowing an error within ±2%.

[0073] Unless otherwise specified, "room temperature" in the present invention refers to 25°C ± 5°C.

[0074] The standards of carnosic acid (purity: 98.70%) and carnosol (purity: 99.73%) were both purchased from Chengdu Mansite Biotechnology Co., Ltd.

[0075] The liquid chromatography conditions for the rosemary oil-like product, carnosol product, and carnosic acid product are as follows:

[0076] Chromatographic column: WONDASILTM C18 column (250 mm × 4.6 mm, 5 μm);

[0077] Mobile phase: acetonitrile: 1% acetic acid water = 65:35 (V / V);

[0078] Flow rate: 1 mL / min;

[0079] Detection wavelength: 285 nm;

[0080] Injection volume: 20 μL;

[0081] Sensitivity: 2.000 AUFS;

[0082] Column temperature: 25°C.

[0083] The calculation formula for the product content is as follows:

[0084]

[0085] Wherein, A 样 is the peak area of carnosic acid or carnosol in the sample solution; A 对 is the peak area of carnosic acid or carnosol in the reference solution; W 样 is the weighed amount of the sample (mg); W 对 is the weighed amount of the standard (mg); V 样 is the volume of the sample solution (mL); V 对is the volume of the reference solution (mL); K is the purity of the standard.

[0086] The preparation method of the reference solution is as follows:

[0087] Accurately weigh 2-3 mg of carnosic acid standard or carnosol standard, place in a 25 mL volumetric flask, add 20 mL of methanol and ultrasonically dissolve, cool to room temperature, make up to volume with methanol, shake well, and set aside.

[0088] The sample solution was prepared as follows:

[0089] Accurately weigh 50 mg of rosemary oil product or carnosine product or carnosic acid product (the weighing weight can be adjusted appropriately according to the sample content), place it in a 25 mL volumetric flask, add an appropriate amount of methanol to dissolve it by ultrasonication, cool to room temperature, make up to volume with methanol, shake well, filter through a 0.45 μm filter membrane, and set aside.

[0090] Example 1

[0091] This embodiment provides a method for preparing a fat-soluble rosemary extract, and the specific process is as follows:

[0092] S1. Weigh 10 kg of dry rosemary leaves in an extraction tank, add 100 L of 50% acidic ethanol aqueous solution (citric acid adjusted to pH 4.0), and reflux extract 2 times (60 min / time) at 100 ° C to obtain an extract. After the extract is double-filtered, it is transferred to a storage tank with a stirring paddle. The total volume of the extract obtained by two reflux extractions is about 170 L. Add 170 g of activated carbon to the extract, stir and decolorize for 60 min, and then filter. The filtrate is transferred to a vacuum concentrator and recovered under reduced pressure at 60 ° C to 70 ° C until there is no alcohol taste to obtain a concentrated solution. After the concentrated solution is centrifuged, the filter residue and the filtrate are collected respectively. The filter residue is dried at 65 ° C to obtain 1.85 kg of crude fat-soluble rosemary extract.

[0093] S2. The crude fat-soluble rosemary extract obtained in step S1 is crushed, passed through a 40-mesh sieve, and then 30 L of extraction reagent A is added. After extraction at room temperature for 60 min, the filtrate is filtered and concentrated under reduced pressure to be free of organic solvent, thereby separating 1.52 kg of rosemary oil product and a solid-phase fat-soluble extract. After drying the fat-soluble extract, 0.36 kg of a high-content fat-soluble rosemary extract is obtained.

[0094] The extraction solvent A is obtained by mixing sunflower oil, ethyl acetate, petroleum ether and octane in a volume ratio of 1:1:15:10.

[0095] S3. Add 0.72 L of extraction reagent B to the high-content fat-soluble rosemary extract, dissolve it in a water bath at 50 °C, and perform hot filtration while it is hot. Dry the filter residue to obtain 0.03 kg of carnosol product; refrigerate the filtrate at 4 °C for 12 h to precipitate crystals, filter, and dry the crystals to obtain 0.28 kg of rosmarinic acid product.

[0096] Among them, extraction solvent B is obtained by mixing ethyl acetate and petroleum ether in a volume ratio of 1:2.

[0097] Example 2

[0098] This example provides a preparation method of a fat-soluble rosemary extract, and the specific process is as follows:

[0099] S1. Weigh 10 kg of dry rosemary leaves into an extraction tank, add 100 L of 70% acidic ethanol aqueous solution (adjusted to pH 3.0 with hydrochloric acid), and reflux and extract at 90 °C for 2 times (60 min / time) to obtain an extract. After the extract is filtered through a double filter, transfer it to a storage tank with a stirring paddle. The total volume of the extract obtained from the two reflux extractions is about 175 L. Add 175 g of activated carbon and 175 g of diatomaceous earth to the extract, stir and decolorize for 60 min, then perform suction filtration. Transfer the filtrate to a vacuum concentrator, and recover it under reduced pressure at 60 °C - 70 °C until there is no alcohol smell to obtain a concentrated solution. After centrifuging the concentrated solution, collect the filter residue and the filtrate separately. Dry the filter residue at 65 °C to obtain 1.89 kg of crude fat-soluble rosemary extract.

[0100] S2. Crush the crude fat-soluble rosemary extract obtained in step S1, pass it through a 60-mesh sieve, add 18.9 L of extraction reagent A, stir and extract at room temperature for 60 min, perform suction filtration. After the filtrate is concentrated under reduced pressure until there is no organic solvent, separate to obtain 1.15 kg of rosemary oil-like product and a solid-phase fat-soluble extract. After drying the fat-soluble extract, obtain 0.39 kg of high-content fat-soluble rosemary extract.

[0101] Among them, extraction solvent A is obtained by mixing pumpkin seed oil, ethyl acetate, n-hexane, and pentane in a volume ratio of 1:2:12:8.

[0102] S3. Add 1.17 L of extraction reagent B to the high-content fat-soluble rosemary extract, dissolve it in a water bath at 60 °C, and perform hot filtration while it is hot. Dry the filter residue to obtain 0.03 kg of carnosol product; refrigerate the filtrate at 4 °C for 12 h to precipitate crystals, filter, and dry the crystals to obtain 0.31 kg of rosmarinic acid product.

[0103] Among them, extraction solvent B is obtained by mixing ethyl acetate, cyclohexane, and pentane in a volume ratio of 1:3:3.

[0104] Example 3

[0105] This embodiment provides a method for preparing a fat-soluble rosemary extract, and the specific process is as follows:

[0106] S1. Weigh 10 kg of dry rosemary leaves in an extraction tank, add 100 L of 80% acidic ethanol aqueous solution (malic acid adjusted to pH 5.0), and extract at 90 ° C for 2 times (60 min / time) to obtain an extract. After the extract is double-filtered, it is transferred to a storage tank with a stirring paddle. The total volume of the extract obtained by two reflux extractions is about 174 L. Add 435 g of diatomaceous earth to the extract, stir and decolorize for 60 min, and then filter. The filtrate is transferred to a vacuum concentrator and recovered under reduced pressure at 60 ° C to 70 ° C until there is no alcohol taste to obtain a concentrated solution. After the concentrated solution is centrifuged, the filter residue and the filtrate are collected respectively. The filter residue is dried at 65 ° C to obtain 1.93 kg of crude fat-soluble rosemary extract.

[0107] S2, the crude fat-soluble rosemary extract obtained in step S1 was crushed, passed through an 80-mesh sieve, 38.6 L of extraction reagent A was added, and the mixture was stirred and extracted at room temperature for 60 min, and filtered, and the filtrate was concentrated under reduced pressure to be free of organic solvent, and then separated to obtain 1.62 kg of rosemary oil product and a solid-phase fat-soluble extract, and the fat-soluble extract was dried to obtain 0.37 kg of a high-content fat-soluble rosemary extract.

[0108] The extraction solvent A is obtained by mixing rapeseed oil, ethyl acetate and cyclohexane in a volume ratio of 1:2:30.

[0109] S3. Add 1.48L of extraction reagent B to the high-fat-soluble rosemary extract, dissolve in a 50°C water bath, and filter while hot. Dry the residue to obtain 0.03kg of carnosyl phenol product; refrigerate the filtrate at 4°C for 12h, precipitate crystals, filter, and dry the crystals to obtain 0.30kg of carnosic acid product.

[0110] The extraction solvent B is obtained by mixing ethyl acetate, n-hexane and octane in a volume ratio of 1:2:2.

[0111] Example 4

[0112] This embodiment provides a method for preparing a fat-soluble rosemary extract, and the specific process is as follows:

[0113] S1. Weigh 10 kg of dry rosemary leaves in an extraction tank, add 100 L of 90% acidic ethanol aqueous solution (L-ascorbic acid adjusted to pH 6.0), and extract at 80 ° C for 2 times (60 min / time) to obtain an extract. After the extract is double-filtered, it is transferred to a storage tank with a stirring paddle. The total volume of the extracted liquid obtained by two reflux extractions is about 175 L. Add 350 g of activated carbon to the extract, stir and decolorize for 60 min, and then filter. The filtrate is transferred to a vacuum concentrator and recovered under reduced pressure at 60 ° C to 70 ° C until there is no alcohol taste to obtain a concentrated solution. After the concentrated solution is centrifuged, the filter residue and the filtrate are collected respectively. The filter residue is dried at 65 ° C to obtain 1.79 kg of crude fat-soluble rosemary extract.

[0114] S2. The crude fat-soluble rosemary extract obtained in step S1 is crushed, passed through a 60-mesh sieve, and then 27 L of extraction reagent A is added. The mixture is stirred and extracted at room temperature for 60 min, and filtered. The filtrate is concentrated under reduced pressure to be free of organic solvent, and then 1.09 kg of rosemary oil product and a solid-phase fat-soluble extract are separated. The fat-soluble extract is dried to obtain 0.36 kg of a high-content fat-soluble rosemary extract.

[0115] The extraction solvent A is obtained by mixing peanut oil, ethyl acetate and petroleum ether in a volume ratio of 1:2:35.

[0116] S3. Add 1.8 L of extraction reagent B to the high-fat-soluble rosemary extract obtained in step S2, dissolve in a 60° C. water bath, and filter while hot. Dry the filter residue to obtain 0.03 kg of carnosin product; refrigerate the filtrate at 4° C. for 12 hours, precipitate crystals, filter, and dry the crystals to obtain 0.29 kg of carnosic acid product.

[0117] The extraction reagent B is obtained by mixing ethyl acetate, petroleum ether and pentane in a volume ratio of 1:3:5.

[0118] Comparative Example 1

[0119] This comparative example provides a method for preparing a fat-soluble rosemary extract, which is different from Example 4 only in that:

[0120] (1) In step S2, the mass volume ratio of the crude fat-soluble rosemary extract to the extraction reagent A is 1 kg:8 L;

[0121] (2) In step S3, the mass volume ratio of the high fat-soluble rosemary extract to the extraction reagent B is 1 kg:1 L.

[0122] Comparative Example 2

[0123] This comparative example provides a method for preparing a fat-soluble rosemary extract, which is different from Example 4 only in that:

[0124] (1) In step S2, the mass-volume ratio of the crude fat-soluble rosemary extract to extraction reagent A is 1 kg: 22 L;

[0125] (2) In step S3, the mass-volume ratio of the high-content fat-soluble rosemary extract to extraction reagent B is 1 kg: 6 L.

[0126] Comparative Example 3

[0127] This comparative example provides a method for preparing a fat-soluble rosemary extract. Compared with Example 4, the only difference is that:

[0128] (1) In step S2, extraction reagent A is obtained by mixing peanut oil, ethyl acetate, and petroleum ether in a volume ratio of 1:1:17.5;

[0129] (2) In step S3, the mass-volume ratio of the high-content fat-soluble rosemary extract to extraction reagent B is 1 kg: 2 L;

[0130] (3) In step S3, extraction reagent B is obtained by mixing ethyl acetate and petroleum ether in a volume ratio of 1:1.

[0131] Comparative Example 4

[0132] This comparative example provides a method for preparing a fat-soluble rosemary extract. Compared with Example 4, the only difference is that:

[0133] (1) In step S2, extraction reagent A is obtained by mixing peanut oil, ethyl acetate, and petroleum ether in a volume ratio of 1:2:40;

[0134] (2) In step S3, the mass-volume ratio of the high-content fat-soluble rosemary extract to extraction reagent B is 1 kg: 2 L;

[0135] (3) In step S3, extraction reagent B is obtained by mixing ethyl acetate and petroleum ether in a volume ratio of 1:9.

[0136] Comparative Example 5

[0137] This comparative example provides a method for preparing a fat-soluble rosemary extract. Compared with Example 4, the only difference is that:

[0138] (1) Step S2 is as follows: The crude fat-soluble rosemary extract obtained in step S1 is pulverized, passed through a 60-mesh sieve, then extraction reagent A without peanut oil is added, stirred and extracted at room temperature for 60 min, filtered by suction, peanut oil is added to the filtrate and stirred at room temperature for 10 min, then concentrated under reduced pressure until there is no organic solvent, the rosemary oil-like product and the fat-soluble extract are separated, and after the fat-soluble extract is dried, a high-content fat-soluble rosemary extract is obtained.

[0139] Among them, the dosages of peanut oil, ethyl acetate, and petroleum ether remain unchanged.

[0140] Detection example

[0141] The weights of the rosemary oil products, the carnosic acid contents in the rosemary oil products, the weights of the carnosol products, the carnosol contents in the carnosol products, the weights of the carnosic acid products, and the carnosic acid contents in the carnosic acid products obtained from Examples 1 to 4 and Comparative Examples 1 to 5 were detected, and the detection results are shown in Table 1.

[0142] Table 1

[0143]

[0144]

[0145] Compared with Example 4, in Comparative Example 1, the amount of extraction reagent A in Step S2 was reduced. Since the amount of extraction reagent A was reduced, the weight of the rosemary oil product decreased; the amount of extraction reagent B in Step S3 was reduced, resulting in ineffective separation of carnosol and carnosic acid, and the carnosol content in the carnosol product and the carnosic acid content in the carnosic acid product both decreased.

[0146] Compared with Example 4, in Comparative Example 2, the amount of extraction reagent A in Step S2 was increased. Since the amount of extraction reagent A was increased, the weight of the rosemary oil product increased and the content decreased; the amount of extraction reagent B in Step S3 was increased, and the weight of the carnosic acid product decreased significantly.

[0147] Compared with Example 4, in Comparative Example 3, the content of the composite low-polar organic solvent in extraction reagent A was reduced, resulting in an increase in the weight of the rosemary oil product and a decrease in the content; the volume percentage of the second low-polar organic solvent in extraction reagent B was reduced, and the amount of extraction reagent B was reduced, making it difficult to separate carnosol and carnosic acid, and it was difficult for carnosic acid to precipitate.

[0148] Compared with Example 4, in Comparative Example 4, the content of the composite low-polar organic solvent in extraction reagent A was increased, and the volume ratio of ethyl acetate to the first low-polar organic solvent was reduced, resulting in a decrease in the weight of the rosemary oil product; the volume percentage of the second low-polar organic solvent in extraction reagent B was increased, and the amount of extraction reagent B was reduced, resulting in a significant decrease in the carnosol content in the carnosol product and a decrease in the weight of the carnosic acid product.

[0149] Comparative Example 3 increased the volume percentage of vegetable oil in Extraction Reagent A compared to Comparative Example 4, resulting in an increase in the weight of the rosemary oil-like product and a decrease in the carnosic acid content; it decreased the volume percentage of petroleum ether in Extraction Reagent B, making it difficult to separate carnosol and carnosic acid, and difficult to precipitate carnosic acid. In Comparative Example 4, due to the low volume percentage of vegetable oil in Extraction Reagent A, the weight of the rosemary oil-like product decreased; the volume percentage of petroleum ether in Extraction Reagent B was too high. Although carnosol product and carnosic acid product could be separated, the content of carnosol in the carnosol product was low.

[0150] Comparative Example 5 compared with Example 4, replaced the treatment of Extraction Solvent B with treatment with a composite low-polar organic solvent first and then with vegetable oil, resulting in a significant decrease in the yields of the carnosol product and the carnosic acid product, and also a decrease in the content of the carnosic acid product.

[0151] The embodiments of the present invention have been described in detail above in conjunction with the examples, but the present invention is not limited to the above examples. Within the scope of knowledge possessed by those of ordinary skill in the art, various changes can be made without departing from the gist of the present invention.

Claims

1. A preparation method of a fat-soluble rosemary extract, characterized in that, It consists of the following steps: S1. Extract rosemary with acidic ethanol aqueous solution at 80°C to 100°C to obtain an ethanol extract. Take the liquid phase, remove alcohol, perform the first solid-liquid separation to obtain a first solid phase; S2. Extract the first solid phase with extraction reagent A, perform the second solid-liquid separation to obtain an extract; Remove the organic solvent, separate to obtain an oil phase and a second solid phase, and the oil phase is the oily product; The extraction reagent A consists of vegetable oil and a composite low-polarity organic solvent with a volume ratio of 1:20 to 40; The composite low-polarity organic solvent consists of ethyl acetate and a first low-polarity organic solvent with a volume ratio of 1:10 to 25; The first low-polarity organic solvent includes at least one of petroleum ether, n-hexane, cyclohexane, pentane, and octane; The mass-volume ratio of the first solid phase to the extraction reagent A is 1 kg:10 L to 22 L; S3. Mix extraction reagent B with the second solid phase, heat and dissolve at 50°C to 60°C, and perform the third solid-liquid separation while it is hot to obtain a third solid phase, which is the carnosol product; Crystallize the liquid phase at low temperature, and the precipitated crystals are the rosmarinic acid product; The extraction reagent B consists of ethyl acetate and a second low-polarity organic solvent; The volume ratio of the ethyl acetate to the second low-polarity organic solvent is 1:2 to 8; The second low-polarity organic solvent includes at least one of petroleum ether, n-hexane, cyclohexane, pentane, and octane; The mass-volume ratio of the second solid phase to the extraction reagent B is 1 kg:2 L to 5 L; In step S3, the temperature of the low-temperature crystallization is not higher than 4°C.

2. The preparation method according to claim 1, characterized in that, The pH of the acidic ethanol aqueous solution is 3 to 6.

3. The preparation method according to claim 1, wherein In the acidic ethanol aqueous solution, the volume percentage of ethanol is 50% to 90%.

4. The preparation method according to claim 1, wherein The mass-volume ratio of the rosemary to the acidic ethanol aqueous solution is 1 kg:5 L to 15 L.

5. The preparation method according to claim 1, characterized in that, In step S2, the extraction time is 40 min to 80 min.

6. The preparation method according to claim 1, characterized in that, The vegetable oil includes at least one of sunflower oil, pumpkin seed oil, rapeseed oil, and peanut oil.

7. The preparation method according to claim 1, characterized in that, In step S3, the low-temperature crystallization time is 12 h to 24 h.

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