Low-solvent-residue carnosic acid crystal and preparation method thereof

Abstract

The invention relates to the technical field of natural product separation, in particular to a low-solvent-residue carnosic acid crystal and a preparation method thereof. The preparation method comprises the following steps: S1, extracting, concentrating and crystallizing a carnosic acid crude product by using a first solvent to obtain a first crystal; the first solvent is a mixed solvent containing normal hexane and ethyl acetate; and S2, dissolving the first crystal by using a second solvent containing ethanol, adjusting the pH value of the dissolved solution to 4.5-5.5, concentrating, adding water, and carrying out secondary washing replacement. By optimizing the crystallization purification sequence and the solvent, the solvent residue can be effectively reduced while the carnosic acid crystal is prepared; meanwhile, the preparation method of the low-solvent-residue carnosic acid crystal is simple, easy to operate, high in yield and suitable for large-scale production.

Description

Technical Field

[0001] The invention relates to the technical field of natural product separation, and in particular to a low-soluble carnosic acid crystal and a preparation method thereof. Background Art

[0002] Carnosic acid is a phenolic diterpenoid compound, a colorless to pale yellow powder crystal, easily soluble in oils and fats, insoluble in water, and has the characteristics of safety, non-toxicity, stable structure, and high temperature resistance. Carnosic acid not only has antioxidant properties, but also can play a variety of pharmacological and biological activities such as resisting tumors and inflammation, protecting brain nerves and liver, and reducing body fat. At the same time, due to its excellent properties, it is also widely used in medicines, skin care products, daily necessities, health products, food additives and spices.

[0003] In the prior art, patent CN110283663B discloses a production process for rosemary extract, in which the rosemary extract is dissolved in salad oil containing 1% to 2% solvent and rosemary essential oil. After membrane distillation, the residual solvent in the product is ≤10ppm. Although it can meet the quality requirements to a certain extent, its carnosic acid content is low.

[0004] At present, the purification process of carnosic acid mainly adopts organic solvent recrystallization to purify and remove impurities. The main organic solvents used are n-hexane, ethyl acetate, dichloromethane and other solvents that are difficult to remove, resulting in high residual carnosic acid crystals. It takes a long time to remove the residual by vacuum drying alone, and long-term heating will cause the loss of carnosic acid content.

[0005] How to solve the above problems existing in the prior art has become a difficult research topic that needs to be solved urgently in the field of natural plant extracts. Summary of the invention

[0006] The object of the present invention is to provide a method for producing low-soluble carnosic acid crystals with simple process, easy operation and easy industrialization.

[0007] Based on this, the present invention has the following technical solutions: In a first aspect, the present invention provides a method for preparing carnosic acid crystals, comprising: S1: extracting, concentrating and crystallizing the crude carnosic acid using a first solvent to obtain first crystals; the first solvent is a mixed solvent containing n-hexane and ethyl acetate; S2: using a second solvent including ethanol to dissolve the first crystal, adjusting the pH value of the solution to 4.5-5.5, concentrating and adding water to perform washing and replacement.

[0008] The present invention finds that the first purification treatment can improve the content and yield of the first crystal, and the second purification treatment can reduce the solvent residue, and the two work together to obtain a high-content carnosic acid crystal with high yield and low solvent residue.

[0009] In the S2 of the present invention, the pH value of the dissolved solution can be adjusted to 4.5-5.5 using an alkaline solution such as an aqueous sodium carbonate solution and / or an aqueous sodium hydroxide solution. For example, the pH value can be adjusted to any value among 4.5, 4.6, 4.7, 4.8, 4.9, 5, 5.1, 5.2, 5.3, 5.4, 5.5, or a numerical range with any two of the above values ​​as endpoints.

[0010] In the present invention, the "water" used is pure water.

[0011] Preferably, in S2, an alkali solution is used to adjust the pH of the dissolved solution, and the alkali solution includes an aqueous sodium carbonate solution and / or an aqueous sodium hydroxide solution; more preferably, the concentration of the alkali solution is 5wt% to 15wt%.

[0012] The present invention finds that, during the purification process, alkaline solution is used to adjust the pH value of the dissolved solution to the above range before concentrating and crystallizing, so that carnosic acid can be easily precipitated and powdered during the concentration process, so that the solvent can be quickly removed from the carnosic acid, and carnosic acid crystals with low residual solvent can be obtained, while effectively avoiding the degradation of carnosic acid.

[0013] In the present invention, the crude carnosic acid is an existing known compound, which can be obtained through various channels, and the crude carnosic acid can be of various purities. Preferably, the purity of the crude carnosic acid is 20-30%.

[0014] In an optional embodiment of the present invention, the crude carnosic acid can be obtained by conventional extraction using rosemary leaves as raw materials and ethanol as an extracting solution in the prior art; for example, using rosemary leaves as raw materials and ethanol as an extracting solution, extracting twice or more at a certain temperature of 60-80°C and a solid-liquid ratio of (1:2-4), filtering, mixing the filtrate to concentrate to remove ethanol, and then filtering, and drying the filter cake to obtain the crude carnosic acid.

[0015] In an optional embodiment of the present invention, the crude carnosic acid is produced and provided by Chenguang Biotechnology Group Co., Ltd.

[0016] Preferably, in kg / L, the mass volume ratio of the crude carnosic acid to the first solvent is 1:(15-20), for example, it can be any ratio of 1:15, 1:16, 1:17, 1:18, 1:19 and 1:20, or a ratio range in which any two of the above ratios are endpoints.

[0017] Preferably, in the mixed solvent, the volume ratio of n-hexane to ethyl acetate is (4-5):1.

[0018] The present invention finds that using a mixed solvent of n-hexane and ethyl acetate to perform S1 on carnosic acid can improve the crystallization yield of carnosic acid at the above-mentioned lower material-liquid ratio.

[0019] Preferably, in Kg / L, the mass volume ratio of the first crystal to the second solvent is 1:(2-6), for example, it can be any ratio of 1:2, 1:3, 1:4, 1:5 and 1:6, or a ratio range in which any two of the above ratios are endpoints; a further preferred mass volume ratio is 1:6.

[0020] The present invention finds that ethanol is used as the second solvent to dissolve the first crystals passing through S1, so that ethanol is fully in contact with n-hexane and ethyl acetate, and the n-hexane and ethyl acetate wrapped in the first crystals are dispersed into the ethanol solvent, that is, the n-hexane and ethyl acetate are replaced. In the subsequent concentration process, the n-hexane and ethyl acetate can come out together with the ethanol, and finally the solvent residues of n-hexane and ethyl acetate in the refined product are effectively reduced.

[0021] Preferably, in Kg / L, the mass-to-volume ratio of the first crystal to water is 1:(4-5); a further preferred mass-to-volume ratio is 1:5.

[0022] Preferably, the method for preparing carnosic acid crystals comprises: S1: dissolving the crude carnosic acid in a mixed solvent of n-hexane and ethyl acetate at 40-60°C, concentrating the filtrate to 8-13% of the original volume of the filtrate after filtration, and then cooling to 15-35°C for crystallization, and filtering to obtain the first crystal; S2: Dissolve the first crystals with ethanol at 40-60°C, adjust the pH value of the solution to 4.5-5.5, and add water to wash and replace when the ethanol concentration is 3-7 vol%, and then continue to concentrate and filter, and dry the filter cake.

[0023] Preferably, in S1, the dissolution temperature is 50-60°C, and the dissolution time is 10-30 min; more preferably, the dissolution temperature is 60°C, and the dissolution time is 30 min.

[0024] Preferably, in S1, the temperature of cooling crystallization is 15-35°C; more preferably, 20-25°C.

[0025] Preferably, in S2, the filter cake is air-dried with an air inlet temperature of 180-200°C and an air outlet temperature of 90-105°C.

[0026] More preferably, the air inlet temperature is 190-200°C, and the air outlet temperature is 100-105°C.

[0027] In the present invention, by airflow drying the filter cake and increasing the drying temperature, the residual solvent in the crystal can be quickly taken away during concentration, which greatly shortens the drying and desolventizing time of the crystal and improves the refining efficiency.

[0028] In the present invention, the above-mentioned preparation method can be completed by using a reaction kettle conventionally used in the art.

[0029] In a second aspect, the present invention provides a carnosic acid crystal, which is prepared by the above-mentioned preparation method.

[0030] In the present invention, the total organic residual of the carnosic acid crystals is ≤50 ppm, wherein the residual concentration of ethanol is ≤10 ppm, and the residual concentration of n-hexane is ≤10 ppm.

[0031] The present invention provides a low-residual carnosic acid crystal and a preparation method thereof. By optimizing the purification sequence and the purification solvent, the carnosic acid crystal can be refined while effectively reducing the solvent residue. At the same time, the preparation method of the low-residual carnosic acid crystal provided by the present invention is simple and easy to operate, has a high yield, and is suitable for large-scale production. DETAILED DESCRIPTION

[0032] In order to make the purpose, technical solution and advantages of the present invention clearer, the technical solution of the present invention will be described clearly and completely below. Obviously, the described embodiments are part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0033] Unless otherwise specified, the various raw materials used in the examples and comparative examples are commercially available conventional raw materials, and the technical means used are conventional means well known to those skilled in the art.

[0034] Unless otherwise specified, "%" in the following examples and comparative examples refers to mass percentage.

[0035] In the following examples, the preparation of the crude carnosic acid used is: The raw material is rosemary leaves, and ethanol is used as the extracting liquid. The extraction is performed three times at 70°C according to a certain material-liquid ratio of 1:3, and the filtrate is mixed and concentrated to remove the ethanol, and then filtered. The filter cake is dried to obtain the crude carnosic acid.

[0036] In the present invention, the detection method of solvent residue is: Accurately weigh 0.1 g (accurate to 0.0001 g) of sample, add 4 mL of DMF, seal the tube, vortex to dissolve, and vortex to mix.

[0037] The chromatographic column used was a polyethylene glycol nitrobenzene modified capillary column (column length: 30 m, column inner diameter: 0.25 mm, film thickness: 0.25 µm); the carrier gas was nitrogen; the carrier gas flow rate was 3.0 mL / min; the injection port temperature was 200 °C; the detector temperature was 205 °C; the injection volume was 1 mL of the quantitative loop; the split ratio was 35:1; the column temperature was increased in a gradient manner: the initial column temperature was 60 °C, maintained for 3 min, increased to 65 °C at 5 °C / min, and then increased to 200 °C at 20 °C / min, and maintained for 5 min.

[0038] Headspace injection conditions: headspace bottle temperature is 80℃; quantitative loop temperature is 85℃; transfer line temperature is 100℃; headspace bottle equilibration time is 40min; gas phase circulation time is 30min.

[0039] Example 1 This embodiment provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 1500 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 4:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 150 L, cool to 20°C for crystallization for 8 h, filter, and obtain 32.50 kg of the first crystal.

[0040] S2: Add the first crystal to 66L 95% ethanol, adjust the pH to 4.5 with 5% sodium hydroxide solution, concentrate at 60°C to 5% ethanol, add 130L pure water, continue to concentrate and filter, and dry the filter cake with air flow at an inlet temperature of 190°C and an outlet temperature of 100°C to obtain carnosic acid crystals.

[0041] The yield of carnosic acid crystals obtained in this example is 63.80%, the content of carnosic acid is 73.8%, the total organic residue is 13 ppm, n-hexane is 4 ppm, and ethanol is 8 ppm.

[0042] Example 2 This embodiment provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 1500 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 5:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 150 L, cool to 25°C for crystallization for 8 h, filter, and obtain 32.50 kg of the first crystal.

[0043] S2: Add the first crystal to 195L 95% ethanol, adjust the pH to 5 with 5% sodium hydroxide solution, concentrate at 60°C to 5% ethanol, add 130L water, continue to concentrate and filter, dry the filter cake with air flow at an inlet temperature of 200°C and an outlet temperature of 100°C to obtain carnosic acid crystals.

[0044] The yield of carnosic acid crystals obtained in this example is 62.70%, the content of carnosic acid is 75.60%, the total organic residue is 10 ppm, n-hexane is 6 ppm, and ethanol is 3 ppm.

[0045] Example 3 This embodiment provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 1500 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 5:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 150 L, cool to 25°C for crystallization for 8 h, filter, and obtain 32.60 kg of the first crystal.

[0046] S2: Add the first crystal to 163L 95% ethanol, adjust the pH to 4.5 with 10% sodium carbonate solution, concentrate at 60°C to 5% ethanol, add 140L pure water, continue to concentrate and filter, and dry the filter cake with air flow at an inlet temperature of 200°C and an outlet temperature of 100°C to obtain carnosic acid crystals.

[0047] The yield of carnosic acid crystals obtained in this example is 62.34%, the content of carnosic acid is 73.73%, the total organic residue is 19 ppm, n-hexane is 8 ppm, and ethanol is 8 ppm.

[0048] Example 4 This embodiment provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 2000 L of a mixed solvent of n-hexane and ethyl acetate, with the volume ratio of n-hexane to ethyl acetate being 4:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 252 L, cool to 25°C for crystallization for 8 h, filter, and obtain 33.10 kg of the first crystal.

[0049] S2: Add the first crystal to 163L 95% ethanol, adjust the pH to 4.5 with 15% sodium carbonate solution, concentrate at 60°C to 5% ethanol, add 165L water, filter, and air dry the filter cake at an inlet air temperature of 200°C and an outlet air temperature of 100°C to obtain carnosic acid crystals.

[0050] The yield of carnosic acid crystals obtained in this example is 62.21%, the content of carnosic acid is 71.69%, the total organic residue is 9 ppm, n-hexane is 2 ppm, and ethanol is 5 ppm.

[0051] Example 5 This embodiment provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 2000 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 4:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 180 L, cool to 25°C for crystallization for 8 h, filter, and obtain 33.10 kg of the first crystal.

[0052] S2: Add the first crystal to 163L 95% ethanol, adjust the pH to 4.5 with 15% sodium carbonate solution, concentrate at 60°C to 5% ethanol, add 160L pure water, continue to concentrate and filter, vacuum dry the filter cake at 60°C to obtain carnosic acid crystals.

[0053] The yield of carnosic acid crystals obtained in this example is 63.30%, the content of carnosic acid is 74.30%, the total organic residue is 24 ppm, n-hexane is 4 ppm, and ethanol is 8 ppm.

[0054] Comparative Example 1 This comparative example provides a low-residue carnosic acid crystal, and the preparation method thereof is different from that of Example 1 only in that: in S1, ethyl acetate is replaced by an equal amount of n-hexane.

[0055] The yield of carnosic acid crystals obtained in this comparative example was 49.57%, the content of carnosic acid was 71.20%, the total organic residual was 60 ppm, n-hexane was 36 ppm, and ethanol was 6 ppm.

[0056] Comparative Example 2 This comparative example provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 2000 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 4:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 180 L, cool to 25°C for crystallization for 8 h, filter, and obtain 33.10 kg of the first crystal.

[0057] S2: Add the first crystal to 163L 95% ethanol, adjust the pH to 4 with 15% sodium carbonate solution, concentrate at 60°C to 5% ethanol, add 160L pure water, continue to concentrate and filter, dry the filter cake with air flow at an inlet temperature of 200°C and an outlet temperature of 100°C to obtain carnosic acid crystals.

[0058] The yield of carnosic acid crystals obtained in this comparative example is 61.94%, the content of carnosic acid is 72.78%, the total organic residue is 514 ppm, n-hexane is 324 ppm, and ethanol is 182 ppm.

[0059] Comparative Example 3 This comparative example provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 2000 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 4:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 180 L, cool to 25°C for crystallization for 8 h, filter, and obtain 33.10 kg of the first crystal.

[0060] S2: Add the first crystal to 163L 95% ethanol, adjust the pH to 6 with 15% sodium carbonate solution, concentrate at 60°C to 5% ethanol, add 160L pure water, continue to concentrate and filter, dry the filter cake with air flow, with an inlet air temperature of 200°C and an outlet air temperature of 100°C, to obtain carnosic acid crystals.

[0061] The yield of carnosic acid crystals obtained in this comparative example is 61.20%, the content of carnosic acid is 74.35%, the total organic residue is 530 ppm, n-hexane is 42 ppm, and ethanol is 486 ppm.

[0062] Comparative Example 4 This comparative example provides a low-residue carnosic acid crystal, and the preparation method thereof comprises: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 2000 L of 95% ethanol. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 180 L, cool to 25°C for crystallization for 8 h, filter, and obtain 40.31 kg of the first crystal.

[0063] S2: Add the first crystal into 163L of a mixed solvent of n-hexane and ethyl acetate, with the volume ratio of n-hexane to ethyl acetate being 4:1, and use 15% sodium carbonate solution to adjust the pH to 6. When the concentration is concentrated to 5% at 60°C, add 180L of pure water, continue to concentrate and filter, and dry the filter cake with air flow at an inlet temperature of 200°C and an outlet temperature of 100°C to obtain carnosic acid crystals.

[0064] The yield of carnosic acid crystals obtained in this comparative example was 60.74%, the content of carnosic acid was 48.67%, the total organic residual was 434 ppm, n-hexane was 420 ppm, and ethanol was 18 ppm.

[0065] Comparative Example 5 This comparative example provides a low-residue carnosic acid crystal, and the preparation method thereof comprises the following steps: S1: Add 100 kg of crude carnosic acid with a content of 22.5% to the reactor, and add 2000 L of a mixed solvent of n-hexane and ethyl acetate, with a volume ratio of n-hexane to ethyl acetate of 4:1. Dissolve at 60°C for 30 min, filter, concentrate the filtrate to 180 L, cool to 25°C for crystallization for 8 h, filter, and obtain 33.10 kg of the first crystal.

[0066] S2: Add the first crystal to 163L 95% ethanol, adjust the pH to 4.5 with 15% sodium carbonate solution, concentrate at 60°C to 5% ethanol, continue to concentrate and filter, and dry the filter cake with air flow at an inlet temperature of 200°C and an outlet temperature of 100°C to obtain carnosic acid crystals.

[0067] The yield of carnosic acid crystals obtained in this comparative example was 54.20%, the content of carnosic acid was 70.35%, the total organic residue was 780 ppm, n-hexane was 42 ppm, and ethanol was 723 ppm.

[0068] Comparative Example 6 This comparative example provides a low-residue carnosic acid crystal, and the preparation method thereof is different from that of Example 1 in that n-hexane is replaced by an equal amount of petroleum ether.

[0069] The yield of carnosic acid crystals obtained in this comparative example was 62.4%, the content of carnosic acid was 72.30%, the total organic residue was 52 ppm, n-hexane was 29 ppm, and ethanol was 21 ppm.

[0070] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit it. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art should understand that they can still modify the technical solutions described in the aforementioned embodiments, or make equivalent replacements for some of the technical features therein. However, these modifications or replacements do not deviate the essence of the corresponding technical solutions from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A method for preparing carnosic acid crystals, characterized in that: include: S1: extracting, concentrating and crystallizing the crude carnosic acid using a first solvent to obtain first crystals; the first solvent is a mixed solvent containing n-hexane and ethyl acetate; S2: using a second solvent including ethanol to dissolve the first crystal, adjusting the pH value of the solution to 4.5-5.5, concentrating and adding water to perform washing and replacement.

2. The method for preparing carnosic acid crystals according to claim 1, characterized in that: The purity of the crude carnosic acid is 10-30%.

3. The method for preparing carnosic acid crystals according to claim 1 or 2, characterized in that: In terms of Kg / L, the mass volume ratio of the crude carnosic acid to the first solvent is 1:(15-20).

4. The method for preparing carnosic acid crystals according to any one of claims 1 to 3, characterized in that: In the mixed solvent, the volume ratio of the n-hexane to the ethyl acetate is (4-5):

1.

5. The method for preparing carnosic acid crystals according to any one of claims 1 to 4, characterized in that: In terms of Kg / L, the mass volume ratio of the first crystal to the second solvent is 1:(2-6).

6. The method for preparing carnosic acid crystals according to any one of claims 1 to 5, characterized in that: In Kg / L, the mass volume ratio of the first crystal to water is 1:(4-5).

7. The method for preparing carnosic acid crystals according to any one of claims 1 to 6, characterized in that: In S2, an alkali solution is used to adjust the pH of the dissolved solution, and the alkali solution includes an aqueous sodium carbonate solution and / or an aqueous sodium hydroxide solution; preferably, the concentration of the alkali solution is 5wt%~15wt%.

8. The method for preparing carnosic acid crystals according to claims 1 to 7, characterized in that: include: S1: dissolving the crude carnosic acid in a mixed solvent of n-hexane and ethyl acetate at 40-60°C, concentrating the filtrate to 8-13% of the original volume of the filtrate after filtration, and then cooling to 15-35°C for crystallization, and filtering to obtain the first crystal; S2: Dissolve the first crystals with ethanol at 40-60°C, adjust the pH value of the solution to 4.5-5.5, and add water to wash and replace when the ethanol concentration is 3-7 vol%, and then continue to concentrate and filter, and dry the filter cake.

9. The method for preparing carnosic acid crystals according to claim 8, characterized in that: In S2, the filter cake is air dried with an air inlet temperature of 180-200°C and an air outlet temperature of 90-105°C.

10. A low-residual carnosic acid crystal, characterized in that: It is prepared by the preparation method according to any one of claims 1 to 9; preferably, the total organic residual of the carnosic acid crystals is ≤50ppm, wherein the residual concentration of ethanol is ≤10ppm, and the residual concentration of n-hexane is ≤10ppm.