Purification method of sulforaphane
The composite is formed by β-cyclodextrin and lysate crude product, and the oxazolidinthionine is removed by temperature change precipitation, which solves the problem of difficulty in removing oxazolidinthionine in the prior art, and realizes the simple and efficient preparation of high-purity lysate sulfane.
Patent Information
- Application Number
- CN202510901458.2
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-07-01
- Publication Date
- 2025-08-29
AI Technical Summary
The prior art is difficult to efficiently remove oxazolidinthione in lysate, resulting in low product purity, affecting safety of use, and the conventional methods are complex and costly.
β-cyclodextrin is used to mix with lysate crude product, and the hydrophobic cavity characteristics are used to load lysate sulfanane to form a composite through temperature changes, precipitation is used to remove oxazolidinthione, and then lysate lysate is extracted and separated by dichloromethane.
The efficient removal of oxazolidinthione is achieved, and the purity of lysodium sulfane is increased to more than 90%, which is simple to operate and is suitable for industrial production.
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Figure CN120554262A_ABST
Abstract
Description
Technical Field
[0001] The invention belongs to the technical field of substance purification, and particularly relates to a method for purifying sulforaphane. Background Art
[0002] Isothiocyanates, which have anti-inflammatory, anti-cancer, antioxidant, and anti-aging properties, are primarily derived from cruciferous plants. Sulforaphane (4-methylsulfinylbutyl isothiocyanate) is an important isothiocyanate compound, with the structural formula shown below:
[0003]
[0004] Broccoli is the main source of sulforaphane, but broccoli also contains 2-hydroxy-3-butenyl isothiocyanate, which can easily undergo intramolecular cyclization to form oxazolidinone (oxazolidinone may cause goiter and needs to be removed from sulforaphane). The reaction formula for intramolecular cyclization is shown below:
[0005]
[0006] During solvent extraction of broccoli, 2-hydroxy-3-butenyl isothiocyanate is easily introduced into the target product, sulforaphane. During concentration and other processes, it is cyclized to form oxazolidinone, which poses a problem for subsequent purification and product application. Oxazolidinone, an isothiocyanate derivative, has physical and chemical properties similar to those of sulforaphane, making it difficult to separate and remove from the system. Furthermore, oxazolidinone is significantly more stable than sulforaphane. During storage, oxazolidinone gradually accumulates (isothiocyanate-sulforaphane has poor stability and inevitably decomposes during separation and purification, reducing its content). This results in a continuous increase in the oxazolidinone content of the sulforaphane product, reaching 10% to 40% by weight. This seriously affects the purity of the sulforaphane product and threatens its safety. Therefore, it is necessary to remove oxazolidinone from the crude sulforaphane product.
[0007] Conventional sulforaphane preparation methods easily yield crude sulforaphane with a purity of 40% to 70%. However, sulforaphane is often used in food and pharmaceutical applications and requires purification to a purity exceeding 90%. Currently, the removal of oxazolidinethiones from sulforaphane requires methods such as silica gel column chromatography, preparative chromatography, and molecular distillation; however, these methods are complex and costly. Therefore, a simple purification method that produces high-purity sulforaphane is urgently needed. Summary of the Invention
[0008] In view of this, the object of the present invention is to provide a method for purifying sulforaphane. The method uses β-cyclodextrin to remove oxazolidinethione from sulforaphane, which has simple operation, high oxazolidinethione removal rate, and high purity of the obtained sulforaphane.
[0009] The present invention provides a method for purifying sulforaphane, comprising the following steps:
[0010] β-cyclodextrin, first water and crude sulforaphane are mixed and subjected to a first heating, followed by a first cooling and a first solid-liquid separation to obtain a sulforaphane-β-cyclodextrin complex;
[0011] mixing the sulforaphane-β-cyclodextrin complex with second water and performing a second heating, followed by a second cooling and a second solid-liquid separation to obtain a purified complex;
[0012] mixing the purified complex with dichloromethane for extraction, separating the obtained extract into solid and liquid, and removing the solvent from the obtained liquid to obtain sulforaphane;
[0013] The temperatures of the first heating and the second heating are independently 40 to 80°C.
[0014] Preferably, the dosage ratio of the β-cyclodextrin to the first water is 0.6-2 g:10 mL.
[0015] Preferably, the mass ratio of the β-cyclodextrin to the crude sulforaphane is 1:0.08-0.3.
[0016] Preferably, the temperature of the first heating is 60-65° C.; the first heating is performed under stirring, the stirring speed is 400-600 rpm, and the time is 5-30 minutes.
[0017] Preferably, the first cooling includes a first cooling step and a second cooling step performed sequentially; the temperature after the first cooling step is 10 to 30°C; after the first cooling step, the further step includes: mixing the obtained mixed system with ethanol; the volume content of ethanol in the system after mixing with ethanol is 1% to 20%; the temperature after the second cooling step is -80 to 5°C.
[0018] Preferably, the usage ratio of the sulforaphane-β-cyclodextrin complex to the second water is 0.6-2 g:10 mL.
[0019] Preferably, the temperature of the second heating is 60-65° C.; the second heating is performed under stirring, the stirring speed is 400-600 rpm, and the time is 5-30 minutes.
[0020] Preferably, the second cooling comprises a third cooling step and a fourth cooling step performed sequentially; the temperature after the third cooling step is 10 to 30°C; after the third cooling step, the further step comprises: mixing the obtained mixed system with ethanol; the volume content of ethanol in the system after mixing with ethanol is 1% to 20%; the temperature after the fourth cooling step is -80 to 5°C.
[0021] Preferably, after the second solid-liquid separation, the following steps are repeated: mixing the obtained solid and water, heating them, and then cooling and solid-liquid separation are performed in sequence; the usage ratio of the solid and water is 0.6-2g:10mL; and the heating temperature is 40-80°C.
[0022] Preferably, the number of repetitions is 1 to 4 times.
[0023] Compared with the prior art, the present invention has the following beneficial effects:
[0024] The present invention provides a method for purifying sulforaphane, comprising the following steps: mixing β-cyclodextrin, first water and a crude sulforaphane product and performing a first heating, followed by a first cooling and a first solid-liquid separation to obtain a sulforaphane-β-cyclodextrin complex; mixing the sulforaphane-β-cyclodextrin complex with a second water and performing a second heating, followed by a second cooling and a second solid-liquid separation to obtain a purified complex; mixing the purified complex with dichloromethane and performing extraction, performing solid-liquid separation on the obtained extract, and removing the solvent from the obtained liquid to obtain sulforaphane; the temperatures of the first heating and the second heating are independently 40-80°C.
[0025] The present invention utilizes the hydrophobic interior and relatively hydrophilic exterior of the β-cyclodextrin cavity to load sulforaphane into the cavity, while the relatively hydrophilic oxazolidinethione is difficult to enter the cavity. At the same time, utilizing the large temperature-dependent solubility of β-cyclodextrin, water-soluble β-cyclodextrin forms a complex with sulforaphane at a relatively high temperature (i.e., β-cyclodextrin loaded with sulforaphane), and oxazolidinethione remains in the aqueous solution. After cooling, sulforaphane and β-cyclodextrin are co-precipitated as a complex, thereby achieving the removal of oxazolidinethione from the sulforaphane product. After the β-cyclodextrin loaded with sulforaphane is purified, dichloromethane is used for extraction and separation to obtain sulforaphane. The invention utilizes the hydrophobic cavity of beta-cyclodextrin and the change of solubility with temperature to form a complex with sulforaphane and precipitate to remove oxazolidinethione. The operation is simple, the removal rate of oxazolidinethione is high, the obtained sulforaphane has high purity, and is convenient for industrial scale-up production. BRIEF DESCRIPTION OF THE DRAWINGS
[0026] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.
[0027] Figure 1 This is a liquid chromatogram of the crude sulforaphane in Example 1;
[0028] Figure 2 is the liquid chromatogram of the clear solution in Example 1;
[0029] Figure 3 This is a liquid chromatogram of the sulforaphane product after removing oxazolidinethione in Example 1. DETAILED DESCRIPTION
[0030] The present invention provides a method for purifying sulforaphane, comprising the following steps:
[0031] β-cyclodextrin, first water and crude sulforaphane are mixed and subjected to a first heating, followed by a first cooling and a first solid-liquid separation to obtain a sulforaphane-β-cyclodextrin complex;
[0032] mixing the sulforaphane-β-cyclodextrin complex with second water and performing a second heating, followed by a second cooling and a second solid-liquid separation to obtain a purified complex;
[0033] mixing the purified complex with dichloromethane for extraction, separating the obtained extract into solid and liquid, and removing the solvent from the obtained liquid to obtain sulforaphane;
[0034] The temperatures of the first heating and the second heating are independently 40 to 80°C.
[0035] In the present invention, unless otherwise specified, the materials and equipment used are commercially available products in the art.
[0036] The present invention mixes beta-cyclodextrin, first water and sulforaphane crude product, performs first heating, and then sequentially performs first cooling and first solid-liquid separation to obtain sulforaphane-beta-cyclodextrin complex; the temperature of the first heating is 40-80°C.
[0037] In the present invention, the mass content of sulforaphane in the crude sulforaphane is preferably 40% to 70%, specifically 53% or 66%; the mass content of oxazolidinethione is preferably 10% to 40%, specifically 16% or 21%.
[0038] In the present invention, the method for preparing crude sulforaphane preferably comprises the following steps: extracting broccoli with water, concentrating the resulting extract, and then extracting it with an organic solvent. The solvent is removed from the resulting organic phase to obtain crude sulforaphane. The broccoli preferably comprises seeds, flowers, stems, or leaves of broccoli; the broccoli is preferably pulverized before use; the water is preferably cold water, preferably at a temperature of 4 to 37°C, specifically 20°C; the extraction preferably includes solid-liquid separation, preferably filtration; and the organic solvent preferably comprises dichloromethane or ethyl acetate.
[0039] In the present invention, the usage ratio of the β-cyclodextrin and the first water is preferably 0.6-2 g:10 mL, specifically 1 g:10 mL or 1.2 g:10 mL.
[0040] In the present invention, the mass ratio of the β-cyclodextrin to the crude sulforaphane is preferably 1:0.08 to 0.3, specifically 1:0.12 or 1:0.15. β-cyclodextrin has a hydrophobic cavity and can bind to sulforaphane through hydrophobic interaction. The structural formula of β-cyclodextrin is shown below:
[0041]
[0042] In the present invention, the first heating temperature is preferably 60-65°C; the first heating is preferably performed under stirring, the stirring speed is preferably 400-600 rpm, specifically 500 rpm, and the duration is preferably 5-30 minutes, specifically 5 minutes or 10 minutes. During the first heating process, β-cyclodextrin and sulforaphane form a complex, i.e., β-cyclodextrin loaded with sulforaphane.
[0043] In the present invention, the β-cyclodextrin, the first water, and the crude sulforaphane are preferably mixed by first dissolving the β-cyclodextrin and the first water, followed by adding the crude sulforaphane. The dissolution is preferably performed under stirring and heating conditions, with the stirring speed preferably being 400-600 rpm, specifically 500 rpm. The heating temperature is preferably 60-65°C. Under the stirring and heating conditions, the β-cyclodextrin is completely dissolved in the water. The stirring and heating temperatures are maintained while adding the crude sulforaphane.
[0044] In the present invention, the first cooling step preferably includes a first cooling step and a second cooling step performed sequentially. The temperature after the first cooling step is preferably 10-30°C, specifically room temperature, such as 20°C or 25°C. After the first cooling step, the further step preferably includes: mixing the resulting mixed system with ethanol; the volume content of ethanol in the system after the mixing step is preferably 1%-20%, specifically 10% or 20%; the temperature after the second cooling step is preferably 4-5°C or -80-5°C, specifically 4°C or -20°C; and the temperature of the second cooling step is preferably maintained for 0.5-8 hours, specifically 2 hours. During the first cooling step, the sulforaphane-β-cyclodextrin complex precipitates, while the oxazolidinone remains in the solution, achieving separation of sulforaphane and oxazolidinone. The addition of ethanol in the present invention helps to improve the removal rate of oxazolidinone; the amount of ethanol added in the present invention achieves a high removal rate of oxazolidinone without disrupting the binding of sulforaphane and β-cyclodextrin and affecting the precipitation of the complex.
[0045] In the present invention, the first solid-liquid separation is preferably centrifugation or filtration, and the centrifugation conditions preferably include: 11000 rpm, 4°C, and 30 minutes.
[0046] After obtaining the sulforaphane-β-cyclodextrin complex, the present invention mixes the sulforaphane-β-cyclodextrin complex with second water and performs a second heating, followed by a second cooling and a second solid-liquid separation to obtain a purified complex; the second heating temperature is 40-80°C.
[0047] In the present invention, the usage ratio of the sulforaphane-β-cyclodextrin complex to the second water is preferably 0.6-2 g:10 mL, specifically 1 g:10 mL or 1.2 g:10 mL.
[0048] In the present invention, the second heating temperature is preferably 60-65°C; the second heating is preferably performed under stirring, the stirring speed is preferably 400-600 rpm, specifically 500 rpm, and the duration is preferably 5-30 minutes, specifically 5 minutes or 10 minutes. During the second heating process, the sulforaphane-β-cyclodextrin complex is completely dissolved in water.
[0049] In the present invention, the second cooling step preferably includes a third cooling step and a fourth cooling step, performed sequentially. The temperature after the third cooling step is preferably 10-30°C, specifically room temperature, such as 20°C or 25°C. The third cooling step preferably further comprises: mixing the resulting mixed system with ethanol; the volume content of ethanol in the system after mixing with ethanol is preferably 1%-20%, specifically 10% or 20%; the temperature after the fourth cooling step is preferably 4-5°C or -80-5°C, specifically 4°C or -20°C; and the temperature of the fourth cooling step is preferably maintained for 0.5-8 hours, specifically 2 hours. The second cooling step of the present invention allows the sulforaphane-β-cyclodextrin complex to precipitate. The addition of ethanol in the present invention helps lower the cooling temperature and improve the removal rate of oxazolidinethione. The amount of ethanol added in the present invention achieves a high removal rate of oxazolidinethione without disrupting the binding of sulforaphane and β-cyclodextrin, thereby affecting the precipitation of the complex.
[0050] In the present invention, the second solid-liquid separation is preferably performed by centrifugation or filtration, and the centrifugation conditions preferably include: 11000 rpm, 4°C, and 30 minutes.
[0051] In the present invention, after the second solid-liquid separation, it is preferred that the following steps are repeated: the obtained solid and water are mixed and heated, and then cooled and solid-liquid separated in sequence. The number of repetitions is preferably 1 to 4 times, specifically 2 or 3 times. The amount ratio of the solid and water is preferably 0.6 to 2 g:10 mL, specifically 1 g:10 mL or 1.2 g:10 mL. The heating temperature is preferably 40 to 80 ° C, preferably 60 to 65 ° C; the heating is preferably carried out under stirring, the stirring speed is preferably 400 to 600 rpm, specifically 500 rpm, and the time is preferably 5 to 30 minutes, specifically 5 minutes or 10 minutes.
[0052] After obtaining the purified complex, the present invention mixes the purified complex with dichloromethane for extraction, separates the obtained extract into solid and liquid, removes the solvent from the obtained liquid, and obtains sulforaphane.
[0053] In the present invention, the usage ratio of the purified complex to dichloromethane is preferably 1 g:1-5 mL, specifically 1 g:2 mL.
[0054] In the present invention, the extraction is preferably performed by shaking, and the shaking time is preferably 5 to 10 seconds.
[0055] In the present invention, the solid-liquid separation is preferably filtration.
[0056] Oxazolidinethione is difficult to remove during the purification process of sulforaphane. The present invention uses β-cyclodextrin and sulforaphane to form a host-guest structure in an aqueous solution, and then utilizes the characteristic that the solubility of β-cyclodextrin changes greatly with temperature to achieve co-precipitation of sulforaphane and β-cyclodextrin, thereby efficiently removing oxazolidinethione in the crude sulforaphane product.
[0057] To further illustrate the present invention, the purification method of sulforaphane provided by the present invention is described in detail below with reference to the accompanying drawings and examples, but they should not be construed as limiting the scope of protection of the present invention.
[0058] In the embodiments or comparative examples of the present invention, the preparation method of crude sulforaphane includes the following steps: crushing broccoli and extracting it with cold water (20° C.), filtering out solids and concentrating the solution, extracting sulforaphane with dichloromethane, and evaporating the organic phase to obtain a crude sulforaphane product with a mass content of 40% to 70%.
[0059] In the examples or comparative examples of the present invention, the oxazolidinethione removal rate = (the total mass of the oxazolidinethione remaining in the complex precipitate / the total mass of the oxazolidinethione in the crude sulforaphane) * 100%.
[0060] Example 1
[0061] The crude product of sulforaphane with a mass content of 53% and the mass content of oxazolidinethione with a mass content of 21% ( Figure 1 (This is the liquid chromatogram of crude sulforaphane).
[0062] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of β-cyclodextrin. Stir (500 rpm) and heat to 60°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.12 times the mass of β-cyclodextrin). Continue stirring for 10 minutes before removing from heat.
[0063] 2. After cooling to room temperature, ethanol was added to the solution until the volume content of ethanol was 10%, and the solution was kept at -20°C for 2 hours. The precipitate was collected by centrifugation (11000 rpm, 4°C, 30 minutes) to obtain β-cyclodextrin loaded with sulforaphane (i.e., sulforaphane-β-cyclodextrin complex) and the clear solution ( Figure 2 is the liquid chromatogram of the clear solution).
[0064] 3. Prepare a solution at a ratio of 10 mL of water per 1 g of sulforaphane-loaded β-cyclodextrin. Stir and heat to 60°C until the sulforaphane-loaded β-cyclodextrin is completely dissolved. Continue stirring for 10 minutes and then stop heating.
[0065] 4. After cooling to room temperature, ethanol was added to the solution until the volume content of ethanol was 10%. The solution was kept at -20°C for 2 hours. The precipitate was collected by centrifugation. The process was repeated twice from step 3 to obtain a purified complex with an oxazolidinethione removal rate of 95.8%.
[0066] 5. Extract with 2 mL of dichloromethane per 1 g of the purified complex (shake for 5 seconds), filter out the precipitate, and evaporate the solvent to obtain the sulforaphane product ( Figure 3 (The figure shows the liquid chromatogram of the sulforaphane product after the removal of oxazolidinethione). The purity of sulforaphane is 90.7%. It can be seen that some other impurities are removed while the oxazolidinethione is removed.
[0067] Example 2
[0068] The crude sulforaphane product with a sulforaphane mass content of 66% and the mass content of oxazolidinethione of 16% were selected.
[0069] 1. Prepare a solution at a ratio of 10 mL of water per 1.2 g of β-cyclodextrin. Stir and heat to 65°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of β-cyclodextrin) and continue stirring for 5 minutes before removing from heat.
[0070] 2. After cooling to room temperature, add ethanol to a volume content of 20% in the solution, maintain at -20°C for 2 hours, and collect the precipitate by centrifugation to obtain β-cyclodextrin loaded with sulforaphane.
[0071] 3. Prepare a solution at a ratio of 10 mL of water per 1.2 g of sulforaphane-loaded β-cyclodextrin. Stir and heat to 65°C until the sulforaphane-loaded β-cyclodextrin is completely dissolved. Continue stirring for 5 minutes and then stop heating.
[0072] 4. After cooling to room temperature, ethanol was added to the solution until the volume content of ethanol was 20%. The solution was kept at -20°C for 2 hours. The precipitate was collected by centrifugation. The process was repeated twice from step 3 to obtain a purified complex with an oxazolidinethione removal rate of 98.5%.
[0073] 5. Extraction was performed by adding 2 mL of dichloromethane per 1 g of the purified complex (shaking for 5 seconds). After filtering off the precipitate, the solvent was evaporated to dryness to obtain the sulforaphane product. The purity of the sulforaphane was 92.3%.
[0074] Example 3
[0075] The crude sulforaphane product with a sulforaphane mass content of 66% and the mass content of oxazolidinethione of 16% were selected.
[0076] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of β-cyclodextrin. Stir and heat to 60°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of β-cyclodextrin). Continue stirring for 10 minutes before removing from heat.
[0077] 2. After cooling to room temperature, keep at 4°C for 2 hours, collect the precipitate by centrifugation, and obtain β-cyclodextrin loaded with sulforaphane.
[0078] 3. Prepare a solution at a ratio of 10 mL of water per 1 g of sulforaphane-loaded β-cyclodextrin. Stir and heat to 60°C until the sulforaphane-loaded β-cyclodextrin is completely dissolved. Continue stirring for 10 minutes and then stop heating.
[0079] 4. After cooling to room temperature, the mixture was kept at 4°C for 2 hours, and the precipitate was collected by centrifugation. The process was repeated twice from step 3 to obtain a purified complex with an oxazolidinethione removal rate of 90.3%.
[0080] 5. Extraction was performed by adding 2 mL of dichloromethane per 1 g of the purified complex (shaking for 5 seconds). After filtering off the precipitate, the solvent was evaporated to dryness to obtain the sulforaphane product. The purity of the sulforaphane was 90.1%.
[0081] Compared with Example 1, in this example, ethanol was not added during the precipitation of β-cyclodextrin loaded with sulforaphane, and the removal rate of oxazolidinethione was reduced.
[0082] Comparative Example 1
[0083] The crude sulforaphane product with a sulforaphane mass content of 66% and the mass content of oxazolidinethione of 16% were selected.
[0084] Prepare a solution by adding 10 mL of 10% aqueous ethanol per 1 g of β-cyclodextrin. Stir and heat to 60°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of β-cyclodextrin) and continue stirring for 10 minutes before removing from heat.
[0085] 2. After cooling to room temperature, the mixture was kept at -20°C for 2 hours and the precipitate was collected by centrifugation. The obtained precipitate contained almost no sulforaphane (the mass content of sulforaphane in the precipitate was less than 0.2%). Sulforaphane almost remained in the clear solution and could not be separated from oxazolidinethione.
[0086] Compared with Example 1, ethanol was added when β-cyclodextrin and crude sulforaphane were mixed. The presence of ethanol at high temperature would destroy the host-guest structure, resulting in sulforaphane being unable to precipitate with β-cyclodextrin.
[0087] Comparative Example 2
[0088] The crude sulforaphane product with a sulforaphane mass content of 66% and the mass content of oxazolidinethione of 16% were selected.
[0089] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of β-cyclodextrin. Stir and heat to 60°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of β-cyclodextrin). Continue stirring for 10 minutes before removing from heat.
[0090] 2. After cooling to room temperature, ethanol was added to the solution until the volume content of ethanol was 10%. The solution was kept at -20°C for 2 hours and the precipitate was collected by centrifugation to obtain β-cyclodextrin loaded with sulforaphane. The removal rate of oxazolidinethione was only 78.0%.
[0091] Comparative Example 3
[0092] A crude sulforaphane product with a sulforaphane mass content of 66% was selected, and the mass content of oxazolidinethione was 16%.
[0093] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of β-cyclodextrin. Stir and heat to 60°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of β-cyclodextrin). Continue stirring for 10 minutes before removing from heat.
[0094] 2. After cooling to room temperature, the mixture was kept at 4°C for 2 hours and the precipitate was collected by centrifugation to obtain β-cyclodextrin loaded with sulforaphane. The removal rate of oxazolidinethione was only 67.2%.
[0095] Comparative Examples 2 to 3 did not perform subsequent precipitation operations, and the removal rate of oxazolidinethione was low.
[0096] Comparative Example 4
[0097] A crude sulforaphane product with a sulforaphane mass content of 66% was selected, and the mass content of oxazolidinethione was 16%.
[0098] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of hydroxypropyl-β-cyclodextrin. Stir and heat to 60°C until the hydroxypropyl-β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of hydroxypropyl-β-cyclodextrin). Continue stirring for 10 minutes and then stop heating.
[0099] 2. After cooling to room temperature, the mixture was kept at 4°C for 2 hours. No precipitate was formed, and the separation of sulforaphane and oxazolidinethione was not achieved.
[0100] It can be seen that, using hydroxypropyl-β-cyclodextrin, it is difficult to precipitate the complex by cooling.
[0101] Comparative Example 5
[0102] A crude sulforaphane product with a sulforaphane mass content of 66% was selected, and the mass content of oxazolidinethione was 16%.
[0103] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of methyl-β-cyclodextrin. Stir and heat to 60°C until the methyl-β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of the methyl-β-cyclodextrin) and continue stirring for 10 minutes before removing from heat.
[0104] 2. After cooling to room temperature, the mixture was kept at 4°C for 2 hours. No precipitate was formed, and the separation of sulforaphane and oxazolidinethione was not achieved.
[0105] It can be seen that, even when using methyl-β-cyclodextrin, it is difficult to precipitate the complex by cooling.
[0106] Comparative Example 6
[0107] The crude sulforaphane product with a sulforaphane mass content of 66% and the mass content of oxazolidinethione of 16% were selected.
[0108] 1. Prepare a solution at a ratio of 10 mL of water per 1 g of β-cyclodextrin. Stir and heat to 60°C until the β-cyclodextrin is completely dissolved. While maintaining stirring, add crude sulforaphane (0.15 times the mass of β-cyclodextrin). Continue stirring for 10 minutes before removing from heat.
[0109] 2. After cooling to room temperature, ethanol was added to the solution until the volume content of ethanol was 25%. The solution was kept at -20°C for 2 hours and the precipitate was collected by centrifugation. The precipitate had a sulforaphane content of less than 1% (more than 90% of the sulforaphane was distributed in the clear solution).
[0110] It can be seen that when the volume content of ethanol is greater than 20%, the binding between sulforaphane and cyclodextrin is destroyed, and sulforaphane and oxazolidinethione cannot be separated.
[0111] Although the above embodiments provide a detailed description of the present invention, they are only part of the embodiments of the present invention, not all of the embodiments. People can also obtain other embodiments based on the embodiments of the present invention without creative work, and these embodiments all fall within the scope of protection of the present invention.
Claims
1. A method for purifying sulforaphane, characterized in that: The following steps are involved: β-cyclodextrin, first water and crude sulforaphane are mixed and subjected to a first heating, followed by a first cooling and a first solid-liquid separation to obtain a sulforaphane-β-cyclodextrin complex; mixing the sulforaphane-β-cyclodextrin complex with second water and performing a second heating, followed by a second cooling and a second solid-liquid separation to obtain a purified complex; mixing the purified complex with dichloromethane for extraction, separating the obtained extract into solid and liquid, and removing the solvent from the obtained liquid to obtain sulforaphane; The temperatures of the first heating and the second heating are independently 40 to 80°C.
2. The purification method according to claim 1, wherein The dosage ratio of the β-cyclodextrin and the first water is 0.6-2 g:10 mL.
3. The purification method according to claim 1 or 2, characterized in that The mass ratio of the beta-cyclodextrin to the crude sulforaphane is 1:0.08-0.
3.
4. The purification method according to claim 1, wherein The temperature of the first heating is 60-65° C.; the first heating is performed under stirring, the stirring speed is 400-600 rpm, and the time is 5-30 minutes.
5. The purification method according to claim 1, wherein The first cooling includes a first cooling step and a second cooling step performed sequentially; the temperature after the first cooling step is 10 to 30° C.; after the first cooling step, the further step includes: mixing the obtained mixed system with ethanol; the volume content of ethanol in the system after mixing with ethanol is 1% to 20%; the temperature after the second cooling step is -80 to 5° C.
6. The purification method according to claim 1, wherein The usage ratio of the sulforaphane-β-cyclodextrin complex and the second water is 0.6-2 g:10 mL.
7. The purification method according to claim 1 or 6, characterized in that The second heating temperature is 60-65° C.; the second heating is performed under stirring, the stirring speed is 400-600 rpm, and the time is 5-30 minutes.
8. The purification method according to claim 1, wherein The second cooling includes a third cooling step and a fourth cooling step performed sequentially; the temperature after the third cooling step is 10 to 30° C.; after the third cooling step, the further step includes: mixing the obtained mixed system with ethanol; the volume content of ethanol in the system after mixing with ethanol is 1% to 20%; the temperature after the fourth cooling step is -80 to 5° C.
9. The purification method according to claim 1, wherein After the second solid-liquid separation, the method further comprises repeating the following steps: mixing the obtained solid and water, heating the mixture, and then cooling and solid-liquid separation in sequence; the usage ratio of the solid and water is 0.6-2g:10mL; and the heating temperature is 40-80°C.
10. The purification method according to claim 9, characterized in that The number of repetitions is 1 to 4 times.