Method for extracting isothiocyanate from cured mustard seed cake

Abstract

The invention discloses a method for extracting isothiocyanate from a cured mustard seed cake, which comprises the following steps of: performing high-pressure treatment and superfine grinding on the cured mustard seed cake obtained by performing cold pressing on mustard seeds and curing to obtain micro powder; ultrasonically dispersing the micro powder in water, freezing and redissolving, and treating by a pulsed electric field to obtain a pretreated mixture; and finally, carrying out supercritical extraction on the pretreated mixture to obtain the isothiocyanate. The method disclosed by the invention is green and environment-friendly, and the isothiocyanate is high in yield and good in quality.

Description

Technical Field

[0001] The present invention relates to a method for extracting isothiocyanates from ripened mustard seed cakes, belonging to the technical field of natural extraction. Background Art

[0002] Mustard seeds are an oil crop with a rapeseed oil content of over 30%. The residual mustard seed cakes after oil extraction are rich in protein and are good components for animal feed. Mustard seeds contain 2-propenyl glucosinolate, which hydrolyzes to form isothiocyanates under the action of myrosinase.

[0003] The skeleton of the isothiocyanate component is -N=C=S. The C on the skeleton has high electrophilicity and can undergo nucleophilic addition reactions with nucleophiles (such as amino groups, thiols, carbonyl groups, carboxylic acids, etc.). Therefore, it can react with molecules such as water and alcohols, and its thermal stability is not high.

[0004] Isothiocyanates can participate in various organic reactions and are used to synthesize various types of sulfur, nitrogen, and oxygen-containing compounds, especially heterocyclic compounds. Isothiocyanates can be widely used in the preparation of organic synthesis products, such as pharmaceuticals, cosmetic raw materials, pesticides, dyes, etc. Isothiocyanates can also be used to determine the amino acid sequences in polypeptides and proteins and are used as fluorescent markers. Isothiocyanates have a strong spicy taste, can prevent tooth decay, stimulate the secretion of saliva and gastric juice, and enhance appetite. Moreover, isothiocyanates have very good medicinal value. Research shows that isothiocyanates can prevent cancers caused by DNA damage induced by various carcinogens in food, such as polycyclic aromatic hydrocarbons, heterocyclic amines, and nitrosamines, and can accelerate the excretion of the above carcinogens, thus playing a role in preventing cancer. In addition, isothiocyanates can also prevent blood clots and have a certain effect on the treatment of asthma. Isothiocyanates have very good application prospects in the above many fields. Extracting isothiocyanates from mustard seed cakes not only solves the problem of waste pollution but also realizes the recovery of beneficial components, meeting the current environmental protection concept.

[0005] However, myrosinase is easily denatured and inactivated by heat and will start to lose its activity due to protein denaturation in an environment above 45°C; the product isothiocyanates are also easily volatilized by heat. The hot pressing method used in traditional oil extraction processes will undoubtedly affect the yield of isothiocyanates.

[0006] Patent application CN104710335A discloses a method for extracting mustard oil from radishes, including the following steps: raw material pretreatment, solvent extraction, filtration adsorption, centrifugal extraction, and concentration. This patented technology mainly extracts the isothiocyanate component from radishes through ethanol extraction, with a large consumption of ethanol and being less environmentally friendly. Summary of the Invention

[0007] The object of the present invention is to overcome the deficiencies of the above-mentioned prior art and provide a method for extracting isothiocyanates from ripened mustard seed cake, which is green and environmentally friendly, has a high yield of isothiocyanates and good quality.

[0008] To achieve the above object, the present invention adopts the following technical solutions:

[0009] A method for extracting isothiocyanates from ripened mustard seed cake, the specific steps are as follows:

[0010] (1) First, the ripened mustard seed cake obtained by cold pressing mustard seeds and ripening is subjected to high-pressure treatment and ultrafine grinding to obtain fine powder.

[0011] (2) Then, the fine powder is ultrasonically dispersed in water, subjected to freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture.

[0012] (3) Finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates.

[0013] Preferably, in step (1), a screw oil press is used for cold pressing, and the cold pressing temperature is 40-43 °C.

[0014] Preferably, in step (1), the ripening method is: microwave irradiation at 300-350 W for 2-3 minutes, stopping irradiation for 1 minute, continuing microwave irradiation at 300-350 W for 2-3 minutes, and standing at 10-15 °C for 2-3 days.

[0015] Preferably, in step (1), the high-pressure treatment conditions are: holding pressure at 300-400 MPa for 4-5 minutes.

[0016] Preferably, in step (1), the ultrafine grinding is to 400-500 mesh.

[0017] Preferably, in step (2), the mass ratio of the fine powder to water is 1:2-3.

[0018] Preferably, in step (2), the process conditions for ultrasonic dispersion are: frequency 25-30 kHz, power 300-400 W, and time 10-20 minutes.

[0019] Preferably, in step (2), the specific method for freeze-thaw treatment is: first treating at -10 to -8 °C for 3-4 hours, then treating at -30 to -28 °C for 6-7 hours, and then treating at 20-25 °C for 3-4 hours.

[0020] Preferably, in step (2), the process conditions for pulsed electric field treatment are: electric field strength 20-25 kV, pulse width 15-20 μs, temperature 33-35 °C, and time 3-5 minutes.

[0021] Preferably, in step (3), the supercritical extraction conditions are as follows: pressure 15 - 20 MPa, temperature 40 - 50 °C, carbon dioxide flow rate 35 - 45 L / min, and time 3 - 4 hours.

[0022] Advantages of the present invention:

[0023] The present invention provides a method for extracting isothiocyanates from ripened mustard seed cake. First, the ripened mustard seed cake obtained by cold pressing and ripening the mustard seeds is subjected to high-pressure treatment and ultrafine grinding to obtain fine powder; then the fine powder is ultrasonically dispersed in water, followed by freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture; finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates. The method of the present invention is green and environmentally friendly, with a high yield and good quality of isothiocyanates.

[0024] The raw material ripened mustard seed cake used in the present invention is obtained by cold pressing. The cold pressing temperature is low and does not affect the activity of myrosinase, ensuring the smooth hydrolysis of 2-propenyl glucosinolate to generate isothiocyanates.

[0025] The present invention subjects the ripened mustard seed cake to high-pressure treatment and ultrafine grinding, reducing the difficulty of subsequent treatment and facilitating the release of components in the ripened mustard seed cake.

[0026] The fine powder is dispersed in water and subjected to freeze-thaw treatment, which can promote the full release of 2-propenyl glucosinolate and myrosinase in the ripened mustard seed cake. The subsequent pulsed electric field treatment promotes the enzymatic reaction to improve the yield of isothiocyanates. Combined with the subsequent supercritical extraction, the purity of the obtained isothiocyanates is improved. Specific embodiments

[0027] The present invention will be further elaborated below in conjunction with embodiments. It should be noted that the following description is only for explaining the present invention and does not limit its content.

[0028] Example 1:

[0029] A method for extracting isothiocyanates from ripened mustard seed cake, the specific steps are as follows:

[0030] (1) First, the ripened mustard seed cake obtained by cold pressing and ripening the mustard seeds is subjected to high-pressure treatment and ultrafine grinding to obtain fine powder;

[0031] (2) Then the fine powder is ultrasonically dispersed in water, followed by freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture;

[0032] (3) Finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates.

[0033] In step (1), the dry mustard seeds are cold pressed using a screw oil press, and the cold pressing temperature is 40 °C.

[0034] The ripening method is as follows: irradiate with 300 W microwave for 2 minutes, stop irradiation for 1 minute, continue irradiating with 300 W microwave for 2 minutes, and let it stand at 10 °C for 2 days.

[0035] The high-pressure treatment conditions are: keep the pressure at 300 MPa for 4 minutes.

[0036] Ultra-finely grind to 400 mesh.

[0037] In step (2), the mass ratio of the fine powder to water is 1:2.

[0038] The process conditions for ultrasonic dispersion are: frequency 25 kHz, power 300 W, and time 10 minutes.

[0039] The specific method for freeze-thaw treatment is: first treat at -10 °C for 3 hours, then treat at -30 °C for 6 hours, and then treat at 20 °C for 3 hours.

[0040] The process conditions for pulsed electric field treatment are: electric field strength 20 kV, pulse width 15 μs, temperature 33 °C, and time 3 minutes.

[0041] In step (3), the supercritical extraction conditions are: pressure 15 MPa, temperature 40 °C, carbon dioxide flow rate 35 L / min, and time 3 hours.

[0042] Example 2:

[0043] A method for extracting isothiocyanate from ripened mustard seed cake, the specific steps are as follows:

[0044] (1) First, perform high-pressure treatment and ultra-fine grinding on the ripened mustard seed cake obtained by cold pressing mustard seeds to obtain fine powder;

[0045] (2) Then disperse the fine powder ultrasonically in water, perform freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture;

[0046] (3) Finally, perform supercritical extraction on the pretreated mixture to obtain isothiocyanate.

[0047] In step (1), dry mustard seeds are cold pressed using a spiral oil press, and the cold pressing temperature is 43 °C.

[0048] The ripening method is as follows: irradiate with 350 W microwave for 3 minutes, stop irradiation for 1 minute, continue irradiating with 350 W microwave for 3 minutes, and let it stand at 15 °C for 3 days.

[0049] The high-pressure treatment conditions are: keep the pressure at 400 MPa for 5 minutes.

[0050] Ultra-finely grind to 500 mesh.

[0051] In step (2), the mass ratio of the fine powder to water is 1:3.

[0052] The process conditions for ultrasonic dispersion are: frequency 30 kHz, power 400 W, and time 20 minutes.

[0053] The specific method for freeze-thaw treatment is: first treat at -8°C for 4 hours, then treat at -28°C for 7 hours, and then treat at 25°C for 4 hours.

[0054] The process conditions for pulsed electric field treatment are: electric field strength 25 kV, pulse width 20 μs, temperature 35°C, and time 5 minutes.

[0055] In step (3), the supercritical extraction conditions are: pressure 20 MPa, temperature 50°C, carbon dioxide flow rate 45 L / min, and time 4 hours.

[0056] Example 3:

[0057] A method for extracting isothiocyanates from ripened mustard seed cake, the specific steps are as follows:

[0058] (1) First, subject the ripened mustard seed cake obtained by cold pressing mustard seeds to high-pressure treatment and ultrafine grinding to obtain fine powder;

[0059] (2) Then disperse the fine powder ultrasonically in water, perform freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture;

[0060] (3) Finally, perform supercritical extraction on the pretreated mixture to obtain isothiocyanates.

[0061] In step (1), dry mustard seeds are cold-pressed using a screw oil press, and the cold pressing temperature is 40°C.

[0062] The ripening method is: microwave irradiation at 350 W for 2 minutes, stop irradiation for 1 minute, continue microwave irradiation at 350 W for 2 minutes, and let stand at 15°C for 2 days.

[0063] The high-pressure treatment conditions are: holding pressure at 400 MPa for 4 minutes.

[0064] Ultrafine grind to 500 mesh.

[0065] In step (2), the mass ratio of the fine powder to water is 1:2.

[0066] The process conditions for ultrasonic dispersion are: frequency 30 kHz, power 300 W, and time 20 minutes.

[0067] The specific method for freeze-thaw treatment is: first treat at -10°C for 4 hours, then treat at -30°C for 7 hours, and then treat at 20°C for 4 hours.

[0068] The process conditions for pulsed electric field treatment are: electric field strength 20 kV, pulse width 20 μs, temperature 33 °C, and time 5 minutes.

[0069] In step (3), the supercritical extraction conditions are: pressure 15 MPa, temperature 50 °C, carbon dioxide flow rate 35 L / min, and time 4 hours.

[0070] Example 4:

[0071] A method for extracting isothiocyanates from ripened mustard seed cake, the specific steps are as follows:

[0072] (1) First, the ripened mustard seed cake obtained by cold pressing and ripening mustard seeds is subjected to high-pressure treatment and ultrafine grinding to obtain fine powder;

[0073] (2) Then, the fine powder is ultrasonically dispersed in water, subjected to freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture;

[0074] (3) Finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates.

[0075] In step (1), the dry mustard seeds are cold pressed using a screw oil press, and the cold pressing temperature is 42 °C.

[0076] The ripening method is: microwave irradiation at 300 W for 3 minutes, stopping irradiation for 1 minute, continuing microwave irradiation at 350 W for 2 minutes, and standing at 12 °C for 3 days.

[0077] The high-pressure treatment conditions are: holding pressure at 350 MPa for 5 minutes.

[0078] Ultrafine grinding to 500 mesh.

[0079] In step (2), the mass ratio of the fine powder to water is 1:2.5.

[0080] The process conditions for ultrasonic dispersion are: frequency 30 kHz, power 300 W, and time 15 minutes.

[0081] The specific method for freeze-thaw treatment is: first treat at -9 °C for 3 hours, then treat at -30 °C for 6.5 hours, and then treat at 22 °C for 3.5 hours.

[0082] The process conditions for pulsed electric field treatment are: electric field strength 22 kV, pulse width 18 μs, temperature 34 °C, and time 4 minutes.

[0083] In step (3), the supercritical extraction conditions are: pressure 18 MPa, temperature 45 °C, carbon dioxide flow rate 40 L / min, and time 3.5 hours.

[0084] Comparative Example 1

[0085] A method for extracting isothiocyanates from ripened mustard seed cake, the specific steps are as follows:

[0086] (1) First, the ripened mustard seed cake obtained by cold pressing and ripening mustard seeds is subjected to ultrafine grinding to obtain fine powder;

[0087] (2) Then, the fine powder is ultrasonically dispersed in water, followed by freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture;

[0088] (3) Finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates.

[0089] In step (1), the dry mustard seeds are cold pressed using a screw oil press at a cold pressing temperature of 40°C.

[0090] The ripening method is: microwave irradiation at 300W for 2 minutes, stopping irradiation for 1 minute, continuing microwave irradiation at 300W for 2 minutes, and standing at 10°C for 2 days.

[0091] Ultrafine grind to 400 mesh.

[0092] In step (2), the mass ratio of the fine powder to water is 1:2.

[0093] The process conditions for ultrasonic dispersion are: frequency 25kHz, power 300W, time 10 minutes.

[0094] The specific method for freeze-thaw treatment is: first treat at -10°C for 3 hours, then treat at -30°C for 6 hours, and then treat at 20°C for 3 hours.

[0095] The process conditions for pulsed electric field treatment are: electric field strength 20kV, pulse width 15μs, temperature 33°C, time 3 minutes.

[0096] In step (3), the supercritical extraction conditions are: pressure 15MPa, temperature 40°C, carbon dioxide flow rate 35L / min, time 3 hours.

[0097] Comparative Example 2

[0098] A method for extracting isothiocyanates from ripened mustard seed cake, the specific steps are as follows:

[0099] (1) First, the ripened mustard seed cake obtained by cold pressing and ripening mustard seeds is subjected to high-pressure treatment and ultrafine grinding to obtain fine powder;

[0100] (2) Then, the fine powder is ultrasonically dispersed in water, followed by pulsed electric field treatment to obtain a pretreated mixture;

[0101] (3) Finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates.

[0102] In step (1), the dried mustard seeds are cold-pressed using a screw oil press, and the cold-pressing temperature is 40°C.

[0103] The ripening method is: microwave irradiation at 300W for 2 minutes, stopping irradiation for 1 minute, continuing microwave irradiation at 300W for 2 minutes, and standing at 10°C for 2 days.

[0104] The high-pressure treatment conditions are: holding pressure at 300 MPa for 4 minutes.

[0105] Ultra-finely milled to 400 mesh.

[0106] In step (2), the mass ratio of the fine powder to water is 1:2.

[0107] The process conditions for ultrasonic dispersion are: frequency 25 kHz, power 300 W, and time 10 minutes.

[0108] The process conditions for pulsed electric field treatment are: electric field strength 20 kV, pulse width 15 μs, temperature 33°C, and time 3 minutes.

[0109] In step (3), the supercritical extraction conditions are: pressure 15 MPa, temperature 40°C, carbon dioxide flow rate 35 L / min, and time 3 hours.

[0110] Comparative Example 3

[0111] A method for extracting isothiocyanates from ripened mustard seed cakes, the specific steps are as follows:

[0112] (1) First, the ripened mustard seed cakes obtained by cold-pressing and ripening the dried mustard seeds are subjected to high-pressure treatment and ultra-finely milled to obtain fine powder;

[0113] (2) Then, the fine powder is ultrasonically dispersed in water and subjected to freeze-thaw treatment to obtain a pretreated mixture;

[0114] (3) Finally, the pretreated mixture is subjected to supercritical extraction to obtain isothiocyanates.

[0115] In step (1), the dried mustard seeds are cold-pressed using a screw oil press, and the cold-pressing temperature is 40°C.

[0116] The ripening method is: microwave irradiation at 300W for 2 minutes, stopping irradiation for 1 minute, continuing microwave irradiation at 300W for 2 minutes, and standing at 10°C for 2 days.

[0117] The high-pressure treatment conditions are: holding pressure at 300 MPa for 4 minutes.

[0118] Ultra-finely milled to 400 mesh.

[0119] In step (2), the mass ratio of the fine powder to water is 1:2.

[0120] The process conditions for ultrasonic dispersion are as follows: frequency 25 kHz, power 300 W, and time 10 minutes.

[0121] The specific method for freeze-thaw treatment is as follows: first treat at -10°C for 3 hours, then treat at -30°C for 6 hours, and then treat at 20°C for 3 hours.

[0122] In step (3), the supercritical extraction conditions are: pressure 15 MPa, temperature 40°C, carbon dioxide flow rate 35 L / min, and time 3 hours.

[0123] Calculate the yields of the product isothiocyanate in Examples 1-4 and Comparative Examples 1-3 respectively, and test the purity of the obtained products.

[0124] Among them, the yield (%) = mass of product isothiocyanate / mass of mustard seeds × 100.

[0125] The purity test method is as follows:

[0126] Accurately weigh about 3 g of the product, add it to 30 mL of water, stir and disperse evenly. Accurately add 25.0 mL of hexahydropyridine standard solution, stopper, shake for 3 minutes, let stand for 40 minutes, filter to obtain the filtrate. Using bromocresol green-methyl red as the indicator (obtained by dissolving 0.1000 g of bromocresol green and 0.0200 g of methyl red in 100 mL of anhydrous ethanol), titrate the filtrate with hydrochloric acid standard solution until it turns wine red as the end point. Calculate the isothiocyanate content (calculated as allyl isothiocyanate) according to the formula: isothiocyanate content (%) = 9.915×(c1V1 - c2V2) / W, where c1 is the concentration of hexahydropyridine standard solution (mol / L), V1 is the added volume of hexahydropyridine standard solution (mL), c2 is the concentration of hydrochloric acid standard solution (mol / L), V2 is the volume of hydrochloric acid standard solution consumed during titration (mL), and W is the sample amount of the product (g).

[0127] Preparation of hydrochloric acid standard solution: Measure 4.5 mL of concentrated hydrochloric acid (mass concentration 37%), dissolve it in 1000 mL of distilled water to obtain hydrochloric acid standard solution. Accurately weigh 0.1000 g of anhydrous sodium carbonate that has been dried to constant weight at 280°C, dissolve it with 50 mL of distilled water, add 5 drops of bromocresol green-methyl red indicator, titrate with hydrochloric acid solution until it turns from green to wine red, boil for 2 minutes, cool and continue titrating until it turns wine red, record the volume of hydrochloric acid standard solution consumed, and calculate the accurate concentration of the prepared hydrochloric acid standard solution.

[0128] Preparation of hexahydropyridine standard solution: Measure 2.5 mL of freshly distilled hexahydropyridine, add it to a 500 mL volumetric flask, add anhydrous ethanol to volume, shake well, and use bromocresol green-methyl red as the indicator, titrate with hydrochloric acid standard solution to determine the accurate concentration.

[0129] The results are shown in Table 1.

[0130] Table 1. Yield and Purity

[0131] Yield (%) Purity (%) Example 1 0.63 99.5 Example 2 0.64 99.5 Example 3 0.64 99.6 Example 4 0.65 99.8 Comparative Example 1 0.42 96.3 Comparative Example 2 0.45 95.7 Comparative Example 3 0.33 96.6

[0132] As can be seen from Table 1, the yields of Examples 1 to 4 are high and the purities are good.

[0133] In Comparative Example 1, the high-pressure treatment was omitted. In Comparative Example 2, the freeze-thaw treatment was omitted. In Comparative Example 3, the pulsed electric field treatment was omitted. The yields and purities both decreased significantly, indicating that the synergistic effects of high-pressure treatment, freeze-thaw treatment, pulsed electric field treatment, etc. on the ripened mustard seed cake can improve the yields and purities of isothiocyanates.

[0134] Although the specific implementation manners of the present invention are described above, they do not limit the protection scope of the present invention. Based on the technical solutions of the present invention, various modifications or deformations that can be made by those skilled in the art without creative efforts are still within the protection scope of the present invention.

Claims

1. A method for extracting isothiocyanates from ripened mustard seed cake, characterized in that, The specific steps are as follows: (1) First, subject the cooked mustard seed cake obtained by cold pressing and ripening mustard seeds to high-pressure treatment and ultrafine grinding to obtain fine powder; (2) Then, ultrasonically disperse the fine powder in water, perform freeze-thaw treatment and pulsed electric field treatment to obtain a pretreated mixture; (3) Finally, perform supercritical extraction on the pretreated mixture to obtain isothiocyanate.

2. The method according to claim 1, characterized in that, In step (1), a screw oil press is used for cold pressing, and the cold pressing temperature is 40 - 43°C.

3. The method according to claim 1, wherein In step (1), the ripening method is: irradiate with 300 - 350W microwave for 2 - 3 minutes, stop irradiation for 1 minute, continue irradiating with 300 - 350W microwave for 2 - 3 minutes, and let it stand at 10 - 15°C for 2 - 3 days.

4. The method according to claim 1, wherein In step (1), the high-pressure treatment conditions are: keep the pressure at 300 - 400MPa for 4 - 5 minutes.

5. The method according to claim 1, wherein In step (1), ultrafine grind to 400 - 500 mesh.

6. The method according to claim 1, wherein In step (2), the mass ratio of the fine powder to water is 1:2 - 3.

7. The method according to claim 1, characterized in that, In step (2), the process conditions for ultrasonic dispersion are: frequency 25 - 30kHz, power 300 - 400W, time 10 - 20 minutes.

8. The method according to claim 1, wherein In step (2), the specific method for freeze-thaw treatment is: first treat at -10 - -8°C for 3 - 4 hours, then treat at -30 - -28°C for 6 - 7 hours, and then treat at 20 - 25°C for 3 - 4 hours.

9. The method according to claim 1, wherein In step (2), the process conditions for pulsed electric field treatment are: electric field strength 20 - 25kV, pulse width 15 - 20μs, temperature 33 - 35°C, time 3 - 5 minutes.

10. The method according to claim 1, characterized in that, In step (3), the supercritical extraction conditions are: pressure 15 - 20MPa, temperature 40 - 50°C, carbon dioxide flow rate 35 - 45L / min, time 3 - 4 hours.