Method for extracting and purifying capsaicin

Abstract

The application belongs to an extraction and purification process, and discloses a capsaicin extraction and purification method. The method comprises the following steps: mixing crushed chili powder with a composite enzyme, dispersing the mixture in a plasma active eutectic solvent, and simultaneously performing enzymolysis, extraction and purification; after extraction, centrifuging and collecting the supernatant, and freeze-drying after concentration. The application performs targeted extraction and purification by preparing and applying a plasma active eutectic solvent with high selectivity for capsaicin, and the purity of the extract is high. The yield and purity of capsaicin extracted by the method are 222.04% and 36.36% higher than those of 70% (v / v) ethanol extraction, respectively, and the use of a large amount of organic reagents is avoided, which meets the requirements of green chemistry and is conducive to the industrialized production and application of capsaicin.

Description

Technical Field

[0001] This invention relates to an extraction and purification process, specifically to a method for extracting and purifying capsaicin. Background Technology

[0002] Capsaicin is insoluble in water and is mostly extracted using organic solvents. Traditional extraction methods require large amounts of toxic and harmful organic solvents (alcohols, ethers, and chloroform), involving repeated extractions, resulting in low extraction efficiency and cumbersome operations, which violates the principles of "green chemistry." Therefore, developing green, environmentally friendly, and cost-effective extraction methods is of great significance for expanding the industrial application of capsaicin.

[0003] In view of this, the present invention is proposed. Summary of the Invention

[0004] The purpose of this invention is to provide a method for extracting and purifying capsaicin, which uses a compound enzyme to break down cell walls and simultaneously adds a plasma-active low-euclidean solvent for targeted extraction and purification, thereby improving the extraction efficiency and purity of capsaicin.

[0005] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0006] The method for extracting and purifying capsaicin includes the following steps: mixing pulverized chili powder with a complex enzyme, dispersing the mixture in a plasma-active eutectic solvent, and simultaneously performing enzymatic hydrolysis and extraction purification; after extraction, centrifuging and collecting the supernatant; concentrating the supernatant and then freeze-drying it;

[0007] The plasma-active eutectic solvent is prepared from propofol and diethylene glycol butyl ether in a molar ratio of 1:(3-4).

[0008] Preferably, the formulation of the compound enzyme is as follows: for every 1 g of chili powder, add 3500-4000 U of pepsin and 5500-6000 U of trypsin.

[0009] Preferably, the immersion extraction and purification enrichment are carried out at 40-50 °C, with simultaneous extraction and purification for 30-40 min.

[0010] Preferably, the chili powder is 70-80 mesh.

[0011] Preferably, the centrifugation is performed at a speed of 6000-8000 rpm for 10-20 minutes.

[0012] Preferably, the concentration is a rotary evaporation concentration, performed at 30-40 °C and 100-130 rpm for 40-60 min.

[0013] Preferably, the preparation process of the plasma-active eutectic solvent is as follows: propofol and diethylene glycol butyl ether are mixed and then added to 15%-20% by volume of deionized water, and the mixture is subjected to low-temperature plasma treatment for 6-10 min under the conditions of voltage 5-9 kV and frequency 30-50 kHz.

[0014] The present invention also provides capsaicin obtained by any of the extraction and purification methods described above.

[0015] Compared with the prior art, the present invention has the following beneficial effects:

[0016] This invention utilizes a plasma-active eutectic solvent composed of a complex enzyme, propofol, and diethylene glycol butyl ether to simultaneously extract and purify capsaicin from chili peppers, achieving high extraction efficiency and purity. The complex enzyme, composed of pepsin and trypsin, can decompose and disrupt the cell wall membrane of chili peppers, promoting capsaicin release. The plasma-active eutectic solvent prepared from propofol and diethylene glycol butyl ether is added during the extraction process. Plasma treatment increases the content of H₂O₂ and OH· in the solvent, which is beneficial for further disrupting the plant cell wall membrane. Therefore, compared to conventional eutectic solvents, the plasma-active eutectic solvent can cause more severe damage to the cell structure of the extracted material, thereby promoting mass transfer during the extraction process. Consequently, the plasma-active eutectic solvent exhibits superior extraction and purification effects compared to conventional eutectic solvents.

[0017] This invention provides an efficient and simple method for extracting high-purity capsaicin, which helps to expand the high-value application scenarios of chili peppers. Detailed Implementation

[0018] To make the objectives, technical solutions, and advantages of this invention clearer, the invention will be further described in detail below with reference to specific embodiments. It should be understood that these descriptions are merely exemplary and not intended to limit the scope of the invention. Furthermore, descriptions of well-known structures and techniques are omitted in the following description to avoid unnecessarily obscuring the concept of the invention. Unless otherwise specified, specific conditions in the embodiments are performed under conventional conditions or conditions recommended by the manufacturer. Reagents or instruments used, unless otherwise specified, are all commercially available conventional products.

[0019] It should be noted that the chili powder in this invention is all dried chili powder, that is, chili powder obtained by crushing dried chilies and sieving them.

[0020] It should also be noted that the centrifugation method, concentration method, and freeze-drying method used in this invention are all commonly used methods in the field and do not require special limitation. The specific methods given in the embodiments are examples, and those skilled in the art can select appropriate methods according to the actual situation.

[0021] The preparation process of the plasma-active eutectic solvent used in the embodiments of the present invention is as follows: propofol and diethylene glycol butyl ether are mixed and then added to 15%-20% deionized water by volume, and the mixture is subjected to low-temperature plasma treatment for 6-10 min under the conditions of voltage 5-9 kV and frequency 30-50 kHz.

[0022] To make the technical solution of the present invention clearer, the following examples illustrate the extraction and purification method and effect of capsaicin.

[0023] The measurement or reagent preparation methods used in the embodiments of the present invention are as follows:

[0024] (1) Determination of capsaicin content:

[0025] The capsaicin content in the sample was determined by high-performance liquid chromatography (HPLC). 1 mg of sample was accurately weighed, dissolved in 1 mL of chromatographic methanol, and diluted 20-fold. The diluted sample was filtered through a 0.22 μm organic filter membrane and injected into the HPLC sample vial for analysis. The HPLC detection conditions were as follows: a mobile phase of chromatographic methanol and ultrapure water at a volume ratio of 78%:22%; and 20 μL of sample was injected into a C14 sample vial. 18 The chromatographic column was used, and the mobile phase flow rate was 1 mL / min. Detection was performed at a UV wavelength of 280 nm for 15 min. The peak area at 280 nm was recorded, and the capsaicin content in the corresponding sample was calculated based on the standard curve.

[0026] Accurately weigh 0.2, 0.4, 0.6, 0.8, and 1.0 mg of capsaicin standards (purity ≥98%), dissolve them in 1 mL of chromatographic methanol, filter through a 0.22 μm organic filter membrane, and inject into a liquid chromatography sample vial for analysis. Detect and record the corresponding peak areas using high-performance liquid chromatography (HPLC) according to the capsaicin content determination method in (1). Plot a standard curve with the concentration of capsaicin standards on the x-axis and the peak area of ​​the HPLC on the y-axis.

[0027] The equation of the obtained standard curve is: y = 5718.2x + 37.046, R0 2 = 0.9996, which meets the requirements of the standard curve and is used for subsequent calculation of capsaicin content.

[0028] (2) Determination of capsaicin purity

[0029] The capsaicin content in the extract was determined according to the method described in (1), and the purity of the capsaicin extract was calculated using the formula below:

[0030] Purity (%) = (mass of capsaicin / total mass of extract) × 100%.

[0031] Example 1

[0032] The extraction and purification method of capsaicin includes the following steps: (1) passing chili powder through a 70-mesh sieve; (2) preparing a plasma-active eutectic solvent by mixing propofol and diethylene glycol butyl ether at a molar ratio of 1:3; (3) adding 3500 U of pepsin and 5500 U of trypsin to each 1 g of chili powder, mixing, and then adding the plasma-active eutectic solvent in a "one-pot" manner at a material-to-liquid ratio of 1:20 g / mL, extracting and purifying at 40 °C for 30 min; (4) centrifuging at 6000 rpm for 20 min and collecting the supernatant; (5) rotary evaporating and concentrating at 30 °C and 100 rpm for 60 min to obtain a concentrated extract; (6) freeze-drying the concentrated extract to obtain capsaicin.

[0033] The capsaicin yield in this embodiment was determined to be 12.54 mg / g, with a purity of 65.87%.

[0034] Example 2

[0035] The extraction and purification method of capsaicin includes the following steps: (1) passing chili powder through a 70-mesh sieve; (2) preparing a plasma-active eutectic solvent by mixing propofol and diethylene glycol butyl ether at a molar ratio of 1:3.5; (3) adding 3800 U of pepsin and 5700 U of trypsin to each 1 g of chili powder, mixing, and then adding the plasma-active eutectic solvent in a "one-pot" manner at a material-to-liquid ratio of 1:30 g / mL, extracting and purifying at 45 °C for 40 min; (4) centrifuging at 7000 rpm for 15 min and collecting the supernatant; (5) rotary evaporating and concentrating at 40 °C and 110 rpm for 50 min to obtain a concentrated extract; (6) freeze-drying the concentrated extract to obtain capsaicin.

[0036] The capsaicin yield in this embodiment was determined to be 14.87 mg / g, with a purity of 67.43%.

[0037] Example 3

[0038] The extraction and purification method of capsaicin includes the following steps: (1) passing chili powder through an 80-mesh sieve to obtain dryness; (2) preparing a plasma-active eutectic solvent by mixing propofol and diethylene glycol butyl ether at a molar ratio of 1:4; (3) adding 3900 U of pepsin and 6000 U of trypsin per 1 g of chili powder, mixing, and then adding the plasma-active eutectic solvent in a "one-pot" manner at a material-to-liquid ratio of 1:40 g / mL, extracting and purifying at 50 °C for 40 min; (4) centrifuging at 8000 rpm for 10 min and collecting the supernatant; (5) rotary evaporating and concentrating at 40 °C and 130 rpm for 40 min to obtain a concentrated extract; (6) freeze-drying the concentrated extract to obtain capsaicin.

[0039] The capsaicin yield in this example was determined to be 16.41 mg / g, with a purity of 71.50%.

[0040] Experimental Example 1: Effects of Conventional Eutectic Solvents and Propofol-Diethylene Glycol Butyl Ether Plasma-Activated Eutectic Solvents on Capsaicin Yield and Purity

[0041] Using choline chloride and menthol as hydrogen bond acceptors, several conventional eutectic solvents commonly used for the extraction of active substances were prepared for the extraction of capsaicin, and their extraction effects on capsaicin were compared with those of propofol-diethylene glycol butyl ether of the present invention.

[0042] The preparation method of conventional eutectic solvents is as follows: accurately weigh the hydrogen bond acceptor and hydrogen bond donor (corresponding molar ratio) into a beaker, heat at 80 °C while magnetically stirring at 100 rpm for 40 min, and finally add 20% (v / v) deionized water. Conventional eutectic solvents used for comparison include: choline chloride-malic acid (molar ratio 1:1), choline chloride-oxalic acid (molar ratio 1:1), choline chloride-citric acid (molar ratio 1:1), choline chloride-lactic acid (molar ratio 1:1), choline chloride-ethylene glycol (molar ratio 1:2), menthol-1,2-propanediol (molar ratio 1:2), menthol-glycerol (molar ratio 1:2), menthol-urea (molar ratio 1:2), menthol-glucose (molar ratio 1:1), menthol-thymol (molar ratio 1:2), choline chloride-diethylene glycol butyl ether (molar ratio 1:3), and menthol-diethylene glycol butyl ether (molar ratio 1:3). It should be noted that the molar ratio of hydrogen bond acceptors to hydrogen bond donors in the above conventional eutectic solvents are all experimentally selected to achieve the best molar ratio for capsaicin extraction.

[0043] The extraction and purification method of capsaicin is as follows: chili peppers are passed through an 80-mesh sieve to obtain chili powder; 3600 U of pepsin and 5600 U of trypsin are added to 1 g of dried chili powder; after mixing, the appropriate extraction solvent is added in a "one-pot" manner at a material-to-liquid ratio of 1:40 g / mL, and the mixture is extracted and purified at 45 ℃ for 35 min; the mixture is centrifuged at 8000 rpm for 10 min, and the supernatant is collected; the mixture is concentrated by rotary evaporation at 40 ℃ and 130 rpm for 40 min to obtain a concentrated extract; the concentrated extract is freeze-dried to obtain capsaicin extract.

[0044] Capsaicin was extracted using the above-mentioned conventional eutectic solvents and propofol-diethylene glycol butyl ether (molar ratio 1:3). The final capsaicin yield and purity are shown in Table 1.

[0045] Table 1. Yield and purity of capsaicin extracted with different eutectic solvents

[0046] .

[0047] Experimental Example 2: Effect of Different Plasma-Active Eutectic Solvents on Capsaicin Yield and Purity

[0048] Using propofol as a hydrogen bond acceptor and a jet-type atmospheric low-temperature plasma treatment system, several plasma-active eutectic solvents were prepared for the extraction of capsaicin, and their extraction effects were compared with those of the eutectic solvent using diethylene glycol butyl ether as a hydrogen bond donor in this invention.

[0049] The preparation method of plasma-active eutectic solvent is as follows: Accurately weigh propofol-hydrogen bond donors (corresponding molar ratios) into a beaker, add 20% (v / v) of deionized water, and treat using a jet-type atmospheric low-temperature plasma treatment system at 9 kV and 50 kHz for 10 min to obtain the plasma-active eutectic solvent. The plasma-active eutectic solvents used for comparison include: propofol-malic acid (molar ratio 1:1), propofol-oxalic acid (molar ratio 1:1), propofol-citric acid (molar ratio 1:1), propofol-lactic acid (molar ratio 1:1), propofol-ethylene glycol (molar ratio 1:2), propofol-1,2-propanediol (molar ratio 1:2), propofol-glycerol (molar ratio 1:2), propofol-urea (molar ratio 1:2), propofol-glucose (molar ratio 1:1), propofol-thymol (molar ratio 1:2), and propofol-diethylene glycol butyl ether (molar ratio 1:3). It should be noted that, except for the propofol-diethylene glycol butyl ether solvent, the molar ratios of propofol and hydrogen bond donors in the above plasma-active eutectic solvents were all experimentally selected to achieve the best molar ratio for capsaicin extraction.

[0050] The capsaicin extraction and purification method described in Example 1 was used to extract capsaicin using the solvents described above. The final capsaicin yield and purity are shown in Table 2.

[0051] Simultaneously, a control group was set up using 70% (v / v) ethanol extraction. The specific extraction process was as follows: dried chili peppers were processed in a high-speed grinder for 4 min, and then passed through an 80-mesh sieve to obtain dried chili powder. 1 g of dried chili powder was weighed and 70% (v / v) ethanol was added at a material-to-liquid ratio of 1:40 g / mL. Extraction and purification were carried out at 50 ℃ for 40 min. The mixture was centrifuged at 8000 rpm for 10 min, and the supernatant was collected. The extract was concentrated by rotary evaporation at 40 ℃ and 130 rpm for 40 min to obtain a concentrated extract. The concentrated extract was freeze-dried to obtain capsaicin extract.

[0052] Table 2. Yields and purity of capsaicin extracted with different plasma-active eutectic solvents and 70% (v / v) ethanol.

[0053] .

[0054] Experimental Example 3: Effect of different molar ratios of propofol-diethylene glycol butyl ether plasma-active eutectic solvent on capsaicin yield and purity

[0055] Different molar ratios of hydrogen bond donors and acceptors in the formation of eutectic solvents resulted in significant differences in the extraction efficiency of active substances. To investigate the plasma-active eutectic solvent based on propofol-diethylene glycol butyl ether, the extraction and purification method for capsaicin in Example 1 was used at molar ratios of 1:1, 1:2, 1:3, 1:4, and 1:5. The final yield and purity of capsaicin are shown in Table 3.

[0056] Table 3. Yield and purity of capsaicin extracted by propofol-diethylene glycol butyl ether plasma-active eutectic solvents with different molar ratios.

[0057] .

[0058] In summary, the one-pot extraction method using a composite enzyme synergistic with a plasma-active eutectic solvent yields high capsaicin extraction efficiency and purity. Through screening different types of plasma-active eutectic solvents, a solvent synthesized from propofol and diethylene glycol butyl ether and treated with a jet-type atmospheric low-temperature plasma system was identified as having excellent extraction efficiency and purity for capsaicin. Composite enzyme treatment facilitates the release of capsaicin from cells and from capsaicin bound to plant tissues, thus increasing capsaicin yield. The one-pot extraction method simplifies the extraction process, resulting in capsaicin yield and purity that are 222.04% and 36.36% higher than those extracted with 70% (v / v) ethanol, respectively.

[0059] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A method for extracting and purifying capsaicin, characterized in that, Includes the following steps: The pulverized chili powder was mixed with the compound enzyme and dispersed in a plasma-active eutectic solvent for enzymatic hydrolysis, extraction and purification. After extraction, the mixture was centrifuged and the supernatant was collected. The supernatant was then concentrated and freeze-dried. The plasma-active eutectic solvent is prepared from propofol and diethylene glycol butyl ether in a molar ratio of 1:(3-4). The formula for the compound enzyme is as follows: for every 1 g of chili powder, add 3500-4000 U of pepsin and 5500-6000 U of trypsin. The preparation process of the plasma-active eutectic solvent is as follows: propofol and diethylene glycol butyl ether are mixed and then added to 15%-20% deionized water by volume. The mixture is then subjected to low-temperature plasma treatment for 6-10 min under the conditions of voltage 5-9 kV and frequency 30-50 kHz.

2. The method for extracting and purifying capsaicin according to claim 1, characterized in that, The ratio of chili powder to plasma-active low-efficiency eutectic solvent is 1:20-1:40 g / mL.

3. The method for extracting and purifying capsaicin according to claim 1, characterized in that, Extraction and purification were carried out at 40-50 °C for 30-40 min.

4. The method for extracting and purifying capsaicin according to claim 1, characterized in that, The chili powder is 70-80 mesh.

5. The method for extracting and purifying capsaicin according to claim 1, characterized in that, The centrifugation was performed at 6000-8000 rpm for 10-20 minutes.

6. The method for extracting and purifying capsaicin according to claim 1, characterized in that, The concentration is achieved by rotary evaporation, which is carried out at 30-40 ℃ and 100-130 rpm for 40-60 min.