Method for extracting p-coumaric acid from desert cistanche

Abstract

The invention discloses a method for extracting p-coumaric acid from desert cistanche deserticola, which comprises the following steps: freeze-drying desert cistanche deserticola, and grinding to obtain desert cistanche deserticola powder; adding ethanol into the desert cistanche deserticola powder, carrying out ultrasonic extraction, and carrying out solid-liquid separation to obtain desert cistanche deserticola p-coumaric acid crude extract and residues; adding ethanol into the residue, standing, carrying out heating reflux and ultrasonic-assisted extraction, centrifuging, and carrying out solid-liquid separation to obtain a cistanche deserticola p-coumaric acid residue extracting solution and a residue precipitate; carrying out acid-base hydrolysis-ethyl acetate extraction on the residue precipitate to obtain an extract liquor; combining the extracting solution with the desert cistanche p-coumaric acid crude extracting solution and the residue extracting solution to obtain a combined solution; and filtering and purifying the combined solution through a filter membrane to obtain p-coumaric acid. According to the method, the p-coumaric acid component in the desert cistanche can be effectively extracted, and desert cistanche biological resources are fully utilized.

Description

Technical Field

[0001] The invention relates to the technical field of food processing, and in particular to a method for extracting p-coumaric acid from Cistanche deserticola. Background Art

[0002] p-Coumaric acid (PCMA) is an important phenolic acid compound. p-Coumaric acid can be used as a starting material for a variety of commercially valuable chemicals, such as fragrances, drugs, bio-cosmetics, and health and nutrition products. Modern pharmacological studies have shown that p-coumaric acid has good antioxidant activity and can form resonance-stabilized phenolic free radicals. It has a strong scavenging effect on superoxide free radicals, light free radicals, and peroxynitrite. In addition, it also has antibacterial activity and has inhibitory effects on four different strains of Staphylococcus aureus and Shigella dysenteriae. Therefore, p-coumaric acid has gradually attracted the attention of the public.

[0003] Cistanche deserticola (Cistanche deserticola YCMa) is a perennial herbaceous parasitic plant of the Orobanchaceae family. It is a tonic and edible substance, known as the "desert ginseng". It is mainly distributed in Inner Mongolia, Ningxia, Gansu, Xinjiang and other regions. Cistanche deserticola is warm in nature, sweet and salty in taste, and enters the kidney and large intestine meridians. It has the effects of tonic liver and kidney, benefiting essence and blood, and moistening the intestines and relieving constipation. It can be used for kidney yang deficiency, impotence and infertility, weak muscles and bones, dry intestines and constipation, etc. As a plant for both medicinal and edible purposes, Cistanche deserticola is deeply loved by the Chinese people. If the production process such as soaking or fermentation is adopted, Cistanche deserticola can be used as the main raw material to prepare functional health wine. At present, the research on the extraction and activity of Cistanche deserticola mainly focuses on compounds such as phenylethanol glycosides, iridoid ether terpenoid glycosides, lignan glycosides, and sugars, while there are few related studies on p-coumaric acid. Summary of the invention

[0004] In order to solve the above problems, the present invention provides a method for extracting p-coumaric acid from Cistanche deserticola, which can effectively extract the p-coumaric acid component in Cistanche deserticola and make full use of the biological resources of Cistanche deserticola.

[0005] To achieve the above object, the present invention provides a method for extracting p-coumaric acid from Cistanche deserticola, which comprises the following steps:

[0006] (1) freeze-drying and grinding the deserticola to obtain deserticola powder;

[0007] (2) adding ethanol to the desert Cistanche powder, performing ultrasonic extraction, and performing solid-liquid separation to obtain a desert Cistanche p-coumaric acid crude extract and a residue;

[0008] (3) adding ethanol to the residue and allowing it to stand, then heating and refluxing and performing ultrasound-assisted extraction, centrifuging, and taking the supernatant to obtain a p-coumaric acid residue extract of Cistanche deserticola;

[0009] (4) subjecting the residual precipitate of step (3) to acid-base hydrolysis-ethyl acetate extraction to obtain an extract; combining the extract with the crude p-coumaric acid extract of Cistanche deserticola and the p-coumaric acid residue extract of Cistanche deserticola to obtain a combined solution;

[0010] (5) filtering and purifying the combined solution through a filter membrane to obtain p-coumaric acid.

[0011] According to a method for extracting p-coumaric acid from Cistanche deserticola according to an embodiment of the present invention, the method first freeze-drying and grinding Cistanche deserticola to convert it into an easy-to-operate powder, which lays a good foundation for subsequent extraction. Then, using ethanol as a solvent, combined with ultrasonic technology, the crude extract and residue of Cistanche deserticola p-coumaric acid are effectively extracted from the powder, and ultrasound-assisted extraction effectively improves the extraction efficiency. Subsequently, after adding ethanol to the residue obtained in the previous step and standing, the p-coumaric acid of Cistanche deserticola is further extracted by heating reflux and ultrasound-assisted secondary extraction methods, and the purity of the extract is improved. For the residue obtained in the previous step, acid-base hydrolysis-ethyl acetate extraction method is adopted, and the obtained extract is merged with the previous solution, and the p-coumaric acid component therein is fully extracted, and the maximum utilization of resources is achieved. Finally, the combined solution is filtered through a filter membrane to remove impurities, and then through a purification step, the p-coumaric acid product of Cistanche deserticola is finally obtained. The whole process is reasonably designed and rigorously operated, which not only ensures the extraction efficiency, but also guarantees the purity of p-coumaric acid in Cistanche deserticola, providing reliable technical support for the in-depth research and application of p-coumaric acid in Cistanche deserticola.

[0012] Optionally, in step (1), the freeze-drying temperature is -50°C to -20°C, the freeze-drying time is 24 to 36 hours, the vacuum degree is 25 to 35 Pa, and after the freeze-drying is completed, the temperature is naturally raised to room temperature.

[0013] Optionally, in step (2), the deserticola powder and ethanol are added in a mass volume ratio of 1:20 to 1:30 g / mL, and the concentration of the ethanol is 70%.

[0014] Optionally, in step (2), the temperature of ultrasonic extraction is 40-50°C, the ultrasonic extraction time is 30-60 min, and the ultrasonic power is 400W.

[0015] Optionally, in step (3), the heating reflux temperature is 75 to 85° C., and the heating reflux extraction time is 2 to 4 hours.

[0016] Optionally, in step (3), the ultrasonic-assisted extraction time is 60 to 90 min, the ultrasonic-assisted extraction temperature is 30 to 40° C., and the ultrasonic power is 500 to 800 W.

[0017] Optionally, in step (3), the centrifugation time is 20 to 30 min, and the centrifugation speed is 10,000 to 13,000 rpm / min.

[0018] Optionally, in step (4), the acid-base hydrolysis-ethyl acetate extraction is as follows: add NaOH solution to the residue precipitate, mix thoroughly at 45-55° C., hydrolyze for 4-5 hours in the dark, add HCl solution, adjust the pH value to 1-2, shake thoroughly, and extract with ethyl acetate for 1-3 hours, wherein the volume ratio of the solution to ethyl acetate is 20:1.

[0019] Optionally, in step (5), filtration is performed using a filter membrane with a pore size of 100 to 500 nm.

[0020] Optionally, in step (5), purification is performed using D101 macroporous adsorption resin, the macroporous resin column diameter-to-height ratio is 1:4-6, and includes water washing to remove impurities and elution with ethanol solution.

[0021] Furthermore, in step (5), the washing volume for water washing and impurity removal is 6-7 times the column volume BV, the washing flow rate is 2-3 times the column volume BV / h per hour, the ethanol volume fraction of the ethanol solution elution is 65%-75%, the ethanol solution elution volume is 4-6 times the column volume BV, and the ethanol solution elution flow rate is 5-7 times the column volume BV / h per hour.

[0022] Additional aspects and advantages of the present invention will be given in part in the following description and in part will be obvious from the following description, or will be learned through practice of the present invention. DETAILED DESCRIPTION

[0023] The technical solution of the present invention is described below through specific examples. It should be understood that the one or more method steps mentioned in the present invention do not exclude the existence of other method steps before and after the combination step or the insertion of other method steps between these explicitly mentioned steps; it should also be understood that these embodiments are only used to illustrate the present invention and are not used to limit the scope of the present invention. Moreover, unless otherwise specified, the numbering of each method step is only a convenient tool for identifying each method step, and is not intended to limit the order of arrangement of each method step or to limit the scope of the present invention. The change or adjustment of the relative relationship thereof shall also be regarded as the scope of the present invention without substantially changing the technical content.

[0024] In order to better understand the above technical scheme, the exemplary embodiments of the present invention are described in more detail below. Although exemplary embodiments of the present invention are shown, it should be understood that the present invention can be implemented in various forms and should not be limited by the embodiments set forth herein. On the contrary, these embodiments are provided in order to enable a more thorough understanding of the present invention and to enable the scope of the present invention to be fully communicated to those skilled in the art.

[0025] The test materials used in the present invention are all common commercial products and can be purchased in the market.

[0026] The present invention is described below with reference to specific embodiments. It should be noted that these embodiments are merely illustrative and do not limit the present invention in any way.

[0027] Example 1

[0028] A method for extracting p-coumaric acid from Cistanche deserticola comprises the following steps:

[0029] (1) Raw material pretreatment: 50 g of Cistanche deserticola was freeze-dried at -50°C for 24 h with a vacuum degree of 25 Pa. After freeze-drying, the mixture was naturally heated to room temperature. After being ground into powder, the passing rate under a 50-mesh sieve was 100%;

[0030] (2) Ethanol ultrasonic extraction: 70% ethanol was added to the powder obtained in step (1) at a mass volume ratio of 1:20 g / mL, and ultrasonic extraction was performed at 40° C. for 30 min with an ultrasonic power of 400 W. The crude p-coumaric acid extract of Cistanche deserticola and the residue were obtained by solid-liquid separation;

[0031] (3) Re-extraction of the residue: The residue of step (2) was immersed in a 60% ethanol solution at a solid-liquid ratio of 1:25 g / mL, and allowed to stand for 30 min. The residue was subjected to heating reflux and ultrasonic-assisted extraction, centrifuged, and solid-liquid separation to obtain a p-coumaric acid residue extract of Cistanche deserticola and a residue precipitate. The heating reflux extraction temperature was 75°C, the heating reflux extraction time was 2 h, the ultrasonic-assisted extraction time was 60 min, the ultrasonic-assisted extraction temperature was 30°C, the ultrasonic power was 500 W, the centrifugation time was 20 min, and the centrifugation speed was 10000 rpm / min;

[0032] (4) Acid-base hydrolysis and extraction of the residue: The residue precipitate obtained in step (3) is subjected to acid-base hydrolysis-ethyl acetate extraction method, 4 mol / L NaOH solution is added to the residue, the mixture is thoroughly mixed at 45° C., and the mixture is hydrolyzed for 4 h in the dark, 4 mol / L HCl solution is added, the pH value is adjusted to 1.0, and after thorough shaking, ethyl acetate is added for extraction for 1 h, wherein the volume ratio of the solution to ethyl acetate is 20:1; the obtained extract is combined with the crude p-coumaric acid extract of Cistanche deserticola obtained in step (2) and the residue extract obtained in step (3) to obtain a combined solution;

[0033] (5) Filtration and purification: The combined solution obtained in step (4) is filtered through a filter membrane with a pore size of 100 nm, and then purified. The macroporous resin used is D101 macroporous adsorption resin, and the macroporous resin column diameter-to-height ratio is 1:4; it includes a water washing and impurity removal step and an ethanol solution elution step. In the water washing and impurity removal step, the water washing volume is 6 times the column volume (BV), and the water washing flow rate is 2 times the column volume per hour (BV / h); in the ethanol solution elution step, the ethanol volume fraction is 65%, the ethanol solution elution volume is 4 times the column volume (BV), and the ethanol solution elution flow rate is 5 times the column volume per hour (BV / h), and the extraction rate of coumaric acid from Cistanche deserticola is 15.57%, and the purity of coumaric acid from Cistanche deserticola is 95.87%.

[0034] Example 2

[0035] A method for extracting p-coumaric acid from Cistanche deserticola comprises the following steps:

[0036] (1) Raw material pretreatment: 50 g of Cistanche deserticola was freeze-dried at -40°C for 30 h with a vacuum degree of 30 Pa. After freeze-drying, the mixture was naturally heated to room temperature. After being ground into powder, the passing rate under a 50-mesh sieve was 100%;

[0037] (2) Ethanol ultrasonic extraction: 70% ethanol was added to the powder obtained in step (1) at a mass volume ratio of 1:25 g / mL, and ultrasonic extraction was performed at 45° C. for 45 min with an ultrasonic power of 400 W. The crude p-coumaric acid extract of Cistanche deserticola and the residue were obtained by solid-liquid separation;

[0038] (3) Re-extraction of the residue: The residue of step (2) was immersed in a 60% ethanol solution at a solid-liquid ratio of 1:25 g / mL, left to stand for 30 min, and subjected to heating reflux and ultrasound-assisted extraction, centrifugation, and solid-liquid separation to obtain a Cistanche deserticola p-coumaric acid residue extract and a residue precipitate. The heating reflux extraction temperature was 80°C, the heating reflux extraction time was 3 h, the ultrasound-assisted extraction time was 75 min, the ultrasound-assisted extraction temperature was 35°C, the ultrasound power was 600 W, the centrifugation time was 25 min, and the centrifugation speed was 12000 rpm / min;

[0039] (4) Acid-base hydrolysis and extraction of the residue: The residue precipitate obtained in step (3) is subjected to acid-base hydrolysis-ethyl acetate extraction method, 4 mol / L NaOH solution is added to the residue, the mixture is thoroughly mixed at 50° C., and the mixture is hydrolyzed for 4.5 h in the dark, 4 mol / L HCl solution is added, the pH value is adjusted to 1.5, and after thorough shaking, ethyl acetate is added for extraction for 2 h, wherein the volume ratio of the solution to ethyl acetate is 20:1; the obtained extract is combined with the crude p-coumaric acid extract of Cistanche deserticola obtained in step (2) and the residue extract obtained in step (3) to obtain a combined solution;

[0040] (5) Filtration and purification: The combined solution obtained in step (4) is filtered through a filter membrane with a pore size of 200 nm, and then purified. The macroporous resin used is D101 macroporous adsorption resin, and the macroporous resin column diameter-to-height ratio is 1:5; it includes a water washing and impurity removal step and an ethanol solution elution step. In the water washing and impurity removal step, the water washing volume is 6.5 column volumes (BV), and the water washing flow rate is 2.5 column volumes per hour (BV / h); in the ethanol solution elution step, the ethanol volume fraction is 70%, the ethanol solution elution volume is 5 column volumes (BV), and the ethanol solution elution flow rate is 6 column volumes per hour (BV / h), and the extraction rate of coumaric acid from Cistanche deserticola is 13.92%, and the purity of coumaric acid from Cistanche deserticola is 97.39%.

[0041] Example 3

[0042] A method for extracting p-coumaric acid from Cistanche deserticola comprises the following steps:

[0043] (1) Raw material pretreatment: 50 g of Cistanche deserticola was freeze-dried at -30°C for 36 h with a vacuum degree of 35 Pa. After freeze-drying, it was naturally warmed to room temperature. After being ground into powder, the passing rate under a 50-mesh sieve was 100%;

[0044] (2) Ethanol ultrasonic extraction: 70% ethanol was added to the powder obtained in step (1) at a mass volume ratio of 1:30 g / mL, and ultrasonic extraction was performed at 50° C. for 60 min with an ultrasonic power of 400 W. The crude p-coumaric acid extract of Cistanche deserticola and the residue were obtained by solid-liquid separation;

[0045] (3) Re-extraction of the residue: The residue of step (2) was immersed in a 60% ethanol solution at a solid-liquid ratio of 1:25 g / mL, and allowed to stand for 30 min. Heating reflux and ultrasonic-assisted extraction were performed, and the extract of p-coumaric acid residue of Cistanche deserticola and the residue precipitate were obtained by centrifugation. The heating reflux extraction temperature was 85°C, the heating reflux extraction time was 4 h, the ultrasonic-assisted extraction time was 90 min, the ultrasonic-assisted extraction temperature was 40°C, the ultrasonic power was 700 W, the centrifugation time was 30 min, and the centrifugation speed was 13000 rpm / min;

[0046] (4) Acid-base hydrolysis and extraction of the residue: The residue precipitate obtained in step (3) is subjected to acid-base hydrolysis-ethyl acetate extraction method, 4 mol / L NaOH solution is added to the residue, the mixture is thoroughly mixed at 55° C., and the mixture is hydrolyzed for 5 h in the dark, 4 mol / L HCl solution is added, the pH value is adjusted to 2.0, and after thorough shaking, ethyl acetate is added for extraction for 3 h, wherein the volume ratio of the solution to ethyl acetate is 20:1; the obtained extract is combined with the crude p-coumaric acid extract of Cistanche deserticola obtained in step (2) and the residue extract obtained in step (3) to obtain a combined solution;

[0047] (5) Filtration and purification: The combined solution obtained in step (4) is filtered through a filter membrane with a pore size of 500 nm, and then purified. The macroporous resin used is D101 macroporous adsorption resin, and the macroporous resin column diameter-to-height ratio is 1:6; it includes a water washing and impurity removal step and an ethanol solution elution step. In the water washing and impurity removal step, the water washing volume is 7 times the column volume (BV), and the water washing flow rate is 3 times the column volume per hour (BV / h); in the ethanol solution elution step, the ethanol volume fraction is 75%, the ethanol solution elution volume is 6 times the column volume (BV), and the ethanol solution elution flow rate is 7 times the column volume per hour (BV / h), and the extraction rate of coumaric acid from Cistanche deserticola is 10.21%, and the purity of coumaric acid from Cistanche deserticola is 93.46%.

[0048] Comparative Example 1

[0049] This comparative example provides an effect of non-ultrasound-assisted extraction on the extraction rate and purity of p-coumaric acid from Cistanche deserticola, comprising the following steps:

[0050] (1) Raw material pretreatment: 50 g of Cistanche deserticola was freeze-dried at -40°C for 30 h with a vacuum degree of 30 Pa. After freeze-drying, the mixture was naturally heated to room temperature. After being ground into powder, the passing rate under a 50-mesh sieve was 100%;

[0051] (2) ethanol extraction: adding 70% ethanol at a mass volume ratio of 1:25 g / mL to the powder obtained in step (1) and extracting for 45 min, and separating the solid and liquid to obtain a crude p-coumaric acid extract of Cistanche deserticola and a residue;

[0052] (3) Re-extraction of the residue: The residue of step (2) is immersed in a 60% ethanol solution at a solid-liquid ratio of 1:25 g / mL, left to stand for 30 min, extracted by heating and reflux, centrifuged, and solid-liquid separation is performed to obtain a Cistanche deserticola p-coumaric acid residue extract and a residue precipitate. The heating and reflux extraction temperature is 80°C, the heating and reflux extraction time is 3 h, the centrifugation time is 25 min, and the centrifugation speed is 12000 rpm / min;

[0053] (4) Acid-base hydrolysis and extraction of the residue: The residue precipitate obtained in step (3) is subjected to acid-base hydrolysis-ethyl acetate extraction method, 4 mol / L NaOH solution is added to the residue, the mixture is thoroughly mixed at 50° C., and the mixture is hydrolyzed for 4.5 h in the dark, 4 mol / L HCl solution is added, the pH value is adjusted to 1.5, and after thorough shaking, ethyl acetate is added for extraction for 2 h, wherein the volume ratio of the solution to ethyl acetate is 20:1; the obtained extract is combined with the crude p-coumaric acid extract of Cistanche deserticola obtained in step (2) and the residue extract obtained in step (3) to obtain a combined solution;

[0054] (5) Filtration and purification: The combined solution obtained in step (4) is filtered through a filter membrane with a pore size of 200 nm, and then purified. The macroporous resin used is D101 macroporous adsorption resin, and the macroporous resin column diameter-to-height ratio is 1:5; it includes a water washing and impurity removal step and an ethanol solution elution step. In the water washing and impurity removal step, the water washing volume is 6.5 column volumes (BV), and the water washing flow rate is 2.5 column volumes per hour (BV / h); in the ethanol solution elution step, the ethanol volume fraction is 70%, the ethanol solution elution volume is 5 column volumes (BV), and the ethanol solution elution flow rate is 6 column volumes per hour (BV / h).

[0055] The difference between Comparative Example 1 and Example 2 is that: in step (2), ultrasound-assisted extraction is not performed, and the extraction rate of p-coumaric acid from Cistanche deserticola is 10.73%, and the purity of p-coumaric acid from Cistanche deserticola is 94.27%.

[0056] Comparative Example 2

[0057] The effect of acid-base-free hydrolysis extraction on the extraction rate and purity of p-coumaric acid from Cistanche deserticola includes the following steps:

[0058] (1) Raw material pretreatment: 50 g of Cistanche deserticola was freeze-dried at -40°C for 30 h with a vacuum degree of 30 Pa. After freeze-drying, the mixture was naturally heated to room temperature. After being ground into powder, the passing rate under a 50-mesh sieve was 100%;

[0059] (2) Ethanol ultrasonic extraction: 70% ethanol was added to the powder obtained in step (1) at a mass volume ratio of 1:25 g / mL, and ultrasonic extraction was performed at 45° C. for 45 min with an ultrasonic power of 400 W. The crude p-coumaric acid extract of Cistanche deserticola and the residue were obtained by solid-liquid separation;

[0060] (3) Re-extraction of the residue: The residue obtained in step (2) was immersed in a 60% ethanol solution at a solid-liquid ratio of 1:25 g / mL, and allowed to stand for 30 min. Heating reflux and ultrasonic-assisted extraction were performed, and the extract of p-coumaric acid residue of Cistanche deserticola and the residue precipitate were obtained by solid-liquid separation. The heating reflux extraction temperature was 80°C, the heating reflux extraction time was 3 h, the ultrasonic-assisted extraction time was 75 min, the ultrasonic-assisted extraction temperature was 35°C, the ultrasonic power was 600 W, the centrifugation time was 25 min, and the centrifugation speed was 12000 rpm / min;

[0061] (4) Extraction of the residue: the residue precipitate obtained in step (3) is added with ethyl acetate for extraction for 2 h, and the volume ratio of the solution to ethyl acetate is 20:1; the obtained extract is combined with the p-coumaric acid crude extract of Cistanche deserticola obtained in step (2) and the residue extract obtained in step (3) to obtain a combined solution;

[0062] (5) Filtration and purification: The combined solution obtained in step (4) is filtered through a filter membrane with a pore size of 200 nm, and then purified. The macroporous resin used is D101 macroporous adsorption resin, and the macroporous resin column diameter-to-height ratio is 1:5; it includes a water washing and impurity removal step and an ethanol solution elution step. In the water washing and impurity removal step, the water washing volume is 6.5 column volumes (BV), and the water washing flow rate is 2.5 column volumes per hour (BV / h); in the ethanol solution elution step, the ethanol volume fraction is 70%, the ethanol solution elution volume is 5 column volumes (BV), and the ethanol solution elution flow rate is 6 column volumes per hour (BV / h).

[0063] The difference between Comparative Example 2 and Example 2 is that: in step (4), no acid-base hydrolysis extraction is performed, and the extraction rate of p-coumaric acid from Cistanche deserticola is 13.72%, and the purity of p-coumaric acid from Cistanche deserticola is 90.01%.

[0064] Comparative Example 3

[0065] The effect of replacing the macroporous resin on the extraction rate and purity of p-coumaric acid from Cistanche deserticola includes the following steps:

[0066] (1) Raw material pretreatment: 50 g of Cistanche deserticola was freeze-dried at -40°C for 30 h with a vacuum degree of 30 Pa. After freeze-drying, the mixture was naturally heated to room temperature. After being ground into powder, the passing rate under a 50-mesh sieve was 100%;

[0067] (2) Ethanol ultrasonic extraction: 70% ethanol was added to the powder obtained in step (1) at a mass volume ratio of 1:25 g / mL, and ultrasonic extraction was performed at 45° C. for 45 min with an ultrasonic power of 400 W. The crude p-coumaric acid extract of Cistanche deserticola and the residue were obtained by solid-liquid separation;

[0068] (3) Re-extraction of the residue: The residue of step (2) was immersed in a 60% ethanol solution at a solid-liquid ratio of 1:25 g / mL, left to stand for 30 min, and subjected to heating reflux and ultrasound-assisted extraction, centrifugation, and solid-liquid separation to obtain a Cistanche deserticola p-coumaric acid residue extract and a residue precipitate. The heating reflux extraction temperature was 80°C, the heating reflux extraction time was 3 h, the ultrasound-assisted extraction time was 75 min, the ultrasound-assisted extraction temperature was 35°C, the ultrasound power was 600 W, the centrifugation time was 25 min, and the centrifugation speed was 12000 rpm / min;

[0069] (4) Acid-base hydrolysis and extraction of the residue: The residue precipitate obtained in step (3) is subjected to acid-base hydrolysis-ethyl acetate extraction method, 4 mol / L NaOH solution is added to the residue, the mixture is thoroughly mixed at 50° C., and the mixture is hydrolyzed for 4.5 h in the dark, 4 mol / L HCl solution is added, the pH value is adjusted to 1.5, and after thorough shaking, ethyl acetate is added for extraction for 2 h, wherein the volume ratio of the solution to ethyl acetate is 20:1; the obtained extract is combined with the crude p-coumaric acid extract of Cistanche deserticola obtained in step (2) and the residue extract obtained in step (3) to obtain a combined solution;

[0070] (5) Filtration and purification: The combined solution obtained in step (4) is filtered through a filter membrane with a pore size of 200 nm, and then purified. The macroporous resin used is HPD100 macroporous resin, and the macroporous resin column diameter-to-height ratio is 1:5; it includes a water washing and impurity removal step and an ethanol solution elution step. In the water washing and impurity removal step, the water washing volume is 6.5 column volumes (BV), and the water washing flow rate is 2.5 column volumes per hour (BV / h); in the ethanol solution elution step, the ethanol volume fraction is 70%, the ethanol solution elution volume is 5 column volumes (BV), and the ethanol solution elution flow rate is 6 column volumes per hour (BV / h).

[0071] The difference between Comparative Example 3 and Example 2 is that in step (5), the D101 macroporous adsorption resin is replaced by the HPD100 macroporous adsorption resin. The extraction rate of p-coumaric acid from Cistanche deserticola is 13.45%, and the purity is 95.49%.

[0072] In summary, according to the embodiments of the present invention, the present invention has established a method for extracting p-coumaric acid from Cistanche deserticola, so that Cistanche deserticola can be more fully utilized; when p-coumaric acid in a mixed solution of Cistanche deserticola is extracted by ultrasound-assisted technology, the extraction efficiency is significantly improved; the acid-base hydrolysis-ethyl acetate extraction method is adopted to ensure the maximum utilization and purification of the p-coumaric acid component, while also minimizing the waste of resources.

[0073] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" etc. means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the schematic representation of the above terms should not be understood as necessarily being directed to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described may be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art may combine and combine different embodiments or examples described in this specification.

[0074] Although the embodiments of the present invention have been shown and described above, it is to be understood that the above embodiments are exemplary and are not to be construed as limitations of the present invention. A person skilled in the art may change, modify, replace and vary the above embodiments within the scope of the present invention.

Claims

1. A method for extracting p-coumaric acid from Cistanche deserticola, characterized in that: The following steps are involved: (1) freeze-drying and grinding the deserticola to obtain deserticola powder; (2) adding ethanol to the desert Cistanche powder, performing ultrasonic extraction, and performing solid-liquid separation to obtain a desert Cistanche p-coumaric acid crude extract and a residue; (3) adding ethanol to the residue and allowing it to stand, then heating and refluxing, ultrasonic-assisted extraction, centrifuging, and solid-liquid separation to obtain a p-coumaric acid residue extract of Cistanche deserticola and a residue precipitate; (4) subjecting the residual precipitate of step (3) to acid-base hydrolysis-ethyl acetate extraction to obtain an extract; combining the extract with the crude p-coumaric acid extract of Cistanche deserticola and the p-coumaric acid residue extract of Cistanche deserticola to obtain a combined solution; (5) filtering and purifying the combined solution through a filter membrane to obtain p-coumaric acid.

2. The method according to claim 1, characterized in that In step (1), the freeze-drying temperature is -50°C to -20°C, the freeze-drying time is 24 to 36 hours, the vacuum degree is 25 to 35 Pa, and after the freeze-drying is completed, the temperature is naturally raised to room temperature.

3. The method according to claim 1, characterized in that In step (2), the deserticola powder and ethanol are added in a mass volume ratio of 1:20 to 1:30 g / mL, and the concentration of the ethanol is 70%.

4. The method according to claim 1, characterized in that In step (2), the temperature of ultrasonic extraction is 40-50°C, the ultrasonic extraction time is 30-60min, and the ultrasonic power is 400W.

5. The method according to claim 1, characterized in that In step (3), the residue and ethanol are added in a mass volume ratio of 1:25 g / mL, the concentration of ethanol is 60%, and the mixture is allowed to stand for 30 minutes.

6. The method according to claim 1, characterized in that In step (3), the heating reflux temperature is 75 to 85° C., and the heating reflux extraction time is 2 to 4 hours.

7. The method according to claim 1, characterized in that In step (3), the ultrasonic-assisted extraction time is 60 to 90 minutes, the ultrasonic-assisted extraction temperature is 30 to 40° C., and the ultrasonic power is 500 to 800W.

8. The method according to claim 1, characterized in that In step (3), the centrifugation time is 20 to 30 minutes, and the centrifugation speed is 10000 to 13000 rpm / min.

9. The method according to claim 1, characterized in that In step (4), acid-base hydrolysis-ethyl acetate extraction is as follows: add NaOH solution to the residue precipitate, mix thoroughly at 45-55° C., hydrolyze for 4-5 hours in the dark, add HCl solution, adjust the pH value to 1-2, shake thoroughly, and extract with ethyl acetate for 1-3 hours, wherein the volume ratio of the solution to ethyl acetate is 20:

1.

10. The method according to claim 1, characterized in that In step (5), the filtration is performed using a filter membrane with a pore size of 100 to 500 nm.

11. The method according to claim 1, characterized in that In step (5), the purification is performed using D101 macroporous adsorption resin, the macroporous resin column diameter-to-height ratio is 1:4-6, and includes water washing to remove impurities and elution with ethanol solution.

12. The method according to claim 11, characterized in that In step (5), the washing volume for water washing and impurity removal is 6-7 times the column volume BV, the washing flow rate is 2-3 times the column volume BV / h per hour, the ethanol volume fraction of the ethanol solution elution is 65%-75%, the elution volume of the ethanol solution is 4-6 times the column volume BV, and the elution flow rate of the ethanol solution is 5-7 times the column volume BV / h per hour.