Preparation method of dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases

Abstract

The invention discloses a method for preparing dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases, firstly, an enzyme with cellulase and larch powder of predetermined particle size are mixed and enzymolyzed to obtain larch hydrolyzate; and after the larch hydrolyzate and 81% concentration of ethanol solution are mixed in a ratio of 1-5:3-10, the obtained mixed solution is placed in an ultrasonic extractor for extraction for 62-100 minutes to obtain an extract; then the extract is pressurized and heated in an autoclave, and dihydroquercetin is obtained after the extract is crystallized; finally, the crystallized dihydroquercetin is measured by ultraviolet spectroscopy, and the purity is greater than 90% and is screened to obtain a dihydroquercetin refined product. Thus, the obtained purity of the dihydroquercetin extracted under different conditions can be accurately measured, and then its extraction rate is improved and the purity can be accurately determined.

Description

Technical Field

[0001] The invention relates to the technical field of dihydroquercetin, and in particular to a method for preparing dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases. Background Art

[0002] Dihydroquercetin is a natural medicine with significant antioxidant effect. It can be used as a main and auxiliary raw material of medicine and a bioactive food additive and is widely used in medicines and health functional foods. It is widely used in the fields of medicine, industry and agriculture. Dihydroquercetin belongs to the dihydroflavonoids, vitamin P family, and is a strong antioxidant. Among natural antioxidants, the adsorption of free radicals can reach 32μmol / mg (dihydroquercetin purity 95%). As a natural antioxidant, dihydroquercetin has a significant effect on the scavenging of free radicals and prolonging the life of cells. It can be expected that dihydroquercetin has strong biological activity and medicinal value. In the relevant technology, there are certain differences in the purity of dihydroquercetin extracted under different conditions, resulting in low extraction rate and difficulty in accurately determining the purity. Summary of the invention

[0003] The present invention aims to solve at least one of the technical problems in the related art to a certain extent. To this end, one object of the present invention is to provide a method for preparing dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases, comprising:

[0004] S10, mixing an enzyme containing cellulase and larch powder of a predetermined particle size for enzymolysis to obtain a larch enzymolysis solution;

[0005] S20, mixing the larch enzymatic hydrolyzate and 81% ethanol solution in a ratio of 1-5:3-10, placing the obtained mixed solution into an ultrasonic extractor for extraction for 62-100 minutes to obtain an extract;

[0006] S30, placing the extract in an autoclave to pressurize and heat the extract, and obtaining dihydroquercetin after the extract is crystallized;

[0007] S40, measuring the crystallized dihydroquercetin by ultraviolet spectroscopy, and screening those with a purity greater than 90% to obtain a refined dihydroquercetin product.

[0008] Optionally, the step S10 includes:

[0009] Providing larch powder and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution;

[0010] The enzyme containing cellulase and the mixed solution are mixed and enzymolyzed for 2-2.5 hours, and added into a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 5-10 minutes;

[0011] After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate.

[0012] Optionally, the predetermined particle size is 0.11 mm-0.16 mm.

[0013] Optionally, the frequency of the ultrasonic extractor is 5 MHZ, the pulse cycle frequency is 5 times / second, and the sound beam is 1000 m / second.

[0014] Optionally, the step S30 includes:

[0015] The extract is placed in an autoclave and pressurized to 550-580 MPa;

[0016] After the extract solution is kept under pressure for 12 minutes, the pressure is reduced and the temperature of the extract solution is raised to 92° C. while being continuously stirred;

[0017] After the extract is cooled, it is allowed to stand at 23-28° C. for 4 days to obtain crystalline dihydroquercetin.

[0018] Optionally, the step S40 includes:

[0019] Select 5 samples of crystallized dihydroquercetin, and select 2-7 g from each sample and dissolve it in 99.5% ethanol;

[0020] Each sample is measured using an ultraviolet spectrum of a predetermined wavelength to determine the absorbance intensity of each sample;

[0021] According to the absorbance intensity of each sample, the refined dihydroquercetin with a purity greater than 90% was screened out.

[0022] Optionally, the predetermined wavelength is 235nm-270nm.

[0023] Optionally, the amount of the cellulase used is 2.5-2.9% of the total mass of the larch powder; and the enzyme activity of the cellulase is ≥ 60,000 u / g.

[0024] The preparation method of dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases provided by the present invention comprises the following steps: firstly, an enzyme having cellulase and larch powder of a predetermined particle size are mixed and enzymolyzed to obtain a larch hydrolyzate; and after mixing the larch hydrolyzate and an ethanol solution of 81% concentration in a ratio of 1-5:3-10, the obtained mixed solution is placed in an ultrasonic extractor for extraction for 62-100 minutes to obtain an extract; and then the extract is pressurized and heated in an autoclave to obtain dihydroquercetin after crystallization of the extract; and finally, the crystallized dihydroquercetin is measured by ultraviolet spectroscopy, and the purity is greater than 90% for screening to obtain a refined dihydroquercetin product. Thus, the obtained purity of dihydroquercetin extracted under different conditions can be accurately measured, thereby improving its extraction rate and the purity can be accurately determined.

[0025] 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

[0026] The embodiments of the present invention are described in detail below, which are intended to be used to explain the present invention but should not be construed as limiting the present invention. All other embodiments obtained by ordinary technicians in the field without creative work based on the embodiments of the present invention shall fall within the scope of protection of the present invention.

[0027] The following is a detailed description of the method for preparing dihydroquercetin for anti-cancer, anti-oxidation, and anti-cardiovascular and cerebrovascular diseases according to an embodiment of the present invention.

[0028] According to the method for preparing dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases provided by the present invention, the following first to fifth examples prepare dihydroquercetin under different conditions, and measure the purity of the dihydroquercetin obtained under different conditions.

[0029] First embodiment

[0030] The method for preparing the Phellinus linteus extract provided in the first embodiment of the present invention comprises:

[0031] Providing larch powder with a particle size of 0.12 mm and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution;

[0032] The enzyme and the mixed solution with cellulase were mixed and enzymatically hydrolyzed for 2 hours, and added to a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 6 minutes;

[0033] After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate;

[0034] The larch enzymatic hydrolysate and 81% ethanol solution were mixed in a ratio of 1:4, and the obtained mixed solution was placed in an ultrasonic extractor for extraction for 67 minutes to obtain an extract;

[0035] The extract was placed in an autoclave and pressurized to 550 MPa;

[0036] After the extract solution was kept under pressure for 12 minutes, the pressure was reduced and the temperature of the extract solution was raised to 92° C. while being continuously stirred;

[0037] After the extract is cooled, it is allowed to stand at 24° C. for 4 days to obtain crystalline dihydroquercetin;

[0038] The crystallized dihydroquercetin was used as a sample, and 3 g was selected and dissolved in 99.5% ethanol;

[0039] The sample was measured using a 240nm ultraviolet spectrum to determine the corresponding absorbance intensity.

[0040] Second embodiment

[0041] The second embodiment of the present invention provides a method for extracting dihydroquercetin, comprising:

[0042] Providing larch powder with a particle size of 0.14 mm and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution;

[0043] The enzyme and the mixed solution with cellulase were mixed and enzymatically hydrolyzed for 2 hours, and added to a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 8 minutes;

[0044] After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate;

[0045] larch enzymatic hydrolysate and 81% ethanol solution were mixed in a ratio of 2:9, and the obtained mixed solution was placed in an ultrasonic extractor for extraction for 80 minutes to obtain an extract;

[0046] The extract was placed in an autoclave and pressurized to 570 MPa;

[0047] After the extract solution was kept under pressure for 12 minutes, the pressure was reduced and the temperature of the extract solution was raised to 92° C. while being continuously stirred;

[0048] After the extract is cooled, it is allowed to stand at 27° C. for 4 days to obtain crystalline dihydroquercetin;

[0049] The crystallized dihydroquercetin was used as a sample, and 3 g was selected and dissolved in 99.5% ethanol;

[0050] The sample was measured using a 240nm ultraviolet spectrum to determine the corresponding absorbance intensity.

[0051] Third embodiment

[0052] The third embodiment of the present invention provides a method for extracting dihydroquercetin, comprising:

[0053] Providing larch powder with a particle size of 0.11 mm and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution;

[0054] The enzyme and the mixed solution with cellulase were mixed and enzymatically hydrolyzed for 2 hours, and added to a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 8 minutes;

[0055] After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate;

[0056] larch enzymatic hydrolysate and 81% ethanol solution were mixed in a ratio of 3:10, and the obtained mixed solution was placed in an ultrasonic extractor for extraction for 75 minutes to obtain an extract;

[0057] The extract was placed in an autoclave and pressurized to 570 MPa;

[0058] After the extract solution was kept under pressure for 12 minutes, the pressure was reduced and the temperature of the extract solution was raised to 92° C. while being continuously stirred;

[0059] After the extract is cooled, it is allowed to stand at 27° C. for 4 days to obtain crystalline dihydroquercetin;

[0060] The crystallized dihydroquercetin was used as a sample, and 6 g was selected and dissolved in 99.5% ethanol;

[0061] The sample was measured using a 240nm ultraviolet spectrum to determine the corresponding absorbance intensity.

[0062] Fourth embodiment

[0063] The fourth embodiment of the present invention provides a method for extracting dihydroquercetin, comprising:

[0064] Providing larch powder with a particle size of 0.15 mm and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution;

[0065] The enzyme and the mixed solution with cellulase were mixed and enzymatically hydrolyzed for 2.5 hours, and added to a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 8 minutes;

[0066] After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate;

[0067] larch enzymatic hydrolysate and 81% ethanol solution were mixed in a ratio of 2:7, and the obtained mixed solution was placed in an ultrasonic extractor for extraction for 90 minutes to obtain an extract;

[0068] The extract was placed in an autoclave and pressurized to 580 MPa;

[0069] After the extract solution was kept under pressure for 12 minutes, the pressure was reduced and the temperature of the extract solution was raised to 92° C. while being continuously stirred;

[0070] After the extract is cooled, it is allowed to stand at 25° C. for 4 days to obtain crystalline dihydroquercetin;

[0071] The crystallized dihydroquercetin was used as a sample, and 6 g was selected and dissolved in 99.5% ethanol;

[0072] The sample was measured using a 240nm ultraviolet spectrum to determine the corresponding absorbance intensity.

[0073] Fifth embodiment

[0074] The fifth embodiment of the present invention provides a method for extracting dihydroquercetin, comprising:

[0075] Providing larch powder with a particle size of 0.11 mm and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution;

[0076] The enzyme and the mixed solution with cellulase were mixed and enzymatically hydrolyzed for 2.5 hours, and added to a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 10 minutes;

[0077] After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate;

[0078] larch enzymatic hydrolysate and 81% ethanol solution were mixed in a ratio of 1:9, and the obtained mixed solution was placed in an ultrasonic extractor for extraction for 90 minutes to obtain an extract;

[0079] The extract was placed in an autoclave and pressurized to 580 MPa;

[0080] After the extract solution was kept under pressure for 12 minutes, the pressure was reduced and the temperature of the extract solution was raised to 92° C. while being continuously stirred;

[0081] After the extract is cooled, it is allowed to stand at 25° C. for 4 days to obtain crystalline dihydroquercetin;

[0082] The crystallized dihydroquercetin was used as a sample, and 6 g was selected and dissolved in 99.5% ethanol;

[0083] The sample was measured using a 240nm ultraviolet spectrum to determine the corresponding absorbance intensity.

[0084] Combining the following table with the above-mentioned first to fifth embodiments, the test results are shown in the following table.

[0085] Example First embodiment Second embodiment Third embodiment Fourth embodiment Fifth embodiment Larch powder particle size (mm) 0.12 0.14 0.11 0.15 0.11 Larch enzymatic hydrolysate: ethanol solution (ml) 1:4 2:9 3:10 2:7 1:9 Pressure (Mpa) 550 570 570 580 580 Absorption intensity (L*mol-1*cm-1) 15 18 20 19 22

[0086] From the comparison of the first to fifth embodiments in the above table, it can be seen that in the fifth embodiment, the smaller the particle size of the larch powder and the greater the pressure, the higher the corresponding absorbance intensity, that is, the higher the corresponding purity. Therefore, the conditions in the fifth embodiment can be used to extract dihydroquercetin, thereby improving its extraction rate.

[0087] The preparation method of dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases provided by the present invention comprises the following steps: firstly, an enzyme having cellulase and larch powder of a predetermined particle size are mixed and enzymolyzed to obtain larch hydrolyzate; and after the larch hydrolyzate and 81% ethanol solution are mixed in a ratio of 1-5:3-10, the obtained mixed solution is placed in an ultrasonic extractor for extraction for 62-100 minutes to obtain an extract; and then the extract is pressurized and heated in an autoclave to obtain dihydroquercetin after crystallization of the extract; and finally, the crystallized dihydroquercetin is measured by ultraviolet spectroscopy, and the purity is greater than 90% and is screened to obtain a refined dihydroquercetin product. Thus, the obtained purity of dihydroquercetin extracted under different conditions can be accurately measured, thereby improving its extraction rate and the purity can be accurately determined.

[0088] 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 representations of the above terms do not necessarily refer 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 the different embodiments or examples described in this specification and the features of the different embodiments or examples, without contradiction.

[0089] The above descriptions are only preferred embodiments of the present invention, and are not intended to limit the patent scope of the present invention. Any direct / indirect application in other related technical fields under the inventive concept of the present invention is included in the patent protection scope of the present invention.

Claims

1. A method for preparing dihydroquercetin for anti-cancer, anti-oxidation and anti-cardiovascular and cerebrovascular diseases, characterized in that: include: S10, mixing an enzyme containing cellulase and larch powder of a predetermined particle size for enzymolysis to obtain a larch enzymolysis solution; S20, mixing the larch enzymatic hydrolyzate and 81% ethanol solution in a ratio of 1-5:3-10, placing the obtained mixed solution into an ultrasonic extractor for extraction for 62-100 minutes to obtain an extract; S30, placing the extract in an autoclave to pressurize and heat the extract, and obtaining dihydroquercetin after the extract is crystallized; S40, measuring the crystallized dihydroquercetin by ultraviolet spectroscopy, and screening those with a purity greater than 90% to obtain a refined dihydroquercetin product; The step S10 comprises: Providing larch powder and deionized water and mixing them in a ratio of 1:15 to obtain a mixed solution; The enzyme containing cellulase and the mixed solution are mixed and enzymolyzed for 2-2.5 hours, and added into a centrifuge to be centrifuged at a rotation speed of 6000 r / min for 5-10 minutes; After centrifugation, the supernatant was collected to obtain larch enzymatic hydrolysate; The predetermined particle size is 0.11 mm-0.16 mm; The step S40 comprises: Select 5 samples of crystallized dihydroquercetin, and select 2-7 g from each sample and dissolve it in 99.5% ethanol; Each sample is measured using an ultraviolet spectrum of a predetermined wavelength to determine the absorbance intensity of each sample; According to the absorbance intensity of each sample, the refined dihydroquercetin with a purity greater than 90% was screened out.

2. The method for preparing dihydroquercetin for anti-cancer, anti-oxidation, and anti-cardiovascular and cerebrovascular diseases according to claim 1, characterized in that: The frequency of the ultrasonic extractor is 5 MHZ, the pulse cycle frequency is 5 times / second, and the sound beam is 1000 m / second.

3. The method for preparing dihydroquercetin for anti-cancer, anti-oxidation, and anti-cardiovascular and cerebrovascular diseases according to claim 1, characterized in that: The step S30 comprises: The extract is placed in an autoclave and pressurized to 550-580 MPa; After the extract solution is kept under pressure for 12 minutes, the pressure is reduced and the temperature of the extract solution is raised to 92° C. while being continuously stirred; After the extract is cooled, it is allowed to stand at 23-28° C. for 4 days to obtain crystalline dihydroquercetin.

4. The method for preparing dihydroquercetin for anti-cancer, anti-oxidation, and anti-cardiovascular and cerebrovascular diseases according to claim 1, characterized in that: The predetermined wavelength is 235nm-270nm.

5. The method for preparing dihydroquercetin for anti-cancer, anti-oxidation, and anti-cardiovascular and cerebrovascular diseases according to claim 1, characterized in that: The dosage of the cellulase is 2.5-2.9% of the total mass of the larch powder; and the enzyme activity of the cellulase is ≥60,000 u / g.