CN111481583A - Method for efficiently extracting total flavonoids from sauropus spatulifolius - Google Patents
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Abstract
The invention discloses a method for efficiently extracting total flavonoids from sauropus spatholobus, which comprises the steps of adding citric acid, cellulase and pectinase into sauropus spatholobus, taking out sauropus spatholobus and adding water, adding D-sodium erythorbate, sodium chloride and nisin, and pulping to obtain an extracting solution of sauropus spatholobus; adding fermentation strains, brown sugar, urea, ammonium sulfate and potassium dihydrogen phosphate into the sauropus spatulifolius extract, uniformly stirring, and fermenting; the fermentation strain consists of bacillus marinus, aspergillus niger and candida utilis; heating and refluxing the fermented sauropus spatholobus stem extract, performing suction filtration, performing reduced pressure concentration, purifying the concentrated extract by using a macroporous adsorption resin column, and drying to obtain sauropus spatholobus stem total flavonoids. The method comprises the steps of soaking the sauropus spatulifolius in citric acid, cellulase and pectinase to carry out enzymolysis on the sauropus spatulifolius, fermenting the sauropus spatulifolius with mixed strains, and then carrying out heating reflux extraction, so that leaching of effective components of total flavonoids in the sauropus spatulifolius is facilitated, the extraction amount of the total flavonoids is more than 2.8 g/kg), and the content of the total flavonoids is more than 80%.
Description
Technical Field
The invention relates to the technical field of extraction of effective components of plants, in particular to a method for efficiently extracting total flavonoids from sauropus spatulifolius.
Background
Is dried leaves of Sauropus spatulifolius Beille of the family Euphorbiaceae and mainly distributed in Guangxi, Guangdong, Fujian, etc. The sauropus spatulifolius is a Guangxi characteristic traditional Chinese medicine, has the effects of ventilating, moistening lung to arrest cough, relaxing bowel and the like, and is a common strengthening medicine in clinic. The sauropus spatulifolius is medicinal and edible, and is folk in south China
The herbal tea is often used for dispelling phlegm and eliminating dampness, and can also be decocted with lean pork or pig lung for health promotion for cough due to lung heat, aphonia, laryngalgia, acute bronchitis, upper respiratory tract inflammation, etc.
The Guangxi sauropus spatulifolius has rich resources, the medicinal components of the Guangxi sauropus spatulifolius are deeply researched, and the medicinal development and utilization tool for the Longxiaye sauropus spatulifolius
The method is a natural antioxidant, can remove free radicals in a human body and inhibit the generation of the free radicals, and can be used as a natural food preservative and widely applied to the preservation and processing of products, and has been studied to some extent at present.
The above background disclosure is only for the purpose of assisting understanding of the inventive concept and technical solutions of the present invention, and does not necessarily belong to the prior art of the present patent application, and should not be used for evaluating the novelty and inventive step of the present application in the case that there is no clear evidence that the above content is disclosed at the filing date of the present patent application.
Disclosure of Invention
The invention provides a method for efficiently extracting total flavonoids from sauropus spatulifolius aiming at the defects of the current method for extracting the total flavonoids from sauropus spatulifolius. According to the method, citric acid, cellulase and pectinase are adopted for soaking to perform enzymolysis on sauropus spatulifolius, and then mixed strains are used for fermentation and then heating reflux extraction is performed, so that leaching of effective components of total flavonoids in sauropus spatulifolius is facilitated, and the extraction rate of the total flavonoids is improved; d-sodium erythorbate, sodium chloride and nisin are added in the pulping process, so that the medicinal materials are not easy to oxidize, and the stability of the total flavonoids is improved.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
a method for efficiently extracting total flavonoids from sauropus spatulifolius comprises the following steps:
(1) picking the sauropus, freezing for 24-48h, taking out, thawing with warm water, adding a soaking solution consisting of citric acid, cellulase and pectinase, soaking for 2-6h, and taking out the sauropus;
(2) adding water according to the use amount of L according to the material-liquid ratio of 1:20-25g/m, adding D-sodium erythorbate, sodium chloride and nisin, and pulping to obtain an extracting solution of sauropus spatholobus stem;
(3) adding fermentation strains, brown sugar, urea, ammonium sulfate and potassium dihydrogen phosphate into the sauropus spatulifolius extract, uniformly stirring, and fermenting; the fermentation strain consists of bacillus marinus, aspergillus niger and candida utilis;
(4) heating and refluxing the fermented sauropus spatholobus stem extract, performing suction filtration, and performing reduced pressure concentration to obtain a concentrated extract;
(5) dissolving the concentrated extract in 4-6 times of distilled water, centrifuging, and loading the supernatant onto macroporous adsorbent resin column at a loading flow rate of 1.0-3.0 BV/h; eluting at an elution speed of 1.0-3.0BV/h, eluting with water until colorless and transparent, eluting with ethanol and ethyl alcohol eluent, collecting the eluent, concentrating under reduced pressure, recovering the eluent, and drying to obtain the sauropus spatulifolius total flavonoids.
Further, the mass ratio of the sauropus spatulifolius to the citric acid, the cellulase and the pectinase is 100:1-2:0.5-1: 0.3-0.5.
Further, the mass ratio of the sodium D-isoascorbate, the sodium chloride, the nisin to the sauropus spatulifolius is 0.5-1.0:1-3:0.3-0.6: 100.
Further, the mass ratio of the fermentation strain, the brown sugar, the urea, the ammonium sulfate, the potassium dihydrogen phosphate to the sauropus spatulifolius is 1-3:4-8:0.5-1.0:0.3-0.5:0.1-0.3: 100.
Further, the fermentation strain consists of marine bacillus, aspergillus niger and candida utilis in a mass ratio of 3-5:0.5-2: 1-3.
Further, the heating reflux extraction is reflux extraction at 70-80 deg.C for 3 times, each time for 1.5-2.5 h.
Further, the drying in the step (5) is carried out for 3-5h at the temperature of-25 to-45 ℃, and then the hot air drying is carried out at the temperature of 60-80 ℃ until the water content is 4.0-5.5%.
Further, the macroporous absorption resin column is HPD-100 type, AB-8 type or X-5 type macroporous absorption resin column.
Further, the eluent consists of ethanol and ethyl acetate in a volume ratio of 1-3: 1.
Further, fermenting for 36-72h at 25-32 ℃ in the step (3).
Compared with the prior art, the invention has the advantages and beneficial effects that:
1. according to the method, citric acid, cellulase and pectinase are adopted for soaking to perform enzymolysis on sauropus spatulifolius, and then mixed strains are used for fermentation and then heating reflux extraction is performed, so that leaching of effective components of total flavonoids in sauropus spatulifolius is facilitated, and the extraction rate of the total flavonoids is improved; d-sodium erythorbate, sodium chloride and nisin are added in the pulping process, so that the medicinal materials are not easy to oxidize, and the stability of the total flavonoids is improved.
2. According to the method, a mixed strain consisting of bacillus marinus, aspergillus niger and candida utilis is adopted to ferment and extract the sauropus spatulifolius extract, the synergistic effect among microorganisms is fully utilized, the dissolution of total flavonoids is facilitated, wherein the bacillus marinus has strong capability of producing amylase, protease and the like, and can degrade macromolecular starch and protein substances; aspergillus niger can secrete cellulose and other decomposed cellulose, so that total flavonoids can be extracted; the candida utilis can decompose protein, pectinase and the like, and the decomposed protein can provide nutrient substances for other microorganisms.
3. The extract obtained by the method is absorbed by a macroporous absorption resin column and then eluted by ethanol and ethyl acetate, so that the purity of the total flavone can be improved, and the method is simple and has low cost.
4. The method extracts the total flavone, and then the total flavone is subjected to freeze drying and hot air drying, so that the effective components of the total flavone are not damaged, the drying is thorough, the operation is simple, and the cost is low.
5. The method of the invention freezes the general flavone firstly and then unfreezes the general flavone by warm water, and the two different temperature differences are utilized, thereby being beneficial to the extraction of the general flavone.
6. The method has the advantages that the extraction amount of the total flavonoids from sauropus spatulifolius is more than 2.8g/kg (calculated on the weight of sauropus spatulifolius), the content of the total flavonoids is more than 80%, and the extracted total flavonoids have good application prospects in the field of medicines.
Detailed Description
The present invention will be described in further detail with reference to specific embodiments. It should be emphasized that the following description is merely exemplary in nature and is not intended to limit the scope of the invention or its application.
Example 1
A method for efficiently extracting total flavonoids from sauropus spatulifolius comprises the following steps:
(1) picking 1kg of sauropus spatholobus stem, freezing the sauropus spatholobus stem for 24 hours at a temperature of-50 ℃, taking out the sauropus spatholobus stem, thawing the sauropus spatholobus stem with warm water at a temperature of 80 ℃, adding a soaking solution consisting of 15g of citric acid, 8g of cellulase and 5g of pectinase, soaking the sauropus spatholobus stem for 4 hours, and taking out the sauropus spatholobus stem;
(2) adding water according to the use amount of the material-liquid ratio of 1:20g/m L, adding 8g of D-sodium erythorbate, 20g of sodium chloride and 4g of nisin, and pulping to obtain a sauropus spatholobus stem extract;
(3) adding 25g of fermentation strain, 50g of brown sugar, 8g of urea, 3g of ammonium sulfate and 2g of potassium dihydrogen phosphate into the sauropus spatulifolius extract, uniformly stirring, and fermenting for 48 hours at the temperature of 30 ℃; the fermentation strain consists of marine bacillus, aspergillus niger and candida utilis in a mass ratio of 3:1: 1;
(4) heating and reflux-extracting fermented folium sauropi sauropus extract at 75 deg.C for 3 times, each time for 2 hr, vacuum-filtering, and concentrating under reduced pressure to obtain concentrated extract;
(5) dissolving the concentrated extract in 5 times of distilled water, centrifuging, and loading the supernatant onto AB-8 type macroporous adsorbent resin column at a loading flow rate of 2.0 BV/h; eluting at an elution speed of 2.0BV/h, eluting with water to colorless and transparent, eluting with ethanol and ethyl acetate eluent at a volume ratio of 2:1, collecting eluent, concentrating under reduced pressure to recover the eluent, drying at-35 ℃ for 4h, and drying with hot air at 70 ℃ until the water content is 4.8%, thereby obtaining 3.429g of the total flavonoids sauropus spathulifolius.
Quantitative analysis is carried out by adopting an HP L C-UV method to obtain the contents of quercetin-3-O- β -D-glucoside, quercetin-3-O- β -D-gentiobioside, kaempferol-3-O- β -D-glucoside and kaempferol-3-O- β -D-gentiobiose which are 43.49%, 37.24%, 1.61% and 1.04% respectively, and the content of total flavonoids is 83.38%.
Example 2
A method for efficiently extracting total flavonoids from sauropus spatulifolius comprises the following steps:
(1) picking 1kg of sauropus spatholobus stem, freezing the sauropus spatholobus stem for 36 hours at a temperature of-40 ℃, taking out the sauropus spatholobus stem, thawing the sauropus spatholobus stem with warm water at a temperature of 80 ℃, adding a soaking solution consisting of 12g of citric acid, 5g of cellulase and 4g of pectinase, soaking the sauropus spatholobus stem for 3 hours, and taking out the sauropus spatholobus stem;
(2) adding water according to the use amount of L according to the material-liquid ratio of 1:20g/m, adding 5g of D-sodium erythorbate, 25g of sodium chloride and 5g of nisin, and pulping to obtain a sauropus spatulifolius extracting solution;
(3) adding 10g of fermentation strain, 50g of brown sugar, 6g of urea, 4g of ammonium sulfate and 2g of potassium dihydrogen phosphate into the sauropus spatulifolius extract, uniformly stirring, and fermenting at the temperature of 28 ℃ for 45 hours; the fermentation strain consists of marine bacillus, aspergillus niger and candida utilis in a mass ratio of 4:2: 2;
(4) heating and reflux-extracting the fermented sauropus spatulifolius extract at 70 deg.C for 3 times (each for 2.5 hr), suction-filtering, and concentrating under reduced pressure to obtain concentrated extract;
(5) dissolving the concentrated extract in 5 times of distilled water, centrifuging, and loading the supernatant onto AB-8 type macroporous adsorbent resin column at a loading flow rate of 3.0 BV/h; eluting at an elution speed of 3.0BV/h, eluting with water to colorless and transparent, eluting with ethanol and ethyl acetate eluent at a volume ratio of 3:1, collecting eluent, concentrating under reduced pressure to recover the eluent, drying at-35 ℃ for 4h, and drying with hot air at 65 ℃ until the water content is 4.4%, thereby obtaining 2.891g of the total flavonoids sauropus spathulifolius.
Quantitative analysis is carried out by adopting an HP L C-UV method to obtain the contents of quercetin-3-O- β -D-glucoside, quercetin-3-O- β -D-gentiobioside, kaempferol-3-O- β -D-glucoside and kaempferol-3-O- β -D-gentiobiose which are respectively 40.67 percent, 36.78 percent, 1.54 percent and 1.19 percent, and the content of total flavonoids is 80.18 percent.
Example 3
A method for efficiently extracting total flavonoids from sauropus spatulifolius comprises the following steps:
(1) picking 1kg of sauropus spatholobus stem, freezing the sauropus spatholobus stem for 48 hours at a temperature of-35 ℃, taking out the sauropus spatholobus stem, thawing the sauropus spatholobus stem with warm water at a temperature of 70 ℃, adding a soaking solution consisting of 20g of citric acid, 6g of cellulase and 4g of pectinase, soaking the sauropus spatholobus stem for 5 hours, and taking out the sauropus spatholobus stem;
(2) adding water according to the use amount of L according to the material-liquid ratio of 1:25g/m, adding 7g D-sodium erythorbate, 20g sodium chloride and 4g nisin, and pulping to obtain sauropus spatholobus stem extract;
(3) adding 20g of fermentation strain, 70g of brown sugar, 8g of urea, 3g of ammonium sulfate and 3g of potassium dihydrogen phosphate into the sauropus spatulifolius extract, uniformly stirring, and fermenting for 60 hours at the temperature of 30 ℃; the fermentation strain consists of marine bacillus, aspergillus niger and candida utilis in a mass ratio of 3:2: 3;
(4) heating and reflux-extracting fermented folium sauropi sauropus extract at 80 deg.C for 3 times (each for 2 hr), vacuum-filtering, and concentrating under reduced pressure to obtain concentrated extract;
(5) dissolving the concentrated extract in 4 times of distilled water, centrifuging, and loading the supernatant onto HPD-100 type macroporous adsorbent resin column at a loading flow rate of 2.0 BV/h; eluting at an elution speed of 2.0BV/h, eluting with water to colorless and transparent, eluting with ethanol and ethyl acetate eluent at a volume ratio of 1:1, collecting eluent, concentrating under reduced pressure to recover the eluent, drying at-40 ℃ for 5h, and drying with hot air at 60 ℃ until the water content is 4.3%, thereby obtaining 3.195g of the total flavonoids sauropus spathulifolius.
Quantitative analysis is carried out by adopting an HP L C-UV method to obtain the contents of quercetin-3-O- β -D-glucoside, quercetin-3-O- β -D-gentiobioside, kaempferol-3-O- β -D-glucoside and kaempferol-3-O- β -D-gentiobiose which are 48.19%, 34.18%, 1.84% and 1.19% respectively, and the content of total flavonoids is 85.40%.
The foregoing is a more detailed description of the invention in connection with specific/preferred embodiments and is not intended to limit the practice of the invention to those descriptions. It will be apparent to those skilled in the art that various substitutions and modifications can be made to the described embodiments without departing from the spirit of the invention, and such substitutions and modifications are to be considered as within the scope of the invention.
Claims (10)
1. A method for efficiently extracting total flavonoids from sauropus spatulifolius is characterized by comprising the following steps: the method comprises the following steps:
(1) picking the sauropus, freezing for 24-48h, taking out, thawing with warm water, adding a soaking solution consisting of citric acid, cellulase and pectinase, soaking for 2-6h, and taking out the sauropus;
(2) adding water according to the use amount of L according to the material-liquid ratio of 1:20-25g/m, adding D-sodium erythorbate, sodium chloride and nisin, and pulping to obtain an extracting solution of sauropus spatholobus stem;
(3) adding fermentation strains, brown sugar, urea, ammonium sulfate and potassium dihydrogen phosphate into the sauropus spatulifolius extract, uniformly stirring, and fermenting; the fermentation strain consists of bacillus marinus, aspergillus niger and candida utilis;
(4) heating and refluxing the fermented sauropus spatholobus stem extract, performing suction filtration, and performing reduced pressure concentration to obtain a concentrated extract;
(5) dissolving the concentrated extract in 4-6 times of distilled water, centrifuging, and loading the supernatant onto macroporous adsorbent resin column at a loading flow rate of 1.0-3.0 BV/h; eluting at an elution speed of 1.0-3.0BV/h, eluting with water until colorless and transparent, eluting with ethanol and ethyl alcohol eluent, collecting the eluent, concentrating under reduced pressure, recovering the eluent, and drying to obtain the sauropus spatulifolius total flavonoids.
2. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: the mass ratio of the sauropus spatulifolius to the citric acid, the cellulase and the pectinase is 100:1-2:0.5-1: 0.3-0.5.
3. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: the mass ratio of the D-sodium erythorbate, the sodium chloride, the nisin to the sauropus spatulifolius is 0.5-1.0:1-3:0.3-0.6: 100.
4. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: the mass ratio of the fermentation strain, brown sugar, urea, ammonium sulfate, potassium dihydrogen phosphate to sauropus spatholobus stem leaf is 1-3:4-8:0.5-1.0:0.3-0.5:0.1-0.3: 100.
5. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 4, wherein the method comprises the following steps: the fermentation strain consists of marine bacillus, aspergillus niger and candida utilis in a mass ratio of 3-5:0.5-2: 1-3.
6. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: the heating reflux extraction is reflux extraction at 70-80 deg.C for 3 times, each time for 1.5-2.5 hr.
7. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: and (5) drying at the temperature of minus 25 to minus 45 ℃ for 3 to 5 hours, and then drying by hot air at the temperature of 60 to 80 ℃ until the water content is 4.0 to 5.5 percent.
8. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: the macroporous absorption resin column is HPD-100 type, AB-8 type or X-5 type macroporous absorption resin column.
9. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: the eluent consists of ethanol and ethyl acetate with the volume ratio of 1-3: 1.
10. The method for efficiently extracting total flavonoids from sauropus spatulifolius according to claim 1, wherein the method comprises the following steps: fermenting for 36-72h at 25-32 ℃ in the fermentation step (3).
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| CN101248896A (en) * | 2008-03-31 | 2008-08-27 | 北京市科威华食品工程技术有限公司 | Micro-fermentation haw thorn Chinese wolfberry fruit drink and method of preparing the same |
| CN101358219A (en) * | 2008-07-14 | 2009-02-04 | 天津农学院 | Simultaneous sequential chemical extraction of jujube flavones and jujube polysaccharide by fermentation method |
| CN101715998A (en) * | 2009-12-17 | 2010-06-02 | 李振声 | Production method of primary pulp of fresh jujubes |
| CN106674313A (en) * | 2016-12-30 | 2017-05-17 | 成都普思生物科技股份有限公司 | Method for simultaneously separating quercetin-3-O-gentian diglucoside and kaempferol-3-O-gentian diglucoside from folium sauropi |
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2020
- 2020-03-31 CN CN202010243426.5A patent/CN111481583A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101248896A (en) * | 2008-03-31 | 2008-08-27 | 北京市科威华食品工程技术有限公司 | Micro-fermentation haw thorn Chinese wolfberry fruit drink and method of preparing the same |
| CN101358219A (en) * | 2008-07-14 | 2009-02-04 | 天津农学院 | Simultaneous sequential chemical extraction of jujube flavones and jujube polysaccharide by fermentation method |
| CN101715998A (en) * | 2009-12-17 | 2010-06-02 | 李振声 | Production method of primary pulp of fresh jujubes |
| CN106674313A (en) * | 2016-12-30 | 2017-05-17 | 成都普思生物科技股份有限公司 | Method for simultaneously separating quercetin-3-O-gentian diglucoside and kaempferol-3-O-gentian diglucoside from folium sauropi |
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