CN111606794A - Paeonia suffruticosa root-bark extract as well as preparation method and application thereof - Google Patents
Paeonia suffruticosa root-bark extract as well as preparation method and application thereof Download PDFInfo
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- CN111606794A CN111606794A CN202010476367.6A CN202010476367A CN111606794A CN 111606794 A CN111606794 A CN 111606794A CN 202010476367 A CN202010476367 A CN 202010476367A CN 111606794 A CN111606794 A CN 111606794A
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- 229940084070 paeonia suffruticosa root bark extract Drugs 0.000 title claims abstract description 26
- 238000002360 preparation method Methods 0.000 title claims abstract description 17
- UILPJVPSNHJFIK-UHFFFAOYSA-N Paeonol Chemical compound COC1=CC=C(C(C)=O)C(O)=C1 UILPJVPSNHJFIK-UHFFFAOYSA-N 0.000 claims abstract description 162
- YLTGFGDODHXMFB-UHFFFAOYSA-N isoacetovanillon Natural products COC1=CC=C(C(C)=O)C=C1O YLTGFGDODHXMFB-UHFFFAOYSA-N 0.000 claims abstract description 81
- MLIBGOFSXXWRIY-UHFFFAOYSA-N paeonol Natural products COC1=CC=C(O)C(C(C)=O)=C1 MLIBGOFSXXWRIY-UHFFFAOYSA-N 0.000 claims abstract description 81
- 239000000284 extract Substances 0.000 claims abstract description 37
- 238000000605 extraction Methods 0.000 claims abstract description 30
- 238000000034 method Methods 0.000 claims abstract description 30
- 240000005001 Paeonia suffruticosa Species 0.000 claims abstract description 18
- 235000003889 Paeonia suffruticosa Nutrition 0.000 claims abstract description 18
- 239000000287 crude extract Substances 0.000 claims abstract description 15
- 239000003814 drug Substances 0.000 claims abstract description 9
- 239000002537 cosmetic Substances 0.000 claims abstract description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 110
- 239000002994 raw material Substances 0.000 claims description 53
- 229920001661 Chitosan Polymers 0.000 claims description 47
- 239000002244 precipitate Substances 0.000 claims description 32
- 238000001914 filtration Methods 0.000 claims description 26
- 238000003756 stirring Methods 0.000 claims description 26
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 23
- 102000004190 Enzymes Human genes 0.000 claims description 18
- 108090000790 Enzymes Proteins 0.000 claims description 18
- 229940088598 enzyme Drugs 0.000 claims description 18
- 241000736199 Paeonia Species 0.000 claims description 15
- 238000006243 chemical reaction Methods 0.000 claims description 15
- 235000006484 Paeonia officinalis Nutrition 0.000 claims description 12
- 239000000706 filtrate Substances 0.000 claims description 11
- 238000009210 therapy by ultrasound Methods 0.000 claims description 11
- 239000004382 Amylase Substances 0.000 claims description 8
- 108010065511 Amylases Proteins 0.000 claims description 8
- 102000013142 Amylases Human genes 0.000 claims description 8
- 108090000145 Bacillolysin Proteins 0.000 claims description 8
- 108010059892 Cellulase Proteins 0.000 claims description 8
- 102000035092 Neutral proteases Human genes 0.000 claims description 8
- 108091005507 Neutral proteases Proteins 0.000 claims description 8
- 108010059820 Polygalacturonase Proteins 0.000 claims description 8
- 235000019418 amylase Nutrition 0.000 claims description 8
- 229940106157 cellulase Drugs 0.000 claims description 8
- 108010093305 exopolygalacturonase Proteins 0.000 claims description 8
- 239000007791 liquid phase Substances 0.000 claims description 7
- 230000002378 acidificating effect Effects 0.000 claims description 4
- 238000004140 cleaning Methods 0.000 claims description 4
- 238000001035 drying Methods 0.000 claims description 4
- 238000002156 mixing Methods 0.000 claims description 4
- 230000007935 neutral effect Effects 0.000 claims description 3
- 238000007873 sieving Methods 0.000 claims description 3
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 claims description 2
- 239000000203 mixture Substances 0.000 claims 1
- 238000000746 purification Methods 0.000 abstract description 12
- 229940079593 drug Drugs 0.000 abstract description 5
- 238000000265 homogenisation Methods 0.000 abstract description 5
- 238000000926 separation method Methods 0.000 abstract description 3
- 230000000052 comparative effect Effects 0.000 description 38
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 18
- 239000000047 product Substances 0.000 description 16
- 239000000243 solution Substances 0.000 description 16
- 238000001514 detection method Methods 0.000 description 13
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 10
- 230000000694 effects Effects 0.000 description 7
- 238000001179 sorption measurement Methods 0.000 description 7
- 239000000126 substance Substances 0.000 description 6
- 238000005303 weighing Methods 0.000 description 6
- 239000012535 impurity Substances 0.000 description 5
- 239000008213 purified water Substances 0.000 description 5
- 238000004108 freeze drying Methods 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- 238000001704 evaporation Methods 0.000 description 3
- 238000004128 high performance liquid chromatography Methods 0.000 description 3
- 238000001556 precipitation Methods 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 238000000527 sonication Methods 0.000 description 3
- 238000001256 steam distillation Methods 0.000 description 3
- 241000196324 Embryophyta Species 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 2
- 208000003251 Pruritus Diseases 0.000 description 2
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 230000009286 beneficial effect Effects 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 230000003467 diminishing effect Effects 0.000 description 2
- 230000004054 inflammatory process Effects 0.000 description 2
- 230000007803 itching Effects 0.000 description 2
- 229920002521 macromolecule Polymers 0.000 description 2
- 239000002075 main ingredient Substances 0.000 description 2
- 238000010979 pH adjustment Methods 0.000 description 2
- 239000000049 pigment Substances 0.000 description 2
- 238000001953 recrystallisation Methods 0.000 description 2
- 239000013558 reference substance Substances 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241000124495 Paeonia delavayi Species 0.000 description 1
- 241001106477 Paeoniaceae Species 0.000 description 1
- YKRGDOXKVOZESV-WRJNSLSBSA-N Paeoniflorin Chemical compound C([C@]12[C@H]3O[C@]4(O)C[C@](O3)([C@]1(C[C@@H]42)O[C@H]1[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O1)O)C)OC(=O)C1=CC=CC=C1 YKRGDOXKVOZESV-WRJNSLSBSA-N 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 239000004480 active ingredient Substances 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 238000000975 co-precipitation Methods 0.000 description 1
- 238000004440 column chromatography Methods 0.000 description 1
- 238000001816 cooling Methods 0.000 description 1
- 238000005520 cutting process Methods 0.000 description 1
- 230000006196 deacetylation Effects 0.000 description 1
- 238000003381 deacetylation reaction Methods 0.000 description 1
- 238000004821 distillation Methods 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 230000002255 enzymatic effect Effects 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 239000002024 ethyl acetate extract Substances 0.000 description 1
- 238000003810 ethyl acetate extraction Methods 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 239000000945 filler Substances 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 239000002932 luster Substances 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- GBMDVOWEEQVZKZ-UHFFFAOYSA-N methanol;hydrate Chemical compound O.OC GBMDVOWEEQVZKZ-UHFFFAOYSA-N 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- YTJSFYQNRXLOIC-UHFFFAOYSA-N octadecylsilane Chemical compound CCCCCCCCCCCCCCCCCC[SiH3] YTJSFYQNRXLOIC-UHFFFAOYSA-N 0.000 description 1
- YKRGDOXKVOZESV-UHFFFAOYSA-N paeoniflorin Natural products O1C(C)(C2(CC34)OC5C(C(O)C(O)C(CO)O5)O)CC3(O)OC1C24COC(=O)C1=CC=CC=C1 YKRGDOXKVOZESV-UHFFFAOYSA-N 0.000 description 1
- 239000001814 pectin Substances 0.000 description 1
- 229920001277 pectin Polymers 0.000 description 1
- 235000010987 pectin Nutrition 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 239000012088 reference solution Substances 0.000 description 1
- 239000000377 silicon dioxide Substances 0.000 description 1
- 238000002791 soaking Methods 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000012085 test solution Substances 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 238000002137 ultrasound extraction Methods 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
- C07C45/00—Preparation of compounds having >C = O groups bound only to carbon or hydrogen atoms; Preparation of chelates of such compounds
- C07C45/78—Separation; Purification; Stabilisation; Use of additives
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/33—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing oxygen
- A61K8/35—Ketones, e.g. benzophenone
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P17/00—Drugs for dermatological disorders
- A61P17/04—Antipruritics
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
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- Health & Medical Sciences (AREA)
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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Pharmacology & Pharmacy (AREA)
- Medicinal Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Dermatology (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Emergency Medicine (AREA)
- Birds (AREA)
- Epidemiology (AREA)
- Pain & Pain Management (AREA)
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- Medicines Containing Plant Substances (AREA)
- Cosmetics (AREA)
Abstract
The invention relates to the field of biological medicines, relates to a traditional Chinese medicine extract, and a preparation method and application thereof, and particularly relates to a paeonia suffruticosa root-bark extract, and a preparation method and application thereof. In the process of preparing the paeonia suffruticosa root-bark extract, firstly, carrying out low-temperature homogenization treatment on the paeonia suffruticosa root-bark, then, using ultrasonic to assist in extraction, and then, carrying out separation and purification on target components on a crude extract. The preparation method can be used for extracting and separating the paeonia suffruticosa root-bark extract with high paeonol purity from paeonia suffruticosa, and has the advantages of high extraction rate and simple extraction process. The preparation method and the paeonia suffruticosa root-bark extract can be applied to the preparation and development of related cosmetics and medicines.
Description
Technical Field
The invention relates to the field of biological medicines, relates to a traditional Chinese medicine extract, and a preparation method and application thereof, and particularly relates to a paeonia suffruticosa root-bark extract, and a preparation method and application thereof.
Background
Paeonia yunnanensis (Paeonia delavayi) is a plant of Paeonia suffruticosa group of Paeonia of Paeoniaceae, not only has strong ornamental value, but also can be used as medicine for the whole body, and has ornamental value and medicinal value. The root bark of peony group plant is called moutan bark (cortex moutan), which has been used as a Chinese medicinal material for thousands of years. The cortex moutan contains paeonol and paeoniflorin as main ingredients, and has effects of dredging channels, cooling blood, removing blood stasis, relieving pain, diminishing inflammation and relieving itching.
In the prior art, the extraction of paeonol from the root bark of the paeonia suffruticosa is usually carried out by a distillation method, but the stability of the paeonol is influenced by overhigh extraction temperature (the paeonol is easy to decompose under the high-temperature condition). The paeonol obtained by extraction can be used for obtaining an extract product with high purity and good color only by separation and purification steps, the method for separating and purifying the paeonol in the prior art mainly comprises multiple column chromatography purification or recrystallization, the operation steps are more, the consumed time is long, the efficiency is low, the loss of target components can be caused by the increase of the operation steps, and the extraction rate of the paeonol is low.
Disclosure of Invention
The invention mainly solves the technical problem of providing a preparation method of the paeonia suffruticosa root-bark extract, the paeonia suffruticosa root-bark extract with high paeonol purity can be extracted and separated from paeonia suffruticosa by the preparation method, and the preparation method has the advantages of high extraction rate and simple extraction process.
In order to achieve the purpose, the invention provides the following technical scheme:
a preparation method of a Paeonia suffruticosa root-bark extract comprises the following steps,
step (1): pretreating to obtain cortex moutan raw material;
step (2): firstly, adding water into the cortex moutan raw material in the step (1), then homogenizing the cortex moutan raw material, and then filtering to obtain filter residue;
and (3): adding ethyl acetate into the filter residue obtained in the step (2) to obtain an extraction system; carrying out ultrasonic treatment on the extraction system, then filtering to obtain filtrate, and concentrating the filtrate to obtain a crude cortex moutan extract;
and (4): dispersing the crude extract of cortex moutan in water, adding complex enzyme to obtain an enzymolysis system, and carrying out enzymolysis reaction on the enzymolysis system; after the enzymolysis reaction is finished, adjusting the pH value to 5.0-5.5, adding chitosan, stirring and uniformly mixing to obtain a system A, standing, adjusting the pH value of the system A to 7.0-8.0, standing, filtering and taking precipitate; dispersing the precipitate in ethyl acetate, standing, filtering to obtain liquid phase part, concentrating the extractive solution, and drying to obtain cortex moutan extract.
By adopting the technical scheme, the technical principle is as follows: the method is characterized in that the cortex moutan raw material is homogenized firstly, so that the cell structure of the cortex moutan raw material is initially damaged, and the cells absorb water and swell, thereby being beneficial to the subsequent extraction step. And then, extracting paeonol in the cortex moutan raw material by using ethyl acetate as a solvent under the assistance of ultrasound. Under the action of ultrasonic wave, the cell structure of the raw material of cortex moutan is fully destroyed, paeonol is dissolved out by ethyl acetate, and crude extract of cortex moutan is obtained after concentration treatment. After the steps are completed, the crude extract of the cortex moutan needs to be purified to remove impurities such as pigments, and otherwise, the paeonol extract with high purity and good color cannot be obtained. Dispersing the crude extract of cortex moutan in water, and using complex enzyme to fully degrade substances such as fiber, pectin, protein and the like in the crude extract of cortex moutan to remove interfering substances. And then, adjusting the pH value of the system to be acidic, wherein the chitosan can be dissolved under the acidic condition, and the dissolved chitosan can be adsorbed with paeonol. And then, adjusting the pH value of the system to be alkaline, wherein the solubility of the chitosan is reduced under alkaline or neutral conditions, and precipitation begins to be separated out, and due to the adsorption effect of the chitosan on the paeonol, the paeonol and the chitosan are co-precipitated, so that the paeonol is separated from other soluble substances in the system. Collecting the precipitate, dispersing the precipitate in ethyl acetate, extracting paeonol with ethyl acetate, filtering to obtain liquid phase, concentrating, and drying to obtain cortex moutan extract containing paeonol as main ingredient.
The beneficial effects of the technical scheme are as follows:
(1) the method for extracting and purifying paeonol can obtain high-purity paeonol with good color and luster, and the yield of the paeonol is higher.
(2) The paeonol is purified by utilizing the adsorption effect of the chitosan on the paeonol and the properties of the chitosan such as solubility in an acidic environment and poor solubility in an alkaline neutral environment, and the paeonol extract with higher purity is obtained by a simple purification method, so that the complex steps such as column-passing purification, multiple extraction, recrystallization and the like are avoided. The inventor firstly finds the adsorption effect of chitosan on paeonol and applies the adsorption effect to the extraction and purification of paeonol.
(3) The low temperature homogenization process protects paeonol (poor thermal stability) which is easily volatilized with water vapor. Part of water-soluble substances are also taken away in the homogenizing process, so that the interference of a large amount of water-soluble substances on paeonol extraction is avoided, and the purity of the product is increased.
(4) The whole extraction process does not need high-temperature treatment, and the paeonol is ensured not to be degraded and volatilized, so that the quality of the extract is improved.
(5) Before the chitosan is used for purifying the paeonol, a complex enzyme enzymolysis method is used, so that macromolecular interferents which can form coprecipitation with the chitosan are sufficiently removed, and the content of the paeonol in the finally obtained extract is further improved.
Further, in the step (4), the addition amount of the chitosan is 10-20% of the weight of the cortex moutan raw material.
By adopting the technical scheme, the paeonol can be fully adsorbed by the chitosan with the dosage, and the paeonol is effectively purified from impurities.
Further, in the step (4), the compound enzyme comprises cellulase, neutral protease, amylase and pectinase, and the content of the compound enzyme in an enzymolysis system is 2-4 wt%; the temperature of the enzymolysis reaction is 30-45 ℃, the duration is 2-3h, and the pH value is 7.0-7.5.
By adopting the technical scheme, the complex enzyme with the combination and the dosage and the condition of enzymolysis reaction are adopted, so that macromolecular impurities such as protein, starch and the like in the system can be fully removed, and the influence of the macromolecular substances on the adsorption of paeonol by chitosan is reduced.
Further, in the step (4), after the enzymolysis reaction is finished, adjusting the pH value to 5.0-5.5, adding chitosan, stirring and uniformly mixing to obtain a system A, standing, adjusting the pH value of the system A to 7.0-8.0, standing, filtering and taking the precipitate.
By adopting the technical scheme, after the enzymolysis reaction is finished, the pH is adjusted to 5.0-5.5, so that the chitosan cannot be greatly degraded due to over strong acidity, and cannot be well dissolved due to over weak acidity, and the paeonol cannot be effectively adsorbed. The pH value of the system A is adjusted to 7.0-8.0 after standing, so that chitosan is precipitated, if the pH value is too low, the precipitate cannot be fully separated out, paeonol cannot be fully separated from the system, the pH value is too high, the precipitation separation speed is too high, the obtained precipitate is too compact, the precipitate cannot be effectively dispersed in the subsequent ethyl acetate extraction process, the ethyl acetate cannot be fully contacted with the paeonol, the paeonol cannot be fully extracted, and a large amount of paeonol remains in the precipitate, so that the yield of the paeonol is too low.
Further, in the step (4), ethyl acetate 6-9 times of the weight of the cortex moutan raw material is added into the precipitate.
By adopting the technical scheme, the paeonol in the precipitate can be fully extracted by using the ethyl acetate with the dosage.
Further, in the step (3), the ultrasonic power for performing ultrasonic treatment on the extraction system is 100-200W, and the treatment time is 30 min; and adding ethyl acetate 4-7 times of the weight of the cortex moutan raw material into the filter residue.
By adopting the technical scheme, the cell structure can be damaged by ultrasonic treatment of the parameters, so that the effective components are fully dissolved out.
Further, in the step (2), 0-10 ℃ water which is 1-3 times of the weight of the cortex moutan raw material is added into the cortex moutan raw material.
By adopting the technical scheme, the homogenization process is ensured to be carried out at a low temperature, the cortex moutan is prevented from being degraded or volatilized, and the extraction rate of paeonol is improved.
Further, in the step (1), the cortex moutan raw material is prepared by the following method: taking fresh Yunnan peony root bark, cleaning, crushing the fresh Yunnan peony root bark, and then sieving by a 10-mesh sieve to obtain the cortex moutan raw material.
By adopting the technical scheme, the fresh paeonia suffruticosa root bark is plump in cells, the cell structure is easy to damage, and the active ingredients can be efficiently dissolved out.
Furthermore, the content of paeonol in the extract of the root and bark of the paeonia suffruticosa is more than 90 percent.
By adopting the technical scheme, the paeonia suffruticosa root-bark extract obtained by the method has the characteristics of low impurity content and high paeonol purity.
Further, the extract of the root and bark of the paeonia suffruticosa is applied to cosmetics or medicines.
By adopting the technical scheme, the main component of the paeonia suffruticosa root-bark extract prepared by the scheme is paeonol, has the effects of diminishing inflammation and relieving itching, and can be applied to the preparation of cosmetics and medicines.
Detailed Description
Example 1: the technical scheme is adopted to prepare the extract of the root and bark of the paeonia suffruticosa
1. Process for preparing extract
Step (1): taking fresh Yunnan peony root bark (medicinal material cortex moutan), cleaning, crushing the fresh Yunnan peony root bark, and sieving with a 10-mesh sieve to obtain the cortex moutan raw material.
Step (2): putting 100g of cortex moutan raw material into a homogenizer, adding purified water with 4 ℃ into the cortex moutan raw material, wherein the adding amount of the purified water is 3 times of the weight of the cortex moutan raw material (namely 300g of purified water), homogenizing in the homogenizer for 30min (the rotation speed is 20000rpm, the temperature is 4 ℃) to obtain a homogenate product, and filtering the homogenate product to obtain a filter residue part.
And (3): and adding ethyl acetate into the filter residue, and uniformly stirring and mixing, wherein the adding amount of the ethyl acetate is 6 times of the weight of the cortex moutan raw material (namely 600g of ethyl acetate). And carrying out ice-bath ultrasonic treatment on the ethyl acetate containing the filter residue of the homogenate product, wherein the ultrasonic power is 150W. The number of sonications was 3 times, each for 10 min. Filtering after ultrasonic treatment to obtain filtrate, and concentrating the filtrate under reduced pressure (pressure of 0.05MPa and temperature of 50 deg.C) with rotary evaporator to obtain cortex moutan crude extract (extract state, density of 0.5 g/ml).
And (4): dispersing the crude extract of cortex moutan in water, wherein the addition amount of water is 6 times of the weight of the cortex moutan raw material. And adding a complex enzyme to obtain an enzymolysis system. The complex enzyme comprises pectinase (CAS: 9032-75-1), cellulase (CAS: 9012-54-8), neutral protease (CAS: 9068-59-1) and amylase (CAS: 9000-90-2). Wherein the content of the complex enzyme in the enzymolysis system is 3 weight percent, and the mass ratio of the pectinase, the cellulase, the neutral protease and the amylase is 1:1:1: 1. The enzymolysis reaction is carried out for 3 hours in total, the temperature is 35 ℃, and the pH value is 7.0. After the enzymolysis reaction is finished, 1 mass percent hydrochloric acid solution is used for adjusting the pH value to 5.0, chitosan (CAS: 9012-76-4, the deacetylation degree is more than or equal to 80 percent, and the viscosity is less than 200mPa.s) is added, the adding amount of the chitosan is 15 percent of the weight of the cortex moutan raw material (namely 7.5g of the chitosan is added), and a system A is obtained. Fully stirring the system A for 30min at the stirring speed of 200rpm, standing for 30min after stirring is finished, then adjusting the pH value of the system A to 7.5 by using 1 mass percent sodium hydroxide solution, standing for 30min, and filtering to obtain a precipitate. And dispersing the precipitate in ethyl acetate, wherein the addition amount of the ethyl acetate is 8 times of the weight of the cortex moutan raw material, and fully stirring for 60min to ensure that the precipitate and the ethyl acetate are completely mixed, wherein the stirring speed is 200 rpm. And then standing for 30min, filtering to obtain a liquid phase part, concentrating the extracting solution to dryness, concentrating the extracting solution under reduced pressure by using a rotary evaporator (the pressure is 0.05MPa, the temperature is 50 ℃) until the concentration of the extract is 0.5g/ml, then freeze-drying the product of the concentration under reduced pressure to obtain the extract of the root bark of the paeonia suffruticosa, weighing (1.137g) and storing for later use.
2. Paeonol yield and purity detection
According to the determination method of the content of the cortex moutan radicis medicinal material in the Chinese pharmacopoeia (first section) (2015 edition), the fresh cortex moutan radicis (cortex moutan radicis) used in the embodiment is subjected to paeonol content detection, and the content of paeonol in the cortex moutan radicis (cortex moutan radicis) of the cortex moutan radicis 1.25 percent through detection.
Measuring the content of paeonol in the extract product according to "high performance liquid chromatography standard operating procedure", and collecting 0.01g of root bark extract to prepare a sample to be detected. Taking appropriate amount of paeonol reference substance, precisely weighing, and adding methanol to obtain solution containing 20 μ g of paeonol per 1ml to obtain reference substance solution. The specific parameters of the detection process are as follows: methanol-water (45: 55) is used as a mobile phase; the detection wavelength is 274nm, and HPLC detection is carried out by using octadecylsilane chemically bonded silica as a filler. During the measurement, respectively and precisely sucking 10 μ l of each of the reference solution and the test solution, injecting into a liquid chromatograph, and measuring to obtain the content of paeonol in the sample. According to the HPLC detection result, the following calculation results are obtained: the extraction rate of paeonol prepared from the extract of the root and bark of the paeonia suffruticosa is 88.32%, and the purity of the paeonol is 97.11%.
Example 2:
this embodiment is basically the same as embodiment 1, except that in step (4), the following is specifically mentioned:
and (4): dispersing the crude extract of cortex moutan in water, wherein the addition amount of water is 6 times of the weight of the cortex moutan raw material. And adding a complex enzyme to obtain an enzymolysis system. The complex enzyme comprises pectinase, cellulase, neutral protease and amylase. Wherein the content of the complex enzyme in the enzymolysis system is 4 weight percent, and the mass ratio of the pectinase, the cellulase, the neutral protease and the amylase is 1:1:1: 1. The enzymolysis reaction is carried out for 2 hours in total, the temperature is 30 ℃, and the pH value is 7.0. After the enzymolysis reaction is finished, 1 mass percent hydrochloric acid solution is used for adjusting the pH value to 5.0, and chitosan is added, wherein the adding amount of the chitosan is 20 percent of the weight of the cortex moutan raw material, so as to obtain a system A. Fully stirring the system A for 30min at the stirring speed of 200rpm, standing for 30min after stirring is finished, then adjusting the pH value of the system A to 7.0 by using 1 mass percent sodium hydroxide solution, standing for 30min, and filtering to obtain a precipitate. And dispersing the precipitate in ethyl acetate, wherein the addition amount of the ethyl acetate is 6 times of the weight of the cortex moutan raw material, and fully stirring for 60min to ensure that the precipitate and the ethyl acetate are completely mixed, wherein the stirring speed is 200 rpm. Standing for 30min, filtering to obtain liquid phase part, concentrating the extractive solution, evaporating, concentrating under reduced pressure (pressure of 0.05MPa and temperature of 50 deg.C) with rotary evaporator until the extract concentration is 0.5g/ml, freeze drying the product to obtain cortex moutan extract, weighing, and storing.
Example 3:
this embodiment is basically the same as embodiment 1, except that in step (4), the following is specifically mentioned:
and (4): dispersing the crude extract of cortex moutan in water, wherein the addition amount of water is 6 times of the weight of the cortex moutan raw material. And adding a complex enzyme to obtain an enzymolysis system. The complex enzyme comprises pectinase, cellulase, neutral protease and amylase (CAS: 9000-90-2). Wherein the content of the complex enzyme in the enzymolysis system is 2 weight percent, and the mass ratio of the pectinase, the cellulase, the neutral protease and the amylase is 1:1:1: 1. The enzymolysis reaction is carried out for 3 hours in total, the temperature is 45 ℃, and the pH value is 7.5. After the enzymolysis reaction is finished, 1 mass percent hydrochloric acid solution is used for adjusting the pH value to 5.5, and chitosan is added, wherein the adding amount of the chitosan is 10 percent of the weight of the cortex moutan raw material (namely 7.5g of chitosan is added), so as to obtain a system A. Fully stirring the system A for 30min at the stirring speed of 200rpm, standing for 30min after stirring is finished, then adjusting the pH value of the system A to 8.0 by using 1 mass percent sodium hydroxide solution, standing for 30min, and filtering to obtain a precipitate. And dispersing the precipitate in ethyl acetate, wherein the addition amount of the ethyl acetate is 9 times of the weight of the cortex moutan raw material, and fully stirring for 60min to ensure that the precipitate and the ethyl acetate are completely mixed, wherein the stirring speed is 200 rpm. Standing for 30min, filtering to obtain liquid phase part, concentrating the extractive solution, evaporating, concentrating under reduced pressure (pressure of 0.05MPa and temperature of 50 deg.C) with rotary evaporator until the extract concentration is 0.5g/ml, freeze drying the product to obtain cortex moutan extract, weighing, and storing.
Example 4:
this example is basically the same as example 1, except that step (2) and step (3):
step (2): putting 100g of cortex moutan raw material into a homogenizer, adding 10 deg.C purified water 1 times of cortex moutan raw material, homogenizing in the homogenizer for 30min (rotation speed of 20000rpm, temperature of 4 deg.C) to obtain homogenate product, filtering the homogenate product, and collecting residue.
And (3): and adding ethyl acetate into the filter residue, and uniformly stirring, wherein the adding amount of the ethyl acetate is 4 times of the weight of the cortex moutan raw material. And carrying out ice-bath ultrasonic treatment on the ethyl acetate containing the filter residue of the homogenate product, wherein the ultrasonic power is 200W. The number of sonications was 3 times, each for 15 min. Filtering after ultrasonic treatment to obtain filtrate, and concentrating the filtrate under reduced pressure (pressure of 0.05MPa and temperature of 50 deg.C) with rotary evaporator to obtain cortex moutan crude extract (extract state, density of 0.5 g/ml).
Example 5:
this example is basically the same as example 1, except that step (2) and step (3):
step (2): putting 100g of cortex moutan raw material into a homogenizer, adding 0 deg.C purified water 2 times of cortex moutan raw material, homogenizing in the homogenizer for 30min (rotation speed of 20000rpm, temperature of 4 deg.C) to obtain homogenate product, filtering the homogenate product, and collecting residue.
And (3): and adding ethyl acetate into the filter residue, and uniformly stirring, wherein the adding amount of the ethyl acetate is 7 times of the weight of the cortex moutan raw material. And carrying out ice-bath ultrasonic treatment on the ethyl acetate containing the filter residue of the homogenate product, wherein the ultrasonic power is 100W. The number of sonications was 2 times, each for 25 min. Filtering after ultrasonic treatment to obtain filtrate, and concentrating the filtrate under reduced pressure (pressure of 0.05MPa and temperature of 50 deg.C) with rotary evaporator to obtain cortex moutan crude extract (extract state, density of 0.5 g/ml).
Comparative example 1: preparation of the extract of the root bark of the paeonia suffruticosa by adopting a direct steam distillation method
Taking fresh Yunnan peony root bark (medicinal material cortex moutan), cleaning, and cutting the fresh Yunnan peony root bark into 2cm small sections to obtain the cortex moutan raw material. Taking 100g of cortex moutan raw material, adding 6 times of distilled water, soaking at 40 ℃ for 2h, adding 20ml of absolute ethyl alcohol, carrying out steam distillation, continuously collecting secondary distillate, wherein each distillate is 250ml, refrigerating the distillate overnight, filtering, collecting filtrate, concentrating and drying at 40 ℃ to obtain the paeonia suffruticosa root-bark extract, weighing and storing for later use. The paeonol yield and purity detection is carried out according to the method in the example 1, the paeonol extraction rate of the paeonia suffruticosa root-bark extract prepared in the current time is 78.35%, the paeonol purity is 72.14%, and the paeonia suffruticosa root-bark extract prepared in the comparative example is light yellow in color.
Comparative example 2:
this comparative example is substantially the same as example 1, except that in this example, only the steps (1) to (3) were carried out, and each examination was carried out using the crude extract of cortex moutan obtained.
Comparative example 3:
this comparative example is substantially the same as example 1, except that in step (4), no enzymatic treatment is used, as follows:
and (4): dispersing the crude extract of cortex moutan in water, wherein the addition amount of water is 6 times of the weight of the cortex moutan raw material. And (3) adjusting the pH value to 5.0 by using 1 mass percent of hydrochloric acid solution, and adding chitosan, wherein the adding amount of the chitosan is 15 percent of the weight of the cortex moutan raw material, so as to obtain a system A. Fully stirring the system A for 30min at the stirring speed of 200rpm, standing for 30min after stirring is finished, then adjusting the pH value of the system A to 7.5 by using 1 mass percent sodium hydroxide solution, standing for 30min, and filtering to obtain a precipitate. And dispersing the precipitate in ethyl acetate, wherein the addition amount of the ethyl acetate is 8 times of the weight of the cortex moutan raw material, and fully stirring for 60min to ensure that the precipitate and the ethyl acetate are completely mixed, wherein the stirring speed is 200 rpm. Standing for 30min, filtering to obtain liquid phase part, concentrating the extractive solution, evaporating, concentrating under reduced pressure (pressure of 0.05MPa and temperature of 50 deg.C) with rotary evaporator until the extract concentration is 0.5g/ml, freeze drying the product to obtain cortex moutan extract, weighing, and storing.
Comparative example 4:
this comparative example is substantially the same as example 1 except that in step (4), after the completion of the enzymatic reaction, the pH was adjusted to 6.5 before adding chitosan.
Comparative example 5:
this comparative example is substantially the same as example 1 except that, in step (4), after the completion of the enzymatic hydrolysis reaction, the pH was adjusted to 4.0 using hydrochloric acid before adding chitosan.
Comparative example 6:
this comparative example is substantially the same as example 1 except that, in step (4), pH adjustment using sodium hydroxide after addition of chitosan was carried out to a pH of 10.0.
Comparative example 7:
this comparative example is substantially the same as example 1 except that in step (4), the pH was adjusted to 6.5 with a dilute acid after the addition of chitosan.
Comparative example 8:
this comparative example is substantially the same as example 1 except that in step (2), the temperature of the homogenate was room temperature.
Comparative example 9:
this comparative example is substantially the same as example 1, except that step (2) is not included, and the cortex moutan raw material is directly used for the ultrasonic extraction in step (3).
The extract of the root bark of Paeonia yunnanensis obtained in example 1-example 5, comparative example 1-comparative example 9 was examined as described in example 1, and the results are shown in Table 1. All examples and comparative examples used the same batch of fresh Yunnan peony root bark, so the content of paeonol in the raw material was not measured repeatedly, and the content of paeonol in the fresh Yunnan peony root bark was 1.25%.
Table 1: the results of the paeonol content detection of examples 1 to 5, and comparative examples 1 to 9.
| A(g) | B(g) | C(g) | Extraction ratio (%) | Purity (%) | |
| Example 1 | 1.104 | 1.250 | 1.137 | 88.32 | 97.11 |
| Example 2 | 1.056 | 1.250 | 1.104 | 84.32 | 95.65 |
| Example 3 | 1.069 | 1.250 | 1.110 | 85.52 | 96.31 |
| Example 4 | 1.007 | 1.250 | 1.057 | 80.56 | 95.27 |
| Example 5 | 1.011 | 1.250 | 1.073 | 80.88 | 94.22 |
| Comparative example 1 | 0.979 | 1.250 | 3.357 | 78.32 | 29.16 |
| Comparative example 2 | 1.167 | 1.250 | 3.161 | 93.36 | 36.92 |
| Comparative example 3 | 0.849 | 1.250 | 2.361 | 67.92 | 35.96 |
| Comparative example 4 | N/A | 1.250 | N/A | N/A | N/A |
| Comparative example 5 | N/A | 1.250 | N/A | N/A | N/A |
| Comparative example 6 | N/A | 1.250 | N/A | N/A | N/A |
| Comparative example 7 | N/A | 1.250 | N/A | N/A | N/A |
| Comparative example 8 | 0.758 | 1.250 | 1.423 | 60.64 | 53.27 |
| Comparative example 9 | 0.614 | 1.250 | 1.010 | 49.12 | 60.79 |
In table 1, a represents the mass of paeonol in the extract of the root bark of paeonia suffruticosa every 100g of raw material of the root bark of paeonia suffruticosa; b represents the mass of paeonol in every 100g of cortex moutan raw material (namely fresh Yunnan peony root bark); c represents the quality of the extract of the root bark of the paeonia suffruticosa which is obtained by extracting 100g of the raw material of the root bark of the moutan. Wherein, the calculation mode is as follows: extraction yield (%) - (a/B) × 100%; purity (%) × (a/C) × 100%.
In examples 1 to 5, the paeonol has high extraction rate, the purity of more than 90 percent, white color, fully removed pigment and good quality of the extract of the root bark of the paeonia suffruticosa (the main component is the paeonol). Comparative example 1 used conventional steam distillation, which used high temperature and no purification, resulting in low extraction rate and purity of paeonol, and paeonol was unstable in nature and easily volatilized and decomposed. Comparative example 2 does not undergo a step of chitosan purification, so the purity of paeonol of the paeonia suffruticosa root bark extract is low, and the extraction rate of the paeonol is high but the impurity content is also high because no additional purification step is provided and the paeonol is not lost. The comparison document 3 does not use an enzymolysis method before using chitosan for purification, a large number of macromolecular substances in the system are not degraded, substances such as protein, cellulose and the like interact with chitosan, and the chitosan forms inclusion on the molecules, so that the adsorption efficiency of the chitosan on paeonol is reduced, and the purity is very low. Because the paeonol is subjected to a certain loss after the purification step, the extraction rate is relatively reduced. In comparative example 4, the pH was adjusted to 6.5 before adding chitosan, and in the above pH environment, chitosan could not be sufficiently dissolved, and could not form effective adsorption to paeonol, and could not complete the purification of paeonol, and after subsequently adjusting the pH to alkaline, the amount of paeonol contained in the precipitate was very small, and the detection process could not be performed normally. In comparative example 5, the pH was adjusted to 4.0 before the addition of chitosan, which was too low, and the chitosan was degraded, so that it could not effectively adsorb paeonol, and after the subsequent pH adjustment to be alkaline, the precipitate contained very little paeonol, and the detection process could not be performed normally. In comparative example 6, after adding chitosan, the pH was adjusted to 10.0 with sodium hydroxide, the alkalinity was too strong, the formed precipitate was too rapid and dense, the subsequent use of ethyl acetate could not effectively extract the chitosan in the precipitate, the paeonol was included in the chitosan precipitate, and could not be extracted, and the detection process could not be performed normally. In comparative example 6, the pH was adjusted to 6.5 with dilute acid after the chitosan was added, the precipitation amount was too small, paeonol could not be extracted effectively, and the detection process could not be performed normally. Comparative example 8 homogenization at room temperature resulted in the paeonol being easily destroyed. Comparative example 9 did not include a homogenization step, so that the extraction rate of paeonol was low.
The foregoing is merely an example of the present invention and common general knowledge of known specific structures and features of the embodiments is not described herein in any greater detail. It should be noted that, for those skilled in the art, without departing from the structure of the present invention, several changes and modifications can be made, which should also be regarded as the protection scope of the present invention, and these will not affect the effect of the implementation of the present invention and the practicability of the patent. The scope of the claims of the present application shall be determined by the contents of the claims, and the description of the embodiments and the like in the specification shall be used to explain the contents of the claims.
Claims (10)
1. A preparation method of a paeonia suffruticosa root-bark extract is characterized by comprising the following steps,
step (1): pretreating to obtain cortex moutan raw material;
step (2): firstly, adding water into the cortex moutan raw material in the step (1), then homogenizing the cortex moutan raw material, and then filtering to obtain filter residue;
and (3): adding ethyl acetate into the filter residue obtained in the step (2) to obtain an extraction system; carrying out ultrasonic treatment on the extraction system, then filtering to obtain filtrate, and concentrating the filtrate to obtain a crude cortex moutan extract;
and (4): dispersing the crude extract of cortex moutan in water, adding complex enzyme to obtain an enzymolysis system, and carrying out enzymolysis reaction on the enzymolysis system; after the enzymolysis reaction is finished, adjusting the pH value to be acidic, adding chitosan, stirring and uniformly mixing to obtain a system A, standing, adjusting the pH value of the system A to be alkaline or neutral, standing, filtering and taking precipitate; dispersing the precipitate in ethyl acetate, standing, filtering to obtain liquid phase part, concentrating the extractive solution, and drying to obtain cortex moutan extract.
2. The method for preparing the paeonia suffruticosa root-bark extract according to claim 1, wherein in the step (4), the chitosan is added in an amount of 10-20% of the cortex moutan raw material by weight.
3. The method for preparing the paeonia suffruticosa root-bark extract as claimed in claim 2, wherein in the step (4), the complex enzyme comprises cellulase, neutral protease, amylase and pectinase, and the content of the complex enzyme in an enzymolysis system is 2-4 wt%; the temperature of the enzymolysis reaction is 30-45 ℃, the duration is 2-3h, and the pH value is 7.0-7.5.
4. The method for preparing the paeonia suffruticosa root-bark extract as claimed in claim 3, wherein in step (4), after the completion of the enzymatic hydrolysis reaction, the pH value is adjusted to 5.0-5.5, chitosan is added, the mixture is stirred and mixed uniformly to obtain a system A, the pH value of the system A is adjusted to 7.0-8.0 after standing, and the precipitate is obtained by filtration after standing.
5. The method for preparing the paeonia suffruticosa root-bark extract as claimed in claim 4, wherein in the step (4), ethyl acetate 6-9 times of the weight of the moutan bark is added into the precipitate.
6. The method for preparing the paeonia suffruticosa root-bark extract as claimed in any one of claims 1 to 5, wherein in the step (3), the ultrasonic power for ultrasonic treatment of the extraction system is 100-200W, and the treatment time is 30-50 min; and adding ethyl acetate 4-7 times of the weight of the cortex moutan raw material into the filter residue.
7. The method for preparing the extract of the root bark of paeonia suffruticosa as claimed in any one of claims 1 to 5, wherein in the step (2), water of 0 to 10 ℃ is added to the raw material of moutan bark in an amount of 1 to 3 times the weight of the raw material of moutan bark.
8. The method for preparing the paeonia suffruticosa root-bark extract according to any one of claims 1 to 5, wherein in the step (1), the paeonia suffruticosa root-bark raw material is prepared by the following method: taking fresh Yunnan peony root bark, cleaning, crushing the fresh Yunnan peony root bark, and then sieving by a 10-mesh sieve to obtain the cortex moutan raw material.
9. The paeonia suffruticosa root-bark extract prepared by the preparation method according to any one of claims 1 to 5, wherein the content of paeonol in the paeonia suffruticosa root-bark extract is more than 90%.
10. The use of the paeonia suffruticosa root-bark extract according to claim 9 in cosmetics or pharmaceuticals.
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