CN106565802B - White peony root alcohol extract, 4-O-galloyl white paeoniflorin, and preparation and application thereof - Google Patents
White peony root alcohol extract, 4-O-galloyl white paeoniflorin, and preparation and application thereof Download PDFInfo
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- CN106565802B CN106565802B CN201610850345.5A CN201610850345A CN106565802B CN 106565802 B CN106565802 B CN 106565802B CN 201610850345 A CN201610850345 A CN 201610850345A CN 106565802 B CN106565802 B CN 106565802B
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Abstract
The invention relates to 4-O-galloyl paeoniflorin, a preparation method thereof and a new application thereof in preparing a medicine with β -secretase inhibition effect, which is characterized in that the prepared medicine can be used for treating neurodegenerative diseases such as Alzheimer disease, cerebral apoplexy and the like.
Description
The invention relates to a white peony root alcohol extract, a preparation method thereof and a new application of the obtained 4-O-galloyl paeoniflorin in preparing β -secretase inhibition drugs.
The recent study found that the production and accumulation of insoluble a β is mainly the product of Amyloid Precursor Protein (APP) after sequential passage through β -secretase (1) and gamma-secretase cleavage, which in turn initiates a neurotoxic cascade leading to oxidative stress, inflammatory response and irreversible neurological damage, thus, inhibition of β -secretase and gamma-secretase can alleviate the symptoms of AD, which is highly interesting in developing drugs for treating AD, whereas gamma-secretase inhibitors are interesting in developing drugs for treating AD, which are related to the development of toxic side effects of gamma-secretase inhibitors (β) and thus have been considered as side effects of inhibiting the growth of a certain physiological enzyme inhibitors, which are related to the development of a target site, thus, which are considered as a target-inducing inhibitors, such as a target-inducing inhibitor, which is considered to be a potent inhibitor of AD, and a target-secretase, which are considered to be a target-inducing side effects of AD-3526.
White peony root (Paeonia lactiflora Pall.) is also called Paeonia lactiflora Pall, Paeonia plant of Ranunculaceae, is not only a medicinal plant (the root of the Paeonia lactiflora Pall) but also a functional food, is widely used in Asia, has been used for over 2000 years, has a plurality of pharmacological effects as ornamental flowers and food, and is mainly reflected in the aspects of anticoagulation, depression resistance, oxidation resistance, liver protection and the like
The invention content is as follows:
the purpose of the invention is as follows: the invention provides a white peony root alcohol extract: preparation method of 4-O-galloyl albiflorin; and the application of the compound in preparing novel medicines for treating neurodegenerative diseases such as Alzheimer's disease, cerebral apoplexy and the like.
The technical scheme is as follows:
an alcohol extract of white peony root and 4-O-galloyl white paeony glycoside, which is characterized in that: the 4-O-galloyl paeoniflorin is a compound shown as the following general formula:
the preparation method of the white paeony root alcohol extract and the 4-O-galloyl paeoniflorin is characterized by comprising the following steps of: the method comprises the following steps:
(1) taking a white paeony root medicinal material, and carrying out reflux extraction for 3 times by using ethanol with the volume percentage concentration of 60%, wherein the extraction temperature is 80-90 ℃, and the extraction time is two hours each time;
(2) recovering the ethanol in the step (1) to obtain an extract;
(3) dispersing the extract in water, and extracting the extract;
(4) and (3) performing silica gel column chromatography dichloromethane-methanol gradient elution on the extract in the step (3), separating by SephadexLH-20, and preparing a liquid phase to obtain a compound, namely the 4-O-galloyl paeoniflorin.
The preparation method of the 4-O-galloyl paeoniflorin is characterized by comprising the following steps: the mobile phase component and proportion separated by SephadexLH-20 in the step (4) are dichloromethane: methanol is 1: 1.
The application of the 4-O-galloyl albiflorin in preparing medicines for treating neurodegenerative diseases is provided.
The application of the 4-O-galloyl albiflorin in preparing the medicine for treating the neurodegenerative diseases is characterized in that: the neurodegeneration includes alzheimer's disease and stroke.
The detection method of the inhibition effect of the white paeony root alcohol extract and the 4-O-galloyl paeoniflorin on β -secretase is characterized by comprising the following steps of:
(1) dissolving 10mL of buffer solution, 10mL of BACE1, 10mL of substrate and 10mL of sample to be detected in 10% DMSO, and incubating for 60min at 25 ℃ in a dark place, wherein the sample to be detected is 2, 10, 50 and 100 mu M/L4-O-galloyl paeoniflorin;
(2) measuring the fluorescence intensity of the sample by a microplate fluorescence spectrometer, wherein the exciting light is 545nm, the emitted light is 585nm, and the inhibition percentage is obtained by measurement;
the formula is as follows: percent (%) inhibition of ═ 1 [ (S)60﹣S0)/(C60﹣C0)]×100
Wherein: c60As blank control group: the fluorescence intensity detected after incubation of enzyme, buffer solution and substrate for 60 min; c0Fluorescence intensity measured at the beginning of blank controlDegree; s60Detecting the fluorescence intensity detected by the group after incubation for 60 min; s0To monitor the fluorescence intensity detected at the beginning of the group.
The advantages and effects are as follows: the invention explores the development prospect of the white paeony root, the 4-O-galloyl paeoniflorin and the analogues thereof on the pharmacy of neurodegenerative diseases such as Alzheimer's disease, cerebral apoplexy and the like, and develops a new medicinal field.
Description of the drawings:
FIG. 1 is a structural formula of 4-O-galloyl paeoniflorin of the present invention;
FIG. 2 is a graph showing the results of inhibition of β -secretase by using an extract of white peony root;
FIG. 3 is a graph showing the results of the inhibition of β -secretase by 4-O-galloyl albiflorin.
The specific implementation mode is as follows:
the invention relates to a white paeony root alcohol extract and 4-O-galloyl paeoniflorin, which is characterized in that: the 4-O-galloyl paeoniflorin is a compound shown as the following general formula:
also comprises a method for extracting 4-O-galloyl paeoniflorin from a white paeony root medicinal material, which comprises the following steps:
(1) taking a white paeony root medicinal material, and carrying out reflux extraction for 3 times by using ethanol with the volume percentage concentration of 60%, wherein the extraction temperature is 80-90 ℃, and the extraction time is two hours each time;
(2) recovering the ethanol in the step (1) to obtain an extract;
(3) dispersing the extract in water, and extracting the extract;
(4) and (3) performing silica gel column chromatography dichloromethane-methanol gradient elution on the extract in the step (3), separating by SephadexLH-20, and preparing a liquid phase to obtain a compound, namely the 4-O-galloyl paeoniflorin.
In the preparation method of the 4-O-galloyl paeoniflorin, the mobile phase component separated from the Sephadex LH-20 in the step (4) and the proportion are that dichloromethane: methanol is 1: 1.
Also comprises the application of the 4-O-galloyl albiflorin in the preparation of medicines for treating neurodegenerative diseases.
The application of the 4-O-galloyl albiflorin in preparing the medicine for treating the neurodegenerative diseases is characterized in that: the neurodegeneration includes alzheimer's disease and stroke.
The invention adopts a fluorescence resonance energy transfer method (FRET method) to determine the inhibition effect of the white paeony root alcohol extract and the 4-O-galloyl paeoniflorin on β -secretase (BACE1), and the result shows that the white paeony root alcohol extract and the 4-O-galloyl paeoniflorin have obvious inhibition effect on β -secretase (BACE1), so that the white paeony root alcohol extract and the 4-O-galloyl paeoniflorin can be used for researching and developing novel medicaments for treating neurodegenerative diseases such as Alzheimer's disease, cerebral apoplexy and the like.
The invention is illustrated by the examples:
example 1:
the preparation method of the white paeony root alcohol extract and the 4-O-galloyl paeoniflorin as claimed in claim 1, which is characterized by comprising the following steps: extracting radix paeoniae alba with 60% ethanol at 80-90 deg.C for 3 times (2 hr each time), and recovering ethanol to obtain extract (i.e. radix paeoniae alba ethanol extract); dispersing the extract in water, and extracting the extract; the extract is subjected to silica gel column chromatography, dichloromethane-methanol gradient elution and SephadexLH-20 separation (dichloromethane-methanol is 1:1), a liquid phase is prepared to obtain a compound 1, and the compound is identified as 4-O-galloyl paeoniflorin by combining physical and chemical properties and modern spectral means.
Detection of the inhibition effect of the white paeony root alcohol extract on β -secretase:
10mL of buffer solution, 10mL of BACE1, 10mL of substrate and 10mL of sample to be detected (10, 50, 100, 200mg/mL of white peony root alcohol extract) are dissolved in 10% DMSO, and incubated for 60min at 25 ℃ in the absence of light. The fluorescence intensity of the microplate is measured by a fluorescence spectrometer, the exciting light is 545nm, the emitted light is 585nm, and the inhibition percentage of the microplate is measured. The formula is as follows: percent (%) inhibition of ═ 1 [ (S)60﹣S0)/(C60﹣C0)]×100(C60Represent blank control group [ enzyme, buffer, substrate ]Fluorescence intensity detected after incubation for 60 min; c0Indicating the fluorescence intensity detected at the beginning of the blank control group; s60Indicating the fluorescence intensity detected by the detection group after incubation for 60 min; s0Indicating the fluorescence intensity detected at the beginning of the monitoring group).
The results are shown in FIG. 2, where different concentrations of Paeonia lactiflora alcohol extract inhibited β -secretase activity in a dose-dependent manner.
Detection of inhibition of 4-O-galloyl albiflorin to β -secretase:
10mL of buffer solution, 10mL of BACE1, 10mL of substrate and 10mL of sample to be detected (2, 10, 50, 100. mu.M/L of 4-O-galloyl paeoniflorin) are dissolved in 10% DMSO, and incubated for 60min at 25 ℃ in the absence of light. The fluorescence intensity of the microplate is measured by a fluorescence spectrometer, the exciting light is 545nm, the emitted light is 585nm, and the inhibition percentage of the microplate is measured. The formula is as follows: percent (%) inhibition of ═ 1 [ (S)60﹣S0)/(C60﹣C0)]×100(C60Representing the fluorescence intensity detected after incubation of blank control group [ enzyme, buffer solution and substrate ] for 60 min; c0Indicating the fluorescence intensity detected at the beginning of the blank control group; s60Indicating the fluorescence intensity detected by the detection group after incubation for 60 min; s0Indicating the fluorescence intensity detected at the beginning of the monitoring group).
The results are shown in FIG. 3, where various concentrations of 4-O-galloyl albiflorin inhibited β -secretase activity in a dose-dependent manner.
Claims (2)
- The application of 4-O-galloyl paeoniflorin is characterized in that: the application of the medicine in preparing medicine for treating neurodegenerative diseases, wherein the neurodegenerative diseases are Alzheimer's disease and cerebral apoplexy.
- 2. The use of 4-O-galloyl paeoniflorin according to claim 1, wherein different concentrations of 4-O-galloyl paeoniflorin can dose-dependently inhibit β -secretase activity.
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| CN201610850345.5A CN106565802B (en) | 2016-09-26 | 2016-09-26 | White peony root alcohol extract, 4-O-galloyl white paeoniflorin, and preparation and application thereof |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1714797A (en) * | 2004-07-02 | 2006-01-04 | 中国科学院上海药物研究所 | The medical usage of paeoniflorin |
| CN101053598A (en) * | 2007-05-29 | 2007-10-17 | 深圳市生物谷科技有限公司 | Medicinal composition for treating cardio-cerebralvascular diseases and diabetes |
| CN102125575A (en) * | 2010-01-19 | 2011-07-20 | 张作光 | Application of albiflorin in resisting Parkinson's disease |
| CN102258588A (en) * | 2011-03-10 | 2011-11-30 | 张建军 | Preparation method of peony general glycoside |
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- 2016-09-26 CN CN201610850345.5A patent/CN106565802B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1714797A (en) * | 2004-07-02 | 2006-01-04 | 中国科学院上海药物研究所 | The medical usage of paeoniflorin |
| CN101053598A (en) * | 2007-05-29 | 2007-10-17 | 深圳市生物谷科技有限公司 | Medicinal composition for treating cardio-cerebralvascular diseases and diabetes |
| CN102125575A (en) * | 2010-01-19 | 2011-07-20 | 张作光 | Application of albiflorin in resisting Parkinson's disease |
| CN102258588A (en) * | 2011-03-10 | 2011-11-30 | 张建军 | Preparation method of peony general glycoside |
Non-Patent Citations (4)
| Title |
|---|
| "中药白芍的高效液相色谱指纹图谱及其主要成分含量测定";谭菁菁，等;《国际药学研究杂志》;20140430;第41卷(第2期);第238-243页 * |
| "白芍化学成分研究";谭菁菁，等;《中草药》;20100831;第41卷(第8期);第1245-1248页 * |
| "白芍总苷对阿尔茨海默病的保护作用及机制的研究";李岩，等;《职业与健康》;20140930;第30卷(第17期);第2525-2528页 * |
| 谭菁菁，等."中药白芍的高效液相色谱指纹图谱及其主要成分含量测定".《国际药学研究杂志》.2014,第41卷(第2期),第238-243页. * |
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Effective date of registration: 20190419 Address after: 110016 No. 83, cultural road, Shenhe District, Shenyang, Liaoning Applicant after: General Hospital of the Northern War Zone of the Chinese People's Liberation Army Address before: 110 840 No. 83 Cultural Road, Shenhe District, Shenyang City, Liaoning Province Applicant before: Zhao Qingchun |
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