CN109700800A - Application of the AE as preparation prevention and treatment hyperlipidaemic conditions drug - Google Patents
Application of the AE as preparation prevention and treatment hyperlipidaemic conditions drug Download PDFInfo
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- CN109700800A CN109700800A CN201910030717.3A CN201910030717A CN109700800A CN 109700800 A CN109700800 A CN 109700800A CN 201910030717 A CN201910030717 A CN 201910030717A CN 109700800 A CN109700800 A CN 109700800A
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- ptp
- hyperlipidaemic conditions
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Abstract
A kind of application the present invention provides natural products small molecule AE as preparation prevention and treatment hyperlipidaemic conditions drug, result of study of the present invention shows, AE can be such that GSK-3 β and Akt the protein phosphorylation level on PTP-1B signal path raises, to lower the protein level of PTP-1B, and it is able to suppress the activity of PTP-1B, to play the effect of prevention and treatment hyperlipidaemic conditions.AE has a good application prospect as natural novel PTP-1B signal path regulator.
Description
Technical field
The present invention relates to a kind of active ingredient of natural product AE's (8-C-Asorbyl- (-)-Epigallocatechin)
Pharmaceutical usage more particularly to AE (8-C-Asorbyl- (-)-Epigallocatechin) are in prevention and treatment hyperlipidaemic conditions drug
Using belonging to medicine field.
Technical background
8-C-Asorbyl- (-)-Epigallocatechin (formula 1), earliest by Itsuo Nishioka seminar in
It is extracted from oolong tea within 1989 isolated.2013, Yang Zhen seminar was synthesized with catechin and vitamin C for the first time
It arrives.Studies have shown that 8-C-Asorbyl- (-)-Epigallocatechin has various health cares and pharmacological action to human body, such as
Inhibit pancreatic lipases, AntiHIV1 RT activity, it is antitumor the effects of.
The chemical structure of 1 AE of formula (8-C-Asorbyl- (-)-Epigallocatechin)
Insulin can promote the synthesis and storage of fat, reduce free fatty acid in blood, while inhibiting the decomposition of fat
Oxidation.Insulin deficit can cause fat metabolic disturbance, and depot fat is reduced, and is decomposed and is reinforced, and blood lipid increases, and long can cause to move
Arteries and veins hardening, and then lead to the serious condition of cardiovascular and cerebrovascular；At the same time, insulin deficit will lead to body fat and decompose and reinforces,
A large amount of ketoboidies are generated, ketoacidosis occur.PTP-1B plays important role, pancreas islet in Insulin signaling pathway
Element is combined with insulin receptor (IR), is allowed to phosphorylation, and then cause the phosphorylation of downstream signaling molecule Akt and GSK-3 β, from
And activate insulin signaling pathway.PTP-1B is by making insulin receptor that dephosphorylation occur, so that disabling signal is transduceed, in pancreas
Play the role of negative regulator in the element signal path of island.By inhibiting PTP-1B signal path, increase insulin sensitivity, to reach
To the effect of reducing blood lipid.
At present there is no about 8-C-Asorbyl- (-)-Epigallocatechin act on PTP-1B signal path and
The report for preventing and treating hyperlipidemia.
Summary of the invention
The purpose of the present invention is to provide a kind of active ingredient of natural product 8-C-Asorbyl- (-)-
Epigallocatechin prevention and treatment hyperlipidaemic conditions drug in application, should compound may be used as prevention and treatment hyperlipidemia treatment
Potential drug.
In order to solve the above-mentioned technical problem, technical solution proposed by the present invention is:
A kind of natural products small molecule AE prevents and treats the application of hyperlipidaemic conditions drug, small point of the natural products as preparation
The structure of sub- AE are as follows:
Moreover, the compound is 1-100 μM to the effective dose that PTP-1B signal path in hyperlipidemia is adjusted.
A kind of blood lipid-lowering medicine, including AE, the structure of the natural products small molecule AE are as follows:
Moreover, further including the carrier or excipient pharmaceutically received.
Moreover, the excipient pharmaceutically received include tablet, capsule, granule, injection, pill, syrup,
Powder, paste or liquid preparation.
The advantages and positive effects of the present invention are as follows:
AE (8-C-Asorbyl- (-)-Epigallocatechin) provided by the invention can make PTP-1B signal path
On GSK-3 β and Akt protein phosphorylation level up-regulation, to lower the protein level of PTP-1B, and be able to suppress PTP-1B's
Activity has the function that prevent and treat hyperlipidaemic conditions.Illustrate that the compound may be used as the potential drug of prevention and treatment hyperlipidemia treatment.Institute
PTP-1B protein level can be lowered by stating compound, and be able to suppress the activity of PTP-1B enzyme.
The present invention is detected by cell protein, enzyme inhibition activity detection, the detection of cell Lipid-lowering activities, and lipid-loweringing is living in Mice Body
Property detection method.As a result, it has been found that 8-C-Asorbyl- (-)-Epigallocatechin can make on PTP-1B signal path
The up-regulation of GSK-3 β and Akt protein phosphorylation level, to lower the protein level of PTP-1B, and is able to suppress the work of PTP-1B
Property, have the function that prevent and treat hyperlipidaemic conditions.
Detailed description of the invention
Fig. 1 is AE and NaVO4To the Western-blot experimental result of insulin signaling pathway GAP-associated protein GAP.
Fig. 2 is that p-GSK3 β and GSK3 β are gray analysis quantitative ratio figure in Fig. 1.* P < 0.01 P < 0.05, * * and * * * P
< 0.001 compared with control group.
Fig. 3 is that p-AKT and AKT is gray analysis quantitative ratio figure in Fig. 1.* P < 0.05 and P < 0.01 * * and control
Group is compared.
Fig. 4 is that PTP-1B gray analysis is quantitatively schemed in Fig. 1.P < 0.01 * and P < 0.001 * * * are compared with control group.
Specific embodiment
Essentiality content of the invention is specifically introduced below with reference to embodiment, but protection model of the invention is not limited with this
It encloses.
Embodiment 1
A kind of natural products AE (8-C-Asorbyl- (-)-Epigallocatechin) is used as Protein-tyrosine-phosphatase
Application of the 1B signal path regulator in prevention and treatment hyperlipidaemic conditions drug
One, experimental material
PNPP (pNPP), NaVO4, it is purchased from Sigma-Aldrich (St.Louis, MO)；Recombination
People's PTP-1B albumen is purchased from Abcam (Cambridge, UK)；CHO-K1 cell is obtained from Chinese Academy of Sciences Shanghai life science
Institute's (Chinese Shanghai).
Male mouse of kunming (18-22g) is provided by Chinese People's Liberation Army's Academy of Medical Sciences Experimental Animal Center, through ground
Square animal protection office approval strictly observes local and country the code of ethics.Mouse is maintained in polypropylene cage (5, every cage)
And (18-23 DEG C, 55-60% humidity, 12 hours Dark-light cycles) raising in laboratory conditions, all free diet drinks of mouse
Water.
Two, experimental method
1. external inhibition of enzyme activity experimental method
The inhibitory activity test method of 1.1 couples of PTP-1B
Reaction system be 100 μ L, test buffer pH 6.0, comprising 150mM NaCl, 50mM MES, 1mM EDTA,
1mM DTT and 0.05%NP-40.Experimental group includes the test compound of 10 μ L, the PTP-1B (20 μ g/mL) of 5 μ L, blank group
In only buffer, (30 μM) of sodium orthovanadate be used as positive control.Temperature incubates 5min under the conditions of 30 DEG C of compound of enzyme and test, then
The substrate pNPP of 10 μ L, 100mM is added, continues temperature and incubates 10min, reads absorbance value at 405nm using microplate reader, calculates and survey
Compound is tried to the inhibiting rate of PTP-1B.
2. cellular level mechanism of action evaluation method
2.1 Western-blot experiment
CHO-K1 cell is containing 10% fetal calf serum, the DMEM-F12 culture of 10U/mL penicillin and 10mg/mL streptomysin
In base, in 37 DEG C, 5%CO2Under the conditions of cultivate.The cell in logarithmic growth phase is taken, with 5 × 104The density of cells/mL is inoculated with
In capsule, cultivated for 24 hours under the conditions of 37 DEG C, 5%CO2；Test compound or DMSO is added, (20 μM) of sodium orthovanadate as positive
Control acts on 48h；After 10nM insulin stimulating 10min, cell is collected, 1 × PBS is washed 1 time, and the albumen containing 0.1% is added
Enzyme inhibitor PMSF, cracks 30min on ice, and centrifuging and taking supernatant carries out protein quantification with BCA method.
The separation of SDS-PAGE electrophoresis progress destination protein, 100V, 300mA race glue 100min, cutting pvdf membrane, 15V,
300mA transferring film 100min.1h, 4 DEG C of primary antibody overnight incubations are closed with 5% skimmed milk power, secondary antibody is incubated for 1h, and upper machine sweeps film.
3. lipidemia activity research in animal body
The foundation of 3.1 hyperlipemia mice models
After mouse adaptive feeding 1 week, high lipid food is raised 4 weeks, and 6h is deprived of food but not water, small with the dosage of 100mg/kg
STZ is injected intraperitoneally in mouse, continues high lipid food and feeds.It is arranged 5 groups, i.e., normal group, model group, positive controls, high and low dose of drug
Amount group.
Isometric physiological saline is given in normal group and model group stomach-filling, and positive controls give Lovastatin (50 mg/
Kg), administration group gives drug (50mg/kg) and (100mg/kg), records weight, and daily administration 1 time, successive administration 4 weeks.
The detection of 3.2 Triglycerides in Serum and cholesterol
After successive administration 4 weeks, it is deprived of food but not water 6h, eyeball of mouse takes blood in the EP of the heparin sodium aqua containing 1 μ L 0.2%
Guan Zhong, 5000rpm are centrifuged 10min, take serum, the detection for triglycerides and cholesterol biochemical indicator.
Three, experimental result
3.1. (AE) 8-C-Asorbyl- (-)-Epigallocatechin adjusts PTP-1B signal path
Insulin signaling pathway depends on the activation of insulin receptor, and insulin and insulin receptor (IR) are combined, after making
Person's phosphorylation, and then cause the phosphorylation of downstream signaling molecule, to activate insulin signaling pathway.PTP-1B is by making pancreas
Dephosphorylation occurs for island element receptor, so that disabling signal is transduceed, plays the role of negative regulator in insulin signaling pathway.Thin
In born of the same parents' culture, after drug treatment, p-GSK-3 β (Ser9) expressing quantity in insulin access increases (Fig. 1 and Fig. 2)；p-
The protein expression of Akt (Ser473) also raises (Fig. 1 and Fig. 3)；In addition, the protein level of PTP-1B is inhibited, albumen table
Up to downward (Fig. 1 and Fig. 4).However, 8-C-Asorbyl- (-)-Epigallocatechin is right under 5,20 and 40 μM of concentration
HPTP-1B enzyme has faint inhibitory activity (table 1).These are the results show that 8-C-Asorbyl- (-)-Epigallocatechin
Directly enhancing insulin signal transduction activity reduces PTP-1B protein expression level, and has faint inhibition living hPTP-1B
Property.
Inhibiting effect of 1. 8-C-Asorbyl- (-)-Epigallocatechin of table to hPTP-1B
aThe result is that the average value of independent experiment three times,bReference compound.
3.2. (AE) 8-C-Asorbyl- (-)-Epigallocatechin reduces blood lipid level in Mice Body
8-C-Asorbyl- (-)-Epigallocatechin reduces internal serum total cholesterol and triglyceride levels
As shown in table 2, compared with normal group, serum total cholesterol and triglyceride levels significantly increase model mice.With
Model mice is compared, with the serum total cholesterol of 8-C-Asorbyl- (-)-Epigallocatechin diabetic mice handled
Level has dropped 33.8% (50mg/kg) and 27.3% (100mg/kg), and triglyceride levels have dropped 8.3% (100mg/
kg)。
2. mice serum total cholesterol (TC) of table and triglycerides (TG) are horizontala
aData are expressed as average value ± SD (n=10).* P < 0.05 and * * P < 0.01vs T2D model.
Embodiment 2
The application of 8-C-Asorbyl- (-)-Epigallocatechin
It is used to prepare blood lipid-lowering medicine, including active constituent 8-C-Asorbyl- (-)-Epigallocatechin, is also wrapped
Include pharmaceutically acceptable carrier or excipient, be made pharmaceutically acceptable dosage form pharmaceutically acceptable carrier or
Excipient includes one or more solids, semisolid or Auxiliary Liquid Material；Pharmaceutically acceptable dosage form includes tablet, capsule
Agent, granule, injection, pill, syrup, powder, paste, liquid preparation etc..
Claims (5)
1. a kind of application of natural products small molecule AE as preparation prevention and treatment hyperlipidaemic conditions drug, the natural products small molecule
The structure of AE are as follows:
2. application of the natural products small molecule AE according to claim 1 as preparation prevention and treatment hyperlipidaemic conditions drug,
Be characterized in that: the compound is 1-100 μM to the effective dose that PTP-1B signal path in hyperlipidemia is adjusted.
3. a kind of blood lipid-lowering medicine, it is characterised in that: including AE, the structure of the natural products small molecule AE are as follows:
4. blood lipid-lowering medicine according to claim 3, it is characterised in that: further include the carrier or figuration pharmaceutically received
Agent.
5. blood lipid-lowering medicine according to claim 3, it is characterised in that: the excipient pharmaceutically received includes piece
Agent, capsule, granule, injection, pill, syrup, powder, paste or liquid preparation.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201910030717.3A CN109700800A (en) | 2019-01-14 | 2019-01-14 | Application of the AE as preparation prevention and treatment hyperlipidaemic conditions drug |
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| CN201910030717.3A CN109700800A (en) | 2019-01-14 | 2019-01-14 | Application of the AE as preparation prevention and treatment hyperlipidaemic conditions drug |
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| CN109700800A true CN109700800A (en) | 2019-05-03 |
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| CN201910030717.3A Pending CN109700800A (en) | 2019-01-14 | 2019-01-14 | Application of the AE as preparation prevention and treatment hyperlipidaemic conditions drug |
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1980654A (en) * | 2004-07-05 | 2007-06-13 | 三得利株式会社 | Lipase inhibitors |
-
2019
- 2019-01-14 CN CN201910030717.3A patent/CN109700800A/en active Pending
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1980654A (en) * | 2004-07-05 | 2007-06-13 | 三得利株式会社 | Lipase inhibitors |
Non-Patent Citations (2)
| Title |
|---|
| MASAAKI NAKAI等: "Inhibitory Effects of Oolong Tea Polyphenols on Pancreatic Lipase in Vitro", 《J. AGRIC. FOOD CHEM.》 * |
| SATABDEE MOHAPATRA等: "In silico investigation of black tea components on α-amylase,α-glucosidase and lipase", 《JOURNAL OF APPLIED PHARMACEUTICAL SCIENCE》 * |
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