CN106822071A - It is a kind of for treating coronary heart disease, the effective ingredient in Chinese of hyperlipidemia, preparation method and the therefrom method of separating effective ingredient - Google Patents
It is a kind of for treating coronary heart disease, the effective ingredient in Chinese of hyperlipidemia, preparation method and the therefrom method of separating effective ingredient Download PDFInfo
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Abstract
The invention belongs to medical sci-tech field, extracted specifically related to from the rhizome of Radix Curcumae obtain for treating coronary heart disease, the effective ingredient in Chinese of hyperlipidemia, its preparation method and the therefrom method of separating effective ingredient, dry rhizome with turmeric section plant Radix Curcumae is as medicinal part,, through Solvent Extract methods, extract is separated, purify the active component containing sesquiterpenoids composition for obtaining for medicinal part.Pharmacological experiment study shows：Curcumadiol, procurcumenol, curdione and zedoarondiol and total extract have significantly treatment myocardial ischemia to act on; rat myocardium from injury myocardial infarction area scope caused by ischemic can be obviously reduced, coronary ligation myocardial ischemia in rats is damaged has significant protective effect.The application is the divisional application of the application for a patent for invention of entitled " a kind of effective ingredient in Chinese for treating coronary heart disease, hyperlipidemia and preparation method thereof ".
Description
The application is the Application No. 201410491640.7 that September in 2014 is submitted on the 23rd, and entitled " one kind is used for
The divisional application of the application for a patent for invention for the treatment of coronary heart disease, effective ingredient in Chinese of hyperlipidemia and preparation method thereof ".
Technical field
The invention belongs to medical sci-tech field, and in particular to from the rhizome of Radix Curcumae extract obtain for treating coronary disease
Disease, the effective ingredient in Chinese of hyperlipidemia, its preparation method and the therefrom method of separating effective ingredient.
Background technology
Heart disease, malignant tumour, cerebrovascular disease are 3 kinds of diseases of life of the current serious threat mankind, 90 years last century
Coronary heart disease ranking China Health Killer list first place since generation, with developing rapidly for China's economy, living standards of the people are not
Disconnected to improve and life style, the huge change of dietary structure generation, coronary heart disease is in continuous ascendant trend in the incidence of disease of China.Hat
Worry is a kind of most common heart disease, refers to the myocardial dysfunction and device caused by coronary artery stenosis, blood supply insufficiency
Matter venereal disease becomes, therefore also known as ischemic heart disease (IHD).The Etiological for inducing coronary heart disease has：Hyperlipidemia, hypertension, sugar
The bad life habits such as urine disease, obesity and smoking.HypercholesterolemicRats disorder is the most important cause of coronary heart disease, modern medicine
Research shows to be existed between oxidized ldl cholesterol and the level of apolipoprotein and the danger of coronary event
Close relationship, hyperlipidemia is closely related with atherosclerosis, is the major incentive of coronary heart disease.Patients with coronary heart disease is except having
May cause outside death because coronary artery obstruction causes miocardial infarction, anginal frequent breaking-out will also make its study, ability to work big
Big limited, personal lifestyle can not be taken care of oneself when serious, and white elephant is all brought to family and society.Thus, it is found that exploitation has
The medicine of effect treatment coronary heart diseases and angina pectoris, is one very significant to reduce threat of the angiocardiopathy to human health
Work.
Radix Curcumae is the root of dicotyledon Zingiber curcuma Radix Curcumae (Curcuma aromatica salisb.)
Stem, is distributed in the ground such as China Guangxi, Guizhou, is local Wild Medicinal Plants Resources.Radix Curcumae has various medicinal efficacies, clinical
On by custom for curing mainly sternal rib shouting pain, Amenorrhea, rheumatism shoulder arm pain, tumbling and swelling etc., modern pharmacological research shows that it has
Anti-inflammatory and immunosuppressive action.But its Effect study for being used to treat myocardial ischemia, coronary heart disease and hyperlipidemia has no report.
We have found that Nanning Region Guangxi has and it are treated into facing for the obstruction of qi in the chest with other medicinal material compatibilities when collecting time tested folk prescriptions
Bed application practice, method is succinct and evident in efficacy, be no lack of in the case recorded several beds all the year round can not walk, dizzy, whole body
Patient with severe symptoms out of breath, insomnia powerless, uncomfortable in chest and the prolonged illness that the cardiovascular common drug state of an illness is not obviously improved is taken for many years suffer from
Person.In order to fully excavate the clinical value of the medicinal material, the medicinal material is conducted in-depth research, by clinical experience side
Dismemberment study is carried out, the different active components of other side's Chinese medicine carry out medicine efficacy screening, are carried out using substantial amounts of generally acknowledged animal model
Pharmacodynamics verifies that filter out target effective position and active ingredient, pharmacological research confirms that Radix Curcumae sesquiterpenoids can be bright
The aobvious level for reducing LDH and CK in damaged myocardium cell, reduction myocardial infarct size, protect damaged myocardium, can effectively treat hat
Worry, reduces the blood lipid level in blood, improves prognosis, there is foreseeable clinical Benefit.
The content of the invention
The present invention carries out medicine efficacy screening by the different active components to Radix Curcumae, it was found that it is used for therapeutic purpose coronary disease
Disease and hyperlipidemia active component, by carrying out to sesquiterpenoids composition in active component in depth analyzing and researching comprehensively, therefrom
Four sequiterpene monomer active ingredients such as curcumadiol, procurcumenol, curdione, zedoarondiol are isolated, through physics and chemistry
Constant and Spectrum Analysis, their structural formula are as follows respectively：
In the first aspect of the present invention, there is provided a kind of effective ingredient in Chinese for treating coronary heart disease, hyperlipidemia, its
It is characterised by：Dry rhizome with turmeric section plant Radix Curcumae (Curcuma aromatica salisb.) as medicinal part, medicine
With position through Solvent Extract methods, the active component containing sesquiterpenoids composition that extract is separated, purifying is obtained, effective portion
The content (mass percent) of sesquiterpenoids composition is more than 50% in position.
Specifically, main compound in sesquiterpenoids composition be curcumadiol, procurcumenol, curdione and
The content of zedoarondio, wherein curcumadiol in active component is 10~40%, procurcumenol containing in active component
It is 15~35% to measure, and content of the curdione in active component is 10~30%, zedoarondiol in active component
Content is 5~30%.
The second aspect of the present invention provides a kind of preparation method of effective ingredient in Chinese, it is characterised in that including following step
Suddenly：
(1) Radix Curcumae medicinal part add 2~12 times of (weight ratio) concentration is 50~95% ethanol, methyl alcohol, normal propyl alcohol, different
One kind or any several mixed solvents in propyl alcohol, n-butanol or ethyl acetate carry out refluxing extraction, and extract solution recycling design is dense
Contract to obtain Radix Curcumae crude extract；Or after Radix Curcumae medicinal part is pulverized and sieved, with ethanol as entrainer, put supercritical CO2Extraction,
Extract is Radix Curcumae crude extract；
(2) Radix Curcumae crude extract is extracted with 2~18 times of water-saturated n-butanols of amount, ethyl acetate or petroleum ether-ethyl acetates
Take, extract recycling design is concentrated to give Radix Curcumae active component.
Present invention also offers the preparation method of another effective ingredient in Chinese, it is characterised in that comprise the following steps：
(1) Radix Curcumae medicinal part add 2~12 times of (weight ratio) concentration be 50~95% ethanol, or methyl alcohol, normal propyl alcohol,
One kind in isopropanol, n-butanol, ethyl acetate or any several mixed solvents carry out refluxing extraction, extract solution recycling design,
It is concentrated to give Radix Curcumae crude extract；Or after Radix Curcumae medicinal part is pulverized and sieved, with ethanol as entrainer, put supercritical CO2Extraction
Take, extract is Radix Curcumae crude extract；
(2) by Radix Curcumae crude extract through macroporous resin adsorption, respectively with 1~3 times of water of column volume, 1~3 times of column volume
Concentration is 10~30% alcohol flushing resin post, discards water and dilute alcohol eluent, then parses resin with 80~95% ethanol
The active ingredient of absorption, the ethanol eluate of Fractional Collections 80~95% reclaims ethanol, is concentrated to give Radix Curcumae active component.
Preferably, macroreticular resin is styrene type porous polymeric adsorbent, and polarity is low pole or middle polarity；For example may be used
To select D101, HPD-100, HPD-300, HPD-450, HPD-5000 or H103 type macroreticular resin.
The third aspect of the present invention provide a kind of active component from Radix Curcumae in separating effective ingredient method, comprising with
Lower step：
(1) Radix Curcumae active component is prepared；
(2) by Radix Curcumae active component through silica gel column chromatography, eluent is 8-20:1 petroleum ether-ethyl acetate system,
Collect eluent and obtain curdione crude product；Curdione crude product preparative high performance liquid chromatography purify curdione refine
Product；
(3) eluent is replaced by 4-7:1 petroleum ether-ethyl acetate system, collects eluent and obtains
Zedoarondiol crude products, zedoarondiol crude product preparative high performance liquid chromatographies purify zedoarondiol refine
Product；
(4) eluent is replaced by 3-2:1 petroleum ether-ethyl acetate system, collects eluent and obtains the 3rd flow point, receives
Collecting the 3rd flow point and isolated and purified by preparative high performance liquid chromatography can obtain curcumadiol and procurcumenol.
Beneficial effect：
Find that curcumadiol, procurcumenol, curdione and zedoarondiol and Radix Curcumae active component have first
Significantly the effect for the treatment of myocardial ischemia, can be obviously reduced rat myocardium from injury myocardial infarction area scope caused by ischemic, to coronary ligation
Myocardial ischemia in rats is damaged has significant protective effect, also has good curative effect to coronary heart disease and hyperlipidemia.
Specific embodiment：
Embodiment 1：
(1), Radix Curcumae rhizome removes silt, fibrous root, cleans, and section is dried, and is made medicinal part；
(2), Radix Curcumae rhizome medicine materical crude slice 1800g is taken, plus the alcohol reflux of 6 times of amounts 80% must take twice, merging is extracted twice
Liquid, filtration, filtered solution reclaims ethanol, is concentrated under reduced pressure into relative density for 1.10 (45 DEG C) Zuo You, obtains Radix Curcumae crude extract；
(3), crude extract is extracted 6 times with 2 times of water-saturated n-butanols of amount, merges butanol extraction liquid, and positive fourth is recovered under reduced pressure
Alcohol, concentration, is vacuum dried to obtain active component 121g.
Embodiment 2：
(1), Radix Curcumae rhizome removes silt, fibrous root, cleans, and section is dried, and is made medicinal part；
(2), Radix Curcumae rhizome medicine materical crude slice 2000g is taken, plus the alcohol reflux of 6 times of amounts 75% must take twice, merging is extracted twice
Liquid, filtration, filtered solution reclaims ethanol, is concentrated under reduced pressure into relative density for 1.05~1.10 (45 DEG C) Zuo You, obtains Radix Curcumae and slightly carries
Thing；
(3), crude extract is extracted 6 times with 3 times of ethyl acetate of amount, and combined ethyl acetate extract is recovered under reduced pressure acetic acid second
Ester, concentration, is vacuum dried to obtain active component 116g.
Embodiment 3：
(1), Radix Curcumae rhizome removes silt, fibrous root, cleans, and section is dried, and is made medicinal part；
(2), Radix Curcumae rhizome medicine materical crude slice 2000g is taken, plus the alcohol reflux of 6 times of amounts 75% must take twice, merging is extracted twice
Liquid, filtration, filtered solution reclaims ethanol, is concentrated under reduced pressure into relative density for 1.05~1.10 (45 DEG C) Zuo You, obtains Radix Curcumae and slightly carries
Thing；
(3), crude extract with 3 times amount by petroleum ether：(volume ratio is 3~20 to ethyl acetate：1) the mixed solvent extraction of composition
Take, extract 6 times, merge by petroleum ether：Acetic acid ethyl acetate extract, is recovered under reduced pressure solvent, concentration, is vacuum dried to obtain active component
105g。
Embodiment 4
(1), Radix Curcumae rhizome removes silt, fibrous root, cleans, and section is dried, and is made medicinal part；
(2), Radix Curcumae rhizome medicine materical crude slice 2000g is taken, plus the alcohol reflux of 6 times of amounts 75% must take twice, merging is extracted twice
Liquid, filtration, filtered solution reclaims ethanol, and the ethanol that is concentrated under reduced pressure obtains Radix Curcumae alcohol extract to the greatest extent；
(3), through D101 type macroporous resin adsorptions, resin pillar blade diameter length ratio is 1 to Radix Curcumae alcohol extract：8, applied sample amount is medicinal material：
Resin=3：1, resin pillar is first respectively with the alcohol flushing resin that the concentration of 2 times of water of column volume, 2 times of column volumes is 30%
Post, discards water and dilute alcohol eluent；
(4), resin pillar parses the active ingredient of resin adsorption, Fractional Collections ethanol with 4 times of ethanol of column volume 90% again
Eluent, eluent reclaims ethanol, and concentration is vacuum dried to obtain active component 102g.
Embodiment 5：
(1), Radix Curcumae rhizome removes fibrous root, silt, cleans, and section is dried, and is made medicinal part；
(2) the crushing of Radix Curcumae rhizome medicine materical crude slice, is taken, 10 mesh sieves are crossed, 5Kg is weighed and is put supercritical CO2The extraction kettle of extraction equipment
In；
(2), (500g), extracting pressure and supercritical CO with ethanol as entrainer2Fluid temperature (F.T.) is 35MPa and 50 DEG C, is separated
The pressure and temperature of kettle I is 8MPa and 50 DEG C, and it is 5MPa and 35 DEG C to separate II pressure and temperatures, carries out supercritical extract, circulation extraction
Take 1.5~2 hours, extract is released from separating still I, separation reactor I I discharging openings.
(3), supercritical CO260 DEG C of extract, -0.09MPa be removed under reduced pressure ethanol to the greatest extent, with 2 times amount by petroleum ether：Second
(volume ratio is 3~20 to acetoacetic ester：1) mixed extractant solvent of composition 4 times, combining extraction liquid is recovered under reduced pressure solvent, has been concentrated to give
Effect position 415g.
Embodiment 6：The process recipes of Radix Curcumae sequiterpene monomeric compound：
(1), Radix Curcumae rhizome removes silt, fibrous root, cleans, and section is dried, and is made medicinal part；
(2), Radix Curcumae rhizome medicine materical crude slice 3000g is taken, plus the alcohol reflux of 6 times of amounts 80% must take twice, merging is extracted twice
Liquid, filtration, filtered solution reclaims ethanol, is concentrated under reduced pressure into relative density for 1.10 (45 DEG C) Zuo You, obtains Radix Curcumae crude extract；
(3), crude extract is extracted 6 times with 2 times of water-saturated n-butanols of amount, merges butanol extraction liquid, and positive fourth is recovered under reduced pressure
Alcohol, concentration, is vacuum dried to obtain active component 204g.
(4) active component 204g, is taken, is mixed thoroughly with proper silica gel, dry method upper prop, through silica gel column chromatography, with petroleum ether-acetic acid
Ethyl ester system is eluted, and first (presses 20 with petroleum ether-ethyl acetate:1) elute, obtain curdione crude product eluent, eluent is reclaimed
Solvent to the greatest extent, then with preparation liquid phase (mobile phase is 50% acetonitrile-aqueous solution) be prepared into curdione highly finished product 5.7g, purity
99.8%.Silica gel column chromatography (presses 7 with petroleum ether-ethyl acetate again:1) elute, obtain zedoarondiol crude product eluents, elute
Liquid recycling design is to the greatest extent, then to be prepared into zedoarondiol smart with liquid phase (acetonitrile-aqueous solution of mobile phase 10%-15%) is prepared
Product 2.3g, purity 98.5%.Silica gel column chromatography (presses 2 with petroleum ether-ethyl acetate again:1) elute, obtain curcumadiol and former cowherb
Art alcohol crude product eluent, eluent recycling design is made to the greatest extent with liquid phase (acetonitrile-aqueous solution of mobile phase 15%-50%) is prepared
Standby to obtain curcumadiol highly finished product 2.5g, purity 97.8% merges, recycling design through the mother liquor after HPLC preparative separation curcumadiols
Afterwards, then with liquid phase (acetonitrile-aqueous solution of mobile phase 30%) preparation is prepared procurcumenol highly finished product 6.1g, purity can be obtained
98.9%.
Embodiment 7
Four kinds of compounds are carried out with physicochemical constant and Wave Spectrum analysis：To compound curdione using LC-MS,1H-
NMR、13C-NMR、Dept、1H-1H COSY、13C-1H COSY identification of means, obtains molecular formula C15H24O2, such as chemical structural formula institute
Show.To compound zedoarondiol using LC-MS,1H-NMR、13C-NMR、Dept、1H-1H COSY、13C-1H COSY are identified
Means, obtain molecular formula C15H24O3, as shown in chemical structural formula.To compound curcumadiol using LC-MS,1H-NMR、13C-NMR、
Dept、1H-1H COSY、13C-1H COSY identification of means, obtains molecular formula C15H22O3, structural formula is as shown in chemical structural formula.To changing
Compound procurcumenol using LC-MS,1H-NMR、13C-NMR、Dept、1H-1H COSY、13C-1H COSY identification of means, obtains molecule
Formula C15H22O2, as shown in chemical structural formula.Wherein curdione13C-NMR is13C NMR(400MHz,CDCl3)δ16.57,
18.54,19.86,21.12,26.41,30.00,34.05,44.24,46.81,53.61,55.92,129.90,131.59,
211.14,214.37；Zedoarondiol's13C-NMR is13C NMR(400MHz,CDCl3)δ20.17,21.16,22.06,
22.21,22.38,28.47,39.67,51.83,56.15,60.61,71.28,78.49,135.57,139.18,202.55；Cowherb
Art glycol13C-NMR is13C NMR(400MHz,CDCl3)δ21.93；22.00；22.71；24.09；27.19；36.98；
37.16；60.94；83.19；86.44；127.97；132.89；143.30；151.03；194.09；Procurcumenol13C-NMR
For13C NMR(400MHz,CDCl3)δ20.35；21.52；22.49；23.39；26.02；27.75；39.04；49.62；53.01；
79.39；128.27；135.40；135.88；154.22；198.26.(Shanghai is micro- with carbon-13 nmr spectra database for carbon modal data
Spectrum information Technology Co., Ltd.) in the compound carbon spectrum included it is consistent.
Embodiment 8：The content detection analysis of sesquiterpenoids composition in active component
(1), instrument and reagent
The high performance liquid chromatographs of Dionex UltiMate 3000, LPG-3400SD high-pressure pumps, WPS-3000 (RS) is automatic
Injector, TCC-3x00 (RS) column oven, VWD-3x00 (RS) UV-detector, the work stations of Chromeleon 6.8.
Acetonitrile, TEDIA companies, chromatographically pure；Water is the homemade deionized water of Hi-Tech water purification systems (18.2M Ω).
Reference substance：Curdione, zedoarondiol, curcumadiol, procurcumenol
(2), the preparation of solution
The preparation of reference substance solution：Precision weighs curdione, zedoarondiol, curcumadiol, procurcumenol self-control
Appropriate reference substance, plus methyl alcohol dissolving is made every lmL μ containing curdione, zedoarondiol, curcumadiol, procurcumenol about 100
The reference substance solution of g.
(3), the preparation of need testing solution：The active component that precision weighs preparation is appropriate, plus methyl alcohol dissolving is made every lmL
Need testing solution containing about 2mg.
(4), chromatographic condition：
Using RPLC SymmetryC18(5 μm, 250 × 4.6mm), with water as mobile phase A, be with acetonitrile
Mobile phase B, flow velocity 1.0mL/min, gradient elution program is shown in Table 1, Detection wavelength 230nm, 25 DEG C of column temperature.
The gradient elution program of table 1
(5), determination method：Precision draws reference substance solution and each 20 μ 1 of need testing solution, injects liquid chromatograph, determines,
Four kinds of contents of sesquiterpenoids, four kinds of content and containing for active component of sesquiterpenoids are calculated respectively
Amount.
(6), assay result：See the table below 2
The content of sesquiterpenoids composition in the active component that the different preparation methods of table 2 are obtained
Note：Sesquiterpenoids composition is mainly curdione, zedoarondiol, curcumadiol, original in above active component
Four kinds of rcumenol.
Conclusion：Above testing result shows that present invention process has practical significance.
Test examples：
Present invention also offers the Radix Curcumae active components, curdione, zedoarondiol prepared with 5 kinds of methods,
The application of curcumadiol and procurcumenol in preventing and treating myocardial ischemia, coronary heart disease and hyperlipidemia is prepared.
Pharmacological evaluation shows：Curcumadiol, procurcumenol, curdione, zedoarondiol and Radix Curcumae active component
Expeirmental myocardial ischemia model to SD rat coronary artery caused by ligature has significant therapeutic effect, by rating model animal
Myocardial infarct size and CK and LDH confirm that medicine can be obviously reduced rat myocardium from injury myocardial infarction area scope caused by ischemic,
Coronary ligation myocardial ischemia in rats is damaged has significant protective effect.
Gavage gives Radix Curcumae active component and can substantially reduce Hyperlipidemia Rabbits liver and serum TG, TC values, reduces
HDL-C, LDL-C, raise H/L, can improve its hepatic steatosis degree, and the hepatic steatosis degree of each test group animal is bright
Aobvious to mitigate, its effect for reducing fat is substantially suitable with Simvastatin.
(1), lipid-loweringing experiment：
1st, test sample：Radix Curcumae active component (derives from Shanghai Bajiayi Pharmaceutical Technology Co., Ltd.)
2nd, positive reference substance I：Fufang Yiganling tablets (derive from Jiangsu Zhongxing Pharmaceutical Co., Ltd.)
Lot number：130303161；Specification：0.32g/ pieces, every containing silymarin 21mg
3rd, positive reference substance II：Simvastatin Tablets (derive from Hangzhou Mo Shadong pharmaceutical Co. Ltds)
Lot number：120333；Specification：20mg/ piece × 7 piece/box
4th, negative control：CMC-Na (Chemical Reagent Co., Ltd., Sinopharm Group)
Lot number：F20091021 (is configured to 0.5% solution to use) with test water
5th, experimental animal
Germline：New zealand rabbit (gives birth to prosperous experimental animal cultivation Co., Ltd in Shanghai)
Credit number：SCXK (Shanghai) 2012-0007；Quality certification number：200700071847
6th, experimental technique
Male New Zealand White Rabbit 100, original body mass：2,188.4 ± 227.9g, after adaptability is fed 2 weeks, except normal
Outside control group fed standard feed, remaining animal daily morning quantitatively gives high lipid food 150g.It is continuous to feed to after 33 days, every
Curfew eats, and ear edge vein exploitating blood separates serum and determines TG, TC, HDL-C, LDL-C, to confirm modeling success or not.
After confirming model success, the underproof animal of blood lipid level is rejected, according to These parameters and body weight by animal pattern
Equilibrium divides to model control group, active component administration group obtained in method 1~5 (suitable 10g crude drugs/kg), Fufang Yiganling tablets
Group (30mg/kg), Simvastatin group (3mg/kg), in administration is started after packet, blank control group and model group gavage give
4mL/kg blank solvents, remaining group gavage gives corresponding sample, and once a day, successive administration to positive controls occurs obvious
Effect for reducing fat.Do not stop high lipid food during administration.
Claim a body weight during experiment weekly, dosage is adjusted according to body weight.After being administered 2 weeks, animal fasting overnight is not
Prohibit water 12h, ear vein blood sampling, centrifugation serum detects TG, TC, HDL-C, LDL-C value, and calculate HDL-C/LDL-C ratios
Value；After being administered 3 weeks, fasting blood sampling overnight again determines These parameters, second day 3% yellow Jackets of rabbit after blood sampling
(30mg/kg) intraperitoneal anesthesia, solution takes liver 400mg, stuffing matter extract solution (normal heptane：Isopropanol=2：3.5) 4mL homogenate,
Mechanical shaking extraction lipid, takes supernatant and determines liver TG and TC after centrifugation.
7th, statistical procedures：All continuous datas withRepresent, the statistical analysis of data are checked with t；
8th, experimental result
Each experimental group Rabbit Liver lipid content experimental result is shown in Table 3.Result shows that model group hepatic tissue TG, TC content is equal
Apparently higher than Normal group (p<0.01) modeling success, is illustrated.Radix Curcumae active component each group can obviously reduce hepatic tissue TG
With TC contents.
Each experimental group rabbit anteserum TG, TC experimental result is shown in Table 4.Result shows, before administration, each administration group and model group
Serum TG, TC values highly significant are higher than Normal group (p<0.01)；And the serum TG of each administration group, TC values and model group are basic
Unanimously, prompting modeling success, packet is uniform.
Each experimental group rabbit HDL-C, LDL-C content and ratio (H/L) experimental result are shown in Table 5.Result shows, before administration,
Each experimental group Serum HDL-C and the equal highly significant of LDL-C contents are higher than Normal group (p<0.01)；H/L ratio highly significants
Less than Normal group (p<0.01)；The Serum HDL-C of each administration group, LDL-C and H/L ratios and model group are basically identical, carry
Representation model success animal packet is more uniform.The 2nd week and the 3rd week after administration, HDL-C, LDL-C highly significant of model group are higher than
Normal group (p<0.01), H/L highly significants are less than normal group (p<0.01).
9th, conclusion
Above experimental result is pointed out, after administration terminates for 3 weeks, Radix Curcumae active component (suitable 10g lifes prepared by five kinds of methods
Medicine/kg) and Fufang Yiganling tablets 30mg/kg gastric infusions can substantially reduce Hyperlipidemia Rabbits liver and serum TG, TC values, drop
Low HDL-C, LDL-C, raise H/L, can improve hepatic steatosis degree, and each test group animal of Radix Curcumae active component liver
Dirty steatosis degree mitigates.Simvastatin group 3mg/kg gastric infusions can obviously reduce high fat of blood rabbit anteserum TG, TC,
HDL-C, LDL-C value, raise H/L, also have obvious reduction to act on to liver TG, TC.Result is pointed out on this model, and Radix Curcumae has
Effect position can reduce animal lipid level, and its effect for reducing fat is substantially suitable with Simvastatin.
Each experimental group Rabbit Liver tissue lipid content results of table 3.
*p<0.05；**p<0.01, compare with model group；#p<0.05；##p<0.01, compare with Fufang Yiganling tablets group；△p<
0.05；△△p<0.01, compare with Simvastatin group
Each experimental group rabbit lipids contents result of table 4.
*p<0.05；**p<0.01, compare with model group；#p<0.05；##p<0.01, compare with Fufang Yiganling tablets group；△p<
0.05；△△p<0.01, compare with Simvastatin group.
Each experimental group rabbit anteserum high low density lipoprotein cholesterol level of table 5. and ratio result
*p<0.05；**p<0.01, compare with model group；#p<0.05；##p<0.01, compare with Fufang Yiganling tablets group；△p<
0.05；△△p<0.01, compare with Simvastatin group.
(2), to the therapeutic action of myocardial ischemia-reperfusion rat
1st, tested material
Radix Curcumae active component (the letter below prepared with the inventive method one, method two, method three, method four, method five
Title method one is made, method two is made, method three is made, method four is made, the system of method five) and with the inventive method six prepare four sesquialters
Terpene monomeric compound curcumadiol, procurcumenol, curdione, zedoarondiol, face the used time is configured to CMC-Na solution
The liquid of required concentration, for animal administrable.
2nd, positive drug
DIAOXINXUE KANG JIAONANG (hereinafter referred to as Sodium Ferulate)：Chengdu Di Aozhi Group Co., Ltd produces, specification 100mg/
Grain, lot number 1306022.Face the used time is configured to the liquid of required concentration and is administered for animal with CMC-Na solution.
Isosorbide Mononitrate piece (hereinafter referred to as single nitre ester), the glad health of trade name, Lunan Beite Pharmaceutical Co., Ltd.'s production,
Specification 20mg, lot number 07130313.Face used time grinding, with CMC-Na solution be configured to the liquid of respective concentration for rat oral gavage to
It is medicinal.
3rd, medicine box and reagent
Lactic dehydrogenase (LDH) assay kit, Zhongsheng Beikong Biological Science ＆ Technology Co., Ltd.'s production, lot number
130521。
Rat creatine kinase isozyme (CK-MB) ELISA (ELISA) kit, Bio-Swamp Products,
Lot number 201310.
Rat lactic dehydrogenase (LDH) ELISA (ELISA) kit, Bio-Swamp Products, lot number
201310。
Chloraldurate, Tianjin Kermel Chemical Reagent Co., Ltd., lot number 20120110.
4th, animal
SD rats, SPF grades, male, 253.9 ± 10.4 (230~276) g are had by Beijing dimension tonneau China experimental animal technology
Limit company provides, animal productiong licensing SCXK (capital) 2012-0001.
5th, prepared by model
Rat, under conditions of the chloraldurate 360mg/kg general anesthesias of intraperitoneal injection 12%, dorsal position is fixed on operating table.
Outside operating field unhairing, routine disinfection before chest.Skin, muscle layer are cut, muscle layer row purse string suture, the intercostal space of left side the 4th is opened
Chest, heart is pulled out with loop-hook.Branch of coronary artery position at 3~4mm below left auricle of heart, with 6/0 not damaged silk thread threading,
Juxtaposition diameter 1.6mm nylon wires, are ligatured together with ADC, and the other end is stayed in external standby.Heart is put back to
Thoracic cavity, emptying chest chamber air, closing thoracic cavity (sham-operation group animal only in corresponding coronary artery position threading, is not ligatured, other
Surgical procedure is identical with ligation animal).Carefully removing nylon wire after ligation 30min carries out Reperfu- sion.Suture muscle layer and skin, flesh
Note penicillin, once a day, for three days on end.
Compare with normal II lead electrocardiogram (ECG-II), ST sections of 10min after coronary ligation, ECG-II raises 0.2mV
Above person is used as ischemic success；10min after Reperfu- sion, the ST raised in ECG-II sections decline more than 30% person as Reperfu- sion into
Work(.The animal for being unsatisfactory for above-mentioned ischemic or ischemia-reperfusion standard shows that moulding is unsuccessful, is rejected from experiment.
6th, it is grouped and is administered
Choose the successful animal of moulding and be randomly divided into 13 groups, every group 10.Respectively sham-operation group, model control group, receive
Examination medicine group, and Chinese medicine positive drug group, Western medicine positive drug group.
30min starts gastric infusion after moulding success, once a day, continuous 7 days；Each administered volume is 10ml/kg.It is false
Operation group and model control group give solvent control liquid (CMC-Na solution)；Reagent group give respectively with the inventive method one,
Radix Curcumae active component and four sesquialters with the preparation of the inventive method six prepared by method two, method three, method four, method five
Terpene monomeric compound curcumadiol, procurcumenol, curdione, zedoarondiol, dosage (press active component and monomer are effective
The yield of composition is converted to suitable crude drug amount) it is 9g crude drugs/kg and Western medicine Isosorbide Mononitrate piece, dosage is 0.0062g
(piece weight)/kg (equivalent to clinical equivalent amount)；Sodium Ferulate 0.144g/kg (equivalent to clinical equivalent amount).
7th, infarction size measurement
After experiment terminates, rat heart is taken, with normal saline flushing, -20 DEG C carry out freezing processing.By the heart under ligature
It is dirty to be uniformly cut into 2mm slabs, 37 DEG C of dyeing 10min of 2%TTC are put, account for a left side through Pathologic image analysis system measurement infarcted region
The percentage of ventricle.
8th, Serum fibrosis markers detection
After experiment terminates, through abdominal aortic blood, 4 DEG C, 3000rpm × 10min centrifugations take upper serum, Elisa to rat
Method determines LDH, CK-MB.
9th, statistical procedures
All data are represented as mean ± standard deviationRepresent, matched before and after administration or compare use between two groups
Student ' s t-test, P<0.05 is that difference is statistically significant.
Table 6 to myocardial infarct size after myocardial ischemia-reperfusion injury influence (N=10)
Note：1. compare with sham-operation group：△△△P<0.001；2. compare with model group：*P<0.05；3. with positive drug group ratio
Compared with：P>0.05.
10th, to the influence of Serum fibrosis markers
7 days after model group ischemia-reperfusion, Serum LDH activity is significantly raised.The effective portion of Radix Curcumae prepared by 5 kinds of methods
Behind position and separate four monomer sesquiterpenoids (equivalent to 9g crude drugs/kg) treatment 7d, compared with model group, can show
Writing reduces LDH (P<0.05).The results are shown in Table 7.
Table 7 to myocardium enzyme after myocardial ischemia-reperfusion injury influence (N=10)
Test result indicate that：Medicine can be obviously reduced rat myocardium from injury myocardial infarction area scope caused by ischemic, to coronary artery
Banded Rats treating myocardial ischemia damage has significant protective effect.
Claims (1)
1. curdione, zedoarondiol, curcumadiol or procurcumenol are preparing preventing and treating myocardial ischemia, coronary heart disease or fat high
Application in mass formed by blood stasis medicine.
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