CN101759680A - Kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and preparation method thereof - Google Patents
Kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and preparation method thereof Download PDFInfo
- Publication number
- CN101759680A CN101759680A CN200910248888A CN200910248888A CN101759680A CN 101759680 A CN101759680 A CN 101759680A CN 200910248888 A CN200910248888 A CN 200910248888A CN 200910248888 A CN200910248888 A CN 200910248888A CN 101759680 A CN101759680 A CN 101759680A
- Authority
- CN
- China
- Prior art keywords
- daidzein
- dioxo
- group
- dzv
- acetic acid
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Images
Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention belongs to the field of medical technology and relates to a kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and a preparation method thereof; the compound has the following formula, wherein R1 and R2 can be the same of different and are chosen from H, metal ions, organic cations, basic amino acid cations, C1-C10 alkyl group independently; R3 and R4 can be the same or different and are independently chosen from H or C1-C10 alkyl group; the kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound, and a carrier or excipient which can be acceptable medically are combined to form medicine composite; the compound and the medicine composite has the advantages of cerebral ischemia resistance, oxygen-poor resistance, myocardial ischemia resistance, antagonistic dysmnesia resistance, platelet aggregation resistance and thrombopoiesis resistance and protecting rat cortical neuron damage caused by oxygen deficit.
Description
Technical field
The present invention relates to medical technical field; relate to a kind of kudzu-vine root daidzein-7,4 '; the application in the medicine of rat layer neuronal damage due to anti-cerebral ischemia, anoxic, myocardial ischemia, anoxic, antagonism dysmnesia, platelet aggregation-against and antithrombotic formation, the protection anoxic of 4 '-dioxo acetic acid (DZV), the pharmaceutical composition that comprises this compound and new medical use thereof, particularly this compound and composition thereof.The invention still further relates to and utilize Daidzein to prepare the soluble derivative kudzu-vine root daidzein-7,4 ', the synthetic method of 4 '-dioxo acetic acid compound.
Background technology
Daidzein is fabaceous effective constituents such as the Chinese medicine root of kudzu vine and soya bean, prove that through a large amount of pharmacology and clinical study it has tangible coronary artery dilator, the cerebrovascular, peripheral blood vessel and capillary blood vessel, increase vascular flow and circulation, reduce vascular resistance reducing cholesterol and blood viscosity, strengthen myocardial contraction, reduce myocardial consumption of oxygen and β-adrenoceptor retardance, effects such as isoflavones plant hormone.Thereby effective prophylactic treatment cardiovascular disease, diseases such as climacteric syndrome and osteoporosis.
But Daidzein belongs to isoflavonoid, the constructional feature of himself determines it to have a bigger shortcoming and defect, promptly water-soluble extreme difference, and bioavailability is low, thereby limited route of administration and formulation, greatly influenced clinical efficacy and administration scope.Therefore, Daidzein is carried out the part modify, when keeping or strengthening drug effect, improve that it is water-soluble significant.
In order to solve the problem of Daidzein poorly water-soluble, people attempt to improve by Daidzein is derived that it is water-soluble.
CN1296947A discloses some Daidzein novel derivatives.Among the disclosed therein formula I, 7,4 ' derivative of upward with carboxylic acid group who contains 1-4 carbon atom and corresponding ester thereof Daidzein being derived and obtaining at Daidzein disclosed.But this application does not specifically provide compound of the present invention.And when having the dioxo acetic acid group on 7,4 ' of Daidzein, can significantly improve the water-soluble of parent compound.
CN101255149A discloses a kind of 7,4 '-two (monomester succinate) oxygen oxyethyl group-Daidzein (DZ5), and still, the performance of this invention compound is not remarkable.
Summary of the invention
The object of the present invention is to provide a kind of Daidzein novel derivative kudzu-vine root daidzein-7,4 ', 4 '-dioxo acetic acid compound, it is compared with Daidzein, water-soluble big, the bioavailability height, and its pharmacologically active is big, and toxic side effect is little, the medicine of a kind of novel prevention and treatment diseases of cardiovascular and cerebrovascular systems be can be used to prepare, recessive heart trouble, stenocardia, myocardial infarction, myocardiosclerosis, sudden cardiac death and cerebral apoplexy etc. are used for the treatment of.
The applied kudzu-vine root daidzein-7,4 ' of the present invention, 4 '-dioxo acetic acid compound are to be parent with the Daidzein, adopt semisynthetic method to carry out 7,4 ' and replace the Daidzein novel derivative that forms.General structure is as follows:
Wherein R1 and R2 can be identical or different, and are independently from each other H, metal ion, organic cation, basic aminoacids positively charged ion, C1-C10 alkyl, and preferentially are selected from H
+, Na
+, K
+, Li
+, Mg
2+, Al
3+, Fe
2+, Fe
3+, Ca
2+, Mn
2+, Cu
2+, Zn
2+, Ba
2+, Sn
2+, Methionin, arginine, Histidine, NH4
+, methyl, ethyl, propyl group, butyl, isobutyl-, amyl group, isopentyl, basic, heptyl.R3 and R4 can be identical or different, and are independently from each other H or C1-C10 alkyl, and preferentially are selected from H, methyl, ethyl, propyl group, butyl, isobutyl-, amyl group, isopentyl, basic, heptyl.
Synthetic method step of the present invention is as follows:
(1) in the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 2.76-13.8g, potassiumiodide 0.02-3.32g, ethyl chloride 2.45-6.13g, add among the organic solvent 10-200ml, 30 ℃ of-100 ℃ of water-bath 2-15h, organic solvent is removed in distillation, washing, filter oven dry;
(2) get above-claimed cpd and be dissolved among the dehydrated alcohol 10-100ml, add the acid of equimolar amount, hydrolysis under 0 ℃ of-100 ℃ of water-bath obtains target compound, filters, and drying is used 80% ethyl alcohol recrystallization.
Behind this compound that obtains free acid form, those skilled in the art can convert it into various base addition salt or ester with method well known in the art.
In the inventive method, the used organic solvent of step (1) can use polar solvent or non-polar solvent, preferably uses through the exsiccant solvent.As tetrahydrofuran (THF), ethyl acetate, acetone, pyridine, N, dinethylformamide, methylene dichloride, trichloromethane etc.
The inventor is by mouse normal pressure hypoxia tolerance experimental model, chmice acute cerebral ischemia experimental model, mouse KCN poisoning experimental model, mouse NaNO
2Poisoning experimental model, learning and memory experimental model, mouse bilateral common carotid arteries and vagus nerve ligature experiment model, to the action model of rat layer neuronal damage due to the anoxic, influencing platelet aggregation-against experiment (turbidimetry) model in model, external platelet aggregation-against experiment (turbidimetry) model, the body, the thrombotic model etc. of influencing of rabbit arteria carotis communis carried out pharmacology activity research to DZV to rat heart muscle ischemic due to the Pituitrin.
In order to show the beyond thought excellent results of this compound, the present invention is with itself and Daidzein soluble derivative 7,4 '-two (monomester succinate) oxygen oxyethyl group-Daidzein (DZ5), kudzu-vine root daidzein-7,4 '-oxo acetic acid (DZd), kudzu-vine root daidzein-7,4 ', 4 '-dioxo propionic acid (DZe) compares.The result shows that DZV can the significant prolongation decapitated mice breathe the time, makes KCN poisoning mice survival time significant prolongation, points out this medical instrument that anti-acute cerebral ischemia is arranged, and reduces the effect of brain oxygen consumption power consumption.In addition, this medicine mouse memory obstacle due to can antagonism Scopolamine, effect with hypermnesis.DZV can also significant prolongation normal pressure hypoxia tolerance condition under and Racemic isoproterenol induce the survival time of acute myocardial anoxia model mice, and can resist effectively that Racemic isoproterenol causes that myocardial consumption of oxygen increases and the myocardial ischemia-anoxemia that causes improves heart function.Platelet aggregation test is the result show: DZV can significantly suppress AA inductive platelet aggregation, thereby suppresses hematoblastic activation, plays antithrombotic effect.Simultaneously, also has the effect of rat layer neuronal damage due to the protection anoxic.And its every pharmacology index all is better than or is equivalent to other derivatives under same dose.
The present invention has also investigated the toxicity of DZV and has compared with DZ5, and has investigated DZV in its pharmacokinetics in rats parameter, for clinical application is offered help.
Another object of the present invention provides the pharmaceutical composition of a kind of DZV of containing and pharmaceutical acceptable carrier or vehicle.Described pharmaceutical composition has anti-cerebral ischemia, anoxic, myocardial ischemia, anoxic, and the antagonism dysmnesia, platelet aggregation-against and antithrombotic form, effect such as rat layer neuronal damage due to the protection anoxic.
Compare with compound in the prior art, compound water soluble of the present invention is big, the bioavailability height, and pharmacologically active is big, and toxic side effect is little, and performance significantly improves, and its preparation method is simple, favorable reproducibility.
Description of drawings:
Fig. 1 is a daidzein-7,4 ', the UV scanning collection of illustrative plates of 4 '-dioxo acetic acid sodium (water)
Fig. 2 is a daidzein-7,4 ', the infrared figure of 4 '-dioxo acetic acid sodium
Fig. 3 is a daidzein-7,4 ', the mass spectrum of 4 '-dioxo acetic acid sodium
Fig. 4 is a daidzein-7,4 ', the 13C-NMR spectrum of 4 '-dioxo acetic acid sodium
Fig. 5 is a daidzein-7,4 ', the 1H-NMR spectrum of 4 '-dioxo acetic acid sodium
Embodiment:
Describe the present invention in detail by following embodiment.But should be appreciated that these embodiment just illustrate the present invention, rather than in office where face limits the scope of the invention.
Melting point compound is measured the scanningcalorimeter with Shimadzu DSC-60 differential in this experiment; Nucleus magnetic resonance adopts Bruker ARX-300 type nuclear magnetic resonance analyser, interior mark TSP-d4; Mass spectrum adopts Agilent 1100 SL type ion trap mass spectrometers; The ultraviolet full wavelength scanner adopts Unico 4802HUV/Vis double beam spectrophatometer; Infrared employing Bruker IFS 55 infrared analysis instrument.
Used chromatographic condition is as follows among the following embodiment: chromatographic column: Spherisorb C
18Post (10 ц m, 4.6 * 250mm) the Dalian Chemistry and Physics Institute; Moving phase: methyl alcohol: water: phosphoric acid (63: 37: 0.05v: v: v); Flow velocity: 0.8mlmin
-1Ultraviolet detection wavelength: 275nm; Column temperature: 25 ℃.
Embodiment 1: daidzein-7,4 ', the preparation of 4 '-dioxo acetic acid
In the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 2.80g, potassiumiodide 3.32g, ethyl chloroacetate 3.66g adds in the 100ml anhydrous propanone, back flow reaction 8h, acetone is removed in distillation, and washing filters drying; Get above-claimed cpd and be dissolved in the 30ml dehydrated alcohol, add the HCL of equimolar amount 1mol/L, back flow reaction 5 hours is filtered, and drying obtains white powder, uses 80% ethyl alcohol recrystallization, obtains mp274.91 ℃, white plates crystallization 3.33g, yield 90%.
Embodiment 2: daidzein-7,4 ', the preparation of 4 '-dioxo acetic acid sodium
In the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 1.38g, potassiumiodide 3.32g, ethyl chloroacetate 2.45g adds the anhydrous N of 20ml, in the dinethylformamide, 80 ℃ of water-bath 10h, washing filters, oven dry; Get above-claimed cpd and be dissolved in the dehydrated alcohol, add the hydrochloric acid soln of equimolar amount, obtain daidzein-7,4 ', 4 '-dioxo acetic acid filters drying; Get above-claimed cpd and be dissolved in the 50ml dehydrated alcohol, drip the sodium hydroxide solution of equimolar amount, filter, drying obtains white amorphous powder 3.64g (mp＞350 ℃), yield 88%.UV (water): λ max=249nm, 302nm, IR (KBr) cm
-1: 3419.6 (OH), 1638 (C=O); MS:[M-H]
-: 368.7, [M+H]
+: 370.9;
1H-NMR:4.41-4.85 (m, 4H), 6.77 (d, 1H), 6.90-6.93 (d, 2H), 6.95-6.99 (m, 1H), 7.24-7.27 (d, 2H), 7.83-7.86 (d, 1H), 7.92 (s, 1H).Its ultraviolet, infrared, mass spectrum, nuclear-magnetism figure sees attached sheet.
Embodiment 3: daidzein-7,4 ', the preparation of 4 '-dioxo acetic acid ethyl ester
In the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 2.76g, potassiumiodide 0.25g, ethyl chloroacetate 7.35g adds in the anhydrous trichloromethane of 100ml 70 ℃ of water-bath 10h, washing, filter, oven dry obtains white plates crystallization 3.62g, mp=167.51 ℃, yield 85%.
Embodiment 4: daidzein-7,4 ', the preparation of 4 '-dioxo acetic acid ammonium
In the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 1.38g, potassiumiodide 3.32g, ethyl chloroacetate 2.45g adds in the 150ml methylene dichloride, water-bath back flow reaction 15h, washing filters, oven dry; Get above-claimed cpd and be dissolved in the dehydrated alcohol, add the hydrochloric acid soln of equimolar amount, obtain daidzein-7,4 ', 4 '-dioxo acetic acid filters drying; Get above-claimed cpd and be dissolved in the 100ml dehydrated alcohol, drip the ammoniacal liquor of equimolar amount, filter, drying obtains white amorphous powder 3.64g (mp＞350 ℃), yield 90%.
Embodiment 5: daidzein-7,4 ', the arginic preparation of 4 '-dioxo acetic acid
In the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 6.90g, potassiumiodide 3.32g, ethyl chloroacetate 6.13g adds in the 150ml methylene dichloride, water-bath back flow reaction 15h, washing filters, oven dry; Get above-claimed cpd and be dissolved in the 60ml dehydrated alcohol, add the hydrochloric acid soln of equimolar amount, obtain daidzein-7,4 ', 4 '-dioxo acetic acid filters drying; Get above-claimed cpd and be dissolved in the dehydrated alcohol, drip the arginine solution of equimolar amount, filter, drying obtains white amorphous powder 4.61g (mp＞350 ℃), yield 85%.
By the following examples the present invention is described further in the effect aspect pharmacodynamics and the pharmacokinetics, preceding 7 examples are cerebral ischemia experimental models, and 8 to 11 is myocardial ischemia experimental models:
Experimental animal: Kunming mouse, the male and female dual-purpose, mouse age is 6-8 week, body weight is 20 ± 2g; SD is male, 180-220g, male and female dual-purpose; White big ear rabbit, 2.0-2.5kg, the male and female dual-purpose is purchased the animal center in Shenyang Pharmaceutical University, animal subject pre-treatment: in receptacle, observe a week, room temperature 21-23 ℃, relative humidity 30-70%, edible Shenyang Pharmaceutical University animal center provides mechanism material piece, drink tap water, natural lighting.Embodiment 6:DZV is to the influence of chmice acute cerebral ischemia
60 mouse are divided into 6 groups at random, every group 10, male and female half and half, irritate blank medicine-physiological saline, DZV, DZ5, DZd, DZe300mg/kg, the positive control drug-nimodipine (120mg/kg) of stomach same dose respectively, administration every day 1 time, successive administration 7d, behind the last administration 1hr, break end fast from the mouse ear rear portion, record is from the time that the broken end beginning is once panted and stopped to the end, and T check carrying out group difference relatively.(the results are shown in Tab1.)
Tab?1.Effect?of?DZ?V、DZ5、DZd?and?DZe?on?persistent?time?of?gasping?of?isolated?heads?of?miceby?decapitation.(Mean?±SD，n＝10)
"
*" expression P＜0.05 "
*" P＜0.01 "
* *" P＜0.001
The result shows, positive control drug, and DZV, DZd and DZ5 all can prolong breathing the time length of decapitated mice to some extent, and positive control drug and derivative DZV effect be (P＜0.001) significantly.DZV (P＜0.001) effect significance is greater than DZ5, and DZd organizes, and DZe organizes no prolongation effect.
Embodiment 7:DZV is to the influence of mouse normal pressure hypoxia tolerance
60 mouse are divided into 6 groups at random, grouping, medication are with experiment 1, successive administration 7d, behind the last administration 40min, mouse is put into 250ml ground wide-necked bottle, and (bottle is placed with the 15g sodica calx, loam cake filter paper) the bottle cap lid is tight, the record mouse survival time, carry out group difference relatively with the T method of inspection.(the results are shown in Tab2.)
Tab2.Effect?of?DZV、DZ5、DZd?and?DZe?on?Consurvival?time?of?mice?subjected?to?normobarichypoxia.(Mean±SD，n＝10)
The result shows, positive control drug, DZV and DZ5 group have in various degree prolongation effect to the survival time of mouse under the normobaric hypoxia situation.Wherein DZV group and nimodipine group are comparatively obvious, and DZV group significance is organized greater than DZ5, and DZd group and DZe group do not have the significance effect.
The influence that embodiment 8:DZV poisons to mouse KCN
60 mouse are divided into 6 groups at random, the experiment 1 together of grouping, medication, successive administration 7d, last administration is after 1 hour, and tail vein injection KCN, dosage are 5mg/kg.Record mouse survival time and survival rate are carried out group difference relatively with the T method of inspection.(the results are shown in Tab3.)
Tab3.Effect?of?DZV、DZ5、DZd?and?DZe?on?survival?time?of?mice?by?KCN(Mean±SD，n＝10)
The result shows that the effect of DZV group is (P＜0.001) the most obviously, and positive control drug, DZ5 and DZV, DZd, DZe group all can prolong the survival time of KCN poisoning mice in various degree.DZV group significance is greater than DZ5, DZd, DZe group.The animals survived that 10%--40% is arranged.
Embodiment 9:DZV is to the influence of learning and memory of little mouse
105 mouse are divided into 7 groups at random, 15 every group, be male, be respectively blank 1, blank 2, positive controls, DZV, DZ5, DZd and DZe 300mg/kg group.Begin training behind the gastric infusion 3d, train 10 every every day, trains 3d continuously.30min after the last administration, blank 2, positive controls, administration group be abdominal injection Scopolamine (1ml/kg) respectively, begins test after 30 minutes, surveys 5 times the record mouse wrong times for every.Carry out group difference relatively with the T method of inspection.(the results are shown in Tab4.).
Tab4.Effect?of?DZV、DZ5、DZd?and?DZe?on?the?memory?in?mice
" ^^^ " expression with the blank group than P＜0.001 "
*" expression with model group than P＜0.05 "
*" P＜0.01 "
* *" P＜0.001
The result shows, positive control drug, DZV, DZ5 and DZd have improvement effect in various degree to Scopolamine induced mice dysmnesia, mouse wrong times is reduced, the effect of DZV group and positive controls is (P＜0.001) significantly, significance all greater than DZ5 group and DZd group, points out this medicine to the obvious improvement effect of having of mouse memory obstacle.DZe organizes no obvious improvement effect.
The influence that embodiment 10:DZV poisons and tests mouse NaNO2
Get 60 of mouse, be divided into 6 groups at random, 10 every group, male and female half and half, the experiment 1 together of grouping, medication, ip.NaNO2 800mg/kg after 10 minutes after the last administration, the survival time of observing mouse is dead index with breath stopped.(the results are shown in Tab 5.).
Tab?5.Effects?of?DZ?V、DZ5、DZd?and?DZe?on?survival?time?in?the?intoxicant?mice?of?NaNO
2(Mean±SD，n＝10)
Compare with NS
*P＜0.05,
*P＜0.01
Experimental result shows that DZV group, DZ5 group, DZe group and positive controls be the survival time of energy significant prolongation NaNO2 poisoning mice all, relatively has significant difference with the physiological saline control group.
Embodiment 11:DZV is to the effect of rat layer neuronal damage due to the anoxic
The rat of conceived 17-19d is taken off neck execution, and aseptic condition takes out the tire mouse down, takes out tire mouse brain before the super clean bench, places the DMEM nutrient solution of ice bath, carefully rejects pia mater and blood vessel, the separation pallium.Isolating pallium is shredded, in 37 ℃ of water-baths, digest 5min with 0.25% pancreatin, it is centrifugal that (1000r 5min) abandons supernatant liquor, blows and beats gently with IMDM nutrient solution (containing 10% foetal calf serum) and is dispersed into cell suspension, cell suspension filters, the adjustment cell concn is 1 * 106cellmL-1, with cell inoculation in 96 well culture plates that covered with 0.01% poly-lysine in advance, every hole 100 μ L, put 37 ℃, hatch in the 5%CO2 incubator.Renew the bright IMDM nutrient solution that is added with 1% cytosine arabinoside behind the 18-22h, behind 24h, change 50% again and contain serum I MDM nutrient solution.Later on change the fresh serum I MDM nutrient solution that contains 1 time every 1d, cell cultures is to 5-6d.
Experiment is divided into blank, model group and DZ, DZV, DZ5, DZd and DZe different concns group.Each group all is changed to the IMDM nutrient solution of serum-free during experiment, the administration group adds different concns DZ or DZV places the 5%CO2 incubator to hatch, the blank group is changed to Earle ' the s liquid that contains glucose behind the 24h, the anoxia model group is changed to Earle ' the s liquid that does not contain glucose, then culture plate is placed in the anoxic jar, feed 95%N2 and 5%CO2 mixed gas 20min.Behind the 24h, every hole adds MTT solution 10 μ l, and 37 ℃ of 5%CO2 incubators are hatched 4h, sucking-off stoste, and every hole adds DMSO solution 100 μ l again, and the 10min that vibrates slightly measures optical density value with microplate reader 492nm wavelength.
Experimental data adopts the SPSS13.0 statistical software to analyze, and each organizes data, and (X ± SE) expression adopts One-Way ANOVA to estimate globality difference, and relatively, P＜0.05 is for there being significant difference between organizing with LSD with the mean value standard error.
Tab 6. is seen in influence to normal neurons, and Tab 7. is seen in neuronic influence to anoxic.
Tab?6.Effects?of?DZV、DZ5、DZd?and?DZe?on?the?viability?of?primary?cultured?neurons(Mean±SD)
Tab?7.Effects?of?DZV、DZ5、DZd?and?DZe?on?the?injure?of?primary?cultured?neurons?induced?byhypoxia.(Mean±SD)
#P＜0.05，##P＜0.01，###P＜0.001，compared?with?the?control?group；
*P＜0.05，
**P＜0.01，
***P＜0.001，compared?with?the?model?group
Embodiment 12: mouse bilateral common carotid arteries and vagus nerve ligature experiment
Get 60 of mouse, be divided into 6 groups at random, 10 every group, male and female half and half, grouping, medication are with experiment 1.The survival time of observing mouse was dead index with breath stopped with No. 0 silk thread ligation bilateral common carotid arteries and vagus nerve in 40 minutes after the administration.(the results are shown in Tab 8.).
Tab?8.Effects?of?DZV、DZ5、DZd?and?DZe?on?survival?time?ofmice?subjected?to?bilateral?carotidartery?ligation.(Mean±SD，n＝10)
Compare with NS
*P＜0.05
*P＜0.01
Experimental result shows that DZV group, DZ5 group, DZd group and positive controls be the survival time of energy significant prolongation vagus nerve ligation mouse all, relatively has significant difference with the blank group.And DZV organizes time expand greater than DZ5 group and DZd group, and DZe organizes no obvious prolongation effect.
Embodiment 13: to the influence of rat heart muscle ischemic due to the Pituitrin
Get 50 of healthy SD rats, male and female are not limit, and are divided into 5 groups at random: physiological saline (NS) group, DZ5 group, DZd group, DZe group, DZV 75mg/kg intravenous administration.After 25% urethane 1g/kg ip. anesthesia, face upward the position and be fixed on the mouse platform, sting in four limbs subcutaneously with No. 6 syringe needles, and link to each other with electrocardiograph limb leads electrode.Electrocardiograph choice criteria voltage 1mv=10mm, chart speed 50mm/s.What each organized tail vein iv. respective concentration respectively is subjected to reagent thing 5ml/kg, and administration was annotated in hypogloeeis iv. Pituitrin (Pit) 0.8U/kg, 5s after 10 minutes.With the ECG that reaches injection back 15s, 30s, 45s, 60s, 90s and 3min, 5min, 8min before the electrocardiograph record injection Pit.Observing the T wave height changes and changes in heart rate.T wave height and heart rate are each time point and survey 5 waveforms continuously, get its mean value.No matter the T wave height raises or reduces, and gets the absolute value of variation.Its significance is judged in the t check between experimental result work group.
(1) DZV is to the influence of T wave height variation
The result shows that behind the NS control rats iv.Pit, electrocardio takes place obviously to change, and the first phase, (0～30s) tangible S-T section occurs raised and T wave height towering (P＜0.01); The second phase (after the 30s) presents tangible S-T section and moves down, the low flat or inversion (P＜0.05 or P＜0.01) of T ripple.This experimental result conforms to other bibliographical information.The DZV group is significantly resisted ∑ ST rising or the reduction that Pit causes at each time point, and its action effect is better than the DZ5 group of same dose administration, DZd group and DZe group.(the results are shown in Tab 9.).
Tab 9.Effects of DZV, DZ5, DZd and DZe on the aptitude of T wave of acute myocardialischemia induced by PIT in rats (Mean soil SD, n=10)
Compare with pre-pit
*P＜0.05
*P＜0.01
(2) DZV is to the influence of changes in heart rate
Behind the NS group rat iv.Pit, the rat heart rate is obviously slowed down, not recover yet to 8 minutes.DZV is all obviously alleviated decreased heart rate due to the Pit at each time point, and effect is better than the DZ5 group of same dose administration, and DZd group and DZe organize.(the results are shown in Tab 10.).
Tab 10.Effects of DZ V, DZ5, DZd and DZe on the changes of heart rate of acute myocardialischemia induced by PIT in rats (Mean soil SD, n=10)
Compare with pre-pit
*P＜0.05
*P＜0.01
Embodiment 14: external platelet aggregation-against experiment (turbidimetry)
Rabbit is divided 6 groups at random, 6 every group.Ear edge vein exploitating blood 4.5mL, with 3.8% Sodium Citrate anti-freezing (blood and antithrombotics volume ratio are 9: 1), centrifugal under the room temperature (800kg 3min), gets supernatant liquor and prepares platelet rich plasma (PRP).After isolating PRP, centrifugal again (2000kg 10min) gets supernatant liquor and prepares platelet poor plasma (PPP), and regulating transmittance with PPP is 100%.Get PRP 200 μ L and place in the opacity tube, (control group adds isometric(al) NS for ASP, 100mg/ml) 20 μ L to add DZ V, DZ5, DZd, DZe75mg/ml or acetylsalicylic acid respectively.Behind the incubation 5 minutes, (AA) is inductor with arachidonic acid, traces 10 minutes and assembles curve, measures maximum aggregation intensity.The results are shown in Tab 11.
Calculate the anticoagulant percentage according to following formula:
Tab 11.Effects of DZV, DZ5, DZd and DZe on the platelet aggregation induced by AA inrabits in vitro. (Mean soil SD, n=6)
Compare with NS,
*P＜0.05,
*P＜0.01,
* *P＜0.001
Experimental result shows that DZV has obvious restraining effect external to AA inductive rabbit platelet aggregation, and effect is better than DZ5 group and DZd group.Except that the DZe group, each dosage group and NS control group relatively all have significant difference.The ASP group also has obvious suppression effect (P＜0.01) to the external rabbit platelet aggregation of AA inductive.
Embodiment 15: platelet aggregation-against experiment (turbidimetry) in the body
Rabbit is respectively from auricular vein iv.DZ V, DZ5, DZd, DZe 75mg/kg or ASP 80mg/kg, control group iv. isometric(al) NS.10min carries out the preparation of PRP and PPP from another auricular vein bloodletting respectively by the in vitro tests method after the administration, makes inductor with AA, investigates the influence of medicine to platelet aggregation.The results are shown in Tab12.
Tab 12 Effects of DZV, DZ5, DZd and DZe on the platelet aggregation induced by AA in rabitsin vivo. (Mean soil SD, n=6)
Compare with NS,
*P＜0.05,
*P＜0.01,
* *P＜0.001
Experimental result shows that DZV, ASP group, DZ5, DZd and DZe group all have obvious restraining effect to AA inductive rabbit platelet aggregation.Wherein, DZV, DZ5 significance the highest (P＜0.001), restraining effect is greater than DZd and DZe group.
Embodiment 16: to the thrombotic influence of rabbit arteria carotis communis
Rabbit is divided into 6 groups at random, 8 every group.Press 60mg/kg auricular vein injecting anesthetic with 3% talamo, back of the body position is fixing, separate a side arteria carotis communis,, get thin sewing-needle No. 12 blood vessel being clamped with bulldog clamp in two ends at a distance of about 4cm place, the fine rule that connects one one end knotting, penetrate arteria carotis communis from proximal part,, pass from distal end along blood vessel traveling 3cm, cut off fine rule after the other end also tied a knot, slowly unclamp bulldog clamp then.Respectively from opposite side auricular vein iv. physiological saline, DZV, DZ5, DZd, DZe75mg/kg and ASP80mg/kg.Recover behind the blood flow 2h this section arteria carotis communis of each group rabbit to be cut, vertically cut, take out fine rule, put on the filter paper, inhale and remove surplus blood, and weigh rapidly, deduct the fine rule own wt, promptly get wet weight of thrombus.The results are shown in Tab 13.
Calculate the thrombosis inhibiting rate according to following formula:
Tab 13 Effects of DZV, DZ5, DZd and DZe on the wet weight of thrombosis formated incommon carotid artery in rabit. (Mean soil SD, n=8)
Compare with NS,
*P＜0.05,
*P＜0.01
Experimental result shows that DZV (iv.) has obvious restraining effect to rabbit arteria carotis communis thrombosis, with control group significant difference (P＜0.01) is arranged relatively.ASP group and DZ5 group also have obvious suppression effect (P＜0.01) to rabbit arteria carotis communis thrombosis, and DZd and DZe group do not have obvious restraining effect.
The acute toxicity test of embodiment 17:DZV
Irritate the DZV of stomach maximal dose, mouse all survives, and survivor's outward appearance, action are all no abnormal, fail to measure LD50.Trying to achieve maximum tolerated dose is oral 5g/kg, quiet notes administration LD
505.50g/kg.DZ5 is by acute toxicity test, and trying to achieve maximum tolerated dose is oral 3g/kg, quiet notes administration 3.78g/kg.DZV compares with DZ5, has bigger dosis tolerata, and toxic side effect is little, and security is higher.
Embodiment 18:DZV is at the intravital pharmacokinetic study of rat
Get 5 rats, the male and female dual-purpose carries out vein persistence pipe art,, intersect to do and irritate stomach, the experiment of quiet notes (intravenous injection 15,25,35mgkg
-13 kinds of various dose).Above cross-over experiment is 1 week of interval all.After the administration 5,10,20,30,45,60,90,120,180min respectively gets blood 0.25mL, get blood plasma 0.1mL after centrifugal to insert in-20 ℃ of refrigerators to be measured.Plasma Concentration-time data is with the practical pharmacokinetics computation program 3P97 of Chinese mathematics pharmacology association establishment match Plasma Concentration-time curve automatically on computers, and calculates every pharmacokinetic parameter.
(1) intravenously administrable (iv)
The DZV injection liquid of 3 kinds of various dose of rat intravenous injection, the medicine dynamic behavior all meets two Room open models in the rat body.Its main pharmacokinetic parameter is seen Tab 14.
Tab?14?The?pharmacokinitic?parameters?after?the?jugular?vein?injection?of?3?doses?of?DZV?to?rats(Mean±SD，n＝5.)
By tab5 as seen, at intravenous injection 15,25mgkg
-1During DZV, the t1/2 of 2 kinds of various dose (β), V (c), major impetus mathematic(al) parameters such as CL are very close, learn t check there was no significant difference (P＞0.05) by statistics, and AUC increases and proportional increase with dosage, and the elimination that DZ V in this dosage range is described is a linear kinetics, but as intravenous injection 35mgkg
-1During DZV, t1/2 (β) prolongs (P＜0.05), and CL obviously reduces (P＜0.05), and AUC hypergeometric example increases, and shows heavy dose of 35mgkg
-1After the administration, this product is a nonlinear kinetics in the intravital elimination of rat.
(2) gastric infusion (ig)
5 of rats, gastric infusion DZV 120mgkg
-1After, the machine match as calculated of Plasma Concentration-time data, dynamic behavior meets one compartment open model.Its main pharmacokinetic parameter sees Table tab 15.The result shows that this medicine absorbs, eliminates all very rapid.
Tab?15?The?pharmacokinetic?parameters?of?DZ?V?after?intragastic?adminstration?torats(Mean±SD，n＝5，)
Rat records Plasma Concentration-time data and adopts compartment model and 2 kinds of methods of statistical moment to handle respectively after filling stomach, jugular vein intersection give DZV, and (D: dosage), the result of 2 kinds of methods relatively sees tab16,17 to calculate AUC and AUC/D.
Tab?16?The?bioavailability?of?DZV(compartment?model)
Tab?17?The?bioavailability?of?DZ?V(statistical?algorithm)
By tab16,17 as seen, the absolute bioavailability of rat oral gavage DZ V is 38.66%, 40.09%, 2 kind of method result is close, learns by statistics and handles, and does not have significant difference (P＞0.05) between 2, and the reasonableness of data processed result is described.
In a word, this experiment shows that DZV has preferably anti-cerebral ischemia, anoxic, resists myocardial ischemia, anoxic, the antagonism dysmnesia, and platelet aggregation-against and antithrombotic form, effects such as neuroprotective unit.Drug effect is higher than or is equivalent to other derivatives DZ5 of the Daidzein of same dose, DZd, DZe.In addition, can get from acute toxicity test, substantive features of the present invention and marked improvement are DZV when having above each drug effect, and be safe, and its oral and intravenous dosis tolerata is all greater than DZ5.This research is significant for evaluating drug effect and clinical application, for the further exploitation of DZV provides theoretical foundation.
By the following examples the present invention is described further in the application aspect the preparation.
The regular convention formula of tablet can be the DZV:10-90% weight ratio, starch 89-9% weight ratio, and Magnesium Stearate 1% weight ratio, 50% ethanol is an amount of.The regular convention formula of capsule can be the DZV:10%-100% weight ratio, starch 90-0% weight ratio, and the regular convention formula of granule can be the DZV:5%-100% weight ratio, (sucrose+dextrin) 95-0% weight ratio, sucrose: dextrin=2: 1.Those skilled in the art also can be mixed with other formulations with technology well known in the art and carrier as required simultaneously.
Embodiment 19: tablet:
DZV 30g
Starch 32.5g
Magnesium Stearate 0.5g
50% ethanol 11ml
Compressing tablet promptly behind the said components mixing granulation.
Embodiment 20: capsule:
DZV 60g
Starch 80g
The said components mixing is inserted in the capsulae vacuus, promptly
Embodiment 21: granule:
DZV 80g
Starch 230g
Dextrin 90g
Ethanol 54ml
Said components is mixed, is sieved, and mixing is made wet granular with ethanol, drying, and whole grain obtains granule.
Embodiment 22: dripping pill:
DZV 45g
PEG6000 15g
PEG4000 30g
DZV is joined mixing in the polyoxyethylene glycol auxiliary material of heating and melting; Then melting mixing liquid is moved into the dripping pill machine system of dripping; Medicine liquid droplet cools off to the whiteruss refrigerant; Blot refrigerant with suction cloth, select ball, promptly obtain the DZV dripping pill.
Embodiment 23: injection:
DZV 6g
Sodium hydroxide solution is an amount of
Hydrochloric acid soln is an amount of
Water for injection is to 100ml
After said components mixed, add injection water 80ml, be stirred to dissolving fully after, add 0.1% gac, 60 ℃ of insulated and stirred 30min, filtered while hot is removed gac, benefit adds to the full amount of water for injection.With 0.45 μ m filter membrane coarse filtration, use the filtering with microporous membrane of 0.22 μ m again.Embedding, the 15min that sterilizes in 121 ℃ of circulation steams promptly gets the DZV injection liquid.
Embodiment 24: freeze-dried preparation
DZV 6g
N.F,USP MANNITOL 5g
Sodium hydroxide solution is an amount of
Hydrochloric acid soln is an amount of
Water for injection is to 100ml
Take by weighing each component by recipe quantity, add injection water 80ml, be stirred to dissolving fully after, add 0.1% gac, 60 ℃ of insulated and stirred 30min, filtered while hot is removed gac, benefit adds to the full amount of water for injection.With 0.45 μ m filter membrane coarse filtration, use the membrane filtration of 0.22 μ m again, lyophilize promptly gets injection DZV.
Embodiment 25: oral liquid
DZV 5g
Distilled water 30ml
Simple syrup adds to 100ml
Get DZV and be dissolved in the distilled water, add simple syrup to full dose, promptly.
Embodiment 26: external preparation:
DZV 10g
Acritamer 940 4g
Glycerine 15g
Trolamine is an amount of
Distilled water is to 100ml
After getting carbomer 4g and adding the suitable quantity of water swelling, adding glycerine, grind and make moisteningly, add trolamine and grind to form clear gel matrix. other gets DZV, with the suitable quantity of water dissolving, stirs evenly.Above-mentioned soup is added in the gel matrix, and the limit edged grinds, and adding distil water continues to stir to 100ml, gets the clear gel agent.
Claims (9)
1. the kudzu-vine root daidzein-7,4 ' that has following general formula, 4 '-dioxo acetic acid compound:
Wherein R1 and R2 can be identical or different, and are independently selected from the group that H maybe can make this compound formation pharmacologically acceptable salt or ester; R3 and R4 can be identical or different, and are independently selected from H or alkyl.
2. compound according to claim 1 is characterized in that, described R1 and R2 are independently from each other H, metal ion, organic cation, basic aminoacids positively charged ion, C1-C10 alkyl; R3 and R4 are independently from each other H or C1-C10 alkyl.
3. compound according to claim 1, it is characterized in that it is to be parent with the Daidzein, adopts semisynthetic method to carry out 7,4 ' replaces kudzu-vine root daidzein-7,4 ', the free acid of 4 '-dioxo acetic acid compound or its pharmacologically acceptable salt or the ester that forms.
4. compound according to claim 1 and 2 is characterized in that: R1 and R2 are independently selected from H
+, Na
+, K
+, Li
+, Mg
2+, Al
3+, Fe
2+, Fe
3+, Ca
2+, Mn
2+, Cu
2+, Zn
2+, Ba
2+, Sn
2+, Methionin, arginine, Histidine, NH
4+, methyl, ethyl, propyl group, butyl, isobutyl-, amyl group, isopentyl, hexyl, heptyl; R3 and R4 are independently selected from H, methyl, ethyl, propyl group, butyl, isobutyl-, amyl group, isopentyl, hexyl, heptyl.
5. kudzu-vine root daidzein-7,4 ' as claimed in claim 1, the preparation method of 4 '-dioxo acetic acid compound is characterized in that comprising the steps:
(1) in the 500ml round-bottomed flask, add Daidzein 2.54g, Anhydrous potassium carbonate 2.76-13.8g, potassiumiodide 0.02-3.32g, ethyl chloride 2.45-6.13g, add among the organic solvent 10-200ml, 30 ℃ of-100 ℃ of water-bath 2-15h, organic solvent is removed in distillation, washing, filter oven dry;
(2) get above-claimed cpd and be dissolved among the dehydrated alcohol 10-100ml, add the acid of equimolar amount, hydrolysis under 0 ℃ of-100 ℃ of water-bath obtains target compound, filters, and drying is used 80% ethyl alcohol recrystallization.
6. method according to claim 5, it is characterized in that: the described organic solvent of substitution reaction of step (1) is selected from tetrahydrofuran (THF), ethyl acetate, acetone, pyridine, N, dinethylformamide, methylene dichloride, trichloromethane, reaction must be carried out under anhydrous condition.
7. a pharmaceutical composition is characterized in that it comprises kudzu-vine root daidzein-7,4 ', 4 '-dioxo acetic acid compound and pharmaceutically acceptable carrier or vehicle.
8. kudzu-vine root daidzein-7,4 ', 4 '-dioxo acetic acid compound and composition thereof be in preparation anti-cerebral ischemia, anoxic, resists myocardial ischemia, anoxic anti-dysmnesia, the application in platelet aggregation-against and anti-thrombosis drug or the healthcare products.
9. pharmaceutical composition according to claim 7 is characterized in that, it can further make injection, tablet, capsule, granule, dripping pill, oral liquid or exterior-applied formulation form.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN200910248888A CN101759680A (en) | 2009-12-30 | 2009-12-30 | Kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and preparation method thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN200910248888A CN101759680A (en) | 2009-12-30 | 2009-12-30 | Kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and preparation method thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN101759680A true CN101759680A (en) | 2010-06-30 |
Family
ID=42491076
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN200910248888A Pending CN101759680A (en) | 2009-12-30 | 2009-12-30 | Kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and preparation method thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN101759680A (en) |
Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103145673A (en) * | 2011-12-06 | 2013-06-12 | 安徽贝克生物制药有限公司 | Daidzein derivative and pharmaceutically acceptable salt thereof |
| CN105616399A (en) * | 2016-02-25 | 2016-06-01 | 北京师范大学 | Isoflavone derivative and application thereof to erythrocytic hemolysis prevention |
| CN106008440A (en) * | 2016-05-26 | 2016-10-12 | 淮海工学院 | Genistein 7-O-sodium acetate and preparation method thereof |
-
2009
- 2009-12-30 CN CN200910248888A patent/CN101759680A/en active Pending
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103145673A (en) * | 2011-12-06 | 2013-06-12 | 安徽贝克生物制药有限公司 | Daidzein derivative and pharmaceutically acceptable salt thereof |
| CN103145673B (en) * | 2011-12-06 | 2015-05-20 | 安徽贝克生物制药有限公司 | Daidzein derivative and pharmaceutically acceptable salt thereof |
| CN105616399A (en) * | 2016-02-25 | 2016-06-01 | 北京师范大学 | Isoflavone derivative and application thereof to erythrocytic hemolysis prevention |
| CN105616399B (en) * | 2016-02-25 | 2019-02-05 | 北京师范大学 | Isoflavone derivative and its application on anti-erythrocyte haemolysis |
| CN106008440A (en) * | 2016-05-26 | 2016-10-12 | 淮海工学院 | Genistein 7-O-sodium acetate and preparation method thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| JP6656345B2 (en) | Nitroxyl donors with improved therapeutic index | |
| CN106916177B (en) | A kind of deuterated dipeptide boronic acid or its ester type compound and its synthetic method and purposes | |
| IL195313A (en) | Substituted beta-phenyl-alpha-hydroxy propanoic acid synthesis method and use thereof | |
| CN109928972B (en) | Matrine derivative and application thereof in medicine | |
| EP3563873B1 (en) | Application of combination of polyethylene glycol and local anesthetic in non-narcotic analgesia | |
| ES2625329T3 (en) | Reversible asymmetric neuromuscular blocking agents of ultra-short, short or intermediate duration | |
| CN103193789A (en) | Optically active butylphthalide open-ring derivative, preparation method and medical application | |
| CN107382870A (en) | N substituted imidazole carboxylic acid ester compounds and preparation method thereof and purposes in medicine | |
| CN101759680A (en) | Kudzu-vine root daidzein-7, 4'-dioxo acetic acid compound and preparation method thereof | |
| JPS5857397A (en) | Stable s-adenosylmethionine derivative, manufacture and therapeutical composition containing same as active component | |
| JP6952911B2 (en) | 2- (α-Hydroxypentyl) benzoic acid organic amine ester derivative drug | |
| CN104447406A (en) | Alpha-cyano-4-hydroxy cinnamic acid derivative and preparation method and application thereof | |
| AU2015268575B2 (en) | Derivative of butylphthalide and preparation method and use thereof | |
| CA3140231C (en) | 3-aryloxyl-3-five-membered heteroaryl-propylamine compound, and crystal form and use thereof | |
| CN103242337B (en) | Ginkgolide B derivative based on double target spots of PAF (Platelet Activating Factor)/ROS (Reactive Oxygen Species), and preparation method of derivative | |
| CN105153049A (en) | Tanshinol amide derivative and preparation method and application thereof | |
| CN101255149B (en) | 7,4'-di(mono succinate)o-ethoxy-daidzein and novel medical uses thereof | |
| CN116554127A (en) | Piperazine substituted phenol derivative and application thereof | |
| CN109912681B (en) | Derivative containing cyclopentanoperhydrophenanthrene skeleton and application thereof in preparation of medicines for preventing and treating infarct diseases | |
| CN107556251B (en) | A kind of ligustrazine phosphat derivative compound and its pharmaceutical composition | |
| CN106946866B (en) | A kind of drug and preparation method thereof preventing and treating cerebral apoplexy | |
| CN100525759C (en) | Ready-made medicine of traditional chinese medicine for treating ischemic cranial vascular disease | |
| CN108164476B (en) | Isophthalonitrile compound, application thereof and medicine containing compound | |
| CN100422134C (en) | 2-(alpha-n-pentanone) phenmethyl and its preparation process and application | |
| CN106344634A (en) | Mango peel extract and preparation method and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C12 | Rejection of a patent application after its publication | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20100630 |