Summary of the invention
The object of the present invention is to provide a kind of pharmaceutical composition that contains the Folium Helianthi extract.It contains the total flavone valid target that from Folium Helianthi, extracts, and the purity of its effective site is 50~99%.Can also add in total terpenoid effective site of Folium Helianthi and the total organic acids effective site one or both in the said composition.
The compositions that contains the Folium Helianthi extract of the present invention, wherein the preparation raw material of extract can also be selected from sunflower plate, Helianthi stem.
Another object of the present invention is to provide a kind of application of pharmaceutical composition in preparation prevention and treatment hypertension drug that contains the Folium Helianthi extract.
A kind of pharmaceutical composition that contains the Folium Helianthi extract of the present invention is characterized in that said composition through adding the various adjuvants that pharmaceutics allows, processes the various dosage forms that allow on the pharmaceutics; Like tablet, comprise general oral, buccal tablet, dispersible tablet, effervescent tablet, gel film, capsule; Soft capsule, granule, drop pill; Oral liquid, syrup; Injection comprises small-volume injection, big injection, injectable powder, Emulsion, suspension, slow releasing agent, controlled release agent, targeting preparation and other various preparations.
A kind of pharmaceutical composition that contains the Folium Helianthi extract of the present invention; It is characterized in that this pharmaceutical composition contains the Folium Helianthi total flavone valid target; Its preparation technology realizes through following technical scheme: extract Folium Helianthi water or 10~95% ethanol (1) as solvent, extracting solution is concentrated into the medicinal liquid that concentration is 0.5~2g/ml; (2) medicinal liquid is used water elution through macroporous adsorptive resins, and the alcoholic solution of reuse 5%~95% carries out gradient elution, collects alcohol eluen, concentrates, and drying promptly obtains total flavone valid target.See Figure of description 1 for details, accompanying drawing 1 is a Folium Helianthi effective site extraction process flow chart.
The method for preparing of Folium Helianthi total flavone valid target of the present invention is characterized in that: the ethanol elution gradient that macroporous resin adopted is 10%~70%.
The method for preparing of Folium Helianthi total flavone valid target in the pharmaceutical composition of the present invention is characterized in that: the content of described total flavone valid target in Folium Helianthi is 0.5~25%.
The method for preparing of Folium Helianthi total flavone valid target in the pharmaceutical composition of the present invention is characterized in that: described total flavone valid target mainly contains lysionotin (nevadensin, 5,7-dihydroxy-6,8; 4 '-trimethoxyflavone), soda Ji Ting (sudachitin, 5,7,4 '-trihydroxy-6,8; 3 '-trimethoxyflavone), de-methoxy soda Ji Ting (sudachitindemethoxy, 5,7,4 '-trihydroxy-6; 8-dimethoxyflavone), marsh piperonal (hymenoxin, 5,7-dihydroxy-6,8; 3 ', 4 '-tetramethoxyflavone), dinatin (hispidulin, 5,7; 4 '-trihydroxy-6-methoxyflavone), A Xi Rosin (acerosin, 5,7,3 '-trihydroxy-6; 8,4 '-trimethoxyflavone), 7, kind of flavone component surplus 4 '-dihydroxyflavone etc. ten, wherein the content of lysionotin in total flavone valid target is mass fraction 20~60%.The structural formula of above-mentioned flavone component is seen Figure of description 2, and accompanying drawing 2 is the main flavone structural formula parent nucleus figure of Folium Helianthi total flavone valid target.
Lysionotin (R1=OMe, R2=H, R3=OMe, R4=OH, R5=OMe, R6=OH)
Soda Ji Ting (R1=OH, R2=OMe, R3=OMe, R4=OH, R5=OMe, R6=OH)
De-methoxy soda Ji Ting (R1=OH, R2=H, R3=OMe, R4=OH, R5=OMe, R6=OH)
The marsh piperonal (R1=OMe, R2=OMe, R3=OMe, R4=OH, R5=OMe, R6=OH)
Dinatin (R1=OH, R2=H, R3=H, R4=OH, R5=OMe, R6=OH)
The A Xi Rosin (R1=OMe, R2=OH, R3=OMe, R4=OH, R5=OMe, R6=OH)
7.7,4’-dihydroxyflavone(R1=OH,R2=H,R3=H,R4=OH,R5=H,R6=H)
Wherein the content of lysionotin composition in total flavone valid target is mass fraction 0.5~60%.
Pharmaceutical composition of the present invention is characterized in that can also adding in this pharmaceutical composition in total terpenoid effective site of Folium Helianthi and the total organic acids effective site one or both.Wherein the method for preparing of the total terpenoid effective site of Folium Helianthi is following: Folium Helianthi water or 10~95% ethanol are extracted as solvent, and extracting solution is concentrated into the medicinal liquid that concentration is 0.5~2g/ml, uses organic solvent extraction; Combining extraction liquid concentrates, and is water-soluble; Through polyamide column, the ethanol with 5%~95% carries out gradient elution, collects eluent; Concentrate, drying promptly gets.The method for preparing of Folium Helianthi total organic acids effective site is following: the effluent and the water lotion of total flavones step of preparation process (2) macroporous adsorbent resin are merged, through anion exchange resin, with ammonia property ethanol elution, collect eluent, concentrate, drying promptly gets.See Figure of description 1 for details, accompanying drawing 1 is a Folium Helianthi effective site extraction process flow chart.
The method for preparing of the total terpenoid effective site of Folium Helianthi of the present invention is characterized in that: extract concentrates through polyamide column, and the ethanol with 10%~70% carries out gradient elution.
The method for preparing of the total terpenoid effective site of Folium Helianthi of the present invention is characterized in that: the content of described total terpenoid effective site in Folium Helianthi is mass fraction 3~20%.
The method for preparing of the total terpenoid effective site of Folium Helianthi in the pharmaceutical composition of the present invention; It is characterized in that: described total terpenoid effective site mainly contains sesquiterpenoids, sesquiterpene lactones class, like guainane (Guaianolide), germacrane (Garmacranolide), Helianthi alkane (Heliangolide), eudesmane (Edudesmane); Diterpene, diterpene acids are like Colophonium alkane, Trachvlobane type, kaurene chemical compound; Triterpenes components is like oleandrin, (Rearranged 3,4-seco-Tirueallane-Typetriterpenoids) to reset triterpene.
Can also add Folium Helianthi total organic acids effective site in the pharmaceutical composition of the present invention.The method for preparing of Folium Helianthi total organic acids effective site is characterized in that: the content of described total organic acids effective site in Folium Helianthi is mass fraction 2%~25%.
The method for preparing of Folium Helianthi total organic acids effective site in the pharmaceutical composition of the present invention; It is characterized in that: described total organic acids effective site mainly contains chlorogenic acid, citric acid, malic acid, Fumaric acid constituents, and wherein the content of chlorogenic acid composition in total organic acids effective site is mass fraction 10~60%.
Folium Helianthi all has extensive distribution throughout the country, and is a lot of with the research report of changing the active aspect of thing of inducting to this kind chemical constituent abroad, particularly therefrom found the chemical compound of the novel structure of many biologically actives in recent years.But for other activity of Folium Helianthi, Helianthi stem and sunflower plate, the effect of extensively using it for blood pressure lowering especially among the people, this research almost are blank.The present invention is applied to Folium Helianthi that (patent applied for is also obtained the authorization patent name: hypertensive Chinese medicine preparation of a kind of treatment and preparation method thereof in the Chinese medicine blood pressure lowering compound preparation first; The patent No.: ZL200310105398.7); And further be the guide with the biological activity, exploratory development also separates and to have obtained Folium Helianthi and have the effective site of antihypertensive activity, i.e. total flavone valid target, and preparation technology is easy and simple to handle, and production cost is low, is suitable for industrialized great production.In addition, also separate obtaining total terpenoid effective site of Folium Helianthi and total organic acids effective site, in these two kinds of effective sites one or both joined in the total flavone valid target, pharmaceutical composition also have significant antihypertensive effect.In addition, show, in sunflower plate, the Helianthi stem through experimental study; Also contain the flavone similar, organic acid and ter penoids with Folium Helianthi; Measure through extraction separation, the composition basically identical of gained effective part extract and Folium Helianthi effective part extract, therefore; The compositions that contains the Folium Helianthi extract of the present invention, wherein the preparation raw material of extract can also be selected from sunflower plate, Helianthi stem.The present invention contains the pharmaceutical composition of Folium Helianthi extract, can prepare prevention and treat hypertensive Chinese medicine preparation, and is quality controllable, and determined curative effect is safe.
The dosage that the suggestion clinical patients uses the present invention to contain the pharmaceutical composition of Folium Helianthi extract is 0.3~0.8g/ time, 3 times on the one.
The pharmaceutical composition that contains the Folium Helianthi extract provided by the invention proves through the renal hypertensive rat blood pressure is influenced result of the test, administration after-contraction pressure and diastolic pressure obviously descend (P<0.05).Above-mentioned result of the test has pointed out the pharmaceutical composition that contains the Folium Helianthi extract to can be used as active component prevented and treated hypertensive medicine in preparation application.
The specific embodiment
Through specific embodiment the preparation of drug combination method and the pharmacologically active that contain the Folium Helianthi extract are done further to detail below.
Embodiment 1: the preparation of Folium Helianthi total flavone valid target
Folium Helianthi 20kg is put in the extraction pot, add 12 times of amounts of 80% ethanol, extract secondary; Each 1 hour, merge extractive liquid, filtered; Filtrating is concentrated into the medicinal liquid that concentration is 1g/ml, through the ADS-17 macroporous adsorptive resins, earlier with 3 bv of water elution; 3 bv of each eluting of reuse 10% and 30% ethanol collect the alcohol moiety eluent, and concentrate drying obtains total flavone valid target 0.32kg.
Embodiment 2: the preparation of Folium Helianthi total flavone valid target
Folium Helianthi 20kg is put in the extraction pot, add 12 times of amounts of 50% ethanol, extracted 2 hours, medicinal residues add 10 times of amounts of 50% ethanol again; Extracted 1 hour, merge extractive liquid, filters; Filtrating is concentrated into the medicinal liquid that concentration is 0.8g/ml, through the ADS-F8 macroporous adsorptive resins, earlier with 3 bv of water elution; 3 bv of each eluting of reuse 10% and 70% ethanol collect the alcohol moiety eluent, and concentrate drying obtains total flavone valid target 0.29kg.
Embodiment 3: the preparation of sunflower plate total flavone valid target
Sunflower plate 20kg is put in the extraction pot, add 10 times of amounts of 50% ethanol, extracted 2 hours, medicinal residues add 10 times of amounts of 50% ethanol again; Extracted 1 hour, merge extractive liquid, filters; Filtrating is concentrated into the medicinal liquid that concentration is 0.8g/ml, through the LS-300 macroporous adsorptive resins, earlier with 3 bv of water elution; 3 bv of each eluting of reuse 10% and 70% ethanol collect 70% alcohol moiety eluent, and concentrate drying obtains total flavone valid target 0.21kg.
Embodiment 4: the preparation of the total terpenoid effective site of Folium Helianthi
Folium Helianthi 40kg is put in the extraction pot, add 70% ethanol extraction secondary, all add 12 times of amounts at every turn, each 2 hours, merge extractive liquid, filtered, and filtrating is concentrated into the medicinal liquid that concentration is 1g/ml; With the chloroform extraction of equivalent 5 times, combining extraction liquid concentrates through polyamide column, with washing 1bv, and 30% ethanol elution 3bv, 70% ethanol elution 2bv merges eluent, and concentrate drying promptly gets total terpenoid effective site 0.51kg.
Embodiment 5: the preparation of Folium Helianthi total organic acids effective site
Folium Helianthi 50kg is put in the extraction pot, add 30% ethanol extraction secondary, be 10 times of amounts at every turn, each 2 hours; Merge extractive liquid, filters, and filtrating is concentrated into the medicinal liquid that concentration is 1g/ml, through the AB-8 macroporous adsorptive resins; With 3 bv of water elution, merge effluent and water lotion, through the 201X7 strong-basicity styrene series anion exchange resin, with 20% ammonia ethanol elution 4bv; Collect eluent, concentrate drying obtains total organic acids effective site 0.62kg.
Embodiment 6: content of total flavone is measured in the Folium Helianthi total flavone valid target
Adopt content of total flavone in the determined by ultraviolet spectrophotometry total flavone valid target of the present invention:
Lysionotin reference substance 2mg is got in the preparation of reference substance solution, accurate claims surely, puts in the 10ml measuring bottle, is dissolved in water and is settled to scale, shakes up.
The preparation precision of standard curve is measured reference substance solution 0.1ml, 0.2ml, 0.3ml, 0.4ml; 0.5ml 0.6ml puts respectively in the 25ml measuring bottle, the accurate respectively 1% aluminum chloride aqueous solution 5ml that adds; Shake up, the accurate 1mol/l potassium acetate aqueous solution 8ml that adds shakes up, and water is settled to scale; Shake up, as blank,, measure absorbance in the wavelength of 228nm according to ultraviolet visible spectrophotometry (an appendix V of Chinese Pharmacopoeia version in 2005 A) with corresponding reagent; With the absorbance is vertical coordinate, and concentration is abscissa, the drawing standard curve.
Algoscopy is got these article fine powder 3mg, and accurate the title decides, and puts in the 10ml measuring bottle, adds 70% ethanol 8ml; Supersound process (power 100W, frequency 40kHz) 30 minutes is put coldly, adds 70% ethanol to scale; Shake up, filter, precision is measured subsequent filtrate 1.0ml, puts in the 25ml measuring bottle; The accurate respectively 1% aluminum chloride aqueous solution 5ml that adds shakes up, and the accurate 1mol/l sodium acetate aqueous solution 8ml that adds shakes up; Water is settled to scale, shakes up, and as blank, shines ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia version in 2005) with corresponding reagent; Wavelength at 228nm is measured absorbance, from the amount that standard curve is read lysionotin the need testing solution, calculates, and the every 1g of these article contains total flavones and counts 620mg with lysionotin.
Embodiment 7: the assay of lysionotin in the Folium Helianthi total flavone valid target
Adopt the HPLC method to measure the content of lysionotin in the total flavone valid target of the present invention:
Chromatographic condition and system suitability test are filler with the octadecyl silane; With acetonitrile-0.4% Triammonium citrate (40: 60) is mobile phase; The detection wavelength is 284nm.
It is an amount of that the lysionotin reference substance is got in the preparation of reference substance solution, and accurate the title decides, and adds methanol and processes the solution that every 1ml contains 50 μ g, promptly gets.
These article fine powder 15mg is got in the preparation of need testing solution, accurate claims surely, puts in the 10ml measuring bottle, adds 70% ethanol 8ml, and supersound process (power 100W, frequency 40kHz) 30 minutes is put coldly, adds 70% ethanol to scale, shakes up, and filters, and promptly gets.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, and promptly get.
It is 20mg that the every 1g of these article contains lysionotin.
Embodiment 8: contain the hypotensive effect of the pharmaceutical composition of Folium Helianthi extract to renal hypertensive rat
Through the hypotensive effect evidence to renal hypertensive rat, the pharmaceutical composition that contains the Folium Helianthi extract has hypotensive effect.
1, test material:
The blood pressure lowering sample is (Folium Helianthi total flavone valid target, human dosage are 0.25g/ people/day) 1., chocolate brown powder.
The blood pressure lowering sample is (mixture of Folium Helianthi total flavone valid target and Folium Helianthi total organic acids effective site, human dosage are 0.35g/ people/day) 2., the dark-brown powder.
The blood pressure lowering sample is (mixture of the total terpenoid effective site of Folium Helianthi total flavone valid target and Folium Helianthi, human dosage are 0.40g/ people/day) 3., the dark-brown powder.
The blood pressure lowering sample is (mixture of Folium Helianthi total flavone valid target, the total terpenoid effective site of Folium Helianthi and Folium Helianthi total organic acids effective site, human dosage is 0.5g/ people/day) 4., the dark-brown powder.
Positive reference substance: compound Herba Apocyni veneti (Folium Apocyni Veneti) I (6 of day for human beings clothes), 100 slices/bottle.
2, laboratory animal and equipment:
25 of renal hypertensive rats, the cleaning level, body weight 357 ± 42g is provided credit number by Lukang Medical Co., Ltd., Shandong's animal center: SCXK Shandong 20080002.
Key instrument equipment: noinvasive arteria caudalis blood pressure determination analytical system, Shanghai Alcott Bioisystech Co., Ltd.
3, test method
25 of renal hypertensive rats are divided into 5 groups at random, and promptly 1. the blood pressure lowering sample is organized 100mg/kg, blood pressure lowering sample and is organized 2. that 3. 150mg/kg, blood pressure lowering appearance organize 50mg/kg, 4. the blood pressure lowering sample organizes 200mg/kg, and positive control drug compound Herba Apocyni veneti (Folium Apocyni Veneti) I organizes 3/kg.Every group 5.Before administration, measuring rat blood pressure with noinvasive arteria caudalis blood pressure determination analytical system is worth before as medicine.Disposable then gastric infusion.Each administration group is irritated the medicinal liquid 2ml/200g Mus that stomach gives variable concentrations respectively.After administration 2.0 hours, measure rat arteria caudalis blood pressure with noinvasive arteria caudalis blood pressure determination analytical system.
4, data statistics and result judge:
Experimental data is all with mean ± standard deviation (X ± s) expression.Relatively adopt t-check carrying out statistical analysis before and after self.
5, result of the test: the result sees table 1.
The disposable gastric infusion of table 1 blood pressure lowering sample is to the hypotensive effect of renal hypertensive rat (X ± s)
Annotate: with comparison before the administration,
*P<0.05
The result shows that 1,3 pairs of renal hypertensive rat diastolic pressures of blood pressure lowering sample and mean arterial pressure have obvious hypotensive effect, though systolic pressure is not had obvious statistical significance, reduction trend are arranged.2,4 pairs of renal hypertensive rat blood pressures of blood pressure lowering sample all have significant hypotensive effect.
Embodiment 9: contain the preparation of the pharmaceutical composition tablet of Folium Helianthi extract
Get Folium Helianthi total flavone valid target 205g in the foregoing description 1, add 95g starch, mixing is granulated, and sieves, and adds the 2g microcrystalline Cellulose, the 0.5g magnesium stearate, and mixing is pressed into 1000, is tablet of the present invention.
Embodiment 10: contain the preparation of the medicament composition granule agent of Folium Helianthi extract
Get the foregoing description 1,2 Sino-Japan certain herbaceous plants with big flowers leaf flavonoids and total organic acids effective site 405g, add lactose 200g, dextrin 400g granulates, and granulate sieves, and processes granule 1000g, granule of the present invention.
Embodiment 11 Folium Helianthi total flavone valid target tablets are to liver-Yang sthenia type hyperpietic's clinical observation test
1, physical data
46 routine patients all derive from the outpatient service of intracardiac section, are divided into test group and matched group at random.Test group 26 examples, wherein male 12 examples, women 14 examples, 55.14 ± 11.45 years old mean age, average course of disease 10.87 ± 7.13 years; Matched group 20 examples, wherein male 11 examples, women 9 examples, 55.97 ± 10.85 years old mean age, average course of disease 11.46 ± 6.98 years.Two groups of pressure values before sex, age, the course of disease, treatment, the traditional Chinese medical science individual event symptom aspect such as relatively all do not have significant difference (P>0.05).
2, Therapeutic Method
Test group is taken Folium Helianthi total flavone valid target tablet, and each 2, every day 3 times; Matched group is taken Herba Apocyni veneti (Folium Apocyni Veneti) Tabellae, and every day 3 times, each 4, two groups was 1 course of treatment with 4 weeks all, after finishing for 1 course of treatment, and statistics blood pressure level and curative effect.
3, efficacy assessment standard
Efficacy of antihypertensive treatment evaluation criteria: with reference to the standard of national cardiovascular epidemiology in 1979 with crowd prevention and treatment forum formulation; The criterion of tcm symptom total effects: the regulation with reference in " the new Chinese medicine clinical research guideline " of Ministry of Public Health promulgation in 2002 is evaluated.
4, statistical procedures adopts Ridit to analyze the t check.
5, (1) two group of efficacy of antihypertensive treatment of therapeutic outcome and blood pressure lowering amplitude are relatively seen table 2~4.
Table 2 liang group efficacy of antihypertensive treatment relatively
Analyze through Ridit, P<0.05, test group is superior to matched group.
Table 3 liang group systolic pressure amplitude is (kPa) relatively
*Represent and preceding relatively P<0.01 of treatment that # representes to compare P<0.01 with matched group.
Table 4 liang group diastolic pressure amplitude is (kPa) relatively
*Represent and preceding relatively P<0.01 of treatment that # representes to compare P<0.01 with matched group.
Can know by table 2, produce effects 10 examples in test group 26 examples, effective 12 examples are 84.6% effectively always, are superior to matched group, two groups of curative effects are compared through the Ridit analysis has significant difference (P<0.05).Systolic pressure and diastolic pressure by before and after table 3, the visible two groups of patient treatments of table 4 self more all have utmost point significant difference (P<0.01) respectively, and treatment back test group is compared with matched group also has utmost point significant difference (P<0.01).
(2) curative effect of traditional Chinese medical science disease:
Test group is being improved aspect the traditional Chinese medical science individual event symptom; Except that " being irritable and getting angry easily " and " blurred vision "; Other all are superior to matched group like dizzy, headache, dizziness and distending sensation of the head, stiffness of the neck, flushing conjunctival congestion, nervous, feeling of oppression and pain in the chest, insomnia, tinnitus, have compared significant difference (P<0.05 or P<0.01) for two groups.The result shows that Folium Helianthi total flavone valid target tablet of the present invention in effective blood pressure lowering, also can obviously improve symptom; And do not see untoward reaction during the treatment.