CN1039198C

CN1039198C - Process for preparing diterpene alkaloid and its medicinal usage

Info

Publication number: CN1039198C
Application number: CN93110509A
Authority: CN
Inventors: 刘静涵; 后德辉; 王秋娟; 张陆勇; 赵守训; 范炳尧
Original assignee: Shenzhen Daphne Pharmaceuticals Co ltd; China Pharmaceutical University
Current assignee: Jilin Aodong Zhaonan Pharmaceutical Co., Ltd.; China Pharmaceutical University
Priority date: 1993-01-01
Filing date: 1993-01-01
Publication date: 1998-07-22
Anticipated expiration: 2013-01-01
Also published as: CN1088827A

Abstract

The present invention relates to a method for preparing diterpene alkaloid and medicinal purposes of the diterpene alkaloid. A solvent is used for extracting the Chinese herbal medicine of aconitum coreanum; extract is obtained by recovering the extracting solution; the extract is dissolved in acid water; the acid water is made to stand to separate gum; the precipitation is filtered; an organic solvent is used for degreasing the extract; after the organic solvent is removed, the acid water is alkalized until the pH value is from 5 to 10; then, the organic solvent is used for extracting the alkalized liquid; the extraction liquid is collected; after the organic solvent is recovered, a crude product of guanfu base A is obtained; the crude product of the guanfu base A is repeatedly recrystallized by the organic solvent or refined by column chromatography and is dried at the temperature of 50 to 100DEG C; then, the highly finished product of the guanfu base A is obtained. The guanfu base A with low toxicity and a natural structure and a salt thereof capable of being accepted by physiology can be made into various medicament forms for treating or preventing diseases of the cardiovascular system.

Description

A kind of preparation method of diterpene alkaloid

The present invention relates to from Chinese herbal medicine, extract separation of C ₂₀The method of hetisine type (hetisine) chemical compound is especially extracted, separated to the method for diterpene alkaloid and to the influence of cardiovascular system from Ranunculaceae Radix Aconiti Coreani Aconitum coreanum (Levl.) Rapaics.

Cardiovascular system diseases is all classified frequently-occurring disease as in China and many in the world developed countries.Common cardiovascular disease is an arrhythmia.Existing anti-arrhythmic as: though quinidine, lignocaine, amiodarone, verapamil hydrochloride etc. have certain curative effect, also have various shortcoming, for example: quinidine belongs to I _aThe class anti-arrhythmic, main moderate suppresses the O phase rate of climb and prolongs the multipole time.Fully, heart is had untoward reaction though the oral post-absorption of quinidine is fast, one is asystole and conduction block, and another is the myocardial abnormality excitement, and often with the intestines and stomach untoward reaction, therapeutic index is low, and untoward reaction takes place about 1/3 patient.Verapamil hydrochloride prevention angina pectoris or paroxysmal supraventricular tachycardia, but toxic and side effects is big, has 4/5 patient that sinus standstill takes place, and medication is dangerous.Amiodarone belongs to the medicine of over reach current potential, and curative effect is better, but it is big to take toxic and side effects for a long time, causes that easily pulmonary fibrosis, thyroid function change, so should not take for a long time, cardiac sudden death (bringing out the chamber quivers) is restricted for preventing and treating.The lignocaine arrhythmia is effective, but can not be oral, and the anti-chamber effect strong (" Pharmacopoeia of People's Republic of China, two ones, clinical application notice " Chinese Medicine science and technology publishing house, 1990 the 1st version 129～158 pages) of quivering.

The purpose of this invention is to provide a kind of from Radix Aconiti Coreani the method for extraction separation hetisine type chemical compound, and the low new type natural structural compounds of toxicity that obtains be used for the treatment of or prevent cardiovascular system diseases.

The present invention be a kind ofly from Chinese herbal medicine Ranunculaceae Radix Aconiti Coreani, extract, the method for isolating active composition hetisine type chemical compound Guan-fubase A (Guan-Fu BaseA); the chemistry of Guan-fubase A is called 2 α; 13 β-diacetyl; 11 α; 14 alpha-dihydroxy-s are conspicuous for new (2 α; 13 β-diacetyl, 11 α, 14 α-dihydrohetisine).Specific operation process is, use the solvent extraction Radix Aconiti Coreani, reclaim extracting solution, obtain extractum, the extractum commentaries on classics is dissolved in the sour water, transfer pH2-4, leave standstill sour water liquation glue, precipitate in the filtering sour water, the acid liquid of filtering post precipitation lipotropy organic solvent degreasing is removed the lipotropy organic solvent layer, the alkalization acid liquid is to pH5～10, the water liquid of reuse lipotropy organic solvent extraction alkalization is collected extract, normal pressure and/or reclaim under reduced pressure organic solvent, obtain the Guan-fubase A crude product, with organic solvent recrystallization and/or use silica gel repeatedly, aluminium oxide or macroporous resin column chromatography are refining, 50～100 ℃ of dryings, obtain the Guan-fubase A highly finished product.

The organic solvent of defat or extraction usefulness can be for example chloroform, a dichloromethane of halogenated hydrocarbons, and ethyl acetate, methyl acetate, petroleum ether or benzene.

The alkali of alkalization acid liquid can be 1～5%NaOH water liquid, 1-5%KOH water liquid, strong aqua ammonia or Na ₂CO ₃

The used organic solvent of recrystallization can be methanol, ethanol, benzene, ether or acetone, preferably adopts methanol or dehydrated alcohol.

During with the organic solvent recrystallization, can add acid and transfer pH3-5, make the salt of physiologically acceptable Guan-fubase A.Available acid has hydrochloric acid, hydrogen chloride gas, hydrobromic acid, nitric acid, phosphoric acid, preferably uses 1～5% hydrochloric acid.

Guan-fubase A or its salt can with general adjuvant and/or mixed with excipients, make various dosage forms, be used for the treatment of or prevent cardiovascular system diseases, normally be used for the treatment of or prevent arrhythmia, be preferably used in treatment or prevention supraventricular tachycardia and/or atrial fibrillation.

Pharmaceutical excipient is meant the prescription adjuvant of solid, semisolid, filler or liquid diluent and every type.

Tablet, dragee, capsule, pill, granule, stick agent or solution all is alternative pharmaceutical formulation agent.

Tablet, dragee, capsule, pill and granule can contain one or more usual excipients, for example: and filler and bulking agent, as starch, lactose, sucrose, glucose, mannitol and silicon dioxide; Binding agent is as carboxymethyl cellulose, alginate, gelatin and polyvinylpyrrolidone; Humectant is as glycerol; Disintegrating agent, as agar, calcium carbonate and sodium bicarbonate; Dissolving delayer such as paraffin; Absorb accelerator again, as quaternary ammonium compound; Wetting agent is as hexadecanol, glycerol monostearate; Adsorbent is as Kaolin and bentonite; Lubricant, as Pulvis Talci, calcium stearate, magnesium stearate and solid polyethylene glycol.Tablet, dragee, capsule, pill and granule often can be made into coating and the shell that has the lucifuge agent, these dosage forms only also can be mixed with or preferentially discharge active component or delayed release of active elements optionally in certain part of intestinal.

Solution can contain general solubilizing agent, solvent, antiseptic, emulsifying agent, for example: water, ethanol, isopropyl alcohol, benzyl alcohol, benzyl benzoate, propylene glycol, glycerol, Polyethylene Glycol or sorbitan fatty acid fat.

For parenterai administration, solution and Emulsion all are aseptic and isoosmotic with blood.

The said medicine preparation is with known common method preparation, for example can be with active component and one or more mixed with excipients.

The test of the efficacy of Guan-fubase A and salt thereof comprises isolated test, in body test, electrophysiologic testing and toxicity comparative test.

One, isolated test

Material

Guan-fu base A hydrochloride injection (Injection Guan-fu base AHydrochloride).Verapamil (Verapamil) is given by Tianjin Inst. of Materia Medica, and lignocaine is Taixing, a Jiangsu county pharmaceutical factory product, and ouabain is institute for drug control, Shanghai medicine inspection standard substance, and normal Lip river quinoline is given by Shanghai 16 pharmaceutical factories.

All rats, mice, rabbit, mongrel are provided by animal housing of China Medicine University and Chinese Academy of Sciences's Experimental Animal Center.

Method referring to

(1)Allen?J.D.et?al?Br.J.Pharmacol?45：561，1972。

(2) uncle pharmacological experiment methodology Xu cloud, card is as Lian Chen Xiu chief editor People's Health Publisher 1982:400

(3) cardiovascular drug pharmacodynamics specification requirement (original text on probation), cardiovascular pharmacological branch of Chinese Pharmacological Society, 1991.4. compiles in State Pharmaceutical Administration new drug research administrative center

(1) isolated rat heart ligation arteria coronaria is brought out ARR protective effect

15 of rats, body weight 191 ± 21g, ♂ ♀ dual-purpose, be divided into three groups at random, be matched group, Guan-fu base A hydrochloride group and lignocaine group, the langendorff method is carried out the isolated rat heart perfusion, the easypro tension curve that contracts of the heart is traced on JL-3 type three-channel recorder logical O by tonotransducer ₂Rockwell liquid perfusion 5min, change Rockwell liquid perfusion 5min respectively with benefit caine (5 μ g/ml) and Guan-fu base A hydrochloride (50 μ g/ml), use eyeless suture needle (line 4 * 2 4/0) ligation anterior descending coronary initial part (being equivalent to left auricle point place) then, occur the incidence rate of ventricular arrhythmia after the observation ligation in the 10-30min.

The result shows the chamber speed that Guan-fu base A hydrochloride (50 μ g/ml) brings out isolated rat heart ligation arteria coronaria, and quivering in the chamber all has significant protective effect and lignocaine no significant difference (seeing Table 1).Show that Guan-fu base A hydrochloride has protective effect to the myocardial ischemia arrhythmia.

Table 1 pair isolated rat heart ligation arteria coronaria brings out ARR protective effect

Concentration ventricular arrhythmia incidence rate (%) group number of animals

(μ g/ml) chamber is the chamber speed chamber matched group 50 100 100 60 lignocaine groups 5 50 60 0 of quivering early ^*0 ^*Guan-fu base A hydrochloride group 5 50 50 0 ^*0 ^*

*: p＜0.05, * *: p＜0.01 (comparing) by direct short cut technique and matched group

(2) high calcium is brought out the protective effect of isolated rat perfusion fibrillation of heart

47 of male rats, body weight 240-380g, the back of hitting unconsciously is taken out heart rapidly, presses Langendorff method perfusion, passes to 95%O ₂+ 5%CO ₂Krebs liquid constant-flow perfusion (5ml/min).Change no K behind the matched group perfusion 25min ⁺High Ca ⁺⁺(KCl is 0 to liquid, CaCl ₂6H ₂O 4.92).Trace heart surface electrograph before and after the perfusion, the time of origin of (VF) incidence rate and VT and VF quivers for observation ward's speed (VT) and chamber.Change medicinal liquid into behind the administration group Krebs liquid perfusion 10min, change no K behind the 15min ⁺High Ca ⁺⁺Liquid.Matched group is with after causing the liquid perfusion heart that quivers, the VT incidence rate is 55%, time of origin is 5.3 ± 4.1min, the VF incidence rate is 91%, time of origin is 7 ± 5min, and the VT incidence rate was 66.7% when the verapamil group was 2 μ g/ml when concentration, and time of origin is 5.6 ± 1.3min, do not have significant difference with matched group, and VF incidence rate and time of origin have slightly than matched group and alleviate.When concentration is 3 μ g/ml, VT and VF all do not take place, the pyrozoline group: when concentration was 5 μ g/ml, VT and VF incidence rate did not have change, when concentration is 10 μ gml, VT incidence rate no change, and the incidence rate of VF obviously reduces, the Guan-fu base A hydrochloride group: during 20 μ g/ml, VT and VF incidence rate obviously reduce, when concentration is 30 μ g/ml, VF does not take place, the time of origin of VT is obviously postponed.The result shows that Guan-fu base A hydrochloride (20-30 μ g/ml) can suppress that high calcium brings out isolated rat perfusion heart chamber speed and quiver in the chamber, acts on similar to verapamil (2-3 μ g/ml).

Two, test at body

(1) the electricity irritation man rabbit heart arrhythmia of bringing out

The Cor Leporis electricity is caused the influence of ventricular fibrillation threshholds

12 body weight 2.2 ± 0.2kg of rabbit are divided into two groups, and 6 every group, ♂ ♀ half and half.One group is the Guan-fu base A hydrochloride group, and another group is the lignocaine group.Urethane 1.2g/kg ear edge iv anesthesia.Under the artificial respiration, open breast and expose heart, and be connected in pressure transducer, trace the easypro curve that contracts of heart from JL-3 type three road physiographs with Gu Shi lever aroused in interest.Stimulating electrode is placed apex (positive pole) and left side, atrioventricular junction place (negative pole) and is connected in DGQ-2 type stimulator, and the square wave stimulation 10s with the wide 1ms of ripple, frequency 20Hz stimulates once every 2min, increases stimulus intensity (mV) one by one.Record the Cor Leporis chamber of the causing value value in contrast of quivering.Then, inject Guan-fu base A hydrochloride or lignocaine respectively by rabbit ear vein, the various dose administration 10min of being separated by, 5min measures and causes the threshold that quivers after each administration, and compares (seeing Table 2) before the administration.The result show iv Guan-fu base A hydrochloride and lignocaine each 3,6,12mg/kg all can obviously improve the Cor Leporis chamber of the causing value (P＜0.01) of quivering, and acts on reinforcement with the dosage increase.Illustrating to quiver to be significantly improved in chamber that Guan-fu base A hydrochloride causes the electricity irritation Cor Leporis causes the threshold effect that quivers, and does not have significant difference (P＜0.05) with lignocaine.Two groups all cause the threshold that quivers and return back to level before the administration again in the 30-40min behind iv12mg/kg.

The influence (N=6) of table 2 pair Cor Leporis electricity threshold of ventricular fibrillation current

Electricity causes the threshold that quivers (X ± SD mv) group

Dosage (mg/kg) before the administration

36 12 Guan-fu base A hydrochloride group 9.8 ± 0.1 15.5 ± 1.1 ^*18.3 ± 1.4 ^*21.2 ± 1.6 ^*Lignocaine group 9.7 ± 0.4 14.7 ± 2.4 ^*17.4 ± 2.2 ^*20.6 ± 2.3 ^**: P＜0.01 (with comparing before the administration)

(2) arrhythmia that ligation branch of coronary artery forms (the second phase ligation of Canis familiaris L. arteria coronaria)

13 of hybrid dogs, body weight 6-12kg, ♂ ♀ does not limit, and is divided into Guan-fu base A hydrochloride group (n=7) and quinidine group (n=6).Animal is anaesthetized with pentobarbital sodium 30mg/kg intravenous injection, the field of operation unhairing, and routine disinfection adopts Harris second phase ligation dog left anterior descending coronary artery to cause ventricular arrhythmia under aseptic condition.Art finishes the local gentamycin 80,000 unit prevention infection of injecting.About postoperative 20 hours, animal administration under waking state.Adopt summation intravenous injection Guan-fu base A hydrochloride 10mg/kg, 20mg/kg and 40mg/kg or positive control drug quinidine 2.5mg/kg, 5mg/kg and 10mg/kg.About 2 minutes of each administration time.With the II lead electrocardiogram of different time in 30 minutes before the administration of electrocardiograph record and after the administration, every 5 minutes records once, each continuous record 1 minute.The isopyknic normal saline of injection compares before the administration, in 30 minutes heart rate, incidence of arrhythmia is not had obvious influence.

The results are shown in Table 3-table 5.Because the influence of each dog arteria coronaria side Zhi Xunhuan etc., the ARR order of severity of each dog has nothing in common with each other, ventricular rhythm and heart rate than about 15%-100%.Guan-fu base A hydrochloride 10mg/kg, 20mg/kg and 40mg/kg can both significantly reduce ventricular rhythm, increase sinus rhythm.80% following person, Guan-fu base A hydrochloride has stronger inhibition ventricular rhythm effect to ventricular rhythm, and 20-40mg/kg can suppress ventricular rhythm fully, makes to change into sinus rhythm.Nearly 100% to ventricular rhythm, inhibitory action a little less than.40mg/kg is to the average suppression ratio about 50% of ventricular rhythm.Quinidine 5mg/kg, 10mg/kg significantly suppress ventricular rhythm, and increase sinus rhythm.Similar to Guan-fu base A hydrochloride, quinidine also is effective to the low person of ventricular rhythm percentage rate.Adopt in the more approaching several routine animal of ventricular rhythm percentage rate, each dosage returns the average suppression ratio of ventricular rhythm, and Guan-fu base A hydrochloride suppresses the ED of ventricular arrhythmia ₅₀ED for the 19.5mg/kg quinidine ₅₀Be 6.2mg/kg.Guan-fu base A hydrochloride also has remarkable decreased heart rate effect, and average heart rate slows down less than 20% during 40mg/kg.Quinidine does not make significant difference to heart rate at used dosage.

The above shows that Guan-fu base A hydrochloride has the ventricular arrhythmia that significant inhibition Harris second phase ligation dog coronary artery causes, increases sinus rhythm, and has certain decreased heart rate effect.Guan-fu base A hydrochloride suppresses the ED of this ventricular rhythm ₅₀Be about three times of quinidine, because the arrhythmia before the medication of Guan-fu base A hydrochloride group is serious than the quinidine group, so the ratio of the antiarrhythmic effect intensity of the former with the latter on this model may be greater than 1/3.

(3) drug-induced arrhythmia

1. aconitine brings out arrhythmia

(1) iv Guan-fu base A hydrochloride injection brings out the protective effect of rat ventricular arrhythmia to aconitine

50 of rats, body weight 238 ± 31g, ♂ ♀ dual-purpose is divided into 5 groups at random, after 1.2g/kg urethane ip anesthesia, respectively from femoral vein iv quinidine 15mg/kg, matched group femoral vein iv equivalent normal saline.Inject Aconitine Nitrate solution (1 μ g/0.2ml/min) with WSQ-A type trace transfusion device by the external jugular vein constant speed after 2 minutes, and monitor that by electrocardioscope electrocardio changes, and the record chamber of appearance morning, chamber speed, the consumption of aconitine when quiver in the chamber.

The result shows that the Guan-fu base A hydrochloride injection of height, middle low dosage can obviously increase Aconitine Nitrate and bring out the dosage that quivers chamber morning, chamber speed, chamber, and its effect increases with increasing of Guan-fu base A hydrochloride injection dosage.Illustrate that iv Guan-fu base A hydrochloride injection has significant protective effect to the ventricular arrhythmia that aconitine brings out.(table 3)

Table 3 IV Guan-fu base A hydrochloride injection brings out aconitine

The protective effect X of rat ventricular arrhythmia ± SD N=10 * * P＜0.01 (comparing) with matched group

Cause the chamber early Aconitum carmichjaelii Debx. cause that chamber speed Aconitum carmichjaelii Debx. causes the chamber Aconitum carmichjaelii Debx. group alkali consumption alkali consumption alkali consumption that quivers

(μ g/kg) (μ g/kg) (μ g/kg) matched group 23.5 ± 3.3 28.4 ± 4.2 42.3 ± 5.4 Guan-fu base A hydrochloride injection 32.7 ± 3.2 ^*38.7 ± 2.6 ^*51.3 ± 5.4 ^*(10mg/kg) the Guan-fu base A hydrochloride injection 43.8 ± 3.2 ^*49.5 ± 3.0 ^*65.0 ± 4.3 ^*(25mg/kg) the Guan-fu base A hydrochloride injection 56.1 ± 5.4 ^*63.5 ± 4.8 ^*82.6 ± 8.3 ^*(40mg/kg) quinidine hydrochloride (15mg/kg) 33.2 ± 3.2 ^*38.5 ± 2.3 ^*52.4 ± 6.8 ^*

(2) the iv Guan-fu base A hydrochloride brings out the protective effect of rat ventricular arrhythmia to aconitine.

38 of rats, body weight 259 ± 38g, ♂ ♀ half and half is divided into 5 groups at random, after 1.2g/kg urethane ip anesthesia, gives Guan-fu base A hydrochloride 10,25 by external jugular vein respectively, 40mg/kg and lignocaine 15mg/kg.Matched group iv equivalent normal saline.Inject Aconitine Nitrate solution (1 μ g/0.2ml/min) with WSQ-A type trace transfusion device by the external jugular vein constant speed after 2 minutes; and early by the electrocardioscope supervision chamber of appearance; chamber speed; the consumption of record aconitine when quiver in the chamber; the result shows height; in; the Guan-fu base A hydrochloride of low dosage can obviously increase aconitine and bring out the chamber early; chamber speed; the dosage that quivers in the chamber; its effect increases with the increasing of Guan-fu base A hydrochloride dosage, illustrates that the iv Guan-fu base A hydrochloride has significant protective effect to the ventricular arrhythmia that aconitine brings out.(table 4).

Table 4 IV Guan-fu base A hydrochloride injection brings out aconitine

The protective effect of rat ventricular arrhythmia

X ± SD * P＜0.05 * * P＜0.01 (comparing) with matched group

Animal cause the chamber early Aconitum carmichjaelii Debx. cause that chamber speed Aconitum carmichjaelii Debx. causes that the chamber Aconitum carmichjaelii Debx. group of quivering counts alkali consumption alkali consumption alkali consumption

(only) (μ g/kg) (μ g/kg) (μ g/kg) matched group 10 23.4 ± 5.2 27.9 ± 4.9 35.7 ± 4.8 Guan-fu base A hydrochloride injection 10 37.7 ± 3.5 ^*45.4 ± 4.7 ^*52.0 ± 3.5 ^*(10mg/kg) the Guan-fu base A hydrochloride injection 6 43.6 ± 4.4 ^*49.6 ± 6.0 ^*59.8 ± 14.7 ^*(25mg/kg) the Guan-fu base A hydrochloride injection 6 80.5 ± 10.2 ^*84.1 ± 10.7 ^*106.0 ± 13.6 ^*(40mg/kg) lignocaine (15mg/kg) 6 27.6 ± 3.1 34.1 ± 7.6 ^*48.1 ± 11.5 ^*

(3) the lumbar injection aconitine brings out rat ventricular

The ip Guan-fu base A hydrochloride brings out the protective effect of rat ventricular arrhythmia to aconitine.Operation is the same.23 of rats, body weight 268 ± 31g, ♂ ♀ half and half is divided into 5 groups at random, after the urethane 1.2g/kg ip anesthesia, difference ip Guan-fu base A hydrochloride 10,25,40mg/kg and lignocaine 15mg/kg, matched group ip equivalent normal saline, write down the chamber of appearance early by last method behind the 20min, chamber speed, the consumption of injection Aconitine Nitrate when quiver in the chamber.The result shows that high, medium and low dosage Guan-fu base A hydrochloride can obviously increase aconitine and bring out the dosage that quivers chamber morning, chamber speed, chamber; its effect strengthens with dosage and strengthens, and illustrates that the ventricular arrhythmia that the ip Guan-fu base A hydrochloride brings out aconitine also has significant protective effect (seeing Table 5).

Table 5 ip Guan-fu base A hydrochloride injection brings out aconitine

The protective effect of rat ventricular arrhythmia

X ± SD * P＜0.05 * * P＜0.01 (comparing) with matched group

(only) (μ g/kg) (μ g/kg) (μ g/kg) matched group 5 28.3 ± 7.3 32.1 ± 4.1 52.8 ± 8.2 Guan-fu base A hydrochloride injection 5 34.7 ± 4.2 ^*52.2+ ± 15.2 ^*72.3 ± 11.6 ^*(10mg/kg) the Guan-fu base A hydrochloride injection 5 38.6 ± 6.0 ^*57.0 ± 6.9 ^*82.7 ± 14.1 ^*(25mg/kg) the Guan-fu base A hydrochloride injection 4 57.2 ± 11.2 ^*68.2 ± 8.4 ^*103.1 ± 25.6 ^*(40mg/kg) lignocaine (15mg/kg) 4 42.8 ± 0.9 ^*50.8 ± 5.7 ^*64.6 ± 15.6

2. the calcium chloride arrhythmia of bringing out

To CaCl ₂Bring out the rat ventricular protective effect

44 of SD rats, body weight 172 ± 13g, pentobarbital sodium ip anesthesia, rat is divided into iv verapamil or Guan-fu base A hydrochloride group and normal saline matched group.Quick (5s) iv CaCl of 5min after the administration ₂130mg/kg observes II helical pitch ECG and changes.

The results are shown in Table 6.12 rats of matched group, iv CaCl ₂Quiver in back generation chamber rapidly, 11/12 Mus is dead in 1-2min.The dosage of verapamil is 2 and 3.8mg/kg, 5min after the administration, and the rat heart rate reduces by 16 and 32% respectively, and 2mg/kg organizes 2/8 Mus and takes place dead.And 3.8mg/kg organizes none death.Also occur decreased heart rate behind iv Guan-fu base A hydrochloride 20 and the 30mg/kg, reduce by 25 and 28% respectively, can lower CaCl equally ₂Cause that quiver in the chamber and dead incidence rate.

Table 6 IV verapamil or Guan-fu base A hydrochloride are to CaCl ₂

Cause ARR protective effect

The group number of rats ectopic rhythm chamber death toll of quivering during dosage 5min

(mg/kg) change rhythm of the heart matched group 12 0 10,/12 12,/12 11 verapamil groups 2.0 8-16% 2/8 2/8 2

3.8 8-32% 1/8 1/8 0 Guan-fu base A hydrochloride group 20 8-25% 6/8 4/8 4

30 8 -28％ 2/8 1/8 0

Above result shows, Guan-fu base A hydrochloride has the antagonism calcium chloride similar to verapamil to bring out the chamber to quiver and reduce mortality rate.

3. the ouabain arrhythmia of bringing out

Ouabain is brought out the effect of Canis familiaris L. ventricular arrhythmia

(1) to anaesthetizing the effect of Canis familiaris L.

5 of adult dog, body weight are 9.5 ± 0.5kg, pentobarbital sodium (30mg/kg) iv anesthesia.Record II helical pitch electrocardiogram, slowly inject ouabain solution with method gradation such as Allen by femoral vein, first dose is 40 μ g/kg, and iv 20 μ g/kg again behind the 30min are later on every 15min, iv 5 μ g/kg, until sVT occurring, then, the iv Guan-fu base A hydrochloride, transfer hole to by chamber speed, calculate its effective dose.The result shows that the mean dose that 5 Canis familiaris L. iv ouabains bring out ventricular tachycardia is 58.1 ± 3.3 μ g/kg.Can make the chamber quick-recovery behind the iv Guan-fu base A hydrochloride in the 1-3min is sinus rhythm, and its mean effective dose is 11.3 ± 1.3mg/kg.Show that Guan-fu base A hydrochloride causes cardiotoxicity to ouabain and brings out arrhythmia therapeutical effect is arranged.

(2) to the effect of clear-headed Canis familiaris L.

Get 5 clear-headed taming Canis familiaris L.s, body weight is 10 ± 0.5kg, record II electrocardiogram, and after forelimb iv ouabain 60 μ g/kg brought out lasting ventricular tachycardia, slowly iv Guan-fu base A hydrochloride 9mg/kg transferred sinus rhythm to by chamber speed behind the 1-2min.Continue to observe 30-60min and arrhythmia do not occur.The result shows that Guan-fu base A hydrochloride causes the arrhythmia that cardiotoxicity brings out to ouabain therapeutical effect is arranged.

4. to bring out anesthesia Cor Canitis rule not normal for acetylcholine

Acetylcholine is brought out the protective effect of Canis familiaris L. atrial fibrillation

Cause the dog atrial fibrillation with reference to the Klevans method.5 of dogs, body weight 12 ± 1kg.Pentobarbital sodium iv anesthesia, under the artificial respiration, the 4th intercostal is opened breast from the right side, a bipolar electrode is fixed in right atrium, the recording right atrial electrograph.Drip in right atrium with 10% acecoline 0.05ml, gently press with blade holder then, bring out the sustainable 4-16min of atrial fibrillation.Attack once the record atrial fibrillation time with acetylcholine every 15min.Measuring secondary before the administration is worth in contrast.10min after the second time, acetylcholine was attacked has annotated in the iv Guan-fu base A hydrochloride 10-20mg/kg, 3min.

The result is in 10 control experiments of 5 dogs, and bringing out the atrial fibrillation persistent period for the first time is 7.6 ± 1.7min, and the atrial fibrillation persistent period is 8.7 ± 4.3min for the second time.

2 dogs, atrial fibrillation does not take place in iv Guan-fu base A hydrochloride 10mg/kg in the 15min, just take place to 20min.For the second time repeat iv 10mg/kg, still identical effect can occur.3 dogs in addition, iv 20mg/kg, suppressing the effect that atrial fibrillation takes place can reach about 35min.The result shows that the iv Guan-fu base A hydrochloride has obvious inhibition acetylcholine and brings out atrial fibrillation, and the dosage increase, and effect is more remarkable.

5. resist CaCl ₂-Ach liquid brings out the mice room and pounces on (atrial fibrillation) effect

60 of mices, body weight 25 ± 2g, pentobarbital sodium 60mg/kg ip anesthesia, be divided into normal saline group and five various dose groups (two adjacent groups dosage ratio is 0.85) at random, 10 every group, ♂ ♀ half and half, 1.5min behind iv Guan-fu base A hydrochloride or the equivalent normal saline, ivCaCl ₂-Ach liquid capacity is 10ml/kg (CaCl ₂6mg/kg, Ach30 μ g/ml face use preceding preparation), in 10s, annotated, with 6501 type electrocardiographic recorder II helical pitch ECG, observe f or F pounces on as the room or the positive index of atrial fibrillation.Do not appear as fully after the medication effectively, compare, calculate ED with simplifying probit's general formula by direct short cut technique and matched group ₅₀(Ye Dingzhong, Zhu Siming applied physiology magazine 1986; 2:64).The result shows that Guan-fu base A hydrochloride can obviously resist CaCl at 13.4mg/kg ₂-Ach liquid brings out the mice room and pounces on (quivering).Suppress 40%, 21mg/kg resists (seeing Table 7), its ED fully ₅₀Be 12.4 ± 1.5mg/kg.Therapeutic index is 13.8.

Table 7 antagonism CaCl ₂-Ach liquid brings out the mice room and pounces on (quivering) effect N=10 group dosage (mg/kg) effective percentage (%) ED ₅₀(mg/kg) normal saline group 0 Guan-fu base A hydrochloride group 1 21.0 100 ^*Guan-fu base A hydrochloride group 2 16.8 70 ^*Guan-fu base A hydrochloride group 3 13.4 40 ^*12.4 ± 1.5 Guan-fu base A hydrochloride groups 4 10.8 30 Guan-fu base A hydrochloride groups 5 8.6 20

* P＜0.05; * P＜0.01 (comparing) with the normal saline group

Isolated test shows

1. Guan-fu base A hydrochloride (50 μ g/ml) arrhythmia that the isolated rat heart coronary artery ligation is brought out has protective effect, and is similar to lignocaine (50 μ g/ml) effect.

2. Guan-fu base A hydrochloride (20-30 μ g/ml) can suppress that low potassium high calcium brings out isolated rat heart chamber speed and quiver in the chamber, and is similar to verapamil (2-3 μ g/ml) effect.

Show in the body test

1.iv Guan-fu base A hydrochloride (3,6,12mg/kg) the electricity irritation Cor Leporis is caused to quiver to be significantly improved in the chamber and cause quiver threshold effect and lignocaine (3,6,12mg/kg) effect is similar.

2.iv or the ip Guan-fu base A hydrochloride (10,25,40mg/kg) the rat ventricular arrhythmia that all aconitine is brought out has significant protective effect.The rat ventricular arrhythmia that iv Guan-fu base A hydrochloride injection (10,25,40mg/kg) all brings out aconitine has significant protective effect, and its effect is better than quinidine hydrochloride.

3. (mean effective dose is to make ouabain cause that property process aroused in interest in chamber transfers regular sinus rhythm to behind 11.3 ± 1.3mg/kg in the 1-3min to the attached first element of anesthesia Canis familiaris L. iv hydrochloric acid first.

Guan-fu base A hydrochloride (20,30mg/kg) brings out the rat chamber and quivers and reduce mortality rate 4.iv can resist calcium chloride.Effect is similar to verapamil (2,3,8mg/kg).

5.iv Guan-fu base A hydrochloride (10,20mg/kg) have the effect that obvious inhibition acetylcholine brings out the Canis familiaris L. atrial fibrillation, can keep 10-35min.

6.iv Guan-fu base A hydrochloride can resist CaCl ₂-Ach liquid brings out the mice room and pounces on (quivering), its ED ₅₀Be that 12.4 ± 1.5mg/kg therapeutic index is 13.8.

7. Guan-fu base A hydrochloride 10,20,40mg/kg have the ventricular arrhythmia that significant inhibition Harris second phase ligation dog coronary artery causes, increase sinus rhythm, and have certain decreased heart rate effect.Guan-fu base A hydrochloride suppresses the ED of this ventricular rhythm ₅₀Be about three times of quinidine.

Three, electric physiological action mechanism

Observe the influence of Guan-fu base A hydrochloride with cardiovascular pharmacodynamics technical manual required standard glass microelectrode method to the guinea pig papillary muscle action potential.

The making Cavia porcellus of Cavia porcellus nipple specimen, body weight 300-400g, ♀ ♂ is regardless of, and heart is taken out in the back of hitting unconsciously, at logical 95%O ₂+ 5%CO ₂Tyrode ' s liquid in prepare papillary muscles of right ventricle, specimen is fixed in horizontal direction in the water bath of 1ml, with 37 ± 0.5 ℃ Tyrode ' s liquid circulation perfusion, flow velocity is 4ml/min, pH 7.2-7.4.Tyrode ' s liquid composition following (mmol/L): NaCl 136.7, and KCl 5.4, CaCl ₂1.8, MgCl ₂1.05, NaH ₂PO ₄1.1, NaHCO ₃11.9, Glucose5.5.

Stimulating electrode is the platinum wire bipolar electrode, stimulus frequency 1Hz, and the wide 1ms of stimulus wave, stimulus intensity are 150% of diastolic threshold.Specimen begins experiment after stablizing 1hr.

Effective refractory period is measured: at basic stimulus frequency is on the basis of 1Hz, stimulates before per 8 stimulations inserted for 1 coming half year.The ripple of stimulus wave is wide before phase is the twice that basilic rhythm stimulates with intensity.Interval between stimulation and basilic rhythm stimulate before the change phase can cause that in the hope of preceding stimulation the shortest time that AP produces is spaced apart ERP.

The dynamic process that frequency dependence suppresses: the train with different frequency comes body to move specimen (ISI=4800,2400,1200,600,300,200ms), uses first _MaxValue and new stable state _MaxBetween difference account for first _MaxPercentage recently weigh the size of this rate dependent block (RDB) effect.The recovery dynamic process of research RDB is to be used in to provide the method for premature beat to carry out with different intervals after train is ended.

The making rabbit of sinuatrial node specimen, body weight 2.0-3.0Kg, ♀ ♂ is regardless of, and the back of hitting unconsciously is taken out heart rapidly, puts into through 95%O ₂+ %CO ₂In the saturated Tyrode ' s liquid, cut off right atrium, cut sinuatrial node part (DE Carvalhe AP, DE Mello WC, Hoffman BF.Am J Physiol 1959 along boundary's ridge; 196:483), the specimen endocardial surface is fixed in temperature and is in 36 ± 0.5 ℃ the thermostatic bath up, begins experiment behind the balance 1hr.

Contrast and pharmaceutically-active observation are all finished in all experiments in same cell or same specimen, the result adopts the paired t-test statistical disposition.Verapamil hydrochloride (the favorable to the people pharmaceutical factory in Guangdong Province), quinidine, Lopantcol (Hubei the medical professionals provide), mexiletine (Guangzhou medicine inspecting institute provides).

1. Guan-fu base A hydrochloride and quinidine, mexiletine, Lopantcol are to the influence of guinea pig papillary muscle fast response AP accumulation dose regimen, and drug level increases progressively with 0.5logmol/L, delivery time 30min.Guan-fu base A hydrochloride 0.86 μ mol/L, quinidine 0.84 μ mol/L, mexiletine 0.84 μ mol/L, Lopantcol 0.81 μ mol/L can significantly reduce _Max, they are to _MaxAll present obvious concentration dependent with the reduction effect of APA.Except that Lopantcol can be raised RP when the high concentration, other three medicine did not have obvious influence to RP.Guan-fu base A hydrochloride (0.26-8.6 μ mol/L), quinidine (0.28-28 μ mol/L) make APD ₅₀, APD ₉₀, ERP all prolongs, mexiletine (2.8-28 μ mol/L) makes APD ₅₀, APD ₉₀Shorten, and ERP prolongs relatively.Lopantcol (0.27-8.6 μ mol/L) is to APD ₅₀, APD ₉₀, ERP do not make significant difference.V _MTFor the rapid depolarization starting point to _MaxRequired time and 0 ratio of depolarization persistent period mutually, this value can be used as weighs Na ⁺The index of accelerator during channel opener.Experimental result shows that Guan-fu base A hydrochloride, quinidine, mexiletine, Lopantcol all can make Vmt significantly increase according to concentration, and _MaxPairing transmembrane potential V _MvConstant on-25mV left and right sides potential level.

To ERP, Guan-fu base A hydrochloride also presents not same-action to Guan-fu base A hydrochloride to the influence of effective refractory period, and in above-mentioned low strength range, ERP is absolute prolongation: during high concentration, ERP does not have obvious change.But because APD ₉₀Shorten ERP/APD ₉₀Ratio but enlarge markedly, illustrate that ERP prolongs relatively.

Guan-fu base A hydrochloride, quinidine, Lopantcol, mexiletine are to _MaxThe influence of the dynamic process that frequency dependence suppresses:

At identical ISI (300ms), produce under the situation of roughly the same RDB the relatively beginning dynamic process of four medicines.When their concentration is followed successively by 32 μ mol, 16 μ mol, 1 μ mol, during 25 μ mol, the RDB that their produce is respectively 40%, 48%, 42% and 48%, but the speed that they produce RDB is obviously different.Except that mexiletine, the speed of the RDB of other three medicines is obviously different, and the beginning process of RDB meets the rule of single index curve.To reach each before the stable state _MaxDeduct stable state _MaxFor Δ ', with Δ ' be standardized as Δ=Δ '/first _Max, this Δ value meets following equation:

N is the sequence number of train pulse herein, with 1/ τ _oWeigh RDB and begin dynamic (dynamical) speed, promptly RDB opens dynamic (dynamical) speed constant covered with clouds.The relation of Δ value and n number is used

Carry out process of fitting treatment, 1/ τ _oValue is: Guan-fu base A hydrochloride 0.1333AP ^-1, quinidine 0.1689AP ^-1, Lopantcol 0.0598AP ^-1It is too fast that the RDB of mexiletine begins the dynamic process generation, can not weigh with said method.

When ISI=600ms, compare the recovery process that four medicines produce RDB.If first of train _MaxBe peak value _Max, make θ=(peak value _Max-premature beat _Max)/peak value _Max, then θ meets following equation:

, on behalf of train, t be terminated to the time that premature beat stimulates beginning, τ _ReBe _MaxFrom the time constant that RDB recovers, θ value and t time relation are used

Carry out process of fitting treatment, get τ _ReValue is: Guan-fu base A hydrochloride 45 (s), Lopantcol 26 (s), quinidine 18 (s), mexiletine are 2.6 (s).

2. Guan-fu base A hydrochloride, verapamil (Ver) is pressed characteristics and zone (Bleeker WK, Mackaay AJC, the Mireille MP of literature method according to excellent easypro pacemaker cell to the influence of rabbit sinuatrial node (SAN) advantage pacemaker cell AP, Bouman LN, BeckerAE.Circ Res 1980; 46:11.Brown HF.Physiol Rev 1982; 62:505), at first detect the advantage pacemaker cell and draw spontaneous AP, stablize 15min after, the record AP in contrast, add medicine then.

(1) behind adding Guan-fu base A hydrochloride (the 50 μ mol/L) 30min, the APA of SAN action potentials of cells does not have significant change, the 0 very fast rate (MRD) of being divided by, diastolic depolarization speed (RDD) and average repolarization speed (MRR) are reduced to 82.6% before the medication respectively, 87.7% and 86.11% (N=10, p＜0.01), spontaneous activity frequency (SFF) reduces to 92% (N=10, p＜0.001), relaxing period interval (DI), total duration of diastole (DDT), 0 phase depolarization time (DT) and APD (the needed time from the action potential peak to maximum diastolic potential) all prolong.

(2) add Ver 0.5 μ mol/L similar effect is also arranged.Behind the 15min, and compare before the administration, APA drops to 74% (N=10 before the medication, p＜0.001), MRD and RDD are respectively 36.4%, 87% (N=10, p＜0.05) before the medication, and MRR is reduced to 58.1% (N=10, p＜0.001), SFF reduces to 84% (N=10, p＜0.001), and DDT extends to 123% preceding (N=10 of medication, p＜0.05), DI, DT and APD all prolong.

Electrophysiologic study is the result show: Guan-fu base A hydrochloride can reduce in concentration dependent ground _MaxAnd APA, in low concentration (the 0.26-8.6 μ mol/L) scope, APD ₅₀, APD ₉₀Prolong APD during high concentration (26 μ mol/L) with ERP ₅₀, APD ₉₀Shorten, and ERP does not have obvious change, but ERP/APD ₉₀Ratio significantly increase.And to _MaxInhibition tangible frequency dependence and potential dependent are arranged.To fast Na ⁺Passage does not influence myocardial excitability when the specificity retardance is arranged, and this helps clinical practice.At scarce O ₂, high K ⁺In the myocardial cell of low pH, Guan-fu base A hydrochloride suppresses _Max, APA is more obvious with the effect of relative prolongation ERP.This is useful to treating quick type and ischemic arrhythmia.Thereby the inhibition antidromic conduction prolongs refractory stage, eliminates unidirectional conduction block or makes it become bidirectional block, and the reciprocal excitation that this will help blocking the ischemia diseased region benefits the ARR generation of prevention.

Guan-fu base A hydrochloride is by suppressing the sinus node cells long response time electrical activity spontaneous frequency of sinuatrial node that slows down, rather than by beta-2 adrenoceptor and M cholinoceptor and also be different from calcium channel blocker, is to directly act on sinuatrial node.Its negative inotropic action is also irrelevant with beta receptor.The advantage of Guan-fu base A hydrochloride is that its negative chronotropic action is strong about 9 times than negative inotropic action, and verapamil hydrochloride has only 1.7 times, and sinus standstill does not take place Guan-fu base A hydrochloride again, and verapamil hydrochloride has 4/5 sinus standstill takes place.So than verapamil hydrochloride come safety.It seems to have Na simultaneously at present ^-, Ca ⁺⁺The medicine of antagonism has significant treatment advantage.And Guan-fu base A hydrochloride is compared with quinidine, mexiletine and Lopantcol, and it is to I _NaInhibiting onset concentration is close.Produce the beginning dynamic characteristic of RDB according to them, Guan-fu base A hydrochloride and quinidine are close.

The toxicity comparative test of Guan-fu base A hydrochloride and other anti-arrhythmic

Guan-fu base A hydrochloride and lidocaine hydrochloride, verapamil hydrochloride, propranolol are to the comparison of cardiovascular effect

To anesthetized rat blood pressure, heart rate, cardiac electrical influence

Get 35 of rats, body weight is 220 ± 15g, ♂ ♀ dual-purpose.Be divided into the normal saline matched group, lidocaine hydrochloride group, propranolol group, verapamil hydrochloride group and Guan-fu base A hydrochloride group.Urethane 1.2g/kgip anesthesia.With the administration of WSQ type trace transfusion device femoral vein constant infusion (0.18-0.19ml/min).The carotid artery intubate is measured blood pressure, measures II helical pitch electrocardiogram with ECG-6501 type electrocardiograph.Every kind of medicine is got blood pressure, heart rate, the electrocardio that six dosage are observed these six dosage, and writes down the lethal dose of every kind of medicine.

The mean lethal dose (MLD) verapamil hydrochloride of rat (N=7) is 20.2 ± 3.1mg/kg, and propranolol is 46.0 ± 15.2mg/kg, and lidocaine hydrochloride is 67.2 ± 14.8mg/kg, and Guan-fu base A hydrochloride is 343.8 ± 52.1mg/kg.The toxicity maximum that shows verapamil hydrochloride, Guan-fu base A hydrochloride toxicity minimum.The effect of verapamil hydrochloride decreased heart rate is the strongest, but hypotensive effect and toxicity are too big.Increase with verapamil hydrochloride dosage, engender auriculoventricular block (the QRS ripple comes off), the P ripple disappears, and Guan-fu base A hydrochloride is in decreased heart rate, and its toxicity is relative with hypotensive effect less.

Example 1:

The preparation of Guan-fu base A hydrochloride

With industrial ethanol percolate extraction Radix Aconiti Coreani 1000 grams, medical material carries out general pulverizing routinely, collects the ethanol liquid of percolation, reclaim ethanol, obtain extractum, add 1% dissolving with hydrochloric acid, acid liquid is transferred pH3, leaves standstill sour water liquation glue, precipitates in the filtering acid liquid, use defat with petroleum ether, divide and remove petroleum ether layer, alkalize acid liquid to pH5 with dense ammonia deuterium, the reuse chloroform extraction, collect chloroform solution, the reclaim under reduced pressure chloroform obtains the Guan-fubase A crude product, use dissolve with ethanol, filter, pure liquid adds hydrochloric acid and transfers to pH4, places crystallize, sucking filtration, get white crystals 3 grams, reuse dehydrated alcohol recrystallization, dry that white crystals 2.1 restrain at 100 ℃.Yield 2.1%0, MP.301 ℃.

Elementary analysis: C ₂₄H ₃₁NO ₆HCl.

Value of calculation (%): C 61.86; H 6.92; N 3.01; O 20.6; Cl l7.61.

Experiment value (%): C 61.86; H 6.98; N 3,12; O 22.2.

High resolution mass spectrum: m/z 465.97 (M ⁺)

Molecular weight: 465.97

Example 2

The preparation of hydrobromic acid Guan-fubase A

Soak routinely, 25 kilograms of percolation Radix Aconiti Coreanis, medical material is conventional to be pulverized, and soaks 5 days earlier with industrial alcohol, after carry out diafiltration, collect pure liquid, reclaim ethanol, obtain extractum, add 2% dissolving with hydrochloric acid extractum, transfer to pH3, inclining acid liquid, leaves standstill and analyses glue, the filtering precipitation, use the chloroform defat, divide and remove chloroform layer, acid liquid reuse 2%NaOH-water liquid transfers to pH10, use dichloromethane extraction, collect dichloromethane solution, the reclaim under reduced pressure dichloromethane obtains the Guan-fubase A crude product, add 95% dissolve with ethanol, alcohol liquid adds hydrobromic acid, transfers to pH4.5, places crystallize, sucking filtration, get white crystals 300 grams, use the dehydrated alcohol recrystallization, dry that white crystals 37.5 restrain at 90 ℃, yield 1.5 ‰, MP285 ℃.

Free alkali elementary analysis: C ₂₄H ₃₁NO ₆

Value of calculation (%): C 67.1; H 7.28; N 3.26; O 22.4.

Experiment value (%): C 67.29; H 7.33; N 3.36; O 21.98.

High resolution mass spectrum: m/z 429.2072 (M ⁺)

Molecular weight: 429.2072 (value of calculation 429.2142)

Example 3

The preparation of Guan-fubase A

The gram Radix Aconiti Coreanis are conventional pulverizes with 2000, with 80% soak with ethanol 4 days, collect soak, recovery ethanol, get extractum, add diluted hydrochloric acid dissolution, transferring pH is 4, leaches acid liquid, with chloroform extraction three times, remove chloroform solution, acid liquid is transferred pH to 9 with strong aqua ammonia, uses dichloromethane extraction, the combined dichloromethane extract reclaims filtrate to doing, and gets the Guan-fubase A crude product, with 95% dissolve with ethanol crude product, recrystallization, reuse acetone recrystallization, dry that Guan-fubase A 4 restrains yield 2 ‰, MP.198-199 ℃ at 70 ℃.

Elementary analysis: C ₂₄H ₃₁NO ₆

Value of calculation (%): C 67.11; H 7.28; N 3.26; O 22.37.

Experiment value (%): C 67.15; H 7.33; N 3.36; O 22.16.

High resolution mass spectrum: m/z 429.2074 (M ⁺)

Molecular weight: 429.2074 (value of calculation 429.2142)

Example 4

The preparation of Guan-fubase A

The same example of previous action is the Guan-fubase A crude product, through alumina column chromatography, and routine operation, (neutral alumina, III-IV level) uses the cyclohexane extraction of different proportion then successively, cyclohexane/ethyl acetate, methanol gradient elution, through thin layer chromatography inspection stream part, stream part 30～54 is a chromatography speckle, merge eluent, be recycled to driedly, use acetone recrystallization, at 60 ℃ of dry Guan-fubase As, MP.199 ℃

Elementary analysis: C ₂₄H ₃₁NO ₆

Value of calculation (%): C 67.1; H 7.28; N 3.26; O 22.4.

Experiment value (%): C 66.90; H 7.30; N 3.40; O 22.40.

High resolution mass spectrum: m/z 429.2080 (M ⁺)

Molecular weight: 429.2080 (value of calculation 429.2142)

Example 5

Injection

With Guan-fu base A hydrochloride 100.0 gram and sodium chloride for injection 77.2 grams, add 70% water for injection of full dose, stirring makes dissolving, adds to the full amount of water for injection 2000 milliliters.Add 1.5% pin activated carbon, stirred coarse filtration deactivation carbon 10 minutes at 50 ℃.(0.8 μ m) is filtered to clear and bright with microporous filter membrane.Survey pH value (should be 3.0-4.5).Fill N ₂, embedding.100 ℃ of circulation steam sterilizations 30 minutes promptly get the Guan-fu base A hydrochloride injection.Per 2 milliliters of Guan-fu base A hydrochlorides that contain 100.0 milligrams.

Example 6

Sterile powder for injection

Guan-fu base A hydrochloride 100.0 gram and sodium chloride for injection 77.2 are restrained,, put coldly, send into aseptic subpackaged chamber and be distributed into 1000 aseptic powder injections, 100.0 milligrams of every hydrochloric Guan-fubase As in 100 ℃ of vacuum dryings 6 hours.

Claims

1. from Chinese herbal medicine, use solvent extraction for one kind, reclaim extracting solution, obtain extractum, the extractum commentaries on classics is dissolved in the sour water, transfer pH2～4, sour water liquation glue, precipitation prepares the method for Guan-fubase A in the filtering acid liquid, it is characterized in that used Chinese herbal medicine is Ranunculaceae Radix Aconiti Coreani Aconitum coreanum (Levl) Rapaics, and acid liquid leaves standstill analyses glue, the acid liquid of filtering post precipitation lipotropy organic solvent degreasing, remove the lipotropy organic solvent layer, the alkalization acid liquid is to pH5～10, the water liquid of reuse lipotropy organic solvent extraction alkalization, collect extract, reclaim organic solvent, obtain the Guan-fubase A crude product, refining repeatedly, 50～100 ℃ of dryings, obtain the Guan-fubase A highly finished product.

2. according to the described method for preparing Guan-fubase A of claim 1, it is characterized in that the organic solvent of defat or extraction usefulness is halogenated hydrocarbons, ethyl acetate, methyl acetate, petroleum ether, cyclohexane extraction or benzene.

3. according to the described method for preparing Guan-fubase A of claim 1, the alkali of the acid liquid that it is characterized in that alkalizing is 1～5% sodium hydroxide water liquid, 1～5% potassium hydroxide water liquid, strong aqua ammonia or Na ₂CO ₃

4. according to the described method for preparing Guan-fubase A of claim 1, the organic solvent that it is characterized in that recrystallization is methanol, ethanol, benzene, ether or acetone.

5. according to the described method for preparing Guan-fubase A of claim 1, it is characterized in that when operating recrystallization routinely, adding acceptable acid on the human physiology, transferring to pH3～5, make the salt of Guan-fubase A with organic solvent.