CN102008521B - Application of Chinese trumpet creeper extract to preparation of medicament for treating and resisting cerebral ischemia - Google Patents
Application of Chinese trumpet creeper extract to preparation of medicament for treating and resisting cerebral ischemia Download PDFInfo
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- CN102008521B CN102008521B CN2010105764916A CN201010576491A CN102008521B CN 102008521 B CN102008521 B CN 102008521B CN 2010105764916 A CN2010105764916 A CN 2010105764916A CN 201010576491 A CN201010576491 A CN 201010576491A CN 102008521 B CN102008521 B CN 102008521B
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Abstract
The invention relates to application of Chinese trumpet creeper extract to preparation of a medicament for treating and resisting cerebral ischemia, which can effectively solve the problem of the treatment of the cerebral ischemia. The technical scheme for solving the problem comprises the following steps of: adding 60 to 90 mass percent ethanol in an amount which is 10 times the weight of the Chinese trumpet creeper into the Chinese trumpet creeper, soaking for 30 minutes, extracting for one hour and filtering to obtain liquid medicament; adding 60 to 90 mass percent ethanol in an amount which is 8 times the weight of the Chinese trumpet creeper into medicament dregs, extracting for one hour, and filtering to obtain liquid medicament; and merging the liquid medicament obtained twice, concentrating in a water bath, performing vacuum drying to obtain the Chinese trumpet creeper extract which can be effectively used for preparing the medicament for treating and resisting the cerebral ischemia. The Chinese trumpet creeper extract is easy to prepare, has low cost and is innovation of the medicament for treating and resisting the cerebral ischemia.
Description
One, technical field
The present invention relates to field of medicaments, the application of particularly a kind of Flos Campsis extract in preparation treatment anti-cerebral ischemia drugs.
Two, background technology
Flos Campsis is the Chinese medicine that the addiction effect is gone in promoting the circulation of blood that has of China's tradition use.Flos Campsis Flos Capsis is Bignoniaceae plant the reach the clouds dried floral of C.radicans (L) Seem. of Campsis grandiflora (Thunb.) K.sehum. and America of reaching the clouds.Nature and flavor are sweet, sour, cold, return liver, pericardium channel, removing heat from blood is arranged, change addiction, the effect of dispeling the wind is mainly used in menoxenia, amenorrhea disease trace, puerperal swelling of the breast, rubella is rubescent, skin expectorant is itched, seat skin ulcer etc.
The modern study report: Flos Campsis contains triterpenes (oleanane type triterpene, ursane type triterpene, phenylpropanol glycoside), flavonoid, anthocyan, alkaloids, volatile oil etc.Flos Campsis can suppress people's phthalidyl coenzyme A-cholesterol phthalidyl transferring enzyme-1 activity: the arjunolic acid in the Flos Campsis, mountain santenic acid, 23-hydroxyl ursolic acid and the acid of laughable Soviet Union; Inhibition with high relatively human acyl coenzyme A-cholesterol acidic group transferring enzyme-1 (h-ACAT) is active.Can suppress the unpregnancy uterine contraction, strengthen pregnant uterine contraction: Flos Campsis and trumpet creeper decocting fluid power highly significant ground suppress the unpregnancy Mouse Uterus and shrink; Acetone in the Flos Campsis ethanol extract: methanol (1: 1) position can strengthen the contraction intensity of the pregnant Mouse Uterus flesh bar that exsomatizes extremely significantly.Have antioxidation, antiinflammatory and analgesia itching-relieving action: Flos Campsis 50% ethanol extraction can have significant protective effect to the HSF to resistive to hydrogen peroxide, and the lactic acid dehydrogenase that can also significantly suppress hydrogen peroxide-induced discharges and dna break.Flos Campsis is processed spray, and coating medicine-feeding causes the increase of mouse skin capillary permeability to histamine has remarkable inhibitory action, gastric infusion can make hot plate method mice chronic pain model bitterly the value of cutting off from increase significantly, have the effect of slight inhibition mice autonomic activities.Can improve blood circulation, reduce blood flow, suppress thrombosis: Flos Campsis decocting liquid centrifugal evaporate to dryness behind the ethanol precipitate with ethanol; Be dissolved in and process 2% crude extract solution in 0.9% sodium chloride injection; Gastric infusion can be accelerated the old rats VPV, increases the capillary network cross point; Expand little blood vessels caliber, inhibiting erythrocyte aggregation.The Flos Campsis methanolic extract reduces egg-white lysozyme sensitized mice blood flow has significant improvement effect.Flos Campsis decocting liquid has the usefulness that suppresses contraction to isolated pig coronary artery bar; 50% Flos Campsis, leaf decoct have inhibitory action to shigella flexneri and Bacillus typhi.
Cerebral ischemia belongs to the scope ischemic cerebrovascular of the traditional Chinese medical science " ischemia apoplexy "; It is one of principal disease that threatens human health and existence; It is common ailment of middle-aged and old; Characteristics with high incidence, high prevalence, high mortality, high disability rate, high relapse rate are a kind of diseases of present keypoint control.Angiostenosis or obturation during cerebral infarction; Cause partial ischemia of cerebral tissue or necrosis, its pathogeny belongs to the traditional Chinese medical science " blood stasis " category, and the traditional Chinese medical science adopts the kidney invigorating method (remove free radical, improve hemorheology) for a long time; Activating blood and removing stasis Method (expelling wind and activating blood circulation, promoting tissue regeneration by removing blood stasis, blood circulation promoting and blood stasis dispelling); The purging FU-organs method of reducing phlegm is that the main rule of treatment is treated primary disease, uses corresponding decoction for the treatment cerebral ischemia and has obtained effect preferably, has obtained widely certainly.But do not see the open report or the related data of useful Flos Campsis extract treatment cerebral ischemia so far.
Three, summary of the invention
To above-mentioned situation; For overcoming the defective of prior art, the object of the invention just provides the application of a kind of Flos Campsis extract in preparation treatment anti-cerebral ischemia drugs, can effectively solve the treatment problem of cerebral ischemia; The technical scheme of its solution is: it is 60%～90% ethanol that Flos Campsis is added 10 times of weight, mass concentration; Soaked 30 minutes, extracted 1 hour, cross and filter medicinal liquid; Medicinal residues add again 8 times of Flos Campsis weight, mass concentration is 60%～90% ethanol, extracts 1 hour, cross and filter medicinal liquid; Merge medicinal liquid twice, water-bath concentrates, vacuum drying, gets Flos Campsis extract, is effective to preparation treatment anti-cerebral ischemia drugs, and Flos Campsis extract preparation of the present invention is simple, and cost is low, is the innovation of treating on the anti-cerebral ischemia drugs.
Four, the specific embodiment
Below in conjunction with embodiment and concrete condition, specific embodiments of the invention elaborates.
In practical implementation; Flos Campsis extract of the present invention is to get Flos Campsis dry product (dried floral) 10g, and add 10 times of weight mass concentrations and the be 70% ethanol 100g (or 100ml) of (or 60%, 80%, 90% all can) soaked 30 minutes; Extracted 1 hour, and crossed and filter medicinal liquid; Medicinal residues add 8 times of Flos Campsis weight mass concentrations be 70% ethanol 80g (or 80ml) of (or 60%, 80%, 90% all can) again, extract 1 hour, cross and filter medicinal liquid; Merge medicinal liquid twice, place in the evaporating dish and in water-bath, concentrate vacuum drying; Get Flos Campsis extract; Be effective to preparation treatment anti-cerebral ischemia drugs, this extract fully proves the effect with anti-cerebral ischemia through zoopery; Can be used in the preparation treatment anti-cerebral ischemia drugs, relevant experimental data is following:
1, experiment material
1.1 medicine and reagent
A, Flos Campsis extract of the present invention:
Flos Campsis Flos Capsis is Bignoniaceae plant Campsis grandiflora (Thunb.) K.sehum. that reaches the clouds, dried floral; Get the Flos Campsis medical material, add the ethanol of 10 times of amounts 70% (or 60%, 80%, 90% all can), soak 30min, extract 1h, released liquor; Medicinal residues add the ethanol of 8 times of amounts 70% (or 60%, 80%, 90% all can) again, soak 30min, extract 1h.Merge for the first time and secondary medicinal liquid, the medicinal liquid after merging is placed in the evaporating dish in water-bath, concentrate, vacuum drying, get Flos Campsis extract;
B, dexamethasone sodium phosphate injection production, lot number 0907183; Jiangsu Lianshui Pharmaceutical Co., Ltd. produces, and YANGXUE QINGNAO KELI is produced lot number Z10960082 by Tianjin Tasly Pharmaceutical Co., Ltd; The NOS testing cassete, Nanjing is built up bio-engineering research and is provided, lot number 20100626; The NO testing cassete, Nanjing is built up bio-engineering research and is provided, lot number 20100626; The ATP detection kit, building up bio-engineering research by Nanjing provides, lot number 20100114,20100613; The Coomassie brilliant blue testing cassete, Nanjing is built up bio-engineering research and is provided, lot number 20100626; Nimodipine tablet, universal love Pharmaceutical Co in Xi'an provides, lot number 090117; HUATUO ZAIZAO WAN, Qixing Pharmaceutical Co., Ltd., Guangzhou provides, lot number 7438.
12 laboratory animals
White mice, cleaning level, the Kunming kind is provided by Hebei province's Experimental Animal Center, 18～22g, male and female half and half, the quality certification numbers 912102; Animal quality certification numbering: 28～32g, male, the quality certification numbers 1006019.
1.3 experimental apparatus
Automatic blood flow graph (LBY-N6B&LBY-N6K), Pulisheng Instruments Co., Ltd., Beijing produces; The UV-2000 ultraviolet-uisible spectrophotometer, UNICO(Shanghai) Instruments Co., Ltd. produces; The TGL-16G desk centrifuge, Anting Scientific Instrument Factory, Shanghai produces; Electronic analytical balance, Ao Haosi (Shanghai) company produces; DZKW-4 type electronic thermostatic water-bath, the bright forever Medical Instruments in Beijing factory produces; DY89-1 type electric driven glass refiner, the new sesame bio tech ltd in Ningbo is produced; Adjustable pipette, Lei Bo Analytical Instrument Co., Ltd in Shanghai produces.
2. experimental technique and result
2.1 influence to mice normal pressure anoxia enduring
Get 50 of Kunming mouses, male and female half and half, body weight 19～22g; Be divided into 5 groups at random, 10 every group, irritate the Flos Campsis ethanol extract suspension (0.2g/ml that obeys large, medium and small dosage respectively; 0.1g/ml, 0.05g/ml; 0.2ml/10g), (0.1g/ml is 0.2ml/10g) and with the normal saline (0.2ml/10g) of volume for the YANGXUE QINGNAO KELI suspension.Administration every day 1 time, successive administration 5 days.After the last administration 1 hour, mice is placed the 150mL wide mouthed bottle of demarcating through capacity, the built-in 10g sodica calx of bottle is in order to absorbing carbon dioxide; Bottle cap is coated with rapidly with the vaseline sealing, makes it air tight, immediately timing; With the respiratory arrest is index, the record time-to-live of mice in airtight port grinding bottle.The result sees table 1.
Table 1 Flos Campsis alcohol extract is to the influence
of mice normal pressure anoxia enduring
* represent with model group than P＜0.05, * * represent with model group than (P＜0.01) following with
Can find out that from last table with blank group ratio, but the equal significance of large, medium and small dosage Flos Campsis extract group and YANGXUE QINGNAO KELI group improves mice normal pressure hypoxia-bearing capability (P＜0.01), act as excellent with big or middle dosage Flos Campsis extract group.
2.2 influence to the mouse brain ischemia-reperfusion injury model
Get 60 of Kunming mouses, male and female half and half, body weight 19～22g is divided into 6 groups at random, 10 every group; Wherein make the cerebral ischemia re-pouring model for 5 groups, other has 1 group to be the blank group.5 groups of Flos Campsis ethanol extract suspension (0.2g/ml that irritate the large, medium and small dosage of clothes respectively of modeling type; 0.1g/ml, 0.05g/ml, 0.2ml/10g), YANGXUE QINGNAO KELI suspension (0.1g/ml; 0.2ml/10g) and with the normal saline (0.2ml/10g) of volume; The blank group is given the normal saline (0.2ml/10g) with volume, administration every day 1 time, successive administration 5 days.In last administration 1h (water 12h is can't help in fasting), 10% chloral hydrate intraperitoneal injection of anesthesia mice after mice is weighed faces upward and is fixed on the little rat holder; The cervical region sterilization, median incision, passivity is separated; Expose bilateral common carotid arteries, recover to sew up behind the perfusion behind the blocking-up bilateral common carotid arteries blood flow 30min, after each mice recovers to pour into 24h again; Take off cervical vertebra and put to death, get cerebral tissue rapidly and weigh, add 9 times of abundant homogenate of normal saline; High speed low temperature centrifugal machine is centrifugal, gets supernatant; Say requirement mensuration cerebral tissue protein content and ATP enzyme activity according to test kit.The result sees table 2.
Attaches 1 Coomassie brilliant blue protein determination Operations Sequence Chart
Mixing left standstill 10 minutes, in the 595nm place, and the 1cm optical path, the distilled water zeroing is surveyed and is respectively managed the OD value
Protein content (g/L)=(measuring pipe OD value-blank pipe OD value) * standard pipe concentration ÷ (standard pipe OD value-blank pipe OD value)
Attaches atpase assay Operations Sequence Chart in 2 brain homogenates
Mixing, 37 ℃ of water-baths 30 minutes are cooled to room temperature, at 660nm, 1cm optical path distilled water zeroing colorimetric
Annotate: the A pipe is control tube; The B pipe is Na
+K
+-ATP enzyme pipe; The C pipe is Mg
++-ATP enzyme pipe; The D pipe is Ca
++-ATP enzyme pipe;
Diluted sample multiple (2.5) * 6 ÷ sample protein content (mgprot/ml) in ATP vigor in the tissue (μ molPi/mgprot/hour)=(measuring pipe OD value-control tube OD value) ÷ standard pipe OD value * standard pipe concentration (0.5 μ mol/ml) * reaction system
Table 2 Flos Campsis extract is to the influence
of mouse brain ischemia model brain homogenate ATP enzyme activity
* represent with model group than (P＜0.01)
Can find out from last table, with blank group ratio, model group Na
+-K
+-ATP, Mg
++-ATP, Ca
++-ATP and Ca
++-Mg
++-ATP vigor significantly reduces (P＜0.01), explains and makes the cerebral ischemic model success.With the model group ratio, but the equal significance of large, medium and small dosage Flos Campsis extract group and YANGXUE QINGNAO KELI group improves mouse brain ischemia model brain homogenate Na
+-K
+-ATP, Mg
++-ATP, Ca
++-ATP and Ca
++-Mg
++-ATP vigor (P＜0.01).Act as excellent with big or middle dosage Flos Campsis extract group.
2.3 the mice blood stasis is merged the influence of cerebral ischemia re-pouring model
Get 216 of Kunming mouses, male, body weight 29～32g is divided into 9 groups at random, 24 every group; Wherein 1 group is that blank group, 1 group are that blood stasis model sham operated rats, 1 group are not hemopoietic stasis of blood model surgery group, and the 6 groups of whole hemopoietic stasis of blood of mice models add the operation hemopoietic stasis of blood and merge cerebral ischemic model in addition.The hemopoietic stasis of blood merges 6 groups of cerebral ischemic models; Irritate the Flos Campsis ethanol extract suspension (0.2g/ml of the large, medium and small dosage of clothes respectively; 0.1g/ml, 0.05g/ml, 0.2ml/10g), the HUATUO ZAIZAO WAN suspension (0.1g/ml, 0.2ml/10g), nimodipine suspension (0.02g/ml; 0.2ml/10g) reaching normal saline (0.2ml/10g) with volume, blank group, blood stasis model sham operated rats and not hemopoietic stasis of blood model surgery group are all irritated the normal saline (0.2ml/10g) with volume.Administration every day 1 time, successive administration 16 days; Administration in the 1st day simultaneously, 7 groups of equal back leg intramuscular injection of dexamethasone 0.8mg/kg every day (2 groups of injection equal-volume normal saline of not hemopoietic stasis of blood model) of hemopoietic stasis of blood model, 15d continuously.1h after the 16d administration (water 12h is can't help in fasting), each organizes mice through 10% chloral hydrate 0.033ml/10g intraperitoneal anesthesia, lies on the back and is fixed on the operating-table; The throat median incision, passivity is separated bilateral common carotid arteries, and blank group and blood stasis model sham operated rats are done sham-operation; All the other each all close bilateral common carotid arteries 10min with arteriole folder folder, recover perfusion 10min, reuse arteriole folder folder closes bilateral common carotid arteries 10min; Recover blood supply, sew up mice.Each is organized mice 24h eye socket after operation and gets blood survey hemorheology, and the result sees table 3; Put to death mice then, get cerebral tissue rapidly and weigh, add the abundant homogenate of normal saline, high speed low temperature centrifugal machine is centrifugal, gets supernatant, says according to test kit and surveys the ATP enzyme activity, surveys NO and NOS level.The result sees table 4, table 5.
WBV operating procedure: at first adjust the instrument level.Turn on the power switch subsequently, get into preheat mode, temperature value reaches 37 when spending on the screen to be shown, the beginning application of sample.Get the 1ml whole blood with sample injector, the liquid bath inwall is close in the sample injector outlet is slowly injected downwards, in order to avoid sneak into the aeration measurement result.Click the whole blood test menu, make an experiment, detect finish after, write down that whole blood is high, normal, basic to be cut.Click clear button, after the cleaning, get into next sample test.
Attaches NO mensuration sequence list in 3 brain homogenates
NO content in the tissue (μ mol/gprot)=(measuring the pipe absorbance-blank pipe of mensuration absorbance) * standard substance concentration ÷ (the blank pipe of standard pipe absorbance-standard absorbance) ÷ protein content (mgprot/ml)
Attaches NOS mensuration sequence list in 4 brain homogenates
TNOS vigor (U/ml)=(TNOS measures the blank pipe of pipe-TNOS) ÷ colour generation thing nanomole extinction coefficient * (reactant liquor cumulative volume ÷ gets liquid measure) ÷ (colorimetric optical path * response time) ÷ sample protein content (mggrot/L)
INOS vigor (U/ml)=(iNOS measures the blank pipe of pipe-iNOS) ÷ colour generation thing nanomole extinction coefficient * reactant liquor cumulative volume ÷ gets liquid measure * [1 ÷ (colorimetric optical path * response time)] ÷ sample protein content (mggrot/L)
Table 3 Flos Campsis extract merges the influence
of cerebral ischemic model whole blood viscosity to the mice blood stasis
▲ ▲Expression and blank group be than P＜0.01, * * represent with model group than P＜0.01
Can find out from last table; With blank group ratio; Not hemopoietic stasis of blood surgical groups, blood stasis not surgical groups and blood stasis add that the surgical groups blood flow is high, medium and low cuts equal significance and raise (P＜0.01), and it is the most serious to add surgical groups with blood stasis, and it is maximum that death condition also adds surgical groups with blood stasis; Explain and make the brain ischemia-reperfusion injury model and hemopoietic stasis of blood model all can make the mouse blood viscosity significantly increase, both act on stackable.Add the surgical groups ratio with blood stasis; Each administration group dead mouse situation all reduces; Large, medium and small dosage Flos Campsis extract group, HUATUO ZAIZAO WAN group and nimodipine group all can make blood stasis add the high, medium and low index of cutting of hemorheology that operation raises due to the modeling significantly to reduce (P＜0.01), act as excellent with heavy dose of Flos Campsis extract group.
Table 5 Flos Campsis extract merges the influence
of cerebral ischemia re-pouring model brain homogenate NO and NOS level to the mice blood stasis
▲ ▲Expression and blank group be than P＜0.01, * * represent with model group than P＜0.01
Can find out from last table; With blank group ratio; Not hemopoietic stasis of blood surgical groups, blood stasis not surgical groups and blood stasis add the equal significance of surgical groups brain homogenate NO, INOS and TNOS level and raise (P＜0.01), and it is the most serious to add surgical groups with blood stasis, and it is maximum that death condition also adds surgical groups with blood stasis; Explain and make the brain ischemia-reperfusion injury model and hemopoietic stasis of blood model all can make mouse brain homogenate NO, INOS and TNOS level significantly increase, both act on stackable.Add the surgical groups ratio with blood stasis; Each administration group dead mouse situation all reduces; Large, medium and small dosage Flos Campsis extract group, HUATUO ZAIZAO WAN group and nimodipine group all can make blood stasis add that rising brain homogenate NO, INOS and TNOS level significantly reduce (P＜0.01) due to the operation modeling, act as excellent with big or middle dosage Flos Campsis extract group.
Table 4 Flos Campsis extract merges the influence
of cerebral ischemic model brain homogenate ATP enzyme activity to the mice blood stasis
▲ ▲Expression and blank group be than P＜0.01, * * represent with model group than P＜0.01
Can find out that from last table with blank group ratio, not hemopoietic stasis of blood surgical groups, blood stasis not surgical groups and blood stasis add surgical groups blood flow brain homogenate Ca
++-ATP enzyme, Mg
++-ATP enzyme and Ca
++-Mg
++The equal significance of-ATP vigor reduces (P＜0.01); It is the most serious to add surgical groups with blood stasis; It is maximum that death condition also adds surgical groups with blood stasis, explains to make the brain ischemia-reperfusion injury model and hemopoietic stasis of blood model all can make the mouse blood viscosity significantly increase, and both act on stackable.Add the surgical groups ratio with blood stasis, each administration group dead mouse situation all reduces, and large, medium and small dosage Flos Campsis extract group, HUATUO ZAIZAO WAN group and nimodipine group all can make blood stasis add the brain homogenate Ca that operation reduces due to the modeling
++-ATP enzyme, Mg
++-ATP enzyme and Ca
++-Mg
++-ATP vigor significantly raises (P＜0.01), act as excellent with heavy dose of Flos Campsis extract group.
3, conclusion:
But Flos Campsis extract significant prolongation mice normal pressure hypoxia-bearing capability; Significantly improve the energy metabolism of mice ischemia-reperfusion injury model brain homogenate, significantly reduce the mice blood stasis and merge the cerebral ischemic model hemorheology and high, medium and lowly cut level, significantly reduce brain homogenate NO, INOS and TNOS level, significantly raise brain homogenate Ca++-ATP enzyme, Mg++-ATP enzyme and Ca++-Mg++-ATP vigor.Explaining that Flos Campsis extract has good anti-cerebral ischemia and cerebral ischemia reperfusion injury effect, effectively as the treatment anti-cerebral ischemia drugs, opened up the medical value of Flos Campsis, is the innovation on the treatment brain ischemia medicament.
Claims (1)
1. the Flos Campsis extract application in preparation treatment anti-cerebral ischemia drugs, described Flos Campsis extract be, with Flos Campsis add 10 times of weight, mass concentration is 60%～90% ethanol, soaks 30 minutes, extracts 1 hour, mistake filters medicinal liquid; Medicinal residues add again 8 times of weight of Flos Campsis, mass concentration is 60%～90% ethanol, extracts 1 hour, cross and filter medicinal liquid; Merge medicinal liquid twice, water-bath concentrates, vacuum drying, gets Flos Campsis extract.
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|---|---|---|---|
| CN2010105764916A CN102008521B (en) | 2010-12-07 | 2010-12-07 | Application of Chinese trumpet creeper extract to preparation of medicament for treating and resisting cerebral ischemia |
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|---|---|---|---|
| CN2010105764916A CN102008521B (en) | 2010-12-07 | 2010-12-07 | Application of Chinese trumpet creeper extract to preparation of medicament for treating and resisting cerebral ischemia |
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| KR102221353B1 (en) * | 2019-11-18 | 2021-03-02 | 대구한의대학교산학협력단 | Composition comprising the extract of Campsis grandiflora and Hydrangea macrophylla for preventing or treating neuro degenerative disease |
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