CN1202832C - High-activity ginkgo leaf extract preparation and application - Google Patents
High-activity ginkgo leaf extract preparation and application Download PDFInfo
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Abstract
The present invention discloses a freeze-dried powder preparation of ginkgo leaf extract, which is prepared from ginkgo leaf extract with high activity. The present invention simultaneously discloses a method for preparing ginkgo leaf extract with high activity, a formulation for the freeze-dried powder preparation and a method for preparing the freeze-dried powder preparation. In addition, the present invention also discloses an application of the freeze-dried powder preparation in the prevention and the treatment of cardiovascular and cerebrovascular diseases.
Description
Technical field
The present invention relates to a kind of extract formulation and purposes of Folium Ginkgo, relate to a kind of Folium Ginkgo extract powder injection formulation and purposes or rather.
Background technology
In the prior art, Folium Ginkgo extract sees the TEBONIN of German Schwabe drugmaker the earliest as the medicine of cardiovascular and cerebrovascular disease, developed Ginaton later on again, it is a kind of liquid drugs injection powder, its leading indicator is to contain total flavonoid glycoside more than 24% in the Folium Ginkgo extract, total lactone is more than 6%, and ginkgoic acid is less than 5ppm.But effective ingredient is on the low side in said preparation, and bilobalide is almost lost, and bibliographical information is arranged, and Folium Ginkgo extract is used for the treatment of its total flavones of cardiovascular and cerebrovascular disease and total lactone plays synergism, and damage has protective effect to bilobalide to ischemic tissue of brain; The film that can suppress hypoxia inducible in the brain decomposes; Improve cyanide and handle the neuronic viability of back Embryo Gallus domesticus; induce and improve CA1 cone innervation; suppress MPN and reduce butyrine level and glutamate decarboxylase activity in Hippocampus and the cerebral cortex, have the protective effect of nervus centralis and remove effect such as free radical.In order to obtain ginkgo leaf extract preparation more efficiently, just must obtain a kind of high Folium Ginkgo extract that contains total flavonoid glycoside and total lactone, simultaneously as injection, also should make the impurity of extract few as much as possible.This is that people are seeking always, but the target that does not reach as yet so far.
We find by in Folium Ginkgo primary extract subtractive process now, with flavone component and lactone component separating, refining respectively, obtain two kinds of Folium Ginkgo extract, their high respectively total flavonoid glycoside and total lactones of containing, and, obtained the highly active Folium Ginkgo extract that a kind of total flavonoid glycoside and total lactone have certain proportioning, and prepare highly active preparation with it by pharmacological research.
Purpose of the present invention just provides a kind of highly active ginkgo leaf extract preparation and is used for the treatment of cardiovascular and cerebrovascular disease.
Summary of the invention
High-activity ginkgo leaf extract preparation of the present invention is a kind of lyophilized injectable powder, it is characterized in that this lyophilized injectable powder contains high-activity ginkgo leaf extract, this high-activity ginkgo leaf extract is to be mixed by Folium Ginkgo total flavones glucoside extract (abbreviating extract A as) and ginkgo biloba leaf total terpene lactone extract (abbreviating extract B as), it is more than 65% that extract A contains total flavonoid glycoside, extract B contains total lactone more than 75%, wherein the content of bilobalide in total lactone is more than 40 percent, ratio is (4.20-5.0): (1.05-1.80), contain total flavonoid glycoside and total lactone more than 65% through mixed extract, ginkgoic acid content is less than 5ppm;
High-activity ginkgo leaf extract lyophilized injectable powder of the present invention has following feature:
(1) to contain high total flavonoid glycoside and total lactone and content of bilobalide be more than 40 percent to the extract in the highly active ginkgo leaf extract freeze dried powder injection, and available following method is measured.
The total flavonoid glycoside assay is measured according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2000 D).
Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Methanol-0.4% phosphoric acid solution (50: 50) is a mobile phase; The detection wavelength is 360nm.Number of theoretical plate calculates by the Quercetin peak should be not less than 3000.
The preparation of reference substance solution is accurate respectively to take by weighing through dry Quercetin, kaempferide, the isorhamnetin reference substance of crossing of phosphorus pentoxide, adds a little dissolve with methanol, adds mobile phase and makes the solution that every 1ml contains 0.036mg, 0.020mg, 0.006mg respectively.
The lyophilized injectable powder content is got in the preparation of need testing solution, and porphyrize is mixed, and gets 0.2g, and accurate the title decides, add 7.5% hydrochloric acid 20ml, add 70% ethanol 20ml, 95 ℃ were refluxed 2 hours, cooling, be transferred in the 50ml measuring bottle, and add dehydrated alcohol, shake up, promptly to scale.
Accurate respectively reference substance solution and each the 20 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, and calculate the content of three kinds of flavone aglycone respectively, are converted into the content of total flavonoid glycoside with following formula.
Total flavonoid glycoside content=(quercetin content+kaempferide content+different Fructus rhamni (Rhamnus davurica Pall.) content) * 2.51
Lactone content is measured according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2000 D) and is measured.
Chromatographic condition and system suitability test are filler with octadecylsilane chemically bonded silica; Water-methanol-oxolane (75: 20: 10) is a mobile phase, flow velocity 1.0ml/min; Alltech 500 ELSD are detector, and the detector drift tube temperature is 115 ℃, and nitrogen flow rate is 2.74SLPM, and number of theoretical plate calculates by the bilobalide peak should be not less than 9000.
The preparation of reference substance solution is learnt from else's experience and is equipped with the phosphorus pentoxide desiccant, the reference substance of 60 ℃ of vacuum drying 6h is an amount of, add water-methanol-oxolane (44: 50: 6) solvent and make every 1ml bilobalide-containing J:0.135mg, ginkalide C: 0.355mg, bilobalide 0.660mg, ginkalide A: 0.400mg, ginkalide B: 0.102mg, be reference substance solution I; Get reference substance solution I 3ml and 2ml, put respectively in the 5ml measuring bottle, add above-mentioned solvent dilution, be reference substance solution II, III to scale.
The lyophilized injectable powder content is got in the preparation of need testing solution, the porphyrize mixing is got 0.52g, and accurate the title decides, put in the conical flask, add the 20ml aqueous solution, regulate pH to 3-4, be added on polyamide column (the internal diameter 1.5cm that has handled well with 2% hydrochloric acid, the polyamide of long 5cm, spend the night with 70% soak with ethanol in advance, ethanol is abandoned in filter, the dress post, wash with water to there not being the alcohol flavor, standby) on, add water 40ml eluting, collect eluent, eluent adds sodium chloride 1.8g, with ethyl acetate extraction 3 times, be respectively 40ml, 35ml, 30ml, merge ethyl acetate liquid; Evaporated under reduced pressure, residue add water-methanol-oxolane (44: 50: 6) solvent makes dissolving, quantitatively changes and holds to the 5ml measuring bottle, shakes up, and crosses the filter membrane of 0.45 μ m, gets subsequent filtrate, promptly.
Accurate respectively each the 20 μ l of reference substance solution I, II, III and need testing solution that draw of algoscopy inject high performance liquid chromatograph, measure.Calculate with the external standard line-of-sight course after concentration and peak area all taken from right logarithm, obtain the natural logrithm of test sample peak area, and convert the content of test sample to, promptly.
The content of bilobalide in Ginkgo total lactones is by calculating.
(2) total flavonoid glycoside and total lactone also have following finger printing feature in the highly active ginkgo leaf extract freeze dried powder injection:
Description of drawings
A) the finger printing feature of flavones ingredient (seeing accompanying drawing 1);
The peak number (relative retention time) at total peak: 1 (0.424), 2 (0.544), 3 (0.651), 4 (1.000), 5 (1.151), 6 (1.571), 7 (1.845).
The ownership peak 4 at total peak is a Quercetin; Peak 6 is a nimbecetin; Peak 7 is an isorhamnetin; Peak 5 is for having the flavones ingredient of the same parent nucleus of Quercetin.
Peak area ratio 4 (S): 6: 7=(1.000): (0.561～0.841): (0.136～0.252)
Above-mentioned technical characterictic is measured by following method:
According to " the high-performance liquid chromatography method of an appendix VI of Chinese pharmacopoeia version in 2000 D.
Instrument: the HP1100 Series of Hewlett-Packard.
Chromatographic condition and system suitability test: (Alltech) Platinum EPS C
18100A 3 μ 150mm * 4.6mm; Methanol-0.4% phosphate aqueous solution (50: 50) is a mobile phase, flow velocity 0.7ml/min; Detect wavelength side 370nm.Slope Senesitivity：1、peakwidth：0.5、Area Reject：0.5、heightReject 0.5、Shoulders off，0.000 Integration OFF，3.464 Integration ON、4.890Baseline Now ON。Number of theoretical plate is pressed Quercetin and is calculated, and should be not less than 3200.
The preparation of need testing solution: get lyophilized injectable powder content 0.2g of the present invention, the accurate title, decide, and adds 7.5% hydrochloric acid 20ml, dissolving, add 70% ethanol 20ml, 95 ℃ were refluxed 2 hours, cooling, be transferred in the 50ml measuring bottle, and add dehydrated alcohol to scale, and shake up, cross the filter membrane of 0.45 μ m, get subsequent filtrate, promptly.
The object of reference formulations prepared from solutions: it is an amount of to get the Quercetin reference substance, and accurate the title decides, and adds a little methanol supersound process and makes dissolving, adds mobile phase and makes the solution that every 1ml contains 0.026mg, in contrast product solution.
Assay method: accurate respectively reference substance solution and each 20 μ l of need testing solution of drawing, inject chromatograph of liquid, write down 45 minutes chromatograms, promptly.With the retention time of the chromatographic peak (S peak) of Quercetin and peak area is 1 to calculate relative retention time and peak area ratio.
B) lactone ingredients fingerprint feature (seeing accompanying drawing 2):
The peak number (relative retention time) at total peak: 1 (0.713), 2 (0.884), 3 (1.000), 4 (1.296), 5 (1.701)
Ownership peak 1, total peak is a bilobalide J; Peak 2 is a ginkalide C; Peak 3 is a bilobalide; Peak 4 is a ginkalide A; Peak 5 is a ginkalide B.
2: 3 (S): 4=(0.166～0.276) of peak area ratio: (1.000): (0.362～0.544)
Above-mentioned technical characterictic is measured by following method:
According to " the high-performance liquid chromatography method of an appendix VI of Chinese pharmacopoeia version in 2000 D.
Instrument; Tianjin, island LC-10ATvp of company, monitor C-R6A; Detector: Alltech 500 ELSD of U.S. Kodak technology company.
Chromatographic condition and system suitability test: Phenomenex
Hypersil 5 μ C
18(ODS), 250mm * 4.6mm; Water-methanol-oxolane (75: 20: 10) is a mobile phase, flow velocity: 1.0ml/min; The detector drift tube temperature: 115 ℃, nitrogen flow rate: 2.74SLPM; ATTEN:6, SLOPE:500, MIN.AREA:5000, chart speed 5mm/min.Number of theoretical plate calculates by the bilobalide peak should be not less than 3500.
Lyophilized injectable powder 0.52g of the present invention is got in the preparation of need testing solution, and accurate the title decides, and puts in the conical flask, add the 20ml water dissolution, regulate pH to 3～4, be added on polyamide column (the internal diameter 1.5cm that has handled well with 2% hydrochloric acid, the polyamide of long 5cm, spend the night with 70% soak with ethanol in advance, ethanol is abandoned in filter, the dress post, wash with water to there not being the alcohol flavor, standby) on, add water 40ml eluting, collect eluent, eluent adds sodium acetate 1.8g, with ethyl acetate extraction 3 times, be respectively 30ml, 20ml, 20ml merges ethyl acetate extraction liquid, wash extracted twice liquid with water, each 15ml, evaporated under reduced pressure, residue adds water-methanol-oxolane (44: 50: 6) solvent makes dissolving, quantitatively change molten to the 5ml measuring bottle, cross the filter membrane of 0.45 μ m, get subsequent filtrate, promptly.
It is an amount of that the bilobalide reference substance is got in the preparation of reference substance solution, and accurate the title decides, and adds water-methanol-oxolane (44: 50: 6) solvent and makes the solution that every 1ml contains 0.40mg, promptly.
Assay method: accurate respectively reference substance solution and each 20 μ l of testing liquid of drawing, inject chromatograph of liquid, write down 30 minutes chromatograms, and be 1 to calculate relative retention time and peak area ratio with the retention time of the chromatographic peak 3 (S peak) of bilobalide and peak area.
(3) the high-activity ginkgo leaf extract lyophilized injectable powder has following outstanding pharmacodynamic profiles
A) high-activity ginkgo leaf extract can dwindle MCAO rat cerebral infarction area, reduces the nervous symptoms scoring, reduces brain water content and cerebral index, reduces the rat capillary permeability, compares the significant meaning of its performance difference tool with reference substance.
B) high-activity ginkgo leaf extract can increase the anesthetized cat cerebral blood flow, reduces vascular resistance, compares its performance difference tool significance meaning with reference substance.
C) high-activity ginkgo leaf extract can reduce the rat platelet aggregation rate, the prolong rats carotid artery thrombosis time, improve the brinase lytic activity, and reduce MDA content, compare its performance difference tool significance meaning with matched group.
D) high-activity ginkgo leaf extract can prolong clotting time of mice, and platelet count is not had obvious influence.
E) high bilobalide (BB) content, damage has protective effect to ischemic tissue of brain; The film that can suppress hypoxia inducible in the brain decomposes; Improve cyanide and handle the neuronic viability of back Embryo Gallus domesticus; induce and improve CA1 cone innervation; suppress MPN and reduce butyrine level and glutamate decarboxylase activity in Hippocampus and the cerebral cortex, have the protective effect of nervus centralis and remove effect such as free radical.
Above-mentioned character can confirm with following experiment:
Experiment material and instrument
1, animal: the SD rat, the animal rank: cleaning level, west, Shanghai City pul---must provide by triumphant laboratory animal company limited the quality certification number: the moving word of doctor 02-49-2 number.Kunming mice, Jiangxi Medical College's animal center provides, the animal quality certification number: the moving word 021-9609 of doctor.
2, medicine:
A kind of test medicine that injection high-activity ginkgo leaf extract lyophilized injectable powder (be called for short injection Folium Ginkgo (lyophilizing)) prepares by preparation process of the present invention, lot number 980827, it is standby to be mixed with respective concentration with distilled water during experiment;
Triphenyltetrazolium chloride Shanghai chemical reagent head factory;
Azovan blue Shanghai chemistry degree agent purchasing and supply station, lot number 820805;
Bio-engineering research institute is built up in malonaldehyde testing cassete Nanjing.
3, instrument: 307-6 type table electric dental engine, Shanghai Dental Medical Apparatus and Instrument Factory; 1422 type high frequency electro surgical units, Shanghai Medical Electric Instrument Factory; The GMIAS image analyzer, BJ University of Aeronautics ﹠ Astronautics; The MFA-1200 electromagnetic flowmeter, Japanese Nihokohden company; TYXN-91A intelligence blood agglutometer, Shanghai General Machinery ﹠ Electric technology Inst.'s system; 721 type spectrophotometers, Shanghai the 3rd instrument plant; TG328A type analysis balance, balance equipment factory in Shanghai produces; WMT-01 type digital thermometer, Shanghai Medical Instrument and Meter Factory; The experimental thrombus in vivo of BT87-3 type forms instrument, and supervise hospital attached to a medical college cardiovascular research chamber, packet header.
Test the influence of the focal type cerebral ischemia of a pair of rat infarct size
Get 70 of SD rats, male and female half and half, body weight 280 ± 30g, be divided into sham operated rats at random by the male and female body weight, the cerebral ischemic model group, dosage group (extract 10mg/kg) in injection Folium Ginkgo (lyophilizing) high dose group (extract 15mg/kg), injection Folium Ginkgo (lyophilizing), injection silver leaf (lyophilizing) low dose group (extract 5mg/kg).With chloral hydrate 350mg/kg; Ip anesthesia, except that sham operated rats is not blocked the middle cerebral artery, all the other each groups are pressed the Tamara method
[1]Under aseptic condition, pyrrole and right ear canal line mid point are made the stringer otch outside right eye, separate and cut off masseter and temporalis, remove zygomatic arch, expose squamosal bone, open cranium at roots of zygoma and squamosal bone the place ahead with dental drill boring, expose middle cerebral artery, electricity consumption is closed with fixed attention with fixed attention and is positioned at tractus olfactorius place tremulous pulse sections, wound surface is placed gelfoam, layer-by-layer suture muscle skin.Each administration group of postoperative is all by above-mentioned dosage intravenous injection, and sham operated rats, cerebral ischemic model group give the isometric(al) normal saline quiet notes, once a day, and continuous 2 days.After 48 hours, use the etherization rat, broken end is got brain, parallelly in the about backward 2mm in optic chiasma place place do crown section (thickness 5mm), place 2%TTC liquid [37 ℃] lucifuge to hatch 30 minutes, normal structure is dyed redness, blocking tissue is a white, with 10% formalin fixed, on the CMIAS image analyzer, measure cerebral infarct size, calculate right side cerebrum block area percentage [right side cerebral infarct size/right side brain area * 100%].The results are shown in Table 1.
Test the influence of two pairs of focal cerebral infarction rat nervous symptoms
Get 70 of SD rats, male and female half and half, body weight 260 ± 40g, be divided into sham operated rats at random by the sex body weight, cerebral ischemic model group, injection Folium Ginkgo (lyophilizing) high dose group, dosage group in the injection Folium Ginkgo (lyophilizing), injection Folium Ginkgo (lyophilizing) low dose group.Except that sham operated rats, all the other all capable right side of each group middle cerebral artery Interruptions.Art each group the previous day is by corresponding dose of iv.qd.Each group of postoperative 2h is pressed same dosage iv.qd,
Table 1 injection Folium Ginkgo (lyophilizing) is to the influence (X ± SD of intraluminal middle cerebral artery occlusion in rats barrier cerebral infarct size
n＝10)
| Group | Right side brain average area (mm 2) | Average infarct size (the mm of right side brain 2) | Right side cerebral infarct size percentage ratio (%) | Right side infarct size slip (%) |
| Sham operated rats | 118.82±9.61 | |||
| The cerebral ischemic model group | 121.20±12.26 | 29.20±4.63 | 23.85±4.53 | |
| Injection Folium Ginkgo (lyophilizing) high dose group | 109.74±9.49 | 14.08± 5.32 *** | 12.96± 4.56 *** | 51.78 |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | 122.59±11.95 | 16.69± 6.23 *** | 13.60± 4.87 *** | 42.84 |
| Injection Folium Ginkgo (lyophilizing) low dose group | 114.07±8.75 | 19.80± 6.15 ** | 16.27± 4.24 ** | 31.88 |
Annotate: compare * * P＜0.01 * * * P＜0.001 with the cerebral ischemic model group
Table 2 injection Folium Ginkgo (lyophilizing) is to the influence of focal cerebral infarction rat nervous symptoms scoring
( X±SD n＝10)
| Grouping | Dosage | 24h | 48h | 72h |
| Sham operated rats | N.S 4ml/kg | 0 | 0 | 0 |
| The cerebral ischemic model group | N.S 4ml/kg | 6.20±1.64 | 6.05±1.20 | 5.80±1.75 |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 15mg/kg | 4.25± 1.48 * | 3.25±1.47 ** | 2.25±2.01 ** |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 10mg/kg | 4.15± 1.73 * | 3.30±1.66 ** | 2.60±2.15 ** |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 5mg/kg | 4.95±1.67 | 4.05±1.94 * | 3.55±2.09 * |
Annotate: with cerebral ischemic model group * P＜0.05 * * P＜0.01 clinical symptoms of observing 24h, 48h after the last administration, 72h relatively, determine the nervous symptoms standards of grading by document: (1) is mentioned the Mus tail and is observed in the left fore and receive, and left side shoulder inward turning is chosen as the 1-4 branch; (2) rat is placed on the flat board, push away right shoulder and be moved to the left,, be chosen as the 1-3 branch with left side resistance drop low degree; (3) look left fore muscular strength decline degree and be chosen as the 1-3 branch.Total points is 10 minutes, adopts above-mentioned standards of grading, carries out statistical procedures, the results are shown in Table 2.
Test the influence of three pairs of focal cerebral ischemia in rats brain water contents, cerebral index
Grouping of SD rat and medication are the same, after observing nervous symptoms 72h after the last administration, use etherization, open cranium and get brain, take by weighing full cutaneous horn with precision balance and weigh,, take by weighing dry weight in 100 ℃ of oven for baking 24h, by formula calculate brain water content (weight in wet base-dry weight/weight in wet base * 100%), and by formula calculate cerebral index (heavy * 100/ body weight of brain).The results are shown in Table 3.
The influence of four pairs of rat brain capillary permeabilities of experiment
Get 60 of male SD rats, body weight 220～255g, be divided into sham operated rats at random, the cerebral ischemic model group, dosage group in injection Folium Ginkgo (lyophilizing) high dose group, injection Folium Ginkgo (lyophilizing), injection Folium Ginkgo (lyophilizing) low dose group, each group is by its dosage penile vein drug administration by injection, qd * 2d, last administration 30min is by intravenous injection 20% azovan blue 0.3ml/100g, behind the 5min, except that sham operated rats, each organizes the ligation bilateral common carotid arteries, and broken end is got brain behind the 4h, claim full cutaneous horn heavy, use 0.5%Na
2SO
43ml and acetone 7ml, make 10% brain homogenate, be statically placed in 12h in the refrigerator, centrifugal 3000r/min 20min inserts 12h in the refrigerator again, centrifugal 3000r/min 20min, get supernatant, with 721 spectrophotometers (wavelength 620nm) colorimetric,, the results are shown in Table 4 according to OD value and standard curve computation organization azovan blue content.
Test the influence of five pairs of anesthetized cat cerebral blood flows and cerebral vascular resistance
36 of healthy cats, the male and female dual-purpose, body weight 2.4～3.8kg, be divided into the normal saline matched group at random, injection Folium Ginkgo (lyophilizing) high dose group (extract 7.5mg/kg), group (extract 5mg/kg) in the dosage in the injection Folium Ginkgo (lyophilizing), injection Folium Ginkgo (lyophilizing) low dose group (extract 2.5mg/kg).
Table 3 injection Folium Ginkgo (lyophilizing) is to the influence of focal cerebral ischemia in rats brain water content, cerebral index
( X±SD n＝10)
| Grouping | Dosage | Brain water content (%) | Cerebral index |
| Sham operated rats | N.S 4ml/kg | 76.40±1.26 △△△ | 0.60±0.04 △△ |
| The cerebral ischemic model group | N.S 4ml/kg | 78.96±1.07 | 0.69±0.06 |
| Injection Folium Ginkgo (lyophilizing) powder pin high dose group | Extract 15mg/kg | 77.04±1.01 ** | 0.62±0.05 * |
| Dosage group in injection Folium Ginkgo (lyophilizing) the powder pin | Extract 10mg/kg | 77.15±1.05 ** | 0.63±0.05 * |
| Injection Folium Ginkgo (lyophilizing) powder pin low dose group | Extract 5mg/kg | 77.72±1.18 * | 0.64±0.07 |
Annotate: administration group and model group be * P＜0.05 * * P＜0.01 relatively
The sham operated rats model group relatively
△ △P＜0.01
△ △ △P＜0.001
Table 4 injection Folium Ginkgo (lyophilizing) is to the influence of rat brain capillary permeability
( X±SD n＝10)
| Grouping | Dosage | Azovan blue content (ug/g) |
| Sham operated rats | N.S 4ml/kg | 14.23±2.95 *** |
| The cerebral ischemic model group | N.S 4ml/kg | 24.86±5.01 |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 15mg/kg | 17.56±3.63 ** |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 10mg/kg | 17.98±3.04 ** |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 5mg/kg | 19.39±4.34 * |
Annotate: compare with the cerebral ischemic model group
*P＜0.05
*P＜0.01 usefulness, 1.5% chloralose+25% urethane (2: 1) solution 3.5～5ml/kg ip anesthesia.Separate right carotid artery, upwards separate external carotid artery, internal carotid artery, separate downwards, separate vertebral artery downwards at the inside turning of external jugular vein along right carotid artery.Ligation right side neck place tremulous pulse and vertebral artery.Be fit to probe with electromagnetic flowmeter and measure right side internal carotid artery per minute mean blood flow.Carry out femoral arteriography in addition, directly detect instrumentation and decide blood pressure, carry out femoral venous catheter simultaneously, with the slow intravenous drip of normal saline with hydrargyrum, after treating blood pressure stabilization 20min, write down administration respectively before, ICAF amount, the blood pressure of 10min, 30min, 60min, 90min after the administration.Experiment is won full brain and is weighed, by formula after finishing
[3]Calculate cerebral blood flow and vascular resistance, the results are shown in Table 5-1,5-2.
The influence of six pairs of rat platelet aggregations of experiment
60 of SD male rats, body weight 240 ± 30g, be divided into the normal saline group at random, injection Folium Ginkgo (lyophilizing) high dose group, dosage group in the injection Folium Ginkgo (lyophilizing), injection Folium Ginkgo (lyophilizing) low dose group, each group is pressed its dosage by the penile vein drug administration by injection, once a day, for three days on end, behind the last administration 30min, with 30% sodium pentobarbital 30mg/kg ip anesthesia, through abdominal aortic blood, with 3.28% sodium citrate (anticoagulant is 1: 9 with the ratio of whole blood), with 500rpm * 3min, preparation platelet rich plasma (PRP), and the preparation platelet poor plasma (PPP, 3000rpm * 10min), adopt PPP test cup to regulate zero point, with ADP is derivant (concentration 4 μ m, consumption 11ul), calculates platelet aggregation rate with TYXN-91A type intelligence blood agglutometer by turbidimetry, and calculate platelet aggregation inhibition rate=matched group aggregation rate-medicine group aggregation rate/matched group aggregation rate * 100%, the results are shown in Table 6.
The influence of seven pairs of rat carotid artery thrombus formation time of experiment
Get 50 of SD rats, male and female half and half, body weight 260～330g is divided into dosage group, injection Folium Ginkgo (lyophilizing) low dose group in normal saline group, injection Folium Ginkgo (lyophilizing) high dose group, the injection Folium Ginkgo (lyophilizing) at random.Each group is by its dosage intravenous injection qd * 2d, and the last administration is after 15 minutes, with penta crust ratio
Table 5-1 injection Folium Ginkgo (lyophilizing) is to the influence (X ± SD n=6) of anesthetized cat cerebral blood flow
Cerebral blood flow (ml/100g/min)
The average blood of (min) 30min cerebral blood flow 30min after the administration before the group administration
10 ' 30 ' 60 ' 90 ' increment rate (%) Kpa
Normal saline group 64.38 ± 9.71 69.75 ± 10.93 65.81 ± 9.66 63.53 ± 8.45 63.92 ± 11.29 2.22 17.18 ± 2.01
Injection Folium Ginkgo (lyophilizing) high dose group 66.17 ± 9.34 94.57 ± 16.03
*109.15 ± 21.51
* *101.71 ± 16.01
*88.89 ± 16.81
*64.95 18.53 ± 1.57
Dosage group 60.59 in the injection Folium Ginkgo (lyophilizing) ± 10.04 83.80 ± 15.79
*92.35 ± 21.60
*88.56 ± 24.45
*78.23 ± 12.90
*52.42 17.30 ± 1.54
Injection Folium Ginkgo (lyophilizing) low dose group 64.12 ± 5.76 84.90 ± 19.67
*92.16 ± 25.69
*84.79 ± 19.50
*79.18 ± 17.19
*43.73 17.22 ± 0.84
Ginaton group 57.20 ± 9.19 73.38 ± 12.74
*85.68 ± 13.49
*76.95 ± 16.28
*69.93 ± 15.19
*49.53 18.63 ± 2.17
Puerarin group 70.35 ± 9.45 91.04 ± 23.87
*97.51 ± 19.30
*91.18 ± 20.40
*84.30 ± 17.42 38.60 17.28 ± 2.14
Annotate: each organizes cerebral blood flow comparison before the day part cerebral blood flow and administration after the administration
*P＜0.05
*P＜0.01
* *P＜0.001; Day part cerebral blood flow and 30min mean blood pressure injection Folium Ginkgo (lyophilizing) are low
Dosage group and Ginaton be P＞0.05 relatively
Table 5-2 ginkgo leaf powder is at the influence (X ± SD n=6) of anesthetized cat brain blood resistance
Cerebral blood flow (ml/100g/min)
(min) 30min cerebral vascular resistance after the administration before the group administration
10 ' 30 ' 60 ' 90 ' rate of descent (%)
Normal saline group 0.27 ± 0.02 0.26 ± 0.03 0.27 ± 0.03 0.28 ± 0.03 0.26 ± 0.03
Injection Folium Ginkgo (lyophilizing) high dose group 0.29 ± 0.04 0.21 ± 0.03
*0.17 ± 0.02
* *0.19 ± 0.02
*0.22 ± 0.02
*41.38
Dosage group 0.29 in the injection Folium Ginkgo (lyophilizing) ± 0.04 0.22 ± 0.03
*0.19 ± 0.03
*0.21 ± 0.05
*0.23 ± 0.04
*34.48
Injection Folium Ginkgo (lyophilizing) low dose group 0.27 ± 0.03 0.21 ± 0.04
*0.20 ± 0.05
*0.21 ± 0.05
*0.23 last 0.04
*25.92
Ginaton group 0.31 ± 0.03 0.25 ± 0.04
*0.23 ± 0.05
*0.25 ± 0.04
*0.25 ± 0.04
*25.81
Puerarin group 0.25 ± 0.03 0.19 ± 0.05
*0.19 ± 0.04
*0.21 ± 0.05
*0.22 ± 0.04 24.00
Annotate: each organizes cerebral vascular resistance comparison before the day part cerebral vascular resistance and administration after the administration
*P＜0.05
* *P＜0.01
Table 6 injection Folium Ginkgo (lyophilizing) is induced the influence of rat platelet aggregation rate to ADP
( X±SD n＝10)
| Grouping | Dosage | Aggregation rate (%) | Assemble suppression ratio (%) |
| The normal saline group | 4ml/kg | 47.54±6.00 | |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 15mg/kg | 30.38± 11.74 ** | 36.10 |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 10mg/kg | 32.09± 12.78 ** | 32.50 |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 5mg/kg | 36.80± 10.64 * | 22.59 |
Annotate: compare with the normal saline group
*P＜0.05
*P＜0.01
Table 7 injection Folium Ginkgo (lyophilizing) is to the influence of rat carotid artery thrombus formation time
( X±SD n＝10)
| Grouping | Dosage | Thrombus formation time (min) | Rate elongation (%) |
| The normal saline group | 4ml/kg | 17.78±4.28 | |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 15mg/kg | 29.29±7.69 ** | 64.73 |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 10mg/kg | 27.32±8.02 ** | 53.66 |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 5mg/kg | 23.47±6.06 * | 32.00 |
Annotate: write physiology saline group relatively
*P＜0.05
*P＜0.01 appropriate sodium 30ml/kg ip anesthesia.Separate right carotid artery, after WMY-01 type digital thermometer measurement carotid artery vascular surface temperature, reuse BT87-3 type thrombus in vivo forms instrument with 1.5mv galvanic stimulation 7min, survey a carotid artery surface temperature every 2min, observation from electricity irritation begin to the time that temperature descends suddenly be thrombus formation time in the rat carotid artery, the results are shown in Table 7.
Test eight pairs of active influences of rat fibrinoclase
Get 50 of male SD rats, body weight 220 ± 20g is divided into 4 groups at random.Each group is anaesthetized through pentobarbital sodium 30mg/kg ip by corresponding dosage intravenous injection qd * 2d before the experiment.By abdominal aortic blood 2ml, add 3.8% sodium citrate 0.2ml, with 2500r/min centrifugal 10 minutes, make platelet poor plasma, get blood plasma 0.5ml, add acidifying water 9.5ml (1% glacial acetic acid 11ml is diluted to 1000ml).Inserted refrigerator 30 minutes, and centrifugal 15 minutes again, removed supernatant with 2500r/min, boric acid solution with PH9.0 makes resolution of precipitate, adds the 0.5ml/40M calcium chloride solution, and liquid in pipe is solidified, the record EGCT by formula calculates the fibrinoclase activity
[6], the results are shown in Table 8.
Test the influence of nine pairs of clotting time of mice and platelet count
Get 75 of healthy kunming mices, body weight 21 ± 3g, male and female half and half are divided into 4 groups at random.Each group is carried out tail vein injection by dosage in the table, qd * 2d, behind the last administration 15min, extract eyeball of mouse rapidly, drip one and bleed in cleaning slide one end, provoke from the edge of bleeding with syringe needle every 30s, record blood coagulation time of occurrence is drawn 10ul blood with suction pipe simultaneously, places in the platelet diluent and leaves standstill 15min, with high power lens meter platelet count, the results are shown in Table 9.
Test the influence that MDA measures in ten pairs of rat cerebral ischemia tissues
Get 50 of SD rats, body weight 238 ± 17.5g, the male and female dual-purpose is divided into 4 groups at random.Respectively group is pressed dosage intravenous injection in the table, qd * 2d, and behind the last administration 15min, except that sham operated rats, all capable bilateral carotid arteries ligation (not ligation vagus nerve) of all the other groups.Each is organized after 3 hours broken end and gets brain and weigh, and makes 10% brain homogenate with normal saline, and 3000r/min 10min abandons supernatant and stays the precipitation tissue, adds distilled water, mixes at vortex
Table 8 injection Folium Ginkgo (lyophilizing) is to the active influence of rat fibrinoclase
( X±SD n＝10)
| Grouping | Dosage | Plasma clot dissolution time (min) | Fibrinoclase activity (μ) |
| The normal saline group | 4ml/kg | 338.2±51.03 | 30.12±4.40 |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 15mg/kg | 253.9±47.03 ** | 40.63±7.55 ** |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 10mg/kg | 264.9±48.57 ** | 38.79±6.80 ** |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 5mg/kg | 287.8±39.07 * | 35.31±4.65 * |
Annotate: compare with the normal saline group
*P＜0.05
*P＜0.01
Table 9 injection Folium Ginkgo (lyophilizing) is to the influence of clotting time of mice and platelet count
( X±SD n＝10)
| Grouping | Dosage | Clotting time (s) | Platelet count (10 9/L) |
| The normal saline group | 20ml/kg | 73.66±28.52 | 201.04±30.35 |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 30mg/kg | 123.80± 35.69 ** | 210.09±28.21 |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 20mg/kg | 117.83± 29.25 ** | 193.37±30.30 |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 10mg/kg | 109.06± 28.96 * | 211.10±36.69 |
Annotate: compare with the normal saline group
*P＜0.05
*P＜0.01
Table 10 injection Folium Ginkgo (lyophilizing) is to the influence of rats with cerebral ischemia cerebral tissue MDA content
( X±SD n＝10)
| Grouping | Dosage | MDA(nmol/mg -1) |
| Sham operated rats | N.S 4ml/kg | 0.97±0.24 *** |
| The cerebral ischemic model group | N.S 4ml/kg | 2.15±0.57 |
| Injection Folium Ginkgo (lyophilizing) high dose group | Extract 15mg/kg | 1.39±0.37 *** |
| Dosage group in the injection Folium Ginkgo (lyophilizing) | Extract 10mg/kg | 1.45±0.31 ** |
| Injection Folium Ginkgo (lyophilizing) low dose group | Extract 5mg/kg | 1.62±0.39 * |
Annotate: compare with the cerebral ischemic model group
*P＜0.05
*P＜0.01
* *P＜0.001
The comparison of (%) of total lactone of table ten gingko leaf preparation and the total lactone of BB/
Mixing on the even device makes the brain cell dissolving.With 95% ethanol extracting MDA, with the chloroform protein precipitation.Press the malonaldehyde kit test method, respectively the production standard pipe, measure pipe, blank pipe, form the principle of red product by MDA and thiobarbituricacid complexation, adopt 721 spectrophotometer 532nm places to carry out colorimetric, calculate MDA content by above-mentioned tee pipe absorbance, the results are shown in Table 10.
| The name of an article | Manufacturer | Total lactone (%) | The total lactone of BB/ (%) |
| Ginaton injection | The Germany big pharmaceutical factory of Weil-McLain Shu Pei | 4.27 | 5.35 |
| Injection Folium Ginkgo (lyophilizing) | Jiangxi Province's drug research and Haizheng Medicine Stock Co., Ltd., Zhejiang Prov's development | 14.35 | 40.35 |
Table ten one be with Ginaton injection in bilobalide (BB) content relatively, kept higher content in the injection of high activity extract preparation of the present invention.
Preparation of the present invention is the same disease that is used to prevent and treat cardiovascular and cerebrovascular vessel with ordinary preparation on using.
The invention provides the preparation method of high-activity ginkgo leaf extract.
The high-activity ginkgo leaf extract preparation method adopt flavone compound and terpene lactones chemical compound are extracted respectively refining, blended by a certain percentage again method.Prove from prior art, flavone compound has different character chemical compounds with terpene lactones chemical compound two classes, two class materials extract simultaneously and are difficult to obtain simultaneously the extract that height contains this two classes material, prior art also proves, extract refining two kinds of extracts that can obtain high level respectively, but they all are different from purpose of the present invention, and the content of total flavonoid glycoside is the highest can only to reach 48%, the present invention further improves prior art, make that the content of total flavonoid glycoside reaches more than 65% in extract A, total lactone content reaches more than 75% in extract B, then with extract A and extract B conventional method, mix obtaining highly active Folium Ginkgo extract by a certain percentage, the content of mixed total flavonoid glycoside and total lactone is reached more than 65%.
We adopt following technical scheme to realize above-mentioned target:
The preparation of extract A: the first extract of Folium Ginkgo (being called for short EGb) is added the water supersound extraction, get supernatant, last polyamide column, wash eluting with water, discard the water of retention volume, collect all lactone stream parts, for extracting Folium Ginkgo total lactones usefulness, reuse 60% ethanol elution, with the reaction of HCl-Mg powder is index, collect eluent and at concentrating under reduced pressure below 60 ℃, spray drying, the yellowish-brown powder, this powder adds the acetone supersound extraction 2 times, extracting liquid filtering is concentrated into driedly, and dry substance adds the ethyl acetate stir process, filter, get the filtering residue vacuum drying, porphyrize gets pale yellow powder and is goal object.In the present invention, adopt concentrating under reduced pressure, spray drying, avoid the active component pyrolytic, the yellowish-brown powder that obtains behind polyamide column acetone supersound extraction 2 times, extracting liquid filtering is removed insoluble matter, and filtrate is concentrated into dried, dry substance adds the ethyl acetate stir process, is the key that improves the total flavonoid glycoside content in the extract A.
The preparation of extract B: get the polyamide washing stream part in the extractive of general flavone preparation process, concentrating under reduced pressure below has two kinds of method: a. by ethyl acetate extraction, the extracting solution vacuum concentration, and drying, porphyrize is goal object; B. concentrated solution washing, cooling, crystallization, filter, get white lactone crystallization thing, the lactone crystallization thing is through vacuum drying, porphyrize is standby, mother solution reuse ethyl acetate extraction is got ethyl acetate extraction liquid, concentrates, vacuum drying, porphyrize gets the powder of ivory buff, adds above-mentioned lactone crystallization thing, and mix homogeneously is goal object.
Highly active Folium Ginkgo extract preparation: extract A and extract B are pressed (4.20-5.0): (1.05-1.80) mix homogeneously, obtain highly active Folium Ginkgo extract, total flavonoid glycoside and total lactone content are greater than 65%, and ginkgoic acid content is below the 5ppm.
The present invention also provides a kind of freeze-dried powder preparation method: get high-activity ginkgo leaf extract and lactose, add proper amount of water for injection again, make it dissolving with ultrasonic method, drip the sodium hydroxide solution of 1mol/L, the limit edged fully stirs, and is adjusted to pH6.5-7.1, adds the injection water to capacity, fully mix, aseptic filtration, packing is put in the freeze dryer, be lyophilized into loose block, and vacuum state lower cover sealing in freeze dryer.The most important thing is in the above technology that dissolving high-activity ginkgo leaf extract and sig water with ultrasonic method regulates pH and be controlled in certain scope, it and the common dissolved method comparison of heating, bilobalide is not destroyed.Bilobalide is that a kind of ischemic rat brain is damaged has protective effect; the film that can suppress hypoxia inducible in the brain decomposes; improve cyanide and handle the neuronic viability of back Embryo Gallus domesticus; induce and improve CA1 cone innervation; suppress MPN and reduce butyrine level and glutamate decarboxylase activity in Hippocampus and the cerebral cortex, have the protective effect of nervus centralis and remove effect such as free radical.The bilobalide that fully keeps as effective ingredient in lyophilized injectable powder also is characteristics of the present invention.
In order to understand the present invention better, the explanation that below will give an actual example, but this does not mean it is limitation of the present invention.
At first noun is made an explanation and the method for inspection is described:
Flavone is meant the various flavonoid glycosides that exist in the Semen Ginkgo and the general name of flavone aglycone compounds, mainly to be Quercetin, kaempferide and isorhamnetin exist with the form of glycosides contained flavone in extract of the present invention, alleged flavonoid glycoside content is meant that extract is behind hydrochloric acid hydrolysis, flavonoid glycoside wherein is hydrolyzed into aglycon, pressing the HPLC method again measures, the content of the flavonoid glycoside that calculates, total lactone is meant various Semen Ginkgo diterpenoid-lactones and sesquiterpene lactones summation.
Total flavonoid glycoside content is calculated as follows;
Total flavonoid glycoside content=(quercetin content+kaempferide content+isorhamnetin content) * 2.51.
All expressions by weight percentage of the content of said total flavonoid glycoside and total lactone content among the present invention.
Supersound extraction is to make the effective ingredient stripping by ultrasound wave.
The specific embodiment
Embodiment 1
The preparation method of a extract A:
Get Semen Ginkgo primary extract (EGb total flavonoid glycoside content＞24%, total lactone content＞6%) 100 grams, add the ultrasonic 45min of water, residue adds water supersound process 30min, and (the polyamide amount is 10 times of Folium Ginkgo extract to get polyamide column on the supernatant.Before the upper prop, newly with polyamide need be soaked in water spend the night after, the flushing of the water of 5 times of retention volumes, standby), the water eluting discards and is equivalent to polyamide retention volume water lotion, continues the water eluting, collects the water of about 8 times of volumes of polyamide weight, be concentrated into 1/8 volume, for extracting total lactone, standby.Polyamide column behind the water eluting is used 60% ethanol elution, is index with the reaction of HCl-Mg powder, collect alcohol eluen, vacuum concentration, spray drying (inlet temperature: 160 ℃～180 ℃, 80 ℃～90 ℃ of outlet pathogenic wind-warm degree) get the yellowish-brown dried powder, the acetone that adds 10 times of dry powder amounts, supersound extraction 2 times filters, merging filtrate, evaporated under reduced pressure, dry substance stirs (100r/min) extraction 2 times, sucking filtration with the ethyl acetate of 10 times of amounts, get filtering residue, vacuum drying, dry thing was pulverized 100 mesh sieves, again in 50 ℃ of vacuum dryings 24 hours, get extract 7.5 grams of yolk color, be extract A, the content of total flavonoid glycoside is 71.8%, and extract A is 7.5% to the EGb yield.
The preparation of b extract B
In the extract A preparation process, be concentrated into 1/8 volume for extracting the concentrated solution that total lactone is used, ethyl acetate extraction with concentrated solution 2/3 volume, behind the extracting solution water washing secondary, No. 3 sand stamen funnel filters, 75 ℃ of rotary evaporations of filtrate are removed ethyl acetate, the foaming thing of yellowish white, 80 ℃ of vacuum dryings 0.5 hour, room temperature vacuum drying 3 hours, pulverized 100 mesh sieves, 50 ℃ of vacuum dryings 24 hours obtain loose yellowish white total lactone extract 4.7 grams, and total lactone content is 80.6%, the total lactone ratio of BB/ is 46.44%, is 4.7% to the yield of EGb.
The preparation of c high-activity ginkgo leaf extract
With extract A 6.6 grams, extract B 1.5 grams, ratio is 4.4: 1, and mix homogeneously gets high-activity ginkgo leaf extract 8.1 grams, and total flavonoid glycoside and total lactone content are 73.4%, and ginkgoic acid content is below the 5ppm.
Get the high activity extract 8.1g of embodiment 1, lactose 130g adds injection water 1.9L, ultrasonic 30min makes dissolving, drip 1mol/L NaOH, the limit edged fully stirs, and regulates pH7, add the injection water to 2000ml, shake well, 0.8mm film coarse filtration, 0.2mm filter membrane fine straining, the lyophilizing of Spain Telstar L-100 freeze dryer is put in fill (2ml puts in the cillin bottle of 8ml).Lyophilisation condition is: pre-freeze keeps 3h to-40 ℃; Sublimation drying: be warming up to 3.0 ℃ with 45min, be warming up to 5.0 ℃,, be warming up to 35 ℃ with 3h in 5.0 ℃ of maintenance 6h with 6h; Dry again: as to keep 7h in 35.0 ℃.Extract the interior air of bottle out and jump a queue the bundle aluminium lid promptly.Every bottle contains total flavonoid glycoside 4.64mg, contains total lactone 1.14mg, and the total lactone of BB/ is 40.35%; The content of total flavonoid glycoside and total lactone is 71.36%.
Claims (4)
1. ginkgo leaf extract freeze dried powder injection, it is characterized in that this lyophilized injectable powder contains the Folium Ginkgo extract, this Folium Ginkgo extract is to be mixed by Folium Ginkgo total flavones glucoside extract A and ginkgo biloba leaf total terpene lactone extract B, it is more than 65% that extract A contains total flavonoid glycoside, extract B contains total lactone more than 75%, wherein the content of bilobalide in total lactone is more than 40 percent, extract A and extract B mixed proportion are 4.20-5.0: 1.05-1.80, contain total flavonoid glycoside and total lactone more than 65% through mixed extract, ginkgoic acid content is less than 5ppm;
Wherein Folium Ginkgo extract A and Folium Ginkgo extract B are by following method preparation:
The preparation of extract A: get the apricot yellow ketoside of argentiferous greater than 24% and bilobalide greater than the first extract of 6% Folium Ginkgo, add the water supersound extraction, get supernatant, last polyamide column, wherein just the ratio of extract and polyamide is 1: 8-15, the water eluting, discard the water of retention volume, collect all lactone stream parts, use for extracting Folium Ginkgo total lactones, continue with 60% ethanol elution, just the alcoholic acid ratio of extract and eluant 60% is 1: 80-120, eluent concentrating under reduced pressure, spray drying, get the yellowish-brown powder, this powder adds the acetone supersound extraction 2 times, extracting liquid filtering, be evaporated to driedly, dry substance adds the ethyl acetate stir process, filter, the filtering residue vacuum drying, porphyrize gets pale yellow powder and is extract A;
The preparation of extract B: get lactone stream part of the polyamide water eluting in the total flavonoid glycoside extract preparation process, concentrating under reduced pressure, ethyl acetate extraction, ethyl acetate extraction liquid vacuum concentration, drying, porphyrize get extract B; Or the lactone of polyamide water eluting stream part is through concentrated, cooling, crystallization, filtration, get white lactone crystallization thing, the lactone crystallization thing is through vacuum drying, porphyrize is standby, mother solution reuse ethyl acetate extraction is got ethyl acetate extraction liquid, the powder that concentrate, vacuum drying, porphyrize gets ivory buff, add above-mentioned lactone crystallization thing, mix homogeneously is extract B.
2. according to the lyophilized injectable powder of claim 1, it is characterized in that every bottle of this lyophilized injectable powder contains Folium Ginkgo extract 7.5-10mg.
3. according to the lyophilized injectable powder of claim 1, it is characterized in that this lyophilized injectable powder prepares with the following method:
(1) prescription: Folium Ginkgo extract 7.5-10 gram
Lactose 110-140 gram
Water for injection 2000ml
(2) technical process: Folium Ginkgo extract and the lactose of getting formula ratio add the injection water, use ultrasonic dissolution, and the hydro-oxidation sodium solution is regulated pH to 7, add the injection water to capacity, aseptic filtration, and packing, lyophilization gets flaxen loose block.
4. be used to prepare the purposes of the medicine that prevents and treat ischemic cerebrovascular, the decline of senile cognitive function and senile dementia disease according to the lyophilized injectable powder of claim 1.
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| CN102552340A (en) * | 2010-12-15 | 2012-07-11 | 中国医药集团总公司四川抗菌素工业研究所 | Preparation method of ginkgolide monomer and total ginkgo flavone-glycoide |
| WO2019113861A1 (en) * | 2017-12-13 | 2019-06-20 | 巴卫松 | Ginkgo biloba extract medicinal raw material and preparation method therefor |
| CN109793233A (en) * | 2018-07-25 | 2019-05-24 | 上海药辰生物科技有限公司 | A kind of deep working method of ginkgo nut endosperm |
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