CN1935176A - Method for preparing red flower extract containing total red flower uranidin - Google Patents
Method for preparing red flower extract containing total red flower uranidin Download PDFInfo
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Abstract
The present invention relates to a preparation method of carthamus extract. Said method includes the following steps: soaking carthamus flower in warm water to obtain extract A; concentrating the extract A to obtain concerrtrate, separating said concentrate by means of macroporous adsorbing resin, then successively using water and alcohol to make elution, collecting eluted portion by using alcohol, reeovering eluent until the alcohol content is below 25% so as to obtain eluent B; using ceramic membrane to make ultrafiltration so as to obtain ultrafiltrate C; decompressing ultrafiltrate, recovering solvent to obtain heavy paste, drying so as to obtain the invented product. Said extract is characterized by that total carthamus yellow content is greater than 80%, hydroxyl carthamus yellow A content is greater than 20% and kaempferol-3-0-rutinoside content is greater than 0.6%.
Description
Technical field
The present invention relates to a kind of preparation method of Flos Carthami extract, specifically, is Flos Carthami extract that contains total Carthamus yellow and preparation method thereof, and the pharmaceutical composition and the application thereof that contain this extract.
Background technology
Chinese medicine safflower is the dried floral of feverfew Carthamus tinctorius L., it is traditional drug for invigorating blood circulation and eliminating stasis, effect with blood circulation promoting and blood stasis dispelling, inducing menstruation to relieve menalgia, many disturbance of blood circulation diseases such as prescription treatment coronary heart disease, angina pectoris of utilizing Flos Carthami clinically or containing Flos Carthami have good curative effect, simultaneously, total Carthamus yellow is because of having the very strong stability of good colorability, as food coloring and extensive use.Since the seventies, very deep to the chemical constitution study of Chinese medicine safflower, and clear and definite its effective ingredient is the total Carthamus yellow of water solublity, its main effective monomer component is a hydroxyl safflor yellow A etc.The preparation method of relevant total Carthamus yellow has a lot of reports, and early stage method commonly used mainly is a water extract-alcohol precipitation, gets extract [Zhou Yufang etc., the extraction of safflower effective ingredients, Xinjiang Polytechnical College's journal, 1997,18 (4): 270-272] after the extracting solution drying; In recent years many waters or rare pure lixiviate or heating extraction, extracting solution carries out purification by polyamide or macroporous adsorbent resin, gets extract after the drying [referring to CN 1101424C; CN 1085674C].Above-mentioned two patents adopt the polar solvent lixiviate, method with the adsorbents adsorb effective ingredient after the extracting solution simple process is carried out separation and purification, obtain extract after the drying, this method has improved the quality of Flos Carthami extract to a certain extent, but the amount of the composition of the extract effective ingredient that makes of method and impurity is all indeterminate thus, cause the preparation stability and the safety that form by its exploitation relatively poor, especially be not suitable for injection, along with Chinese medicine extensive use clinically, report a lot [Mo Binbin etc. by the untoward reaction that Chinese medicine causes, the document statistical analysis of Chinese medicine untoward reaction, Central-South pharmacy, 2003,1 (3): 184-185], great majority are to make with extra care the degree deficiency and cause owing to continuing to use traditional extraction process, so its drug safety is a problem worried in the industry and that pay close attention to.How to develop a kind of sophisticated preparation technology, make the effective ingredient of Flos Carthami extract and content is clear and definite, purity is high, impurity is few, and can remove polymerization macromole and pyrogen simultaneously, improve the preparation stability of the Flos Carthami extract that contains total Carthamus yellow, make it to be directly used in injection, and then guarantee the especially safety of injection of Flos Carthami preparation, become the problem that the insider endeavours to solve.
Summary of the invention
The inventor passes through great deal of experimental, obtained the effective ingredient of Flos Carthami extract, it is a kind of extract that contains total Carthamus yellow (also can be described as Carthamus yellow), this extract is that the Flos Carthami crude extract makes through making with extra care, and wherein the content of total Carthamus yellow is very high, has solved the low defective of prior art effective ingredient, reached the purpose of the composition of clear and definite safflower effective ingredients, and improved its content, thus improved bioavailability, increased therapeutic effect.
The object of the present invention is to provide the above-mentioned preparation method that contains the Flos Carthami extract of total Carthamus yellow, this method comprises the extraction subtractive process, especially adopt the technology of HPD600 type or HP-20 type macroporous adsorbent resin and ceramic membrane ultrafitration, make active constituent content height in the Flos Carthami extract, impurity few, more help providing its application on preparation.
The present invention also aims to provide a kind of Flos Carthami extract, it is made with extra care by conventional extraction separation and ceramic membrane ultrafitration and forms, and this extract component is clear and definite, is more suitable for being directly used in the injection of Flos Carthami extract.
The invention provides a kind of preparation method of Flos Carthami extract, comprising: the Chinese crude drug Flos Carthami is obtained extracting solution with at least 2 times of water temperature lixiviates; Concentrate, then macroporous adsorbent resin on the concentrated solution is separated, with water and low concentration alcohol (concentration is about 20-50%) eluting, collect pure eluting part successively, reclaim eluent to pure content below 20%; By ceramic membrane ultrafitration, the ultrafiltrate drying.
The benefit that flos carthami adopts temperature to carry in the inventive method is, some effective ingredient that contain in the Flos Carthami extract not exclusively when merceration (method that cold water soak is extracted), and might make effective ingredient destroyed because of oxidation or decomposition under the situation of heating and refluxing extraction, the screening experiment of just carrying proves that also the effect that temperature is carried is better than merceration and Re Ti.2-14 times of water temperature put forward the extracting solution that obtains preferably is fit to Flos Carthami extract by the process screening HPD600 type or HP-20 type macroporous adsorbent resin and ultrafilter membrane (ceramic membrane) isolation technics, the productive rate and the content of effective ingredient have not only been improved, simultaneously also removed polymerization macromole and the pyrogen in the Flos Carthami extract effectively, it can be applied in the injection type safely and effectively.
It is 40-90 ℃ that said method warm macerating wherein extracts temperature, preferred 50-80 ℃, and most preferably 65-75 ℃; Wherein separation steps comprises earlier with 1-2 bed volume washing, the blade diameter length ratio of described post is 1: 6-1: 10, discard water liquid to remove water-solubility impurities such as polysaccharide, reuse low concentration (20%-50%, preferred 30%) a pure eluting 3-5 bed volume, the collection eluent is recycled to determining alcohol and is lower than 20%; The molecular cut off of described ceramic membrane is preferably 10,000, if take all factors into consideration factor such as yield, also can be 6000-5 ten thousand, is preferably 1-3 ten thousand, so that improve purity and yield, and more effectively remove polymerization macromole and pyrogen in the described Flos Carthami extract.
The kind that is used for the membrane ultrafiltration of Chinese medicine extraction field of refinement has a lot, for example polysulfones organic membrane, polyamide membrane, cellulose acetate film, ceramic membrane, tetrafluoroethylene etc., so long as define the ultrafilter membrane of identical molecular cut off scope, the effect of its ultrafiltration should be identical in theory.But in actual mechanical process to the refining ultrafiltration of Flos Carthami extract, the applicant finds, for the injection preparation that preparation contains total Carthamus yellow, the ultrafiltration effect of ceramic membrane is best, wherein index performances such as the stability of this film, resistance to acids and bases, permeability are good, for filtering the high strong Chinese medicine extract of this toughness of total carthamus tinctorius yellow color content, greatly minimizing, workable of corresponding loss, obviously be better than the effect that other membrane ultrafiltration can reach, the contrast and experiment of handling for same amount medical material and identical molecular cut off is as follows:
The present invention also screens the kind model of adsorbent resin, and concrete screening technique is: get dissimilar resins such as HPD600, HPD100, Dianion HP-20 and 860021 types, at first carry out pretreatment, get 1L adsorbent resin dress post, and stand-by; With Flos Carthami extract (recording wherein, hydroxyl safflor yellow A content is 7.628mg/ml) upper prop, whether TLC checks saturated, wash with water earlier, reuse Different concentrations of alcohol eluting, collected post residual liquid, water lotion, ethanol elution respectively, measure wherein hydroxyl safflor yellow A content, and calculate relevant characterisitic parameter, the results are shown in Table 7.
Table 7 different model resin absorption purification characterisitic parameter
| The resin model | Than the plain A of upper column quantity medical material hydroxyl safflower yellow (g/g) (mg/g) | Than adsorbance hydroxyl safflor yellow A (mg/g) | Than elution amount (hydroxyl safflor yellow A meter, mg/g) 30% ethanol |
| HPD 600 (polarity) HP-20 (nonpolar) HPD100 (nonpolar) 860021 (Semi-polarity) | 0.215 13.45 0.207 11.35 0.110 6.86 0.298 18.65 | 11.08 10.21 1.82 15.82 | 11.50 10.39 2.18 2.29 |
By table 7 result as can be known, be index components with the hydroxyl safflor yellow A, Flos Carthami extract has different absorption, elute effect on different resins, and wherein HPD100 type resin is more less than upper column quantity; The ratio upper column quantity of HPD600, HP-20,860021 model resins and all bigger than adsorbance, but the ratio elution amount during 860021 usefulness, 30% ethanol elution is little, and promptly sample loss is serious.The ratio upper column quantity of HP-20 and HPD 600 type resins is big and all bigger than elution amount, and the absorb-elute performance is fine.Therefore with respect to the more heavy-gravity Flos Carthami extract of extract, preferably adopt HP-20 and HPD 600 type resins to carry out separation and purification, more preferably HPD 600 type resins.
Determine that by testing upward column condition is:
Blade diameter length ratio: 1: 6-1: 10, preferred 1: 8
Adsorption temp: room temperature (＜35 ℃)
Liquor strength: be equivalent to the 80-130g crude drug/liter, preferred 110g/L.
Extracting solution after said extracted is refining can obtain the prepared solids extraction thing of the inventive method through super-dry after testing, and drying mode can be any conventional drying method, preferably spray drying.
Preparation method of the present invention is compared with the disclosed preparation method of prior art, because the concrete scheme difference that the separation and purification step adopts, the effective ingredient that the extract that obtains wherein contains shows evident difference, refining as having adopted repeatedly column chromatography in the extract for preparing in patent 01131268.8 and 03157479.3, the content of hydroxyl safflor yellow A is all greater than 70, other flavochrome compounds all are used as impurity and remove, and result of study of the present invention shows, the too high levels of hydroxyl safflor yellow A, curative effect and active high better not as good as the content of total this index of Carthamus yellow on the contrary, therefore the extract by the inventive method preparation has better clinical application value.
Innovation part of the present invention is by using specific macroporous adsorbent resin and ceramic membrane isolation technics, improved the content of total Carthamus yellow in the Flos Carthami extract, make that simultaneously preparation (especially injection type) stability that contains this extract improves greatly, removed polymerization macromole and the pyrogen in the plant extract effectively, can directly be prepared into injection, and preparation security has obtained effective assurance.
The present invention also provides a kind of Flos Carthami extract, it is to make with above-mentioned preparation method, after testing, the content of total Carthamus yellow is at least 60% in this extract, preferred content is (in the hydroxyl safflor yellow A) more than 80%, wherein the content of hydroxyl safflor yellow A is at least 15%, and is preferred more than 20%.
Because the total Carthamus yellow of effective ingredient that is contained in the extract of the present invention is a mixture, the clear and definite after testing active compound composition that it contained comprises the hydroxyl safflor yellow A, hydroxyl safflower yellow B, kaempferol-3-O-rutinoside, Quercetin-3-O-glucoside etc., so far there is not document to report openly what Chinese medicine safflower is at the reasonable composition of effective ingredient aspect the treatment cardiovascular and cerebrovascular disease, how many preferred therapeutic amounts is, certainly will cause occurring dose-effect relationship like this is not easy to hold, defective characteristics such as curative effect instability, its clinical experiment effect neither be very desirable, especially for the Chinese medicine injection dosage form, constituent is definite, content accurately is easier to hold the quality of the pharmaceutical preparations.The applicant finds, definite curative effect chemical compound is the hydroxyl safflor yellow A of (but being not only) treatment effective dose in the extract that preparation method of the present invention obtains, other compositions except the hydroxyl safflor yellow A (kaempferol-3-O-rutinoside for example even, Quercetin-3-O-glucoside) also progradation is played in treatment, so having carried out a large amount of experiments, the present invention detects, the result confirms, the content of total Carthamus yellow is at least 60% in the Flos Carthami extract, and wherein the content of hydroxyl safflor yellow A is at least 15%, it is best that the content of kaempferol-3-O-rutinoside is at least its therapeutic effect of Flos Carthami extract of 0.8%, obviously is better than the Flos Carthami extract with the method preparation of documents.
The content assaying method of above-mentioned total Carthamus yellow is the ultraviolet light absorption photometry, and operating procedure is:
It is an amount of that precision takes by weighing hydroxyl safflor yellow A reference substance, adds water and make reference substance solution.The extract of getting preparation as stated above during mensuration is an amount of, and accurate the title decides, be dissolved in water as need testing solution, and according to spectrophotography (2000 editions one appendix VA spectrophotography of Chinese Pharmacopoeia), be blank with water.Measure reference substance and absorption of sample degree at 332nm wavelength place, calculate the amount that is equivalent to the hydroxyl safflor yellow A in the need testing solution by external standard method, promptly.
The assay of hydroxyl safflor yellow A adopts high effective liquid chromatography for measuring, the concrete operations step:
With octadecylsilane chemically bonded silica is filler; The sodium dihydrogen phosphate of mobile phase A: 0.05mol/L (it is 4.0 that phosphoric acid is regulated pH value).Mobile phase B: acetonitrile.Flow velocity is 1.0ml/min, and the detection wavelength is 403nm.The separating degree of hydroxyl safflor yellow A and other impurity peaks should be up to specification, and theoretical cam curve is pressed hydroxyl safflor yellow A peak and calculated, and should be not less than 3000.
The preparation of reference substance solution: it is an amount of that precision takes by weighing reference substance hydroxyl safflor yellow A, is dissolved in water to be diluted to debita spissitudo product solution in contrast.
The preparation of need testing solution: it is an amount of to get extract, is diluted to debita spissitudo with water dissolution and both gets.
Algoscopy: accurate reference substance solution and each the 20 μ l of need testing solution of drawing of difference, inject hplc determination, press external standard method with calculated by peak area, promptly.
Kaempferol-3-O-rutinoside assay adopts high effective liquid chromatography for measuring, and operating procedure is:
With octadecylsilane chemically bonded silica is filler; The phosphoric acid of mobile phase A: 0.05mol/L.Mobile phase B: acetonitrile.Flow velocity is 1.0ml/min, and the detection wavelength is 365nm.The separating degree of kaempferol-3-O-rutinoside and other impurity peaks should be up to specification, and theoretical cam curve is pressed kaempferol-3-O-rutinoside peak and calculated, and should be not less than 3000.
The preparation of reference substance solution: it is an amount of that precision takes by weighing reference substance kaempferol-3-O-rutinoside, is dissolved in water to be diluted to debita spissitudo product solution in contrast.
The preparation of need testing solution: it is an amount of to get extract, is diluted to debita spissitudo with water dissolution and both gets.
Algoscopy: accurate reference substance solution and each the 20 μ l of need testing solution of drawing of difference, inject hplc determination, press external standard method with calculated by peak area, promptly.
The present invention also provides a kind of pharmaceutical composition, and it contains Flos Carthami (Carthamus yellow) extract and the conventional medicine adjuvant of clinical treatment effective dose, and it is 60-240mg/ day that this clinical drug is recommended effective dose.
Aforementioned pharmaceutical compositions comprises peroral dosage form and injection type, and optimizing injection comprises infusion solutions, freeze-dried powder and little liquid drugs injection.In said medicine, as the auxiliary element that is used with effective ingredient, according to different concrete injections, at least a in mannitol, glucose, dextran or the sodium chloride selected in suggestion for use.For example, for lyophilized injectable powder, can adopt conventional freeze-drying, with water is solvent, get extract and add an amount of caffolding agent lyophilizing and form, at least a as auxiliary caffolding agent in mannitol, glucose, the dextran selected in suggestion for use, forms jointly with said extract; Wherein preferred mannitol is as caffolding agent.For oozing the medicine of infusion preparation form, then can select at least a in mannitol, glucose, the sodium chloride for use as auxiliary element for the grade of directly using; Wherein excellent serves as preferred to adopt sodium chloride as auxiliary element.During for preparation small-volume injection injection and great transfusion preparation, can add antioxidant, antioxidant can be one or more in ethylenediaminetetraacetic acid, disodium EDTA, ascorbic acid, sodium ascorbate, sodium thiosulfate and the sodium pyrosulfite.
The present invention also provides the above-mentioned application of pharmaceutical composition in the medicine of preparation treatment cardiovascular and cerebrovascular disease that contains Flos Carthami extract.
Carthamus yellow extract test of pesticide effectiveness result of the present invention is as follows:
(1) research of the anti-myocardial ischemia effect of Carthamus yellow intravenously administrable
1. mice is closed the influence of trachea electrocardio extinction time:
Tried mice with 60 and be divided into 6 groups at random, every group of 10 male and female half and half, mice be will be tried and urethane (1.2g/kg) intraperitoneal injection of anesthesia, fixing, the also free trachea of throat portion surgical exposure used, link to each other with the mice extremity through pin type electrode with MS302 multimedization bio signal record analysis system, gather mark II lead electrocardiogram, the 1st group of matched group injecting normal saline of tail venipuncture drug administration by injection.2nd, 3,4 groups is Carthamus yellow senior middle school low dose group, and dosage is respectively 120mg/kg, 60mg/kg, and 30mg/kg, the 5th group is Carthamus yellow gastric infusion group, dosage is 60mg/kg, in closing preceding 30 minutes gastric infusions of trachea.Each is organized the administration volume and is 0.2ml/10g, and injection speed is that 0.02ml/s is mixed with desired concn with 0.9%NaCl before test.Use small artery clip closure trachea behind the drug administration by injection at once, write down every group every mice and close trachea to the electrocardio extinction time, the testing data significance test is handled with the t-test of pairing data, result of the test is as shown in table 1, and the Carthamus yellow intravenous administration obviously prolongs the electrocardio extinction time and matched group relatively has highly significant difference in selected dosage range; Carthamus yellow 60mg/kg group gastric infusion is had prolongation trend to trying the mice electrocardio, and does not have significant difference between matched group.
The influence of trachea electrocardiogram extinction time is closed in the intravenous injection of table 1 Carthamus yellow to mice
Each group compares with matched group
*P＜0.05
*P＜0.01
2. to the influence of mice anoxia enduring:
Contain soda lime (calcium hydroxide factory of Beipiao Mine Administration, 991001), with in the bottleneck air-tight bottle, regulate CY-2 type oxygen analyser, it is linked to each other, writes down the death time (min) that oxygen content in the container (%) changes and write down mice with hermetic container with vaseline.The testing data significance test is handled with the t-test of pairing data, result of the test is as shown in table 2, in selected dosage range, the Carthamus yellow intravenous administration obviously reduces oxygen consumption and prolongs the mice time-to-live, relatively has significantly and highly significant difference with matched group.Carthamus yellow 60mg/kg dosage group is with gastric infusion, and the mice oxygen consumption reduces and prolonged survival period has certain effect to being tried, but and do not have significant difference between matched group.
The intravenous injection of table 2 Carthamus yellow is to the influence of white mice oxygen consumption (%) time-to-live (min)
| Group | Dosage (mg/kg) | Number of animals (only) | Oxygen content (%) | Death time (min) | ||
| 5min | 10min | 15min | ||||
| Matched group Carthamus yellow iv Carthamus yellow iv Carthamus yellow iv Carthamus yellow ig | 0.9％NaCl (0.2ml/kg) 120 60 30 60 | 10 10 10 10 10 | 13.3±0.8 15.6±1.2 ** 14.9±1.1 ** 14.2±1.1 13.6±1.4 | 8.3±0.5 11.3±1.4 ** 10.8±1.2 ** 9.6±1.1 ** 8.9±1.3 | 6.4±0.5 7.9±1.0 ** 7.6±1.2 * 6.9±1.1 6.8±1.1 | 16.6±1.6 25.2±3.8 ** 23.1±3.3 ** 22.7±3.5 ** 17.3±4.1 |
Each administration group and matched group are relatively
*P＜0.05
*P＜0.01
(2) Carthamus yellow antiplatelet aggregation and thrombotic effect
1. to the poly-influence that acts on of ADP induced platelet:
60 male Wister rats are divided into 6 groups at random, matched group intravenous injection 0.9%NaCl Carthamus yellow intravenous injection group, dosage is respectively 80mg/kg, 40mg/kg, 20mg/kg, Carthamus yellow gastric infusion group dosage is 40mg/kg, each is organized the medicine group and prepare the desired concn medicinal liquid with 0.9% sodium chloride before administration, the intravenous administration volume is 5ml/kg, and injection speed is 2.0ml/min.The gastric infusion volume is 20ml/kg, 1 successive administration of administration every day 7 days, after the last administration 1 hour through vena orbitalis posterior clump blood sampling with 0.38% sodium citrate anticoagulant (1: 9), with the centrifugal 600rpm of anticoagulation, 10min prepares rich blood plate blood plasma (PRP) and blood plasma usefulness under She was prepared platelet poor plasma (PPP) in 3000rpm15 minute.
PAM-3 type dual pathways platelet aggregation instrument is given hot 30min.When treating 37 ℃ of machine temperature meter pointed, in the cleaning cuvette, move into 200 μ lPPP, stir and give hot 5min, transfer the PPP potentiometer with micropipette, make monitor pointed 20, take out cuvette, get the same blood sample PRP of 200 μ l again, pour into than in the turbid cup, give heat, reach 37 ℃.Transfer the PRP potentiometer, make monitor pointed 80, add the ADP working solution (2.2 μ M) of the new preparation of 20 μ l this moment in PRP, curve of platelet aggregation is traced in the variation of observation recorder pointer, and according to platelet aggregation rate (%), converse platelet aggregation inhibition rate, compare significance test, result of the test shows, shifting to an earlier date administration 7 days at selected dosage has obvious inhibitory action to ADP induced platelet aggregation rate, has between each administration group and matched group significantly or highly significant difference.Detailed ginseng table 3.
The quiet notes of table 3 Carthamus yellow to ADP induce rat platelet aggregation influence (
N=10)
| Group | Dosage (mg/kg) | Platelet aggregation rate (%) | Suppress (%) |
| Matched group Carthamus yellow iv Carthamus yellow iv Carthamus yellow iv Carthamus yellow ig | 0.9NaCl 5ml/kg 80 40 20 40 | 51.2±4.7 20.7±3.2 ** 30.6±3.8 ** 36.7±4.4 ** 31.2±4.1 ** | 59.6 40.3 28.3 39.1 |
Compare with matched group
*P＜0.05,
*P＜0.01
2. the influence that platelet dependency blood plate is formed
(male body weight 2.0-3.0kg) divides 6 groups at random with 36 rabbit, and 6 every group, matched group, 3 dosage groups of Carthamus yellow intravenous injection, dosage is respectively 44mg/kg, 22mg/kg, 11mg/kg; Carthamus yellow is irritated the stomach group, and dosage is 22mg/kg, positive control drug FUFANG DANSHEN ZHUSHEYE group, and dosage is 1500mg/kg.Matched group intravenous injection 0.9%NaCl, each administration group is all prepared the desired concn medicinal liquid with 0.9%NaCl before administration, and the intravenous injection volume is 2ml/kg.The gastric infusion volume is 5ml/kg.1 successive administration of administration every day 7 days uses pentobarbital sodium with 30mg/kg auricular vein injecting anesthetic after the last administration.
Animal subject back of the body position is fixing, throat preserved skin, operation, free common carotid artery, with little fine needle 1 trumpeter's art silk thread is penetrated about 3cm in the side common carotid artery, after intra-arterial is preserved 2 hours, cut common carotid artery, removal of thromboses claims weight in wet base behind the removal silk thread, is observation index with the wet weight of thrombus, compare between organizing, significance test, result of the test show, administration in advance 7 days has obvious inhibitory action to rabbit common carotid artery thrombosis in selected dosage range, each administration group and matched group relatively have significantly or highly significant difference, detailed ginseng table 4.
Table 4 Carthamus yellow intravenously administrable to the thrombotic influence of rabbit common carotid artery (
Mg)
| Group | Dosage mg/kg | Number of animals only | Wet weight of thrombus mg | Thrombosis suppression ratio % |
| Matched group Carthamus yellow iv Carthamus yellow iv Carthamus yellow iv Carthamus yellow ig | 0.9％NaCl 2.0ml 44 22 11 22 | 6 6 6 6 6 | 18.4±4.3 ** 9.7±1.6 ** 11.9±2.3 ** 13.2±1.7 * 13.4±1.2 * | 47.3 35.3 28.3 27.2 |
Compare with matched group
*P＜0.05
*P＜0.01
The extract that relates in the pharmacological evaluation of the present invention makes according to the method for embodiment 1.
The specific embodiment
Following examples are in order to illustrate in greater detail the present invention, are not that the present invention is construed as limiting.
The preparation of embodiment 1 flooding contains the Flos Carthami extract of total Carthamus yellow
Get flos carthami 2000g, add 10 times water (medical material weight), stir to extract 2 times in 70 ℃, each 1 hour, filter, merge secondary raffinate, be cooled to room temperature after.According to upper prop sample size and amount of resin (1: 10/W: ratio V), cross the HP-20 macroporous adsorbent resin column chromatography, the blade diameter length ratio of post is 1: 7, water is with 1-2ml/cm
21 bed volume of the flow velocity eluting of/min discards, and continues with 25% ethanol 1-2ml/cm
24 bed volumes of the flow velocity of/min, 30% ethanol elution partly is total Carthamus yellow part, 70 ℃ of decompression and solvent recoveries, be about 1.05 concentrated solution to proportion, the ceramic membrane of above-mentioned solution being crossed molecular cut off 8000 carries out ultrafiltration, collect the filtrate after drying and both got the total Carthamus yellow of extract, can be directly used in the preparation injection.
After testing, total carthamus tinctorius yellow color content is 85% in this extract, and hydroxyl safflor yellow A content is 25%, kaempferol-3-O-rutinoside content 0.7%.
The preparation of embodiment 2 floodings contains the Flos Carthami extract of total Carthamus yellow
Get flos carthami 2000g, add 8 times water (medical material weight), stir to extract 2 times in 75 ℃, each 1 hour, filter, merge secondary raffinate, be cooled to room temperature after.According to upper prop sample size and amount of resin (1: 8/W: ratio V), add HPD 600 macroporous adsorbent resin column chromatography, the blade diameter length ratio of post is 1: 8, water is with 1.5-2.5ml/cm
21 bed volume of the flow velocity eluting of/min discards, and continues with 40% ethanol 1-2ml/cm
24 bed volumes of the flow velocity of/min, 40% ethanol elution partly is total Carthamus yellow part, 70 ℃ of decompression and solvent recoveries, be about 1.02 concentrated solution to proportion, the ceramic membrane of above-mentioned solution being crossed molecular cut off 10000 carries out ultrafiltration, collects the filtrate after drying and promptly gets the total Carthamus yellow of extract.
After testing, total carthamus tinctorius yellow color content is 81% in this extract, and hydroxyl safflor yellow A content is 23%, kaempferol-3-O-rutinoside content 0.65%.
Embodiment 3 rare alcohol extraction preparations contain the Flos Carthami extract of total Carthamus yellow
Get flos carthami 2000g, add 10 times 30% ethanol (medical material weight), reflux, extract, 2 times each 1 hour, filters, merge secondary raffinate, reclaim extracting solution to there not being alcohol, according to upper prop sample size and amount of resin (1: 10/W: ratio V), adding HP-20 macroporous adsorbent resin column chromatography, the blade diameter length ratio of post is 1: 10, and water is with 1-2ml/cm
21 bed volume of the flow velocity eluting of/min discards, and continues with 30% ethanol 1-2ml/cm
24 bed volumes of the flow velocity of/min, 35% ethanol elution partly is the Carthamus yellow part, and 70 ℃ of decompression and solvent recoveries are about 1.05 concentrated solution to proportion, the organic membrane of above-mentioned solution being crossed molecular cut off 10000 carries out ultrafiltration, collects the filtrate after drying and gets the extract Carthamus yellow.
After testing, total carthamus tinctorius yellow color content is 88% in this extract, and hydroxyl safflor yellow A content is 27%, kaempferol-3-O-rutinoside content 0.72%.
The preparation of embodiment 4 (always) Carthamus yellow freeze-dried powder
Take by weighing the 310g dextran, be dissolved in 4500 milliliters of waters for injection, take by weighing Carthamus yellow extract 300g again, be dissolved in the above-mentioned solution by the inventive method preparation.Sodium hydroxide solution with 20% is regulated pH to 7.0 ± 0.2.The 0.1% needle-use activated carbon heated and stirred 15 minutes that adds amount of preparation, sucking filtration, filtrate replenishing adds water for injection to 5000 milliliter.Fine straining, intermediate inspection, fill, lyophilizing, gland, finished product inspection, packing.
The preparation of embodiment 5 (always) Carthamus yellow freeze-dried powder
Take by weighing 300g mannitol, be dissolved in 4500 milliliters of waters for injection, take by weighing Carthamus yellow extract 300g again, be dissolved in the above-mentioned solution by the inventive method preparation.Sodium hydroxide solution with 20% is regulated pH to 7.0 ± 0.2.This solution molecular cut off is 10,000 ceramic membrane ultrafitration, moisturizing 8000ml ultrafiltration at twice.Ultrafiltrate is done the intermediate inspection, fill, lyophilizing, gland, finished product inspection, packing.
The preparation of embodiment 6 (always) Carthamus yellow infusion solutions
Get the Carthamus yellow extract 320g that makes as stated above, add the water for injection of 834.7g sodium chloride and 90L, stirring and dissolving, it is an amount of to add 0.1mol/L NaOH, transfer pH to be about 7.0, replenish water for injection, add the needle-use activated carbon of liquid measure 0.15% again to capacity, stirred 15 minutes, the sucking filtration de-carbon, measure content, pH value qualified after, fine straining packing again, (105 ℃) pressure sterilizing, promptly.
The preparation of the little liquid drugs injection of embodiment 7 (always) Carthamus yellow
Get the Carthamus yellow extract 250g that makes as stated above, add the water for injection of 80.0g sodium chloride and 10L, stirring and dissolving, it is an amount of to add 0.1mol/L NaOH, transfers pH to be about 7.0, adds 10g antioxidant sodium thiosulfate, replenish water for injection to capacity, the needle-use activated carbon that adds liquid measure 0.1% again stirred the sucking filtration de-carbon 15 minutes, measure content, pH value qualified after, fine straining packing again, (105 ℃) pressure sterilizing, promptly.
More than described the preferred embodiment for the present invention, so it is not in order to limit the present invention.Those skilled in the art can not depart from the improvement and the variation of category of the present invention and spirit to embodiment disclosed herein.
Claims (10)
1, a kind of preparation method that contains the Flos Carthami extract of total Carthamus yellow, comprising:
(1) Flos Carthami is obtained extracting solution I with at least 2 times of water temperature lixiviates;
(2) extracting solution concentrate concentrated solution, then macroporous adsorbent resin on the concentrated solution is separated, successively with the pure eluting of water and 20%-50%, collect pure eluting part, reclaim eluent to pure content below 20%, obtain eluent II;
(3) by ceramic membrane ultrafitration, obtain ultrafiltrate III;
(4) ultrafiltrate III is 1.05-1.20 through being recycled to relative density, gets final product.
2, the described preparation method of claim 1, wherein warm macerating extraction temperature is 40-90 ℃.
3, the described preparation method of claim 1, wherein macroporous adsorbent resin is HPD600 type or HP-20 type in the step (2), its blade diameter length ratio is 1: 6-1: 10, described separation comprises earlier with 1-2 bed volume washing, discard water liquid, the pure eluting 3-5 bed volume of reuse concentration 20%-50%, the eluent of collecting pure eluting part is recycled to determining alcohol below 20%.
4, the described preparation method of claim 1, wherein the molecular cut off of the ceramic membrane described in the step (3) is 1-3 ten thousand.
5, the described preparation method of claim 1, wherein the alcohol used of the eluting described in the step (2) is 30% ethanol.
6, the described preparation method of claim 1, wherein the determining alcohol of the eluent II that obtains of step (2) is 5%-15%.
7, a kind of Flos Carthami extract that contains total Carthamus yellow that makes with the described preparation method of claim 1 is characterized by total carthamus tinctorius yellow color content＞80%, hydroxyl safflor yellow A content＞20%, kaempferol-3-O-rutinoside content＞0.6%.
8, a kind of pharmaceutical composition is characterized in that containing described Flos Carthami extract and the pharmacy acceptable auxiliary that contains total Carthamus yellow of the claim 7 for the treatment of effective dose.
9, the described compositions of claim 8, wherein this pharmaceutical composition is an injection.
10, the described compositions of claim 9, wherein said injection contain the described Flos Carthami extract 1-3 of claim 7 part, mannitol, dextran or sodium chloride 0.8-5 part.
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| CN2005101053878A CN1935176B (en) | 2005-09-23 | 2005-09-23 | Method for preparing red flower extract containing total red flower uranidin |
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| CN2005101053878A CN1935176B (en) | 2005-09-23 | 2005-09-23 | Method for preparing red flower extract containing total red flower uranidin |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101275167B (en) * | 2008-05-13 | 2010-11-10 | 中国人民解放军第二军医大学 | Screening method for hydroxyl carthamin yellow A-containing safflower |
| CN102119984A (en) * | 2011-03-13 | 2011-07-13 | 浙江大学 | Preparation method and application of carthamus tinctorius L. active ingredient containing hydroxysafflor yellow A |
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| CN101891723B (en) * | 2007-12-04 | 2011-11-02 | 哈药集团中药二厂 | Method for extracting hydroxysafflor yellow A |
| CN103479702A (en) * | 2012-06-14 | 2014-01-01 | 北京星昊医药股份有限公司 | Extraction method and application of safflower yellow pigment |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1085674C (en) * | 1998-11-24 | 2002-05-29 | 北京市心肺血管疾病研究所 | Safflower total uranidin, its preparation and application |
| CN1101424C (en) * | 2000-01-21 | 2003-02-12 | 浙江大学 | Method for extracting natural edible pigment safflor yellow |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101891723B (en) * | 2007-12-04 | 2011-11-02 | 哈药集团中药二厂 | Method for extracting hydroxysafflor yellow A |
| CN101275167B (en) * | 2008-05-13 | 2010-11-10 | 中国人民解放军第二军医大学 | Screening method for hydroxyl carthamin yellow A-containing safflower |
| CN102119984A (en) * | 2011-03-13 | 2011-07-13 | 浙江大学 | Preparation method and application of carthamus tinctorius L. active ingredient containing hydroxysafflor yellow A |
| CN102125592A (en) * | 2011-03-13 | 2011-07-20 | 浙江大学 | Preparation method and use of active ingredients of safflower |
| CN102125592B (en) * | 2011-03-13 | 2012-11-28 | 浙江大学 | Preparation method and use of active ingredients of safflower |
| CN102119984B (en) * | 2011-03-13 | 2013-03-13 | 浙江大学 | Preparation method and application of carthamus tinctorius L. active ingredient containing hydroxysafflor yellow A |
| CN103479702A (en) * | 2012-06-14 | 2014-01-01 | 北京星昊医药股份有限公司 | Extraction method and application of safflower yellow pigment |
| EP2829547A4 (en) * | 2013-02-07 | 2015-11-18 | Zhejiang Yongning Pharmaceutical Co Ltd | Sodium hydroxysafflor yellow a and production method and pharmaceutical use thereof |
| CN116139344A (en) * | 2023-02-20 | 2023-05-23 | 武汉理工大学 | Bone repair material for promoting osteoblast formation and preparation method thereof |
| CN116139344B (en) * | 2023-02-20 | 2024-08-23 | 武汉理工大学 | Bone repair material for promoting osteoblast formation and preparation method thereof |
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| CN1935176B (en) | 2010-07-14 |
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