CN101837037B - Artemisia rupestris L pill and production method and application thereof - Google Patents
Artemisia rupestris L pill and production method and application thereof Download PDFInfo
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Abstract
The invention relates to an artemisia rupestris L pill and a production method and application thereof, the method comprises the following steps: step I: CommonSt.John'swortHerb and artemisia rupestris L with equal quantity are mixed and are crushed into coarse powder, ethanol is added for backflow extraction, the extracting solution is filtered, cooled and centrifuged, and then supernate is concentrated; step II: the concentrated solution is purified by macroporous absorption resin, and effluent liquid is collected in different segments, the effluent liquid is concentrated and is dried to obtain extract powder for spare; step III: the dry extract powder is mixed with polyethylene glycol according to proportion, the mixture is melted by heating, and then pills are produced by dropping. The process in the invention is stable and reliable, the contents of effective ingredients in the pills are stable; the invention solves the problem that residual organic solvent caused by the extraction of organic solvents such as petroleum ether, ethyl acetate and the like is harm to human bodies. Compared with the prior art, the method in the invention is suitable industrial production; the artemisia rupestris L pill obtained in the invention has obvious anti-inflammatory analgesic activities which are proved by experiments in pharmacodynamics and is used for treatment of rheumatism and rheumatoid arthritis.
Description
Technical field
The present invention relates to medical technical field, is a kind of artemisia rupestris L pill and production method thereof and application, and it relates to a kind of pure Chinese medicinal preparation; Particularly relate to a kind of artemisia rupestris L pill of treating rheumatism and rheumatoid arthritis; It adopts macroporous resin technology and drops according to the characteristics of Chinese traditional treatment rheumatism and rheumatoid arthritis, processes a kind of drop pill; It is fast to have curative effect, the characteristics that cure rate is high.
Background technology
Rheumatism, rheumatoid arthritis be a kind of be main chronic autoimmune disease with the arthropathy, be commonly encountered diseases, the frequently-occurring disease of serious harm human health, classified as clinical difficult miscellaneous diseases, this disease is called as " cancer that lives ".Data show, global rheumatisant is about 400,000,000, is one type of the hugest in medical domain illness.It is characterized in that chronic inflammatory arthropathy becomes, and get involved with a plurality of systems of whole body.Rheumatism without systematic treating can be delayed for many years repeatedly, finally causes joint deformity, afunction, causes very big misery to the patient.In China, there is people more than 200,000,000 suffering from rheumatic arthritis or rheumatoid arthritis in varying degrees, wherein comparatively serious basic disability has 8,200 ten thousand people, and comparatively serious life can't be taken care of oneself basically has 1,100 ten thousand people.Above-mentioned this 9,300 ten thousand people has not only had a strong impact on the progress of social productive forces, consumes a large amount of resource medicines, brings great burden to me and family simultaneously.Because rheumatism, rheumatoid arthritis sickness rate and disability rate are high, the healthy of the mankind in serious harm, brings huge misery for countless families, receives whole world medical circle and pays much attention to, and caused whole society's extensive concern.
The medicine of western medical treatment rheumatism and rheumatoid arthritis can be divided into universal medication and special-purpose medicaments two big classes by its scope clinically at present.Universal medication is applicable to multiple rheumatism, and special-purpose medicaments only is applicable to indivedual diseases.Can be divided into NSAID by chemical constitution and pharmacology characteristics, the steroidal anti-inflammatory medicine acts on antirheumatic slowly, cell toxicity medicament and immunomodulator.These medicines only can symptomatic treatment, is prone to recurrence after the drug withdrawal, the course of treatment is long, toxic and side effects is big, and serious gastrointestinal irritation is arranged, and it is big to take side effect for a long time, and patient is difficult for accepting.Undoubtedly, research and development are efficient, the antirheumatic medicine of low toxicity is significant.Therefore, utilize China's traditional medicine, from the traditional plant medicine, excavate a kind of determined curative effect of exploitation, new Chinese medicine that toxic and side effects is low is the direction of current numerous medical research person growing interests for thousands of years in the crystallization of human disease preventing and treating.
In the trend that the world today goes back to nature; Chinese medicine just worldwide attracts great attention and is progressively accepted, like the tripterygium glycosides sheet of using clinically, tripterygium total terpenoids sheet; The Radix Tripterygii Wilfordii microencapsule tablet; The Radix Tripterygii Wilfordii drop pill, Tripterygium Hypoglaucum Hutch Tablet etc. are all having better curative effect aspect treatment rheumatism and the rheumatoid arthritis.China is the development that continues to advance the modernization of Chinese medicine and natural drug at present; List one of main task of pharmaceuticals industry in, wherein the modernization of the national medicines of characteristics such as the exploitation of wind resistance damp disease class medicine and propelling Tibetan, illiteracy, dimension, the Dai Nationality, Yi nationality, Miao Ethnomedicine is one of key content of natural drug development.Uighur medicine is the important component part of motherland's traditional medicine, existing more than 2000 year history, and its theoretical unique and clinical effectiveness is confirmed in practice, has accumulated a lot of abundant and valuable medication experiences aspect the difficult miscellaneous diseases preventing and treating.Therefore; On the basis of analyzing uighur ' s culture medicine ancient books and records and single proved recipe among the people, in conjunction with the unique curative effect in the application among the people, modern study and the exploitation of tieing up medicine are to promote the dimension medicine to move towards modernization; Make the dimension medicine by the market of moving towards among the people, the effective way that benefits the various nationalities masses.
Summary of the invention
The present invention provides a kind of artemisia rupestris L pill and production method and application, has overcome the deficiency of prior art, has solved the organic solvent residual that causes with organic solvent extractions such as petroleum ether, the ethyl acetate problems such as harm to human body; Adopted modern novel purification with macroreticular resin technology; The gained artemisia rupestris L pill contains rupestonic acid and total flavones is higher, and active constituent content is stable in the gained drop pill, and the obvious anti-inflammatory and anti analgesic activity is arranged; Can be used for the treatment of rheumatism and rheumatoid arthritis; And evident in efficacy, the production method of this artemisia rupestris L pill is easy to operate, and production cost is low.
One of technical scheme of the present invention obtains in the following manner: a kind of artemisia rupestris L pill, its raw material Herba Hyperici Monogyni and Herba Achilleae medical material by weight, ratio is 1: 1; This artemisia rupestris L pill contains 10% to 20% rupestonic acid and 20% to 40% total flavones by weight percentage; This artemisia rupestris L pill obtains by following step:
The first step: Herba Hyperici Monogyni and the Herba Achilleae medical material of getting equivalent mix; Be ground into coarse powder; Add 8 to 12 times of amount 60% to 80% alcohol reflux 2 to 3 times, each 1 to 2 hour, merge extractive liquid; Extracting liquid filtering, put coldly, centrifugal, supernatant concentration obtains every milliliter of concentrated solution that contains crude drug 0.20 gram to 0.25 gram;
Second step: get above-mentioned concentrated solution, last macroporous adsorbent resin carries out purification, and the macroporous resin type is one or more among HPD100, HPD400, HPD100A and the HPD400A, the Fractional Collections effluent, and effluent concentrates, drying is processed extract powder, and is subsequent use;
The 3rd step: get above-mentioned dry extract, mix by a certain percentage with Polyethylene Glycol, heating and melting drips and processes drop pill.
Be further optimization and/or selection below to one of technique scheme:
Above-mentioned second step is undertaken by following step: get above-mentioned concentrated solution, and the macroporous adsorbent resin of one or more among mistake HPD100, HPD400, HPD100A and the HPD400A, the resin path height ratio is 1: 4 to 1: 10; Adsorption rate 1.0BV to 3BV/h, elution speed 1.0BV to 3BV/h is earlier with the flow velocity eluting of 3BV to 5BV water with 3BV to 6BV/h; The flush away water-solubility impurity; With 30% ethanol elution 4BV to 6BV, 70% ethanol elution 4BV to 6BV collects 30% and 70% ethanol elution again; Decompression recycling ethanol and to be concentrated into relative density be 1.22 to 1.25 clear paste under 60 ℃; Dry down 60 ℃ of temperature in vacuum drying oven again, get 100 purpose extract powders, subsequent use; Take by weighing the medicinal substrate Polyethylene Glycol, its weight and 100 purpose extract powder ratios are counted 2: 1 to 3: 1 by weight, after complete fusion on 70 ℃ to the 100 ℃ water-baths; Add extract powder, behind the stirring and evenly mixing, pour in the charge chute of drop pill machine; Stir, make fluid temperature reach 95 ℃, be incubated after 30 minutes; 8 centimetres to 10 centimetres drip apart under, splash in the liquid coolant with 20 droplets/minute to 40 droplets/minute speed, treat that drop pill shrinks the condensation molding fully after; Take out, remove the liquid coolant on drop pill surface and promptly get with getting rid of oily machine.
Two of technical scheme of the present invention obtains in the following manner: a kind of production method of artemisia rupestris L pill, its raw material Herba Hyperici Monogyni and Herba Achilleae medical material by weight, ratio is 1: 1; This artemisia rupestris L pill contains 10% to 20% rupestonic acid and 20% to 40% total flavones by weight percentage; This artemisia rupestris L pill is undertaken by following step:
The first step: Herba Hyperici Monogyni and the Herba Achilleae medical material of getting equivalent mix; Be ground into coarse powder; Add 8 to 12 times of amount 60% to 80% alcohol reflux 2 to 3 times, each 1 to 2 hour, merge extractive liquid; Extracting liquid filtering, put coldly, centrifugal, supernatant concentration obtains every milliliter of concentrated solution that contains crude drug 0.20 gram to 0.25 gram;
Second step: get above-mentioned concentrated solution, last macroporous adsorbent resin carries out purification, and the macroporous resin type is one or more among HPD100, HPD400, HPD100A and the HPD400A, the Fractional Collections effluent, and effluent concentrates, drying is processed extract powder, and is subsequent use;
The 3rd step: get above-mentioned dry extract, mix by a certain percentage with Polyethylene Glycol, heating and melting drips and processes drop pill.
Be two further optimization and/or selection below to technique scheme:
Above-mentioned second step is undertaken by following step: get above-mentioned concentrated solution, and the macroporous adsorbent resin of one or more among mistake HPD100, HPD100A, HPD400 and the HPD400A, the resin path height ratio is 1: 4 to 1: 10; Adsorption rate 1.0BV to 3BV/h, elution speed 1.0BV to 3BV/h is earlier with the flow velocity eluting of 3BV to 5BV water with 3BV to 6BV/h; The flush away water-solubility impurity; With 30% ethanol elution 4BV to 6BV, 70% ethanol elution 4BV to 6BV collects 30% and 70% ethanol elution again; Decompression recycling ethanol and to be concentrated into relative density be 1.22 to 1.25 clear paste under 60 ℃; Dry down 60 ℃ of temperature in vacuum drying oven again, get 100 purpose extract powders, subsequent use; Take by weighing the medicinal substrate Polyethylene Glycol, its weight and 100 purpose extract powder ratios are counted 2: 1 to 3: 1 by weight, after complete fusion on 70 ℃ to the 100 ℃ water-baths; Add extract powder, behind the stirring and evenly mixing, pour in the charge chute of drop pill machine; Stir, make fluid temperature reach 95 ℃, be incubated after 30 minutes; 8 centimetres to 10 centimetres drip apart under, splash in the liquid coolant with 20 droplets/minute to 40 droplets/minute speed, treat that drop pill shrinks the condensation molding fully after; Take out, remove the liquid coolant on drop pill surface and promptly get with getting rid of oily machine.
One of technical scheme of the present invention obtains in the following manner: a kind of above-mentioned artemisia rupestris L pill is applied to the Chinese medicine as treatment rheumatism and rheumatoid arthritis.
The artemisia rupestris L pill of gained of the present invention not only contains 10% to 20% rupestonic acid, also contains 20% to 40% flavones ingredient.The production method of this artemisia rupestris L pill has adopted the purification with macroreticular resin technology; Rupestonic acid and total flavonoid composition in ability while enrichment Herba Achilleae and the Herba Hyperici Monogyni; This composition all is that process stabilizing is reliable for the effective composition of anti-inflammatory and antalgic, and active constituent content is stable in the gained drop pill.This production method equipment needed thereby is simple; The material safety environmental protection of using; And can reclaim through resin regeneration and ethanol and recycle; Practiced thrift production cost greatly, solved simultaneously the organic solvent residual that causes with organic solvent extractions such as petroleum ether, ethyl acetate problems such as harm again human body.The artemisia rupestris L pill of gained of the present invention proves the effect with tangible anti-inflammatory and antalgic through pharmacodynamic experiment, can be used for the treatment of rheumatism and rheumatoid arthritis.
Description of drawings
The chromatogram that accompanying drawing 1 is measured for the rupestonic acid reference substance.Accompanying drawing 2 is chromatogram before the purifying products of the present invention.
Accompanying drawing 3 is chromatogram behind the purifying products of the present invention.Accompanying drawing 4 is the total flavones UV scanning figure of product of the present invention.
Accompanying drawing 5 is the total flavones canonical plotting of product of the present invention.
The specific embodiment
The present invention does not receive the restriction of following embodiment, can confirm concrete embodiment according to the technical scheme and the practical situation of the invention described above.
Below in conjunction with embodiment the present invention is done further argumentation:
Embodiment 6, are with the difference of embodiment 1 or 2 or 3: the artemisia rupestris L pill of embodiment 6 contains 20% rupestonic acid and 40% total flavones by weight percentage.
Embodiment 7, are with the difference of embodiment 1 or 2 or 3: the artemisia rupestris L pill of embodiment 7 contains 10% to 20% rupestonic acid and 20% to 40% total flavones by weight percentage;
Experimental apparatus: SPD-10AVP type high performance liquid chromatograph (institute is made in a day island proper Tianjin)
Experiment medicine: rupestonic acid reference substance (Chinese Academy Of Sciences Xinjiang physics & chemistry Technology Research Institute, lot number: 030401); Artemisia rupestris L pill.
Experimental technique:
Chromatographic condition: adopt Shimpack ODSC
18Post (4.6 * 250mm, 5 μ m); Mobile phase: methanol-0.4% phosphoric acid (50: 50), detect wavelength X=241nm; Column temperature: 35 ℃; Flow velocity: 1.0 μ gmL
-1Sample size is 10 μ L.
The preparation of reference substance solution: take by weighing rupestonic acid reference substance 4.86mg with loss of weight method precision, put in the 25mL measuring bottle, with dissolve with methanol and be diluted to scale, shake up, promptly make rupestonic acid reference substance stock solution (194.4 μ gmL
-1).
The preparation of need testing solution: get artemisia rupestris L pill sample 20 balls in the above-mentioned enforcement row, the accurate title, decide, and porphyrize is got in 0.g to the 25mL measuring bottle, and dehydrated alcohol is diluted to scale, and supersound process 20min filters, and filtrating is as need testing solution.
Assay: draw reference substance solution and need testing solution respectively,, calculate according to external standard method according to recording peak area.Chromatogram is seen accompanying drawing 1 to 3.
The result: rupestonic acid content is all in 10% to 20% scope in the artemisia rupestris L pill sample of above-mentioned enforcement row 1 to 7 gained.
The artemisia rupestris L pill of above-mentioned enforcement row gained is carried out pharmacodynamic experiment, and its result and process are following:
Test row 1: artemisia rupestris L pill of the present invention is to the inhibitory action of adjuvant arthritis rats constitutional pathological changes
Get 60 of Wistar male rats, body weight 180 ± 20g is divided into the artemisia rupestris L pill group of distilled water (DW) matched group, model group, safe and sound group of positive control drug rheumatism, three dosage according to body weight, 10 every group at random.Earlier measure every group of right back sufficient sole of the foot thickness of rat with micrometer callipers.Each administration group is by setting dosage ig administration, every day 1 time, continuous 10 days, the DW of matched group and model group ig equivalent, after the first administration 1 hour, except that matched group, all the other each organize all at right back sufficient plantar subcutaneous injection Freund ' s Freund's complete adjuvant 0.1ml.From injecting the back the 2nd day, measure rat right back sufficient sole of the foot thickness with micrometer callipers every day, surveys altogether 10 times.Calculate the suppression ratio of each treated animal, relatively the drug effect of each administration group at different time swelling degree of the paw and medicine.
The scorching front foot sole of the foot thickness of the scorching metapedes sole of the foot thickness of swelling degree=cause-cause
As a result, right back sufficient sole of the foot injection adjuvant swelling next day of model group and administration treated animal is obvious, and increases the weight of day by day, alleviates to some extent after 3 days, and the 8th day obvious once again.In safe and sound of rheumatism and the artemisia rupestris L pill, heavy dose of group all has the obvious suppression effect to the rat constitutional inflammatory reaction that is caused by Freund ' s Freund's complete adjuvant.Each treated animal is seen table 1 and table 2 at different time swelling degree of the paw and suppression ratio.
Test row 2: artemisia rupestris L pill of the present invention is to the therapeutical effect of adjuvant arthritis rats Secondary cases pathological changes
Get 60 of Wistar male rats, body weight 180 ± 20g divides into groups to measure every group of right back sufficient sole of the foot thickness of rat with micrometer callipers earlier with 2.1.Except that matched group, all the other each groups cause scorching back and began each administration group on the 18th day by setting dosage ig administration, the DW of matched group and model group ig equivalent, continuous 12 days all at right back sufficient plantar subcutaneous injection Freund ' s Freund's complete adjuvant 0.1ml.Each group was all measured right back sufficient sole of the foot thickness to the 30 days with micrometer callipers every other day after the administration.Calculate the suppression ratio of each treated animal, relatively the drug effect of each administration group at different time swelling degree of the paw and medicine.
The scorching front foot sole of the foot thickness of the scorching metapedes sole of the foot thickness of swelling degree=cause-cause
Statistical result shows, safe and sound of rheumatism with in, heavy dose of artemisia rupestris L pill all has the obvious treatment effect to the rat secondary inflammation that is caused by Freund ' s Freund's complete adjuvant.Each treated animal is seen table 3 and table 4 at different time swelling degree of the paw and suppression ratio.
Test row 3: artemisia rupestris L pill on Carrageenan of the present invention causes the influence of rat paw edema
Get 50 of Wistar rats, male and female half and half, body weight 120-150g is divided into the artemisia rupestris L pill group of model group, safe and sound group of rheumatism, three dosage at random according to the body weight sex.Each administration group is by setting dosage ig administration, and model group gives equivalent DW, every day 1 time, continuous 5 days.Behind time administration 1h, cause inflammation for the left back sufficient plantar aponeurosis injected 1% carrageenin sterile solution 50ul of rat.Measure respectively cause scorching before and cause scorching back 1h, 2h, 4h, 6h, the left back sufficient sole of the foot thickness of 8h so that before and after scorching the difference of sufficient sole of the foot thickness as the swelling degree.Calculate and respectively organize rat paw edema degree average and suppression ratio, organize a mean t check, statistical result shows that artemisia rupestris L pill can suppress carrageenin to some extent and cause rat paw edema.The result sees table 5 and table 6.
Test row 4: artemisia rupestris L pill of the present invention is to the bullate influence of rat granuloma
Get 50 of Wistar rats, male and female half and half, body weight 200 ± 20g.Divide into groups with 2.3.With the cotton balls autoclaving of 20 ± 1mg, add ampicillin (10mg/ml) 0.1ml/, in 50 ℃ of oven dry.Under the ether light anaesthesia, the sterile working, the axillary incision in the rat both sides, it is subcutaneous that two sterilization cotton balls are implanted both sides axillary region respectively, skin suture.Postoperative each group on the same day begins by setting dosage ig administration, model group ig equivalent distilled water, every day 1 time, continuous 7 days.Drew cervical vertebra to put to death rat on the 8th day, take out cotton balls, pick the degrease tissue, warp is dried to constant weight for 60 ℃, and it is heavy to deduct former cotton balls, is granuloma heavy (the mg/100g body weight is represented).Calculate and respectively organize granuloma weight, organize a mean t check, statistical result shows, compares with model group, and bullate generation all has the obvious suppression effect to artemisia rupestris L pill to rat granuloma.The result sees table 7.
Test row 5: artemisia rupestris L pill of the present invention is to the influence of mouse peritoneal capillary permeability
Get 50 of Kunming mouses, male and female half and half, body weight 20 ± 2g; The artemisia rupestris L pill group that is divided into model group, safe and sound group of rheumatism, three dosage at random, each administration group is by setting dosage ig administration, model group ig equivalent DW; Every day 1 time, for three days on end, 1.5h respectively organizes the azovan blue normal saline solution 0.1ml/10g of mouse tail vein injection 0.5% after the last administration; And the acetic acid NS solution of ip 0.6% immediately, 0.2ml/ is only.Put to death mice behind the 20min, cut off abdominal part, with 6ml NS gradation washing abdominal cavity; The sucking-off flushing liquor merges the back and is diluted to 10ml with NS, the centrifugal 15min of 3000rpm; Get supernatant and survey the trap value in the 590nm place; Organize mean t check, each is organized OD primary system meter result and shows, in safe and sound of rheumatism and the artemisia rupestris L pill, heavy dose all can obviously suppress the mouse peritoneal capillary permeability and increase.The result sees table 8.
Test row 6: artemisia rupestris L pill Dichlorodiphenyl Acetate of the present invention causes the influence of mouse writhing reaction
Get 50 of Kunming mouses, male and female half and half, body weight 20 ± 2g divides into groups with 2.5.Each administration group is pressed and is set dosage ig administration, model group ig equivalent distilled water, and every day 1 time, for three days on end.2h after the last administration, every Mus ip 0.6% acetic acid NS solution 0.2ml/ are only.Turn round the body number of times in the observed and recorded mice 15min, organize a mean t check, and calculate suppression ratio.Statistical result shows, in safe and sound of rheumatism and the artemisia rupestris L pill, heavy dose all can obviously reduce acetic acid and cause the mouse writhing reaction times.The result sees table 9.
Test row 7: artemisia rupestris L pill of the present invention is to 2, and the 4-dinitrochlorobenzene causes the influence of Mice Auricle skin delayed hypersensitivity
Get 50 of NIH mices, male and female half and half, body weight 25 ± 1g divides into groups with 2.5.Each administration group is by setting dosage ig administration, model group ig equivalent distilled water, every day 1 time, continuous 9 days.After administration, respectively organized in the 10th day mice left side ear smear 0.25% 2,4-dinitrochlorobenzene acetone solution 0.02ml/ only, the offside auricle is smeared the acetone solution of same volume and is done contrast; Put to death mice behind the 24h, cut the Mus ear, downcut the equal area auricle in left and right ear with card punch from the basal part of the ear; Weigh respectively, the difference of calculating left and right sides auricle weight is organized a mean t check as the swelling degree; And calculating suppression ratio; Statistical result shows, in safe and sound of rheumatism and the artemisia rupestris L pill, heavy dose all can obviously suppress 2,4-dinitrochlorobenzene to Mice Auricle delayed hypersensitivity.The result sees table 10.
Test row 8: the influence that artemisia rupestris L pill of the present invention generates the mice serum hemolysin
The preparation of sheep red blood cell (SRBC) (SRBC) and GPS: by the aseptic blood 50ml that gets of healthy sheep jugular vein, put in the sterilization conical flask that fills bead, shake 10min and defibrinate to remove; Add the A Shi liquid that is equivalent to Sanguis caprae seu ovis volume twice; It is subsequent use to shake up rearmounted 4 ℃ of preservations, face with preceding with aseptic NS wash 3 times (1500rpm, 5min); Abandon supernatant, do 3: 5 (V/V) dilution with NS again.Getting blood from healthy guinea pig carotid artery prepares serum in addition, faces the time spent and does dilution in 1: 10 with aseptic NS.
Get 50 of NIH mices, male and female half and half, body weight 20 ± 2g divides into groups with 2.5.Each administration group is by setting dosage ig administration, matched group ig equivalent distilled water, every day 1 time, continuous 7 days.Only after the last administration, respectively organized the equal ip 10%SRBC of mice suspension 0.2ml/ in 2 hours, the immunity back was got blood on the 6th day, place centrifugal behind the 1h (2000rpm 10min), gets serum, with according to the form below application of sample after 500 times of the NS dilutions:
Each is managed behind the application of sample in 37 ℃ of water bath heat preservation 10min, ice bath cessation reaction, centrifugal (2000rpm; 5min), get supernatant 1ml, add Dou Shi liquid 3ml; Other gets 10%SRBC suspension 0.25ml and 4ml Dou Shi liquid mixing, surveys absorption value behind the 10min as SRBC HD50 absorption value, and each sample cell is blank with the control tube; Measure absorption value, calculation sample HC in 540nm
50Statistical result shows, in safe and sound of rheumatism and the artemisia rupestris L pill, heavy dose of generation that all can obviously suppress the mice serum hemolysin.The result sees table 11.
HC
50=absorption of sample degree average/SRBC HD50 absorption value * serum diluting multiple
Test row 9: artemisia rupestris L pill of the present invention is to the lymphopoietic influence of T
Conventional preparation splenocyte suspension: get the NIH mice and pluck the eyeball sacrificed by exsanguination, the aseptic spleen of getting is rejected fat and connective tissue; Place on the 100 order nylon cloths to shred, contain on the culture dish of the cold HBSS liquid of 10ml in nylon is arranged, shred; Use homogenizer homogenate, collecting cell liquid, centrifugal (1000rpm; 10min), abandon supernatant, through hypotonic (0.8% NH
4Cl handles) remove erythrocyte, do the cell counting, the adjustment cell concentration is 1 * 10
6/ ml, above process is all carried out at 0～4 ℃.Sample and ConA are made into desired concn with RPMI-1640 liquid, in 96 orifice plate according to the form below application of samples:
Culture plate is put 37 ℃, 5%CO
2Take out after cultivating 72h in the incubator, cultivate 4～6h again after adding MTT (10ul/ hole) mixing, inhale and abandon supernatant, add DMSO (100ul/ hole), the Shi Jia Za product that vibrates slightly fully dissolves, and measures each hole OD with ELIASA
570nmValue.Statistical result shows that ConA cellular control unit propagation degree is apparently higher than the blank group; Compare with the ConA group, artemisia rupestris L pill can obviously promote the lymphocytic propagation of T in concentration is 1000～2000ug/ml scope.The result sees table 12.
Test row 10: artemisia rupestris L pill of the present invention is to the influence of bone-marrow-derived lymphocyte propagation
Conventional preparation splenocyte suspension method is the same.Sample and LPS are made into desired concn with RPMI-1640 liquid, in 96 orifice plate according to the form below application of samples:
Culture plate is put 37 ℃, 5%CO
2Take out after cultivating 72h in the incubator, cultivate 4～6h again after adding MTT (10ul/ hole) mixing, inhale and abandon supernatant, add DMSO (100ul/ hole), the Shi Jia Za product that vibrates slightly fully dissolves, and measures each hole OD with ELIASA
570nmValue.Experimental result shows that LPS group cell proliferation degree is apparently higher than the blank group; Compare with the LPS group, artemisia rupestris L pill can suppress the propagation of bone-marrow-derived lymphocyte in concentration is 250～2000ug/ml scope.The result sees table 13.
Artemisia rupestris L pill causes to rat assist agent arthritis, carrageenin that rat paw edema and rat granuloma are swollen all to have a good inhibitory effect; Can reduce the mouse peritoneal capillary permeability, reduce acetic acid and cause the mouse writhing reaction times, and suppress Mice Auricle skin delayed hypersensitivity; Can promote simultaneously the mice serum hemolysin to generate, in concentration is 1000～2000ug/ml scope, can promote the lymphocytic propagation of T, in concentration is 250～2000ug/ml scope, suppress the propagation of bone-marrow-derived lymphocyte.The prompting artemisia rupestris L pill has good antiinflammatory, analgesia and immunoregulation effect, for it is used to treat rheumatism and rheumatoid arthritis provides the pharmacology foundation.
Claims (3)
1. an artemisia rupestris L pill is characterized in that raw material Herba Hyperici Monogyni and Herba Achilleae medical material by weight, and ratio is 1:1; This artemisia rupestris L pill contains 10% to 20% rupestonic acid and 20% to 40% total flavones by weight percentage; This artemisia rupestris L pill obtains by following step:
The first step: Herba Hyperici Monogyni and the Herba Achilleae medical material of getting equivalent mix; Be ground into coarse powder; Add 8 to 12 times of amount 60% to 80% alcohol reflux 2 to 3 times, each 1 to 2 hour, merge extractive liquid; Extracting liquid filtering, put coldly, centrifugal, supernatant concentration obtains every milliliter of concentrated solution that contains crude drug 0.20 gram to 0.25 gram;
Second step: get above-mentioned concentrated solution, last macroporous adsorbent resin carries out purification, and the macroporous resin type is more than one among HPD100, HPD400, HPD100A and the HPD400A, the Fractional Collections effluent, and effluent concentrates, drying is processed extract powder, and is subsequent use;
The 3rd step: get above-mentioned dry extract, mix by a certain percentage with Polyethylene Glycol, heating and melting drips and processes drop pill;
Wherein, above-mentioned second step is undertaken by following step: get above-mentioned concentrated solution, excessively the macroporous adsorbent resin of more than one among HPD100, HPD400, HPD100A and the HPD400A; The resin path height ratio is 1:4 to 1:10, adsorption rate 1.0BV to 3BV/h, elution speed 1.0BV to 3BV/h; With the flow velocity eluting of 3BV to 5BV water with 3BV to 6BV/h, the flush away water-solubility impurity is again with 30% ethanol elution 4BV to 6BV earlier; 70% ethanol elution 4BV to 6BV collects 30% and 70% ethanol elution, decompression recycling ethanol and to be concentrated into relative density be 1.22 to 1.25 clear paste under 60 ℃; Dry down 60 ℃ of temperature in vacuum drying oven again, get 100 purpose extract powders, subsequent use; Take by weighing the medicinal substrate Polyethylene Glycol, its weight and 100 purpose extract powder ratios are counted 2:1 to 3:1 by weight, after complete fusion on 70 ℃ to the 100 ℃ water-baths; Add extract powder, behind the stirring and evenly mixing, pour in the charge chute of drop pill machine; Stir, make fluid temperature reach 95 ℃, be incubated after 30 minutes; 8 centimetres to 10 centimetres drip apart under, splash in the liquid coolant with 20 droplets/minute to 40 droplets/minute speed, treat that drop pill shrinks the condensation molding fully after; Take out, remove the liquid coolant on drop pill surface and promptly get with getting rid of oily machine.
2. the production method of an artemisia rupestris L pill is characterized in that raw material Herba Hyperici Monogyni and Herba Achilleae medical material by weight, and ratio is 1:1; This artemisia rupestris L pill contains 10% to 20% rupestonic acid and 20% to 40% total flavones by weight percentage; This artemisia rupestris L pill is undertaken by following step:
The first step: Herba Hyperici Monogyni and the Herba Achilleae medical material of getting equivalent mix; Be ground into coarse powder; Add 8 to 12 times of amount 60% to 80% alcohol reflux 2 to 3 times, each 1 to 2 hour, merge extractive liquid; Extracting liquid filtering, put coldly, centrifugal, supernatant concentration obtains every milliliter of concentrated solution that contains crude drug 0.20 gram to 0.25 gram;
Second step: get above-mentioned concentrated solution, last macroporous adsorbent resin carries out purification, and the macroporous resin type is more than one among HPD100, HPD400, HPD100A and the HPD400A, the Fractional Collections effluent, and effluent concentrates, drying is processed extract powder, and is subsequent use;
The 3rd step: get above-mentioned dry extract, mix by a certain percentage with Polyethylene Glycol, heating and melting drips and processes drop pill;
Wherein, above-mentioned second step is undertaken by following step: get above-mentioned concentrated solution, excessively the macroporous adsorbent resin of more than one among HPD100, HPD400, HPD100A and the HPD400A; The resin path height ratio is 1:4 to 1:10, adsorption rate 1.0BV to 3BV/h, elution speed 1.0BV to 3BV/h; With the flow velocity eluting of 3BV to 5BV water with 3BV to 6BV/h, the flush away water-solubility impurity is again with 30% ethanol elution 4BV to 6BV earlier; 70% ethanol elution 4BV to 6BV collects 30% and 70% ethanol elution, decompression recycling ethanol and to be concentrated into relative density be 1.22 to 1.25 clear paste under 60 ℃; Dry down 60 ℃ of temperature in vacuum drying oven again, get 100 purpose extract powders, subsequent use; Take by weighing the medicinal substrate Polyethylene Glycol, its weight and 100 purpose extract powder ratios are counted 2:1 to 3:1 by weight, after complete fusion on 70 ℃ to the 100 ℃ water-baths; Add extract powder, behind the stirring and evenly mixing, pour in the charge chute of drop pill machine; Stir, make fluid temperature reach 95 ℃, be incubated after 30 minutes; 8 centimetres to 10 centimetres drip apart under, splash in the liquid coolant with 20 droplets/minute to 40 droplets/minute speed, treat that drop pill shrinks the condensation molding fully after; Take out, remove the liquid coolant on drop pill surface and promptly get with getting rid of oily machine.
3. an artemisia rupestris L pill according to claim 1 is applied to prepare the Chinese medicine as treatment rheumatism and rheumatoid arthritis.
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| CN103127196A (en) * | 2013-03-18 | 2013-06-05 | 中国科学院新疆理化技术研究所 | Preparation method and application of total extractive of artemisia rupestris L |
| CN107280005A (en) * | 2017-06-20 | 2017-10-24 | 安徽阳光药业有限公司 | Sustained-release dropping pill containing Cordyceps militaris and okra and preparation method thereof |
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