CN1931272A - Quality control method for gynecopathy treating medicine capsule - Google Patents
Quality control method for gynecopathy treating medicine capsule Download PDFInfo
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- CN1931272A CN1931272A CN 200510086429 CN200510086429A CN1931272A CN 1931272 A CN1931272 A CN 1931272A CN 200510086429 CN200510086429 CN 200510086429 CN 200510086429 A CN200510086429 A CN 200510086429A CN 1931272 A CN1931272 A CN 1931272A
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Abstract
The present invention is the quality control method for gynecopathy treating medicine, and the method includes identifying angelica, Dangshen and berberine hydrochloride and determining the content of andrographis herb. The quality control method can identify the medicine preparation simply and intuitively and ensure the curative effect of the medicine.
Description
Technical field
The present invention relates to a kind of method of quality control for the treatment of the medicine for gynecopathy capsule, be specifically related to a kind of discrimination method and content assaying method for the treatment of the medicine for gynecopathy capsule.
Background technology
As the good medicine for the treatment of gynaecopathia such as morbid leukorrhea, stomachache, capsule of the present invention is applied for multinomial product and process patent and is dropped into commercial production for many years by the applicant, is extended to market, determined curative effect.
Along with the development of traditional Chinese medicine technology, improve drug effect and enhance productivity, improve the controllability of drug quality, reducing cost is a great problem that the applicant will solve.
Through long-term theory research and industrial practice, the applicant has found method of quality control such as best preparation consumption and preparation scheme and discriminating, assay.
Summary of the invention
The purpose of this invention is to provide a kind of method of quality control for the treatment of the medicine for gynecopathy capsule, comprising discrimination method and content assaying method.
Treatment medicine for gynecopathy capsule of the present invention is prepared by following method:
Take by weighing following raw materials according: 16 parts of Radix Flemingiae Philippinensiss, 16 parts of Radix Rosae Laevigataes, 9 parts of Herba Andrographis, 16 parts of Caulis Mahoniaes, 9 parts of Fructus Zanthoxyli Dissiti, 9 parts of Radix Angelicae Sinensis, 16 parts of Caulis Spatholobis, 9 parts of Radix Codonopsis.
A, Herba Andrographis is broken into coarse powder, extracts with ethanol refluxing process and make clear paste, medicinal residues are standby;
B, Radix Angelicae Sinensis is broken into coarse powder, makes clear paste according to the percolation under fluid extract and the extractum item, medicinal residues are standby;
C, Caulis Mahoniae and Fructus Zanthoxyli Dissiti two flavor medical material decocting in water are extracted twice, after filtering the merging of both filtrates is made clear paste;
D, with Radix Flemingiae Philippinensis, Radix Rosae Laevigatae, Caulis Spatholobi, Radix Codonopsis four flavors, after-filtration of extracting in water with the Radix Angelicae Sinensis medicinal residues of technology A and the Herba Andrographis medicinal residues decocting in water of technology B, filters the medicinal residues after filtering, merging filtrate is also made clear paste;
E, the above-mentioned 4 kinds of clear paste of merging through mixing, are granulated, drying, and granulate incapsulates, and makes capsule, promptly.
Method of quality control of the present invention comprises discrimination method and assay two parts at the above-mentioned capsule that makes, and wherein discrimination method comprises one or more in the following discriminating:
The discriminating of Radix Angelicae Sinensis: get described capsule 's content 1-3g, add ethanol 10-30ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 1-3ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 0.5-1.5g, shines medical material solution in pairs with legal system; Test according to thin layer chromatography, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (7-11: 0.5-1.5) be developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color;
The discriminating of Radix Codonopsis: get described capsule 's content 0.5-1.5g, add methanol 10-30ml, supersound process 30 minutes filters, filtrate is put evaporate to dryness in the water-bath, residue adds the about 1-3ml of methanol makes dissolving, is added on the neutral alumina post of having handled well, with 10-100% methanol 50-150ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 1-3ml makes dissolving, as need testing solution; Other gets Radix Codonopsis control medicinal material 0.5-1.5g, shines medical material solution in pairs with legal system; Test according to thin layer chromatography, draw above-mentioned two kinds of each 2ul of solution, put respectively on same silica gel g thin-layer plate, with n-butyl alcohol-butanols-water (10-20: 1-5: 1-3) be developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color;
The discriminating of berberine hydrochloride: get described capsule 's content 0.5-3g, add hydrochloric acid-alcohol mixed solution (0.5-1.5: 80-120) 10-30ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 1-3ml makes dissolving, as need testing solution; Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution; Test according to thin layer chromatography, draw above-mentioned reference substance solution 2ul, need testing solution 4ul, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-methanol-isopropyl alcohol-water (4-8: 1-5: 0.5-2.5: 0.5-2.5: be developing solvent 0.1-0.5), put in the chromatography cylinder of ammonia saturated with vapor, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
Its preferred discrimination method comprises one or more in the following discriminating:
A, get described capsule 's content 2g, add ethanol 20ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 1g, shines medical material solution in pairs with legal system.According to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B) test, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (9: 1) is developing solvent, launches, and takes out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color.
B, get described capsule 's content 1g, add methanol 20ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds the about 2ml of methanol makes dissolving, be added on neutral alumina post (100～200 orders of having handled well, 5g is on the internal diameter 10～15mm), with 40% methanol 100ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Codonopsis control medicinal material 1g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned two kinds of each 2ul of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with n-butyl alcohol-butanols-water (15: 3: 2), launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color.
C, get described capsule 's content 1g, add hydrochloric acid-alcohol mixed solution (1-100) 20ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned reference substance solution 2ul, need testing solution 4ul, putting respectively on same silica gel g thin-layer plate, is developing solvent with benzene-ethyl acetate-methanol-isopropyl alcohol-water (6: 3: 1.5: 1.5: 0.3), puts in the chromatography cylinder of ammonia saturated with vapor, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
Method of quality control of the present invention also comprises following assay:
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get described capsule 's content 0.1-1g, accurately claim surely, put in the tool plug conical flask, precision adds methanol 15-35ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (44-64: 36-56: 0.1-2) be mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, described capsule 's content contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
Its preferred content assay method is as follows:
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get described capsule 's content 0.5g, accurately claim surely, put in the tool plug conical flask, precision adds methanol 25ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version D) are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (54: 46: 1) is a mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, described capsule 's content contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
Discrimination method in the method for quality control of the present invention is done contrast with Radix Angelicae Sinensis, Radix Codonopsis control medicinal material and berberine hydrochloride reference substance, directly compares spot size and position, makes operation easier, and is directly perceived; Content assaying method detects with the andrographolide reference substance, makes quality more secure, and drug effect is more definite.
The capsule of the present invention that utilizes method of quality control monitoring of the present invention to make, drug effect is definite, and effect is stable.Below utilize experimental example to specify.
Experiment condition:
Animal: cleaning level Kunming mouse, BALB/C mice, SD rat.
Medicine: capsule 0.4g/ grain of the present invention, every g capsule 's content contains crude drug 8.33g; Limited company provides by zhuzhou,hunan a thousand pieces of gold Pharmaceutical;
Method for preparation of drug and laboratory animal grouping: laboratory animal is divided into 4 groups.First group is matched group, gives the equal-volume distilled water; Second and third, four groups be respectively the high, medium and low dosage group of capsule of the present invention.
Capsule of the present invention is got content, respectively with distilled water be made into 14.56,29.12, the suspension solution of 58.24g crude drug/100ml, use for mice; Be made into 12.6,25.2 respectively, the suspension solution of 50.4g crude drug/100ml, use for rat.Capsule mice dosage of the present invention is respectively 3.64,7.28 and 14.56g crude drug/kg, and rat dosage is respectively 2.52,5.04 and 10.08g crude drug/kg (press the body surface area conversion, be equivalent to the equivalent, 2 times and 4 times of people's one consumption per day respectively).Unless otherwise indicated, more than all every day gastric infusion once, laboratory temperature 20-25 ℃.
Experimental example 1: antiinflammatory action
1, to the influence of Oleum Tiglii induced mice ear swelling
60 of mices, male, body weight 24-26g is divided into 4 groups (n=10) at random, administration as stated above, totally seven days once a day.After the last administration 1 hour, every Mus auris dextra is coated with 2% Oleum Tiglii (2% Oleum Tiglii, 20% dehydrated alcohol, 5% distilled water and 73% ether) 0.1ml, and left ear is made blank.Be administered once again after 2 hours, be coated with Oleum Tiglii after 4 hours cervical vertebra dislocation put to death mice, cut two ears along the auricle baseline, be that the card punch of 8mm is laid auricle and weighed with diameter, as every group of mice swelling degree, the results are shown in Table 1 with two ear weight differences.
Table 1, to the influence of Oleum Tiglii induced mice ear swelling (X ± SD, n=10)
| Group | Dosage (g/kg) | Auris dextra heavy (mg) | Left side ear heavy (mg) | Swelling degree (mg) | Suppression ratio (mg) |
| Model control group | Equal-volume water | 31.3±3.7 | 9.7±4.0 | 21.6±4.1 | |
| The present invention | 3.64 | 26.6±3.9 | 10.4±4.3 | 16.2±4.6 ** | 25.0 |
| The present invention | 7.28 | 26.2±3.4 | 10.1±3.7 | 16.1±3.9 *** | 25.5 |
| The present invention | 14.56 | 26.2±4.1 | 10.3±4.2 | 15.9±4.4 *** | 26.4 |
Compare with model control group
*P＞0.05,
*P＜0.05,
* *P＜0.01
The result shows that each administration group compares with model control group respectively, all can obviously suppress Oleum Tiglii induced mice ear swelling.
2, to the influence of rat paw edema due to the chondrus ocellatus Holmes
60 of SD rats, male, body weight 180-200g is divided into 4 groups (n=10), administration as stated above, once a day, and totally seven days, 30min after the last administration, injection 1% carrageenin 0.1ml/ only is administered once after 2 hours again under rat left hind foot aponeurosis (aponeuroses).Measurement cause scorching before and after the rat paw size, per hour survey 1 time (is that the tape measure of mm is measured sufficient pawl to the ankle girth with scale), survey altogether 6 times after giving chondrus ocellatus Holmes, calculating swelling degree (cause scorching metapedes sole of the foot girth-cause scorching front foot sole of the foot girth) the results are shown in Table 2.
Table 2, to the influence of rat paw edema due to the chondrus ocellatus Holmes (X ± SD, n=10)
| Group | Dosage (g/Kg) | Experiment front foot size (cm) | Cause scorching back swelling degree (cm) | |||||
| 1 hour | 2 hours | 3 hours | 4 hours | 5 hours | 6 hours | |||
| Matched group | Equal-volume water | 2.23± 0.09 | 0.52± 0.09 | 0.53± 0.10 | 0.88± 0.14 | 0.90± 0.12 | 0.70± 0.17 | 0.67±0.13 |
| The present invention | 2.52 | 2.27± 0.06 | 0.36± 0.09 *** | 0.36± 0.08 *** | 0.59± 0.13 *** | 0.56± 0.12 *** | 0.52± 0.09 *** | 0.51± 0.09 *** |
| The present invention | 5.04 | 2.21± 0.08 | 0.34± 0.14 *** | 0.36± 0.07 *** | 0.58± 0.20 *** | 0.55± 0.22 *** | 0.49± 0.11 *** | 0.45± 0.12 *** |
| The present invention | 10.08 | 2.29± 0.08 | 0.27± 0.13 *** | 0.30± 0.07 *** | 0.49± 0.18 *** | 0.50± 0.14 *** | 0.46± 0.14 *** | 0.43± 0.13 *** |
Compare with model control group
*P＞0.05
*P＜0.05
* *P＜0.01
The result shows, each administration group respectively with model control group relatively, all can obviously suppress rat paw edema due to the chondrus ocellatus Holmes,
3, to the influence of rat uterus inflammation
60 of SD rats, female, body weight 180-200g is divided into 4 groups (n=10), with 25% urethane ip 0.4ml/100g anesthetized rat, then rat is lain on the back in operating-table, cut off the hypogastric region hair, with 75% alcohol disinfecting skin, disinfecting action, long in hypogastric region median incision 2cm, expose the uterus, make a transverse incision along 1cm place on the cornua uteri of right side, with a plastic tube (caliber 2mm, long 0.5cm, heavy 2mg, alcohol disinfecting) be positioned over intrauterine, with the uterine incision sutured, with slip-off preventing, wound splashes into 0.2mg penicillin+streptomycin (being dissolved in the 0.2ml water for injection), the anti-infection, postoperative treated that rat began by 2.1 method administrations after clear-headed in 6 hours, once a day, totally seven days, after the last administration 24 hours, rat is put to death in the cervical vertebra dislocation, take out the uterus, both sides, heavily be inflammation swelling degree, calculate swelling rate and suppression ratio (not causing scorching uterus weight=swelling rate) with swelling degree ÷ with uterus, right side Utero-left side; ([the average swelling rate of the model control group-average swelling rate of administration group] average swelling rate of ÷ model control group is suppression ratio) the results are shown in Table 3.
Table 3, FUKE QIANJIN JIAONANG to the influence of rat uterus inflammation (X ± SD, n=10)
| Group | Dosage (g/kg) | The right side, uterus is heavy | The left side, uterus is heavy | Swelling rate (%) | Suppression ratio (%) |
| Matched group | Equal-volume water | 390.1±21.2 | 301.1±17.8 | 29.6±6.2 | |
| The present invention | 2.52 | 373.3±20.9 | 309.0±21.0 | 20.8±3.8 *** | 29.7 |
| The present invention | 5.04 | 355.5±24.2 | 298.0±27.6 | 19.3±3.2 *** | 34.8 |
| The present invention | 10.08 | 359.8±19.8 | 307.0±24.4 | 17.2±2.7 *** | 41.9 |
Compare with model control group
* *P＜0.01.
The result shows that each administration group compares with model control group respectively, and inflammation of uterus is all had the obvious suppression effect,
4, to the bullate influence of rat granuloma
50 of SD rats, male, anaesthetize with 25% urethane lumbar injection 0.4ml/100g, then position, abdominal cavity skin is cut an osculum, with two sterilization cotton balls (each heavy 50 ± 1mg, autoclavings, each adds ampicillin 1mg/0.1ml/, 50 ℃ of dry for standby) it is subcutaneous to implant rat both sides axillary fossa respectively, treats that animal is divided into 3 groups (n=10) after clear-headed, and postoperative began by 2.1 method administrations the same day.Once a day, totally seven days.Cervical vertebra dislocation in the 8th day is put to death, and takes out cotton balls, and 60 ℃ were dried by the fire 12 hours, weighed, and calculated the granuloma degree, represented with mg granuloma/100g body weight, the results are shown in Table 4.
Table 4, FUKE QIANJIN JIAONANG to the bullate influence of rat granuloma (X ± SDmg, n=10)
| Group | Dosage (g/kg) | Granulation heavy (mg/100g body weight) | Suppression ratio (%) |
| Matched group | Equal-volume water | 130.7±28.7 | |
| Capsule of the present invention | 5.04 | 96.9±23.8 ** | 25.9 |
| Capsule of the present invention | 10.08 | 95.7±25.6 *** | 35.0 |
Compare with model control group
*P＜0.05
* *P＜0.01
The result shows that each administration group compares with model control group respectively, and rat granuloma is swollen all has the obvious suppression effect.
Experimental example 2: influence 60 of mices to what lumbar injection acetic acid induced mice abdominal cavity capillary permeability increased, the male and female dual-purpose, body weight 20-22g, be divided into 4 groups (n=10), administration as stated above, once a day, totally seven days, after the last administration 1 hour, tail vein injection 0.5% azovan blue normal saline solution 0.1ml/10g, lumbar injection 0.6% acetic acid 0.2ml/ is only simultaneously, put to death mice behind the 20min, cut off the muscle of skin of abdomen, wash the abdominal cavity at twice, collect cleaning mixture with the 2ml normal saline, it is centrifugal in 3000rpm to merge the back, get supernatant, on the TU-1221 ultraviolet spectrophotometer, survey trap, the results are shown in Table 5 in the 590nm place.
The influence that table 5, Dichlorodiphenyl Acetate induced mice abdominal cavity capillary permeability increase (X ± SD, n=10)
| Group | Dosage (g/kg) | Trap (* 100) | Suppression ratio (%) |
| Matched group | Equal-volume water | 35.9±7.6 | |
| Capsule of the present invention | 3.64 | 24.5±5.1 ** | 31.8 |
| Capsule of the present invention | 7.28 | 24.0±5.9 *** | 33.1 |
| Capsule of the present invention | 14.56 | 23.8±4.7 *** | 33.7 |
Compare with matched group
* *P＜0.01
The result shows that each administration group compares with matched group respectively, all can obviously suppress the effect that acetic acid induced mice abdominal cavity capillary permeability increases.
Experimental example 3: to the influence of the type blood deficiency mice that loses blood
60 of mices, male and female dual-purpose, body weight 20-22g, be divided into 5 groups (n=10), at first get the normal value that blood is surveyed mice RBC and Hb, then except that the normal control group, every mice of other group is from eye socket venous plexus blood-letting 0.5ml, get blood after 24 hours again and survey the value of mice RBC and Hb, then by 2.1 method administrations, once a day, totally seven days, after the last administration 24 hours, the mouse orbit venous plexus was got blood and is surveyed RBC and Hb value, the results are shown in Table 6.
Table 6, FUKE QIANJIN JIAONANG to the influence of the influence of the type blood deficiency mice that loses blood (X ± SD, n=10)
| Group | Dosage (g/kg) | RBC (* 1012/L) | Hb(g/L) | ||||
| Before losing blood | After losing blood | After the treatment | Before losing blood | After losing blood | After the treatment | ||
| The normal control group | Equal-volume water | 6.56± 0.59 * | 6.38± 0.81 * | 6.42± 0.47 *** | 124.7± 7.1 * | 120.1± 8.1 | 125.0± 10.7 *** |
| Model control group | Equal-volume water | 6.47± 0.38 | 3.45± 0.54 | 4.79±0.62 | 121.3± 9.6 | 81.5± 6.4 | 96.9±7.9 |
| Capsule of the present invention | 3.64 | 6.35± 0.87 * | 3.65± 0.57# | 5.40± 0.59 ** | 118.6± 12.1 * | 77.5± 7.7# | 109.1± 7.8 *** |
| Capsule of the present invention | 7.28 | 6.63± 0.75 * | 3.75± 0.57# | 5.51± 0.59 ** | 122.9± 10.5 * | 79.6± 9.9# | 110.9± 10.4 *** |
| Capsule of the present invention | 14.56 | 6.48± 0.77 * | 3.71± 0.60# | 5.72± 0.54 ** | 119.6±15. 2 * | 78.2±8.2 # | 113.4±7.3 *** |
Compare with model control group
*P＞0.05,
*P＜0.05,
* *P＜0.01
Compare #P＜0.01 with the normal control group
The result shows that model group and normal control group compare, and mice RBC and Hb level obviously descend after the blood-letting, through giving capsule treatment of the present invention seven days, each administration group compares with model control group respectively, and its RBC and Hb value all are significantly higher than model group, and prompting this product has blood tonification effect.
Experimental example 4: to effect of immunologic function
1, the influence that caused by cyclophosphamide immunologic hypofunction mice serum hemolytic antibody is formed
Get 60 of BALB/C mice, body weight 20-22g, male and female half and half are divided into 4 groups of (n=10) administrations as stated above, and in administration the 4th day, every Mus ip 5% chicken erythrocyte suspension 0.2ml/ only carried out immunity, continued administration.Except that the normal control group, all the other each treated animals are ip cyclophosphamide 20mg/Kg/ days respectively in the 6th, seven, eight day of administration, administration is three days again, after the last administration 1 hour, get blood from the mouse orbit venous plexus, separation of serum, the method for pressing document on the TU-1221 ultraviolet spectrophotometer in the 540nm place, survey hemolytic antibody in its serum, the results are shown in Table 7.
Table 7, the influence that immunologic hypofunction mice hemolytic antibody due to the Cy is formed
(X±SD，n＝10)
| Group | Dosage (g/kg) | Optical density (* 100) | Suppression ratio (%) |
| The normal control group | Equal-volume water | 35.2±10.1 *** | |
| Model control group | Equal-volume water | 20.3±5.7 | |
| Capsule of the present invention | 7.28 | 33.3±10.0 *** | 64.0 |
| Capsule of the present invention | 14.56 | 34.7±8.9 *** | 70.9 |
Compare with model control group
* *P＜0.01
The result shows, cyclophosphamide model group and normal control group relatively, cyclophosphamide can obviously suppress the generation of mice hemolytic antibody, each administration group respectively with model group relatively, all can obviously resist the effect that immune function of mice that cyclophosphamide causes suppresses.
2, to the influence of immunologic hypofunction macrophage phagocytosis of mice due to the cyclophosphamide (Cy)
Get 60 of BALB/C mice, body weight 20-22g, male and female half and half are divided into 4 groups (n=10) by 2.1 method administrations, except that the normal control group, all the other each treated animals in the 7th, eight, Ninth Heaven ip cyclophosphamide 20mg/Kg respectively, continue administration three days.After the last administration 1 hour, every ip in mice 5% chicken red blood cell 0.5ml, put to death mice after 4 hours, wash the abdominal cavity with the 1ml normal saline, collecting cell, smear, dyeing, every chicken red blood cell number that calculates 200 phagocyte and engulf calculates phagocytic percentage and phagocytic index under the oil mirror, the results are shown in Table 8.
Table 8, to the influence of the macrophage phagocytosis of mice of immunologic hypofunction due to the Cy
(X±SD)
| Group | Dosage (g/kg) | Phagocytic percentage | Raising rate (%) | Phagocytic index (a) | Raising rate (%) |
| The normal control group | Equal-volume water | 36.6±4.3 *** | 0.55±0.06 *** | ||
| Model control group | Equal-volume water | 15.1±2.1 | 0.23±0.03 | ||
| The present invention | 7.28 | 21.6±2.5 *** | 43.0 | 0.32±0.07 *** | 39.1 |
| The present invention | 14.56 | 22.9±2.3 *** | 51.7 | 0.34±0.03 *** | 47.8 |
Compare with model control group
* *P＜0.01 (n=10)
The result shows, model group and normal control group are relatively, the cyclophosphamide group can obviously suppress the phagocytic function of mouse macrophage, and each administration group compares with model group respectively, all can obviously improve phagocytic percentage and the phagocytic index of caused by cyclophosphamide immunologic hypofunction mice.
The protective effect of coli-infection in 5 pairs of mice bodies of experimental example
Get 140 of Kunming mouses, the male and female dual-purpose, body weight 20-22g, be divided into 8 groups (n=10), first group is normal control group (not giving escherichia coli), second group is escherichia coli matched group (not giving medicine), and two groups all gavage distilled water, and the 3rd to the 8th group gives capsule of the present invention, all ig administrations, after 2 hours, every ip in mice 0.5ml escherichia coli cultured solution of broth (amount of bacteria is 109CFU/ml, contains 10% gastric Mucin powder), behind the mouse infection antibacterial the 8th, 16 and 24 hour, respectively be administered once again, observed seven days, record animal dead number.The results are shown in Table 10.
Table 10, to the protective effect of coli-infection mice (X ± SD, n=10)
| Group | Dosage (g/Kg) | Number of animals (only) | Survival number (only) | Survival rate (%) |
| The normal control group | Equal-volume water | 10 | 10 | 100 |
| Model control group | Equal-volume water | 10 | 0 | 0 |
| Capsule of the present invention | 3.64 | 10 | 0 | 0 |
| Capsule of the present invention | 7.28 | 10 | 1 | 10 |
| Capsule of the present invention | 14.56 | 10 | 4 | 40 |
| Capsule of the present invention | 29.12 | 10 | 4 | 40 |
| Capsule of the present invention | 58.24 | 10 | 6 | 60 |
| Capsule of the present invention | 116.48 | 10 | 7 | 70 |
The result shows that capsule of the present invention has the certain protection effect to the coli-infection mice.
Experimental example 5 content assaying method precision tests of the present invention
Get the reference substance solution of same concentration, " assay " method of pressing among the embodiment 3 is measured 5 times, and record peak area integrated value the results are shown in Table 11.
Table 11, Precision test result
| Peak area | 12366 | 12196 | 12234 | 12041 | 11996 |
| RSD(％) | 1.23 | ||||
The result shows: content assaying method precision of the present invention is good.
Experimental example 6 content assaying method stability tests of the present invention
Press " assay " method mensuration among the embodiment 3, same duplicate samples solution was measured at 0,1,2,3,4,5 hour respectively, the results are shown in Table 12.
Table 12, stability test result
| Time (hour) | 0 | 1 | 2 | 3 | 4 | 5 |
| Peak area | 12486 | 12475 | 12439 | 12448 | 12415 | 12657 |
| RSD(％) | 0.69 |
The result shows, and is basicly stable in 5 hours.
Experimental example 7 content assaying method replica tests of the present invention
Get 5 parts in the sample (n=2) of same lot number, press " assay " method mensuration among the embodiment 3, the results are shown in Table 13.
Table 13, repeated experiment result
| Content (mg/g) | 2.123 | 2.137 | 2.200 | 2.184 | 2.162 |
| RSD(％) | 1.47 | ||||
The result shows: content assaying method good reproducibility of the present invention.
Experimental example 8 content assaying method average recovery tests of the present invention
It is 5 parts in 2.146mg/g sample that precision takes by weighing known content, and the accurate respectively dehydrorographolide reference substance 0.732mg that adds presses " assay " method mensuration among the embodiment 3, the results are shown in Table 14.
Table 14, determination of recovery rates result
| Sampling amount (g) | Sample size (mg) | The amount (mg) of adding | Measure content (mg) | The response rate (%) | Average recovery rate (%) | RSD(％) |
| 0.2478 | 0.5352 | 0.732 | 1.2621 | 99.30 | ||
| 0.2802 | 0.6052 | 0.732 | 1.3364 | 99.89 | ||
| 0.2469 | 0.5333 | 0.732 | 1.2516 | 98.12 | 98.94 | 0.73 |
| 0.2345 | 0.5065 | 0.732 | 1.2263 | 98.33 |
| 0.3196 | 0.6903 | 0.732 | 1.4156 | 99.08 |
The result shows that this method response rate is better.
The specific embodiment
Following examples are used to illustrate the present invention, but are not used for limiting the scope of the invention.
The preparation method of embodiment 1 medicine capsule of the present invention
Take by weighing following raw materials according: Radix Flemingiae Philippinensis 16g, Radix Rosae Laevigatae 16g, Herba Andrographis 9g, Caulis Mahoniae 16g, Fructus Zanthoxyli Dissiti 9g, Radix Angelicae Sinensis 9g, Caulis Spatholobi 16g, Radix Codonopsis 9g.
A, Herba Andrographis is broken into coarse powder, extracts with ethanol refluxing process and make clear paste, medicinal residues are standby;
B, Radix Angelicae Sinensis is broken into coarse powder, makes clear paste according to the percolation under fluid extract and the extractum item, medicinal residues are standby;
C, Caulis Mahoniae and Fructus Zanthoxyli Dissiti two flavor medical material decocting in water are extracted twice, after filtering the merging of both filtrates is made clear paste;
D, with Radix Flemingiae Philippinensis, Radix Rosae Laevigatae, Caulis Spatholobi, Radix Codonopsis four flavors, after-filtration of extracting in water with the Radix Angelicae Sinensis medicinal residues of technology A and the Herba Andrographis medicinal residues decocting in water of technology B, filters the medicinal residues after filtering, merging filtrate is also made clear paste;
E, the above-mentioned 4 kinds of clear paste of merging through mixing, are granulated, drying, and granulate incapsulates, and makes capsule, promptly.
The discrimination method of embodiment 2 medicine capsules of the present invention
A, get capsule content 2 grams of the method preparation of the foregoing description 1, add 60% ethanol 20ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 2ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 1g, shines medical material solution in pairs with legal system.According to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B) test, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (9: 1) is developing solvent, launches, and takes out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color.
B, get capsule content 1 gram of the method preparation of the foregoing description 1, add 40% methanol 20ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds the about 2ml of methanol makes dissolving, be added on neutral alumina post (200 orders of having handled well, 5g, internal diameter 15mm) on, with 40% methanol 100ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 2ml makes dissolving, as need testing solution.Other gets Radix Codonopsis control medicinal material 1g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned two kinds of each 2ul of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with n-butyl alcohol-butanols-water (15: 3: 2), launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color.
C, get capsule content 1 gram of the method preparation of the foregoing description 1, add hydrochloric acid-alcohol mixed solution (1: 100) 20ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 2ml makes dissolving, as need testing solution.Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned reference substance solution 2ul, need testing solution 4ul, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-methanol-isopropyl alcohol-water (6: 3: 1.5: 1.5: 0.3) is developing solvent, puts in the chromatography cylinder of ammonia saturated with vapor, launches, take out, dry, put under the ultra-violet lamp (365nm) and inspect in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
The discrimination method of embodiment 3 medicine capsules of the present invention
A, get capsule content 3 grams of the method preparation of the foregoing description 1, add 95% ethanol 30ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 1.5g, shines medical material solution in pairs with legal system.According to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B) test, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (11: 1.5) is developing solvent, launches, and takes out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color.
B, get capsule content 0.5 gram of the method preparation of the foregoing description 1, add 80% methanol 10ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds the about 3ml of methanol makes dissolving, be added on neutral alumina post (100 orders of having handled well, 5g, internal diameter 10mm) on, with 60% methanol 50ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 3ml makes dissolving, as need testing solution.Other gets Radix Codonopsis control medicinal material 0.5g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned two kinds of each 2ul of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with n-butyl alcohol-butanols-water (10: 1: 1), launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color.
C, get capsule content 3 grams of the method preparation of the foregoing description 1, add hydrochloric acid-alcohol mixed solution (0.5: 80) 10ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 1ml makes dissolving, as need testing solution.Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned reference substance solution 2ul, need testing solution 4ul, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-methanol-isopropyl alcohol-water (8: 5: 2.5: 0.5: 0.5) is developing solvent, puts in the chromatography cylinder of ammonia saturated with vapor, launches, take out, dry, put under the ultra-violet lamp (365nm) and inspect in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
The discrimination method of embodiment 4 medicine capsules of the present invention
A, get capsule content 3 grams of the method preparation of the foregoing description 1, add 40% ethanol 10ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 3ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 1.5g, shines medical material solution in pairs with legal system.According to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B) test, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (7: 1.5) is developing solvent, launches, and takes out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color.
B, get capsule content 1.5 grams of the method preparation of the foregoing description 1, add 50% methanol 30ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds the about 3ml of methanol makes dissolving, be added on neutral alumina post (200 orders of having handled well, 5g, internal diameter 10mm) on, with 60% methanol 150ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 3ml makes dissolving, as need testing solution.Other gets Radix Codonopsis control medicinal material 0.5g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned two kinds of each 2ul of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with n-butyl alcohol-butanols-water (10: 5: 3), launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color.
C, get capsule content 3 grams of the method preparation of the foregoing description 1, add hydrochloric acid-alcohol mixed solution (0.5: 120) 10ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 1ml makes dissolving, as need testing solution.Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned reference substance solution 2ul, need testing solution 4ul, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-methanol-isopropyl alcohol-water (8: 5: 0.5: 2.5: 0.5) is developing solvent, puts in the chromatography cylinder of ammonia saturated with vapor, launches, take out, dry, put under the ultra-violet lamp (365nm) and inspect in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
The discrimination method of embodiment 5 medicine capsules of the present invention
A, get capsule content 1 gram of the method preparation of the foregoing description 1, add 40% ethanol 100ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 3ml makes dissolving, as need testing solution.Other gets Radix Angelicae Sinensis control medicinal material 0.5g, shines medical material solution in pairs with legal system.According to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B) test, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (7: 0.5) is developing solvent, launches, and takes out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color.
B, get capsule content 1.5 grams of the method preparation of the foregoing description 1, add 60% methanol 30ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds the about 1ml of methanol makes dissolving, be added on neutral alumina post (100 orders of having handled well, 5g, internal diameter 15mm) on, with 20% methanol 150ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 3ml makes dissolving, as need testing solution.Other gets Radix Codonopsis control medicinal material 1.5g, shines medical material solution in pairs with legal system.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned two kinds of each 2ul of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with n-butyl alcohol-butanols-water (20: 5: 3), launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color.
C, get capsule content 0.5 gram of the method preparation of the foregoing description 1, add hydrochloric acid-alcohol mixed solution (1.5: 120) 10ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 3ml makes dissolving, as need testing solution.Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia nineteen ninety-five version B), draw above-mentioned reference substance solution 2ul, need testing solution 4ul, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-methanol-isopropyl alcohol-water (4: 1: 0.5: 2.5: 0.1) is developing solvent, puts in the chromatography cylinder of ammonia saturated with vapor, launches, take out, dry, put under the ultra-violet lamp (365nm) and inspect in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
The content assaying method of embodiment 6 medicine capsules of the present invention
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get capsule content 0.5 gram of the method preparation of the foregoing description 1, accurately claim surely, put in the tool plug conical flask, precision adds 40% methanol 25ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005) are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (54: 46: 1) is a mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, this pharmaceutical composition contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
The content assaying method of embodiment 7 medicine capsules of the present invention
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get capsule content 1 gram of the method preparation of the foregoing description 1, accurately claim surely, put in the tool plug conical flask, precision adds 50% methanol 35ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005) are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (64: 36: 2) is a mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, this pharmaceutical composition contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
Embodiment 8 content assaying methods of the present invention
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get capsule content 0.1 gram of the method preparation of the foregoing description 1, accurately claim surely, put in the tool plug conical flask, precision adds 60% methanol 15ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography (appendix VID of Chinese Pharmacopoeia version in 2005) are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (44: 56: 0.1) is a mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, this pharmaceutical composition contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
Claims (5)
1, a kind of method of quality control of the treatment medicine for gynecopathy capsule of making by Radix Flemingiae Philippinensis, Radix Rosae Laevigatae, Herba Andrographis, Caulis Mahoniae, Fructus Zanthoxyli Dissiti, Radix Angelicae Sinensis, Caulis Spatholobi, Radix Codonopsis, it is characterized in that this method comprises discriminating and assay two parts, wherein differentiate and comprise employing thin layer chromatography, the discriminating of carrying out with Radix Angelicae Sinensis control medicinal material, Radix Codonopsis control medicinal material and berberine hydrochloride reference substance respectively; Assay is determined to contain Herba Andrographis, with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
2, method of quality control as claimed in claim 1 is characterized in that described discriminating comprises one or more in the following discriminating:
The discriminating of Radix Angelicae Sinensis: get described capsule 's content 1-3g, add ethanol 10-30ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 1-3ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 0.5-1.5g, shines medical material solution in pairs with legal system; Test according to thin layer chromatography, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (7-11: 0.5-1.5) be developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color;
The discriminating of Radix Codonopsis: get described capsule 's content 0.5-1.5g, add methanol 10-30ml, supersound process 30 minutes filters, filtrate is put evaporate to dryness in the water-bath, residue adds the about 1-3ml of methanol makes dissolving, is added on the neutral alumina post of having handled well, with 10-100% methanol 50-150ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 1-3ml makes dissolving, as need testing solution; Other gets Radix Codonopsis control medicinal material 0.5-1.5g, shines medical material solution in pairs with legal system; Test according to thin layer chromatography, draw above-mentioned two kinds of each 2ul of solution, put respectively on same silica gel g thin-layer plate, with n-butyl alcohol-butanols-water (10-20: 1-5: 1-3) be developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color;
The discriminating of berberine hydrochloride: get described capsule 's content 0.5-3g, add hydrochloric acid-alcohol mixed solution (0.5-1.5: 80-120) 10-30ml, supersound process 30 minutes, filter, filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 1-3ml makes dissolving, as need testing solution; Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution; Test according to thin layer chromatography, draw above-mentioned reference substance solution 2ul, need testing solution 4ul, put respectively on same silica gel g thin-layer plate, with benzene-ethyl acetate-methanol-isopropyl alcohol-water (4-8: 1-5: 0.5-2.5: 0.5-2.5: be developing solvent 0.1-0.5), put in the chromatography cylinder of ammonia saturated with vapor, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
3, method of quality control as claimed in claim 2 is characterized in that described discriminating comprises one or more in the following discriminating:
The discriminating of Radix Angelicae Sinensis: get described capsule 's content 2g, add ethanol 20ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds ethanol 2ml makes dissolving, as need testing solution; Other gets Radix Angelicae Sinensis control medicinal material 1g, shines medical material solution in pairs with legal system; Test according to thin layer chromatography, draw above-mentioned two kinds of each 4ul of solution, put respectively on same silica gel g thin-layer plate, with normal hexane-ethyl acetate (9: 1) is developing solvent, launches, and takes out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the fluorescence speckle of same color;
The discriminating of Radix Codonopsis: get described capsule 's content 1g, add methanol 20ml, supersound process 30 minutes filters, filtrate is put evaporate to dryness in the water-bath, residue adds the about 2ml of methanol makes dissolving, is added on the neutral alumina post of having handled well, with 40% methanol 100ml eluting, collect eluent, put evaporate to dryness in the water-bath, residue adds methanol 2ml makes dissolving, as need testing solution; Other gets Radix Codonopsis control medicinal material 1g, shines medical material solution in pairs with legal system; Test according to thin layer chromatography, draw above-mentioned two kinds of each 2ul of solution, putting respectively on same silica gel g thin-layer plate, is developing solvent with n-butyl alcohol-butanols-water (15: 3: 2), launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, about 5 minutes of 105 ℃ of bakings, in the test sample chromatograph, with control medicinal material chromatograph relevant position on show the speckle of same color;
The discriminating of berberine hydrochloride: get described capsule 's content 1g, add hydrochloric acid-alcohol mixed solution (1 → 100) 20ml, supersound process 30 minutes filters, and filtrate is put evaporate to dryness in the water-bath, and residue adds methanol 2ml makes dissolving, as need testing solution; Other gets the berberine hydrochloride reference substance, adds methanol and makes the solution that contains 1mg among every 1ml, in contrast product solution; Test according to thin layer chromatography, draw above-mentioned reference substance solution 2ul, need testing solution 4ul, putting respectively on same silica gel g thin-layer plate, is developing solvent with benzene-ethyl acetate-methanol-isopropyl alcohol-water (6: 3: 1.5: 1.5: 0.3), puts in the chromatography cylinder of ammonia saturated with vapor, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect, in the test sample chromatograph, with the corresponding position of reference substance chromatograph on show the fluorescence speckle of same color.
4, method of quality control as claimed in claim 1 is characterized in that described assay comprises:
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get described capsule 's content 0.1-1g, accurately claim surely, put in the tool plug conical flask, precision adds methanol 15-35ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (44-64: 36-56: 0.1-2) be mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, described capsule 's content contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
5, method of quality control as claimed in claim 4 is characterized in that described assay comprises:
A, precision take by weighing dehydrorographolide reference substance 5mg, put in the 100ml measuring bottle, add dissolve with methanol and are diluted to scale, shake up, and make reference substance solution, contain dehydrorographolide 50ug among wherein every 1ml;
B, get described capsule 's content 0.5g, accurately claim surely, put in the tool plug conical flask, precision adds methanol 25ml, claims decide weight, and supersound process 40 minutes is put coldly, supplies weight, filtration, and filtrate is as need testing solution;
C, photograph high performance liquid chromatography are measured, and chromatographic condition and system suitability test are: with octadecylsilane chemically bonded silica is filler; Methanol-water-glacial acetic acid (54: 46: 1) is a mobile phase; The detection wavelength is 250nm, and number of theoretical plate is pressed the dehydrorographolide peak and calculated, and should be not less than 1000; Accurate respectively reference substance solution and each 10ul of need testing solution of drawing injects chromatograph of liquid, measures, promptly;
D, described capsule 's content contain Herba Andrographis with dehydrorographolide (C
20H
28O
4) meter, must not be less than 0.50mg.
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|---|---|---|---|
| CN2005100864298A CN1931272B (en) | 2005-09-15 | 2005-09-15 | Quality control method for gynecopathy treating medicine capsule |
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|---|---|---|---|
| CN2005100864298A CN1931272B (en) | 2005-09-15 | 2005-09-15 | Quality control method for gynecopathy treating medicine capsule |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102721782A (en) * | 2012-07-02 | 2012-10-10 | 涂瑶生 | Method for detecting quality of philippine flemingia root formula granules |
| CN109470788A (en) * | 2018-11-14 | 2019-03-15 | 株洲千金药业股份有限公司 | A kind of method of quality control of FUKE QIANJIN PIAN |
| WO2020098831A1 (en) * | 2018-11-14 | 2020-05-22 | 株洲千金药业股份有限公司 | Gynecological qianjin capsule and quality control method therefor |
-
2005
- 2005-09-15 CN CN2005100864298A patent/CN1931272B/en active Active
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102721782A (en) * | 2012-07-02 | 2012-10-10 | 涂瑶生 | Method for detecting quality of philippine flemingia root formula granules |
| CN102721782B (en) * | 2012-07-02 | 2014-03-26 | 涂瑶生 | Method for detecting quality of philippine flemingia root formula granules |
| CN109470788A (en) * | 2018-11-14 | 2019-03-15 | 株洲千金药业股份有限公司 | A kind of method of quality control of FUKE QIANJIN PIAN |
| WO2020098831A1 (en) * | 2018-11-14 | 2020-05-22 | 株洲千金药业股份有限公司 | Gynecological qianjin capsule and quality control method therefor |
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| CN1931272B (en) | 2011-01-26 |
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