CN1739598A - Chinese medicine injection for treating cancer and its prepn process and quality control method - Google Patents
Chinese medicine injection for treating cancer and its prepn process and quality control method Download PDFInfo
- Publication number
- CN1739598A CN1739598A CNA2005100431342A CN200510043134A CN1739598A CN 1739598 A CN1739598 A CN 1739598A CN A2005100431342 A CNA2005100431342 A CN A2005100431342A CN 200510043134 A CN200510043134 A CN 200510043134A CN 1739598 A CN1739598 A CN 1739598A
- Authority
- CN
- China
- Prior art keywords
- solution
- water
- present
- reference substance
- adds
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Pending
Links
Landscapes
- Medicines Containing Plant Substances (AREA)
Abstract
The present invention relates to one kind of Chinese medicine injection for treating cancer and its preparation process and quality control method. The Chinese medicine injection is prepared with red ginseng, astragalus root, toad cake and cantharides. The quality control method includes the thin-layer identification of red ginseng and astragalus root, toad cake; the detection of pH value, total solid, ignition residue, heavy metal, As salt, protein, tanning, resin, oxalate, potassium ion and pyretogen; and the content measurement of total polysaccharide, toad cake and cantharides.
Description
Technical field
The present invention relates to a kind of Chinese medicine preparation and method for making thereof and method of quality control, particularly a kind of Chinese medicine and method for making and method of quality control that is used for the treatment of hepatocarcinoma.
Background technology
The people's health and lives in the cancer serious harm, and ranks first place with liver cancer mortality among the cancer mortality crowd at home.December in 2004 Chinese patent gazette on the 8th discloses the name of being declared by the applicant, and to be called " a kind of Chinese patent medicine that is used for the treatment of hepatocarcinoma ", publication number be 1552434 patent application, it mainly is prepared from by Radix Ginseng Rubra, the Radix Astragali, Bufo siccus, Mylabris, now, we are on the basis of original prescription, by a large amount of experiments, be made into injection and scientifically carry out quality control, clinical pharmacodynamic experiment effect is remarkable.
Summary of the invention
The objective of the invention is to: Chinese medicine injection and method for making and method of quality control that a kind of obvious results treatment hepatocarcinoma is provided.
Technical solution of the present invention is achieved in that
One, prescription:
Radix Ginseng Rubra 100g, Radix Astragali 300g, Venenum Bufonis 0.4g, Mylabris 2g.
Two, method for making:
More than four the flavor, Venenum Bufonis is ground into fine powder, it is an amount of to add water, makes pasty state, adds ethanol 10ml, soaked 24 hours, the filtration, filtrate recycling ethanol, standby; Radix Ginseng Rubra is made the solvent heating and refluxing extraction three times with 65% ethanol, and 1.5 hours for the first time, each 1 hour of second and third time, merge extractive liquid, reclaims ethanol, and it is an amount of to add the injection water, stir evenly, regulate pH value to 12 with the calcium hydroxide saturated solution, put coldly, pass to sulfur dioxide and regulate pH value to 4.2～5.0, leave standstill, filter, filtrate decompression is concentrated into 100ml, and is standby; Powder of cantharide is broken into fine powder, decocts with water secondary, and each 0.5 hour, collecting decoction, filter, filtrate is regulated pH value to 12 with the calcium hydroxide saturated solution, puts coldly, passes to sulfur dioxide and regulates pH value to 6.5～8.0, leave standstill, filter, filtrate decompression is concentrated into 20ml, and is standby; Radix Astragali section decocts with water secondary, and each 1 hour, collecting decoction, filter, filtrate is regulated pH value to 12 with the calcium hydroxide saturated solution, puts coldly, passes to sulfur dioxide and regulates pH value to 4.2～5.0, leave standstill, filter, filtrate decompression is concentrated into 300ml, merges with above-mentioned concentrated solution, mixing, cold preservation (4～10 ℃) 12 hours filters, filtrate adds the injection water to 1000ml, and mixing filters with microporous filter membrane, embedding, sterilization, promptly.
Three, character:
This product is fallow clear liquid.
Four, differentiate:
(1) gets this product 10ml, extract 2 times, each 20ml with water saturated n-butyl alcohol jolting, merge n-butanol extracting liquid, evaporate to dryness, residue water 3ml dissolving, be added in neutral alumina post (100～120 orders, 5g, internal diameter 0.9cm) on, with 40% methanol 50ml eluting, collect eluent, evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution.Other gets ginsenoside Re's reference substance, ginsenoside Rg
1Reference substance, astragaloside reference substance add methanol and make the mixed solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, lower floor's solution of placing below 10 ℃ with chloroform-methanol-water (13: 7: 2) is developing solvent, launches, and takes out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, puts respectively under daylight and the ultra-violet lamp (365nm) and inspects.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, the speckle of apparent same color under the daylight; The fluorescence speckle of apparent same color under the ultraviolet light.
(2) get this product 30ml, extract 2 times with the chloroform jolting, each 30ml merges chloroform extraction liquid, and evaporate to dryness, residue add chloroform 0.5ml makes dissolving, as need testing solution.Other gets bufogenin reference substance and cinobufagin reference substance, adds chloroform and makes the mixed solution that every 1ml contains 1mg, in contrast product solution.Test according to thin layer chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 B), draw need testing solution 10 μ l, reference substance solution 5 μ l, put respectively on same silica gel g thin-layer plate, with chloroform-acetone-cyclohexane extraction-ethyl acetate (4: 2.2: 5: 1) be developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
Five, check:
(1) pH value: should be 6.5～8.5 (an appendix VII of Chinese Pharmacopoeia version in 2005 G).
(2) total solid: precision is measured this product 10ml, puts in the evaporating dish that is dried to constant weight, and evaporate to dryness 105 ℃ of dryings 3 hours, was put in the exsiccator cooling 30 minutes, claimed to decide weight rapidly.The every 1ml of this product contains total solid must not be less than 20mg.
(3) residue on ignition: precision is measured this product 2ml, puts in the crucible of ignition to constant weight, and evaporate to dryness, from " slowly blazing " to carbonization fully, inspection (an appendix IX of Chinese Pharmacopoeia version in 2005 J) in accordance with the law, residue must not cross 0.65%.
(4) heavy metal: get the residue under the residue on ignition inspection item, check (an appendix IX of Chinese Pharmacopoeia version in 2005 E second method) in accordance with the law.Contain heavy metal and must not cross 10/1000000ths.
(5) arsenic salt: get this product 1ml, adding calcium hydroxide 1g, mixing, evaporate to dryness earlier makes carbonization with little heated is bright, blazingly makes complete ashing at 500～600 ℃ again, puts coldly, and residue adds hydrochloric acid 5ml and water 23ml makes dissolving, as need testing solution.Get need testing solution, check (an appendix IX of Chinese Pharmacopoeia version in 2005 F first method) in accordance with the law.Arsenic content must not cross 2/1000000ths.
(6) protein: get this product 1ml, add 30% sulfosalicylic acid test solution 1ml of new preparation, mixing was placed 5 minutes, must not become turbid.
(7) tannin: get this product 1ml, check (an appendix IX of Chinese Pharmacopoeia version in 2005 S) in accordance with the law.Should be up to specification.
(8) resin: get this product 5ml, check (an appendix IX of Chinese Pharmacopoeia version in 2005 S) in accordance with the law.Should be up to specification.
(9) oxalates: it is an amount of to get this product, regulates pH value to 1～2 with dilute hydrochloric acid, filters, and gets filtrate 2ml, regulates pH value to 5～6 with sodium hydroxide test solution, adds 3% calcium chloride solution 2-3 and drips, and places 10 minutes, must not become turbid or precipitate.
(10) potassium ion: precision is measured this product 2ml, checks (an appendix IX of Chinese Pharmacopoeia version in 2005 S) in accordance with the law.Should be up to specification.
(11) pyrogen: get this product, check that in accordance with the law (an appendix XIII of Chinese Pharmacopoeia version in 2005 A) checks, dosage is by the every 1kg body weight injection of rabbit 3ml.Should be up to specification.
(12) other: should meet every regulation relevant under the injection item (appendix IU of Chinese Pharmacopoeia version in 2005).
(13) n-butanol extract: precision is measured this product 25ml, puts in the separatory funnel, extracts 3 times with the chloroform jolting, each 30ml merges chloroform extraction liquid, washes with water 2 times, each 10ml, washing liquid is incorporated in the above-mentioned aqueous solution, extracts 3 times with water saturated n-butyl alcohol jolting, each 30ml merges n-butanol extracting liquid, puts in the evaporating dish that is dried to constant weight, evaporate to dryness 105 ℃ of dryings 3 hours, was put in the exsiccator cooling 30 minutes, claim rapidly to decide weight, promptly.
This product contains n-butanol extract must not be less than 0.15%.
Six, assay:
(1) total polysaccharides
The preparation of reference substance solution: be taken at 80 ℃ of anhydrous glucose reference substance 80mg that are dried to constant weight, the accurate title, decide, put in the 100ml measuring bottle, with water dissolution and be diluted to scale, shake up, precision is measured 5ml, put in the 50ml measuring bottle, add water to scale, shake up, promptly get (every 1ml contains anhydrous glucose 80 μ g).
The preparation of standard curve: precision is measured reference substance solution 0.2,0.4,0.6,0.8,1.0ml, put respectively in the 10ml tool plug test tube, precision adds water and makes into 1ml in each pipe, and the accurate sulphuric acid anthrone solution that adds (is got anthrone 50mg, added water 25ml and sulphuric acid 75ml makes dissolving in ice bath.Face and use preceding preparation) 4ml, shake up, put in the boiling water bath and heated 10 minutes, cooled off 40 minutes; With the reagent corresponding is blank.According to ultraviolet visible spectrophotometry (appendix VA of Chinese Pharmacopoeia version in 2005), measure trap at 625nm wavelength place, be that vertical coordinate, concentration are abscissa drawing standard curve with the trap.
The preparation of need testing solution: precision is measured this product 50ml, puts and is concentrated into about 5ml in the water-bath, puts coldly, is transferred in the 10ml measuring bottle, adds water to scale, shakes up.Precision is measured 1ml, the accurate dehydrated alcohol 10ml that adds, and close plug shakes up, and leaves standstill 4 hours, and is centrifugal, abandoning supernatant, precipitation is used water dissolution, is transferred in the 50m1 measuring bottle, is diluted with water to scale, shakes up, precision is measured 5ml, puts in the 50ml measuring bottle, adds water to scale, shakes up, promptly.
Algoscopy: precision is measured need testing solution 1ml, puts in the 10ml tool plug test tube method under the sighting target directrix curve item, from " the accurate sulphuric acid anthrone solution 4ml that adds in ice bath ", measure trap in accordance with the law, the amount of reading polysaccharide the need testing solution from standard curve, promptly.
The every 1ml of this product contains total polysaccharides with anhydrous glucose (C
6H
12O
6) meter, must not be less than 3.5mg.
(2) Venenum Bufonis is measured according to high performance liquid chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 D).
Chromatographic condition and system suitability test: with the octadecylsilane chemically bonded silica is filler; With normal propyl alcohol-methanol-0.04% potassium dihydrogen phosphate (2: 3: 7) is mobile phase; The detection wavelength is 300nm.Number of theoretical plate calculates by the cinobufagin peak should be not less than 2500.
The preparation of reference substance solution: get the bufogenin reference substance respectively and the cinobufagin reference substance is an amount of, accurately claim surely, add methanol and make the mixed solution that every 1ml contains 40 μ g, promptly.
The preparation of need testing solution: precision is measured this product 20ml, put in the separatory funnel, extract 4 times with the chloroform jolting, each 30ml, extracting solution filters with the filter paper that is covered with a small amount of anhydrous sodium sulfate, filtrate merges, evaporate to dryness, residue is with dissolve with methanol and be transferred in the 5ml measuring bottle, adds methanol to scale, shake up, promptly.
Algoscopy: accurate respectively reference substance solution and each 10 μ l of need testing solution of drawing, inject chromatograph of liquid, measure, promptly.
The every 1ml of this product contains Venenum Bufonis with bufogenin (C
24H
32O
4) and cinobufagin (C
26H
34O
6) the total amount meter, should be 12～18 μ g.
(3) Mylabris is measured according to gas chromatography (an appendix VI of Chinese Pharmacopoeia version in 2005 E).
Chromatographic condition and system suitability test: with methyl silicone rubber (SE-30) is fixative, and coating concentration is 3.5%; Column temperature is 165 ± 10 ℃.Number of theoretical plate calculates by the cantharidin peak should be not less than 2500.
The preparation of reference substance solution: it is an amount of to get the cantharidin reference substance, and accurate the title decides, and adds chloroform and makes the solution that every 1ml contains 0.32mg, promptly.
The preparation of need testing solution: precision is measured this product 50ml, regulates pH value to 3～4 with 5% sulfuric acid solution, extracts 5 times with water saturated chloroform jolting, each 40ml, merge chloroform extraction liquid, filter through the filter paper that is covered with a small amount of anhydrous sodium sulfate, filtrate merges, put and evaporate chloroform in the water-bath below 60 ℃ to about 10ml, naturally wave again and be dissipated to 2～3ml, be transferred in the 5ml measuring bottle, add chloroform to scale, shake up, promptly.
Algoscopy: accurate respectively reference substance solution 2 μ l and the need testing solution 4 μ l of drawing, inject gas chromatograph is measured, promptly.
The every 1ml of this product contains Mylabris with cantharidin (C
10H
12O
4) meter, should be 14～18 μ g.
Seven, function and curing mainly:
QI invigorating is set upright ， Xiao Disorder eliminating stagnation.Be used for advanced primary liver cancer qi deficiency and Stagnation card, disease is seen right latus palpatable abdominal mass, unmovable pain, abdominal distention and little diet, fatigue and weakness.
Eight, usage and consumption:
Intravenous drip.This product 40～60ml is used with 500ml5% glucose injection or 0.9% sodium chloride injection dilution back, drip speed and be advisable to be no more than 60 of per minutes, 1 time on the one, 45 is a course of treatment.Drug withdrawal one Zhou Houke carries out the treatment of next course of treatment, or follows the doctor's advice.Per course of treatment, consumption first reduced by half, and the ratio that this product is diluted to medicinal liquid and diluent was not less than 1: 20, dripped a fast per minute and was no more than 15.As have no adverse reaction, can increase gradually after half an hour and drip speed.
Nine, note:
This product be must guard against direct intravenous injection.This product and other medicine mixed venous must not be instiled.This product contains cantharidin and cinobufagin, forbids directly to use without dilution; Must not add to drip in the kettle and splash into.Generally speaking, the ratio of this product and diluent should be not less than 1: 10.Reduce the feed liquor amount if desired, the ratio of this product and diluent is the highest must not be greater than 1: 5, at the medicinal liquid that uses 1: 10 above dilution ratio after 2 days, when not seeing any untoward reaction, can use the medicinal liquid of 1: 5 dilution ratio, a speed of this concentration liquid is no more than 50 with per minute and is advisable.Should note detecting liver, renal function during the medication.If uncomfortable in chest, cardiopalmus, reaction such as breathe hard, drug withdrawal immediately, and do conventional the disposal.The symptom that may occur urine urgency-frequency after the small number of patients medication, accidental hematuria and albuminuria, above-mentioned situation occurring should discontinue medication, should be during the reuse medicine by 1: 20 dilution proportion medicinal liquid, or slow down and drip speed (being no more than 40 of per minutes), or polydipsia water.Hepatorenal damage may occur after the small number of patients medication, accidentally feel sick, vomiting, abdominal distention.
Chinese medicine injection of the present invention has carried out following pharmacodynamic experiment:
One, to H
22Inhibiting research in the Hepar Mus JEG-3 body
1 ascites tumor goes down to posterity
The sterilization of lotus ascites tumor Mus abdominal part, suction ascites, normal saline dilution in 1: 3.Wait to plant the mouse web portion sterilization, lumbar injection diluent 0.5ml.When plantation Mus abdominal circumference obviously increases, gather ascites and carry out the solid tumor plantation.
Plantation of 2 solid tumors and processing
50 of homology kunming mices, about body weight 20g, male and female half and half, the right fore skin degerming, oneself dilutes ascites 0.1ml (normal saline dilution in 1: 3) subcutaneous injection.Inoculate the 8th day and be divided into 5 groups (10 every group) at random, be that (0.2ml/ only for normal saline, the blank group), dosage group 753mg/kg, the present invention's heavy dose are organized 1412.5mg/kg, administration every other day 1 time, totally 10 times among 5-FU group 40mg/kg, small dose group 452mg/kg of the present invention, the present invention.1 week of drug withdrawal, take off cervical vertebra and put to death mice, measure tumor volume, tumor heavy (w), calculate tumour inhibiting rate.Tumour inhibiting rate=(W
Matched group-W
Experimental group)/W
Matched group* 100%.The results are shown in Table 1.
Table 1 couple H
22Inhibitory action in the Hepar Mus JEG-3 body
| Group | n | Dosage mg/kg | Tumor heavy (g) | Tumour inhibiting rate (%) |
| The heavy dose of group of dosage group the present invention among the small dose group the present invention of the present invention of matched group 5-FU group | 10 10 10 10 10 | 40 452 753 1412.5 | 1.76±0.31 0.68±0.42** 1.40±0.33* 1.10±0.30** 0.73±0.37** | 61.4 20.5 37.5 58.5 |
Compare * P＜0.05 * * P＜0.05 with matched group
3 result of the tests
The result shows that the large, medium and small dosage group of the present invention all has tumor-inhibiting action in various degree, compares with matched group, and small dose group has significant difference (P＜0.05=; In, heavy dose of group is to H
22The Hepar Mus cancer has the tumor-inhibiting action (P＜0.01) of highly significant.The tumour inhibiting rate of the heavy dose of group of the present invention is 58.5%, is only second to 5-FU group (61.4%), illustrates that the present invention is to H
22The Hepar Mus JEG-3 has good inhibition effect.
Two, to mice S
180The inhibiting research of solid tumor
Get 50 of kunming mices, about body weight 20g, male and female half and half are given the subcutaneous injection of mice one side armpit S
180Ascites diluent 0.2ml (contains oncocyte 10 approximately
6), be divided into 5 groups (10 every group) next day at random, be that (0.2ml/ only for normal saline, the blank group), dosage group 753mg/kg, the heavy dose of group of the present invention 1412.5mg/kg among cyclophosphamide (cy) group 25mg/kg, small dose group 452mg/kg of the present invention, the present invention, administration every day 1 time, continuous 7 days.Put to death mice next day after the drug withdrawal, peels off the subcutaneous tumors piece, claims tumor heavy, calculates tumour inhibiting rate, compares the difference between administration group and the matched group, and carry out test of significance.The results are shown in Table 2.
Table 2 couple mice S
180The inhibiting research of solid tumor
| Group | Dosage mg/kg | n | Tumor heavy (g) | Tumour inhibiting rate (%) |
| The heavy dose of group of dosage group the present invention among the small dose group the present invention of the present invention of matched group cyclophosphamide group | - 25 452 753 1412.5 | 10 10 10 10 10 | 2.75±0.72 0.31±0.24*** 2.25±0.89 1.62±1.02* 1.51±0.98** | 88.7 18.2 41.1 45.1 |
Compare * P＜0.05 * * P＜0.01 * * * P＜0.001 with matched group
The result shows that the large, medium and small dosage group of the present invention all has tumor-inhibiting action in various degree, compares with matched group, and the dosage group can suppress mice S significantly among the present invention
180Solid tumor (P＜0.05).The inhibition mice S of the heavy dose of group energy of the present invention highly significant
180Solid tumor (P＜0.01).
Three, to the influence of NK cells in mice poison
Get 8-20 week Bab1/c mouse inbred lines, be divided into 5 groups (10 every group) at random, be that (0.2ml/ only for the normal saline group, the blank group), dosage group 753mg/kg, heavy dose of 1412.5mg/kg, administration every day 1 time, continuous 21 days organized of the present invention among interferon group 5000u/kg, small dose group 452mg/kg of the present invention, the present invention, after the last administration 12 hours, put to death mice, spleen lymphocyte is made the individual cells suspension, use Tris-NH with aseptic apparatus
4The molten cell of CL, counting is made into 2 * 10
7/ ml is stand-by.
With the 125I-udR trophophase 1,500,000 YAC-1 cells of taking the logarithm, labelling is after 2 hours, gives a baby a bath on the third day after its birth time with PBS, and it is stand-by that counting is made into 100,000/ml.
Add in the 96 hole circle chassis according to the dead target ratio of difference, and establish nature release, maximum release, put CO
2Behind the incubator 18 hours, get supernatant, measure ray, calculate and kill and wound percentage rate, the results are shown in Table 3.
Table 3 the present invention is to the influence of NK cells in mice poison
| Group | Dosage | n | NK cell toxicant (%) |
| The heavy dose of group of dosage group the present invention among the small dose group the present invention of the present invention of normal saline group interferon group | - 5000u/kg 452mg/kg 753mg/kg 1412.5mg/kg | 10 10 10 10 10 | 25.0 58.2 55.5 66.5* 78.9** |
Annotate: compare * P＜0.05 * * P＜0.01 with the normal saline group
The result show the present invention little, in, heavy dose of group all has the cytotoxic effect of increase NK in various degree, with matched group relatively, among the present invention, heavy dose of group can increase NK cytotoxicity (P＜0.05) significantly.
Four, the removing of india ink is tested
Get the healthy mice of body weight between 18-22g, male and female half and half, be divided into 5 groups immediately, 10 every group, normal saline group (blank 0.5ml/ is only), dosage group 1130mg/kg, the heavy dose of 2260mg/kg of the present invention among interferon group (positive drug 4000 υ), small dose group 565mg/kg of the present invention, the present invention, administration every day 1 time, successive administration 15 days, after the last administration 1 hour, 0.1% india ink 0.1ml is injected from white mice tail vein, inject back 3 minutes (t
1) and 13 minutes (t
2), get blood 2.5ml from the socket of the eye vein respectively and be dissolved in 2ml0.1%NaHCO
3In the solution, under 650nm, measure optical density OD1 (3 minutes) and OD2 (13 minutes) respectively, calculate phagocytic index K value with 721 type spectrophotometers.Experiment is carried out three times altogether.Experimental result sees Table 4
The scavenging action of table 4 pair india ink (X ± SD)
| Group | n | Dosage (/kg) | OD1 | OD2 | The K value |
| The heavy dose of group of dosage group the present invention among the small dose group the present invention of the present invention of normal saline group interferon | 10 10 10 10 10 | 4000υ 565mg 1130mg 2260mg | 1.305±0.097 1.134±0.095 1.069±0.106 1.179±0.098 1.100±0.091 | 1.090±0.93 0.578±0.186 0.621±0.280 0.780±0.172 0.573±0.178 | 0.00786 0.02915** 0.02353** 0.01798** 0.02826** |
Compare * P＜0.05 * * P＜0.01 with the normal saline group
The result shows: compare with the blank group, the large, medium and small dosage of the present invention has tangible potentiation (P＜0.01) to the phagocytic function of white mice reticuloendothelial system.
Five, to the influence of Turnover of Mouse Peritoneal Macrophages phagocytic function
By the sterile working, vein is got blood under the chicken wing, puts into the triangular flask that Ah's test solution is housed immediately, puts in the 0-4 ℃ of refrigerator and preserves.With physiological saline solution centrifuge washing three times, be made into 5% cell suspension with normal saline standby before using.
Get the healthy mice of body weight between 18-22g, male and female half and half, be divided into 5 groups immediately, every group 10, normal saline group (blank 0.5ml/ only), interferon group (positive drug 4000 υ), small dose group 565mg/kg of the present invention, dosage group 1130mg/kg among the present invention, the heavy dose of 2260mg/kg of the present invention, administration every day 1 time, successive administration 15 days is in collecting peritoneal macrophage preceding 10 hours, every mouse peritoneal is injected 5% chicken erythrocyte suspension 0.4ml, put to death mice, the sterilization abdominal part is cut off skin, exposes peritoneum, inject Han Shi liquid 2.5ml, soft abdominal part is in the hope of obtaining more macrophage.Extract peritoneal scissors one osculum with tweezers, draw peritoneal fluid 2ml with the long-neck suction pipe and place test tube, behind the mixing, a little splashes into (about 2 * the 1.5cm of drop size on the microscope slide sucking-off
2), lie in the enamel tray that contains wet sand dripping sheet, put in 37 ℃ of incubators and hatched 30 minutes, brush away the chicken red blood cell that floats on the sheet in normal saline after, cold wind dries up, and methanol is fixed 5 minutes, dyeed 5-10 minute with Ji Mu Sa-Rui Teshi dye liquor, the water flushing is dried, and uses oily sem observation, calculates macrophage phagocytic percentage rate and phagocytic index.The results are shown in Table 5.
The influence of table 5 pair Turnover of Mouse Peritoneal Macrophages phagocytic function (X ± SD)
| Group | n | Dosage (/kg) | Phagocytic percentage (%) | Phagocytic index |
| The heavy dose of group of dosage group the present invention among the small dose group the present invention of the present invention of normal saline group interferon | 10 10 10 10 10 | 4000υ 565mg 1130mg 2260mg | 40.1±16.09 62.00±13.4** 50.01±11.54 59.61±13.24** 69.18±14.09*** | 1.00±0.53 2.24±0.66** 1.42±0.58 1.98±0.47** 2.45±0.68*** |
Compare * P＜0.05 * * P＜0.01 * * P＜0.001 with the normal saline group
The result shows: with the normal saline group relatively, the large, medium and small dosage group of the present invention all has in various degree enhancing intraperitoneal mouse macrophage to the phagocytic function of chicken red blood cell.Learn by statistics and handle, there is significant difference P＜0.01 and P＜0.001.
Above-mentioned pharmacodynamic experiment shows: the present invention is to H
22Hepar Mus JEG-3 and mice S
180Solid tumor all has significant inhibitory effect, has effects such as enhance immunity in addition.
Claims (3)
1, a kind of Chinese medicine injection that is used for the treatment of hepatocarcinoma, it is characterized in that it makes by the following method: take by weighing raw material Radix Ginseng Rubra 100g, Radix Astragali 300g, Venenum Bufonis 0.4g, Mylabris 2g, Venenum Bufonis is ground into fine powder, it is an amount of to add water, makes pasty state, adds ethanol 10ml, soaked 24 hours, filter, filtrate recycling ethanol, standby; Radix Ginseng Rubra is made the solvent heating and refluxing extraction three times with 65% ethanol, and 1.5 hours for the first time, each 1 hour of second and third time, merge extractive liquid, reclaims ethanol, and it is an amount of to add the injection water, stir evenly, regulate pH value to 12 with the calcium hydroxide saturated solution, put coldly, pass to sulfur dioxide and regulate pH value to 4.2～5.0, leave standstill, filter, filtrate decompression is concentrated into 100ml, and is standby; Powder of cantharide is broken into fine powder, decocts with water secondary, and each 0.5 hour, collecting decoction, filter, filtrate is regulated pH value to 12 with the calcium hydroxide saturated solution, puts coldly, passes to sulfur dioxide and regulates pH value to 6.5～8.0, leave standstill, filter, filtrate decompression is concentrated into 20ml, and is standby; The Radix Astragali decocts with water secondary, and each 1 hour, collecting decoction filtered, filtrate is regulated pH value to 12 with the calcium hydroxide saturated solution, puts coldly, passes to sulfur dioxide and regulates pH value to 4.2～5.0, leaves standstill, filter, filtrate decompression is concentrated into 300ml, merges with above-mentioned concentrated solution, mixing, 4～10 ℃ of cold preservation 12 hours filters, filtrate adds the injection water to 1000ml, and mixing filters with microporous filter membrane, embedding, sterilization, promptly.
2, the method for making of the Chinese medicine injection of treatment hepatocarcinoma as claimed in claim 1, it is characterized in that: take by weighing raw material Radix Ginseng Rubra 100g, Radix Astragali 300g, Venenum Bufonis 0.4g, Mylabris 2g, Venenum Bufonis is ground into fine powder, it is an amount of to add water, makes pasty state, adds ethanol 10ml, soaked 24 hours, filter, filtrate recycling ethanol, standby; Radix Ginseng Rubra is made the solvent heating and refluxing extraction three times with 65% ethanol, and 1.5 hours for the first time, each 1 hour of second and third time, merge extractive liquid, reclaims ethanol, and it is an amount of to add the injection water, stir evenly, regulate pH value to 12 with the calcium hydroxide saturated solution, put coldly, pass to sulfur dioxide and regulate pH value to 4.2～5.0, leave standstill, filter, filtrate decompression is concentrated into 100ml, and is standby; Powder of cantharide is broken into fine powder, decocts with water secondary, and each 0.5 hour, collecting decoction, filter, filtrate is regulated pH value to 12 with the calcium hydroxide saturated solution, puts coldly, passes to sulfur dioxide and regulates pH value to 6.5～8.0, leave standstill, filter, filtrate decompression is concentrated into 20ml, and is standby; The Radix Astragali decocts with water secondary, and each 1 hour, collecting decoction filtered, filtrate is regulated pH value to 12 with the calcium hydroxide saturated solution, puts coldly, passes to sulfur dioxide and regulates pH value to 4.2～5.0, leaves standstill, filter, filtrate decompression is concentrated into 300ml, merges mixing with above-mentioned concentrated solution, 4～10 ℃ of cold preservation 12 hours,, filter, filtrate adds the injection water to 1000ml, and mixing filters with microporous filter membrane, embedding, sterilization, promptly.
3, the method for quality control of the Chinese medicine injection of treatment hepatocarcinoma as claimed in claim 1 is characterized in that described method is one or more the combination in the following method:
(1) thin layer of Radix Ginseng, the Radix Astragali is differentiated: get 10ml of the present invention, extract 2 times with water saturated n-butyl alcohol jolting, each 20ml merges n-butanol extracting liquid, evaporate to dryness, residue water 3ml dissolving, being added in 100～120 orders, 5g, internal diameter is on the 0.9cm neutral alumina post, with 40% methanol 50ml eluting, collect eluent, evaporate to dryness, residue add methanol 1ml makes dissolving, as need testing solution; Other gets ginsenoside Re's reference substance, ginsenoside Rg
1Reference substance, astragaloside reference substance add methanol and make the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to the Chinese Pharmacopoeia thin layer chromatography, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica gel g thin-layer plate, with place below 10 ℃, proportioning is that chloroform-methanol-water lower floor solution of 13: 7: 2 is developing solvent, launches, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing at 105 ℃, puts respectively under the outer light modulation of daylight and purple 365nm and inspects; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, the speckle of apparent same color under the daylight, the fluorescence speckle of apparent same color under the ultraviolet light.
(2) thin layer of Venenum Bufonis is differentiated: get 30ml of the present invention, extract 2 times with the chloroform jolting, each 30ml merges chloroform extraction liquid, and evaporate to dryness, residue add chloroform 0.5ml makes dissolving, as need testing solution; Other gets bufogenin reference substance and cinobufagin reference substance, adds chloroform and makes the mixed solution that every 1ml contains 1mg, in contrast product solution; Test according to the Chinese Pharmacopoeia thin layer chromatography, draw need testing solution 10 μ l, reference substance solution 5 μ l, put respectively on same silica gel g thin-layer plate, with proportioning is 4: 2.2: 5: chloroform-acetone of 1-cyclohexane extraction-ethyl acetate is developing solvent, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the speckle of same color.
(3) inspection of pH value:, should be 6.5～8.5 according to Chinese Pharmacopoeia pH value algoscopy.
(4) inspection of total solid: precision is measured 10ml of the present invention, puts in the evaporating dish that is dried to constant weight, and evaporate to dryness 105 ℃ of dryings 3 hours, was put in the exsiccator cooling 30 minutes, claimed to decide weight rapidly; The every 1ml of the present invention contains total solid must not be less than 20mg.
(5) inspection of residue on ignition: precision is measured 2ml of the present invention, puts in the crucible of ignition to constant weight evaporate to dryness, slowly blazing to carbonization fully, put and be chilled to room temperature, add sulphuric acid 0.5-1ml make moistening, after low-temperature heat to sulfuric acid vapor eliminates, blazingly make complete carbonization at 700-800 ℃, move in the exsiccator, put and be chilled to room temperature, after accurate title is fixed, at 700-800 ℃ of ignition to constant weight, residue must not cross 0.65% again.
(6) inspection of heavy metal: precision is measured 2ml of the present invention, puts in the crucible of ignition to constant weight evaporate to dryness, slowly blazing carbonization is extremely fully put and is chilled to room temperature, adds sulphuric acid 0.5-1ml and makes moistening, after low-temperature heat to sulfuric acid vapor eliminates, blazingly make complete carbonization at 500-600 ℃, move in the exsiccator, put and be chilled to room temperature, after accurate title is fixed, at 500-600 ℃ of ignition to constant weight, check again, contain heavy metal and must not cross 10/1000000ths according to Chinese Pharmacopoeia heavy metal inspection technique.
(7) inspection of arsenic salt: get 1ml of the present invention, adding calcium hydroxide 1g, mixing, evaporate to dryness earlier makes carbonization with little heated is bright, blazingly makes complete ashing at 500～600 ℃ again, puts coldly, and residue adds hydrochloric acid 5ml and water 23ml makes dissolving, as need testing solution; Get need testing solution, check that according to Chinese Pharmacopoeia arsenic salt inspection technique arsenic content must not cross 2/1000000ths.
(8) proteinic inspection: get 1ml of the present invention, add 30% sulfosalicylic acid test solution 1ml of new preparation, mixing was placed 5 minutes, must not become turbid.
(9) inspection of tannin: get 1ml of the present invention, check according to injection related substance inspection technique, should be up to specification.
(10) inspection of resin: get 5ml of the present invention, check according to injection related substance inspection technique, should be up to specification.
(11) inspection of oxalates: it is an amount of to get the present invention, regulates pH value to 1～2 with dilute hydrochloric acid, filters, and gets filtrate 2ml, regulates pH value to 5～6 with sodium hydroxide test solution, adds 3% calcium chloride solution 2-3 and drips, and places 10 minutes, must not become turbid or precipitate.
(12) inspection of potassium ion: precision is measured 2ml of the present invention, checks according to injection related substance inspection technique, should be up to specification.
(13) inspection of pyrogen: get the present invention, check according to the Chinese Pharmacopoeia pyrogen test, dosage should be up to specification by the every 1kg body weight injection of rabbit 3ml.
(14) inspection of n-butanol extract: precision is measured 25ml of the present invention, put in the separatory funnel, extract 3 times with the chloroform jolting, each 30ml, merge chloroform extraction liquid, wash with water 2 times, each 10ml, washing liquid is incorporated in the above-mentioned aqueous solution, extract 3 times with water saturated n-butyl alcohol jolting, each 30ml merges n-butanol extracting liquid, put in the evaporating dish that is dried to constant weight, evaporate to dryness 105 ℃ of dryings 3 hours, was put in the exsiccator cooling 30 minutes, claim rapidly to decide weight, the present invention contains n-butanol extract must not be less than 0.15%.
(15) assay of total polysaccharides: be taken at 80 ℃ of anhydrous glucose reference substance 80mg that are dried to constant weight, the accurate title, decide, put in the 100ml measuring bottle, with water dissolution and be diluted to scale, shake up, precision is measured 5ml, put in the 50ml measuring bottle, add water to scale, shake up, promptly get the reference substance solution that every 1ml contains anhydrous glucose 80 μ g; Get anthrone 50mg, add water 25ml and sulphuric acid 75ml makes dissolving, promptly get sulphuric acid anthrone solution; Precision is measured reference substance solution 0.2,0.4,0.6,0.8,1.0ml, puts respectively in the 10ml tool plug test tube, and precision adds water and makes into 1ml in each pipe, the accurate sulphuric acid anthrone solution 4ml that faces with preceding preparation that adds in ice bath, shake up, put in the boiling water bath and heated 10 minutes, cooled off 40 minutes; With the reagent corresponding is blank; According to the Chinese Pharmacopoeia ultraviolet visible spectrophotometry, measure trap at 625nm wavelength place, be that vertical coordinate, concentration are abscissa drawing standard curve with the trap; Precision is measured 50ml of the present invention, puts to be concentrated into about 5ml in the water-bath, puts coldly, is transferred in the 10ml measuring bottle, adds water to scale, shakes up; Precision is measured 1ml, the accurate dehydrated alcohol 10ml that adds, and close plug shakes up, and leaves standstill 4 hours, and is centrifugal, abandoning supernatant, precipitation is used water dissolution, is transferred in the 50ml measuring bottle, is diluted with water to scale, shakes up, precision is measured 5ml, puts in the 50ml measuring bottle, adds water to scale, shakes up, and promptly gets need testing solution; Precision is measured need testing solution 1ml, put in the 10ml tool plug test tube, the accurate sulphuric acid anthrone solution 4ml that adds in ice bath, shake up, put in the boiling water bath and heated 10 minutes, cooled off 40 minutes, measure trap at 625nm wavelength place, from the amount that standard curve is read polysaccharide the need testing solution, it is C with the molecular formula that the every 1ml of the present invention contains total polysaccharides
6H
12O
6The anhydrous glucose meter, must not be less than 3.5mg.
(16) assay of Venenum Bufonis: with the octadecylsilane chemically bonded silica is filler; With proportioning is that normal propyl alcohol-methanol-0.04% potassium dihydrogen phosphate of 2: 3: 7 is a mobile phase; The detection wavelength is 300nm; Number of theoretical plate calculates by the cinobufagin peak should be not less than 2500; Get the bufogenin reference substance respectively and the cinobufagin reference substance is an amount of, accurately claim surely, add methanol and make the mixed solution that every 1ml contains 40 μ g, promptly get reference substance solution; Precision is measured 20ml of the present invention, put in the separatory funnel, extract 4 times with the chloroform jolting, each 30ml, extracting solution filters with the filter paper that is covered with a small amount of anhydrous sodium sulfate, filtrate merges, evaporate to dryness, residue is with dissolve with methanol and be transferred in the 5ml measuring bottle, adds methanol to scale, shake up, promptly get need testing solution; According to the Chinese Pharmacopoeia high performance liquid chromatography, accurate respectively reference substance solution and each 10 μ l of need testing solution of drawing inject chromatograph of liquid, measure, and it is C with the molecular formula that the every 1ml of the present invention contains Venenum Bufonis
24H
32O
4Bufogenin and molecular formula be C
26H
34O
6The total amount meter of cinobufagin, should be 12～18 μ g.
(17) assay of Mylabris: with the methyl silicone rubber is fixative, and coating concentration is 3.5%; Column temperature is 165 ± 10 ℃; Number of theoretical plate calculates by the cantharidin peak should be not less than 2500; It is an amount of to get the cantharidin reference substance, and accurate the title decides, and adds chloroform and makes the solution that every 1ml contains 0.32mg, promptly gets reference substance solution; Precision is measured 50ml of the present invention, regulates pH value to 3～4 with 5% sulfuric acid solution, extracts 5 times with water saturated chloroform jolting, each 40ml, merge chloroform extraction liquid, filter through the filter paper that is covered with a small amount of anhydrous sodium sulfate, filtrate merges, put and evaporate chloroform in the water-bath below 60 ℃ to about 10ml, naturally wave again and be dissipated to 2～3ml, be transferred in the 5ml measuring bottle, add chloroform to scale, shake up, promptly get need testing solution; According to the Chinese Pharmacopoeia gas chromatography, accurate respectively reference substance solution 2 μ l and the need testing solution 4 μ l of drawing, inject gas chromatograph is measured, and it is C with the molecular formula that the every 1ml of the present invention contains Mylabris
10H
12O
4The cantharidin meter, should be 14～18 μ g.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2005100431342A CN1739598A (en) | 2005-08-23 | 2005-08-23 | Chinese medicine injection for treating cancer and its prepn process and quality control method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNA2005100431342A CN1739598A (en) | 2005-08-23 | 2005-08-23 | Chinese medicine injection for treating cancer and its prepn process and quality control method |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN1739598A true CN1739598A (en) | 2006-03-01 |
Family
ID=36092223
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNA2005100431342A Pending CN1739598A (en) | 2005-08-23 | 2005-08-23 | Chinese medicine injection for treating cancer and its prepn process and quality control method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1739598A (en) |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100386089C (en) * | 2006-03-02 | 2008-05-07 | 上海慈瑞医药科技有限公司 | Compound lentinan preparation and its preparing method |
| CN102085220A (en) * | 2010-11-02 | 2011-06-08 | 河北以岭医药研究院有限公司 | Thin-layer identification method with fluorescence intensifier detection for traditional Chinese medicines |
| CN102749414A (en) * | 2012-07-09 | 2012-10-24 | 新疆天山莲药业有限公司 | Quality control method for Uighur medicine spikenard strength replenishing preparation |
| CN103278466A (en) * | 2013-05-02 | 2013-09-04 | 广西北斗星动物保健品有限公司 | Quality control method for toad venom injection |
| CN105510257A (en) * | 2015-12-28 | 2016-04-20 | 上海景峰制药有限公司 | Method for measuring content of total sugar in tanshinol and ligustrazine glucose injection |
| CN105606556A (en) * | 2016-01-28 | 2016-05-25 | 广西壮族自治区中医药研究院 | Ultraviolet spectrophotometry for polysaccharide content in medicine material fokien angiopteris rhizomes |
| CN107085045A (en) * | 2017-04-06 | 2017-08-22 | 广西壮族自治区药用植物园 | The assay method of cantharidin in green side blister beetle |
| CN109668887A (en) * | 2019-02-26 | 2019-04-23 | 云南中医学院 | A kind of inspection method of trace protein of Chinese medicine injection |
| CN110702834A (en) * | 2019-09-27 | 2020-01-17 | 石家庄平安医院有限公司 | Non-interference rapid thin-layer identification method for kidney-tonifying toxin-expelling granules |
-
2005
- 2005-08-23 CN CNA2005100431342A patent/CN1739598A/en active Pending
Cited By (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100386089C (en) * | 2006-03-02 | 2008-05-07 | 上海慈瑞医药科技有限公司 | Compound lentinan preparation and its preparing method |
| CN102085220A (en) * | 2010-11-02 | 2011-06-08 | 河北以岭医药研究院有限公司 | Thin-layer identification method with fluorescence intensifier detection for traditional Chinese medicines |
| CN102085220B (en) * | 2010-11-02 | 2011-12-28 | 河北以岭医药研究院有限公司 | Thin-layer identification method with fluorescence intensifier detection for traditional Chinese medicines |
| CN102749414A (en) * | 2012-07-09 | 2012-10-24 | 新疆天山莲药业有限公司 | Quality control method for Uighur medicine spikenard strength replenishing preparation |
| CN102749414B (en) * | 2012-07-09 | 2015-01-07 | 新疆天山莲药业有限公司 | Quality control method for Uighur medicine spikenard strength replenishing preparation |
| CN103278466A (en) * | 2013-05-02 | 2013-09-04 | 广西北斗星动物保健品有限公司 | Quality control method for toad venom injection |
| CN105510257A (en) * | 2015-12-28 | 2016-04-20 | 上海景峰制药有限公司 | Method for measuring content of total sugar in tanshinol and ligustrazine glucose injection |
| CN105606556A (en) * | 2016-01-28 | 2016-05-25 | 广西壮族自治区中医药研究院 | Ultraviolet spectrophotometry for polysaccharide content in medicine material fokien angiopteris rhizomes |
| CN105606556B (en) * | 2016-01-28 | 2018-09-21 | 广西壮族自治区中医药研究院 | The ultraviolet spectrophotometry of polyoses content in fokien angiopteris rhizome medicinal material |
| CN107085045A (en) * | 2017-04-06 | 2017-08-22 | 广西壮族自治区药用植物园 | The assay method of cantharidin in green side blister beetle |
| CN109668887A (en) * | 2019-02-26 | 2019-04-23 | 云南中医学院 | A kind of inspection method of trace protein of Chinese medicine injection |
| CN110702834A (en) * | 2019-09-27 | 2020-01-17 | 石家庄平安医院有限公司 | Non-interference rapid thin-layer identification method for kidney-tonifying toxin-expelling granules |
| CN110702834B (en) * | 2019-09-27 | 2021-08-10 | 石家庄平安医院有限公司 | Non-interference rapid thin-layer identification method for kidney-tonifying toxin-expelling granules |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN1739598A (en) | Chinese medicine injection for treating cancer and its prepn process and quality control method | |
| US20110059124A1 (en) | The quality control method and application of a kind of ganoderma lucidum spore oil fat emulsion | |
| CN106937959B (en) | A kind of antitumor extra large Blatta seu periplaneta extract and its preparation technology and detection method and purposes | |
| CN100486586C (en) | Imature bitter orange total flavone and its preparing method for its preparation and quality control method | |
| CN103222988A (en) | Periplaneta americana extract and its preparation method and use | |
| CN113848278A (en) | Quality control method for standard decoction of radix Cudraniae | |
| CN1799605A (en) | 'Shengmai' infusion and its preparation process | |
| CN102028859A (en) | Quality control method for Chinese medicinal preparation for treating asthma | |
| CN106324161A (en) | Quality detection method for traditional Chinese medicine composition capable of treating diabetic nephropathy | |
| CN106072626A (en) | A kind of towards doctor's compound recipe antler health-care food and its production and use | |
| CN101829224A (en) | Traditional Chinese medicine composition preparation and preparation method and quality control method | |
| CN102488722B (en) | Preparation method and quality detection method of bupleurum oral solution | |
| CN102526656B (en) | Medicament or health-care food composition for preventing or/and treating erythremia | |
| CN101439083B (en) | Detection method of Chinese medicine soft capsules for clearing wind heat and clearing nasal passage | |
| CN101077372A (en) | Chinese medicinal materials composition for preventing and treating iron-deficiency anemia and method of making the same | |
| CN1324040C (en) | Medicament for treating diabetes, diabetes complication, its preparation and novel use | |
| CN101926929B (en) | Traditional Chinese medicine for treating vascular dementia and preparation method thereof | |
| CN1739682A (en) | Nudiflorous beautyberry soft capsule for body cavity and its prepn | |
| CN1823933A (en) | Ginseng aconite injection liquid and its preparation method | |
| CN1895396A (en) | Chinese-medicinal preparation with Yin-nourishing, intestin-moistening and bowls-relaxing functions | |
| CN107582639A (en) | A kind of vinegar corydalis tuber granule and preparation method thereof | |
| CN1323676C (en) | Method for preparing traditional Chinese medicine capsule for treating weakness of spleen and stomach, and its quality control method | |
| CN111214474A (en) | Application of Stichopus japonicus sterol sulfate in anti-atherosclerosis product | |
| CN1202832C (en) | High-activity ginkgo leaf extract preparation and application | |
| CN101053587A (en) | Medicinal composition for treating tumor and preparation method and quality control method thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |