CN102319268B - Preparation method of injection used for treating chronic congestive cardiac failure - Google Patents
Preparation method of injection used for treating chronic congestive cardiac failure Download PDFInfo
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- CN102319268B CN102319268B CN2011102300755A CN201110230075A CN102319268B CN 102319268 B CN102319268 B CN 102319268B CN 2011102300755 A CN2011102300755 A CN 2011102300755A CN 201110230075 A CN201110230075 A CN 201110230075A CN 102319268 B CN102319268 B CN 102319268B
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Abstract
The invention discloses an injection used for treating chronic congestive cardiac failure, and a preparation method thereof. The medicine composition comprises components of, by weight: 3% to 6% of cockroach extract, 10% to 30% of polyethylene glycol, 0.7% to 1.0% of sodium chloride, and balance of water for injection. The preparation method comprises steps that: first, the cockroach extract is prepared through steps that: dried cockroach polypides are crushed into powder, the powder is processed from ethanol refluxing, and the extract is concentrated with reduced pressure, such that a thick paste is obtained; purified water is added to the paste for carrying out an oil-water separation process; medicine in the water layer is collected, and is processed through active carbon absorption, packing, and n-butyl alcohol elution; the eluant is concentrated with reduced pressure, and is processed through a secondary ethanol extraction; the obtained filtrate is concentrated with reduced pressure, and is processed through water extraction, centrifugation, filtration, decoloration, ultrafiltration, and concentration with reduced pressure, such that the cockroach extract thick paste is obtained; then, the medicine injection is prepared through steps that: the cockroach extract, medicinal polyethylene glycol and medicinal sodium chloride are mixed according to the formulation; the pH value is regulated to 5.4-6.4; the mixture is diluted to 1000ml by using the water for injection; the obtained solution is filtered, encapsulated, sterilized, lamp inspected, and packaged, such that cockroach extract injections are obtained.
Description
Technical field
The invention belongs to biological pharmacy technical field, be specifically related to a kind of injection that chronic heart failure levies and preparation method thereof for the treatment of.
Background technology
Cockroach, having another name called periplaneta americana (being commonly called as Blatta seu periplaneta) is volume maximum in the Blattidae, becomes long 29～35 millimeters of polypide.The cockroach epidermis contains sclerotin and chitin, the elements such as bromine, zinc, nickel, manganese, potassium, calcium, titanium, chlorine, sulfur, silicon, aluminum, magnesium, the muscle hydrolysis can get 13 seed amino acids, the also in store plain B1 of little life, B2, nicotinic acid and ascorbic acid etc. in the health contain trehalose, trehalase, glycoprotein, inositol, protocatechuic acid Fructus Vitis viniferae glycoside etc. in the lymph; All contain ergothioneine, 1-methyl-2-pyridinium carboxylate, trigonelline, glycine, betanin, anus alkali, trimethylamine, adenine etc.Modern scientific research shows that cockroach extractive has very strong biological activity, but also there is significant limitation in prior art for the application of cockroach extractive, and the cockroach extractive application potential awaits further development and utilization.
Summary of the invention
The first purpose of the present invention is for the deficiencies in the prior art, and a kind of injection that chronic heart failure is levied for the treatment of is provided; The second purpose is to provide a kind of preparation method of this this injection.
The first purpose of the present invention is achieved in that
The cockroach alcohol extract that contains weight ratio 3～6% in the described injection, 10～30% Polyethylene Glycol and 0.7～1% sodium chloride, surplus is water for injection.
Its prioritization scheme contains the cockroach extractive of weight ratio 4～5% in the described injection, 15～20% Polyethylene Glycol and 0.8～1% sodium chloride, and surplus is water for injection.
Described Polyethylene Glycol is 400 type Polyethylene Glycol.
The second purpose of the present invention is achieved in that the preparation method of described injection comprises:
Cockroach alcohol extract preparation: the dried polypide of cockroach is ground into coarse powder, and through 90～95% ethanol percolate extraction of 5～10 times of weight 8～12 hours, extracting solution got thick paste through concentrating under reduced pressure; Adding 4～6 times of purified water fully stirs and leaves standstill and made oil-water separation in 12～24 hours again, remove precipitate and oils and fats, collect the water layer medicinal liquid, add medicinal carbon and leave standstill 1～2 hour absorption medicinal liquid to colourless, the dress post, the water-saturated n-butanol eluting, eluent adds 8～10 times of weight behind concentrating under reduced pressure 65～85% ethanol carry out the secondary alcohol extraction, cold preservation was filtered after 12～24 hours, filtrate is behind concentrating under reduced pressure, the purified water that adds again 8～10 times of weight portions is carried out water extraction, after cold preservation 12～24 hours, low-temperature centrifugation filtered to get water extraction liquid, and water extraction liquid is with the needle-use activated carbon decolouring of 0.1～0.5% weight ratio, the limit edged stirs, and after being heated to 60～80 ℃ static 0.5～1 hour; Concentrating under reduced pressure gets the cockroach extractive thick paste after film filter filtration, ultrafiltration;
The preparation of drug injection: by the compositions proportioning cockroach extractive, medical polyethylene glycol and medical sodium chloride are mixed, be diluted to 1000ml with water for injection, filter, embedding, sterilization, lamp inspection, packing namely get the injection of cockroach extractive.
Its prioritization scheme,
In the described cockroach extractive preparation, 90～95% alcohol dipping of 5～10 times of weight ratios of cockroach coarse powder adding 48～60 hours are done the slow diafiltration of solvent with 90～95% ethanol again and are little yellow to filtrate, and filtrate gets thick paste through concentrating under reduced pressure.
Temperature when centrifugal in the described cockroach extractive preparation is 2～6 ℃.
Refrigerated storage temperature is 4～8 ℃ in the described cockroach extractive preparation.
In the described cockroach extractive preparation, the concentrating under reduced pressure temperature is 60～80 ℃.
In the described cockroach extractive preparation, the relative density of thick paste is 1.2～1.3.
Cockroach extractive of the present invention and preparation thereof can promote myocardial cell Ca
2+Interior stream, gentleness increase myocardial contraction enduringly, and blood vessel dilating reduces pulmonary artery pressure, pulmonary capillary wedge; Coronary artery dilating, increase coronary flow, the myocardial damage of inhibition Mediated by Free Radicals; Nephrectasia blood vessel, increase renal blood flow, diuresis; Improve microcirculation; The correction neuroendocrine is unbalance.
Description of drawings
Accompanying drawing is the standard finger-print of injection of the present invention.
The specific embodiment
The present invention is further illustrated below in conjunction with embodiment, but never in any form the present invention is limited, and any conversion based on training centre of the present invention is done all belongs to protection scope of the present invention.
The cockroach alcohol extract that contains weight ratio 3～6% in the injection of the present invention, 10～30% Polyethylene Glycol and 0.7～1% sodium chloride, surplus is water for injection.
Its preferred implementation contains the cockroach extractive of weight ratio 4～5% in the described injection, 15～20% Polyethylene Glycol and 0.8～1% sodium chloride, and surplus is water for injection.
Described Polyethylene Glycol is 400 type Polyethylene Glycol.
The preparation method of injection of the present invention comprises:
Cockroach alcohol extract preparation: the dried polypide of cockroach is ground into coarse powder, and through 90～95% ethanol percolate extraction of 5～10 times of weight 8～12 hours, extracting solution got thick paste through concentrating under reduced pressure; Adding 4～6 times of purified water fully stirs and leaves standstill and made oil-water separation in 12～24 hours again, remove precipitate and oils and fats, collect the water layer medicinal liquid, add medicinal carbon and leave standstill 1～2 hour absorption medicinal liquid to colourless, the dress post, the water-saturated n-butanol eluting, eluent adds 8～10 times of weight behind concentrating under reduced pressure 65～85% ethanol carry out the secondary alcohol extraction, cold preservation was filtered after 12～24 hours, filtrate is behind concentrating under reduced pressure, the purified water that adds again 8～10 times of weight portions is carried out water extraction, after cold preservation 12～24 hours, low-temperature centrifugation filtered to get water extraction liquid, and water extraction liquid is with the needle-use activated carbon decolouring of 0.1～0.5% weight ratio, the limit edged stirs, and after being heated to 60～80 ℃ static 0.5～1 hour; Concentrating under reduced pressure gets the cockroach extractive thick paste after film filter filtration, ultrafiltration;
The preparation of drug injection: by the compositions proportioning cockroach extractive, medical polyethylene glycol and medical sodium chloride are mixed, be diluted to 1000ml with water for injection, filter, embedding, sterilization, lamp inspection, packing namely get the injection of cockroach extractive.
Its preferred implementation,
In the described cockroach extractive preparation, 90～95% alcohol dipping of 5～10 times of weight ratios of cockroach coarse powder adding 48～60 hours are done the slow diafiltration of solvent with 90～95% ethanol again and are little yellow to filtrate, and filtrate gets thick paste through concentrating under reduced pressure.
Temperature when centrifugal in the described cockroach extractive preparation is 2～6 ℃.
Refrigerated storage temperature is 4～8 ℃ in the described cockroach extractive preparation.
In the described cockroach extractive preparation, the concentrating under reduced pressure temperature is 60～80 ℃.
In the described cockroach extractive preparation, the relative density of thick paste is 1.2～1.3.
Embodiment 1
Get the dried polypide 1000g of cockroach and be ground into coarse powder, add 90% alcohol dipping 48 hours of 5 times of weight, again with 90% ethanol percolate extraction 8 hours, it is 1.2～1.3 thick paste that extracting solution gets relative density at 60 ℃ of lower concentrating under reduced pressure; Adding 4 times of purified water fully stirs and leaves standstill and made oil-water separation in 12 hours again, remove precipitate and oils and fats, collect the water layer medicinal liquid, add medicinal carbon and leave standstill 1 hour absorption medicinal liquid to colourless, the dress post, the water-saturated n-butanol eluting, eluent adds 8 times of weight behind concentrating under reduced pressure 65% ethanol carries out the secondary alcohol extraction, filters after 12 hours 4 ℃ of lower cold preservations, and filtrate is behind 60 ℃ of lower concentrating under reduced pressure, the purified water that adds again 8 times of weight portions is carried out water extraction, after 12 hours, get water extraction liquid 2 ℃ of lower low temperature centrifugal filtrations 4 ℃ of lower cold preservations, water extraction liquid decolours with the needle-use activated carbon of 0.1% weight ratio, the limit edged stirs, and after being heated to 60 ℃ static 0.5 hour; Through film filter filter, after the ultrafiltration concentrating under reduced pressure to get relative density be 1.2～1.3 cockroach extractive thick paste;
Cockroach alcohol extract by 3%, 10% Polyethylene Glycol and 0.7%% sodium chloride, surplus is that the ratio of water for injection cooperates, and through filtration, embedding, sterilization, lamp inspection, packing, namely gets described injection.
The dried polypide of cockroach is ground into coarse powder 1000g, adds 95% alcohol dipping 60 hours of 10 times of weight, and then with 95% reflux, extract, 12 hours, and it is 1.2～1.3 thick paste that extracting solution gets relative density at 80 ℃ of lower concentrating under reduced pressure; Adding 6 times of purified water fully stirs and leaves standstill and made oil-water separation in 24 hours again, remove precipitate and oils and fats, collect the water layer medicinal liquid, add medicinal carbon and leave standstill 2 hours absorption medicinal liquids to colourless, the dress post, the water-saturated n-butanol eluting, eluent adds 0 times of weight behind 80 ℃ of lower concentrating under reduced pressure 85% ethanol carries out the secondary alcohol extraction, filters after 24 hours 8 ℃ of lower cold preservations, and filtrate is behind 80 ℃ of lower concentrating under reduced pressure, the purified water that adds again 10 times of weight portions is carried out water extraction, after 24 hours, get water extraction liquid 6 ℃ of lower low temperature centrifugal filtrations 8 ℃ of lower cold preservations, water extraction liquid decolours with the needle-use activated carbon of 0.5% weight ratio, the limit edged stirs, and after being heated to 80 ℃ static 1 hour; Through film filter filter, after the ultrafiltration concentrating under reduced pressure to get relative density be 1.2～1.3 cockroach extractive thick paste;
Cockroach alcohol extract by 6%, 30% Polyethylene Glycol and 1.0% sodium chloride, surplus is that the ratio of water for injection cooperates, and through filtration, embedding, sterilization, lamp inspection, packing, namely gets described injection.
Embodiment 3
The dried polypide of cockroach is ground into coarse powder 1000g, and through 95% alcohol dipping of 8 times of weight 55 hours, and then with 95% reflux, extract, 10 hours, it is 1.2～1.3 thick paste that extracting solution gets relative density at 70 ℃ of lower concentrating under reduced pressure; Adding 5 times of purified water fully stirs and leaves standstill and made oil-water separation in 18 hours again, remove precipitate and oils and fats, collect the water layer medicinal liquid, add medicinal carbon and leave standstill 1.5 hours absorption medicinal liquids to colourless, the dress post, the water-saturated n-butanol eluting, eluent adds 9 times of weight behind 70 ℃ of lower concentrating under reduced pressure 75% ethanol carries out the secondary alcohol extraction, filters after 18 hours 6 ℃ of lower cold preservations, and filtrate is behind 70 ℃ of lower concentrating under reduced pressure, the purified water that adds again 9 times of weight portions is carried out water extraction, after 18 hours, get water extraction liquid 4 ℃ of lower low temperature centrifugal filtrations 6 ℃ of lower cold preservations, water extraction liquid decolours with the needle-use activated carbon of 0.3% weight ratio, the limit edged stirs, and after being heated to 70 ℃ static 0.7 hour; Through film filter filter, after the ultrafiltration concentrating under reduced pressure to get relative density be 1.2～1.3 cockroach extractive thick paste;
Cockroach alcohol extract by 5%, 20% Polyethylene Glycol and 0.8% sodium chloride, surplus is that the ratio of water for injection cooperates, and through filtration, embedding, sterilization, lamp inspection, packing, namely gets described injection.
Embodiment 4
With the cockroach extractive thick paste of preparation among the embodiment 1, the cockroach extractive by 4%, 15% Polyethylene Glycol and 0.8% sodium chloride, surplus is water for injection, through filtration, embedding, sterilization, lamp inspection, packing, namely gets described injection.
With the cockroach extractive thick paste of preparation among the embodiment 1, the cockroach extractive by 5%, 20% Polyethylene Glycol and 1.0% sodium chloride, surplus is water for injection, through filtration, embedding, sterilization, lamp inspection, packing, namely gets described injection.
Injection of the present invention is yellow clear liquid, by the following method injection and cockroach extractive is differentiated, is checked and measure:
1, differentiates
(1) thin layer chromatography test:
Get injection 1ml, thin up is to 5ml, as need testing solution.
Get cockroach dry product 1g, add n-butyl alcohol-glacial acetic acid (4:1) mixed liquor 50ml, supersound process was filtered after 30 minutes, and with the filtrate evaporate to dryness, residue adds water 1ml dissolving, as the medical material contrast solution.
According to thin layer chromatography (" an appendix VI of Chinese pharmacopoeia B) test, draw each 5 μ l of above-mentioned two kinds of solution, put respectively on same silica GF254 lamellae, launch with n-butyl alcohol-glacial acetic acid-water (4:1:1) mixed liquor, take out, dry, put under the ultra-violet lamp (254nm) and inspect, in the test sample chromatograph, with the corresponding position of control medicinal material solution chromatograph on, should show the speckle of 3 same colors.Spray the ninhydrin solution with 2%, it is clear to be heated to the speckle colour developing under 105 ℃ again, with the corresponding position of control medicinal material chromatograph on, should show the speckle of 1 same color.
(2) high performance liquid chromatography test:
Get injection 1ml, thin up is to 5ml, by strong alkalinity anion exchange glucosan chromatographic column (QAE-Sephadex A-25 type, internal diameter 1.5cm, the high 5cm of post), use the 20ml water elution, discard water elution liquid, use again 0.02mol/L hydrochloric acid eluting, discard just eluent 8ml, collect continuous eluent 20ml, as need testing solution.
According to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia D) test, take octadecylsilane chemically bonded silica as filler, chromatographic column (column length 150mm, column internal diameter are 4.6mm); So that 0.05mol/L sodium-acetate buffer (pH 5.0)-methanol (90:10) mixed liquor is as mobile phase, (DAD) detects with diode array detector, and the detection wavelength is 246nm, and flow velocity is per minute 0.5ml, and column temperature is 30 ℃.The accurate need testing solution 5 μ l that draw, the injection liquid chromatography is measured, and records the chromatogram in 12 minutes.The DAD ultraviolet detection spectrogram of main peak in the test sample chromatogram (maximum peak area) has absorption maximum at the wavelength place of 246nm and 286nm.
2, check
Color and luster: get this product 1ml, add water to 10ml, shake up, according to " an appendix XI of Chinese pharmacopoeia A first method checks, compares with yellow No. 8 standard color solutions, must not be darker.
PH value: should be 5.4～6.4(" an appendix VII of Chinese pharmacopoeia G).
Protein: get injection 1ml, add freshly prepared 30% sulfosalicylic acid solution 1ml, mixing was placed 5 minutes, muddiness must not occur.
Resin: get injection 5ml, add 1 of hydrochloric acid, placed 30 minutes, should separate out without the resin-like thing.
Oxalates: get injection 2ml, add 2～3 of 3% calcium chloride test solutions, placed 10 minutes, muddiness or precipitation must not occur.
Potassium ion: get injection 2ml, burn to carbonization with little heated first, 500～600 ℃ of blazing extremely fully ashing, add 6% acetic acid and make dissolving again.Put in the 25ml measuring bottle, thin up is to scale, mixing, measure 1ml and place the 10ml nessler colorimetric tube, add 12 of alkaline formaldehyde solution (formalin is transferred pH to 8.0～9.0 with the 0.1mol/L sodium hydroxide solution), 2 of 3% editic acid sodium solutions, 3% tetraphenyl borate sodium solution 0.5ml adds water to 10ml as the test sample pipe.Get an amount of porphyrize of potassium sulfate, be dried to constant weight in 110 ℃, precision takes by weighing 2.330g, adds water and makes in right amount and be dissolved into 1000ml, and the accurate 10ml thin up of drawing becomes 100ml, namely gets standard potassium ion solution; The accurate potassium ion solution of label taking (100 μ g/ml) 0.8ml places the 10ml nessler colorimetric tube, and the same operation is QC in contrast, gets the test sample pipe and contrasts QC and estimate than turbid, and the test sample pipe must not be more muddy.
Heavy metal: get injection 2ml, according to " an appendix IX of Chinese pharmacopoeia E the second method inspection contains heavy metal and must not cross 5/1000000ths.
Arsenic salt: get injection 1.0ml, slowly heat carbonization, incomplete such as carbonization, add a small amount of sulphuric acid moistening after, slowly heating makes complete carbonization, again in 500～600 ℃ of blazing ashing, let cool, add hydrochloric acid 5ml, add water 23ml, according to " an appendix IX of Chinese pharmacopoeia F first method checks, should be up to specification 0.0002%.
Pyrogen: get injection according to " an appendix X of Chinese pharmacopoeia III A checks that dosage is by the every kg injection of rabbit body weight 1ml, and its result should be up to specification.
Undue toxicity: get injection, add sodium chloride injection among every 1ml and be diluted to 10ml, according to " two appendix XI of Chinese pharmacopoeia C checks that press intravenous administration, its result should be up to specification.
3, hypersensitive test
The test liquid preparation: get injection, every 1ml adds the chlorination sodium injection and is diluted to 4ml as test sample.
Get 6 of body weight 250～350g healthy guinea pigs, the next day every each lumbar injection test sample 0.5ml, totally 3 times, carry out sensitization.Then it is divided into 2 groups, 3 every group, injecting rear 14 days and 21 days first respectively, 1ml attacks by the intravenous injection test sample.In rear 30 minutes of injection, anaphylaxis must not appear.If any perpendicular hair, sneeze, retch, continuously cough 3 and the phenomenon such as dyspnea in more than 2 kinds or 2 kinds, or have a convulsion, the one of shock, the phenomena of mortality, judge that then test sample is against regulation.
Haemolysis and agglutination test:
Prepare first 2% red blood cell suspension.Get a Sanguis Leporis seu oryctolagi number milliliter, put into the conical flask that fills bead, Fibrinogen is removed in jolting 10 minutes, makes into defibrinated blood, the physiological sodium chloride solution that adds 10 times of amounts, shake up, centrifugal, remove supernatant, the regeneration of erythrocytes reason sodium chloride solution washing of precipitation 2～3 times is till the not aobvious redness of supernatant; The erythrocyte of gained is mixed with 2% red blood cell suspension with physiological sodium chloride solution, and get final product.
| The test tube numbering | 1 | 2 | 3 | 4 | 5 |
| 2% red blood cell suspension (ml) | 2.5 | 2.5 | 2.5 | 2.5 | 2.5 |
| Physiological sodium chloride solution (ml) | 2.2 | 2.2 | 2.2 | 2.5 | - |
| Distilled water (ml) | - | - | - | - | 2.5 |
| For examination injection (ml) | 0.3 | 0.3 | 0.3 | 0.0 | 0.0 |
By upper method inspection, in 3 hours haemolysis and red blood cell condensation must not appear for the examination injection.
Finger printing: get injection 1ml, thin up is to 5ml, by strong-base anion-exchange resin chromatographic column (Dowex2, Cl-type, 200 orders, internal diameter 1.5cm, high 2.5cm, wet method dress post, chromatographic column pretreatment adopt 0.5mol/L sodium hydroxide solution 10ml eluting, again water 10ml eluting, for subsequent use), water 10ml eluting discards water elution liquid, use again 0.25mol/L acetum eluting, discard just eluent 5ml, collect continuous eluent 25ml, shake up, as need testing solution.It is an amount of to get the inosine reference substance, accurately weighed, adds water and is mixed with the solution that every 1ml contains inosine 50 μ g, in contrast product solution.According to " high effective liquid chromatography for measuring of an appendix VI of Chinese pharmacopoeia D, take octadecylsilane chemically bonded silica as filler, chromatographic column (column length 150mm, column internal diameter are 4.6mm); Gradient elution, take 0.05 mol/L sodium-acetate buffer (pH 5.0) as mobile phase A, take methanol as Mobile phase B, the detection wavelength is 254nm, and flow velocity is per minute 0.6ml, and column temperature is 30 ℃.Number of theoretical plate calculates by the inosine peak should be not less than 5000.
| Time (minute) | Mobile phase A (%) | Mobile phase B (%) |
| 0～5 | 90 | 10 |
| 5～15 | 90→74 | 10→26 |
| 15～18 | 74 | 26 |
Precision is drawn reference substance solution and each 5 μ l of need testing solution respectively, and the injection liquid chromatography records the chromatogram in 25 minutes.The test sample chromatogram should be basically identical with standard finger-print.The test sample chromatogram is imported similarity evaluation (2004,1.0B version), compare with standard finger-print, calculate similarity.Similarity should be not less than 0.90.
4, effective component content is measured in the cockroach extractive
The mensuration of compound nucleoside base: precision is measured cockroach extractive 1ml, puts in the 50ml measuring bottle, with the dissolving of glycine-hydrochloride buffer and be diluted to scale, shake up, precision is measured 5ml and is put in the 100ml measuring bottle, adds above-mentioned buffer to scale, shake up, namely get need testing solution.According to ultraviolet visible spectrophotometry (" appendix VA of Chinese pharmacopoeia), measure trap at 254nm wavelength place, be calculated as follows the compound nucleoside base contents C(mg of every ml cockroach extractive injection)
?
In the formula: A is the need testing solution trap
490 when being 1% for compound nucleoside base class concentration in the cockroach extractive at the average absorption degree at 254nm wavelength place.
N is extension rate
Measurement result is to contain compound nucleoside base among the every 1ml of cockroach extractive to should be 10.0mg～18.0mg.
In conjunction with amino acid whose mensuration:
The preparation standard curve.Precision takes by weighing glutamic acid reference substance 10mg, adds the heat of solution of about 160ml water temperature, puts in the 200ml volumetric flask, adds water to scale, shakes up, and namely gets the reference substance solution that contains 0.05mg among every 1ml.Get 6 in tool plug test tube, accurate reference substance solution 0.2,0.4,0.6,0.8, the 1.0ml of adding adds water to respectively 1.0ml respectively, respectively adds 0.2mol/L citrate buffer solution (pH5.0) 1.0ml take 1.0ml water as blank.Ninhydrin reagent 1.0ml shakes up, and heating is 15 minutes in 100 ℃ of water-baths, take out, use water cooling, placed 5～10 minutes, each adds 60% ethanol 3.0ml, shakes up, and measures trap at 570nm wavelength place according to ultraviolet visible spectrophotometry (" an appendix V of Chinese pharmacopoeia A).Take aminoglutaric acid concentration as abscissa, trap is vertical coordinate drawing standard curve.
The preparation need testing solution.Precision is measured cockroach extractive 1.0ml, in the 50ml measuring bottle, adds water to scale, shakes up, and is the test sample diluent.Precision is measured 1.0ml test sample diluent, put in the sky ampoule, add 1.0ml hydrochloric acid, sealing was in 110 ℃ of hydrolysis 8 hours, taking-up lets cool, all forward content to evaporating dish mid-boiling water bath evaporate to dryness, add water and make in right amount in dissolving and the immigration 10ml measuring bottle, add water to scale, shake up, be the test sample hydrolyzed solution.
Measure.Precision is measured test sample hydrolyzed solution 0.1ml, adds water to 1.0ml, places test tube A, is test sample hydrolyzed solution not; Getting test sample hydrolyzed solution 1.0ml places test tube B many.Take 1.0ml water as blank, according to the method under the above-mentioned standard curve preparation from " respectively adding 0.2mol/L citrate buffer solution (pH5.0) 1.0ml ", measure respectively the trap of not hydrolysis and hydrolyzed solution, find its amino acid concentration from standard curve, calculate, namely get every gram cockroach extractive Determination of Free Amino Acids (A) and total amino acids content (B), by (B-A) namely get in every gram cockroach extractive in conjunction with amino acid content.
The every 1ml of this product contains in conjunction with aminoacid with glutamic acid (C
5H
9NO
4) meter, should be 15.0 mg～25.0mg.
The mensuration of uracil, hypoxanthine and inosine:
According to " the high effective liquid chromatography for measuring of an appendix VI of Chinese pharmacopoeia D.
Chromatographic condition and system suitability.Take octadecylsilane chemically bonded silica as filler.Mobile phase A is 0.05mol/L sodium-acetate buffer (pH5.0); Mobile phase B is methanol-mobile phase A (80:20).According to the form below carries out gradient elution; The detection wavelength is 254nm, and flow velocity is 0.5ml/min, and number of theoretical plate calculates by uracil, hypoxanthine and inosine peak respectively all should be not less than 3000.Separating degree between uracil, hypoxanthine and the inosine should meet the requirements.
| Time (minute) | Mobile phase A (%) | Mobile phase B (%) |
| 0～5 | 90 | 10 |
| 5～15 | 90→70 | 10→30 |
| 15～18 | 70 | 30 |
The preparation of reference substance solution.It is an amount of that precision takes by weighing uracil, hypoxanthine and inosine reference substance, adds mobile phase A and make and contain the solution that uracil, hypoxanthine and inosine are respectively 30 μ g, 40 μ g, 70 μ g among every 1ml, shakes up, and get final product.
The preparation of need testing solution.Precision is measured cockroach extractive 1ml, puts in the 50ml measuring bottle, is diluted to scale with mobile phase A, shakes up, and get final product.
Measure.Precision is drawn reference substance solution and each 10 μ l of need testing solution respectively, and the injection liquid chromatography is measured, and be get final product.
Contain uracil (C among the every ml of cockroach extractive
4H
4N
2O
2) should be 1.0mg～1.8mg, hypoxanthine (C
5H
5N
4O) should be 1.5mg～2.5mg, inosine (C
10H
12N
4O
5) should be 3.5mg～5.0mg.
The results of pharmacodynamic test of pharmaceutical preparation of the present invention:
Gained preparation of the present invention has benefiting QI for activating blood circulation, activating YANG for promoting diuresis.Be used for gas sun two void, cardiopalmus, tachypnea, edema, dim complexion, lip cyanosis due to the blood-stasis internal-depression; Chronic heart failure, chronic pulmonary heart disease are seen above-mentioned patient.Below carry out the summary summary of II clinical trial phase in Shanghai Univ. of Traditional Chinese Medicine clinical pharmacology base for this injection: according to the II phase clinical research plan of this injection for treating congestive heart failure, press each 100 example of 1:1 randomized controlled trial, open trial 208 examples, total 408 examples.Result of the test is: treatment group obvious effective rate 37%, effective percentage 89%; Open trial group obvious effective rate 55.29%, effective percentage 87.5%, treatment group and matched group (dobutamine) compare, statistics, and significant difference (P ﹤ 0.01) is arranged.This injection to right heart failure, decline whole-heartedly, the curative effect of cardiac function II level and III level is obvious, all is better than the dobutamine matched group.Two groups of cardiac function onset time no difference of science of statistics (P＞0.05), treatment group cardiac function onset time is 3.31 days.Treatment group is respectively 93.35% and 88.46% to Qi deficiency blood stasis type and motive YIN-deficiency type curative effect, and there were significant differences (P＜0.01 and P＜0.05) with matched group, and treatment group has obvious curative effects to breathing, heart rate and edema.Heart rate onset time comparison is inhaled in two group callings, and significant difference (P＜0.01 and P＜0.05) is all arranged.Traditional Chinese medical science disease integral contrast, treatment group is improved significantly better than matched group (P＜0.01) cardiopalmus, tachypnea, weak and fear of cold integration.Electrocardiogram, rabat, blood pressure, two groups of no difference of science of statistics, security inspection has no obvious adverse reaction.Result of the test shows that this injection can improve cardiac function, alleviates the diseases such as cardiopalmus, tachypnea, edema, and is obvious to Qi deficiency blood stasis type and motive YIN-deficiency type curative effect.This injection for treating congestive heart failure curative effect is reliable, uses safety.
Claims (6)
1. treat the injection preparation method that chronic heart failure is levied for one kind, contain the cockroach alcohol extract of weight ratio 3～6% in the described injection, 10～30% Polyethylene Glycol and 0.7～1% sodium chloride, surplus is water for injection; Described Polyethylene Glycol is 400 type Polyethylene Glycol; The preparation method that it is characterized in that described injection comprises:
Cockroach alcohol extract preparation: the dried polypide of cockroach is ground into coarse powder, and through 90～95% ethanol percolate extraction of 5～10 times of weight 8～12 hours, extracting solution got thick paste through concentrating under reduced pressure; Adding 4～6 times of purified water fully stirs and leaves standstill and made oil-water separation in 12～24 hours again, remove precipitate and oils and fats, collect the water layer medicinal liquid, add medicinal carbon and leave standstill 1～2 hour absorption medicinal liquid to colourless, the dress post, the water-saturated n-butanol eluting, eluent adds 8～10 times of weight behind concentrating under reduced pressure 65～85% ethanol carry out the secondary alcohol extraction, cold preservation was filtered after 12～24 hours, filtrate is behind concentrating under reduced pressure, the purified water that adds again 8～10 times of weight portions is carried out water extraction, after cold preservation 12～24 hours, low-temperature centrifugation filtered to get water extraction liquid, and water extraction liquid is with the needle-use activated carbon decolouring of 0.1～0.5% weight ratio, the limit edged stirs, and after being heated to 60～80 ℃ static 0.5～1 hour; Concentrating under reduced pressure gets the cockroach extractive thick paste after film filter filtration, ultrafiltration;
The preparation of drug injection: by the compositions proportioning cockroach extractive, medical polyethylene glycol and medical sodium chloride are mixed, be diluted to 1000ml with water for injection, filter, embedding, sterilization, lamp inspection, packing namely get the injection of cockroach extractive.
2. the preparation method of injection according to claim 1, it is characterized in that: in the described cockroach extractive preparation, 90～95% alcohol dipping of 5～10 times of weight ratios of cockroach coarse powder adding 48～60 hours, do the slow diafiltration of solvent with 90～95% ethanol again and be little yellow to filtrate, filtrate gets thick paste through concentrating under reduced pressure.
3. the preparation method of injection according to claim 1 is characterized in that: the temperature when centrifugal in the described cockroach extractive preparation is 2～6 ℃.
4. the preparation method of injection according to claim 1 is characterized in that: refrigerated storage temperature is 4～8 ℃ in the described cockroach extractive preparation.
5. the preparation method of injection according to claim 1 is characterized in that: in the described cockroach extractive preparation, the concentrating under reduced pressure temperature is 60～80 ℃.
6. the preparation method of injection according to claim 1 is characterized in that: in the described cockroach extractive preparation, the relative density of thick paste is 1.2～1.3.
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| CN105832659A (en) * | 2016-04-29 | 2016-08-10 | 芜湖杨燕制药有限公司 | Production technology of traditional Chinese medicine injection |
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| CN1124141A (en) * | 1994-12-09 | 1996-06-12 | 大理医学院 | Xinmailong extract and Xinmailong medical preparation for curing angiocardiopath |
| CN101019893A (en) * | 2007-03-20 | 2007-08-22 | 大理学院 | Process of enriching effective antitumor part of periplaneta americana with polyamide |
| CN101057872A (en) * | 2007-03-20 | 2007-10-24 | 大理学院 | Anti tumor active part of American cockroach prepared with reverse phase material and medical application |
| CN101156875A (en) * | 2007-04-25 | 2008-04-09 | 深圳信立泰药业股份有限公司 | Cockroach extractive for treating cardiovascular disease, preparation method and its composition |
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Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1124141A (en) * | 1994-12-09 | 1996-06-12 | 大理医学院 | Xinmailong extract and Xinmailong medical preparation for curing angiocardiopath |
| CN101019893A (en) * | 2007-03-20 | 2007-08-22 | 大理学院 | Process of enriching effective antitumor part of periplaneta americana with polyamide |
| CN101057872A (en) * | 2007-03-20 | 2007-10-24 | 大理学院 | Anti tumor active part of American cockroach prepared with reverse phase material and medical application |
| CN101156875A (en) * | 2007-04-25 | 2008-04-09 | 深圳信立泰药业股份有限公司 | Cockroach extractive for treating cardiovascular disease, preparation method and its composition |
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