CN102879495A - Antidotal capsule of Tibetan medicine compound and quality detection method of preparation of antidotal capsule - Google Patents

Antidotal capsule of Tibetan medicine compound and quality detection method of preparation of antidotal capsule Download PDF

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CN102879495A
CN102879495A CN2012103655078A CN201210365507A CN102879495A CN 102879495 A CN102879495 A CN 102879495A CN 2012103655078 A CN2012103655078 A CN 2012103655078A CN 201210365507 A CN201210365507 A CN 201210365507A CN 102879495 A CN102879495 A CN 102879495A
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CN102879495B (en
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马振元
陈芹利
任松鹏
王海苹
董玉波
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JINHE TIBETAN MEDICINE CO., LTD.
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Shandong Arura Pharmaceutical Research & Development Co Ltd
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Abstract

The invention discloses an antidotal capsule of a Tibetan medicine compound and a quality detection method of a preparation of the antidotal capsule. Content of strychnine of semen strychni is controlled by a high performance liquid chromatography (HPLC) process; iron content in iron powder is controlled by means of titration; cholic acid in calculus bovis artifactus and hydroxy saffionin A in safflower are detected quantitatively by the high performance liquid liquid-phase chromatography; so that efficacy is guaranteed while hidden dangers to health of patients when the patients take excessive antidotal capsules are avoided, and product quality is improved, product safety, stability and quality controllability are guaranteed.

Description

The quality determining method of a kind of Tibetan medicinal composition detoxicating capsule and preparation thereof
Technical field
The present invention relates to a kind of quality determining method of Tibetan medicinal composition preparation, particularly the quality determining method of a kind of Tibetan medicinal composition detoxicating capsule and preparation thereof.
Background technology
Detoxicating capsule comes from Tibetan medicine's ancient books and records " sweet dew storehouse ", is the good recipe that the Tibetan medicine treats various nosotoxicosis.Record in Chinese patent drug provincial standard rising national standard part, standard number: WS-10537(ZD-0537)-2002.Detoxicating capsule is comprised of 35 flavor medicines such as vomiting nut (system), safflower, iron powder (system), calculus bovis factitius, Phytolacca acinosa, dry and soft, tsaoko, hooker winghead roots, has heat reliving and toxin-eliminating, the effect of putrefaction-removing granulation-promoting.Be used for various toxications, outmoded pyreticosis, some contact dermatitis etc.
Detoxicating capsule contains Strychnos nux-vomica subclass toxicity medicinal material, and its toxic component is mainly the alkaloids composition, and have and remove obstruction in channels to relieve pain, the effect of mass dissipating and swelling eliminating, but overdose may cause poisoning, even various bad reactions such as dead should not too much be taken or clothes of a specified duration.Contain iron powder in the prescription simultaneously, " Kaibao Bencao " record iron powder has tranquilizing mind, hard marrow, the effects such as profit skin.Iron is the trace element of human body, has the vital role such as the red blood cell of promotion generation.Used iron powder is to be used as medicine after concocting in Tibetan medicinal composition detoxicating capsule and the preparation prescription thereof, more is conducive to absorb.But the iron Excessive Intake may cause acute iron poisoning, is detrimental to health.
Detoxicating capsule proper mass standard only has the discriminating of cholic acid thin layer, gallic acid content to measure item, and to vomiting nut Poisoning composition alkaloid, iron content does not carry out quality control, makes said preparation have potential safety hazard.
Summary of the invention
For the deficiencies in the prior art, the purpose of this invention is to provide the quality determining method of a kind of Tibetan medicinal composition detoxicating capsule and preparation thereof.
Summary of the invention
The invention provides the control method of iron content in the peace strychnia, iron powder of vomiting nut sergeant in a kind of Tibetan medicinal composition detoxicating capsule and the preparation thereof; the content assaying method of hydroxyl radical carthamin yellow carthamus A in cholic acid, the safflower in the calculus bovis factitius; guarantee that strychnine, strychnia and iron content are all inexcessive; make said preparation in the treatment disease, guaranteed the safe and effective of medication.
The term explanation:
Detoxicating capsule is the nomenclature of drug of national standard for traditional Chinese medicines compilation (Chinese patent drug provincial standard rising national standard part) record.
Detoxicating capsule and preparation thereof comprise with other preparations of detoxicating capsule bulk drug prescription preparation, comprise detoxicating capsule.The proportioning of each bulk drug can be identical with the detoxicating capsule proportioning, also can suitably adjust in the drug effect scope.
Vomiting nut (system), borax (system), pearl (system), lazurite (system), turquoise (system), iron powder (system), sound copper (system), parmelia saxatilis (system), it is the routine techniques term of Tibetan medicine, " system " in the bracket all concocted according to the inner method of " Qinghai Province's Tibetan medicine concocted specification " (food and medicine Surveillance Authority in Qinghai Province's compiles, the versions in 2010 that the Qinghai People's Press publishes).
For achieving the above object, the present invention adopts following technical proposals:
The quality determining method of a kind of Tibetan medicinal composition detoxicating capsule and preparation thereof; the bulk drug of said preparation consists of the animal treasured; Radix Brassicae rapae; earthworm fruit mustard; vomiting nut (system); German tamarisk; borax (system); pearl (system); coral; lazurite (system); turquoise (system); iron powder (system); ring copper (system); parmelia saxatilis (system); ichor cream; nutmeg; tsaoko; Tabasheer; safflower; cloves; the myrobalan; Phytolacca acinosa; santal; santal; Moschus; calculus bovis factitius; whorlleaf oxytropis herb; grass-leaved sweetflag; cardamom; hooker winghead root; west szechwan pyrethrum; the rhizome of davallia; rhizoma nardostachyos; Corallodiscus flabellatus (Franch.) Burtt; alpine gentian herb; muscicolous woodbetony root; it is characterized in that, quality determining method comprises one or more in the following content assaying method:
A. the assay of vomiting nut: get respectively the strychnine reference substance, the strychnia reference substance is contrast, octadecylsilane chemically bonded silica is filling agent; Take volume parts than the acetonitrile of 15-25:85-75-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; Detecting wavelength is the high performance liquid chromatography of 250-270nm;
B. the assay of iron powder: after Chinese medicine preparation ashing to be determined, with the concentrated hydrochloric acid dissolving, add the methyl orange indicator solution, drip stannous chloride solution with Fe 3+Be reduced to Fe 2+, solution becomes pale red, then take diphenylamine sulfonic acid sodium salt as indicator, is titrated to solution with the potassium dichromate standard solution and is purple and is terminal point, calculates the content of iron powder with the amount of known potassium dichromate standard solution;
C. the assay of safflower: get hydroxyl safflower anthocyanidin A reference substance and add volume parts and be mixed with reference substance solution than 25% methanol aqueous solution, get preparation adding volume parts to be detected and after ultrasonic dissolution, removal of impurities, be mixed with need testing solution than 25% methanol aqueous solution, take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 20-30:80-70 than 0.5% aqueous formic acid as mobile phase, detecting wavelength is the high performance liquid chromatography of 400-410nm;
D. the assay of calculus bovis factitius: get the cholic acid reference substance and add methyl alcohol and be mixed with reference substance solution, get preparation to be detected and add methyl alcohol and after ultrasonic dissolution, removal of impurities, be mixed with need testing solution, take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 80-70:20-30 than 1% glacial acetic acid aqueous solution as mobile phase, the high performance liquid chromatography of evaporative light-scattering detector.
As a further improvement on the present invention, the assay concrete steps of described vomiting nut are:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 15-25:85-75-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; The detection wavelength is 250-270nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g);
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 10-14g, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 5-10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
As a further improvement on the present invention, the content assaying method concrete steps of described iron powder are:
Get Tibetan medicinal composition detoxicating capsule content or preparation powder 6 ~ 10g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 0.5-2ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, in 500-700 ℃ of blazing 3-5 hour, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, at 60-70 ℃ of left and right sides heating for dissolving 3-5 hour, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 0-15ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 2-10ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.
As a further improvement on the present invention, the concrete steps of described safflower are:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 20-30:80-70 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 400-410nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 6-10 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 4-6g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 20-30ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 5-10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
As a further improvement on the present invention, the concrete steps of described calculus bovis factitius assay are:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 80-70:20-30 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.4-0.5mg, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 6-8g, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 40-60ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 20-30ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
Bulk drug formed below the present invention was preferred detoxicating capsule and preparation thereof: precious 4.0 weight portions of animal; Radix Brassicae rapae 4.0 weight portions; earthworm fruit mustard 30.4 weight portions; vomiting nut (system) 1.6 weight portions; German tamarisk 30.4 weight portions; borax (system) 0.8 weight portion; pearl (system) 3.2 weight portions; coral 1.6 weight portions; lazurite (system) 1.6 weight portions; turquoise (system) 1.6 weight portions; iron powder (system) 2.4 weight portions; ring copper (system) 0.8 weight portion; parmelia saxatilis (system) 0.8 weight portion; ichor cream 4.0 weight portions; nutmeg 1.6 weight portions; tsaoko 1.6 weight portions; Tabasheer 1.6 weight portions; safflower 1.6 weight portions; cloves 1.6 weight portions; myrobalan's 2.4 weight portions; Phytolacca acinosa 1.6 weight portions; santal 30.4 weight portions; santal 27.9 weight portions; Moschus 1.6 weight portions; calculus bovis factitius 2.4 weight portions; whorlleaf oxytropis herb 30.4 weight portions; grass-leaved sweetflag 1.6 weight portions; cardamom 1.6 weight portions; hooker winghead root 30.4 weight portions; west szechwan pyrethrum 30.4 weight portions; the rhizome of davallia 3.2 weight portions; rhizoma nardostachyos 1.6 weight portions; Corallodiscus flabellatus (Franch.) Burtt 30.4 weight portions; alpine gentian herb 30.4 weight portions; muscicolous woodbetony root 30.4 weight portions.
For above-mentioned preferred formulation, described quality determining method is:
A. the assay of vomiting nut:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 20:80-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; The detection wavelength is 260nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g);
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 12g, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
B. the assay of iron powder:
Get Tibetan medicinal composition detoxicating capsule content or preparation powder 8g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, 65 ℃ of left and right sides heating for dissolving 4 hours, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.
C. the assay of safflower:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 25:75 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 403nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 5g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 25ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
D. the assay of calculus bovis factitius:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 76:24 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 7g, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 50ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 25ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
In the quality determining method to preferred formulation, vomiting nut content is with strychnine (C in the described vomiting nut content 31H 22N 2O 2) meter, should be 0.043mg/g-0.080mg/g; With strychnia (C 23H 26N 2O 1) meter, must not be less than 0.028mg/g.Iron content is 0.409mg/g-0.682mg/g in the assay of described iron powder.Safflower content was with hydroxyl radical carthamin yellow carthamus A (C during described safflower content was measured 27H 30O 15) meter, must not be less than 0.027mg/g.Calculus bovis factitius content is with cholic acid in the described calculus bovis factitius assay
(C 24H 40O 5) meter, must not be less than 0.252mg/g.
The unit corresponding relation of the weight portion described in this instructions and parts by volume is g/ml or kg/l.
Safe and effective for medication in order to guarantee the patient, primary standard is demanded urgently and need to be improved, the toxic component alkaloid that vomiting nut in the detoxicating capsule is contained carries out content control, and the content of iron in the iron powder is controlled comprehensively, increases simultaneously the assay of effective constituent cholic acid, hydroxyl radical carthamin yellow carthamus A.
The invention discloses the quality determining method of a kind of Tibetan medicinal composition detoxicating capsule and preparation thereof; iron content in vomiting nut sergeant's peace strychnia content, the iron powder has been carried out quality control; hydroxyl radical carthamin yellow carthamus A in cholic acid, the safflower in the calculus bovis factitius has been carried out quantitative detection; not only guaranteed drug effect but also guaranteed to be unlikely to take too much to cause healthy hidden danger; improve product quality, guaranteed product safety, stable, quality controllable.This law also can be used for other preparations of Tibetan medicinal composition detoxicating capsule simultaneously, such as detoxification particles, detoxifying pills, Jiedu Tablets etc.
Following experimental example and embodiment are used for further specifying but are not limited to the present invention.
Experimental example 1: the assay experiment of vomiting nut
1. instrument, reagent and test sample
Instrument: L-2100 type Hitachi high performance liquid chromatograph, AuW220D type Shimadzu electronic analytical balance.
Reference substance: strychnine reference substance (Nat'l Pharmaceutical ﹠ Biological Products Control Institute), lot number: 110705-200306; Strychnia reference substance (Nat'l Pharmaceutical ﹠ Biological Products Control Institute), lot number: 110706-200505.
Sample: detoxicating capsule (the Qinghai gold is scolded Tibetan medicine medicine company incorporated company and provided), lot number: 20100701,20100702,20100703.
2. detect the selection of wavelength
Get strychnine, strychnia mixing reference substance solution, scan in the 190-400nm wavelength coverage, according to ultraviolet absorpting spectrum, selected 260nm is for detecting wavelength.
3. mobile phase is selected
Selecting acetonitrile-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) is mobile phase.Investigated respectively acetonitrile-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than the 10% phosphorus acid for adjusting pH value 2.8) separating effect when the different volumes ratio is mobile phase, volume parts all can reach better separating effect during for mobile phase than the acetonitrile of 15-25:85-75-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8), and acetonitrile-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) volume parts is than for 20:80 the time, strychnine peak retention time is 21min, strychnia peak retention time is 17min, and separating effect is best.
4. system suitability and negative interference test
Under above-mentioned chromatographic condition, the accurate absorption mixed contrast solution, need testing solution, each 10 μ l of negative control solution, injection liquid chromatography, record chromatogram respectively.The result shows, mixes strychnine, strychnia are adjacent chromatographic peak in reference substance, the test sample chromatogram degree of separation all greater than 1.5, negative control solution at strychnine, that strychnia goes out the peak position is noiseless.The results are shown in Figure 1, Fig. 2 and Fig. 3.
5. mix the reference substance preparation
Get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g).
6. test sample preparation
6.1 extract the investigation of quantity of solvent
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, respectively accurate methenyl choloride 20ml, 25ml, the 50ml of adding.The content of strychnine, strychnia determines to add the methenyl choloride amount as index in every gram medicine.The results are shown in Table 1.
Table 1 extracts quantity of solvent and investigates test findings
Figure BDA00002201111000061
The result shows, the content that adds strychnine, strychnia in methenyl choloride 20ml, 25ml, the every gram medicine of 50ml gained is basically identical, extract fully for guaranteeing strychnine in the medicine, strychnia, and be 25ml therefore select methenyl choloride to extract quantity of solvent.
6.2 the investigation of extraction time
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, put respectively in the water-bath refluxing extraction 1 hour, 2 hours, 3 hours.The content of strychnine, strychnia is determined extraction time as index in every gram medicine.The results are shown in Table 2.
Table 2 extraction time investigation test findings
Figure BDA00002201111000062
The result shows, extracted 2 hours and 3 hours every gram medicines of gained in the content of strychnine, strychnia basic identical, be 2 hours therefore select reflux extracting time.
7. quantitative limit
Get the strychnine reference substance an amount of, accurately weighed, add methyl alcohol and make the solution that every 1ml contains strychnine 0.630 μ g.Get the strychnia reference substance an amount of, accurately weighed, add methyl alcohol and make the solution that every 1ml contains strychnia 0.315 μ g.Precision is drawn 10 μ l, injection liquid chromatography respectively.The result shows, the signal to noise ratio (S/N ratio) of strychnine, strychnia chromatographic peak all is about 10:1, then strychnine quantitatively be limited to 6.30ng, strychnia quantitatively be limited to 3.15ng.
8. the investigation of the preparation of typical curve and linear relationship
Precision measures the mixing reference substance stock solution solution of strychnine, strychnia, and (strychnine content is 25.2 μ g/ml, strychnia content is 12.6 μ g/ml) 1ml, 3ml, 5ml, 8ml, 10ml, put respectively in the 10ml volumetric flask, methyl alcohol is diluted to scale, shake up, each accurate sample introduction 10 μ l carries out linear regression with peak area (A) to reference substance concentration (C), get strychnine regression equation: A=9174.6C+174.47, related coefficient: R=0.9998; Strychnia regression equation: A=9170.1C+633.28, related coefficient: R=0.9999.The result shows, strychnine is in 2.52 μ g/ml-25.2 μ g/ml scopes, and the peak area of strychnine (A) is good with reference substance concentration (C) linear relationship; Strychnia is in 1.26 μ g/ml-12.6 μ g/ml scopes, and the peak area of strychnia (A) is good with reference substance concentration (C) linear relationship.The results are shown in Table 3, table 4.
Table 3 strychnine linear relationship is investigated the result
Figure BDA00002201111000071
Table 4 strychnia linear relationship is investigated the result
Figure BDA00002201111000072
9. precision test
The accurate mixing reference substance solution 10 μ l that draw strychnine, strychnia, the injection liquid chromatography, each METHOD FOR CONTINUOUS DETERMINATION 6 times, the record peak area also calculates relative standard deviation.The result shows, instrument precision is good.The results are shown in Table 5, table 6.
Table 5 strychnine Precision test result
Figure BDA00002201111000073
Table 6 strychnia Precision test result
Figure BDA00002201111000074
Figure BDA00002201111000081
10. stability test
After the need testing solution preparation is finished, the accurate 10 μ l that draw, the injection liquid chromatography, the record peak area was measured once every 2 hours later on, investigated 8 hours, and the record peak area also calculates relative standard deviation.The result shows, strychnine, strychnia measurement result in 8 hours is stable in the need testing solution.The results are shown in Table 7, table 8.
Table 7 strychnine stability test result
Figure BDA00002201111000082
Table 8 strychnia stability test result
11. replica test
Get with a collection of detoxicating capsule sample (product batch number: 20100701) 12g, accurately weighed, totally 6 parts, prepare need testing solution by the method under the preparation of need testing solution, precision is drawn 10 μ l, injection liquid chromatography, the content of strychnine, strychnia in the calculation sample respectively.The result shows, this analytical approach repeatability is good.The results are shown in Table 9, table 10.
Table 9 strychnine replica test result
Table 10 strychnia replica test result
Figure BDA00002201111000085
Figure BDA00002201111000091
12. recovery test
Precision takes by weighing that (product batch number: 20100701) 6 parts, each accurate strychnine, strychnia reference substance of adding is measured its content, calculate recovery rate with a collection of detoxicating capsule sample.The result shows, this assay method measurement result is accurate.The results are shown in Table 11, table 12.
Table 11 strychnine recovery test result
Figure BDA00002201111000092
Table 12 strychnia recovery test result
Figure BDA00002201111000093
13. sample determination
Get three batches of Tibetan medicinal composition detoxicating capsules, measure and calculate strychnine, strychnia content.The results are shown in Table 13.
Table 13 sample size measurement result
Figure BDA00002201111000094
Experimental example 2: the assay experiment of iron powder
1. instrument, reagent and test sample
Instrument: BT125D type Sai Duolisi electronic analytical balance (German Sai Duolisi sartorius company); 2.5-10 molding box formula resistance furnace (Beijing is bright medical apparatus and instruments factory forever); HH-4 type digital display thermostat water bath (Changzhou Rong Guan experimental analysis instrument plant).
Main agents: (take by weighing stannous chloride and be dissolved in the 40ml concentrated hydrochloric acid, thin up is to 100ml for concentration expressed in percentage by weight 5% stannous chloride solution.Dilute with water is one times before use, and get final product); Methyl orange indicator solution (get methyl orange 0.1g and be dissolved in the 100ml water, and get final product); Diphenylamine sulfonic acid sodium salt indicator solution (get diphenylamine sulfonic acid sodium salt 0.2g and be dissolved in the 100ml water, and get final product); Mixture of sulfuric phosphoric acid solution (the 150ml mass percentage concentration 95%-98% concentrated sulphuric acid is under agitation slowly injected 500ml water, add again 150ml phosphoric acid after the cooling, be diluted with water to 1000ml, mixing, and get final product).(standard reagent is stored in the exsiccator in 150 ~ 160 ℃ of dryings 2 hours the potassium dichromate standard solution, accurately takes by weighing the 0.0980g potassium dichromate in the 250ml beaker, quantitatively change in the 1000ml volumetric flask after adding the suitable quantity of water dissolving, be diluted with water to scale, shake up, its concentration is 0.002mol/l).
Sample: precious imperial restoring consciouness capsule (the Qinghai gold is scolded Tibetan medicine medicine company incorporated company and provided), lot number is respectively: 20090516,20090908,20110426.
2. the ashing of sample
The crucible that fills sample is put on the electric furnace slowly vehement burning (avoiding burning, expanding, overflow), blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature.Drip the mass percentage concentration 95%-98% concentrated sulphuric acid an amount of, make carbide all moistening, take 0.5ml to 2ml as excellent, wherein 1ml is optimum.Continue to be heated to steam and eliminate, white cigarette emitted to the greatest extent, in 500-700 ℃ of blazing 3-5 hour.Wherein take 600 ℃ blazing 4 hours as optimum.Use the calcination charing, then add sulfuric acid, the method purpose of ashing is again: remove the interference of other flavour of a drug in the Tibetan medicinal preparation, improved simultaneously the content of iron, make iron be converted into Fe3+, be convenient to measure.
3. the selection of concentration of hydrochloric acid
Reaction is carried out in hydrochloric acid (HCl) medium, reduction Fe 3+The time hydrochloric acid (HCl) concentration with 4mol/l for well, Sn during greater than 6mol/l 2+The methyl orange of then reducing first is colourless, makes it can't indicate Fe 3+Reduction, Cl simultaneously -Excessive concentration also may consume K 2Cr 2O 7, HCl concentration is lower than 2mol/l, and then Methyl Orange Discoloration is slow.
4. the selection of molten sample temperature
Molten sample temperature has considerable influence to the measurement result of iron content.When molten sample temperature was lower than 55 ℃, molten sample was slow; Surpass 75 ℃ of HCl volatilizations too fast, concentration reduces, and molten sample is incomplete.Experiment shows that molten sample temperature is controlled at 65 ℃ of left and right sides best results.
5. the control of redox temperature
When reducing with stannous chloride, temperature should be controlled at 70-80 ℃, and constantly shakes.And in last oxidimetric titration, solution temperature should be controlled at 20～30 ℃ for well, so that reaction can comparatively fast be carried out.
6. the selection of mixture of sulfuric phosphoric acid solution
The Fe that generates in the titration process 3+Be yellow, affect the observation of terminal point, if in solution, add phosphoric acid (H 3PO 4), H 3PO 4With Fe 3+Generate colourless Fe (HPO4) 2-, can shelter Fe 3+Simultaneously because Fe (HPO4) 2-Generation so that Fe 3+/ Fe 2+The right conditional potential of electricity reduces, and titration jump increases, and indicator can variable color in mutation range, thereby reduces titration error.
7. principle
This product sample is behind sample ashing, dissolving with hydrochloric acid, and iron wherein is converted into Fe 3+Under strong acidic condition, Fe 3+Can pass through SnCl 2Be reduced to Fe 2+Sn 2+With Fe 3+Reduce complete after, methyl orange also can be by Sn 2+Be reduced into hydrogenation methyl orange and fade, thereby methyl orange can be indicated Fe 3+The reduction terminal point.Sn 2+Can also continue to make hydrogenation methyl orange to be reduced into N, N-dimethyl-p-phenylenediamine and sodium sulfanilate.Its reaction equation is:
(CH 3) 2NC 6H 4N=NC 6H 4SO 3Na+2e -+2H +→(CH 3) 2NC 6H 4NH-NHC 6H 4SO 3Na
(CH 3) 2NC 6H 4NH-NHC 6H 4SO 3Na＋2e -＋2H +→(CH 3) 2NC 6H 4NH 2+NH 2C 6H 4SO 3Na
So, slightly excessive Sn 2+Also be eliminated.Because above-mentioned reaction is irreversible, so the reduzate of methyl orange does not consume K 2Cr 2O 7After having reacted, take diphenylamine sulfonic acid sodium salt as indicator, use K 2Cr 2O 7Standard solution is titrated to solution and is purple and is terminal point, and the key reaction formula is as follows:
2FeCl 4 -+SnCl 2-+2Cl -=2FeCl 4 2-+SnCl 6 2-
6Fe 2++Cr 2O 7 2-+14H +=6Fe 3++2Cr 3++7H 2O
8. the assay of iron in the sample
Get Tibetan medicinal composition detoxicating capsule content powder 14g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning (avoiding burning, expanding, overflow), blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, (frequently shook in 4 hours 65 ℃ of left and right sides heating for dissolving, avoid boiling), after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, put in the 250ml conical flask, add mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, add 6 methyl orange indicator solutions after, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, be cooled to room temperature with cold water rapidly after, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.The results are shown in Table 14.
Iron content measurement result in table 14 detoxicating capsule
Figure BDA00002201111000111
9. reappearance test
Get with a collection of detoxicating capsule sample (product batch number: 20100701) accurately weighed, totally 6 parts, measure according to the iron content determination method respectively.The result shows, the method repeatability is good.The results are shown in Table 15.
Iron content is measured replica test in table 15 detoxicating capsule
Figure BDA00002201111000112
10. recovery test
Do not contain the sample of iron powder in the ratio autogamy of Tibetan medicinal composition detoxicating capsule prescription consumption, get approximately 14g of this blank sample, accurately weighed, the standard iron powder of accurate adding known content is 0.1g approximately, carries out ashing and processes, and measures as stated above content, and calculate recovery rate.The result shows, this assay method measurement result is accurate.The results are shown in Table 16.
Table 16 recovery test measurement result
Figure BDA00002201111000121
Experimental example 3: the assay experiment of safflower
1. instrument, reagent and test sample
Instrument: 1220 type Agilent high performance liquid chromatographs; AuW220D type Shimadzu electronic analytical balance.
Reference substance: hydroxyl radical carthamin yellow carthamus A reference substance (Nat'l Pharmaceutical ﹠ Biological Products Control Institute), lot number: 111637-200905.
Sample: detoxicating capsule (the Qinghai gold is scolded Tibetan medicine medicine company incorporated company and provided), lot number: 20100701,20100702,20100703.
2. detect the selection of wavelength
Get the hydroxyl radical carthamin yellow carthamus A reference substance solution, scan in the 190-500nm wavelength coverage, according to ultraviolet absorpting spectrum, selected 403nm is for detecting wavelength.
3. mobile phase is selected
Research finds, during as mobile phase, all can reach good chromatographic resolution effect than 0.5% aqueous formic acid than methyl alcohol-volume parts of 20-30:80-70 take volume parts.Wherein optimum as 25:75 as mobile phase than the volume ratio of 0.5% formic acid solution take methyl alcohol-volume parts, about relative retention time 10min.
4. system suitability and negative interference test
Under above-mentioned chromatographic condition, precision is drawn reference substance solution, need testing solution, each 10 μ l of negative control solution, injection liquid chromatography, record chromatogram respectively.The result shows, the degree of separation that hydroxyl radical carthamin yellow carthamus A is adjacent chromatographic peak in reference substance, the test sample chromatogram is all greater than 1.5, and it is noiseless that negative control solution goes out the peak position at hydroxyl radical carthamin yellow carthamus A.The results are shown in Figure 4, Fig. 5 and Fig. 6.
5. reference substance preparation
Get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product.
6. test sample preparation
6.1 the investigation of extracting method
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, ultrasonic, backflow, Zhen Oscillating processed 40 minutes respectively.The content of hydroxyl radical carthamin yellow carthamus A is determined extracting method as index in every gram medicine.The results are shown in Table 17.
Table 17 extracting method is investigated test findings
Figure BDA00002201111000122
The result shows, the content of hydroxyl radical carthamin yellow carthamus A is the highest in the every gram medicine of ultrasonic extraction gained, is ultrasonic extraction therefore select extracting method.
6.2 extract the investigation of solvent
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, precision adds entry, volume parts than 25% methanol aqueous solution, each 25ml of methyl alcohol respectively.The content of hydroxyl radical carthamin yellow carthamus A determines to extract solvent as index in every gram medicine.The results are shown in Table 18.
Table 18 extracts solvent and investigates test findings
Figure BDA00002201111000131
The result shows, the hydroxyl radical carthamin yellow carthamus A content that water, volume parts are surveyed than 25% methanol aqueous solution and methyl alcohol is basically identical, extracts the solvent filter difficulty owing to using water as, and pure methyl alcohol toxicity is large, and to extract solvent be that volume parts is than 25% methanol aqueous solution therefore select.
6.3 the investigation of extraction time
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, ultrasonic processing is 30 minutes, 40 minutes, 50 minutes respectively.The content of hydroxyl radical carthamin yellow carthamus A is determined extraction time as index in every gram medicine.The results are shown in Table 19.
Table 19 extraction time investigation test findings
Figure BDA00002201111000132
The result shows, the content of hydroxyl radical carthamin yellow carthamus A is basically identical in ultrasonic extraction 40 minutes and the 50 minutes every gram medicines of gained, is 40 minutes therefore select extraction time.
7. the investigation of the preparation of typical curve and linear relationship
Precision measures hydroxyl radical carthamin yellow carthamus A reference substance stock solution solution (hydroxyl radical carthamin yellow carthamus A content is 16.2 μ g/ml) 1ml, 3ml, 5ml, 8ml, 10ml, put respectively in the 10ml volumetric flask, volume parts is diluted to scale than 25% methanol aqueous solution, shake up, each accurate sample introduction 10 μ l, with peak area (A) reference substance concentration (C) is carried out linear regression, get hydroxyl radical carthamin yellow carthamus A regression equation: A=21.006C-4.0763, related coefficient: R=0.9998.The result shows, hydroxyl radical carthamin yellow carthamus A is in 1.62 μ g/ml-16.2 μ g/ml scopes, and the peak area of hydroxyl radical carthamin yellow carthamus A (A) is good with reference substance concentration (C) linear relationship.The results are shown in Table 20.
Table 20 hydroxyl radical carthamin yellow carthamus A linear relationship is investigated the result
Figure BDA00002201111000133
8. precision test
The accurate hydroxyl radical carthamin yellow carthamus A reference substance solution 10 μ l that draw, the injection liquid chromatography, each METHOD FOR CONTINUOUS DETERMINATION 6 times, the record peak area also calculates relative standard deviation.The result shows, instrument precision is good.The results are shown in Table 21.
Table 21 hydroxyl radical carthamin yellow carthamus A Precision test result
Figure BDA00002201111000134
Figure BDA00002201111000141
9. stability test
After the need testing solution preparation is finished, the accurate 10 μ l that draw, the injection liquid chromatography, the record peak area was measured once every 2 hours later on, investigated 8 hours, and the record peak area also calculates relative standard deviation.The result shows, hydroxyl radical carthamin yellow carthamus A measurement result in 8 hours is stable in the need testing solution.The results are shown in Table 22.
Table 22 hydroxyl radical carthamin yellow carthamus A stability test result
Figure BDA00002201111000142
10. replica test
Get with a collection of detoxicating capsule sample (product batch number: 20100701) 5g, accurately weighed, totally 6 parts, prepare need testing solution by the method under the preparation of need testing solution, precision is drawn 10 μ l, injection liquid chromatography, the content of hydroxyl radical carthamin yellow carthamus A in the calculation sample respectively.The result shows, this analytical approach repeatability is good.The results are shown in Table 23.
Table 23 hydroxyl radical carthamin yellow carthamus A replica test result
Figure BDA00002201111000143
11. recovery test
Precision takes by weighing that (product batch number: 20100701) 6 parts, each accurate hydroxyl radical carthamin yellow carthamus A reference substance that adds is measured its content, calculate recovery rate with a collection of detoxicating capsule sample.The result shows, this assay method measurement result is accurate.The results are shown in Table 24.
Table 24 hydroxyl radical carthamin yellow carthamus A recovery test result
Figure BDA00002201111000151
12. sample determination
Get three batches of Tibetan medicinal composition detoxicating capsules, measure and calculate hydroxyl radical carthamin yellow carthamus A content.The results are shown in Table 25.
Table 25 sample size measurement result
Figure BDA00002201111000152
Experimental example 4: the assay experiment of calculus bovis factitius
1. instrument, reagent and test sample
Instrument: L-2100 type Hitachi high performance liquid chromatograph, AuW220D type Shimadzu electronic analytical balance.
Reference substance: cholic acid reference substance (Nat'l Pharmaceutical ﹠ Biological Products Control Institute) lot number: 100078-200414.
Sample: detoxicating capsule (the Qinghai gold is scolded Tibetan medicine medicine company incorporated company and provided), lot number: 20100701,20100702,20100703.
2. mobile phase is selected
The assay of cholic acid adopts methyl alcohol-glacial acetic acid system more, research is found all to reach content inspection requirements than 1% glacial acetic acid aqueous solution as mobile phase than methyl alcohol-volume parts of 80-70:20-30 take volume parts, wherein methyl alcohol-volume parts is more best than 1% glacial acetic acid aqueous solution (76:24) separating effect, about relative retention time 13min.
4. system suitability
Under above-mentioned chromatographic condition, precision is drawn reference substance solution, need testing solution, each 10 μ l of negative control solution, injection liquid chromatography, record chromatogram respectively.The result shows, the degree of separation that cholic acid is adjacent chromatographic peak in reference substance, the test sample chromatogram is all greater than 1.5, and it is noiseless that negative control solution goes out the peak position at cholic acid.The results are shown in Figure 7, Fig. 8 and Fig. 9.
5. reference substance preparation
It is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product.
6. test sample preparation
6.1 the investigation of extracting method
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, ultrasonic, backflow, Zhen Oscillating processed 30 minutes respectively.The content of cholic acid is determined extracting method as index in every gram medicine.The results are shown in Table 26.
Table 26 extracting method is investigated test findings
Figure BDA00002201111000153
The result shows, the content of cholic acid is the highest in the every gram medicine of ultrasonic extraction gained, is ultrasonic extraction therefore select extracting method.
6.2 extract the investigation of solvent
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, accurate methyl alcohol, ethanol, the volume parts of adding is than each 50ml of 60% ethanol water respectively.The content of cholic acid determines to extract solvent as index in every gram medicine.The results are shown in Table 27.
Table 27 extracts solvent and investigates test findings
Figure BDA00002201111000161
The result shows, the content of cholic acid is the highest in the measured every gram medicine of methyl alcohol, and to extract solvent be methyl alcohol therefore select.
6.3 the investigation of extraction time
By method under the assay item need testing solution is detected.Prepare 3 parts by the method under the preparation of need testing solution, ultrasonic processing is 20 minutes, 30 minutes, 40 minutes respectively.The content of cholic acid is determined extraction time as index in every gram medicine.The results are shown in Table 28.
Table 28 extraction time investigation test findings
Figure BDA00002201111000162
The result shows, the content of cholic acid is basically identical in ultrasonic extraction 30 minutes and the 40 minutes every gram medicines of gained, is 30 minutes therefore select extraction time.
7. the investigation of the preparation of typical curve and linear relationship
Precision measures cholic acid reference substance stock solution solution (cholic acid content is 892.8 μ g/ml) 1ml, 3ml, 5ml, 8ml, 10ml, put respectively in the 10ml volumetric flask, methyl alcohol is diluted to scale, shake up, each accurate sample introduction 10 μ l, with the logarithm (Log A) of peak area the logarithm (Log C) of reference substance concentration is carried out linear regression, get the cholic acid regression equation: Log A=1.0191Log C+3.4350, related coefficient: R=0.9995.The result shows, cholic acid is in 89.28 μ g/ml-892.8 μ g/ml scopes, and the logarithm of cholic acid peak area (LogA) is good with logarithm (Log C) linear relationship of reference substance concentration.The results are shown in Table 29.
Table 29 cholic acid linear relationship is investigated the result
Figure BDA00002201111000163
8. precision test
The accurate cholic acid reference substance solution 10 μ l that draw, the injection liquid chromatography, each METHOD FOR CONTINUOUS DETERMINATION 6 times, the record peak area also calculates relative standard deviation.The result shows, instrument precision is good.The results are shown in Table 30.
Table 30 cholic acid Precision test result
9. stability test
After the need testing solution preparation is finished, the accurate 10 μ l that draw, the injection liquid chromatography, the record peak area was measured once every 2 hours later on, investigated 8 hours, and the record peak area also calculates relative standard deviation.The result shows, cholic acid measurement result in 8 hours is stable in the need testing solution.The results are shown in Table 31.
Table 31 cholic acid stability test result
10. replica test
Get with a collection of detoxicating capsule sample (product batch number: 20100701) 7g, accurately weighed, totally 6 parts, prepare need testing solution by the method under the preparation of need testing solution, respectively accurate 10 μ l, injection liquid chromatography, the content of cholic acid in the calculation sample drawn.The result shows, this analytical approach repeatability is good.The results are shown in Table 32.
Table 32 cholic acid replica test result
Figure BDA00002201111000172
11. recovery test
Precision takes by weighing that (product batch number: 20100701) 6 parts, each accurate cholic acid reference substance that adds is measured its content, calculate recovery rate with a collection of detoxicating capsule sample.The result shows, this assay method measurement result is accurate.The results are shown in Table 33.
Table 33 cholic acid recovery test result
12. sample determination
Get three batches of Tibetan medicinal composition detoxicating capsules, measure and calculate cholic acid content.The results are shown in Table 34.
Table 34 sample size measurement result
Figure BDA00002201111000181
Following embodiment all can realize the described effect of above-mentioned experimental example.
Description of drawings
Fig. 1 is that strychnine, strychnia mix the contrast high-efficient liquid phase chromatogram;
Fig. 2 is Tibetan medicinal composition detoxicating capsule vomiting nut test sample high-efficient liquid phase chromatogram;
Fig. 3 is that the Tibetan medicinal composition detoxicating capsule lacks vomiting nut negative control high-efficient liquid phase chromatogram;
Fig. 4 is hydroxyl radical carthamin yellow carthamus A contrast high-efficient liquid phase chromatogram;
Fig. 5 is Tibetan medicinal composition detoxicating capsule safflower test sample high-efficient liquid phase chromatogram;
Fig. 6 is that the Tibetan medicinal composition detoxicating capsule lacks safflower negative control high-efficient liquid phase chromatogram;
Fig. 7 is cholic acid contrast high-efficient liquid phase chromatogram;
Fig. 8 is Tibetan medicinal composition detoxicating capsule calculus bovis factitius test sample high-efficient liquid phase chromatogram;
Fig. 9 is Tibetan medicinal composition detoxicating capsule shortage of staff worker cow-bezoar negative control high-efficient liquid phase chromatogram;
Wherein, the horizontal ordinate of Fig. 1, Fig. 2, Fig. 3, Fig. 7, Fig. 8, Fig. 9 is the time, unit: minute (Minutes); Ordinate is voltage, unit: volt (Volts).The horizontal ordinate of Fig. 4, Fig. 5, Fig. 6 is the time, unit: minute (min); Ordinate is absorbance (mAU).
Embodiment
Below in conjunction with embodiment the present invention is done detailed elaboration, but be not limited to the embodiment of these concrete records.The Tibetan medicinal composition detoxicating capsule that detects is that the Qinghai gold is scolded Tibetan medicine medicine company incorporated company and produced and to sell.
Embodiment 1: the detoxicating capsule quality determining method
Detoxicating capsule comprises the discriminating of cholic acid thin layer, gallic acid content mensuration (also can not comprise these mensuration items), also comprises following mensuration item:
A. the assay of vomiting nut:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 20:80-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; The detection wavelength is 260nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g);
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content powder 12g, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product vomiting nut content is with strychnine (C 31H 22N 2O 2) meter, should be 0.043mg/g-0.080mg/g; With strychnia (C 23H 26N 2O 1) meter, must not be less than 0.028mg/g.
B. the assay of iron powder:
Get Tibetan medicinal composition detoxicating capsule content powder 8g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, 65 ℃ of left and right sides heating for dissolving 4 hours, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.
This product iron content is 0.409mg/g-0.682mg/g.
C. the assay of safflower:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 25:75 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 403nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content powder 5g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 25ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product safflower content is with hydroxyl radical carthamin yellow carthamus A (C 27H 30O 15) meter, must not be less than 0.027mg/g.
D. the assay of calculus bovis factitius:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 76:24 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content powder 7g, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 50ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 25ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
This product calculus bovis factitius content is with cholic acid (C 24H 40O 5) meter, must not be less than 0.252mg/g.
Embodiment 2: the detoxification particles quality determining method
Detoxification particles comprises the discriminating of cholic acid thin layer, gallic acid content mensuration (also can not comprise these mensuration items), also comprises following mensuration item:
A. the assay of vomiting nut:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 20:80-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; The detection wavelength is 260nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g);
The preparation of need testing solution: get Tibetan medicinal composition detoxification particles powder 12g, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product vomiting nut content is with strychnine (C 31H22N 2O 2) meter, should be 0.043mg/g-0.080mg/g; With strychnia (C 23H 26N 2O 1) meter, must not be less than 0.028mg/g.
B. the assay of iron powder:
Get Tibetan medicinal composition detoxification particles powder 8g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, 65 ℃ of left and right sides heating for dissolving 4 hours, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.
This product iron content is 0.409mg/g-0.682mg/g.
C. the assay of safflower:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 25:75 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 403nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxification particles powder 5g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 25ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product safflower content is with hydroxyl radical carthamin yellow carthamus A (C 27H 30O 15) meter, must not be less than 0.027mg/g.
D. the assay of calculus bovis factitius:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 76:24 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxification particles powder 7g, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 50ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 25ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
This product calculus bovis factitius content is with cholic acid (C 24H 40O 5) meter, must not be less than 0.252mg/g.
Described Tibetan medicinal composition detoxification particles refers to the precious 4.0g of detoxicating capsule bulk drug prescription animal; Radix Brassicae rapae 4.0g; earthworm fruit mustard 30.4g; vomiting nut (system) 1.6g; German tamarisk 30.4g; borax (system) 0.8g; pearl (system) 3.2g; coral 1.6g; lazurite (system) 1.6g; turquoise (system) 1.6g; iron powder (system) 2.4g; ring copper (system) 0.8g; parmelia saxatilis (system) 0.8g; ichor cream 4.0g; nutmeg 1.6g; tsaoko 1.6g; Tabasheer 1.6g; safflower 1.6g; cloves 1.6g; myrobalan 2.4g; Phytolacca acinosa 1.6g; santal 30.4g; santal 27.9g; Moschus 1.6g; calculus bovis factitius 2.4g; whorlleaf oxytropis herb 30.4g; grass-leaved sweetflag 1.6g; cardamom 1.6g; hooker winghead root 30.4g; west szechwan pyrethrum 30.4g; rhizome of davallia 3.2g; rhizoma nardostachyos 1.6g; Corallodiscus flabellatus (Franch.) Burtt 30.4g; alpine gentian herb 30.4g; muscicolous woodbetony root 30.4g; add conventional auxiliary material, according to the granule of common process preparation.
Embodiment 3: the detoxifying pills quality determining method
Detoxifying pills comprises the discriminating of cholic acid thin layer, gallic acid content mensuration (also can not comprise these mensuration items), also comprises following mensuration item:
A. the assay of vomiting nut:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 20:80-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; The detection wavelength is 260nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g);
The preparation of need testing solution: get Tibetan medicinal composition detoxifying pills powder 12g, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product vomiting nut content is with strychnine (C 31H 22N 2O 2) meter, should be 0.043mg/g-0.080mg/g; With strychnia (C 23H 26N 2O 1) meter, must not be less than 0.028mg/g.
B. the assay of iron powder:
Get Tibetan medicinal composition detoxifying pills powder 8g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, 65 ℃ of left and right sides heating for dissolving 4 hours, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.
This product iron content is 0.409mg/g-0.682mg/g.
C. the assay of safflower:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 25:75 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 403nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxifying pills powder 5g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 25ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product safflower content is with hydroxyl radical carthamin yellow carthamus A (C 27H 30O 15) meter, must not be less than 0.027mg/g.
D. the assay of calculus bovis factitius:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 76:24 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxifying pills powder 7g, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 50ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 25ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
This product calculus bovis factitius content is with cholic acid (C 24H 40O 5) meter, must not be less than 0.252mg/g.
Described Tibetan medicinal composition detoxifying pills refers to the precious 4.0g of detoxicating capsule bulk drug prescription animal; Radix Brassicae rapae 4.0g; earthworm fruit mustard 30.4g; vomiting nut (system) 1.6g; German tamarisk 30.4g; borax (system) 0.8g; pearl (system) 3.2g; coral 1.6g; lazurite (system) 1.6g; turquoise (system) 1.6g; iron powder (system) 2.4g; ring copper (system) 0.8g; parmelia saxatilis (system) 0.8g; ichor cream 4.0g; nutmeg 1.6g; tsaoko 1.6g; Tabasheer 1.6g; safflower 1.6g; cloves 1.6g; myrobalan 2.4g; Phytolacca acinosa 1.6g; santal 30.4g; santal 27.9g; Moschus 1.6g; calculus bovis factitius 2.4g; whorlleaf oxytropis herb 30.4g; grass-leaved sweetflag 1.6g; cardamom 1.6g; hooker winghead root 30.4g; west szechwan pyrethrum 30.4g; rhizome of davallia 3.2g; rhizoma nardostachyos 1.6g; Corallodiscus flabellatus (Franch.) Burtt 30.4g; alpine gentian herb 30.4g; muscicolous woodbetony root 30.4g; add conventional auxiliary material, according to the pill of common process preparation.
Embodiment 4: the Jiedu Tablets quality determining method
Jiedu Tablets comprises the discriminating of cholic acid thin layer, gallic acid content mensuration (also can not comprise these mensuration items), also comprises following mensuration item:
A. the assay of vomiting nut:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 20:80-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution (with volume parts than 10% phosphorus acid for adjusting pH value 2.8) as mobile phase; The detection wavelength is 260nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up.Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and namely gets (every 1ml contains strychnine 12 μ g, strychnia 6 μ g);
The preparation of need testing solution: get Tibetan medicinal composition Jiedu Tablets powder 12g, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product vomiting nut content is with strychnine (C 31H 22N 2O 2) meter, should be 0.043mg/g-0.080mg/g; With strychnia (C 23H 26N 2O 1) meter, must not be less than 0.028mg/g.
B. the assay of iron powder:
Get Tibetan medicinal composition Jiedu Tablets powder 8g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, 65 ℃ of left and right sides heating for dissolving 4 hours, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used dichromate titration liquid (0.002mol/l) titration immediately.Every 1ml dichromate titration liquid (0.002mol/l) is equivalent to the iron of 0.6702mg.
This product iron content is 0.409mg/g-0.682mg/g.
C. the assay of safflower:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 25:75 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 403nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition Jiedu Tablets powder 5g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 25ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
This product safflower content is with hydroxyl radical carthamin yellow carthamus A (C 27H 30O 15) meter, must not be less than 0.027mg/g.
D. the assay of calculus bovis factitius:
Measure according to high performance liquid chromatography (" an appendix VI of Chinese pharmacopoeia version in 2010 D);
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 76:24 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition Jiedu Tablets powder 7g, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 50ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 25ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
This product calculus bovis factitius content is with cholic acid (C 24H 40O 5) meter, must not be less than 0.252mg/g.
Described Tibetan medicinal composition Jiedu Tablets refers to the precious 4.0g of detoxicating capsule bulk drug prescription animal; Radix Brassicae rapae 4.0g; earthworm fruit mustard 30.4g; vomiting nut (system) 1.6g; German tamarisk 30.4g; borax (system) 0.8g; pearl (system) 3.2g; coral 1.6g; lazurite (system) 1.6g; turquoise (system) 1.6g; iron powder (system) 2.4g; ring copper (system) 0.8g; parmelia saxatilis (system) 0.8g; ichor cream 4.0g; nutmeg 1.6g; tsaoko 1.6g; Tabasheer 1.6g; safflower 1.6g; cloves 1.6g; myrobalan 2.4g; Phytolacca acinosa 1.6g; santal 30.4g; santal 27.9g; Moschus 1.6g; calculus bovis factitius 2.4g; whorlleaf oxytropis herb 30.4g; grass-leaved sweetflag 1.6g; cardamom 1.6g; hooker winghead root 30.4g; west szechwan pyrethrum 30.4g; rhizome of davallia 3.2g; rhizoma nardostachyos 1.6g; Corallodiscus flabellatus (Franch.) Burtt 30.4g; alpine gentian herb 30.4g; muscicolous woodbetony root 30.4g; add conventional auxiliary material, according to the tablet of common process preparation.

Claims (8)

1. the quality determining method of a Tibetan medicinal composition detoxicating capsule and preparation thereof; the bulk drug of said preparation consists of the animal treasured; Radix Brassicae rapae; earthworm fruit mustard; Semen Strychni (processed); German tamarisk; borax processed; pearl processed; coral; lazurite processed; turquoise processed; Magnetite (processed); system is rung copper; parmelia saxatilis processed; ichor cream; nutmeg; tsaoko; Tabasheer; safflower; cloves; the myrobalan; Phytolacca acinosa; santal; santal; Moschus; calculus bovis factitius; whorlleaf oxytropis herb; grass-leaved sweetflag; cardamom; hooker winghead root; west szechwan pyrethrum; the rhizome of davallia; rhizoma nardostachyos; Corallodiscus flabellatus (Franch.) Burtt; alpine gentian herb; muscicolous woodbetony root; it is characterized in that, quality determining method comprises one or more in the following content assaying method:
A. the assay of vomiting nut: get respectively the strychnine reference substance, the strychnia reference substance is contrast, octadecylsilane chemically bonded silica is filling agent; Take volume parts than the acetonitrile of 15-25:85-75-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution and with volume parts than 10% phosphorus acid for adjusting pH value 2.8 as mobile phase; Detecting wavelength is the high performance liquid chromatography of 250-270nm;
B. the assay of iron powder: after Chinese medicine preparation ashing to be determined, with the concentrated hydrochloric acid dissolving, add the methyl orange indicator solution, drip stannous chloride solution with Fe 3+Be reduced to Fe 2+, solution becomes pale red, then take diphenylamine sulfonic acid sodium salt as indicator, is titrated to solution with the potassium dichromate standard solution and is purple and is terminal point, calculates the content of iron powder with the amount of known potassium dichromate standard solution;
C. the assay of safflower: get hydroxyl safflower anthocyanidin A reference substance and add volume parts and be mixed with reference substance solution than 25% methanol aqueous solution, get preparation powder adding volume parts to be detected and after ultrasonic dissolution, removal of impurities, be mixed with need testing solution than 25% methanol aqueous solution, take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 20-30:80-70 than 0.5% aqueous formic acid as mobile phase, detecting wavelength is the high performance liquid chromatography of 400-410nm;
D. the assay of calculus bovis factitius: get the cholic acid reference substance and add methyl alcohol and be mixed with reference substance solution, get preparation powder to be detected and add methyl alcohol and after ultrasonic dissolution, removal of impurities, be mixed with need testing solution, take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 80-70:20-30 than 1% glacial acetic acid aqueous solution as mobile phase, the high performance liquid chromatography of evaporative light-scattering detector.
2. quality determining method as claimed in claim 1 is characterized in that, the assay concrete steps of described vomiting nut are:
According to high effective liquid chromatography for measuring;
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 15-25:85-75-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution and with volume parts than 10% phosphorus acid for adjusting pH value 2.8 as mobile phase; The detection wavelength is 250-270nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up; Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and get final product;
The preparation of need testing solution: get preparation powder 10-14g to be detected, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product.
Determination method: draw respectively each 5-10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
3. quality determining method as claimed in claim 1 is characterized in that, the content assaying method concrete steps of described iron powder are:
Get preparation powder 6 ~ 10g to be detected, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 0.5-2ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, in 500-700 ℃ of blazing 3-5 hour, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, at 60-70 ℃ of left and right sides heating for dissolving 3-5 hour, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 0-15ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, can add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 2-10ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions are used 0.002mol/l potassium dichromate solution titration titration immediately.
4. quality determining method as claimed in claim 1 is characterized in that, the concrete steps of described safflower are:
According to high effective liquid chromatography for measuring;
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 20-30:80-70 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 400-410nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 6-10 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get preparation powder 4-6g to be detected, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 20-30ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 5-10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product.
5. quality determining method as claimed in claim 1 is characterized in that, the concrete steps of described calculus bovis factitius assay are:
According to high effective liquid chromatography for measuring;
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 80-70:20-30 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.4-0.5mg, and get final product;
The preparation of need testing solution: get preparation powder 6-8g to be detected, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 40-60ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 20-30ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
6. quality determining method as claimed in claim 1; it is characterized in that, the bulk drug of described preparation consists of: precious 4.0 weight portions of animal; Radix Brassicae rapae 4.0 weight portions; earthworm fruit mustard 30.4 weight portions; Semen Strychni (processed) 1.6 weight portions; German tamarisk 30.4 weight portions; borax 0.8 weight portion processed; pearl 3.2 weight portions processed; coral 1.6 weight portions; lazurite 1.6 weight portions processed; turquoise 1.6 weight portions processed; Magnetite (processed) 2.4 weight portions; system is rung copper 0.8 weight portion; parmelia saxatilis 0.8 weight portion processed; ichor cream 4.0 weight portions; nutmeg 1.6 weight portions; tsaoko 1.6 weight portions; Tabasheer 1.6 weight portions; safflower 1.6 weight portions; cloves 1.6 weight portions; myrobalan's 2.4 weight portions; Phytolacca acinosa 1.6 weight portions; santal 30.4 weight portions; santal 27.9 weight portions; Moschus 1.6 weight portions; calculus bovis factitius 2.4 weight portions; whorlleaf oxytropis herb 30.4 weight portions; grass-leaved sweetflag 1.6 weight portions; cardamom 1.6 weight portions; hooker winghead root 30.4 weight portions; west szechwan pyrethrum 30.4 weight portions; the rhizome of davallia 3.2 weight portions; rhizoma nardostachyos 1.6 weight portions; Corallodiscus flabellatus (Franch.) Burtt 30.4 weight portions; alpine gentian herb 30.4 weight portions; muscicolous woodbetony root 30.4 weight portions.
7. quality determining method as claimed in claim 6 is characterized in that, described quality determining method is:
A. the assay of vomiting nut:
According to high effective liquid chromatography for measuring;
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than the acetonitrile of 20:80-0.01mol/l sodium heptanesulfonate and 0.02mol/l potassium dihydrogen phosphate mixed in equal amounts solution and with volume parts than 10% phosphorus acid for adjusting pH value 2.8 as mobile phase; The detection wavelength is 260nm; Number of theoretical plate calculates by the strychnine peak should be not less than 5000;
The preparation of reference substance solution: get strychnine reference substance 6mg, strychnia reference substance 3mg, accurately weighed, put respectively in the 100ml measuring bottle, add methenyl choloride and make in right amount dissolving and be diluted to scale, shake up; Precision measures 2ml respectively, puts in the same 10ml measuring bottle, is diluted to scale with methyl alcohol, shakes up, and get final product;
The preparation of need testing solution: get preparation powder 12g to be detected, accurately weighed, put in the tool plug conical flask, hydro-oxidation sodium test solution 10ml, mixing was placed 30 minutes, the accurate methenyl choloride 25ml that adds, close plug, weighed weight was put in the water-bath refluxing extraction 2 hours, let cool, weighed weight is again supplied the weight of less loss with methenyl choloride, shake up, divide and get methenyl choloride liquid, filter with the filter paper that is covered with a small amount of anhydrous sodium sulfate, discard just filtrate, precision measures subsequent filtrate 5ml, put in the 10ml measuring bottle, add methyl alcohol to scale, shake up, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product;
B. the assay of iron powder:
Get Tibetan medicinal composition detoxicating capsule content or preparation powder 8g, accurately weighed, the crucible that fills sample is put on the electric furnace slowly vehement burning, blazingly be black, and no longer smolder to the whole charings of test sample, let cool to room temperature; Drip mass percentage concentration 95%-98% concentrated sulphuric acid 1ml, make carbide all moistening, continuing to be heated to steam eliminates, white cigarette emits to the greatest extent, blazing 4 hours in 600 ℃, let cool, move in the 250ml tool plug conical flask with 50ml mass percentage concentration 36%-38% concentrated hydrochloric acid gradation washing, 65 ℃ of left and right sides heating for dissolving 4 hours, after the cooling solution is moved in the 250ml volumetric flask, with a small amount of moisture time washing container, washing lotion is incorporated in the same measuring bottle, thin up is to scale, shake up, filter, precision measures subsequent filtrate 25ml, puts in the 250ml conical flask, adds mass percentage concentration 36%-38% concentrated hydrochloric acid 5ml, be heated to 70-80 ℃, after adding 6 methyl orange indicator solutions, drip while hot concentration expressed in percentage by weight 5% stannous chloride solution to solution while shaking and become pale pink, take off if shake rear pink colour, then add 1 methyl orange to solution and present stable pale pink, after being cooled to room temperature with cold water rapidly, add distilled water 50ml, mixture of sulfuric phosphoric acid solution 4ml, 15 of diphenylamine sulfonic acid sodium salt indicator solutions, fixed with 0.002mol/l dichromate titration drop immediately;
C. the assay of safflower:
According to high effective liquid chromatography for measuring;
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 25:75 than 0.5% aqueous formic acid as mobile phase; The detection wavelength is 403nm; Number of theoretical plate calculates by the hydroxyl radical carthamin yellow carthamus A peak should be not less than 3000;
The preparation of reference substance solution: get hydroxyl safflower anthocyanidin A reference substance an amount of, accurately weighed, put in the brown bottle, add volume parts and make the solution that every 1ml contains 8 μ g than 25% methanol aqueous solution, and get final product;
The preparation of need testing solution: get Tibetan medicinal composition detoxicating capsule content or preparation powder 5g, accurately weighed, put in the tool plug conical flask, the accurate volume parts that adds is than 25% methanol aqueous solution 25ml, close plug, weighed weight, ultrasonic 40 minutes, let cool, weighed weight is again supplied the weight of less loss than 25% methanol aqueous solution with volume parts, shake up, filter, get subsequent filtrate, and get final product;
Determination method: draw respectively each 10 μ l of reference substance solution and need testing solution, the injection liquid chromatography is measured, and be get final product;
D. the assay of calculus bovis factitius:
According to high effective liquid chromatography for measuring;
Chromatographic condition and system suitability: take octadecylsilane chemically bonded silica as filling agent; Take volume parts than methyl alcohol-volume parts of 76:24 than 1% glacial acetic acid aqueous solution as mobile phase; Evaporative light-scattering detector; Number of theoretical plate calculates by the cholic acid peak should be not less than 5000;
The preparation of reference substance solution: it is an amount of that precision takes by weighing the cholic acid reference substance, adds methyl alcohol and make the solution that every 1ml contains 0.45mg, and get final product;
The preparation of need testing solution: get preparation powder 7g to be detected, accurately weighed, put in the conical flask of tool plug the accurate methyl alcohol 50ml that adds, close plug, weighed weight, ultrasonic 30 minutes, let cool, weighed weight is again supplied the weight of less loss with methyl alcohol, shake up, filter, get subsequent filtrate 25ml, 40 ℃ of following recovered under reduced pressure are transferred in the 5ml measuring bottle, add methanol constant volume to scale, filter, get subsequent filtrate, and get final product;
Determination method: precision is drawn reference substance solution 10 μ l, 20 μ l respectively, need testing solution 10 μ l, and the injection liquid chromatography is measured respectively, calculates with external standard two-point method logarithmic equation, and get final product.
8. quality determining method as claimed in claim 7 is characterized in that, vomiting nut content is with strychnine (C in the described vomiting nut content 31H 22N 2O 2) meter, should be 0.043mg/g-0.080mg/g; With strychnia (C 23H 26N 2O 1) meter, must not be less than 0.028mg/g; Iron content is 0.409mg/g-0.682mg/g in the assay of described iron powder; Safflower content was with hydroxyl radical carthamin yellow carthamus A (C during described safflower content was measured 27H 30O 15) meter, must not be less than 0.027mg/g; Calculus bovis factitius content is with cholic acid (C in the described calculus bovis factitius assay 24H 40O 5) meter, must not be less than 0.252mg/g.
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CN103529141A (en) * 2013-10-18 2014-01-22 山东阿如拉药物研究开发有限公司 Method for testing content of bilirubin in inclusion calculus bovis factitius
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CN105092725B (en) * 2014-05-16 2016-09-21 成都中医药大学 The HPLC detection method of Herba Gentianae Algidae
CN109932434A (en) * 2017-12-18 2019-06-25 河北万邦复临药业有限公司 The detection method of cholic acid content in a kind of Xihuang capsules
CN109060809A (en) * 2018-08-28 2018-12-21 西藏民族大学 A kind of Tibet red sandalwood quality standard and its detection method
CN109541131A (en) * 2019-01-07 2019-03-29 长春黄金研究院有限公司 The analysis method of copper and iron METHOD FOR CONTINUOUS DETERMINATION in a kind of ore

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