CN101244240B - Quantitative and qualitative analysis method for four turmeric soup preparations - Google Patents
Quantitative and qualitative analysis method for four turmeric soup preparations Download PDFInfo
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Abstract
The invention discloses a quantitative and qualitative analysis method for a tibetan traditional medicine of four-species curcuma longa decoction preparation, in particular to a quantitative and qualitative analysis method for the powder, pills, granules, capsules and tablets of the four-species curcuma longa decoction, wherein, the four-species curcuma longa decoction is prepared with curcuma longa, skin of barberry, fruit of emblic leafflower and puncture vine. The quality control method is characterized in that the curcuma longa, skin of barberry, fruit of emblic leafflower, and puncture vine in the compound preparation are identified with the method of thin layer chromatography; the content of curcumin is determined with the method of high-efficiency liquid chromatography; the content of berberine is determined with the method of capillary electrophoresis. The quality control method has the advantages of ensuring the rapid, accurate and advanced quality detection for four-species curcuma longa decoction, facilitating for the effective control of the internal quality of four-species curcuma longa decoction preparation, and enabling to be used as the index of investigating process stability.
Description
Technical field
The present invention relates to a kind of tibetan traditional medicine---the qualitative and quantitative analysis method of four turmeric soup preparations, be particularly related to the qualitative and quantitative analysis method of four turmeric soup powder, pill, granule, capsule and tablet, belong to the technical field of medicine being carried out quality control.
Technical background
SIWEI JIANGHUANG TANGSAN is the heat clearing away of recording in first of " Drug Standard of Ministry of Public Health of the Peoples Republic of China " Tibetan medicine that has, the diuresis function, cure mainly urethritis, frequent micturition, the tibetan traditional medicine of urgent micturition, other preparations of four turmeric soup are to carry out agent to change the medicine that obtains on the powder basis, comprise preparation series such as pill, granule, capsule, tablet.Its prescription is made up of Chinese crude drug Rhizoma Curcumae Longae 15g, Radix Berberidis Amurensis 12.5g, Fructus Phyllanthi 25g, Fructus Tribuli 25g.Former preparation quality standard only has the inspection of powder general rule, is not enough to control drug quality.
The present invention has proposed qualitative and quantitative analysis method to change products product pill, granule, capsule, tablet etc. of four turmeric soup powder mediating recipe, and the quality standard after the raising can effectively be controlled medicine, really embodies the safe and effective, quality controllable of medicine.
Summary of the invention
The objective of the invention is to: the qualitative and quantitative analysis method that a kind of four turmeric soup preparations is provided.Said preparation comprises powder, pill, granule, capsule and tablet.It is too simple to the present invention is directed to original preparation SIWEI JIANGHUANG TANGSAN quality standard, the unmanageable shortcoming of product quality, qualitative and quantitative analysis method to this preparation is studied, having set up the TLC thin layer of Rhizoma Curcumae Longae, Radix Berberidis Amurensis, Fructus Phyllanthi, Fructus Tribuli four medical materials differentiates, the high performance liquid chromatography quantitative analysis method of curcumin and the capillary electrophoresis quantitative analysis method of berberine hydrochloride have been increased, improve the quality control standard of four turmeric soup preparations, thereby guaranteed the clinical efficacy of this preparation.
Four turmeric soup preparations described in the present invention is all according to the prescription ratio preparation of SIWEI JIANGHUANG TANGSAN in first of " Drug Standard of Ministry of Public Health of the Peoples Republic of China " Tibetan medicine, the preparation method of its pill is: above four flavors or part flavour of a drug (all the other flavour of a drug waters decoct), be ground into fine powder, sieve mixing.Every 100g powder adds an amount of water or decocting liquid flooding ball with refined honey 35～50g, and drying is made water-honeyed pill; Or add refined honey 80～110g and make small honey pill or big honeyed pills, promptly; The preparation method of other dosage forms is: all or part of medical material water or aqueous alcohol reflux, extract, 1-3 time, filter, merging filtrate, concentrate or reclaim ethanol and become thick paste, with above-mentioned thick paste and part medical material fine powder or appropriate amount of auxiliary materials mixing, be prepared into granule, capsule or tablet according to diverse ways again.
Preparation of the present invention can add the medicine acceptable carrier as required when making preparation, these carriers can be any carriers that is fit to make preparation of the present invention.
Qualitative and quantitative analysis method of the present invention can be used for the tibetan traditional medicine preparation that above method is made, its step comprises or two in discriminating, two projects of assay, wherein differentiate comprise with Rhizoma Curcumae Longae control medicinal material, Radix Berberidis Amurensis control medicinal material and berberine hydrochloride reference substance, Fructus Phyllanthi control medicinal material, Fructus Tribuli control medicinal material be the thin layer of contrast differentiate part or all of; Assay comprises that with curcumin and/or berberine hydrochloride reference substance be the content assaying method of contrast.
Qualitative and quantitative analysis method of the present invention comprises:
Differentiate:
1. get four turmeric soup preparations, porphyrize adds in ethanol, methanol or the aqueous alcohol (all 〉=70%) any, and reflux filters, and filtrate is as need testing solution; Other gets the Rhizoma Curcumae Longae control medicinal material, shines medical material solution in pairs with legal system; Draw above-mentioned 2 kinds of solution respectively, put in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with benzene (toluene)-chloroform-ethanol (4-6: 2-4: 0.1-0.3) be developing solvent, launch, take out, dry, put under the ultra-violet lamp (365 or 254nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the fluorescence speckle of same color.
2. get the reflux extracting liquid under 1, evaporate to dryness, residue add ammonia solution 10-20ml makes dissolving, and reuse chloroform 7-15ml extraction divides and gets chloroform liquid, volatilizes, and residue adds ethanol makes dissolving, as need testing solution.Other gets the Radix Berberidis Amurensis control medicinal material, adds alcohol reflux, filters, and filtrate is medical material solution in contrast.Get the berberine hydrochloride reference substance again, add dissolve with ethanol, in contrast product solution; Draw above-mentioned 3 kinds of solution respectively, put in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with n-butyl alcohol-36% acetic acid-water (5-7: 1-3: 0.15-0.35) be developing solvent, launch, take out, dry, put under the ultra-violet lamp (365 or 254nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot.
3. get four turmeric soup preparations, porphyrize, any in adding ethanol, methanol or the aqueous alcohol (all 〉=70%), supersound process filters, and filtrate evaporate to dryness, residue add ethanol or methanol makes dissolving, as need testing solution.Other gets the Fructus Phyllanthi control medicinal material, shines medical material solution in pairs with legal system.Draw above-mentioned 2 kinds of solution respectively, the point in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid (4-6: 3-5: 0.5-2) be developing solvent, launch, take out, dry, spray is with 1-5% ferric chloride alcoholic solution, and it is clear that 100-110 ℃ of heating or hot blast blow to speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue spot.
4. get four turmeric soup preparations, porphyrize adds water boil, filters, and filtrate adds hydrochloric acid 0.5-2ml, heating in water bath 1 hour, and cooling is immediately extracted with chloroform 10-30ml, and extracting solution evaporate to dryness, residue add methanol makes dissolving, as need testing solution.Other gets the Fructus Tribuli control medicinal material, shines medical material solution in pairs with legal system.Draw above-mentioned 2 kinds of solution respectively, the point in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with cyclohexane extraction-ethyl acetate (10-20: 0.5-2) be developing solvent, launch, take out, dry, spray is with the 5-10% ethanol solution of sulfuric acid, and it is clear that 100-110 ℃ of heating or hot blast blow to speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color (purple); Put ultra-violet lamp (365 or 254nm) inspection down, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical fluorescent orange speckle.
Curcumin (C
21H
20O
6) assay
According to an appendix VI of Chinese Pharmacopoeia version in 2005 D high effective liquid chromatography for measuring.
The test of chromatographic condition and system suitability: with octadecylsilane chemically bonded silica is filler, with the second eyeball: 4% glacial acetic acid (43-53: 47-57) be mobile phase, detection wavelength 430 ± 2nm, number of theoretical plate should be not less than 4000 by the calculating of curcumin peak;
It is an amount of that the curcumin reference substance is got in the reference substance solution preparation, and accurate the title decides, and adds methanol or ethanol and makes the solution that every 1ml contains 2-20 μ g, promptly.
The need testing solution preparation: get four turmeric soup preparations, porphyrize is got about 0.02-0.2g, the accurate title, decide, and accurate any 10-50ml that adds in methanol, ethanol or the aqueous alcohol (all 〉=70%) claims to decide weight, supersound process (power 250W, frequency 40kHz) 30-90 minute, put coldly, claim to decide weight again, supply the weight that subtracts mistake with same solvent, shake up, high speed centrifugation or filter with 0.45 μ m filter membrane, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
In this four turmeric soup preparations, every 1g contains Rhizoma Curcumae Longae with curcumin (C
21H
20O
6) meter, must not be less than 0.40mg (capsule and tablet can be converted to every or every content according to different size weight).
Berberine (C
20H
18ClNO
4) assay
Measure according to an appendix VI of Chinese Pharmacopoeia version in 2005 D high performance capillary electrophoresis.
Electrophoretic separation condition: fused-silica capillary column: 58.5cm (effective length 50cm) * 75 μ mID; Running buffer: 0.2M sodium acetate: methanol=6-10: 1-3; Pressure sample introduction 10-30kPas; Working voltage 10-25kV; Detect λ=265 ± 2nm on the post; 20-40 ℃ of capillary tube column temperature; Wash capillary tube 1,2,3min respectively with 0.1M NaOH, water, running buffer between each sample introduction; Inner mark solution: every 1ml methanol or ethanol contain iodate methyl triphenyl phosphorus (methyl-triphenylphosphoniumiodide, content 97%) 4-10mg, press 5% of test liquid constant volume during use and add.
It is an amount of that the preparation precision of reference substance solution takes by weighing the berberine hydrochloride reference substance, and the accurate inner mark solution that adds is an amount of, makes the solution that every 1ml contains 20-60 μ g berberine hydrochloride and 200-500 μ g internal standard substance matter with methanol or ethanol, shakes up, promptly.
Four turmeric soup preparations 0.1-0.3g is got in the need testing solution preparation, the accurate title in the 10-50ml measuring bottle, accurate any 8-45ml that adds in inner mark solution 0.5-2.5ml and hydrochloric acid-methanol or the hydrochloric acid-ethanol (1: 100), supersound process (power 200-300W, frequency 30-50kHz) 30-90 minute, put cold, be settled to scale, shake up, high speed centrifugation or filter with 0.45 μ m filter membrane, get supernatant or filtrate, promptly.
Algoscopy places injector with need testing solution to be measured, and the setting operation parameter is measured.
In the four turmeric soup preparations, every 1g contains berberine with berberine hydrochloride (C
20H
18ClNO
4) meter, must not be less than 0.4mg (capsule and tablet can be converted to every or every content according to different size weight).
4 kinds of medical materials are differentiated described optimum developing solvent in the above-mentioned four turmeric soup preparations: Rhizoma Curcumae Longae is benzene (toluene)-chloroform one ethanol (methanol) (5: 3: 0.2); Radix Berberidis Amurensis is n-butyl alcohol-36% acetic acid-water (6: 2: 0.25); Fructus Phyllanthi is toluene-ethyl acetate-formic acid (5: 4: 1); Fructus Tribuli is cyclohexane extraction-ethyl acetate (15: 1).
Because crude drug source, the place of production, growth year and collecting season etc. there are differences, cause the content difference of preparation bigger, the present invention fixes tentatively minimum, and the content limit of actual preparation can be adjusted according to the data of many batches of production samples under fixing medical material place of production condition.
Above-mentioned test item can be selected partly or entirely to carry out as required, and its step need can also not carried out conventional sense simultaneously according to sequencing:
Character:
For powder: this product is yellow coarse powder; Feeble QI perfume (or spice), bitter in the mouth, slightly sweet.
For pill: this product is that yellowish-brown is to tan water-honeyed pill, small honey pill or big honeyed pills; Feeble QI, bitter in the mouth, slightly sweet.
For granule: this product is that yellowish-brown is to tan granule; Feeble QI, bitter in the mouth.
For capsule: this product content is that yellowish-brown is to tan powder or granule; Feeble QI, bitter in the mouth.
For tablet: this product is a Film coated tablets, is yellowish-brown to sepia after removing coating; Feeble QI, bitter in the mouth.
Check: should meet relevant every regulation under each dosage form item of general rule of Chinese Pharmacopoeia.
Characteristics of the present invention and advantage are: set up the determination methods that Rhizoma Curcumae Longae, Radix Berberidis Amurensis, Fructus Phyllanthi, Fructus Tribuli four Chinese medicine material have or not in the four turmeric soup preparations first; Set up the HPLC quantitative analysis method and the standard of effective ingredient curcumin in the four turmeric soup preparations first; Set up the HPCE quantitative analysis method and the standard of effective ingredient berberine in the four turmeric soup preparations first; The standardization of technology and the stable and controllable of quality have been guaranteed; In addition, from operation, simple, the accurate advanced person of qualitative and quantitative analysis method of the present invention.
Further narrate technical scheme of the present invention and effect below in conjunction with case study, the present invention is not limited only to described case study.
1. the thin layer Study on Identification of SIWEI JIANGHUANG TANGSAN
1.1 get this product 5g, add 70% ethanol 20ml, reflux 1 hour is put coldly, filters, and filtrate is as need testing solution.Other gets the negative control product 4g that lacks Rhizoma Curcumae Longae, makes negative control product solution with method.Get Rhizoma Curcumae Longae control medicinal material 0.5g again, add 70% ethanol 10ml, shine medical material solution in pairs with legal system.Draw above-mentioned 3 kinds of each 3ul of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with benzene-chloroform-ethanol (5: 3: 0.2), toluene-chloroform-methanol (5: 3: 0.2), chloroform-methanol (96: 4) and chloroform-methanol-formic acid (96: 4: 0.7) respectively, launch, take out, dry, put under the ultra-violet lamp (365 or 254nm) and inspect.The result shows, four kinds of development systems all obtain similar result, but when being developing solvent with benzene-chloroform-ethanol (5: 3: 0.2), in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot, and main speckle separating degree is good, negative control is noiseless, and the method favorable reproducibility is so list the quality standard text in.
1.2 get 70% ethanol extract 10ml under the Rhizoma Curcumae Longae discriminating item, evaporate to dryness, residue add ammonia solution 15ml makes dissolving, reuse chloroform 10ml extraction divides and gets chloroform liquid, volatilizes, and residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets the negative control product 1.8g that lacks Radix Berberidis Amurensis, adds 70% ethanol 10ml, makes negative control product solution with method.Get Radix Berberidis Amurensis control medicinal material 0.5g again, add ethanol 20ml, reflux 1 hour is put coldly, filters, and filtrate is medical material solution in contrast.Get the berberine hydrochloride reference substance again, add ethanol and make the solution that every 1ml contains 0.2mg, in contrast product solution.Draw each 2 μ l of above-mentioned 4 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, respectively with n-butyl alcohol-36% acetic acid-water (6: 2: 0.25), benzene-ethyl acetate-methanol-isopropyl alcohol-liquor ammoniae fortis (6: 3: 1.5: 1.5: 0.5, the ammonia vapo(u)rous) is developing solvent, launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot, the negative control product that lack Radix Berberidis Amurensis are noiseless.Two kinds of development system results are similar, but developing solvent n-butyl alcohol-36% acetic acid-water (6: 2: 0.25) composition is simple, and toxicity is little, so list the quality standard text in.
1.3 get this product 2g, add 70% ethanol 20ml, supersound process 15 minutes filters, and filtrate evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets the negative control product 2g that lacks Fructus Phyllanthi, makes negative control product solution with method.Get Fructus Phyllanthi control medicinal material 0.5g again, shine medical material solution in pairs with legal system.Draw each 2 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with toluene-ethyl acetate-formic acid (5: 4: 1), ethyl acetate-formic acid-water (8: 2: 2) respectively, launch, take out, dry, spray is with 1% ferric chloride alcoholic solution, and it is clear that hot blast blows to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue spot.The negative control product that lack Fructus Phyllanthi are noiseless.Two kinds of development system results are similar, but toluene-ethyl acetate-formic acid (5: 4: 1) developing solvent gained chromatogram Rf value is moderate, so list the quality standard text in.
1.4 get this product 3g, add water 30ml, boiled 30 minutes, supply moisture, filter with Cotton Gossypii, filtrate adds hydrochloric acid 1ml, heating in water bath 1 hour, cooling is immediately extracted with chloroform 20ml, and extracting solution evaporate to dryness, residue add methanol 0.5ml makes dissolving, as need testing solution.Other gets the negative control product 2.2g that lacks Fructus Tribuli, makes negative control product solution with method.Get Fructus Tribuli control medicinal material 1g again, shine medical material solution in pairs with legal system.Draw each 3 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with cyclohexane extraction-ethyl acetate (15: 1), normal hexane-ether (8: 2) respectively, launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color (purple); Put inspection under the ultra-violet lamp (365nm), in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical fluorescent orange speckle.Two kinds of development system results are similar, but cyclohexane extraction-ethyl acetate (15: 1) developing solvent gained chromatogram Rf value is moderate, so list the quality standard text in.
2. the high performance liquid chromatography content assaying method of curcumin research in the four turmeric soup ball
2.1 instrument and chromatographic condition high performance liquid chromatograph (Agilent 1100 series); Phenomenex Luna C
18Post (250mm * 4.6mm, 5 μ m); With second eyeball-4% glacial acetic acid (48: 52) is mobile phase; Detect wavelength 430nm.Number of theoretical plate is pressed the curcumin peak and is calculated, and should be not less than 4000.With this understanding, curcumin and other compositions reach baseline separation, and negative sample is noiseless (seeing accompanying drawing 1).
2.2 reagent and medicine four turmeric soup ball are purchased to Dardo County Tibetan medicine hospital; Rhizoma Curcumae Longae control medicinal material and curcumin reference substance (for assay with) all purchase in Nat'l Pharmaceutical ﹠ Biological Products Control Institute; Methanol, acetonitrile are chromatographically pure, and water is high purity water; All the other reagent are analytical pure.
2.3 linear relationship is investigated precision and is taken by weighing curcumin reference substance 5.6mg, puts in the 100ml measuring bottle, adds the dissolve with methanol standardize solution, shakes up, and promptly gets the reference substance stock solution.Precision is measured stock solution 0.4,0.8,1.2,1.6,2.0,3.2ml, put respectively in the 10ml measuring bottle, add methanol and be prepared into the reference substance solution that concentration is 2.24,4.48,6.72,8.96,11.2,17.92 μ g/ml, the accurate respectively 10 μ l that draw inject chromatograph of liquid.With the peak area integrated value sample introduction concentration is returned, get regression equation Y=121.62447X+0.7296053.The result shows: sample size is good linear relationship (r=0.9997) in 0.0224～0.179 μ g scope.
2.4 the required Cmin that enters chromatographic column calculated when minimum detectable activity was measured and be able to be produced the signal of 3 times of noises with detector.By baseline record and curcumin reference substance solution dilution sample introduction, determine that finally the minimum detectable activity of curcumin reference substance solution is 0.015 μ g/ml in the experiment.
2.5 the same curcumin reference substance solution of the accurate absorption of instrument precision test, repeat auto injection 5 times by above-mentioned chromatographic condition, record the peak area integrated value and be respectively 580.6,583.3,581.5,583.1,582.7, RSD=0.198% (n=5) illustrates that the sample introduction of instrument is good with detection precision.
2.6 the need testing solution study on the stability is got same need testing solution and is placed in room temperature condition, in 0,2,4,6,8,20, the 24h sample introduction measures its peak area, the peak area of measuring for 7 times is respectively 588.2,588.2,588.3,592.4,595.1,592.3,603.3 as a result, RSD is 0.919%, and it is basicly stable to illustrate that the need testing solution room temperature is placed 24h.
2.7 replica test is got with 5 parts of a collection of test samples, by " 2.9 " method operation down, and by " a 2.1 " chromatographic condition test down, as a result the content of 5 duplicate samples be respectively 0.943,0.937,0.981,0.973,0.964mg/g, average content is 0.960mg/g, and RSD is 1.98%.
2.8 the recovery test amount of reducing by half takes by weighing 9 parts of the same batch samples of known content, the accurate title, decide, respectively by the accurate a certain amount of curcumin reference substance solution (48.2 μ g/ml) that adds of gradient (0.8 times, 1 times, 1.2 times), press the pre-treating method processing down of " 2.9 " item, measure by chromatographic condition under " 2.1 " item.Average recovery rate is 99.4% as a result, and RSD is 1.46% (seeing Table 1).
2.9 it is an amount of that sample determination is got the four turmeric soup ball, porphyrize is got about 0.05g, the accurate title, decide, and the accurate methanol 10ml that adds claims to decide weight, supersound process (power 250W, frequency 40kHz) 60 minutes is put cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, high speed centrifugation, get supernatant, by chromatographic condition under " 2.1 " item, sample introduction 10 μ l measure.The every 1g of 3 batch samples contains that curcumin is respectively 0.960,1.05,0.918mg.
Table 2.1 sample pipetting volume recovery test (mg)
3. the capillary electrophoresis content assaying method of berberine research in the SIWEI JIANGHUANG TANGSAN
3.1 reagent and medicine reference substance berberine hydrochloride are available from SIGMA (calculating its content greater than 98% according to area normalization method) under this experiment condition, the Radix Berberidis Amurensis control medicinal material is available from Nat'l Pharmaceutical ﹠ Biological Products Control Institute.The SIWEI JIANGHUANG TANGSAN self-control; Water is high purity water (purchasing in southwestern physics Institute); Internal standard substance iodate methyl triphenyl phosphorus (methyl-triphenylphosphonium iodide, content 97%) is purchased the ACROS ORGANICS company in the U.S.; All the other reagent are analytical pure.
3.2 instrument and experiment condition Hewlett-Packard
3DCE capillary electrophoresis apparatus (diode array detector and HP chromatographic work station), elastic quartz capillary tube 58.5cm (effective length 50cm) * 75 μ m (Hebei sharp Feng chromatograph Yongnian device company limited); Detect wavelength 265nm; Pressure sample introduction 20kPas; Working voltage 15kV, 25 ℃ of column temperatures, buffer: 0.2molL
-1Sodium acetate-methanol (8: 2).Use 0.1molL respectively before each sample introduction
-1Sodium hydroxide solution, water, running buffer flushing capillary tube 1,2,3min.
Under this HPCE condition, berberine and other composition good separation, negative sample is noiseless (seeing accompanying drawing 2).
3.3 berberine hydrochloride is at 0.2molL in the ultra-violet absorption spectrum of berberine hydrochloride usefulness online detection reference substance of diode array detector (DAD) and the sample
-1Spectrum in sodium acetate-methanol (8: 2) buffer system, berberine hydrochloride has maximum absorption band at the 265nm place, so adopt the 265nm wavelength to detect.
3.4 linear relationship and lowest detectable limit precision take by weighing berberine hydrochloride reference substance 19.90mg, are settled to 50ml with dissolve with methanol, accurate respectively absorption 0.25,0.5,1.0,2.0,3.0 4.0ml places the 10ml volumetric flask, adds inner mark solution (0.6917g is dissolved in 100ml 70% methanol) 0.5ml, with hydrochloric acid-methanol (100: 1)] standardize solution, being mixed with concentration respectively is 9.95,19.9,39.8,79.6,119.4,159.2 μ gml
-1Reference substance solution, with the relative peak area Y (being the peak area ratio of measured object and internal standard substance) of berberine hydrochloride to berberine hydrochloride reference substance solution concentration C (mgL
-1) make standard curve.Its regression equation is:
Y＝0.0244C+0.0121 r＝0.9999
When sample size is 20kPas, equal 3 in signal to noise ratio, the lowest detection of berberine is limited to 1.9 μ gml
-1
3.5 condition determination optimization
3.5.1 the best applied voltage value of the selection separation system of working voltage is relevant with length with buffer concentration (ionic strength) and capillary inner diameter, does not produce too much Joule heat in order to use high voltage as far as possible, we have made the Ohm's law curve.Get 0.2molL
-1Sodium acetate-methanol (8: 2) buffer solution is by the different working voltages (3,5,8,10 that are provided with, 12,15,18,20kV) test its current value, draw the Ohm's law curve, the result begins the departs from linear section during greater than 15kV when voltage, and its corresponding electric current is bigger than normal, so definite 15kV is a working voltage.
3.5.2 the SIWEI JIANGHUANG TANGSAN sample solution is got in cleaning capillaceous between sample introduction, directly washes and use 0.1molL with running buffer between 2 sample introductions
-1Sodium hydroxide solution, water, running buffer clean 1,2 respectively, sample introduction behind the 3min, and result's kind mode later on cleans the berberine hydrochloride peak shape symmetry of mensuration, and the former hangover.So determine to use 0.1molL respectively between 2 sample introductions
-1Sodium hydroxide solution, water, running buffer flushing 1,2,3min.
3.6 it is 39.8 μ gml that concentration is got in the test of sample introduction precision
-1Berberine hydrochloride reference substance solution continuous sample introduction 5 times respectively, recording berberine hydrochloride relative peak area meansigma methods is 1.38, RSD=1.2%.
3.7 replica test takes by weighing the about 0.2g of SIWEI JIANGHUANG TANGSAN, totally 5 parts, presses operation under the sample determination item, its content of berberine average out to 3.14mgg
-1, RSD=1.65%.Measured this batch SIWEI JIANGHUANG TANGSAN once more at the 10th day in addition, its average content is 3.19mgg
-1(n=3, RSD=1.41%).More than 2 same date sample determination RSD=1.71% of average (n=8) as a result not, illustrate that this method repeatability is good.
3.8 it is 39.8 μ gml that stability experiment is got concentration
-1The berberine hydrochloride reference substance solution preparation same day 0,1,2,3,4,5,6,7,8h respectively measures 1 time, its relative peak area meansigma methods is 1.27, RSD=1.61% (n=9) illustrate that reference substance room temperature on the same day places the basic no change of mensuration concentration in the 8h.Standard solution with new preparation is measured the reference substance solution that refrigerator is placed January, and its compound concentration and mensuration concentration RSD=1.4% are basicly stable.
Other got the SIWEI JIANGHUANG TANGSAN sample solution the same day the 0th, 3,8, and 9h measures respectively 4 times, and its relative peak area meansigma methods is 1.31, RSD=1.58%; Measure once again next day (24h), its in the daytime the RSD of relative peak area be 1.67% (n=5).
3.9 application of sample reclaims experiment and gets known content (3.14mgg
-1) SIWEI JIANGHUANG TANGSAN 0.1g, accurate claim that 5 parts of operation repetitives add 119.4 μ gml in the 25ml measuring bottle
-1Berberine hydrochloride reference substance solution 3ml, inner mark solution (0.6917g is dissolved in 100ml 70% methanol) 1.25ml operates by 3.10 sample determination items method down, the average average recovery of result is 98.7%, RSD=2.4%.
3.10 sample determination is got SIWEI JIANGHUANG TANGSAN 0.2g, the accurate title, accurately add inner mark solution 1.25ml in the 25ml measuring bottle, with hydrochloric acid-methanol (1: 100) 20ml supersound extraction 60min, put to room temperature, be settled to scale, get about 1.5ml high speed centrifugation (14000rmin
-1) 10min, supernatant behind 0.45 μ m filtering with microporous membrane as sample solution.Measure by the experiment condition under 3.1.As a result three batch sample content of berberine hydrochloride be respectively 3.14,3.31,3.24mgg
-1
Description of drawings
Fig. 1 is a curcumin assay specificity HPLC spectrogram of the present invention;
Fig. 2 is a berberine hydrochloride content specificity HPCE spectrogram of the present invention.
Among Fig. 1, A-lacks Rhizoma Curcumae Longae feminine gender, B-sample, C-Rhizoma Curcumae Longae control medicinal material, D-Rhizoma Curcumae Longae raw medicinal material, E-curcumin reference substance
Among Fig. 2, A-lacks mark+Radix Berberidis Amurensis control medicinal material, the interior mark+berberine hydrochloride reference substance of E-in mark+Radix Berberidis Amurensis feminine gender in Radix Berberidis Amurensis feminine gender, the B-, the interior mark+sample of C-, the D-
The specific embodiment
The particulate qualitative and quantitative analysis method of embodiment 1 four turmeric soup
[prescription] Rhizoma Curcumae Longae 774.2g Radix Berberidis Amurensis 645.2g Fructus Phyllanthi 1290g Fructus Tribuli 1290g.
[method for making] above four flavors add 10 times of water gagings and soaked 30 minutes, decoct and extract 3 times; each 1.5 hours; decocting liquid merges, and filters, and filtrate is condensed into the clear paste that relative density is 1.10～1.15 (80 ℃ of heat are surveyed); be divided into two parts; portion is spray dried to extract powder, and as the parent nucleus of granulating, all the other clear paste are as adhesive; adopt one-step-granulating method to make the 1000g granule, promptly.
[character] this product is that yellowish-brown is to tan granule; Feeble QI, bitter in the mouth.
This product 5g is got in [discriminating] (1), and porphyrize adds 70% ethanol 20ml, and reflux 1 hour is put coldly, filters, and filtrate is as need testing solution.Other gets Rhizoma Curcumae Longae control medicinal material 0.5g, adds 70% ethanol 10ml, shines medical material solution in pairs with legal system.Draw each 3 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with toluene-chloroform-methanol (5: 3: 0.2), launch, taking-up is dried, and puts under the ultra-violet lamp (365nm) and inspects.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot.
(2) get Rhizoma Curcumae Longae and differentiate following 70% an ethanol extract 10ml, evaporate to dryness, residue add ammonia solution 15ml makes dissolving, reuse chloroform 10ml extraction, and branch is got chloroform liquid, volatilizes, and residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Berberidis Amurensis control medicinal material 0.5g, adds ethanol 20ml, and reflux 1 hour is put coldly, filters, and filtrate is medical material solution in contrast.Get the berberine hydrochloride reference substance again, add ethanol and make the solution that every 1ml contains 0.2mg, in contrast product solution.Draw each 1～2 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with n-butyl alcohol-36% acetic acid-water (6: 2: 0.25), launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot.
(3) get this product 2g, add 70% ethanol 20ml, supersound process 15 minutes filters, and filtrate evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Fructus Phyllanthi control medicinal material 0.5g, shines medical material solution in pairs with legal system.Draw each 1～2 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with toluene-ethyl acetate-formic acid (5: 4: 1), launch, take out, dry, spray is with 1% ferric chloride alcoholic solution, and it is clear that hot blast blows to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue spot.
(4) get this product powder 3g, add water 30ml, boiled 30 minutes, supply moisture, filter with Cotton Gossypii, filtrate adds hydrochloric acid 1ml, heating in water bath 1 hour, and cooling is immediately extracted with chloroform 20ml, and extracting solution evaporate to dryness, residue add methanol 0.5ml makes dissolving, as need testing solution.Other gets Fructus Tribuli control medicinal material 1g, shines medical material solution in pairs with legal system.Draw each 1～2 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with cyclohexane extraction-ethyl acetate (15: 1), launch, take out, dry, spray is with 10% ethanol solution of sulfuric acid, and it is clear to be heated to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the speckle of same color (purple); Put inspection under the ultra-violet lamp (365nm), in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical fluorescent orange speckle.
[inspection] should meet every regulation (appendix IC) relevant under granule item of Chinese Pharmacopoeia.
[assay] is according to an appendix VI of Chinese Pharmacopoeia version in 2005 D high effective liquid chromatography for measuring.
Chromatographic condition and system suitability test are filler with the octadecylsilane chemically bonded silica; With the second eyeball: 4% glacial acetic acid (48: 52) is a mobile phase, detects wavelength 430nm, and number of theoretical plate calculates by the curcumin peak should be not less than 4000;
It is an amount of that the preparation precision of reference substance solution takes by weighing the curcumin reference substance, adds methanol and make the solution that every 1ml contains 10 μ g, shakes up, promptly.
The four turmeric decoction particles is got in the need testing solution preparation, porphyrize, and it is fixed to get the accurate title of about 0.05g, the accurate methanol 10ml that adds, claim to decide weight, supersound process (power 250W, frequency 40kHz) 60 minutes, put cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with methanol, high speed centrifugation or filter with 0.45 μ m filter membrane, promptly.
Accurate respectively reference substance solution and each the 10 μ l of need testing solution of drawing of algoscopy inject chromatograph of liquid, measure, promptly.
The every 1g granule of this product contains Rhizoma Curcumae Longae with curcumin (C
21H
20O
6) meter, must not be less than 2.0mg.
The capsular qualitative and quantitative analysis method of embodiment 2 four turmeric soups
[prescription] Rhizoma Curcumae Longae 258.6g Radix Berberidis Amurensis 215.5g Fructus Phyllanthi 431g Fructus Tribuli 431g.
[method for making] above four flavors add 10 times of water gagings and soaked 30 minutes, decoct and extract 3 times; each 1.5 hours, decocting liquid merged, and filtered; filtrate is condensed into the clear paste that relative density is 1.10～1.15 (80 ℃ of heat are surveyed), is divided into two parts, and portion is spray dried to extract powder; as the parent nucleus of granulating; all the other clear paste adopt one-step-granulating method to make granule as adhesive, incapsulate; make 1000, promptly.
[character] this product content is that yellowish-brown is to tan powder or granule; Feeble QI, bitter in the mouth.
[discriminating] is with embodiment 1.
[inspection] should meet every regulation (appendix IL) relevant under capsule item of Chinese Pharmacopoeia.
[assay] measured according to an appendix VI of Chinese Pharmacopoeia version in 2005 D high performance capillary electrophoresis.
Electrophoretic separation condition: fused-silica capillary column: 58.5cm (effective length 50cm) * 75 μ mID; Running buffer: 0.2M sodium acetate: methanol=8: 2; Pressure sample introduction 20kPas; Working voltage 15kV; Detect λ=265nm on the post; 30 ℃ of capillary tube column temperatures; Wash capillary tube 1,2,3min respectively with 0.1M NaOH, water, running buffer between each sample introduction; Inner mark solution: every 1ml methanol or ethanol contain iodate methyl triphenyl phosphorus (methyl-triphenylphosphonium iodide, content 97%) 4mg, press 5% of test liquid constant volume during use and add.
It is an amount of that the preparation precision of reference substance solution takes by weighing the berberine hydrochloride reference substance, and the accurate inner mark solution that adds is an amount of, makes the solution that every 1ml contains 20 μ g berberine hydrochloride and 200 μ g internal standard substance matter with methanol or ethanol, shakes up, promptly.
Four turmeric soup preparations 0.2g is got in the need testing solution preparation, the accurate title in the 25ml measuring bottle, accurate any 20ml that adds in inner mark solution 1.25ml and hydrochloric acid-methanol or the hydrochloric acid-ethanol (1: 100), supersound process (power 250W, frequency 40kHz) 60 minutes, put cold, be settled to scale, shake up, high speed centrifugation or filter with 0.45 μ m filter membrane, get supernatant or filtrate, promptly.
Algoscopy places injector with need testing solution to be measured, and the setting operation parameter is measured.
Every of this product contains Radix Berberidis Amurensis with berberine hydrochloride (C
20H
18ClNO
4) meter, must not be less than 1.8mg.The qualitative and quantitative analysis method of embodiment 3 four turmeric soup tablets
[prescription] Rhizoma Curcumae Longae 258.6g Radix Berberidis Amurensis 215.5g Fructus Phyllanthi 431g Fructus Tribuli 431g.
[method for making] above four flavors add 10 times of water gagings and soaked 30 minutes, decoct and extract 3 times, and each 1.5 hours, decocting liquid merged, and filtered, and filtrate is concentrated in right amount, and drying adds right amount of auxiliary materials, makes granule, and drying is pressed into 1000, the bag film-coat, promptly.
[character] this product is a Film coated tablets, removes to show yellowish-brown behind the coating to sepia; Feeble QI, bitter in the mouth.
[discriminating] is with embodiment 1.
[inspection] should meet every regulation (appendix ID) relevant under tablet item of Chinese Pharmacopoeia.
[assay] method is with embodiment 1.
Per 1 of this product contains Rhizoma Curcumae Longae with curcumin (C
21H
20O
6) meter, must not be less than 2.0mg.
Per 1 of this product contains Radix Berberidis Amurensis with berberine hydrochloride (C
20H
18ClNO
4) meter, must not be less than 1.8mg.
The qualitative and quantitative analysis method of embodiment 4 four turmeric soup balls
[prescription] Rhizoma Curcumae Longae 193.5g Radix Berberidis Amurensis 161.2g Fructus Phyllanthi 322.6g Fructus Tribuli 322.6g
[method for making] above four flavors are ground into fine powder, sieve mixing.Every 100g powder adds an amount of water pill with refined honey 35g, and drying is made water-honeyed pill 1000g, or adds refined honey 80～110g and make small honey pill or big honeyed pills, promptly.
[character] this product is that yellowish-brown is to tan water-honeyed pill, small honey pill or big honeyed pills; Feeble QI, bitter in the mouth, slightly sweet.
This product 5g is got in [discriminating] (1), and porphyrize adds 70% ethanol 20ml, reflux 1 hour, put coldly, filter, filtrate is got this product 5g, porphyrize, add 70% ethanol 20ml, reflux 1 hour is put cold, filter, get filtrate 10ml, evaporate to dryness, residue adds ammonia solution 15ml makes dissolving, and reuse chloroform 10ml extraction divides and gets chloroform liquid, volatilize, residue adds ethanol 1ml makes dissolving, as need testing solution.Other gets Radix Berberidis Amurensis control medicinal material 0.5g, adds ethanol 20ml, and reflux 1 hour is put coldly, filters, and filtrate is medical material solution in contrast.Get the berberine hydrochloride reference substance again, add ethanol and make the solution that every 1ml contains 0.2mg, in contrast product solution.Draw each 1～2 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with n-butyl alcohol-36% acetic acid-water (6: 2: 0.25), launch, take out, dry, put under the ultra-violet lamp (365nm) and inspect.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot.
(2) get this product 2g, porphyrize adds 70% ethanol 20ml, and supersound process 15 minutes filters, and filtrate evaporate to dryness, residue add ethanol 1ml makes dissolving, as need testing solution.Other gets Fructus Phyllanthi control medicinal material 0.5g, shines medical material solution in pairs with legal system.Draw each 1～2 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, be developing solvent with toluene-ethyl acetate-formic acid (5: 4: 1), launch, take out, dry, spray is with 1% ferric chloride alcoholic solution, and it is clear that hot blast blows to the speckle colour developing.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue spot.
[inspection] should meet every regulation (appendix IA) relevant under pill item of Chinese Pharmacopoeia.
[assay] is with embodiment 1.
The every 1g pill of this product contains Rhizoma Curcumae Longae with curcumin (C
21H
20O
6) meter, must not be less than 1.9mg.
The qualitative and quantitative analysis method of embodiment 5 SIWEI JIANGHUANG TANGSAN
[prescription] Rhizoma Curcumae Longae 193.5g Radix Berberidis Amurensis 161.2g Fructus Phyllanthi 322.6g Fructus Tribuli 322.6g
[method for making] above four flavors are ground into coarse powder, sieve, and mixing, promptly.
[character] this product is yellow coarse powder; Feeble QI perfume (or spice), bitter in the mouth, slightly sweet.
[discriminating] is with embodiment 2.
[inspection] should meet every regulation (appendix IB) relevant under powder item of Chinese Pharmacopoeia.
[assay] is with embodiment 1.
The every 1g powder of this product contains Rhizoma Curcumae Longae with curcumin (C
21H
20O
6) meter, must not be less than 1.9mg.
Claims (1)
1. the qualitative and quantitative analysis method of a tibetan traditional medicine four turmeric soup preparations, described preparation is powder, pill, granule, capsule or tablet, it is characterized in that: described qualitative and quantitative analysis method is by differentiating and two item designs of assay; Wherein differentiate be selected from Rhizoma Curcumae Longae control medicinal material, Radix Berberidis Amurensis control medicinal material and berberine hydrochloride reference substance, Fructus Phyllanthi control medicinal material, Fructus Tribuli control medicinal material be the thin layer of contrast differentiate part or all of; Assay is for being the content assaying method of contrast with curcumin reference substance and/or berberine hydrochloride reference substance; Wherein:
A, discriminating are selected from one or more in the following method:
(1) the Rhizoma Curcumae Longae medical material is differentiated with thin layer chromatography and is got four turmeric soup preparations 3-7g in the said preparation, porphyrize, add ethanol, methanol or 〉=any 10-50ml in 70% aqueous alcohol, reflux 30-90 minute, put coldly, filter, filtrate is as need testing solution; Other gets Rhizoma Curcumae Longae control medicinal material 0.3-0.7g, shines medical material solution in pairs with legal system; Test according to an appendix VI of Chinese Pharmacopoeia version in 2005 B thin layer chromatography, draw each 1～5 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with benzene-chloroform-ethanol 4-6: 2-4: 0.1-0.3, toluene-chloroform-methanol 4-6: 2-4: 0.1-0.3, chloroform-methanol 93-99: 3-5 and chloroform-methanol-formic acid 93-99: 3-5: any among the 0.5-0.9 is developing solvent, launch, take out, dry, put 365 or the 254nm ultra-violet lamp under inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot;
(2) the Radix Berberidis Amurensis medical material is differentiated the reflux extracting liquid 5-15ml that gets under (1) item with thin layer chromatography in the said preparation, and evaporate to dryness, residue add ammonia solution 10-20ml makes dissolving, reuse chloroform 7-15ml extraction divides and gets chloroform liquid, volatilizes, residue adds ethanol 1-2ml makes dissolving, as need testing solution; Other gets Radix Berberidis Amurensis control medicinal material 0.3-0.7g, add ethanol, methanol or 〉=any 10-30ml in 70% aqueous alcohol, reflux 30-90 minute, put coldly, filter, filtrate is medical material solution in contrast; Get the berberine hydrochloride reference substance again, add ethanol and make the solution that every 1ml contains 0.1-0.3mg, in contrast product solution; Test according to an appendix VI of Chinese Pharmacopoeia version in 2005 B thin layer chromatography, draw each 1-3 μ l of above-mentioned 3 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with n-butyl alcohol-36% acetic acid-water 5-7: 1-3: 0.15-0.35, benzene-ethyl acetate-methanol-isopropyl alcohol-liquor ammoniae fortis 5-7: 1-4: 1-2: 1-2: any among the 0.3-0.7 is developing solvent, launch, take out, dry, put 365 or the 254nm ultra-violet lamp under inspect, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical yellow fluorescence principal spot;
(3) the Fructus Phyllanthi medical material is differentiated with thin layer chromatography and is got four turmeric soup preparations 1-3g in the said preparation, add ethanol, methanol or 〉=any 10-50ml in 70% aqueous alcohol, supersound process 15-45 minute, filter, filtrate evaporate to dryness, residue add ethanol 1-2ml makes dissolving, as need testing solution; Other gets Fructus Phyllanthi control medicinal material 0.3-0.7g, shines medical material solution in pairs with legal system; Test according to an appendix VI of Chinese Pharmacopoeia version in 2005 B thin layer chromatography, draw each 1-3 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with toluene-ethyl acetate-formic acid 4-6: 3-5: 0.5-2, ethyl acetate-formic acid-water 6-10: 1-3: any among the 1-3 is developing solvent, launch, take out, dry, spray is with 1-5% ferric chloride alcoholic solution, it is clear that 100-110 ℃ of heating or hot blast blow to speckle colour developing, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical blue spot;
(4) the Fructus Tribuli medical material is got four turmeric soup preparations 1-5g with the thin layer chromatography discriminating in the said preparation, add water 10-50ml, boiled 10-60 minute, supply moisture, filter with Cotton Gossypii, filtrate adds hydrochloric acid 0.5-2ml, heating in water bath 1 hour, cooling immediately, extract with chloroform 10-30ml, extracting solution evaporate to dryness, residue add methanol 0.5-1ml makes dissolving, as need testing solution; Other gets Fructus Tribuli control medicinal material 0.5-2g, shines medical material solution in pairs with legal system; Draw each 1-5 μ l of above-mentioned 2 kinds of solution, put respectively in same be on the silica gel g thin-layer plate of adhesive with the sodium carboxymethyl cellulose, with cyclohexane extraction-ethyl acetate 10-20: 0.5-2, normal hexane-ether 6-10: 1-3 is developing solvent, launches, and takes out, dry, spray is with the 5-10% ethanol solution of sulfuric acid, and it is clear that 100-110 ℃ of heating or hot blast blow to the speckle colour developing, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show the purple dot of same color; Put 365 or the 254nm ultra-violet lamp under inspection, in the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, show identical fluorescent orange speckle;
B, assay are selected from one or both in the following method:
(1) with high effective liquid chromatography for measuring curcumin C
21H
20O
6The content chromatographic condition: with octadecylsilane chemically bonded silica is filler, and with the second eyeball: 4% glacial acetic acid 43-53: 47-57 is a mobile phase, detects wavelength 420-440nm, and number of theoretical plate calculates by the curcumin peak should be not less than 4000; Reference substance solution: every 1ml methanol or ethanol contain 2-20 μ g curcumin; The need testing solution preparation: get four turmeric soup preparations, porphyrize is got about 0.02-0.2g, the accurate title, decide, accurate add methanol, ethanol or 〉=any 10-50ml in 70% aqueous alcohol, claim to decide weight, power 200-300W, frequency 30-50kHz supersound process 30-90 minute, put cold, claim again to decide weight, supply the weight that subtracts mistake, shake up with same solvent, high speed centrifugation or filter with 0.45 μ m filter membrane, promptly; Accurate respectively reference substance solution and each 5-20 μ l of need testing solution of drawing injects hplc determination content, and in this four turmeric soup preparations, every 1g contains Rhizoma Curcumae Longae with curcumin C
21H
20O
6Meter must not be less than 0.40mg, and capsule and tablet can be converted to every or every content according to different size weight;
(2) with the berberine electrophoretic separation condition in the high performance capillary electrophoresis mensuration four turmeric soup preparations: fused-silica capillary column: 58.5cm * 75 μ mID, effective length 50cm; Running buffer: 0.2M sodium acetate: methanol=6-10: 1-3; Pressure sample introduction 10-30kPas; Working voltage 10-20kV; Detect λ=255-275nm on the post; 20-40 ℃ of capillary tube column temperature; Wash capillary tube 1,2,3min respectively with 0.1M NaOH, water, running buffer between each sample introduction; Inner mark solution: every 1ml methanol or ethanol contain 97% iodate methyl triphenyl phosphorus 4-10mg, press 5% of test liquid constant volume during use and add; Reference substance solution: every 1ml methanol or the hydrochloric berberine 20-60 of ethanol μ g and 97% iodate methyl triphenyl phosphorus 200-500 μ g; Need testing solution: get four turmeric soup preparations 0.1-0.3g, the accurate title in the 10-50ml measuring bottle, accurate any 8-45ml that adds in inner mark solution 0.5-2.5ml and hydrochloric acid-methanol or the hydrochloric acid-ethanol 1: 100, power 200-300W, frequency 30-50kHz supersound process 30-90 minute, put coldly, be settled to scale, shake up, high speed centrifugation or filter with 0.45 μ m filter membrane is got supernatant or filtrate, promptly; Algoscopy: need testing solution to be measured is placed injector, and the setting operation parameter is measured, and in the four turmeric soup preparations, every 1g contains Radix Berberidis Amurensis with berberine hydrochloride C
20H
18ClNO
4Meter must not be less than 0.4mg, and capsule and tablet can be converted to every or every content according to different size weight;
Described preparation is made by following Chinese medicine raw materials by weight proportion: Rhizoma Curcumae Longae 15g, Radix Berberidis Amurensis 12.5g, Fructus Phyllanthi 25g, Fructus Tribuli 25g.
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| CN102879515A (en) * | 2012-08-07 | 2013-01-16 | 何述金 | Mass control method for medicinal composition for treating hepatitis |
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| CN111579671A (en) * | 2020-05-26 | 2020-08-25 | 上海市农业科学院 | Method for determining content of three curcumins in turmeric |
| CN113813354A (en) * | 2021-10-28 | 2021-12-21 | 西藏神猴药业有限责任公司 | Tibetan medicine four-flavor turmeric soup dropping pill and preparation method thereof |
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