Summary of the invention: the objective of the invention is to: YUGAN dragon pharmaceutical preparation and preparation method thereof and method of quality control that treatment hepatitis is provided; This pharmaceutical preparation mainly comprises capsule, the drop pill with characteristics such as covering bitterness, easy to carry, good mouthfeel, absorption are fast, steady quality.
The present invention constitutes like this: it is mainly by Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g or the effervescent tablet that is made with their extract of corresponding weight portion, injection, powder pin, freeze-dried powder, gel, dispersible tablet, capsule, soft capsule, microcapsule, granule, pill, micropill, powder, drop pill, slow releasing preparation, controlled release preparation, gel, oral liquid, soft extract, extractum and membrane.Concrete preparation can be capsule, drop pill.
The preparation method of described YUGAN dragon pharmaceutical preparation: after XIAONIEGEN decocts with water 2 hours, adding Herba Artemisiae Scopariae, Radix Bupleuri, Radix Scutellariae, Herba Taraxaci are heated to the back of boiling and add Radix Arnebiae (Radix Lithospermi), decoct secondary, be followed successively by 3 hours, 2 hours, gradation filters, merging filtrate, left standstill 12 hours, and drew supernatant, drying, be ground into fine powder, make different preparations then respectively.
Capsule in the described preparation prepares like this: after XIAONIEGEN decocts with water 2 hours, adding Herba Artemisiae Scopariae, Radix Bupleuri, Radix Scutellariae, Herba Taraxaci are heated to the back of boiling and add Radix Arnebiae (Radix Lithospermi), decoct secondary, are followed successively by 3 hours, 2 hours, gradation filters, merging filtrate left standstill 12 hours, drew supernatant, get about 3/4 medicine liquid spray drying, make extract powder, get the residue medicinal liquid again and do binding agent, boiling granulating in fluid bed: feed liquor speed 40~45ml/min; Atomisation pressure 0.15Mpa; 50~55 ℃ of temperature of charge; 58~68 ℃ of inlet temperature; 50~55 ℃ of leaving air temps incapsulate, promptly.
The preparation method of described YUGAN dragon pharmaceutical preparation can also be carried out like this: Herba Artemisiae Scopariae, radix bupleuri extract volatile oil, and standby; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add the medicinal residues after Radix Scutellariae, Herba Taraxaci and Herba Artemisiae Scopariae, the radix bupleuri extract volatile oil again, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter, collecting decoction and above-mentioned aqueous solution, concentrating under reduced pressure, drying sprays into volatile oil or volatile oil clathrate compound, mixing is made different preparations then respectively.
Capsule in the described preparation also can prepare like this: Herba Artemisiae Scopariae, radix bupleuri extract volatile oil; Get β-CD, add 6 times of water gagings, under the ultrasound wave of 35KHz, in β-CD: the ratio of volatile oil=8: 1 slowly drips the mixed volatilization oil solution with ethanol dilution, and ultrasonic 40min takes out drying; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add the medicinal residues after Radix Scutellariae, Herba Taraxaci and Herba Artemisiae Scopariae, the radix bupleuri extract volatile oil again, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter collecting decoction and above-mentioned aqueous solution, get about 3/4 medicine liquid spray drying, make extract powder, get the residue medicinal liquid again and do binding agent, boiling granulating in fluid bed: feed liquor speed 40~45ml/min; Atomisation pressure 0.15Mpa; 50~55 ℃ of temperature of charge; 58~68 ℃ of inlet temperature; 50~55 ℃ of leaving air temps are granulated, and add volatile oil clathrate compound, incapsulate, promptly.
Drop pill in the described preparation prepares like this: Herba Artemisiae Scopariae, radix bupleuri extract volatile oil; Get β-CD, add 6 times of water gagings, under the ultrasound wave of 35KHz, in β-CD: the ratio of volatile oil=8: 1 slowly drips the mixed volatilization oil solution with ethanol dilution, and ultrasonic 40min takes out drying; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add Radix Scutellariae again, Herba Taraxaci and Herba Artemisiae Scopariae, medicinal residues after the radix bupleuri extract volatile oil, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter collecting decoction and above-mentioned aqueous solution, medicine liquid spray drying, mix with volatile oil clathrate compound, with the Macrogol 4000 is substrate, and according to medicine: the part by weight of substrate=1: 3 adds Macrogol 4000, mixing, the employing internal diameter is 3.0mm, external diameter is the dropper of 4.0mm, drip 50 ℃ of system temperature, dripping speed is 20~30d/min, dripping apart from being 6cm, splash in the long cooling column of 120cm, is liquid coolant again with the methyl-silicone oil, pill, promptly.
The method of quality control of YUGAN dragon of the present invention pharmaceutical preparation, it comprises assay, discriminating etc.; Wherein assay is index with the baicalin, differentiates and adopts berberine hydrochloride, Radix Bupleuri control medicinal material to be contrast.Specifically: assay is such: with methanol: 0.025mol/L phosphoric acid=45: 55 is mobile phase; The detection wavelength is 278nm; 40 ℃ of column temperatures; It is an amount of that precision takes by weighing the baicalin reference substance, adds methanol and make the solution that every 1ml contains 0.039mg, promptly gets reference substance solution; Get this product 15g, porphyrize is got 1g, accurate claims surely, adds methanol 20ml, supersound process, and power 250W, frequency 33kHz, 30 minutes, put coldly, filter, filtrate moves in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly gets need testing solution; Accurate respectively reference substance solution and the need testing solution drawn of algoscopy injects chromatograph of liquid, measures, promptly.Discriminating is such: (1) gets this product 30g, and porphyrize adds methanol 50ml, supersound process 20 minutes, put coldly, filter, filtrate evaporate to dryness, residue add water 20ml makes dissolving, in the dislocation separatory funnel, extract 2 times with the ether jolting, each 20ml discards ether solution, extract 2 times with water saturated n-butyl alcohol jolting, each 30ml merges n-butanol extracting liquid, wash 2 times with ammonia solution, each 20ml discards ammonia solution, get n-butyl alcohol liquid evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 2g, adds water 40ml, decocts 30 minutes, filters, and gets filtrate, presses the need testing solution preparation method from " extracting 2 times with the ether jolting " operation, makes control medicinal material solution; According to the thin layer chromatography test, draw above-mentioned two kinds of each 5﹠amp of solution; Micro; L puts respectively on same silica gel g thin-layer plate, and with chloroform: methanol: water=65: 35: 10 lower floor's solution is developing solvent, launches, and takes out, and dries, and spray is heated to clear spot with 40% sulphuric acid ethanol liquid of 2% paradime thylaminobenzaldehyde at 60 ℃.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, should show the principal spot of same color; (2) get [assay] item need testing solution 20ml down, evaporate to dryness, residue add ethanol 2ml makes dissolving, gets supernatant as need testing solution.Other gets the berberine hydrochloride reference substance, add ethanol and make the solution that every 1ml contains 0.2mg, product solution according to the thin layer chromatography test, is drawn above-mentioned two kinds of solution in contrast, put respectively on same silica gel g thin-layer plate, with benzene: ethyl acetate: isopropyl alcohol: methanol: dense ammoniacal liquor=12: 6: 3: be developing solvent at 3: 1, launches under the saturated ammonia steam, takes out, dry, put under the ultra-violet lamp 365nm and inspect; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.
Compared with prior art, the YUGAN of this treatment hepatitis provided by the invention dragon novel formulation has the comparison notable therapeutic effect for hepatitis disease; Can be used for treatment of conditions such as acute, chronic hepatitis, first cirrhosis, edema; That the medicament form of pharmaceutical preparation that provides has is easy to carry, cover bitterness, absorb and can wait characteristics well soon; Overcome the problem that prior art, product exist; Preparation method that provides and method of quality control can reasonablely instruct enterprise to carry out the quality and the curative effect of suitability for industrialized production, assurance product; Reached the purpose of invention.
The applicant under study for action, in order to prevent volatile component scattering and disappearing in storage process in the preparation, increase stability of drug, improve curative effect of medication etc., volatile oil is made the beta-schardinger dextrin-(clathrate of β-CD), optimize the key factor that influences the ultrasonic method enclose (ratio, the ultrasonic time of the ratio of supersonic frequency, water and β-CD, oil and β-CD) by experiment, worked out the working condition of suitable this product.In research process, find, the baicalin character instability of one of effective ingredient in the product, easy oxidation deterioration, so when molding, want strict control temperature, therefore the applicant has carried out a series of experiments, selects the supplementary product kind of preparation process condition, use of the most suitable pharmaceutical preparation of the present invention and consumption, ratio etc.; Guarantee its science, reasonable, feasible; The preparation that obtains has superperformance and therapeutic effect.
Experimental example 1: Study on Forming
(1) spray drying
1. supersonic frequency/KHz oil: β-CD/mL: g water: β-CD/mL: g time/h inclusion rate %
30????????????????1∶6???????????????4∶1??????????????40????????85.3
30????????????????1∶6???????????????4∶1??????????????45????????78.9
35????????????????1∶6???????????????6∶1??????????????50????????90.1
35????????????????1∶8???????????????6∶1??????????????40????????95.2
40????????????????1∶8???????????????8∶1??????????????45????????88.3
40????????????????1∶8???????????????8∶1??????????????50????????86.4
The result gets β-CD as can be known, adds 6 times of water gagings, and under the ultrasound wave of 35KHz, in β-CD: the ratio of volatile oil=8: 1 slowly drips the mixed volatilization oil solution with ethanol dilution, ultrasonic 40min.
(2) capsule
1. the fluidized granulating important technological parameters is investigated
Feed liquor speed (ml/min) | ??55~60 | ??50~55 | ??40~45 | ??35~40 |
Atomisation pressure (Mpa) steam pressure (Mpa) temperature of charge (℃) EAT (℃) leaving air temp (℃) granulation situation scutelloside loss late % | 0.15 0.5 65~60 98~88 50~45 particles are inhomogeneous, easily bond 22% | 0.15 0.5 60~55 80~100 60~65 uniform particles easily granulate 15% | 0.15 0.5 50~55 58~68 50~55 uniform particles easily granulate 4% | 0.15 0.5 50~45 68~58 35~30 uniform particles, but granulation time long 3% |
Experimental result as can be known, optimised process: feed liquor speed 40~45ml/min; Atomisation pressure 0.15Mpa; 50~55 ℃ of temperature of charge; 58~68 ℃ of inlet temperature; 50~55 ℃ of leaving air temps, the high loss of effective components of temperature is big, and it is long that it's the low granulation time is past temperature.
(3) drop pill
Drip apart from, drip the selection of speed, temperature: evaluation index: the heavy qualification rate of ball is by mass discrepancy requirement of Pharmacopoeia of the People's Republic of China version in 2000: meet ± 7.5% within.
Group temperature/℃ drip a distance/cm drips the ball weight qualification rate/% baicalin loss rate % of speed/(dmin-1)
1????????70?????????4??????????20~30????????????76.7??????????????10.3
2????????70?????????6??????????30~40????????????87.8??????????????12.2
3????????70?????????8??????????40~60????????????83.4??????????????10.7
4????????60?????????4??????????30~40????????????91.2??????????????13.8
5????????60?????????6??????????40~60????????????90.1??????????????17.9
6????????60?????????8??????????20~30????????????92.3??????????????19.6
7????????50?????????4??????????40~50????????????90.2??????????????12.8
8????????50?????????6??????????20~30????????????95.6??????????????5.6
9????????50?????????8??????????30~40????????????91.0??????????????14.2
The result as can be known, best art for coating: with the Macrogol 4000 is substrate, according to medicine: the part by weight of substrate=1: 3 adds Macrogol 4000, mixing, the employing internal diameter is that 3.0mm, external diameter are the dropper of 4.0mm, and dripping system temperature 50 ℃, droplet speed is that 20~30d/min, a distance are 6cm, splashes in the long cooling column of 120cm, be liquid coolant with the methyl-silicone oil again, loss of effective components is little.
(4) study on the stability (accelerated tests)
Loss of volatile oil rate %
Group March 2 month January
Volatile oil is enclose drop pill 11.5 13.6 14.7 not
Drop pill 2.4 3.5 3.8 of the present invention
Volatile oil is enclose capsule 15.6 18.2 18.9 not
Capsule 2.8 3.9 4.1 of the present invention
Syrup 41.4 61.2 79.7
Granule 21.0 25.4 36.8
The result as can be known, technology of the present invention can effectively reduce the loss of volatile oil rate.
Experimental example 2: pharmacodynamic experiment
1, to the inhibiting research of HBV
Use the P2.0HBV plasmid, through enzyme action, electrophoresis, the dna fragmentation of collection 7.0kb, this fragment contains the full gene of HBV of 2 connected head-to-tail 3.2kb and the PBR322 carrier DNA of 0.6kb, collect dna fragmentation, quantitative through electrophoresis method, under aseptic condition, be dissolved in respectively in the TE buffer, concentration is the 1mg/L packing, and microinjection is treated in-20 ℃ of preservations.Get the pure strain mice, injection contains the full geneome plasmid of HBV, and the mice that is produced is G0 generation, with G0 for integrate positive mice with the normal female Mus copulation of strain, the birth G1 for mice, detect positive person again with the G2 generation of being given birth to the copulation of strain normal female mice, so obtain G3 for mice.Get G3 for 250 of mices, 6~8 ages in week are about body weight 20g.At first organize screening, the positive person of tissue detection HBVDNA waits to do to detect in the further blood, and HBVDNA is positive in the blood, is the HBVDNA transgenic mouse.250 mices filter out the transgenic mice that contains HBVDNA in 40 serum.Get with 8 of strain normal mouses, as the normal control group, other 40 are equally divided into 5 groups, are respectively model group, commercially available YUGANLONG TANGJIANG group, commercially available YUGANLONG KELI group, Capsules group of the present invention, soft capsule group of the present invention.The treatment group is irritated stomach and is given the relative medicine medicinal liquid, is equivalent to medicine 50g/kg.Normal control group and model group are irritated the equal-volume normal saline, once a day, and around being total to.
After the last medication 2 hours, eyeball is put to death after getting blood, and this separation of blood sampling serum is to be checked.Cut open the belly rapidly and get the mouse liver piecemeal, preserve in-70 ℃ to be checked behind the liquid nitrogen freezing respectively.Detect transgenic mice serum HBV DNA content with the PCR quantitative method, the cutoff of detection is 1.0 * 10
5Copy/ml.Adopt the content of detected by dot blot hybridization murine liver tissue HBVDNA; The rapid extraction of recombiant plasmid HBV is pressed alkaline lysis; Dot blot hybridization: get 40 μ la-32P-DNA probe mark liquid point films, in degeneration liquid, soak, the degeneration caudacoria is put 80 ℃ of baking boxs dried in 2 hours,, judge positive degree with every speckle OD value through pre-paying, wash film, intermediate plate.
1. the influence that HBV DNA in the HBVTGM blood is changed, concrete outcome sees the following form.
HBVDNA content (1.0 * 10 in the HBVTGM blood
5Copy/ml) variation
The group n HBV dna content in the routine number blood of turning out cloudy
Normal control group 8 0-
Model group 80 8.896 ± 0.97
YUGANLONG TANGJIANG group 82 6.595 ± 2.34
YUGANLONG KELI group 83 6.218 ± 3.68
Capsules group 83 5.482 ± 2.55 of the present invention
Soft capsule group 84 5.967 ± 1.98 of the present invention
The explanation of above experimental result, use commercially available YUGANLONG TANGJIANG, commercially available YUGANLONG KELI, capsule of the present invention, soft capsule of the present invention after, all have the HBV DNA of mice to turn out cloudy, and serum HBV DNA reduces than the model group significance.
2. to the influence of HBVTGM hepatic tissue HBV dna content, concrete outcome sees the following form
Influence to HBVTGM hepatic tissue HBV dna content
The group n routine number HBV DNA dot blot hybridization of turning out cloudy
Normal control group 800
Model group 80 0.468 ± 0.138
YUGANLONG TANGJIANG group 82 0.312 ± 0.105
YUGANLONG KELI group 83 0.303 ± 0.085
Capsules group 83 0.281 ± 0.069 of the present invention
Soft capsule group 84 0.275 ± 0.109 of the present invention
Above presentation of results, the content of back hepatocyte HBV DNA is starkly lower than model group in 4 week of medication.And the content of HBV DNA can obviously reflect the content of hepatocyte virus.That is to say that YUGANLONG TANGJIANG, YUGANLONG KELI, capsule of the present invention, soft capsule of the present invention all can effectively suppress hepatitis B virus, and the effect of product of the present invention is better than YUGANLONG TANGJIANG and YUGANLONG KELI.
The specific embodiment:
Embodiments of the invention 1: Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g
After XIAONIEGEN decocts with water 2 hours, adding Herba Artemisiae Scopariae, Radix Bupleuri, Radix Scutellariae, Herba Taraxaci are heated to the back of boiling and add Radix Arnebiae (Radix Lithospermi), decoct secondary, are followed successively by 3 hours, 2 hours, gradation filters, merging filtrate left standstill 12 hours, drew supernatant, get about 3/4 medicine liquid spray drying, make extract powder, get the residue medicinal liquid again and do binding agent, boiling granulating in fluid bed: feed liquor speed 40~45ml/min; Atomisation pressure 0.15Mpa; 50~55 ℃ of temperature of charge; 58~68 ℃ of inlet temperature; 50~55 ℃ of leaving air temps incapsulate, and promptly get capsule, this product oral, three times on the one, each 2.
Embodiments of the invention 2: Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g
Herba Artemisiae Scopariae, radix bupleuri extract volatile oil; Get β-CD, add 6 times of water gagings, under the ultrasound wave of 35KHz, in β-CD: the ratio of volatile oil=8: 1 slowly drips the mixed volatilization oil solution with ethanol dilution, and ultrasonic 40min takes out drying; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add the medicinal residues after Radix Scutellariae, Herba Taraxaci and Herba Artemisiae Scopariae, the radix bupleuri extract volatile oil again, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter collecting decoction and above-mentioned aqueous solution, get about 3/4 medicine liquid spray drying, make extract powder, get the residue medicinal liquid again and do binding agent, boiling granulating in fluid bed: feed liquor speed 40~45ml/min; Atomisation pressure 0.15Mpa; 50~55 ℃ of temperature of charge; 58~68 ℃ of inlet temperature; 50~55 ℃ of leaving air temps are granulated, and add volatile oil clathrate compound, incapsulate, and promptly get capsule.
Embodiments of the invention 3: Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g
Herba Artemisiae Scopariae, radix bupleuri extract volatile oil; Get β-CD, add 6 times of water gagings, under the ultrasound wave of 35KHz, in β-CD: the ratio of volatile oil=8: 1 slowly drips the mixed volatilization oil solution with ethanol dilution, and ultrasonic 40min takes out drying; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add Radix Scutellariae again, Herba Taraxaci and Herba Artemisiae Scopariae, medicinal residues after the radix bupleuri extract volatile oil, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter collecting decoction and above-mentioned aqueous solution, medicine liquid spray drying, mix with volatile oil clathrate compound, with the Macrogol 4000 is substrate, and according to medicine: the part by weight of substrate=1: 3 adds Macrogol 4000, mixing, the employing internal diameter is 3.0mm, external diameter is the dropper of 4.0mm, drip 50 ℃ of system temperature, dripping speed is 20~30d/min, dripping apart from being 6cm, splash in the long cooling column of 120cm, is liquid coolant again with the methyl-silicone oil, pill promptly gets drop pill.
Embodiments of the invention 4: Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g
After XIAONIEGEN decocted with water 2 hours, adding Herba Artemisiae Scopariae, Radix Bupleuri, Radix Scutellariae, Herba Taraxaci were heated to the back of boiling and add Radix Arnebiae (Radix Lithospermi), decoct secondary, are followed successively by 3 hours, 2 hours, gradation filters, and merging filtrate left standstill 12 hours, draws supernatant, drying is ground into fine powder, adds distilled water, promptly gets oral liquid.
Embodiments of the invention 5: Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g
Herba Artemisiae Scopariae, radix bupleuri extract volatile oil, standby; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add the medicinal residues after Radix Scutellariae, Herba Taraxaci and Herba Artemisiae Scopariae, the radix bupleuri extract volatile oil again, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter, collecting decoction and above-mentioned aqueous solution, concentrating under reduced pressure, drying sprays into volatile oil or volatile oil clathrate compound, add carbomer, make gel.
Embodiments of the invention 6: Herba Artemisiae Scopariae 500g, XIAONIEGEN 500g, Radix Bupleuri 300g, Herba Taraxaci 250g, Radix Scutellariae 150g, Radix Arnebiae (Radix Lithospermi) 33g
Herba Artemisiae Scopariae, radix bupleuri extract volatile oil, standby; Aqueous solution after distillation device is in addition collected, and is standby; Radix Berberidis Amurensis was decocted 2 hours, add the medicinal residues after Radix Scutellariae, Herba Taraxaci and Herba Artemisiae Scopariae, the radix bupleuri extract volatile oil again, boil the back and add Radix Arnebiae (Radix Lithospermi), decoct secondary, 3 hours for the first time, 2 hours for the second time, filter, collecting decoction and above-mentioned aqueous solution, concentrating under reduced pressure, drying, spray into volatile oil or volatile oil clathrate compound, add sodium carboxymethyl cellulose, tabletting is made dispersible tablet.
Embodiments of the invention 7: assay
With methanol: 0.025mol/L phosphoric acid=45: 55 is mobile phase; The detection wavelength is 278nm; 40 ℃ of column temperatures; It is an amount of that precision takes by weighing the baicalin reference substance, adds methanol and make the solution that every 1ml contains 0.039mg, promptly gets reference substance solution; Get this product 15g, porphyrize is got 1g, accurate claims surely, adds methanol 20ml, supersound process, and power 250W, frequency 33kHz, 30 minutes, put coldly, filter, filtrate moves in the 25ml measuring bottle, adds methanol and is diluted to scale, shakes up, and filters, and gets subsequent filtrate, promptly gets need testing solution; Accurate respectively reference substance solution and the need testing solution drawn of algoscopy injects chromatograph of liquid, measures, promptly.
Embodiments of the invention 8: differentiate
(1) get this product 30g, porphyrize adds methanol 50ml, supersound process 20 minutes, put coldly, filter, filtrate evaporate to dryness, residue add water 20ml makes dissolving, in the dislocation separatory funnel, extract 2 times with the ether jolting, each 20ml discards ether solution, extract 2 times with water saturated n-butyl alcohol jolting, each 30ml merges n-butanol extracting liquid, wash 2 times with ammonia solution, each 20ml discards ammonia solution, get n-butyl alcohol liquid evaporate to dryness, residue adds methanol 1ml makes dissolving, as need testing solution.Other gets Radix Bupleuri control medicinal material 2g, adds water 40ml, decocts 30 minutes, filters, and gets filtrate, presses the need testing solution preparation method from " extracting 2 times with the ether jolting " operation, makes control medicinal material solution; According to the thin layer chromatography test, draw above-mentioned two kinds of each 5﹠amp of solution; Micro; L puts respectively on same silica gel g thin-layer plate, and with chloroform: methanol: water=65: 35: 10 lower floor's solution is developing solvent, launches, and takes out, and dries, and spray is heated to clear spot with 40% sulphuric acid ethanol liquid of 2% paradime thylaminobenzaldehyde at 60 ℃.In the test sample chromatograph, with the corresponding position of control medicinal material chromatograph on, should show the principal spot of same color;
(2) get [assay] item need testing solution 20ml down, evaporate to dryness, residue add ethanol 2ml makes dissolving, gets supernatant as need testing solution.Other gets the berberine hydrochloride reference substance, add ethanol and make the solution that every 1ml contains 0.2mg, product solution according to the thin layer chromatography test, is drawn above-mentioned two kinds of solution in contrast, put respectively on same silica gel g thin-layer plate, with benzene: ethyl acetate: isopropyl alcohol: methanol: dense ammoniacal liquor=12: 6: 3: be developing solvent at 3: 1, launches under the saturated ammonia steam, takes out, dry, put under the ultra-violet lamp 365nm and inspect; In the test sample chromatograph, with the corresponding position of reference substance chromatograph on, show the fluorescence speckle of same color.