CN105403628A - Detection method for berberine hydrochloride content in stomach regulating and pain stopping capsule - Google Patents
Detection method for berberine hydrochloride content in stomach regulating and pain stopping capsule Download PDFInfo
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- CN105403628A CN105403628A CN201410473310.5A CN201410473310A CN105403628A CN 105403628 A CN105403628 A CN 105403628A CN 201410473310 A CN201410473310 A CN 201410473310A CN 105403628 A CN105403628 A CN 105403628A
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Abstract
The present invention discloses a detection method for the berberine hydrochloride content in a stomach regulating and pain stopping capsule, and belongs to the technical field of pharmaceutical component detection. According to the detection method, the chromatographic conditions comprise that an octadecylsilane chemically bonded silica gel is adopted as a filler, an acetonitrile-0.033 mol/L potassium dihydrogen phosphate solution (30:70) is adopted as a mobile phase, and the detection wavelength is 265 nm; and the method comprises: precisely weighing a berberine reference substance, placing into a measuring flask, adding a hydrochloric acid-methanol (1:100) solution, diluting, uniformly shaking, diluting with the mobile phase, uniformly shaking, taking a subject stomach regulating and pain stopping capsule content, uniformly mixing, precisely adding hydrochloric acid-methanol (1:100), precisely weighing, heating for 15 min in a 60 DEG C water bath, taking out, carrying out heating ultrasonic treatment for 30 min, placing overnight at a room temperature, precisely weighing, supplementing the lost weight with hydrochloric acid-methanol (1:100), uniformly shaking, diluting with the mobile phase, respectively and precisely sucking the reference substance solution and the subject solution, injecting into a liquid chromatograph, recording the chromatogram, and calculating the peak area according to an external standard method so as to obtain the content.
Description
Technical field
The present invention relates to drug ingedient detection technique field, be specifically related to the detection method with content of berberine hydrochloride in stomach and alleviating pain capsule.
Background technology
Add taste optimization with stomach and alleviating pain capsule according to " Zuojin Wan " in Yuan Dynasty's " danxi's experiential therapy " to form, through fundamental research for many years and clinical verification, effect such as quickly alleviating pain, anti-inflammatory antiulcer that existing Western medicine stomach medicine has, have the effect of Chinese medicine across-the-board regulation functions of intestines and stomach concurrently, the spinoff of Western medicine can be overcome again, remarkable to gastrointestinal disease result for the treatment of.Show through clinical verification with stomach and alleviating pain capsule: the effect with clearing liver and stomach, stopping nausea and vomiting by lowering the adverse flow of QI, regulating qi-flowing for relieving pain, to alleviation liver-stomach disharmony type stomach trouble, the symptoms such as gastral cavity pain, belch acid regurgitation have good result for the treatment of.This product because of its prescription unique, evident in efficacy, therefore since launch products, to be popular in users.Wherein, containing in a kind of daily life in the composition with stomach and alleviating pain capsule, common curative drug, the main composition containing Berberine hydrochloride in this medicine.Though formulated the detection method of content to Berberine hydrochloride in this medicine in standard, accuracy is low, poor sensitivity, effectively can not embody the quality of preparation.
Such as, existing with the detection method of content of berberine hydrochloride in stomach and alleviating pain capsule be: adopt thin layer scanning method, namely utilize sample to recycle thin-layer chromatogram scanner after launching through thin layer plate and chromatographic scan is carried out to sample point, cubage is carried out by the intensity of chromatographic absorption, this method affects comparatively large by extraneous factor, as temperature when the thickness of thin layer plate, sample launch etc., all can produce larger impact to result, accuracy is poor, and needs to consume a large amount of man power and materials.
Application number is in the Chinese patent of " 201310588401.9 ", disclose the patent of invention that name is called the detection method of Berberine hydrochloride " in the eight taste swertia pill ", its technical scheme is as follows: precision takes the Berberine hydrochloride reference substance scheduled volume of dry 24h in phosphorus pentoxide vacuum drying apparatus, and the methanol hydrochloride solution adding 1% makes the reference substance solution of every 1mL containing 0.01mg; Get eight taste swertia pill porphyrizes and get 2.5g, add the methanol hydrochloride solution scheduled volume of 1%, reflux heating 1 hour, by extracting liquid filtering in 100ml measuring bottle, with the methanol hydrochloride solution washing residue of 1%, be diluted to scale and shake up; Get extract 5ml, be placed on alumina column; With ethanol elution, collect eluent, put in 25ml measuring bottle, add ethanol and be diluted to scale and shake up, get in 2ml to 10ml measuring bottle, the methanol hydrochloride solution with 1% is diluted to scale and shakes up, and through 0.45 μm of filtering with microporous membrane, obtains need testing solution; Draw contrast solution and each 10ul of need testing solution, injection liquid chromatography measures.Although the present invention has the advantages such as accuracy is high, highly sensitive, specificity is strong, the preparation of its need testing solution is too loaded down with trivial details, spends a large amount of resources and energy, and need testing solution preparation efficiency is too low, and accuracy is not high.
Application number is in the Chinese patent of " 201310056321.9 ", disclose the patent of invention that name is called " pre-treating method that a kind of Berberine Hydrochloride Tablets carries out HPLC detection ", the technical scheme wherein relating to mobile phase is as follows: described mobile phase is that phosphate solution and acetonitrile are formulated according to volume ratio 60:40, described phosphate solution is that the potassium dihydrogen phosphate of 0.05mol/L and the sodium heptanesulfonate of 0.05mol/L mix with volume ratio 1:1, containing 0.2% triethylamine, and by phosphoric acid adjust ph to 3.0.Although produce the mobile phase obtained in this technical scheme there is colour developing and dilution function, chromatographic purity of its colour developing is poor, after the chromatogram that develops the color can there is other assorted peaks interference.
Summary of the invention
In order to solve the problem, the invention discloses the detection method with content of berberine hydrochloride in stomach and alleviating pain capsule, it has the features such as accuracy is high, highly sensitive, specificity is strong, and its technological process is more energy-efficient.
Detection method with content of berberine hydrochloride in stomach and alleviating pain capsule, comprises the following steps:
A. chromatographic condition and system suitability test: take octadecylsilane chemically bonded silica as filling agent, with acetonitrile-0.033mol/L potassium dihydrogen phosphate (30:70) for mobile phase, determined wavelength is 265nm, and theoretical cam curve calculates should be not less than 3000 by Berberine hydrochloride peak value;
B. the preparation of control sample solution: precision takes Berberine hydrochloride reference substance and is about 12.5mg, be positioned in 50mol measuring bottle, add hydrochloric acid-methanol (1:100) solution and be diluted to scale, shake up, then precision measures in the measuring bottle of dilution 2ml to 25ml, be diluted to scale with mobile phase, shake up, to obtain final product;
C. the preparation of need testing solution: get and stomach and alleviating pain capsule 's content, mixing, get about 0.8g accurately weighed, precision adds hydrochloric acid-methanol (1:100) 50ml, accurately weighed weight, and heats 15 minutes in 60 DEG C of water-baths, take out, heating (50 DEG C-60 DEG C) ultrasonic process (power 250W, frequency 25KHz) 30 minutes, left at room temperature over night, accurately weighed weight again, supplement the weight of less loss with hydrochloric acid-methanol (1:100), shake up, filter, precision measures in subsequent filtrate 2ml to 25ml measuring bottle, be diluted to scale with mobile phase, shake up, to obtain final product;
D. determination method: accurate absorption reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography, record chromatogram, by external standard method with calculated by peak area, to obtain final product.
Further, the concrete grammar of steps d determination method is as follows:
(1) precision draws reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography;
(2) judge in peak table according to the display figure of chromatogram in liquid chromatograph, whether have other Component peak area to overlap;
(3) calculate with peak face amount by external standard method.
The invention has the advantages that: mobile phase adopts acetonitrile-0.033mol/L potassium dihydrogen phosphate (30:70), have the advantages that chromatographic purity is high, meanwhile, adopt exact instrument to take sample, its accuracy is high, adopt ultrasonic process to heat, heating and temperature control is accurate, can save manpower man-hour, improve and produce efficiency, prepare gained test sample highly sensitive, therefore the inventive method has the features such as accuracy is high, highly sensitive, specificity is strong, and the feature such as its technological process is more energy-efficient.
Accompanying drawing explanation
In order to be illustrated more clearly in the embodiment of the present invention or technical scheme of the prior art, be briefly described to the accompanying drawing used required in embodiment or description of the prior art below, apparently, accompanying drawing in the following describes is only some embodiments of the present invention, for those of ordinary skill in the art, under the prerequisite not paying creative work, other accompanying drawing can also be obtained according to these accompanying drawings.
Fig. 1 is the HPLC chromatogram of reference substance solution;
Fig. 2 is the HPLC chromatogram of need testing solution;
Fig. 3 is the HPLC chromatogram of negative controls solution.
Embodiment
The invention discloses the detection method with content of berberine hydrochloride in stomach and alleviating pain capsule, it has the features such as accuracy is high, highly sensitive, specificity is strong, and its technological process is more energy-efficient.
Below in conjunction with the accompanying drawing in the present invention, be clearly and completely described the technical scheme in the present invention, obviously, described is only the present invention's part embodiment, instead of whole embodiments.Based on the embodiment in the present invention, the every other embodiment that those of ordinary skill in the art obtain under the prerequisite of not making creative work, all belongs to the scope of protection of the invention.
Detection method with content of berberine hydrochloride in stomach and alleviating pain capsule, comprises the following steps:
A. chromatographic condition and system suitability test: take octadecylsilane chemically bonded silica as filling agent, with acetonitrile-0.033mol/L potassium dihydrogen phosphate (30:70) for mobile phase, determined wavelength is 265nm, and theoretical cam curve calculates should be not less than 3000 by Berberine hydrochloride peak value;
B. the preparation of control sample solution: precision takes Berberine hydrochloride reference substance and is about 12.5mg, be positioned in 50mol measuring bottle, add hydrochloric acid-methanol (1:100) solution and be diluted to scale, shake up, then precision measures in the measuring bottle of dilution 2ml to 25ml, be diluted to scale with mobile phase, shake up, to obtain final product;
C. the preparation of need testing solution: get and stomach and alleviating pain capsule 's content, mixing, get about 0.8g accurately weighed, precision adds hydrochloric acid-methanol (1:100) 50ml, accurately weighed weight, and heats 15 minutes in 60 DEG C of water-baths, take out, heating (50 DEG C-60 DEG C) ultrasonic process (power 250W, frequency 25KHz) 30 minutes, left at room temperature over night, accurately weighed weight again, supplement the weight of less loss with hydrochloric acid-methanol (1:100), shake up, filter, precision measures in subsequent filtrate 2ml to 25ml measuring bottle, be diluted to scale with mobile phase, shake up, to obtain final product;
D. determination method: accurate absorption reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography, record chromatogram, by external standard method with calculated by peak area, to obtain final product.
Wherein, the concrete grammar of steps d determination method is as follows:
(1) precision draws reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography;
Judge in peak table according to the display figure of chromatogram in liquid chromatograph, whether have other Component peak area to overlap;
(2) calculate with peak face amount by external standard method.
Be below specific embodiment:
1, instrument and reagent
High performance liquid chromatograph (AgilentTechnologies1260infinity), Ultraviolet Detector (AgilentTechnologies), 100,000/balance, ten thousand/balance, Yi Lite chromatographic column (SinoChromODS-BP), Berberine hydrochloride reference substance solution and stomach and alleviating pain capsule 's content, acetonitrile-0.033mol/L potassium dihydrogen phosphate (30:70), deionized water.
2, test process and result
System suitability: get reference substance solution, need testing solution and negative sample solution, analyze, the results are shown in Figure 1, Fig. 2, Fig. 3, and wherein, Fig. 1 is control sample solution Berberine hydrochloride chromatographic peak; The retention time of Berberine hydrochloride is about 13 minutes, and the degree of separation of Berberine hydrochloride and other component peaks is all greater than 1.5, and negative controls solution is noiseless to mensuration.
A, linear relationship are investigated: precision takes Berberine hydrochloride reference substance and is about 10.2mg(in 86.7%), put in 50ml measuring bottle, add hydrochloric acid ?methyl alcohol (1: 100) and dissolve and be diluted to scale, shake up, as stock solution.Accurate absorption reference substance solution 1,2,3,4,5mL, puts respectively in 25mL volumetric flask, adds mobile phase to scale, shake up, and each precision draws 20 μ L, respectively injection liquid chromatography, measures peak area.Take peak area as ordinate, take mass concentration as horizontal ordinate drawing standard curve, the equation of linear regression of Berberine hydrochloride is: y=795.6x-0.29r=0.999(n=5), shows that Berberine hydrochloride is good with peak area linear relationship within the scope of 0.0071 ~ 0.0354mg/mL.
B, stability test: draw same need testing solution 20 μ L, measure in 0,1,3,6,8h.Data as shown in Table, results peaks area RSD=0.7%(n=5), show that need testing solution 8 hours at normal temperatures internal stabilities are good.
C, precision test: the accurate reference substance solution 20 μ L drawing 0.0354mg/mL, continuous sample introduction 6 times.Data as shown in Table, the RSD=0.2%(n=6 of results peaks area), show that instrument precision is good.
D, replica test: get same batch sample 6 parts, by need testing solution preparation method, prepare solution and measure.As shown in Table, Berberine hydrochloride average content is 8.21mg/ grain to result, RSD=0.2%(n=6), show that the precision that method operates is good.
E, average recovery are tested: precision takes 5 parts, the sample that known content is (20.02mg/g) and stomach and alleviating pain capsule, every part of about 0.3g, accurately add Berberine hydrochloride reference substance solution (3.0mg/ml) each 2ml respectively, prepare solution by need testing solution preparation method and measure, calculate the recovery, as shown in Table, average recovery rate is 98.8% to result, RSD=0.6%, shows that the accuracy of the method is higher.
F, sample size measure
By the obtained 3 batches of need testing solutions respectively of method under need testing solution preparation, accurate absorption need testing solution 20 μ L respectively, sample introduction successively, measures.Every batch of need testing solution measures 3 times, averages, and calculate the content of Berberine hydrochloride according to following computing formula by the integrating peak areas value recorded:
The peak area ÷ test sample weight of the heavy ÷ reference substance solution of peak area × extension rate × grain of test sample content=reference substance solution concentration × need testing solution
Measurement result is in Table.
Berberine hydrochloride content result (n=3) in table sample product
Known by describing above, the invention has the advantages that: mobile phase adopts acetonitrile-0.033mol/L potassium dihydrogen phosphate (30:70), have the advantages that chromatographic purity is high, simultaneously, exact instrument is adopted to take sample, its accuracy is high, ultrasonic process is adopted to heat, heating and temperature control is accurate, manpower man-hour can be saved, improve and produce efficiency, prepare gained test sample highly sensitive, therefore the inventive method has the features such as accuracy is high, highly sensitive, specificity is strong, and the feature such as its technological process is more energy-efficient.
Above content is in conjunction with concrete preferred implementation further description made for the present invention, can not assert that specific embodiment of the invention is confined to these explanations.For general technical staff of the technical field of the invention, without departing from the inventive concept of the premise, some simple deduction or replace can also be made, all should be considered as belonging to protection scope of the present invention.
Claims (2)
1., with the detection method of content of berberine hydrochloride in stomach and alleviating pain capsule, it is characterized in that comprising the following steps:
A, chromatographic condition and system suitability test: take octadecylsilane chemically bonded silica as filling agent, with acetonitrile-0.033mol/L potassium dihydrogen phosphate (30:70) for mobile phase, determined wavelength is 265nm, and theoretical cam curve calculates should be not less than 3000 by Berberine hydrochloride peak value;
The preparation of b, control sample solution: precision takes Berberine hydrochloride reference substance and is about 12.5mg, be positioned in 50mol measuring bottle, add hydrochloric acid-methanol (1:100) solution and be diluted to scale, shake up, then precision measures in the measuring bottle of dilution 2ml to 25ml, be diluted to scale with mobile phase, shake up, to obtain final product;
The preparation of c, need testing solution: get and stomach and alleviating pain capsule 's content, mixing, get about 0.8g accurately weighed, precision adds hydrochloric acid-methanol (1:100) solution 50ml, accurately weighed weight, is placed in 60 DEG C of water-bath heating 15 minutes, take out, heating (50 DEG C-60 DEG C) ultrasonic process (power 250W, frequency 25KHz) 30 minutes, left at room temperature over night, accurately weighed weight again, supply the weight of less loss with hydrochloric acid-methanol (1:100), shake up, filter, precision measures in subsequent filtrate 2ml to 25ml measuring bottle, be diluted to scale with mobile phase, shake up, to obtain final product;
D, determination method: accurate absorption reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography, record chromatogram, by external standard method with calculated by peak area, to obtain final product.
2. the detection method of content of berberine hydrochloride according to claim 1 and stomach and alleviating pain capsule, it is characterized in that, the concrete grammar of steps d determination method is as follows:
(1) precision draws reference substance solution and each 20 μ l of need testing solution respectively, injection liquid chromatography;
(2) judge in peak table according to the display figure of chromatogram in liquid chromatograph, whether have other Component peak area to overlap;
(3) calculate with peak area value by external standard method.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110187030A (en) * | 2019-06-12 | 2019-08-30 | 贵州联盛药业有限公司 | The measuring method of content of berberine hydrochloride in a kind of gram of leaching open capsule |
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| CN101721530A (en) * | 2009-11-18 | 2010-06-09 | 广西汇科药物研究有限责任公司 | Method for controlling quality of Jinji preparation |
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| 余少婷 等: "HPLC测定黄连上清片中盐酸小檗碱的含量", 《中成药》 * |
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN110187030A (en) * | 2019-06-12 | 2019-08-30 | 贵州联盛药业有限公司 | The measuring method of content of berberine hydrochloride in a kind of gram of leaching open capsule |
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