CN104950053A - Method for measuring content of impurities in cefuroxime axetil tablet through high performance liquid chromatography - Google Patents
Method for measuring content of impurities in cefuroxime axetil tablet through high performance liquid chromatography Download PDFInfo
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Abstract
The invention discloses a method for measuring the content of impurities in a cefuroxime axetil tablet through high performance liquid chromatography. The method includes the following steps: a high performance liquid chromatograph is characterized in that a fixing phase serves as an octadecyl silane chromatographic column, and the flowing phase is that the ratio of a 0.1-0.2 mol/L ammonium dihydrogen phosphate solution to methyl alcohol is 60:40-64:36. The column temperature ranges from 25 DEG C to 35 DEG C, and the detection wave length ranges from 278 nm to 280 nm. 20 microliters of a blank solution, 20 microliters of an impurity solution and 20 microliters of a test solution are respectively sucked to be injected into the liquid chromatograph, and a chromatogram map is recorded. Through calculation, the impurity reference substance solution concentration value is X, the corresponding peak area value is Y, linear fitting is carried out, a linear regression equation is obtained, and a remarkable linear relation is formed between the linear range and the peak response value of the linear range when the impurity ranges from 2.0136 micrograms per milliliter to 7.5510 micrograms per milliliter, the impurity B ranges from 1.4826 micrograms per milliliter to 5.5599 micrograms per milliliter, the impurity C ranges from 0.5050 micrograms per milliliter to 1.8936 micrograms per milliliter, the impurity D ranges from 0.5155 micrograms per milliliter to 1.9330 micrograms per milliliter, and the impurity E ranges from 0.4993 micrograms per milliliter to 1.8723 micrograms per milliliter. The method is used for measuring the impurity A, the impurity B, the impurity C, the impurity D and the impurity E in cefuroxime axetil bulk pharmaceutical chemicals, the separating efficiency is high, the analysis speed is high, and the detection flexibility is high.
Description
Technical field
The present invention relates to high-efficient liquid phase chromatogram technology field, be specifically related to the method for impurity content in a kind of high-performance liquid chromatogram determination cefuroxime axetil tablets.
Background technology
Cefuroxime axetil tablets is produced, and owing to can be subject to the impact of different condition in storage, may degrade and produce different impurity, need carry out limit handling to it.Therefore we establish the HPLC-UV determination method of impurity content in cefuroxime axetil tablets, better detect the impurity that may exist in cefuroxime axetil tablets, better to control the quality of cefuroxime axetil tablets.
Summary of the invention
The object of the invention is to set up a kind of method measuring impurity content in cefuroxime axetil tablets, better the impurity that may exist in cefuroxime axetil tablets is detected, better to control the quality of cefuroxime axetil tablets.
Technical scheme of the present invention is: the method for impurity content in high-performance liquid chromatogram determination cefuroxime axetil tablets, it comprises the steps:
The preparation of impurity storing solution: get CEFUROXIME AXETIL, impurity A, impurity B, impurity C, impurity D, impurity E methyl alcohol dissolves respectively and be diluted to every 1mL respectively about containing the solution of CEFUROXIME AXETIL 0.250mg, impurity A 0.50mg, impurity B 0.37mg, impurity C 0.12mg, impurity D 0.12mg, impurity E 0.12mg.
The preparation of impurity test solution: get respectively and be settled to scale with mobile phase in above-mentioned each impurity storing solution 1ml to 100ml measuring bottle and shake up, to obtain final product.
The preparation of need testing solution: it is appropriate that precision takes cefuroxime axetil tablets, adds mobile phase after dissolving be settled to scale with methyl alcohol, makes every 1mL about containing the solution of CEFUROXIME AXETIL 0.250mg.
The preparation of reference substance solution: precision measures need testing solution 1ml, puts in 100ml measuring bottle, adds mobile phase and is diluted to scale, shake up.
The preparation of blank solution: get right amount of auxiliary materials, adds mobile phase after dissolving be settled to scale with methyl alcohol.
The detecting device of described high performance liquid chromatograph is ultraviolet absorption detector, and determined wavelength is 278nm.
Described mobile phase is 0.2mol/L biphosphate ammonium salt solution ﹕ methyl alcohol=62 ﹕ 38.
The invention has the beneficial effects as follows: impurity content in cefuroxime axetil tablets surveyed by the present invention's application high performance liquid chromatography, and separation efficiency is high, analysis speed is fast, detection sensitivity is high.By detecting impurity content in cefuroxime axetil tablets, controlling that impurity A in every gram of CEFUROXIME AXETIL must not be greater than 2.0%, impurity B must not be greater than 1.5%, other single impurity must not be greater than 1.0%, is conducive to the quality better controlling cefuroxime axetil tablets.
Accompanying drawing illustrates: Fig. 1 is impurity test solution chromatogram;
Fig. 2 is blank auxiliary test color spectrogram;
Fig. 3 is the linear regression graph of CEFUROXIME AXETIL.
Form is described in further detail content of the present invention more by the following examples, but should not be interpreted as in the above-mentioned subject area of the present invention at this point and be only limitted to following examples.Do not departing under the above-mentioned technology prerequisite of the present invention, the corresponding replacement made according to ordinary skill knowledge and customary means or the amendment of change, include within the scope of the invention
.
Embodiment 1: the determination of chromatographic column.
Instrument: high performance liquid chromatograph Waters e2695-2998, diode array detector;
Chromatographic column: octadecyl silane is filler (4.6 × 150mm 5 μ),
Flow velocity: 1.0 ml/min;
Sample size: 20 μ l;
Column temperature: 30 DEG C;
Wavelength: 278nm;
Mobile phase: 0.2mol/L biphosphate ammonium salt solution ﹕ methyl alcohol=62:38.
The preparation of impurity test solution: get CEFUROXIME AXETIL, impurity A, impurity B, impurity C, impurity D, impurity E methyl alcohol dissolves respectively and be diluted to every 1mL respectively about containing the solution of CEFUROXIME AXETIL 0.00250mg, impurity A 0.0050mg, impurity B 0.0037mg, impurity C 0.0012mg, impurity D 0.0012mg, impurity E 0.0012mg.
Get impurity test solution sample introduction record chromatogram.
Impurity A went out peak, impurity B and went out peak, impurity C at 33.070,40.637 minutes and go out at 3.686 minutes that peak, impurity D went out peak at 8.128 minutes, impurity E went out peak at 9.658 minutes, main peak went out peak at 16.185,18.614 minutes, saw Fig. 1 at 22.177 minutes.
Conclusion: impurity A, B, C, D, E and main peak can be separated completely under the process conditions.
Embodiment 2: the determination of determined wavelength.
Instrument: high performance liquid chromatograph Waters e2695-2998, diode array detector;
Chromatographic column: octadecyl silane is filler (4.6 × 150mm 5 μ);
Flow velocity: 1.0ml/min;
Sample size: 20 μ l;
Column temperature: 30 DEG C;
Wavelength: 190 ~ 400nm;
Mobile phase: 0.2mol/L biphosphate ammonium salt solution ﹕ methyl alcohol=62:38.
The preparation of impurity test solution: get CEFUROXIME AXETIL, impurity A, impurity B, impurity C, impurity D, impurity E methyl alcohol dissolves respectively and be diluted to every 1mL respectively about containing the solution of CEFUROXIME AXETIL 0.00250mg, impurity A 0.0050mg, impurity B 0.0037mg, impurity C 0.0012mg, impurity D 0.0012mg, impurity E 0.0012mg.
Get impurity test solution sample introduction record chromatogram.
The absorption maximum of impurity A is 283.5nm, the absorption maximum of impurity B is 277.5nm and 275.1nm, the absorption maximum of impurity C is 264.5nm, the absorption maximum of impurity D is 272.8nm, the absorption maximum of impurity E is 277.5nm, the absorption maximum of main peak is 279.9nm.
Conclusion: under the process conditions, goes out peak by main peak in 279.9nm, therefore adopts the wavelength detecting near 278 ~ 280nm.
Embodiment 3: blank auxiliary interference test.
Get blank auxiliary solution sample introduction record chromatogram, see Fig. 2.
The peak measured without impact in chromatogram occurs.
Conclusion: show that auxiliary material is noiseless to determination of foreign matter.
Embodiment 4: system suitability is tested.
Instrument: LC-10ATvp/SPD-10Avp N2000 workstation;
Chromatographic column: 4.6 × 150mm 5 μ octadecyl silane chromatographic column;
Flow velocity: 1.0ml/min;
Sample size: 20 μ l;
Column temperature: 35 DEG C;
Wavelength: 278nm.
Get same part sample solution, continuous sample introduction 5 times, record chromatogram is tried to achieve relative standard deviation and should be not more than 2.0%, the results are shown in Table 1.
Conclusion: test shows that true qualities spectra system precision is good.
Embodiment 5: linear and scope.
Getting concentration is the cefuroxime ester solution being respectively 60.795 μ g/ml, 121.590 μ g/ml, 182.385 μ g/ml, 243.180 μ g/ml, 303.975 μ g/ml, 340.452 μ g/ml, each 20 μ l, injection liquid chromatography respectively, record chromatogram, take peak area as ordinate, concentration is that horizontal ordinate carries out linear regression, the results are shown in Table 2, Fig. 3.
It is C=2E-05S+10.506 r=0.9994 that the regression equation of CEFUROXIME AXETIL obtains regression equation.
Result shows, cefuroxime ester concentration respectively within the scope of 60.795 ~ 34.452 μ g/ml sample introduction concentration and peak area value have good linear relationship.
Embodiment 6: detectability.
Get certain density cefuroxime ester solution, stepwise dilution obtains the solution of series concentration, gets series concentration solution each 20ul injection liquid chromatography respectively, and record chromatogram, observe signal to noise ratio (S/N ratio), when signal to noise ratio (S/N ratio) is 3:1, detectable concentration is 62.5ng/ml.
Conclusion: result shows, minimum detectable activity is 62.5ng/ml.
Embodiment 7: replica test.
Get 6 parts, the sample of same lot number, accurately weighed respectively, measure impurity, the results are shown in Table 3.
Embodiment 8: recovery test.
Accuracy is the recovery gained that 80%, 100%, 120% 3 each impurity of variable concentrations by adding index in test sample records.The accuracy of known impurities is the impurity adding known quantity, then measures the ratio (recovery) between the measurement result of known impurities in loaded sample and theoretical value, represents, require that the recovery is between 70.0%-130.0% with percent %.
Recovery computing formula
Test shows that the accuracy of this method is good, the results are shown in Table 4.
Conclusion: better by the visible recovery of table.
Embodiment 9: stability of solution.
Get with a need testing solution, respectively get 20 μ l, injection liquid chromatography at 0,2,4,6,8,12,18,24 hour, record chromatogram, the results are shown in Table 5.
Conclusion: from table, its peak area of time lengthening placed along with solution reduces gradually, and related substance then increases gradually, show that solution impurity area in 24 hours has comparatively significant change, proof sample is unstable, therefore should be noted when measuring, and namely need testing solution should join and namely survey.
Embodiment 10: durability is tested.
In order to study the influence degree of subtle change to test findings of liquid phase chromatogram condition, carry out durability experiment.Major influence factors has buffer salinity, determined wavelength, mobile phase ratio, column temperature, flow velocity, chromatographic column.Get system testing liquid sample introduction respectively, investigate degree of separation, the results are shown in Table 6.
Experimental result shows, the method measures reliable, and durability is better.
Claims (4)
1. the method for its related substances in high-performance liquid chromatogram determination cefuroxime axetil tablets, it is characterized in that, it comprises the steps:
(1) preparation of impurity storing solution: get CEFUROXIME AXETIL, impurity A, impurity B, impurity C, impurity D, impurity E methyl alcohol dissolves respectively and be diluted to every 1mL respectively about containing the solution of CEFUROXIME AXETIL 0.250mg, impurity A 0.50mg, impurity B 0.37mg, impurity C 0.12mg, impurity D 0.12mg, impurity E 0.12mg;
(2) preparation of impurity test solution: get respectively and be settled to scale with mobile phase in above-mentioned each impurity storing solution 1ml to 100ml measuring bottle and shake up, to obtain final product;
(3) preparation of need testing solution: it is appropriate that precision takes cefuroxime axetil tablets fine powder, adds mobile phase after dissolving be settled to scale with methyl alcohol, makes every 1mL about containing the solution of CEFUROXIME AXETIL 0.250mg;
(4) preparation of reference substance solution: precision measures need testing solution 1ml, puts in 100ml measuring bottle, adds mobile phase and is diluted to scale, shake up;
(5) preparation of blank solution: methyl alcohol;
(6) measure: high performance liquid chromatograph take Stationary liquid as octadecyl silane chromatographic column, and mobile phase is 0.1 ~ 0.2mol/L ammonium dihydrogen phosphate: methyl alcohol=60:40 ~ 64:36;
Column temperature is 25 DEG C ~ 35 DEG C, and determined wavelength is that 278 ~ 280nm draws reference substance solution and each 15 ~ 25 μ l of need testing solution respectively, injects high performance liquid chromatograph, reads data;
(7) value of each dirt solution concentration and the equation of linear regression of respective peaks area value is calculated, related coefficient and 0.99 should be not less than, reference substance solution peak shape is symmetrical, theoretical cam curve more than 2000, in impurity test solution chromatogram, each magazins' layout degree should be not less than 1.5, blank solution chromatogram, occur without the peak identical with impurity test solution chromatogram, namely blank solution is noiseless;
(8) in every gram of cefuroxime axetil tablets, impurity A must not be greater than 2.0%, impurity B must not be greater than 1.5%, other single impurity must not be greater than 1.0%.
2. method according to claim 1, high performance liquid chromatograph take Stationary liquid as octadecyl silane chromatographic column, mobile phase is 0.2mol/L ammonium dihydrogen phosphate: methyl alcohol=62:38, column temperature is 30 DEG C, determined wavelength is that 278nm draws reference substance solution and each 20 μ l of need testing solution respectively, inject high performance liquid chromatograph, read data.
3. method according to claim 1, high performance liquid chromatograph take Stationary liquid as octadecyl silane chromatographic column, mobile phase is 0.1mol/L ammonium dihydrogen phosphate: methyl alcohol=60:40, column temperature is 25 DEG C, determined wavelength is that 278nm draws reference substance solution and each 15 μ l of need testing solution respectively, inject high performance liquid chromatograph, read data.
4. method according to claim 1, high performance liquid chromatograph take Stationary liquid as octadecyl silane chromatographic column, mobile phase 0.2mol/L ammonium dihydrogen phosphate: methyl alcohol=64:36, column temperature is 35 DEG C, determined wavelength is that 280nm draws reference substance solution and each 25 μ l of need testing solution respectively, inject high performance liquid chromatograph, read data.
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| CN110146621A (en) * | 2019-06-13 | 2019-08-20 | 国药集团致君（深圳）制药有限公司 | The content assaying method of polymer in cephalosporin analog antibiotic drug |
| CN111948318A (en) * | 2020-08-16 | 2020-11-17 | 江苏正大清江制药有限公司 | Method for determining high-molecular polymer in cefuroxime axetil tablets |
| CN114354800A (en) * | 2021-12-31 | 2022-04-15 | 山东大学 | Method for analyzing acetyl bromide content in cefuroxime axetil |
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| CN111948318B (en) * | 2020-08-16 | 2022-04-01 | 江苏正大清江制药有限公司 | Method for determining high-molecular polymer in cefuroxime axetil tablets |
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Application publication date: 20150930 |