CN101592637A - A kind of detection method of new compound CTX sodium-tazobactam sodium - Google Patents
A kind of detection method of new compound CTX sodium-tazobactam sodium Download PDFInfo
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Abstract
A kind of new high performance liquid chromatography (HPLC) method, can detect the content and the relative substance of two kinds of single components in the CTX sodium-tazobactam sodium compound simultaneously, two kinds of composition non-interference and influence, operation is simple for this method, specificity is strong, and is highly sensitive, and the range of linearity is big, good stability can be used for the detection of compound preparation and raw material.
Description
Background of invention:
Cefotaxime Sodium (cefotaxime sodium, CTX) be first third-generation cephalosporin of German Hoechst AG in 1977 development, be used for the treatment of the infectious diseases due to multiple gram-positive bacteria and the negative bacterium, has has a broad antifungal spectrum, antibacterial activity is strong, Tissue distribution is few, characteristics such as low toxicity.Nearly 30 years of this launch, use a plurality of countries and regions in the whole world, is subjected to extensive favorable comment, is still a clinical line medication so far.
Because a large amount of and clinical practice widely, even the appearance of abuse in recent years, produced the drug-fast bacteria of anti-CTX clinically.These class drug-fast bacteria great majority work by producing beta-lactam enzymatic degradation CTX, have reduced the antibacterial activity of Cefotaxime Sodium like this.Can effectively suppress drug-resistance of bacteria after share at the beta-lactamase inhibitor of this situation exploitation and microbiotic, recover antibiotic activity, therefore, adopt the compound of " microbiotic+beta-lactamase inhibitor " to make up the resistance problem that can suppress zymogenic bacteria effectively.Compound preparation by this thinking exploitation is successful clinically as " amoxicillin and clavulanate ", " cefoperazone sodium sulbactam sodium ", " piperacillin-sulbactam sodium " etc.From the angle of preparation, because Cefotaxime Sodium contains certain moisture, and sodium-tazobactam is met the water degraded, unstable, if simply that both are admixed together, then can't solve the stability problem of product, must on preparation process, solve the stability problem of product.Simultaneously owing to there is not compound preparation before, all be that single component is done detection, both need set up new detection method at mixed compound check, such detection method not only requirement can detect the content and the relative substance of single component in the compound, simultaneously, this detection method can't be subjected to the influence of another composition to the mensuration of single component, and can reach exclusive sensitivity, requirements such as easy operation, and can check product quality in preparation factory and various places testing agency or hospital, require us to set up the method for a new detection compound CTX sodium-tazobactam sodium at such.
Summary of the invention:
We adopt in common high performance liquid chromatographies such as factory, institute for drug control, hospital (HPLC) by optimizing indexs such as moving phase, flow velocity, detection wavelength, set up a kind of new detection method, this method can detect the content and the related substance of two kinds of single components in the compound CTX sodium-tazobactam sodium, do not influence each other and disturb and two kinds of compositions can not occur, method is simple, operation steps is few for this, the method specificity is strong, highly sensitive, the range of linearity is big, good reproducibility etc., can be used for the conventional sense of the preparation and the raw material of compound.
The patent example:
The Preliminary detection of embodiment one, compound raw material and preparation
" cefotaxime sodium for injection sodium-tazobactam " adopted in experiment, and ratio is 6: 1, and specification is 1.0g, and simple loading amount on preparation changes because of other proportionings and specification, and its raw material sources, preparation technology all do not have significant difference.So study with this proportioning, specification sample.
Test specimen is white, off-white color or yellowish white powder, carries out following various test.
1, discrimination test:
Two kinds of component Tazobactam Sodiums and CTX all exist with sodium-salt form, should present the flame reaction of sodium salt during calcination.Get platinum filament, after usefulness hydrochloric acid is moistening, dips in and get this product, burn in colourless flame, flame promptly shows foresythia.
Two kinds of component sodium-tazobactams are different with Cefotaxime Sodium polarity, can separate by the HPLC method, and contrast discriminating with reference substance.Get the standard items of this product and Tazobactam Sodium and CTX, make the solution that every 1ml contains 0.48mg Cefotaxime Sodium and 0.08mg sodium-tazobactam with moving phase respectively, according to high performance liquid chromatography test under this patent assay item described below, the test sample peak is consistent with the reference substance peak as a result.(test method is seen embodiment two, three)
2, detect test
Test specimen acidity is checked: get test specimen and add the injection water and make the solution that contains CTX 0.1g among every 1ml, measure with acidometer, the pH of test specimen is specification product between 4.5-6.5 as a result.
Test sample solution clarity is checked: get 5 bottles of test specimens, add water respectively and make the solution that contains this product 0.1g among every 1ml approximately, compare with No. 1 turbidity standard, check clarity.Test specimen is a clear liquid as a result, does not surpass No. 1 turbidity standard, and clarity meets relevant regulations, is qualified samples.
The test sample solution colour of solution: get 5 bottles of test specimens, be dissolved in water, place the nessler colorimetric tube of 25ml, thin up compares with the yellow tone standard color solution to 10ml.Test specimen is a clear liquid as a result, and solution colour does not surpass yellow or No. 9 standard color solutions of yellow green, meets relevant regulations, is qualified samples.
Test specimen moisture is checked, gets test specimen 1g, measures by the water content detection method, and the test specimen moisture all in 3% left and right sides conformance with standard, is qualified samples as a result.
Because said preparation is by aseptic raw material direct packaging, so, just the same on the composition of raw material and preparation, the content, there is not difference.The detection method that adopts this patent obtains the result identical with preparation to the detection that raw material carries out.
The analysis and the method for quality control of embodiment two, compound raw material and preparation
For the quality of control product, must analyze and research to the impurity that might occur in the product.Before owing to the experience that two materials is not mixed use, so existing data also has only the method for inspection to relative substance in the single Cefotaxime Sodium (or sodium-tazobactam).
Though such method of inspection is to the one matter check effectively, but the relative substance in whether can test mixture is not clear, because be prepared into after the mix preparation whether can influence each other the other side's the detection of two kinds of principal ingredients, the detection whether two kinds of impurity also can influence the other side has a lot of uncertain factors, need carry out a large amount of tests and verify.So we have set up corresponding detection method to impurity in the compound material according to Cefotaxime Sodium, sodium-tazobactam one matter detection method.
1, the relative substance material detects:
Because the HPLC method can perform well in two kinds of single composition detection, so we adopt the HPLC method to set up the method for detecting impurities of compound.
Destructive test is at first carried out in test, carried out 1 respectively according to the medicine stability test guide) alkali destruction: it is an amount of to get Cefotaxime Sodium, sodium-tazobactam and cefotaxime sodium for injection sodium-tazobactam respectively, in the 10ml measuring bottle, dissolve with 0.1mol/l sodium hydroxide solution 1ml, solution colour becomes little yellow, with the neutralization of 0.1mol/l hydrochloric acid, is diluted with water to scale, shake up, as need testing solution.2) adopt acid to destroy, then: it is an amount of to get Cefotaxime Sodium, sodium-tazobactam and cefotaxime sodium for injection sodium-tazobactam respectively, in the 10ml measuring bottle, with the 0.1mol/l dissolve with hydrochloric acid solution and be diluted to scale, shakes up, as need testing solution.3) adopt oxidative degradation, again: it is an amount of to get Cefotaxime Sodium, sodium-tazobactam and cefotaxime sodium for injection sodium-tazobactam respectively, and in the 10ml measuring bottle, the water dissolving adds 2 in hydrogen peroxide, is diluted with water to scale, shakes up, as need testing solution.
The above-mentioned three kinds of need testing solutions of learning from else's experience, under following chromatographic condition, test, whether detection sodium-tazobactam, Cefotaxime Sodium have obvious degradation, with the catabolite peak separate with main peak＞2.0, Cefotaxime Sodium catabolite and sodium-tazobactam main peak degree of separation＞1.5 are the judge index of criterion of acceptability.
We at first adopt following chromatographic condition to screen best detection method:
Chromatographic condition (1): Waters 515 pumps, Waters2487 UV-detector are adopted in test; Chromatographic column: UltimateXB-NH2 positive analytical column; Specification: 5 μ m, 4.6*50mm; Moving phase: normal hexane-chloroformic solution (2: 1); Detect wavelength: 220 nm-260nm; Flow velocity: 0.1～3ml/min; Sample size: 10～30 μ l.
Chromatographic condition (2): Waters 515 pumps, Waters2487 UV-detector are adopted in test; Chromatographic column: octadecylsilane chemically bonded silica post, Kromasil C
185 μ m 4.6mm * 20cm; Moving phase: acetonitrile-potassium dihydrogen phosphate (0.015mol/L)-tetrabutylammonium hydroxide sodium solution (10%) (180-200: 785-795: 35-5) (is 4.0 with the phosphoric acid adjust pH); Detect wavelength: 220nm-260nm; Flow velocity: 0.1～3ml/min; Sample size: 10～30 μ l.
Chromatographic condition (3): Waters 515 pumps, Waters2487 UV-detector are adopted in test; Chromatographic column: octadecylsilane chemically bonded silica post, Kromasil C
185 μ m 4.6mm * 20cm; Moving phase: acetonitrile-potassium dihydrogen phosphate (0.015mol/L)-methyl alcohol (10%) (180-200: 785-795: 35-5) (is 4.0 with the phosphoric acid adjust pH); Detect wavelength: 220nm-260nm; Flow velocity: 0.1～3ml/min; Sample size: 10～30 μ l.
Chromatographic condition (4): Waters 515 pumps, Waters2487 UV-detector are adopted in test; Chromatographic column: octadecylsilane chemically bonded silica post, Kromasil C
185 μ m 4.6mm * 20cm; Moving phase: acetonitrile-potassium dihydrogen phosphate (0.015mol/L)-ethanol (20%) (180-200: 785-795: 35-5) (is 4.0 with the phosphoric acid adjust pH); Detect wavelength: 220nm-260nm; Flow velocity: 0.1～3ml/min; Sample size: 10～30 μ l.
Adopt said method to detect and find, it is all inseparable less than 1.0, two components that chromatographic condition (1), (3), (4) though also can be used to detects both degree of separation, and the degree of separation of chromatographic condition (2) is better, but also need optimize.Sample sodium-tazobactam, Cefotaxime Sodium under chromatographic condition (2) test condition through three kinds of destructive tests all do not have obvious degradation, Cefotaxime Sodium impurity peaks and sodium-tazobactam main peak degree of separation＞1.5.
Which whether methodology inadequately sensitivity do not detect impurity? do not still itself impurity just occur? to this, we at first adopt LC-MS that sample is checked, find sample through above-mentioned three kinds of failure tests, also not finding under the LC-MC testing conditions has impurity to occur, and illustrates that this method of inspection is reliable.
Further, whether we adopt extreme means that above-mentioned failure condition is strengthened 2-10 doubly, at first detect to guarantee occurring impurity with LC-MS, observe this test condition and can detect.Found that, after adopting stronger failure condition, Cefotaxime Sodium and sodium-tazobactam all degraded occurring in varying degrees, such degraded just occurs under the condition of the acid destruction of 3 times of reinforcements, impurity level is very low, can detect the appearance of impurity with LC-MS, stronger failure condition can produce more impurity.We adopt the detection method of above-mentioned foundation, impurity analysis just occurred under the condition of destroying for the acid of 3 times of reinforcements, and this method can be found the appearance of impurity as a result.
We, optimize the HPLC detection method that we set up as detected object with " solution that has just occurred impurity under the condition that the acid of 3 times of reinforcements destroys ", have determined that at last following method is as best authentication method:
Chromatographic condition: Waters 515 pumps, Waters2487 UV-detector are adopted in test; Chromatographic column: octadecylsilane chemically bonded silica post, Kromasil C
185 μ m 4.6mm * 20cm; Moving phase: acetonitrile-potassium dihydrogen phosphate (0.03mol/L)-tetrabutylammonium hydroxide sodium solution (10%) (190: 795: 15) (is 4.0 with the phosphoric acid adjust pH); Detect wavelength: 230nm; Flow velocity: 1ml/min; Sample size: 20 μ l.
We detect check sample with the above-mentioned method of inspection, and the result is as follows:
Sample solution preparation: it is an amount of to get this product, accurately claims surely, makes the solution that contains Cefotaxime Sodium 0.48mg and sodium-tazobactam 0.08mg among every 1ml with the moving phase dissolving, as need testing solution.Precision is measured 0.33ml to 10ml measuring bottle, is diluted to scale with moving phase, in contrast solution.Precision is measured need testing solution 1ml, puts in the 100ml measuring bottle, adds water to scale, shakes up, in contrast solution.Get contrast solution 20 μ l and inject liquid chromatograph, regulate detection sensitivity, making the major component peak height is 20～25% of full scale, gets need testing solution 20 μ l again and injects liquid chromatograph, and the record chromatogram is to 2 times of major component peak retention time.
Experimental examination three batches of production sample related substance check results all up to specification.The results are shown in Table 1 and Fig. 1 ~ 3
Table 1 related substance check result
2, CTX sodium polymer
Because Cefotaxime Sodium also has polymkeric substance to produce, if the polymer content height can cause the generation of bad reaction, need control the content of polymkeric substance in product, must detect polymkeric substance according to related request.In the past, only in the folk prescription Cefotaxime Sodium, carry out the detection of polymkeric substance, also do not mix the research report of carrying out polymer determination in the back about compound CTX sodium-tazobactam, the situation of the condition measured of folk prescription in can the accurate response compound whether, in other words, whether the compound potpourri can influence the accuracy of the detection of original condition, and to this, we also study.
We have at first carried out the test of CTX polymkeric substance chromatographic condition and assay method.
Chromatographic condition and system suitability test: with sephadex G-10 (40 ~ 120 μ m) is filling agent; Glass column internal diameter 1.3～1.5cm, bed volume 50～60ml; Mobile phase A is phosphate buffer (get sodium hydrogen phosphate 21.85g and sodium dihydrogen phosphate 5.38g, add water 1000ml and make dissolving, mixing is regulated pH value to 7.0), and Mobile phase B is 0.01% sodium dodecyl sulfate solution; Flow velocity is per minute 1ml; The detection wavelength is 254nm.With the mobile phase A is moving phase, 200 μ l measure with 1mg/ml blue dextran 2000 solution sample introductions, number of theoretical plate calculates with blue dextran 2000 peaks should be not less than 900, tailing factor should be between 0.75～1.5, with the Mobile phase B is moving phase, get reference substance solution, repeat sample introduction 200 μ l, the relative standard deviation of peak area value should be less than 5%.
The sodium-tazobactam interference test: it is an amount of to get the Tazobactam Sodium sodium raw materials, and accurate the title decides, and adds the mobile phase A dissolving and makes the solution that every 1ml contains Tazobactam Sodium 20mg, measures according to said method.Chromatogram is seen accompanying drawing 7.The result shows that sodium-tazobactam is noiseless to this product CTX polymer determination.
Detection limit: get the CTX contrast solution, be diluted to variable concentrations successively, detect with above-mentioned chromatographic condition.Detected level is 41ng, and detection limit is about 0.001% of CTX amount.
The preparation of reference substance solution: it is an amount of to get the CTX reference substance, and accurate the title decides, and adds water and makes the solution that contains CTX 100 μ g among every 1ml, shakes up.
Determination method: sample thief, the accurate title, decide, and adds water and make the solution that contains 20mg among every 1ml, shakes up, and sample introduction 200 μ l are that moving phase is measured with the mobile phase A immediately, the record chromatogram; Other gets reference substance solution 200 μ l and injects liquid chromatograph, is moving phase with the Mobile phase B, and the record chromatogram calculates by external standard method.
The result: three batch sample CTX polymer determination chromatograms are seen accompanying drawing 4-79-12, the results are shown in Table 5.
Table 2 CTX sodium polymer check result
| Lot number | wm0401 | wm0402 | wm0403 |
| Polymkeric substance % | 0.035 | 0.040 | 0.041 |
According to the quality standard requirement of test result and cefotaxime sodium for injection, draft this product CTX polymkeric substance in the quality standard and must not cross 1.0% in CTX.
In order further to verify such situation, we have adopted the LC-MS checking for said method, found that, adopt the result of said method mensuration and the result of LC-MS method mensuration to can be good at correspondence, though it is potpourri that compound medicine is described, such potpourri does not influence the specificity and the sensitivity of original mensuration polymkeric substance method.
Embodiment three, the foundation of new detection method that is used for this compound preparation and formulation content are measured checking
Be the quality of control product, it is an important indicator that the content of two principal ingredients in the compound preparation is controlled, and has had many research methods to set up for single Cefotaxime Sodium, two kinds of content detection of sodium-tazobactam.But, after being prepared into potpourri, such method still can be used in the detection of composition in the potpourri? in other words, two materials detection of the other side that can influence each other in the potpourri? still nobody studies such problem, wherein also have a lot of uncertain factor demand sides to and explain.To this, we carry out detection method for content how to carry out two kinds of compositions in the compound preparation and study.
Because the HPLC method can perform well in two kinds of single composition detection, therefore, we adopt the HPLC method to set up the detection method of two component contents of compound.
Chromatographic condition is selected for use: Waters 515 pumps, Waters2487 UV-detector are adopted in test; Chromatographic column: octadecylsilane chemically bonded silica post, Kromasil C
185 μ m 4.6mm * 20cm; Moving phase: acetonitrile-potassium dihydrogen phosphate (0.03mol/L)-tetrabutylammonium hydroxide sodium solution (10%) (190: 795: 15) (is 4.0 with the phosphoric acid adjust pH); Detect wavelength: 230nm; Flow velocity: 1ml/min; Sample size: 20 μ l.
Wavelength is selected: get CTX, the Tazobactam Sodium reference substance is an amount of, add the moving phase dissolving respectively, scan in 200～300nm wavelength coverage, the result shows: CTX has absorption maximum at 235nm wavelength place, all has more by force to absorb in 220～260nm wavelength coverage; Tazobactam Sodium solution does not have tangible maximum absorption band, in the following scope of 230nm wavelength absorption is arranged; CTX and Tazobactam Sodium have at 230nm wavelength place suitable absorption are all arranged, proportion is less in this product to consider Tazobactam Sodium, and selected wavelength also is 230nm in the assay HPLC method of Tazobactam Sodium sodium raw materials, so this law selects 230nm as detecting wavelength.
The selection of moving phase: this product is a compound preparation, the appearance time of two main peaks is mainly considered in the selection of moving phase, the separation case of separating effect and main peak and impurity peaks, when selecting acetonitrile-potassium dihydrogen phosphate (0.015mol/L)-methyl alcohol (10%) for use, the degree of separation of two components is too little, and the retention time of Cefotaxime Sodium is long, is unsuitable for measuring.Suitably regulating moving phase forms.For avoiding the CTX retention time long, adopt the moving phase of acetonitrile-potassium dihydrogen phosphate (0.03mol/L)-TBAH solution (10%) (190: 795: 15) (is 4.0 with the phosphoric acid adjust pH) as this product, this moving phase can make two components in the sample effectively detect, degree of separation can obtain preferable separating effect all greater than 1.5.
On the basis of above-mentioned research, we adopt selected chromatographic condition at first to carry out the system suitability test, and under above-mentioned chromatographic condition, the about 3min of solvent peak retention time does not disturb major component to measure.Take by weighing CTX sodium raw materials and Tazobactam Sodium sodium raw materials respectively, the sample solution for preparing one-component and blending ingredients according to content assaying method is measured according to above-mentioned chromatographic condition as need testing solution.
We further study the range of linearity of detection method:
The CTX linear relationship: it is an amount of to get the CTX reference substance, and accurate the title decides, and dilute with water is made every 1ml and contained the solution that CTX is 265.1,353.5,441.9,530.2,618.6 μ g.Get 20 μ l respectively, inject liquid chromatograph, record chromatogram 8-12 the results are shown in Table 7, CTX concentration-peak area linear relationship chart See Figure.
The relation of table 7 CTX concentration and peak area (seeing accompanying drawing 29)
Conclusion: under this chromatographic condition, in CTX concentration 265～619 μ g/ml scopes, its peak area A and concentration C are the good linear relation.
The Tazobactam Sodium linear relationship is investigated: it is an amount of to get the Tazobactam Sodium reference substance, and accurate the title decides, and is diluted with water to every 1ml and contains the solution that Tazobactam Sodium is 40.27,53.69,67.11,80.54,93.96 μ g.Get 20 μ l respectively, injecting chromatograph, record chromatogram (seeing accompanying drawing 13-17) the results are shown in Table 8, Tazobactam Sodium na concn-peak area linear relationship chart See Figure.
The relation of table 8 Tazobactam Sodium concentration and peak area (seeing accompanying drawing 30)
The result shows, under this chromatographic condition, the Tazobactam Sodium na concn is between 40 μ g/ml ~ 94 μ g/ml the time, and its peak area A and concentration C are the good linear relation.
Further again, we also carry out replica test: take by weighing Cefotaxime Sodium and sodium-tazobactam in the prescription ratio, with moving phase dissolving and be diluted to scale, repeat sample introduction five times by above-mentioned chromatographic condition, write down chromatogram, calculate the relative standard deviation of peak area respectively.The results are shown in Table 11.
Table 11 replica test result
| |
1 | 2 | 3 | 4 | 5 | Average peak area | RSD％ |
| CTX | 20631.646 | 20625.404 | 20575.443 | 20566.861 | 20571.779 | 20594.23 | 0.15 |
| Tazobactam Sodium | 409.157 | 407.785 | 407.087 | 406.050 | 406.190 | 407.26 | 0.31 |
We have carried out recovery test again: take by weighing each 6 parts of Cefotaxime Sodium and sodium-tazobactams by recipe quantity, put respectively in the 50ml measuring bottle, add the moving phase dissolving and be diluted to scale, get the solution sample introduction-, other gets the reference substance solution sample introduction, uses regression equation calculation content, and calculate recovery rate.The results are shown in Table 9 and accompanying drawing 18-22.
Table 9 recovery test result
As seen from table: the recovery of two compositions is all better in this product.
Show that from top test findings this content assaying method that we set up has sensitivity, good stability, repeatability is high.It is the method that two compositions of compound medicine are checked in desirable being used to.
Because said preparation is by aseptic raw material direct packaging, so, just the same on the composition of raw material and preparation, the content, there is not difference.The detection method that adopts this patent obtains the result identical with preparation to the detection that raw material carries out.
Embodiment four, the new detection method of employing detect compound CTX sodium-tazobactam sodium sample
Further, we adopt this new method of inspection that the test specimen content of producing under three batches of GMP conditions is measured.Carry out according to aforementioned content assaying method, it is an amount of to get this product, accurately claims surely, adds the moving phase dissolving and is diluted to the solution that every ml contains Cefotaxime Sodium 0.48mg and contains sodium-tazobactam 0.06mg, and precision is measured 20 μ l, injecting chromatograph; Other gets Cefotaxime Sodium and the sodium-tazobactam reference substance is an amount of, with method dilution and mensuration, by the content of sodium-tazobactam and Cefotaxime Sodium in the external standard method calculating test sample.
Illustrate: W is right: the sample weighting amount of reference substance, and the W sample: the sample weighting amount of sample, W dress: the average loading amount of sample, the A mark: the peak area of reference substance, the A sample: the peak area of sample, to %: the purity of reference substance, labelled amount: the labelled amount of sample
Three batch sample assays the results are shown in Table 8, and chromatogram is seen Figure 23-28.
Table 8 three batch sample assay results
Because said preparation is by aseptic raw material direct packaging, so, just the same on the composition of raw material and preparation, the content, there is not difference.The detection method that adopts this patent obtains the result identical with preparation to the detection that raw material carries out.
Embodiment five, new detection method are to the compound raw material of different proportionings and the detection of preparation
The detection method that adopts above-mentioned example to set up, we detect the compound preparation of the CTX sodium-tazobactam sodium of different proportionings, to observe the detection whether such method can be used for the compound preparation of different proportionings, found that, for CTX sodium/tazobactam sodium 20: 1,10: 1,8: 1,6: 1,5: 1,4: 1,3: 1,2: 1,1: 1,1: 2,1: 3,1: 4,1: 5,1: 6,1: 8,1: 10, the detection of the sample of 17 different proportionings such as 1: 20 grade, the new method that this patent is set up all can well be measured two constituents, and this method has good sensitivity and reliability.The be determined at influence that 20: 1～1: 20 scope in be not subjected to proportioning of this method to the CTX sodium-tazobactam is described.
Because said preparation is by aseptic raw material direct packaging, so, just the same on the composition of raw material and preparation, the content, there is not difference.The detection method that adopts this patent obtains the result identical with preparation to the detection that raw material carries out.Embodiment six, adopt this kind method of inspection to be judged as the zooperal result that carries out of qualified preparation
Carry out the preparation of three batch samples according to the GMP standard, method according to this patent example one, two, three is tested to three batch samples, adopt the qualified formulation samples of above-mentioned standard test to carry out animal experiment, check whether the sample that is up to the standards by above-mentioned new detection method can meet zooperal requirement.
Test specimen at first carries out sterility test.In desinfection chamber, carry out sterile working, get each 6 of three batches of test specimens, add 100ml 0.9% aseptic sodium chloride solution respectively and make dissolving, shake up, open the filter lid, make bottleneck pass through flame, pour into immediately in the filter at the other stopper of opening the sample test liquid of flame, and close filter lid, open the valve of discharge opeing frame, start vacuum pump and carry out decompress filter, after waiting to do, wash filter membrane 3 times with sterile saline according to aforesaid operations, each 100ml.Open bottle gas outlet of bleeding, filtrator is taken off, with the sterilization tweezers filter membrane is taken out and put into the two dish of sterilization.With the sterilization scissors filter membrane is divided into 3 parts, places the container that respectively contains 50ml THIOGLYCOLLIC ACID salt fluid nutrient culture media and improvement Martin nutrient culture media respectively, a copy of it is done positive control.Temperature was in accordance with regulations cultivated 14 days.Day by day observe between culture period and write down whether bacteria growing is arranged.The result does not have bacterial growth, and sterility test is up to specification, is specification product.
Test specimen carries out the undue toxicity inspection: get three batches of test specimens, add the chlorination sodium injection and make the solution that contains 0.15g among every 1ml, get 15 mouse and be divided into three groups, the solution 0.5ml of every group of 5 same lot numbers of tail vein injection, the interior no abnormal reaction of observing 48 hours does not have death yet, and the result is specification product.
Test specimen carries out pyrogen test: get three batches of test specimens, add sterilized water for injection and make the solution that contains 0.15g among every 1ml, get 9 rabbit and be divided into 3 groups, 3 every group, measure after its normal body temperature in 15 minutes, dosage is exempted from the every 1kg injection of body weight 1ml by family, slowly inject prescribed dose and be warmed to 38 ℃ need testing solution from ear vein, measured its body temperature 1 time every 30 minutes then, survey altogether 6 times, with the highest normal body temperature that once deducts in 6 body temperature, be the rising temperature of this rabbit body temperature.Each rabbit fervescence all is lower than 0.6 ℃, and every group of rabbit body temperature rising summation be lower than 1.4 ℃, and it is up to specification that therefore three batch samples can be judged as heat source check, and three batch samples are specification product.
The results are shown in Table 9
Table 9: pyrogen test result
Description of drawings:
Fig. 1 is that to criticize related substance controlling chart spectrogram 2 be that wm0402 criticizes related substance and checks collection of illustrative plates to wm0401
Fig. 3 is that to criticize related substance controlling chart spectrogram 4 are polymkeric substance contrast collection of illustrative plates to wm0403
Fig. 5 is that wm0401 batch sample polymer determination figure Fig. 6 is wm0402 batch sample polymer determination figure
Fig. 7 is that wm0401 batch sample polymer determination figure Fig. 8 is a Cefotaxime Sodium linear test collection of illustrative plates 1
Fig. 9 is that Cefotaxime Sodium linear test collection of illustrative plates 2 Figure 10 are Cefotaxime Sodium linear test collection of illustrative plates 3
Figure 11 is that Cefotaxime Sodium linear test collection of illustrative plates 4 Figure 12 are Cefotaxime Sodium linear test collection of illustrative plates 5
Figure 13 is that linear collection of illustrative plates 1 Figure 14 of Tazobactam Sodium is the linear collection of illustrative plates 2 of Tazobactam Sodium
Figure 15 is that linear collection of illustrative plates 3 Figure 16 of Tazobactam Sodium are the linear collection of illustrative plates 4 of Tazobactam Sodium
Figure 17 is that linear collection of illustrative plates 5 Figure 18 of Tazobactam Sodium are recovery test collection of illustrative plates 1
Figure 19 is that recovery test collection of illustrative plates 2 Figure 20 are recovery test collection of illustrative plates 3
Figure 21 is that recovery test collection of illustrative plates 4 Figure 22 are recovery test collection of illustrative plates 5
Figure 23 is that to criticize assay collection of illustrative plates 1 Figure 24 be that wm0401 criticizes assay collection of illustrative plates 2 to wm0401
Figure 25 is that to criticize assay collection of illustrative plates 1 Figure 26 be that wm0402 criticizes assay collection of illustrative plates 2 to wm0402
Figure 27 is that to have criticized assay collection of illustrative plates 1 Figure 28 be that wm0403 has criticized assay collection of illustrative plates 2 to wm0403
Figure 29 is that graph of a relation Figure 30 of CTX na concn and peak area is the relation of Tazobactam Sodium concentration and peak area.
Claims (8)
1, a kind of new detection method that is used for Cefotaxime Sodium and sodium-tazobactam compound raw material and preparation.
2, method according to claim 1, it is characterized in that this method adopts high performance liquid chromatography, the moving phase that is adopted can be normal hexane, chloroform, methylene chloride, tetrabutylammonium hydroxide amine, acetonitrile, potassium dihydrogen phosphate, methyl alcohol, water etc. wherein preferably adopt tetrabutylammonium hydroxide amine, potassium dihydrogen phosphate and acetonitrile.
3, according to claim 1,2 described methods, it is characterized in that the detection wavelength that this method adopts can be 210 ~ 260nm, preferred wavelength adopts 230nm.
4, according to claim 1,2,3 described methods, it is characterized in that the flow rate of mobile phase that this method adopts can be 0.1 ~ 3.0ml/min, preferably flow velocity adopts 1.0ml/min;
5,, it is characterized in that the chromatographic column that this method adopts can be the amido post, the cyano group post according to claim 1,2,3,4 described methods, the octadecylsilane chemically bonded silica post, eight alkyl silane bonded silica gel posts, tetraalkyl silane group silicagel column, phenyl silane bonded silica gel post etc.; Preferably adopting octadecylsilane chemically bonded silica is the chromatographic column (C18 post) of filling agent.
6, according to claim 1,2,3,4,5 described methods, it is characterized in that this method all can detect for the CTX sodium-tazobactam sodium compound of different proportion, both components in proportions of CTX sodium-tazobactam sodium can comprise 1: 50 ~ 50: 1, are preferably used for 1: 20 ~ 20: 1.
7, according to claim 1,2,3,4,5,6 described methods, it is characterized in that this method can be used for the raw material detection of CTX sodium-tazobactam sodium compound, the preparation that also can be used for this compound detects.
8, according to claim 1,2,3,4,5,6,7 described methods, the method of this detection CTX sodium-tazobactam sodium compound, can detect well two components contents and impurity, and such check is not subjected to the interference of another composition, good sensitivity, specificity, simple and easy to do are arranged, this method can be used for drugmaker in the raw material of this compound and preparation production run for the detection of product quality, also can be used for of the monitoring of units such as medicine inspecting institute, hospital to the quality of the pharmaceutical preparations.
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| CN101822682A (en) * | 2010-05-04 | 2010-09-08 | 南京大海药物研究有限公司 | Injectable drug combining cefotaxime and tazobactam with proportion of 6:1 |
| CN101912403A (en) * | 2010-08-02 | 2010-12-15 | 陶灵刚 | Cefoperazone sodium and tazobactam sodium medicinal composition microsphere injection |
| CN108802206A (en) * | 2017-05-05 | 2018-11-13 | 江苏灵豹药业股份有限公司 | Method that is a kind of while measuring main component and the content of major impurity in cefotaxime sodium and tazobactam sodium for injection |
| CN108802241A (en) * | 2017-05-05 | 2018-11-13 | 江苏灵豹药业股份有限公司 | Liquid chromatography-tandem mass spectrometry method for qualitative analysis in relation to substance in a kind of cefotaxime sodium and tazobactam sodium for injection |
| RU2687493C1 (en) * | 2018-10-08 | 2019-05-14 | Федеральное казённое учреждение здравоохранения "Ставропольский научно-исследовательский противочумный институт" Федеральной службы по надзору в сфере защиты прав потребителей и благополучия человека | Method for determining cefotaxime by reversed-phase high-performance liquid chromatography |
| CN116858975A (en) * | 2022-03-28 | 2023-10-10 | 南京优科生物医药股份有限公司 | Method for detecting impurities of cefotaxime sodium tazobactam sodium polymer |
| CN116930390A (en) * | 2022-04-02 | 2023-10-24 | 南京优科生物医药股份有限公司 | Two-dimensional liquid chromatography-tandem mass spectrometry analysis method for detecting related substances in cefotaxime sodium tazobactam sodium |
| CN118243828A (en) * | 2024-05-29 | 2024-06-25 | 南京优科制药有限公司 | Detection method of tazobactam impurity A in cefotaxime sodium tazobactam sodium for injection |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101822682A (en) * | 2010-05-04 | 2010-09-08 | 南京大海药物研究有限公司 | Injectable drug combining cefotaxime and tazobactam with proportion of 6:1 |
| CN101912403A (en) * | 2010-08-02 | 2010-12-15 | 陶灵刚 | Cefoperazone sodium and tazobactam sodium medicinal composition microsphere injection |
| CN101912403B (en) * | 2010-08-02 | 2012-01-11 | 陶灵刚 | Cefoperazone sodium and tazobactam sodium medicinal composition microsphere injection |
| CN108802206A (en) * | 2017-05-05 | 2018-11-13 | 江苏灵豹药业股份有限公司 | Method that is a kind of while measuring main component and the content of major impurity in cefotaxime sodium and tazobactam sodium for injection |
| CN108802241A (en) * | 2017-05-05 | 2018-11-13 | 江苏灵豹药业股份有限公司 | Liquid chromatography-tandem mass spectrometry method for qualitative analysis in relation to substance in a kind of cefotaxime sodium and tazobactam sodium for injection |
| RU2687493C1 (en) * | 2018-10-08 | 2019-05-14 | Федеральное казённое учреждение здравоохранения "Ставропольский научно-исследовательский противочумный институт" Федеральной службы по надзору в сфере защиты прав потребителей и благополучия человека | Method for determining cefotaxime by reversed-phase high-performance liquid chromatography |
| CN116858975A (en) * | 2022-03-28 | 2023-10-10 | 南京优科生物医药股份有限公司 | Method for detecting impurities of cefotaxime sodium tazobactam sodium polymer |
| CN116930390A (en) * | 2022-04-02 | 2023-10-24 | 南京优科生物医药股份有限公司 | Two-dimensional liquid chromatography-tandem mass spectrometry analysis method for detecting related substances in cefotaxime sodium tazobactam sodium |
| CN116930390B (en) * | 2022-04-02 | 2024-10-29 | 南京优科生物医药股份有限公司 | Two-dimensional liquid chromatography-tandem mass spectrometry analysis method for detecting related substances in cefotaxime sodium tazobactam sodium |
| CN118243828A (en) * | 2024-05-29 | 2024-06-25 | 南京优科制药有限公司 | Detection method of tazobactam impurity A in cefotaxime sodium tazobactam sodium for injection |
| CN118243828B (en) * | 2024-05-29 | 2024-10-29 | 南京优科制药有限公司 | Detection method of tazobactam impurity A in cefotaxime sodium tazobactam sodium for injection |
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