Method for determining impurities in ceftriaxone sodium for injection

Abstract

The invention provides a method for determining impurities in ceftriaxone sodium for injection, which is characterized in that an RP-HPLC method is adopted, and a system applicable solution is prepared by taking 1g of ceftriaxone sodium for injection, putting the ceftriaxone sodium into a 10ml measuring flask, dissolving with a phosphate buffer solution with the pH value of 7.0, diluting to a scale, uniformly mixing, sealing, placing in a drying oven with the temperature of 20-50 DEG C for less than 20 hours, and diluting with water to prepare a solution containing 2mg per 1ml; chromatographic conditions comprise that a mobile phase A is an ammonium formate solution, and a mobile phase B is mobile phase A / acetonitrile; performing linear gradient elution; the detection wavelength is 254 nm; the sample injection volume is 5 microliters. The method has good specificity and is suitable for quality control of polymer impurities.

Description

Technical Field

[0001] The present invention relates to the field of analytical chemistry, and in particular to a method for determining impurities in ceftriaxone sodium for injection. Background Art

[0002] In traditional qualitative and quantitative analysis, the TSK method is often used for impurity analysis. However, research shows that the TSK method has poor specificity in analyzing impurities in ceftriaxone polymers for injection and cannot effectively achieve the separation and quantitative analysis of impurities. Summary of the invention

[0003] The present invention aims to overcome the above defects, and uses the RP-HPLC method to analyze ceftriaxone polymer impurities, which has good specificity and is suitable for quality control of polymer impurities; the ceftriaxone positive sample solution can be used as a positioning solution for the analysis of ceftriaxone polymer impurities, and the polymer response of the positive sample solution and the actual detection analysis of the test solution are combined, and the ceftriaxone polymer 1-6 is used as a pointer polymer to control the polymer content. Further investigation of the method specificity and durability shows that the method is suitable for the quality control of ceftriaxone polymer impurities.

[0004] The present invention discloses a method for determining impurities in ceftriaxone sodium for injection, which is used for determining a sample to be tested and is characterized in that: RP-HPLC method is adopted,

[0005] The system suitability solution is as follows: 1 g of ceftriaxone sodium for injection is placed in a 10 ml volumetric bottle, dissolved and diluted to the mark with pH 7.0 phosphate buffer, mixed evenly, sealed, placed in an oven at 20-50°C for less than 20 hours, and diluted with water to make a solution containing 2 mg per 1 ml;

[0006] The chromatographic conditions include: mobile phase A is ammonium formate solution, mobile phase B is mobile phase A / acetonitrile, linear gradient elution, detection wavelength is 254 nm, injection volume is 5 μl, column temperature is 33° C. to 37° C., and flow rate is 0.9 mL / min to 1.1 mL / min.

[0007] For linear gradient elution, the initial proportion of mobile phase B was 8% to 12%.

[0008] Furthermore, the pH is preferably between 2.4 and 2.6.

[0009] The differences measured under the above conditions were all between 0.01% and 0.02%, all less than 0.05%. Ceftriaxone showed a good linear relationship in the concentration range of 0.1839 μg / ml to 3.6780 μg / ml, with a linear equation of y=0.2620x-0.0157 and a correlation coefficient of r=0.9998

[0010] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0011] The mobile phase A is 0.01 mol / L, pH 2.5 ammonium formate solution.

[0012] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0013] The above linear gradient elution conditions are as follows:

[0014]

[0015] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0016] A Diamonsil Plus C18, 4.6×250 mm chromatographic column was used.

[0017] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0018] The types of impurities that can be separated by the above method are as follows:

[0019]

[0020]

[0021] In addition, the present invention also provides a carrier for determining impurities in ceftriaxone sodium for injection, characterized in that: it comprises a determination module and an analysis module;

[0022] The above-mentioned determination module obtains the retention time of each peak in the sample to be tested according to the above-mentioned method;

[0023] The above analysis module determines the impurities and their contents in the sample to be tested according to the retention time of each peak and based on the impurity characteristics shown in the above table.

[0024] Furthermore, the present invention also provides a method for determining impurities in ceftriaxone sodium for injection, which is used to identify and distinguish various impurities in ceftriaxone sodium, and is characterized in that it comprises the following steps:

[0025] S1. Enrichment of polymer impurities of ceftriaxone for injection using TSK method;

[0026] S2. Qualitative analysis of polymer impurities in ceftriaxone sodium was performed using RP-HPLC-Qtof method.

[0027] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0028] In S1, the selected failure condition is:

[0029] The diluent was pH = 7 phosphate buffer;

[0030] The concentration of the destruction solution is: 10-100 mg / mL;

[0031] Storage conditions are 20-50℃;

[0032] The placement time is 2-20 hours.

[0033] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0034] In S1, the chromatographic conditions are:

[0035] Mobile phase A was disodium hydrogen phosphate solution / sodium dihydrogen phosphate solution (50 / 50)-acetonitrile;

[0036] The flow rate was 0.6 mL / min, the injection volume was 5-20 μL, the column temperature was 30 °C, the detection wavelength was 254 nm, and the elution was isocratic.

[0037] Furthermore, the present invention provides a method for determining impurities in ceftriaxone sodium for injection, characterized in that:

[0038] In S2, the chromatographic conditions are:

[0039] Mobile phase A was ammonium formate solution, mobile phase B was mobile phase A / acetonitrile; linear gradient elution; detection wavelength was 254 nm; injection volume was 5 μl;

[0040] The process of the linear gradient elution is as shown in claim 3. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 . Overlay of different diluents at the same concentration and temperature for equivalent destruction time;

[0042] Figure 2 .Superimpose the graphs of different concentrations in the same diluent and temperature for the same destruction time;

[0043] Figure 3 .Destruction of overlays at different temperatures in the same diluent and concentration;

[0044] Figure 4 . Overlay the destruction temperature at different times with the same diluent and concentration;

[0045] Figure 5.Typical spectrum of HPSEC positive samples;

[0046] Figure 6 .Enriched impurity confirmation overlay;

[0047] Figure 7 .TSK method enrichment of impurities, trans-cefotaxime, and positive sample solution overlay;

[0048] Figure 8 .The first-level mass spectrum of the target peak in the HPSEC-3 enrichment solution;

[0049] Fig. 9 . Primary mass spectrum of target peak in positive sample solution;

[0050] Fig.10 . Primary mass spectrum of target peak in enriched solution;

[0051] Fig.11 . Primary mass spectrum of target peak in positive sample solution;

[0052] Fig.12 . Primary mass spectrum of target peak in enriched solution

[0053] Fig.13 . Primary mass spectrum of target peak in positive sample solution

[0054] Fig.14 . Primary mass spectrum of target peak in enriched solution

[0055] Fig.15 . Primary mass spectrum of target peak in positive sample solution

[0056] Fig.16 . Primary mass spectrum of target peak in HPSEC-1 enrichment solution

[0057] Fig.17 . Primary mass spectrum of target peak in HPSEC-2 enrichment solution

[0058] Fig.18 . Primary mass spectrum of target peak in HPSEC-3 enriched solution

[0059] Fig.19 . Primary mass spectrum of target peak in positive sample solution

[0060] Fig. 20 . Primary mass spectrum of target peak in positive sample solution

[0061] Fig.21 .Trans-cefotaxime primary mass spectrum

[0062] Fig. 22 . Primary mass spectrum of target peak in positive sample solution

[0063] Fig.23 .Second-order mass spectrum of the target peak in the positive sample solution (m / z 950.0928)

[0064] Fig.24 . Primary mass spectrum of target peak in HPSEC-4 enrichment solution

[0065] Fig.25 .Secondary mass spectrum of target peak in enriched solution HPSEC-4 (m / z 950.0907)

[0066] Fig.26 . Primary mass spectrum of target peak in positive sample solution

[0067] Fig. 27 .Second-order mass spectrum of the target peak in the positive sample solution;

[0068] Fig.28 . Primary mass spectrum of target peak in HPSEC-1 enrichment solution

[0069] Fig.29 . Primary mass spectrum of target peak in HPSEC-3 enriched solution

[0070] Fig.30 . Primary mass spectrum of target peak in positive sample solution

[0071] Fig.31 . Primary mass spectrum of target peak in HPSEC-4 enrichment solution

[0072] Fig.32 . Primary mass spectrum of target peak in positive sample solution

[0073] Fig.33 .Second-order mass spectrometry of target peak in positive sample solution

[0074] Fig.34 .Typical chromatogram of positive sample solution;

[0075] Fig.35 . System suitability solution chromatogram; (for all impurities that destroy the solution)

[0076] Fig.36 .Sensitivity solution chromatogram;

[0077] Fig.37 .Typical chromatogram of system suitability solution (for polymer impurities 1, 2, 3, 5, 6)

[0078] Fig.38 Typical chromatogram of the test solution DETAILED DESCRIPTION

[0079] The present invention is capable of various modifications and various embodiments, and therefore specific embodiments are illustrated and described in the accompanying drawings. However, this is not intended to limit the present invention to specific implementations, but should be understood to include all modifications, equivalents, and even substitutes that fall within the concept and technical scope of the present invention.

[0080] Example 1. Method for identifying impurities in ceftriaxone sodium for injection

[0081] S1.1. Enrichment of polymer impurities of ceftriaxone for injection using TSK method

[0082] 1. Screening of positive sample preparation solutions

[0083] This example was screened under the following 8 conditions:

[0084]

[0085]

[0086] like Figure 1 As shown, when different diluents are used at the same concentration, temperature and destruction time, there is no obvious difference in the number of impurities produced by salt destruction and water destruction. The impurity content produced by salt destruction is relatively large, so salt solution is temporarily selected as the diluent.

[0087] like Figure 2 As shown, when different concentrations are used in the same diluent, at the same temperature and for the same destruction time, there is no significant difference in the number of impurities produced. The impurity content produced by the high concentration of 100 mg / mL destruction is relatively large, so the concentration of the destruction solution is temporarily set at 100 mg / mL.

[0088] like Figure 3 As shown, under the same diluent and the same concentration of destruction at different temperatures, the number of impurities produced is basically the same. There is no obvious difference in the impurity content produced by destruction at 50°C. Based on rapid preparation, the destruction temperature is selected to be 50°C.

[0089] like Figure 4 As shown, at different times, under the same diluent, the same concentration and the same destruction temperature, the number of impurities produced is basically the same. As the destruction time increases, the impurity content increases, reaching the maximum value at 20 hours of destruction, and the main peak area percentage is less than 80%, which is excessive degradation. Based on rapid preparation, the destruction time is tentatively set at 2 hours.

[0090] In summary, through the comparison of various degradation conditions, the preparation conditions of positive samples are determined as follows:

[0091] Take about 1 g of ceftriaxone sodium for injection, weigh accurately, place in a 10 mL volumetric bottle, dissolve and dilute to the scale with phosphate solution, shake well, and leave 50 bottles under the same conditions for 2 hours as the destruction solution stock solution.

[0092] Positive sample screening solution (0.5 mg / mL): Take 5 μL of ceftriaxone sodium destruction solution and place it in a sample vial, add 0.995 mL of diluent, and shake well.

[0093] 2. Chromatographic parameters are as follows

[0094]

[0095] 3. Typical spectrum of HPSEC positive samples Figure 5 As shown, the specific impurities are classified as follows

[0096] name Retention time (min) Relative retention time HPSEC-1 9.963 0.83 HPSEC-2 10.153 0.85 HPSEC-3 10.433 0.87 HPSEC-4 10.937 0.91 Ceftriaxone 11.990 1.00 HPSEC-5 13.943 1.16

[0097] 4. By increasing the concentration of the positive test sample (injection concentration 10 mg / mL) and the injection volume (20 μl), enrich the relevant impurities and confirm the naming as follows Figure 6 shown.

[0098] S1.2. Qualitative analysis of polymer impurities in ceftriaxone sodium using RP-HPLC-Qtof method

[0099] 1. Chromatographic conditions

[0100]

[0101] 2. Solution Preparation

[0102] Ceftriaxone sodium positive sample solution: the solution of the above-mentioned ceftriaxone sample No. 5, which has been placed for 2 hours;

[0103] Positive sample (2 mg / mL): Take 20 μL of ceftriaxone sodium destruction solution and place it in a sample vial, add 0.98 mL of diluent, and shake well.

[0104] Test solution (2.2 mg / mL): Weigh 22 mg of each batch of ceftriaxone sodium for injection into a centrifuge tube and add 10 mL of diluent to dissolve.

[0105] Trans-cefotaxime (2 μg / mL): Take 2 mg of trans-cefotaxime, add 10 mL of water to dissolve, mix well, then take 10 μL, add 990 μL of water to dilute, and mix well to obtain the product.

[0106] TSK method enriched impurity solution: that is, each enriched impurity solution.

[0107] 3. Experimental results

[0108] The HPSEC impurities enriched by the TSK method and the positive test solution were scanned by RP-HPLC. In addition to the main component, the positive sample had ten characteristic peaks. The correlation of the remaining HPSEC enriched impurities is as follows: Figure 7The specific corresponding relationship is shown in the following table:

[0109]

[0110]

[0111] Peak 1 mass spectrum analysis

[0112] Peak 1 elutes at 3.796 min in the RP-HPLC method and appears in the HPSEC-3 enriched impurity by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-3 is before ceftriaxone, so it is suspected to be a polymer impurity. In the positive ion mode, the primary spectra of the target peak at the retention time in the positive sample solution and the TSK enriched sample are basically the same, which is the same substance. Figure 8 and 9 As shown in the figure, there are characteristic ion peaks of m / z 243.05, m / z 265.03, and m / z 281.01, which are [M+H] + 、[M+Na] + , [M+K] + Peak, estimated molecular weight is 242. It is a small molecule impurity, non-polymer impurity, and its structure needs further analysis.

[0113] Peak 2 mass spectrum analysis

[0114] Peak 2 elutes at 5.163 min in the RP-HPLC method and appears in the impurity HPSEC-5 enriched by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-5 is after ceftriaxone, so it should be a small molecule impurity in theory. Under the positive ion mode, the primary spectra of the target peak at this retention time in the positive sample solution and the TSK enriched sample are basically the same, indicating that they are the same substance. Fig.10 and 11 As shown in the figure, there is a characteristic ion peak of m / z 555.05 in the primary mass spectrum, which is [M+H] + The peak has a predicted molecular weight of 554, which is consistent with the molecular weight of ceftriaxone, and is speculated to be a ceftriaxone isomer.

[0115] Peak 3 mass spectrum analysis

[0116] Peak 3 elutes at 5.743 min in the RP-HPLC method and appears in the impurity HPSEC-5 enriched by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-5 is after ceftriaxone, so it should be a small molecule impurity in theory. Under the positive ion mode, the primary spectra of the target peak at this retention time in the positive sample solution and the TSK enriched sample are basically the same, indicating that they are the same substance. Fig.12 and13 As shown in the figure, there is a characteristic ion peak of m / z 160.0176 in the primary mass spectrum, which is [M+H] + The peak has a presumed molecular weight of 159 and is a small molecule impurity. Based on the molecular weight and retention time, it is presumed to be ceftriaxone impurity C.

[0117]

[0118] Peak 4 mass spectrum analysis

[0119] Peak 4 elutes at 7.683 min in the RP-HPLC method and appears in the impurity HPSEC-5 enriched by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-5 is after ceftriaxone, so it should be a small molecule impurity in theory. Under the positive ion mode, the primary spectra of the target peak at this retention time in the positive sample solution and the TSK enriched sample are basically the same, indicating that they are the same substance. Fig.14 and 15 As shown in the first-level mass spectra, there is a molecular ion peak of m / z 414.05, which is [M+H] + The peak has an estimated molecular weight of 413, which is a small molecule impurity and is estimated to be the impurity B lactone ring opening impurity.

[0120]

[0121] Peak 5 mass spectrum analysis

[0122] Peak 5 elutes at 12.369 min in the RP-HPLC method and appears in the impurities HPSEC-1, HPSEC-2 and HPSEC-3 enriched by the TSK method. According to the retention principle of the TSK method, the peaks of HPSEC-1 to HPSEC-3 elute before ceftriaxone. Theoretically, it may be a polymer impurity with a molecular weight greater than ceftriaxone. In the positive ion mode, the primary spectra of the target peak at this retention time in the positive sample solution and the TSK enriched sample are basically the same, indicating that they are the same substance. Figure 16-19 As shown in the first-level mass spectra, there is a molecular ion peak of m / z 667.07, and the estimated molecular weight is 666, which is [M+H] + The peak is inferred to have the following structure:

[0123]

[0124] Peak 6 mass spectrum analysis

[0125] Peak 6 was found at 18.303 min in the RP-HPLC method and was not found in the impurities enriched by the TSK method. According to the known impurities, its retention time was consistent with that of trans-cefotaxime. The target peak in the positive sample solution and the primary mass spectrometry of trans-cefotaxime in the reference solution were compared. Fig. 20 and 21 Comparison shows that there is a molecular ion peak of m / z 555.05, which is [M+H] + The peak had a predicted molecular weight of 554, which was confirmed to be trans-cefotaxime.

[0126]

[0127] Peak 7 mass spectrum analysis

[0128] Peak 7 was eluted at 19.083 min in the RP-HPLC method and was not found in the impurities enriched by the TSK method. The target peak produced molecular ion peaks m / z 475.5 and m / z 950 in the positive ion mode. The mass-to-charge ratio difference of adjacent bar graphs of m / z 475.5 was 0.5. It was judged that m / z 475.5 was a doubly charged ion generated by m / z 950. The quasi-molecular ion of the target peak was determined to be m / z 950, and the nitrogen on the thiazole ring did not react. The impurity cleavage process in the secondary mass spectrometry produced ceftriaxone m / z 555, indicating that the structure of ceftriaxone contained in the structure did not change. Figure 22-23 The fragment ion of m / z 730 is produced by the desorption of the triazine ring, 1 carboxyl group and 1 O from the peak 7 impurity.

[0129] The possible structure is as follows:

[0130]

[0131] Peak 8 mass spectrum analysis

[0132] Peak 8 elutes at 20.196 min in the RP-HPLC method and appears in the HPSEC-4 enriched impurity by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-4 is before ceftriaxone. Theoretically, it may be a polymer impurity with a molecular weight greater than ceftriaxone. The primary spectra of the target peak at this retention time in the positive sample solution and the TSK enriched sample are basically the same, indicating that they are the same substance. In the positive ion mode, in the primary mass spectrum, there are a series of summed ion peaks with molecular ion peaks of m / z 950.0908, m / z 972.0720, and m / z 988.0460, which are [M+H] + 、[M+Na] + , [M+K] +, its molecular weight is estimated to be 949, m / z 475.5489 is a doubly charged ion generated by m / z 950.0908, and the nitrogen on the thiazole ring has not reacted; in the secondary mass spectrum, the fragment ion of m / z 730 is generated by the peak 8 impurity detriazine ring, 1 carboxyl group and 1 O. Figure 24-27 As shown, the possible structure is as follows:

[0133]

[0134] Peak 9 mass spectrum analysis

[0135] Peak 9 elutes at 24.608 min in the RP-HPLC method, appears in the impurity HPSEC-1 enriched by the TSK method, and is not found in the positive sample solution. According to the retention principle of the TSK method, the peak position of HPSEC-1 is before ceftriaxone, and theoretically it may be a polymer impurity with a molecular weight greater than ceftriaxone. In the positive ion mode, in the primary mass spectrum, there is a molecular ion peak of m / z889.0107, which is [M+H] + peak, its molecular weight is estimated to be 888. Fig.28 The structure is to be inferred.

[0136] Peak 10 mass spectrum analysis

[0137] Peak 10 elutes at 25.529 min in the RP-HPLC method and appears in the impurity HPSEC-3 enriched by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-3 is before ceftriaxone. Theoretically, it may be a polymer impurity with a molecular weight greater than ceftriaxone. In the positive ion mode, the primary spectra of the target peak at this retention time in the positive sample solution and the TSK enriched sample are basically the same, indicating that they are the same substance. In the primary mass spectrum, Figure 29-30 As shown, there are a series of molecular ion peaks m / z 1096.0939, m / z 1118.0521, and m / z 1134.0204, which are [M+H] + 、[M+Na] + , [M+K] + , its molecular weight is estimated to be 1095, and m / z 548 is judged to be a doubly charged ion generated by m / z 1096. The estimated structure is as follows:

[0138]

[0139] Peak 11 mass spectrum analysis

[0140] Peak 11 elutes at 27.949 min in the RP-HPLC method and appears in the HPSEC-4 enriched impurities by the TSK method. According to the retention principle of the TSK method, the peak position of HPSEC-4 is before ceftriaxone. Theoretically, it may be a polymer impurity with a molecular weight greater than that of ceftriaxone. In the positive ion mode, the primary spectra of the target peak at the retention time in the positive sample solution and the TSK enriched sample are basically consistent, and they are the same substance. In the primary mass spectrum, there are molecular ion peaks m / z 475.5 and m / z 950. It is judged that m / z 475.5 is a doubly charged ion generated by m / z 950, and the quasi-molecular ion of peak 11 is determined to be m / z 950, and the nitrogen on the thiazole ring has not reacted; in the secondary mass spectrum, m / z 396 is the chemical bond breakage of the polymerization after peak 11 removes the triazine ring, producing two molecules of the same ceftriaxone detriazine ring structure; m / z 324 is m / z 396 losing a molecule of CO2 and CO is produced. Figure 31-33 The inferred structure is as follows:

[0141]

[0142] Summary and conclusions of mass spectrometry studies

[0143]

[0144]

[0145] Research conclusion: The TSK method has poor specificity in determining polymer impurities. Small molecular impurities are contained in the polymer peak before the main peak, which affects the accurate quantification of the polymer. Based on the existing RP-HPLC method, the related impurities that appear before the main peak in the TSK method are studied. The polymer impurities can be effectively detected in the RP-HPLC method with good specificity, which is suitable for the quality control of polymer impurities.

[0146] Example 2. Systemic Solutions and Sensitivity

[0147] 2.1. Chromatographic conditions: Since the main peak of the 2.2 mg / mL test solution was overloaded when the injection volume was 20 μL, the injection volume was changed to 5 μL, the modified method was confirmed, and multiple batches of samples were scanned.

[0148]

[0149] 2.2. Solution preparation

[0150] Diluent: Purified water

[0151] Test solution (2.2 mg / mL): Weigh 22 mg of ceftriaxone sodium for injection into a 10 mL volumetric flask and add diluent to dissolve to the mark.

[0152] 0.1% self-control solution: Measure 1.0mL of the test solution, place it in a 100mL volumetric flask, dilute it to the mark with diluent, shake well, take 1.0mL of the above solution, place it in a 10mL volumetric flask, dilute it to the mark with diluent, shake well, as the 0.1% self-control solution.

[0153] Buffered saline solution: weigh 0.18 g of disodium hydrogen phosphate and 0.72 g of sodium dihydrogen phosphate into the mobile phase bottle, add 50 ml of water, mix well, filter, and degas by ultrasonication.

[0154] System suitability solution (2 mg / mL): Weigh approximately 1.0 g of ceftriaxone sodium for injection, place in a 10 mL volumetric flask, dissolve and dilute to the mark with phosphate buffer, shake well, and place at 50 °C for 2 h to prepare the system suitability solution stock solution (100 mg / mL); Accurately measure 1 mL of the system suitability solution stock solution, place in a 50 mL volumetric flask, dissolve and dilute to the mark with purified water, shake well, and prepare the system suitability solution.

[0155] Quantitation limit solution: Pipette 1.0 mL of 0.1% self-control solution into a 10 mL volumetric flask, dilute to the mark with diluent, and shake well.

[0156] 2.3 Experimental Results

[0157] The results of the system suitability sample solution and sensitivity solution are as follows Fig.35 and 36 As shown, the separation between the polymer peak and the adjacent peaks in the system suitability solution is greater than 1.5, and the signal-to-noise ratio S / N of the sensitivity solution is greater than 10, both of which meet the acceptance criteria.

[0158] Example 3. Test sample detection

[0159] 3.1. Solution configuration:

[0160] Phosphate buffer (pH 7.0): weigh 0.18 g of disodium hydrogen phosphate and 0.72 g of sodium dihydrogen phosphate, dissolve them by ultrasonication in water, dilute to 50 mL, and mix well to obtain the solution.

[0161] Test solution Take an appropriate amount of ceftriaxone sodium for injection, accurately weigh 22.0 mg and place it in a 10 ml volumetric flask, add water to dissolve and quantitatively dilute to the scale, and shake well.

[0162] 0.1% Self-control Solution Take an appropriate amount of the test solution, accurately measure 1 ml and place it in a 100 ml volumetric flask, dilute it to the mark with water, shake well, then accurately measure 1 ml and place it in a 10 ml volumetric flask, dilute it to the mark with water, shake well.

[0163] System suitability solution: Take about 1 g of ceftriaxone sodium for injection, weigh accurately, place in a 10 ml volumetric flask, dissolve and dilute to the scale with phosphate buffer (pH 7.0), mix well, seal, place in a 50°C oven for 2 h, and dilute with water to make a solution containing about 2 mg per 1 ml.

[0164] 3.2. Chromatographic conditions: Octadecylsilane bonded silica gel was used as filler (Diamonsil Plus C18, 4.6 mm × 250 mm, 5 μm); mobile phase A was 0.01 mol / L ammonium formate solution (pH adjusted to 2.5 with formic acid), and mobile phase B was mobile phase A-acetonitrile (20:80); column temperature was 35 °C; linear gradient elution was performed according to the table below; flow rate was 1.0 ml per minute; retention time of ceftriaxone was about 12 minutes; detection wavelength was 254 nm; injection volume was 5 μl.

[0165]

[0166] 3. Determination method

[0167] Accurately measure the system suitability solution, the test solution and the 0.1% self-control solution, inject them into the liquid chromatograph respectively, and record the chromatogram. The impurity peaks in the system suitability chromatogram are identified and analyzed by the polymer impurity structure, and polymer impurity 1, polymer impurity 2, polymer impurity 3, polymer impurity 5, and polymer impurity 6 are determined to emerge in sequence. The retention time of the main peak of ceftriaxone is about 12-14 minutes, and the separation degree of the main peak of ceftriaxone and the polymer impurity peak meets the requirements of the pharmacopoeia. Fig.34 This is a typical chromatogram of a positive solution during mass spectrometry analysis. Fig.37 is a typical chromatogram of a system suitability solution. Fig.38 This is a typical chromatogram of the test solution. The relative retention time of each polymer impurity peak is shown in the following table:

[0168] name Relative retention time Ceftriaxone 1.00 Polymer Impurities 1 0.89 Polymer Impurity 2 1.41 Polymer Impurity 3 1.52 Polymer Impurities 5 1.93 Polymer Impurities 6 2.06

[0169] 4. Representative sample test results

[0170]

[0171]

[0172] Results of accelerated test and 3-month long-term test of 3 batches of homemade preparations under the original method chromatographic conditions Results of accelerated test of 3 batches of homemade preparations

[0173]

[0174]

[0175] Long-term test results of 3 batches of homemade preparations (3 months)

[0176]

Claims

1. A method for determining impurities in ceftriaxone sodium for injection, characterized in that: Using RP-HPLC method, The system suitability solution is as follows: 1 g of ceftriaxone sodium for injection is placed in a 10 ml volumetric bottle, dissolved and diluted to the mark with pH 7.0 phosphate buffer, mixed evenly, sealed, placed in an oven at 20-50°C for less than 20 hours, and diluted with water to make a solution containing 2 mg per 1 ml; The chromatographic conditions included: mobile phase A was ammonium formate solution, mobile phase B was mobile phase A / acetonitrile; column temperature was 33° C. to 37° C., flow rate was 0.9 mL / min to 1.1 mL / min; linear gradient elution; detection wavelength was 254 nm; injection volume was 5 μl.

2. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 1, characterized in that: The mobile phase A is 0.01 mol / L ammonium formate solution with a pH of 2.

5.

3. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 1, characterized in that: The linear gradient elution conditions are as follows:

4. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 1, characterized in that: A Diamonsil Plus C18, 4.6×250 mm chromatographic column was used.

5. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 1, characterized in that: The types of impurities that can be separated by the method are as follows:

6. A carrier for determining impurities in ceftriaxone sodium for injection, characterized in that: It includes a determination module and an analysis module; The determination module obtains the retention time of each peak in the sample to be tested according to any one of claims 1 to 3; The analysis module determines the impurities and their contents in the sample to be tested according to the retention time of each peak and based on the impurity characteristics as claimed in claim 4.

7. A method for determining impurities in ceftriaxone sodium for injection, characterized in that: The following steps are included: S1. Enrichment of polymer impurities of ceftriaxone for injection using TSK method; S2. Qualitative analysis of polymer impurities in ceftriaxone sodium was performed using RP-HPLC-Qtof method.

8. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 7, characterized in that: In S1, the selected failure condition is: The diluent was pH = 7 phosphate buffer; The concentration of the destruction solution is: 10-100 mg / mL; Storage conditions are 20-50℃; The placement time is 2-20 hours.

9. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 7, characterized in that: In S1, the chromatographic conditions are: Mobile phase A was disodium hydrogen phosphate solution / sodium dihydrogen phosphate solution (50 / 50)-acetonitrile; Flow rate 0.6mL / min, injection volume 5-20μL, column temperature 30℃, detection wavelength 254nm, isocratic elution.

10. A method for determining impurities in ceftriaxone sodium for injection as claimed in claim 7, characterized in that: In S2, the chromatographic conditions are: Mobile phase A was ammonium formate solution, mobile phase B was mobile phase A / acetonitrile; linear gradient elution; detection wavelength was 254 nm; injection volume was 5 μl; The process of linear gradient elution is as shown in claim 3.

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