Separation and purification method of cefuroxime sodium degradation impurities

Abstract

The invention provides a method for degrading cefuroxime sodium so as to enrich an impurity A and an impurity H, and the impurity A and the impurity H are collected by a liquid phase preparation method. The method is simple and easy to implement, the purity is relatively high, the prepared impurity A and impurity H are relatively large in amount, and the quality research of the cefuroxime sodium impurity reference substance is met.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceutical analysis technology, specifically relating to a method for separating and purifying impurities in cefuroxime sodium, applicable to the preparation and quality control of drug impurities. Background Technology

[0002] Cefuroxime sodium is a second-generation broad-spectrum cephalosporin, developed and manufactured by GlaxoSmithKline in the UK. It was first marketed in the UK in 1978, and subsequently in many countries and regions including the US, Italy, Japan, France, and China. It is widely used for respiratory tract infections, ear, nose, and throat infections, urinary tract infections, skin and soft tissue infections, and other infections. Its chemical name is (6R,7R)-7-[2-(furan-2-yl)-2-(methoxyimino)acetamido]-3-carbamoyloxymethyl-8-oxo-5-thia-1-azabicyclo[4.2.0]oct-2-ene-2-carboxylate sodium salt, chemical formula I.

[0003]

[0004] Cefuroxime sodium is listed in the Chinese Pharmacopoeia CP2020, the European Pharmacopoeia EP11, the United States Pharmacopoeia USP42, and the Japanese Pharmacopoeia JP17. The Chinese Pharmacopoeia specifies the following related substances for cefuroxime sodium: the peak area of ​​a single impurity must not exceed the area of ​​the main peak in the control solution (1.0%), and the sum of the peak areas of all impurities must not exceed three times the area of ​​the main peak in the control solution (3.0%). It does not control any single specific impurity. The European Pharmacopoeia controls specific impurities A, B, C, D, E, F, G, H, and I.

[0005] Europe is one of the important consumer markets for cefuroxime sodium, accounting for approximately 25% of the global market. Sales of cefuroxime sodium in China have grown by about 15% over the past five years, making it one of the fastest-growing markets globally. The European market is also an important export destination for Chinese raw material pharmaceutical manufacturers. Meeting the quality standards of the European Pharmacopoeia is one of the basic requirements for entering the European market; therefore, it is necessary to conduct research and control of related substances for cefuroxime sodium in accordance with the quality requirements of the European Pharmacopoeia.

[0006] Cefuroxime sodium has a relatively complex chemical structure, containing a tetra-amide ring, a sulfur-containing six-membered ring, ester bonds, and other active groups, making it relatively unstable. In recent years, Chinese pharmaceutical researchers have conducted some studies on the impurities of cefuroxime sodium. Deng Guifu (Study on the Impurity Spectrum of Cefuroxime Sodium for Injection, Chinese Journal of Antibiotics Impurities, February 2019, Vol. 44, No. 2, pp. 228-232) conducted a comparative analysis of the impurity spectra of domestically produced and imported cefuroxime sodium injections. His research suggests adding controls for specific degradation impurities of cefuroxime sodium, especially impurities A, E, and H, to the Chinese Pharmacopoeia.

[0007]

[0008] Based on the chemical structure, impurity A is a decarbamoyl degradation impurity, impurity E is a C=N double bond cis-trans isomerization impurity, and impurity H is obtained by further intramolecular cyclization of impurity A.

[0009] In existing technologies, to address impurities in active pharmaceutical ingredients (APIs) or formulations, LCMS is often used to scan the API or formulation to determine the molecular weight of potential impurities. Based on possible degradation pathways and literature review, the potential molecular structure is then inferred, and a synthetic route is designed for targeted synthesis. This method is time-consuming, costly, and often prone to reasoning errors. This can result in synthesized impurities that are only similar in mass spectrometry to the actual potential impurities in the sample, but have different chemical structures, leading to a waste of resources.

[0010] The present invention relates to two specific degradation impurities, A and H. Currently, there are no literature reports on the synthesis and preparation methods of these two impurities. In order to accurately and comprehensively carry out quality control, the preparation of high-purity impurity reference standards is of positive practical significance for the quality research of cefuroxime sodium raw material and its preparations. Summary of the Invention

[0011] This invention provides a method for degrading cefuroxime sodium to enrich impurities A and H, and for collecting impurities A and H by a preparative liquid chromatography method. This method is simple and easy to implement, yields high purity, and produces a large quantity of impurities A and H, meeting the quality requirements for cefuroxime sodium impurity reference standards.

[0012] Common degradation experiments for active pharmaceutical ingredients (APIs) or formulations include acid degradation, alkali degradation, high-temperature degradation, photodegradation, and oxidative degradation. According to the degradation test conditions in the Chinese Pharmacopoeia CP2020, under photodegradation conditions (4500±500 Lx) for 48 hours, both impurities A and H showed a significant increase, but neither exceeded 0.5%, which is unfavorable for impurity preparation. This invention provides a method for the catalytic photodegradation of cefuroxime sodium: cefuroxime sodium API and 0.5%–2% photodegradation catalyst are photodegraded in a solvent at 4500±500 Lx.

[0013] serial number Photodegradation catalysts ratio to substrate mass solvent Related substances 1 none none none 1.65％ 2 Zinc oxide 1％ water 22.41％ 3 Zinc sulfide 1％ water 14.32％ 4 Cerium oxide 1％ water 8.69％ 5 Zinc sulfide 1％ dilute acid 34.21％ 6 Zinc sulfide 0.5％ water 6.98％ 7 Zinc sulfide 2％ water 21.37％

[0014] Studies have shown that:

[0015] 1) Without using a photodegradation catalyst, the total degradation of the active pharmaceutical ingredient after 48 hours of degradation testing is no more than 2%, and the specific degradation impurities do not exceed 0.5%.

[0016] 2) Three photodegradation catalysts: zinc oxide, zinc sulfide, and cerium oxide. Under zinc oxide degradation conditions, the degradation degree was too high, with target impurities exceeding 6%. However, the increase in other related impurities was also significant, potentially interfering with subsequent preparation and separation, and increasing the difficulty of separation. Zinc sulfide showed good degradation efficiency, and the increase in target impurities was significant, all exceeding 4%, with no major impurity interference. Under cerium oxide degradation conditions, the increase in target impurities was significant, all exceeding 2%, with no significant interference from related impurities. Comparing the three photodegradation catalysts, the photodegradation catalyst is selected from any one of zinc oxide, zinc sulfide, and cerium oxide or a combination thereof. Zinc sulfide is the preferred photodegradation catalyst, followed by cerium oxide, as zinc oxide catalysis is too vigorous.

[0017] 3) Under zinc sulfide catalysis, compared with water and dilute hydrochloric acid as solvents, the degradation test showed that under dilute hydrochloric acid degradation test conditions, there were more and more impurities in the degradation, which may involve reactions such as amide four-membered ring opening and hydrolysis. Water is the preferred solvent with a concentration range of 0.1g / ml to 1g / ml, preferably 0.5g / ml.

[0018] 4) Under zinc sulfide catalysis, the effects of 0.5% and 2% catalyst dosages were investigated. Studies showed that at 0.5% catalyst dosage, the target degradation impurities exceeded 3%, and at 2% catalyst dosage, the target degradation impurities reached 8%. Therefore, the zinc sulfide dosage range is 0.5%–2.0%, with dosages of 0.5%, 0.8%, 1.0%, 1.2%, 1.4%, 1.6%, 1.8%, and 2.0%, preferably 1.0%.

[0019] The method for related substances testing of cefuroxime sodium in the Chinese Pharmacopoeia CP2020 shows good resolution; a typical HPLC chromatogram is shown in the appendix. Figure 1 Related substances test method for cefuroxime sodium: Stationary material: octylsilane-bonded silica gel; Mobile phase A: acetate buffer (0.68 g sodium acetate, 5.8 g glacial acetic acid, diluted with water to 1000 ml, pH adjusted to 3.4 with glacial acetic acid). Mobile phase B: acetonitrile.

[0020] The mobile phase contains a large amount of inorganic salts, requiring a cumbersome desalting process even after separation, making it unsuitable for direct liquid phase preparation. Furthermore, impurities E, C, B, and H are relatively similar, hindering the collection of impurity H. This invention provides a mobile phase system free of inorganic salts, comprising the following steps:

[0021] Step a) Sample preparation: Add the cefuroxime sodium photodegradation sample to an acetonitrile / water mixed solvent, filter and set aside;

[0022] Step b) Using the Hanbang Technology NP7100C system, the chromatographic column is YMC-Pack C8 (50×250mm, 10μm), the eluent is phase A: containing 0.1% trifluoroacetic acid aqueous solution, and phase B is acetonitrile, gradient elution;

[0023] Step c) Collect the fractions of impurities A and H separately, freeze-dry them to obtain impurity A and impurity H, with HPLC purity greater than 95%;

[0024] Furthermore,

[0025] In step a), the concentration of cefuroxime sodium solution is 5 mg to 50 mg / ml, preferably 20 mg / ml; the acetonitrile / water mixed solution is a 20% to 35% acetonitrile aqueous solution, preferably a 25% acetonitrile aqueous solution;

[0026] In step b), a gradient elution program was used: 0–15 min (5% B → 15% B), 15–25 min (15% B → 30% B), 25–35 min (30% B → 70% B);

[0027] Flow rate: 50–100 mL / min, preferably 80 mL / min;

[0028] Injection volume: 0.5–5 ml / time

[0029] Step c) Freeze-dry to obtain the product.

[0030] In step a), the concentration of cefuroxime sodium test sample is preferably 20 mg / ml to match the column capacity, which is within the conventional concentration range and improves the preparation efficiency;

[0031] The use of acetonitrile / water mixed solvent in step a) further improves the peak shape during the preparation process, reduces the diffusion effect, facilitates the preparation and separation of impurities, reduces the interference between impurities caused by the diffusion effect, and thus enriches a larger amount of impurity A and impurity H.

[0032] Step b) is an optimized elution procedure that further improves the enrichment efficiency of impurity H. The gradient method of Chinese Pharmacopoeia CP2020 is used. Impurities E, C, B and H are adjacent and have no value in preparing impurity H.

[0033] In all steps of this invention, the solvent is removed by freeze-drying, rather than by the conventional method of vacuum concentration. Because the sample processing and the resulting fraction contain a large proportion of water, direct concentration to remove the solvent requires high temperatures and long times; generally, the temperature for concentrating aqueous solutions needs to be close to 100°C. This concentration process leads to secondary degradation of impurities, which is not conducive to enriching high-purity impurities A and H.

[0034] The photocatalytic degradation method for cefuroxime sodium provided by this invention significantly accelerates the degradation of cefuroxime sodium, shortens the degradation time, and specifically improves the enrichment of impurities A and H, laying a good material foundation for the efficient preparation of impurities A and H. Furthermore, it provides a method for separating and purifying the degradation products of cefuroxime sodium, obtaining impurity reference standards with a chromatographic purity greater than 95%, which meets the requirements for quality research and quality control of active pharmaceutical ingredients and formulations, and has positive practical significance. Attached Figure Description

[0035] Figure 1 Typical chromatograms of related substances of cefuroxime sodium in Chinese Pharmacopoeia CP2020;

[0036] Figure 2 HPLC chromatogram of impurity A after preparation and separation

[0037] Figure 3 MS spectrum of impurity A after preparation and separation

[0038] Figure 4 HPLC chromatogram of impurity H after preparation and separation

[0039] Figure 5 MS spectrum of impurity H after preparation and separation Detailed Implementation

[0040] To further illustrate the present invention, the method for separating and purifying impurities A and H in cefuroxime sodium using preparative liquid chromatography (HPLC) provided by the present invention is described in detail below with reference to embodiments. However, these descriptions should not be construed as limiting the scope of protection of the present invention. The cefuroxime sodium, reagents, preparative liquid chromatograph, electronic analytical balance, etc., used in the specific embodiments of the present invention are all known products.

[0041] Comparative Example 1: Degradation test of cefuroxime sodium (2 days, without photodegrading agent)

[0042] Take 10g of cefuroxime sodium raw material, spread it evenly in a petri dish, keeping the thickness not exceeding 0.5cm, and place it in a light test chamber with an illuminance of 4500±500Lx for 2 days. According to HPLC purity test, the chromatographic purity of the raw material decreased from the initial 99.27% ​​to 98.35%, of which impurity A and impurity H were not greater than 0.5%.

[0043] Comparative Example 2: Degradation test of cefuroxime sodium (10 days, without photodegrading agent)

[0044] Take 10g of cefuroxime sodium raw material, spread it evenly in a petri dish, keeping the thickness not exceeding 0.5cm, and place it in a light test chamber with an illuminance of 4500±500Lx for 10 days. According to HPLC purity test, the chromatographic purity of the raw material decreased from the initial 99.27% ​​to 96.35%, of which impurity A and impurity H were both not greater than 1.0%.

[0045] Example 1: Degradation test of cefuroxime sodium (2 days, zinc oxide)

[0046] Take 10g of cefuroxime sodium raw material, add 0.1g of zinc oxide (1%) to a 100ml volumetric flask, add 50ml of purified water, and place it in a light test chamber with an illuminance of 4500±500Lx for 2 days. After HPLC purity detection, the chromatographic purity of the raw material decreased from the initial 99.27% ​​to 77.59%, of which impurity A and impurity H were both greater than 6%.

[0047] Example 2: Degradation test of cefuroxime sodium (2 days, zinc sulfide)

[0048] Take 10g of cefuroxime sodium raw material, add 0.1g of zinc sulfide (1%) to a 100ml volumetric flask, add 50ml of purified water, and place in a light test chamber with an illuminance of 4500±500Lx for 2 days. According to HPLC purity detection, the chromatographic purity of the raw material decreased from the initial 99.27% ​​to 85.68%, of which impurity A and impurity H were both greater than 3.0%.

[0049] Example 3: Degradation test of cefuroxime sodium (2 days, zinc sulfide)

[0050] Take 10g of cefuroxime sodium raw material, add 0.05g of zinc sulfide (0.5%) to a 100ml volumetric flask, add 50ml of purified water, and place in a light test chamber with an illuminance of 4500±500Lx for 2 days. According to HPLC purity detection, the chromatographic purity of the raw material decreased from the initial 99.27% ​​to 93.02%, of which impurity A and impurity H were both greater than 1.5%.

[0051] Example 4: Degradation test of cefuroxime sodium (2 days, zinc sulfide)

[0052] Take 10g of cefuroxime sodium raw material, add 0.2g of zinc sulfide (2%) to a 100ml volumetric flask, add 50ml of purified water, and place in a light test chamber with an illuminance of 4500±500Lx for 2 days. According to HPLC purity detection, the chromatographic purity of the raw material decreased from the initial 99.27% ​​to 78.63%, of which impurity A and impurity H are both greater than 4%.

[0053] Example 5: Preparation and separation of impurities A and H in cefuroxime sodium

[0054] Take the degradation product of cefuroxime sodium after 2 days (Example 2), add 500 ml of chromatographic acetonitrile / water to dissolve it, filter it through a 0.22 μm filter membrane to remove foreign matter, and obtain the test solution;

[0055] Using the Hanbang Technology NP7100C system, with a YMC-Pack C8 column (50×250mm, 10μm) and eluent phase A:

[0056] Aqueous solution containing 0.1% trifluoroacetic acid, phase B being acetonitrile, gradient elution;

[0057] Gradient elution program: 0–15 min (5% B → 15% B), 15–25 min (15% B → 30% B), 25–35 min (30% B → 70% B);

[0058] Flow rate: 80 mL / min;

[0059] Injection volume: 2 ml / time

[0060] The collected solutions rich in impurities A and H were taken separately, and the portions with HPLC purity greater than 95% were combined and lyophilized, yielding 246 mg of impurity A and 137 mg of impurity H. The HPLC purity was determined, and the structures were confirmed by mass spectrometry. The results showed that the HPLC purity was greater than 95%, and the mass spectrometry results matched the structures of impurities A and H.

Claims

1. A method for preparing cefuroxime sodium impurities A and H, wherein impurities A and H are as follows: Its features are, Includes the following steps: (1) Cefuroxime sodium was degraded in aqueous solution under the presence of a photodegradation catalyst; (2) The degradation sample was prepared by preparative liquid phase method and freeze-dried to obtain the final product.

2. The method for preparing cefuroxime sodium impurity A and impurity H according to claim 1, wherein the photodegradation catalyst in step (1) is selected from any one of zinc sulfide, zinc oxide, titanium oxide, or a combination thereof.

3. The method for preparing cefuroxime sodium impurity A and impurity H according to claim 2, characterized in that the amount of zinc sulfide photodegradation catalyst used in step (1) is 1.0%.

4. The method for preparing cefuroxime sodium impurity A and impurity H according to claim 1, characterized in that, in step (1), the photodegradation intensity is 4500±500Lx, the degradation time is 48 hours, and the concentration of cefuroxime sodium aqueous solution is 0.1g / ml~1.0g / ml.

5. The method for preparing cefuroxime sodium impurities A and H according to claim 1, characterized in that the preparation steps in step (2) are as follows: a) Sample preparation: Add the photodegraded cefuroxime sodium sample to an acetonitrile / water mixed solvent, filter and set aside; b) Using the Hanbang Technology NP7100C system, the chromatographic column was YMC-Pack C8 (50×250mm, 10μm), and the eluent was phase A: containing 0.1% trifluoroacetic acid aqueous solution, and phase B was acetonitrile, with gradient elution; c) Collect the fractions of impurities A and H, freeze-dry them to obtain impurity A and impurity H with HPLC purity greater than 95%.

6. The method for preparing cefuroxime sodium impurities A and H according to claim 5, characterized in that, in step a), the concentration of the cefuroxime sodium solution is 5 mg to 50 mg / ml; and the acetonitrile / water mixed solvent is a 20% to 35% acetonitrile aqueous solution.

7. The method for preparing cefuroxime sodium impurity A and impurity H according to claim 5, characterized in that the gradient elution in step b) is: 0-15 min (5% B → 15% B), 15-25 min (15% B → 30% B), 25-35 min (30% B → 70% B).

8. The method for preparing cefuroxime sodium impurity A and impurity H according to claim 5, characterized in that, in step b), the flow rate is 50-100 mL / min and the injection volume is 0.5-5 mL / time.

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