A method for detecting related substances of thiamphenicol or its preparations

Abstract

The invention discloses a method for detecting a substance related to thiamphenicol or its preparation, and belongs to the technical field of pharmaceutical and chemical detection. The detection method of the invention adopts an octadecyl bonded silica gel as a filler chromatographic column, and a phosphate aqueous solution-acetonitrile as a mobile phase; and separates and determines the substance related to thiamphenicol or its preparation in a reverse phase chromatography system. The detection method of the invention can effectively separate and determine the substance related to thiamphenicol or its preparation, and has the advantages of good separation, high theoretical plate number, reliable accuracy, good repeatability, etc.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical and chemical detection technology, and specifically relates to a method for detecting thiamphenicol or related substances in its preparations Background Art

[0002] Thiamphenicol, chemical name: [R-(R*,R*)]N-[1-(hydroxymethyl)-2-hydroxy-2-[4-(methylsulfonyl)phenyl]ethyl]-2,2-dichloroacetamide, has the following structural formula:

[0003]

[0004] Thiamphenicol is also known as thiamphenicol, and its chemical name is 1-p-methylsulfonyl-phenyl-2-dichloroacetamide-1,3-propanediol. It is a chloramphenicol antibiotic and belongs to the amide alcohol class of drugs. Its antibacterial spectrum and antibacterial effect are similar to those of chloramphenicol, and it has a broad-spectrum antibacterial effect, but the methylsulfonyl group of thiamphenicol replaces the nitro group of chloramphenicol, so its toxicity is reduced. Thiamphenicol is widely used in the treatment of typhoid fever, dysentery, intestinal infection, brucellosis, meningitis and surgical infection in clinical practice. In animal husbandry, it is mainly used for the treatment of bacterial infections of poultry (such as Escherichia coli, Salmonella, Salmonella typhi, etc.), and can also be used to prevent and treat bacterial diseases of aquatic organisms such as fish. my country began to trial it in 1971. At present, the thiamphenicol preparations widely used in clinical practice are all oral solid preparations, such as enteric-coated tablets, capsules, granules, etc.

[0005] Foreign pharmacopoeias include thiamphenicol, but do not control its related substances.

[0006] The Chinese Pharmacopoeia includes thiamphenicol and its preparations, and uses high performance liquid chromatography to control impurities, requiring that a single impurity should not exceed 1.0% and the total impurity should not exceed 2.0%. However, the detection ability is poor, some impurities cannot be effectively detected, there are problems such as impurity omissions, poor peak shape, and low response, which cannot meet the detection of related substances of thiamphenicol or its preparations.

[0007] In the current prior art, patent document CN 117686628 A discloses a method for detecting genotoxic impurities of thiamphenicol. This method is used for detecting dichloroacetic acid in thiamphenicol. When its chromatographic conditions are used for detecting thiamphenicol related substances, the specificity is poor, the blank solvent peak interferes with the detection of impurities, and some impurities overlap, so that each impurity cannot be effectively separated and detected. And another patent document CN 106353437 A discloses a method for detecting thiamphenicol residues in a sample. When its chromatographic conditions are used for detecting thiamphenicol related substances, the specificity is poor, the blank solvent peak interferes with the detection of impurities, the peak shape of some impurities is poor, and some impurities have double peaks. It cannot meet the detection of thiamphenicol or its preparation related substances.

[0008] Therefore, in order to ensure the quality of drugs and the safety of drug use, it is necessary to develop a more sensitive method for detecting related substances. Summary of the invention

[0009] [Technical issues]

[0010] Provided is a method for achieving effective separation and detection of thiamphenicol or its preparation-related substances and thiamphenicol, as well as thiamphenicol or its preparation-related substances, thereby ensuring higher drug quality and better drug safety.

[0011] [Technical solution]

[0012] In order to solve the above problems, the present invention provides a method for detecting thiamphenicol or related substances of its preparations. The method has strong specificity, good repeatability, precision, tolerance, and high sensitivity, and can effectively detect the contents of 6 organic impurities in thiamphenicol, further ensuring the safety, effectiveness, and controllability of clinical drug use.

[0013] Specifically:

[0014] The present invention provides a method for detecting thiamphenicol or a substance related to its preparation, the method comprising the following steps:

[0015] Preparation of test solution: Take an appropriate amount of thiamphenicol or its preparation to prepare the test solution;

[0016] Chromatographic conditions: Octadecyl bonded silica gel was used as the filler column, and phosphate aqueous solution-acetonitrile was used as the mobile phase;

[0017] Determination method: Take the test sample solution and inject it into the liquid chromatograph for determination.

[0018] In one embodiment of the present invention, the thiamphenicol or its preparation-related substance comprises the structure shown below:

[0019]

[0020] In one embodiment of the present invention, the method for preparing the test solution is: take thiamphenicol or its preparation, dissolve and dilute it with the mobile phase to obtain a test solution with a concentration of 0.1 to 3 mg / mL. Further preferably, the method for preparing the test solution is: take an appropriate amount of thiamphenicol or its preparation, accurately weigh it, put it in a volumetric flask, dissolve it with the mobile phase and dilute it to make a 1 mg / mL solution.

[0021] In one embodiment of the present invention, the mobile phase is 70-90:30-10 phosphate aqueous solution: acetonitrile solution. Further preferably, the mobile phase is 75-85:25-15 phosphate aqueous solution: acetonitrile solution; most preferably, the mobile phase is 80:20 phosphate aqueous solution: acetonitrile solution.

[0022] In one embodiment of the present invention, the concentration of the phosphate aqueous solution is 0.001 mol / L-0.1 mol / L.

[0023] In one embodiment of the present invention, the phosphate is dihydrogen phosphate or dihydrogen phosphate.

[0024] In one embodiment of the present invention, the phosphate aqueous solution is a 0.001mol / L-0.1mol / L dihydrogen phosphate solution or a dihydrogen phosphate solution. Further optimized, the phosphate aqueous solution is a 0.01mol / L-0.05mol / L dihydrogen phosphate aqueous solution; most preferably, the phosphate aqueous solution is a 0.05mol / L potassium dihydrogen phosphate aqueous solution.

[0025] In one embodiment of the present invention, the detection wavelength is 210-240nm, the column temperature is 20-30°C, the flow rate is 0.5-2mL / solution and min, and the injection volume is 1-30μL; further preferably, the detection wavelength is 225nm, the column temperature is 25°C, the flow rate is 1.0mL / min, and the injection volume is 10μL.

[0026] In one embodiment of the present invention, the preparation includes capsules, tablets, and granules.

[0027] The present invention also provides application of the method in drug quality control.

[0028] Beneficial effects:

[0029] The detection method of the present invention adopts octadecyl bonded silica gel as a filler chromatographic column, phosphate aqueous solution-acetonitrile as a mobile phase; and separates and determines thiamphenicol or its preparation related substances in a reverse phase chromatographic system. The analytical method provided by the present invention can effectively separate and determine thiamphenicol or its preparation related substances, and has the advantages of good separation degree, high theoretical plate number, reliable accuracy, good repeatability, etc.

[0030] Compared with the existing reported thiamphenicol impurity detection method, the detection method of the present invention has the following advantages: ① separation of thiamphenicol and related impurities is achieved; ② interference of blank solvent peak with impurity detection is eliminated; ③ impurity leakage is effectively prevented. BRIEF DESCRIPTION OF THE DRAWINGS

[0031] Figure 1 It is the chromatogram of blank solution in Example 1.

[0032] Figure 2 The chromatogram of the test solution in Example 1 is shown in FIG.

[0033] Figure 3 This is the solution chromatogram of the system suitability in Example 1.

[0034] Figure 4 It is the system suitability solution chromatogram in Comparative Example 1 (Chinese Pharmacopoeia 2020 Edition).

[0035] Figure 5 This is the chromatogram of the blank solution in Comparative Example 2 (CN 117686628 A patent).

[0036] Figure 6 This is the solution chromatogram of the system suitability in Comparative Example 2 (CN 117686628 A patent).

[0037] Figure 7 This is the chromatogram of the blank solution in Comparative Example 3 (CN 106353437 A patent).

[0038] Figure 8 This is the solution chromatogram of the system suitability in Comparative Example 3 (CN 106353437 A patent). DETAILED DESCRIPTION

[0039] The exemplary embodiments of the present invention will be described in more detail below with reference to the accompanying drawings. Although the exemplary embodiments of the present invention are shown in the accompanying drawings, it should be understood that the present invention can be implemented in various forms and should not be limited by the embodiments set forth herein. On the contrary, these embodiments are provided in order to enable a more thorough understanding of the present invention and to enable the scope of the present invention to be fully communicated to those skilled in the art.

[0040] Example 1

[0041] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0042] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0043] Detection wavelength: 225nm;

[0044] Column temperature: 25°C;

[0045] Flow rate: 1.0 mL / min;

[0046] Preparation of blank solution: mobile phase;

[0047] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0048] Preparation of test solution: Weigh about 50 mg of thiamphenicol test sample into a 50 mL volumetric flask, accurately weigh, dissolve and dilute to the scale with mobile phase.

[0049] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph, and record the chromatogram. Figure 1-3 .

[0050] The test results show that the separation between thiamphenicol and its related impurities in the system suitability solution is greater than 1.5, the blank solution does not interfere with the detection, and the method has good specificity. At the same time, it can effectively separate thiamphenicol and its related impurities in the test sample, preventing impurities from being missed and achieving higher requirements for drug quality control.

[0051] Example 2

[0052] Chromatographic conditions and system suitability experiments

[0053] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0054] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0055] Detection wavelength: 225nm;

[0056] Column temperature: 20°C;

[0057] Flow rate: 1.0 mL / min;

[0058] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0059] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0060] Example 3

[0061] Chromatographic conditions and system suitability experiments

[0062] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0063] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0064] Detection wavelength: 225nm;

[0065] Column temperature: 45°C;

[0066] Flow rate: 1.0 mL / min;

[0067] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0068] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0069] Example 4

[0070] Chromatographic conditions and system suitability experiments

[0071] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0072] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0073] Detection wavelength: 225nm;

[0074] Column temperature: 25°C;

[0075] Flow rate: 0.5 mL / min;

[0076] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0077] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0078] Example 5

[0079] Chromatographic conditions and system suitability experiments

[0080] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0081] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0082] Detection wavelength: 225nm;

[0083] Column temperature: 25°C;

[0084] Flow rate: 2.0 mL / min;

[0085] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0086] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0087] Example 6

[0088] Chromatographic conditions and system suitability experiments

[0089] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0090] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (75:25);

[0091] Detection wavelength: 225nm;

[0092] Column temperature: 25°C;

[0093] Flow rate: 1.0 mL / min;

[0094] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0095] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0096] Example 7

[0097] Chromatographic conditions and system suitability experiments

[0098] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0099] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (85:15);

[0100] Detection wavelength: 225nm;

[0101] Column temperature: 25°C;

[0102] Flow rate: 1.0 mL / min;

[0103] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0104] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0105] Example 8

[0106] Chromatographic conditions and system suitability experiments

[0107] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0108] Mobile phase: 0.01 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0109] Detection wavelength: 225nm;

[0110] Column temperature: 25°C;

[0111] Flow rate: 1.0 mL / min;

[0112] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0113] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0114] Example 9

[0115] Chromatographic conditions and system suitability experiments

[0116] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0117] Mobile phase: 0.1 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0118] Detection wavelength: 225nm;

[0119] Column temperature: 25°C;

[0120] Flow rate: 1.0 mL / min;

[0121] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0122] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0123] Example 10

[0124] Chromatographic conditions and system suitability experiments

[0125] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0126] Mobile phase: 0.05 mol / L sodium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0127] Detection wavelength: 225nm;

[0128] Column temperature: 25°C;

[0129] Flow rate: 1.0 mL / min;

[0130] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0131] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0132] Embodiment 11

[0133] Chromatographic conditions and system suitability experiments

[0134] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0135] Mobile phase: 0.05 mol / L ammonium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0136] Detection wavelength: 225nm;

[0137] Column temperature: 25°C;

[0138] Flow rate: 1.0 mL / min;

[0139] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0140] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0141] Example 12

[0142] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0143] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0144] Detection wavelength: 225nm;

[0145] Column temperature: 25°C;

[0146] Flow rate: 1.0 mL / min;

[0147] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0148] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0149] Example 13

[0150] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0151] Mobile phase: 0.05 mol / L sodium hydrogen phosphate aqueous solution: acetonitrile (80:20);

[0152] Detection wavelength: 225nm;

[0153] Column temperature: 25°C;

[0154] Flow rate: 1.0 mL / min;

[0155] Preparation of system suitability solution: Take appropriate amounts of thiamphenicol and related impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0156] Preparation of test solution: Weigh about 50 mg of the test sample into a 50 mL volumetric flask, accurately weigh, ultrasonically dissolve and dilute to the scale with mobile phase.

[0157] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0158] Embodiment 14

[0159] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0160] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0161] Detection wavelength: 225nm;

[0162] Column temperature: 25°C;

[0163] Flow rate: 1.0 mL / min;

[0164] Preparation of system suitability solution: Take appropriate amount of thiamphenicol enteric-coated tablets and relevant impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0165] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0166] Embodiment 15

[0167] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0168] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0169] Detection wavelength: 225nm;

[0170] Column temperature: 25°C;

[0171] Flow rate: 1.0 mL / min;

[0172] Preparation of system suitability solution: Take appropriate amount of thiamphenicol capsules and relevant impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0173] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0174] Example 16

[0175] Chromatographic column: chromatographic column with octadecyl bonded silica gel as filler;

[0176] Mobile phase: 0.05 mol / L potassium dihydrogen phosphate aqueous solution: acetonitrile (80:20);

[0177] Detection wavelength: 225nm;

[0178] Column temperature: 25°C;

[0179] Flow rate: 1.0 mL / min;

[0180] Preparation of system suitability solution: Take appropriate amount of thiamphenicol granules and relevant impurities, weigh accurately, and use mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL.

[0181] Determination method: Accurately measure 10 μL of each of the above solutions, inject them into the liquid chromatograph respectively, and record the chromatogram.

[0182] The test results showed that the separation between thiamphenicol and its related impurities in the system suitability solution was greater than 1.5, the blank solution did not interfere with the detection, and the method had good specificity.

[0183] The system suitability solutions were tested according to the above 16 examples, and the separation degree, theoretical plate number, and tailing factor were recorded. The results are shown in Table 1.

[0184] Table 1 Test results of Examples 1-16 - Degree of separation

[0185]

[0186]

[0187] The above results indicate that the above Examples 1-16 can all achieve the separation and detection effect of the detection method of the present invention.

[0188] The detection method of the present invention uses octadecyl bonded silica gel as a filler chromatographic column and phosphate aqueous solution-acetonitrile as a mobile phase; thiamphenicol or its preparation related substances are separated and determined in a reverse phase chromatography system. The detection method can effectively separate and determine thiamphenicol or its preparation related substances, and has the advantages of good separation, high theoretical plate number, reliable accuracy, good repeatability, etc.

[0189] Comparative Example 1

[0190] According to the method of the 2020 edition of the Chinese Pharmacopoeia, take appropriate amounts of thiamphenicol and related impurities, weigh them accurately, and use the mobile phase to make a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL. The mobile phase is water-acetonitrile (4:1).

[0191] Test results such as Figure 4 As shown in the liquid chromatogram, it can be seen that this method cannot detect impurities 3 and 4, and the peak shapes of other impurities are poor, which cannot meet the requirements of the detection of thiamphenicol or related substances in its preparations.

[0192] Comparative Example 2

[0193] According to the method of patent CN 117686628 A, appropriate amounts of thiamphenicol and related impurities were taken, accurately weighed, and prepared into a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL using a mobile phase. The mobile phase was a phosphoric acid aqueous solution-acetonitrile gradient elution, and the gradient program was as follows:

[0194]

[0195] Test results such as Figure 5 , 6 As shown in the liquid chromatogram, it can be seen that impurity 4 and impurity 5 overlap, and impurity 1 has a poor peak shape and is interfered by the blank. This shows that this method cannot meet the requirements for the detection of thiamphenicol or related substances in its preparations.

[0196] Comparative Example 3

[0197] According to the patent method of CN 106353437 A, appropriate amounts of thiamphenicol and related impurities were taken, accurately weighed, and prepared into a solution containing 1 mg of thiamphenicol, about 10 μg of impurities 1, 3, 4, 5, and 6, and about 4 μg of impurity 2 per 1 mL using a mobile phase. The mobile phase was methanol-0.05 mol / L potassium dihydrogen phosphate solution-triethylamine (20:80:0.05), and after mixing, the pH was adjusted to 3.0 with phosphoric acid.

[0198] Test results such as Figure 7 , 8 As shown in the liquid chromatogram, it can be seen that impurity 1 and impurity 3 have poor peak shapes and are both interfered by the blank, and impurity 6 is a double peak. This shows that this method cannot meet the detection of thiamphenicol or related substances in its preparations.

[0199] It can be seen that compared with the existing reported thiamphenicol impurity detection method, the detection method of the present invention has the following advantages: ① It realizes the separation of thiamphenicol and various related impurities; ② It eliminates the interference of blank solvent peak with impurity detection; ③ It effectively prevents the missed detection of impurities.

[0200] The embodiments provided above are not intended to limit the scope of the present invention, and the steps described are not intended to limit the execution order thereof. Those skilled in the art may make obvious improvements to the present invention in combination with existing common knowledge, which also fall within the scope of protection defined by the claims of the present invention.

Claims

1. A method for detecting a substance related to thiamphenicol or its preparation, characterized in that: The method comprises the following steps: Preparation of test solution: Take an appropriate amount of thiamphenicol or its preparation to prepare the test solution; Chromatographic conditions: Octadecyl bonded silica gel was used as the filler column, and phosphate aqueous solution-acetonitrile was used as the mobile phase; Determination method: Take the test sample solution and inject it into the liquid chromatograph for determination.

2. The detection method according to claim 1, characterized in that: The thiamphenicol or its preparation-related substances include the following structure:

3. The detection method according to claim 1, characterized in that: The preparation method of the test solution is: dissolving thiamphenicol or its preparation with a mobile phase and diluting it to obtain a test solution with a concentration of 0.1 to 3 mg / mL.

4. The detection method according to claim 1, characterized in that: The mobile phase is a 70-90:30-10 phosphate aqueous solution:acetonitrile solution.

5. The detection method according to claim 1, characterized in that: The mobile phase is a phosphate aqueous solution: acetonitrile solution in a ratio of 80:

20.

6. The detection method according to claim 1, characterized in that: The concentration of the phosphate aqueous solution is 0.001 mol / L-0.1 mol / L.

7. The detection method according to claim 1, characterized in that: The phosphate is dihydrogen phosphate or distear hydrogen phosphate.

8. The detection method according to claim 1, characterized in that: The mobile phase is 80:20 0.05 mol / L potassium dihydrogen phosphate solution: acetonitrile solution.

9. The detection method according to any one of claims 1 to 8, characterized in that: The chromatographic conditions are: detection wavelength of 200-240 nm, column temperature of 20-45° C., flow rate of 0.5-2 mL / min, and injection volume of 1-30 μL.

10. The detection method according to any one of claims 1 to 8, characterized in that: The chromatographic conditions are as follows: detection wavelength of 225 nm, column temperature of 25° C., flow rate of 1.0 mL / min, and injection volume of 10 μL.

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