Method for detecting residual solvent of dibekacin

Abstract

The invention discloses a method for detecting a residual solvent of dibekacin. The method comprises the following steps: respectively preparing a blank solution, a reference solution and a test solution; chromatographic conditions are as follows: an instrument used is a gas chromatograph with a hydrogen flame ionization detector, a chromatographic column is a capillary column taking polydimethylsiloxane as stationary liquid, and headspace sample injection is adopted to inject into the gas chromatograph; respectively carrying out sample injection detection to obtain a chromatogram of the reference substance solution and a chromatogram of the test solution, carrying out detection analysis by adopting an external standard method, and calculating the residual quantity of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile in the dibekacin. According to the method, chromatographic detection conditions are optimized, and a plurality of residual solvents are placed in the same system for detection under the condition of ensuring good theoretical plate and separation degree, so that the detection efficiency is greatly improved. According to the detection conditions of the method, ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile are completely separated, and the linear relationship is good in the investigated range.

Description

Technical Field

[0001] The invention relates to the technical field of drug analysis, and in particular to a method for detecting residual solvents in dibekacin. Background Art

[0002] Dibekacin is a semisynthetic aminoglycoside antibiotic with a similar structure to gentamicin. Its chemical name is (2R, 3R, 4S, 5S, 6R)-4-amino-2-[(1S, 2S, 3R, 4S, 6R)-4,6-diamino-3-[(2R, 3R, 6S)-3-amino-6-(aminomethyl)oxacyclohexane-2-yl]oxy-2-hydroxycyclohexyl]oxy-6-(hydroxymethyl)oxane-3,5-diol. Its structural formula is as follows: .

[0003] Residual solvents in drugs refer to organic volatile compounds produced or used in the production of raw materials or excipients, as well as in the preparation of preparations. Since residual solvents not only have no therapeutic effect, but may also increase the toxic side effects of drugs and affect the stability of drugs, organic solvents should be removed as much as possible. In order to protect patients from the harm of residual organic solvents in drugs, it is necessary to accurately measure the residual amount of organic solvents introduced during the drug production process.

[0004] Chinese patent document CN 111704636A (application number 202010236161.6) discloses a method for preparing an aminoglycoside drug intermediate, wherein the intermediate is 1,3,2',6',3''-penta-N-p-toluenesulfonyl kanamycin B, which is a key intermediate in the synthesis of 3',4'-dideoxy-kanamycin B, i.e., dibekacin. The preparation method comprises the following steps: adding kanamycin B, an acid binding agent, and a mixed solvent into a reactor, adding p-toluenesulfonyl chloride at 0 to 30°C, and stirring the reaction at 0 to 30°C for 8 to 18 hours; after the reaction, adding an inert solvent, stirring and crystallizing, filtering, and drying to obtain 1,3,2',6',3''-penta-N-p-toluenesulfonyl kanamycin B.

[0005] The mixed solvent is a solvent system composed of three or more solvents, including water and alcohol, and one or more of dichloromethane, ethyl acetate, and chloroform; the inert solvent is a combination of one or more of hexane, heptane, ethyl ether, and methyl tert-butyl ether; the alcohol constituting the mixed solvent is methanol, ethanol, or isopropanol.

[0006] In the above-mentioned preparation process of the dibekacin intermediate, organic solvents such as ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile are involved; these organic solvents may be brought into dibekacin.

[0007] According to the requirements of the ICH guidelines: the residual limits of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile are 0.5%, 0.5%, 0.06% and 0.041% respectively. Therefore, the determination of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile in dibekacin is of great significance to the drug quality of dibekacin and the health of patients.

[0008] Currently, there is no record of the detection method for the content of residual solvents in dibekacin, and it is necessary to establish a detection method with high sensitivity and accuracy. Summary of the invention

[0009] The technical problem to be solved by the present invention is to provide a method for detecting residual solvent of dibekacin with high sensitivity and accuracy.

[0010] The technical solution for achieving the purpose of the present invention is a method for detecting residual solvent of dibekacin, comprising the following steps: ① Prepare blank solution, reference solution and test solution respectively.

[0011] The blank solution is a diluent, and the diluent is a mixed solution of water and N,N-dimethylformamide, or N-methylpyrrolidone.

[0012] The reference substance solution consists of ethanol, methyl tert-butyl ether, dichloromethane, acetonitrile and a diluent.

[0013] The test solution is obtained by dissolving the dibekacin to be tested in a diluent.

[0014] ② Sample injection test.

[0015] The chromatographic conditions are as follows: the instrument used is a gas chromatograph with a hydrogen flame ionization detector, the chromatographic column is a capillary column with polydimethylsiloxane as the stationary liquid, the gas chromatograph is injected by headspace injection, the carrier gas is nitrogen, the injection port temperature is 200±10℃, the column temperature is set to rise in gradient, and the temperature of the hydrogen flame ionization detector is 250℃~260℃.

[0016] The chromatograms of the reference solution and the test solution were obtained by injection testing respectively, and the external standard method was used for detection and analysis to calculate the residual amounts of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile in dibekacin.

[0017] Optionally, when the diluent is a mixed solution of water and N,N-dimethylformamide, the volume ratio of water:N,N-dimethylformamide is 3:2.

[0018] In the above step ②, the carrier gas flow rate is 1.5 mL / min.

[0019] In the above step ②, the split ratio of the split direct injection mode is 2:1 to 5:1.

[0020] Optionally, in the above step ②, when the column temperature is increased in a gradient manner, the starting temperature is 40°C, maintained for 5 to 15 minutes, and then increased to 200 to 240°C at a rate of 20°C per minute and maintained for 5 to 6 minutes.

[0021] Optionally, in the above step ②, the injection order is 1 injection of blank solution, 5 injections of reference solution, and 2 injections of test solution.

[0022] Optionally, the chromatographic column is Agilent DB-624, Agilent HP-1, Agilent DB-1 or other similar chromatographic columns.

[0023] Optionally, the gas chromatography column has a specification of 30 m*0.53 mm, 1 μm; 30 m*0.53 mm, 3 μm; or 60 m*0.32 mm, 5 μm.

[0024] The above detection method can simultaneously detect the residual amounts of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile.

[0025] The present invention has positive effects: (1) The present invention optimizes the chromatographic detection conditions. While ensuring that the theoretical plates and separation are good, multiple residual solvents are placed in the same system for detection, which greatly improves the detection efficiency.

[0026] (2) According to the detection conditions of the present invention, ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile were completely separated and had a good linear relationship within the range investigated.

[0027] The sensitivity and accuracy of the method of the invention both meet the detection requirements for residual organic solvents, and the method is suitable for detecting residual solvents such as ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile in dibekacin raw materials. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 It is the gas chromatogram of the blank solution in Example 1.

[0029] Figure 2 It is the gas chromatogram of the reference substance solution in Example 1.

[0030] Figure 3 It is the gas chromatogram of the test solution in Example 1.

[0031] Figure 4 It is the gas chromatogram of the blank solution in Example 2.

[0032] Figure 5It is the gas chromatogram of the reference substance solution in Example 2.

[0033] Figure 6 It is the gas chromatogram of the test solution in Example 2. DETAILED DESCRIPTION

[0034] (Example 1) The detection method of dibekacin residual solvent of the present embodiment comprises the following steps: ① Prepare blank solution, control stock solution, reference solution and test solution respectively.

[0035] The blank solution is a diluent, and the diluent is water:N,N-dimethylformamide in a volume ratio of 3:2.

[0036] The preparation method of the control stock solution is as follows: accurately weigh 201.31 mg of acetonitrile and 301.08 mg of dichloromethane in a 50 mL volumetric flask, add N,N-dimethylformamide to the scale, shake well, accurately measure 1 mL, add 51.91 mg of methyl tert-butyl ether and 508.69 mg of ethanol to a 100 mL volumetric flask, dilute to the scale with N,N-dimethylformamide, mix well, and use as the control stock solution.

[0037] The preparation method of the reference solution is as follows: accurately measure 2 mL of the reference stock solution into a headspace bottle, then accurately measure 3 mL of water, seal with a cap, shake well, and prepare five parallel portions as the reference solution.

[0038] The preparation method of the test solution is as follows: accurately weigh 202.04 mg of dibekacin raw material into a headspace bottle, add 3 ml of water to dissolve it, then accurately measure 2 mL of N,N-dimethylformamide, cover and seal, shake well, and use this as the test solution.

[0039] ② Sample injection test.

[0040] The chromatographic conditions were as follows: The instrument used is a gas chromatograph with a hydrogen flame ionization detector, which in this embodiment is an Agilent-7890B gas chromatograph.

[0041] The chromatographic column is a capillary column with polydimethylsiloxane as the stationary liquid, and the model can be selected from Agilent DB-624, Agilent HP-1, Agilent DB -1 or other similar chromatographic columns, and the gas chromatographic column specifications are 30m*0.53mm, 1μm; 30m*0.53mm, 3μm; or 60m*0.32mm, 5μm. In this embodiment, it is Agilent DB-624, with specifications of 30m*0.53mm, 3μm.

[0042] The injector was Agilent-7697A headspace injector.

[0043] The carrier gas was nitrogen with a flow rate of 1.5 mL / min. The split ratio of the split direct injection mode was 2:1 to 5:1.

[0044] Headspace injection was used, the injection port temperature was 200±10°C (200°C in this embodiment), the column temperature was set to a gradient temperature increase, the starting temperature was 40°C, maintained for 15 minutes, and then increased to 240°C at a rate of 20°C per minute and maintained for 6 minutes; the detector temperature was 260°C, and the injection volume was 1 mL.

[0045] Accurately pipette 1 mL of blank solution, reference solution, and test solution, and inject into the gas chromatograph, with 1 injection of blank solution, 5 injections of reference solution, and 2 injections of test solution, for determination, and record the chromatogram. According to the chromatogram, calculate the residual amounts of ethanol, methyl tert-butyl ether, dichloromethane, and acetonitrile in dibekacin by external standard method.

[0046] Figure 1 is the gas chromatogram of the blank solution in Example 1, Figure 2 is the gas chromatogram of the reference substance solution in Example 1, Figure 3 It is the gas chromatogram of the test solution in Example 1.

[0047] Depend on Figure 1 It can be seen that the blank solution does not interfere with the detection of the solvent peak.

[0048] Depend on Figure 2 It can be seen that in the reference solution, the separation degree of the solvent peak is greater than 1.5, achieving complete separation and meeting the requirements. The peak shape is good, the peak purity is good, and the theoretical plate number is greater than 3000, which meets the standard.

[0049] Depend on Figure 3 It can be seen that methyl tert-butyl ether and dichloromethane were found in the test sample, and the contents were 0.02% and 0.0003% calculated by external standard method, which are lower than the limits specified in the ICH guidelines and meet the requirements. It also shows that the method of the present invention is a detection method with high sensitivity and accuracy.

[0050] (Example 2) The detection method of dibekacin residual solvent of the present embodiment comprises the following steps: ① Prepare blank solution, control stock solution, reference solution and test solution respectively.

[0051] The blank solution is a diluent, and the diluent is N-methylpyrrolidone.

[0052] The preparation method of the control stock solution is as follows: accurately weigh 392.0 mg of acetonitrile and 127.2 mg of dichloromethane in a 50 mL volumetric flask, add N-methylpyrrolidone to the scale, shake well, accurately measure 5 mL, add 499.7 mg of methyl tert-butyl ether and 5000.3 mg of ethanol to a 100 mL volumetric flask, dilute to the scale with N-methylpyrrolidone, mix well, and use as the control stock solution.

[0053] The preparation method of the reference solution is as follows: accurately measure 5 mL of the reference stock solution into a 25 mL volumetric flask and make up to volume with N-methylpyrrolidone; measure 1 mL and add it to the headspace bottle, then add 4 mL of water, cover and seal, mix well, and make five parallel copies as the reference solution.

[0054] The preparation method of the test solution is as follows: accurately weigh 206.0 mg of dibekacin raw material into a headspace bottle, add 4 mL of water, ultrasonically dissolve, and seal to use as the test solution.

[0055] ② Sample injection test.

[0056] The chromatographic conditions were as follows: The instrument used was Agilent-7890B gas chromatograph.

[0057] The chromatographic column is Agilent DB-1, with specifications of 30m*0.53mm, 1μm.

[0058] The injector was Agilent-7697A headspace injector.

[0059] The carrier gas was nitrogen with a flow rate of 1.5 mL / min. The split ratio of the split direct injection mode was 2:1 to 5:1.

[0060] Headspace injection was used with an injection port temperature of 200°C. The column temperature was set to a gradient temperature increase, with the starting temperature at 40°C, maintained for 5 minutes, then increased to 200°C at a rate of 40°C per minute and maintained for 5 minutes; the detector temperature was 250°C and the injection volume was 1 mL.

[0061] During the injection test, 1 mL of blank solution, reference solution, and test solution were accurately drawn and injected into the gas chromatograph, with 1 injection of blank solution, 5 injections of reference solution, and 2 injections of test solution. The chromatogram was recorded. Based on the chromatogram, the residual amounts of ethanol, methyl tert-butyl ether, dichloromethane, and acetonitrile in dibekacin were calculated using the external standard method.

[0062] Depend on Figure 4 It can be seen that the blank solution does not interfere with the detection of the solvent peak.

[0063] Depend on Figure 5It can be seen that in the reference solution, the separation degree of the solvent peak is greater than 1.5, achieving complete separation and meeting the requirements. The peak shape is good, the peak purity is good, and the theoretical plate number is greater than 3000, which meets the standard.

[0064] Depend on Figure 6 It can be seen that the sample was found to contain methyl tert-butyl ether and dichloromethane, and the content calculated by external standard method was 0.024% and 0.0005%, which is lower than the limit specified in the ICH guidelines and meets the requirements.

Claims

1. A method for detecting residual solvent of dibekacin, characterized in that The following steps are involved: ① Prepare blank solution, reference solution and test solution respectively; The blank solution is a diluent, and the diluent is a mixed solution of water and N,N-dimethylformamide, or N-methylpyrrolidone; The reference substance solution is composed of ethanol, methyl tert-butyl ether, dichloromethane, acetonitrile and a diluent; The test solution is obtained by dissolving the dibekacin to be tested in a diluent; ② Sampling test; The chromatographic conditions were as follows: the instrument used was a gas chromatograph with a hydrogen flame ionization detector, the chromatographic column was a capillary column with polydimethylsiloxane as the stationary liquid, headspace injection was used for injection into the gas chromatograph, the carrier gas was nitrogen, the injection port temperature was 200±10°C, the column temperature was set to a gradient temperature increase, and the hydrogen flame ionization detector temperature was 250°C to 260°C; The chromatograms of the reference solution and the test solution were obtained by injection testing respectively, and the external standard method was used for detection and analysis to calculate the residual amounts of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile in dibekacin.

2. The method for detecting residual solvent of dibekacin according to claim 1, characterized in that: When the diluent is a mixed solution of water and N,N-dimethylformamide, the volume ratio of water:N,N-dimethylformamide is 3:

2.

3. The method for detecting residual solvent of dibekacin according to claim 1, characterized in that: In step ②, the carrier gas flow rate is 1.5 mL / min.

4. The method for detecting residual solvent of dibekacin according to claim 3, characterized in that: In step ②, the split ratio of the split direct injection mode is 2:1 to 5:

1.

5. The method for detecting residual solvent of dibekacin according to claim 1, characterized in that: In step ②, when the column temperature is increased in a gradient manner, the starting temperature is 40°C, maintained for 5 to 15 minutes, and then increased to 200 to 240°C at a rate of 20°C per minute, and maintained for 5 to 6 minutes.

6. The method for detecting residual solvent of dibekacin according to claim 1, characterized in that: In step ②, the injection order is 1 injection of blank solution, 5 injections of reference solution, and 2 injections of test solution.

7. The method for detecting residual solvent of dibekacin according to claim 1, characterized in that: The chromatographic column is Agilent DB-624, Agilent HP-1, Agilent DB-1 or other chromatographic columns of the same type.

8. The method for detecting residual solvent of dibekacin according to claim 7, characterized in that: The gas chromatography column specifications are 30m*0.53mm, 1μm; 30m*0.53mm, 3μm; or 60m*0.32mm, 5μm.

9. The method for detecting residual solvent of dibekacin according to claim 1, characterized in that: The residual amounts of ethanol, methyl tert-butyl ether, dichloromethane and acetonitrile can be detected simultaneously.

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