CN116183741B - Method for detecting chloroacetyl chloride in fluconazole - Google Patents
Method for detecting chloroacetyl chloride in fluconazole Download PDFInfo
- Publication number
- CN116183741B CN116183741B CN202211542631.7A CN202211542631A CN116183741B CN 116183741 B CN116183741 B CN 116183741B CN 202211542631 A CN202211542631 A CN 202211542631A CN 116183741 B CN116183741 B CN 116183741B
- Authority
- CN
- China
- Prior art keywords
- mobile phase
- solution
- liquid chromatography
- fluconazole
- standard
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/04—Preparation or injection of sample to be analysed
- G01N30/06—Preparation
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/62—Detectors specially adapted therefor
- G01N30/72—Mass spectrometers
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N30/00—Investigating or analysing materials by separation into components using adsorption, absorption or similar phenomena or using ion-exchange, e.g. chromatography or field flow fractionation
- G01N30/02—Column chromatography
- G01N30/86—Signal analysis
- G01N30/8675—Evaluation, i.e. decoding of the signal into analytical information
- G01N30/8679—Target compound analysis, i.e. whereby a limited number of peaks is analysed
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Physics & Mathematics (AREA)
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Engineering & Computer Science (AREA)
- Library & Information Science (AREA)
- Investigating Or Analyzing Non-Biological Materials By The Use Of Chemical Means (AREA)
- Other Investigation Or Analysis Of Materials By Electrical Means (AREA)
Abstract
The application relates to the technical field of chemical detection, and particularly discloses a method for detecting chloroacetyl chloride in fluconazole, which comprises the following steps: preparing a solution: respectively preparing a standard solution and a sample solution; and (3) measuring: and detecting the standard solution and the sample solution by utilizing liquid chromatography-mass spectrometry, and drawing a standard curve of the standard solution to obtain the content of chloracetyl chloride in the sample solution. The detection method provided by the application has the advantages of simple operation method, high detection speed, high sensitivity, high precision, high repeatability and high accuracy.
Description
Technical Field
The application relates to the technical field of chemical detection, in particular to a method for detecting chloroacetyl chloride in fluconazole.
Background
As a novel triazole antifungal drug, fluconazole (CAS number 86386-73-4) is a broad-spectrum antifungal drug, and has the advantages of quick absorption, wide in-vivo distribution, long half-life, good tolerance, small adverse reaction and the like; in addition, fluconazole has better effect of treating fungal infection of human and animals, so the application of fluconazole is very wide.
As can be seen from the synthetic route of fluconazole, chloroacetyl chloride (CAS number 79-04-9) is the starting material for the reaction and is often present in fluconazole drugs. Chloroacetyl chloride carries a warning structural group, is a potentially genotoxic impurity, and may be genotoxic and carcinogenic. In order to ensure the safety and effectiveness of the fluconazole drug, it is necessary to detect the chloroacetyl chloride impurity in the fluconazole and further control the chloroacetyl chloride impurity in the fluconazole.
At present, no mature detection method is available for trace detection of chloroacetyl chloride in fluconazole. Therefore, it is necessary to establish a highly sensitive method for detecting chloroacetyl chloride impurity in fluconazole.
Disclosure of Invention
In order to detect trace impurity chloroacetyl chloride in fluconazole and improve the precision and accuracy of the detection method, the application provides a detection method of chloroacetyl chloride in fluconazole.
The application provides a method for detecting chloracetyl chloride in fluconazole, which adopts the following technical scheme:
the method for detecting chloracetyl chloride in fluconazole specifically comprises the following steps:
preparing a solution: respectively preparing a standard solution and a sample solution;
and (3) measuring: and detecting the standard solution and the sample solution by utilizing liquid chromatography-mass spectrometry, and drawing a standard curve of the standard solution to obtain the content of chloracetyl chloride in the sample solution.
By using the detection method, a linear regression equation is obtained by drawing a standard curve of a chloracetyl chloride standard solution; and substituting the test result of the test solution into a standard curve to quickly obtain the content of chloracetyl chloride in the test solution. The detection method has the advantages of high sensitivity, high repeatability, high accuracy and high precision.
The method adopts liquid chromatography-mass spectrometry (LC-MS) as a separation system of chloracetyl chloride and other substances in a sample to be detected, and mass spectrometry as a chloracetyl chloride detection system. The liquid chromatography combines the advantages of high separation capacity of liquid chromatography on a sample to be detected, high selectivity and high sensitivity with mass spectrum and capability of providing relative molecular mass and structural information.
The liquid chromatography has the characteristic of stronger separation degree for a sample to be detected, can effectively improve the separation effect of chloracetyl chloride and other substances by selecting proper chromatographic columns, column temperatures, mobile phases, elution procedures and other parameter conditions, not only can provide enough ions for mass spectrum test analysis, but also can primarily separate impurities interfering with mass spectrum, avoid the influence of interference impurities on subsequent mass spectrum, and further obtain good peak type parameters, thereby effectively improving the accuracy and precision of detection results.
Preferably, in the liquid chromatography-mass spectrometry: the column used for liquid chromatography was an Agilent InfinityLabPorosiwell 120SB-AQ column, 3.0X100 mm,2.7 μm.
The SB-Aq chromatographic column has hydrophilic surface, so that collapse of the stationary phase can be effectively prevented; the SB-Aq chromatographic column surface is porous core-shell filler particles, which can provide higher flux and higher separation degree. Therefore, the SB-Aq column is adopted to separate the sample to be detected, and good repeatability and excellent peak shape can be obtained.
Preferably, in the liquid chromatography-mass spectrometry: the test conditions for liquid chromatography were: the column temperature is 25-35 ℃, the sample injection volume is 15-25 mu L, the flow rate is 0.2-0.4mL/min, the mobile phase A is buffer solution, and the mobile phase B is organic solvent.
Preferably, in the liquid chromatography-mass spectrometry: separating the solution to be detected by liquid chromatography by adopting a mobile phase gradient elution method;
the mobile phase gradient elution method comprises the following steps: at 0-1.0min, mobile phase A is 95% and mobile phase B is 5%;1.0-4.0min, mobile phase A decreases linearly from 95% to 5%, mobile phase B increases linearly from 5% to 95%;4.0-6.0min, mobile phase A of 5% and mobile phase B of 95%; at 6.0-6.1min, mobile phase A increases linearly from 5% to 95%, mobile phase B decreases linearly from 95% to 5%; at 6.1-10.0min, mobile phase A is 95% and mobile phase B is 5%; mobile phase a and mobile phase B were both in volume percent.
Further, the mobile phase A is formic acid-water solution; the volume concentration of formic acid in the formic acid-water solution is 0.07-0.13%.
Further, the mobile phase B is acetonitrile.
According to the application, the formic acid-water solution is selected as the mobile phase A, and the volume concentration of the formic acid in the formic acid-water solution is controlled within the range, so that the sample to be measured can be maintained in a proper pH environment, the stability of the sample to be measured is improved, and the dissociation of the sample to be measured is effectively avoided.
Further, the mobile phase A is acetic acid-water solution; the acetic acid concentration by volume in the acetic acid-water solution is 0.09-0.15%.
Preferably, in the liquid chromatography-mass spectrometry: the mass spectrum test conditions are as follows: the ionization form is ESI, the polarity is negative, the ionization voltage is-4500V, the collision air pressure is 8psi, the atomization air pressure is 40psi, the auxiliary heating air pressure is 40psi, the air curtain air pressure is 20psi, and the drying air temperature is 280-320 ℃.
Preferably, in the liquid chromatography-mass spectrometry: the MRM scan parameters are: the parent ion was 93.0, the daughter ion was 35.0, and the residence time was 200ms.
Preferably, the preparation method of the standard solution comprises the following steps: respectively and precisely transferring 0.10mL, 0.25mL, 0.40mL, 0.50mL, 0.75mL and 1.00mL of chloracetyl chloride stock solution into different volumetric flasks, respectively adding acetonitrile to complement the volume to 1mL, diluting to scale with 0.01% ammonia water solution, and swirling for 1min to obtain a series of standard solutions with different concentrations.
In summary, the technical scheme of the application has the following effects:
the application provides a detection method with high detection speed and simple operation method by detecting chloracetyl chloride in fluconazole through a liquid chromatography-mass spectrometry technology.
The method for detecting the chloracetyl chloride in the fluconazole has the advantages of low detection limit, high precision, high repeatability and high accuracy by screening and optimizing the relevant test conditions of liquid chromatography and mass spectrum in the liquid chromatography-mass spectrometry.
Drawings
FIG. 1 is a standard graph of the standard solution of chloroacetyl chloride in example 1 of the present application.
Detailed Description
The application provides a method for detecting chloroacetyl chloride in fluconazole, which specifically comprises the following steps:
preparing a solution: respectively preparing a standard solution and a sample solution;
and (3) measuring: and detecting the standard solution and the sample solution by utilizing liquid chromatography-mass spectrometry, and drawing a standard curve of the standard solution to obtain the content of chloracetyl chloride in the sample solution.
The testing conditions of the liquid chromatography in the liquid chromatography-mass spectrometry are as follows: the chromatographic column is Agilent InfinityLabPorosiwell 120SB-AQ column, 3.0X100 mm,2.7 μm; the column temperature is 25-35 ℃, the sample injection volume is 15-25 mu L, the flow rate is 0.2-0.4mL/min, the mobile phase A is buffer solution, and the mobile phase B is organic solvent;
separating the solution to be detected by liquid chromatography by adopting a mobile phase gradient elution method; the procedure of the mobile phase gradient elution method is as follows: at 0-1.0min, mobile phase A is 95% and mobile phase B is 5%;1.0-4.0min, mobile phase A decreases linearly from 95% to 5%, mobile phase B increases linearly from 5% to 95%;4.0-6.0min, mobile phase A of 5% and mobile phase B of 95%; at 6.0-6.1min, mobile phase A increases linearly from 5% to 95%, mobile phase B decreases linearly from 95% to 5%; at 6.1-10.0min, mobile phase A is 95% and mobile phase B is 5%; mobile phase a and mobile phase B were both in volume percent.
Further, mobile phase a is a formic acid-water solution; the volume concentration of formic acid in the formic acid-water solution is 0.07-0.13%; mobile phase B was acetonitrile.
Further, mobile phase a may also be an acetic acid-water solution; the volume concentration of acetic acid in the acetic acid-water solution is 0.09-0.15%.
The mass spectrum test conditions in the liquid chromatography-mass spectrometry are as follows: the ionization form is ESI, the polarity is negative, the ionization voltage is-4500V, the collision air pressure is 8psi, the atomization air pressure is 40psi, the auxiliary heating air pressure is 40psi, the air curtain air pressure is 20psi, and the drying air temperature is 280-320 ℃.
Specifically, the MRM scanning parameters in the liquid chromatography-mass spectrometry are as follows: the parent ion was 93.0, the daughter ion was 35.0, and the residence time was 200ms.
Further, the preparation method of the standard solution comprises the following steps: respectively and precisely transferring 0.10mL, 0.25mL, 0.40mL, 0.50mL, 0.75mL and 1.00mL of chloracetyl chloride stock solution into different volumetric flasks, respectively adding acetonitrile to complement the volume to 1mL, diluting to scale with 0.01% ammonia water solution, and swirling for 1min to obtain a series of standard solutions with different concentrations.
In the application, the used instrument liquid chromatograph-tandem mass spectrometer is a liquid chromatograph-tandem mass spectrometer, and the model is ABSCIEX TripleQuad5500+ (ABSCIEX of manufacturer); the fluconazole materials and sources thereof are shown in table 1; the remaining materials, reagents, solvents, and the like are commercially available.
Wherein, the chloracetyl chloride in the fluconazole has a warning structure, and the limit is 3.75ppm by referring to the ICH guiding principle M7 part.
Table 1 raw materials and sources thereof used in the examples
The application is described in further detail below in connection with examples and performance testing experiments, which are not to be construed as limiting the scope of the application as claimed.
Examples
Example 1
Example 1 provides a standard curve for chloroacetyl chloride standard solution.
The specific steps of this embodiment are as follows:
s1: preparing standard solution
Taking a chloracetyl chloride stock solution containing chloracetyl chloride 151.2894ng/mL as a standard solution, respectively and precisely transferring 0.10mL, 0.25mL, 0.40mL, 0.50mL, 0.75mL and 1.00mL into different volumetric flasks, respectively adding acetonitrile to complement the volume to 1mL, diluting with 0.01% ammonia water solution to scale, and swirling for 1min to obtain a series of standard solutions with different concentrations, wherein the concentrations of the standard solutions are respectively 1.51ng/mL, 3.78ng/mL, 6.05ng/mL, 7.56ng/mL, 11.35ng/mL and 15.13ng/mL.
S2: test conditions for LC-MS
S2.1: liquid chromatography test conditions
The chromatographic column is Agilent InfinityLab Poroshell SB-AQ column (3.0X100 mm,2.7 μm), and the number of the chromatographic column is GB-L-20-09-121; the column temperature is 30 ℃; the sample injection volume is 20 mu L; mobile phase a is 0.1% formic acid-water solution, mobile phase B is acetonitrile; the flow rate is 0.30mL/min; eluting the sample solution to be detected by adopting a mobile phase gradient elution method;
the mobile phase gradient elution conditions are specifically as follows: at 0-1.0min, mobile phase A is 95% and mobile phase B is 5%;1.0-4.0min, mobile phase A decreases linearly from 95% to 5%, mobile phase B increases linearly from 5% to 95%;4.0-6.0min, mobile phase A of 5% and mobile phase B of 95%; at 6.0-6.1min, mobile phase A increases linearly from 5% to 95%, mobile phase B decreases linearly from 95% to 5%; at 6.1-10.0min, mobile phase A is 95% and mobile phase B is 5%; mobile phase a and mobile phase B were both in volume percent.
S2.2: mass spectrometry conditions
The mass spectrum test conditions are as follows: the ionization form is ESI, the polarity is negative, the ionization voltage is-4500V, the collision air pressure is 8psi, the atomization air pressure is 40psi, the auxiliary heating air pressure is 40psi, the air curtain air pressure is 20psi, and the drying air temperature is 280-320 ℃.
S2.3: mass Spectrometry Multi-reaction monitoring (MRM) scanning parameters for chloroacetyl chloride
The MRM scan parameters for chloroacetyl chloride are shown in table 2.
TABLE 2 MRM scanning parameters of chloroacetyl chloride
S3: measurement results and analysis
And (3) testing a series of standard solutions with different concentrations in the step (S2) by using the test conditions of the LC-MS in the step (S3), and drawing a standard curve according to the test result.
S3.1: standard curve and regression equation
Regression analysis is carried out on the corresponding concentration of the chloroacetyl chloride according to the response value of the chloroacetyl chloride in the standard solution, and a standard curve is obtained as shown in figure 1.
As can be seen from fig. 1, the regression equation of the standard curve of chloroacetyl chloride is y= 1609.53064x-459.40944, and the correlation coefficient is r= 0.99234, which indicates that the concentration of chloroacetyl chloride and the peak area have a good linear relationship, and the standard curve of chloroacetyl chloride provided by the application can be used for quantitatively detecting the content of chloroacetyl chloride in fluconazole.
S3.2: detection limit of standard curve
The detection limit is set by detecting a response signal to noise ratio of about 3:1, i.e., the detection limit of chloroacetyl chloride obtained at a 3-fold signal to noise ratio (S/N).
The detection Limit (LOD) of the chloracetyl chloride detection method is 0.76ng/mL (0.38 ppm or 0.38 ug/g), which shows that the detection method has higher sensitivity.
S3.3: quantitative limit of standard curve
The limit of quantification is determined by detecting a response signal to noise ratio of about 10:1, i.e. the quantification limit of chloroacetyl chloride obtained at a 10-fold signal to noise ratio (S/N). Then, 6 parts of a standard solution having a concentration of a quantitative limit was prepared and tested, and the Relative Standard Deviation (RSD) of the peak area was calculated to examine the reproducibility of the quantitative limit.
Detection result: as shown in table 3.
TABLE 3 quantitative limited reproducibility assay results
From the detection results of Table 3, it was found that the limit of quantitation (LOQ) of the chloroacetyl chloride detection method was 1.51ng/mL; and the RSD of the peak area of the chloracetyl chloride in the 6 parts of quantitative limiting solution is 7.4%, which shows that the quantitative limiting solution in the chloracetyl chloride detection method provided by the application has good repeatability.
S3.4: repeatability and precision of standard curve
Preparing a test sample solution: respectively precisely weighing 20mg of fluconazole raw material to 0.01mg, placing into a 10mL volumetric flask, accurately adding 1mL of acetonitrile for vortex dissolution, then adding 0.01% ammonia water solution for dilution and volume fixing to scale, and vortex for 1min to obtain a sample solution. And (3) measuring the sample solution by using the test conditions of the LC-MS in the step S2.
Preparing a labeled sample solution with the concentration of 7.56ng/mL and 7.66ng/mL by using the sample solution and the standard solution provided in the step S1, detecting the labeled sample solution by using the LC-MS detection condition provided in the step S2, and calculating the labeled recovery rate of the labeled sample solution according to the labeled concentration and the detection concentration result of the labeled sample solution.
Comparing the detected concentration of the marked sample with the marked concentration, calculating the average recovery rate of the chloracetyl chloride, and examining the repeatability of the detection method. And (3) carrying out parallel measurement on each concentration level for 6 times to obtain the relative standard deviation, and examining the precision of the detection method.
Detection result: as shown in Table 4
TABLE 4 repeatability and precision test results
As can be seen from Table 4, the labeled sample solution with the concentration of 7.56ng/mL has a labeled recovery rate in the range of 102.4-109.4% and a relative standard deviation of 2.6%; the labeled sample solution with the concentration of 7.66ng/mL is tested, the labeled recovery rate is in the range of 93.9-105.1%, and the relative standard deviation is 4.3%. In conclusion, the standard adding recovery rate in the chloracetyl chloride detection method provided by the application is close to 100%, which shows that the chloracetyl chloride detection method provided by the application has higher repeatability; the relative standard deviation is less than 5.0%, which indicates that the method for detecting the chloracetyl chloride provided by the application has higher precision.
S3.5: accuracy of standard curve
Preparing a test sample solution: respectively precisely weighing 20mg of fluconazole raw material to 0.01mg, placing into a 10mL volumetric flask, accurately adding 1mL of acetonitrile for vortex dissolution, then adding 0.01% ammonia water solution for dilution and volume fixing to scale, and vortex for 1min to obtain a sample solution. And (3) measuring the sample solution by using the test conditions of the LC-MS in the step S2.
And (2) preparing a 50% limit concentration labeling test sample solution, a 100% limit concentration labeling test sample solution and a 150% limit concentration labeling test sample solution (the concentrations are respectively 3.78ng/mL, 7.56ng/mL and 11.35 ng/mL) by using the test sample solution and the standard solution provided in the step S1, detecting the labeling test sample solution by using the LC-MS detection conditions provided in the step S2, and calculating the labeling recovery rate of the labeling test sample solution according to the labeling concentration and the detection concentration result of the labeling test sample solution. ( And (3) injection: the limiting concentration of chloroacetyl chloride was 3.75ppm, i.e., 7.56ng/mL )
Comparing the detected concentration of the marked sample with the marked concentration, calculating the average recovery rate of the chloracetyl chloride, measuring each concentration level for 3 times in parallel, calculating the relative standard deviation, and examining the accuracy of the detection method.
Detection result: as shown in table 5.
TABLE 5 accuracy test results
As shown by the detection results in the table 5, the labeling recovery rate of the labeled sample solution is between 99.6 and 108.3 percent, and the labeling recovery rate is close to 100 percent, which indicates that the detection method of the chloracetyl chloride provided by the application has higher accuracy.
Example 2
The embodiment provides a method for detecting chloroacetyl chloride in fluconazole raw material.
According to the method for preparing the test sample solution provided in example 1, three batches (batch numbers are 11#, 12#, 13#) of 20mg (accurate to 0.01 mg) of fluconazole raw material are respectively precisely weighed, placed in a 10mL volumetric flask, accurately added with 1mL of acetonitrile for vortex dissolution, then added with 0.01% ammonia solution for dilution and volume fixing to scale, and vortex for 1min, thus obtaining the test sample solution.
Under the LC-MS detection condition provided in the embodiment 1, respectively and precisely measuring 20 mu L of the sample solution to be detected in a liquid chromatography-mass spectrometry, and then comparing the detection result with a standard curve to obtain the detection result of the chloroacetyl chloride impurity in the fluconazole raw material.
Detection result: as shown in table 6.
Example 3
The embodiment provides a method for detecting chloracetyl chloride in fluconazole sodium chloride injection.
And respectively precisely transferring 25mL of fluconazole sodium chloride injection in three batches (batch numbers are 21#, 22#, 23#) into a freeze-drying cup, freeze-drying, adding 50% acetonitrile for re-dissolution, and transferring to a 5mL volumetric flask for constant volume to obtain the sample solution.
Under the LC-MS detection condition provided in the embodiment 1, respectively precisely measuring 20 mu L of the sample solution to be detected in an injection mass spectrometer, and comparing the detection result with a standard curve to obtain the detection result of chloracetyl chloride impurity in the fluconazole sodium chloride injection.
Detection result: as shown in table 6.
Detection result
The detection results of chloracetyl chloride in the fluconazole materials and chloracetyl chloride in the fluconazole sodium chloride injection obtained in example 2 and example 3 are shown in table 6.
Table 6 results of detection of chloroacetyl chloride in fluconazole starting material and fluconazole sodium chloride injection
As can be seen from the above table, the method for detecting chloracetyl chloride in fluconazole detects chloracetyl chloride impurities in the commercial fluconazole raw material and the commercial fluconazole sodium chloride injection, and the detection result shows that the commercial fluconazole raw material and the commercial fluconazole sodium chloride injection do not contain chloracetyl chloride impurities.
Example 4
The embodiment provides a method for detecting chloroacetyl chloride in fluconazole.
Preparing a solution to be tested: the 150% limiting concentration labeled sample solution, 100% limiting concentration labeled sample solution, 50% limiting concentration labeled sample solution, quantitative limiting solution, and detection limiting solution (concentrations 11.35ng/mL, 7.56ng/mL, 3.78ng/mL, 1.51ng/mL, and 0.76ng/mL, respectively) in example 1 were taken as 5 test solutions.
(1-2) measurement: the above 5 parts of the solution to be measured were measured in parallel 3 times by using the measurement method and standard curve provided in example 1, and the recovery rate and relative standard deviation of the solution to be measured were calculated.
Detection result: as shown in table 7.
TABLE 7 detection results of the detection method in example 4
Comparative example
The comparative example provides a method for detecting chloroacetyl chloride in fluconazole.
This comparative example was tested for chloroacetyl chloride in fluconazole in reference application number 202110530984.4.
The specific method comprises the following steps:
(1) Preparing a control solution: about 150mg of chloroacetyl chloride control was precisely weighed, and a chloroacetyl chloride control solution having a concentration of 1.51. Mu.g/ml was prepared using methanol as a diluent.
Preparing a solution to be tested: respectively precisely weighing 20mg of fluconazole raw material to 0.01mg, placing into a 10mL volumetric flask, accurately adding 1mL of methanol, and vortex-dissolving to obtain a sample solution. And 5 parts of to-be-detected solutions with standard adding concentrations of 11.35ng/mL, 7.56ng/mL, 3.78ng/mL, 1.51ng/mL and 0.76ng/mL are prepared by using the chloracetyl chloride control solution.
(2) Measurement conditions for gas chromatography
The chromatographic column is filled with modified polyethylene glycol (ZebronzB-FFAP 30m×0.32mm,0.5 μm chromatographic column); column temperature: the mixture is kept at 60 ℃ for 20 minutes; sample inlet temperature: 200 ℃ detector temperature: 220 ℃; sample injection amount: 3 μl; carrier gas: n2; split ratio: 2:1 line rate: 50cm/sec.
(3) Measurement
After the system is stable, 1 needle of methanol solution, 5 needles of control solution and 1 needle of solution to be detected are added, and the chromatogram is recorded.
And detecting the 5 parts of the solution to be detected, respectively carrying out parallel measurement for 3 times, and calculating the recovery rate and the relative standard deviation of the solution to be detected.
The detection concentration of chloracetyl chloride in the solution to be detected=the peak area of chloracetyl chloride in the solution map to be detected/5 is aimed at the average peak area of chloracetyl chloride in the reference solution map multiplied by the concentration of chloracetyl chloride in the control solution.
Detection result: as shown in table 8.
Table 8 results of the test method in comparative example
When the detection method provided by the comparative example is used for detecting chloracetyl chloride in fluconazole by combining the detection results of the table 7 and the table 8, and when the concentration of chloracetyl chloride in the solution to be detected is 0.76ng/mL, the detection result is undetected; the detection limit of the detection method provided by the application is as low as 0.76ng/mL, and further proves that the detection method provided by the application has higher sensitivity.
When the concentration of chloracetyl chloride in the solution to be detected is the same level (11.35 ng/mL, 7.56ng/mL, 3.78ng/mL, 1.51 ng/mL), compared with the detection method provided by the comparison example 1, the recovery rate of the detection result obtained by the detection method is closer to 100%, and the relative standard deviation is smaller, so that the detection method provided by the application has higher accuracy and precision.
In conclusion, the method for detecting the chloracetyl chloride in the fluconazole has the advantages of high detection speed, simple operation method, low detection limit, good quantitative limit repeatability, high precision, high repeatability and high accuracy.
While the application has been described in detail in the foregoing general description and with reference to specific embodiments thereof, it will be apparent to one skilled in the art that modifications and improvements can be made thereto. Accordingly, such modifications or improvements may be made without departing from the spirit of the application and are intended to be within the scope of the application as claimed.
Claims (2)
1. The method for detecting chloracetyl chloride in fluconazole is characterized by comprising the following steps of:
preparing a solution: respectively preparing a standard solution and a sample solution;
and (3) measuring: detecting the standard solution and the sample solution by utilizing liquid chromatography-mass spectrometry, and drawing a standard curve of the standard solution to obtain the content of chloracetyl chloride in the sample solution;
in the liquid chromatography-mass spectrometry: the chromatographic column adopted by the liquid chromatography is Agilent InfinityLab Poroshell, SB-AQ column, 3.0X100 mm,2.7 μm; the test conditions for liquid chromatography were: the column temperature is 25-35 ℃, the sample injection volume is 15-25 mu L, the flow rate is 0.2-0.4mL/min, the mobile phase A is formic acid-water solution with the formic acid volume concentration of 0.07-0.13%, and the mobile phase B is acetonitrile;
the liquid chromatography adopts a mobile phase gradient elution method to separate a solution to be detected, and the mobile phase gradient elution method comprises the following procedures: at 0-1.0min, mobile phase A is 95% and mobile phase B is 5%;1.0-4.0min, mobile phase A decreases linearly from 95% to 5%, mobile phase B increases linearly from 5% to 95%;4.0-6.0min, mobile phase A of 5% and mobile phase B of 95%; at 6.0-6.1min, mobile phase A increases linearly from 5% to 95%, mobile phase B decreases linearly from 95% to 5%; at 6.1-10.0min, mobile phase A is 95% and mobile phase B is 5%; mobile phase a and mobile phase B are both in volume percent;
the mass spectrum test conditions are as follows: the ionization form is ESI, the polarity is negative, the ionization voltage is-4500V, the collision air pressure is 8psi, the atomization air pressure is 40psi, the auxiliary heating air pressure is 40psi, the air curtain air pressure is 20psi, and the drying air temperature is 280-320 ℃;
in the liquid chromatography-mass spectrometry: the MRM scan parameters are: the parent ion was 93.0, the daughter ion was 35.0, and the residence time was 200ms.
2. The method for detecting chloroacetyl chloride according to claim 1, wherein the standard solution is prepared by the following steps: respectively and precisely transferring 0.10mL, 0.25mL, 0.40mL, 0.50mL, 0.75mL and 1.00mL of chloracetyl chloride stock solution into different volumetric flasks, respectively adding acetonitrile to complement the volume to 1mL, diluting to scale with 0.01% ammonia water solution, and swirling for 1min to obtain a series of standard solutions with different concentrations.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211542631.7A CN116183741B (en) | 2022-12-03 | 2022-12-03 | Method for detecting chloroacetyl chloride in fluconazole |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202211542631.7A CN116183741B (en) | 2022-12-03 | 2022-12-03 | Method for detecting chloroacetyl chloride in fluconazole |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN116183741A CN116183741A (en) | 2023-05-30 |
| CN116183741B true CN116183741B (en) | 2023-08-29 |
Family
ID=86447803
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202211542631.7A Active CN116183741B (en) | 2022-12-03 | 2022-12-03 | Method for detecting chloroacetyl chloride in fluconazole |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN116183741B (en) |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2021047565A1 (en) * | 2019-09-11 | 2021-03-18 | 苏州同力生物医药有限公司 | Method for preparing (r)-praziquantel and chiral intermediate thereof, and composition |
| CN113219098A (en) * | 2021-05-18 | 2021-08-06 | 珠海润都制药股份有限公司 | Method for detecting chloroacetyl chloride in diclofenac sodium |
| CN115184531A (en) * | 2021-04-07 | 2022-10-14 | 浙江康恩贝制药股份有限公司 | Method for simultaneously determining contents of 3 impurities in tadalafil |
-
2022
- 2022-12-03 CN CN202211542631.7A patent/CN116183741B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2021047565A1 (en) * | 2019-09-11 | 2021-03-18 | 苏州同力生物医药有限公司 | Method for preparing (r)-praziquantel and chiral intermediate thereof, and composition |
| CN115184531A (en) * | 2021-04-07 | 2022-10-14 | 浙江康恩贝制药股份有限公司 | Method for simultaneously determining contents of 3 impurities in tadalafil |
| CN113219098A (en) * | 2021-05-18 | 2021-08-06 | 珠海润都制药股份有限公司 | Method for detecting chloroacetyl chloride in diclofenac sodium |
Non-Patent Citations (1)
| Title |
|---|
| Fully Automated Chemical Synthesis: Toward the Universal Synthesizer;Nathan Collins 等;Organic Process Research & Development;第24卷;全文 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN116183741A (en) | 2023-05-30 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN111766317B (en) | Method for measuring GS-441524 content in preparation by using polyethylene glycol and water | |
| CN114019062B (en) | Method for detecting related substances in rifampicin | |
| CN113295805B (en) | Method for detecting hydrazine hydrate in medicine | |
| CN109900830B (en) | Method for separating and determining sulfonamide impurities in celecoxib by adopting HPLC (high performance liquid chromatography) and application | |
| CN111965273B (en) | HPLC (high Performance liquid chromatography) method for detecting genotoxic impurities in candesartan cilexetil | |
| CN116183741B (en) | Method for detecting chloroacetyl chloride in fluconazole | |
| CN111208215B (en) | Method for detecting impurity 2-mercaptobenzothiazole in ceftriaxone sodium | |
| CN114166982B (en) | Method for simultaneously determining dimer, trimer and caprolactam in aminocaproic acid injection | |
| CN114264752B (en) | Detection method of methyl bromoacetate | |
| CN114324703B (en) | Method for simultaneously detecting multiple amino acids | |
| CN115616133A (en) | Method for detecting cysteine in compound amino acid injection and application thereof | |
| CN115774061A (en) | Method for detecting acetic acid in 1-cyclohexyl piperazine | |
| CN117074578B (en) | LC-MS/MS quantitative detection method of 2- (methylamino) -ethanol | |
| CN115015457B (en) | Method for analyzing bromoacetic acid impurity in selapage by high performance liquid chromatography | |
| CN114200067B (en) | High performance liquid chromatography analysis method for 6-bromo-3-hydroxy pyrazine-2-carboxamide and impurities | |
| CN114280191B (en) | Method for detecting related substances in double-cysteine and preparation thereof | |
| CN112526038B (en) | Carprofen and related substance detection method | |
| CN115128184B (en) | Method for determining thiourea content in pramipexole dihydrochloride raw material by using HPLC external standard method | |
| CN117630202A (en) | Method for detecting buspirone hydrochloride intermediate impurities | |
| CN116263436A (en) | Detection method of basic toxin impurities in milbegron | |
| CN116359371A (en) | Method for measuring urea content by high performance liquid chromatography | |
| CN115876926A (en) | Method for detecting BOC butyric acid in water and algae culture medium by using HPLC-MS/MS | |
| CN115711958A (en) | Method for detecting cyclohexanone in fluconazole sodium chloride injection | |
| CN116297921A (en) | Isomer detection method of tedizolid phosphate intermediate | |
| CN117110447A (en) | Method for measuring acid residues in benzothiopyrone compound by HPLC (high Performance liquid chromatography) |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |