A method for separating and detecting related substances in p-bromofluorobenzene

Abstract

The present invention relates to a method for separating and detecting related substances in p-bromofluorobenzene, and belongs to the field of analytical chemistry. In the method, gas chromatography separation technology is adopted, a chromatographic column with a strong polar stationary phase as filler is adopted during detection, a suitable solvent is selected to dissolve the sample according to the structure and physicochemical properties of the impurities to be analyzed, and a strong polar polyethylene glycol capillary chromatographic column and a detector are selected. The method of the present invention is simple and fast to operate, has good separation degree, and the result is accurate and reliable.

Description

Technical Field

[0001] The invention belongs to the field of analytical chemistry, and particularly relates to a method for separating and detecting related substances in p-bromofluorobenzene. Background Art

[0002] Escitalopram oxalate has the effect of enhancing the 5-hydroxytryptamine (5-HT) energy of the central nervous system. It is a selective serotonin reuptake inhibitor and is clinically used for the treatment of major depression and generalized anxiety disorder. p-Bromofluorobenzene is the starting material used in the synthesis of escitalopram. Due to the process technology, p-Bromofluorobenzene will inevitably produce some related substances, including fluorobenzene (CB), bromobenzene (BB), chlorobenzene (CB), p-Bromochlorobenzene (BCB), o-Bromofluorobenzene (2-BFB) and / or m-Bromofluorobenzene (3-BFB). These six related substances have similar structures and properties, and are difficult to separate and detect at the same time. If the related substances in the starting material p-Bromofluorobenzene cannot be accurately detected and analyzed to control them, it will most likely lead to the presence of multiple related substances in the final product escitalopram oxalate or the quality will be difficult to detect and control.

[0003] The adverse reactions of drugs in clinical use are not only related to the pharmacological activity of the drugs themselves, but sometimes also closely related to the related substances in the drugs. In order to ensure the quality and safety of drugs, the drugs that are registered for application require the research and effective control of the related substances introduced in the process. Therefore, it is of great significance for the safety of patients to study and control the related substances or related substances in p-bromofluorobenzene and make the quality of the final product escitalopram oxalate controllable.

[0004] At present, the United States Pharmacopoeia USP, the European Pharmacopoeia EP, the Japanese Pharmacopoeia JP and the Chinese Pharmacopoeia Ch.P. do not include the corresponding separation and detection methods, and there is no prior art to disclose the detection methods. In order to better and more accurately detect the related substances in the control product and ensure the quality of the drug, it is necessary to study a method for analyzing and detecting the related substances in the starting material of escitalopram, p-bromofluorobenzene. Summary of the invention

[0005] The object of the present invention is to provide a method for separating and detecting related substances in p-bromofluorobenzene by gas chromatography, so as to achieve the separation and detection of these related substances such as fluorobenzene (CB), bromobenzene (BB), chlorobenzene (CB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and / or m-bromofluorobenzene (3-BFB), thereby ensuring the quality and safety of medicines. The method provided by the present invention can effectively separate and determine the related substances in p-bromofluorobenzene, is simple and fast to operate, has good separation degree, and the result is accurate and reliable, and can be used for the control of related substances in medicines.

[0006] The method for separating and detecting related substances in p-bromofluorobenzene described in the present invention comprises: using gas chromatography separation technology, selecting a suitable solvent to dissolve a p-bromofluorobenzene sample and a related substance reference substance according to the structure and physicochemical properties of the related substances to be analyzed, preparing a related substance peak identification solution, a test sample solution and a test sample spiked solution, and selecting a capillary chromatographic column and a detector with a polyethylene glycol as the stationary phase.

[0007] The related substances include or are at least one of chlorobenzene, bromobenzene, fluorobenzene, p-bromochlorobenzene, o-bromofluorobenzene, and m-bromofluorobenzene. In some embodiments, the related substances include or are chlorobenzene, bromobenzene, fluorobenzene, p-bromochlorobenzene, o-bromofluorobenzene, and m-bromofluorobenzene.

[0008] The solvent comprises at least one selected from chloroform, n-propanol, butanone, acetonitrile and toluene. In some embodiments, the solvent is acetonitrile.

[0009] The polyethylene glycol capillary chromatographic column is selected from one of HP-INNOWAX, DB-FFAP, HP-20M, DB-WAX, and 007-20M. In some embodiments, the chromatographic column model is DB-WAX.

[0010] The detector is a hydrogen flame ionization detector.

[0011] The method for separating and detecting related substances in p-bromofluorobenzene may include the following steps or be implemented according to the following steps:

[0012] 1) Diluent / blank solution: acetonitrile;

[0013] 2) Test solution: Weigh the p-bromofluorobenzene sample and dissolve it in a diluent to prepare a p-bromofluorobenzene test solution;

[0014] 3) Peak identification stock solution: weigh the reference substances of the relevant substances, add diluent to dissolve, and prepare the peak identification stock solution;

[0015] 4) Peak identification solution: pipette the peak identification stock solution obtained in step 3) respectively, add diluent to dilute, and prepare peak identification solutions of each relevant substance;

[0016] 5) Test sample spike solution: Weigh the p-bromofluorobenzene sample, add the peak identification stock solution obtained in step 3) respectively, dilute with diluent, and prepare the test sample spike solution;

[0017] 6) Setting instrument parameters: injection port temperature 200℃-300℃, carrier gas flow rate 1.3mL / min-1.7mL / min, programmed temperature method, the temperature program is initial temperature 33-37℃, maintain 10min-15min, then increase the temperature to 80℃ at a rate of 10℃-25℃ per minute, maintain 5min-15min, and finally increase the temperature to 140℃ at a rate of 5℃-15℃ per minute, maintain 5min-15min, detector temperature 200℃-300℃, split ratio 6:1-10:1;

[0018] 7) Take blank solution, peak identification solution, test sample solution, and test sample spiked solution and perform gas chromatography analysis under the above conditions and record the chromatogram.

[0019] Preferably, the carrier gas used for gas chromatography separation is nitrogen, and the column flow rate is 1.5 mL / min.

[0020] Preferably, the temperature rising program of the gas chromatography separation is an initial temperature of 35°C, maintained for 12 minutes, then heated to 80°C at a rate of 20°C per minute, maintained for 10 minutes, and finally heated to 140°C at a rate of 10°C per minute, maintained for 10 minutes.

[0021] Preferably, the injection port temperature of the gas chromatography separation method is 250°C.

[0022] Preferably, the detector is a hydrogen flame ionization detector (FID), wherein the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min.

[0023] Preferably, the detector temperature of the gas chromatography separation method is 250°C.

[0024] Preferably, the split ratio of the gas chromatography separation method is 8:1.

[0025] Preferably, the chromatographic column used in the gas chromatography separation method is a DB-WAX capillary chromatographic column, which has an inner diameter of 0.32 mm, a length of 30 m, and a liquid film thickness of 0.5 μm.

[0026] In some embodiments, the method for separating and detecting related substances in p-bromofluorobenzene comprises the following steps:

[0027] 1) Diluent / blank solution: acetonitrile;

[0028] 2) Test solution: Weigh the p-bromofluorobenzene sample and dissolve it in a diluent to prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0029] 3) Peak identification stock solution: weigh each related substance reference substance, add diluent to dissolve, and prepare 1 mg / mL peak identification stock solution of each related substance;

[0030] 4) Peak identification solution: pipette the peak identification stock solution obtained in step 3), add diluent to dilute, and prepare 8 μg / mL chlorobenzene, 8 μg / mL bromobenzene, 8 μg / mL fluorobenzene, 8 μg / mL p-bromochlorobenzene, 8 μg / mL o-bromofluorobenzene, 32 μg / mL m-bromofluorobenzene peak identification solutions respectively;

[0031] 5) Test sample spike solution: Weigh the p-bromofluorobenzene sample, add the peak identification stock solutions obtained in step 3) respectively, dilute with diluent, and prepare a test sample spike solution containing 40 mg / mL p-bromofluorobenzene, 8 μg / mL chlorobenzene, 8 μg / mL bromobenzene, 8 μg / mL fluorobenzene, 8 μg / mL p-bromochlorobenzene, 8 μg / mL o-bromofluorobenzene, and 32 μg / mL m-bromofluorobenzene;

[0032] 6) Set instrument parameters:

[0033] Chromatographic column: DB-WAX capillary column, with an inner diameter of 0.32 mm, a length of 30 m, and a liquid film thickness of 0.5 μm;

[0034] Inlet temperature: 250°C;

[0035] Split ratio: 8:1;

[0036] Carrier gas: nitrogen;

[0037] Flow rate mode: constant flow;

[0038] Column flow rate: 1.5 mL / min;

[0039] Injection volume: 1.0 μL;

[0040] Heating program: initial temperature 35°C, maintain for 12 min, then increase the temperature to 80°C at a rate of 20°C per minute, maintain for 10 min, and finally increase the temperature to 140°C at a rate of 10°C per minute, maintain for 10 min;

[0041] Detector: FID, where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0042] Detector temperature: 250°C;

[0043] Sampling frequency: 20Hz / min;

[0044] 7) Take blank solution, peak identification solution, test sample solution, and test sample spiked solution and perform gas chromatography analysis under the above conditions and record the chromatogram.

[0045] In some embodiments, the method for separating and detecting related substances in p-bromofluorobenzene comprises the following steps:

[0046] 1) Diluent / blank solution: acetonitrile;

[0047] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene sample, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0048] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0049] 4) Peak identification solution: Pipette 40 μL of each of CB, BB, FB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0050] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0051] 6) Setting instrument parameters: injection port temperature 200℃-300℃, carrier gas flow rate 1.3mL / min-1.7mL / min, programmed temperature method, the temperature program is initial temperature 33-37℃, maintain for 12min, then increase the temperature to 80℃ at a rate of 10℃-25℃ per minute, maintain for 10min, and finally increase the temperature to 140℃ at a rate of 5℃-15℃ per minute, maintain for 10min, detector temperature 200℃-300℃, split ratio 6:1-10:1;

[0052] 7) Take blank solution, peak identification solution, test sample solution, and test sample spiked solution and perform gas chromatography analysis under the above conditions and record the chromatogram.

[0053] In some embodiments, the method for separating and detecting related substances in p-bromofluorobenzene comprises the following steps:

[0054] 1) Diluent / blank solution: acetonitrile;

[0055] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0056] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0057] 4) Peak identification solution: Pipette 40 μL of each of CB, BB, FB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0058] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0059] 6) Set instrument parameters:

[0060] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph;

[0061] Chromatographic column: DB-WAX capillary column, with an inner diameter of 0.32 mm, a length of 30 m, and a liquid film thickness of 0.5 μm;

[0062] Inlet temperature: 250°C;

[0063] Split ratio: 8:1;

[0064] Carrier gas: nitrogen;

[0065] Flow rate mode: constant flow;

[0066] Column flow rate: 1.5 mL / min;

[0067] Injection volume: 1.0 μL;

[0068] Heating program: initial temperature 35°C, maintain for 12 min, then increase the temperature to 80°C at a rate of 20°C per minute, maintain for 10 min, and finally increase the temperature to 140°C at a rate of 10°C per minute, maintain for 10 min;

[0069] Detector: FID, where the gas flow rate is H 2 :Air:N 2=30:300:30, mL / min;

[0070] Detector temperature: 250°C;

[0071] Sampling frequency: 20Hz / min;

[0072] 7) Take blank solution, peak identification solution, test sample solution, and test sample spiked solution and perform gas chromatography analysis under the above conditions and record the chromatogram.

[0073] The invention adopts a method for separating and determining related substances in p-bromofluorobenzene by gas chromatography, selects a polyethylene glycol capillary chromatographic column, can quickly and effectively separate and determine related substances in p-bromofluorobenzene, has a separation degree greater than 2.0, has good separation degrees between different related substances, can obtain the content of each related substance to be measured in a mixed sample by an area normalization method, has high accuracy and good reproducibility, is not interfered by other components, thereby meeting the control product quality requirements and ensuring the drug safety of escitalopram products.

[0074] Definition of terms

[0075] In the above or below, whether or not the words "about" or "approximately" are used, all the numbers disclosed herein are approximate values. Based on the disclosed numbers, the value of each number may differ by 1%, 2%, 5%, 7%, 8% or 10%.

[0076] The method provided by the present invention can effectively separate the related substances in p-bromofluorobenzene, with a separation degree of 1.5 or more, 2.5 or more, or 5 or more, and complete baseline separation. The method provided by the present invention is simple, fast, accurate and highly sensitive.

[0077] In the present invention, peak area normalization means that the sum of the peak areas of all compound components that can emit peaks in the gas phase is regarded as 1, and the percentage of the peak area of ​​each component represents the component content.

[0078] In the present invention, Air means air, and FID means a hydrogen flame ionization detector.

[0079] In the present invention, μL represents microliter, mL represents milliliter, ℃ represents Celsius, mg / mL represents milliliter, μg / mL represents microgram / milliliter, min represents minute, mL / min represents milliliter / minute, μm represents micrometer, mm represents millimeter, m represents meter, and Hz / min represents Hertz / minute. BRIEF DESCRIPTION OF THE DRAWINGS

[0080] Figure 1 The blank solution spectrum in Example 1 is shown;

[0081] Figure 2 The sample solution spectrum in Example 1 is shown;

[0082] Figure 3 The graph of the spiked solution of the test sample in Example 1 is shown;

[0083] Figure 4 The chlorobenzene solution spectrum of the related substance reference substance in Example 1 is shown;

[0084] Figure 5 The fluorobenzene solution spectrum of the related substance reference substance in Example 1 is shown;

[0085] Figure 6 The bromobenzene solution spectrum of the related substance reference substance in Example 1 is shown;

[0086] Figure 7 The reference substance p-bromochlorobenzene solution spectrum of the related substance in Example 1 is shown;

[0087] Figure 8 The spectrum of the reference substance o-bromofluorobenzene in Example 1 is shown;

[0088] Fig. 9 The spectrum of the reference substance m-bromofluorobenzene in Example 1 is shown;

[0089] Fig.10 The specificity experimental spectrum in Example 1 is shown;

[0090] Fig.11 The blank solution spectrum in Example 2 is shown;

[0091] Fig.12 The graph of the spiked solution of the test sample in Example 2 is shown;

[0092] Fig.13 The blank solution spectrum in Example 3 is shown;

[0093] Fig.14 The graph of the spiked solution of the test sample in Example 3 is shown;

[0094] Fig.15 The blank solution spectrum in Example 4 is shown;

[0095] Fig.16 The graph of the spiked solution of the test sample in Example 4 is shown;

[0096] Fig.17 The blank solution spectrum in Example 5 is shown;

[0097] Fig.18 The graph of the spiked solution of the test sample in Example 5 is shown;

[0098] Fig.19 The blank solution spectrum in Example 6 is shown;

[0099] Fig. 20 The graph of the spiked solution of the test sample in Example 6 is shown;

[0100] Fig.21 The blank solution spectrum in Example 7 is shown;

[0101] Fig. 22 The graph of the spiked solution of the test sample in Example 7 is shown. DETAILED DESCRIPTION

[0102] In order to enable those skilled in the art to better understand the technical solution of the present invention, some non-limiting embodiments are further disclosed below to further illustrate the present invention in detail.

[0103] The reagents used in the present invention can be purchased from the market or prepared by the method described in the present invention.

[0104] Example 1

[0105] Instruments and conditions

[0106] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0107] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0108] Inlet temperature: 250°C;

[0109] Split ratio: 8:1;

[0110] Carrier gas: nitrogen;

[0111] Flow rate mode: constant flow;

[0112] Column flow rate: 1.5 mL / min;

[0113] Injection volume: 1.0 μL;

[0114] Heating program:

[0115]

[0116]

[0117] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0118] Detector temperature: 250°C;

[0119] Sampling frequency: 20Hz / min.

[0120] Experimental procedures

[0121] 1) Diluent / blank solution: acetonitrile;

[0122] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0123] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0124] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0125] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0126] 6) Low concentration linear stock solution: Pipette 100 μL of the test solution into a 10 mL volumetric flask, dilute to the mark with diluent, and shake well;

[0127] 7) Quantitative limit solution: Take 100 μL of low concentration linear stock solution into a 10 mL volumetric flask, dilute to the mark with diluent, and shake well;

[0128] 8) Detection limit solution: Take 3.0 mL of the quantitation limit solution and transfer it to a 10 mL volumetric flask. Dilute to the mark with diluent and shake well.

[0129] 9) The linear solution is prepared as follows:

[0130] Table 1: Linear solution 1 preparation table

[0131]

[0132] Table 2: Linear solution 2 preparation table

[0133]

[0134]

[0135] 10) Take blank solution, quantitative limit solution, test solution, test sample spiked solution, each related substance peak identification solution, detection limit solution, each related substance limit level linear solution (0.01% to 0.15%), and each concentration level linear solution of the test solution (80% to 120%) and perform gas chromatography analysis according to Table 3, record the chromatogram, and the results are as follows: Figure 1-Figure 10 shown.

[0136] Table 3: Injection sequence table

[0137] Solution name Number of injection needles blank solution 1 Quantitative limit solution 1 Test solution 1 Test sample spike solution 1 blank solution 1 Chlorobenzene identification solution 1 Fluorobenzene identification solution 1 Bromobenzene identification solution 1 4-Bromochlorobenzene identification solution 1 o-Bromochlorobenzene identification solution 1 m-Bromochlorobenzene identification solution 1 blank solution 1 Detection limit solution 3 Quantitative limit solution 3 0.01% related substance limit solution 1 0.02% related substance limit solution 1 0.04% related substance limit solution 1 0.08% related substance limit solution 1 0.12% related substance limit solution 1 0.15% related substance limit solution 1 blank solution 1 80% test solution 1 90% test solution 1 100% test solution 1 110% test solution 1 120% test solution 1 blank solution 1 Test sample spike solution 1 1 Test sample spike solution 2 1 Test sample spike solution 3 1 Test sample spike solution 4 1 Test sample spike solution 5 1 Test sample spike solution 6 1

[0138] Result analysis:

[0139] Table 4: System suitability results

[0140]

[0141]

[0142] Table 5: Specificity results

[0143]

[0144]

[0145] Table 6: Repeatability results

[0146]

[0147] Table 7: Limit of quantitation results

[0148]

[0149] Table 8: Detection limit results

[0150]

[0151]

[0152] Table 9: Linearity-range results (LOQ ~ 0.15%)

[0153]

[0154] Table 10: Linear-range results (test sample concentration 80% to 120%)

[0155]

[0156] Example 2

[0157] Instruments and conditions

[0158] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0159] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0160] Inlet temperature: 250°C;

[0161] Split ratio: 8:1;

[0162] Carrier gas: nitrogen;

[0163] Flow rate mode: constant flow;

[0164] Column flow rate: 1.3 mL / min;

[0165] Injection volume: 1.0 μL;

[0166] Heating program:

[0167]

[0168] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0169] Detector temperature: 250°C;

[0170] Sampling frequency: 20Hz / min.

[0171] Experimental procedures

[0172] 1) Diluent / blank solution: acetonitrile;

[0173] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0174] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0175] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0176] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0177] 6) Take the blank solution and the test sample spiked solution and perform gas chromatography analysis under the above conditions, record the chromatogram, and the results are as follows: Fig.11 , Fig.12 shown.

[0178] Table 11: Retention time of related substances and separation degree of adjacent peaks in Example 2

[0179] Related substances Retention time (min) Separation Chlorobenzene (CB) 17.45 45.3 Bromobenzene (BB) 23.83 7.8 p-Bromochlorobenzene(BCB) 30.88 3.6 2-Bromofluorobenzene(2-BFB) 26.06 6.5 3-Bromofluorobenzene(3-BFB) 22.03 29.4 4-Bromofluorobenzene(BFB) 24.66 3.0

[0180] Example 3

[0181] Instruments and conditions

[0182] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0183] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0184] Inlet temperature: 250°C;

[0185] Split ratio: 8:1;

[0186] Carrier gas: nitrogen;

[0187] Flow rate mode: constant flow;

[0188] Column flow rate: 1.7 mL / min;

[0189] Injection volume: 1.0 μL;

[0190] Heating program:

[0191]

[0192] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0193] Detector temperature: 250°C;

[0194] Sampling frequency: 20Hz / min.

[0195] Experimental procedures

[0196] 1) Diluent / blank solution: acetonitrile;

[0197] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0198] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0199] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0200] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0201] 6) Take the blank solution and the test sample spiked solution and perform gas chromatography analysis under the above conditions, record the chromatogram, and the results are as follows: Fig.13 , Fig.14 shown.

[0202] Table 12: Retention time of related substances and separation degree of adjacent peaks in Example 3

[0203] Related substances Retention time (min) Separation Chlorobenzene (CB) 16.13 212.4 Bromobenzene (BB) 21.23 7.2 p-Bromochlorobenzene(BCB) 29.87 2.7 2-Bromofluorobenzene(2-BFB) 23.72 6.4 3-Bromofluorobenzene(3-BFB) 19.79 26.8 4-Bromofluorobenzene(BFB) 21.95 2.6

[0204] Example 4

[0205] Instruments and conditions

[0206] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0207] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0208] Inlet temperature: 250°C;

[0209] Split ratio: 6:1;

[0210] Carrier gas: nitrogen;

[0211] Flow rate mode: constant flow;

[0212] Column flow rate: 1.5 mL / min;

[0213] Injection volume: 1.0 μL;

[0214] Heating program:

[0215]

[0216] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0217] Detector temperature: 250°C;

[0218] Sampling frequency: 20Hz / min.

[0219] Experimental procedures

[0220] 1) Diluent / blank solution: acetonitrile;

[0221] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0222] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0223] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0224] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0225] 6) Take the blank solution and the test sample spiked solution and perform gas chromatography analysis under the above conditions, record the chromatogram, and the results are as follows: Fig.15 , Fig.16 shown.

[0226] Table 13: Retention time of related substances in Example 4 and separation degree from adjacent peaks

[0227] Related substances Retention time (min) Separation Chlorobenzene (CB) 16.72 8.6 Bromobenzene (BB) 22.38 7.1 p-Bromochlorobenzene(BCB) 30.31 3.2 2-Bromofluorobenzene(2-BFB) 24.96 6.5 3-Bromofluorobenzene(3-BFB) 20.78 27.8 4-Bromofluorobenzene(BFB) 23.21 2.7

[0228] Example 5

[0229] Instruments and conditions

[0230] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0231] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0232] Inlet temperature: 250°C;

[0233] Split ratio: 10:1;

[0234] Carrier gas: nitrogen;

[0235] Flow rate mode: constant flow;

[0236] Column flow rate: 1.5 mL / min;

[0237] Injection volume: 1.0 μL;

[0238] Heating program:

[0239]

[0240] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0241] Detector temperature: 250°C;

[0242] Sampling frequency: 20Hz / min.

[0243] Experimental procedures

[0244] 1) Diluent / blank solution: acetonitrile;

[0245] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0246] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0247] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0248] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0249] 6) Take the blank solution and the test sample spiked solution and perform gas chromatography analysis under the above conditions, record the chromatogram, and the results are as follows: Fig.17 , Fig.18 shown.

[0250] Table 14: Retention time of related substances in Example 5 and separation degree from adjacent peaks

[0251] Related substances Retention time (min) Separation Chlorobenzene (CB) 16.70 28.0 Bromobenzene (BB) 22.34 7.6 p-Bromochlorobenzene(BCB) 30.31 12.6 2-Bromofluorobenzene(2-BFB) 24.94 6.8 3-Bromofluorobenzene(3-BFB) 20.75 28.0 4-Bromofluorobenzene(BFB) 23.09 2.6

[0252] Example 6

[0253] Instruments and conditions

[0254] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0255] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0256] Inlet temperature: 250°C;

[0257] Split ratio: 8:1;

[0258] Carrier gas: nitrogen;

[0259] Flow rate mode: constant flow;

[0260] Column flow rate: 1.5 mL / min;

[0261] Injection volume: 1.0 μL;

[0262] Heating program:

[0263]

[0264]

[0265] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0266] Detector temperature: 250°C;

[0267] Sampling frequency: 20Hz / min.

[0268] Experimental procedures

[0269] 1) Diluent / blank solution: acetonitrile;

[0270] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0271] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0272] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0273] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0274] 6) Take the blank solution and the test sample spiked solution and perform gas chromatography analysis under the above conditions, record the chromatogram, and the results are as follows: Fig.19 , Fig. 20 shown.

[0275] Table 15: Retention time of related substances in Example 6 and separation degree from adjacent peaks

[0276] Related substances Retention time (min) Separation Chlorobenzene (CB) 16.96 28.3 Bromobenzene (BB) 22.62 7.5 p-Bromochlorobenzene(BCB) 30.43 2.9 2-Bromofluorobenzene(2-BFB) 25.18 6.6 3-Bromofluorobenzene(3-BFB) 21.03 28.3 4-Bromofluorobenzene(BFB) 23.40 2.6

[0277] Example 7

[0278] Instruments and conditions

[0279] Gas chromatograph model: Agilent 7890A-7693 gas chromatograph

[0280] Chromatographic column: DB-WAX capillary column (30m*0.32mm, 0.5μm)

[0281] Inlet temperature: 250°C;

[0282] Split ratio: 8:1;

[0283] Carrier gas: nitrogen;

[0284] Flow rate mode: constant flow;

[0285] Column flow rate: 1.5 mL / min;

[0286] Injection volume: 1.0 μL;

[0287] Heating program:

[0288]

[0289] Detector: FID (flame ionization detector), where the gas flow rate is H 2 :Air:N 2 =30:300:30, mL / min;

[0290] Detector temperature: 250°C;

[0291] Sampling frequency: 20Hz / min.

[0292] Experimental procedures

[0293] 1) Diluent / blank solution: acetonitrile;

[0294] 2) Test solution: Weigh about 200 mg of p-bromofluorobenzene, place in a 5 mL volumetric flask, dissolve and dilute with diluent, shake well, and prepare a 40 mg / mL p-bromofluorobenzene test solution;

[0295] 3) Peak identification stock solution: weigh about 25 mg of each of chlorobenzene (CB), bromobenzene (BB), fluorobenzene (FB), p-bromochlorobenzene (BCB), o-bromofluorobenzene (2-BFB), and m-bromofluorobenzene (3-BFB) reference substances, place them in 25 mL volumetric flasks, add diluent to dissolve, make up to volume, shake well, and prepare 1 mg / mL peak identification stock solution;

[0296] 4) Peak identification solution: Pipette 40 μL of each of CB, FB, BB, BCB, and 2-BFB stock solutions and 160 μL of 3-BFB stock solution into 6 different 5 mL volumetric flasks, add diluent to dissolve, make up to volume, and shake well to obtain peak identification solutions of the relevant substances;

[0297] 5) Test sample spike solution: Take about 200 mg of p-bromofluorobenzene, weigh it, put it into a 5 mL volumetric flask, add 40 μL of CB, BB, BCB stock solutions, 160 μL of 3-BFB stock solution, dilute to the scale with diluent, shake well to obtain the test sample spike solution;

[0298] 6) Take the blank solution and the test sample spiked solution and perform gas chromatography analysis under the above conditions, record the chromatogram, and the results are as follows: Fig.21 , Fig. 22 shown.

[0299] Table 16: Retention time of related substances in Example 7 and separation degree from adjacent peaks

[0300] Related substances Retention time (min) Separation Chlorobenzene (CB) 16.42 27.8 Bromobenzene (BB) 22.07 7.4 p-Bromochlorobenzene(BCB) 30.19 3.0 2-Bromofluorobenzene(2-BFB) 24.72 6.9 3-Bromofluorobenzene(3-BFB) 20.47 27.8 4-Bromofluorobenzene(BFB) 22.86 2.6

[0301] The method of the present invention has been described through preferred embodiments, and relevant personnel can obviously modify or appropriately change and combine the methods and applications described herein within the content, spirit and scope of the present invention to implement and apply the technology of the present invention. Those skilled in the art can refer to the content of this article and appropriately improve the process parameters. It is particularly important to point out that all similar replacements and modifications are obvious to those skilled in the art, and they are all considered to be included in the present invention.

Claims

1. A method for separating and detecting related substances in p-bromofluorobenzene by gas chromatography, characterized in that: The p-bromofluorobenzene sample and the related substance reference substance are dissolved in a solvent to prepare a related substance peak identification solution, a test solution and a test spike solution, and a capillary chromatographic column and a detector with a polyethylene glycol as the stationary phase are selected, and the related substances include or are chlorobenzene, bromobenzene, fluorobenzene, p-bromochlorobenzene, o-bromofluorobenzene and m-bromofluorobenzene; The following steps are involved: 1) Diluent / blank solution: acetonitrile; 2) Test solution: Weigh the p-bromofluorobenzene sample and dissolve it in a diluent to prepare a p-bromofluorobenzene test solution; 3) Peak identification stock solution: weigh the reference substances of the relevant substances, add diluent to dissolve, and prepare the peak identification stock solution; 4) Peak identification solution: pipette the peak identification stock solution obtained in step 3) respectively, add diluent to dilute, and prepare peak identification solutions of each relevant substance; 5) Sample spike solution: Weigh the p-bromofluorobenzene sample, add the peak identification stock solution obtained in step 3) respectively, dilute with diluent, and prepare the sample spike solution; 6) Set the instrument parameters: injection port temperature 200℃-300℃, carrier gas flow rate 1.3mL / min-1.7mL / min, programmed temperature method, the heating program is the initial temperature 33-37℃, maintain for 10min-15min, then increase the temperature at a rate of 10℃-25℃ per minute to 80℃, maintain for 5min -15min, finally, increase the temperature to 140℃ at a rate of 5℃-15℃ per minute, maintain for 5min-15min, detector temperature 200℃-300℃, split ratio 6:1-10:1; 7) Take blank solution, peak identification solution, test solution, and test sample spiked solution and perform gas chromatography analysis according to the above parameters and record the chromatogram.

2. The method according to claim 1, wherein the solvent comprises at least one of chloroform, n-propanol, butanone, acetonitrile and toluene.

3. The method according to claim 1, wherein the polyethylene glycol capillary chromatographic column is selected from one of HP-INNOWAX, DB-FFAP, HP-20M, DB-WAX and 007-20M. The method according to claim 1 , wherein the detector is a hydrogen flame ionization detector.

5. The method according to claim 1, wherein the carrier gas used for gas chromatography separation is nitrogen, and the flow rate is 1.5 mL / min.

6. The method according to claim 1, wherein the heating program is an initial temperature of 35°C, maintained for 12 minutes, then heated to 80°C at a rate of 20°C per minute, maintained for 10 minutes, and finally heated to 140°C at a rate of 10°C per minute, maintained for 10 minutes.

7. The method according to claim 1, wherein the chromatographic column is a DB-WAX capillary chromatographic column having an inner diameter of 0.32 mm, a length of 30 m, and a liquid film thickness of 0.5 μm.

8. The method according to claim 1, wherein the injection port temperature is 250°C, the detector temperature is 250°C, and the split ratio is 8:

1.

9. The method according to any one of claims 1 to 8, characterized in that: The method comprises the following steps: 1) Diluent / blank solution: acetonitrile; 2) Test solution: Weigh the p-bromofluorobenzene sample and dissolve it in a diluent to prepare a 40 mg / mL p-bromofluorobenzene test solution; 3) Peak identification stock solution: weigh each related substance reference substance, add diluent to dissolve, and prepare 1 mg / mL peak identification stock solution of each related substance; 4) Peak identification solution: pipette the peak identification stock solution obtained in step 3), add diluent to dilute, and prepare 8 μg / mL chlorobenzene, 8 μg / mL bromobenzene, 8 μg / mL fluorobenzene, 8 μg / mL p-bromochlorobenzene, 8 μg / mL o-bromofluorobenzene, 32 μg / mL m-bromofluorobenzene peak identification solutions respectively; 5) Test sample spike solution: Weigh the p-bromofluorobenzene sample, add the peak identification stock solutions obtained in step 3) respectively, dilute with diluent, and prepare a test sample spike solution containing 40 mg / mL p-bromofluorobenzene, 8 μg / mL chlorobenzene, 8 μg / mL bromobenzene, 8 μg / mL fluorobenzene, 8 μg / mL p-bromochlorobenzene, 8 μg / mL o-bromofluorobenzene, and 32 μg / mL m-bromofluorobenzene; 6) Set instrument parameters: Chromatographic column: DB-WAX capillary column, with an inner diameter of 0.32 mm, a length of 30 m, and a liquid film thickness of 0.5 μm; Inlet temperature: 250°C; Split ratio: 8:1; Carrier gas: nitrogen; Flow rate mode: constant flow; Column flow rate: 1.5 mL / min; Injection volume: 1.0 μL; Heating program: initial temperature 35°C, maintain for 12 min, then increase the temperature to 80°C at a rate of 20°C per minute, maintain for 10 min, and finally increase the temperature to 140°C at a rate of 10°C per minute, maintain for 10 min; Detector: FID, where the gas flow rate ratio is H2:Air:N2=30:300:30, mL / min; Detector temperature: 250°C; Sampling frequency: 20Hz / min; 7) Take blank solution, peak identification solution, test solution, and test sample spiked solution and perform gas chromatography analysis according to the above parameters and record the chromatogram.

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