CN113984921A - Gas chromatography-mass spectrometry combined method for determining genotoxic impurity 1, 3-dichloro-2-propanol - Google Patents
Gas chromatography-mass spectrometry combined method for determining genotoxic impurity 1, 3-dichloro-2-propanol Download PDFInfo
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Abstract
The invention discloses a gas chromatography-mass spectrometry combined analysis method of genotoxic impurity 1, 3-dichloro-2-propanol, which comprises the following steps: (1) pretreating a sample to prepare a sample solution, wherein perfluoropropionic acid is used as a derivatization reagent; (2) and detecting the sample solution by adopting a gas chromatography-mass spectrometer to determine the content of the 1, 3-dichloro-2-propanol impurity in the sample. The method has the advantages of good linearity, strong specificity, good precision, high accuracy and high sensitivity, and can quickly and accurately measure the content of the 1, 3-dichloro-2-propanol.
Description
Technical Field
The invention belongs to the field of drug analysis and detection, and particularly relates to a gas chromatography-mass spectrometry combined method for determining genotoxic impurity 1, 3-dichloro-2-propanol.
Background
Linezolid (Linezolid) is an artificially synthesized oxazolidinone antibiotic, and acts on bacterial 50S ribosomal subunit to inhibit the connection of mRNA and ribosome during the initiation stage of the translation system, thereby preventing the formation of 70S initiation complex and inhibiting the synthesis of bacterial protein. US FDA approval for treatment of gram-positive (G) was obtained in 2000+) Infections caused by cocci. The structural formula is shown as the following formula I:
in the synthesis process of linezolid, an intermediate linezolid epoxide (formula II) is used, epichlorohydrin is used as a starting material for synthesizing the intermediate, and 1, 3-dichloro-2-propanol is produced in the industrial preparation process of epichlorohydrin. In 2017, 10 and 27, the list of carcinogens published by the international cancer research institution of the world health organization is preliminarily collated for reference, and the 1, 3-dichloro-2-propanol is in the list of 2B carcinogens.
Since 1, 3-dichloro-2-propanol has a low limit (15ppm), has no ultraviolet absorption, cannot be directly detected by liquid chromatography, is not easily ionized, and is directly detected by liquid chromatography or gas chromatography, the signal is generally poor.
The domestic and international detection shows that 1, 3-dichloro-2-propanol is mostly in the field of food, the food treatment method needs to add adsorbents such as diatomite and the like, and then the enrichment treatment is carried out by utilizing column chromatography, the sample pretreatment steps are various, the sample usage amount is large, and the food sample treatment method is not suitable for the pretreatment of a medicine solution (Zhou Xiang Ju, Xiejing Jing, Zhao Yuqi, and the like. gas chromatography-mass spectrometry is used for measuring chloropropanol compounds [ J ] in soy sauce, Chinese seasoning, 2011(05): 88-90.).
CN201910616208.9 discloses a method for detecting residual chloropropanol compounds in ornidazole, which is to use headspace gas chromatography to detect by using a hydrogen Flame Ionization Detector (FID), wherein the sensitivity of the instrument is poor (the detection limit concentration is 150 ng/mL: 7.5 mug/mL of reference substance stock solution is prepared, then 0.2mL to 10mL of volumetric flask is taken for constant volume to obtain the detection limit concentration of 150ng/mL), the usage amount of a sample is large, and the pretreatment of the sample is complicated (about 1.0g of ornidazole is taken and precisely weighed, a 10mL iodine measuring flask is put, 4.0mL of mixed solvent, namely acetone-n-hexane (1:9) is added and sealed, the mixture is oscillated for 50 times per minute by a speed-regulating oscillator for 10min, an ice water bath is 30min, a 0.22μm nylon membrane is filtered, and the continuous filtrate is taken and recovered to room temperature to be used as a sample solution).
Therefore, the development of a method which has high sensitivity and small sample dosage and can quickly and accurately detect the genotoxic impurity 1, 3-dichloro-2-propanol is a problem to be solved by the technical personnel in the field.
Disclosure of Invention
The invention provides a method for detecting genotoxic impurity 1, 3-dichloro-2-propanol, which has the advantages of good linearity, strong specificity, good precision, high accuracy and high sensitivity, and can quickly and accurately determine the content of 1, 3-dichloro-2-propanol.
A gas chromatography-mass spectrometry combined detection method for genotoxic impurity 1, 3-dichloro-2-propanol is realized by the following scheme: (1) sample pretreatment: mixing a sample to be tested with perfluoropropionic acid (formula IV), and carrying out derivatization reaction on 1, 3-dichloro-2-propanol (formula III) in the sample and the perfluoropropionic acid (formula IV), wherein a catalyst of the derivatization reaction is a sulfuric acid solution, and the concentration of the sulfuric acid solution is 15-50%, preferably 25-40%, most preferably 40%, and the unit is v/v
(2) And (3) gas chromatography-mass spectrometry detection: and detecting by using a gas chromatography-mass spectrometer to determine the content of the 1, 3-dichloro-2-propanol impurity in the sample.
In a specific embodiment, the time of the derivatization reaction in step (1) is 10min to 30min, preferably 15min to 25min, and most preferably 20 min.
In particular embodiments, the temperature of the derivatization reaction in step (1) is from 50 ℃ to 70 ℃, preferably from 55 ℃ to 65 ℃, and most preferably 60 ℃.
In a specific embodiment, a diluent is further used in the derivatization reaction in step (1), and the diluent is isopentane, cyclohexane, or n-heptane, preferably n-heptane.
In a specific embodiment, the method comprises:
(1) sample pretreatment: mixing a sample to be tested with perfluoropropionic acid, a diluent and 25-40% sulfuric acid solution for derivatization reaction, reacting at 50-70 ℃ for 10-30 min, adding water and the diluent for extraction, and taking supernatant to obtain a sample solution.
(2) And (3) gas chromatography-mass spectrometry detection: and (2) detecting the sample solution in the step (1) by adopting a gas chromatography-mass spectrometer, recording a spectrogram of the sample solution, and calculating the content of the 1, 3-dichloro-2-propanol impurity in the sample to be detected according to an external standard method according to a pre-obtained standard curve of the 1, 3-dichloro-2-propanol.
In a specific embodiment, the standard curve of 1, 3-dichloro-2-propanol in step (2) is obtained by the following method: dissolving a reference substance of 1, 3-dichloro-2-propanol in a diluent to prepare a linear solution stock solution of 1, 3-dichloro-2-propanol with a linear concentration range; respectively mixing the stock solution of the linear solution of 1, 3-dichloro-2-propanol with perfluoropropionic acid, a diluent and 25-40% sulfuric acid solution for derivatization reaction, adding water and the diluent for extraction after the reaction is carried out for 10-30 min at 50-70 ℃, and taking supernatant as the linear solution; and (3) carrying out gas chromatography-mass spectrometry detection on the linear solution, and drawing a standard curve by taking the concentration of the linear solution as a horizontal coordinate and the peak area as a vertical coordinate.
In a particular embodiment, the standard curve of 1, 3-dichloro-2-propanol previously obtained according to the invention is: 91.1711x +638.7254, R2＝0.9992。
In a specific embodiment, the chromatography column for GC-MS detection in step (2) is DB-5MS, HP-5MS, SH-Rxi-5Sil MS, preferably SH-Rxi-5Sil MS, with a specification of 30m 250 μm,0.25 μm.
In a specific embodiment, the conditions of the GC assay in step (2) are as follows: gas phase conditions: the carrier gas is high-purity helium; the flow rate of the carrier gas is 0.6-1.5mL/min, preferably 1.2 mL/min; the split ratio is 3: 1-5: 1, preferably 3: 1; sample inlet temperature: 230 to 280 ℃, preferably 250 ℃; the temperature raising program is that after the temperature is kept at 50-80 ℃ for 2min, the temperature is raised to 100-150 ℃ at 15-25 ℃/min and kept for 0-2 min, then the temperature is raised to 240 ℃ at 40 ℃/min and kept for 2min, preferably, after the temperature is kept at 60 ℃ for 2min, the temperature is raised to 100 ℃ at 20 ℃/min and kept for 2min, and then the temperature is raised to 240 ℃ at 40 ℃/min and kept for 2 min.
In a specific embodiment, the conditions of the GC-MS detection in the step (2) are mass spectrometry conditions: the ionization source is an EI source, and the temperature of the ionization source is 200-250 ℃, preferably 230 ℃; an analyzer: and the single quadrupole mass analyzer has a monitoring mode of selecting an ion monitoring mode SIM and extracts ions of 110 and 225.
In a specific embodiment, the conditions for detecting the combination of gases in step (2) are as follows
a) Gas phase conditions:
a chromatographic column: DB-5MS, HP-5MS, SH-Rxi-5Sil MS, preferably SH-Rxi-5Sil MS, with specification of 30m 250 μm,0.25 μm;
carrier gas: high purity helium gas;
flow rate of carrier gas: 0.6mL/min-1.5mL/min, preferably 1.2 mL/min;
the split ratio is 3: 1-5: 1, preferably 3: 1;
sample inlet temperature: 230 to 280 ℃, preferably 250 ℃;
sample introduction amount: 1.0-3.0. mu.L, preferably 1.0. mu.L;
temperature rising procedure: after keeping the temperature at 50-80 ℃ for 2min, heating to 100-150 ℃ at 15-25 ℃/min for 2min, and then heating to 240 ℃ at 40 ℃/min for 2 min; preferably, after keeping the temperature at 60 ℃ for 2min, heating to 100 ℃ at 20 ℃/min for 2min, and then heating to 240 ℃ at 40 ℃/min for 2 min;
the sample introduction mode is direct sample introduction;
b) mass spectrum conditions: the ionization source is an EI source, and the temperature of the ionization source is 200-250 ℃, preferably 230 ℃; an analyzer: single quadrupole mass analyser, monitoring mode is selected ion monitoring mode SIM, extracted ions are 110, 225, solvent delay: 3 min.
In specific embodiments, the GC/MS instrument of the invention is a SHIMADZU GCMS QP2020 or Agilent 7890B-5977B.
In a specific embodiment, the sample to be tested is linezolid or a linezolid intermediate, preferably, the linezolid intermediate is a compound represented by formula II
In a particular embodiment, the concentration of the sample solution of the invention is calculated according to the following method: the concentration of the sample solution was equal to the concentration of linear 3/limit of 1, 3-dichloro-2-propanol, more specifically, the limit of 1, 3-dichloro-2-propanol was 15ppm, the concentration of linear 3 was 300ng/mL, and the concentration of the sample solution was calculated to be 300ng/mL-1/15ppm＝20mg/mL。
The reagent and solvent used in the present invention are not particularly limited, and commercially available conventional reagents and solvents can be used.
It should be emphasized that the values or numerical end-points referred to in the claims are not limited to the numbers per se, and those skilled in the art will appreciate that they include the allowable error ranges that are well accepted in the art, such as experimental errors, measurement errors, statistical errors, random errors, etc., and that such error ranges are included in the scope of the invention.
Compared with the prior art, the invention has the following beneficial effects, but it should not be understood that the detection method of the invention only has the following effects:
(1) the detection method has the advantages of high sensitivity, small sample dosage, less pretreatment steps, and detection limit (1.5ppm) and quantification limit (4.5ppm) far lower than the limit (15ppm) of genotoxic impurity 1, 3-dichloro-2-propanol.
(2) The detection method has good linear relation in the range of 60 ng/mL-600 ng/mL and the linear correlation coefficient R2Is 0.9992.
(3) The detection method is accurate and feasible, the sample recovery rate is 106.09%, the RSD is 7.08%, and the instrument precision RSD is 2.05%.
(4) The invention develops a new method for detecting genotoxic impurity 1, 3-dichloro-2-propanol, realizes effective control of genotoxic impurity 1, 3-dichloro-2-propanol in linezolid or an intermediate thereof for the first time, and ensures the medication safety of patients.
Description of the drawings:
FIG. 1 is a chromatogram of a 1, 3-dichloro-2-propanol control of example 1
FIG. 2 is a standard curve of a linear equation for 1, 3-dichloro-2-propanol from example 2
FIG. 3 is a chromatogram of the quantitative limiting solution of example 2
FIG. 4 is a chromatogram of the detection limiting solution of example 2
FIG. 5 is a chromatogram of the sample solution of example 3
FIG. 6 is a chromatogram of the sample solution of example 3
Detailed Description
The following examples are intended to illustrate the present invention in further detail, but it should not be construed that the scope of the above-described subject matter of the present invention is limited to the following examples.
Linezolid epoxy (compound of formula II) used in the examples of the present invention was purchased from Jiangsu alpha pharmaceuticals, Inc.
The 1, 3-dichloro-2-propanol control used in the examples of the invention was purchased from aladdin and had a purity of 97%.
The instruments and models adopted in this embodiment are: and 4, directly injecting the sample by using a SHIMADZU GCMS QP2020 sampling mode.
The preparation method of the following solution used in the embodiment of the invention specifically comprises the following steps:
25% sulfuric acid solution: remove 7.5mL of water into a 20mL beaker, slowly add 2.5mL of concentrated sulfuric acid, and mix well.
40% sulfuric acid solution: 6.0mL of water was removed from a 20mL beaker, and 4.0mL of concentrated sulfuric acid was added slowly and mixed well.
60% sulfuric acid solution: 4.0mL of water was removed from a 20mL beaker, and 6.0mL of concentrated sulfuric acid was added slowly and mixed well.
Example 1: location experiments
Gas phase conditions: a chromatographic column: SH-Rxi-5Sil MS,30m × 250 μm,0.25 μm; carrier gas: high purity helium gas; flow rate of carrier gas: 1.2mL/min, the split ratio is 5: 1, sample introduction amount: 1.0 μ L, injection port temperature: at 250 ℃, programmed temperature increases were used: after keeping the temperature at 60 ℃ for 2min, the temperature is raised to 120 ℃ at 20 ℃/min, and then the temperature is raised to 240 ℃ at 40 ℃/min and kept for 2 min.
Mass spectrum conditions: the ionization source is an EI source, and the ion source temperature: 230 ℃, analyzer: a single quadrupole mass analyzer, full sweep range of 20-400, solvent delay: 3 min.
Diluent agent: n-heptane
Control stock solution a: accurately weighing 1, 3-dichloro-2-propanol reference substance 30mg to 100mL in a volumetric flask, adding a diluent to dissolve, diluting to a scale, and uniformly mixing; precisely transferring the solution into a volumetric flask with the volume ranging from 1.0mL to 10mL, diluting the solution to a scale with a diluent, and uniformly mixing the solution to obtain a reference substance stock solution a with the concentration of 30 mu g/mL.
Control solution: respectively transferring concentrated sulfuric acid solution (100 mu L), reference substance stock solution a (100 mu L) and perfluoropropionic acid (100 mu L) into a 10mL glass tube with a plug scale, adding water (5mL) and n-heptane (0.9mL) after water bath at 60 ℃ for 20min, standing for layering, taking supernatant to obtain reference substance solution with the concentration of 3.0 mu g/mL, injecting 1.0ul, and recording a map (figure 1). It can be seen from the figure that there is no interference peak around the control, and the method can be used for detecting the impurity.
Example 2: methodology investigation
Gas phase conditions: a chromatographic column: SH-Rxi-5Sil MS,30m × 250 μm,0.25 μm; carrier gas: high purity helium gas; flow rate of carrier gas: 1.2mL/min, the split ratio is 3: 1, sample introduction amount: 1.0 μ L, injection port temperature: at 250 ℃, programmed temperature increases were used: after keeping the temperature at 60 ℃ for 2min, heating to 100 ℃ at 20 ℃/min for 2min, and then heating to 240 ℃ at 40 ℃/min for 2 min.
Mass spectrum conditions: the ionization source is an EI source, and the ion source temperature: 230 ℃, analyzer: single quadrupole mass analyser, monitoring mode is selected ion monitoring mode (SIM), extracted ions are 110, 225, solvent delay: 3 min.
Diluent agent: n-heptane.
Control stock solution b: accurately weighing 1, 3-dichloro-2-propanol reference substance 30mg to 100mL in a volumetric flask, adding a diluent to dissolve, diluting to a scale, and uniformly mixing; precisely transferring the solution into a volumetric flask with the volume ranging from 1.0mL to 50mL, diluting the solution to a scale with a diluent, and uniformly mixing the solution to obtain a reference substance stock solution b with the concentration of 6.0 mu g/mL.
1. Linearity and range
1-5 parts of linear solution stock solution: precisely transferring the reference substance stock solutions b into 10mL volumetric flasks with 1.0mL, 3.0mL, 5.0mL, 7.0mL and 10.0mL to 5 different volumes respectively, diluting the volumetric flasks to a scale with a diluent, and uniformly mixing the volumetric flasks to obtain linear solution stock solutions 1 to 5, wherein the concentrations of the stock solutions are as follows: 0.6. mu.g/mL, 1.8. mu.g/mL, 3.0. mu.g/mL, 4.2. mu.g/mL, 6.0. mu.g/mL.
Linear solution: respectively transferring 40% sulfuric acid solution (100 mu L) + each linear solution stock solution (100 mu L) + perfluoropropionic acid (100 mu L) into a 10mL glass tube with a plug, adding water (5mL) after 60 ℃ water bath (20min), shaking with n-heptane (0.9mL), standing for layering, taking supernate to obtain linear 1-linear 5 solutions with the concentrations of 60ng/mL, 180ng/mL, 300ng/mL, 420ng/mL and 600ng/mL, respectively injecting 1.0ul, recording the map, drawing a standard curve (figure 2) by taking the concentration as an abscissa and the peak area as an ordinate, and obtaining a standard curve equation y of 91.1711x +638.7254, wherein R is 91.1711x +638.72542The linearity was found to be good in the range of 60ng/mL to 600ng/mL (0.9992).
2. Detection limit and quantification limit
Quantitative limiting solution stock solution: transfer control stock solution b 1.5mL into 10mL volumetric flask, dilute to the mark with diluent, mix well.
Detection limit solution stock solution: 0.5mL of the control solution stock solution b is transferred into a 10mL volumetric flask, diluted to the mark with diluent, and mixed evenly.
Quantitative limiting solution: transferring 40% sulfuric acid solution (100ul), quantitative limiting solution stock solution (100uL) and perfluoropropionic acid (100uL) into a 10mL glass tube with a plug, adding water (5mL) and n-heptane (0.9mL) after water bath (20min) at 60 ℃, standing for layering, taking supernatant to obtain quantitative limiting solution with the concentration of 90ng/mL, injecting 1.0ul each, and recording a map (figure 3).
Detection limiting solution: transferring 40% sulfuric acid solution (100ul), stock solution of the detection limit solution (100uL) and perfluoropropionic acid (100uL) into a 10mL glass tube with a plug, adding water (5mL) and n-heptane (0.9mL) after water bath (20min) at 60 ℃, standing for layering, taking supernatant to obtain the detection limit solution with the concentration of 30ng/mL, injecting 1.0ul of sample, and recording a map (figure 4).
The signal-to-noise ratio of the detection limit solution is greater than 3, and the signal-to-noise ratio of the quantification limit solution is greater than 10, which indicates that the detection limit and the quantification limit meet the test requirements.
3. Sample recovery rate
Precisely weighing 9 parts of 20mg of linezolid epoxy, respectively placing the 9 parts into a 10mL glass tube with a plug scale, dividing the glass tube into three groups (3 parts in each group), respectively adding 40% sulfuric acid solution (100 mu L) and quantitative limiting solution stock solution/linear solution stock solution 2/linear solution stock solution 3(100 mu L respectively) and perfluoropropionic acid (100 mu L) in each group, adding water (5mL) after water bath at 60 ℃ for 20min, shaking up n-heptane (0.9mL), standing for layering, taking supernate, preparing a quantitative limiting solution standard adding solution with the concentration of 90ng/mL, a linear 2 solution standard adding solution with the concentration of 180ng/mL and a linear 3 solution standard adding solution with the concentration of 300ng/mL into each 3 parts, respectively injecting 1.0ul, recording a graph, and calculating the recovery rate (n is 9) to be 106.09% and the RSD to be 7.08%.
4. Precision degree
Weighing 6 parts of 20mg of linezolid epoxy, respectively placing the 6 parts into 10mL glass tubes with plugs, respectively adding 40% sulfuric acid solution (100 mu L), linear solution stock solution 3(100 mu L), perfluoropropionic acid (100 mu L), water (5mL) after 60 ℃ water bath (20min), shaking uniformly with n-heptane (0.9mL), standing for layering, taking supernatant, preparing linear 3 solution with the concentration of 300ng/mL, adding standard solution, respectively injecting 1.0ul, recording a map, and calculating the recovery rate (n is 6) to be 110.03% and the RSD to be 2.05%.
Example 3: sample detection
Gas phase conditions: a chromatographic column: SH-Rxi-5Sil MS,30m × 250 μm,0.25 μm; carrier gas: high purity helium gas; flow rate of carrier gas: 1.2mL/min, the split ratio is 3: 1, sample introduction amount of 1.0 μ L, sample inlet temperature: at 250 ℃, programmed temperature increases were used: after maintaining at 60 deg.C for 2min, heating to 100 deg.C at 20 deg.C/min for 2min, and heating to 240 deg.C at 40 deg.C/min for 2 min.
Mass spectrum conditions: the ionization source is an EI source, and the ion source temperature: 230 ℃, analyzer: single quadrupole mass analyser, monitoring mode is selected ion monitoring mode (SIM), extracted ions are 110, 225, solvent delay: 3 min.
Preparing a sample solution: weighing 20mg of linezolid epoxy, respectively placing 2 parts of the linezolid epoxy into 10mL glass tubes with plugs, respectively adding 40% sulfuric acid solution (100 mu L) + n-heptane (100 mu L) + perfluoropropionic acid (100 mu L), adding water (5mL) after water bath (20min) at 60 ℃, uniformly shaking the n-heptane (0.9mL), standing for layering to prepare 20mg/mL sample solution, respectively injecting 1.0uL, recording a map (figures 5-6), and if the result is not detected, reporting that the sample is less than the detection limit (1.5 ppm).
Example 4: selection of concentrated sulfuric acid as catalyst
Gas phase conditions: a chromatographic column: SH-Rxi-5Sil MS,30m × 250 μm,0.25 μm; carrier gas: high purity helium gas; flow rate of carrier gas: 1.2mL/min, the split ratio is 5: 1, sample introduction amount of 1.0 μ L, sample inlet temperature: at 250 ℃, programmed temperature increases were used: after maintaining at 60 deg.C for 2min, heating to 100 deg.C at 20 deg.C/min for 2min, and heating to 240 deg.C at 40 deg.C/min for 2 min.
Mass spectrum conditions: the ionization source is an EI source, and the ion source temperature: 230 ℃, analyzer: single quadrupole mass analyser, monitoring mode is selected ion monitoring mode (SIM), extracted ions are 110, 225, solvent delay: 3 min.
Diluent agent: n-heptane
Control stock solution c: accurately weighing 1, 3-dichloro-2-propanol reference substance 30mg to 100mL in a volumetric flask, adding a diluent to dissolve, diluting to a scale, and uniformly mixing; precisely transferring the solution to a volumetric flask of 1.0mL to 100mL, diluting the solution to a scale with a diluent, uniformly mixing, precisely transferring the solution to a volumetric flask of 5.0mL to 10mL, diluting the solution to a scale with the diluent, and uniformly mixing to obtain a reference substance stock solution c with the concentration of 1.5 mu g/mL.
Control solution: adding a reference substance stock solution c (100 mu L) + perfluoropropionic acid (100 mu L) + concentrated sulfuric acid (100 mu L)/non-concentrated sulfuric acid into a 10mL glass tube with a plug, adding water (5mL) and n-heptane (0.9mL) after water bath at 60 ℃ (20min), standing for layering to prepare control solutions with the concentration of 150ng/mL and added with concentrated sulfuric acid and non-concentrated sulfuric acid, respectively injecting 1.0ul, recording a map, and processing data shown in Table 1.
Adding a standard solution to a sample: weighing 4 parts of linezolid epoxy, namely 10mg, respectively placing the 4 parts of linezolid epoxy into 10mL glass tubes with plugs, respectively adding a reference substance stock solution c (100 muL) + perfluoropropionic acid (100 muL) + concentrated sulfuric acid (100 muL)/sulfuric acid without adding, adding water (5mL) after a water bath (20min) at 60 ℃, shaking uniformly n-heptane (0.9mL), standing for layering, preparing 2 parts of sample addition standard solutions with the concentration of 150ng/mL, respectively injecting 1.0ul, recording a map, and processing data shown in Table 1, wherein the result shows that concentrated sulfuric acid is not added, the response is lower, the later-stage quantitative limit is increased, the detection interference possibly exists in item investigation such as the detection limit and the like, and in order to meet the limit requirement, the sample amount needs to be increased, but the sample amount can cause interference on the detection of the reference (1, 3-dichloro-2-propanol); the response is higher when concentrated sulfuric acid is directly added, but the recovery rate exceeds the requirement of a laboratory (the recovery rate is 50-150% required by gas chromatography-mass spectrometry).
TABLE 1 results of concentrated sulfuric acid sample application experiments
Example 5: screening test for sulfuric acid solution concentration
Gas phase conditions: a chromatographic column: SH-Rxi-5Sil MS,30m × 250 μm,0.25 μm; carrier gas: high purity helium gas; flow rate of carrier gas: 1.2mL/min, the split ratio is 3: 1, sample introduction amount of 1.0 μ L, sample inlet temperature: at 250 ℃, programmed temperature increases were used: after maintaining at 60 deg.C for 2min, heating to 100 deg.C at 20 deg.C/min for 2min, and heating to 240 deg.C at 40 deg.C/min for 2 min.
Mass spectrum conditions: the ionization source is an EI source, and the ion source temperature: 230 ℃, analyzer: single quadrupole mass analyser, monitoring mode is selected ion monitoring mode (SIM), extracted ions are 110, 225, solvent delay: 3 min.
Control stock b and diluent: the same as in example 2.
1-3 of a control solution stock solution: precisely transferring the control stock solutions b into 1.5mL, 2.5mL and 5.0mL to 3 different 10mL volumetric flasks respectively, diluting the flasks to the scale with a diluent, and uniformly mixing the flasks to obtain control solution stock solutions 1 to 3, wherein the concentrations of the control solution stock solutions are as follows: 900ng/mL, 1500ng/mL, 3000 ng/mL.
Control solution of 25% sulfuric acid concentration: transferring 25% sulfuric acid solution (100 muL) + control solution stock solution 1/control solution stock solution 2/control solution stock solution 3(100 muL each) + perfluoropropionic acid (100 muL) into a 10mL glass tube with a plug, adding water (5mL) after water bath (20min) at 60 ℃, shaking up n-heptane (0.9mL), standing for layering, taking supernatant to obtain control 1-control 3 solutions with the concentrations of 90ng/mL, 150ng/mL and 300ng/mL, and preparing 2 parts in parallel.
25% sulfuric acid concentration sample addition standard solution: weighing 20mg of linezolid epoxy, respectively placing 6 parts of the linezolid epoxy into 10mL glass tubes with plugs, respectively adding 25% sulfuric acid solution (100 mu L), control solution stock solution 1, control solution stock solution 2, control solution stock solution 3(100 mu L respectively), perfluoropropionic acid (100 mu L), adding water (5mL) after water bath at 60 ℃ for 20min, shaking up n-heptane (0.9mL), standing for layering, taking supernate to obtain control 1 solution adding standard solution with the concentration of 90ng/mL, control 2 solution adding standard solution with the concentration of 150ng/mL and control 3 solution adding standard solution with the concentration of 300ng/mL, respectively 2 parts of each standard solution.
Control solution of 40% sulfuric acid concentration: transferring 40% sulfuric acid solution (100 muL) + control solution stock solution 1/control solution stock solution 2/control solution stock solution 3(100 muL each) + perfluoropropionic acid (100 muL) into a 10mL glass tube with a plug, adding water (5mL) after water bath (20min) at 60 ℃, shaking up n-heptane (0.9mL), standing for layering, taking supernatant to obtain control 1-control 3 solutions with the concentrations of 90ng/mL, 150ng/mL and 300ng/mL, and preparing 2 parts in parallel.
Sample with 40% sulfuric acid concentration added with standard solution: weighing 20mg of linezolid epoxy, respectively placing 6 parts of the linezolid epoxy into 10mL glass tubes with plugs, respectively adding 40% sulfuric acid solution (100 mu L), control solution stock solution 1, control solution stock solution 2, control solution stock solution 3(100 mu L respectively), perfluoropropionic acid (100 mu L), adding water (5mL) after water bath at 60 ℃ for 20min, shaking up n-heptane (0.9mL), standing for layering, taking supernate to obtain control 1 solution adding standard solution with the concentration of 90ng/mL, control 2 solution adding standard solution with the concentration of 150ng/mL and control 3 solution adding standard solution with the concentration of 300ng/mL, respectively 2 parts of each standard solution.
Control solution of 60% sulfuric acid concentration: transferring 60% sulfuric acid solution (100 muL) + control solution stock solution 1/control solution stock solution 2/control solution stock solution 3(100 muL each) + perfluoropropionic acid (100 muL) into a 10mL glass tube with a plug, adding water (5mL) after water bath at 60 ℃ (20min), shaking with n-heptane (0.9mL), standing for layering, taking supernatant to obtain control 1-control 3 solutions with concentrations of 90ng/mL, 150ng/mL and 300ng/mL, and preparing 2 parts in parallel.
60% sulfuric acid concentration sample addition standard solution: weighing 20mg of linezolid epoxy, respectively placing 6 parts of the linezolid epoxy into 10mL glass tubes with plugs, respectively adding 60% sulfuric acid solution (100 mu L), control solution stock solution 1, control solution stock solution 2, control solution stock solution 3(100 mu L respectively), perfluoropropionic acid (100 mu L), adding water (5mL) after water bath at 60 ℃ for 20min, shaking up n-heptane (0.9mL), standing for layering, taking supernate to obtain control 1 solution adding standard solution with the concentration of 90ng/mL, control 2 solution adding standard solution with the concentration of 150ng/mL and control 3 solution adding standard solution with the concentration of 300ng/mL, respectively 2 parts of each standard solution.
The solutions are injected with 1.0uL of sample respectively, a graph is recorded, data processing is shown in table 2, results show that after 25% -40% of concentrated sulfuric acid is added, response is high, recovery rate meets requirements, and response value of 40% of sulfuric acid added and sample recovery rate are optimal.
TABLE 2 results of recovery test of sulfuric acid of different concentrations
Claims (10)
1. A gas chromatography-mass spectrometry combined detection method of 1, 3-dichloro-2-propanol is characterized in that,
(1) sample pretreatment: mixing a sample to be detected with perfluoropropionic acid (formula IV) to perform derivatization reaction on 1, 3-dichloro-2-propanol (formula III) in the sample and the perfluoropropionic acid (formula IV);
(2) and (3) gas chromatography-mass spectrometry detection: detecting by adopting a gas chromatography-mass spectrometer to determine the content of the 1, 3-dichloro-2-propanol impurity in the sample;
wherein, the catalyst of the derivatization reaction in the step (1) is sulfuric acid solution, and the concentration of the sulfuric acid solution is 15-50%, preferably 25-40%, most preferably 40%, and the unit is v/v.
2. The gas chromatography-mass spectrometry combined detection method according to claim 1, wherein the time of the derivatization reaction in step (1) is 10min to 30 min.
3. The gas chromatography-mass spectrometry combined detection method according to claim 1 or 2, wherein the temperature of the derivatization reaction in the step (1) is 50 ℃ to 70 ℃.
4. The gas chromatography-mass spectrometry detection method as claimed in any one of claims 1 to 3, wherein a diluent is further used in the derivatization reaction in step (1), and the diluent is isopentane, cyclohexane, or n-heptane.
5. The gas chromatography-mass spectrometry detection method of any one of claims 1-4, wherein the method comprises:
(1) sample pretreatment: mixing a sample to be tested with perfluoropropionic acid, a diluent and 25-40% sulfuric acid solution for derivatization reaction, reacting at 50-70 ℃ for 10-30 min, adding water and the diluent for extraction, and taking supernatant to obtain a sample solution.
(2) And (3) gas chromatography-mass spectrometry detection: and (2) detecting the sample solution in the step (1) by adopting a gas chromatography-mass spectrometer, recording a spectrogram of the sample solution, and calculating the content of the 1, 3-dichloro-2-propanol impurity in the sample to be detected according to an external standard method according to a pre-obtained standard curve of the 1, 3-dichloro-2-propanol.
6. The gas chromatography-mass spectrometry detection method of any one of claims 1 to 5, wherein the standard curve of 1, 3-dichloro-2-propanol in step (2) is obtained by: dissolving a reference substance of 1, 3-dichloro-2-propanol in a diluent to prepare a linear solution stock solution of 1, 3-dichloro-2-propanol with a linear concentration range; respectively mixing the stock solution of the linear solution of 1, 3-dichloro-2-propanol with perfluoropropionic acid, a diluent and a 25-40% sulfuric acid solution for derivatization reaction, reacting at 50-70 ℃ for 10-30 min, adding water and the diluent for extraction, and taking the supernatant to obtain the linear solution; and (3) carrying out gas chromatography-mass spectrometry detection on the linear solution, and drawing a standard curve by taking the concentration of the linear solution as a horizontal coordinate and the peak area as a vertical coordinate.
7. The GC MS/MS detection method of any one of claims 1 to 6, wherein the GC MS column in step (2) is DB-5MS, HP-5MS, SH-Rxi-5Sil MS, preferably SH-Rxi-5Sil MS, with a specification of 30m 250 μm,0.25 μm.
8. The GC MS/MS detection method according to any one of claims 1 to 7, wherein the GC MS detection in step (2) is performed under the following conditions: gas phase conditions: the carrier gas is high-purity helium; the flow rate of the carrier gas is 0.6-1.5mL/min, preferably 1.2 mL/min; the split ratio is 3: 1-5: 1; sample inlet temperature: 230 to 280 ℃, preferably 250 ℃; the temperature raising program is that after the temperature is kept at 50-80 ℃ for 2min, the temperature is raised to 100-150 ℃ at 15-25 ℃/min and kept for 0-2 min, then the temperature is raised to 240 ℃ at 40 ℃/min and kept for 2min, preferably, after the temperature is kept at 60 ℃ for 2min, the temperature is raised to 100 ℃ at 20 ℃/min and kept for 2min, and then the temperature is raised to 240 ℃ at 40 ℃/min and kept for 2 min.
9. The GC MS/MS detection method according to any one of claims 1 to 8, wherein the GC MS detection in step (2) is performed under the following conditions: mass spectrum conditions: the ionization source is an EI source, and the temperature of the ionization source is 200-250 ℃, preferably 230 ℃; an analyzer: and the single quadrupole mass analyzer has a monitoring mode of selecting an ion monitoring mode SIM and extracts ions of 110 and 225.
10. The gas chromatography-mass spectrometry combined detection method according to any one of claims 1 to 9, wherein the sample to be detected is linezolid or a linezolid intermediate, preferably a linezolid intermediate, and further preferably a compound represented by formula II
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