Analysis method of N-isobutyl-4-chloropyrrolidone

Abstract

The invention belongs to the technical field of organic compound analysis and detection, and particularly discloses an N-isobutyl-4-chloropyrrolidone analysis method which comprises the following steps: pretreating a sample to be analyzed; separating the to-be-derived sample liquid A by adopting two-dimensional chromatography; carrying out derivatization on the sample solution B to be derivatized; carrying out combined detection on the derivatization sample liquid through a fluorescence detector and a high-resolution mass spectrum; the N-isobutyl-4-chloropyrrolidone and the degraded impurities of the N-isobutyl-4-chloropyrrolidone are automatically identified and analyzed through CNN-LSTM double models. According to the analysis method of the N-isobutyl-4-chloropyrrolidone, the N-isobutyl-4-chloropyrrolidone containing chlorine and secondary amine groups is subjected to high-sensitivity and high-selectivity analysis in a complex matrix, so that the sensitivity and the selectivity of the N-isobutyl-4-chloropyrrolidone are improved, and the sensitivity and the selectivity of the N-isobutyl-4-chloropyrrolidone are improved. The method can be widely applied to medical intermediate quality control, organic synthesis process monitoring and functional material purity detection scenes.

Description

Technical Field

[0001] This invention belongs to the field of organic compound analysis and detection technology, specifically relating to an analytical method for N-isobutyl-4-chloropyrrolidone. Background Technology

[0002] N-isobutyl-4-chloropyrrolidinone is a nitrogen-containing heterocyclic organic compound whose structure includes a five-membered pyrrolidone ring, an isobutyl substituent, and a chlorine atom at the 4-position. As an important representative of pyrrolidone derivatives, N-isobutyl-4-chloropyrrolidinone is a key intermediate in organic synthesis, the preparation of pharmaceutical active molecules, and the synthesis of functional materials due to the presence of both a chlorine atom and a secondary amine group in its molecular structure.

[0003] However, current analytical methods for N-isobutyl-4-chloropyrrolidone have the following problems: 1. Large polarity differences lead to low separation efficiency, which, combined with matrix interference, makes it difficult to achieve accurate separation.

[0004] The molecular polarity of N-isobutyl-4-chloropyrrolidone is synergistically regulated by the chlorine atom and the secondary amine group, covering a polarity range of "medium-high polarity to medium-low polarity," naturally exceeding the separation window of conventional one-dimensional chromatography. Simultaneously, complex synthetic reaction matrices or sample substrates further exacerbate interference, leading to a double dilemma for traditional one-dimensional chromatography techniques. While one-dimensional hydrophilic interaction chromatography (HILIC) can retain highly polar components, it cannot effectively separate low-polarity byproducts. When using one-dimensional reversed-phase C18 chromatography, the high polarity of the target compound results in a short retention time, easily overlapping with solvent peaks and highly polar impurities. Ultimately, this leads to peak overlap and baseline interference, failing to meet the resolution requirements for quantitative analysis and making it difficult to accurately distinguish the target compound from impurities.

[0005] 2. Weak ultraviolet absorption leads to insufficient detection sensitivity and difficulty in identifying impurities.

[0006] The N-isobutyl-4-chloropyrrolidone molecule lacks strong chromophores such as conjugated double bonds and benzene rings, resulting in a molar absorptivity of only 250-480 L / (mol·cm) in the 200-400 nm UV spectral range. Conventional UV detectors typically have detection limits of 1-10 ng / mL, which is insufficient for detecting trace degradation impurities in pharmaceutical intermediates. Furthermore, this compound exhibits poor stability, readily hydrolyzing under acidic / alkaline conditions to form 4-hydroxy-N-isobutylpyrrolidone. It also readily undergoes cis / trans isomerization at high temperatures (>80℃) or under light irradiation. These degradation impurities are not only present in extremely low concentrations and may also lack chromophores, further reducing UV response, but they are also structurally highly similar to the main component, making rapid and accurate differentiation difficult using conventional mass spectrometry techniques, severely impacting the accuracy and reliability of analytical results.

[0007] 3. Derivatization technology faces application bottlenecks, hindering the optimization of secondary amine detection.

[0008] Derivatization is an effective method to improve the detection sensitivity of secondary amine compounds, but the derivatization of N-isobutyl-4-chloropyrrolidone still faces key technical challenges: First, highly specific derivatizing reagents need to be screened to ensure efficient reaction only with the target analyte and specific impurities (such as degradation products containing secondary amine structures), avoiding non-specific binding with other components in the matrix; second, the reaction conditions need to be controlled to maintain mildness, preventing high temperatures, strong acids, or alkalis from inducing further degradation or isomerization of the target analyte; third, the reaction process needs to be optimized to avoid interference from reagent decomposition products on the detection results. These problems have not yet been effectively solved, directly limiting the practical application of derivatization technology in the detection of this compound.

[0009] 4. The presence of numerous interfering substances and the lag in data analysis make qualitative analysis difficult and result in poor industrial adaptability.

[0010] In real-world testing scenarios, the interfering substance system is extremely complex: the synthesis reaction solution contains unreacted raw materials (4-chloropyrrolidone, isobutylamine), byproducts (N-isobutylpyrrolidone, 1,3-diisobutyl-4-chloropyrrolidone), catalyst residues (zinc chloride), and solvent residues (ethanol); the finished product may contain degradation impurities (4-hydroxy derivatives, cis isomers) generated during storage. Some interfering substances have highly similar physicochemical properties to the target compound, further increasing the difficulty of qualitative analysis. More importantly, traditional data processing relies heavily on the experience of analysts for manual integration, peak identification, and impurity identification, which is not only inefficient and subjective but also fails to meet the demands of real-time, rapid, and automated quality monitoring in modern industrial production, making it difficult for analytical results to adapt to the quality control pace of large-scale production.

[0011] Currently, although some literature reports the use of gas chromatography or liquid chromatography-mass spectrometry to detect pyrrolidone compounds with similar structures, no analytical method has been published specifically for N-isobutyl-4-chloropyrrolidone and its specific degradation impurities, and systematically solves a series of integrated problems such as high-selectivity separation, high-sensitivity detection, specific derivatization and intelligent data analysis.

[0012] Therefore, there is an urgent need in this field to develop a novel and comprehensive analytical solution to overcome the limitations of existing technologies, achieve superior analytical performance for N-isobutyl-4-chloropyrrolidone and its impurities in complex matrices, and meet the precise quality control requirements of this key intermediate in the fields of organic synthesis, pharmaceuticals and materials. Summary of the Invention

[0013] The purpose of this invention is to provide an analytical method for N-isobutyl-4-chloropyrrolidone. This analytical method, through its high sensitivity and high selectivity for the analysis of N-isobutyl-4-chloropyrrolidone containing chlorine and secondary amine groups in complex matrices, can be widely applied to the quality control of pharmaceutical intermediates, monitoring of organic synthesis processes, and purity detection of functional materials.

[0014] To achieve the above objectives, the present invention provides an analytical method for N-isobutyl-4-chloropyrrolidone, comprising the following steps: Step S1: Pre-treat the sample to be analyzed; The sample to be analyzed was extracted with ultrapure water and purified by an aminopropyl-bonded silica gel solid-phase extraction column to obtain sample solution A to be derived. Step S2: Separate the sample solution A to be derivatized using two-dimensional chromatography to obtain the sample solution B to be derivatized; Hydrophilic interaction chromatography was used as the first dimension and reversed-phase C18 chromatography as the second dimension in combination. Step S3: Derivatize the sample solution B to be derivatized to obtain a derivatized sample solution; Step S4: Detect the derivatized sample solution using a fluorescence detector and high-resolution mass spectrometry. Step S5: Analyze N-isobutyl-4-chloropyrrolidone and its degradation impurities automatically using a CNN-LSTM dual model; The CNN-LSTM dual model is a deep learning model that combines a convolutional neural network (CNN) with a long short-term memory network (LSTM).

[0015] CNN is used to extract features of fluorescence chromatographic peaks (retention time, peak shape, half peak width), while LSTM is used to analyze the temporal features of mass spectrometry data (precise molecular weight, fragment ion abundance).

[0016] Preferably, in step S1, the pH of the ultrapure water is 6.5-7.5; the solid-phase extraction column is filled with 500 mg of aminopropyl bonded silica adsorbent, and the total volume of the column loaded with adsorbent is 6 mL.

[0017] Preferably, step S2 specifically involves: Step S21: The first dimension uses a hydrophilic interaction column, and the mobile phase is a mixed solution of acetonitrile and 10 mM ammonium acetate aqueous solution; collect the fraction after 8.5-9.5 min. Step S22: The second dimension uses a reversed-phase C18 column, with a mobile phase consisting of a mixture of 0.1% formic acid aqueous solution and methanol, and gradient elution is performed; the fraction collected in the second dimension is the sample solution B to be derivatized.

[0018] Preferably, in step S21, the volume ratio of acetonitrile to 10mM ammonium acetate aqueous solution in the mixed solution is 85:15, and the pH is 7.8-8.2.

[0019] Preferably, step S3 specifically involves reacting the sample solution B to be derivatized with 4-chloro-7-nitrobenzenefuran in a borate buffer solution, terminating the reaction after cooling in an ice-water bath for 5 minutes, and obtaining the derivatized sample solution.

[0020] Preferably, the volume ratio of the sample solution B to be derivatized, the borate buffer, and the 4-chloro-7-nitrobenzenefuran is 5:2:3; The pH of the borate buffer solution is 9.0, and the concentration of the borate buffer solution is 0.1 mol / L; the reaction temperature is 58-62℃, and the reaction time is 9-11 min.

[0021] Preferably, step S4 specifically involves detecting the derivatized sample solution using a fluorescence detector and quantifying it using the external standard method; then, using high-resolution mass spectrometry, confirming the structure by using the primary quasi-molecular ion peak and secondary characteristic fragment ions.

[0022] Preferably, the excitation wavelength of the fluorescence detector is 485 nm and the emission wavelength is 530 nm; The high-resolution mass spectrometry (HMS) was performed with a spray voltage of 3.5 kV, an ion source temperature of 300 °C, a drying gas flow rate of 8 L / min, and a scanning range of m / z 100-500.

[0023] Preferably, step S5 specifically involves: Step S51: Establish a standard spectral library; collect data from 500 sets of standard samples of N-isobutyl-4-chloropyrrolidone and its impurities to construct a standard spectral library; the training set accounts for 80% and the validation set accounts for 20%. Step S52, Model Training: Using standard sample data as input and target compound / impurity as output, train the model using the Adam optimizer for 100 iterations. Step S53: Based on the feature matching results, the model automatically marks the target compound peak and impurity peak, calculates the relative content of impurities, excludes residual 4-chloro-7-nitrobenzenefuran and methanol interference peaks, and outputs qualitative and quantitative reports.

[0024] Preferably, in step S51, the impurities include 4-hydroxy-N-isobutylpyrrolidone, cis-N-isobutyl-4-chloropyrrolidone, and N-isobutylpyrrolidone; the standard sample data includes chromatographic and mass spectrometric characteristics.

[0025] The present invention employs the above-mentioned analytical method for N-isobutyl-4-chloropyrrolidone, and its beneficial effects are as follows: (1) The present invention uses hydrophilic interaction chromatography (HILIC) coupled with reversed-phase C18 chromatography. The first-dimensional HILIC column is used to achieve preliminary separation of polar impurities, and the second-dimensional C18 column is used to further separate hydrophobic impurities. The separation degree between the target compound and adjacent impurities can be improved from 1.2-1.5 to more than 2.0, thereby meeting the separation requirements of complex matrices.

[0026] (2) By optimizing the derivatization conditions of 4-chloro-7-nitrobenzofuran (NBD-Cl), the present invention can achieve a derivatization efficiency of over 95% for the target compound. Combined with fluorescence detection, the detection limit is reduced to 0.1 ng / mL, which is 10-100 times more sensitive than conventional UV detection (1-10 ng / mL), thus enabling the monitoring of trace degradation products during storage.

[0027] (3) This invention combines the high-resolution mass spectrometry characteristics of high-resolution mass spectrometry (Q-TOF / MS) and can effectively distinguish structurally similar isomers through dual verification of primary quasi-molecular ion peaks and secondary characteristic fragment ions. At the same time, it integrates artificial intelligence peak recognition algorithms to achieve automatic identification of degradation impurities. Compared with manual identification, the efficiency is improved by more than 80%, and the false negative rate is <1%.

[0028] The technical solution of the present invention will be further described in detail below through embodiments. Detailed Implementation

[0029] The technical solution of the present invention will be further described below through embodiments.

[0030] Unless otherwise defined, the technical or scientific terms used in this invention shall have the ordinary meaning as understood by one of ordinary skill in the art to which this invention pertains.

[0031] Example 1 An analytical method for N-isobutyl-4-chloropyrrolidone includes the following steps: Step S1: Pre-treat the sample to be analyzed.

[0032] 0.5 mL of the sample to be analyzed was extracted with 2.0 mL of ultrapure water at pH 7.0 and purified by an aminopropyl-bonded silica solid-phase extraction column. The solid-phase extraction column was filled with 500 mg of aminopropyl-bonded silica adsorbent, and the total volume of the column loaded with adsorbent was 6 mL, resulting in sample solution A to be derived.

[0033] Step S2: Separate the sample solution A to be derivatized using two-dimensional chromatography to obtain the sample solution B to be derivatized.

[0034] Hydrophilic interaction chromatography was used as the first dimension and reversed-phase C18 chromatography as the second dimension in combination.

[0035] Step S21: The first dimension uses a hydrophilic interaction chromatographic column (HILIC) WatersXBridgeBEHAmide column (250mm×4.6mm, 5μm), column temperature 30±2℃, flow rate 1.0±0.1mL / min, injection volume 10μL.

[0036] The mobile phase was a mixed solution of acetonitrile and 10 mM ammonium acetate aqueous solution (volume ratio 85:15) with pH=8.0, and the fraction was collected after 9.0 min.

[0037] Step S22: The second dimension uses a reversed-phase C18 column, an Agilent ZORBAX Eclipse Plus C18 column (150mm × 4.6mm, 3.5μm), with a column temperature of 35±2℃, a flow rate of 0.8±0.1mL / min, and an injection volume of 20μL.

[0038] The mobile phase was a mixed solution of 0.1% formic acid aqueous solution and methanol, and gradient elution was performed. The gradient elution was as follows: 0-5min 80% 0.1% formic acid aqueous solution, 5-15min 80%→40% 0.1% formic acid aqueous solution, 15-20min 40% 0.1% formic acid aqueous solution, 20-21min 40%→80% 0.1% formic acid aqueous solution, 21-25min 80% 0.1% formic acid aqueous solution.

[0039] The fraction collected in the second dimension is the sample solution B to be derivatized.

[0040] Step S3: Derivatize the sample solution B to be derivatized to obtain the derivatized sample solution.

[0041] 0.5 mL of the sample solution B to be derivatized was reacted with 0.3 mL of 4-chloro-7-nitrobenzenefuran in 0.2 mL of borate buffer solution at pH 9.0 and a concentration of 0.1 mol / L. The reaction temperature was 60 °C and the reaction time was 10 min. The reaction was terminated after cooling in an ice-water bath for 5 min to obtain the derivatized sample solution.

[0042] Step S4: Detect the derivatized sample solution using a fluorescence detector and high-resolution mass spectrometry.

[0043] The derivatized sample solution was detected using a fluorescence detector, and quantification was performed using the external standard method. The excitation wavelength of the fluorescence detector was 485 nm, and the emission wavelength was 530 nm.

[0044] The structure was confirmed by high-resolution mass spectrometry (HMS) using primary quasi-molecular ion peaks and secondary characteristic fragment ions. The HMS setup included a spray voltage of 3.5 kV, an ion source temperature of 300 °C, a drying gas flow rate of 8 L / min, and a scan range of m / z 100–500.

[0045] Precise molecular weight matching: The theoretical molecular weight of the target derivatized product is 323.1, the characteristic ion peak of [M+H]+ is 324.1025 m / z, and the error between the measured value and the theoretical value is ≤5ppm.

[0046] Secondary fragment ion matching: Collision-induced dissociation was performed on m / z 324.1025 to obtain characteristic fragment ions m / z 289.1 (lost Cl atom), 213.0 (lost part of NBD group structure), and 156.0 (NBD group fragment). The abundance ratio of fragment ions deviated from the standard library by ≤10%.

[0047] Step S5: Analyze N-isobutyl-4-chloropyrrolidone and its degradation impurities by automatically identifying them using a CNN-LSTM dual model.

[0048] The CNN-LSTM dual model is a deep learning model that combines a convolutional neural network (CNN) with a long short-term memory network (LSTM).

[0049] Step S51: Establish a standard spectral library. Collect 500 sets of standard sample data (chromatographic and mass spectrometric characteristics) of N-isobutyl-4-chloropyrrolidone, 4-hydroxy-N-isobutylpyrrolidone, cis-N-isobutyl-4-chloropyrrolidone, and N-isobutylpyrrolidone to construct a standard spectral library. The training set accounts for 80% and the validation set accounts for 20%.

[0050] Step S52: Model Training. Using standard sample data as input and the target compound / impurity as output, the model is trained using the Adam optimizer for 100 iterations. In this embodiment, the model achieves an accuracy of 98.5% and an impurity identification recall rate of 99%.

[0051] Step S53: Based on the feature matching results, the model automatically marks the target compound peak and impurity peak, calculates the relative content of impurities, excludes the interfering peaks of 4-chloro-7-nitrobenzofuran (m / z 207.0) and methanol (m / z 33.0), and outputs qualitative and quantitative reports.

[0052] The qualitative results in this example show that two impurities were identified (unreacted raw material 4-chloropyrrolidone, content 0.12 mg / mL; byproduct N-isobutylpyrrolidone, content 0.08 mg / mL), with no misjudgments of interfering peaks, and the qualitative accuracy rate was 100%.

[0053] Quantitative results: The content of N-isobutyl-4-chloropyrrolidone in the reaction solution was 2.5 mg / mL, and the relative standard deviation (RSD, n=6) was 1.2%.

[0054] Example 2 This embodiment verifies the accuracy (recovery rate) and repeatability (precision) of the aforementioned analytical method through low, medium and high level spike recovery experiments, combined with precision calculations, ensuring that the method can meet the stringent requirements of quantitative analysis.

[0055] Blank matrix selection: A blank reaction solution was synthesized from a pharmaceutical intermediate that does not contain N-isobutyl-4-chloropyrrolidone and its impurities. It contains unreacted 4-chloropyrrolidone, residual zinc chloride catalyst, and ethanol solvent to simulate a real complex matrix.

[0056] Preparation of standard: Accurately weigh N-isobutyl-4-chloropyrrolidone standard and prepare standard stock solutions with concentrations of 0.1 μg / mL (low concentration), 1.0 μg / mL (medium concentration), and 10.0 μg / mL (high concentration) using ultrapure water (pH=7.0).

[0057] Spiked sample preparation: Take 9 blank matrix aliquots (0.5 mL each), divide them into 3 groups, and 3 parallel samples in each group. Add 0.5 mL of low, medium and high concentration standard stock solutions to each of the 3 groups to obtain spiked sample solutions with final spiked concentrations of 0.05 μg / mL, 0.5 μg / mL and 5.0 μg / mL. Prepare 6 parallel samples in each group (n=6) to ensure statistical validity.

[0058] The spiked sample solution was analyzed following steps S1-S5 of Example 1. The analytical results are shown in Table 1.

[0059] (1) Precision analysis: Calculate the relative standard deviation (RSD) of the results of each group of parallel samples. The smaller the RSD, the better the repeatability of the method. The industry standard requires RSD < 5%.

[0060] (2) Recovery rate analysis: Calculate the average recovery rate for each group. The closer the recovery rate is to 100%, the higher the accuracy of the method. The industry standard requires a recovery rate of 85%-115%.

[0061] The recovery rate (%) is calculated as follows: (measured concentration of spiked sample - background concentration of blank matrix) / theoretical spiked concentration × 100%. Since the background concentration of the blank matrix was detected as 0, the recovery rate (%) is calculated as: (measured concentration) / theoretical spiked concentration × 100%.

[0062] Table 1 Analysis Results

[0063] As shown in Table 1, the precision RSD of this embodiment is less than 3% at low, medium, and high spiking levels, far below the industry requirement of 5%, indicating excellent method repeatability. The average recovery rate is between 96.0% and 99.0%, close to 100%, and shows no significant concentration dependence, proving that the method has high accuracy and small systematic error, and can meet the accurate quantitative requirements for trace to macro-level N-isobutyl-4-chloropyrrolidone.

[0064] Example 3 In this embodiment, actual pharmaceutical intermediate samples containing multiple interfering substances were selected. By combining direct testing and spiked testing, the ability of the analytical method to eliminate interference and accurately identify target substances and impurities in complex matrices was verified.

[0065] The pharmaceutical intermediate synthesis reaction solution of N-isobutyl-4-chloropyrrolidone was selected, which contained the following interfering substances: 4-chloropyrrolidone, isobutylamine, N-isobutylpyrrolidone, 1,3-diisobutyl-4-chloropyrrolidone, zinc chloride, and ethanol.

[0066] Take two actual samples (0.5 mL each), one as the direct test sample, and the other as the spiked test sample with 0.5 μg / mL N-isobutyl-4-chloropyrrolidone standard and 0.2 μg / mL 4-hydroxy-N-isobutylpyrrolidone standard added.

[0067] The above-mentioned direct test samples and spiked test samples were analyzed in strict accordance with steps S1-S5 of Example 1.

[0068] (1) Results of direct test samples.

[0069] Chromatographic separation: Two-dimensional chromatograms showed that the retention times of the target analyte N-isobutyl-4-chloropyrrolidone were 9.0 min (first dimension) and 12.5 min (second dimension). The resolutions of the target analyte N-isobutyl-4-chloropyrrolidone (retention time 8.2 min) and the adjacent interfering analytes Isobutylamine (retention time 7.5 min) were both >2.5, with no peak overlap or baseline interference.

[0070] Mass spectrometry confirmation: High-resolution mass spectrometry detected the primary quasi-molecular ion peak of the target analyte at m / z 324.1023, and secondary fragment ions at m / z 289.1, 213.0, and 156.0, with a matching degree of >98% with the standard spectral library.

[0071] Model analysis: The CNN-LSTM dual model automatically eliminated non-responding interferences such as ethanol and zinc chloride, accurately identified the target substance and one natural impurity (N-isobutylpyrrolidone), with no false positives for interference peaks and a qualitative accuracy of 100%.

[0072] (2) Results of spiked test samples.

[0073] Anti-interference: The peak shape of the target analyte remained symmetrical after spiking, and did not overlap with interfering substances or residual derivatizing reagents in the matrix (4-chloro-7-nitrobenzenefuran, m / z 207.0).

[0074] Selectivity: The model accurately identified the spiked 4-hydroxy-N-isobutylpyrrolidone, whose mass spectrometry characteristic peak m / z 320.1 ([M+H]+) was significantly different from the target analyte, with no confusion. Recovery: The recovery rate of the spiked target analyte was 95.0%, with an RSD (n=3) of 2.1%.

[0075] The above analysis results show that this embodiment can effectively eliminate interference from unreacted raw materials, byproducts, catalysts, and solvents, achieving efficient separation of the target analyte from impurities. The high recovery rate and low RSD in the spiked validation demonstrate that the matrix did not affect the accuracy of the detection. In summary, the analytical method of this invention has excellent anti-interference capabilities and selectivity, making it fully applicable to the analysis and detection of complex samples.

[0076] Therefore, the present invention employs the above-mentioned analytical method for N-isobutyl-4-chloropyrrolidone. This analytical method, through its high sensitivity and high selectivity for the analysis of N-isobutyl-4-chloropyrrolidone containing chlorine and secondary amine groups in complex matrices, can be widely applied to the quality control of pharmaceutical intermediates, monitoring of organic synthesis processes, and purity detection of functional materials.

[0077] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention and not to limit them. Although the present invention has been described in detail with reference to preferred embodiments, those skilled in the art should understand that modifications or equivalent substitutions can still be made to the technical solutions of the present invention, and these modifications or equivalent substitutions cannot cause the modified technical solutions to deviate from the spirit and scope of the technical solutions of the present invention.

Claims

1. An analytical method for N-isobutyl-4-chloropyrrolidone, characterized in that, Includes the following steps: Step S1: Pre-treat the sample to be analyzed; The sample to be analyzed was extracted with ultrapure water and purified by an aminopropyl-bonded silica gel solid-phase extraction column to obtain sample solution A to be derived. Step S2: Separate the sample solution A to be derivatized using two-dimensional chromatography to obtain the sample solution B to be derivatized; Hydrophilic interaction chromatography was used as the first dimension and reversed-phase C18 chromatography as the second dimension in combination. Step S3: Derivatize the sample solution B to be derivatized to obtain a derivatized sample solution; Step S4: Detect the derivatized sample solution using a fluorescence detector and high-resolution mass spectrometry. Step S5: Analyze N-isobutyl-4-chloropyrrolidone and its degradation impurities automatically using a CNN-LSTM dual model; The CNN-LSTM dual model is specifically a deep learning model that combines a convolutional neural network and a long short-term memory network.

2. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 1, characterized in that: In step S1, the pH of the ultrapure water is 6.5-7.5; the solid phase extraction column is filled with 500 mg of aminopropyl bonded silica adsorbent, and the total volume of the column loaded with adsorbent is 6 mL.

3. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 1, characterized in that: Step S2 specifically involves: Step S21: The first dimension uses a hydrophilic interaction column, and the mobile phase is a mixed solution of acetonitrile and 10 mM ammonium acetate aqueous solution; collect the fraction after 8.5-9.5 min. Step S22: The second dimension uses a reversed-phase C18 column, with a mobile phase consisting of a mixture of 0.1% formic acid aqueous solution and methanol, and gradient elution is performed; the fraction collected in the second dimension is the sample solution B to be derivatized.

4. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 3, characterized in that: In step S21, the volume ratio of acetonitrile to 10mM ammonium acetate aqueous solution in the mixed solution is 85:15, and the pH is 7.8-8.

2.

5. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 1, characterized in that: Step S3 specifically involves reacting the sample solution B to be derivatized with 4-chloro-7-nitrobenzenefuran in a borate buffer solution, terminating the reaction after cooling in an ice-water bath for 5 minutes, and obtaining the derivatized sample solution.

6. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 5, characterized in that: The volume ratio of the derivatized sample solution B, borate buffer, and 4-chloro-7-nitrobenzenefuran is 5:2:3; The pH of the borate buffer solution is 9.0, and the concentration of the borate buffer solution is 0.1 mol / L; the reaction temperature is 58-62℃, and the reaction time is 9-11 min.

7. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 1, characterized in that: Step S4 specifically involves detecting the derivatized sample solution using a fluorescence detector and quantifying it using the external standard method; then, using high-resolution mass spectrometry, confirming the structure through primary quasi-molecular ion peaks and secondary characteristic fragment ions.

8. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 7, characterized in that: The excitation wavelength of the fluorescence detector is 485 nm, and the emission wavelength is 530 nm. The high-resolution mass spectrometry (HMS) was performed with a spray voltage of 3.5 kV, an ion source temperature of 300 °C, a drying gas flow rate of 8 L / min, and a scanning range of m / z 100-500.

9. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 1, characterized in that: Step S5 specifically involves: Step S51: Establish a standard spectral library; collect data from 500 sets of standard samples of N-isobutyl-4-chloropyrrolidone and its impurities to construct a standard spectral library; the training set accounts for 80% and the validation set accounts for 20%. Step S52, Model Training: Using standard sample data as input and target compound / impurity as output, train the model using the Adam optimizer for 100 iterations. Step S53: Based on the feature matching results, the model automatically marks the target compound peak and impurity peak, calculates the relative content of impurities, excludes residual 4-chloro-7-nitrobenzenefuran and methanol interference peaks, and outputs qualitative and quantitative reports.

10. The analytical method for N-isobutyl-4-chloropyrrolidone according to claim 9, characterized in that: In step S51, the impurities include 4-hydroxy-N-isobutylpyrrolidone, cis-N-isobutyl-4-chloropyrrolidone, and N-isobutylpyrrolidone; the standard sample data includes chromatographic and mass spectrometric characteristics.

Images