Separation qualitative and quantitative detection method for L-lactide, D-lactide and Meso-lactide

Abstract

The invention relates to a separation qualitative and quantitative detection method for L-lactide, D-lactide and Meso-lactide, and belongs to the field of lactide optical purity detection.The separation qualitative and quantitative detection method comprises the following steps that firstly, various lactide standard curves of different configurations are established respectively, the concentration range of the standard curves is 10-1000 mg / L, and the concentration range of the standard curves is 10-1000 mg / L; the plurality of lactides with different configurations comprise one or more of L-lactide, D-lactide and Meso-lactide; and the lactide with different configurations comprises one or more of the L-lactide, the D-lactide and the Meso-lactide. 2, detecting by using a gas chromatograph-mass spectrometer, and quantitatively calculating a lactide substance to be detected, wherein the concentration range of the lactide substance to be detected is 200-1000mg / L; according to the invention, separation and quantitative detection of three lactide optical isomers are realized by using an analysis method; the influence of impurities in the to-be-detected substance of lactide on the accuracy of the quantitative result of lactide is reduced; qualitative and semi-quantitative information of the impurities in the to-be-detected substance is provided.

Description

Technical Field

[0001] This invention relates to the field of optical purity detection of lactide, specifically to a method for the separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide. Background Technology

[0002] Since the 1950s, the global production of plastic products has exceeded 7.8 billion tons. Approximately 79% of these plastic products are landfilled or carelessly discarded, causing serious soil and marine pollution problems. Only about 21% has been properly disposed of through recycling and incineration. With the increasingly severe situation of "white pollution," countries around the world are actively promoting various laws and regulations to limit or ban plastics. Bio-based biodegradable materials to replace traditional plastics have become one of the new research hotspots in the global academic and industrial communities. Polylactic acid (PLA) is a bio-based biodegradable aliphatic polyester. Its unstable ester bonds can be hydrolyzed through the combined action of acid-base catalysis and microbial degrading enzymes, ultimately producing carbon dioxide and water. PLA possesses excellent mechanical properties, physical properties, biocompatibility, and air permeability, making it widely available in fields such as express packaging, textiles and apparel, biomedicine, and biodegradable agricultural films.

[0003] Ring-opening polymerization of lactide is currently the mainstream process for the synthesis of polylactic acid (PLA). It involves the dehydration and polycondensation of lactic acid to low molecular weight poly(D,L-lactic acid). Under high temperature and high vacuum conditions, this oligomeric D,L-lactic acid undergoes a reverse depolymerization reaction catalyzed by a tin-based catalyst to generate lactide. After purification by distillation, the lactide undergoes ring-opening polycondensation to produce the PLA product. Lactide is an important polymerization intermediate in ring-opening polymerization, and its chemical and optical purity directly affects the performance indicators and production costs of PLA products. During depolymerization, the randomly arranged L-lactic acid and D-lactic acid in the oligomeric lactide chain produce three optical isomers: L-lactide, D-lactide, and meso-lactide. Taking polylactic acid (PLLA) as an example, D-lactide and MESO-lactide impurities in L-lactide raw materials can reduce the crystallinity and crystallization rate of PLA, thereby weakening its mechanical properties, processing properties, melting point, glass transition temperature, and biodegradation rate. Therefore, the optical purity detection of lactide is of great significance for controlling the performance of PLA products and the industrial production cost.

[0004] Currently, the methods for detecting the optical purity of lactide mainly fall into the following categories:

[0005] CN113292531A and CN104837889A utilize nuclear magnetic resonance (NMR) to quantitatively integrate the methyl or methylene peaks of L-lactide and MESO-lactide based on their chemical shift differences, thereby obtaining the ratio of L-lactide to MESO-lactide. The disadvantages of this method are: 1H NMR cannot separate enantiomers L-lactide and D-lactide with the same chemical environment, and the quantitative accuracy is affected by the magnetic field strength and integration errors caused by the overlap of impurity peaks with the target peak. The methylene quartet of L-lactide and MESO-lactide partially overlaps at 5.2 ppm, making it unsuitable for precise quantification; although the methyl peak shows good separation at 1.5 ppm, it is easily interfered with by methyl signals from oligolactic acid, water, and impurities, resulting in larger quantitative errors for lactide samples with low chemical purity.

[0006] CN106153418A and CN114478470A utilize methyl polysiloxane and polyethylene glycol-based chromatographic columns for GC-FID quantitative analysis of L-lactide and MESO-lactide. The disadvantage of this method is that, according to comparative tests, its separation ability is poor, and a shoulder peak appears between the L-lactide and MESO-lactide signal peaks. The shoulder peak is qualitatively identified as lactide using mass spectrometry fragmentation information, and the integral assignment of the shoulder peak severely affects the quantitative accuracy. Specific rotation is another commonly used method for L / D-lactide ratio analysis, often used to supplement the above methods and compensate for their weak enantiomer separation ability. However, the optical rotation of oligolactic acid impurities in lactide affects the accuracy of specific rotation measurements. Therefore, existing analytical methods have poor separation effects and their quantitative accuracy is greatly affected by impurity content, making it difficult to meet the requirements for detecting the optical purity of lactide.

[0007] In summary, developing a method that can simultaneously and accurately determine the relative proportions and contents of L-lactide, D-lactide, and MESO-lactide is a technical problem that urgently needs to be solved in the research and development of lactide processes and product quality control. Summary of the Invention

[0008] To simultaneously and accurately determine the relative proportions and contents of L-lactide, D-lactide, and MESO-lactide, this invention proposes a method for the separation, qualitative, and quantitative detection of L-lactide, D-lactide, and Meso-lactide. This method effectively solves the problems of high separation difficulty of D-lactide, significant influence of lactide impurity content on quantitative accuracy, and limited qualitative information on impurities provided by existing technologies. It offers advantages such as high separation degree of lactide optical isomers, short detection time, low limit of quantitation, and high method reproducibility.

[0009] This invention is the first to use gas chromatography-mass spectrometry (GC-MS) to accurately determine the content of optical isomers of lactide. Compared with existing technologies, GC-MS has extremely strong structural identification and quantitative detection capabilities, and is more suitable for the precise quantitative analysis of lactide and the qualitative analysis of impurities.

[0010] The objective of this invention can be achieved through the following technical solutions:

[0011] A method for the separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide, characterized by comprising the following steps:

[0012] Step 1: Establish standard curves for various lactide configurations, with a concentration range of 10-1000 mg / L. The various lactide configurations include one or more of L-lactide, D-lactide, and Meso-lactide.

[0013] Step 2: Use gas chromatography-mass spectrometry to detect and quantitatively calculate the lactide analyte. The concentration range of the lactide analyte is 200-1000 mg / L.

[0014] The conditions for the gas chromatography-mass spectrometry technique are as follows:

[0015] The chromatographic column is a fully methylated β-cyclodextrin column with dimensions of (25-50)m × 0.25mm × 0.25μm.

[0016] Septum purge flow rate: 3 mL / min;

[0017] The column temperature is increased as follows: the initial temperature is 50-120℃, and the temperature is increased to 180-200℃ at a rate of 10-20℃ per minute, and held for 5-15 minutes.

[0018] Column equilibration time: 0.5-2 min;

[0019] Inlet temperature: 180-200℃;

[0020] The carrier gas is helium; the carrier gas flow rate is 1.0-2.0 mL / min;

[0021] The split ratio is 20-50:1;

[0022] The injection volume is 0.1-1 μL;

[0023] Mass spectrometry scanning mode: Total ion scan mode (TIC) or Selected ion scan mode (SIM);

[0024] Mass spectrometer transfer line temperature: 200-220℃;

[0025] Ion source temperature: 230℃; Quadrupole temperature: 150℃;

[0026] Solvent delay time: 4-6 min;

[0027] Scan quality range: 12-600 m / z;

[0028] Scanning speed: 1562u / s;

[0029] Select the ion scanning mode, and choose the mass-to-charge ratio (m / z) of the quantitative ion as 56, and the auxiliary quantitative ions as 144 and 45.

[0030] The ion residence time is 80-100ms.

[0031] Furthermore, in step one, when the relative proportion of a certain configuration of lactide is less than 1%, the concentration range of its standard curve is adjusted to 10-200 mg / L to reduce the fitting error of the lactide standard curve.

[0032] Furthermore, in step two, when the chemical purity P of the lactide analyte... chemical or the content of the main optical configuration P major optical When the concentration is below 50%, the concentration range of the lactide analyte is increased to 1 / P of the original concentration range. chemical Or 1 / P major optical times.

[0033] Furthermore, in step two, when the concentration of lactide analyte is below 7.0 mg / L, the mass spectrometry scanning mode adopts selected ion scanning mode.

[0034] Furthermore, in step two, the inner diameter of the chromatographic column is 0.25 mm, the length of the chromatographic column is 50 m, and the thickness of the stationary phase film is 0.25 μm.

[0035] Furthermore, the best solvent for dissolving various lactide configurations and lactide analytes is one or a mixture of two of methanol, ethanol, ethyl acetate, acetonitrile, acetone, and chloroform. Preferably, any one of acetonitrile, acetone, and chloroform is the most effective.

[0036] Furthermore, the solutions of various lactide compounds with different configurations and lactide analytes dissolved in a good solvent are filtered to remove insoluble impurities.

[0037] Furthermore, an organic filter membrane is selected during the filtration process, which is one or more of nylon membrane, polypropylene membrane, and polytetrafluoroethylene membrane with a filter diameter of 0.22-0.45 μm.

[0038] Furthermore, in step two, when organic acid residues are present in the lactide analyte, 20-50 μL of N-methyl-N-(trimethylsilane)trifluoroacetamide containing 1% trimethylchlorosilane is added during the dissolution of the lactide analyte, and a constant temperature water bath at 40-50°C is used for 5-15 minutes.

[0039] Furthermore, the lactide analyte is crude lactide that has not yet been purified by distillation.

[0040] Compared with the prior art, the present invention has the following beneficial effects:

[0041] (1) The present invention uses an analytical method to achieve the separation and quantitative detection of three optical isomers of lactide; reduces the influence of impurities in the lactide analyte on the accuracy of the lactide quantitative results; and provides qualitative and semi-quantitative information on impurities in the analyte.

[0042] (2) During the research process, the inventors discovered that under high temperature conditions, lactic acid oligomers in lactide will depolymerize during the test, causing lactide peak tailing and quantitative results to be too high. This invention reduces the depolymerization of lactic acid oligomers during the detection process by optimizing and screening gas chromatography conditions, thereby improving the accuracy of quantitative results.

[0043] (3) During the research process, the inventors discovered that the use of derivatization reagents, such as N-methyl-N-(trimethylsilane)trifluoroacetamide containing 1% trimethylchlorosilane, can greatly improve the retention behavior of organic acids on fully methylated β-cyclodextrin chromatographic columns. The present invention can also quantitatively measure lactic acid, a monomer synthesized from lactide.

[0044] (4) The present invention is more suitable for purity testing of crude lactide that has not yet been purified by distillation. The analytical method provided by the present invention has unique advantages in terms of data support for lactide process optimization and product quality control. Attached Figure Description

[0045] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the accompanying drawings required in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the accompanying drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0046] Figure 1 This is the standard curve for L-lactide using the external standard method.

[0047] Figure 2 This is the standard curve for D-lactide using the external standard method.

[0048] Figure 3 This is the standard curve for the Meso-lactide external standard method.

[0049] Figure 4 The TIC chromatograms for the separation of lactide samples using a cyclodextrin column in Example 1 are shown, from left to right: Meso-lactide, L-lactide, and D-lactide.

[0050] Figure 5 This is the mass spectrum of the Meso-lactide sample from Example 1;

[0051] Figure 6 This is the mass spectrum of the L-lactide sample from Example 1;

[0052] Figure 7 This is the mass spectrum of the D-lactide sample from Example 1;

[0053] Figure 8 The TIC chromatogram of the lactide sample separated using a polyethylene glycol column in Comparative Example 1;

[0054] Figure 9 for Figure 8 Mass spectra of shoulder peaks appearing from 4.2 minutes to 25.7 minutes on the horizontal axis;

[0055] Figure 10 This is a GCMS TIC plot of polylactic acid thermal pyrolysis.

[0056] Figure 11 for Figure 10 The corresponding mass spectrum and spectral library search results for TIC in China;

[0057] Figure 12 Example 2: Polylactic acid thermal pyrolysis-GCMS TIC diagram;

[0058] Figure 13 for Figure 12 The mass spectrum corresponding to TIC and the results of the spectral library search. Detailed Implementation

[0059] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments. These embodiments are implemented based on the technical solution of the present invention, providing detailed implementation methods and specific operating procedures. However, the scope of protection of the present invention is not limited to the following embodiments.

[0060] Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without inventive effort are within the scope of protection of this invention. Where specific techniques or conditions are not specified in the embodiments, they shall be performed according to the techniques or conditions described in the literature in this field or the conditions recommended by the manufacturer. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels. Unless otherwise specified, the experimental methods in the following embodiments are conventional methods. Unless otherwise specified, the experimental materials used in the following embodiments are all commercially available products.

[0061] The endpoints and any values ​​of the ranges disclosed in this invention are not limited to the precise ranges or values, and these ranges or values ​​should be understood to include values ​​close to these ranges or values. For numerical ranges, the endpoint values ​​of the various ranges, the endpoint values ​​of the various ranges and individual point values, and individual point values ​​can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed in this invention.

[0062] To overcome the challenges of high separation difficulty of D-lactide, significant influence of lactide impurity content on quantitative accuracy, and limited qualitative information on impurities provided by existing technologies, this invention provides a method for the simultaneous and accurate separation, qualitative, and quantitative detection of L-lactide, D-lactide, and MESO-lactide, including their relative proportions and contents. This method comprises the following steps:

[0063] Step 1: Establish standard curves for lactide with different configurations, with a concentration range of 10-1000 mg / L.

[0064] Among them, the various configurations of lactide include one or more of L-lactide, D-lactide, and Meso-lactide;

[0065] Specifically, establishing a standard curve involves the following steps:

[0066] (1) Weigh a quantitative amount of a lactide standard with a certain configuration, add a good solvent, vortex to dissolve, and transfer to a volumetric flask and dilute to volume. Prepare six concentration gradient lactide standard solutions by stepwise dilution. The good solvent for dissolving the various lactide configurations is one or a mixture of methanol, ethanol, ethyl acetate, acetonitrile, acetone, and chloroform. Preferably, any one of acetonitrile, acetone, and chloroform is the most effective.

[0067] (2) The lactide standard solution obtained in step (1) is filtered through an organic filter membrane and the filtrate is collected. The organic filter membrane is one of nylon membrane, polypropylene membrane, or polytetrafluoroethylene membrane with a filter diameter of 0.22-0.45 μm.

[0068] (3) Use gas chromatography-mass spectrometry to detect the filtrate in step (2) and establish a standard curve of lactide through data processing; the concentration range of the lactide standard curve is 10-1000 mg / L. When the relative proportion of a certain configuration of lactide is less than 1%, the concentration range of its standard curve is adjusted to 10-200 mg / L to reduce the fitting error of the lactide standard curve.

[0069] Step 2: Use gas chromatography-mass spectrometry to detect and quantitatively calculate the lactide analyte. The concentration range of the lactide analyte is 200-1000 mg / L.

[0070] Specifically, the detection and quantitative calculation of lactide analytes include the following steps: weighing a quantitative amount of lactide analyte, adding a quantitative amount of good solvent to dissolve it by vortexing, filtering it with an organic filter membrane, collecting the filtrate, detecting it with gas chromatography-mass spectrometry and performing quantitative calculations. The selection criteria for good solvents and organic filter membranes are consistent with the above conditions.

[0071] Among them, when the concentration range of the lactide analyte is 200-1000 mg / L, and when the chemical purity P of the lactide analyte is... chemical or the content of the main optical configuration P major optical When the concentration is below 50%, the concentration range of the lactide analyte is increased to 1 / P of the original concentration range. chemical Or 1 / P major optical times.

[0072] The conditions for gas chromatography-mass spectrometry (GC-MS) are as follows:

[0073] Gas chromatography column: Fully methylated β-cyclodextrin column (25-50m × 0.25mm × 0.25μm);

[0074] Septum purge flow rate: 3 mL / min;

[0075] The initial temperature of the chromatographic column is 80-120℃, and it is increased to 180-200℃ at a rate of 10-20℃ per minute, and held for 5-15 minutes.

[0076] Column equilibration time: 0.5-2 minutes;

[0077] Inlet temperature: 180-200℃;

[0078] The carrier gas is helium;

[0079] The carrier gas flow rate is 1.0-2.0 mL / min;

[0080] The split ratio is 20-50:1;

[0081] The injection volume is 0.1-1 μL;

[0082] Mass spectrometry scanning mode: Total ion scan mode (TIC) or Selected ion scan mode (SIM);

[0083] Mass spectrometer transfer line temperature: 200-220℃;

[0084] Ion source temperature: 230℃;

[0085] Quadrupole temperature: 150℃;

[0086] Solvent delay time: 4-6 minutes;

[0087] Scan quality range: 12-600 m / z;

[0088] Scanning speed: 1562u / s;

[0089] SIM mode, quantitative ion selection mass-to-charge ratio (m / z): 56, auxiliary quantitative ions: 144, 45;

[0090] The ion residence time is 80-100ms.

[0091] In a preferred embodiment of the present invention, when the concentration of lactide to be measured is less than 7.0 mg / L, the mass spectrometry scanning mode adopts selected ion scanning mode.

[0092] In a preferred embodiment of the present invention, the inner diameter of the chromatographic column is 0.25 mm, the length of the chromatographic column is 50 m, and the thickness of the stationary phase film is 0.25 μm.

[0093] In a preferred embodiment of the present invention, when organic acid residue is present in the analyte, 20-50 μL of N-methyl-N-(trimethylsilane)trifluoroacetamide containing 1% trimethylchlorosilane is added during the dissolution of the analyte and the solution is dissolved in a constant temperature water bath at 40-50°C for 5-15 minutes, preferably 10 minutes.

[0094] In a preferred embodiment of the present invention, the lactide analyte is crude lactide that has not yet been purified by distillation.

[0095] During the research process, the inventors discovered that under high temperature conditions, lactic acid oligomers in lactide undergo depolymerization during testing, resulting in lactide peak tailing and overestimation of quantitative results. This invention reduces the depolymerization of lactic acid oligomers during detection by optimizing and screening gas chromatography conditions, thereby improving the accuracy of quantitative results.

[0096] Specifically, the optimization of this invention is determined by combining the GC-MS overflow gas analysis results of low molecular weight polylactic acid, i.e., a lactide synthesis intermediate, with the boiling point of lactide, such as... Figure 10As shown, the oligolactic acid sample was thermally pyrolyzed in the pyrolysis chamber at an initial temperature of 50°C and a heating rate of 40°C / min. After pyrolysis, the fragments were directly separated by mass spectrometry without being separated by a chromatographic column. Figure 11 The mass spectrum and library search results corresponding to TIC confirm that the main decomposition product of oligolactic acid is lactide.

[0097] In existing technologies, the vaporization temperature at the injection port is typically set at 220-250℃. Figure 10 Peak elution time calculations show that when the pyrolysis chamber temperature reaches 250℃, 11.34% of oligolactic acid depolymerizes to form lactide. The lactide produced by this depolymerization directly leads to falsely high analyte quantification results. Therefore, lowering the heating temperature can effectively reduce the amount of lactic acid oligomers decomposed. When the temperature is lowered to 180℃, only 0.92% of oligolactic acid depolymerizes. Furthermore, a lower injection port temperature is not always better. The lower limit of the injection port temperature should ensure sufficient lactide vaporization. Injection port temperature optimization was achieved by gradually decreasing the injection port temperature from 250℃ in increments of 10℃ and observing the stability of the analyte peak effect.

[0098] The methodological verification is as follows: :

[0099] This invention validates the above-mentioned detection method in terms of linear range, limit of detection and limit of quantitation, injection precision, method validation, and spiked recovery rate.

[0100] Chromatographic and mass spectrometry conditions:

[0101] Gas chromatography column: Agilent CP-Cyclodextrin-β-2,3,6-M-19 (50m × 0.25mm × 0.25μm);

[0102] Septum purge flow rate: 3 mL / min;

[0103] The initial temperature of the chromatographic column was 120℃, held for 2 minutes, and then increased to 180℃ at a rate of 10℃ per minute, held for 15 minutes.

[0104] Column equilibration time: 0.5 minutes;

[0105] Inlet temperature: 200℃;

[0106] The carrier gas is helium; the carrier gas flow rate is 2 mL / min.

[0107] The split ratio is 50:1;

[0108] The injection volume was 1.0 μL;

[0109] Mass spectrometry scanning mode: Total ion scan mode (TIC);

[0110] Mass spectrometer transfer line temperature: 200℃;

[0111] Ion source temperature: 230℃;

[0112] Quadrupole temperature: 150℃;

[0113] Scan quality range: 12-600 m / z;

[0114] Scanning speed: 1562u / s.

[0115] (1) Linearity and range test

[0116] Solution preparation: Accurately weigh 50.0 mg of L-lactide, D-lactide, and MESO-lactide and place them separately in beakers. Add 20.00 mL of acetonitrile and stir to dissolve. Transfer the solution to a 50.00 mL volumetric flask, make up to volume, and shake to mix to obtain a 1000 mg / L standard stock solution. Prepare standard solutions of 500.0, 200.0, 100.0, 50.0, and 10.00 mg / L by serial dilution.

[0117] Determination method: Inject 1.0 μL of each of the above solutions once and record the chromatographic and mass spectrometric data.

[0118] Experimental results: Within the linear range of 10.00-1000 mg / L,

[0119] The linear relationship between L-lactide concentration and peak area is excellent, R 2 The value is greater than 0.999, and the linear equation is y = 10104.22x + 119395.90;

[0120] D-lactide exhibits excellent linearity between concentration and peak area, R 2 The value is greater than 0.999, and the linear equation is y = 10550.59x + 74092.93;

[0121] Meso-lactide exhibits excellent linearity between concentration and peak area, R 2 The value is greater than 0.999, and the linear equation is y = 8905.57x + 124059.54. (See...) Figure 1-3 .

[0122] (2) Limit of detection and limit of quantitation test

[0123] Solution preparation: Take 10.00 mg / L L-lactide standard solution and dilute it stepwise with acetonitrile to a final volume.

[0124] Determination method: Inject 1 μL of each of the above solutions once and record the chromatographic and mass spectrometric data. The signal-to-noise ratio (SNR) value of L-lactide was calculated using the signal-to-noise ratio (SNR) calculation function of Agilent Masshunter 10.0 software.

[0125] The solution with an SNR value of approximately 10 was used as the limit of quantitation solution, and the solution with an SNR value of approximately 3 was used as the limit of detection solution.

[0126] Experimental results: The calculated SNR value of the standard solution with an L-lactide concentration of 7.00 mg / L was 10.3, and the calculated SNR value of the standard solution with an L-lactide concentration of 2.00 mg / L was 3.1.

[0127] Therefore, the detection limit of the detection method provided by the present invention is 2.00 mg / L and the quantitation limit is 7.00 mg / L. This shows that the present invention has high sensitivity and can meet the quantitative requirements for trace isomers in lactide.

[0128] (3) Sample injection precision test

[0129] Solution preparation: Accurately weigh 250.0 mg of lactide sample, place it in a beaker, add 50.00 mL of acetonitrile and stir to dissolve; transfer the solution to a 250.0 mL volumetric flask, make up to volume, and shake to mix to obtain a 1000 mg / L test solution.

[0130] Determination method: Inject 1.0 μL of each of the above solutions, and inject continuously for 6 times, recording the chromatographic and mass spectrometric data.

[0131] Experimental results: The test results are shown in Table 1. The D-lactide content in the sample was lower than the limit of quantification, so the RSD% was higher than that of the other two configurations. This shows that the injection precision of the present invention is good, and the RSD% of the lactide peak area is less than 2%.

[0132] Table 1

[0133]

[0134] (4) Spike recovery test

[0135] Solution preparation: Six lactide samples were randomly selected. Two 25.0 mg portions of each sample were accurately weighed as the spiked recovery group and the background group. The spiked recovery group was placed in a beaker and 10.0 mg of L-lactide and 50.00 mL of acetonitrile were added and stirred to dissolve. The other portion was placed in a beaker and 50.00 mL of acetonitrile was added and stirred to dissolve.

[0136] Determination method: Inject 1.0 μL of each of the above solutions once and record the chromatographic and mass spectrometric data.

[0137] Experimental results: The test results are shown in Table 2. The average recovery rate of L-lactide spiked was 101.7±6.47%, which proves that the method is accurate and reliable.

[0138] Table 2

[0139]

[0140]

[0141] Example 1: Crude L-lactide sample

[0142] This embodiment analyzes the content of three lactide optical isomers in a crude L-lactide sample, which was synthesized by the inventors using commercially available L-lactic acid raw material via ring-opening polymerization.

[0143] Chromatographic and mass spectrometry conditions:

[0144] Gas chromatography column: Agilent CP-Cyclodextrin-β-2,3,6-M-19 (50m × 0.25mm × 0.25μm);

[0145] Septum purge flow rate: 3 mL / min;

[0146] The initial temperature of the chromatographic column was 50℃, held for 2 minutes, and then increased to 180℃ at a rate of 10℃ per minute, held for 15 minutes.

[0147] Column equilibration time: 0.5 minutes;

[0148] Inlet temperature: 200℃;

[0149] The carrier gas is helium;

[0150] The carrier gas flow rate is 1 mL / min;

[0151] The split ratio is 50:1;

[0152] The injection volume was 1.0 μL;

[0153] Mass spectrometry scanning mode: Total ion scan mode (TIC);

[0154] Mass spectrometer transfer line temperature: 200℃;

[0155] Ion source temperature: 230℃;

[0156] Quadrupole temperature: 150℃;

[0157] Scan quality range: 33-550 m / z;

[0158] Scanning speed: 1562u / s.

[0159] Solution preparation: Accurately weigh 50.0 mg of lactide sample, place it in a beaker, add 50.00 mL of acetonitrile and stir to dissolve; transfer the solution to a 50.00 mL volumetric flask, make up to volume, and shake to mix to obtain a 1000 mg / L test solution.

[0160] Determination method: Inject 1.0 μL of each of the above solutions once and record the chromatographic and mass spectrometric data.

[0161] Experimental results: See the spectrum. Figure 4-7 The quantitative results of the samples are shown in Table 3.

[0162] Table 3

[0163]

[0164] Example 2: Refined lactide

[0165] This embodiment analyzes the content of three lactide optical isomers in a purified L-lactide sample. The purified lactide sample was prepared by the inventors using commercially available L-lactic acid raw material through ring-opening polymerization and purified by distillation.

[0166] Chromatographic and mass spectrometry conditions: The chromatographic and mass spectrometry conditions are the same as those in Example 1.

[0167] Solution preparation: Accurately weigh 50.0 mg of lactide sample, place it in a beaker, add 50.00 mL of acetonitrile and stir to dissolve; transfer the solution to a 50.00 mL volumetric flask, make up to volume, and shake to mix to obtain a 1000 mg / L test solution.

[0168] Determination method: Inject 1.0 μL of each of the above solutions once and record the chromatographic and mass spectrometric data.

[0169] Experimental results: The quantitative results of the samples are shown in Table 4.

[0170] Table 4

[0171]

[0172] Comparative Example 1

[0173] In this comparative example, a polyethylene glycol column was used to analyze the lactide sample, and the chromatographic conditions were consistent with the method described in CN114478470A.

[0174] Chromatographic and mass spectrometry conditions:

[0175] Gas chromatography column: Agilent HP-INNOWAX (60m × 0.25mm × 0.25μm);

[0176] Septum purge flow rate: 3 mL / min;

[0177] The initial temperature of the chromatographic column was 100℃, and it was increased to 140℃ at a rate of 4℃ per minute, and then increased to 180℃ at a rate of 8℃ per minute.

[0178] Column equilibration time: 0.5 minutes;

[0179] Inlet temperature: 180℃; carrier gas: helium;

[0180] The carrier gas flow rate is 2 mL / min; the split ratio is 50:1.

[0181] The injection volume was 1.0 μL;

[0182] Mass spectrometry scanning mode: Total ion scan mode (TIC);

[0183] Mass spectrometer transfer line temperature: 200℃;

[0184] Ion source temperature: 230℃;

[0185] Quadrupole temperature: 150℃;

[0186] Scan quality range: 12-600 m / z;

[0187] Scanning speed: 1562u / s.

[0188] Determination method: Inject 1.0 μL of each of the above solutions once and record the chromatographic and mass spectrometric data.

[0189] Experimental results: See attached spectrum. Figure 8-9 , Figure 8 A distinct acromion appeared between 24.2 and 25.7 minutes, such as... Figure 9 As shown, the mass-to-charge ratio and relative abundance of shoulder peak fragments in mass spectrometry are consistent with those of lactide, and the attribution of shoulder peaks seriously affects the accuracy of quantitative integration.

[0190] Example 2

[0191] This embodiment analyzes an L-lactide sample containing lactic acid, which was synthesized by the inventors using commercially available L-lactic acid raw material via ring-opening polymerization.

[0192] Chromatographic and mass spectrometry conditions:

[0193] Gas chromatography column: Agilent CP-Cyclodextrin-β-2,3,6-M-19 (50m × 0.25mm × 0.25μm);

[0194] Septum purge flow rate: 3 mL / min;

[0195] The initial temperature of the chromatographic column was 80℃, held for 2 minutes, and then increased to 180℃ at a rate of 10℃ per minute, held for 15 minutes.

[0196] Column equilibration time: 0.5 minutes;

[0197] Inlet temperature: 200℃;

[0198] The carrier gas is helium;

[0199] The carrier gas flow rate is 1.5 mL / min;

[0200] The split ratio is 50:1;

[0201] The injection volume was 1.0 μL;

[0202] Mass spectrometry scanning mode: Total ion scan mode (TIC);

[0203] Mass spectrometer transfer line temperature: 200℃;

[0204] Ion source temperature: 230℃;

[0205] Quadrupole temperature: 150℃;

[0206] Scan quality range: 28-550 m / z;

[0207] Scanning speed: 1562u / s.

[0208] Solution preparation: Accurately weigh two 50.0 mg lactide samples, place them in sample bottles, add 5.00 mL of acetonitrile to each sample and stir to dissolve. Take one sample and add 100 μL of N-methyl-N-(trimethylsilane)trifluoroacetamide solution containing 1% trimethylchlorosilane and stir magnetically at 70 °C for 45 minutes to obtain lactide standard and lactide derivatized sample.

[0209] Assay method: Inject 1.0 μL of each of the above solutions once, and record the chromatographic and mass spectrometric data. See [link to data]. Figure 12 .

[0210] like Figure 12-13 As shown, the lactic acid retention behavior of the lactide sample (black) without the addition of BSTFA derivatization reagent was poor, and its decomposition products formed a broad band between 8 and 10 minutes. In contrast, the lactide sample (red) after derivatization showed a well-shaped methylsilanized lactic acid peak at 8.55 minutes, and the M-lactide and L-lactide peaks at 14.56 minutes and 16.26 minutes remained unchanged. This indicates that the derivatization pretreatment process does not cause degradation of the lactide sample. Therefore, this example proves that the lactide GC-MS test method can be combined with BSTFA derivatization for lactic acid analysis.

[0211] In the description of this specification, references to terms such as "an embodiment," "example," "specific example," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0212] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention in any other way. Any person skilled in the art may make changes or modifications to the above-disclosed technical content to create equivalent embodiments. However, any simple modifications, equivalent changes, and modifications made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the protection scope of the present invention.

Claims

1. A method for the separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide, characterized in that, Includes the following steps: Step 1: Establish standard curves for various lactide configurations, with a concentration range of 10-1000 mg / L. The various lactide configurations include one or more of L-lactide, D-lactide, and Meso-lactide. Step 2: Use gas chromatography-mass spectrometry to detect and quantitatively calculate the lactide analyte. The concentration range of the lactide analyte is 200-1000 mg / L. The conditions for the gas chromatography-mass spectrometry technique are as follows: The chromatographic column is a fully methylated β-cyclodextrin column with dimensions of (25-50)m × 0.25mm × 0.25μm. Septum purge flow rate: 3 mL / min; The column temperature is increased as follows: the initial temperature is 50-120℃, and the temperature is increased to 180-200℃ at a rate of 10-20℃ per minute, and held for 5-15 minutes. Column equilibration time: 0.5-2 min; Inlet temperature: 180-200℃; The carrier gas is helium; The carrier gas flow rate is 1.0-2.0 mL / min; The split ratio is 20-50:1; The injection volume is 0.1-1 μL; Mass spectrometry scanning mode: Total ion scan mode or Selected ion scan mode; Mass spectrometer transfer line temperature: 200-220℃; Ion source temperature: 230℃; Quadrupole temperature: 150℃; Solvent delay time: 4-6 min; Scan quality range: 12-600 m / z; Scanning speed: 1562u / s; Select ion scanning mode, mass-to-charge ratio of quantitative ions: 56, auxiliary quantitative ions: 144, 45; The ion residence time is 80-100ms.

2. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, In step one, when the relative proportion of a certain configuration of lactide is less than 1%, the concentration range of its standard curve is adjusted to 10-200 mg / L to reduce the fitting error of the lactide standard curve.

3. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, In step two, when the chemical purity P of the lactide analyte is... chemical or the content of the main optical configuration P majoroptical When the concentration is below 50%, the concentration range of the lactide analyte is increased to 1 / P of the original concentration range. chemical Or 1 / P majoroptical times.

4. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, In step two, when the concentration of lactide analyte is below 7.0 mg / L, the mass spectrometry scanning mode is selected ion scanning mode.

5. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, In step two, the inner diameter of the chromatographic column is 0.25 mm, the length of the chromatographic column is 50 m, and the thickness of the stationary phase film is 0.25 μm.

6. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, A good solvent for dissolving various lactide compounds with different configurations and lactide analytes is one or a mixture of two of methanol, ethanol, ethyl acetate, acetonitrile, acetone, and chloroform.

7. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 6, characterized in that, Various lactide compounds with different configurations and lactide analytes were dissolved in a good solvent, and the resulting solutions were filtered to remove insoluble impurities.

8. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 7, characterized in that, An organic filter membrane is selected during the filtration process. The organic filter membrane is one or more of nylon membrane, polypropylene membrane, and polytetrafluoroethylene membrane with a filter diameter of 0.22-0.45 μm.

9. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, In step two, when organic acid residues are present in the lactide analyte, 20-50 μL of N-methyl-N-(trimethylsilane)trifluoroacetamide containing 1% trimethylchlorosilane is added during the dissolution of the lactide analyte, and a constant temperature water bath at 40-50°C is used for 5-15 minutes.

10. The method for separation, qualitative and quantitative detection of L-lactide, D-lactide and Meso-lactide according to claim 1, characterized in that, The lactide analyte was crude lactide that had not yet been purified by distillation.

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