Pre-column derivatization-gas chromatography detection of related substances in D-(+)-gluconic acid delta-lactone

Abstract

The invention provides precolumn derivatization-gas chromatography detection of related substances in D-(+)-gluconic acid delta-lactone, and relates to the technical field of pharmaceutical analysis. The detection method comprises the following steps: dissolving an impurity standard substance in a solvent to prepare an impurity stock solution; adding a derivatization reagent and carrying out ultrasonic treatment to obtain a reference solution; mixing a to-be-detected sample with the impurity stock solution, adding a derivatization reagent, and performing ultrasonic treatment to obtain a resolution solution; mixing a to-be-detected sample with a solvent, adding a derivatization reagent, and performing ultrasonic treatment to obtain a test solution; respectively detecting and analyzing the contrast solution, the resolution solution and the test solution by adopting GC-FID (Gas Chromatography-Flame Ionization Detector), and recording chromatograms; according to the present invention, the sample treatment is simple, the specificity, the sensitivity, the precision and the durability of the method are good, and the various impurities contained in the D-glucose-delta-lactone can be effectively and simultaneously detected so as to achieve the quality research on the liflozin starting material D-(+)-gluconic acid delta-lactone and the quality control on the finished product.

Description

Technical Field

[0001] The invention relates to the technical field of drug analysis, and in particular to a pre-column derivatization-gas chromatography detection of related substances in D-(+)-glucono delta-lactone. Background Art

[0002] D-(+)-Glucono-δ-lactone (abbreviated as glucono-δ-lactone, GDL) is a white crystal or crystalline powder, which is easily soluble in water, slightly soluble in ethanol, and insoluble in ether. Its aqueous solution slowly hydrolyzes to form gluconic acid at room temperature or lower temperature, and the hydrolysis speeds up as the temperature rises. GDL is widely used in the food field as an additive such as a coagulant, acidifier, and improver. In addition, it is also widely used as a starting material for the synthesis of hypoglycemic drugs such as glibenclamide.

[0003] Glycemic control drugs are sodium-glucose co-transporter 2 (SGLT2) inhibitors used to treat type 2 diabetes. Typical examples include dapagliflozin, empagliflozin and canagliflozin. GDL is the key starting material for the preparation of Glycemic Control API (CAS No.: 90-80-2), and its structural formula is as follows:

[0004]

[0005] Industrial preparation of GDL is mostly based on the oxidation of glucose to gluconic acid, which is converted through an acidification process. During the preparation of GDL, the following impurities may remain:

[0006]

[0007] These impurities may remain or be derived and transferred into the API, affecting the safety of the drug. In order to ensure the safety and effectiveness of the clinical use of gliflozin APIs, a method for the determination of GDL-related substances with high sensitivity and good selectivity is needed for the impurities in GDL.

[0008] At present, the main methods for detecting GDL-related substances that have been disclosed are as follows:

[0009] The national standard method GB7657-2020 discloses a method for detecting reducing sugars, but this method has low sensitivity and poor specificity and does not meet current quality research requirements.

[0010] Chinese patent CN 117147736 A provides a method for detecting related substances of D-glucose-δ-lactone, using a HILIC column analysis method, and only studies two related substances, D-glucono-1,4-lactone and D-gluconic acid.

[0011] Chinese patent CN 116242951 A discloses an analytical method for determining related substances of gluconolactone, (1) preparing a test solution: taking a test sample and adding a diluent to dissolve it, quantitatively diluting it to a set concentration to obtain a test solution; (2) setting chromatographic conditions: using a chromatographic column with a triple-bonded amide-bonded silica gel as a filler, using a Hilic separation mode, using an acidic aqueous solution as mobile phase A, an organic solvent as mobile phase B, performing isocratic elution, and detecting a wavelength of 205nm-220nm; (3) determination and analysis: taking 10μL-50μL of the test solution and injecting it into a high performance liquid chromatograph, recording a chromatogram, and calculating the content of gluconolactone and its related substances gluconolactone and gluconic acid by the area normalization method. This method can detect a variety of related substances, but the method solution has poor stability, and the use of a CAD detector is expensive, which is not conducive to implementation in general laboratories.

[0012] In summary, the research on GDL related substances is mainly analyzed by high performance liquid chromatography using detectors such as ultraviolet and CAD. The existing technology still has many defects and is difficult to meet the quality research needs of general laboratories. Developing a method for detecting related substances in D-(+)-glucono delta-lactone with high sensitivity and strong applicability is the research focus of researchers in this field. Summary of the invention

[0013] In view of the above problems, the present invention provides a gas chromatography detection method for related substances in D-(+)-glucono delta-lactone, which is a pre-column derivatization-gas chromatography method. By optimizing the diluent, derivatization conditions, chromatographic conditions, etc., the content of impurities such as D-glucose, D-gluconic acid, sucrose, D-glucono-1,4-lactone in D-(+)-glucono delta-lactone is detected, and the operation is simple and the sensitivity is high.

[0014] To achieve the above purpose, the technical solution adopted by the present invention is as follows:

[0015] In one aspect, the present invention provides a method for detecting related substances in D-(+)-glucono delta-lactone, which is a pre-column derivatization-gas chromatography detection method, comprising the following steps:

[0016] S1: Preparation of impurity stock solution: D-glucose, D-gluconic acid, sucrose, and D-gluconic acid-1,4-lactone are dissolved in a solvent to prepare an impurity stock solution;

[0017] S2: Preparation of control solution: Take the impurity stock solution, add the derivatization reagent, react for 10-50 minutes, and obtain the control solution;

[0018] S3: Preparation of separation solution: Mix the D-(+)-glucono δ-lactone sample to be tested and the impurity stock solution, add a derivatization reagent, and react for 10-50 minutes to obtain a separation solution;

[0019] S4: Preparation of test solution: Take the D-(+)-glucono delta-lactone sample to be tested and mix it with the solvent.

[0020] Add derivatization reagent and react for 10-50 minutes to obtain the test solution;

[0021] S5: Take the control solution, separation solution and test solution, respectively, and use GC-FID to detect and analyze them, and record the chromatograms;

[0022] The chromatographic conditions are:

[0023] Chromatographic column: capillary column with 100% dimethylpolysiloxane as stationary liquid;

[0024] Inlet temperature: 230-300℃;

[0025] Carrier gas: helium, flow rate 1.3-1.7 mL / min;

[0026] FID temperature: 280-350℃;

[0027] Injection volume: 1-2 μL;

[0028] The split ratio is 50-100:1;

[0029] Heating program:

[0030] Initial temperature: 150-165°C, hold for 0-3 minutes;

[0031] Heat to 170-190°C at a rate of 1-5°C / min and hold for 0-5min;

[0032] The temperature was raised to 280-320°C at a heating rate of 15-30°C / min and maintained for 1-10 min.

[0033] Preferably, in S1 and S4, the solvent is selected from at least one of N,N-dimethylformamide, acetonitrile and dichloromethane; further preferably, in S1 and S4, the solvent is N,N-dimethylformamide.

[0034] Preferably, in S2 and S3, the volume of the impurity stock solution is 1-3 mL; further preferably, in S2 and S3, the volume of the impurity stock solution is 1 mL;

[0035] Preferably, in S4, the volume of the solvent is 1-3 mL; further preferably, in S4, the volume of the solvent is 1 mL;

[0036] Preferably, in S2-S4, the derivatization reagent is a mixture of N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA), trimethylchlorosilane (TMCS) and pyridine;

[0037] Preferably, the volume ratio of N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA), trimethylchlorosilane (TMCS) and pyridine is 6-9: 0.5-1.5: 0.1-0.5; further preferably, the volume ratio of N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA), trimethylchlorosilane (TMCS) and pyridine is 7-9: 0.5-1.0: 0.3-0.5; more preferably, the volume ratio of N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA), trimethylchlorosilane (TMCS) and pyridine is 9: 1: 0.5.

[0038] Preferably, in S2-S4, the volume of the derivatization reagent is 1-3 mL; further preferably, in S2-S4, the volume of the derivatization reagent is 1 mL;

[0039] Preferably, in S2-S4, the reaction is ultrasound, the ultrasound time is 10-40 min, and the power is 35-45 KHz; further preferably, in S2-S4, the reaction is ultrasound, the ultrasound time is 10-30 min, and the power is 35-40 KHz; more preferably, in S2-S4, the reaction is ultrasound, the ultrasound time is 30 min, and the power is 40 KHz.

[0040] Preferably, in S2-S4, the derivatization reaction is carried out in a 20 mL headspace bottle, and the headspace bottle is capped.

[0041] Preferably, in S1, the impurity stock solution contains 0.02-0.04 mg of D-glucose and sucrose, and 0.12-0.17 mg of D-glucono-1,4-lactone and D-gluconic acid per 1 mL;

[0042] Further preferably, in S1, the impurity stock solution contains 0.03 mg of D-glucose and sucrose, 0.15 mg of D-glucono-1,4-lactone and D-gluconic acid per 1 mL;

[0043] Preferably, in S3 and S4, the mass of the D-(+)-glucono δ-lactone sample to be tested is 10-50 mg; further preferably, in S3 and S4, the mass of the D-(+)-glucono δ-lactone sample to be tested is 30 mg.

[0044] Preferably, in S5, the chromatographic conditions are:

[0045] Chromatographic column: HP-1 (30m×0.32mm, 0.25μm) or DB-1 (30m×0.32mm, 0.25μm);

[0046] Inlet temperature: 230-280℃;

[0047] Carrier gas: helium, flow rate 1.3-1.6mL / min;

[0048] FID temperature: 300-330℃;

[0049] Injection volume: 1.2-1.8 μL;

[0050] The split ratio is 60-90:1;

[0051] Heating program:

[0052] Initial temperature, 155-160°C, hold for 0-1min;

[0053] Increase the temperature to 175-185°C at a rate of 1-3°C / min and hold for 0-3min;

[0054] The temperature was raised to 280-300°C at a heating rate of 25-30°C / min and maintained for 3-8 minutes.

[0055] More preferably,

[0056] In S5, the chromatographic conditions are:

[0057] Chromatographic column: HP-1 (30m×0.32mm, 0.25μm);

[0058] Inlet temperature: 250°C;

[0059] Carrier gas: helium, flow rate 1.5 mL / min;

[0060] Detector temperature: 320°C;

[0061] Injection volume: 1.5 μL;

[0062] The split ratio is 80:1;

[0063] Heating program: initial temperature, 160°C, hold for 0 min, increase to 180°C at a heating rate of 2°C / min, hold for 0 min, increase to 300°C at a heating rate of 20°C / min, hold for 5 min.

[0064] Compared with the prior art, the present invention has the following beneficial effects:

[0065] 1. The present invention solves the problem that the GDL test solution is unstable and that GDL and its related substances cannot be directly detected by gas chromatography due to their high boiling points by optimizing the diluent and derivatization conditions.

[0066] 2. The present invention realizes the detection of the content of impurities such as D-glucose, D-gluconic acid, sucrose, D-glucono-1,4-lactone in GDL by optimizing chromatographic conditions. The method has high sensitivity and strong specificity.

[0067] 3. The present invention uses conventional analytical instruments and reagents, is simple to operate, and has high method precision, which is suitable for general laboratories to carry out quality research work. BRIEF DESCRIPTION OF THE DRAWINGS

[0068] Figure 1 This is the chromatogram of the control solution of Example 1.

[0069] Figure 2 The separation solution chromatogram of Example 1.

[0070] Figure 3 The chromatogram of the test solution of Example 1.

[0071] Figure 4 The linear graphs of the derivatization experiments for GDL samples of 10-50 mg for Examples 6-9 are shown.

[0072] Figure 5 It is the chromatogram of the gas chromatography detection method of Example 10.

[0073] Figure 6 The blank solution chromatogram of the test example.

[0074] Figure 7 The chromatogram of the quantitative limit solution of the test case.

[0075] Figure 8 The detection limit solution chromatogram of the test example.

[0076] Fig. 9 The chromatogram is the gas chromatography detection method of Comparative Example 1.

[0077] Fig.10 It is the chromatogram of the gas chromatography detection method of Comparative Example 2.

[0078] Fig.11 The chromatogram is the gas chromatography detection method of Comparative Example 3.

[0079] Fig.12 The chromatogram is the gas chromatography detection method of Comparative Example 4.

[0080] Fig.13 It is the chromatogram of the gas chromatography detection method of Comparative Example 5.

[0081] Fig.14 This is the chromatogram of the gas chromatography detection method of Comparative Example 6.

[0082] Fig.15 This is the chromatogram of the gas chromatography detection method of Comparative Example 7.

[0083] Note: Among them, impurity 1 is D-gluconic acid-1,4-lactone, impurity 2 is D-glucose, impurity 3 is D-gluconic acid, and impurity 4 is sucrose. DETAILED DESCRIPTION

[0084] In order to make the technical means, creative features, purpose and effect of the present invention easy to understand, the present invention is further explained below in conjunction with specific embodiments, but the following embodiments are only preferred embodiments of the present invention, not all. Based on the embodiments in the implementation mode, other embodiments obtained by those skilled in the art without making creative work all belong to the protection scope of the present invention. It is worth noting that the raw materials used in the present invention are all common commercial products, and their sources are not specifically limited. The technology and scientific terms used in the embodiments have the meanings commonly understood by those of ordinary skill in the art to which the present invention belongs.

[0085] The materials and instruments used in the present invention are as follows:

[0086] Table 1. Reference material information (impurity standard)

[0087] Impurity Name factory batch number content Sodium D-Gluconate Aladdin J2227319 ＞99.0％ sucrose Aladdin F2206278 ≥99.5％ D-Glucono-1,4-lactone Aladdin C2224278 ＞99.0％ D-Glucose Runjie Chemical 20150301 ＞99.0％

[0088] Table 2. Instrument information

[0089] Instrument Name Instrument Model factory Balance XS205DU Mettler GC Agilent7890B Agilent Ultrasound KQ5200E Kunshan Shumei

[0090] Example 1

[0091] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0092] S1: Preparation of impurity stock solution: Take appropriate amount of D-glucose, D-gluconic acid, sucrose, and D-gluconic acid-1,4-lactone, dissolve in DMF, and dilute to prepare an impurity stock solution containing 0.3 mg of D-glucose and sucrose, 0.15 mg of D-gluconic acid and D-gluconic acid-1,4-lactone per mL;

[0093] S2: Preparation of control solution: Take 1.0 mL of impurity stock solution in a 20 mL headspace bottle, add 1.0 mL of derivatization reagent, cover, and ultrasonically react for 30 min at a power of 40 kHz to obtain a control solution;

[0094] S3: Preparation of separation solution: Take 30 mg of D-(+)-glucono δ-lactone sample to be tested in a 20 mL headspace bottle, add 1.0 mL of impurity stock solution to dissolve, add 1.0 mL of derivatization reagent, cover, and ultrasonically react for 30 min at a power of 40 KHz to obtain the separation solution.

[0095] S4: Preparation of the test solution: Take 30 mg of the D-(+)-glucono delta-lactone sample to be tested in a 20 mL headspace bottle, add 1.0 mL of DMF to dissolve, add 1.0 mL of the derivatization reagent, cover, and react by ultrasound for 30 min at a power of 40 KHz to obtain the test solution;

[0096] The derivatization reagents described in S2-S4 are N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA), trimethylchlorosilane (TMCS), and pyridine (the volume ratio of the three is 9:1:0.5);

[0097] S5: Take the control solution, the resolution solution and the test solution, respectively, and use GC-FID for detection and analysis, and record the chromatogram; the control solution is used to locate the impurities, the resolution solution is used to examine the separation between the main component and each impurity, the test solution is deducted from the system peak, and the content of the main component and each impurity is calculated by the area normalization method;

[0098] The chromatographic conditions are:

[0099] Chromatographic column: HP-1 (30m×0.32mm, 0.25μm);

[0100] Inlet temperature: 250°C;

[0101] Carrier gas: helium, constant flow rate of 1.5 mL / min;

[0102] Detector temperature: 320°C;

[0103] Injection volume: 1.5 μL;

[0104] The split ratio is 80:1;

[0105] Heating program: initial temperature, 160°C, hold for 0 min, increase to 180°C at a heating rate of 2°C / min, hold for 0 min, increase to 300°C at a heating rate of 20°C / min, hold for 5 min.

[0106] like Figure 1 As shown, it is the chromatogram of the control solution of Example 1, it can be seen that the retention time of D-glucose-1,4-lactone is 11.494min, the retention time of D-glucose is 11.665min, the retention time of D-gluconic acid is 12.961min, and the retention time of sucrose is 16.573min; Figure 2As shown, it is the separation degree solution chromatogram of Example 1, it can be seen that D-(+)-glucono-δ-lactone and each impurity can be separated, and the minimum separation degree is 2.1 of D-(+)-glucono-δ-lactone and D-glucono-1,4-lactone; Figure 3 As shown, it is the chromatogram of the test solution of Example 1, and it can be seen that only two impurities, D-gluconic acid and D-gluconic acid-1,4-lactone, are detected in the test sample.

[0107] Embodiment 2-5

[0108] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0109] Compared with Example 1, only the derivatization time was changed to 10 min, 20 min, 40 min and 50 min, and the rest of the operation was the same as Example 1. (It should be noted that the analysis solution is the test solution)

[0110] As shown in Table 3, when the test solution was ultrasonicated for 10-50 min, there was no obvious difference in the peak area and peak area percentage of the main component and each impurity, indicating that the derivatization reaction can be completed by ultrasonication for 10 min. In order to improve the reproducibility of the method, ultrasonication for 30 min in Example 1 is preferred.

[0111] Table 3. Content of each substance in Examples 1-5

[0112]

[0113] Embodiment 6-9

[0114] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0115] Compared with Example 1, only the sample amount of D-(+)-glucono delta-lactone was changed to 10 mg, 20 mg, 40 mg and 50 mg, and the rest of the operation was the same as Example 1. (It should be noted that the analysis solution is the test solution).

[0116] Table 4. GDL sample size 10-50 mg derivatization test results

[0117]

[0118] From Table 4 and Figure 4 It can be seen that the GDL sample size is 10-50 mg with good linearity. 2 It is 0.9979, indicating that the selected sample size can complete derivatization within 10-50 mg, the impurity detection level is basically the same, and the preferred sample size is 30 mg.

[0119] Example 10

[0120] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0121] Compared with Example 1, only the chromatographic column was changed to DB-1 (30m*0.32mm*0.25μm) and the filler was 100% dimethyl polysiloxane;

[0122] The remaining operations are the same as those in Example 1. (It should be noted that the solution analyzed is the separation solution)

[0123] Figure 5 It is the chromatogram of the gas chromatography detection method of Example 6. Figure 5 and Figure 3 By comparison, it can be seen that both DB-1 and HP-1 can separate all impurities, and the separation effects are similar.

[0124] Test example:

[0125] Methodological verification of the method in Example 1:

[0126] 1. Exclusivity

[0127] Preparation of blank solution: Take 1.0 mL of DMF in a 20 mL headspace bottle, add 1.0 mL of derivatization reagent, cover it, and ultrasonically react for 30 min at a power of 40 kHz to obtain a blank solution.

[0128] The separation solution was prepared in the same manner as in Example 1.

[0129] GC-FID was used for detection and analysis. One injection of blank solution and one injection of separation were injected, and the chromatogram was recorded.

[0130] Depend on Figure 6 It can be seen that the blank solution has no interference with the detection; as can be seen from Table 5, the separation between GDL and various impurities, and between impurities are all greater than 1.5, with good separation effect and strong specificity.

[0131] Table 5. Separation results

[0132] Impurity Name Retention time (min) Minimum resolution of adjacent peaks GDL 11.366 2.1 Impurity 1 11.506 2.1 Impurity 2 11.666 3.2 Impurity 3 12.957 3.4 Impurity 4 16.569 3.4

[0133] 2. Limit of Detection and Limit of Quantitation

[0134] The LOQ concentration and LOD concentration of each impurity were calculated according to the signal-to-noise ratio of each impurity obtained from the control solution, and a LOQ stock solution containing 0.0075 mg / ml impurity 1, 0.0061 mg / ml impurity 2, 0.0041 mg / ml impurity 3 and 0.0037 mg / ml impurity 4 was prepared. 1.0 mL of the LOQ stock solution was placed in a 20 mL headspace bottle, 1.0 mL of the derivatization reagent was added, the bottle was covered, and the ultrasonic reaction was performed for 30 min at a power of 40 KHz to obtain the LOQ solution; a LOD stock solution containing 0.0030 mg / ml impurity 1, 0.0024 mg / ml impurity 2, 0.0016 mg / ml impurity 3 and 0.0015 mg / ml impurity 4 was prepared. 1.0 mL of the LOD stock solution was placed in a 20 mL headspace bottle, 1.0 mL of the derivatization reagent was added, the bottle was covered, and the ultrasonic reaction was performed for 30 min at a power of 40 KHz to obtain the LOD solution.

[0135] Take the LOQ solution and LOD solution for detection and analysis using GC-FID, and record the chromatogram.

[0136] As can be seen from Table 6, the quantitative limits of each impurity are all less than 0.05%, and the method has high sensitivity. Figure 6-7 It is the LOQ and LOD detection spectrum.

[0137] Table 6. LOQ and LOD test results of each impurity

[0138]

[0139] 3. Precision

[0140] Preparation of test solution: Take about 30 mg of D-(+)-glucono delta-lactone sample to be tested in a 20 mL headspace bottle, add 1.0 mL DMF to dissolve, add 1.0 mL derivatization reagent, cover, and ultrasonically react for 30 minutes at a power of 40 KHz to obtain the test solution. Prepare 6 portions in parallel to obtain test solution 1, test solution 2, test solution 3, test solution 4, test solution 5 and test solution 6, and use GC-FID for detection and analysis, and record the chromatogram.

[0141] It can be seen from Table 7 that the RSD of each impurity is less than 10%, and the method has good precision.

[0142] Table 7. Precision test results

[0143]

[0144]

[0145] 4. Solution stability study

[0146] Preparation of the test solution: Take about 30 mg of the D-(+)-glucono delta-lactone sample to be tested in a 20 mL headspace bottle, add 1.0 mL of DMF to dissolve, add 1.0 mL of derivatization reagent, cover, and ultrasonically react for 30 minutes at a power of 40 kHz to obtain the test solution.

[0147] Take the test solution for detection and analysis by GC-FID at 0h, 8h, 24h and 48h, and record the chromatogram.

[0148] It can be seen from Table 8 that the test solution is stable within 48 hours at room temperature.

[0149] Table 8. Solution stability results

[0150]

[0151] It can be seen from the test examples that the gas chromatography detection method for related substances in D-(+)-glucono delta-lactone of the present invention has good specificity, sensitivity, precision and stability.

[0152] Comparative Examples 1-4

[0153] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0154] Compared with Example 1, only the derivatization reagent was changed;

[0155] The remaining operations are the same as in Example 1. (It should be noted that the analysis solution is the test solution)

[0156] Table 9. Derivatization reagents for comparative examples 1-4

[0157] Group Derivatization reagents Comparative Example 1 N,O-bis(trimethylsilyl)trifluoroacetamide:pyridine=10:0.5 Comparative Example 2 Trimethylsilyl chloride: pyridine = 10: 0.5 Comparative Example 3 N,O-bis(trimethylsilyl)trifluoroacetamide:trimethylchlorosilane:pyridine=5:5:0.5 Comparative Example 4 N,O-bis(trimethylsilyl)trifluoroacetamide:trimethylchlorosilane:pyridine=7:3:0.5

[0158] like Figure 8-11 As shown in the figure, compared with Example 1, the byproducts of GDL increased and the derivatization was incomplete when the derivatization reagent was changed, which would affect the specificity and quantification of impurities such as D-glucose. It was proved that only under the specific derivatization reagent of the present invention, the complete derivatization of GDL and the minimum byproduct generation can be achieved, and the effect of detecting four impurities such as D-glucose can be met.

[0159] Comparative Examples 5-7

[0160] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0161] Compared with Example 1, only the type of chromatographic column was changed;

[0162] The remaining operations are the same as those in Example 1. (It should be noted that the analytical solution is the separation solution)

[0163] Table 10. Chromatographic columns of comparative examples 5-7

[0164]

[0165] like Figure 12-15 As shown, compared with Example 1, the chromatographic column is changed, and the four impurities cannot be completely separated; in Comparative Example 5, the two impurities D-glucose-1,4-lactone and D-glucose cannot be separated, in Comparative Example 6, the two impurities D-glucose-1,4-lactone and D-gluconic acid cannot be separated, and in Comparative Example 7, GDL and D-glucose-1,4-lactone cannot be separated; It proves that the specific chromatographic column of the present invention can achieve the effect of simultaneously detecting four impurities D-glucose, D-gluconic acid, sucrose, and D-gluconic acid-1,4-lactone.

[0166] Comparative Examples 8-9

[0167] A gas chromatography detection method for related substances in D-(+)-glucono delta-lactone comprises the following steps:

[0168] Compared with Example 1, only the carrier gas was changed to N2 and H2;

[0169] The remaining operations are the same as those in Example 1. (It should be noted that the analytical solution is the separation solution)

[0170] It can be seen from Table 11 that He is better than N2 as a carrier gas in separating impurities; He is similar to H2 as a carrier gas in separating effects, but the use of H2 is somewhat dangerous, so He is the optimal carrier gas.

[0171] Table 11. Comparison of carrier gas between comparative examples 8 and 9 and the embodiment

[0172]

[0173]

[0174] Finally, it should be noted that the above content is only used to illustrate the technical solution of the present invention, rather than to limit the scope of protection of the present invention. Simple modifications or equivalent substitutions of the technical solution of the present invention by ordinary technicians in this field do not deviate from the essence and scope of the technical solution of the present invention.

Claims

1. A method for detecting related substances in D-(+)-glucono delta-lactone, It is characterized in that It is a pre-column derivatization-gas chromatography detection method, comprising the following steps: S1: Preparation of impurity stock solution: D-glucose, D-gluconic acid, sucrose, and D-gluconic acid-1,4-lactone are dissolved in a solvent to prepare an impurity stock solution; S2: Preparation of control solution: Take the impurity stock solution, add the derivatization reagent, react for 10-50 minutes, and obtain the control solution; S3: Preparation of separation solution: Mix the D-(+)-glucono δ-lactone sample to be tested and the impurity stock solution, add a derivatization reagent, and react for 10-50 minutes to obtain a separation solution; S4: Preparation of test solution: Take the D-(+)-glucono delta-lactone sample to be tested and mix it with the solvent, add the derivatization reagent, react for 10-50 minutes, and obtain the test solution; S5: Take the control solution, separation solution and test solution, respectively, and use GC-FID to detect and analyze them, and record the chromatograms; The chromatographic conditions are: Chromatographic column: capillary column with 100% dimethylpolysiloxane as stationary liquid; Inlet temperature: 230-300℃; Carrier gas: helium, flow rate 1.3-1.7 mL / min; FID temperature: 280-350℃; Injection volume: 1-2 μL; The split ratio is 50-100:1; Heating program: Initial temperature: 150-165°C, hold for 0-3 minutes; Heat to 170-190°C at a rate of 1-5°C / min and hold for 0-5min; The temperature was raised to 280-320°C at a heating rate of 15-30°C / min and maintained for 1-10 min.

2. The detection method according to claim 1, It is characterized in that In S1 and S4, the solvent is selected from at least one of N,N-dimethylformamide, acetonitrile and dichloromethane.

3. The detection method according to claim 1, It is characterized in that In S2 and S4, the derivatization reagent is a mixture of N,O-bis(trimethylsilyl)trifluoroacetamide, trimethylchlorosilane and pyridine.

4. The detection method according to claim 3, It is characterized in that The volume ratio of N,O-bis(trimethylsilyl)trifluoroacetamide, trimethylchlorosilane and pyridine is 6-9: 0.5-1.5：0.1-0.5。 5. The detection method according to claim 4, It is characterized in that The volume ratio of N,O-bis(trimethylsilyl)trifluoroacetamide, trimethylchlorosilane and pyridine is 7-9: 0.5-1.0：0.3-0.5。 6. The detection method according to claim 1, It is characterized in that In S2-S4, the reaction is ultrasound, the ultrasound time is 10-40 minutes, and the power is 35-45KHz.

7. The detection method according to claim 6, It is characterized in that In S2-S4, the reaction is ultrasound, the ultrasound time is 10-30 minutes, and the power is 35-40KHz.

8. The detection method according to claim 1, It is characterized in that In S1, the impurity stock solution contains 0.02-0.04 mg of D-glucose and sucrose, and 0.12-0.17 mg of D-glucono-1,4-lactone and gluconic acid per 1 mL.

9. The detection method according to claim 1, It is characterized in that In S5, the chromatographic conditions are: Chromatographic column: HP-1 (30m×0.32mm, 0.25μm) or DB-1 (30m×0.32mm, 0.25μm); Inlet temperature: 230-280℃; Carrier gas: helium, flow rate 1.3-1.6mL / min; FID temperature: 300-330℃; Injection volume: 1.2-1.8 μL; The split ratio is 60-90:1; Heating program: Initial temperature, 155-160°C, hold for 0-1min; Increase the temperature to 175-185°C at a rate of 1-3°C / min and hold for 0-3min; The temperature was raised to 280-300°C at a heating rate of 25-30°C / min and maintained for 3-8 minutes.

10. The detection method according to claim 1, It is characterized in that In S5, the chromatographic conditions are: Chromatographic column: HP-1 (30m×0.32mm, 0.25μm); Inlet temperature: 250°C; Carrier gas: helium, flow rate 1.5 mL / min; Detector temperature: 320°C; Injection volume: 1.5 μL; The split ratio is 80:1; Heating program: Initial temperature, 160°C, hold for 0 min; Heating was increased to 180°C at a rate of 2°C / min and maintained for 0 min; The temperature was raised to 300°C at a rate of 20°C / min and maintained for 5 min.

Images