CN115420819A

CN115420819A - Method for detecting carbon disulfide in esomeprazole sodium intermediate

Info

Publication number: CN115420819A
Application number: CN202210988679.4A
Authority: CN
Inventors: 宋更申; 王康; 李赛雷
Original assignee: Beijing Youcare Kechuang Pharmaceutical Technology Co ltd
Current assignee: Beijing Youcare Kechuang Pharmaceutical Technology Co ltd
Priority date: 2022-08-17
Filing date: 2022-08-17
Publication date: 2022-12-02

Abstract

The invention belongs to the technical field of drug detection, and particularly relates to a detection method for determining carbon disulfide in an esomeprazole sodium or an intermediate of an isomer of the esomeprazole sodium by using a headspace gas chromatography. According to the method, the carbon disulfide in the esomeprazole sodium or the intermediate of the isomer of the esomeprazole sodium is detected under the specific chromatographic condition and the headspace condition, the accuracy and the repeatability of the detection of the carbon disulfide content in the esomeprazole sodium or the intermediate of the isomer of the esomeprazole sodium are obviously improved, and the method has an extremely low quantification limit and an extremely low detection limit.

Description

Method for detecting carbon disulfide in esomeprazole sodium intermediate

Technical Field

The invention belongs to the technical field of drug detection, and particularly relates to a method for determining carbon disulfide (CS) in an esomeprazole sodium intermediate by using headspace gas chromatography ₂ ) The method of (1).

Background

Esomeprazole sodium is a new generation of proton pump inhibitor, is the levorotatory isomer of omeprazole, and has significant efficacy in the maintenance treatment of gastroesophageal reflux and healed erosive esophagitis. The drug was approved for marketing in the United states and European countries in 2001, and was approved for marketing in China in 2003. The medicine has strong and lasting acid inhibition effect and certain protection effect on gastric mucosa, so that the medicine is the first choice medicine for treating acid-related diseases at present.

The intermediates for synthesizing esomeprazole sodium include 2-mercapto-5-methoxybenzimidazole and 2-chloromethyl-4-methoxy-3,5-lutidine hydrochloride. Wherein, 2-mercapto-5-methoxybenzimidazole is used as a key intermediate substance, and carbon disulfide which is a toxic solvent is used in the synthesis process, so that the monitoring of the carbon disulfide needs to be increased in the internal control quality standard of 2-mercapto-5-methoxybenzimidazole. From PubChem database, the half Lethal Dose (LD) of carbon disulfide is inquired ₅₀ ) 3188mg/kg, the allowable daily exposure (PDE) of carbon disulfide is calculated to be 0.016 mg/day according to the ICH Q3 (C) residual solvent guiding principle and the cleaning and verification guidelines in bulk drug factories issued by the European Committee for pharmaceutical products, and the carbon disulfide is controlled to be below 50ppm according to the maximum daily dose of esomeprazole sodium.

At present, methods for detecting carbon disulfide mainly comprise diethylamine-copper acetate photometry, high performance liquid chromatography-ultraviolet detection (HPLC-UV method), and gas chromatography-flame photometry (GC-FPD method). The spectrophotometry is complicated to operate, low in detection limit and large in interference; the HPLC-UV method and the GC-FPD method also have problems such as low detection limit (GC-FPD method or HPLC-UV method, low sensitivity, detection sensitivity of 100 to 1000ppm in general). Although the detection can be carried out by adopting a gas chromatography-mass spectrometry combined detection method (GC-MS method), the method is complex in development, long in detection period and high in cost, and is not suitable for detection and popularization of large-scale production samples.

We retrieve the following, compare document 1: di Li. Headspace chromatography for measuring carbon disulfide in urine, journal of Chinese sanitary inspection, 2001,11 (3): 321-322, wherein a packed column is used as a chromatographic column, and is combined with a manual sample injection method to measure carbon disulfide in urine, the packed column is gradually eliminated due to poor column efficiency and sensitivity, and the measurement result of manual sample injection is lower in accuracy.

Comparison document 2: helen M. Burbank, michael C. Qian, volume sulfur compounds in chemical chemistry determined by nucleic acid soluble-phase microextraction and gas Chromatography-pulsed fluorescence detection, journal of Chromatography A,1066 (2005) 149-157, which only detected sulfides in cheese.

Therefore, no method for detecting carbon disulfide in esomeprazole sodium intermediate is disclosed in the prior art and patents.

Disclosure of Invention

In order to control the quality of the intermediate of the raw material medicine, the invention provides a detection method for determining carbon disulfide in esomeprazole sodium or the isomer intermediate thereof by using a headspace gas chromatography (HS-GC-ECD).

In the present invention, the phrase "esomeprazole sodium or its isomer intermediate" refers to 2-mercapto-5-methoxybenzimidazole unless otherwise specified.

The "headspace gas chromatography", also known as liquid-above gas chromatography, is a combined operation technique, mainly divided into three types, static headspace analysis, dynamic headspace analysis, and headspace-solid phase microextraction. A headspace chromatographic sampler (sample injection needle) is usually used for extracting and adsorbing solid, liquid, gas and the like under certain conditions and temperature, and then desorption and injection are carried out on a gas chromatographic analyzer, so that volatile components in the liquid or solid sample are directly introduced into the gas chromatographic instrument for separation and detection. (Liu Dong, research on residual solvent in drugs by headspace gas chromatography, beijing university of traditional Chinese medicine)

The invention provides a method for detecting carbon disulfide in an esomeprazole sodium intermediate by adopting a full-automatic headspace gas chromatography, and by adopting specific chromatographic conditions and headspace conditions, 1) the detection specificity of the carbon disulfide is strong, and the characteristic peak separation effect is obvious; 2) The quantitative limit and the detection limit of the carbon disulfide are obviously reduced, and the sensitivity can be as low as 5ng/mL; the lowest detection limit compared with the requirement of the general standard is reduced by 1000 times; 3) The detection method has higher accuracy and wider detection range; 4) The precision is greatly improved, and compared with the general standard requirement, the RSD of the invention is reduced by more than 5 times; 5) The durability is greatly increased, and compared with the RSD standard of a plurality of detection results of common measurement, the RSD of the detection method is reduced by more than 7 percent.

The method has the advantages of high analysis speed, good reproducibility, convenience for detecting and monitoring the quality of the esomeprazole sodium intermediate, universality and suitability for popularization and application in pharmaceutical supervision departments and enterprise production.

Specifically, the invention provides the following technical scheme.

A method for detecting carbon disulfide in an esomeprazole sodium or an intermediate of an isomer thereof by headspace gas chromatography,

which comprises the following steps:

1) Solution preparation: preparing a test solution and a reference solution respectively by using a diluent as a solvent; wherein the test sample comprises esomeprazole sodium or an intermediate of an isomer thereof, and the reference sample is carbon disulfide;

2) And (3) detection: precisely measuring a test solution and/or a reference solution, respectively placing in a headspace bottle, injecting into a gas chromatograph, and detecting under the following chromatographic conditions and headspace conditions;

wherein, the chromatographic column selected in the chromatographic condition is a capillary column taking 100 percent dimethyl polysiloxane as a stationary liquid;

in some embodiments, the temperature programming process comprises: the initial temperature is 30-50 ℃, the temperature is maintained for 5-10 minutes, and the temperature is increased to 150 ℃ at the rate of 10-30 ℃ per minute;

in some preferred embodiments, the temperature-programmed process comprises: the initial temperature was 40 ℃, maintained for 10 minutes, and the temperature was raised to 150 ℃ at a rate of 10 ℃ per minute;

in some embodiments, the headspace conditions have a headspace equilibrium temperature of 65 to 75 ℃, and in some preferred embodiments, the headspace equilibrium temperature is 70 ℃.

In some embodiments, the headspace equilibration time is from 25 to 40min; in some preferred embodiments, the headspace equilibration time is 30min.

In some embodiments, wherein the chromatographic conditions have a sample split ratio of sample exiting the injector to sample entering the column of 2:1 to 1; in some preferred embodiments, the split ratio of sample exiting the injector to sample entering the column is 2:1.

In some embodiments, wherein, in the chromatographic conditions, the sample inlet temperature is 130 to 170 ℃; in some preferred embodiments, the injection port temperature is 150 ℃.

In some embodiments, wherein, in the chromatographic conditions, the detector temperature is 250 to 280 ℃; in some preferred embodiments, the detector temperature is 260 ℃.

In some preferred embodiments, wherein the chromatographic conditions the gas chromatograph is equipped with a detector that is an electron capture ECD detector.

In some embodiments, wherein the chromatographic column is a DB-1 chromatographic column.

In some preferred embodiments, the column length is 30m, the column diameter is 0.35mm, and the liquid film thickness is 3 μm.

In some embodiments, the concentration of the test solution is 5-20 mg/mL; in some preferred embodiments, the concentration of the test solution is 10mg/mL.

In some embodiments, wherein the concentration of the control solution is 0.25 to 1 μ g/mL; in some preferred embodiments, the concentration of the control solution is 0.5. Mu.g/mL.

In some preferred embodiments, wherein the diluent is dimethyl sulfoxide.

In some preferred embodiments, the carrier gas in the chromatographic conditions is nitrogen.

In some embodiments, wherein, in the chromatographic conditions, the column flow rate of the gas chromatography column is 2 to 5mL/min; in some preferred embodiments, the column flow rate is 3mL/min.

In some embodiments, wherein the intermediate of esomeprazole sodium or an isomer thereof is 2-mercapto-5-methoxybenzimidazole.

In some embodiments, wherein the isomer of esomeprazole sodium is omeprazole sodium.

The invention also provides a detection method for determining carbon disulfide in a drug or an intermediate thereof by headspace gas chromatography, which comprises the solution preparation method, the chromatographic conditions and the headspace conditions in any one of the technical schemes, wherein a test sample comprises the drug or the intermediate thereof, and the intermediate does not comprise an intermediate of esomeprazole sodium or an isomer thereof.

The invention has the following beneficial effects:

1. the invention provides a method for detecting carbon disulfide in esomeprazole sodium or an intermediate of an isomer of esomeprazole sodium by adopting a full-automatic headspace gas chromatography. The detection method of the invention has (1) strong specificity and obvious separation effect of the characteristic peak of the carbon disulfide; (2) the quantitative analysis can be carried out on the carbon disulfide in the esomeprazole sodium or the intermediate of the isomer of the esomeprazole sodium, the quantitative limit and the detection limit are obviously reduced, and the detection limit is reduced by 1000 times compared with the lowest detection limit required by the general standard; (3) the accuracy of the detection method is greatly improved, and the detection range is wider; the accuracy of the assay is improved by two orders of magnitude compared to that required by the general standard; (4) the precision degree is greatly increased, and compared with the general standard requirement, the RSD of the invention is reduced by more than 5 times, and the excellent accuracy is shown; (5) the durability is obviously improved, and compared with the RSD standard of multiple detection results of common determination, the RSD of the detection method is reduced by more than 7%.

2. The method of the invention is verified by methodology that the following indexes all meet the detection requirements and meet the requirements of Chinese drug regulatory agencies on drug detection:

(1) the method has good specificity, the measured solvent peak and the characteristic peak of the carbon disulfide in the sample do not interfere with each other, the separation degree is far greater than 1.5, and the separation effect is obvious;

(2) the quantitative limit of carbon disulfide is as low as 5ng/mL (corresponding to 1% of the limit concentration); the lowest detection limit is 2.5ng/mL; compared with the lowest detection limit (mu g grade) required by the general standard, the lowest detection limit (2.5 ng/mL) of the invention is reduced by more than 1000 times, and the detection limit reduction effect is obvious;

(3) the carbon disulfide is in a concentration range of 5 ng/mL-1000 ng/mL, a good linear relation is shown between the peak area of a characteristic absorption peak and the concentration of the carbon disulfide, a linear equation is y =908.1749x-2908.7572, a correlation coefficient r =0.9999, and the linear relation is good; compared with the correlation coefficient r =0.99 required by the general standard, the accuracy of the determination is improved by two orders of magnitude; the detection method has higher accuracy and wider detection range;

(4) in the process of measuring the carbon disulfide, the precision is good (RSD is less than or equal to 2.1 percent), the recovery rate of accuracy is between 96 and 105 percent, the requirement is met (80 to 120 percent), and the RSD of the recovery rates of different samples is less than or equal to 2.0 percent; compared with the standard that the recovery rate RSD required by the general standard is less than or equal to 10.0 percent, the RSD of the detection method is reduced by more than 5 times, and excellent accuracy is shown;

(5) detecting carbon disulfide in a sample solution under different conditions, wherein RSD of a detection result is less than or equal to 3.0% under the conditions that the flow rate of a chromatographic condition is 2.5-3.5 mL/min, the temperature of a sample inlet is 145-155 ℃, the temperature of a detector is 255-265 ℃, the headspace equilibrium temperature is 65-75 ℃ and chromatographic columns of different batches are replaced; compared with the standard that the RSD of the general determination result is less than or equal to 10.0 percent, the RSD of the detection method is reduced by more than 7 percent; the invention can realize good and accurate quantitative detection effect of carbon disulfide within the condition range, and the detection result is not influenced by the condition change within the condition range, and the carbon disulfide shows good and remarkable durability.

3. The method of the invention can be used for quality control of other medicaments containing carbon disulfide.

Drawings

FIG. 1a is a gas chromatogram of a blank solvent provided in example 1 of the present invention;

FIG. 1b is a gas chromatogram of the test sample labeling solution provided in example 1 of the present invention;

figure 2a is a gas chromatogram obtained from the detection of carbon disulfide in esomeprazole sodium intermediate by chromatography column (DB-1 capillary column (30m x 0.53mm,3 μm)) in example 1 of the present invention;

figure 2b is a gas chromatogram obtained from the detection of carbon disulfide in esomeprazole sodium intermediate by chromatography column (DB-624 capillary column (30m x 0.53mm,3 μm)) in example 1 of the present invention;

figure 2c is a gas chromatogram obtained from the detection of carbon disulphide in the esomeprazole sodium intermediate by means of a chromatographic column (HP-5 capillary column (30m x 0.32mm,3 μm)) in example 1 according to the invention;

figure 2d is a gas chromatogram obtained from the detection of carbon disulphide in the esomeprazole sodium intermediate by means of a chromatographic column (DB-Wax capillary column (30m x 0.32mm,3 μm)) in example 1 according to the invention;

FIG. 3 is a gas chromatogram obtained by detecting carbon disulfide in esomeprazole sodium intermediate using the temperature programmed method of example 1;

FIG. 4 is a gas chromatogram obtained by detecting carbon disulfide in esomeprazole sodium intermediate using the temperature programming in example 3;

figure 5 is a linear regression plot of carbon disulfide as provided in example 7 of the present invention;

FIG. 6 is a graph representing the poor sensitivity provided in example 6 of the present invention;

FIG. 7 is a plot of the headspace equilibrium temperature versus the peak area of carbon disulfide at vapor phase conditions as found in example 4 of the present invention;

FIG. 8 is a plot of the correlation of headspace equilibrium time with carbon disulfide peak area under vapor phase conditions in accordance with example 5 of the present invention;

figure 9a is a gas chromatogram obtained from the detection of carbon disulfide in the esomeprazole sodium intermediate of comparative example 1;

figure 9b is a gas chromatogram obtained from the detection of carbon disulfide in the esomeprazole sodium intermediate of comparative example 2;

figure 9c is a gas chromatogram obtained from the detection of carbon disulfide in the esomeprazole sodium intermediate of comparative example 3.

Fig. 10 is a gas chromatogram obtained by detecting carbon disulfide in the esomeprazole sodium intermediate in comparative example 4.

Detailed Description

The following examples further illustrate the present invention but are not to be construed as limiting the invention. Modifications or substitutions to methods, procedures, or conditions of the invention may be made without departing from the spirit and scope of the invention.

Unless otherwise specified, the technical means used in the examples are conventional means well known to those skilled in the art, and the raw materials added in the examples are commercially conventional raw materials unless otherwise specified.

The term "product" in the present invention refers to 2-mercapto-5-methoxybenzimidazole, which is an intermediate of esomeprazole sodium or its isomer, unless otherwise specified, and the gas chromatograph used in the present invention is mainly equipped with an automatic Headspace Sampler (HS) and an Electron Capture Detector (ECD).

Example 1: specialization inspection

1. Chromatographic conditions

2. Head space conditions

Head space equilibrium temperature	70℃
		Time of equilibrium	30min
Sample Ring temperature	100℃
		Temperature of transmission line	110℃

3. Solution preparation

Blank solvent (diluent): dimethyl sulfoxide;

control solution: weighing a proper amount of carbon disulfide closely, adding a diluent to prepare an impurity reference solution, wherein the concentration of the carbon disulfide in the reference solution is 0.5 mu g/mL;

test solution: weighing an esomeprazole sodium intermediate, adding a diluent to prepare a test solution, wherein the concentration of the test solution is 10mg/mL;

adding a standard solution into a test sample: respectively weighing esomeprazole sodium intermediate and carbon disulfide, mixing, adding a diluent to prepare a sample adding standard solution, wherein the concentration of the raw material drug intermediate in the sample adding standard solution is 10mg/mL, and the concentration of the carbon disulfide is 0.5 mu g/mL.

4. Measurement method

And precisely measuring the solution, respectively performing headspace sample injection, and recording the chromatogram map of the test.

5. Conclusion

As can be seen from the gas chromatogram of the control solution, the baseline is flat, and a characteristic absorption peak appears when the retention time is about 7min, while a characteristic peak of carbon disulfide does not appear in the blank solvent. The blank solvent does not interfere the detection of the carbon disulfide, the separation degree of the solvent peak and the carbon disulfide characteristic peak in the reference solution is far larger than 1.5, and the solvent does not interfere the detection of the component carbon disulfide to be detected. As the carbon disulfide (containing no carbon disulfide) is not actually detected in the test sample solution (namely, the bulk drug intermediate), the method designs a standard addition experiment of the test sample, and the result shows that the characteristic peak of the carbon disulfide is successfully detected in the standard addition test sample, which indicates that the method has good specificity. The specificity is shown in FIG. 1a and FIG. 1b.

Example 2: selection of chromatography columns

On the basis of the normal condition of the embodiment 1, the chromatographic columns with different polarities are replaced, other detection parameters are the same as those in the embodiment 1, and the applicability of the method is examined.

And (4) conclusion:

when the chromatographic column (DB-1 capillary column (30m x 0.53mm,3 μm)) and the method of example 1 of the present invention were used to detect carbon disulfide in the esomeprazole sodium intermediate, the separation of carbon disulfide from the blank solvent was good.

Under the same condition, when the chromatographic columns with different polarities in the embodiment 2 are used for detecting carbon disulfide in the esomeprazole sodium intermediate, the separation degree and the peak shape of the carbon disulfide and a blank solvent are poor, namely the blank solvent interferes with the detection of the component to be detected, and the requirement of quantitative determination cannot be met. A comparison of column selection is shown in FIGS. 2a, 2b, 2c and 2d.

Therefore, the DB-1 chromatographic column in the method can realize the obvious separation of the carbon disulfide in the esomeprazole sodium intermediate, and other chromatographic columns cannot realize the separation effect.

Example 3: determination of temperature programmed mode

On the basis of the normal condition of the embodiment 1, the temperature programming mode is changed, other detection parameters are the same as the embodiment 1, and the applicability of the method is examined. The modified temperature programming mode is as follows: the starting temperature was 60 ℃ for 2 minutes, and the temperature was raised to 150 ℃ at a rate of 20 ℃ per minute.

And (4) conclusion:

when the carbon disulfide in the esomeprazole sodium intermediate is detected by adopting the temperature programming mode (the initial temperature is 40 ℃, the temperature is maintained for 10 minutes, and the temperature is raised to 150 ℃ at the rate of 10 ℃ per minute) in the embodiment 1 of the invention, the complete separation of the characteristic peaks of the carbon disulfide and the blank solvent is fully realized, the matrix in the test spectrogram does not interfere with the detection of the component to be detected, the quantitative determination requirement is fully met, and the detection effect is excellent. The spectrum of the good separation degree is shown in FIG. 3.

When the programmed temperature raising method of example 3 (initial temperature 60 ℃, holding time 2 minutes, raising temperature to 150 ℃ at a rate of 20 ℃ per minute) is adopted to detect carbon disulfide in the esomeprazole sodium intermediate, carbon disulfide and a blank solvent cannot be completely separated, a matrix interferes with the detection of a component to be detected, and a baseline is not stable, so that the quantitative determination requirement cannot be met. The spectrum of the separation difference is shown in FIG. 4.

Therefore, by adopting the method, the initial temperature in the programmed heating process is controlled to be 30-50 ℃, the temperature is maintained for 5-10 minutes, and the temperature is raised to 150 ℃ at the rate of 10-30 ℃ per minute, so that the carbon disulfide in the esomeprazole sodium intermediate can be obviously separated; when the initial temperature is 40 ℃, the temperature is maintained for 10 minutes, and the temperature is increased to 150 ℃ at the rate of 10 ℃ per minute, and the separation effect is best.

Example 4: determination of headspace equilibrium temperature

The headspace equilibrium temperature was changed based on the normal conditions of example 1, and the analysis was performed by equilibration at 50 ℃, 55 ℃, 60 ℃, 65 ℃,70 ℃, 75 ℃ and 80 ℃ for 30min, and other detection parameters were the same as those of example 1. The headspace equilibrium temperature is related to peak area as shown in the following table and figure 7.

And (4) conclusion:

the increase of the equilibrium temperature can increase the concentration of the substance to be detected in the equilibrium gas phase, which is beneficial to improving the sensitivity of the method, but too high equilibrium temperature can cause the degradation of the test sample, bring much interference and influence the detection effect. In principle, the larger the peak area response value of the component to be measured, the higher the sensitivity of the method. As can be seen from FIG. 7, the peak area gradually increases from 50 to 70 ℃, the peak area of 70 to 80 ℃ does not change obviously, and reaches the plateau phase, and the 70 ℃ is the most obvious and appropriate headspace equilibrium temperature in consideration of the factors such as the pressure-resistant safety factor of the headspace bottle and more interference caused by high temperature.

According to experimental results, the method disclosed by the invention has the advantages that when the equilibrium temperature in the headspace condition is controlled to be 65-75 ℃, the effect of separating carbon disulfide from the esomeprazole sodium intermediate is obvious; the separation was best when the headspace equilibrium temperature was 70 ℃.

Example 5: determination of headspace equilibrium time

On the basis of the normal condition of the embodiment 1, the headspace is balanced for 10 min, 20min, 25 min, 30min, 40min, 50 min and 60min respectively at the headspace balance temperature of 70 ℃, other detection parameters are the same as the embodiment 1, and the relationship between the balance time and the peak area is shown in the following table and fig. 8.

Conclusion

As can be seen from FIG. 8, within 10-30 min, the peak area is continuously increased along with the increase of the equilibrium time, and after 30min, the peak area has a tendency to decrease, and the analysis may be caused by the decrease of the airtightness of the headspace bottle due to too long equilibrium time. Based on the principle that the higher the peak area response value of the component to be measured is, the higher the sensitivity of the method is, in addition, considering the timeliness of the experiment, the optimal headspace balance time is selected to be 30min.

According to experimental results, when the method is adopted and the balance time in the headspace condition is controlled to be 25-40 min, the separation detection of the carbon disulfide in the esomeprazole sodium intermediate can be achieved; when the headspace equilibrium time is 30min, the separation effect is optimal.

Example 6: effect of headspace equilibrium temperature and time on Signal-to-noise ratio

On the basis of the conditions of the embodiment 1, the balance temperature and the balance time in the headspace parameters were changed, and other detection parameters were the same as those of the embodiment 1. Headspace parameters were adjusted as follows:

parameter(s)	Example 1	Practice ofExample 6
			Head space equilibrium temperature	70℃	60℃
Head space balance time	30min	20min
			Signal to noise ratio	S/N＞100	S/N＝3
Signal to noise ratio standard	S/N≥10	S/N≥10

And (4) conclusion:

by comparison of inventive example 1 with example 6, the headspace parameters were varied, with the equilibrium temperature ranging from 70 ℃→ 60 ℃; equilibration time 30min → 20min.

In the embodiment 1, the signal-to-noise ratios of carbon disulfide response values detected by the gas chromatograph ECD are all S/N & gt 100 and are far higher than the requirement that the quantitative limit S/N is more than or equal to 10, and the signal intensity is far higher than the noise intensity, so that the detection effect is more accurate, the influence factors caused by other noises are smaller, and the detection sensitivity is obvious.

In example 6, after the headspace parameter is adjusted, the signal-to-noise ratio S/N =3, which is much smaller than the requirement that the quantitative limit S/N is greater than or equal to 10, the difference between the signal strength and the signal strength is small, and the signal strength of the characteristic absorption peak of carbon disulfide is easily affected by the noise strength, which cannot meet the requirement of quantitative determination. The less sensitive representative map is shown in FIG. 6.

Example 7: study of linearity

Preparation of stock solution: precisely weighing 200mg of carbon disulfide, placing the carbon disulfide into a 100mL measuring flask, diluting the carbon disulfide to a scale with a diluent, shaking up, precisely weighing 1mL again, placing the carbon disulfide into a 200mL measuring flask, diluting the carbon disulfide to a scale with the diluent, and shaking up to obtain the carbon disulfide.

Preparation of linear solution: respectively transferring stock solutions with different volumes according to the following table for dilution, and preparing linear solutions with corresponding concentrations, which are recorded as L1-L6; wherein the concentration of carbon disulfide in L4 is determined by LD ₅₀ The content standard of 50ppm (fifty parts per million of the test article content) prepared (half of the lethal dose) is used as a limit concentration value, and the limit concentration is specified to be 100%.

Precisely measuring 2mL of the linear solutions L1-L6, placing in a 20mL headspace bottle, sealing, injecting into a gas chromatograph, and recording the chromatogram. The chromatographic conditions and headspace conditions were the same as in example 1. Figure 5 is a linear regression plot of carbon disulfide. The linearity results are given in the following table:

and (4) conclusion:

the carbon disulfide is in a concentration range of 5 ng/mL-1000 ng/mL, a linear equation between a peak area and the carbon disulfide concentration (ng/mL) is y =908.1749x-2908.7572, wherein a correlation coefficient (r) is 0.9999, the linear relation is good, and compared with the correlation coefficient r =0.99 required by a general standard, the accuracy of determination is improved by two orders of magnitude; the accuracy of the detection method is greatly improved, and the detection range is wider. The quantitative detection Limit (LOQ) of the carbon disulfide by the instrument can be as low as 5ng/mL, and an extremely low quantitative detection line proves that the detection method has obvious detection sensitivity on the carbon disulfide.

Example 8: quantitative and detection Limit research

Blank solvent (diluent): dimethyl sulfoxide (DMSO).

Taking a proper amount of the stock solution, adding a solvent to dilute step by step, preparing a series of concentration samples, precisely measuring 2mL respectively, placing in a 20mL headspace bottle, sealing, injecting into a gas chromatograph, and recording the chromatogram. Inspecting the signal-to-noise ratio (S/N) of a chromatographic peak, and determining the signal-to-noise ratio as a detection limit when the signal-to-noise ratio is more than or equal to 3; when the signal-to-noise ratio is more than or equal to 10, the quantitative limit is obtained. The chromatographic conditions and headspace conditions were the same as in example 1. The results are given in the following table:

and (4) conclusion:

the quantitative limit of carbon disulfide is 5ng/mL, the peak area RSD of 6 parts of quantitative limit solution is 4.1 percent and is less than 15.0 percent; the signal-to-noise ratios of the 6 parts of quantitative limiting solution are all more than or equal to 10; the detection limit is 2.5ng/mL, and the signal-to-noise ratio of 3 parts of detection limit solution is more than or equal to 3. According with the standard requirement, in the detection method, the quantitative limit of the carbon disulfide is 5ng/mL, and the detection limit is as low as 2.5ng/mL, which shows that the method has obvious sensitivity effect. Compared with the lowest detection limit (2.5 mug/mL) required by the general standard, the lowest detection limit (2.5 ng/mL) of the invention is reduced by 1000 times, and the detection limit reduction effect is obvious.

Example 9: study of accuracy

Diluent agent: dimethyl sulfoxide (DMSO).

Preparation of recovery stock solution: precisely weighing 200mg of carbon disulfide, placing the carbon disulfide into a 100mL measuring flask, diluting the carbon disulfide to a scale with a diluent, shaking up, precisely weighing 1mL, placing the carbon disulfide into a 200mL measuring flask, diluting the carbon disulfide to a scale with the diluent, and shaking up to obtain the carbon disulfide.

Control solution (0.5. Mu.g/mL): precisely measuring 1mL of the reference stock solution, placing the reference stock solution in a 20mL measuring flask, diluting the reference stock solution to a scale with a diluent, precisely measuring 2mL of the reference stock solution, placing the reference stock solution in a 20mL headspace flask, and sealing.

Background solution: taking about 100mg of the product, accurately weighing, placing in a 10mL measuring flask, adding a diluent to dissolve and dilute to a scale, accurately weighing 2mL, placing in a 20mL headspace bottle, and sealing.

50% recovery solution: weighing 200mg of the product, accurately weighing, placing in a 20mL measuring flask, accurately adding 0.5mL of recovery stock solution, diluting with diluent to scale, shaking, accurately weighing 2mL, placing in a 20mL headspace flask, and sealing. The 50% recovery solution was prepared in parallel in 3 portions, which were respectively designated 50% -1, 50% -2, 50% -3.

100% recovery solution: weighing 200mg of the product, accurately weighing, placing in a 20mL measuring flask, accurately adding 1mL of recovery stock solution, diluting with diluent to scale, shaking, accurately weighing 2mL, placing in a 20mL headspace flask, and sealing. 3 parts of the 100% recovery rate solution are prepared in parallel and are respectively marked as 100-1, 100-2 and 100-3.

150% recovery solution: weighing 200mg of the product, accurately weighing, placing in a 20mL measuring flask, accurately adding 1.5mL of recovery stock solution, diluting with diluent to scale, shaking, accurately weighing 2mL, placing in a 20mL headspace flask, and sealing. The 150% recovery rate solution was prepared in parallel in 3 portions, which were respectively designated 150% -1, 150% -2, 150% -3.

And (4) injecting the solution into a gas chromatograph, and recording a chromatogram. The chromatographic conditions and headspace conditions were the same as in example 1, and the carbon disulfide recovery results are given in the following table:

and (4) conclusion:

the test results of the carbon disulfide recovery rate in the table show that the carbon disulfide recovery rate is between 96% and 105%, the recovery rate distribution range is concentrated and is close to 100%, and the quantitative determination requirement on carbon disulfide can be accurately met. Wherein, in the quantitative determination process, the carbon disulfide can also reach 100% recovery rate, which shows that the method has good accuracy. The RSD percent of the recovery rate of 9 samples does not exceed 2.0 percent, and compared with the standard that the RSD percent of the recovery rate required by the general standard is less than or equal to 10.0 percent, the RSD of the detection method is reduced by more than 5 times, and the accuracy of the detection result is obviously improved.

Example 10: study of precision

Diluent agent: dimethyl sulfoxide (DMSO).

Preparation of a control stock solution: precisely weighing 200mg of carbon disulfide, placing the carbon disulfide in a 100mL measuring flask, diluting the carbon disulfide to a scale with a diluent, shaking the carbon disulfide, precisely weighing 1mL, placing the carbon disulfide in a 200mL measuring flask, diluting the carbon disulfide to a scale with the diluent, and shaking the carbon disulfide to obtain the carbon disulfide. (parallel preparation 2 portions)

Adding standard test solution to the limit of 100%: weighing 200mg of the product, accurately weighing, placing in a 20mL measuring flask, accurately adding 1mL of reference stock solution, diluting with diluent to scale, shaking, accurately weighing 2mL, placing in a 20mL headspace flask, and sealing. The 100% limit added standard test sample solution is prepared into 6 parts in parallel, and the parts are respectively recorded as: a test sample-1, a test sample-2, a test sample-3, a test sample-4, a test sample-5 and a test sample-6.

And respectively injecting the 100% limit standard sample solution into a gas chromatograph, and recording the chromatogram. The chromatographic conditions and headspace conditions were the same as in example 1, and the reproducibility results are given in the following table:

and (4) conclusion:

as can be seen from the results of the carbon disulfide repeatability tests in the above table, by quantitatively detecting the content of carbon disulfide in 6 parts of the same 100% limit standard sample solution, the RSD of the 6 times of measurement results is less than or equal to 2.1%, which meets the standard requirements, and shows that the method has good repeatability and remarkable precision effect.

Example 11: study of durability

The solution was prepared as in example 7.

The chromatographic conditions were the same as in example 1.

The durability of the detection method is examined under the conditions of different flow rates, injection port temperatures, detector temperatures, headspace equilibrium temperatures and different batches of chromatographic columns of the same brand and model.

Durability results are given in the following table:

and (4) conclusion:

according to the durability test results of the embodiment 11 and the embodiment 1 under different conditions, the RSD of the chromatographic conditions provided by the invention is less than or equal to 3.0% under the conditions that the flow rate is 2.0-4.0 mL/min, the injection port temperature is 140-160 ℃, the detector temperature is 250-270 ℃, the headspace equilibrium temperature is 65-75 ℃ and chromatographic columns of different batches are replaced, and compared with the standard that the RSD of the general determination result is less than or equal to 10.0%, the RSD of the detection method is reduced by more than 7%; the invention can realize good and accurate quantitative detection effect of carbon disulfide within the condition range, and the measurement result is not influenced by the condition change within the condition range, so that the durability of chromatographic conditions is greatly improved.

Wherein, when the chromatographic conditions are that the flow rate is 3.0mL/min, the temperature of the sample inlet is 150 ℃, the temperature of the detector is 260 ℃ and the head space equilibrium temperature is 70 ℃, the quantitative detection effect of the carbon disulfide is optimal.

Comparative examples 1 to 3:

as represented by example 1 of the present invention, comparative analysis was conducted on the same as in comparative documents 1, 2 and 3. Wherein, the first and the second end of the pipe are connected with each other,

comparison document 1: di Li headspace chromatography for carbon disulfide in urine, journal of Chinese sanitation inspection, 2001,11 (3): 321-322, (see pages 321-322, part 1 and tables 1 and 2);

comparison document 2: journal of Chromatography A,1066 (2005) 149-157 (see pages 150 to 151, parts 2.2 and 3);

comparison document 3: mass spectra, 2007;21 (see pages 1505-1509).

Specific comparisons are shown in the following table:

table 1 comparison of research directions of this application with comparison documents

Table 2 comparison of chromatographic conditions of the present application with the reference

And (4) conclusion:

according to the comparison between the present embodiment 1 and the comparison documents 1, 2, and 3 in the above table, the main differences between the present embodiment 1 and the comparison document 1 are as follows: 1) A chromatographic column: the chromatographic column used in this example 1 was a capillary column with specification 30m x 0.53mm,3 μm; the column used in comparative example 1 was a packed column (which was gradually eliminated due to poor column efficiency and sensitivity). 2) Headspace sampler: in this example 1, a full-automatic headspace sampler is used, and the comparison document 1 adopts manual sampling. In addition, the method established in this embodiment 1 is verified by a complete methodology, and has significant effects on linearity (correlation coefficient r = 0.9999), precision (RSD = 2.1%), and accuracy (recovery rate 96% -105%).

The detection in the comparison document 2 is sulfide in cheese, the chromatographic column type, the temperature programming mode and the sample injection mode are all different from the application, and the difference between the sample injection port temperature and the detector temperature is obvious from the application. Meanwhile, the comparison document 3 is used for detecting the 2-mercaptobenzimidazole and the 2-thiouracil derivative, and different analysis and detection methods are adopted, and parameters of the detection method are also obviously different.

In the detection processes of comparative examples 1 to 3, the carbon disulfide in the test solution and the standard solution of the test article of example 1 of the present invention was detected under the detection conditions of the above-mentioned comparative documents 1 to 3, respectively.

The detection results show that (figure 9a (comparative example 1), figure 9b (comparative example 2) and figure 9c (comparative example 3), the characteristic peak of carbon disulfide can not be detected in the chromatograms of comparative examples 1 to 3, therefore, the separation detection conditions of comparative documents 1 to 3 can not realize accurate detection and analysis of carbon disulfide in esomeprazole sodium intermediate.

Comparative example 4:

this was compared with a comparative document 4, which is representative of example 1 of the present invention. Wherein the content of the first and second substances,

comparison document 4: wang Yongping, li Xinyun, zhang Junhua, wang Xincheng, shi Tingxiu carbon disulfide, environmental chemistry, 1994,13 (4): 359-362, (see pages 359-361, sections 1, 5 and 7);

solution preparation:

adding a standard solution into a No. 1 sample: respectively weighing esomeprazole sodium intermediate and carbon disulfide, mixing, adding a diluent to prepare a sample adding standard solution, wherein the concentration of the raw material drug intermediate in the sample adding standard solution is 10mg/mL, and the concentration of the carbon disulfide is 2.5 mu g/mL.

Sample No. 2 adding standard solution: respectively weighing esomeprazole sodium intermediate and carbon disulfide, mixing, adding a diluent to prepare a sample adding standard solution, wherein the concentration of the raw material drug intermediate in the sample adding standard solution is 10mg/mL, and the concentration of the carbon disulfide is 5.5 mu g/mL.

The carbon disulfide determination was performed using the conditions of the comparison document 4 headspace gas chromatography (HS-GC-ECD), the specific comparison is given in the following table:

table 3 comparison of the present application with reference 4

In the detection method of the reference 4, the chromatographic column is a packed column, and chromatographic conditions such as column temperature, injection port temperature, detector temperature and the like are significantly different from those of the present application.

The carbon disulfide in the sample solution 1 is detected under the condition of the comparative example 4, the detection result shows that 10 (the comparative example 4) is shown, the characteristic peak of the carbon disulfide cannot be detected in the chromatogram of the comparative example 4, the sensitivity is poor, and the detection limit under the condition of the comparative example 4 (5.5 mu g/L) is obviously lower than that of the application (2.5 ng/mL). Therefore, the separation detection conditions of the reference 4 cannot realize accurate detection and analysis of carbon disulfide in the esomeprazole sodium intermediate.

And (3) detecting the carbon disulfide in the sample solution added with the standard solution of No. 2 by adopting the conditions of the comparative example 4, repeating the detection for 8 times, and inspecting the accuracy.

Table 4 accuracy comparison of this application with reference 4

And (4) conclusion:

from the carbon disulfide repeatability test results in the table, by quantitatively detecting the content of carbon disulfide in 8 parts of the same standard sample solution, the RSD of the measurement result in example 1 is 1.1% (standard requirement is less than or equal to 10.0%), the method meets the standard requirement, and the method has good repeatability and shows remarkable precision effect.

The RSD of the measurement result of example 4 is 13.1%, which exceeds the standard requirement, and after repeated experiments, the relative standard deviation recorded in the reference 4 can not be reached to the standard of less than 7%, which indicates that the method of example 4 has poor precision, can not realize accurate quantification, and has poor repeatability.

Although the invention has been described in detail hereinabove by way of general description, specific embodiments and experiments, it will be apparent to those skilled in the art that many modifications and improvements can be made thereto based on the invention. Accordingly, such modifications and improvements are intended to be within the scope of the invention as claimed.

Claims

1. A detection method for determining carbon disulfide in an esomeprazole sodium or an intermediate of esomeprazole isomer by headspace gas chromatography comprises the following steps:

wherein, the chromatographic column selected in the chromatographic condition is a capillary column taking 100 percent dimethyl polysiloxane as stationary liquid;

the temperature programming process comprises the following steps: the initial temperature is 30-50 ℃, the temperature is maintained for 5-10 minutes, and the temperature is increased to 150 ℃ at the rate of 10-30 ℃ per minute;

preferably, the temperature programming process includes: the initial temperature was 40 ℃, maintained for 10 minutes, and the temperature was raised to 150 ℃ at a rate of 10 ℃ per minute;

in the headspace condition, the headspace equilibrium temperature is 65-75 ℃, and the preferred headspace equilibrium temperature is 70 ℃; the headspace balance time is 25-40 min, and the preferred headspace balance time is 30min.

2. The detection method according to claim 1, wherein in the chromatographic conditions, the sample split ratio of the discharge sample injector to the sample entering the chromatographic column is 2:1-20; the preferred split ratio of sample exiting the injector to sample entering the column is 2:1.

3. The detection method according to claim 1, wherein in the chromatographic conditions, the sample inlet temperature is 130-170 ℃; the preferred injection port temperature is 150 ℃.

4. The detection method according to claim 1, wherein in the chromatographic condition, the temperature of a detector is 250-280 ℃; the preferred detector temperature is 260 ℃.

5. The detection method according to claim 1, wherein in the chromatographic condition, the detector equipped with the gas chromatograph is an electron capture ECD detector.

6. The detection method according to claim 1, wherein in the chromatographic conditions, the carrier gas is nitrogen.

7. The detection method according to claim 1, wherein in the chromatographic conditions, the column flow rate of the chromatographic column is 2 to 5mL/min; a preferred column flow rate is 3mL/min.

8. The method according to claim 1, wherein the column is a DB-1 column.

9. The detection method according to claim 1 or 8, wherein the column length of the column is 30m, the column diameter is 0.35mm, and the liquid film thickness is 3 μm.

10. The detection method according to claim 1, wherein the concentration of the sample solution is 5 to 20mg/mL, preferably 10mg/mL.

11. The test method according to claim 1, wherein the concentration of the control solution is 0.25 to 1 μ g/mL, preferably the concentration of the control solution is 0.5 μ g/mL.

12. The detection method according to claim 1, wherein the diluent is dimethyl sulfoxide.

13. The assay according to any one of claims 1 to 12, wherein the intermediate of esomeprazole sodium or its isomer is 2-mercapto-5-methoxybenzimidazole.

14. The assay according to any one of claims 1 to 13, wherein the isomer of esomeprazole sodium is omeprazole sodium.

15. A detection method for determining carbon disulfide in a drug or an intermediate thereof by headspace gas chromatography, comprising the solution formulation method, chromatographic conditions and headspace conditions of any one of claims 1 to 14, wherein the test sample comprises the drug or the intermediate thereof, and the intermediate does not comprise an intermediate of esomeprazole sodium or an isomer thereof.