Analysis method for determining tromethamine content in lornoxicam for injection

Abstract

The invention relates to the technical field of pharmaceutical analysis, and particularly discloses a gas chromatographic analysis method for determining tromethamine content in lornoxicam for injection. A DB-624 chromatographic column is adopted, the initial column temperature is 150 DEG C, the temperature is increased to 240 DEG C at the speed of 30 DEG C / min, and the temperature is kept for 8 minutes; a sample inlet of the hydrogen flame ionization detector is 240 DEG C, the split ratio is 10: 1, the carrier gas flow velocity is 4.0 ml / min, and the sample size is 2 microliters. Key pretreatment: preparing a test solution and a reference solution by using a 20% glacial acetic acid methanol solution as a diluent; a blank solution is a mixed solution of a 20% glacial acetic acid methanol solution and water, and matrix interference is effectively eliminated. And quantifying through an external standard method. The method is high in specificity, and organic amine impurities can be separated; the linear range is wide, the precision is high and the accuracy is good. The method overcomes the defects of low sensitivity of a titration method and poor specificity of HPLC, and is simple and convenient to operate, rapid in analysis and suitable for quality control of the preparation.

Description

Technical Field

[0001] An analytical method for determining the content of tromethamine in lornoxicam for injection, relating to the field of pharmaceutical analysis technology. Background Technology

[0002] In the field of pharmaceutical analysis, especially the accurate determination of excipient content in injectable formulations, has always been a crucial link in ensuring drug quality and patient safety. Trometamol, as an important pharmaceutical excipient, is widely used in injectable formulations. Its main functions include adjusting pH, maintaining the stability of drug solutions, and preventing the degradation of active ingredients (such as lornoxicam) due to pH fluctuations. Trometamol is included in the 2020 edition of the Chinese Pharmacopoeia, but its content determination method is mainly titration. However, titration relies on the endpoint determination of the chemical reaction, which has significant limitations when determining low-content samples: when the trometamol content is low (such as a typical concentration of about 3 mg / ml in injectable lornoxicam), titration cannot provide sufficient sensitivity and accuracy, is easily affected by matrix interference, leading to large deviations in the determination results, and cannot meet the high precision requirements of modern pharmaceutical quality control.

[0003] In existing technologies, high-performance liquid chromatography (HPLC) is a commonly used alternative method in pharmaceutical analysis, widely applied for the quantitative detection of excipients and active ingredients due to its high separation efficiency and wide applicability. For example, HPLC is often used for the determination of buffers, but this method may face insufficient specificity when dealing with impurities similar to tromethorphan (such as other organic amines), making it difficult to achieve complete baseline separation and thus affecting quantitative accuracy. Furthermore, HPLC methods typically involve complex mobile phase optimization and long analysis times, resulting in shortcomings in terms of environmental friendliness and ease of operation. Specifically, for the specific drug lornoxicam for injection, there are no technical documents or reports (including HPLC methods) specifically addressing the determination of its tromethorphan content, reflecting a technological gap in this field. Because lornoxicam injection contains a small amount of tromethorphan and has a complex matrix (containing other excipients and active ingredients), existing titration methods or general HPLC methods cannot effectively solve problems such as poor specificity, large fluctuations in recovery rate (for example, the recovery rate of conventional methods may be lower than 90% or higher than 108%), and poor reproducibility (RSD is often higher than 5%). There is an urgent need to develop a more accurate, efficient and environmentally friendly analytical method.

[0004] In summary, existing technologies (such as titration and HPLC) have significant shortcomings in the determination of tromethamine in lornoxicam for injection: the analytical methods lack specificity, have low adaptability to different drug types, are prone to interference from solution composition (such as diluents and matrices), and their performance indicators (such as linear range, precision, and accuracy) are difficult to meet pharmacopoeia standards. These limitations not only affect drug quality control but may also jeopardize medication safety, highlighting the urgent need for innovative analytical technologies. This invention aims to fill this technological gap by introducing a unique gas chromatography (GC) scheme to solve the aforementioned problems and increase the likelihood of patent grants. Summary of the Invention

[0005] The technical problem to be solved by this invention is to overcome the shortcomings of the prior art and provide a high-precision, highly specific, environmentally friendly and simple analytical method to solve the key problems in the determination of tromethamine content in lornoxicam for injection.

[0006] Titration has low sensitivity and cannot accurately detect low-content samples (typical concentration of about 3 mg / ml), and is easily affected by matrix interference, leading to result deviation.

[0007] HPLC has poor specificity, making it difficult to separate similar impurities (such as other organic amines), and it is also complex to operate and takes a long time to analyze.

[0008] Existing methods have large fluctuations in recovery rates (common methods often have recovery rates below 90% or above 108%) and poor repeatability (RSD often exceeds 5%), failing to meet the stringent requirements of pharmacopoeia standards for drug quality control.

[0009] The technical solution adopted by this invention to solve its technical problem is: an analytical method for determining the content of tromethamine in lornoxicam for injection, using gas chromatography for detection, with a DB-624 60m x 0.32mm x 1.80μm column, a temperature program of initial column temperature 150℃, increasing to 240℃ at a rate of 30℃ per minute, and maintaining for 8 minutes, a flame ionization detector with a detector temperature of 250℃, an injection port temperature of 240℃, a split ratio of 10:1, an injection volume of 2μl, and external standard method for quantitative analysis. This invention achieves efficient separation of impurities with properties similar to tromethamine (such as organic amines), ensuring high specificity. At the same time, the external standard method combined with optimized parameters improves quantitative accuracy, making the method simple to operate, with short analysis time, and environmentally friendly, avoiding the complex mobile phase of HPLC.

[0010] Preferably, the analytical method for determining the content of tromethamine in lornoxicam for injection further includes the preparation of a pre-detection solution for gas chromatography. This solution comprises a blank solution, a diluent, a test solution, and a reference solution. The blank solution is prepared by mixing 20% ​​glacial acetic acid methanol solution and water at a volume ratio of 1:1, and the diluent is a 20% glacial acetic acid methanol solution. This specific solution composition effectively eliminates matrix interference, ensuring a pure detection signal. Simultaneously, the diluent simplifies sample processing steps, improves the reproducibility and environmental friendliness of the method, reduces the amount of organic solvent used, and supports high accuracy (97.0% recovery, RSD 4.5%) and precision (RSD 1.5%), which is superior to conventional methods.

[0011] Preferably, the test solution is prepared by measuring 5.0 ml of lornoxicam injection solution into a 10 ml volumetric flask, adding diluent to dilute to the mark, shaking well, filtering, and using the filtrate as the test solution. Quantitatively measuring 5.0 ml of the solution and diluting it to 10 ml optimizes the sample concentration, ensuring it falls within the optimal linear range of this method (1.52-4.55 mg / ml), avoiding overload or insufficient sensitivity issues; using the filtrate removes particle interference and improves detection stability.

[0012] Preferably, the concentration of tromethamine in the test solution is 3 mg / ml. This fixed concentration matches the midpoint of the linear range of the method, maximizing the utilization of detection sensitivity; the response value is stable at this concentration (peak area approximately 53.995), ensuring the accuracy of quantitative analysis, while simplifying the sample pretreatment process.

[0013] Preferably, the preparation method of the reference solution is as follows: weigh 27mg-33mg of tromethamine reference standard, place it in a 10ml volumetric flask, add 5ml of water, sonicate to dissolve, then dilute to the mark with diluent and shake well. Sonication and dilution ensure complete dissolution and uniform dispersion of the reference standard, reducing sources of error; adding 5ml of water first and sonicating improves dissolution efficiency and avoids precipitation.

[0014] Preferably, the concentration range of tromethamine in the reference solution is 2.7 mg / ml to 3.3 mg / ml. This concentration range covers the typical concentration of the test sample (3 mg / ml), ensuring that the reference standard and the sample are in the same response range, thus improving the matching degree of the external standard method.

[0015] Preferably, the flow rate of the gas chromatograph is 4.0 ml / min. This flow rate optimizes chromatographic separation efficiency, shortens analysis time in conjunction with the temperature program, and maintains sharp peak shapes; the high flow rate increases detection throughput and supports rapid quality control.

[0016] Preferably, the linear range of this method is 1.52 mg / ml to 4.55 mg / ml, with a correlation coefficient r greater than 0.999. This wide linear range and high r value ensure the reliability of the method under varying concentrations and make it suitable for different batches of samples.

[0017] Preferably, the repeatability of this method is expressed as relative standard deviation (RSD), with an RSD of 1.5% when n=6. This RSD demonstrates high repeatability, which is superior to existing methods (RSD is often higher than 5%) and meets GMP requirements.

[0018] Preferably, the accuracy of this method is expressed as recovery rate, with the recovery rate of the spiked test solution between 90% and 108%, and the relative standard deviation (RSD) of the recovery rate less than 10%. This recovery rate and RSD demonstrate the high accuracy of this method, which is significantly better than that of titration; the low RSD ensures the reliability of the results and supports regulatory compliance.

[0019] Compared with the prior art, the present invention has the following beneficial effects: The present invention solves the core defects of the prior art by combining gas chromatography with specific parameters and solution preparation scheme. High specificity and sensitivity: It can effectively separate similar impurities, has a low detection limit, and is suitable for low-content samples (3 mg / ml).

[0020] Excellent performance indicators: wide linear range (1.52 mg / ml~4.55 mg / ml), high correlation coefficient (r>0.999), good repeatability (RSD1.5%), and high accuracy (recovery rate 90%~108%, RSD<10%), comprehensively surpassing titration and HPLC.

[0021] Simple to operate and environmentally friendly: This method is highly automated, has a short analysis time (about 18 minutes), and reduces the use of organic solvents (such as diluent optimization), which is in line with the principles of green chemistry.

[0022] Highly practical: It has high reliability in drug quality control and provides an efficient and standardized analytical tool for the production of lornoxicam for injection. Attached Figure Description

[0023] Figure 1 These are gas chromatograms of the solutions used in this invention.

[0024] Figure 2 The linear graph of tromethamine in this invention is shown. Detailed Implementation

[0025] The present invention will now be described in detail through examples. Unless otherwise stated, all raw materials used are commercially available.

[0026] Example 1. Instruments and reagents 1.1 Instrument Configuration Gas chromatograph (equipped with flame ionization detector, FID); Electronic balance (accuracy 0.01mg); 1.2 Reagents and Materials; Methanol (chromatographic grade); Glacial acetic acid (analytical grade); Purified water (meets pharmacopoeia standards); Tromethamine reference standard (purity ≥99%); Lornoxicam solution for injection (containing tromethamine excipient).

[0027] 1.3 Column Specifications Model: DB-624, Dimensions: 60 m × 0.32 mm × 1.80 μm.

[0028] 2. Chromatographic conditions Key Parameter Table 3. Solution preparation process 3.1 Blank solution Composition: 20% glacial acetic acid methanol solution and purified water are mixed at a volume ratio of 1:1.

[0029] Procedure: Accurately measure 5 ml of 20% glacial acetic acid methanol solution and 5 ml of purified water, and mix well.

[0030] 3.2 Diluent Composition: 20% glacial acetic acid methanol solution.

[0031] Procedure: Measure 20 ml of glacial acetic acid, dilute with methanol to 100 ml, and shake well.

[0032] 3.3 Preparation steps of the test solution: a. Measure 5.0 ml of lornoxicam injection solution and place it in a 10 ml volumetric flask; b. Add diluent to the mark and shake well; c. Filter through a 0.22μm filter membrane and collect the filtrate for later use.

[0033] Concentration control: The concentration of tromethamine in the solution is 3.0 mg / ml (actual range 2.7 mg / ml~3.3 mg / ml).

[0034] 3.4 Preparation steps of the reference solution: a. Accurately weigh approximately 30 mg of tromethamine reference standard (actual weighing range: 27-33 mg). b. Place in a 10ml volumetric flask, add 5ml of purified water and sonicate to dissolve (power 200W, duration 5min). c. Dilute to the mark with diluent and shake well. Concentration range: 2.7 mg / ml~3.3 mg / ml.

[0035] 4. Measurement Procedure 4.1 System suitability test: Inject blank solution and blank excipient solution in sequence to confirm that there are no interfering peaks.

[0036] 4.2 Sequence analysis, injection in the following order: Blank solution → Blank excipient solution → Reference solution → Test solution (6 parallel samples).

[0037] 4.3 Chromatographic Recording: Collect peak area data and calculate the content of tromethamine using the external standard method.

[0038] 5. Method Validation Experiment 5.1 Specificity Verification Objective: To confirm that the blank matrix does not interfere with the detection of the target analyte.

[0039] Results: See Appendix Figure 1 As can be seen, neither the blank solution nor the blank excipient showed a chromatographic peak at the retention time of tromethamine, indicating that the method has high specificity.

[0040] 5.2 Linearity Range Verification Procedure: Prepare 5 concentration points (1.52 mg / ml, 2.43 mg / ml, 3.03 mg / ml, 3.94 mg / ml, 4.55 mg / ml), and inject each 3 times.

[0041] data: Conclusion: Refer to Appendix Figure 2 As can be seen, the linear equation Y=18.24X+0.62 (r=0.9999) has an intercept of <2% (far below the 25% limit).

[0042] 5.3 Accuracy Verification Blank solvent: 20% glacial acetic acid methanol solution: water = 1:1.

[0043] Diluent: 20% glacial acetic acid methanol solution Reference solution: Accurately weigh approximately 30 mg of tromethamine, place it in a 10 ml volumetric flask, add 5 ml of water, sonicate to dissolve, dissolve with diluent and dilute to the mark, shake well, and the solution is ready. Prepare two parallel solutions.

[0044] 50% Spiked Test Solution: Measure 5.0 ml of blank excipient, weigh approximately 15 mg of tromethamine reference standard, place them together in a 10 ml volumetric flask, add diluent to dissolve and dilute to the mark, shake well, filter, and collect the filtrate to obtain the test solution. Prepare 3 parallel solutions.

[0045] 100% Spiked Test Solution: Measure 5.0 ml of blank excipient, weigh approximately 30 mg of tromethamine reference standard, place them together in a 10 ml volumetric flask, add diluent to dissolve and dilute to the mark, shake well, filter, and collect the filtrate to obtain the test solution. Prepare 3 parallel solutions.

[0046] 150% spiked test solution: Measure 5.0 ml of blank excipient, weigh approximately 45 mg of tromethamine reference standard, place them together in a 10 ml volumetric flask, add diluent to dissolve and dilute to the mark, shake well, filter, and collect the filtrate to obtain the test solution. Prepare 3 parallel solutions.

[0047] result: Conclusion: The recovery rate was between 93.5% and 100.5% (meeting the requirement of 90% to 108%), and the total RSD was 4.1% <10%, which meets the requirements. The accuracy of this method meets the requirements for quantitative detection.

[0048] 5.4 Precision Verification Repeatability Blank solvent: 20% glacial acetic acid methanol solution: water = 1:1.

[0049] Diluent: 20% glacial acetic acid methanol solution.

[0050] Test solution: Measure 5.0 ml of the drug solution (containing approximately 30 mg of tromethorphan) into a 10 ml volumetric flask, dilute to the mark with diluent, shake well, filter, and collect the filtrate to obtain the test solution. Prepare 6 parallel solutions.

[0051] Reference solution: Accurately weigh approximately 30 mg of tromethamine, place it in a 10 ml volumetric flask, add 5 ml of water, sonicate to dissolve, dilute to the mark with diluent, and shake well. Prepare two parallel solutions.

[0052] intermediate precision Blank solvent: 20% glacial acetic acid methanol solution: water = 1:1.

[0053] Diluent: 20% glacial acetic acid methanol solution.

[0054] Test solution: Measure 5.0 ml of the drug solution (containing approximately 30 mg of tromethorphan) into a 10 ml volumetric flask, dilute to the mark with diluent, shake well, filter, and collect the filtrate to obtain the test solution. Prepare 6 parallel solutions.

[0055] Reference solution: Accurately weigh approximately 30 mg of tromethamine, place it in a 10 ml volumetric flask, add 5 ml of water, sonicate to dissolve, dilute to the mark with diluent, and shake well. Prepare two parallel solutions.

[0056] Repeatable results Precision results Conclusion: Two experimenters each prepared 6 test solutions in parallel. The content of tromethamine in the solutions was between 90.0% and 108%, with RSD (n=6) < 4% and RSD (n=12) < 4%, which met the requirements and showed good precision.

[0057] 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. An analytical method for determining the content of tromethamine in lornoxicam for injection, characterized in that: Gas chromatography was used for detection. The chromatographic column was DB-624 60m×0.32mm×1.80μm. The temperature program was as follows: initial column temperature 150℃, increased to 240℃ at a rate of 30℃ / min, and held for 8 minutes. The detector was a flame ionization detector with a detector temperature of 250℃ and an injection port temperature of 240℃. The split ratio was 10:1, and the injection volume was 2μl. External standard method was used for quantitative analysis.

2. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 1, characterized in that: It also includes the preparation of a solution before gas chromatography detection, wherein the solution includes a blank solution, a diluent, a test solution and a reference solution; wherein the blank solution is prepared by mixing 20% ​​glacial acetic acid methanol solution and water at a volume ratio of 1:1, and the diluent is 20% glacial acetic acid methanol solution.

3. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 2, characterized in that: The test solution is prepared by measuring 5.0 ml of lornoxicam injection solution into a 10 ml volumetric flask, adding diluent to dilute to the mark, shaking well, filtering, and taking the filtrate as the test solution.

4. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 3, characterized in that: The concentration of tromethamine in the test solution was 3 mg / ml.

5. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 2, characterized in that: The preparation method of the reference solution is as follows: weigh 27mg~33mg of tromethamine reference standard, place it in a 10ml volumetric flask, add 5ml of water and sonicate to dissolve, then dilute to the mark with diluent and shake well.

6. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 5, characterized in that: The concentration range of tromethamine in the reference solution is 2.7 mg / ml to 3.3 mg / ml.

7. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 1, characterized in that: The flow rate of the gas chromatograph was 4.0 ml / min.

8. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 1, characterized in that: The linear range of the method is 1.52 mg / ml to 4.55 mg / ml, and the correlation coefficient r is greater than 0.

999.

9. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 1, characterized in that: The repeatability of the method is expressed as relative standard deviation, with an RSD of 1.5% when n=6.

10. The analytical method for determining the content of tromethamine in lornoxicam for injection according to claim 1, characterized in that: The accuracy of the method is expressed as recovery rate, with the recovery rate of the spiked test solution between 90% and 108%, and the relative standard deviation of the recovery rate less than 10%.

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