Method for detecting nitrite in metronidazole bulk drug

Abstract

The invention discloses a method for detecting nitrite in a metronidazole bulk drug. According to the method, an ultraviolet spectrophotometric method is adopted, diazotization with sulfanilamide is carried out, then color development with N-(1-naphthyl) ethylenediamine hydrochloride is carried out, and the content of nitrite is detected at the wavelength of 530-534 nm. According to the detection method, the content of nitrite in the metronidazole bulk drug can be rapidly and accurately detected, and the method is high in recovery rate and sensitivity and good in precision, repeatability and linearity.

Description

Technical Field

[0001] The invention belongs to the field of chemical analysis and detection technology, specifically relating to a method for detecting nitrite in metronidazole raw material. Background Technology

[0002] Nitrites are substances containing nitrite anions (NO2). - Nitrite is a salt of nitrite and has strong oxidizing properties. When secondary or tertiary amine structures are present in a drug, nitrosamine impurities may form, such as NDMA and NDEAD, which are highly carcinogenic substances. Controlling the nitrite content in raw materials can indirectly control the formation of nitrosamine impurities in drugs.

[0003] Metronidazole tablets, metronidazole injections, and other products are prone to degradation of their raw material, metronidazole (a nitroimidazole compound), during sterilization, light exposure, or storage, producing nitrite (NO2). - Since nitrite in metronidazole affects product quality, it is necessary to develop a rapid and accurate detection method. Summary of the Invention

[0004] To address the above problems, this invention provides a method for detecting nitrite in metronidazole raw material.

[0005] The specific steps of the detection method are as follows: (1) Blank solution: Take 2 ml of p-aminobenzenesulfonic acid solution, put it in a 50 ml volumetric flask, add an appropriate amount of solvent, shake, let stand for 5 min, then add 1 ml of naphthylethylenediamine hydrochloride solution, dilute to the mark with solvent, shake well, and let stand for 15 min.

[0006] (2) Reference stock solution: Weigh an appropriate amount of sodium nitrite accurately, place it in a 200ml volumetric flask, add an appropriate amount of water and sonicate to dissolve the sodium nitrite, dilute with water to the mark, and shake well. Accurately measure 5ml, place it in a 100ml volumetric flask, dilute with water to the mark, and shake well.

[0007] (3) Reference solution: Accurately measure 2 ml of the reference stock solution, place it in a 50 ml volumetric flask, add 2 ml of p-aminobenzenesulfonic acid solution, shake well, add an appropriate amount of solvent, shake well, let stand for 5 min, then add 1 ml of naphthylethylenediamine hydrochloride solution, dilute to the mark with solvent, shake well, and let stand for 15 min.

[0008] (4) Test solution: Take an appropriate amount of metronidazole, accurately weigh it, put it in a 50ml volumetric flask, add 2ml of p-aminobenzenesulfonic acid solution, shake well, add an appropriate amount of solvent, shake well, let stand for 5min, add 1ml of hydrochloric acid naphthylethylenediamine solution, dilute to the mark with solvent, shake well, and let stand for 15min.

[0009] (5) Determination of the content of the test solution: Take the test solution and the reference solution and measure the absorbance by ultraviolet-visible spectrophotometry. Calculate the content of nitrite in the test solution according to the external standard method.

[0010] In steps (1), (2), (3) and (4), the p-aminobenzenesulfonic acid solution needs to be prepared using 20% ​​hydrochloric acid solution with a concentration of 3.6 mg / mL to 4.4 mg / mL.

[0011] In steps (1), (2), (3) and (4), the naphthylethylenediamine hydrochloride solution is an aqueous solution with a concentration of 1.8 mg / mL to 2.2 mg / mL.

[0012] The solvent added in steps (1), (3) and (4) is water.

[0013] In step (2), the amount of sodium nitrite sample is 67.5 mg to 82.5 mg.

[0014] In step (4), the amount of metronidazole weighed is 450mg to 550mg.

[0015] In step (5), the ultraviolet-visible spectrophotometric detection wavelength is 530nm to 534nm. Attached Figure Description

[0016] Figure 1 This is a linear graph of nitrite. Detailed Implementation

[0017] The following description, in conjunction with specific embodiments, further illustrates the present invention, but the scope of protection of the present invention is not limited thereto.

[0018] (1) Specificity and system applicability The solution preparation is shown in the table below:

[0019] The absorbance of the blank solution and the reference solution was measured at a wavelength of 532 nm. The results are shown in the table below:

[0020] Conclusion: The absorbance of the blank solution at 532 nm was 0.000, which did not interfere with the detection. Five consecutive measurements of the absorbance of reference solution 1 showed an RSD of 0.085%, and the recoveries of the response factors for both reference solutions were 100.2%. This method demonstrates good specificity and system suitability.

[0021] (2) Repeatability Blank solution: Take 2 ml of p-aminobenzenesulfonic acid solution, place it in a 50 ml volumetric flask, add an appropriate amount of water, shake, let stand for about 5 minutes, then add 1 ml of naphthylethylenediamine hydrochloride solution, dilute with water to the mark, shake well, and let stand for 15 minutes to obtain the blank solution.

[0022] Reference solution: Accurately weigh approximately 75 mg of sodium nitrite and place it in a 200 ml volumetric flask. Add an appropriate amount of water and sonicate for 10 min to dissolve the sodium nitrite. Dilute with water to the mark and mix well. Accurately measure 5 ml of the solution and place it in a 100 ml volumetric flask. Dilute with water to the mark and mix well. Accurately measure 2 ml of the solution and place it in a 50 ml volumetric flask. Add 2 ml of p-aminobenzenesulfonic acid solution and mix well. Add an appropriate amount of water and mix well. Let stand for 5 min. Then add 1 ml of naphthylethylenediamine hydrochloride solution and dilute with water to the mark. Mix well and let stand for 15 min. (Prepare two parallel portions) Test solution: Accurately weigh approximately 500 mg of this product and place it in a 50 ml volumetric flask. Add 2 ml of p-aminobenzenesulfonic acid solution, shake well, add an appropriate amount of water, shake well, and let stand for about 5 minutes. Add 1 ml of naphthylethylenediamine hydrochloride solution, dilute with water to the mark, shake well, and let stand for 15 minutes. (Prepare 6 parallel solutions) Take the blank solution, reference solution, and test solution, and measure the absorbance at a wavelength of 532 nm. The results are shown in the table below:

[0023] Conclusion: The absolute value of the range of nitrite content in the six test solutions was 0.000001%, indicating that the method has good repeatability.

[0024] (3) Intermediate precision Blank solution: Take 2 ml of p-aminobenzenesulfonic acid solution, place it in a 50 ml volumetric flask, add an appropriate amount of water, shake, let stand for about 5 minutes, then add 1 ml of naphthylethylenediamine hydrochloride solution, dilute with water to the mark, shake well, and let stand for 15 minutes to obtain the blank solution.

[0025] Reference solution: Accurately weigh approximately 75 mg of sodium nitrite and place it in a 200 ml volumetric flask. Add an appropriate amount of water and sonicate for 10 min to dissolve the sodium nitrite. Dilute with water to the mark and mix well. Accurately measure 5 ml of the solution and place it in a 100 ml volumetric flask. Dilute with water to the mark and mix well. Accurately measure 2 ml of the solution and place it in a 50 ml volumetric flask. Add 2 ml of p-aminobenzenesulfonic acid solution and mix well. Add an appropriate amount of water and mix well. Let stand for 5 min. Then add 1 ml of naphthylethylenediamine hydrochloride solution and dilute with water to the mark. Mix well and let stand for 15 min. (Prepare two parallel portions) Test solution: Accurately weigh approximately 500 mg of this product and place it in a 50 ml volumetric flask. Add 2 ml of p-aminobenzenesulfonic acid solution, shake well, add an appropriate amount of water, shake well, and let stand for about 5 minutes. Add 1 ml of naphthylethylenediamine hydrochloride solution, dilute with water to the mark, shake well, and let stand for 15 minutes. (Prepare 6 parallel solutions) Take the blank solution, reference solution, and test solution, and measure the absorbance at a wavelength of 532 nm. The results are shown in the table below:

[0026] Conclusion: The absolute value of the range of nitrite content in the 12 test solutions from the two experiments was 0.000029%, indicating that the method has good precision.

[0027] (4) Linear The solution preparation is shown in the table below:

[0028] The absorbance of each linear solution was measured at a wavelength of 532 nm, and the results are shown in the table below:

[0029] Conclusion: Within the concentration range of 0 μg / ml (0%) to 0.75434 μg / ml (150%), the linear equation for nitrite is y = 1.0187x + 0.0096, the correlation coefficient r is 0.9997, the percentage of the residual standard deviation relative to the 100% response value is 1.5%, and the sum of squared residuals is 0.0002427. The linear relationship is good under this method.

[0030] (5) Limit of detection and limit of quantitation According to the "Guideline for Validation of Analytical Methods 9101" in the 2020 edition of the Chinese Pharmacopoeia, the limits of detection and quantitation were calculated using the residual standard deviation and slope of the standard curve under the linearity and range section, based on the standard deviation of the response value and the slope of the standard curve. The results are shown in the table below:

[0031] Conclusion: The detection limit for nitrite is 0.02523 μg / ml, which is equivalent to 5.0% of the limit concentration and 0.00025% of the concentration of the test solution; the quantitation limit is 0.07646 μg / ml, which is equivalent to 15.3% of the limit concentration and 0.00076% of the concentration of the test solution. This method has high sensitivity.

[0032] (6) Accuracy The solution preparation is shown in the table below:

[0033] The absorbance of blank solution, reference solution, and accuracy solution was measured at a wavelength of 532 nm. The results are shown in the table below:

[0034] Conclusion: The recovery rate at the 25% concentration level (n=3) was 101.83%–101.87%, with an RSD of 0.030%; the recovery rates at other concentration levels (n=6) were 95.90%–97.71%, with an RSD of 0.96%. This indicates that the method has good accuracy and can meet the experimental requirements.

Claims

1. A method for detecting a nitrite salt of metronidazole drug substance, characterized by, It comprises the following steps: (1) Blank solution: take 2ml of p-aminobenzenesulfonic acid solution, put it in a 50ml volumetric flask, add appropriate amount of solvent, shake, stand for 5min, then add 1ml of naphthyl ethylenediamine hydrochloride solution, dilute to the mark with solvent, shake well, stand for 15min; (2) Control solution stock solution: take appropriate amount of sodium nitrite, accurately weigh, put it in a 200ml volumetric flask, add appropriate amount of water to dissolve sodium nitrite by ultrasonic, dilute to the mark with water, shake well. Accurately take 5ml, put it in a 100ml volumetric flask, dilute to the mark with water, shake well; (3) Control solution: accurately take 2ml of control solution stock solution, put it in a 50ml volumetric flask, add 2ml of p-aminobenzenesulfonic acid solution, shake well, add appropriate amount of solvent, shake well, stand for 5min, then add 1ml of naphthyl ethylenediamine hydrochloride solution, dilute to the mark with solvent, shake well, stand for 15min; (4) Test solution: take appropriate amount of metronidazole, accurately weigh, put it in a 50ml volumetric flask, add 2ml of p-aminobenzenesulfonic acid solution, shake well, add appropriate amount of solvent, shake well, stand for 5min, add 1ml of naphthyl ethylenediamine hydrochloride solution, dilute to the mark with solvent, shake well, stand for 15min; (5) Determination of the content of the test solution: take the test solution and the control solution to determine the absorbance by ultraviolet-visible spectrophotometry, and calculate the content of nitrite in the test solution according to the external standard method.

2. The method of claim 1, wherein, The p-aminobenzenesulfonic acid solution in steps (1), (2), (3) and (4) needs to be prepared with 20% hydrochloric acid solution, with a concentration of 3.6mg / mL-4.4mg / mL.

3. The detection method according to claim 1, characterized in that, The naphthyl ethylenediamine hydrochloride solution in steps (1), (2), (3) and (4) is a 1.8mg / mL-2.2mg / mL aqueous solution.

4. The method of claim 1, wherein, The solvent added in steps (1), (3) and (4) is water.

5. The detection method according to claim 1, characterized in that, The amount of sodium nitrite in step (2) is 67.5mg-82.5mg.

6. The method of claim 1, wherein, The amount of metronidazole in step (4) is 450mg-550mg.

7. The detection method according to claim 1, characterized in that, The detection wavelength of ultraviolet-visible spectrophotometry in step (5) is 530nm-534nm.

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