Method for determining content of red light impurities in paracetamol bulk drug

Abstract

The invention discloses a method for determining the content of red light impurities in a paracetamol raw material medicine. The method comprises the following steps: (1) preparing a solution: (1.1) preparing a diluent with the pH value of 11-14 from alkali and pure water; (1.2) preparing a contrast solution of the red light impurity indophenol by using a diluent; (1.3) preparing a test solution by using a diluent; (2) setting detection conditions: detecting by adopting an ultraviolet and visible spectrophotometer; the spectrum scanning range is that spectrum scanning is carried out in the range of 200-800 nm, and the maximum absorption wavelength of indophenol is 629 nm + / -3 nm; (3) respectively measuring the contrast solution and the test solution under the spectrum condition in the step (2) to obtain absorbance values of the contrast solution and the test solution; and (4) calculating the content of the red light impurity indophenol in the test sample through a reference substance comparison method. The method disclosed by the invention can be used for rapidly, accurately and stably measuring the content of the red light impurity indophenol in paracetamol.

Description

Technical Field

[0001] This invention relates to a method for determining the content of red light impurities in acetaminophen raw materials, belonging to the field of pharmaceutical analysis technology. Background Technology

[0002] Paracetamol is an excellent antipyretic and analgesic. The most representative method for its preparation involves a high-temperature acylation reaction of p-aminophenol with acetic acid in an acetic acid-water solvent, followed by recrystallization and decolorization. During the preparation of paracetamol, both the starting materials and the finished product contain highly reactive groups such as hydroxyl and amino groups. Since the preparation and post-processing are carried out at high temperatures, numerous impurities are generated during production, post-processing, and storage. These impurities, in addition to those specified by European and American standards and the Chinese Pharmacopoeia Commission, also include some color-changing impurities, especially a reddish one. This impurity causes the outer layer of freshly prepared white paracetamol to gradually turn reddish after prolonged exposure to air, and this red color is glossy; hence, it is called a reddish impurity.

[0003] There are existing literature reports on the study of red-light impurities. *Qilu Pharmaceutical Affairs*, 1984, 3(02): 34-35, reports that this red-light impurity is indophenol. Publication number CN111635333A describes the preparation of a mixture of red-light impurities in paracetamol, with indophenol being the main chromogenic impurity. Regarding the detection of the red-light substance indophenol, *Chinese Drug Standards*, 2004(6): 24-25+60 reports that the presence of red-light impurities can be confirmed visually, but this method only meets qualitative requirements. Publication number CN 111610158A reports that indophenol, a red-light impurity in paracetamol, can be determined by HPLC. Since the maximum absorption wavelength of indophenol is 629 nm, when the wavelength of the instrument used for HPLC determination is adjusted to this range, the baseline noise is very large. Furthermore, since indophenol is relatively stable under alkaline conditions, especially at pH greater than 10, these alkaline conditions exceed the pH range applicable to ordinary HPLC columns (generally pH 2-8). Therefore, the detection results obtained by HPLC determination are not accurate enough and can only be used to detect the presence of red-light impurity indophenol in paracetamol, but cannot be used to accurately determine the content of red-light impurity indophenol. Summary of the Invention

[0004] Purpose of the invention: The purpose of this invention is to provide a method for accurately, rapidly, and stably determining the content of the red-light impurity indophenol in acetaminophen.

[0005] Technical solution: The method for determining the content of red light impurities in acetaminophen raw material according to the present invention includes the following steps: (1) Preparation of solution: (1.1) Prepare a diluent with a pH of 11-14 using alkali and pure water; (1.2) Prepare a control solution of red light impurity indophenol using a diluent. The concentration of red light impurity indophenol in the control solution is 2.5 µg of red light impurity indophenol per 1 mL of control solution. (1.3) Prepare the test solution with a diluent. The concentration of the test solution is: 10 mg of test sample per 1 mL of test solution; (2) Set the detection conditions: Detection was performed using a UV-Vis spectrophotometer; spectral scanning range: 200–800 nm, with the maximum absorption wavelength of indophenol at 629 nm ± 3 nm; slit width: 2.0 nm; integration time: 0.1 seconds; detector unit: standard; scanning speed: medium speed; sampling interval: 0.5; scanning mode: single. (3) The prepared diluent, control solution and test solution are measured under the spectral conditions in step (2) to obtain the absorbance values ​​of the control solution and the test solution; (4) The content of the red-light impurity indophenol in the test sample was calculated by the reference standard comparison method. The calculation formula used is as follows: ; In the formula, X% represents the content (%) of the red-light impurity indophenol in the test sample; C R : Concentration of red-light impurity indophenol in the control solution (µg / mL); C X : Concentration of the test solution (µg / mL); A X : Absorbance value of indophenol, a red-light impurity, in the test solution; A R : Absorbance value of indophenol, a red light impurity in the control solution.

[0006] In step (1), the alkali is sodium hydroxide or potassium hydroxide, preferably sodium hydroxide; the pH of the diluent is 13.

[0007] In step (2), the spectral scanning range is 400~800 nm; the maximum absorption wavelength of indophenol is 629 nm±1 nm; and the UV-Vis spectrophotometer is a Shimadzu UV-2700 UV-Vis spectrophotometer or a Shimadzu UV-2550 UV-Vis spectrophotometer.

[0008] Since the maximum absorption wavelength of the red-light impurity indophenol is about 629 nm, while the maximum absorption wavelengths of acetaminophen raw material and other impurities are all in the ultraviolet region and below 400 nm, they will not interfere with the determination of the red-light impurity indophenol. The absorption peak formed at 629 nm will only be the red-light impurity indophenol. Therefore, this invention can achieve rapid and accurate quantitative detection of unstable red-light impurity indophenol in acetaminophen.

[0009] Beneficial Effects: Compared with existing technologies, this invention has the following significant advantages: This invention can be used for the quantitative detection of indophenol, a red-light impurity, in paracetamol raw materials, thereby helping to quantitatively analyze and control the formation of unstable red-light impurity indophenol during the preparation of paracetamol raw materials. The method of this invention uses ultraviolet spectrophotometry, which has the advantages of being rapid, accurate, stable, highly repeatable, and highly sensitive. Attached Figure Description

[0010] Figure 1 The relationship between the pH of the test solution and the maximum absorption wavelength; Figure 2 The images show the UV spectra of the diluent, control solution, and test solution; where a is the diluent, b is the control solution, and c is the test solution. Figure 3 This is a linear regression curve. Detailed Implementation

[0011] The source of the red-light impurity indophenol involved in the embodiments of this invention is: red-light impurity indophenol, Nanjing Xize Pharmaceutical Technology Co., Ltd., with a purity of 96.5%. Paracetamol raw material is selected from the following three batches: batch numbers CW-2305087, CW-2305088, and CW-2305089; crude paracetamol is selected from the following four batches: batch numbers 22040095, 22050105, 22050106, and 22050107, all manufactured by Lianyungang Kangle Pharmaceutical Co., Ltd.

[0012] Example 1 The present invention provides a method for determining the content of red light impurities in acetaminophen raw material, comprising the following steps: (1) Preparation of solution: (1.1) Prepare a diluent with a pH of 13 using sodium hydroxide and pure water; Preparation of pH=13 diluent: Weigh 0.4g of sodium hydroxide accurately and place it in a 100mL volumetric flask. Dissolve and dilute to the mark with pure water, mix well, and prepare a solution containing 4.0 mg of sodium hydroxide per 1mL of solution. This is the pH=13 diluent. (1.2) Prepare a reference solution of red light impurity indophenol using a diluent. Specifically, take 50 mg of red light impurity indophenol reference standard, accurately weigh it and place it in a 200 mL volumetric flask. Dissolve and dilute it to the mark with a diluent, mix well, then accurately pipette 1 mL of the solution into a 100 mL volumetric flask, dilute it to the mark with a diluent, mix well, and finally prepare a solution with 2.5 µg of red light impurity indophenol per 1 mL of solution as the reference solution. (1.3) Prepare the test solution with diluent as follows: accurately weigh 100 mg of the test sample and place it in a 10 mL volumetric flask. Dissolve and dilute to the mark with diluent, mix well, and prepare a solution in which 10 mg of the test sample is contained in 1 mL of solution. (2) Set the detection conditions: Detection was performed using a UV-Vis spectrophotometer; spectral scanning range: 400–800 nm, with the maximum absorption wavelength of indophenol at 629 nm ± 1 nm; slit width: 2.0 nm; integration time: 0.1 seconds; detector unit: standard; scanning speed: medium speed; sampling interval: 0.5; scanning mode: single. (3) The prepared diluent, control solution and test solution are measured under the spectral conditions in step (2), and the ultraviolet spectra are recorded to obtain the absorbance values ​​of the control solution and the test solution; (4) The content of the red-light impurity indophenol in the test sample was calculated by the reference standard comparison method. The calculation formula used is as follows: (I) In the formula, X% represents the content (%) of the red-light impurity indophenol in the test sample; C R : Concentration of red-light impurity indophenol in the control solution (µg / mL); C X : Concentration of the test solution (µg / mL); A X : Absorbance value of indophenol, a red-light impurity, in the test solution; A R : Absorbance value of indophenol, a red light impurity in the control solution.

[0013] Example 2 The content of the red-light impurity indophenol in six batches of samples was determined using the method described in Example 1. Specifically: Take three batches of crude paracetamol (batch numbers: 22050105, 22050106, 22050107) and three batches of paracetamol raw materials (batch numbers: CW-2305087, CW-2305088, CW-2305089), and prepare diluent (pH=13 diluent), control solution (2.5µg / mL) and test solution (10mg / mL) according to the method of step (1) in Example 1. Spectral conditions were measured according to the method of step (2) in Example 1, and absorbance values ​​were recorded. The content of red light impurity indophenol in the test solution was calculated using formula (I). The results of the determination of the content of red light impurity indophenol are shown in Table 1.

[0014] Table 1. Determination results of indophenol content (a red light impurity) in 6 batches of test samples

[0015] As shown in Table 1, the red-light impurity indophenol in the three batches of crude paracetamol with batch numbers 22050105, 22050106, and 22050107 were 0.011%, 0.011%, and 0.010%, respectively. The red-light impurity indophenol was not detected in the three batches of paracetamol raw materials with batch numbers CW-2305087, CW-2305088, and CW-2305089.

[0016] Example 3 To prepare a diluent with a pH of 1-14, the specific steps are as follows: Preparation of diluent for pH=14: Accurately weigh 4.0 g of sodium hydroxide and place it in a 100 mL volumetric flask. Dissolve and dilute to the mark with pure water, mix well, and prepare a solution containing 40 mg of sodium hydroxide per mL. This is the diluent for pH=14. Preparation of diluent for pH=13: Accurately weigh 0.4 g of sodium hydroxide and place it in a 100 mL volumetric flask. Dissolve and dilute to the mark with pure water, mix well, and prepare a solution containing 4.0 mg of sodium hydroxide per mL. This is the diluent for pH=13.

[0017] Preparation of diluents with pH=1~12: Take 20 mL of diluent with pH=14, add an appropriate amount of hydrochloric acid, and dilute to prepare diluent solutions with pH=1~12 respectively.

[0018] Using the diluents with pH values ​​of 1-14 as solvents, 250 mg of crude paracetamol (batch number 22040095) was diluted to prepare test solutions (25 mg / mL) with pH values ​​of 1-14 according to step (1) of Example 1. The maximum absorption wavelength and absorbance of the test solutions were measured under the set spectral conditions (spectral scanning range: 400-800 nm; maximum absorption wavelength of indophenol: 629 nm ± 1 nm; slit width: 2.0 nm; integration time: 0.1 seconds; detector unit: standard; scanning speed: medium speed; sampling interval: 0.5; scanning mode: single). The results showed that within the wavelength range of 400-800 nm, in the test solutions prepared with diluents with pH values ​​of 12-14, the red-light impurity indophenol exhibited a stable characteristic absorption peak at 629 nm. When the test solution was prepared with diluents with pH value of 13, the absorbance of the red-light impurity indophenol was the highest at 629 nm. Therefore, the method of this invention selects a sodium hydroxide solution with pH=13 as the diluent for the test solution. Its ultraviolet spectrum is shown below. Figure 1 .

[0019] Example 4 Prepare the diluent (pH=13), control solution (2.5µg / mL), and test solution (10mg / mL) according to step (1) of Example 1; take the diluent, control solution, and test solution, and measure them according to the spectral conditions set in step (2) of Example 1, and record the spectra; see the UV spectra of the diluent, control solution, and test solution. Figure 2 .

[0020] Depend on Figure 2 It is known that the maximum absorption wavelength of the red light impurity indophenol is about 629 nm, while the maximum absorption wavelengths of acetaminophen raw material and other impurities are all in the ultraviolet region, and the maximum absorption wavelengths are all below 400 nm. They do not interfere with the determination of the red light impurity indophenol, indicating that the method of the present invention has good specificity.

[0021] Example 5 Prepare the diluent (pH=13 diluent) according to step (1) of Example 1; at the same time, prepare the impurity reference stock solution with the diluent; specifically: take 50 mg of red light impurity indophenol reference standard, accurately weigh it and put it in a 10 mL volumetric flask, dissolve and dilute it to the mark with the diluent, mix well, then accurately pipette 1 mL into a 100 mL volumetric flask, dilute it to the mark with the diluent, mix well, and prepare a solution containing 50 μg of red light impurity indophenol per 1 mL of solution, as the impurity stock solution.

[0022] Linear solutions 1 to 9 were prepared by adding different amounts of diluent to the impurity stock solution, as shown in Table 2.

[0023] Table 2

[0024] Take the prepared diluent and linear solutions 1-9 as described above, and measure them according to the spectral conditions set in step (2) of Example 1, and record the spectra. Perform linear regression with absorbance (A) as the ordinate and the concentration of the red light impurity indophenol (C) as the abscissa to obtain the regression equation, see [link to relevant documentation]. Figure 3 The results of the linear experiment are shown in Table 3.

[0025] Table 3

[0026] As shown in Table 3, the concentration of the red light impurity indophenol in the range of 0.01001 to 10.0046 µg / mL showed a good linear relationship between concentration and absorbance (n=9), with r>0.999.

[0027] Based on the slope of the linear regression equation described above, the detection limit was calculated to be 1.48 ng / mL. Therefore, the detection limit of this method is 4.45 ng / mL, and the quantitation limit is 14.8 ng / mL. This indicates that the method of the present invention has good sensitivity.

[0028] Example 6 The control solution (2.5 µg / mL) was prepared according to the method of step (1) in Example 1. The absorbance was measured 6 times consecutively under the spectral conditions of step (2) in Example 1, and the absorbance value A was recorded. The results of the repeatability test are shown in Table 4.

[0029] Table 4

[0030] As shown in Table 4, the RSD (n=6) of the absorbance value of the colorimetric impurity indophenol in the control solution is 0.109%, indicating that the method of the present invention has good repeatability.

[0031] Example 7 Two researchers and two ultraviolet spectrometers conducted experiments under the spectral conditions set in step (2) of Example 1, and recorded the spectra. The content of the red-light impurity indophenol in the test solution was calculated based on formula (I), and the results are shown in Table 5.

[0032] Table 5

[0033] As shown in Table 5, the RSD (n=4) of the colorimetric impurity indophenol determined by the two researchers was 1.148%, indicating good intermediate precision.

[0034] Example 8 Prepare the diluent (pH=13), control solution (2.5µg / mL), and test solution (10 mg / mL) of batch number 22040095 according to step (1) of Example 1. Place the diluent, control solution, and test solution at room temperature for 24 h, and take samples at 0, 3, 6, 9, 12, 15, 18, 21, and 24 h respectively. Spectroscopic measurements are performed under the conditions described in step (2) of Example 1, and the chromatograms are recorded. The absorbance of the red-light impurity indophenol in the control solution and test solution is statistically analyzed. The stability test results of the control solution and test solution are shown in Tables 6 and 7.

[0035] Table 6 Results of stability test of control solution

[0036] As shown in Table 6, after the control solution was placed at room temperature for 24 h, the absorbance value of the colorimetric impurity indophenol did not change significantly compared with 0 h, and the RSD (n=9) of the absorbance value was 0.526%, indicating that the control solution was basically stable within 24 h at room temperature.

[0037] Table 7 Results of the stability test of the test sample solution

[0038] As shown in Table 7, after the test solution was placed at room temperature for 24 h, the absorbance value of the red light impurity indophenol did not change significantly compared with 0 h. The RSD (n=9) of the absorbance value was 0.964%, indicating that the test solution was basically stable within 24 h at room temperature.

[0039] Example 9 Prepare the diluent (pH=11) and the test solution (10 mg / mL) of batch number 22040095 according to the method of step (1) in Example 1; place the above diluent and test solution at room temperature for 24 h respectively, and take samples at 0, 3, 6, 9, 12, 15, 18, 21 and 24 h respectively, and determine them according to the spectral conditions of step (2) in Example 1, and record the chromatograms. Statistically analyze the absorbance of the red light impurity indophenol in the test solution, and the results of the stability test of the test solution are shown in Table 8.

[0040] Table 8

[0041] As shown in Table 8, after the test solution was placed at room temperature for 24 h, the absorbance of the red-light impurity indophenol showed a slow decreasing trend over time. The relative standard deviation (RSD) of the absorbance measurements at each time point (n=9) was 5.00%, indicating that the stability of the test solution prepared with pH=11 and the test solution prepared with pH=13 decreased significantly within 24 h.

[0042] Example 10 Prepare the diluent (pH=13 diluent) and control solution (2.5µg / mL) according to step (1) of Example 1. Prepare the impurity control stock solution (50µg / mL) according to the method of Example 5. Accurately weigh 100mg of the test sample with batch number 22040095, and accurately weigh 9 portions in parallel. Place each portion in a 10mL volumetric flask. Divide the 9 volumetric flasks into 3 groups. Add 0.8mL, 1mL, and 1.2mL of the impurity control stock solution to each group of volumetric flasks, respectively. Then dissolve and dilute to the mark with the diluent (pH=13), mix well, so that each 1mL of the final solution contains 4µg, 5µg, and 6µg of indophenol reference standard, respectively, thus obtaining 3 portions of recovery solutions with limit concentrations of 80%, 100%, and 120%. Take the diluent, control solution, and the above 9 recovery solutions and measure them under the spectral conditions of step (2) of Example 1. Record the absorbance values. The recovery test results are shown in Table 9.

[0043] Table 9

[0044] As shown in Table 9, the recovery rate of the colorimetric impurity indophenol in the test solution was between 99.32% and 100.57%, and the RSD (n=9) was 0.387%, indicating that the method of the present invention has good accuracy in determining the content of the red-light impurity indophenol in acetaminophen raw material.

Claims

1. A method for determining the content of red light impurity in paracetamol drug substance, characterized in that, Comprising the following steps: (1) Preparation of solution: (1.1) Preparation of diluent with pH of 11-14 by alkali and pure water; (1.2) Preparation of control solution of red impurity indophenol with diluent, the concentration of red impurity indophenol in the control solution is: 2.5 μg of red impurity indophenol per 1 mL of control solution; (1.3) Preparation of test sample solution with diluent, the concentration of test sample solution is: 10 mg of test sample per 1 mL of test sample solution; (2) Setting detection conditions: Detection is performed by ultraviolet visible spectrophotometer; spectral scanning range: spectral scanning is performed in the range of 200-800 nm, the maximum absorption wavelength of indophenol is 629 nm±3 nm; slit width: 2.0 nm; integration time: 0.1 second; detector unit: standard; scanning speed: medium speed; sampling interval 0.5; scanning mode: single; (3) The prepared diluent, control solution and test sample solution are respectively detected under the spectral conditions of step (2) to obtain the absorbance values of the control solution and the test sample solution; (4) The content of red impurity indophenol in the test sample is calculated by control comparison method, and the calculation formula used is: ； In the formula, X%: content of red light impurity indophenol in the test sample (%); C R : concentration of red light impurity indophenol in the control solution (µg / mL); C X : concentration of the test sample solution (µg / mL); A X : absorbance value of red light impurity indophenol in the test sample solution; A R : absorbance value of red light impurity indophenol in the control solution.

2. A method of determining the content of red light impurity in paracetamol drug substance as claimed in claim 1, wherein: In step (1), the pH of the diluent is 13.

3. A method of determining the content of red light impurity in paracetamol drug substance as claimed in claim 1, wherein: In step (1), the alkali is sodium hydroxide or potassium hydroxide.

4. A method of determining the content of red light impurity in paracetamol drug substance as claimed in claim 1, wherein: In step (2), the spectral scanning range is 400-800 nm.

5. A method of determining the content of red light impurity in paracetamol drug substance as claimed in claim 1, wherein: In step (2), the maximum absorption wavelength of indophenol is 629 nm±1 nm.

6. A method of determining the content of red light impurity in paracetamol drug substance as claimed in claim 1, wherein: In step (2), the ultraviolet visible spectrophotometer is Shimadzu UV-2700 type ultraviolet visible spectrophotometer or Shimadzu UV-2550 type ultraviolet visible spectrophotometer.

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