A method for determining the content of sulfamethoxazole

Abstract

This invention belongs to the field of sulfonamide antibiotic detection, specifically relating to a method for determining the content of sulfamethoxazole. The method includes the following steps: 1) Sulfamethoxazole standard and water are mixed and sonicated until dissolved to obtain a sulfamethoxazole stock solution with a concentration not exceeding 10 mg / L; standard solutions of different concentrations are prepared using the sulfamethoxazole stock solution and water, absorbance is measured, a standard curve is plotted, and a linear regression equation is obtained; 2) Sulfamethoxazole solid preparation is mixed with water, sonicated to aid dissolution, and filtered to prepare a test solution; or the water sample to be tested is filtered to prepare a test solution; the absorbance of the test solution is measured, and the content of sulfamethoxazole is determined according to the linear regression equation. This invention utilizes ultraviolet spectrophotometry, using water as a solvent, to determine the content of the active ingredient in sulfamethoxazole preparations or the concentration of sulfamethoxazole in environmental water samples. It offers high accuracy, significantly reduces testing costs, and shortens testing time.

Description

Technical Field

[0001] This invention belongs to the field of detection of sulfonamide antibiotics, specifically relating to a method for determining the content of sulfamethoxazole. Background Technology

[0002] Antibiotics are among the most widely used drugs, primarily for the prevention and treatment of bacterial and fungal diseases. In recent years, water pollution caused by antibiotic overuse has drawn attention in some European and American countries. Sulfonamides are one of the most widely used classes of antibiotics, and are derivatives containing the p-aminobenzenesulfonamide structure. These drugs have a broad antibacterial spectrum, inhibiting most Gram-positive and Gram-negative bacteria. However, long-term overuse of sulfonamides can lead to the proliferation of antibiotic-resistant bacteria (ARBs) in animals and the environment, inducing the production of antibiotic resistance genes (ARGs). Once these genes are transferred to human pathogens, they reduce the chances of curing infectious diseases and have a serious impact on human health. Therefore, sensitive, accurate, and rapid analytical methods for residual antibiotics in the aquatic environment are essential for studying their environmental behavior and ecological effects.

[0003] Sulfamethoxazole is a type of sulfonamide antibiotic that has played a significant role in the history of human antibacterial treatment. Due to its broad-spectrum antibacterial properties, stability, low cost, and ease of use, it is widely used in animal husbandry for the treatment and prevention of diseases, and is added to animal feed. Therefore, the detection of sulfamethoxazole formulation content and its concentration in water bodies is of great importance for its rational use and pollution control.

[0004] Existing analytical techniques for antibiotics mainly include ultra-high performance liquid chromatography (UHPLC) with tandem ultraviolet detection, high performance liquid chromatography with tandem fluorescence detection, enzyme-linked immunosorbent assay (ELISA), capillary electrophoresis, gas chromatography-tandem mass spectrometry (GC-MS), liquid chromatography (LC), and GC-MS with tandem mass spectrometry (MS-MS). GC-MS combines the high separation performance of chromatography with the high discrimination characteristics of mass spectrometry, offering advantages such as a wider analytical range, higher sensitivity, lower detection limits, more reliable qualitative structure analysis, and the ability to simultaneously detect multiple contaminants. It is of great significance for the qualitative and quantitative analysis of trace and ultra-trace components in complex compounds and is the most widely used method in antibiotic analysis. However, current detection methods generally suffer from complex processes, long detection times, environmental pollution due to the use of organic reagents such as acetonitrile, and difficulties in ensuring recovery rates, resulting in low efficiency, poor accuracy, and low stability. Summary of the Invention

[0005] The purpose of this invention is to provide a method for determining the content of sulfamethoxazole, which can significantly reduce testing costs, shorten testing time, reduce the probability of deviations during the experiment, and improve detection efficiency.

[0006] To achieve the above objectives, the technical solution of the present invention is as follows:

[0007] A method for determining the content of sulfamethoxazole includes the following steps:

[0008] 1) Sulfamethoxazole standard and water are mixed and sonicated until dissolved to obtain sulfamethoxazole stock solution with a concentration not greater than 10 mg / L; standard solutions of different concentrations are prepared using the sulfamethoxazole stock solution and water, absorbance is measured, standard curves are plotted, and linear regression equations are obtained.

[0009] 2) Mix sulfamethoxazole solid dosage form with water, dissolve with ultrasonic assistance and filter to prepare a test solution; or filter the water sample to be tested to prepare a test solution; measure the absorbance of the test solution, and determine the content of sulfamethoxazole in the sulfamethoxazole solid dosage form or the water sample to be tested according to the linear regression equation determined in step 1).

[0010] The method for determining the sulfamethoxazole content of the present invention utilizes ultraviolet spectrophotometry, with water as the solvent, to determine the content of the active ingredient in sulfamethoxazole preparations or the concentration of sulfamethoxazole in environmental water samples. It has high accuracy, can significantly reduce testing costs, shorten testing time, and makes pretreatment more convenient and faster. At the same time, it can reduce the probability of deviations during the experiment and improve detection efficiency.

[0011] The sulfamethoxazole solid dosage form uses sulfamethoxazole as the main drug, preferably with a sulfamethoxazole content of 90% or more, more preferably 95% or more.

[0012] Preferably, in step 2), the concentration of the detection solution is below 2 mg / L. In actual wastewater, antibiotics are soluble in water, and using water as the detection solvent allows for faster and more accurate detection. Sulfamethoxazole is extremely poorly soluble in water; controlling the concentration of the detection solution to below 2 mg / L ensures the stability of the dissolution and improves detection accuracy.

[0013] Preferably, in steps 1) and 2), the wavelength for measuring absorbance is 265 nm.

[0014] Preferably, in steps 1) and 2), the ultrasonic power is 120W, the frequency is 40kHz, and the duration is 20min. Using ultrasound assistance can improve the solubility of sulfamethoxazole, and combined with the control of the solubility concentration, the detection process can proceed smoothly.

[0015] Preferably, in step 1), the concentration of the standard solution is no greater than 2 mg / L. More preferably, the concentration range of the standard solution is 0.2–2 mg / L. For example, it can be 0.2 mg / L, 0.4 mg / L, 0.8 mg / L, 1.2 mg / L, and 2 mg / L.

[0016] Preferably, in step 2), the pore size of the filter paper used for filtration is 5 μm. Attached Figure Description

[0017] Figure 1 This is the standard curve obtained in Embodiment 1 of the present invention. Detailed Implementation

[0018] This invention provides a method for determining the content of sulfamethoxazole. Water is used as the detection concentration, and the absorbance of the sample is measured by ultraviolet spectrophotometry at a wavelength of 265 nm. The content of sulfamethoxazole is then calculated using the external standard method. The detection process uses only water as a solvent, which is environmentally friendly, safe, non-toxic, and highly accurate compared to the organic solvents used in conventional methods.

[0019] The beneficial effects of adopting the above technical solution are as follows:

[0020] (1) Fast detection speed and high accuracy: It takes at least half an hour to detect one sample by liquid chromatography, which may affect the production schedule when there are too many samples; while the method of this invention can detect at least 6 samples in the same time, which greatly improves the production efficiency.

[0021] (2) Safety and environmental protection: When using liquid chromatography to detect the content of sulfamethoxazole, toxic organic solvents are required. These solvents are harmful to the human body, and long-term exposure to them can easily damage the health of the testing personnel. However, the method of this invention only uses water as a solvent, which is safe, non-toxic, green and environmentally friendly, fast and accurate.

[0022] (3) HPLC systems also ultimately use VWD or DAD ultraviolet detectors to measure absorbance at a specific wavelength, and the detection principle is the same. Furthermore, ultraviolet detection is more time-efficient than HPLC systems. In actual wastewater, antibiotics are generally water-soluble; using water as the detection solvent, with both systems using the same solvent, further reduces solvent interference and improves detection stability and accuracy.

[0023] The embodiments of the present invention will be further described below with reference to specific examples.

[0024] Specific embodiments of the method for determining the sulfamethoxazole content of the present invention are as follows:

[0025] Example 1

[0026] The method for determining the sulfamethoxazole content in this embodiment includes the following steps:

[0027] 1. Establish standard working curves

[0028] Preparation of sulfamethoxazole standard solution: Accurately weigh 10 mg of sulfamethoxazole standard, mix thoroughly with purified water, transfer to a 1 L volumetric flask, and dilute to the mark. Use an ultrasonic cleaner to sonicate for 20 minutes (ultrasonic power 120 W, frequency 40 kHz) to promote drug dissolution, obtaining a 10 mg / L sulfamethoxazole stock solution. Add 0.5, 1.0, 2.0, 3.0, and 5.0 mL of the sulfamethoxazole stock solution to a series of 25 mL colorimetric tubes, respectively, and dilute to 25 mL with purified water to obtain standard solutions with concentrations of 0.2 mg / L, 0.4 mg / L, 0.8 mg / L, 1.2 mg / L, and 2 mg / L. Measure the absorbance at 265 nm to obtain the standard curve, see [reference needed]. Figure 1 .

[0029] Evaluation criteria for the working curve: regression equation R 2 Not less than 0.999.

[0030] Table 1. Confirmation data of linear range of content

[0031]

[0032] Conclusion: Figure 1 As shown, the standard working curve of sulfamethoxazole is linearly consistent with the standard, and the method validation was successful.

[0033] 2. Precision test

[0034] Different concentrations of sulfamethoxazole standard solutions were taken, and the absorbance was measured at 265 nm. The measurements were repeated 6 times, and the relative standard deviations were calculated as shown in Table 2 below.

[0035] Table 2 Relative standard deviation of the method

[0036]

[0037] As shown in Table 2 above, the spike recovery rate of the method of the present invention is between 97.66% and 100.04%, which meets the measurement requirements. The relative standard deviation of precision is within the range of 0.67% to 2.19%, which meets the precision requirements.

[0038] 3. Determination of sulfamethoxazole content:

[0039] Accurately weigh 10 mg of sulfamethoxazole (active drug) into a 1 L volumetric flask, add purified water and mix well, then add water to the mark. Use an ultrasonic cleaner to sonicate for 20 minutes (ultrasonic power 120 W, frequency 40 kHz) to promote drug dissolution, thus preparing the test solution. After filtering with filter paper, pipette 2.5 ml into a 25 ml colorimetric tube, add purified water to the mark, and measure the absorbance three times at 265 nm using a quartz cuvette. The values ​​are 0.060, 0.061, and 0.061, respectively. Based on the absorbance of the sample solution and the linear regression equation, the content of sulfamethoxazole in the test sample can be calculated to be 96.09%, 96.09%, and 94.53%, with an average of 95.57%.

[0040] Experimental Example 2

[0041] The method for determining the sulfamethoxazole content in this experimental example differs from that in Example 1 in that a laboratory-synthesized simulated wastewater is used instead of ultrapure water to test the feasibility of this method for determining the sulfamethoxazole content in water. The simulated wastewater composition is: C2H3O2Na, 1.282 g / L; (NH4)2SO4, 0.47 g / L; K2HPO4·7H2O, 0.022 g / L; KH2PO4, 0.040 g / L. g / L; CaCl2, 0.046 g / L; MgSO4·7H2O, 0.044 g / L; FeSO4·7H2O, 0.018 g / L; NaHCO3, 0.1 g / L; CoCl2·6H2O, 250 μg / L; CuSO4·5H2O, 250 μg / L; MnSO4·H2O, 250 μg / L; NiCl·6H2O, 250 μg / L, solvent: water.

[0042] Water samples containing different amounts of sulfamethoxazole were filtered with filter paper, and then the absorbance was measured three times at 265 nm using a cuvette. Based on the absorbance of the water samples and the linear regression equation, the content of sulfamethoxazole in the water samples could be calculated.

[0043] Table 3. Precision and Spiking Recovery of this Method for Measuring Sulfamethoxazole Content in Water Bodies

[0044]

[0045]

[0046] As shown in Table 3 above, when the method of the present invention is used to measure the content of sulfamethoxazole in water, the spiked recovery rate is between 93.97% and 104.82%, which meets the measurement requirements. The relative standard deviation of precision is within the range of 0.80% to 2.30%, which meets the precision requirements.

[0047] In summary, the results obtained by the method of the present invention are relatively stable and have high accuracy. Therefore, the present invention, a method for determining the content of sulfamethoxazole using ultraviolet spectrophotometry, can be applied to the determination of the content of sulfamethoxazole preparations and the analysis and detection of sulfamethoxazole content in water bodies.

Claims

1. A method for determining the content of sulfamethoxazole, characterized in that, Includes the following steps: 1) Sulfamethoxazole standard and water are mixed and sonicated until dissolved to obtain sulfamethoxazole stock solution with a concentration not greater than 10 mg / L; standard solutions of different concentrations are prepared using the sulfamethoxazole stock solution and water, absorbance is measured, standard curves are plotted, and linear regression equations are obtained. 2) Mix sulfamethoxazole solid dosage form with water, dissolve with ultrasonic assistance and filter to prepare a test solution; or filter the water sample to be tested to prepare a test solution; measure the absorbance of the test solution, and determine the content of sulfamethoxazole in the sulfamethoxazole solid dosage form or the water sample to be tested according to the linear regression equation determined in step 1).

2. The method for determining the content of sulfamethoxazole as described in claim 1, characterized in that, In step 2), the concentration of the test solution is below 2 mg / L.

3. The method for determining the content of sulfamethoxazole as described in claim 1, characterized in that, In steps 1) and 2), the wavelength for measuring absorbance is 265 nm.

4. The method for determining the content of sulfamethoxazole as described in claim 1, characterized in that, In steps 1) and 2), the ultrasonic power is 120W, the frequency is 40kHz, and the duration is 20min.

5. The method for determining the sulfamethoxazole content according to any one of claims 1 to 4, characterized in that, In step 1), the concentration of the standard solution is no greater than 2 mg / L.

6. The method for determining the content of sulfamethoxazole as described in claim 5, characterized in that, The concentration range of the standard solution is 0.2–2 mg / L.

7. The method for determining the content of sulfamethoxazole as described in any one of claims 1 to 4, characterized in that, In step 2), the filter paper used for filtration has a pore size of 5 μm.

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