Amoxicillin solubility visual detection method based on quantitative color scale

Abstract

The application discloses a kind of amoxicillin solubility visual detection methods based on quantitative color scale, specifically related to drug detection field, the present application aims at solving the problems of large standard gradient interval, unstable color development and lack of correction in the existing visual comparison method, method includes: with 0.2M potassium dihydrogen phosphate solution as solvent, preparation amoxicillin saturated solution;Linear regression equation is established, by precisely preparing 9 concentration gradient amoxicillin standard solution, after adding copper sulfate solution, form stable yellow-green turbidity gradient, and draw linear regression equation with absorbance value;Sample determination, the sample liquid is compared with standard gradient and turbidimetric analysis is carried out, the concentration is determined, and the solubility is calculated by combining dilution multiple, the present application converts visual estimation into accurate quantification by high-density gradient standard and linear regression model, significantly reduces matching error, improves anti-interference ability and accuracy, and is suitable for rapid detection in drug research and development and quality control.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of drug detection, more particularly, the present application relates to a kind of amoxicillin solubility visual detection method based on quantitative color step. BACKGROUND

[0002] In the field of amoxicillin solubility determination and similar substance concentration quantitative analysis, the determination idea based on visual comparison has been applied early, and its core logic is to set a series of standard samples with known concentration, form a visual gradient that can be distinguished directly, then compare the sample to be tested with the standard sample visually, select the standard sample with the closest color or turbidity, and use its concentration to approximate the concentration of the sample to be tested, and finally derive the solubility data. This method is simple to operate and does not require complex instruments, so it is widely used in primary laboratories and on-site rapid detection scenarios. The conventional operation process is as follows: first, prepare standard solutions with gradient concentrations, add specific color developing agents after reaction, and form a stable and obviously different visual gradient system. Then, the same color developing process is performed on the amoxicillin saturated solution to be tested, and the sample to be tested is compared with the standard gradient sample one by one with the help of a color comparison device to determine the standard sample concentration closest to the visual characteristics, and the solubility is calculated by combining the dilution factor.

[0003] However, the existing visual comparison-based method has the following significant defects: 1. The standard gradient setting is not reasonable, most schemes only set 3-5 concentration gradients, and the gradient interval is too large, making it difficult to accurately match the sample to be tested with the standard sample, and introducing large calculation deviations due to visual judgment errors; 2. Poor color developing stability, the reaction of the commonly used color developing agent with amoxicillin is significantly affected by temperature and humidity, and the color developing product is prone to fading or precipitation, making it difficult to maintain the standard gradient system stable for a long time, affecting the accuracy of comparison; 3. Lack of control correction mechanism, no blank control tube and positive control tube are set, which cannot eliminate the interference of the color of the solvent and color developing agent on visual judgment, further reducing the reliability of the results. These problems make the existing visual comparison method have insufficient precision in amoxicillin solubility determination, and it is difficult to meet the requirements of data accuracy in drug research and development, quality control, etc. SUMMARY

[0004] In order to overcome the defects of the prior art, the present application provides a kind of amoxicillin solubility visual detection method based on quantitative color step, which solves the problems raised in the background art by the following scheme.

[0005] To achieve the purpose, the present application provides the following technical scheme: a kind of amoxicillin solubility visual detection method based on quantitative color step, comprising:

[0006] S1, sample solution and reagent preparation: (1) preparation of 0.2M potassium dihydrogen phosphate solution: accurately weigh the potassium dihydrogen phosphate powder, dissolve and dilute to prepare 0.2M potassium dihydrogen phosphate solution; (2) preparation of amoxicillin saturated solution: take excess amoxicillin sample powder, add accurate volume of solution to dissolve, and place in 25°C constant temperature shaking for 24 hours to ensure that the dissolution equilibrium is reached to form a saturated solution;

[0007] S2, establishment of linear regression equation: (1) preparation of amoxicillin standard stock solution: accurately weigh the amoxicillin reference substance, dissolve and dilute to prepare amoxicillin standard stock solution with a concentration of 1000μg / mL; (2) preparation of series of standard tubes: accurately measure different volumes of amoxicillin standard stock solution, dilute with solution to obtain a series of standard solutions with concentration gradient; (3) color development and curve establishment: add 0.1% copper sulfate solution to each series of standard tubes, mix well and stand, and each tube will immediately show a clear turbidity gradient from transparent clear liquid to yellow-green, which is the visual linear regression equation for quantitative comparison.

[0008] S3, sample determination and solubility calculation: (1) sample color development: dilute the amoxicillin saturated solution prepared in S1, add equal amount of 0.1% copper sulfate solution, mix well and stand to obtain the test solution; (2) turbidimetric analysis: compare the test solution with the visual linear regression equation established in S2 to determine the closest standard tube concentration corresponding to the turbidity; (3) solubility calculation: calculate the solubility of amoxicillin sample according to the concentration value determined by comparison and the dilution factor.

[0009] Preferably, the preparation process of 0.1% copper sulfate solution is to weigh sufficient copper sulfate pentahydrate into appropriate deionized water, and the obtained solution is unstable in nature and should be stored in the dark. It should be freshly prepared before the experiment to ensure the stability of the reaction activity.

[0010] Preferably, the preparation method of amoxicillin saturated solution is to tightly cap the conical flask containing the sample and solvent, place it in a constant temperature water bath shaker with a pre-set temperature of 25.0±0.5°C, set the shaker speed to 150rpm, and start the shaker for continuous shaking for 24 hours. After 24 hours, carefully take out the conical flask, place it on the experimental table for a while without shaking, and carefully observe the bottom of the bottle from the side and bottom in a good light. If no undissolved solids are observed, it indicates that the initial feed amount is insufficient and saturation has not been reached. At this time, a small amount of sample needs to be added and the shaking continues for several hours until excess solids are confirmed.

[0011] Preferably, the saturated amoxicillin solution needs to use 0.2M potassium dihydrogen phosphate solution as the solvent, and is dissolved at room temperature; in order to ensure that it is completely and quickly dissolved and forms a uniform solution, the following any one of the following methods is used to assist in dissolving: a) mechanical oscillation method: the container is placed on a vortex oscillator, and intermittent or continuous oscillation is carried out at a speed of not less than 2500 rpm until the solution is clear and transparent, and the undissolved powder is no longer reduced; b) ultrasonic treatment method: the container is placed in the water bath of the ultrasonic cleaning machine (power≥300W, frequency 40kHz), and ultrasonic treatment is carried out for 5-10 minutes, or until the solution system is clear and transparent, and the undissolved powder is no longer reduced.

[0012] Preferably, the solvent used for preparing the saturated amoxicillin solution and for diluting the standard solution in the preparation series is 0.2M potassium dihydrogen phosphate solution, so as to ensure the consistency of the system.

[0013] Technical effects and advantages of the present application: 1. High quantitative accuracy: by establishing a high-density gradient standard series and a visual linear regression model, the transition from visual estimation to accurate quantification is realized, the discrete gradient is converted into a linear regression equation, and the matching error caused by the interval of the gradient is significantly reduced;

[0014] 2. Strong color development stability: the standardized phosphate-copper sulfate color development system is used to ensure the reproducibility and durability of the turbidity gradient: specifically, 0.2M potassium dihydrogen phosphate is used as the solvent, and 0.1% copper sulfate is used to form a specific color development system, a stable yellow-green turbidity gradient is formed, and the result is reliable and reproducible.

[0015] 3. Good anti-interference performance: innovative double-control quality control design is adopted to effectively identify and exclude non-specific interference, by setting blank control and positive control, the background color of the system without phosphate and the background turbidity of the system without drug are identified respectively, which significantly improves the specificity of the method and the accuracy of the result. DETAILED DESCRIPTION

[0016] The technical solutions in the embodiments of the present application will be described below in conjunction with the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, not all. Based on the embodiments in the present application, all other embodiments obtained by those skilled in the art without creative labor are within the scope of protection of the present application. Example 1

[0017] S1, sample solution and reagent preparation: (1) prepare 0.2M potassium dihydrogen phosphate solution: accurately weigh potassium dihydrogen phosphate powder, dissolve and dilute to volume, prepare 0.2M potassium dihydrogen phosphate solution; (2) prepare amoxicillin saturated solution: take excess amoxicillin sample powder, add accurate volume of solution to dissolve, constant temperature shaking at 25°C for 24 hours, ensure that the dissolution equilibrium is reached, form a saturated solution.

[0018] Potassium dihydrogen phosphate powder needs to use enough deionized water as solvent, and the dissolution is carried out at room temperature, i.e. 25±5℃, until the solution system is clear and transparent, and it is ensured that there is no undissolved powder; amoxicillin sample uses 0.2M potassium dihydrogen phosphate solution as solvent, and the continuous mild magnetic stirring is carried out under the condition that the environmental temperature is ≤25℃ and the relative humidity is <40%, until the solution powder in the sample is no longer reduced, the upper clear solution is absorbed, and the clear and transparent amoxicillin saturated solution is obtained.

[0019] S2, establishment of linear regression equation: (1) prepare amoxicillin standard stock solution: accurately weigh amoxicillin reference substance, dissolve and dilute to volume, prepare amoxicillin standard stock solution with a concentration of 1000μg / mL; (2) prepare series of standard tubes: accurately measure different volumes of amoxicillin standard stock solution, dilute with solution, and obtain series of standard solutions with concentration gradient; (3) color development and curve establishment: quantitatively add 0.1% copper sulfate solution to each series of standard tubes, mix well and stand, and each tube will immediately show a clear turbidity gradient from transparent clear liquid to yellow-green turbid liquid, which is the visual linear regression equation for quantitative comparison.

[0020] The preparation method of amoxicillin standard stock solution is the same as that of amoxicillin saturated solution, and the concentration of amoxicillin standard stock solution is calculated as follows: C=1000μg / mL, m=μg, P=100%, V=mL; series of amoxicillin standard solutions with concentration gradient, except blank control tube and positive control tube, are set with 9 concentration gradients, and the target concentrations are 100, 200, 300, 400, 500, 600, 700, 800 and 900μg / mL respectively; accurately measure amoxicillin standard stock solution with a concentration of 1000μg / mL, dilute with 0.2M potassium dihydrogen phosphate solution as diluent, and prepare series of standard working solutions with concentration gradient; series of amoxicillin standard solutions with concentration gradient include blank control tube and positive control tube, and the blank control tube is 1000μg / mL amoxicillin solution dissolved with deionized water as solvent, and the blank control tube does not contain potassium dihydrogen phosphate; the positive control tube is 0.2M potassium dihydrogen phosphate solution dissolved with deionized water as solvent, and does not contain amoxicillin.

[0021] Amoxicillin reacts with copper sulfate to generate a colored complex which produces turbidity. The absorbance value of the turbidity at 550 nm wavelength is linear with the concentration of amoxicillin within a certain range. The solution in the blank control tube is yellow-brown and clear after adding 0.1% copper sulfate solution. The solution in the positive control tube is yellow-brown and turbid after adding 0.1% copper sulfate solution, and the precipitation is obvious.

[0022] The visual linear regression equation is established based on the absorbance values of amoxicillin standard solutions with a series of concentration gradients. The absorbance values of each standard tube and sample tube are determined in turn using a UV-visible spectrophotometer at a wavelength of 550 nm, with the blank control tube correcting the instrument zero point. The linear regression equation is plotted with the absorbance values of the standard tubes as the ordinate and the corresponding amoxicillin concentrations as the abscissa. The linear regression equation is y = a x C + b, where y is the absorbance value, C is the amoxicillin concentration with the unit of μg / mL, a is the slope, and b is the intercept, which is obtained by fitting the absorbance values of amoxicillin standard solutions with a series of concentration gradients.

[0023] Table 1

[0024] Amoxicillin standard stock solution addition (ul) Potassium dihydrogen phosphate solution addition (ul) Amoxicillin solution concentration (ug / ml) Blank control tube 1000 0 1000 Tube 1 100 900 100 Tube 2 200 800 200 Tube 3 300 700 300 Tube 4 400 600 400 Tube 5 500 500 500 Tube 6 600 400 600 Tube 7 700 300 700 Tube 8 800 200 800 Tube 9 900 100 900 Positive control tube 0 1000 0

[0025] S3, sample determination and solubility calculation: (1) sample color development: dilute the amoxicillin saturated solution prepared in S1, add an equal amount of 0.1% copper sulfate solution, mix well and stand, to obtain the test solution; (2) turbidimetric analysis: compare the test solution with the visual linear regression equation established in S2 to determine the closest standard tube concentration corresponding to the turbidity; (3) calculate the solubility: calculate the solubility of the amoxicillin sample according to the concentration value determined by comparison and the dilution factor, to finally obtain the solubility of the amoxicillin sample. The solubility of the amoxicillin sample is calculated by the formula S = (C x D) / 1000, where S is the solubility of the amoxicillin sample in the solvent with the unit of mg / mL, C is the original concentration of the amoxicillin sample saturated solution determined by the linear regression equation and calculated by the linear regression equation with the unit of μg / mL, and D is the dilution factor of the saturated solution before turbidimetric analysis. Example 2

[0026] S1, sample solution and reagent preparation: prepare amoxicillin saturated solution: take excess amoxicillin sample powder to be tested, add deionized water as solvent, continuously and gently stir under the condition of ambient temperature ≤ 25°C and relative humidity < 40%, until the solution powder in the sample no longer decreases, and then take the upper clear solution to obtain a yellow-brown clear transparent amoxicillin saturated solution.

[0027] S2, establishment of linear regression equation: (1) preparation of amoxicillin standard stock solution: accurately weighed amoxicillin reference substance was dissolved and diluted to prepare amoxicillin standard stock solution with a concentration of 1000 μg / mL; (2) preparation of series of standard tubes: different volumes of amoxicillin standard stock solution were accurately measured and diluted with solution to obtain standard solutions with a series of concentration gradients; (3) color development and curve establishment: 0.1% copper sulfate solution was added to each series of standard tubes, and after mixing and standing, each tube showed a clear turbidity gradient from transparent clear liquid to yellow-green turbid liquid, which was the visual linear regression equation for quantitative comparison; the series of amoxicillin standard solutions with concentration gradients were set at 9 concentration gradients, and the target concentrations were 100, 200, 300, 400, 500, 600, 700, 800 and 900 μg / mL respectively; the amoxicillin standard stock solution with a concentration of 1000 μg / mL was accurately measured and diluted with 0.2M potassium dihydrogen phosphate solution as diluent to prepare standard working solutions with a series of concentration gradients; the series of amoxicillin standard solutions with concentration gradients included a blank control tube, and the blank control tube contained 1000 μg / mL amoxicillin solution dissolved with deionized water as solvent.

[0028] S3, sample determination and solubility calculation: (1) sample color development: the amoxicillin saturated solution prepared in S1 was diluted, and an equal amount of 0.1% copper sulfate solution was added, mixed and stood to obtain the test solution; (2) turbidimetric analysis: the test solution was compared with the visual linear regression equation established in S2 to determine the closest standard tube concentration corresponding to the turbidity; (3) solubility calculation: the concentration value determined by comparison was combined with the dilution factor to calculate the solubility of the amoxicillin sample. Example 3

[0029] S1, sample solution and reagent preparation: (1) preparation of 1% sodium hexametaphosphate solution: accurately weighed 1% sodium hexametaphosphate powder was dissolved and diluted to prepare 1% sodium hexametaphosphate solution; (2) preparation of amoxicillin saturated solution: an excess amount of amoxicillin sample powder to be tested was added to an accurately measured volume of solution to dissolve, and the solution was kept at 25°C constant temperature for 24 hours to ensure that the dissolution equilibrium was reached to form a saturated solution.

[0030] The 1% sodium hexametaphosphate powder needs to be dissolved with sufficient deionized water as solvent at room temperature, i.e. 25±5°C, until the solution system is clear and transparent, and no undissolved powder is ensured; the amoxicillin sample uses 1% sodium hexametaphosphate solution as solvent, and is continuously and gently stirred by magnetic force under the condition of ambient temperature ≤25°C and relative humidity <40%, until the solution powder in the sample no longer decreases, and the upper clear solution is taken to obtain a clear and transparent amoxicillin saturated solution.

[0031] S2, establishment of linear regression equation: (1) preparation of amoxicillin standard stock solution: accurately weigh amoxicillin reference substance, dissolve and dilute to volume to prepare amoxicillin standard stock solution with a concentration of 1000 μg / mL; (2) preparation of series of standard tubes: accurately take different volumes of amoxicillin standard stock solution, dilute with solution to obtain standard solutions with a series of concentration gradients; (3) color development and establishment of curve: quantitatively add 0.1% copper sulfate solution to each series of standard tubes, mix well and stand, and each tube will immediately show a clear turbidity gradient from transparent clear liquid to yellow-green turbid liquid, which is the visual linear regression equation used for quantitative comparison.

[0032] The preparation method of amoxicillin standard stock solution is the same as that of amoxicillin saturated solution, and the series of amoxicillin standard solutions with concentration gradients are set at 9 concentration gradients, and the target concentrations are 100, 200, 300, 400, 500, 600, 700, 800 and 900 μg / mL respectively; accurately take amoxicillin standard stock solution with a concentration of 1000 μg / mL, use 1% sodium hexametaphosphate solution as diluent, and dilute step by step to prepare standard working solutions with a series of concentration gradients; the series of amoxicillin standard solutions with concentration gradients include blank control tube and positive control tube, the blank control tube is 1000 μg / mL amoxicillin solution dissolved with deionized water, and the blank control tube does not contain 1% sodium hexametaphosphate; the positive control tube is 1% sodium hexametaphosphate solution dissolved with deionized water, and does not contain amoxicillin.

[0033] The visual linear regression equation is established based on the absorbance values of the series of amoxicillin standard solutions with concentration gradients, and the ultraviolet-visible spectrophotometer is used to measure the absorbance values of each standard tube and sample tube at 550 nm wavelength, with the blank control tube correcting the instrument zero point, and the absorbance values of the standard tubes as the ordinate and the corresponding amoxicillin concentrations as the abscissa, to draw the linear regression equation.

[0034] S3, sample determination and solubility calculation: (1) sample color development: after dilution of the amoxicillin saturated solution prepared in S1, an equal amount of 0.1% copper sulfate solution is added, mixed well and stood to obtain the test solution; (2) turbidimetric analysis: compare the test solution with the visual linear regression equation established in S2 to determine the closest standard tube concentration corresponding to the turbidity; (3) calculation of solubility: according to the concentration value determined by comparison, the dilution factor is combined to calculate the solubility of amoxicillin sample. Example 4

[0035] S1, sample solution and reagent preparation: (1) preparation of 1% sodium hydroxide solution: accurately weigh 1% sodium hydroxide powder, dissolve and dilute to volume, and prepare 1% sodium hydroxide solution; (2) preparation of amoxicillin saturated solution: take excess amoxicillin sample powder, add accurate volume of solution to dissolve, and keep constant temperature at 25°C for 24 hours, ensure that the dissolution balance is reached, and form saturated solution.

[0036] 1% sodium hydroxide powder needs to use enough deionized water as solvent, and dissolve at room temperature, i.e. 25±5°C, until the solution system is clear and transparent, and ensure that there is no undissolved powder; amoxicillin sample uses 1% sodium hydroxide solution as solvent, and keeps constant magnetic stirring under the condition of ambient temperature ≤25°C and relative humidity <40%, until the solution powder in the sample no longer decreases, and the upper clear solution is taken to obtain clear and transparent amoxicillin saturated solution.

[0037] S2, establishment of linear regression equation: (1) preparation of amoxicillin standard stock solution: accurately weigh amoxicillin reference substance, dissolve and dilute to volume, and prepare amoxicillin standard stock solution with concentration of 1000 μg / mL; (2) preparation of series of standard tubes: accurately take different volumes of amoxicillin standard stock solution, dilute with solution to obtain series of standard solutions with concentration gradient; (3) color development and curve establishment: to each series of standard tubes, add 1.0 mL of 1M NaOH solution in turn, mix well, keep at room temperature for 10-15 minutes, ensure complete hydrolysis, then add 1.0 mL of 1M HCl solution for neutralization, and then add accurate volume of 0.005M iodine solution, mix well, avoid light for 10 minutes, finally, add 1.0 mL of 1% starch indicator to each tube, dilute to the same volume with water, and mix well; measure absorbance at 580 nm wavelength with water as reference, at this time, absorbance value is negatively correlated with concentration.

[0038] The preparation method of amoxicillin standard stock solution is the same as that of amoxicillin saturated solution, and series of amoxicillin standard solutions with concentration gradient are set with 9 concentration gradients, and the target concentrations are 100, 200, 300, 400, 500, 600, 700, 800 and 900 μg / mL respectively; accurately take amoxicillin standard stock solution with concentration of 1000 μg / mL, dilute with 1% sodium hydroxide solution as diluent to prepare series of standard working solutions with concentration gradient; series of amoxicillin standard solutions with concentration gradient include blank control tube and positive control tube, the blank control tube is 1000 μg / mL amoxicillin solution dissolved with deionized water as solvent, and the blank control tube does not contain 1% sodium hydroxide; the positive control tube is 1% sodium hydroxide solution dissolved with deionized water as solvent, and does not contain amoxicillin.

[0039] The visual linear regression equation is established based on absorbance values of amoxicillin standard solution of a series of concentration gradients, using a UV-visible spectrophotometer, correcting the instrument zero point with a blank control tube, sequentially determining absorbance values of each standard tube and sample tube, taking absorbance values of the standard tube as the ordinate and corresponding amoxicillin concentrations as the abscissa, and drawing a linear regression equation.

[0040] S3, sample determination and solubility calculation: (1) sample color development: diluting the saturated amoxicillin solution prepared in S1, adding an equal amount of 0.1% copper sulfate solution, mixing and standing to obtain a test solution; (2) turbidimetric analysis: comparing the test solution with the visual linear regression equation established in S2 to determine the turbidity corresponding to the closest standard tube concentration; (3) calculating solubility: calculating according to the concentration value determined by comparison and combining with the dilution factor to finally obtain the solubility of the amoxicillin sample.

[0041] Table 2

[0042]

[0043] Example 2 uses deionized water instead of potassium dihydrogen phosphate solution as a solvent, so that the formed precipitate is fast and unstable, causing the color gradient to be destroyed in a short time, and effective comparison cannot be performed; Example 3 uses 1% sodium hexametaphosphate solution instead of potassium dihydrogen phosphate, and the precipitate is produced too fast and unstable, so the operator must complete the comparison in a very short time, otherwise the result is invalid, the fault tolerance is very low, and the operation pressure and misjudgment risk are increased; Example 4 involves multiple necessary and waiting steps such as alkaline hydrolysis, acid neutralization, iodization and starch color development, and the process is complicated and time-consuming, which is not suitable for rapid analysis.

[0044] Table 3

[0045]

[0046] The relative standard deviation is repeated three times to establish the linear regression equation process, the same sample is substituted into three different linear regression equations, the dispersion degree between the measured values is calculated, the smaller the RSD value, the better the repeatability of the method, and the more stable and reliable the results; the deviation from the standard value is the deviation of the absorbance of the amoxicillin solution with the known solubility from the absorbance of the amoxicillin solution with the same solubility measured by the present application.

[0047] Example 2 uses deionized water as a solvent, which cannot provide a stable reaction environment, the formed precipitated particles are uneven in size, and quickly aggregate and settle, and the settlement speed and state of the precipitate can be different each time of measurement, resulting in a sharp fluctuation of the absorbance reading in a short time; Example 3 uses 1% sodium hexametaphosphate solution, which can provide a relatively stable and uniform reaction environment, but sodium hexametaphosphate itself is a strong metal ion chelating agent, which can compete with the chromogenic agent copper sulfate to form a complex, reducing the generation amount of the chromogenic product, resulting in a generally low absorbance value and a large deviation from the standard value; Example 4 has too many operation steps, involving hydrolysis, neutralization, iodization, starch color development and other links, and a small operation error in each link will be accumulated and transferred to the end, resulting in a relatively large difference between the parallel samples, and the iodine solution is volatile and reacts easily under light, so a small change in its concentration will directly affect the color development, and a small loss of control in these links will eventually be converted into an error in concentration calculation.

[0048] Secondly, in the disclosed embodiments, only the structures involved in the disclosed embodiments are involved, other structures can be referred to the usual design, and in the case of no conflict, the same embodiment and different embodiments of the present application can be combined with each other;

[0049] Finally, the above only describes the preferred embodiments of the present application, and is not used to limit the present application, and any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application should be included in the protection scope of the present application.

Claims

1. A visual detection method for amoxicillin solubility based on quantitative color gradations, characterized in that, include: S1. Preparation of sample solution and reagents: (1) Preparation of 0.2M potassium dihydrogen phosphate solution: accurately weigh potassium dihydrogen phosphate powder, dissolve and dilute to volume to prepare 0.2M potassium dihydrogen phosphate solution; (2) Preparation of amoxicillin saturated solution: take excess amoxicillin sample powder to be tested, add an accurate volume of 0.2M potassium dihydrogen phosphate solution to dissolve, shake at 25°C for 24 hours to ensure that dissolution equilibrium is reached and a saturated solution is formed; S2. Establishment of linear regression equation: (1) Preparation of amoxicillin standard stock solution: accurately weigh amoxicillin reference standard, dissolve and dilute to volume, and prepare amoxicillin standard stock solution with a concentration of 1000 μg / mL; (2) Preparation of a series of standard tubes: accurately measure different volumes of amoxicillin standard stock solution, dilute with 0.2M potassium dihydrogen phosphate solution, and obtain a series of standard solutions with concentration gradients; (3) Color development and establishment of curve: quantitatively add 0.1% copper sulfate solution to each series of standard tubes, mix well and let stand. Each tube will then show a clear turbidity gradient from transparent clear liquid to yellow-green. This gradient is the visual linear regression equation used for quantitative comparison. S3. Sample determination and solubility calculation: (1) Sample color development: Dilute the amoxicillin saturated solution prepared in S1, add an equal amount of 0.1% copper sulfate solution, mix and let stand to obtain the test solution; (2) Turbidity analysis: Compare the test solution with the visual linear regression equation established in S2 to determine the concentration of the closest standard tube corresponding to its turbidity; (3) Calculate solubility: Calculate the solubility of the amoxicillin sample based on the concentration value determined by comparison and the dilution factor.

2. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 1, characterized in that: The potassium dihydrogen phosphate powder must be dissolved in sufficient deionized water at room temperature (25±5℃) until the solution is clear and transparent, ensuring that there is no undissolved powder.

3. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 2, characterized in that: The amoxicillin sample and amoxicillin reference standard were subjected to continuous and gentle magnetic stirring under ambient temperature ≤25℃ and relative humidity <40% until the powder no longer decreased. The clear upper layer of solution was then aspirated to obtain a clear and transparent amoxicillin saturated solution and amoxicillin standard stock solution, respectively.

4. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 2, characterized in that: The concentration of the amoxicillin standard stock solution was calculated as follows: Where C is the concentration of amoxicillin standard stock solution in μg / mL, m is the mass of the reference standard weighed in μg, P is the purity of the reference standard, and V is the final volume in mL.

5. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 1, characterized in that: The series of amoxicillin standard stock solutions, excluding blank control tubes and positive control tubes, are configured with nine concentration gradients, with target concentrations of 100, 200, 300, 400, 500, 600, 700, 800, 900, and 1000 μg / mL, respectively. Amoxicillin standard stock solution with a concentration of 1000 μg / mL is accurately measured and diluted stepwise using 0.2 M potassium dihydrogen phosphate solution to prepare a series of standard working solutions with different concentration gradients.

6. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 1, characterized in that: The series of amoxicillin standard stock solutions includes blank control tubes and positive control tubes. The blank control tubes contain a 1000 μg / mL amoxicillin solution dissolved in deionized water, and the solution in the blank control tubes does not contain potassium dihydrogen phosphate. The positive control tubes contain a 0.2 M potassium dihydrogen phosphate solution dissolved in deionized water, and do not contain amoxicillin.

7. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 6, characterized in that: The blank control tube, after adding 0.1% copper sulfate solution, produced a clear, yellowish-brown solution; the positive control tube, after adding 0.1% copper sulfate solution, produced a turbid, yellowish-brown solution with obvious precipitation.

8. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 1, characterized in that: The visual linear regression equation was established based on a series of absorbance values ​​of amoxicillin standard stock solutions. Using a UV-Vis spectrophotometer, the instrument zero point was calibrated at a wavelength of 550 nm with a blank control tube. The absorbance values ​​of each standard tube and sample tube were measured sequentially. The linear regression equation was plotted with the absorbance value of the standard tube as the ordinate and the corresponding amoxicillin concentration as the abscissa.

9. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 1, characterized in that: The linear regression equation is y=a×C+b, where y is the absorbance value, C is the amoxicillin concentration in μg / mL, a is the slope, and b is the intercept, which is obtained by fitting the absorbance values ​​of a series of amoxicillin standard stock solutions.

10. The visual detection method for amoxicillin solubility based on quantitative color gradation according to claim 1, characterized in that: The solubility of amoxicillin sample is calculated using the formula S=(C×D) / 1000, where S is the solubility of amoxicillin sample in solvent, in mg / mL; C is the original concentration of the saturated amoxicillin solution determined by linear regression equation and calculated by linear regression equation, in μg / mL; and D is the dilution factor of the saturated amoxicillin solution before turbidimetric analysis.

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