Method for detecting content of andrographolide total sulfonate
阅读说明：本技术 一种穿心莲内酯总磺化物含量的检测方法 (Method for detecting content of andrographolide total sulfonate ) 是由 邓双炳 方礼 刘佳佳 古丽萍 周舟 刘地发 黄冰峰 魏勇翔 谢华萍 林芳 于 2019-11-15 设计创作，主要内容包括：本发明公开了一种适用于穿心莲内酯总磺化物含量的检测方法,先用过量碱与穿心莲内酯总磺化物反应,然后再用盐酸反滴定的方法来测定穿心莲内酯总磺化物的含量,该方法的精密度高、准确度好。(The invention discloses a method for detecting the content of andrographolide total sulfonate, which comprises the steps of reacting excessive alkali with andrographolide total sulfonate, and then measuring the content of andrographolide total sulfonate by a hydrochloric acid back titration method.)
1. A method for detecting andrographolide total sulfonate is characterized by comprising the following steps:
(1) preparation of a test solution: precisely measuring a liquid preparation containing andrographolide total sulfonate, placing the liquid preparation into a wide-mouth conical flask with a plug, and precisely adding 0.3-30 ml of sodium hydroxide titration solution; adding 0.2-200 ml of water, heating in a water bath for 20-40 minutes, taking out, and cooling to room temperature to obtain a test solution;
(2) determination of the content of andrographolide total sulfonate: taking a test solution, titrating with a hydrochloric acid titration solution according to a potentiometric titration method, and correcting a titration result by using a blank test; and (4) calculating.
2. The detection method according to claim 1, wherein the liquid preparation in step (1) is an injection, an aerosol, a syrup, an oral solution, an oral suspension or a lotion, preferably an injection, an aerosol, an oral suspension.
3. The detection method according to claim 1, wherein the concentration of the sodium hydroxide titration solution in the step (1) is 0.05-0.5 mol/L, preferably 0.1 mol/L.
4. The detection method according to claim 1, wherein the temperature of the water bath in the step (1) is 40 to 80 ℃, preferably 55 to 65 ℃.
5. The detection method according to claim 1, wherein the concentration of the hydrochloric acid titration solution in the step (2) is 0.01-0.1 mol/L, preferably 0.02 mol/L.
6. The detection method according to claim 1, wherein the reaction amount of andrographolide sulfonate and sodium hydroxide titration solution is calculated in step (2), and 1ml of sodium hydroxide (0.1mol/L) is consumed in each reaction, which is equivalent to 45.25mg of andrographolide sulfonate.
7. The detection method according to claim 1, wherein the calculation formula in step (2) is
The total content (mg/ml) of andrographolide sulfonate is (C)1*V1-C2*V2)*452.5/VFor supplying to
In the formula C1: the concentration (mol/L) of the sodium hydroxide titration solution;
V1: the addition amount (ml) of sodium hydroxide titration solution;
C2: hydrochloric acidConcentration of the titration solution (mol/L);
V2: consumption (ml) of hydrochloric acid titration solution at the potential jump point;
Vfor supplying to: sample size (ml);
452.50: with C20H29O5·SO3Molecular weight 452.50 calculated as Na.
Technical Field
The invention relates to a method for detecting the content of andrographolide total sulfonate, belonging to the technical field of drug analysis.
Background
In the national drug standard WS-10863(ZD-0863) -2002-2011Z of Xiyanping injection, the determination method of the total sulfonate is specified as follows:
preparation of control solutions: accurately weighing andrographolide control sample about 0.1g, placing into 25ml measuring flask, adding water 5ml, adding appropriate amount of anhydrous ethanol, sealing, dissolving by ultrasonic, cooling, adding anhydrous ethanol to scale, and shaking to obtain the final product (containing andrographolide 4mg per 1 ml).
Preparation of a standard curve: precisely measuring 1ml, 2ml, 3ml, 4ml and 5ml of reference solution, respectively placing in a 25ml measuring flask, precisely measuring 3ml of 1% m-dinitrobenzene ethanol solution, placing in a 25ml measuring flask as reference solution, and adding 80% ethanol to 5.0ml respectively; adding 3ml of 1% m-dinitrobenzene ethanol solution and 10ml of ethanol solution (taking 240.0g of absolute ethanol, placing the mixture in a 500ml measuring flask, adding water to the scale and shaking uniformly) into each flask precisely, shaking uniformly, placing the mixture in a water bath at 60 ℃, then adding 5ml of potassium hydroxide solution (taking 1.37g of potassium hydroxide and 5.9g of sodium citrate, placing the mixture in the same 100ml measuring flask, adding water to dissolve and dilute the mixture to the scale and shaking uniformly) precisely, shaking uniformly, heating for 40 minutes, cooling, adding the ethanol solution to the scale and shaking uniformly, measuring the absorbance at 407nm wavelength by an ultraviolet-visible spectrophotometry, and drawing a standard curve by taking the absorbance as the ordinate and the concentration as the abscissa.
The determination method comprises the following steps: precisely measuring 5ml of the product, placing the product in a 25ml measuring flask, adding absolute ethyl alcohol to the scale, shaking up, precisely measuring 3ml, placing the product in the 25ml measuring flask, measuring the absorbance by a method according to a standard curve preparation item from the point that 80% ethyl alcohol is added to 5.0ml, reading the content of andrographolide in a test solution from the standard curve, calculating, and multiplying the result by 1.291 to obtain the andrographolide content.
Each 1ml of the product contains andrographolide sulfonate (sodium andrographolide sulfonate C)20H29O5·SO3Na) is 85.0 to 115.0 percent of the marked amount.
The following problems are encountered in treating a sample solution by the assay method described above:
the reference substance of 0.1g is needed for each measurement, a standard curve is also needed to be prepared, and the detection cost is higher; the operation process is slightly careless and has large difference, and the repeatability is not good.
Disclosure of Invention
The invention aims to provide a measuring method with higher precision and accuracy, which is used for detecting the content of the andrographolide total sulfonate liquid preparation. The research structure shows that andrographolide sulfonate contains five-membered unsaturated lactone ring and can react with sodium hydroxide, and according to the principle, an experiment is designed and tested by adopting a potentiometric titration method, wherein excessive alkali is firstly used for reacting with andrographolide sulfonate, and then the content of andrographolide sulfonate is measured by a hydrochloric acid back titration method.
The purpose of the invention is realized by the following technical scheme:
(1) preparation of a test solution: precisely measuring a liquid preparation containing andrographolide total sulfonate, placing the liquid preparation in a wide-mouth conical flask with a plug, and precisely adding 0.3-30 ml of sodium hydroxide titration solution; adding 0.2-200 ml of water, heating in a water bath for 20-40 minutes, taking out, and cooling to room temperature to obtain a test solution;
(2) determination of the content of andrographolide total sulfonate: taking a test solution, titrating with a hydrochloric acid titration solution according to a potentiometric titration method, and correcting a titration result by using a blank test; and (4) calculating.
Preferably, the liquid preparation in step (1) is injection, aerosol, syrup, oral solution, oral suspension or lotion, preferably injection, aerosol and oral suspension.
Preferably, the concentration of the sodium hydroxide titration solution in the step (1) is 0.05-0.5 mol/L, and more preferably 0.1 mol/L.
Preferably, the temperature of the water bath in the step (1) is 40-80 ℃, and more preferably 55-65 ℃.
Preferably, the concentration of the hydrochloric acid titration solution in the step (2) is 0.01-0.1 mol/L, and more preferably 0.02 mol/L.
Preferably, the reaction amount of the andrographolide total sulfonate and the sodium hydroxide titration solution calculated in the step (2) is equal to 45.25mg of andrographolide total sulfonate consumed 1ml of sodium hydroxide (0.1mol/L) per reaction.
Preferably, the calculation formula is
The total content (mg/ml) of andrographolide sulfonate is (C)1*V1-C2*V2)*452.5/VFor supplying to
In the formula C1: the concentration (mol/L) of the sodium hydroxide titration solution;
V1: the addition amount (ml) of sodium hydroxide titration solution;
C2: the concentration (mol/L) of the hydrochloric acid titration solution;
V2: consumption (ml) of hydrochloric acid titration solution at the potential jump point;
Vfor supplying to: sample size (ml);
452.50: with C20H29O5·SO3Molecular weight 452.50 calculated as Na.
The invention has the beneficial effects that: through the improvement of the prior art, the technical scheme of the invention is simple, convenient, economic and practical, greatly improves the accuracy and precision of detecting the content of the sulfonate in the liquid preparation containing the andrographolide active ingredient, can more comprehensively and accurately evaluate the quality of the andrographolide total sulfonate liquid preparation, and can better ensure the quality of the product and the safety and effectiveness of the clinical use of the medicine.
Detailed Description
The technical solutions provided by the present invention are described in detail below with reference to examples, which are only for illustration and do not limit the scope of the present invention.
Example 1: detection method of andrographolide total sulfonate
1. Preparation of a test solution: accurately measuring 10ml of andrographolide total sulfonate injection (self-made), placing the andrographolide total sulfonate injection into a 50ml wide-mouth conical flask with a plug, accurately adding 3ml of sodium hydroxide titration solution (0.1mol/L), adding 20ml of water, placing the mixture in a water bath at 60 ℃ for heating for 30 minutes, taking out the mixture, and cooling the mixture to room temperature.
2. Measuring the content of andrographolide total sulfonate in the test sample: titrating by hydrochloric acid titration solution (0.02mol/L) according to the potentiometric titration method, correcting the titration result by a blank test,the reaction amount of andrographolide total sulfonate and sodium hydroxide titration solution can be calculated, 1ml of sodium hydroxide (0.1mol/L) consumed in each reaction is equivalent to 45.25mg of andrographolide total sulfonate (expressed as C)20H29O5·SO3Molecular weight 452.50 calculated as Na).
The calculation formula is as follows:
the total content (mg/ml) of andrographolide sulfonate is (C)1*V1-C2*V2)*452.5/VFor supplying to
In the formula C1: the concentration (mol/L) of the sodium hydroxide titration solution;
V1: the addition amount (ml) of sodium hydroxide titration solution;
C2: the concentration (mol/L) of the hydrochloric acid titration solution;
V2: consumption (ml) of hydrochloric acid titration solution at the potential jump point;
Vfor supplying to: sample size (ml);
452.50: with C20H29O5·SO3Molecular weight 452.50 calculated as Na.
The calculated total sulfonate content of andrographolide was 24.5 mg/ml.
3. Results of recovery test
The improved method was tested for recovery and 3 samples were prepared at 50%, 100%, and 150% concentration levels for content determination and comparison of accuracy. The results are given in the following table:
the result shows that the recovery rate range of the improved method is 99.80-103.51%, and the RSD is 1.58%, which shows that the method has better accuracy and precision.
Example 2: detection of andrographolide total sulfonate
1. Preparation of a test solution: accurately measuring 10ml of andrographolide total sulfonate injection (self-made), placing the andrographolide total sulfonate injection into a 50ml wide-mouth conical flask with a plug, accurately adding 3ml of sodium hydroxide titration solution (0.1mol/L), adding 20ml of water, heating the mixture in a water bath at 55 ℃ for 40 minutes, taking out the mixture, and cooling the mixture to room temperature.
2. Measuring the content of andrographolide total sulfonate in the test sample: titrating with hydrochloric acid (0.02mol/L) according to potentiometric titration method, correcting titration result with blank test, and calculating reaction amount of andrographolide sulfonate and sodium hydroxide titration solution, wherein 1ml of sodium hydroxide (0.1mol/L) is consumed per reaction and is equivalent to 45.25mg of andrographolide sulfonate (as C)20H29O5·SO3Molecular weight 452.50 calculated as Na).
3. The calculated total sulfonate content of andrographolide was 24.4 mg/ml.
Example 3: detection of andrographolide total sulfonate
1. Preparation of a test solution: accurately measuring 10ml of andrographolide total sulfonate injection (self-made), placing the andrographolide total sulfonate injection into a 50ml wide-mouth conical flask with a plug, accurately adding 3ml of sodium hydroxide titration solution (0.1mol/L), adding 20ml of water, placing the mixture in a water bath at 65 ℃ for heating for 20 minutes, taking out the mixture, and cooling the mixture to room temperature.
2. Measuring the content of andrographolide total sulfonate in the test sample: titrating with hydrochloric acid (0.02mol/L) according to potentiometric titration method, correcting titration result with blank test, and calculating reaction amount of andrographolide sulfonate and sodium hydroxide titration solution, wherein 1ml of sodium hydroxide (0.1mol/L) is consumed per reaction and is equivalent to 45.25mg of andrographolide sulfonate (as C)20H29O5·SO3Molecular weight 452.50 calculated as Na).
3. The calculated total sulfonate content of andrographolide was 24.7 mg/ml.
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