Method for detecting content of edetate disodium in epinephrine hydrochloride injection

Abstract

The invention discloses a method for detecting the content of disodium edetate in an epinephrine hydrochloride injection, which adopts high performance liquid chromatography for detection, and a sample is pretreated according to the following steps before sample injection: adding L-ascorbic acid into the epinephrine hydrochloride injection, then adding a metal ion solution, then diluting with an acid solution, and filtering. The method for detecting the content of EDTA in the epinephrine hydrochloride injection is accurate, efficient, convenient and high in popularization rate, an injection sample is pretreated, L-ascorbic acid and a ferric trichloride solution are added, epinephrine and Fe < 3 + > in the injection can be prevented from generating epinephrine quinone which interferes detection, and the detection accuracy is improved. Ascorbic acid can also form a ternary stable system with iron ions and EDTA, and it is ensured that after liquid phase separation, edetate disodium-metal chelate chromatographic peaks which are symmetrical in peak shape and free of trailing can be detected; and other auxiliary materials such as sodium pyrosulfite and other antioxidants in the injection can be prevented from competitively consuming ferric trichloride so as to ensure that the EDTA complex reaction is complete.

Description

Technical Field

[0001] The invention belongs to the technical field of pharmaceutical preparation analysis, and particularly relates to a method for detecting the content of disodium edetate in epinephrine hydrochloride injection. Background Art

[0002] Epinephrine hydrochloride injection, an important drug in the emergency medical field, is clinically used to treat anaphylactic shock, cardiac arrest, and acute bronchial asthma attacks by stimulating α- or β-receptors. Epinephrine has a phenolic hydroxyl group structure, which is susceptible to oxidation and deterioration when exposed to metal ions during production and storage. Therefore, edetate disodium is added as a stabilizer. However, excessive intake of edetate disodium can lead to hypocalcemia, arrhythmias, or electrolyte imbalances. The safety limit for edetate disodium in pharmaceutical preparations is typically 0.005% to 0.1%. Therefore, accurate measurement of edetate disodium content in injections is essential for ensuring the safety and effectiveness of the preparation.

[0003] The quantitative method for edetate disodium listed in the Chinese Pharmacopoeia is volumetric analysis, which has a high limit of quantification and is not suitable for analyzing the components of injectable excipients. Currently, high-performance liquid chromatography (HPLC) is the most commonly used method for analyzing edetate disodium in injectable solutions due to its wide application and high accuracy. However, edetate disodium interacts with the stainless steel surface of the chromatographic column, resulting in severe tailing of its chromatographic peak, making accurate quantification impossible. Most literature uses the peak area external standard method to determine edetate disodium content by adding metal ions to the formulation to generate edetate disodium-metal chelates that can be quantitatively tested.

[0004] However, the components of epinephrine hydrochloride injection are complex. In addition to the active ingredient epinephrine, it also includes excipients such as sodium metabisulfite, tartaric acid, disodium edetate, hydrochloric acid, sodium hydroxide, and sodium chloride. Among them, epinephrine, sodium metabisulfite, tartaric acid, and disodium edetate can react with metal ion solutions, and hydrochloric acid and sodium hydroxide can affect the pH of the solution, thereby affecting the competitive reaction between the metal ion reagent and various excipients in the complex solution and the peak shape of the metal chelate chromatographic peak. In existing high-performance liquid phase analysis methods, the chelate chromatographic peak has problems such as broadening of the peak shape, inability to accurately quantify or the separation does not meet the analysis requirements, and even the inability to detect the disodium edetate chromatographic peak. Therefore, it is necessary to develop an analytical method for detecting the content of disodium edetate in epinephrine hydrochloride injection. Summary of the Invention

[0005] The present invention comprehensively considers the types of raw materials and auxiliary materials of epinephrine hydrochloride injection and their pH and physicochemical properties, and provides a method for detecting the content of disodium edetate in epinephrine hydrochloride injection, so as to solve the technical problems of broadened peaks of disodium edetate and chelate chromatographic peaks, inability to accurately quantify, and unsatisfactory separation when detecting epinephrine hydrochloride injection.

[0006] According to one aspect of the present invention, a method for detecting the content of edetate disodium in epinephrine hydrochloride injection is provided. The method employs high performance liquid chromatography for detection. Prior to injection, the sample is pretreated according to the following steps: L-ascorbic acid is added to the epinephrine hydrochloride injection, followed by a metal ion solution. The solution is then diluted with an acid solution until the concentration of edetate disodium in the solution is 0.5 to 20 μg / mL, and filtered.

[0007] In some embodiments, the metal ion solution is selected from at least one of a ferric chloride solution, a copper sulfate solution, and a ferrous sulfate solution. Preferably, the metal ion solution is a ferric chloride solution. The ratio of the concentration of disodium edetate to the volume of the metal ion solution is 5:0.5 to 5:2; preferably, the ratio of the concentration of disodium edetate to the volume of the metal ion solution is 5:1. The metal ion solution is prepared by the following steps: weighing a metal reagent and dissolving it in part of water, then adding 0.1% of the prepared volume of concentrated hydrochloric acid and constant to volume; wherein, in the metal ion solution, the metal ion concentration is 0.015 to 0.03 mol / L. Preferably, the metal ion concentration is 0.02 mol / L.

[0008] It should be noted that the point value of the preparation volume is the product of the volume of the solution after the volume is fixed and the value. For example, when preparing 1 L of 0.2 mol / L ferric chloride solution, adding 0.1% of the preparation volume of concentrated hydrochloric acid means adding 0.1% × 1 L = 1 ml of concentrated hydrochloric acid.

[0009] In some embodiments, the acid solution can be selected from at least one of phosphoric acid, hydrochloric acid, and sulfuric acid. More preferably, the acid solution can be selected from a 0.1% (v / v) phosphoric acid aqueous solution. The pH of the test solution and the reference solution is adjusted to 1-4 by dilution with the acid solution.

[0010] In some embodiments, the filtration step is performed using an aqueous membrane filter. Preferably, filtration is performed using a 0.45 μm aqueous membrane filter. This is to remove insoluble particles (e.g., ferric hydroxide colloids, excipient precipitates) that may be present in the sample solution, thereby preventing clogging of the chromatographic column.

[0011] In some embodiments, the mobile phase A used in high-performance liquid chromatography is an ion pair reagent, and the mobile phase B is methanol or acetonitrile. The ratio of mobile phase A to mobile phase B is 50:50 to 95:5, and isocratic elution is used. The ion pair reagent can be selected from 0.05% to 0.10% (V / V) tetrabutylammonium hydroxide solution, sodium lauryl sulfate solution, or ammonium dihydrogen phosphate solution. Preferably, the ratio of mobile phase A to mobile phase B is 70:30, the ion pair reagent is 0.09% (V / V) tetrabutylammonium hydroxide solution, the pH is 1 to 3, more preferably the pH is 2.3, and it is adjusted with phosphoric acid.

[0012] In some embodiments, the chromatographic column used in the high performance liquid chromatography method is an octadecyl bonded silica gel chromatographic column; the octadecyl bonded silica gel chromatographic column can be selected from YMC-Pack ODS-AQ 250×4.6mm, ID S-5μm, 12nm, i.e., the pore size is 12nm and the filler particle size is 5μm.

[0013] In some embodiments, the detection wavelength of the high performance liquid chromatography is 200-500 nm. Preferably, the detection wavelength is 200-300 nm. More preferably, the detection wavelength is 254 nm.

[0014] In some embodiments, the flow rate of the HPLC method is 0.9 to 1.5 mL / min. Preferably, the flow rate is 1 mL / min.

[0015] In some embodiments, the column temperature of the high performance liquid chromatography is 25-35° C. Preferably, the column temperature is 30° C.

[0016] In some embodiments, the injection volume for high performance liquid chromatography is 15 to 25 μL, preferably 20 μL.

[0017] The beneficial effects of the present invention include: (1) The present invention selects high performance liquid chromatography and obtains chromatographic peaks with asymmetry and separation that meet the requirements by screening the chromatographic column, mobile phase composition and pH value, thereby providing an accurate, efficient, convenient and widely used method for detecting the content of disodium edetate in epinephrine hydrochloride injection.

[0018] (2) Considering that the pH of epinephrine hydrochloride injection is about 3.5, the present invention uses ferric chloride solution as the metal ion solution and 0.1% phosphoric acid solution as the diluent to provide an acidic reaction environment for generating a stable EDTA-iron ion chelate and inhibit the reaction of other excipients with iron ions.

[0019] (3) The present invention pre-treats the injection sample by adding L-ascorbic acid and then adding ferric chloride solution to the injection solution, thereby blocking the interaction between adrenaline and Fe 3+ Ascorbic acid can generate adrenaline quinone that interferes with the detection. Ascorbic acid can also form a ternary stable system with iron ions and EDTA, ensuring that a symmetrical and non-tailing edetate disodium-metal chelate chromatographic peak can be detected after liquid phase separation; and it can avoid other excipients in the injection solution (such as antioxidants such as sodium metabisulfite) from competitively consuming ferric chloride to ensure the complete EDTA complexation reaction. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1The HPLC chromatograms of the test solution and the reference solution of Example 1 of the present invention are shown; the upper graph represents the test solution, and the lower graph represents the reference solution; Figure 2 The HPLC chromatograms of the test solution and the reference solution of Comparative Example 1 of the present invention are shown in FIG. 1 ; the upper graph represents the reference solution, and the lower graph represents the test solution; Figure 3 This is a high performance liquid chromatogram related to Comparative Example 2 of the present invention; from top to bottom, it is the mobile phase (blank), the reference solution, and the test solution; Figure 4 The chromatogram of the reference solution obtained under chromatographic condition 1 of the present invention; Figure 5 The relevant chromatograms obtained under the second chromatographic condition of the present invention; the upper figure represents the test solution, and the lower figure represents the reference solution; Figure 6 The figure is the relevant chromatogram obtained under the third chromatographic condition of the present invention; from top to bottom, it is the reference solution of the first optimization of the mobile phase, the reference solution of the second optimization of the mobile phase, and the test solution of the first optimization. DETAILED DESCRIPTION

[0021] The present invention is further described below in detail with reference to the accompanying drawings, but the embodiments of the present invention are not limited thereto. The raw materials and reagents used in the following examples are all commercially available. Among them: The epinephrine hydrochloride injection used in the present invention was manufactured by Guangzhou Baiyunshan Mingxing Pharmaceutical Co., Ltd.; the edetate disodium reference substance was purchased from the China Food and Drug Administration with a purity of 90.4%.

[0022] Considering that the pH of epinephrine hydrochloride injection is about 3.5, the present invention uses ferric chloride solution as the metal ion solution and 0.1% phosphoric acid aqueous solution as the diluent to provide an acidic reaction environment for generating a stable EDTA iron ion chelate and inhibit the reaction of other excipients with iron ions. In addition, the present invention can block the reaction of epinephrine in the injection with Fe by first adding L-ascorbic acid and then adding ferric chloride solution as the complexing metal. 3+ Ascorbic acid generates adrenaline quinone, which interferes with the assay. It also forms a ternary stable system with iron ions and EDTA, ensuring a symmetrical, non-tailing chromatographic peak of disodium edetate-metal chelate after liquid phase separation. This also prevents competitive consumption of ferric chloride by other excipients in the injection, ensuring complete EDTA complexation. Finally, by screening the chromatographic column, mobile phase composition, and pH, a chromatographic peak of the EDTA-iron chelate with satisfactory resolution and an asymmetry of approximately 1.0 was obtained. This provides an accurate, efficient, convenient, and widely available method for the determination of EDTA content in epinephrine hydrochloride injection. Details are provided below.

[0023] 1. Solution preparation method Test solution: Take 0.5 ml of epinephrine hydrochloride injection in a 20 ml volumetric flask, first add about 4 mg of L-ascorbic acid, then add 1 ml of 0.02 mol / L ferric chloride solution, dilute to the scale with 0.1% (v / v) phosphoric acid, and filter through a 0.45 μm water filter membrane to obtain the solution.

[0024] Reference solution: Weigh approximately 20 mg of edetate disodium reference substance into a 100 ml volumetric flask, dilute with water and dissolve to the mark to obtain edetate disodium stock solution. Take 0.5 ml of edetate disodium stock solution into a 20 ml volumetric flask, first add 4 mg of L-ascorbic acid, then add 1 ml of 0.02 mol / L ferric chloride solution, dilute to the mark with 0.1% phosphoric acid, and filter with a 0.45 μm water filter membrane to obtain the solution.

[0025] 2. Chromatographic conditions Chromatographic column: octadecyl bonded silica gel column; mobile phase: mobile phase A is 0.09% tetrabutylammonium hydroxide solution (adjusted to pH 1-3 with phosphoric acid), mobile phase B is methanol or acetonitrile; mobile phase A: mobile phase B = 50:50-95:5; flow rate: 0.9-1.5 mL / min; detection wavelength: 200-500 nm; column temperature: 25-35°C; injection volume: 15-25 μL; elution mode: isocratic elution.

[0026] 3. Determination method Take appropriate amount of test solution and reference solution, inject into chromatograph, record chromatogram, and calculate the content of disodium edetate using external standard method. The calculation formula is: , Where C 供 : Concentration of edetate disodium in epinephrine hydrochloride injection, mg / mL; A 供 : chromatographic peak area of ​​disodium edetate in the chromatogram of the test solution; m: weight of edetate disodium reference substance, mg; A: Content of edetate disodium reference substance, %; V 供 : dilution volume of epinephrine hydrochloride injection, mL; A 对 : Chromatogram of edetate disodium reference solution, chromatographic peak area of ​​edetate disodium; V 对 : Dilution volume of edetate disodium reference substance, mL.

[0027] Example 1 This embodiment provides a method for detecting the content of disodium edetate in epinephrine hydrochloride injection, comprising the following steps: (1) Prepare the test solution and reference solution according to the above solution preparation method; (2) Using high performance liquid chromatography, accurately measure the test solution and the reference solution, inject them into the high performance liquid chromatograph, and test the test solution and the reference solution according to the following chromatographic conditions, and record the chromatograms; Chromatographic conditions included: column: YMC-Pack ODS-AQ 250×4.6 mm l.DS-5 μm, 12 nm; mobile phase: 0.09% tetrabutylammonium hydroxide solution (adjusted to pH 2.3 with phosphoric acid): acetonitrile = 70:30 (V:V); flow rate: 1 mL / min; detection wavelength: 254 nm; column temperature: 30°C; injection volume: 20 μL; elution mode: isocratic elution; (3) Calculate the content of disodium edetate in the test solution by the external standard method.

[0028] Comparative Example 1 This comparative example provides a method for detecting the content of disodium edetate in epinephrine hydrochloride injection, comprising the following steps: (1) Take 0.5 mL of epinephrine hydrochloride injection in a 20 mL volumetric flask, add 1 mL of 0.02 mol / L ferric chloride solution, dilute to the mark with 0.1% phosphoric acid, and filter with a 0.45 μm water filter membrane to obtain the test solution; (2) Weigh approximately 20 mg of edetate disodium reference substance into a 100 mL volumetric flask and dilute with water to the mark. Place 0.5 mL of the above solution into a 20 mL volumetric flask, add 1 mL of 0.02 mol / L ferric chloride solution, and dilute to the mark with 0.1% phosphoric acid. Filter through a 0.45 μm aqueous filter to obtain the reference substance solution. (3) Using high performance liquid chromatography, accurately measure the reference solution and the test solution, inject them into the high performance liquid chromatograph, test the test solution according to the following chromatographic conditions, and record the chromatogram; The chromatographic conditions included: chromatographic column: YMC-Pack ODS-AQ 250×4.6mml.DS-5μm, 12nm; mobile phase: 0.09% tetrabutylammonium hydroxide solution (adjusted to pH 2.3 with phosphoric acid): acetonitrile = 70:30 (V:V); flow rate: 1 mL / min; detection wavelength: 254 nm; column temperature: 30°C; injection volume: 20 μL; elution mode: isocratic elution.

[0029] Comparative Example 2 On the basis of Example 1, the chromatographic conditions were changed and the determination was carried out according to the method of the literature "Determination of the content of disodium edetate in nalmefene hydrochloride injection by high performance liquid chromatography" (Yao Wenjing et al. International Journal of Pharmaceutical Research, 2017 (1)). The specific chromatographic conditions were as follows: chromatographic column: YMC-Pack ODS-AQ 250×4.6mm l.D. S-5μm, 12nm; mobile phase: 0.3% tetrabutylammonium hydroxide solution (adjusted to pH 4.0 with hydrochloric acid)-water-acetonitrile (20:45:35, V:V:V); flow rate: 1mL / min; detection wavelength: 254nm; column temperature: 30℃; injection volume: 20μL; elution mode: isocratic elution.

[0030] The chromatogram of Example 1 is as follows Figure 1 As shown, the chromatogram of Comparative Example 1-2 is as shown Figure 2-3 shown. Figure 1 and Figure 2 By comparison, it can be seen that when ascorbic acid was not added, the test solution showed an obvious front peak; after adding ascorbic acid, the obtained test solution was a colorless and transparent solution, which can effectively reduce the background noise, and no front peak appeared in the chromatographic peak of disodium edetate-metal chelate, thereby improving the sensitivity of the chromatographic analysis method. Figure 3 This shows that the analytical method of Comparative Example 2 can detect the chromatographic peak of disodium edetate-metal chelate in the reference solution, but cannot accurately quantify the disodium edetate chelate in the test solution of epinephrine hydrochloride injection.

[0031] The following describes the development process and methodology verification of the detection method for the content of disodium edetate in the epinephrine hydrochloride injection of the present invention.

[0032] 1. Chromatographic conditions 1 Test solution: Take 0.5 mL of epinephrine hydrochloride injection in a 20 mL volumetric flask, add 1 mL of 0.02 mol / L ferric chloride solution, and make up to volume with water to obtain a solution containing 5 μg of edetate disodium per 1 mL. Filter and inject.

[0033] Reference solution: Weigh approximately 20 mg of edetate disodium reference substance into a 100 mL volumetric flask, add 1 mL of 0.02 mol / L ferric chloride solution, and dilute to volume with water to obtain a solution containing 5 μg of edetate disodium per 1 mL. Filter and inject.

[0034] Chromatographic conditions: chromatographic column: YMC-Pack ODS-AQ 250×4.6mm l.D. S-5μm, 12nm; mobile phase: 0.1mol / L phosphoric acid aqueous solution-methanol (95:5); column temperature: 30℃; injection volume: 10μL; detection wavelength: 254nm.

[0035] Record the chromatogram and calculate the content of disodium edetate by the peak area according to the external standard method. The result is as follows: Figure 4 As shown. Figure 4 It can be seen that when the reference solution is prepared using aqueous solution, the chromatographic peak of disodium edetate-metal chelate cannot be detected.

[0036] 2. Chromatographic conditions 2 Test solution: Take 0.5 mL of epinephrine hydrochloride injection in a 20 mL volumetric flask, add 1 mL of 0.02 mmol / L ferric chloride solution, and make up to volume with 0.1% (V / V) phosphoric acid aqueous solution to obtain a solution containing 5 μg of edetate disodium per 1 mL. Filter and inject.

[0037] Reference solution: Weigh approximately 20 mg of edetate disodium reference substance into a 100 mL volumetric flask, add 1 mL of 0.02 mol / L ferric chloride solution, and dilute to volume with 0.1% (v / v) aqueous phosphoric acid solution to obtain a solution containing 5 μg of edetate disodium per 1 mL. Filter and inject.

[0038] The above test solution and reference solution were injected and tested according to the above chromatographic conditions, and the chromatogram was recorded. The results are as follows. Figure 5 As shown, Figure 5 From top to bottom, they are test solution and reference solution. Figure 4 and Figure 5 Comparison shows that compared to preparation with aqueous solution, the edetate disodium-metal chelate chromatographic peak can be detected when the test solution and the reference solution are prepared with 0.1% phosphoric acid aqueous solution. However, when phosphoric acid aqueous solution and methanol are used as the mobile phase, the retention time of the edetate disodium-metal chelate chromatographic peak in the chromatographic column is short. By adding an ion pair reagent, the retention of the edetate disodium-metal chelate chromatographic peak is enhanced, achieving the purpose of effectively separating this chromatographic peak.

[0039] 3. Chromatographic Condition 3: Optimization of Mobile Phase (1) First optimization of mobile phase Chromatographic conditions: YMC-Pack ODS-AQ 250×4.6mm l.D.S-5μm, 12nm; mobile phase: ammonium dihydrogen phosphate solution-methanol (90:10), column temperature 30°C; injection volume 10μl; detection wavelength 254nm. The ammonium dihydrogen phosphate solution was prepared by weighing 5g of ammonium dihydrogen phosphate, dissolving it in 1000ml of water, and adjusting the pH to 2.5 with phosphoric acid.

[0040] Prepare the test solution and reference solution according to the method in "Chromatographic Condition 2", then inject and test them respectively according to the above chromatographic conditions and record the chromatograms.

[0041] (2) Second optimization of mobile phase Chromatographic conditions: YMC-Pack ODS-AQ 250×4.6mm l.D.S-5μm, 12nm; mobile phase: ammonium dihydrogen phosphate solution-methanol (90:10), column temperature 30°C; injection volume 10μl; detection wavelength 254nm. The ammonium dihydrogen phosphate solution was prepared by weighing 15g of ammonium dihydrogen phosphate, dissolving it in 1000ml of water, and adjusting the pH to 2.5 with phosphoric acid.

[0042] Prepare the test solution and reference solution according to the method in "Chromatographic Condition 2", then inject and test them respectively according to the above chromatographic conditions and record the chromatograms.

[0043] The chromatograms of the two mobile phase optimizations can be found in Figure 6 ,from Figure 6 As can be seen, the chromatographic peak of disodium edetate is preceded by the peak of ammonium dihydrogen phosphate, and the peak area increases with increasing ammonium dihydrogen phosphate concentration. Furthermore, the resolution between the ammonium dihydrogen phosphate peak and the chromatographic peak of disodium edetate-metal chelate is poor. To avoid interference from the mobile phase peaks, tetrabutylammonium hydroxide was used as the ion-pairing reagent to improve the resolution of the chelate peaks.

[0044] 4. Chromatographic Condition 4: Optimization of Mobile Phase and Injection Volume Chromatographic conditions: YMC-Pack ODS-AQ 250×4.6mm l.D. S-5μm, 12nm; mobile phase: 0.09% tetrabutylammonium hydroxide solution (adjusted to pH 2.3 with phosphoric acid): acetonitrile = 70:30; column temperature 30°C; injection volume 20μl; detection wavelength 254nm.

[0045] Prepare the test solution and reference solution according to the method in "Chromatographic Condition 2", then inject the test solution and reference solution into the sample according to the above chromatographic conditions and record the chromatogram. The results are as follows: Figure 2 As shown, from Figure 2 It can be seen that there is an obvious front peak in the chromatogram, which may be caused by substances in the sample solution that interfere with quantification, and the chromatographic conditions need to be further optimized.

[0046] 5. Chromatographic Conditions 5 Chromatographic conditions: YMC-Pack ODS-AQ 250×4.6mm l.D. S-5μm, 12nm; mobile phase: 0.09% tetrabutylammonium hydroxide solution (adjusted to pH 2.3 with phosphoric acid): acetonitrile = 70:30; column temperature 30°C; injection volume 20μl; detection wavelength 254nm.

[0047] Prepare the test solution and reference solution according to the method in "Solution Preparation Method", inject the sample for testing, and record the chromatogram. Figure 1As shown in the figure, after adding ascorbic acid, the obtained test solution is a colorless and transparent solution, which can effectively reduce the background noise. No front peak appears in the chromatographic peak of disodium edetate-metal chelate, which improves the sensitivity of the chromatographic analysis method.

[0048] 3. Methodological Validation Prepare 6 aliquots of the test solution according to the solution preparation method and inject them for testing. The RSD value of the content calculated by the external standard method should be no greater than 2.0%.

[0049] The repeatability results of the test solution disodium edetate content are shown in Table 1. As shown in Table 1, the repeatability results meet the acceptance criteria.

[0050] Table 1 Repeatability results of disodium edetate content in test solution

[0051] In summary, the present invention determines a method suitable for sample pretreatment of epinephrine hydrochloride injection by adding L-ascorbic acid, screening the concentration ratio of 0.02 mol / L ferric chloride solution and disodium edetate in the test solution, preparing the sample with 0.1% phosphoric acid, and filtering with a 0.45 μm water filter membrane, so that the disodium edetate in the sample can be fully complexed with metal ions and accurately determined.

[0052] In addition, the present invention determines that the type of ionic reagent added to mobile phase A is tetrabutylammonium hydroxide, the pH of mobile phase A is controlled at 2.3, and a YMC-Pack ODS-AQ 250×4.6mm l.D.S-5μm, 12nm chromatographic column is used. Under the chromatographic conditions of the present invention, disodium edetate in a test solution can be effectively separated. After liquid phase injection, the asymmetry of the chromatographic peak of disodium edetate-metal chelate measured is 1.02. The content of disodium edetate in the test solution can be accurately determined using a reference substance external standard method, thereby providing an accurate, efficient, convenient, and widely used method for detecting the EDTA content in epinephrine hydrochloride injection.

[0053] The above are only some embodiments of the present invention. For those skilled in the art, several modifications and improvements can be made without departing from the inventive concept of the present invention, which all fall within the scope of protection of the present invention.

Claims

1. A method for detecting the content of disodium edetate in epinephrine hydrochloride injection, characterized in that: High performance liquid chromatography was used for detection. Samples were pretreated as follows before injection: L-ascorbic acid was added to the epinephrine hydrochloride injection, followed by the addition of a metal ion solution. The solution was then diluted with an acid solution until the concentration of disodium edetate in the solution was 0.5-20 μg / mL, and then filtered.

2. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to claim 1, wherein The metal ion solution is selected from at least one of ferric chloride solution, copper sulfate solution, and ferrous sulfate solution; the metal ion concentration in the metal ion solution is 0.015-0.03 mol / L.

3. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to claim 2, wherein The ratio of the concentration of disodium edetate to the volume of the metal ion solution is 5:0.5 to 5:

2.

4. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to claim 1, wherein The acid solution is selected from at least one of phosphoric acid, hydrochloric acid and sulfuric acid.

5. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to claim 1, wherein The specific step of filtration is filtering with a water filter membrane.

6. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to any one of claims 1 to 5, characterized in that: The mobile phase A used in the high performance liquid chromatography method is an ion pair reagent, the mobile phase B is methanol or acetonitrile, and the ratio of the mobile phase A to the mobile phase B is 50:50-95:

5.

7. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to claim 6, wherein: The ion pair reagent is selected from tetrabutylammonium hydroxide solution, sodium lauryl sulfate solution or ammonium dihydrogen phosphate solution.

8. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to claim 7, wherein: The ion pair reagent is a 0.09% (V / V) tetrabutylammonium hydroxide solution with a pH of 1-3.

9. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to any one of claims 1 to 5, characterized in that: The chromatographic column used in the high performance liquid chromatography method is an octadecyl bonded silica gel chromatographic column.

10. The method for detecting the content of disodium edetate in the epinephrine hydrochloride injection according to any one of claims 1 to 5, characterized in that: The detection wavelength of the high performance liquid chromatography method is 200-500 nm; the flow rate is 0.9-1.5 mL / min; the column temperature is 25-35° C.; and the injection volume is 15-25 μL.

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