A method for determining the content of carboprost tromethamine injection

Abstract

The invention relates to the technical field of drug detection, and in particular to a method for determining the content of a carboprost tromethamine injection. The method comprises the following steps: S1, taking a test solution, preparing a reference solution and a system adaptability solution; the reference solution contains a carboprost tromethamine reference; the system adaptability solution contains carboprost tromethamine and carboprost 5,6-trans isomer; S2, using the system adaptability solution to adjust chromatographic conditions until the separation degree between a carboprost 5,6-trans isomer peak and a carboprost peak is greater than 1.5, and the carboprost peak tailing factor is less than 2.0, thereby obtaining an adaptability chromatographic condition; the detection wavelength of the adaptability chromatographic condition is 210nm; S3, subjecting the test solution and the reference solution to high performance liquid chromatography analysis according to the adaptability chromatographic condition, calculating by peak area according to an external standard method (the conversion coefficient between carboprost and carboprost tromethamine is 0.7526), ​​thereby obtaining the content of the carboprost tromethamine injection.

Description

Technical Field

[0001] The invention relates to the technical field of drug detection, and in particular to a method for determining the content of carboprost tromethamine injection. Background Art

[0002] The content of carboprost tromethamine injection disclosed at home and abroad is usually determined by the following method (high performance liquid chromatography, Appendix VD of Part II of the Chinese Pharmacopoeia 2010 Edition):

[0003] Chromatographic conditions and system suitability test: Use silica gel as filler, water-1,3-butanediol-dichloromethane (0.25:3.5:496) as mobile phase, detection wavelength is 254nm. The number of theoretical plates calculated based on the carboprost peak should not be less than 2000; the separation between the carboprost peak and the internal standard substance peak should meet the requirements.

[0004] Preparation of internal standard solution: Take an appropriate amount of guaifenesin, add it to the mobile phase to dissolve and dilute it to make a solution containing about 0.3 mg per 1 ml.

[0005] Determination method: operate in dark. Accurately measure an appropriate amount of this product (equivalent to approximately 0.5 mg of carboprost), place it in a stoppered centrifuge tube, accurately add 20 mL of dichloromethane and 1 mL of citric acid buffer (take 10.5 g of citric acid, add an appropriate amount of water to dissolve, adjust the pH value to 4.0 with 5 mol / L sodium hydroxide solution, then dilute to 100 mL with water and shake well), shake vigorously for 10 minutes, centrifuge, discard the water layer, accurately measure 5 mL of the lower layer solution, blow dry with nitrogen, add 100 μl of freshly prepared 2% α-bromo-2-acetonaphthyl acetonitrile solution L and 50 μL of acetonitrile solution of 1% diisopropylethylamine, rotate four times to dissolve the residue, place at 30-35°C for more than 15 minutes, blow dry with nitrogen, then accurately add 2mL of internal standard solution to dissolve, shake well, take 20 μL and inject into liquid chromatograph, record chromatogram; take appropriate amount of carboprost tromethamine reference substance and benzyl alcohol, accurately weigh, add water to dissolve and quantitatively dilute to make a solution containing about 0.332mg of carboprost tromethamine and 9mg of benzyl alcohol per 1mL, accurately measure 2mL, and determine in the same way. Calculate by peak area according to internal standard method (the conversion coefficient of carboprost and carboprost tromethamine is 0.7526), ​​that is,

[0006] Since the content of carboprost tromethamine in carboprost tromethamine injection is low and its ultraviolet absorption is weak, the method recorded in the above pharmacopoeia uses a pre-column derivatization method to improve the ultraviolet absorption of carboprost tromethamine. However, the operation process of this method is extremely complicated: after multiple steps of extraction, drying, derivatization and re-drying, the internal standard substance is added, and the internal standard substance added in the last step cannot calibrate and eliminate the error caused by the fluctuation of the previous multiple steps of operating conditions, so this method is complicated to operate and has poor repeatability. Summary of the invention

[0007] In order to overcome the above-mentioned defects of the prior art, the technical problem to be solved by the present invention is to provide a method for quickly determining the content of carboprost tromethamine injection.

[0008] In order to solve the above technical problems, the technical solution adopted by the present invention is: a method for determining the content of carboprost tromethamine injection, comprising the following steps:

[0009] S1. Take carboprost tromethamine injection as the test solution, and prepare the reference solution and system suitability solution;

[0010] Wherein, the reference substance solution contains carboprost tromethamine reference substance;

[0011] The system adaptability solution contains carboprost tromethamine and carboprost 5,6-trans isomer;

[0012] S2. Using the system adaptability solution, the chromatographic conditions are adjusted until the separation degree between the peak of carboprost 5,6-trans isomer and the peak of carboprost is greater than 1.5, and the tailing factor of the peak of carboprost is less than 2.0, thereby obtaining the adaptive chromatographic conditions;

[0013] Wherein, the detection wavelength of the adaptive chromatographic condition is 210nm;

[0014] S3. Performing HPLC analysis on the test solution and the reference solution according to the chromatographic conditions, and calculating the peak area by the external standard method to obtain the content of carboprost tromethamine injection.

[0015] The beneficial effects of the present invention are as follows: the method for determining the content of carboprost tromethamine injection provided by the present invention can quickly and accurately determine the content of carboprost tromethamine injection, and the method is simple and highly repeatable. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 Shown is a chromatogram overlay in the investigation of specificity of a detection example in a specific implementation manner of the present invention. DETAILED DESCRIPTION

[0017] In order to explain the technical content, achieved objectives and effects of the present invention in detail, the following is an explanation in combination with the implementation modes and the accompanying drawings.

[0018] A method for determining the content of carboprost tromethamine injection comprises the following steps:

[0019] S1. Take carboprost tromethamine injection as the test solution, and prepare the reference solution and system suitability solution;

[0020] Wherein, the reference substance solution contains carboprost tromethamine reference substance;

[0021] The system adaptability solution contains carboprost tromethamine and carboprost 5,6-trans isomer;

[0022] S2. Using the system adaptability solution, the chromatographic conditions are adjusted until the separation degree between the peak of carboprost 5,6-trans isomer and the peak of carboprost is greater than 1.5, and the tailing factor of the peak of carboprost is less than 2.0, thereby obtaining the adaptive chromatographic conditions;

[0023] Wherein, the detection wavelength of the adaptive chromatographic condition is 210nm;

[0024] S3. The test solution and the reference solution are subjected to HPLC analysis according to the adaptive chromatographic conditions, and the content of the carboprost tromethamine injection is obtained by calculating the peak area by the external standard method (the conversion coefficient of carboprost to carboprost tromethamine is 0.7526).

[0025] The inventors found in experiments that by optimizing the chromatographic conditions and selecting a specific detection wavelength in combination with a systemic adaptive solution, it is possible to simplify the existing carboprost tromethamine injection content detection process while improving the accuracy and repeatability of the carboprost tromethamine injection content detection process.

[0026] Preferably, the concentration of the carboprost tromethamine reference substance in the reference substance solution is 0.33 mg / mL.

[0027] Preferably, the system adaptability solution contains 0.0025 mg / mL of carboprost 5,6-trans isomer and 0.25 mg / mL of carboprost. Wherein, the carboprost 5,6-trans isomer is carboprost tromethamine impurity A, i.e. (E)-7-[(1R,2R,3R,5S)-3,5-dihydroxy-2-[(E)-(3S)-3-hydroxy-3-methyl-1-octenyl]cyclopentyl]-5-heptenoic acid, Cas: 76498-29-8, and the specific structural formula is as follows:

[0028]

[0029] Preferably, the adaptive chromatographic conditions are:

[0030] Chromatographic column: octadecylsilane bonded silica gel column or its equivalent column;

[0031] Mobile phase: acetonitrile-methanol-0.02 mol / L potassium dihydrogen phosphate solution, pH 3.0;

[0032] Flow rate: 0.8 mL / min;

[0033] Column temperature: 30°C;

[0034] Injection volume: 20 μL.

[0035] Wherein, the chromatographic column is preferably a Hypersil BDS C18 4mm×150mm, 3μm or a chromatographic column with equivalent performance.

[0036] Preferably, the volume ratio of acetonitrile, methanol and 0.02 mol / L potassium dihydrogen phosphate in the mobile phase is 20:30:50, and the pH is adjusted to 3.0 with phosphoric acid.

[0037] Example 1

[0038] A method for determining the content of carboprost tromethamine injection comprises the following steps:

[0039] S1. Take carboprost tromethamine injection as the test solution;

[0040] S2. Accurately weigh the carboprost tromethamine reference substance (source: USP reference substance, purity: 98.3%), dissolve it in water and quantitatively dilute it to a solution containing 0.33 mg per 1 mL, thus obtaining the reference substance solution;

[0041] S3. Accurately weigh carboprost tromethamine (same as carboprost tromethamine mentioned above) and carboprost tromethamine impurity A (source: QCS reference substance, purity: 95.84%), dissolve in water and quantitatively dilute to a mixed solution containing 0.25 mg of carboprost and 0.0025 mg of carboprost tromethamine impurity A per 1 mL, to obtain a system adaptability solution;

[0042] S2. Using the system adaptability solution, the chromatographic conditions are adjusted until the separation degree between the carboprost tromethamine impurity A peak and the carboprost peak is greater than 1.5, and the carboprost peak tailing factor is less than 2.0, thereby obtaining adaptive chromatographic conditions;

[0043] Wherein, the adaptive chromatographic conditions are:

[0044] Column: Hypersil BDS C18 4 mm × 150 mm, 3 μm;

[0045] Mobile phase: acetonitrile-methanol-0.02 mol / L potassium dihydrogen phosphate solution (volume ratio: 20:30:50), phosphoric acid adjusted to pH 3.0;

[0046] Flow rate: 0.8 mL / min;

[0047] Column temperature: 30°C;

[0048] Injection volume: 20 μL;

[0049] Detection wavelength: 210nm;

[0050] S3. The test solution and the reference solution are subjected to HPLC analysis according to the adaptive chromatographic conditions, and the content of the carboprost tromethamine injection is obtained by calculating the peak area by the external standard method (the conversion coefficient of carboprost to carboprost tromethamine is 0.7526).

[0051] Test example

[0052] The effects of detection wavelength (200 nm and 210 nm) on method linearity and recovery were investigated.

[0053] A series of linear concentration reference solutions with concentrations of 70%, 80%, 100%, 110% and 130% were prepared by the aforementioned carboprost tromethamine reference solution preparation method, and analyzed at 200nm and 210nm detection wavelengths according to the adaptive chromatographic conditions shown in Example 1, respectively, and the linear equation was calculated based on each concentration and peak area, and the linearity of the method was evaluated by the correlation coefficient R and intercept of the linear equation. The linear data are shown in Table 1.

[0054] Take 80%, 100%, 120% of the prescription amount of carboprost tromethamine, add 100% of the prescription amount of auxiliary materials, respectively, to prepare recovery sample solutions (carboprost in each sample solution is about 200μg / ml, 250μg / ml, 300μg / ml, respectively, and the auxiliary material content is about 9mg / ml of sodium chloride, 9.45mg / ml of benzyl alcohol, and 83μg / ml of tromethamine), each group of samples is prepared in triplicate, and analyzed at 200nm and 210nm detection wavelengths according to the adaptive chromatographic conditions shown in Example 1, and the recovery of carboprost tromethamine in the recovery sample is calculated by the external standard method. The recovery data are shown in Table 2.

[0055] Table 1

[0056]

[0057] Table 2

[0058]

[0059]

[0060] As can be seen from Table 1, when the wavelength is 200nm, r=0.9981, indicating that the linearity does not meet the requirements (r should be greater than 0.999), while when the wavelength is 210nm, r=0.9999, which meets the linearity requirements. And when the wavelength is 200nm, the intercept of the linear equation is 1.86% (1534766.96 / 8236337.8) of the 100% concentration response value, which does not meet the linearity requirements (should be less than 2%), while when the wavelength is 210nm, the intercept of the linear equation is 0.5% of the 100% concentration response value, which meets the linearity requirements.

[0061] As can be seen from Table 2, when the wavelength is 200nm, 80% to 120% recovery samples were prepared, and the recovery rates of 9 samples were between 98.15% and 103.24%, with an average recovery rate of 100.37%, and RSD (n=9) was 1.82%. When the wavelength is 210nm, 80% to 120% recovery samples were prepared, and the recovery rates of 9 samples were between 99.92% and 100.36%, with an average recovery rate of 100.19%, and RSD (n=9) was 0.16%. At the same time, the results show that the sample recovery rate measured at 200nm wavelength shows a decreasing trend with the decrease in sample concentration.

[0062] The above linear test and recovery test results both show that setting the wavelength to 210 nm is better than 200 nm.

[0063] Exclusive inspection

[0064] Each sample was prepared according to Table 3, and chromatographic analysis was performed using the adaptive chromatographic conditions shown in Example 1. The results are shown in Tables 4 and Figure 1 shown.

[0065] Table 3

[0066]

[0067]

[0068] Table 4

[0069]

[0070] from Figure 1 It can be seen that the blank solvent, blank excipients and known impurities (impurity A and impurity B) do not interfere with the main peak under the aforementioned adaptive chromatographic conditions test, and the main peak purity angle of carboprost tromethamine is less than the purity threshold, indicating that there is no co-elution in the main peak, and the chromatographic conditions have good specificity.

[0071] Repeatability test

[0072] Two operators prepared the reference substance and the test substance respectively according to Table 5, and each prepared 6 solutions in parallel. The samples were injected and analyzed by different instruments at different times. The chromatographic conditions were the adaptive chromatographic conditions shown in Example 1. The content of the test substance was calculated, and the results are shown in Table 6.

[0073] Table 5

[0074]

[0075] Table 6

[0076]

[0077]

[0078] As can be seen from Table 6, the average content of the 12 test samples prepared by the two operators was 99.4%, the RSD (n=6) was less than 1%, and the RSD (n=12) was 0.73%, that is, the precision of the determination of carboprost tromethamine content by this analytical method was good.

[0079] In summary, the method for determining the content of carboprost tromethamine injection provided by the present invention can quickly and accurately determine the content of carboprost tromethamine injection, and the method is simple and highly repeatable.

[0080] The above descriptions are merely embodiments of the present invention and are not intended to limit the patent scope of the present invention. Any equivalent transformations made using the contents of the present invention's specification and drawings, or directly or indirectly applied in related technical fields, are also included in the patent protection scope of the present invention.

Claims

1. A method for determining the content of carboprost tromethamine injection, characterized in that: The steps include: S1. Take carboprost tromethamine injection as the test solution, and prepare the reference solution and system suitability solution; Wherein, the reference substance solution contains carboprost tromethamine reference substance; The system suitability solution contains carboprost 5,6-trans isomer at a concentration of 0.0025 mg / mL and carboprost at a concentration of 0.25 mg / mL; S2. Using the system adaptability solution, the chromatographic conditions are adjusted until the separation degree between the peak of carboprost 5,6-trans isomer and the peak of carboprost is greater than 1.5, and the tailing factor of the peak of carboprost is less than 2.0, thereby obtaining the adaptive chromatographic conditions; Wherein, the detection wavelength of the adaptive chromatographic condition is 210nm; S3, subjecting the test solution and the reference solution to HPLC analysis according to the adaptive chromatographic conditions, and calculating the peak area by the external standard method to obtain the content of carboprost tromethamine injection; The adaptive chromatographic conditions are: Chromatographic column: octadecylsilane bonded silica gel column; Mobile phase: acetonitrile-methanol-0.02 mol / L potassium dihydrogen phosphate solution, pH 3.0; Flow rate: 0.8 mL / min; Column temperature: 30°C; Injection volume: 20 μL; The volume ratio of acetonitrile, methanol and 0.02 mol / L potassium dihydrogen phosphate in the mobile phase is 20:30:

50.

2. The measuring method according to claim 1, characterized in that: The concentration of the carboprost tromethamine reference substance in the reference substance solution is 0.33 mg / mL.

3. The measuring method according to claim 1, characterized in that: The carboprost 5,6-trans isomer is carboprost tromethamine impurity A.

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