Abstract: Objective To establish a method for determining the residual amount of chloroform, acetic acid and ethyl acetate in isosorbide nitrate APIs. Methods The chromatographic column was a PEG-20M stainless steel column (4m × 2mm). The temperature of the column was programmed. The initial column temperature was maintained at 50 ℃ for 3min, then increased to 180 ℃ at a rate of 10 ℃ / min for 5min, and then at 10 ℃ / The rate of min rose to 280 ° C for 10 min. FID detector, detector temperature 320 ℃, inlet temperature 320 ℃, nitrogen as carrier gas, flow rate 50ml / min, hydrogen flow rate .30ml / min, air flow rate 400ml / min. Results The three organic solvents were completely separated. Chloroform showed a good linear relationship in the concentration range of 0.0005 to 0.003 mg / ml. The recovery rate was 100.3% (RSD <1.0%, ethyl acetate was in the concentration range of 0.005 to 0.03 mg / ml. There is a good linear relationship (r = 0.9994), the recovery rate is 100.5%, RSD <1.0%, acetic acid has a good linear relationship (r = 0.9997) in the concentration range of 0.005 ~ 0.03mg / ml, the recovery rate is 98.7 %, (RSD <1.0%). Conclusion: This method is sensitive and accurate, suitable for the detection of organic residues in isosorbide nitrate.
Keywords: isosorbide dinitrate, chloroform, ethyl acetate, acetic acid, high performance gas chromatography
Isosorbide dinitrate is an organic nitrate drug, and its preparation (tablet) is clinically used for the treatment of coronary heart disease and angina pectoris, and is one of the most widely used cardiovascular drugs. In the synthesis process, organic reagents such as chloroform, acetic acid, and ethyl acetate are used. Chloroform is harmful to the human body. Acetic acid can easily cause the decomposition of the main drug in the preparation. Ethyl acetate has a bad smell, so it is necessary to control its residual amount. Simultaneous determination of three organic solvents in isosorbide nitrate raw materials by gas chromatography is sensitive, accurate and reliable.
1 Instruments and reagents
Shimadzu GC-14C gas chromatography system, isosorbide nitrate (Shandong Linuo Pharmaceutical Factory), chloroform, acetic acid, ethyl acetate and DMF are all analytically pure.
2 Experimental part
2.1 Preparation of solution
2.1.1 Preparation of standard solution: Take appropriate amount of chloroform, acetic acid and ethyl acetate, dilute with DMF to 0.01mg per 1ml containing chloroform and 0.1mg containing ethyl acetate, shake well to obtain the standard stock solution. Take appropriate amount of standard stock solution and dilute to obtain reference solution.
2.1.2 Preparation of test solution: Take 1 g of isosorbide dinitrate, weigh accurately, place in a 10 ml volumetric flask, add DMF to dissolve and dilute to the mark, shake to obtain the test solution.
2.2 Chromatographic conditions and content calculation method
The meteorological chromatography column is a PEG-20M stainless steel column (4m × 2mm). The column temperature is programmed to increase the temperature. The initial column temperature is maintained at 50 ℃ for 3min, then increased to 180 ℃ at a rate of 10 ℃ / min for 5min, and then at 10 ℃ / The rate of min rises at 280 ° C for 10 min. FID detector, detector temperature 320 ℃, inlet temperature 320 ℃, nitrogen as carrier gas, flow rate 50ml / min, hydrogen flow rate 30ml / min, air flow rate 400ml / min. The theoretical plate number is not less than 1600 in terms of ethyl acetate, and the injection volume of the reference substance and the test sample solution is 2μl. The external standard peak area method is used to calculate the content.
3 Results and discussion
3.1 Investigation of linear relationship
Precisely measure the standard stock solution 0.5, 1.0, 1.5, 2.0, 2.5, 3.0ml to 10ml volumetric flask, add DMF to dilute to the mark, shake well to obtain the series of standard solutions â… -â…¥. For sample injection measurement, linear regression was performed with the concentration (C) of each solution as the abscissa and the corresponding peak area (A) as the ordinate. The linear equations for chloroform and ethyl acetate were Y = 38.01 + 3535X, (r = 0.9994 ), Y = 9.51 + 2926.76X, (r = 0.9997), Y = -5.34478 + 1198.35821X, (r = 0.9999), indicating that chloroform is in the range of 0.0005 to 0.003 mg / ml and the concentration of acetic acid and ethyl acetate is 0.005 to The linear relationship is good in the range of 0.03mg / ml.
3.2 Precision test
Take 1 reference solution and inject 5 times. As a result, the RSD of the chloroform peak area = 2.6%, the RSD of the acetic acid peak area = 3.6%, and the RSD of the ethyl acetate peak area = 3.9%.
3.3 Recovery rate test
Precisely weigh the same batch of known residual amount of isosorbide nitrate sample 0.5g (3 parts), (containing chloroform 6.5 × 0.0001mg, containing acetic acid, ethyl acetate 8.5 × 0.001mg, placed in a 10ml measuring bottle, respectively Add standard stock solutions 0.05ml, 0.10ml, 0.15ml, then dissolve with DMF, dilute to the mark, shake well. Inject samples according to the above chromatographic conditions, calculate the recovery rate, the recovery rates of chloroform, acetic acid and ethyl acetate are: 100.3 %, 98.7% and 100.5%.
3.4 Determination of detection limit
Dilute the standard solution in 2.1.1 step by step and inject samples according to the above chromatographic conditions. The detection limits (N = 3) of chloroform, acetic acid, and ethyl acetate are 30 mg, 50 mg, and 100 mg, respectively.
3.5 Sample determination
Accurately measure 2μl each of the control solution and the test solution, inject samples according to the above chromatographic conditions, and calculate the organic residue in each sample according to the peak area according to the external standard method. The results show that the three batches of samples all meet the requirements of the organic residue standard .
3.6 Selection of experimental conditions
Since chloroform is a highly polar substance, a packed column of PEG-20M stainless steel column with strong polar fixative is selected; due to the difference in boiling points of chloroform, acetic acid and ethyl acetate, the constant temperature separation time is long and the peak shape is poor and easy to match The solvents overlap, so the temperature-programmed mode is used; and because the experiment requires lower detection limits and are all carbon-containing compounds, a hydrogen flame detector with higher sensitivity is used.
references
1 ChP (Chinese Pharmacopoeia) (Part 2) Appendix, 64
2 China Pharmaceutical Standards, 2002, 3 (4) 29
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