Abstract: Ephedrine hydrochloride is a white needle crystal or crystalline powder, which is the hydrochloride salt of ephedrine. It can be used for bronchial asthma, whooping cough, hay fever and other allergic diseases. It can also fight blood pressure reduction and pupil dilation caused by spinal anesthesia. It can also be used for myasthenia gravis, dysmenorrhea and other diseases. It can also be used as a central nervous system stimulant.
Objective To establish a method for determination of ephedrine hydrochloride in Chuankening tablets. Methods Reversed-phase high-performance liquid chromatography was used, using Hanbang C18 (4.6 × 250mm, 5μm) chromatographic column, with acetonitrile-0.1% phosphoric acid (5:95) as mobile phase, flow rate 0.8ml / min, detection wavelength 207nm, column Temperature 30 ℃. Results Under this condition, ephedrine hydrochloride showed a good linear relationship in the range of 0.01174 to 0.1174 μg (r = 1.0000), with an average sample recovery rate of 98.3% and an RSD of 0.7% (n = 6). Conclusion This method is simple to operate and has good reproducibility, and can be used as a method for determining the content of this preparation.
Chuankening tablets are compound preparations composed of 5 Chinese herbal medicines, such as bellflower, pig bile, licorice, bitter almond cream, and ephedra. The original quality standards are contained in "Fourth Volume of Chinese Medicine Prescriptions of the Ministry of Health." In this prescription, ephedra is the main medicine, which has the functions of sweating and dispelling cold, relieving lungs and relieving asthma, and diminishing swelling. It is used for diseases such as cold, cold, chest tightness, cough, fengshui edema, and bronchial asthma. The main active ingredient in ephedra is ephedrine hydrochloride. There is no method for content determination in the original standard, and its intrinsic quality cannot be effectively controlled. In order to control the intrinsic quality of the preparation more effectively, this experiment uses reversed-phase high performance liquid chromatography to determine ephedra The content of ephedrine hydrochloride in the method is easy to operate, has good reproducibility, is reasonable and applicable, and can be used as a quantitative indicator to control the internal quality of Chuankening tablets.
1. Materials and methods 1.1 Instruments and test drugs Shimadzu LC-2010AHT high-performance liquid chromatograph, ultraviolet detector; KQ-100 ultrasonic cleaner; Shimadzu UV-2450 ultraviolet spectrophotometer; ephedrine hydrochloride reference substance ( Lot No. 171242-200404, purchased from China National Institute for the Control of Pharmaceutical and Biological Products); acetonitrile is chromatographically pure reagent, water is purified water, and other reagents are analytically pure; Chuankening tablets (produced by a pharmaceutical industry in Jilin Province).
1.2 Method 1.2.1 Preparation of reference substance solution Weigh an appropriate amount of ephedrine hydrochloride reference substance, add methanol to make a solution containing 11.74 μg per 1 ml.
1.2.2 Preparation of the test product solution Take 10 tablets of this product (lot number 20051103), remove the sugar coating, determine the average tablet weight, grind it, take 1g, accurately weigh it, put it in a conical flask with a stopper, and add 25ml of methanol in the precision, Weigh the weight, sonicate for 40 min, let cool, weigh it again, make up the lost weight with methanol, shake well and filter. Accurately measure 1ml of the continuous filtrate, place it on a neutral alumina column (100-200 mesh, 1.5g, inner diameter 1cm), elute with 50% methanol, collect the eluent about 9ml, place in a 10ml volumetric flask, add 50% Methanol to the mark, shake well, that is.
1.2.3 Chromatographic conditions 1.2.3.1 Selection of measurement wavelength Take the reference solution and scan it with Shimadzu UV-2450 ultraviolet spectrophotometer in the range of 200 ～ 400nm. As a result, ephedrine hydrochloride has maximum absorption at 207nm wavelength. Choose 207nm as the measurement wavelength of this experiment.
1.2.3.2 The system suitability test uses chromatographic columns: (1) Hanbang C18 (4.6 × 250mm, 5μm); (2) KromasilC18 (4.6 × 250mm, 5μm), with acetonitrile-0.1% phosphoric acid (5:95) The mobile phase has a detection wavelength of 207 nm and a column temperature of 30 ° C. Accurately draw 5μl of the reference substance solution and 10μl of the test substance solution into the liquid chromatography. The theoretical plates of the ephedrine hydrochloride peaks of the reference substance and the test substance are: (1) 15711, 15366; (2 ) 11064,11118. Based on this, a reasonable number of theoretical plates should be determined. The theoretical number of plates should be no less than 4000 based on the chromatographic peak of ephedrine hydrochloride.
1.2.4 Methodological investigation 1.2.4.1 Preparation of standard curve Precision pipette reference solution 1, 4, 6, 8, 10 μl, respectively, injected into the liquid chromatograph, with the injection volume as the abscissa and the peak area as the ordinate, draw the standard Curve, the calculation regression equation is Y = 1641856X-205.245 (r = 1.0000), the results show that ephedrine hydrochloride in the range of 0.01174 ~ 0.1174μg, showing a good linear relationship.
1.2.4.2 Negative control test According to the preparation method of the test product solution, a negative control solution containing no ephedra medicinal materials is prepared. Pipette 10μl of this solution into liquid chromatography and determine it according to the law. As a result, no interference peak appeared near the retention time corresponding to the chromatographic peak of ephedrine hydrochloride, thus proving the method is reasonable and feasible.
1.2.4.3 Stability test Take the same test solution and inject 10μl at 0, 5, 10, 15, 20h respectively. Based on the peak area of ​​ephedrine hydrochloride, the average peak area is 125535 and the RSD is 0.9%. The results showed that the content of ephedrine hydrochloride in the test sample was stable within 20 hours.
1.2.4.4 Precision test Take 10 μl of the same test solution and repeat the injection 6 times. The average peak area of ​​ephedrine hydrochloride is 125490 and the RSD is 0.8%. The measurement results show that the precision of the method is good.
1.2.4.5 Reproducibility test Take the same test sample, prepare 6 copies in parallel according to the method of preparation of the test sample solution, inject 5μl of the reference solution, and inject 10μl of the test solution, and measure ephedrine hydrochloride in the sample according to the law The average value of the content is 1.12mg / g and the RSD is 1.0%, indicating that the method has good reproducibility.
1.2.4.6 Recovery rate test Weigh accurately 6 samples of known content 0.5g (batch number 20051103, content 1.114mg / g) determined by the same method, and add freshly prepared ephedrine hydrochloride reference solution (24.96) μg / ml) 25ml, prepared according to the preparation method of the test solution, the control solution was injected 5μl, the test solution was injected 10μl, measured according to the method, and the recovery rate was calculated. The average recovery rate of ephedrine hydrochloride was 98.3%, RSD 0.7%. It shows that this method is accurate and feasible.
1.2.4.7 Sample measurement results Take 5 batches of Chuankening tablets samples, respectively prepared according to the preparation method of the test solution, the control solution injection 5μl, the test solution injection 10μl, each batch determination 2 times, ephedrine hydrochloride The average content is shown in Table 1. Table 1 Sample measurement results 2. Discussion In this experiment, the preparation method of the test solution was investigated. Two methods have been used, one is the method of the upper neutral alumina column described in this article, and the other is the method of adding methanol to ultrasonic treatment and directly loading the sample after filtration. The results measured by the two methods are basically the same, but it can be seen from the comparison that the test sample without the treatment of the neutral alumina column shows that the impurity is obviously too much, and there is interference of impurity peaks, which is easy to damage the column Therefore, the method of using a neutral alumina column is selected.
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