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Through methodological investigation, the linear range, precision, stability, repeatability and sample recovery rate of the method were all good, and a reliable quality control method was established for the quality analysis of the preparation.
1 Instruments and reagents Shimadzu CS-930 thin layer scanner, S10200A ultrasonic cleaner, berberine hydrochloride reference product provided by China Biological Products Certification Institute. The reagents used were chloroform, methanol benzene, ethyl acetate and isopropanol. Thin layer chromatography was produced with silica gel G (60 type) China Qingdao Marine Chemical Group Co., Ltd.
2 Methods and results
2.1 Chromatography and chromatographic conditions The thin layer plate is a silica gel G plate with a plate thickness of 0.5 mm, 110 ° C for 30 min, room temperature of 24 to 28 ° C, and a relative humidity of 56% to 72%. The developing agent: benzene-ethyl acetate-isopropyl alcohol-methanol-water (20:10:5:5:1), the concentrated ammonia test solution was pre-equilibrated for 15 min, and the thin layer plate was simultaneously pre-equilibrated. Upward expansion.
2.2 Linear range inspection Precision weighing berberine hydrochloride reference substance (batch number: 0713-9906) 10,01mg, placed in a 25mL volumetric flask, add methanol to dissolve and dilute to the mark, shake well, accurately measure 1mL set lOmL amount Add methanol to the mark and shake well to make a solution containing 0.04004 mg per 1 mL.
Accurately draw 0.5, 1, 2, 4, 8 μl, respectively, on the same silica gel G thin layer plate, and operate under the above chromatographic conditions. The integral value of the fluorescence intensity of the reference substance was determined, and the regression equation was linear regression, Y=56922.79X+107.45, r=0.9999.
It can be seen that the linear amount of berberine hydrochloride in the range of 0.2002-0.32 μg is good.
2.3 Preparation of the test solution Take 5 bags of this product, grind finely, take about 1Og of powder, accurately weigh, add 25mL of water, sonicate for 3~5min, adjust the pH value by 9~10 with concentrated ammonia test solution, extract with chloroform 5 times (30, 30, 30, 20, 20mL) combined extracts, evaporated to dryness, the residue was added with methanol to dissolve, and transferred to a 5mL volumetric flask, add methanol to the mark, shake well, that is.
2.4 Precision test The sample with the batch number of 200103 was taken according to the method under the text [Content Determination]. The same method was used to continuously scan 5 points, and the RSD of the measurement result was 0.90%.
2.5 Stability test Investigate 5μl of the test solution, and measure the integral value of fluorescence intensity at time intervals of 0, 1, 2, 3, 4, 5h. The results show that there is no significant change in the integral value of fluorescence intensity measured within 5 hours. The RSD is 1.2%. The chemical properties of berberine hydrochloride in the test sample are stable.
2.6 Determination of sample content Take 5 batches of samples of the same batch number and prepare the test solution according to the preparation method of the test solution. Another appropriate amount of berberine hydrochloride reference substance was added, and methanol was added to make a solution containing 0.04 mg per 1 mL. As a reference solution.
Accurately draw 5 μl of the test solution, 1 μl and 3 μl of the reference solution, respectively, on the same silica gel G thin plate, with benzene-ethyl acetate-isopropanol-methanol-water (20:10:5:5: 1) As a developing agent, pre-equilibrated with concentrated ammonia test solution for 15 min (thin layer is simultaneously pre-equilibrated). Sample content determination results: 0.1036 mg, 0.1022 mg, 0.1048 mg, 0.1046 mg, 0.1029 mg per bag.
2.7 Recovery test Take the known accurate content of the sample (batch number: 200103 content is 0.008706 mg / g) about 5g, accurately weighed, add 25mL of water, sonicated for 5min, precision addition of berberine hydrochloride reference solution (0.04004mg /g) 1.2 mL, operated according to the content measurement method, the measurement result was x = 102.42 (n = 5), and the RSD was 0.91%.
3 Discussion This test is in the investigation of the preparation method of the test solution. According to the solubility properties of berberine hydrochloride, this experiment used hydrochloric acid-methanol solution (1:100) as the extraction solvent, heated in a 60 °C water bath for 15 min, sonicated for 30 min, added methanol to volume, filtered, and the filtrate was taken as a test. The solution of the product, the sample amount is relatively small, but the test sample TLC has background interference, and then changed to the preparation method of the test solution in the text, dissolved in water, alkalized and extracted with chloroform to reduce impurity interference, The clarity of the chromatogram was greatly improved. Under the selected TLC conditions, the berberine hydrochloride spot and the other components in the test sample were well separated from the baseline, and the resolution reached the Chinese Pharmacopoeia 2000 edition. Claim.
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Determination of Berberine Hydrochloride in Shuxin Chongji
Key words: thin-layer scanning method; Shuxin granules; berberine hydrochloride content research Shuxin granules, which are made by extracting and processing 5 kinds of Chinese medicinal materials such as Salvia miltiorrhiza , Radix Scutellariae and Phellodendron chinense . It has the effect of promoting blood circulation, nourishing yin and qi, and removing phlegm. Fluorescence scanning of berberine hydrochloride was carried out by TLC scanning.