Study On Quality Of The Related Substances Of Diosmin

Object: To optimize the HPLC conditions of examination the related substances of Diosmin(DASM), and establish the standard of DASM according to the content of the related substances.Methods: Referred to EP8.0 and related literatures and combined the DASM synthesis process, we established and optimized the HPLC method to examine the related substances of DASM. By adjusting the Chromatography column, mobile phase, column temperature and the flow rate, we found the best chromatographic conditions. Use the new established method to examine the content of related substances of DASM. The detecting wavelength was set as 347 nm when examining the content of DASM, and set as 275 nm when examining the related substances. Impurities A, C and D were positioned by relative retention time; External standard method was carried out to detecting impurities B,E, F, diosmetin-7-O-?-D-glucopyranoside(impurity X) and DASM. At last, we studied the stability of DASM by detecting the content of each related substances.Results: The best chromatographic conditions of examining the related substances were: use DMSO as solvent, chromatography column choose Welch(C18,4.6 mm×100mm,3 ?m), mobile phase: glacial acetic acid aqueous solution(6:66)-methanol-acetonitrile(67:32:1), the column temperature was 30?, the flow rate was 1.0ml/min, detection wavelength was 275 nm. The content of impurity A was less than 1%, impurity B was less than 5%, impurity C was less than 3%, impurity D was less than 1%,impurity E was less than 3%, impurity F was less than 3%, impurity diosmetin-7-O-?-D-glucopyranoside(impurity X) was less than 0.8%, the other impurities were totally not more than 1%. The content of DASM was between 90%-102%. Under the environment of high temperature and high humidity, DASM easier to create impurity A and impurity X, its color could be turned into gray-yellow from bright-yellow.Conclusion: The newly optimized method can welly separate each related substance, and determine its content accurately. The content of each related substance of DASM can be limited by using the result of this article, and the DASM stand drafting according to it should be more reliable.