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This paper describes the analysis of anticancer drug hexamethylene bisacetamide (HMBA) in biological fluids and tissues by RP-HPLC. The samples of plasmas, urine and tissues from cancer mice and rabbits were first used with methanol for pretreatment to remove protein impurity, then analyzed by RP-HPLC on an ODS column. The mobile phase was methanol/water (70/30, V/V), UV detection is employed at 210 nm. 153 samples were analyzed and good results were obtained. The recoveries of 93.3%-104% with a relative standard deviation of 0.04%. The regression equation is: Y = 0.32X - 1.07, with a linear correlation coefficient of 0.998 and the linear range of HMBA is 0 to 1000 mg/L. A sample analysis can be completed within 13 minutes. This method could be satisfactorily applied not only for the analysis of HMBA but probably also for the analysis of other drugs in biological fluids and tissues.
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