In this study the authors report a new method to determine estazolam in blood and urine by high pressure liquid chromatography (HPLC). This compound is a triazolobenzodiazepine and shows an interesting psychopharmacological activity. The extraction from biological fluids was carried out using a mixture of ethylenechloride, methylenechloride, and ethylacetate (1:1:8). The HPLC analysis was carried out with a C18 column, using methanol:phosphate buffer (pH 7.5, 0.011M):acetonitrile (65:33:2) as mobile phase. The detector was set at lambda = 240 nm. The method shows good repeatability and a linear response in the range of 0.6 to 10 micrograms estazolam/mL both in serum and urine. A case of non-lethal acute intoxication in which the method was applied is also reported.
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