Single procedure for detection, confirmation, and quantification of benzodiazepines in serum by liquid chromatography with photodiode-array detection.

We developed a reversed-phase chromatographic procedure for detecting benzodiazepines and other drugs in serum. A liquid-liquid extraction step with hexane/ethyl acetate isolates the drugs from serum; absolute recoveries are generally greater than 85%. Reconstituted extracts are chromatographed on a 4-microns (particle size) C18 column; 14 drugs and an internal standard (flunitrazepam) are separated in 8 min. Peak detection, purity checking, and identification are performed with a computerized photodiode-array detector. Run-to-run imprecision (CV) for many benzodiazepines is less than 3%. In a study of 126 specimens from Emergency Department patients, the procedure showed excellent agreement with a gas-chromatographic method involving either mass-spectrometric or flame-ionization detection. This single procedure provides rapid and accurate detection, quantification, and confirmation of benzodiazepines in serum.