A quantitative method using high-performance liquid chromatography-mass spectrometry (LC-MS, ion trap) after matrix supported liquid-liquid extraction is described for the simultaneous determination in whole blood of 33 benzodiazepines including metabolites and benzodiazepine-like substances. The limits of detection (LOD) range from 0.0001 to 0.0126 mg/l. Linearity is satisfactory for all compounds. The extraction recoveries for the benzodiazepines in whole blood are between 60 and 91%, desmethyldiazepam, OH-bromazepam and brotizolam excepted. Selectivity, accuracy and precision are satisfactory for clinical and forensic purposes.