Abstract The objective of this study was to perform biological monitoring of subjects who are occupationally exposed to pyrethroids. The study group consisted of 30 pest control operators exposed to cyfluthrin, cypermethrin or permethrin. After exposure, 24-h urine samples were collected and 20 ml of blood was drawn. The pyrethroid metabolites cis- and trans-3-(2,2-dichlorovinyl)-2,2-dimethylcyclopropanecarboxylic acid, 3-phenoxybenzoic acid and fluorophenoxybenzoic acid were determined in the urine samples (limit of detection: 0.5 μg/l) by GC MS and the pyrethroids in plasma (limit of detection: 5 μg/l) by GC-ECD. The concentrations of metabolites in the urine of the pest control operators ranged between <0.5 μg/l and 277 μg/l urine. The concentrations of cyfluthrin, cypermethrin and permethrin in the plasma were below the limits of detection (< 5 μg/l). To test if the metabolites are specific for pyrethroid exposure, they were determined in the urine of non-exposed subjects ( n = 40). In no case could pyrethroid metabolites be detected. A cyfluthrin elimination experiment showed that cyfluthrin metabolites are eliminated following first-order kinetics ( t 1 2 = 6.4 h ). Storage experiments demonstrate that frozen urine samples (− 21°C) show no significant losses of metabolites within a year. In contrast, pyrethroids stored in plasma are susceptible to further biodegeneration.