Zebrafish (Danio rerio) early life stages provide an important model for chemical risk assessment due to their genetic similarity to humans and the availability of well-established high-throughput techniques. Cells isolated from zebrafish conserve all these advantages and can, in addition, be multiplied in the laboratory to an unlimited extend. However, the biotransformation capacities of both systems have not been fully charac-terized. One biotransformation pathway, which is of utmost importance for the clearance of electrophilic compounds and phase I biotransformation products, is the mercapturic acid pathway. Its first step is the conjugation of the electrophile with glutathione, catalyzed by glutathione S-transferases (GSTs). The gluta-thione conjugates are usually further biotransformed within the mercapturic acid pathway by sequential re-moval of the glutamyl- and glycyl-moieties and an acetylation step followed by elimination of the mercap-turate. Considering the significance of this biotransformation route for the outcome of toxicological investi-gations, this thesis aims to close some major knowledge gaps regarding the protein expression of GSTs and the functionality of the mercapturic acid pathway in two test systems, zebrafish early life stages and the embryo-derived cell line PAC2. To investigate the repertoire of cytosolic GSTs and their expression dynamics, a targeted proteomics meth-od using electrospray ionization (ESI) was developed. Zebrafish embryos showed a basal expression of GST isoenzymes belonging to the classes alpha, mu, pi and rho as early as 4 hours post fertilization, indi-cating maternal transfer. After hatching, all cytosolic GST classes could be detected in the free-swimming larvae. The embryo-derived cell line PAC2 also expressed all cytosolic GST classes, except class alpha. Motivated by the abundant expression of GSTs in the zebrafish models, focus was set on the potential of GSTs to biotransform a model substrate (1-chloro-2,4-dinitrobenzene, CDNB) and to initiate its further bio-transformation within the mercapturic acid pathway. Since CDNB has no acid/base properties, an atmos-pheric pressure chemical ionization (APCI) method based on electron capture was developed for CDNB analysis at and below the nontoxic concentrations. For the determination of CDNB biotransformation prod-ucts of the mercapturic acid pathway, an ESI method was developed. In both test systems, the expression of cytosolic GSTs was not affected by non-toxic concentrations of the model substrate CDNB. Further-more, both test systems disclosed a fully functional mercapturic acid pathway with the first (glutathione conjugate) and last (mercapturate) biotransformation product being produced and excreted in zebrafish early life stages and PAC2 cells. To conclude, this thesis reveals the expression of a large GST repertoire and a functional mercapturic acid pathway in zebrafish early life stages and PAC2 cells. The presence of this important chemical activa-tion/deactivation and clearance route supports the application of these test systems as alternative models in toxicology and chemical hazard assessment.