Presence of antineoplastic compounds in the marine environment could cause adverse effects even at very low concentrations. The bulk of the produced literature at present has focused on the detection of these compounds in wastewater samples, while only a few papers tackled the more complex solid samples and until now, to the best of our knowledge, none has investigated marine organisms. This work presents the development, optimisation and application of an analytical method for the extraction and determination of the four antineoplastic compounds; cyclophosphamide, vincristine, vinblastine and tamoxifen, from fish tissues by microwave assisted extraction and ultra-high-performance liquid chromatography coupled to tandem mass spectrometry. Under optimal conditions, recoveries ranged between 60% and 120%, while infra- and inter-day precision were in the 5.0-19.4% and 12.0-20.4% ranges, respectively. Method detection and quantification limits were in the ranges 0.8-1.3 and 2.7-4.3 ng.g(-1) dry weight for tamoxifen and cyclophosphamide, and 46.3-536 and 154-1787 ng.g(-1) dry weight for vinblastine and vincristine, respectively. The developed analytical method was successfully applied for the extraction and determination of target compounds from muscle and liver tissues belonging to three different fish species. The results did not highlight any contamination by target antineoplastic molecules in the investigated fish tissues.