The determination of trace pollutants in seawater is challenging, and sampling is a crucial step in the entire analytical process. Passive samplers combine in situ sampling and preconcentration, thus limiting the tedious treatment steps of the conventional sampling methods. Their use to monitor water quality in confined marine environment could bring several advantages. In this work, the presence of organic contaminants at trace and ultra-trace levels was assessed in the Genoa Aquarium supply-and-treated water using Polar Organic Integrative Samplers (POCIS). Both untargeted gas chromatography-mass spectrometry and targeted liquid chromatography-tandem mass spectrometry were employed. The untargeted approach showed the presence of hydrocarbons, diphenyl sulfone and 2,4-di-tert-butyl-phenol. Only hydrocarbons were detected in all the samples. Nineteen emerging contaminants, belonging to different classes (pharmaceuticals, UV-filters, hormones and perfluorinated compounds), were selected for the target analysis. Thirteen analytes were detected, mainly in supply water, even though the majority of them were below the quantitation limit. It is worthy to note that two of the detected UV-filters had never been reported in seawater using the POCIS samplers. The comparison of the analytes detected in supply and treated water indicated a good performance of the Aquarium water treatment system in the abatement of seawater contaminants.