As there are conceivably more pharmaceuticals than the 5 included in targeted monitoring that enter into the environment, it is important to determine which other pharmaceuticals may be present, to gain a more holistic idea of the pharmaceutical loading in Luxembourgish surface waters. The presence of pharmaceuticals in the aquatic environment poses a threat to human and environmental health due to exposure to either the pharmaceuticals themselves or their metabolites and TPs, which may still possess bioactivity. These chemicals have potential negative impacts on human health and the environment through different routes of exposures. There are many approaches to account for the presence of xenobiotics in the environment, but recently increasing effort has been in the use of non-targeted analysis (NTA) and/or suspect screening using high resolution mass spectrometry (HRMS) specifically to support risk assessment efforts and regulatory institutions. HRMS enables measurement of known pollutants, discovery of contaminants of emerging concern as well as retrospective screening. However, setting up analyses, both experimentally and computationally, is no trivial matter. Despite these challenges, the information that can be obtained from such analyses has a wide breadth of utility, especially for environmental studies. NTA and suspect screening are effective techniques for the monitoring and discovery of xenobiotics in the aquatic environment. Nevertheless, the interpretation of HRMS data presents challenges that highlight the need for computational tools to enable the proper identification and annotation of the chemical components in environmental matrices. MetFrag () is an open source tool for compound identification, including in silico fragmentation, mass spectral matching, and metadata functions. 26 is an open chemistry database at the National Institutes of Health (NIH) containing more than 110M compounds. While such a large database provides access to many chemicals, it can lead to (tens of) thousands of candidates per unknown when performing non-target screening of hundreds of masses. For this work, an early version of PubChemLite was used, which contains ~300,000 compounds selected to be highly relevant for environmental investigations based on annotation content, including information relevant for pharmaceuticals. PubChemLite has been shown to outperform other databases such as the whole of PubChem and CompTox for well-known chemicals and delivers important metadata that can be used during identification with MetFrag. PubChem and PubChemLite also contain information on environmental TPs contributed via the NORMAN Suspect List Exchange (). This information can be exploited programmatically during the environmental screening of hundreds of compounds, together with their transformation products.