Lübeck et al. Environ Sci Eur (2020) 32:78 https://doi.org/10.1186/s12302-020-00353-2 RESEARCH Open Access GC GC–HRMS nontarget fngerprinting of organic× micropollutants in urban freshwater sediments Josephine S. Lübeck* , Guilherme L. Alexandrino and Jan H. Christensen Abstract Background: Sediments are sinks for organic micropollutants, which are traditionally analysed by gas chromatogra- phy–mass spectrometry (GC–MS). Although GC–MS and GC–tandem MS (MS/MS) are preferred for target screening, they provide only limited chromatographic resolution for nontarget screening. In this study, a comprehensive two- dimensional GC–high-resolution MS method (GC GC–HRMS) was developed for nontarget screening and source identifcation of organic micropollutants in sediments× from an urban channel and adjacent lake in Copenhagen, Denmark. The GC GC–HRMS data were processed by pixel-based chemometric analysis using baseline subtraction, alignment, normalisation,× and scaling before principal component analysis (PCA) of the pre-processed GC GC– HRMS base peak ion chromatograms (BPCs). The analysis was performed to identify organic micropollutants× of high abundance and relevance in the urban sediments and to identify pollution sources. Tentative identifcations were based on match factors and retention indices and tagged according to the level of identifcation confdence. Results: The channel contained both a signifcantly higher abundance of micropollutants and a higher diversity of compounds compared to the lake. The PCA models were able to isolate distinct sources of chemicals such as a natural input (viz., a high relative abundance of mono-, di- and sesquiterpenes) and a weathered oil fngerprint (viz., alkanes, naphthenes and alkylated polycyclic aromatic hydrocarbons). A dilution efect of the weathered oil fngerprint was observed in lake samples that were close to the channel. Several benzothiazole-like structures were identifed in lake samples close to a high-trafc road which could indicate a signifcant input from asphalt or tire wear particles. In total, 104 compounds and compound groups were identifed. Conclusions: Several chemical fngerprints of diferent sources were described in urban freshwater sediments in Copenhagen using a pixel-based chemometric approach of GC GC–HRMS BPCs. Various micropollutants of anthro- pogenic origin were identifed. Tailored pre-processing and careful× interpretation of the identifcation results is inevita- ble and still requires further research for an automated workfow. Keywords: Chemical fngerprint, Sediment analysis, Source identifcation, GC GC–HRMS, Pixel-based analysis, Prioritisation × Background Anthropogenic pollution of freshwater ecosystems via agricultural, industrial and urban activities is ubiquitous. Te World Water Assessment Programme estimated in 2017 that approximately 80% of all wastewater, globally, *Correspondence: [email protected] is discharged into the environment without treatment Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, Thorvaldsensvej 40, 1871 Frederiksberg, [1]. However, environmental awareness is increasing and Denmark thus, the desire for a better understanding of what types © The Author(s) 2020. This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creat iveco mmons .org/licen ses/by/4.0/. Lübeck et al. Environ Sci Eur (2020) 32:78 Page 2 of 16 of pollutants are ending up in our environment. Anthro- NTS have been described in the literature, e.g., by inten- pogenic micropollutants comprise numerous chemicals sity, specifc isotopic patterns or as part of a homologous and their degradation products such as pharmaceuticals, series [6, 16], but few methods have been suggested for detergents, pesticides and chemicals from consumer sample prioritisation. Even though chemometric tools, care products [2, 3]. Tese can be introduced to fresh- such as principal component analysis (PCA) [6] or hier- water ecosystems via, e.g., household or industrial waste archical clustering analysis [17], have already been incor- efuents; littering, road runof and car exhaust. Com- porated in NTS workfows to prioritise specifc chemical pounds with low water solubility and high octanol/water profles, applications in environmental monitoring are partition coefcient (log K OW) mostly deposit in sedi- still limited, particularly for GC × GC–(HR)MS. How- ments and are only released slowly to the water where ever, some benefts of chemometrics in NTS are appar- they could cause adverse efects to the local fauna. Te ent, e.g., the extraction of the most relevant chemical European Parliament and Council identifed 45 prior- patterns or diferent sources of pollution can be visual- ity substances in 2013 that every member state ought to ised from the entire chemical fngerprint of the samples. monitor in surface waters at least once a year [4]. Te list In the so-called pixel-based approach, the data analy- includes several heavy metals, pesticides, halogenated sis is performed directly on the chromatographic pixels compounds, polycyclic aromatic hydrocarbons (PAHs) [18]. Terefore, the main variance in the data is displayed and phenols. directly on the chromatographic space without prior Gas chromatography (GC) coupled with mass spec- peak-picking. Te pixel-based approach facilitates the trometry (MS) is the conventional solution for target interpretation of structured chromatograms commonly analysis of volatile and semi-volatile persistent organic found in GC × GC–HRMS data, such as homologous pollutants (POPs) in the environment [5]. Te use of series. While hierarchical clustering analysis is useful extracted ion chromatograms (EICs) facilitates the for assessing the overall chemical similarity among sam- identifcation of known compounds; however, moni- ples, the role of the individual variables, in this case, the toring unknown chemicals or chemicals of emerging chromatographic pixels that explain a similarity profle concern (CECs) that have previously not been included in the dataset cannot be evaluated. For example, a study on any regulatory list, is a more challenging task. For a by Alexandrino et al. successfully implemented the pixel- more comprehensive chemical impact assessment of our based analysis for forensic investigations of diesel spills in environment, suspect screening and nontarget screen- the environment based on GC × GC–HRMS data [19]. ing (NTS) are used in combination with target analyses Tis study aimed to characterise freshwater lake and [2, 6]. Te identifcation of unknown compounds in an channel sediments in an urban area (Copenhagen, NTS workfow is usually made by comparing experi- Denmark) based on an NTS analytical workfow using mental mass spectra with MS-libraries such as NIST for GC × GC–HRMS. First of all, the overall chemical vari- GC–MS with electron ionisation [7]. An adequate chro- ation between and within two sampling sites was inves- matographic resolution that provides mass spectra of tigated with pixel-based PCA. Second, a PCA for each the unidentifed peaks free of chemical interferences is sampling site intended to give a refned insight into the key to increase the reliability of the spectral matching. sources of pollution and chemical fngerprints. Subse- Terefore, GC–MS may not provide sufcient resolu- quently, distinct samples were prioritised based on the tion for NTS of complex environmental matrices such as pixel-based PCA for tentative compound identifcation. sediments. Comprehensive two-dimensional GC with either low or Materials and methods Sampling high-resolution mass spectrometry detection (GC × GC– (HR)MS) provides higher peak capacities than those Figure 1 shows the two sampling sites from the sampling obtained by one-dimensional GC both for target analy- campaign in September to November 2017: the lake sis of, e.g., PAHs, PAH derivatives and organochlorine Utterslev Mose (UTM) and the adjacent fortress chan- pesticides [8]; and for NTS where more unambiguous nel (FSK as in Fæstningskanalen) in Copenhagen, Den- identifcation of POPs has been obtained based on, e.g., mark. Te lake is part of a protected nature park in the high-resolution neutral loss of halogens, isotopic patterns western part of Copenhagen and is, among others, fed by and mass defect calculations [9, 10]. Additional applica- the fortress channel which surrounds the Danish capi- tions of GC × GC–(HR)MS for environmental purposes tal. Sampling was performed with a Kajak sediment core have also been reported [3, 11–14]. Data mining in NTS sampler (KC Denmark A/S) with acrylic sample tubes is still labour-intense, and prioritisation and identifcation (length: 47.5 cm, i.d.: 0.46 cm, wall thickness: 0.4 cm). of thousands of peaks can be challenging, especially with Sediment samples (0–30 cm) were obtained