High-Throughput Effect-Directed Analysis Using Downscaled in Vitro Reporter Gene Assays to Identify Endocrine Disruptors in Surface Water

Nick Zwart*, Shan Li Nio, Corine J. Houtman, Jacob De Boer, Jeroen Kool, Timo Hamers, Marja H. Lamoree

*Corresponding author for this work

Research output: Contribution to JournalArticleAcademicpeer-review


Effect-directed analysis (EDA) is a commonly used approach for effect-based identification of endocrine disruptive chemicals in complex (environmental) mixtures. However, for routine toxicity assessment of, for example, water samples, current EDA approaches are considered time-consuming and laborious. We achieved faster EDA and identification by downscaling of sensitive cell-based hormone reporter gene assays and increasing fractionation resolution to allow testing of smaller fractions with reduced complexity. The high-resolution EDA approach is demonstrated by analysis of four environmental passive sampler extracts. Downscaling of the assays to a 384-well format allowed analysis of 64 fractions in triplicate (or 192 fractions without technical replicates) without affecting sensitivity compared to the standard 96-well format. Through a parallel exposure method, agonistic and antagonistic androgen and estrogen receptor activity could be measured in a single experiment following a single fractionation. From 16 selected candidate compounds, identified through nontargeted analysis, 13 could be confirmed chemically and 10 were found to be biologically active, of which the most potent nonsteroidal estrogens were identified as oxybenzone and piperine. The increased fractionation resolution and the higher throughput that downscaling provides allow for future application in routine high-resolution screening of large numbers of samples in order to accelerate identification of (emerging) endocrine disruptors.

Original languageEnglish
Pages (from-to)4367-4377
Number of pages11
JournalEnvironmental Science and Technology
Issue number7
Early online date16 Mar 2018
Publication statusPublished - 3 Apr 2018


The described research was funded by the Dutch Technology Foundation (STW), Project Number 12396. The authors acknowledge Dr. Michael Denison for providing the VM7Luc4E2 and H4IIE DR-Luc cell lines, Dr. Iida Mitsuru and Dr. Hiroyuki Kojima for providing the AR-EcoScreen, and the TIPTOP project funded by CEFIC-LRI (Eco023) for providing passive sampler extracts.

FundersFunder number
Stichting voor de Technische Wetenschappen12396


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