Elsevier

Environment International

Volume 99, February 2017, Pages 120-130
Environment International

Review article
Analysis of the sensitivity of in vitro bioassays for androgenic, progestagenic, glucocorticoid, thyroid and estrogenic activity: Suitability for drinking and environmental waters

https://doi.org/10.1016/j.envint.2016.12.014Get rights and content

Highlights

  • Reviewed suitability of in vitro assays to detect endocrine activity in water

  • Included androgenic, progestagenic, glucocorticoid, thyroid and estrogenic activity

  • Some assays are sensitive enough to detect activity in most environmental waters.

  • Others appear not sensitive enough to detect activity even in polluted samples.

  • Findings provide guidance on bioassay selection and required sample enrichment.

Abstract

The presence of endocrine disrupting chemicals in the aquatic environment poses a risk for ecosystem health. Consequently there is a need for sensitive tools, such as in vitro bioassays, to monitor endocrine activity in environmental waters. The aim of the current study was to assess whether current in vitro bioassays are suitable to detect endocrine activity in a range of water types. The reviewed assays included androgenic (n = 11), progestagenic (n = 6), glucocorticoid (n = 5), thyroid (n = 5) and estrogenic (n = 8) activity in both agonist and antagonist mode. Existing in vitro bioassay data were re-evaluated to determine assay sensitivity, with the calculated method detection limit compared with measured hormonal activity in treated wastewater, surface water and drinking water to quantify whether the studied assays were sufficiently sensitive for environmental samples. With typical sample enrichment, current in vitro bioassays are sufficiently sensitive to detect androgenic activity in treated wastewater and surface water, with anti-androgenic activity able to be detected in most environmental waters. Similarly, with sufficient enrichment, the studied mammalian assays are able to detect estrogenic activity even in drinking water samples. Fewer studies have focused on progestagenic and glucocorticoid activity, but some of the reviewed bioassays are suitable for detecting activity in treated wastewater and surface water. Even less is known about (anti)thyroid activity, but the available data suggests that the more sensitive reviewed bioassays are still unlikely to detect this type of activity in environmental waters. The findings of this review can help provide guidance on in vitro bioassay selection and required sample enrichment for optimised detection of endocrine activity in environmental waters.

Introduction

There is increasing concern about the presence of endocrine disrupting chemicals in the environment due to their potential effects on both human and ecosystem health (Bergman et al., 2013). Environmental waters in particular can be impacted by endocrine disrupting chemicals, with both point sources, such as wastewater effluent, and diffuse sources, such as agriculture and industry, contributing to the chemical load (Vethaak et al., 2005). In addition to natural and synthetic hormones, a wide range of environmental chemicals, including industrial compounds, pesticides and UV filters, have been identified as known or suspected endocrine disrupting chemicals (Bergman et al., 2012). As a result of the wide range of potential endocrine disrupting chemicals and the fact that they will be present in water as a complex mixture of contaminants, chemical analysis alone is insufficient to monitor endocrine disrupting chemicals. Instead, in vitro bioassays indicative of hormonal activity, including androgenic, progestagenic, glucocorticoid, thyroid and estrogenic activity, can be applied to assess endocrine activity in environmental waters. Such assays have been applied widely to wastewater effluent and surface water (e.g. Bain et al., 2014, Schiliro et al., 2012, Scott et al., 2014, Thomas et al., 2002), with less focus on cleaner water sources, such as advanced treated or drinking water (e.g. Brand et al., 2013, Conley et al., 2016). Much of the literature concentrates on the detection of agonistic activity in water, but some environmental contaminants can act as antagonists, which, if present in a sample, can reduce the agonistic response in vitro (Ihara et al., 2014), emphasising the importance of evaluating both agonism and antagonism in environmental samples.

As many endocrine disrupting chemicals are present in the aquatic environment at low concentrations, there is a need for sensitive methods to detect both endocrine agonists and antagonists at environmentally relevant levels. In addition to the inherent sensitivity of the assay, factors such as sample enrichment, typically by solid phase extraction (SPE), and sample dilution in the assay, which is dependent on assay solvent tolerance, will affect the overall assay sensitivity. The aim of the current study was to review the sensitivity of a range of specific hormonal activity in vitro bioassays to determine whether they are suitable to detect endocrine activity in environmental water samples. The reviewed bioassays focus on androgenic, progestagenic, glucocorticoid, thyroid and estrogenic activity in both agonist and antagonist mode and include a range of assay types, such as receptor binding assays, yeast reporter gene assays, mammalian reporter gene assays and cell proliferation assays. Between 5 and 11 assays were selected for each endpoint, with all assays identified as in at least moderate use for water quality testing (Global Water Research Coalition, 2006, Global Water Research Coalition, 2012). A literature search in ScienceDirect (31st March 2016) was conducted to determine the number of studies that mentioned each assay specifically (Fig. 1). While this approach may not capture all publications, it does reveal that the different endpoints have received different levels of attention, with the majority of work focusing on estrogenic activity, followed by androgenic activity. In contrast, progestagenic, glucocorticoid and thyroid activity have received much less attention. However, these hormonal systems still play an important role for maintenance of sexual development, growth and homeostasis, thus there is still a need for sensitive methods to detect these less studied endpoints in environmental waters.

Section snippets

Experimental approach

The reviewed assays included 11 (anti)androgenic assays, 6 (anti)progestagenic assays, 5 (anti) glucocorticoid assays, 5 (anti)thyroid assays and 8 (anti)estrogenic assays. Assay sensitivity was determined by re-evaluating existing reference compound concentration-effect curves from the literature in order to determine the concentration causing 10% effect (EC10) for each assay. The EC10 was selected as the assay limit of detection based on Escher et al. (2014). Ideally, an assay reference

Agonist mode

Eleven in vitro assays for androgenic activity covering different assays types, including receptor binding assays, yeast reporter gene assays, mammalian reporter gene assays and cell proliferation assays, were considered. Dihydrotestosterone (DHT) was used as the reference compound in agonist mode. The re-analysed concentration-effect curves are shown in the Supplementary Material in Fig. S1, while the assay sensitivity parameters including reported concentration causing 50% effect (EC50),

Conclusions

Through analysis of the available literature, we can conclude that based on assay sensitivity and typical enrichment factors (1000 ×), current in vitro bioassays are sufficiently sensitive to detect androgenic activity in treated wastewater and surface water, anti-androgenic activity in most water samples, some progestagenic and glucocorticoid activity in most treated wastewater and surface water and estrogenic activity in most samples. Understanding (anti)thyroid activity in environmental

Acknowledgements

This project (Global Water Research Coalition project # 2013-05) was funded by PUB (Singapore), the Foundation for Applied Water Research (STOWA), Water Research Australia, the Water Technology Center (TZW), the Water Research Foundation (WRF) and the Global Water Research Coalition (GWRC).

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