In Vitro Cytotoxicity and Adaptive Stress Responses to Selected Haloacetic Acid and Halobenzoquinone Water Disinfection Byproducts

Abstract

The process of disinfecting drinking water inadvertently leads to the formation of numerous disinfection byproducts (DBPs). Some of these are mutagenic, genotoxic, teratogenic, and cytotoxic, as well as potentially carcinogenic both in vivo and in vitro. We investigated the in vitro biological activity of five DBPs: three monohaloacetic acids (monoHAAs) [chloroacetic acid (CAA), bromoacetic acid (BAA), and iodoacetic acid (IAA)] and two novel halobenzoquinones (HBQs) [2,6-dichloro-p-benzoquinone (DCBQ) and 2,6-dibromo-p-benzoquinone]. We focused particularly on cytotoxicity and induction of two adaptive stress response pathways: the oxidative stress responsive Nrf2/ARE and DNA-damage responsive p53 pathways. All five DBPs were cytotoxic to the Caco-2 cell line after a 4 h exposure, and all DBPs induced both of the adaptive stress response pathways, Nrf2/ARE and p53, in the micromolar range, as measured by two β-lactamase-based reporter gene assays. The decreasing order of potency for all three endpoints for the five DBPs was IAA ∼ BAA > DCBQ ∼ DBBQ > CAA. Induction of oxidative stress was previously proposed to be the molecular initiating event (MIE) for both classes of DBPs. However, comparing the levels of activation of the two pathways uncovered that the Nrf2/ARE pathway was the more sensitive endpoint for HAAs, whereas the p53 pathway was more sensitive in the case of HBQs. Therefore, the DNA damage-responsive p53 pathway may be an important piece of information to fill in a gap in the adverse outcome pathway framework for the assessment of HBQs. Finally, we cautiously compared the potential risk of the two novel HBQs using a benchmarking approach to that of the well-studied CAA, which suggested that their relative risk may be lower than that of BAA and IAA.

Bioanalytical assessment of adaptive stress responses in drinking water: A predictive tool to differentiate between micropollutants and disinfection by-products

“…Electrophilic chemicals and chemicals that produce reactive oxygen species can induce the oxidative stress response as the presence of these chemicals releases transcription factor Nrf2 from negative regulator Keap1, which in turn activates the antioxidant response element (Zhang 2006). Environmental water samples (Escher et al 2012), micropollutants (Martin et al 2010, Escher et al 2013 and DBPs (Procházka et al 2015, Stalter et al 2016a) have been shown to activate the oxidative stress response, with 23% of analysed chemicals in the US EPA ToxCast database reported to induce oxidative stress (US EPA 2015). The p53 transcription factor responds to DNA damage and will initiate repair proteins, alter the cell cycle or induce apoptosis (Knight et al 2009).…”

“…The p53 transcription factor responds to DNA damage and will initiate repair proteins, alter the cell cycle or induce apoptosis (Knight et al 2009). The p53 response assay can detect genotoxic compounds and has previously been applied to individual DBPs (Procházka et al 2015, Stalter et al 2016a) and water samples (Yeh et al 2014, Neale et al 2015a. NF-κB is an important transcription factor associated…”

“…for effect-based trigger values to differentiate between what is an acceptable or unacceptable response(Escher et al 2015). There have been a number of different approaches proposed in the literature to derive effect-based trigger values.…”

“…There have been a number of different approaches proposed in the literature to derive effect-based trigger values. For example,Brand et al (2013) proposed trigger values based on the acceptable daily intake of the assay reference compound using equivalent concentrations and accounting for some in vitro to in vivo toxicokinetic differences Escher et al (2015). used an alternative approach by reading across from existing drinking water guidelines to derive bioanalytical trigger values.…”

Maternal Swimming Pool Exposure during Pregnancy in Relation to Birth Outcomes and Cord Blood DNA Methylation among Private Well Users

“…There are growing concerns regarding potential epigenetic, cytotoxic and genotoxic effects of compounds formed by consumer products such as halobenzoquinones . These compounds, which are not typical found in drinking water, may affect neural stem cells (Fu et al, 2017;Procházka et al, 2015;. In addition, natural humic substances as well as consumer products may generate DBPs with endocrine disrupting activities (Itoh et al, 2009).…”