Ozonation attenuates the steroidogenic disruptive effects of sediment free oil sands process water in the H295R cell line Yuhe He a , Steve B. Wiseman a, * , Xiaowei Zhang a , Markus Hecker a,b , Paul D. Jones a , Mohamed Gamal El-Din c , Jonathan W. Martin d , John P. Giesy a,e,f,g,h a Department of Veterinary Sciences and Toxicology Centre, University of Saskatchewan, Saskatoon, Saskatchewan, Canada b ENTRIX Inc. Saskatoon, Saskatchewan, Canada c Department of Civil and Environmental Engineering, University of Alberta, Edmonton, Alberta, Canada d Department of Laboratory Medicine and Pathology, Division of Analytical and Environmental Chemistry, University of Alberta, Edmonton, Alberta, Canada e Department of Biology and Chemistry, State Key Laboratory in Marine Pollution, City University of Hong Kong, Tat Chee Avenue, Kowloon, Hong Kong SAR, People’s Republic of China f Department of Zoology, Centre for Integrative Toxicology, Michigan State University, E. Lansing, MI, USA g School of Environment, Nanjing University, Nanjing, China h Key Laboratory of Marine Environmental Science, College of Oceanography and Environmental Science, Xiamen University, Xiamen, People’s Republic of China article info Article history: Received 10 December 2009 Received in revised form 16 March 2010 Accepted 6 April 2010 Available online 13 May 2010 Keywords: OSPW Steroidogenesis Estradiol Testosterone Naphthenic acids Petroleum hydrocarbons abstract There is concern regarding oil sands process water (OSPW) produced by the oil sands industry in Alberta, Canada. Little is known about the potential for OSPW, and naphthenic acids (NAs), which are the primary persistent and toxic constituents of OSPW, to affect endocrine systems. Although ozonation significantly reduces concentrations of NAs and OSPW toxicity, it was hypothesized that oxidation of OSPW might pro- duce hydroxylated products with steroidogenic activity. Therefore, untreated and ozone treated OSPW were examined for effects on sex steroid production using the H295R Steroidogenesis Assay. Untreated OSPW significantly decreased testosterone (T) and increased 17b-estradiol (E2) concentrations at OSPW dilutions greater or equal to 10-fold. This effect was mainly due to decreased E2 metabolism. Analysis of CYP19A (aromatase) mRNA abundance and enzyme activity suggested that induction of this enzyme activity may have also contributed to these effects. Reduction of parent NA concentrations by 24% or 85% decreased the effect of OSPW on E2 production. Although T production remained significantly reduced in cells exposed to ozone treated OSPW, the effect was diminished. Aromatase mRNA abundance and enzyme activity were significantly greater in cells exposed to ozone treated OSPW, however the mag- nitude was less than in cells exposed to untreated OSPW. No change of E2 metabolism was observed in cells exposed to ozone treated OSPW, which may account for recovery of E2 levels. The results indicate that OSPW exposure can decrease E2 and T production, but ozonation is an effective treatment to reduce NA concentrations in OSPW without increasing affects on steroidogenesis. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction The oil sands deposits in the Athabasca Basin, located in north- eastern Alberta, Canada, are one of the largest reserves of petro- leum in the world (Hunt, 1979), and are increasingly being developed by a rapidly growing oil sands industry. In 2000, Alber- ta’s oil sands industry produced approximately 604,700 barrels of marketable bitumen and crude oil per day (Alberta Energy and Utilities Board, 2004–2005), and production increased to an aver- age of 1,184,000 barrels per day in 2008 (Alberta Energy and Util- ities Board, 2007–2008). With the costs associated with bitumen recovery from oil sands decreasing due to technological advances, and a decline in global conventional oil supplies, development of the major reserves in the Alberta Basin, which are estimated to be at least 173.2 billion barrels, is expected to continue (Wlliams, 2003; Alberta Energy and Utilities Board, 2006–2007). In the surface mining oil sands industry, extraction of bitumen from oil sands involves the Clarke hot water extraction method. This process results in the production of large volumes of pro- cess-affected waters that contain sand, clay, unrecoverable bitu- men, hydrocarbons, and a water soluble organic acid fraction known commonly as naphthenic acids (NAs). This wastewater is commonly referred to oil sands process water (OSPW). OSPW is stored on-site in active settling basins, or tailing ponds, where the clarified fraction can eventually be recycled back into the extraction plant for further use. In accordance with a zero dis- charge policy OSPW is not intentionally released (MacKinnon, 1989; FTFC, 1995) and in 2006 it was estimated that greater than 0045-6535/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.chemosphere.2010.04.018 * Corresponding author. Address: University of Saskatchewan, Toxicology Centre, 44 Campus Drive, Saskatoon, SK, Canada, S7 N 5B3. Fax: +1 306 970 4796. E-mail address: steve.wiseman@usask.ca (S.B. Wiseman). Chemosphere 80 (2010) 578–584 Contents lists available at ScienceDirect Chemosphere journal homepage: www.elsevier.com/locate/chemosphere