http://informahealthcare.com/nan ISSN: 1743-5390 (print), 1743-5404 (electronic) Nanotoxicology, Early Online: 1–12 ! 2013 Informa Healthcare USA, Inc. DOI: 10.3109/17435390.2013.848949 ORIGINAL ARTICLE Temperature-dependent toxicities of nano zinc oxide to marine diatom, amphipod and fish in relation to its aggregation size and ion dissolution Stella W. Y. Wong and Kenneth M. Y. Leung The Swire Institute of Marine Science and School of Biological Sciences, The University of Hong Kong, Pokfulam, Hong Kong, China Abstract This study, for the first time, concurrently investigated the influence of seawater temperature, exposure concentration and time on the aggregation size and ion dissolution of nano zinc oxides (nZnO) in seawater, and the interacting effect of temperature and waterborne exposure of nZnO to the marine diatom Skeletonema costatum, amphipod Melita longidactyla and fish Oryzias melastigma, respectively. Our results showed that aggregate size was jointly affected by seawater temperature, nZnO concentration and exposure time. Among the three factors, the concentration of nZnO was the most important and followed by exposure time, whereas temperature was less important as reflected by their F values in the three-way analysis of variance (concentration: F 3, 300 ¼ 247.305; time: F 2, 300 ¼ 20.923 and temperature: F 4, 300 ¼ 4.107; All p values 5 0.001). The aggregate size generally increased with increasing nZnO concentration and exposure time. The release of Zn ions from nZnO was significantly influenced by seawater temperature and exposure time; the ion dissolution rate generally increased with decreasing temperature and increasing exposure time. Growth inhibition of diatoms increased with increasing temperature, while temperature and nZnO had an interactional effect on their photosynthesis. For the amphipod, mortality was positively correlated with temperature. Fish larvae growth rate was only affected by temperature but not nZnO, while the two factors interactively modulated the expression of heat shock and metallothionein proteins. Evidently, temperature can influence aggregate size and ion dissolution and thus toxicity of nZnO to the marine organisms in a species-specific manner. Keywords Chlorophyll a fluorescence, heat shock protein, metallothionein, nanomaterial, toxicity History Received 15 November 2012 Revised 16 September 2013 Accepted 23 September 2013 Published online 13 November 2013 Introduction The US Environmental Protection Agency has recently published a report on the evaluation of vulnerability of aquatic ecosystems exposed to multiple stressors, in particular, interactions of global climate change with anthropogenic influences (USEPA, 2011). The potential releases of engineered nanomaterials (NMs) into the aquatic environment have received much concern from govern- ments and scientists, and their ultimate behavior and fate may be determined by various physicochemical conditions including temperature, pH and salinity. Several in vitro projects concerning the cellular uptake of NMs at different temperatures have been undertaken over the recent decade (Arsianti et al., 2010; Bejjani et al., 2005). However, as the purpose of most cellular researches was to evaluate the role of NMs as drug delivery carriers, the temperatures used for comparison were normally 4  C (where energy-dependent endocytosis ceases almost completely and cell membrane becomes too rigid for passive diffusion) and 37  C (normothermia) (Arsianti et al., 2010). Moreover, Rispoli et al. (2010) reported that higher temperatures led to decreased aggregate size of nano zero-valent copper in nutrient broth medium, and thus aggravated its toxicity toward Escherichia coli. On the other hand, in vivo experiments with aquatic plants and animals are relatively rare, with only two documented in vivo studies. Tao et al. (2009) observed that mortality and development of the freshwater flea Daphnia magna exposed to 0.2 mg L 1 of fullerene (C 60 ) was temperature-independent. Oukarroum et al. (2012) also observed that temperature had an influence on the aggregate formation and zeta potential of nano silver, which subsequently affected their inhibitory effect on the freshwater microalgae Chlorella vulgaris and Dunaliella tertiolecta. Our laboratory has previously documented a study comparing the toxicity of nano zinc oxide (nZnO), bulk ZnO and Zn ion to five marine organisms at room temperature (25  C; Wong et al., 2010); while nanoparticulate effects were observed, the toxicity of nZnO to the marine organisms was mainly attributed to dissolved Zn ions. This study, for the first time, was designed to concurrently examine the influence of temperature on the ion dissolution and size aggregation of nZnO in seawater and the interacting effect of temperature and waterborne exposure of nZnO to three selected marine organisms, namely, the diatom Skeletonema costatum, amphipod Melita longidactyla and fish Oryzias melastigma. These species represent the primary produ- cer, primary consumer and secondary consumer, respectively. Our experimental results showed that temperature can signifi- cantly influence aggregate size and ion dissolution and thus toxicity of nZnO, although the interacting effect of temperature and nZnO is also highly species dependent. Correspondence: Professor Kenneth M. Y. Leung, School of Biological Sciences, The University of Hong Kong, Pokfulam, Hong Kong, China. Tel: +852 22990607. Fax: +852 25176082. E-mail: kmyleung@hku.hk Nanotoxicology Downloaded from informahealthcare.com by University of Hong Kong Libraries on 11/13/13 For personal use only.