Copper and Zinc Tolerance of TwoTropical Microalgae After Copper Acclimation Hilary L. Johnson, 1,2 Jenny L. Stauber, 1 Merrin S. Adams, 1 Dianne F. Jolley 2 1 Centre for Environmental Contaminants Research, CSIRO Land and Water, Private Mail Bag 7, Bangor, New South Wales 2234, Australia 2 GeoQuEST, Department of Chemistry, University of Wollongong, New South Wales 2522, Australia Received 13 June 2006; revised 24 January 2007; accepted 28 January 2007 ABSTRACT: Current toxicity tests with microalgae are often criticized as being overly sensitive to metals because algae are cultured in metal-deficient media. If such bioassays overestimate copper toxicity in surface waters, the relevance of water quality guidelines derived from these tests is questionable. In this study, the effect of acclimation to copper at environmentally relevant concentrations, on the sensitivity of the marine diatom Nitzschia closterium and the freshwater green alga Chlorella sp. to copper and zinc was examined. N. closterium was acclimated in culture medium containing 5 or 25 g Cu L 1 for 200 days, while Chlorella sp. was acclimated in medium containing 2 g Cu L 1 for 100 days. Changes in algal growth rates and copper and zinc tolerance were monitored using standard growth inhibition toxicity tests in minimal medium over 72 h. Neither of the two acclimated N. closterium cultures had increased zinc or copper tolerance compared with that of the nonacclimated algae, nor were there any changes in control growth rates. Similarly, no changes in copper tolerance or control growth rates were observed for the acclimated Chlorella sp. culture. This was supported by measurements of intracellular and extracellular copper which confirmed that there were no differences in copper accumulation in either acclimated or nonacclimated algae. These results suggest that these algae grown in standard culture media are gener- ally no more sensitive than algae grown in a metal-enriched medium. This supports the continued use of current laboratory bioassays with microalgae, as part of a suite of tests for assessing metal bioavailability, for use in ecological risk assessments and for providing data for the derivation of water quality guidelines. # 2007 Wiley Periodicals, Inc. Environ Toxicol 22: 234–244, 2007. Keywords: copper; zinc; algae; tolerance; acclimation; growth rate INTRODUCTION Algal bioassays are currently used to assess the impacts of contaminants on aquatic ecosystems as well as to assist in the development of water quality guidelines. Chronic algal toxicity tests typically measure the decrease in growth rate or cell biomass after a 72-h exposure to the contaminant. Algae have been found to be particularly sensitive to metals due to their high surface-to-volume ratio and the variety of membrane metal-ion binding sites, which differ in both affinity and specificity (Megharaj et al., 2003). Both copper and zinc are essential elements required for the normal functioning of enzyme systems within algae. However, both metals are toxic when algae are exposed to concentrations exceeding those required for optimal growth. Both metals disrupt photosynthesis, respiration, ATP production, and pigment synthesis, as well as inhibit cell division (De Filippis et al., 1981; Sunda and Huntsman, 1983; Stauber and Florence, 1987, 1990; Cid et al., 1996). Both biotic and abiotic factors affect the sensitivity of algae to copper and zinc (Stauber and Davies, 2000). Algal responses to metals depend on the particular species (with Correspondence to: J. L. Stauber; e-mail: jenny.stauber@csiro.au Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/tox.20265  C 2007 Wiley Periodicals, Inc. 234