RESEARCH PAPER Evaluation of the toxicity of ZnO nanoparticles to Chlorella vulgaris by use of the chiral perturbation approach Hui Zhou & Xiaojun Wang & Ying Zhou & Hongzhou Yao & Farooq Ahmad Received: 30 September 2013 /Revised: 13 March 2014 /Accepted: 17 March 2014 # Springer-Verlag Berlin Heidelberg 2014 Abstract The toxicity of ZnO nanoparticles (NPs) has been widely investigated because of their extensive use in consumer products. The mechanism of the toxicity of ZnO NPs to algae is unclear, however, and it is difficult to differentiate between particle-induced toxicity and the effect of dissolved Zn 2+ . In the work discussed in this paper we investigated particle- induced toxicity and the effects of dissolved Zn 2+ by using the chiral perturbation approach with dichlorprop (DCPP) as chiral perturbation factor. The results indicated that intracellular zinc is important in the toxicity of ZnO NPs, and that ZnO NPs cause oxidative damage. According to dose–response curves for DCPP and the combination of ZnO NPs with (R)-DCPP or (S)-DCPP, the toxicity of DCPP was too low to perturb the toxicity of ZnO NPs, so DCPP was suitable for use as chiral perturbation factor. The different glutathione (GSH) content of algal cells exposed to (R)-DCPP or (S)-DCPP correlated well with different production of reactive oxygen species (ROS) after exposure to the two enantiomers. Treatment of algae with ZnO NPs and (R)-DCPP resulted in reduced levels of GSH and the glutathione/oxidized glutathione (GSH/GSSG) ratio in the cells compared with the control. Treatment of algae with ZnO NPs and (S)-DCPP, how- ever, resulted in no significant changes in GSH and GSH/GSSG. Moreover, trends of variation of GSH and GSH/GSSG were different when algae were treated with ZnSO 4 ·7H 2 O and the two enantiomers. Overall, the chiral perturbation approach revealed that NPs aggravat- ed generation of ROS and that released Zn 2+ and NPs both contribute to the toxicity of ZnO NPs. Keywords ZnO nanoparticles . nanotoxicity . Chiral perturbation . Reactive oxygen species . Zn 2+ . Algae Introduction Zinc oxide nanoparticles (ZnO NPs) are important nanomaterials (NMs) that have been extensively used in in- dustrial and consumer products, resulting in the increasing presence of NMs in the environment [1]. For example, it is estimated that 1,000 tons of skincare products containing ZnO NPs are produced annually for the global market [2]. Italian researchers have calculated that at least 25 % of the amount of sunscreen applied to the skin is washed off during bathing and swimming [3]. This implies that approximately 250 tons of these nanomaterials are potentially discharged into the water environment [4]. Compared with normal materials, the risks to health and the environment of the novel properties of NPs are unknown. The toxicity of ZnO NPs in aquatic ecosystems has, therefore, aroused much concern. ZnO NPs have been shown to be toxic to algae [5–8], crustaceans [9, 10], bacteria [11], and fish [12, 13]. Studies of the biotoxicity of ZnO NPs suggest several mechanisms of action. Overproduction of reactive oxygen species (ROS) is believed to be a major mechanism of the toxicity of NPs [14]. Because of the large specific surface area of NPs, which endows them with high reactivity and electron density, they can interact with biomolecules [15]. During this process, chemical reactions occur and result in increased formation of Published in the topical collection Euroanalysis XVII (The European Conference on Analytical Chemistry) with guest editor Ewa Bulska. Electronic supplementary material The online version of this article (doi:10.1007/s00216-014-7773-0) contains supplementary material, which is available to authorized users. H. Zhou : X. Wang : Y. Zhou : H. Yao : F. Ahmad College of Chemical Engineering and Materials Science, Zhejiang University of Technology, Hangzhou, China Y. Zhou (*) Research Center of Analysis and Measurement, Zhejiang University of Technology, 18 Chaowang Road, Hangzhou 310032, Zhejiang Province, China e-mail: yingzhou@zjut.edu.cn Anal Bioanal Chem DOI 10.1007/s00216-014-7773-0