Contents lists available at ScienceDirect Ecotoxicology and Environmental Safety journal homepage: www.elsevier.com/locate/ecoenv The evaluation of the potential ecotoxicity of pyroligneous acid obtained from fast pyrolysis Gabriel Goetten de Lima a,b,1 , Camila Mendes c,1 , Gustavo de Marchi c , Taynah Vicari c , Marta Margarete Cestari c , Monike F. Gomes d , Wanessa Algarte Ramsdorf d , Washington Luiz Esteves Magalhães e , Fabricio Augusto Hansel e , Daniela Morais Leme c,* a Graduate Program in Engineering and Science of Materials – PIPE, Federal University of Paraná - UFPR, 81.531-990 Curitiba PR, Brazil b Materials Research Institute, Athlone Institute of Technology, Athlone, Ireland c Department of Genetics, Federal University of Paraná, Curitiba, PR, Brazil d Laboratory of Ecotoxicology, Federal University of Technology – Paraná, Curitiba, PR, Brazil e Embrapa Forestry – Brazilian Agricultural Research Agency, Colombo, PR, Brazil ARTICLE INFO Keywords: Pyroligneous acid Acute toxicity DNA damages Allium cepa Daphnia magna RTG-2 fish cell line ABSTRACT Pyroligneous acid (PA) is a by-product of bio-oil, which is obtained by pyrolysis of the wood. This product has been tested for use in several areas, such as agriculture, as a promising green herbicide; however, there are few scientific data regarding its environmental impacts. For this study, an ecotoxicity testing battery, composed of Daphnia magna acute toxicity test, Allium cepa test and in vitro Comet assay with the rainbow trout gonad-2 cell fish line (RTG-2) were used to evaluate the acute toxicity and genotoxicity of PA obtained from fast pyrolysis of eucalyptus wood fines. The PA presented acute toxicity to D. magna (microcrustacea) with EC 50 of 26.12 mg/L, and inhibited the seed germination (EC 50 5.556 g/L) and root development (EC 50 3.436 g/L) of A. cepa (higher plant). No signs of genotoxicity (chromosomal aberrations and micronuclei in A. cepa and primary DNA lesions in RTG-2 cells) were detected to this product. The acute toxicity and absence of genotoxicity may relate to the molecules found in the PA, being the phenolic fraction the key chemical candidate responsible for the toxicity observed. In addition, daphnids seem to be more sensitivity to the toxicity of PA than higher plants based on their EC 50 values. This first ecotoxicological evaluation of PA from fast pyrolysis pointed out the need of de- termining environmental exposure limits to promote the safer agriculture use of this product, avoiding impacts to living organisms. 1. Introduction Bio-oil is a product that can be derived from pyrolysis of wood (Meier and Faix, 1999; Mohan et al., 2006; Roberts, 1970), which consists in decomposition of the biomass by heat under air controlled environment (Mohan et al., 2007). The resultant process is formed by vapor condensation. Bio-oil has many unique characteristics that make this product valuable in a number of applications, such as crop pro- tection agent (Shihadeh and Hochgreb, 2000). For the production of bio-oil, two main methods are employed (slow and fast pyrolysis) and they differ in the percentage of gas, char and liquid products obtained (Grewal et al., 2018). Slow pyrolysis consists of slow heating rates and yields equal quantities of gas, char and liquid while is heated at temperatures of 300 °C. Contrary, fast pyrolysis, which consists of high heating rates, usually yields larger quantity of liquid phase (60–75% of liquid bio-oil) heating at temperatures of 500 °C. The derived product via pyrolysis can be separated via dis- tillation of the condensed liquid (Souza et al., 2012) and, although research have been focusing on the energy combustion of this product (pyroligneous tar) (Bridgwater, 2003; Honnery et al., 2008), the aqu- eous part (pyroligneous acid - PA) is used in agriculture. The promising agriculture usage of PA relates to their antimicrobial, antioxidant and pesticidal activities; however, this product has not yet been properly investigated towards its safety to environmental organisms (e.g., non- target plants and aquatic life) (Kadota et al., 2002; Ma et al., 2013; Mathew and Zakaria, 2015; Mmojieje, 2016; Murayama et al., 1995; Wei et al., 2010). The two main routes to obtain PA, have its own attractiveness https://doi.org/10.1016/j.ecoenv.2019.05.058 Received 31 January 2019; Received in revised form 16 May 2019; Accepted 17 May 2019 * Corresponding author. Federal University of Paraná, Av. Cel. Francisco H. dos Santos, 81531-980, Curitiba, PR, Brazil. E-mail addresses: danielamoraisleme@gmail.com, daniela.leme@ufpr.br (D.M. Leme). 1 These authors contributed equally to this manuscript. Ecotoxicology and Environmental Safety 180 (2019) 616–623 0147-6513/ © 2019 Published by Elsevier Inc. T