The quantification and monitoring of

The quantification and monitoring of ChEs enzymatic activities have traditionally been associated to environmental studies in order to diagnose the exposure of non-target organisms to specific anti-cholinesterasic agents (Nunes, 2011). The importance of these enzymes as biomarkers of environmental contamination is a consequence of their sensitivity towards a multiplicity of environmental contaminants, being present in a large number of distinct species. In addition, the methodology by which it is possible to determine cholinesterasic activity has a relatively low cost, simple determination and good reproducibility (Nunes, 2011). These features lead to the use of cholinesterasic inhibition as an effect criterion in a large number of field studies, or to test the toxicity of several xenobiotic compounds under laboratory conditions (Nunes et al., 2004, Nunes et al., 2015, Solé et al., 2009, Oliveira et al., 2015). The potential for ChE inhibition is a common feature among several classes of environmental contaminants, namely pesticides, which were designed and chemically synthetized to attain this purpose. In fact, such ChEs inhibitors bind covalently to cholinesterases, specifically at the amino Methoxy-X04 serine, which is located at the active site of the enzyme. This inhibitory effect is ecologically relevant considering the widespread environmental contamination by anticholinesterase pesticides, namely anticholinesterasic insecticides from the organophosphates and carbamates classes. In addition, several pharmaceutical drugs were also designed to attain a similar effect, namely rivastigmine, donepezil, pyridostigmine and neostigmine (Čolović et al., 2013). Recently, evidences showed the involvement of other compounds (metals) in cholinesterasic inhibition. Cunha et al. (2007) assessed the effects of metals on cholinesterase activities of the marine gastropods Monodonta lineata and Nucella lapillus; this study showed that copper was able to cause in vitro inhibition of cholinesterase activity. The study performed by De Lima et al. (2012) showed the inhibitory effects of copper, lead, iron and cadmium, both in vivo and in vitro, on cholinesterases of the fish species Danio rerio. Despite the in vitro anticholinesterase effects observed for all metals, only lead and cadmium were able to cause an in vivo inhibition of cholinesterase activity. Nunes et al. (2014) evaluated the chronic effects of the metals lead, copper, cadmium and zinc in the European eel Anguilla anguilla; the authors did not observe alterations in cholinesterase activity. The anticholinesterase effects of metals may be strongly conditioned by the metal itself, the type of exposure and also by characteristics of the tested species. In addition, the issue of inhibition of cholinesterases by metallic compounds is still a matter of dispute, since the most commonly used determination methodology, the Ellman\'s assay, is prone to suffer already known interferences and artifacts (metals can react with the chemicals present in the reaction media and thus cause a false enzymatic activity inhibition) that may occur, especially in in vitro experiments (Frasco et al., 2005). However, and for specific metals, ChEs inhibition seems to be a real effect (Nunes, 2011). Knowing the type of cholinesterase found in each tissue is of the uttermost importance when studying the putative toxic effects of the previously mentioned contaminants on cholinesterases (Nunes, 2011). Only then the activity of these enzymes can be successfully used as biomarkers of environmental impacts. The multiplicity of cholinesterasic forms in aquatic organisms has been already demonstrated. Several studies characterizing distinct forms have already been carried out with aquatic species. Sturm et al. (1999) characterized the cholinesterases of several species of teleost fish, such as Gasterosteus aculeatus, Limanda limanda, Platichthys flesus and Serranus cabrilla. Cholinesterases from other aquatic organisms have also been characterized, as Gambusia holbrooki (Nunes et al., 2005), Lepomis gibbosus (Rodrigues et al., 2011), Corbicula fluminea (Ramos et al., 2012), Diopatra neapolitana and Solen marginatus (Nunes and Resende, 2017), Palaemonetes pugio (Key and Fulton, 2002), Oreochromis niloticus (Rodríguez-Fuentes and Gold-Bouchot, 2004), Haemulon plumieri (Leticia and Gerardo, 2008), and Poecilia reticulata (Garcia et al., 2000). In addition, Monteiro et al. (2005) made the characterization of cholinesterases (and analyzed the feasibility of using the characterized forms in biomonitoring programs) of the estuarine fish Pomatoschistus microps.