Results were statistically analyzed t-test,
with significance of 0.05. The bloom extract showed the presence of 2 MCs (Fig. 1), while the mass spectrometry analyses on reflective mode showed the mass components at 995.577 Da and 908.961 Da, suggesting the presence of Microcystin-LR and Microcystin-LA respectively (Fig. 2). The mass spectrometry analyses in lift mode confirmed the presence of MCs by showing in the fragment pattern the ion 135.0, an ADDA fragment exclusive to microcystin toxins (Data not shown). The concentration of total MCs estimated in the extract was 138.30 μg mL−1. Separately, the concentrations were 41.0 μg mL−1 and Etoposide 97.9 μg mL−1 for MC-LR and MC-LA respectively. Neither mortality nor visible adverse effects were observed in the two exposure routes during the experiments. Doses of 13.80 μg kg−1 bw for 72 h in the ip assay and 103.72 μg L−1 for 72 h in the exposure assay were considered the maximum tolerated doses, because previously tested higher doses caused toxic signals such as refusing food, scale loss, alteration in the behavior and mortality. The LC50 was greater than 103.72 μg L−1 of microcystin in the body exposure route and the LD50 was greater than 13.80 μg kg−1 bw
via ip. The micronucleus test showed that only the highest concentration of 103.72 μg L−1, Selleck 5-FU via body exposure, was statistically significant (P = 0.040, Table 1). Treatment by intraperitoneal injection did not induce MN ( Table 1). The highest treatment via body exposure and both treatments via intraperitoneal injection significantly increased the index of DNA damage ( Table 2). Apoptosis-necrosis test showed that at lower concentrations more apoptosis was found. On the other hand, at higher concentrations more necrosis was found ( Table 3). Gaudin et al. (2008) reported DNA damage in different tissues through single cell gel electrophoresis (comet assay)
in mice after single acute oral administration and intraperitoneal injection of MCs. An extract of cyanobacteria containing MCs from a water source in China was mutagenic to Salmonella typhimurium (Ames test), induced DNA damage in rat hepatocytes (comet assay) and induced micronucleated polychromatic erythrocytes in mouse bone marrow nearly ( Ding et al., 1999). Zegura et al. (2004) demonstrated in human hepatoma HepG2 cells that MC-LR increases the levels of reactive oxygen species (ROS), providing evidence that the observed genotoxicity was mediated by ROS. MC-LR and -RR increased the antioxidant enzymatic activities of superoxide dismutase and catalase in tilapia fish Oreochromis sp ( Jos et al., 2005 and Prieto et al., 2006). In our previous study in Astyanax bimaculatus (Pisces: characidae), this same batch of microcystin induced comets and MN ( Silva et al., 2010). Thus, there is evidence that MCs are genotoxic to different aquatic species and mammals, including human cells. In contrast, Abramsson-Zetterberg et al.
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