Superoxide dismutase determination kit (SOD, 19160-1KT-F), Glutathione Peroxidase Cellular Activity Assay Kit (GPx, CGP1-1KT), and Protein Quantification Kit-Rapid (51254-KT) were purchased from Merck, Germany. All the analyses were performed according to the manufacturer's instructions. Malondialdehyde (MDA) levels were assessed by thiobarbituric acid-reactive assay following a preestablished work protocol [41 (link)].
At the end of chronic exposure period, each fish was individually well homogenized in a 10 volume of ice-cold saline (0.90% NaCl). Each sample was centrifuged at 5500 rpm for 10 min in accordance with the already established protocols by Jin et al. [42 (link)] and Ni et al. [43 (link)]. The supernatant obtained was aliquoted into 2 ml Eppendorf tubes for the subsequent determination of oxidative biomarkers levels. A spectrophotometer at distinct wavelengths (Specord 210 Plus producer Analytik Jena, Germany) was used to assess each enzyme activity.
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