Gsh px activity assay kits

At the end of the restraint, mice in each group were sacrificed by decapitation. The brain was removed and the hippocampus was quickly dissected on an ice-cold surface. Brain tissue was shortly homogenized in cold 0.9% NaCl solution to give 10% homogenate (w/v). After being centrifuged at 2500 rpm/min for 10 min at 4 °C to remove nuclei and debris, the supernatants were separated for further analysis.
The superoxide dismutase (SOD) and glutathione peroxidase (GSH-PX) activity assay kits were purchased from Nanjing Jiancheng Bio-engineering Institute (Nanjing, China). Total SOD activity and GSH-PX activity were assayed with the guidance of the instructions described in the commercial kits [38 (link)]. One unit of SOD activity is calculated by the amount of enzyme in each ml of the reaction solution at 50% SOD inhibition at 37 °C. One unit of GSH-PX activity is calculated by the net amount of enzyme capable of hydrolyzing 1 μmol of GSH (Glutathione) per min at 37 °C. Protein concentration was determined in hippocampal homogenates using bovine serum albumin as a standard.

All the processes were strict in accordance with the instructions of the superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) activity assay kits (Jiancheng, Nanjing, China), as well as the glutathione (GSH) and malondialdehyde (MDA) level measurement kits (Jiancheng). The disrupted cell or tissue lysate was centrifuged at 12000g for 5 min, and the supernatant was mixed with detection solution and incubated for 40 min at 95°C in a water bath. After cooling, the samples were centrifuged at 4000g for 10 min. The optical density values of each group were measured and recorded at 532 nm with a 1 cm light path. The SOD and GSH-Px activities as well as the GSH and MDA levels were calculated based on the optical density values.