Biochemical analyzer bs 200

At the end of the experimental period, the animals were fasted for 12-15 hours, then
anesthetized with an intramuscular injection of ketamine (50 mg/kg) and xylazine (0.5
mg/kg), and euthanized by decapitation. Blood samples were collected and the serum
was separated by centrifugation at 3,000 X g for 15 minutes at 4°C, and stored at
-80°C until further analysis. The serum was analyzed for levels of glucose,
triglycerides (TG), total cholesterol (T-Chol), high-density lipoprotein cholesterol
(HDL), low-density lipoprotein cholesterol (LDL), insulin, and
leptin. Glucose, TG, T-Chol, HDL, and LDL were measured with an automatic enzymatic
analyzer system (Biochemical analyzer BS-200, Mindray, China). Leptin and insulin
levels were determined by enzyme-linked immunosorbent assay (ELISA) method using
commercial kits (Linco Research Inc., St. Louis, MO, USA).

Blood samples were collected from the abdominal artery and centrifuged at 5000 rpm for 10 minutes (Eppendorf Centrifuge 5804-R, Hamburg, Germany) in dry tubes, and the plasma was collected and stored in Eppendorf tubes in the freezer at -80°C (Thermo Fisher Scientific LLC, Asheville, NC, USA). The tissues used for further analysis were collected and frozen immediately with liquid nitrogen and then packaged and stored in the freezer at -80ºC (ColdLab Ultra Freezer CL374-86V, Piracicaba, São Paulo, Brazil). Plasma was evaluated by levels of total cholesterol (T-Chol) and high-density cholesterol (HDL), and hormones (insulin and leptin). T-Chol and HDL were measured with an automatic enzymatic analyzer system (Biochemical analyzer BS-200, Mindray, China). Leptin levels were determined with the enzyme-linked immunosorbent assay (ELISA) method using commercial kits (Linco Research Inc., St. Louis, MO, USA).

Serum was separated by low‐speed centrifugation (3000 r/min) for 15 minutes. The levels of MDA, SOD, and TAC in serum were detected with an automatic analyzer (Biochemical analyzer BS-200, Mindray, China). The operation process was carried out according to the manufacturer's instructions.