Antioxidant Enzymes and MDA Determination

The REC was measured using the immersion method [57 (link)]. Pro content was assayed following the methods described by Bates et al. [58 (link)]. Briefly, 0.5 g leaf samples were added in 5 mL of 3% sulfosalicylic acid and then placed in boiling water for 10 min. After filtering, the supernatant (2 mL) was mixed with 2 mL of acetic acid glacial and 2 mL of ninhydrin. Subsequently, the mixture was maintained in boiling water for 30 min. After cooling, 4 mL of methylbenzene was added. Absorbance was measured at 520 nm using methylbenzene as blank. MDA content was determined using a plant MDA assay kit (Nanjing Jiancheng, Nanjing, China) in accordance with the manufacturer’s instructions. The activities of antioxidant enzymes (SOD, POD, and CAT) were determined following previously described procedures [48 (link)].