*2.6. Proline*

Proline content was determined according to [28]. A total of 0.5 g of fresh leaves was extracted in 10 mL of aqueous sulfosalicylic acid (3%). A total of 2 mL of the extract was taken and mixed with 2 mL of acid ninhydrin reagent (Alpha, Mumbai, 400 002, India) and 2 mL of glacial acetic acid for one h at 100 ◦C. After cooling, 4 mL of toluene was added to the reaction mixture to extract the proline content. The absorbance was recorded using a spectrophotometer (VEB, Carl-Zeiss-Promenade, Jena, Germany) at 520 nm using toluene as a blank. Free proline was determined from the standard curve of *L*-proline.

#### *2.7. Total Phenolics*

Total phenolic content in dry leaves was estimated by the method described by [29,30]. One milliliter of the extract was added to ten drops of concentrated HCl in a boiling water bath for ten min and cooled. Then 1 mL of Folin–Ciocalteau reagent and 1.5 mL of 14% sodium carbonate were added to the extract. The mixture was completed with distilled water up to 5 mL, shaken well, and then kept in a boiling water bath for five minutes. The absorbance at 650 nm was noted, and the data were represented as mg g−<sup>1</sup> FW using a pyrogallol standard curve.
