Uv vis

Leaves or needles (0.2 g) were homogenized using a mortar and pestle in 2 mL of 0.1% thiobarbituric acid (TBA). The extracts were centrifuged at 10,000× g for 20 min, and the resulting supernatants were used to determine lipid peroxidation and H₂O₂.
The level of lipid peroxidation in the leaves and needles of the selected species was evaluated by measuring malondialdehyde (MDA) using the thiobarbituric acid method, which yields a colored product [58 ]. A mixture of supernatant and 0.5% thiobarbiturate acid (TBA) in 20% trichloroacetic acid (TCA) was heated at 95 °C for 30 min and then cooled in an ice bath. After centrifugation at 10,000× g for 10 min, the absorbance of the supernatant was read at 532 nm, and the correction for unspecific turbidity was conducted by subtracting the absorbance at 600 nm on the LaboMed UV-VIS. A total of 0.25% TBA in 10% TCA served as the blank. The MDA content was calculated according to its extinction coefficient of 155 mM⁻¹ cm⁻¹ and expressed as nmol g⁻¹ FW [58 ,59 (link)].
To determine hydrogen peroxide (H₂O₂), a 100 mM potassium phosphate buffer and potassium iodide were added to the supernatant. The absorbance of the mixture was read at 390 nm on the LaboMed UV-VIS. The molar extinction coefficient for H₂O₂ is 0.28 µM⁻¹ cm⁻¹, and the amount of H₂O₂ is expressed as µmol per gram of fresh weight of leaves or needles (µmol g⁻¹ FW) [58 ,59 (link)].

Leaves or needles (0.1 g) were homogenized using a mortar and pestle in 10 mL of chilled 80 % acetone. The chlorophyll extract was refrigerated at 4 °C for 24 h. The samples were then centrifuged at 4000× g for 10 min, and the absorbance of the supernatant was measured using a spectrophotometer (LaboMed UV-VIS) at 663 nm and 645 nm. The content of photosynthetic pigments is expressed as µg of photosynthetic pigments per gram of fresh weight of leaves or needles (µg g⁻¹ FW) [57 (link)].

Plant samples used in the foliar nutrition analysis were oven-dried at 80 °C for 24 h. Approximately 0.3 g of sub-samples were digested using a mixture of 30% H₂O₂ and 65% HNO₃ for plants and 0.3 g of the elemental analyzer (LECO CNS-2000, St. Joseph, MI, USA) [56 ]. The concentration of nutrients was determined as follows: N using an elemental analyzer P with a spectrophotometer (LaboMed UV-VIS, Los Angeles, CA, USA) and Ca, P, Mg, and K using an atomic absorption spectrophotometer (Perkin Elmer AAS AAnalyst 700, Waltham, MA, USA) [56 ].