Quantifying Leaf Pigments: Chlorophyll, Carotenoids, and Anthocyanins

The contents of chlorophyll, carotenoid, and anthocyanin of the QHP, ZSY, and L2025 leaves were determined based on a previously described method (Tian et al., 2021b (link)). Fresh leaves were used to measure total chlorophyll content based on the method described by Wu et al. (2020) (link). Leaf tissue (0.1 g) was ground into powder and extracted in 5 mL of 95% ethanol at 50°C for 2 h. The mixture was vortexed and centrifuged at 5,000 rpm for 5 min. The absorbance of the supernatant was measured at 470, 649, and 665 nm using an ultraviolet–visible spectrophotometer (Wu et al., 2020 (link); Zhuang et al., 2021b (link)). Fresh leaves were used to determine the carotenoid content in accordance with the method used by (Gao et al., 2021 (link)). Approximately 100 mg of fresh leaf tissue were cut into pieces with scissors, and extracted in 10 mL of 1% (v/v) HCl–ethanol at 60°C for 30 min. The mixture was vortexed and centrifuged at 13,000 × g for 5 min. The absorbance of the supernatant was measured at 530, 620, and 650 nm using an ultraviolet-visible spectrophotometer (Gao et al., 2021 (link)).