3.6.2. Phenylalanine Ammonia Lyase (PAL) Activity

PAL activity was measured as described by Zhan et al. [58], wherein 50 mg of fresh plant material was homogenized in 1000 µL phosphate buffer (Na e K, 50 mM, pH 8.0) and centrifuged (20,000× g, 20 min, 4 °C). Five hundred microliters of L-phenylalanine (50 mM) were mixed with 100 µL of supernatant and 1400 µL of the buffer. The mixture was incubated at 37 °C for 1 h. The optical density was measured at 290 nm and the enzyme activity was expressed as μmol cinnamic acid per hour, per milligram of protein (μmol_CA/mg_protein/h). The protein content of enzyme extracts was evaluated using the Bradford [56] method.