叶绿素含量测定

叶绿素含量测定 To measure pigment amounts per unit leaf area, we homogenized a frozen 1-cm-diameter disc of leaf tissue by hand using a plastic pestle in liquid nitrogen in a 1.5-mL tube, mixed by vortexing with 500 μ L of cold 80% acetone, and extracted overnight at 4 ? C in the dark. After each tube was vortexed again the next morning to expose any settled pigments to the acetone solution and was centrifuged at 18 000 × g for 15 min at 4? C, 100 μ L of each solution was added to 900 μ L of 80% acetone in a 1-mL cuvette. Absorbance was measured at 663 nm, 647 nm, and 470 nm using a UVmini-1240 spectrophotometer (Shimadzu Scientific Instruments, Columbia,Maryland, USA). Formulas from Lichtenthaler (1987) were used to estimate the concentrations ( μ g/mL) of chlorophyll a and chlorophyll b individually, as well as total chlorophyll ( a + b ) and total carotenoids (xanthophylls and β -carotene).