Subcellular Cr Fractionation in Leaf and Root

The subcellular fractionation of Cr in leaf and root tissues was quantified employing the procedure of Sheng et al. [108 (link)]. Briefly, fresh leaf and root material (0.5 g) was extracted in 10 mL of extraction buffer (1.0 mM DL-dithioerythritol, 0.25 mM sucrose, 5 mM AsA, and 50 mM Tris-HCl). The extract was filtered by using a nylon fabric with a mesh size of 100 mm and the filtrate was labelled as “cell wall fraction” (CWF) and subjected to centrifugation (15,000 rpm) for 45 min. The aliquot solution and resultant pellet were labelled as “soluble fraction” (SF) and “organelle fraction” (OF), respectively. These procedures were carried out at 4 °C. Both CWF and OF were transferred to an Erlenmeyer flask (100 mL) filled with deionized water, dried, and digested with HNO₃ (5 mL) [109 (link)]. Thereafter, Cr concentrations were determined with an ICP-OES (Perkin Elmer Optima 5300 DV).