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Field-collected, young plants of Ni hyperaccumulator Stackhousia tryonii, grown in a glasshouse for 20 weeks, were exposed to low- (available Ni concentration in the native serpentine soil, i.e. 60 μg g⁻¹ dry soil) and high- (external application of 1000 ppm) Ni concentrations in the substrate. Nickel concentration in the freeze-dried leaf tissues increased from 3700 μg g⁻¹ to 13 700 μg g⁻¹ with soil Ni supplementation, of which >60% was extracted with dilute acid (0.025 M HCl). Nickel supplementation also elicited a 575%, 211%, and 37% increase in the final concentrations of oxalic, citric, and malic acids, respectively, in leaf tissues. Malic acid was the dominant organic acid, followed by citric and oxalic acids. The molar ratio of Ni to malic acid was 1.0, consistent with a role for malate as a ligand for Ni in hyperaccumulating plants, supporting detoxification/transport and storage of this heavy metal in S. tryonii. The total amino acid concentrations in the xylem sap did not change with Ni supplementation (21.7±3.7 mM and 17.9±5 mM, respectively, for low- and high-nickel-treated plants). Glutamine was the major amino acid in both the low- and high-Ni-treated plants. The concentration of glutamine decreased by >60%, with a corresponding increase in alanine, aspartic acid, and glutamic acid, on exposure to high Ni. A role of amino acids in Ni complexation and transport in S. tryonii is not immediately apparent.