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Indole-3-acetic acid (IAA) is a potential mediator in the protection of plants from copper (Cu) toxicity and the enhancement of Cu tolerance. In this paper, spinach (Spinacia oleracea L.) seedlings were cultivated in soil containing 700 mg kg−1 Cu and the leaves of seedlings were sprayed with different concentrations of IAA. Exogenous IAA treatment reduced the malondialdehyde (MDA) concentrations in Cu-stressed seedlings and increased biomass, proline content, and the activities of antioxidant enzymes. Exogenous IAA treatment also increased the levels of nitrogen (N) assimilation compounds and the activities of N-metabolizing enzymes, but reduced NH4+ content. Notably, lower concentrations of IAA (10–40 mg L−1) increased the Cu concentrations in roots and reduced the Cu concentrations in leaves, while higher concentrations of IAA (50 mg L−1) reduced the Cu concentrations in both roots and leaves to the lowest levels. The findings indicated that the application of IAA reduced Cu accumulation, alleviated Cu toxicity, and enhanced Cu tolerance in spinach seedlings. IAA application could be used as an alternative strategy for reducing Cu accumulation in vegetable crops and for remediating Cu-contaminated soil, in turn reducing the hazardous effects of heavy metal contamination on human health and the environment.

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