| Summary: | Water shortages greatly threaten global food security and limit crop production. Hence, increasing crop water stress tolerance is a critical way to secure agricultural production. 24-Epibrassinolide (EBL) and spermine (Spm) are closely involved in plant growth and development, as well as stress tolerance. In this study, the potential role of 0.1 mg L<sup>−1</sup> EBL and/or 25 mg L<sup>−1</sup> Spm foliage applications in improving the tolerance of maize to water-deficit conditions (50% and 75% field capacity) was investigated. We found that EBL, either alone or in combination with Spm, plays a major role in maize drought tolerance through upregulating the expression of both regulatory genes (<i>ZmBZR1</i> and <i>ZmBES1</i>) of the brassinosteroid signal transduction pathway and gene-encoding antioxidant defense enzymes <i>ZmSOD</i>, <i>ZmCAT</i>, <i>ZmAPX</i>, <i>ZmMDHAR</i>, <i>ZmDHAR</i>, and <i>ZmGR.</i> Moreover, exogenous treatments alleviated the inhibition of maize plant growth and productivity and mitigated drought-induced oxidative stress by improving antioxidant enzyme (superoxide dismutase, catalase, ascorbate peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase, glutathione reductase) activity, enhancing antioxidant molecule (ascorbate, glutathione) content, preventing reactive oxygen species accumulation, and maintaining cell membrane integrity. These findings reveal that the application of EBL, either individually or in combination with Spm, can be a good strategy for ameliorating water stress in sustainable agricultural systems.
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