Comparative Analysis of Phytochemicals and Gene Expression in Soybean (Glycine max) Under Acute Moderated and Severe Elevated Ozone: Unravelling the Role of Antioxidant Defence

The increasing concentrations of ground-level ozone (O₃) resulting from industrialisation and anthropogenic activities present a substantial environmental threat to agricultural productivity, particularly affecting O₃-sensitive crops such as soybeans. The effects of acute O₃ exposure on soybean yield attributes and seed quality and whether soybean showed different detoxification mechanisms in response to moderate and severe O₃ stress are not extensively explored. In this study, soybean seedlings were exposed to moderate (80 nmol mol⁻¹) and acute severe (200 nmol mol⁻¹) O₃ stress, and then growth parameters, yield attributes, reactive oxygen species (ROS) levels, enzymatic and non-enzymatic antioxidant properties and associated gene expression in the leaves were assessed. The results revealed that moderate O₃ exposure enhanced growth parameters but reduced the 100-grain weight, while acute severe exposure sharply depressed growth parameters, yield attributes and the 100-grain weight. Moderate O₃ fumigation significantly increased hydrogen peroxide (H₂O₂) levels and catalase (CAT) activity from 4 to 32 h. Acute severe O₃ stress induced the overproduction of superoxide anions (O₂^.−) and H₂O₂ during nearly the whole experiment period, but only enhanced superoxide dismutase (SOD) activity at 32 h, and showed no stimulatory effects on CAT activity. Additionally, the relative expression levels of the SOD and CAT gene family in soybean leaves exposed to elevated O₃ were upregulated, peaking at 8 h. Moderate O₃ treatment enhanced reduced glutathione (GSH) and ascorbate (AsA) levels and increased the activities of AsA–GSH cycle-related enzymes. In contrast, acute severe O₃ exposure inhibited GSH and AsA contents and markedly suppressed AsA–GSH cycle-related enzymes, particularly from 8 to 32 h. Redundancy analysis indicated that CAT and AsA play crucial roles in scavenging O₃-induced ROS under moderate stress, while ascorbate peroxidase (APX) and GSH were more effective under acute severe stress conditions. These findings provide insights into the differential impacts of acute O₃ stress on soybeans, emphasising the importance of considering both crop yield and grain quality in assessing O₃ risks to crops.