Microscale Spatiotemporal Variation of Reactive Oxygen Species in the Charosphere: Underlying Formation Mechanism and Their Role in CO2 Emission

Charosphere, a highly active zone between biochar and surrounding soil, is widely present in agricultural and wildfire-affected soils, yet whether reactive oxygen species (ROS) are produced within the charosphere remains unclear. Herein, the production and spatiotemporal evolution of charosphere ROS were explored. In situ ROS capture visualized a gradual decrease in ROS production with increasing distance from the biochar/soil interface. Temporally, O2•- and H2O2 contents initially increased and then declined with increasing incubation time, peaking at 3.04 and 5.40 μmol kg-1, respectively, while •OH content decreased continuously. High-throughput sequencing revealed that dissolved biochar (DBC) facilitated ROS production by promoting the growth of bacteria with electron-releasing capacity, such as Bacteroidetes, Acidobacteria, Actinobacteria, and Chloroflexi. Additionally, adding electron transfer-weakened DBC significantly decreased ROS contents (ANOVA, P < 0.05), demonstrating that DBC also served as the electron shuttle and electron-storing materials to promote ROS production by accelerating electron transfer. This was further confirmed via fluorescence imaging, which visually showed stronger electron transfer ability near the soil/biochar surface. Inhibition and isotope experiments revealed the critical role of charosphere ROS in CO2 emissions, primarily from soil organic carbon. This study highlights the charosphere as a prevalent yet overlooked ROS hotspot, advancing our understanding of organic carbon turnover in soils.