Fluorescence excitation-emission matrix (EEM) analysis was applied to characterize seasonal variation of dissolved organic matter (DOM) in arsenic-affected shallow aquifers of the Jianghan Plain, central China, to better understand the effect of DOM on arsenic transformation during dry season and wet season. It was identified that there are three components of DOM, of which C1 and C3 are fulvic acid-like substances and C2 is humic acid substance, and there is no obvious seasonal variability in C2, and the concentration is slightly higher in the wet season than the dry season. C1 and C3 have the opposite change rules in different seasons, which is mainly reflected in the shallow unconfined water.