Wheat arabinoxylan (AX) exhibits characteristics of high molecular weight (Mw), strong water absorption and water holding capacity, which significantly influence the hydration and solubility properties of wheat flour. However, challenges existed in visualizing the distribution of AX and quantifying water content accurately. This study investigated the effect of fluorescent labeling on the physicochemical properties of AX with different Mw. AX was initially degraded using xylanase, which resulted in the formation of AX fragments with different Mw. As the xylanase concentration increased, the Mw, degree of aggregation and apparent viscosity of AX decreased, whereas the water binding capacity, solubility and hydrophilicity increased. Enzymatically hydrolyzed AX was covalently linked to fluorescein isothiocyanate (FITC) through reductive amination modification, yielding AX-tyramine-FITC. Compared to AX, AXTF exhibited a slightly higher Mw, with a slight increase in water binding capacity and hydrophilicity, a significant increase in solubility. The degree of aggregation and apparent AX viscosity also increased slightly. The fluorescent labeling had a minor effect on AX properties, and the trend in AXTF properties with respect to Mw was consistent with that of AX. This study established a basis for the effect of AX with different Mw on the water migration pattern in the flour system, and provided a theoretical basis for the application of AX fluorescent labeling for visualization purposes.