This study systematically investigated the correlations between fluorescence distributions characterized by the excitation-emission matrix (EEM) and hydrophobic/hydrophilic composition of dissolved organic matter (DOM) in membrane bioreactors (MBRs). On the basis of samples from 10 full-scale MBRs, we performed point-to-point comparisons among different components using an EEM fluorescence quotient (FQ) method and obtained a hydrophobic/hydrophilic fluorophore distribution map via Wilcoxon signed rank test. Hydrophobic acids/bases (HOA/HOB) concentrated in the low-wavelength region [excitation wavelength (Ex) < 235 nm], while hydrophilic substances (HIS) were enriched in the region of Ex > 235 nm [especially with emission wavelength (Em) = 300-360 nm]. Quantitatively, EEM regional contribution to whole wavelength fluorescence was found to significantly correlate with the hydrophobic/hydrophilic proportions of DOM, with Pearson's coefficients of 0.94 and 0.78 (p < 0.01) for HOA and HIS, respectively. We established a linear regression model showing the HOA proportion as a function of the EEM regional contribution at (Ex, Em) = (200-285, 340-465 nm), with R2 = 0.876, which was validated via leave-one-out cross-validation and Monte Carlo simulation. This study shows a statistically hydrophobicity-dependent fluorescence property across different MBRs, and it might be applied to provide a quick estimation of hydrophobic/hydrophilic composition of DOM in wastewater treatment systems based on EEM monitoring.