In the present study, fluorescence spectra of the dissolved organic matter (DOM) extracted from early stage leachate samples obtained from biologically pretreated leachate fed to RO tertiary treatment, the permeated, the concentrate, and liquids collected after cleaning the membrane with acid and then with base were determined. The results of synchronous fluorescence spectra analysis showed the RO membrane effectively removed the high content of organic matter corresponding to short wavelengths of 280, 340, and 370 nm of the feed. Liquids collected after cleaning membrane with acid and base showed obvious influence on the organic pollutant matters in the range of 300～420 nm. The results of three-dimensional excitation-emission matrix (3D-EEM) fluorescence spectra showed three protein-like peaks, namely low-excitation wavelength, tyrosine-like high-excitation wavelength, tyrosine-like high-excitation wavelength, tryptophan-like and two fulvic-like peaks, visible and ultra visible fulvic-like were found in the feed. The permeated contains two peaks with higher intensity than the feed, low-excitation wavelength tyrosine-like and high-excitation wavelength tyrosine-like, while the other three peaks were shown in the concentrate. The acid and the base cleaning had great influence on the molecule chemical structure of the organic pollutants on the RO membrane and caused obvious location shifts. It can be concluded that the RO mainly separated the fulvic matter in the early stage leachate and the fouling consisted of fulvic-like matter together with the protein-like, and low-excitation wavelength tyrosine-like.