The composition of isolated natural organic matter (NOM) fractions was studied by means of high pressure size-exclusion chromatography (HP-SEC), specific ultraviolet absorbance (SUVA) at 254 nm, total luminescence spectroscopy (TLS) and synchronous scanning fluorescence measurements. The HP-SEC, SUVA and TLS studies revealed that humic acids (HA) and hydrophobic acids (HOA) are composed of complex, aromatic and high molecular weight subunits. Hydrophobic neutrals (HON) consist of few groups of compounds with relatively low molecular sizes and a degree of condensed aromatic moieties. Similar characteristics were observed for hydrophilic acids (HIA) and bases (HIB). Obtained results also demonstrated that hydrophilic neutrals (HIN) are made of non-humic, aliphatic and low molecular weight components. Furthermore, the synchronous excitation-emission spectra indicated the presence of proteinlike constituents in HIB, HIN and HON fractions as well as lignin-derived constituents with relatively low molecular sizes in HIA.