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Abstract In this study, we utilized a size exclusion chromatography (SEC) system that was equipped with a fluorescence emission scanning mode to explore the heterogeneous distribution of the humification index (HIX) values within bulk dissolved organic matter (DOM). The HIX-based SEC chromatograms showed that the molecular weight (MW)-dependent distribution of aromatic condensation was heavily affected by the DOM sources and the progress of biodegradation. The HIX heterogeneity across different MW was more pronounced for terrestrial versus aquatic DOM sources. Microbial incubation of leaf litter DOM led to the initial enhancement of the HIX at a relatively low MW, followed by a gradual increase at larger MW values. The dynamic changes of the HIX can be attributed to (1) the preferential removal of non-aromatic or less-aromatic molecules by microorganisms, (2) the production of microbial metabolites, (3) microbial humification, and (4) self-assemblage of humic-like molecules. From the SEC chromatograms, the HIX-based average molecular weight (or MW HIX) was proposed as a unifying surrogate to represent an MW that was highly associated with aromatic condensation. The MW HIX discriminated four different DOM sources and described well the biodegradation-induced changes. The MW HIX also presented a good positive correlation with pyrene organic carbon-normalized binding coefficients (K oc). The prediction capability of the MW HIX for pyrene K oc was higher than those based on the single descriptors of bulk DOM, such as HIX and MW, which revealed its superior linkage with the DOM reactivity related to both MW and HIX. Graphical abstract Unlabelled Image Highlights • A SEC system with multi emission scan was applied to propose a new DOM descriptor. • The new index, MW HIX , represents a MW value associated with aromatic condensation. • MW HIX was useful to describe the structural changes of DOM upon biodegradation. • MW HIX proved its effectiveness for the prediction of pyrene binding affinity. [ABSTRACT FROM AUTHOR]