The complexity of natural organic matter (NOM) motivates determination of how specific components in a NOM mixture interact with and affect nanoparticle (NP) behavior. The effects of two Suwannee River NOM fractions (separated by a 100,000 g/mol ultrafiltration membrane) on gold NP aggregation are compared. The weight-average molecular weight, Mw, for the unfractionated NOM was 23,300 g/mol, determined by size exclusion chromatography with multiangle light scattering. The NOM was comprised of ~1.8 wt % of >100,000 g/mol retentate (NOMr, Mw = 691,000 g/mol) and 98 wt % of filtrate (NOMf, Mw = 12,800 g/mol). Ten ppm of NOMr provided significantly better NP stability against aggregation than 10 ppm of NOMf in 100 mM NaCl due to steric effects. In the unfractionated NOM, the relative importance of the two components was concentration-dependent. For a low concentration of unfractionated NOM (10 ppm), both fractions contributed to the NOM effects; for a high concentration (560 ppm), NP stability was controlled by the small amount (10 ppm) of NOMr present, rather than the higher amount (550 ppm) of NOMf. Therefore, large humic aggregates in a heterogeneous NOM sample can have disproportionately strong effects, and characterization of Mw distributions (rather than average Mw) may be required to explain NOM effects on NP behavior.