Sorption mechanisms of an antibiotic sulfamethazine (SMT) to humin (HM) isolated from a peat soil and its subfractions after sequential treatments were examined. The treatments of HM included removal of ash, O-alkyl carbon, lipid, and lignin components. The HF/HCl de-ashing treatment removed a large amount of minerals (mainly silicates), releasing a fraction of hydrophobic carbon sorption domains that previously were blocked, increasing the sorption of SMT by 33.3%. The de-O-alkyl carbon treatment through acid hydrolysis greatly reduced polarity of HM samples, thus weakening the interaction between sorbents with water at the interfaces via H-bonding, leaving more effective sorption sites. Sorption of SMT via mechanisms such as van der Waals forces and π-π interactions was enhanced by factors of 2.04–2.50. After removing the lipid/lignin component with the improved Soxhlet extraction/acid hydrolysis, the organic carbon content-normalized sorption enhancement index Eoc was calculated. The results demonstrated that the Eoc-lipid for SMT (16.9%) was higher than Eoc-lignin (10.1%), implying that removal of unit organic carbon mass of lipid led to a higher increase in sorption strength than that of lignin. As each component was progressively removed from HM, the sorption strength and isotherm nonlinearity of the residual HM samples for SMT were gradually enhanced. The Koc values of SMT by HM samples were positively correlated with their aromatic carbon contents, implying that π-π electron donor-acceptor interactions between the benzene ring of sorbate and the aromatic domains in HM played a significant role in their interactions.