The adsorption of persistent organic pollutants such as pentachlorophenol (PCP) on soil matrices plays a crucial role in determining their environmental persistence and ecological risk. This study provides molecular-level insights into the sorption mechanisms of pentachlorophenol on biochar-amended soil organic matter, combining experimental and computational approaches to unravel the fundamental interactions that govern pollutant immobilization. The research employed Fourier Transform Infrared (FTIR) and solid-state Nuclear Magnetic Resonance (NMR) spectroscopy to characterize functional group interactions, complemented by Density Functional Theory (DFT) modeling to elucidate binding configurations at the molecular scale. Results revealed that biochar amendment significantly enhanced PCP sorption capacity compared to unamended soils, primarily due to the introduction of ?–? electron donor–acceptor interactions, hydrogen bonding, and hydrophobic partitioning between aromatic moieties of biochar and the chlorinated phenol molecule. Spectroscopic analyses confirmed the formation of stable surface complexes involving carboxyl, hydroxyl, and aromatic carbon sites. DFT simulations further identified energetically favorable adsorption geometries, with binding energies consistent with experimental thermodynamic trends. These findings indicate that biochar modifies soil organic matter structure to create heterogeneous, high-affinity sorption domains that reduce PCP bioavailability and mobility. The integration of spectroscopic validation with computational modeling provides a mechanistic understanding of how biochar mediates molecular interactions, advancing knowledge in the field of sustainable soil remediation. The study establishes a foundation for designing biochar materials with tailored surface chemistries for the efficient immobilization of hydrophobic organic contaminants. Ultimately, these insights contribute to the development of predictive sorption models and climate-resilient remediation strategies for persistent pollutants in terrestrial ecosystems.