The use of density functional methods for the elucidation of the structure of radiation-induced bio-radicals by comparison of computed and experimental EPR properties is discussed. Three case studies, radiation induced radicals of the amino acid alanine, steroid hormones and β-d-fructose, with increasing degree of uncertainty about the proposed radical structures, are investigated. Next to the analysis of the isotropic and anisotropic components of the hyperfine tensor, also the direction cosines of the principal axes of this tensor were investigated in greater detail in the case of the β-d-fructose radicals. Since all radicals considered in this contribution are formed in a solid matrix, also the question as to how to incorporate the effect of the molecular environment is addressed. It is concluded that the methodology outlined represents a powerful tool to aid experimentalists in the assignment of the contributions of various radicals contributing to the observed EPR spectra. Copyright © 2004 John Wiley & Sons, Ltd.