For the design of ligands specific for iron(III), it is important to explore the molecular conformations of both the ligand and the metal,complex since they are responsible for the thermodynamics of the chelation process. The conformational behavior of ferric hexadentate N,N,N-tris[2-(3-hydroxy-2-oxo-1,2-dihydropyridin-1-yl)-acetamido]ethylamine complex (coded-as FeCP130) was investigated both in vacuo and in aqueous solvent. Two conformations are of interest in this complex. One is referred to as "in" conformation, where the tertiary amine N atom and the free electron pair point toward the metal atom and the other, the "out" conformation, where the same N atom points "out", away from the metal atom. Both molecular mechanics and molecular dynamics simulation studies show that the "in" conformation is more stable than the "out" Conformation. By use of the free energy perturbation method, the relative free energy difference between the two conformations was determined and contributions from both enthalpy and entropy were evaluated. An analogue of FeCP130 was modeled to evaluate the role played by the intramolecular hydrogen bonds between amide protons and chelating oxygen atoms in determining the conformation and the stability of FeCP130. The lack of preorganization of hexadentate 3-hydroxy-2(1H)-pyridinone CP130 was revealed by simulating the process : protonated free ligand --> chelated ligand. The present study offers an explanation for the observation that at high concentrations, hexadentate 3-hydroxy-2-(1H)-pyridinones are weaker iron(III) binders than their bidentate analogues. The work presents the first computational study of the relative free energy of complexation of a siderophore-type ligand with iron(III).