The iron coordination chemistry of two macrocyclic dihydroxamate siderophores, alcaligin (AG) and bisucaberin (BR), has been investigated thermodynamically and structurally. Alcaligin is a siderophore of freshwater bacteria as well as mammalian pathogens, including the bacterium that causes whooping cough in humans, while bisucaberin, a structural analogue of alcaligin, is produced by marine bacteria. Both alcaligin and bisucaberin form 1:1 ferric complexes (FeL(+)) in acidic conditions and 2:3 ferric complexes (Fe(2)L(3)) at and above neutral pH. The stability constants of these macrocyclic dihydroxamate siderophores differ significantly from that of rhodotorulic acid (RA), a linear dihydroxamate siderophore. Notably, K(FeL) of alcaligin is 32 times greater than that of rhodotorulic acid, while the subsequent stepwise formation constant for Fe(2)L(3) is 3 times less. The Fe(III) complexes of alcaligin are stereospecific; the absolute configuration of the Fe(2)L(3) complex (circular dichroism and X-ray structure) is Lambda. The structure of the Fe(2)L(3) alcaligin complex is a topological alternative to the triple-helicate structure of the rhodotorulic complex Fe(2)(RA)(3). The structures of the free ligand and the bisbidentate ligand in the FeL complex are essentially identical, indicating that alcaligin is highly preorganized for metal ion binding. This explains the difference in K(FeL) between alcaligin and rhodotorulic acid, as well as explaining the monobridged topology of the Fe(2)L(3) alcaligin complex. The protonation constants (log K(a1) and log K(a2)) are 9.42(5) and 8.61(1) for alcaligin and 9.49(2) and 8.76(3) for bisucaberin. The stepwise formation constants of the Fe(III) complexes (log K(ML) and log K(M)()2(L)()3) are 23.5(2) and 17.7(2) for alcaligin and 23.5(5) and 17.2(5) for bisucaberin. The overall formation constants (log beta(230)) of alcaligin and bisucaberin are 64.7(1) and 64.3(1). The solution chemistry of Fe(III) and alcaligin was further investigated at a lower ligand to metal ratio (1:1). At high pH, a novel 2:2 ferric bis-&mgr;-oxo-bridged complex of alcalagin forms (Fe(2)L(2)O(2)(2)(-)) with a log beta(22)(-)(4) of 16.7(2). This species exhibits behavior consistent with an iron bis-&mgr;-oxo complex, including antiferromagnetic coupling. Crystal data: Fe(2)(AG)(3).25H(2)O crystallizes in the orthorhombic space group P2(1)2(1)2(1) with a =13.3374(4) Å, b = 16.1879(5) Å, c = 37.886(1) Å, V = 8179.7(4), Z = 4. For 5512 reflections with F(o)(2) > 3sigma(F(o)(2)) the final R (R(w)) = 0.053(0.068).