Hemozoin (Hz) is a heme crystal produced upon the digestion of hemoglobin (Hb) by blood-feeding organisms as a main mechanism of heme disposal. The structure of Hz consists of heme dimers bound by reciprocal iron-carboxylate interactions and stabilized by hydrogen bonds. We have recently described heme crystals in the blood fluke, Schistosoma mansoni, and in the kissing bug, Rhodnius prolixus. Here, we characterized the structures and morphologies of the heme crystals from those two organisms and compared them to synthetic b-hematin (bH). Synchrotron radiation X-ray powder diffraction showed that all heme crystals share the same unit cell and structure. The heme crystals isolated from S. mansoni and R. prolixus consisted of very regular units assembled in multicrystalline spherical structures exhibiting remarkably distinct surface morphologies compared to bH. In both organisms, Hz formation occurs inside lipid droplet-like particles or in close association to phospholipid membranes. These results show, for the first time, the structural and morphological characterization of natural Hz samples obtained from these two blood-feeding organisms. Moreover, Hz formation occurring in close association to a hydrophobic environment seems to be a common trend for these organisms and may be crucial to produce very regular shaped phases, allowing the formation of multicrystalline assemblies in the guts of S. mansoni and R. prolixus.

The multi-frequency high-field electron paramagnetic resonance (HFEPR) was used to study the magnetic properties of malarial pigment hemozoin and its synthetic analogue, β-hematin. (FeIII−protoporphyrin−IX)2 dimers containing five-coordinate high-spin FeIII, S = 5/2, are the building blocks of these pigments. The fit of EPR spectra that were acquired in an unprecedented wide range of microwave frequencies of 34 and 94 GHz for hemozoin and 27−500 GHz for β-hematin yielded a complete set of intrinsic spin Hamiltonian parameters:  D = +5.85(1) cm-1, E = 0, g ⊥ = 1.95(1), g ∥ = 2.00(1). These results point to the existence of largely axial symmetry of the iron environment in the bulk phase of hemozoin and β-hematin.