Increase on the Initial Soluble Heme Levels in Acidic Conditions Is an Important Mechanism for Spontaneous Heme Crystallization In Vitro

Background: Hemozoin (Hz) is a heme crystal that represents a vital pathway for heme disposal in several blood-feeding organisms. Recent evidence demonstrated that beta-hematin (beta H) (the synthetic counterpart of Hz) formation occurs under physiological conditions near synthetic or biological hydrophilic-hydrophobic interfaces. This seems to require a heme dimer acting as a precursor of Hz crystals that would be formed spontaneously in the absence of the competing water molecules bound to the heme iron. Here, we aimed to investigate the role of medium polarity on spontaneous beta H formation in vitro. Methodology/Principal Findings: We assessed the effect of water content on spontaneous beta H formation by using the aprotic solvent dimethylsulfoxide (DMSO) and a series of polyethyleneglycols (PEGs). We observed that both DMSO and PEGs (3.350, 6.000, 8.000, and 22.000) increased the levels of soluble heme under acidic conditions. These compounds were able to stimulate the production of beta H crystals in the absence of any biological sample. Interestingly, the effects of DMSO and PEGs on beta H formation were positively correlated with their capacity to promote previous heme solubilization in acidic conditions. Curiously, a short chain polyethyleneglycol (PEG 300) caused a significant reduction in both soluble heme levels and beta H formation. Finally, both heme solubilization and beta H rmation strongly correlated with reduced medium water activity provided by increased DMSO concentrations. Conclusions: The data presented here support the notion that reduction of the water activity is an important mechanism to support spontaneous heme crystallization, which depends on the previous increase of soluble heme levels.