Phospholipid membranes are thought to be one of the main inducers of hemozoin formation in Plasmodia and other blood-feeding parasites. The "membrane surrounding hemozoin" has been observed in infected cells but has not been observed in in vitro experiments. This study focused on observing the association of phospholipid membranes and synthetic β-hematin, which is chemically identical to hemozoin, and on a further exploration into the mechanism of phospholipid membrane-induced β-hematin formation. Our results showed that β-hematin formation was induced by phospholipids in the fluid phase but not in the gel phase. The ability of phospholipids to induce β-hematin formation was inversely correlated with gel-to-liquid phase transition temperatures, suggesting an essential insertion of heme into the hydrocarbon chains of the phospholipid membrane to form β-hematin. For this study, a cryogenic transmission electron microscope was used to achieve the first direct observation of the formation of a monolayer of phospholipid membrane surrounding β-hematin.