Preparation of Nitrogen Analogues of Ceramide and Studies of Their Aggregation in Sphingomyelin Bilayers.

Ceramides can regulate biological processes probably through the formation of laterally segregated and highly packed ceramide-rich domains in lipid bilayers. In the course of preparation of its analogues, we found that a hydrogen-bond-competent functional group in the C1 position is necessary to form ceramide-rich domains in lipid bilayers [Matsufuji; 2018]. Hence, in the present study, we newly synthesized three ceramide analogues: CerN, CerNH, and CerNHAc, in which the 1-OH group of ceramide is substituted with a nitrogen functionality. CerNH and CerNHAc are capable of forming hydrogen bonds in their headgroups, whereas CerN is not. Fluorescent microscopy observation and differential scanning calorimetry analysis disclosed that these ceramide analogues formed ceramide-rich phases in sphingomyelin bilayers, although their thermal stability was slightly inferior to that of normal ceramides. Moreover, wide-angle X-ray diffraction analysis showed that the chain packing structure of ceramide-rich phases of CerNHAc and CerN was similar to that of normal ceramide, while the CerNH-rich phase showed a slightly looser chain packing due to the formation of CerNH. Although the domain formation of CerN was unexpected because of the lack of hydrogen-bond capability in the headgroup, it may become a promising tool for investigating the mechanistic link between the ceramide-rich phase and the ceramide-related biological functions owing to its Raman activity and applicability to click chemistry.