Polarized infrared spectroscopy imaging applied to structural analysis of bilirubin aggregate at liquid-liquid interface.

Fourier transform infrared spectroscopy imaging (FTIRSI) combined with spectral analysis and polarization approach was creatively used to investigate both structures of bilirubin (BR) precipitate and BR aggregate at liquid-liquid interface. It was found by spectral analysis that the internal hydrogen bonds of BR molecules all broke and the dihedral angles increased during the formation of BR aggregate at liquid-liquid interface. And the BR molecule might be of layer assembly along the long axis direction of CD half-group to form J-type aggregates, which could be parallel to the direction of the transition dipole moment of BR aggregate. The further study of polarized imaging/anisotropy revealed that the absorbance of 1570 and 1703 cm bands of BR aggregate changed periodically at intervals of 90°, which were not shown in BR precipitate case, indicating that the CC of the corresponding lactam ring and the CO of the adjacent carboxyl groups formed ordered arrangement in BR aggregate. It also suggested that the two positions might be the active sites which J-type aggregates assembled on. The combined technique was firstly applied in interfacial aggregate research, which was helpful for further understanding and controlling the aggregation as well as structural transformation of BR molecules so as to decrease physiological hazard and facilitate the wide spread application in biomedicine.