Ice Recrystallization Inhibition and Ice-nucleating Promotion Activity of Bile Salts

The long-term storage of cells, tissues, and organs relies on cryopreservation, in which cryoprotectants are used to improve effectiveness. Ice recrystallization inhibitors are novel cryoprotectants. Low molecular weight ice recrystallization inhibitors have recently aroused interest in the cryopreservation field due to their potential to penetrate cells and tissues. Previous studies in this lab found a group of natural compounds – nanocellulose – contains potent ice recrystallization inhibition (IRI) activities and hypothesized its mechanism based on a structural amphiphilicity. Herein, the exploration was extended into a group of natural compounds found in the human body – bile salts. The major hypothesis is that the bile salts possess IRI activity based on their structural amphiphilicity. The aim of this thesis is to explore the IRI activity and ice-nucleating modification ability of bile salts and to understand the factors influencing such activity. Two representative bile salts were selected from each primary and secondary bile salts.

The IRI activities of four bile salts were quantified using the “splat” assay. Particularly, NaLC completely stopped ice crystal growth as low as 25 mM. In addition to IRI, the ice-nucleating promotion activity (INPA) of bile salts has been confirmed. The most IRI-active NaLC exhibited the highest INPA; however, the least IRI-active NaC also raised the ice nucleation temperature to a high extent. A series of parameters such as pH and ionic strength that could alter the amphiphilicity and micellization behavior of bile salts, influenced the IRI activity and ice-nucleating promotion activity. Ionic strength is crucial for bile salts to possess potent IRI activity but diminishes the activity of INPA. The IRI activity of bile salts examined here may provide a new direction for further cryopreservation study.