Using Fluorescent Probes to Study the Location and Interaction of Disaccharides within Lipid Bilayers
Barker, Morgan Elizabeth
It is well known that certain organisms, such as the tardigrade, are capable of surviving extreme conditions including dehydration, freezing and / or oxygen deficiency. The tardigrades ability to withstand such extreme conditions is associated with a decrease in its internal water content and the excess production of a disaccharide known as trehalose. Studies have focused on better understanding this organism survival mechanism in order to help aid in the preservation of biological cells. Researchers have attempted to replicate this phenomenon in vitro through a freeze-drying technique known as lyopreservation and have shown trehalose to be an effective lyoprotecting agent. However, lyopreservation is very technically challenging so some studies have focused on trehalose has a potential cryoprotectant. Cryopreservation involves storing hydrated cells or other materials at very low temperatures in a frozen state. Unfortunately, trehalose did not show effectiveness as a cryoprotecting agent. This may be due to trehalose being a highly hydrophilic molecule which does not penetrate into the membrane to offer protection during freezing. Our research aims to investigate other disaccharides with varying hydrophilicity as potential cryoprotectants for hydrated cells. In order to study the effectiveness of certain disaccharides as cryoprotectants, liposomes were used as a model membrane system. The location and interactions of both sucralose and sucrose within these membranes were monitored through fluorescence and ultra-violet visible spectrophotometry using two specific fluorophores. The first of these, diphenylhexatriene, is a hydrophobic probe quenched by water that exhibits strong fluorescence when incorporated into a cell membrane. The second, merocyanine 540, is a negatively charged cyanine dye located slightly above the glycerol backbone of the phospholipids and is sensitive to any structural variations within the membrane. The data obtained with each respective probe indicates that the packing efficiency and polarity of the membrane is decreased in the presence of sucralose, but not in the presence of sucrose. We hypothesize that this is due to different hydrophobic properties of the sugars allowing sucralose to penetrate further into the bilayer while sucrose does not. This study indicates that sucralose has a significant effect on both the packing efficiency and the polarity of the bilayer and thus its usefulness as a potential cryoprotectant for biological cells is of great interest and needs to be investigated.
Barker, Morgan Elizabeth. (April 2016). Using Fluorescent Probes to Study the Location and Interaction of Disaccharides within Lipid Bilayers (Master's Thesis, East Carolina University). Retrieved from the Scholarship. (http://hdl.handle.net/10342/5336.)
Barker, Morgan Elizabeth. Using Fluorescent Probes to Study the Location and Interaction of Disaccharides within Lipid Bilayers. Master's Thesis. East Carolina University, April 2016. The Scholarship. http://hdl.handle.net/10342/5336. February 18, 2020.
Barker, Morgan Elizabeth, “Using Fluorescent Probes to Study the Location and Interaction of Disaccharides within Lipid Bilayers” (Master's Thesis., East Carolina University, April 2016).
Barker, Morgan Elizabeth. Using Fluorescent Probes to Study the Location and Interaction of Disaccharides within Lipid Bilayers [Master's Thesis]. Greenville, NC: East Carolina University; April 2016.
East Carolina University