TY - CHAP

T1 - From Water to Ice

T2 - Investigation of the Effect of Ice Crystal Reduction on the Stability of Frozen Large Unilamellar Vesicles

AU - Siow, Lee Fong

AU - Rades, Thomas

AU - Lim, Miang Hoong

PY - 2010/5/14

Y1 - 2010/5/14

N2 - The solid phase of water, ice, is generally detrimental to cells because it causes freeze injury of cells during cryopreservation. Nonpermeable cryoprotective agents (CPAs) such as sucrose, trehalose, and glucose and/or permeable CPAs such as dimethyl sulfoxide (DMSO) and ethylene glycol (EG) are often added to freezing solutions to reduce such effects. The effect of ice formation on the stability of phospholipid bilayers was investigated using a model membrane: 1,2 - dipalmitoyl - rac - glycero - 3 -phosphocholine (DPPC) large unilamellar vesicles (LUVs) encapsulated with carboxyfl uorescein (CF) solution. The LUV dispersion was added with a CPA, cooled to 40 ° C, and heated to 20 ° C at 10 ° C/min. LUV stability was described by the degree of CF leakage. In the presence of sugars, DMSO, or EG, ice formation was reduced. Simultaneous to the ice reduction, an increase of unfrozen fraction prevented LUV aggregation and leakage. The leakage was observed to decrease above a defi ned sugar concentration and as the concentration of DMSO or EG increased. Below the defi ned sugar concentration, LUV leakage increased with increasing sugar concentrations. Nonpermeable CPAs were more effective for the frozen LUVs compared to permeable CPAs. The current result suggests that LUVs were spaced out from one another in the presence of nonpermeable CPAs. On the other hand, LUVs in 10% (wt/wt) of permeable CPAs could probably undergo structural destabilization.

AB - The solid phase of water, ice, is generally detrimental to cells because it causes freeze injury of cells during cryopreservation. Nonpermeable cryoprotective agents (CPAs) such as sucrose, trehalose, and glucose and/or permeable CPAs such as dimethyl sulfoxide (DMSO) and ethylene glycol (EG) are often added to freezing solutions to reduce such effects. The effect of ice formation on the stability of phospholipid bilayers was investigated using a model membrane: 1,2 - dipalmitoyl - rac - glycero - 3 -phosphocholine (DPPC) large unilamellar vesicles (LUVs) encapsulated with carboxyfl uorescein (CF) solution. The LUV dispersion was added with a CPA, cooled to 40 ° C, and heated to 20 ° C at 10 ° C/min. LUV stability was described by the degree of CF leakage. In the presence of sugars, DMSO, or EG, ice formation was reduced. Simultaneous to the ice reduction, an increase of unfrozen fraction prevented LUV aggregation and leakage. The leakage was observed to decrease above a defi ned sugar concentration and as the concentration of DMSO or EG increased. Below the defi ned sugar concentration, LUV leakage increased with increasing sugar concentrations. Nonpermeable CPAs were more effective for the frozen LUVs compared to permeable CPAs. The current result suggests that LUVs were spaced out from one another in the presence of nonpermeable CPAs. On the other hand, LUVs in 10% (wt/wt) of permeable CPAs could probably undergo structural destabilization.

KW - Cryopreservation - usage in biopreservation, preserving organisms at low temperature

KW - Cryoprotective agents (CPAs) addition - improving cell survival during cryopreservation

KW - Effect of ice crystal reduction - stability of frozen large unilamellar vesicles

KW - Ice - detrimental to cells, freezing injury of cells during cryopreservation

KW - Increase of unfrozen fraction - preventing LUV aggregation and leakage

KW - Permeable CPAs - suppressing intracellular ice formation

KW - Successful cryopreservation - survival of the frozen organisms after warming ice

UR - http://www.scopus.com/inward/record.url?scp=84886140096&partnerID=8YFLogxK

U2 - 10.1002/9780470958193.ch49

DO - 10.1002/9780470958193.ch49

M3 - Book chapter

AN - SCOPUS:84886140096

SN - 9780813812731

SP - 551

EP - 562

BT - Water Properties in Food, Health, Pharmaceutical and Biological Systems

PB - Wiley-Blackwell

ER -