Kuchkov V. Distribution of endocellular cryoprotectants and water state in suspensions of mammalian erythrocytes prior to cryopreservation

The research object was physical and chemical processes in suspensions of mammalіаn erythrocytes, which take place after addition of cryoprotectants. The aim was studying water activity in the suspensions of human, bovine and equine erythrocytes and partition of dimethylsulfoxide, 1,2-propanediol and glycerol between the cells and extracellular medium. Research methods were cryoscopy, spectrophotometry, EPR spin probes, parametric and nonparametric analysis. The partition coefficients of cryoprotectants when it’s distribution between erythrocytes and extracellular medium are defined. This has allowed to establish concentration range wherein the essential contribution in partition is led by adsorption processes. This range studying has shown an existence of concentration range of cryoprotectants where the water activity in extracellular medium increases as a result of its replacement from hydrated coats of erythrocytes cellular components. The interaction mechanism of cryoprotectants with hydrated coats of erythrocytes components is offered. According to this, the cryoprotectants force out water molecules from hydrated coats of biomacromolecules, directly interacting with them. It is shown that the cryoprotectants addition to human, bovine and equine erythrocytes with concentration of 10 % w. leads to microviscosity decrease and change of a profile of temperature dependent rearrangements of dynamic cytosol structure. It is shown that increase of general water content in suspensions of human erythrocytes reduces hemolysis after effect endocellular cryoprotectants. The quantity of cryoprotectant which remains in suspension after washing out of cells is estimated.