| Freeze-drying provides an ideal method for long-term preservation of human platelets. By now, there have been a lot of experimental studies optimizing the lyophilization of human platelets, the cell morphology, ultra microstructure and some functions of hydrated platelets after freeze-dying still deviated from those of fresh cells, besides, the lyophilization of large volume of human platelets encountered inhomogeneous problems in heat and mass transfer. Thus, the thermophysical problemes are needed to be investigated. Aiming at the freezing and pre-hydration step of lyophilization of human platelets, the following thermophysical studies were carried out:1. The freezing temperatures Tm of ternary and quaternary solutions composed of water, NaCl and lyoprotectants including trehalose, BSA or HES were measured. The Polynomial Summation Method (PSM) and Osmotic Virial Equation (OVE) were used to calculate the Tm values of above solutions, and the differences between the calculated and measured values were smaller for PSM compared with OVE. A method of using the modified coeffient by fitting the Tm data of water/NaCl/non-electrolyte ternary solutions to calculated the Tm value of quaternary solutions were proposed, the average standard deviation was 0.46℃.2. The eutectic temperatures and concentrations, Teu and Weu, glass transition temperatures Tg, concentrations for maximumly freeze-concentrated solutions Wg’ and corresponding glass transition temperatures Tg’, ante-melting temperatures Tm’ of the above ternary and quaternary solutions were measured. The proportional relationship between the unfrozen water content and solute content at maximumly freeze-concentrated point was found. The assumption that the unfrozen water content corresponding to a specified solute was unaffected by other solutes in multi-component solutions was proposed, and the method to calculate Wg’, Tg’ and Tm’ of multi-component lyoprotectant solutions was put forward. The average standard deviation between calculated and measured Tg and Tm’ were 1.95℃ and 3.23℃ respectively.3. The mathematical model for freezing process of human platelet based on equations of phase diagrams of lyoprotectant solutions was established. The impacts of freezing rate, composition of lyoprotectant solution, loading concentrantion of intracelluar trehalose and concentration of permeable cryoprotectants, etc on the intracellular solution state were analysed:increasing initial osmolality of lyoprotectant solution would decrease the supercooling degree of intracellular solution; increasing intracellular concentration of trehalose would increase the supercooling degree and residual water content of intracellular solution; increasing concentration of DMSO would also increase the residual water content, and decrease the glass transition temperature of intracellular solution.4. The moisture sorption characteriscs of freeze-dried human platelets were studied, and the platelet cells which accounting for 5% of dry matter volume could bring significant influcence to the moisture sorption isotherm and isosteric heat of moisture sorption, the optimized average water content after pre-hydration was equal to the monolayer moisture content of freeze-dried plateltes. The moisture diffusion coefficient of freeze-dried platelets in vial was fitted with experimental data, and the impacts of enviromental temperature, humidity, thickness of freeze-dried platelets on pre-hydration process were modeled. Decreasing the relative humidity from 85% to 65% at 37℃ would increase the pre-hydration time from 48min to 93min, but the final moisture content distribution would be more uniform.The phase digram for multi-component lyoprotectant solutions, the simulated results of freezing process for human platelets, the moisture sorption characteristics of freeze-dried human platelets, and the numerical results of pre-hydration process were obtained in this thesis, thery were valuable for optimizing the freeze-drying and rehydration processes of human platelets. |
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