Research Initiation Award: Protein Separation Utilizing Utilizing Temperature-Sensitive Mecroemulsions

该提案研究旨在开发一种提取 从水溶液中回收蛋白质技术 利用温度敏感的微乳液相。 的 技术可用作初级回收步骤， 大规模生物分离。 利用液相质量 转移，初级隔离的停留时间可以是 最小化并消除蛋白酶作用。 微乳液 与两相水溶液相比， 用于这种和提取技术的系统，包括易于 阶段脱离（消除对资本的需求- 高强度离心机）和低粘度材料（允许 用于更快的质量传递）。 通过利用非离子 表面活性剂，温和的温度升高可用于 以浓缩形式回收蛋白质，而不是 利用目前用于离子的化学加成， 表面活性剂反胶束体系。 非离子温度- 敏感的微乳液需要掺入 亲和配体以获得高度特异性 需要的话 模型酶的可逆抑制剂，>- 半乳糖苷酶，将化学连接到乙烯 用于形成非离子表面活性剂的氧化物加合物 温敏微乳液。 的分割>- 将半乳糖苷酶加入配制的微乳液中， 作为温度的函数进行研究。 增溶 预期该酶在低温（4 ℃）下发生， 而破乳和酶回收将在 更高的温度（35 ℃），其远低于变性 - 半乳糖苷酶的温度（50%变性发生在 57>C）。 然后将该微乳液体系用于 研究温度敏感的加工优势， 在实验室规模的连续分离单元上进行微乳液的分离。

The proposal research is directed at developing an extraction technique to recover proteins from an aqueous solution utilizing a temperature-sensitive microemulsion phase. The technique may be utilized as a primary recovery step for large-scale bioseparations. By utilizing liquid phase mass transfer, residence times for primary isolation can be minimized and protease action eliminated. Microemulsions possess several processing advantages over two-phase aqueous systems for such and extraction technique, including ease of phase disengagement (eliminating the need for capital- intensive centrifuges) and low viscosity materials (allowing for faster mass transfer). By utilizing a nonionic surfactant, an mild temperature increase can be utilized to recover the protein in a concentrated form instead of utilizing chemical addition as currently practiced for ionic surfactant reverse micellar systems. Nonionic temperature- sensitive microemulsions require the incorporation of an affinity ligand to obtain the high degree of specificity desired. A reversible inhibitor of a model enzyme, >- galactosidase, will be chemically attached to the ethylene oxide adduct of the nonionic surfactant employed to form the temperature-sensitive microemulsion. The partitioning of >- galactosidase into the formulated microemulsion will be investigated as a function of temperature. Solubilization of the enzyme is expected to occur at a low temperature (4>C), while demulsification and enzyme recovery will occur at a higher temperature (35>C) which is well below the denaturation temperature for >-galactosidase (50% denaturation occurs at 57>C). This microemulsion system will then be utilized to study the processing advantages of a temperature-sensitive microemulsion on a bench-scale continuous separation unit.