IN VIVO INVESTIGATIONS USING MICRODIALYSIS SAMPLING

该提案的目标是开发微透析采样， 液相色谱作为生物传感器能够连续监测 在体内的生物化学过程中， 系统 微透析采样是由一个短的透析长度 tubing（管）200 μ m OD），溶液被缓慢地泵送通过该孔。 小 样品中的分子扩散到探针中并被输送到 在收集小瓶中，较大的分子被膜排除在外。 在此期间，将研究微透析的四种应用 项目 首先，微透析技术将被开发用于 药代动力学研究。 通过在实验动物体内植入几个探针 药物在几个部位的药代动力学将同时 测定 例如，探针将被植入肝脏， 肌肉，静脉注射，以研究血液水平与 全身分布和肝脏代谢。 第二，微透析将 用于研究药物与血液蛋白的相互作用。 只有 药物的游离部分通过微透析取样， 药物不能通过透析膜扩散。 三、多探头 也将用于研究药物在血脑中的转运 屏障 最后，使用皮下植入的探针，透皮药物 交付将被调查。 该项目还将包括公共交通的基本方面 在微透析条件下穿过膜。 的质量 分析数据取决于对这些跨膜的理解。 流程. 分析物、膜、探针设计和 将对样品基质进行研究。 属性的影响，例如 电荷、疏水性和分子量对运输的影响将是 测定 其他参数，如样品粘度和离子强度， 跨膜的温度梯度、透析管长度和 直径也将进行研究。 最后，微透析分析系统将实现自动化。 经特殊 设计的双进样阀将用于连续样品采集 和注射。 这将使实验者从样品处理中解脱出来 任务，以便将全部注意力集中在生物方面， 本实验 为了进一步提高实验的分辨率， 微透析将直接与质谱联用。 的 微透析探针将提供适合于直接注射到微透析系统中的样品。 质谱仪将提供选择性来确定药物 及其代谢物。

The goal of this proposal is to develop microdialysis sampling coupled to liquid chromatography as a biosensor capable of continuously monitoring biochemical processes in vivo with minimal perturbation of the biological system. Microdialysis sampling is consists of a short length of dialysis tubing (ca. 200 mum OD) through which solution is slowly pumped. Small molecules in the sample diffuse into the probe and are transported to a collection vial, while larger molecules are excluded by the membrane. Four applications of microdialysis will be investigated during this project. First, microdialysis techniques will be developed for pharmacokinetic studies. By implanting several probes into a test animal the pharmacokinetics of a drug at several sites will be simultaneously determined. For example, probes will be implanted in the liver, the muscle, and intravenously to study the relationship of blood levels to systemic distribution and hepatic metabolism. Second, microdialysis will be used to study the interaction of drugs with blood proteins. Only the free fraction of the drug is sampled by microdialysis because protein bound drug can not diffuse through the dialysis membrane. Third, multiple probes will also be used to study transport of drugs across the blood-brain barrier. Finally, using probes implanted subcutaneously, transdermal drug delivery will be investigated. The project will also encompass fundamental aspects of mass transport across membranes under microdialysis conditions. The quality of the analytical data depends on an understanding of these transmembrane processes. Parameters of the analyte, the membrane, the probe design and the sample matrix will be investigated. The effect of properties such as charge, hydrophobicity and molecular weight on transport will be determined. Other parameters, such as sample viscosity and ionic strength, temperature gradients across the membrane, and dialysis tubing length and diameter will also be studied. Finally, the microdialysis analysis system will be automated. A specially designed dual injection valve will be used for continuous sample collection and injection. This will relieve the experimenter from sample handling tasks so that total attention can be focused on the biological aspects of the experiment. To further enhance the resolution of the experiment, microdialysis will be directly coupled to mass spectrometry. The microdialysis probe will provide a sample amenable to direct injection into the mass spectrometer which will provide the selectivity to determine drugs and their metabolites.