A Device for Automating Titration Experiments in High-Resolution NMR Spectroscopy

描述(由申请人提供)：该项目的总体目标是开发一种坚固耐用且成本相对较低的设备，该设备可以使用现有的核磁共振探针和硬件对珍贵的核磁共振样品进行微型分析滴定。液态高分辨率核磁共振波谱已被证明是生物医学科学中非常有价值的工具，其应用包括生物大分子的结构和生物物理表征、小分子的物理化学表征以及复杂生物样品的分析，如人血清、尿液和匀浆组织提取液。一种广泛使用的实验方案涉及在二次化学或生物成分的逐步滴定过程中收集生物样品上的连续的核磁共振谱。常见的例子包括pH滴定中的酸或碱的滴定以测量大分子中特定位置的pKa值，标记的生物分子(例如蛋白质)与小分子或另一大分子的滴定以进行配体结合的生物物理表征，或在筛选药物开发的治疗靶点时滴定化学混合物。然而，目前进行这些核磁共振滴定的方法要么对大多数学术环境来说昂贵得令人望而却步，要么在实验工作或消耗往往非常珍贵的核磁共振样品材料方面效率低下。作为回应，我们建议开发一种低成本的设备，使用单个核磁共振样本自动进行核磁共振滴定，该设备可以适用于任何现有的商业高分辨率核磁共振光谱仪。我们的设计将两个电机驱动的非磁性注射器安装在标准核磁共振管架(目前用于将样本插入高场磁铁的中心孔)的边缘，并将包括两段连接到各自注射器的小直径管。在一种新的应用中，非磁压电马达将控制注射器，一个包含滴定剂的浓缩溶液，另一个专门用于样品混合。这种设备的可获得性将对各种各样的生物医学研究人员具有极高的价值，并将在基本上所有人类疾病的研究中得到应用。

DESCRIPTION (provided by applicant): The overall goal of the proposed project is to develop a robust and relatively low-cost device that can perform analytical micro-scale titrations on precious NMR samples using pre-existing NMR probes and hardware. Liquid-state, high resolution NMR spectroscopy has proven to be an extremely valuable tool in the biomedical sciences, with applications including the structural and biophysical characterization of biologic macromolecules, physicochemical characterization of small molecules, and the analysis of complex biologic specimens such as human serum, urine and homogenized tissue extracts. A widely utilized experimental protocol involves the collection of successive NMR spectra on a biological specimen during a stepwise titration of a secondary chemical or biological component. Common examples include titration of acid or base in a pH titration to measure site-specific pKa values in macromolecules, titration of a labeled biomolecule (e.g. protein) with a small molecule or another macromolecule for biophysical characterization of ligand-binding, or titrations of chemical mixtures during screening of therapeutic targets for drug development. However, current methodologies for performing these NMR titrations are either prohibitively costly for most academic settings or inefficient in terms of experimental effort or the consumption of often highly precious NMR sample material. In response, we are proposing to develop a low-cost device for automating NMR titrations using a single NMR sample, which could be adapted to any existing, commercial high-resolution NMR spectrometer. Our design fits two motor-driven, non-magnetic syringes to the edges of a standard NMR tube holder (currently used to insert the sample into central bore of the high field magnet), and will include two lengths of small diameter tubing connected to their respective syringes. In a novel application, non-magnetic Piezo-electric motorswill control the syringes, one containing a concentrated solution of titrant and the other dedicated to sample mixing. The availability of such a device would be highly valuable to a wide diversity of biomedical researchers and would have applications in the investigation of essentially all human diseases.