Capillary Electrophoresis: Bioanalytical Applications

描述(申请人提供)：这项研究的主要目标是发展毛细管电泳法(CE)作为一种分析技术来研究抗生素万古霉素(Van)、瑞索西汀(Ris)和替考拉宁(TEL)的物理化学参数。它的重点是演示原理和开发可用于分析受体-配体相互作用和微型反应的有用的生物分析方法。抗生素耐药细菌的进化使得VAN类抗生素的研究和开发成为研究的关键领域。因此，通过化学修饰来探索这些抗生素的物理化学参数的研究是非常必要的。这项工作中将使用两种特殊的毛细管电泳法：柱上微反应器技术和亲和毛细管电泳法。本研究所采用的技术包括化学修饰和表征、利用激光诱导荧光(LIF)和紫外/可见光(UV/VIS)检测方法和高效液相色谱(HPLC)对小分子进行化学分离和鉴定，以及酶动力学。这项研究将开发新的生物分析技术，并将重点放在检查涉及疾病预防和公共卫生问题的小生物分子上。这一理解将加强CE的设计、准备和应用的科学基础，以解决一系列与健康有关的问题。 本研究的具体目的是：1.利用柱上微反应器技术和ACE估计抗生素与配体之间的结合常数。用柱上微反技术测定抗生素的电荷和PL。用反应性化学试剂考察抗生素的衍生化反应动力学。IV.估算连接的抗生素与多肽的结合常数。进行D-丙氨酸-D-乳糖终止肽与VAN类抗生素结合的实验研究。 拟议的研究将：展示CE在确定抗生素的物理化学参数方面的多功能性；展示与ACE偶联的受体和配体的高通量衍生化；提供多个药物靶标的快速合成和准确分析；与补充分析技术相比，需要减少样本量；以及减少样品浪费和处置。

DESCRIPTION (provided by applicant): The broad goals of this research are to develop capillary electrophoresis (CE) as an analytical technique to study the physicochemical parameters of the antibiotics vancomycin (Van), ristocetin (Ris), and teicoplanin (Tel). Its focus is on demonstrating principles and on developing useful bioanalytical procedures that can be applied to the analysis of receptor-ligand interactions and microscale reactions. The evolution of antibiotic resistant bacteria has made the study and development of Van-like antibiotics a critical area of research focus. Hence, studies probing the physicochemical parameters of these antibiotics via chemical modification are highly warranted. Two specific CE techniques will be utilized in this work: on-column microreactor techniques and affinity capillary electrophoresis (ACE). Techniques utilized in this study include chemical modification and characterization, chemical separation and identification of small molecules by CE utilizing both laser-induced fluorescence (LIF) and ultraviolet/visible (UV/VIS) detection schemes and high performance liquid chromatography (HPLC), and enzyme kinetics. The research will develop new bioanalytical techniques and will focus on examining small biomolecules involved in the prevention of disease and, hence, to the issue of public health. This understanding will strengthen the scientific base underlying the design, preparation, and application of CE towards a host of health-related problems. The specific aims of the research are to I. Utilize On-Column Microreactor Techniques and ACE in the Estimation of Binding Constants Between Antibiotics and Ligands. II. Determine the Charge and pl of Antibiotics Using On-Column Microreactor Techniques. III. Examine the Kinetics of Derivatization of Antibiotics Using Reactive Chemical Reagents. IV. Estimate Binding Constants of Linked Antibiotics to Peptides. V. Conduct Experimental Studies on D-Ala-D-Lac Terminus Peptide Binding to Van-Group Antibiotics. The proposed research will: Demonstrate the versatility of CE in determining physicochemical parameters of antibiotics; Demonstrate high-throughput derivatization of receptors and ligands coupled to ACE; Provide for rapid synthesis and accurate analysis of multiple drug targets simultaneously; Require reduced sample volumes compared to complementary analytical techniques, and; Reduce sample waste and disposal.