CARBOHYDRATE FRACTIONATION FACILITY

本提案的目的是建立一个碳水化合物分馏 采用最新的HPLC技术快速分离和 定量单糖、低聚糖和 具有最大分辨率和灵敏度的糖肽。 该设施将 由三个HPLC系统组成，旨在分离分析或 基于电荷、大小、凝集素的碳水化合物的定量， 在高pH值下的结合性能和阴离子性能。 目前可供小组使用的资金将用于离职， 收费和规模。 要求Perkin Elmer HPLC系统专用于 用于凝集素亲和HPLC，其根据以下分离碳水化合物： 它们结合不同植物凝集素的能力 第三个系统在 该设施将是Dionex碳水化合物梯度HPLC系统，一种新的 最近开发的方法，它允许快速测定少至 0.1n摩尔复合碳水化合物和低聚糖的分离 相同组合物仅在单个糖苷键上不同。 的 Dionex系统包括脉冲安培检测器（PAD）， 非常灵敏，将检测10-100皮摩尔的碳水化合物。 第二 要求PAD与Rainin HPLC系统配合使用。 的这种方法 检测不能应用于监测凝集素亲和HPLC，因为高 在缓冲液中使用不同浓度的糖。 因此，连续 流动Berthold辐射探测器要求有效监测 这些柱子。 所述HPLC系统和检测器的组合 代表了“国家的最先进的”技术，在快速和灵敏的分析， 真核碳水化合物的复杂混合物， 有助于本文所述的研究计划。

The aim of this proposal is to establish a Carbohydrate Fractionation Facility that uses the latest HPLC techniques for rapidly separating and quantitating complex mixtures of monosaccharides, oligosaccharides and glycopeptides with maximal resolution and sensitivity. The facility will consist of three HPLC Systems designed to separate analytical or quantitative amounts of carbohydrates on the basis of charge, size, lectin- binding properties and anionic properties at high pH. A Rainin HPLC system presently available to the group will be used for separations based on charge and size. A Perkin Elmer HPLC system is requested to be dedicated for use in lectin-affinity HPLC which separates carbohydrates according to their abilities to bind different plant lectins. The third system in the facility will be the Dionex Carbohydrate Gradient HPLC system, a novel method developed recently, which allows rapid determination of as little as 0.1n mole complex carbohydrates and the separation of oligosaccharides with identical composition that differ only in a single glycosidic linkage. The Dionex system includes a pulsed amperometric detector (PAD) that is extremely sensitive and will detect 10-100 pmoles carbohydrate. A second PAD is requested for use with the Rainin HPLC system. This method of detection cannot be applied to monitoring lectin-affinity HPLC since high concentrations of sugars are used in the buffers. Therefore, a continuous flow Berthold radiation detector is requested for efficient monitoring of these columns. The combination of HPLC systems and detectors described represents "state-of-the-art" technology in rapid and sensitive analysis of complex mixtures of eukaryotic carbohydrates and would be enormously helpful to the research programs described herein.