Upgrade and repair of high performance liquid chromatography

高效液相色谱升级改造

• 批准号: RTI-2018-00367
• 负责人: Ro, DaeKyun
• 金额: $ 1.45万
• 依托单位: University of Calgary
• 依托单位国家: 加拿大
• 项目类别: Research Tools and Instruments
• 财政年份: 2017
• 资助国家: 加拿大
• 起止时间: 2017-01-01 至 2018-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=642174
• 关键词: Upgrade; repair; performance; liquid; chromatography

Isoprenoids are the largest class of natural products with more than 70,000 known structures. Despite their structural complexities, all isoprenoids are biosynthesized by a simple building block, five-carbon isopentenyl diphosphate (IPP). As many isoprenoids have direct implications in human society as pharmaceuticals, nutraceuticals, flavors, and polymers, how this simple IPP can be transformed to a diverse array of isoprenoids has been intensively studied. My research program focuses on elucidating biochemical mechanisms for the formations of diverse isoprenoids with an aim at providing knowledge for biotechnological productions of naturally scarce isoprenoids. Two major isoprenoids of interest in my laboratory are the biopolymer of natural rubber and pharmaceutically important sesquiterpene lactones. Natural rubber is a poly-cis-isoprene biopolymer, comprised of more than a thousand IPP monomers. Despite progresses in polymer chemistry, synthetic rubber derived from petroleum cannot fully replace natural rubber in manufacturing industries because the physical properties of synthetic rubber cannot surpass those of natural rubber (e.g., elasticity, heat dissipation, and abrasion resistance). Natural rubber is, therefore, an irreplaceable biomaterial synthesized from plant. On the other hand, sesquiterpene lactones form an important group of small isoprenoids, members of which have been used as anti-malarial (e.g., artemisinin) and anti-prostate cancer (e.g., thapsigargin) drug. However, most sesquiterpene lactones are present at low quantities in nature and their chemical synthesis is economically not viable due to their stereochemical complexities. The enzymatic synthesis of sesquiterpene lactones in surrogate hosts has emerged as an alternative biological production. However, the biochemical mechanism of how these isoprenoids are synthesized remains elusive, limiting biotechnological productions of these compounds. My research has identified a novel protein complex necessary for natural rubber biosynthesis, using lettuce as a model system. We have also elucidated a complete biosynthetic pathway of the simplest sesquiterpene lactone, costunolide. Advancing the biochemical knowledge for natural rubber and sesquiterpene lactone has been dependant on the routine use of an analytical instrument, high performance liquid chromatography (HPLC) coupled with a photodiode array detector (for sesquiterpene lactone analysis) and an evaporative light scattering detector (for natural rubber analysis). Reliable detection, quantitation, and purification of small or polymeric isoprenoids using HPLC are essential analytical practices. Timely upgrade and repair of HPLC are critically required to continue the metabolic studies of the isoprenoids.

类异型是最大的天然产品，具有超过70,000个已知结构。尽管具有结构上的复杂性，但所有类吸引者都是由简单的构建块（五碳等苯基二磷酸盐（IPP））生物合成的。由于许多类药物在人类社会中具有直接的影响，因为药物，营养素，口味和聚合物如何将这种简单的IPP转化为多种类异on-的类异on-，已经深入研究了。我的研究计划着重于阐明各种类异型素质形成的生化机制，目的是为自然稀缺的类异on子的生物技术生产提供知识。在我的实验室中，两个主要感兴趣的原子能是天然橡胶和药品重要的倍半萜烯内酯的生物聚合物。天然橡胶是一种聚-CIS-异丙烯生物聚合物，由超过一千个IPP单体组成。尽管聚合物化学的进展，但从石油衍生的合成橡胶不能完全替代制造业中的天然橡胶，因为合成橡胶的物理特性无法超过天然橡胶的物理特性（例如，弹性，散热和耐磨性）。因此，天然橡胶是从植物中合成的不可替代的生物材料。另一方面，倍半萜烯内酯构成了重要的小类吸异形，它们的成员已被用作抗癌症（例如青蒿素）和抗攻击癌（例如Thapsigargin）药物。但是，大多数倍半萜烯内酯在性质中存在低量，由于其立体化学复杂性，它们的化学合成在经济上是不可行的。替代宿主中倍半萜内酯的酶促合成已成为一种替代生物产生。但是，这些类化学的生化机制仍然难以捉摸，这限制了这些化合物的生物技术生产。我的研究已经确定了一种新型的蛋白质复合物，即使用生菜作为模型系统所必需的天然橡皮生物合成所需的蛋白质复合物。我们还阐明了最简单的倍半萜内酯，即Costunolide的完整生物合成途径。促进天然橡胶和倍半萜内酯的生物化学知识取决于分析仪器的常规使用，高性能液相色谱（HPLC）以及光电二极管阵列探测器（用于倍序烯烯烯烯烃分析）和蒸发光散射检测器（用于自然磨牙分析）。使用HPLC对小型或聚合物类异戊二烯的可靠检测，定量和纯化是必不可少的分析实践。迫切需要及时的升级和HPLC的修复才能继续进行类异急动物的代谢研究。