Rapid Identification of Active Agents and Metabolomics of Botanical Supplements

描述(申请人提供)：这项申请解决了高通量方法的未得到满足的需求，以确定植物膳食补充剂中的活性成分，并通过代谢组学研究确定其作用机制。超滤质谱(PUF-MS)是本实验室发明的，在筛选、表征、分离和鉴定植物膳食补充剂或其他天然产品混合物中具有重要药理意义的配体方面，它比传统的生物测定引导的分离更有效。我们建议使用一种新的超快速PUF-MS(UPUF-MS)方法将这项技术的生产率提高100倍，并开发新的经过验证的基于UHPLC-MS-MS的植物膳食补充剂标准化分析方法，其速度至少比以前快10倍。我们将把UHPLC-MS应用于代谢组学研究中，对来自临床试验的血清和尿液样本进行研究，研究植物性膳食补充剂黑升麻、红三叶、啤酒花、百忧解和安慰剂对更年期妇女潮热的影响。我们的假设是，这种代谢组学方法将有助于发现指示血管运动症状的生物标记物，如潮热和盗汗。然后，这些生物标志物可以用于客观、定量地确定植物性膳食补充剂对潮热的影响，而不是依赖于临床试验中受试者对潮热的主观自我报告。总体而言，这些研究将解决这一中心假设，即植物膳食补充剂含有药理活性化合物，可以使用UPUF-MS鉴定，用于UHPLC-MS-MS的化学标准化，以及这些活性成分的体内效应可以使用UHPLC-MS-MS与代谢组学相结合来检测。

DESCRIPTION (provided by applicant): This application addresses the unmet need for high-throughput methods to identify active components in botanical dietary supplements and to determine their mechanisms of action through metabolomics studies. Ultrafiltration mass spectrometry (PUF-MS), which was invented in this laboratory, is more efficient than conventional bioassay-guided fractionation for the screening, characterization, isolation, and identification of pharmacologically significant ligands contained in botanical dietary supplements or other natural product mixtures. We propose to enhance the productivity of this technology 100-fold using a new ultrafast PUF-MS (UPUF-MS) approach and to develop new validated assays for the standardization of botanical dietary supplements based on UHPLC-MS-MS that are at least 10-fold faster than previously possible. We will incorporate UHPLC-MS into metabolomics studies of archived serum and urine samples from clinical trials of the effects of botanical dietary supplements black cohosh, red clover, hops, Prempro, and placebo on hot flashes in menopausal women. It is our hypothesis that this metabolomics approach will facilitate the discovery of biomarkers indicative of vasomotor symptoms such as hot flashes and night sweats. Then, these biomarkers may be used for the objective, quantitative determination of the effects of botanical dietary supplements on hot flashes instead of relying on subjective self-reporting of hot flashes by subjects in clinical trials. Overall, these studies will address te central hypothesis that botanical dietary supplements contain pharmacologically active compounds that can be identified using UPUF-MS, used for chemical standardization with UHPLC-MS-MS and that the in vivo effects of these active constituents can be detected using UHPLC-MS-MS in combination with metabolomics.