Rapid Identification of Active Agents and Metabolomics of Botanical Supplements

描述（由申请人提供）：本申请解决了对高通量方法的未满足需求，以鉴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。 超滤质谱法（PUF-MS），这是在这个实验室发明的，是更有效的筛选，表征，分离和鉴定植物膳食补充剂或其他天然产物混合物中所含的重要配体比传统的生物测定指导分馏。我们建议使用新的超快PUF-MS（UPUF-MS）方法将该技术的生产率提高100倍，并开发新的经验证的测定方法，用于基于UHPLC-MS-MS的植物膳食补充剂的标准化，其速度至少比以前快10倍。我们将把超高效液相色谱-质谱结合到存档血清和尿液样本的代谢组学研究中，这些样本来自植物膳食补充剂黑升麻、红三叶草、啤酒花、Prempro和安慰剂对绝经期妇女潮热影响的临床试验。我们的假设是，这种代谢组学方法将有助于发现指示血管痉挛症状（如潮热和盗汗）的生物标志物。然后，这些生物标志物可用于客观、定量测定植物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，这些研究将解决中心假设，即植物膳食补充剂含有可使用UPUF-MS鉴定的生物活性化合物，用于UHPLC-MS-MS的化学标准化，并且这些活性成分的体内作用可使用UHPLC-MS-MS结合代谢组学检测。

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.