IN VITRO STUDIES OF METABOLISM, ABSORPTION, AND TOXICITY

van Breeman的项目将通过评估活性植物化合物的体外和体内肝脏代谢、毒性、药代动力学和血脑屏障通透性，与该植物中心的所有其他项目和核心相互作用。这一信息将有助于为安全有效地使用植物性膳食补充剂以促进妇女健康提供合理依据。在Specific Aim 1中，Farnsworth和Bolton以及Analytical Core项目中鉴定的活性化合物的代谢将在体外使用人类和大鼠肝微粒体和肝细胞进行研究。对这些代谢产物的形成负责的人类酶的特定同工酶将被确定，这些化合物或它们的代谢产物是否抑制这些酶将被确定。这些研究将表明这些化合物有可能引起植物与药物的相互作用。在特定目标2中，本实验室开发的超滤LC-MS-MS分析将用于确定植物提取物中化合物的任何亲电性和可能有毒的代谢物是否可以

Project by van Breeman will interact with all the other Projects and Cores within this Botanical Center by evaluating in vitro and in vivo the hepatic metabolism, toxicity, pharmacokinetics, and blood-brain barrier permeability of active botanical compounds. This information will help provide a rational basis for the safe and effective use of botanical dietary supplements for women's health. In Specific Aim 1, the metabolism of active compounds identified in Projects by Farnsworth and Bolton and Analytical Core will be investigated in vitro using human and rat liver microsomes and hepatocytes. The specific isozymes of the human enzymes that are responsible for the formation of these metabolites will be identified and whether or not these compounds or their metabolites inhibit these enzymes will be determined. These studies will indicate the potential of these compounds to cause botanical-drug interactions. In Specific Aim 2, an ultrafiltration LC-MS-MS assay developed in this laboratory will be used to determine whether any electrophilic and possibly toxic metabolites of compounds in the botanical extracts can be formed in the liver. These studies will help predict toxicity at high doses. In addition, a new assay based on alkylation of the protein Keap1 and subsequent disruption of the Keap1-Nrf2 complex will be developed as a complementary assay. Since disruption of the Keap1-Nrf2 complex produces up-regulation of a variety of Phase II detoxification enzymes, exposure to low levels of botanical compounds that bind to Keap1 might actually produce a protective response. In Specific Aim 3, compounds in botanical extracts that are determined in Project by Bolton to have potential CNS effects will be assayed for blood-brain barrier permeability using a bovine brain epithelial cell monolayer model system. Finally in Specific Aim 4, the in vivo metabolism, bioavailability, and pharmacokinetics of active compounds will be investigated in rats and in a Phase I clinical trial in healthy post-menopausal women. These studies will provide information regarding the safety and pharmacokinetics of the active constituents in hops. Overall, these in vitro and in vivo studies of metabolism, metabolic activation, permeability, bioavailability, and pharmacokinetics will facilitate the design of future Phase II and III clinical trials of the safety and efficacy of these botanical dietary supplements.