Countercurrent chromatography (CCC), CCC/MS and LC/MS techniques in studies of phytoestrogens in plant extracts and human urine

• Main Author: Chen, Lijuan
• Published: Swansea University 2004
• Subjects: 572.072
• Online Access: https://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.636240

Phytoestrogens are members of classes of polyphenolic compounds synthesized by plants. They include isoflavones and other flavonoids, lignans, coumestanes and zearalenones. Photoestrogens are weak estrogen-like plant compounds that act similarly to the hormone estrogen, Over the last twenty years, interest in the physiological effects of phyto-estrogen containing plant substances has increased significantly, since it was demonstrated that the phytoestrogen components may play a potential role in the prevention and treatment of hormonally-dependent diseases, including breast and prostate cancer. This thesis describes studies of High Speed Countercurrent Chromatography and combined Liquid Chromatography-Mass Spectrometry techniques in the separation, purification, identification and quantitation of phytoestrogens in plants and human fluid. Chapter One provides a brief history and introduction to the principles and applications of high-speed countercurrent chromatography, liquid chromatography, mass spectrometry, and their combination of interfacing systems. Chapter Two investigates the separation and purification the flavonoids from the ethyl acetate extract from the seeds of O, indicum using high-speed countercurrent chromatography. Five flavonoids are successfully separated and purified with high purities (above 92 %) and two components are for the first time separated and identified from the plant. Mass spectrometry, Ultraviolet and Nuclear Magnetic Resonance are used for the structural elucidations of flavonoids. Chapter Three and Chapter Four compare different coil volumes of high-speed countercurrent chromatography instruments in the separation and purification of phytoestrogens. On line high speed countercurrent chromatography-mass spectrometry is studied for the separation and identification of flavones and isoflavones in plants. Chapter five develops a simple and accurate method for the quantitative determination of phytoestrogens in soy food supplements using on line liquid chromatography-mass spectrometry. Nine isoflavones are determined and liquid chromatography-tandem mass spectrometry is also investigated to the structural elucidation of some unknown components. Chapter Six develops a sensitive on line liquid chromatography-mass spectrometry method to determine the concentration of isoflavones and their major metabolites in human urine after consumption of soy-based supplements. The structural elucidations of some unknown metabolite isoflavones are also explored using liquid chromatography-tandem mass spectrometry.