published_or_final_version / Chemistry / Master / Master of Philosophy

To Kin Wah. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (p. 293-345). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

by Fan Kai Leung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references (leaves [87-96]). / Abstract / Acknowledgement / Chapter Chapter 1 --- Introduction / Chapter 1.0 --- Foreword / Chapter 1.1 --- Project Objective / Chapter 1.2 --- Historical Background / Chapter 1.3 --- The properties of Radial Pulse Wave-An Engineering View / Chapter 1.4 --- The development in RPW acquisition and research-An Overview / Chapter 1.5 --- Problems to be solved for RPW data acquisition system / Chapter 1.6 --- The contribution of this project / Chapter Chapter 2 --- System Realisation / Chapter 2.0 --- Foreword / Chapter 2.1 --- System Development-An Overview / Chapter Chapter 3 --- Signal Preprocessing Unit / Chapter 3.0 --- Foreword / Chapter 3.1 --- Earlier Methods for offset Cancellation / Chapter 3.2 --- Feedback control of the offset and d.c. transient cancellation / Chapter 3.3 --- The level shifting circuit / Chapter 3.4 --- Amplifier / Chapter 3.6 --- The 50Hz notch filter / Chapter 3.7 --- The 4th order low pass filter / Chapter 3.8 --- Circuit for controlling valves and pump / Chapter Chapter 4 --- Signal Processing for RPW Acquisition / Chapter 4.0 --- Foreword / Chapter 4.1 --- Artefact and signal noise / Motion artefact / Respiratory Artefact / Base line shift due to tissue displacement / Electromagnetic Interference and circuit noise / Chapter 4.2 --- Artefact Correction by the Switch Capacitive Device(MF-6) / Operating principle / Result and Discussion / Chapter 4.3 --- Artefact Correction by Recursive Moving Average(RMA) / The RMA Operation / Software Implementation of RMA / Chapter 4.4 --- Further Improvement of the RMA baseline estimator / Chapter 4.5 --- Signal Noise Suppression by Non-linear filtering / Chapter 4.6 --- An arithmetic cycle detector for RPW and ECG / Chapter Chapter 5 --- The Software for the DAS / Chapter 5.0 --- Foreword / Chapter 5.1 --- The present DAS software / Chapter 5.2 --- The software design / Chapter 5.3 --- Important points about the software / Signal input / Waveform display and smooth scrolling / Dynamic display area allocation / The software FIFO data buffer / Chapter Chapter 6 --- Recapitulation and Topics for Further Investigation / Chapter 6.0 --- Foreword / Chapter 6.1 --- Recapitulations / Objective / Difficulties / System Realisation / Signal Preprocessing Unit / Signal Processing / The software / Chapter 6.2 --- Direction for further investigation / Improvement to the DAS / Data analysis and modelling research / Appendix A The Comparasion of the RMA and SMA Characteristics / Appendix B List of Publications

by Kok Dick Shun, Louis. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references. / Acknowledgements --- p.I / Table of Contents --- p.II / Abbreviations --- p.V / Aim and Scope of This Dissertation --- p.IX / Abstract --- p.X / Chapter Chapter One： --- General Introduction --- p.1 / Chapter 1.1 --- An Overview of the Immune System --- p.2 / Chapter 1.1.1 --- Innate Immunity --- p.2 / Chapter 1.1.2 --- Adaptive Immunity --- p.3 / Chapter 1.1.2.1 --- Humoral antibody immune response --- p.4 / Chapter 1.1.2.2 --- Cell- mediated immune response --- p.5 / Chapter 1.2 --- Immunomodulation --- p.6 / Chapter 1.3 --- An overview of the Host-mediated response against tumours --- p.9 / Chapter 1.3.1 --- T and B lymphocytes --- p.9 / Chapter 1.3.2 --- M acrophages --- p.14 / Chapter 1.3.3 --- Natural killer cells --- p.17 / Chapter 1.3.4 --- Lymphokines-activated killer cells --- p.20 / Chapter 1.3.5 --- Tumour infiltrating lymphocytes --- p.22 / Chapter 1.3.6 --- Cytokines --- p.23 / Chapter 1.4 --- Carbohydrates as Potential Immunostimulating agents --- p.33 / Chapter 1.5 --- General Properties of Bupleuri radix (B.R.) --- p.35 / Chapter Chapter Two： --- Materials and Methods --- p.36 / Chapter 2.1 --- Materials --- p.37 / Chapter 2.1.1 --- Animals --- p.37 / Chapter 2.1.2 --- Bupleuri radix --- p.37 / Chapter 2.1.3 --- "Buffers, culture media and chemicals" --- p.37 / Chapter 2.1.4 --- Cell lines --- p.48 / Chapter 2.2 --- Methods --- p.49 / Chapter 2.2.1 --- Extraction and fractionation of Bupleuri radix --- p.49 / Chapter 2.2.2 --- Purification of Bupleuri radix --- p.54 / Chapter 2.2.3 --- Characterization of Bupleuri radix --- p.55 / Chapter 2.2.4 --- In vivo Drug Treatment --- p.59 / Chapter 2.2.5 --- Isolation and preparation of cells --- p.59 / Chapter 2.2.6 --- Assays for the immunomodulatory activities of Bupleuri radix --- p.62 / Chapter 2.2.7 --- Assays for the immunorestorative properties of Bupleuri radix --- p.74 / Chapter 2.2.8 --- Assays for the anti-tumour activities of Bupleuri radix --- p.75 / Chapter 2.2.9 --- Statistical analysis --- p.83 / Chapter Chapter Three： --- "Fractionation, Purification and Characterization of Bioactive Compounds from Bupleuri radix" --- p.84 / Chapter 3.1 --- Results / Chapter 3.1.1 --- Extraction and Fractionation of Bupleuri radix --- p.85 / Chapter 3.1.2 --- Purification of Bupleuri radix --- p.85 / Chapter 3.1.3 --- Carbohydrate and Protein Contents of B.R. Fractions --- p.87 / Chapter 3.1.4 --- Lack of cytotoxicity of Bupleuri radix to Mouse Splenocytes --- p.91 / Chapter 3.1.5 --- LC50 of B.R. Fractions determined by Brine Shrimp Bioassay --- p.91 / Chapter 3.1.6 --- Heat stability of B.R. Fractions --- p.93 / Chapter 3.1.7 --- "Uronic Acid Content of BRIai, BRIaii, BRIbi and BRIbii" --- p.93 / Chapter 3.2 --- Discussion --- p.93 / Chapter Chapter Four： --- The Immunomodulatory Activities of Bupleuri radix --- p.96 / Chapter 4.1 --- Results / Chapter 4.1.1 --- Effect of Bupleuri radix on the Specific and Nonspecific Immunity --- p.97 / Chapter 4.1.1.1 --- Mitogenic effect of B.R. Fractions on Murine Splenocytes in vitro --- p.97 / Chapter 4.1.1.2 --- Mitogenic effect of B.R. Fractions on Murine Splenocytes ex vivo --- p.97 / Chapter 4.1.1.3 --- In vitro Mitogenic effect of B.R. Fractions treated with Periodate --- p.103 / Chapter 4.1.1.4 --- In vitro Mitogenic effect of B.R. Fractions treated with Acetic Acid --- p.103 / Chapter 4.1.1.5 --- In vitro Co -mitogenic effect of B.R. Fractions with Polymyxin B Sulphate --- p.107 / Chapter 4.1.1.6 --- Effect of B.R. Fractions on Lymphocyte sub-populations --- p.107 / Chapter 4.1.1.7 --- Primary Humoral Immune Response to SRBC in B.R.-treated mice --- p.107 / Chapter 4.1.1.8 --- Activity of cytotoxic T cells in B.R-treated mice --- p.111 / Chapter 4.1.1.9 --- Effect of B.R. Fractions on Interleukin-1 - like Factors Production --- p.111 / Chapter 4.1.1.10 --- Effect of B.R. Fractions on Interleukin-2 Production --- p.116 / Chapter 4.1.1.11 --- Effect of B.R. Fractions on Interleukin-2 Receptor Expression on Murine Splenocytes --- p.116 / Chapter 4.1.1.12 --- Effect of B.R. Fractions on GM-CSF Production --- p.119 / Chapter 4.1.1.13 --- Immunopotentiating effects of B.R. Fractions on Macrophages: --- p.119 / Chapter 4.1.1.13.1 --- In vivo Migration of Macrophages in B.R.-treated mice --- p.119 / Chapter 4.1.1.13.2 --- Effect of B.R. Fractions on the Fc Receptor Expression on Murine Resident Peritoneal Exudate Cells --- p.123 / Chapter 4.1.2 --- Immunorestorative Properties of Bupleuri radix --- p.123 / Chapter 4.1.2.1 --- Effect of B.R. Fractions on Lymphocyte Blastogenesis in Aged Mice --- p.123 / Chapter 4.1.2.2 --- Effect of B.R. Fractions on Lymphocyte Blastogenesis in Tumour-bearing Mice --- p.125 / Chapter 4.2 --- Discussion --- p.125 / Chapter Chapter Five： --- The Anti-tumour Activities of Bupleuri radix --- p.132 / Chapter 5.1 --- Results / Chapter 5.1.1 --- Cytostatic Effect of B.R. Fractions on Murine Tumour Cell Lines in vitro --- p.133 / Chapter 5.1.2 --- Effect of B.R. Fractions on the Growth of Tumour Ceils in vivo --- p.133 / Chapter 5.1.3 --- Effect of B.R. Fractions on the Survival of EAT-bearing mice --- p.140 / Chapter 5.1.4 --- Ex vivo Induction of Natural Killer Cell Activity by B.R. Fractions --- p.146 / Chapter 5.1.5 --- In vitro Induction of Lymphokine-activated Killer Cell Activity by B.R Fractions --- p.149 / Chapter 5.1.6 --- In vivo Induction of Tumour Infiltrating Lymphocytes by B.R. Fractions --- p.149 / Chapter 5.1.7 --- In vitro Induction of Macrophage-mediated Cytostatic Effect on Tumour Cells by B.R. Fractions --- p.151 / Chapter 5.1.8 --- In vitro Induction of Macrophage-mediated Cytostatic Eifect on Tumour Cells by B.R. Fractions --- p.153 / Chapter 5.1.9 --- Effect of B.R. Fractions on γ-interferon Production in vitro --- p.156 / Chapter 5.2 --- Discussion --- p.156 / Chapter Chapter Six： --- "General Discussion, Conclusion and Future Prospects" --- p.164 / Bibliography --- p.i

Leung Yiu Kwong, Michael. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 155-161). / Acknowledgements --- p.i / Abbreviations --- p.ii / Aim and scope of this dissertation --- p.v / Abstract --- p.vi / Table of contents --- p.viii / Introduction --- p.1 / Chapter 1.1 --- Introduction --- p.2 / Chapter 1.2 --- Tumor Biology --- p.3 / Chapter 1.3 --- The Defence Mechanisms --- p.4 / Chapter 1.3.1 --- Non-specific defence mechanisms --- p.5 / Chapter 1.3.2 --- Specific defence mechanisms --- p.6 / Chapter 1.4 --- Effector Mechanisms in Anti-tumor Immunity --- p.7 / Chapter 1.4.1 --- B-cell --- p.8 / Chapter 1.4.2 --- "Natural killer (NK) cells (Non-T, Non-B)" --- p.8 / Chapter 1.4.3 --- Macrophages --- p.9 / Chapter 1.4.4 --- Cytolytic T-lymphocytes (CTLs) --- p.10 / Chapter 1.5 --- Cancer Treatment --- p.10 / Chapter 1.5.1 --- Surgery --- p.10 / Chapter 1.5.2 --- Radiotherapy --- p.12 / Chapter 1.5.3 --- Drug therapy --- p.12 / Chapter 1.5.4 --- Gene therapy --- p.13 / Chapter 1.5.5 --- lmmunotherapy --- p.13 / Chapter 1.6 --- Non-cytotoxic Antitumor Polysaccharides of Fungi --- p.14 / Chapter 1.6.1 --- Yeast polysaccharides --- p.14 / Chapter 1.6.2 --- Lichen polysaccharides --- p.15 / Chapter 1.6.3 --- Fungal polysaccharides --- p.18 / Chapter 1.7 --- Fungi and their Polysaccharides --- p.20 / Chapter 1.7.1 --- Reserve carbohydrates --- p.20 / Chapter 1.7.2 --- Structural polysaccharides --- p.21 / Chapter 1.8 --- The Architecture of the Fungal Cell Wall --- p.22 / Materials and Methods --- p.26 / Chapter 2.1 --- Materials --- p.27 / Chapter 2.1.1 --- Animals --- p.27 / Chapter 2.1.2 --- Mushrooms --- p.27 / Chapter 2.1.3 --- "Buffers, culture media and chemicals" --- p.27 / Chapter 2.1.4 --- Cell lines --- p.34 / Chapter 2.2 --- Methods --- p.35 / Chapter 2.2.1 --- Screening of β-(l→3)-D-glucan --- p.35 / Chapter 2.2.2 --- Extraction and Fractionation of Flammulina velutipes --- p.35 / Chapter 2.2.3 --- Characterisation of Flammulina velutipes --- p.38 / Chapter 2.2.3.1 --- The determination of carbohydrate content of F.V fractions --- p.38 / Chapter 2.2.3.2 --- The determination of protein content of F.V. fractions --- p.39 / Chapter 2.2.3.3 --- The determination of uronic acid content of F.V fractions --- p.39 / Chapter 2.2.3.4 --- The determination of component sugar units of F.V fractions --- p.39 / Chapter 2.2.3.5 --- Periodate uptake of F.V. fractions --- p.40 / Chapter 2.2.3.6 --- Limulus amebocyte lysate (LAL) coagulation assay --- p.40 / Chapter 2.2.3.7 --- The digestion of F.V. fractions with laminarinase --- p.41 / Chapter 2.2.3.8 --- The Secondary and tertiary structure determination of FH and SFA1 --- p.42 / Chapter 2.2.3.9 --- Molecular weight estimation of FH and SFA1 --- p.43 / Chapter 2.2.3.10 --- Vascular dilation and hemorrhage (VDH) activity of F.V. fractions / Chapter 2.2.4 --- Isolation and preparation of cells --- p.43 / Chapter 2.2.4.1 --- Bone marrow cell --- p.43 / Chapter 2.2.4.2 --- Peritoneal exudate cell (PEC) --- p.44 / Chapter 2.2.4.3 --- Splenocytes --- p.44 / Chapter 2.2.4.4 --- Depleting mouse T-cells by anti-mouse T-cell antigen antibody plus complement treatment --- p.45 / Chapter 2.2.4.5 --- Depleting mouse B-cells by Cedarlane column kit --- p.45 / Chapter 2.2.5 --- Assays for the cytotoxicity of Flammulina velutipes --- p.45 / Chapter 2.2.5.1 --- Brine shrimp assay --- p.45 / Chapter 2.2.5.2 --- In vitro cytotoxicity of FH and SFA1 on bone marrow cells of female BALB/c mice --- p.46 / Chapter 2.2.5.3 --- In vivo cytotoxicity of FH and SFA1 on female BALB/c mice --- p.47 / Chapter 2.2.6 --- "Assays for the immunomodulatory activities of Flamm""lina velutipes" --- p.47 / Chapter 2.2.6.1 --- In vitro mitogenic activities of FH and SFA1 on murine lymphocytes --- p.47 / Chapter 2.2.6.2 --- In vitro mitogenic activities of FH and SFA1 with PMB on murine lymphocytes --- p.48 / Chapter 2.2.6.3 --- In vitro mitogenic activities of FH and on T-cell depleted murine lymphocytes --- p.48 / Chapter 2.2.6.4 --- In vitro mitogenic activities of FH on B-cell depleted murine lymphocytes --- p.49 / Chapter 2.2.6.5 --- In vitro co-mitogenic activitiy of FH and SFA1 on murine lymphocytes --- p.49 / Chapter 2.2.6.6 --- In vitro mitogenic activities of FH and SFA1 on murine bone marrow cells --- p.50 / Chapter 2.2.6.7 --- In vivo mitogenic activities of FH and SFA1 on murine lymphocytes --- p.50 / Chapter 2.2.6.8 --- Effect of FH and SFA1 on the enhancement of first antibody production of SRBC immunised mice --- p.51 / Chapter 2.2.6.9 --- Effect of FM and SFA1 on the in vitro phagocytic activity of murine macrophage --- p.51 / Chapter 2.2.6.10 --- Effect of FM and SFA1 on the in vivo phagocytic activity of murine macrophage --- p.51 / Chapter 2.2.6.11 --- In vivo migration of macrophage in FH- and SFAl-treated mice --- p.53 / Chapter 2.2.6.12 --- Effect of FH and SFA1 on the enhancement of murine PEC cytostatic activity --- p.53 / Chapter 2.2.6.13 --- Effect of FH and SFA1 on the Fc receptor expression of peritoneal exudate cells --- p.54 / Chapter 2.2.6.14 --- Effect of FH and SFA1 on murine serum cytokine level --- p.55 / Chapter 2.2.6.15 --- Effect of FH and SFA1 on murine serum TNF level --- p.55 / Chapter 2.2.6.16 --- Effect of FH and SFA1 on the augmentation of SRBC lysing ability of murine serum --- p.56 / Chapter 2.2.7 --- Assays for the anti-tumor activities of Flammulina velutipes --- p.57 / Chapter 2.2.7.1 --- In vitro anti-tumor activity of FH and SFA1 --- p.57 / Chapter 2.2.7.2 --- Effect of FH and SFA1 on the growth of murine transplantable tumor invivo --- p.58 / Chapter 2.2.8 --- Statistical analysis --- p.59 / "Screening, Purification, Fractionation and Characterisation of β-(l→3)-D-glucan(s) from Flammulina velutipes" --- p.60 / Introduction --- p.61 / Results --- p.62 / Chapter 3.1 --- Screening of β-(l→3)-D-Glucan --- p.62 / Chapter 3.2 --- Extraction and Fractionation of Flammulina velutipes --- p.62 / Chapter 3.3 --- The Determination of Carbohydrate Content of F.V. Fractions --- p.65 / Chapter 3.4 --- The Determination of Protein Content of F.V. Fractions --- p.65 / Chapter 3.5 --- The Determination of Uronic Acid Content of F.V. Fractions --- p.69 / Chapter 3.6 --- The Determination of Component Sugar Units of F.V. Fractions --- p.69 / Chapter 3.7 --- Periodate Uptake of F.V. Fractions --- p.69 / Chapter 3.8 --- Limulus Amebocyte Lysate (LAL) Coagulation Assay --- p.73 / Chapter 3.9 --- The Digestion of F.V. Fractions with Laminarinase --- p.73 / Chapter 3.10 --- The Secondary and tertiary Structure Determination of FH and SFA1 --- p.80 / Chapter 3.11 --- Molecular Weight Estimation of FH and SFA1 --- p.82 / Chapter 3.12 --- "Vascular Dilation and Hemorrhage (VDH) Activity of FH, SFA1 and lFA1" --- p.82 / Discussion --- p.90 / The Toxicity of Flammulina velutipes --- p.96 / Introduction --- p.97 / Results --- p.97 / Chapter 4.1 --- Lack of Cytotoxicity of Flammulina velutipes to Brine Shrimp --- p.97 / Chapter 4.2 --- Lack of Cytotoxicity of Flammulina velutipes to Murine Bone Marrow Cells --- p.99 / Chapter 4.3 --- Lack of Cytotoxicity of Flammulina velutipes to Mouse --- p.99 / Discussion --- p.102 / The Immunomodulatory Activities of Flammulina velutipes --- p.103 / Introduction --- p.104 / Results --- p.105 / Chapter 5.1 --- Effect of Flammulina velutipes on Murine Lymphocytes --- p.105 / Chapter 5.2 --- Effect of Flammulina velutipes on Murine Macrophage --- p.115 / Chapter 5.3 --- Effect of Flammulina velutipes on Murine Serum Cytokine and Complement Level --- p.125 / Discussion --- p.133 / The Anti-tumor Activities of Flammulina velutipes --- p.136 / Introduction --- p.137 / Results --- p.137 / Chapter 6.1 --- In Vitro Anti-Tumor Activity of FH and SFA1 --- p.137 / Chapter 6.2 --- Effect of FH and SFAI on the Growth of Murine TransplantableTumors --- p.138 / Discussion --- p.145 / General Discussion --- p.146 / General Discussion and Future Perspectives --- p.147 / References --- p.154

Ha Wai-Yan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (leaves 166-180). / Abstracts in English and Chinese. / Acknowledgements --- p.ii / Abstract --- p.iii / Abbreviations --- p.vi / Table of Contents --- p.vii / Chapter Chapter 1 --- General Introduction --- p.1 / Chapter 1.1 --- "Hstory, cultivation and trade" --- p.2 / Chapter 1.2 --- Botany --- p.4 / Chapter 1.3 --- Chemical Constituents and Pharmacological effects --- p.8 / Chapter 1.4 --- Authentication of Chinese herbal materials --- p.13 / Chapter 1.4.1 --- Morphological marker --- p.15 / Chapter 1.4.2 --- Histological marker --- p.18 / Chapter 1.4.3 --- Chemical marker --- p.20 / Chapter 1.4.4 --- Molecular markers --- p.24 / Chapter 1.4.4.1 --- Protein marker --- p.24 / Chapter 1.4.4.2 --- DNA-based markers --- p.26 / Chapter 1.4.4.2.1 --- PCR-based markers --- p.27 / Chapter 1.4.4.2.1.1 --- Random-primed PCR --- p.28 / Chapter 1.4.4.2.1.2 --- Simple Sequence Repeats (SSR) --- p.30 / Chapter 1.4.4.2.1.3 --- Polymerase Chain Reaction Fragment Length Polymorphism (PCR-RFLP) --- p.31 / Chapter 1.4.4.2.2 --- Hybridization-based markers --- p.33 / Chapter 1.4.4.2.3 --- Sequencing-based markers --- p.35 / Chapter 1.5 --- Objectives and Strategies of the studies --- p.39 / Chapter Chapter 2 --- General Materials and Methods --- p.40 / Chapter 2.1 --- Reagents and Buffers --- p.41 / Chapter 2.1.1 --- Media for bacterial culture --- p.41 / Chapter 2.1.2 --- Reagents for preparation of competent cells --- p.42 / Chapter 2.1.3 --- Reagents for plasmid DNA preparation --- p.42 / Chapter 2.1.4 --- Reagents for agarose gel electrophoresis --- p.43 / Chapter 2.1.5 --- Reagents for polyacrylamide gel electrophoresis --- p.43 / Chapter 2.1.6 --- Reagents for Southern hybridization --- p.44 / Chapter 2.2 --- Agarose Gel electrophoresis of DNA --- p.46 / Chapter 2.3 --- Purification of PCR products --- p.46 / Chapter 2.3.1 --- From agarose gel using Geneclean® II kit --- p.46 / Chapter 2.3.2 --- Using Microspin´ёØ Column --- p.47 / Chapter 2.4 --- End modification of PCR amplified DNA --- p.47 / Chapter 2.5 --- Preparation of Escherichia coli Competent Cells --- p.48 / Chapter 2.6 --- "Ligation and Transformation of E, coli" --- p.49 / Chapter 2.7 --- Plasmid Preparation --- p.50 / Chapter 2.7.1 --- Minipreparation of plasmid DNA --- p.50 / Chapter 2.7.2 --- Preparation of plasmid DNA using Wizard® Plus SV Minipreps DNA Purification Kit (Promega) --- p.50 / Chapter 2.8 --- Screening for the Presence of insert in plasmid --- p.51 / Chapter 2.8.1 --- Rapid alkaline lysis --- p.51 / Chapter 2.8.2 --- PCR screening --- p.52 / Chapter 2.8.3 --- Restriction digestion of plasmid DNA --- p.53 / Chapter 2.9 --- DNA sequencing --- p.53 / Chapter 2.9.1 --- Plasmid sequencing using T7 Sequencing Kit --- p.53 / Chapter 2.9.2 --- Cycle Sequencing from PCR products or plasmid --- p.54 / Chapter 2.10 --- DNA Sequencing electrophoresis --- p.55 / Chapter 2.10.1 --- Preparation of 6 % polyacrylamide gel solution --- p.55 / Chapter 2.10.2 --- Gel casting --- p.55 / Chapter 2.10.3 --- Electrophoresis of Sequencing Gel --- p.56 / Chapter 2.10.4 --- Autoradiography --- p.57 / Chapter 2.11 --- DNA elution from dried sequencing gel --- p.57 / Chapter 2.12 --- Southern blot analysis --- p.58 / Chapter 2.12.1 --- Restriction digestion of genomic DNA --- p.58 / Chapter 2.12.2 --- Purification of digested DNA and agarose gel electrophoresis --- p.58 / Chapter 2.12.3 --- Capillary transfer of DNA to a Hybond´ёØ N+ nylon membrane --- p.59 / Chapter 2.12.4 --- DNA radiolabeling by nick translation --- p.60 / Chapter 2.12.5 --- Purificaiton of radiolabeled probe by NICK® Spin Column --- p.60 / Chapter 2.12.6 --- Hybridization of DNA --- p.61 / Chapter Chapter 3 --- Plant DNA extraction --- p.62 / Chapter 3.1 --- Introduction --- p.63 / Chapter 3.2 --- Reagents and buffer for total DNA extraction --- p.66 / Chapter 3.3 --- Extraction methods --- p.70 / Chapter 3.3.1 --- Sample preparation --- p.70 / Chapter 3.3.2 --- CTAB extraction method --- p.70 / Chapter 3.3.3 --- Potassium acetate/ SDS extraction method --- p.71 / Chapter 3.3.4 --- GIBRO Plant DNAzol® reagent for genomic DNA isolation --- p.72 / Chapter 3.4 --- Qualitative and quantitative analysis of DNA --- p.74 / Chapter 3.5 --- Results --- p.75 / Chapter 3.6 --- Discussion --- p.78 / Chapter Chapter 4 --- Amplified Fragment Length Polymorphism (AFLP) analysis of P. ginseng and P. quinquefolius --- p.81 / Chapter 4.1 --- Introduction --- p.82 / Chapter 4.2 --- Materials and methods --- p.88 / Chapter 4.2.1 --- Plant materials --- p.88 / Chapter 4.2.2 --- Choice of Primers and radiolabeling --- p.89 / Chapter 4.2.3 --- AFLP assay --- p.90 / Chapter 4.2.4 --- Electrophoresis of AFLP fingerprint --- p.91 / Chapter 4.2.5 --- Similarity Index (S.I.) analysis of AFLP profile --- p.91 / Chapter 4.2.6 --- Re-amplification of polymorphic DNA fragments isolated from dried sequencing gel --- p.92 / Chapter 4.2.7 --- Cloning and Sequencing of the AFLP fragments --- p.93 / Chapter 4.2.8 --- Conversion of AFLP marker into Directed Amplification of Minisatellite-region DNA polymorphism (DAMD) marker --- p.93 / Chapter 4.3 --- Results --- p.95 / Chapter 4.4 --- Discussion --- p.102 / Chapter Chapter 5 --- Direct Amplification of Length Polymorphisms (DALP) analysis of P. ginseng and P. quinquefolius --- p.107 / Chapter 5.1 --- Introduction --- p.108 / Chapter 5.2 --- Materials and methods --- p.112 / Chapter 5.2.1 --- Plant materials --- p.112 / Chapter 5.2.2 --- Choice of Primers --- p.113 / Chapter 5.2.3 --- Alternative labelled Amplification reaction --- p.114 / Chapter 5.2.4 --- Electrophoresis of the multi-locus amplification products --- p.114 / Chapter 5.2.5 --- Isolation and Re-amplification of polymorphic DALP fragments from dried sequencing gel --- p.115 / Chapter 5.2.6 --- Cloning and Sequencing --- p.115 / Chapter 5.2.7 --- Conversion of DALP marker to Sequence Tagged Site (STS) marker --- p.116 / Chapter 5.3 --- Results --- p.117 / Chapter 5.4 --- Discussion --- p.135 / Chapter Chapter 6 --- Sequence-characterized amplified region (SCAR): the sequel of random amplified polymorphic DNA (RAPD) --- p.137 / Chapter 6.1 --- Introduction --- p.138 / Chapter 6.2 --- Materials and methods --- p.140 / Chapter 6.2.1 --- Plant materials --- p.140 / Chapter 6.2.2 --- PCR reaction --- p.141 / Chapter 6.2.3 --- Cloning and sequencing --- p.143 / Chapter 6.3 --- Results --- p.144 / Chapter 6.4 --- Discussion --- p.157 / Chapter Chapter 7 --- Outlook --- p.159 / Chapter 7.1 --- Molecular authentication of Chinese medicinal materials --- p.160 / Chapter 7.2 --- Development of molecular markers for Ginseng --- p.161 / Appendix I --- p.164 / Appendix II --- p.165 / References --- p.166

何首烏為寥科多年生藤本植物，而中藥何首烏則為該植物的乾燥塊根。常用於治療白髮及與老年化相關的疾病。然而，何首烏的治療機理卻少有報導。本文對何首烏的粗提物及其主要生物活性成份2,3,5,4' -四羥基二萃乙烯-2-0-ß-D-葡萄糖苷(THSG) 在老鼠及人類的黑色素細胞中影響黑色素生成機理及細胞毒性進行了深入研究。 / 利用高效液相層析連接質譜儀的測定， THSG 在水及乙醇粗提物中含量分別為0.064 及0.75 的百分比。由於THSG 在水及乙醇粗提物中佔有一定份量，所以它對粗提物所產生的生物及生化反應有著重要的影響。在低於細胞毒範圍的劑量內，何首烏粗提物及THSG 能降低老鼠黑色素細胞株melan-a 的左旋多巴(L-DOPA)的轉化反應。在細胞毒性檢測中，水粗提物及THSG 在<100 μg/ml的劑量下均未有對至少5 種黑色素細胞株及黑色素瘤細胞造成傷害。然而，乙醇粗提物的細胞毒卻是水粗提物或THSG 的3-4 倍。 / 在無細胞系統中， THSG 在可逆轉的情況下抑制酪氨酸酶將左旋多巴轉化成黑色素。在細胞系統中，它也能阻止由蛋白激醋A (PKA)引發的黑色素生成反應。THSG 在老鼠及人類黑色素細胞中的中位值抑制率(lC₅₀)分別為123.0 μM及61.5 μM。 THSG 抑制酪氮酸醋的能力展現在黑色素細胞/角質細胞的共培養比在單黑色素細胞培養中更明顯。 / 調控酪氯酸臨可以在脫氧核糖核酸(DNA)轉錄及翻譯後修飾兩方面達成。在DNA 轉錄中，小眼球相關轉錄因數(MITF)的減少導致酪氨酸酶的表達隨著THSG 濃度而減少。翻譯後酪氮酸臨主要依靠蛋白激臨C-ß (PKC-ß)使其磷酸化，從而增加酪氯酸酶/酪氯酸酶相關蛋白-1(TRP-1 )組成複合蛋白。然而THSG 均減少蛋白激酶 C-ß的表達及酪氮酸酪/酪氮酸醋相關蛋白-1 所組成的複合蛋白。 另一方面， THSG 卻沒有影響酪氯酸酶蛋白在內質網/高爾基氏複合體內的糖基化及內涵體與溶酶體間的運輸。 / 總而言之，本文首次展示何首烏粗提物及THSG 在單細胞培養及共培養細胞的系統下抑制黑色素生成。THSG 能在可逆轉的抑制機制下阻止酪氯酸酶作出反應。而在PKA 引發的黑色素生成反應中， THSG 也能在DNA 轉錄及翻譯後修飾等過程中減低酪氯酸酶的活性。 / Radix Polygoni Multiflori, the dried root of Polygonum multiflorum (PM), is well documented for its clinical effects in treating various diseases associated with aging and hair graying, but the evidence based-mechanisms remain largely unknown. In this study, PM was extracted with water and 70% ethanol and a major constituent, 2,3,5,4'-tetrahydroxystilbene-2-0-I3-D-glucoside (THSG) of about 0.064% and 0.75%, respectively, were found in these extracts as analyzed by high-performance liquid chromatography (HPLC) coupled to mass spectrometry. The melanogenic properties and cytotoxicity of the two extracts and THSG were evaluated using murine and human melanocytes. / Both water and ethanol extracts of PM and THSG showed a dose-dependent anti-melanogenic activity in an in vitro murine melan-a melanocyte assay for reduction of L-DOPA conversion by tyrosinase. Of at least 5 melanoma and melanocyte cell lines tested, both water PM extract and THSG were relatively safe, which at doses <100 μg/ml did not demonstrate any significant cytotoxic effects. On the other hand, ethanol PM extract was about 3-4 folds more cytotoxic. / Tyrosinase is the rate-limiting enzyme for melanogenesis. In a cell-free kinetic analysis, THSG inhibited tyrosinase activity in a reversible and non-competitive manner. At the cellular level, this inhibition is mediated through a PKA-dependent melanogenic pathway, as well as in a dose-dependent manner, with IC₅₀ = 123.0 μM and 61.5 μM for murine and human melanocytes, respectively. Tyrosinase was much more sensitive to the inhibitory effect of THSG in the melanocytelkeratinocyte co-culture system than in the melanocyte mono-culture system. / Functional tyrosinase is regulated at both transcriptional and post-translational modification levels. At the transcription level, THSG reduced expression of microphthalmia-associated transcription factor (MITF) esulted in a down-regulation of tyrosinase expression. At the post-translational modification level, THSG inhibited expression of PKC-β which is responsible for tyrosinase phosphorylation, and enhanced tyrosinase/TRP-1 complex formation. On the hand, THSG did not affect glycosylation of tyrosinase nor its trafficking from ER/Golgi to endosomal/ lysosomal compartments. / Taken our results together, the anti-melanogenic property of PM extracts and THSG were firstly demonstrated in both mono- and co-culture system using murine or human melanocytes and keratinocytes. THSG is a reversible and competitive inhibitor, which lowered the tyrosinase activity at both transcription and post-translational modification levels via PKA-mediated melanogenesis. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Cheung, Wing Ki. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 160-174). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Chapter 1 / General introduction / Chapter 1.1 --- Anatomy and Physiology of the Skin --- p.5 / Chapter 1.1.1 --- Epidermis --- p.5 / Chapter 1.1.1.1 --- Stratum basale --- p.6 / Chapter 1.1.1.2 --- Stratum spinosum --- p.6 / Chapter 1.1.1.3 --- stratum granulosum --- p.7 / Chapter 1.1.1.4 --- Stratum corneum --- p.7 / Chapter 1.1.2 --- Dermis --- p.8 / Chapter 1.2 --- Melanogenesis of the Skin --- p.9 / Chapter 1.2.1 --- History of melanogenesis study --- p.9 / Chapter 1.2.2 --- Today's melanogenesis study --- p.10 / Chapter 1.3 --- Hyperpigmentary Disorders --- p.14 / Chapter 1.3.1 --- Malasma --- p.14 / Chapter 1.3.2 --- Lentigines --- p.15 / Chapter 1.3.2.1 --- Lentigo simplex --- p.16 / Chapter 1.3.2.2 --- Lentigo senilis etActinicus --- p.16 / Chapter 1.3.3 --- Post-inflammatory hyperpigmentation --- p.17 / Chapter 1.4 --- Current Available Treatment for Hyperpigmentation --- p.18 / Chapter 1.4.1 --- Topical treatment and their strategies --- p.18 / Chapter 1.4.1.1 --- Inhibition of tyrosinase activity --- p.18 / Chapter 1.4.1.2 --- Antioxidation --- p.21 / Chapter 1.4.1.3 --- Melanosome transfer inhibition --- p.22 / Chapter 1.4.1.4 --- Stimulation of desquamation --- p.22 / Chapter 1.4.2 --- Laser treatment and their strategies --- p.23 / Chapter 1.4.3 --- Sunscreen --- p.24 / Chapter 1.5 --- Theories and the Treatment of Hyperpigmentation with Chinese Herbal Medicine --- p.25 / Chapter 1.6 --- Testing Systems --- p.26 / Chapter 1.7 --- Aims and Objectives of Study --- p.27 / Chapter 2 / Chemical Properties of THSG / Chapter 2.1 --- Introduction --- p.30 / Chapter 2.1.1 --- Radix Polygoni multiflori --- p.30 / Chapter 2.1.2 --- 2,3,5,4'-tetrahydroxystilbene glucoside (THSG) --- p.31 / Chapter 2.1.2.1 --- Stilbene --- p.32 / Chapter 2.1.2.2 --- Chemical properties of THSG --- p.34 / Chapter 2.1.3 --- Objectives --- p.35 / Chapter 2.2 --- Materials and Methods --- p.36 / Chapter 2.2.1 --- Plant materials --- p.36 / Chapter 2.2.2 --- Extraction --- p.36 / Chapter 2.2.3 --- High performance liquid chromatography (HPLC) analysis --- p.36 / Chapter 2.2.4 --- Enzymatic hydrolysis of THSG and salicin --- p.37 / Chapter 2.2.5 --- HPLC/MS analysis --- p.38 / Chapter 2.2.6 --- Benedict's test --- p.38 / Chapter 2.2.7 --- Enzymatic oxidation --- p.38 / Chapter 2.2.8 --- Thin layer chromatography (TLC) analysis --- p.39 / Chapter 2.3 --- Results / Chapter 2.3.1 --- The THSG content in water and alcohol extracts of PM --- p.40 / Chapter 2.3.2 --- The stability of THSG against oxidation --- p.41 / Chapter 2.3.3 --- Enzymatic hydrolysis ofTHSG --- p.42 / Chapter 2.4 --- Discussion --- p.46 / Chapter 3 / The Melanogenic inhibitory mechanisms of Radix Polygonum multiflorum (PM) extracts and THSG in murine melanocyte / Chapter 3.1 --- Introduction --- p.47 / Chapter 3.1.1 --- Murine melanocyte --- p.47 / Chapter 3.1.2 --- Melanogenesis --- p.48 / Chapter 3.1.2.1 --- Factors affecting Melanogenesis --- Oxidation --- p.49 / Chapter 3.1.2.2 --- Factors affecting Melanogenesis --- UV radiation --- p.50 / Chapter 3.1.2.3 --- Factors affecting melanogenesis---- Cellular regulation --- p.51 / Chapter 3.1.2.3.1 --- The regulation of transcription, translation, and post-translational modification of tyrosianse --- p.51 / Chapter 3.1.2.3.2 --- PKA-, PKC-, and PKG- melangenic pathways --- p.56 / Chapter 3.1.3 --- Kinetic analysis of tyrosinase --- p.57 / Chapter 3.1.4 --- Objectives --- p.58 / Chapter 3.2 --- Materials and Methods --- p.59 / Chapter 3.2.1 --- Cell culture --- p.59 / Chapter 3.2.2 --- Kinetic analysis of tyrosinase activity inhibition --- p.60 / Chapter 3.2.3 --- SRB assay --- p.61 / Chapter 3.2.4 --- L -DOPA conversion assay --- p.62 / Chapter 3.2.5 --- Melanin production measurement --- p.62 / Chapter 3.2.6 --- ROS detection by flow cytometer --- p.63 / Chapter 3.2.7 --- In-situ tyrosinase activity assay --- p.63 / Chapter 3.2.8 --- Western blotting (WB) analysis --- p.64 / Chapter 3.2.9 --- Immunofluorescence microscopy --- p.66 / Chapter 3.2.10 --- Glycosylation analysis --- p.67 / Chapter 3.2.11 --- Co-Immunoprecipitation --- p.68 / Chapter 3.2.12 --- Radix Polygonum Multiflorum and THSG metabolite collection from rat serum --- p.69 / Chapter 3.3 --- Results --- p.71 / Chapter 3.3.1 --- Enzyme kinetic study of the catalysis of L-DOPA by murine melanocyte lysate --- p.71 / Chapter 3.3.2 --- Inhibitory effect of crude PM preparations and THSG on tyrosinase activity and melanin synthesis in murine melan-a melanocytes --- p.74 / Chapter 3.3.3 --- Effect of THSG on H₂0₂- induced oxidation --- p.77 / Chapter 3.3.4 --- THSG inhibits PKA-induced melanogenesis --- p.78 / Chapter 3.3.5 --- Reduction of in situ tyrosinase activity in PKA-induced melanogenesis --- p.82 / Chapter 3.3.6 --- Alternation of melanogenic proteins --- p.85 / Chapter 3.3.7 --- THSG does not alter the tyrosinase trafficking in ER and Golgi --- p.89 / Chapter 3.3.8 --- THSG does not alter the tyrosinase trafficking in endosomal lysosomal compartments --- p.93 / Chapter 3.3.9 --- Glycosylation analysis --- p.95 / Chapter 3.3.10 --- Reduction of interaction between tyrosinase and TRP-1 to form heterodimeric complexes --- p.97 / Chapter 3.3.11 --- The metabolite of PM water extract and THSG maintained the in vitro tyrosinase activity --- p.101 / Chapter 3.4 --- Discussion --- p.103 / Chapter 4 / The Inhibitory Effect of THSG on Melanogenesis in Monolayer Culture of Human Melanocytes and in Co-culture of Melanocyte-Keratinocyte / Chapter 4.1 --- Introduction --- p.110 / Chapter 4.1.1 --- Human melanocyte --- p.110 / Chapter 4.1.1.1 --- The origin and the development of melanocyte --- p.110 / Chapter 4.1.1.2 --- Morphology, body site distribution and histological location --- p.111 / Chapter 4.1.1.3 --- In vitro growth of human melanocyte --- p.112 / Chapter 4.1.1.3.1 --- Lifespan vs. culture conditions --- p.113 / Chapter 4.1.1.3.2 --- Lifespan vs. donor age and skin type --- p.114 / Chapter 4.1.1.4 --- Modulation of pigmentation in response to stress --- p.114 / Chapter 4.1.1.5 --- Difference between human and murine TRPs --- p.115 / Chapter 4.1.2 --- Keratinocyte-Melanocyte interaction --- p.117 / Chapter 4.1.2.1 --- Release of melanogenic factors --- p.117 / Chapter 4.1.2.2 --- Release of survival and proliferating factors --- p.118 / Chapter 4.1.2.3 --- Melanosome transfer determines the cutaneous pigmentation --- p.118 / Chapter 4.1.2.3.1 --- Molecular events during melanosome transfer --- p.119 / Chapter 4.1.2.4 --- Others --- p.121 / Chapter 4.1.3 --- Objectives --- p.121 / Chapter 4.2 --- Materials and Methods --- p.122 / Chapter 4.2.1 --- Cell Culture --- p.122 / Chapter 4.2.1.1 --- Human melanocytes isolation and cultivation --- p.122 / Chapter 4.2.1.2 --- Immortalized keratinocytes - HaCaT cells --- p.123 / Chapter 4.2.1.3 --- Co-culture of melanocytes and HaCaT cells --- p.124 / Chapter 4.2.1.3.1 --- Monolayer co-culture --- p.124 / Chapter 4.2.1.3.2 --- Two-layer co-culture --- p.124 / Chapter 4.2.2 --- SRB assay --- p.125 / Chapter 4.2.3 --- L-DOPA conversion assay --- p.125 / Chapter 4.2.4 --- Western blotting (WB) analysis --- p.125 / Chapter 4.2.5 --- Light microscopy and immunofluorescent microscopy --- p.126 / Chapter 4.2.6 --- cAMP immunoassay --- p.126 / Chapter 4.3 --- Results --- p.128 / Chapter 4.3.1 --- The isolation and purification of human melanocytes --- p.128 / Chapter 4.3.2 --- Aging vs. Tyrosinase activity and melanin content --- p.131 / Chapter 4.3.3 --- Inhibitory effect of THSG in tyrosinase activity in human melanocyte --- p.133 / Chapter 4.3.4 --- Alternation of melanogenic proteins --- p.135 / Chapter 4.3.5 --- Sensitization of melanocytes to THSG treatment in co-culture system --- p.138 / Chapter 4.3.6 --- Induction of melanocyte dendricity in co-culture system --- p.140 / Chapter 4.3.7 --- THSG inhibited cAMP induction by forskolin and paracrine factors from keratinocytes --- p.141 / Chapter 4.4 --- Discussion --- p.143 / Discussion / Chapter 5.1 --- Discussion --- p.149 / References --- p.160

Cheng, Baohui. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2013. / Includes bibliographical references (leaves 206-221). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstracts also in Chinese.

Yeung Ming. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 136-162). / Abstracts in English and Chinese. / THESIS COMMITTEE --- p.i / ACKNOWLEDGEMENTS --- p.ii / ABSTRACT (ENGLISH) --- p.iii~v / ABSTRACT (CHINESE) --- p.vi~vii / TABLE OF CONTENTS --- p.viii~xiii / LIST OF TABLES --- p.xiv~xv / LIST OF FIGURES --- p.xvi~xviii / LIST OF ABBREVIATIONS --- p.xix~xx / Chapter CHAPTER ONE: --- INTRODUCTION --- p.1 / Chapter 1.1 --- Different lipoproteins and their functions --- p.1 / Chapter 1.1.1 --- Chylomicrons --- p.4 / Chapter 1.1.2 --- VLDL --- p.4 / Chapter 1.1.3 --- LDL --- p.4 / Chapter 1.1.4 --- HDL --- p.5 / Chapter 1.2 --- Risk factors of coronary heart disease (CHD) --- p.5 / Chapter 1.2.1 --- Background information of CHD --- p.6 / Chapter 1.2.2 --- "Relationship between serum total cholesterol (TC), Low-density lipoprotein (LDL) cholesterol and CHD" --- p.7 / Chapter 1.2.3 --- High-density lipoprotein (HDL) cholesterol and CHD --- p.8 / Chapter 1.2.4 --- Triglyceride and CHD --- p.9 / Chapter 1.3 --- Cholesterol homeostasis --- p.10 / Chapter 1.3.1 --- Roles of HMG-CoA reductase in cholesterol biosynthesis --- p.13 / Chapter 1.3.2 --- Roles of cholesterol 7α-hydroxylase (CYP7A) in cholesterol catabolism…… --- p.15 / Chapter 1.3.3 --- Effects of Short-Chain Fatty Acid (SCFA) --- p.17 / Chapter 1.3.4 --- Related hormone --- p.18 / Chapter 1.4 --- Possible mechanisms of hypolipidemic agents --- p.19 / Chapter 1.4.1 --- Hypolipidemic functional foods --- p.20 / Chapter 1.4.2 --- Pharmacological drugs --- p.26 / Chapter 1.5 --- Edible and medicinal mushrooms --- p.28 / Chapter 1.5.1 --- General introduction --- p.28 / Chapter 1.5.2 --- Hypolipidemic agents from Fungi --- p.31 / Chapter 1.6 --- Animal model --- p.35 / Chapter 1.7 --- Objectives --- p.36 / Chapter CHAPTER TWO: --- MATERIALS AND METHODS --- p.37 / Chapter 2.1 --- Materials --- p.37 / Chapter 2.1.1 --- Mushroom samples and control --- p.37 / Chapter 2.1.1.1 --- Sample introduction --- p.37 / Chapter 2.1.1.2 --- Sample collection --- p.40 / Chapter 2.1.1.3 --- Sample preparation --- p.41 / Chapter 2.1.1.4 --- Moisture content --- p.45 / Chapter 2.1.2 --- Animal diets for different experiments --- p.45 / Chapter 2.1.2.1 --- Basal diet --- p.45 / Chapter 2.1.2.2 --- Diet for preliminary screening --- p.46 / Chapter 2.1.2.3 --- Diet for dosage experiment --- p.46 / Chapter 2.1.2.4 --- Diet for active ingredient experiments --- p.47 / Chapter 2.1.2.5 --- Diet for long-term feeding experiment --- p.47 / Chapter 2.1.3 --- Animal model --- p.49 / Chapter 2.2 --- Methods --- p.49 / Chapter 2.2.1 --- Nutritional components of mushroom samples --- p.49 / Chapter 2.2.1.1 --- Crude protein content (Kjeldahl method) --- p.49 / Chapter 2.2.1.2 --- Total dietary fiber content --- p.50 / Chapter 2.2.1.3 --- Crude lipid content --- p.52 / Chapter 2.2.1.4 --- Ash content --- p.53 / Chapter 2.2.1.5 --- Moisture content --- p.53 / Chapter 2.2.2 --- Animal handling experiments --- p.54 / Chapter 2.2.2.1 --- Feeding experiment standards --- p.54 / Chapter 2.2.2.1.1 --- Feeding experiments of preliminary screening test --- p.54 / Chapter 2.2.2.1.2 --- Feeding experiments of dosage test --- p.55 / Chapter 2.2.2.1.3 --- Feeding experiments of solvent extracts from Agrocybe aegerita (Brig) Sing (AA) --- p.56 / Chapter 2.2.2.1.3.1 --- Fractionation of ethanol & water soluble components of AA --- p.56 / Chapter 2.2.2.1.3.2 --- Feeding experiments of ethanol & water soluble components of AA --- p.57 / Chapter 2.2.2.1.4 --- Feeding experiment of long-term test --- p.58 / Chapter 2.2.2.2 --- Blood sample collection --- p.58 / Chapter 2.2.2.3 --- Serum preparation --- p.58 / Chapter 2.2.2.4 --- Liver sample preparation --- p.58 / Chapter 2.2.2.5 --- Fecal sample preparation --- p.59 / Chapter 2.2.3 --- Determination of serum lipid profiles --- p.59 / Chapter 2.2.3.1 --- Serum total cholesterol (TC) assay --- p.59 / Chapter 2.2.3.2 --- Serum triglyceride (TG) assay --- p.60 / Chapter 2.2.3.3 --- Serum high-density lipoprotein (HDL) cholesterol assay --- p.61 / Chapter 2.2.3.3.1 --- Separation of HDL fraction --- p.61 / Chapter 2.2.3.3.2 --- HDL cholesterol (HDL-c) determination --- p.61 / Chapter 2.2.4 --- Determination of liver lipid profiles --- p.62 / Chapter 2.2.4.1 --- Liver total cholesterol (TC) level determination --- p.62 / Chapter 2.2.4.2 --- Determination of liver total lipid (TL) level --- p.64 / Chapter 2.2.5 --- Quantitative determination of fecal neutral & acidic sterols --- p.64 / Chapter 2.2.5.1 --- Separation of fecal neutral & acidic sterols --- p.64 / Chapter 2.2.5.2 --- Derivatisation of fecal neutral sterols --- p.65 / Chapter 2.2.5.3 --- Derivatisation of fecal acidic sterols --- p.65 / Chapter 2.2.5.4 --- Gas chromatographic analysis of fecal neutral & acidic sterols --- p.66 / Chapter 2.2.6 --- Assays of liver key enzymes in cholesterol metabolism --- p.67 / Chapter 2.2.6.1 --- Preparation of hepatic microsome --- p.67 / Chapter 2.2.6.2 --- Assay of HMG-CoA reductase activity --- p.68 / Chapter 2.2.6.3 --- Assay of CYP7A activity --- p.69 / Chapter 2.3 --- Data statistics --- p.71 / Chapter CHAPTER THREE: --- RESULTS AND DISCUSSION --- p.72 / Chapter 3.1 --- Preliminary screening of eleven mushrooms for their hypolipidemic effect in hyperlipidemic S.D. rats --- p.72 / Chapter 3.1.1 --- Body weight and food intake --- p.73 / Chapter 3.1.2 --- Effect of mushroom supplementation on serum lipid profiles --- p.75 / Chapter 3.1.2.1. --- Effect of mushroom supplementation on serum TC levels --- p.75 / Chapter 3.1.2.2. --- Effect of mushroom supplementation on serum TG levels --- p.77 / Chapter 3.1.2.3. --- Effect of mushroom supplementation on serum HDL levels --- p.79 / Chapter 3.1.2.4 --- Discussion of serum lipid profiles of S.D. rats fed M.S. diets in mushroom screening experiments --- p.83 / Chapter 3.1.3 --- Effect and discussion of mushroom supplementation on hepatic lipid profiles --- p.84 / Chapter 3.1.4 --- Effect and discussion of mushroom supplementation on fecal neutral sterol excretion --- p.87 / Chapter 3.1.5 --- Summary (mushroom screening experiments) --- p.90 / Chapter 3.2 --- Hypolipidemic effect of Agrocybe aegerita (Brig.) Sing (AA) in a dose response study in hyperlipidemic S.D. rats --- p.91 / Chapter 3.2.1 --- Nutritional composition of AA mushroom --- p.91 / Chapter 3.2.2 --- Body weight and food intake --- p.91 / Chapter 3.2.3 --- Effect of three different dosages of AA mushroom supplementation on blood lipid profiles of S.D. rats --- p.93 / Chapter 3.2.3.1 --- Effect of different dosages of AA mushroom supplementation diets on serum TC level --- p.93 / Chapter 3.2.3.2 --- Effect of different dosages of AA mushroom supplementation diets on serum TG level --- p.93 / Chapter 3.2.3.3 --- Effect of different dosages of AA mushroom supplementation diets on serum HDL level --- p.95 / Chapter 3.2.3.4 --- Discussion of different dosages of AA mushroom supplementation diets on serum lipid profiles --- p.97 / Chapter 3.2.4 --- Effect and discussion of three different dosages of AA mushroom supplementation on hepatic lipid profiles --- p.98 / Chapter 3.2.5 --- Effect and discussion of three different dosages of AA mushroom supplementation on fecal neutral & acidic sterol excretion --- p.101 / Chapter 3.2.6 --- Summary (dose response study) --- p.105 / Chapter 3.3 --- Hypolipidemic effect of ethanol extract (E.E.) & water extract (W.E.) from AA in hyperlipidemic S.D. rats --- p.106 / Chapter 3.3.1 --- Extraction yield --- p.106 / Chapter 3.3.2 --- Body weight & food intake --- p.106 / Chapter 3.3.3 --- Effect of AA extract supplementation on serum lipid profiles --- p.107 / Chapter 3.3.3.1 --- Effect of AA extract supplementation on serum TC level --- p.107 / Chapter 3.3.3.2 --- Effect of AA extract supplementation on serum TG level --- p.108 / Chapter 3.3.3.3 --- Effect of AA extract supplementation on serum HDL level --- p.109 / Chapter 3.3.4 --- Effect of AA extract supplementation on hepatic lipid profiles --- p.111 / Chapter 3.3.5 --- Effect of AA extract supplementation on fecal neutral & acidic sterols excretion --- p.111 / Chapter 3.3.6 --- Discussion (active fraction extract study) --- p.113 / Chapter 3.4 --- Long-term evaluation of the hypolipidemic effect of AA supplementation in normolipic S.D. rats --- p.116 / Chapter 3.4.1 --- Body weight & food intake --- p.116 / Chapter 3.4.2 --- Effect of long term AA supplementation on serum lipid profiles --- p.117 / Chapter 3.4.2.1 --- Effect of long term AA supplementation on serum TC level --- p.117 / Chapter 3.4.2.2 --- Effect of long term AA supplementation on serum TG level --- p.118 / Chapter 3.4.2.3 --- Effect of long term AA supplementation on serum HDL level --- p.119 / Chapter 3.4.3 --- Effect of long term AA supplementation on hepatic lipid profiles --- p.119 / Chapter 3.4.4 --- Effect of long term AA supplementation on fecal neutral & acidic sterols excretion --- p.121 / Chapter 3.4.5 --- Effect of long term AA supplementation on hepatic key enzymes of cholesterol metabolism ´ؤ HMG-CoA reductase and CYP7A --- p.123 / Chapter 3.4.5.1 --- Quantitation of hepatic microsomal protein --- p.123 / Chapter 3.4.5.2 --- Effect of long term AA supplementation on HMG-CoA reductase activity in S.D. rats --- p.124 / Chapter 3.4.5.3 --- Effect of long term AA supplementation on CYP7A activity in S.D. rats --- p.124 / Chapter 3.4.7 --- Discussion (long-term study) --- p.126 / Chapter CHAPTER FOUR: --- CONCLUSION AND FUTURE PERSPECTIVES --- p.130 / References --- p.136

Zhang Li. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2003. / Includes bibliographical references (leaves 148-159). / Abstracts in English and Chinese. / Acknowledgements --- p.I / Abstract --- p.II / Abstract (in Chinese) --- p.IV / Publications --- p.V / List of Abbreviations --- p.VI / List of Tables --- p.VII / List of Figures --- p.VIII / Table of Contents --- p.XIII / Chapter Chapter One. --- Introduction --- p.1 / Chapter 1.1 --- Flavonoids --- p.1 / Chapter 1.2 --- Tea --- p.4 / Chapter 1.2.1 --- Composition of green tea catechins (GTC) --- p.4 / Chapter 1.2.2 --- Pharmacological activity --- p.6 / Chapter 1.2.2.1 --- Anticarcinogenic activity --- p.6 / Chapter 1.2.2.2 --- Antioxidative activity --- p.7 / Chapter 1.2.2.3 --- Radical scavenge --- p.7 / Chapter 1.2.2.4 --- Cardiovascular activity --- p.8 / Chapter 1.2.3 --- Pharmacokinetics of GTC --- p.8 / Chapter 1.2.3.1 --- Absorption --- p.10 / Chapter 1.2.3.2 --- Distribution --- p.11 / Chapter 1.2.3.3 --- Elimination --- p.11 / Chapter 1.2.3.4 --- Metabolism --- p.12 / Chapter 1.2.3.4.1 --- Metabolism in the small intestine --- p.12 / Chapter 1.2.3.4.2 --- Metabolism in the liver --- p.13 / Chapter 1.2.3.5 --- Summary of the pharmacokinetics of GTC --- p.13 / Chapter 1.3 --- Hawthorn --- p.14 / Chapter 1.3.1 --- Composition of hawthorn --- p.14 / Chapter 1.3.2 --- Pharmacological activity --- p.16 / Chapter 1.3.2.1 --- Inotonic activity --- p.16 / Chapter 1.3.2.2 --- Antiarrhythmic activity --- p.17 / Chapter 1.3.2.3 --- Hypolipidemic activity --- p.17 / Chapter 1.3.2.4 --- Antihypertensive activity --- p.18 / Chapter 1.3.2.5 --- Antioxidative activity --- p.18 / Chapter 1.3.3 --- Pharmacokinetics of HF --- p.18 / Chapter 1.3.3.1 --- Absorption --- p.19 / Chapter 1.3.3.2 --- Distribution and elimination --- p.21 / Chapter 1.3.3.3 --- Summary of pharmacokinetic of HF --- p.22 / Chapter 1.4 --- Mechanisms of intestinal absorption --- p.22 / Chapter 1.4.1 --- Passive transcellular transport --- p.23 / Chapter 1.4.2 --- Paracellular transport --- p.23 / Chapter 1.4.3 --- Carrier-mediated transport --- p.23 / Chapter 1.5 --- ABC transporters --- p.24 / Chapter 1.5.1 --- Cellular location and tissue distribution --- p.25 / Chapter 1.5.2 --- Substrates and inhibitors of ABC transporters --- p.26 / Chapter 1.6 --- Oral absorption models --- p.31 / Chapter 1.6.1 --- Ussing chamber --- p.31 / Chapter 1.6.2 --- In situ intestinal perfusion model --- p.33 / Chapter 1.6.3 --- Cell culture model --- p.34 / Chapter 1.7 --- Aims of the study --- p.36 / Chapter Chapter Two. --- Transport mechanism of green tea catechins --- p.37 / Chapter 2.1 --- Introduction --- p.37 / Chapter 2.2 --- Materials --- p.38 / Chapter 2.2.1 --- Chemicals --- p.38 / Chapter 2.2.2 --- Materials for cell culture --- p.38 / Chapter 2.2.3 --- Instruments --- p.39 / Chapter 2.3 --- Methods --- p.39 / Chapter 2.3.1 --- Analytical methods --- p.39 / Chapter 2.3.1.1 --- Analytical methods for validation of Caco-2 model --- p.39 / Chapter 2.3.1.1.1 --- Fluorescence analysis of lucifer yellow --- p.39 / Chapter 2.3.1.1.2 --- HPLC analysis of propranolol --- p.39 / Chapter 2.3.1.1.3 --- HPLC analysis of verapamil --- p.40 / Chapter 2.3.1.1.4 --- HPLC analysis of quinidine --- p.40 / Chapter 2.3.1.2 --- Analytical methods for samples contained GTC --- p.41 / Chapter 2.3.1.2.1 --- HPLC analysis for each GTC --- p.41 / Chapter 2.3.1.2.2 --- Preparation of calibration curves for each GTC --- p.42 / Chapter 2.3.1.2.3 --- HPLC/MS analysis of samples containing mixtures of four GTC --- p.42 / Chapter 2.3.1.2.4 --- Preparation of calibration curves for samples containing GTC mixture --- p.43 / Chapter 2.3.1.2.5 --- Validation of the HPLC methods --- p.43 / Chapter 2.3.1.3 --- Identification of metabolites with HPLC/MS --- p.44 / Chapter 2.3.2 --- Determination of stability profile of GTC in phosphate buffer --- p.44 / Chapter 2.3.3 --- Cell culture --- p.45 / Chapter 2.3.4 --- Validation of Caco-2 cell monolayer model --- p.46 / Chapter 2.3.4.1 --- Integrity of Caco-2 cell monolayer at pH 6.0 --- p.46 / Chapter 2.3.4.2 --- Permeability of paracellular and transcellular markers at pH 6.0 --- p.46 / Chapter 2.3.4.3 --- Validation of the existence of P-glycoprotein (P-gp) transporterin Caco-2 monolayer model --- p.46 / Chapter 2.3.4.4 --- Cytotoxicity test --- p.47 / Chapter 2.3.5 --- Transport study of GTC using Caco-2 cell monolayer model --- p.48 / Chapter 2.3.5.1 --- Bi-directional transport experiment --- p.48 / Chapter 2.3.5.2 --- Preparation of different dosing formulations of GTC --- p.48 / Chapter 2.3.5.2.1 --- Preparation of individual pure GTC solutions --- p.48 / Chapter 2.3.5.2.2 --- Preparation of cocktail 1 solution --- p.49 / Chapter 2.3.5.2.3 --- Preparation of green tea extract solution --- p.49 / Chapter 2.3.5.2.4 --- Preparation of cocktail 2 solution --- p.50 / Chapter 2.3.5.3 --- Sample treatment --- p.50 / Chapter 2.3.5.3.1 --- Samples for direct analysis --- p.50 / Chapter 2.3.5.3.2 --- Samples for enzymatic hydrolysis treatment --- p.51 / Chapter 2.3.5.4 --- Further investigation of the transport mechanism of GTC --- p.51 / Chapter 2.3.5.4.1 --- Inhibition transport of EC and EGC --- p.51 / Chapter 2.3.5.4.2 --- Transport mechanism of metabolites of EC and EGC --- p.52 / Chapter 2.3.5.4.3 --- Metabolic competition between EGC and the other GTC --- p.52 / Chapter 2.3.6 --- Calculation --- p.53 / Chapter 2.3.7 --- Data analysis --- p.54 / Chapter 2.4 --- Results --- p.55 / Chapter 2.4.1 --- Validation of the HPLC methods --- p.55 / Chapter 2.4.2 --- Stability of the GTC --- p.55 / Chapter 2.4.3 --- Extract of green tea leaves --- p.55 / Chapter 2.4.4 --- Validation of Caco-2 model --- p.59 / Chapter 2.4.4.1 --- Integrity of Caco-2 cell monolayer --- p.59 / Chapter 2.4.4.2 --- Permeability of paracellular and transcellular markers at pH 6.0 --- p.59 / Chapter 2.4.4.3 --- Validation of P-glycoprotein --- p.60 / Chapter 2.4.4.4 --- Cytotoxicity test --- p.61 / Chapter 2.4.5 --- Transport study of GTC --- p.63 / Chapter 2.4.5.1 --- Bi-directional transport of individual pure GTC --- p.63 / Chapter 2.4.5.2 --- Bi-directional transport of GTC in different dosing formulations --- p.66 / Chapter 2.4.5.2.1 --- Absorption transport profile of GTC in different dosing formulations --- p.66 / Chapter 2.4.5.2.2 --- Secretion transport profile of GTC in different dosing formulations --- p.66 / Chapter 2.4.5.3 --- Identification of metabolites of each GTC formed during the transport in Caco-2 cell model --- p.71 / Chapter 2.4.6 --- Further investigation of the transport mechanism of GTC --- p.82 / Chapter 2.4.6.1 --- Inhibition transport of EC and EGC --- p.82 / Chapter 2.4.6.2 --- Transport mechanism of metabolites of EC and EGC --- p.82 / Chapter 2.4.6.3 --- Metabolic competition between EGC and the other GTC --- p.85 / Chapter 2.4.6.4 --- Contribution of GTC on the metabolism of EGC --- p.89 / Chapter 2.5 --- Discussion --- p.92 / Chapter 2.5.1 --- Stability of the four GTC --- p.92 / Chapter 2.5.2 --- Validation of Caco-2 cell model --- p.92 / Chapter 2.5.3 --- Bi-directional transport of GTC --- p.93 / Chapter 2.5.4 --- Structure related efflux --- p.97 / Chapter 2.5.5 --- Metabolism of GTC --- p.98 / Chapter 2.5.6 --- Relationship between metabolism and efflux transport of GTC --- p.99 / Chapter 2.5.7 --- Bi-directional transport of GTC in different dosing formulations …… --- p.100 / Chapter 2.5.7.1 --- Absorption transport profile of different dosing formulations --- p.100 / Chapter 2.5.7.2 --- Secretion transport profile of different dosing formulations --- p.101 / Chapter 2.6 --- Conclusion --- p.105 / Chapter Chapter Three. --- Transport mechanism of hawthorn flavonoids --- p.106 / Chapter 3.1 --- Introduction --- p.106 / Chapter 3.2 --- Materials --- p.107 / Chapter 3.2.1 --- Chemicals --- p.107 / Chapter 3.2.2 --- Materials for cell culture --- p.108 / Chapter 3.2.3 --- Instruments --- p.108 / Chapter 3.3 --- Methods --- p.109 / Chapter 3.3.1 --- Analytical methods for HF --- p.109 / Chapter 3.3.1.1 --- Analytical methods of individual pure compound of HF --- p.109 / Chapter 3.3.1.1.1 --- HPLC analysis of HP and IQ --- p.109 / Chapter 3.3.1.1.2 --- HPLC analysis of EC --- p.109 / Chapter 3.3.1.2 --- Preparation of calibration curves for individual pure HF --- p.109 / Chapter 3.3.1.3 --- HPLC/MS analysis of three HF in mixture --- p.110 / Chapter 3.3.1.4 --- Preparation of the calibration curves of three HF in mixture --- p.111 / Chapter 3.3.1.5 --- Validation of HPLC methods --- p.111 / Chapter 3.3.2 --- Analytical methods for identification of metabolites with HPLC/MS --- p.111 / Chapter 3.3.3 --- Cell culture --- p.112 / Chapter 3.3.4 --- Cytotoxicity test --- p.113 / Chapter 3.3.5 --- Transport studies of HF using Caco-2 monolayer model --- p.113 / Chapter 3.3.5.1 --- Bi-directional transport experiment --- p.113 / Chapter 3.3.5.2 --- Preparation of loading solutions in different dosing formulations of HF for Caco-2 cell model --- p.114 / Chapter 3.3.5.2.1 --- Preparation of individual pure HF solutions --- p.114 / Chapter 3.3.5.2.2 --- Preparation of cocktail 1 solution --- p.114 / Chapter 3.3.5.2.3 --- Preparation of hawthorn extract solution --- p.114 / Chapter 3.3.5.2.4 --- Preparation of cocktail 2 solution --- p.114 / Chapter 3.3.5.3 --- Sample treatment --- p.115 / Chapter 3.3.5.4 --- Further study of the transport mechanism of HF --- p.115 / Chapter 3.3.5.4.1 --- "Inhibition transport of EC, IQ, and HP" --- p.115 / Chapter 3.3.5.4.2 --- "Transport mechanisms of the metabolites of EC, HP, IQ" --- p.116 / Chapter 3.3.6 --- Calculation --- p.116 / Chapter 3.3.7 --- Data analysis --- p.117 / Chapter 3.4 --- Results --- p.118 / Chapter 3.4.1 --- Validation of the HPLC methods --- p.118 / Chapter 3.4.2 --- Cytotoxicity test --- p.118 / Chapter 3.4.3 --- Transport study of HF --- p.122 / Chapter 3.4.3.1 --- Bi-directional transport of individual pure HF --- p.122 / Chapter 3.4.3.2 --- Bi-directional transport of the HF in different formulations --- p.123 / Chapter 3.4.3.2.1 --- Absorption transport of different formulations of HF --- p.123 / Chapter 3.4.3.2.2 --- Secretion transport of different dosing forms --- p.123 / Chapter 3.4.3.3 --- Identification of metabolites of each HF formed during their transport in Caco-2 model --- p.126 / Chapter 3.4.4 --- Further study on the transport mechanism --- p.136 / Chapter 3.4.4.1 --- "Inhibition transport of EC, HP, IQ" --- p.136 / Chapter 3.4.4.2 --- Transport mechanism of metabolites of HF --- p.136 / Chapter 3.4.4.3 --- Transport profiles of HF metabolites upon the loading of different dosing formulations of HF --- p.138 / Chapter 3.5 --- Discussion --- p.140 / Chapter 3.5.1 --- Bi-directional transport of each HF --- p.140 / Chapter 3.5.2 --- Bi-directional transport of HF in different formulations --- p.141 / Chapter 3.6 --- Conclusion --- p.142 / Chapter Chapter Four. --- Limitations of the current study --- p.143 / Chapter Chapter Five. --- Overall conclusions --- p.146 / References --- p.148 / Appendices --- p.160