Table Of ContentAN ABSTRACT OF THE THESIS OF
Hye-Dong Yoo for the degree of Doctor of Philosophy in
Pharmacy presented on October 13. 1997
Title: Bioactive Secondary Metabolites from Marine Algae and Study of
Oxidized Anandamide Derivatives
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Abstract approved:
William H. Gerwick
My investigations of the natural products of marine algae have resulted in
the discovery of several new secondary metabolites. Bioassay-guided
fractionation led to the isolation of these new compounds and spectroscopic
analysis was utilized in their structural characterization.
Two new and potent antimitotic metabolites, curacins B and C, were
isolated from a Curacao collection of Lyngbya majuscula. In addition, four
curacin A analogs were prepared by semisynthetic methods. The structures of
the new curacins and the curacin A analogs were determined by spectroscopic
analysis in comparison with curacin A. The biological properties of the new
natural products and synthetic derivatives of curacin A were examined.
Investigations of another Curacao collection of L. majuscula revealed a
new cytotoxic lipopeptide, microcolin C. Microcolin C was found to have an
interesting profile of cytotoxicity to human cancer-derived cell lines.
A new metabolite, vidalenolone, was isolated from an Indonesian red
alga Vidalia sp. The structure of this new cyclopentenolone-containing
compound was determined by a combination of spectroscopic methods.
Filamentous cells isolated from female gametophytes of the brown alga
Laminaria saccharina were cultured in flasks or bioreactors. These cultures
produced a variety of w6-lipoxygenase metabolites: 13-hydroxy-9,11
octadecadienoic acid (13-HODE), 13-hydroxy-6,9,11,15-octadecatetraenoic
acid (13-HODTA), and 15-hydroxy-5,8,11,13-eicosatetraenoic acid (15-HETE).
Five oxidized anandamide derivatives were prepared from anandamide
through autoxidation in an exploration of ligand binding to the cannabinoid
receptor. Their structures were determined by a combination of NMR
spectroscopy and GC-MS. The cannabinoid receptor binding affinity of these
derivatives was evaluated. This study revealed the following trend in activity:
anandamide > 15- > 9- > 8- > 11- > 5-hydroxyanandamide.
Copyright by Hye-Dong Yoo
October 13, 1997
All Rights Reserved
Bioactive Secondary Metabolites from Marine Algae and Study of Oxidized
Anandamide Derivatives
by
Hye-Dong Yoo
A THESIS
submitted to
Oregon State University
in partial fulfillment of
the requirements for the
degree of
Doctor of Philosophy
Completed October 13, 1997
Commencement June 1998
Doctor of Philosophy thesis of Hye-Dong Yoo presented on
October 13, 1997
APPROVED:
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Major Professor, representing Pharmacy
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Dean of the College of Pharmacy
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Dean of Grad e School
I understand that my thesis will become part of the permanent collection of
Oregon State University libraries. My signature below authorizes release of my
thesis to any reader upon request.
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Hye-Dong Yoo, Author
ACKNOWLEDGEMENTS
I wish to thank my major advisor, Dr. William H. Gerwick, for his generous
support and guidance throughout my graduate studies. His unlimited patience
and mentorship led me to completion of this project.
I must thank my labmate Mary Ann Roberts for teaching me about algae
and for encouragement during my research.
I would like to thank Dr. George Constantine for his thoughtful advice and
encouragement; Dr. Max Deinzer and Dr. Michael Hyman for accepting to be on
my committee and their time; Dr. Mark Zabriskie and Dr. Philip Proteau for their
insightful suggestions; Dr. Mitchell L. Wise for a long time friendship; Dr. Dale
Nagle for his assistance in my early research years; Dr. Yongfu Li and Dr. Anna
Boulanger for helpful discussions; Brian Arbogast for his mass spectral work;
Roger Kohnert for NMR spectroscopy.
I am also indebted to Dr. Greg Hooper, Mary Roberts, Brian Marquez, Lik
Tan Tong and Namthip Sitachitta for their assistance with this manuscript.
I would like to acknowledge Dr. Pete K. Freeman, Dr. Henry Schaup, and
Dr. Christian Haugen for their unconditional support and friendship.
My past and present labmates are also appreciated for their support and
helpful disscussions.
Finally, I wish to express my gratitude to my family for their patience and
support.
Funding for the work in this thesis was supported by the National Cancer
Institute and Oregon Sea Grant.
CONTRIBUTION OF AUTHORS
Chapters V and VI of my thesis were collaborative efforts performed with
two research groups. Dr. Greg Rorrer research group in the Department of
Chemical Engineering was involved in bioreactor design and start-up and
cultivation of brown alga, Laminaria saccharina female gametophyte cells in
chapter V.
Ken Soderstrom in the laboratory of Dr. Thomas Murray was responsible
for the competitive receptor ligand displacement assay with a variety of
hydroxyanandamides for Chapter VI.
TABLE OF CONTENTS
Page
CHAPTER I. GENERAL INTRODUCTION
1
MARINE MICROORGANISMS
BIOACTIVE METABOLITES FROM CYANOBACTERIA
OXYLIPINS FROM MARINE ALGAE
CHAPTER II. CURACINS, ANTIMITOTIC NATURAL PRODUCTS
FROM THE MARINE CYANOBACTERIUM LYNGBYA MAJUSCULA 23
ABSTRACT 23
INTRODUCTION 24
RESULTS AND DISCUSSION 27
EXPERIMENTAL 50
CHAPTER III. MICROCOLINS, NEW ANTINEOPLASTIC NATURAL
PRODUCTS FROM THE MARINE CYANOBACTERIUM
LYNGBYA MAJUSCULA 59
ABSTRACT 59
INTRODUCTION 60
RESULTS AND DISCUSSION 63
EXPERIMENTAL 76
CHAPTER IV. VIDALENOLONE, A NOVEL SECONDARY
METABOLITE FROM THE INDONESIAN RED MARINE ALGA
VIDALIA SR 79
ABSTRACT 79
INTRODUCTION 80
RESULTS AND DISCUSSION 82
EXPERIMENTAL 89
CHAPTER V. ANALYSIS OF OXYLIPIN PRODUCTS FROM
GAMETOPHYTE CELL CULTURE OF BROWN ALGA
LAMINARIA SACCHARINA
91
ABSTRACT
91
INTRODUCTION 92
RESULTS AND DISCUSSION 97
EXPERIMENTAL 112
CHAPTER VI. PRODUCTION OF OXIDIZED ANANDAMIDE
DERIVATIVES AND EVALUATION OF THEIR BINDING
AFFINITY TO THE CANNABINOID RECEPTOR (CB1) 115
ABSTRACT 115
INTRODUCTION 116
RESULTS AND DISCUSSION 118
EXPERIMENTAL 132
CHAPTER VII. CONCLUSION 135
BIBLIOGRAPHY 138
APPENDIX 146
List of Figures
Figure Page
1.1 Biosynthesis of the products of arachidonic acid in mammals. 15
Structures of selected secondary metabolites from
11.1
Lyngbya majuscula. 25
11.2 Mean graphs of curacins B and C (95:5) mixture obtained
from the National Cancer Institute 60 cell line in vitro screen. 28
11.3 Flow chart of fractionation of Lyngbya majuscula. 29
11.4 Comparison of 1H NMR spectra of curacins A, B, and C. 30
11.5 Geometrical configuration of conjugated diene of curacin B (14)
determined by NOESY in CDCI3. 33
11.6 Thermal isomerization of curacins A, B, and C. 34
11.7 GC-MS traces for thermal isomeration of curacins A, B, and C. 35
11.8 Natural antimitotic agents and synthetic derivatives of
the curacins. 37
11.9 11-1 NMR spectrum of cyclopropyl ring opened curacin A (18)
in C6D6. 39
11.10 1H-1H COSY spectrum of cyclopropyl ring opened
Curacin A (18) in C6D6. 40
11.11 1H NMR spectrum of 19-epi curacin A (19) in C6D6. 41
11.12 1H-1H COSY of 19-epi curacin A (19) in C6D6. 42
11.13 1H NMR spectrum of thiazoline ring opened curacin A
(20) in C6D6. 43
11.14 HMBC plot of thiazoline ring opened curacin A (20) in C6D6. 44
11.15 1H NMR spectrum of derivative 21 in C6D6. 45
11.16 'H NMR spectrum of curadisulfide A (22) in C6D6. 46
Description:My investigations of the natural products of marine algae have resulted in Filamentous cells isolated from female gametophytes of the brown alga.
