ORIGINS AND SHORT-TERM SEDIMENTARY FATE OF GLOBALLY DISTRIBUTED BIOLOGICAL MARKER HYDROCARBONS SIMON JOHN BIRD B.Sc. (Hons.) A thesis submitted to the Council for Academic Awards in partial fulfilment of the requirements for admittance to the degree of: t DOCTOR OF PHILOSOPHY University of Plymouth, Department of Environmental Sciences, Plymouth, Devon, PL4 8AA, U.K. and Plymouth Marine Laboratory, Prospect Place, The Hoe, Plymouth, Devon, PLl 3DH. U.K. Submitted September 1992 "S^j^^iSsiTYO PLYMOUTH LIBRARY SERVICES ^ S-if-7,6\ Hi(^ 90 0136654 4 TO MY FAMILY ORIGINS AND SHORT-TERM SEDIMENTARY FATE OF GLOBALLY DISTRIBUTED BIOLOGICAL MARKER HYDROCARBONS by Simon John Hird ABSTRACT Nearly thirty Cjq, C^, and Ogy highly branched isoprenoid (HBI) hydrocarbons have been detected, sometimes in high concentrations, in recent freshwater, estuarine, coastal and hypersaline sediments, and water column particulate matter from numerous locations worldwide. The parent structures have been proved but only a few of the double bond positions have been established. The assignment of C,,, C22 ^ homologues and other Cjo and isomers, remains tentative. A wide body of evidence suggests that the compounds are biogenic in origin, with algae and possibly bacteria the most likely source organisms. A few of the compounds have been identified in field samples of algae but none have been reported in laboratory cultured biota. The alkenes with more than two double bonds appear to be rapidly removed from the hydrocarbon fraction in most sediments, whereas the alkanes and monoenes seem to be more resistant to biodegradation and hence occur in some more ancient sediments and oils. There is evidence that some of the alkenes react rapidly with sulphur to form either S- containing HBI heterocycles or become bound within macromolecular aggregates both found in sediments and some oils. The compounds, both as hydrocarbons and S-containing analogues, may prove useful environmental indicators once the sources and exact structures of more of them have been established. In the literature the structural elucidation of C25 ^ alkenes has been based mainly on the analysis of their hydrogenation products. However, some authors concluded that the alkenes are cyclic since some could not be fully hydrogenated. The structure of a Cjs HBI diene was proven to be acyclic by hydrogenation studies and GC and GC- MS analyses which showed the HBI compound to be fully saturated. The isolation and characterisation of synthetic alkenes resulted in the assignment, or partial assignment, of structures to four Ca3, six C25 and four €30 monoenes. The formation of novel monoenes via isomerisation reactions has also been achieved. The compounds form a valuable database of chromatographic and spectroscopic information for the assignment of sedimentary alkenes but the importance of isolation and micro-ozonolysis has been emphasised. Synthetic HBI alkenes were used to assign structures and partial structures to naturally occurring HBI hydrocarbons in three sediments. Other monoenes (both with methylene double bonds) were isolated from the sediments and characterised using spectroscopic and micro- ozonolysis data. The widespread occurrence of and C23 HBI hydrocarbons in Tamar sediments and associated algae (macrophytes and diatoms), the large variation in isotopic composition evident for the C^y monoene, and the seasonal sedimentary distribution all suggest two possible sources for the HBI hydrocarbons; microalgae and/or heterotrophic bacteria. Investigation of the distribution of hydrocarbons from the Peru upwelling area confirmed the rapid decrease in concentration of Cjs HBI alkenes with depth. A mixture of HBI monoenes was successfully incorporated into melanoidins but not detected in the humic acid pyrolysate which implied that incorporation of HBI alkenes into accreting humic substances was not a major mechanism of diagenesis of HBI alkenes. This study has extended present knowledge of the structures of HBI monoenes and has suggested two possible biological sources. There is still much to be learned about HBI polyenes and the subject is proving to be a fruitful area for further research into biomarker potential. Some possible future approaches are suggested. Parts of this work have been published (Rowland et ai. (1990), Org- Geochem. 15: 215-218; Hird et ai. (1992), War. Chem. 37: 117-129). ACKNOWLEDGEMENTS I would like thank first and foremost Dr. Steve Rowland for his supervision of this work. I offer my warmest thanks and appreciation for his help, encouragement, patience and continuing support. I am also grateful for the assistance of the following people and organisations who provided funding, samples, loaned equipment or performed specialist analyses, they are: The Local Education Authority of Devon County Council for the award of a Research Assistantship, the Faculty of Science (Polytechnic South West) for continuing the funding after transfer to the independant sector, and Prof. K. Bancroft for extending my contract when I had a knee injury. Dr. M.I. Venkatesan (University of California, U.S.A.) for providing samples of aliphatic hydrocarbons from the Antarctic. Dr. R. Smith (University of Waterloo, Canada), Dr. C. Parrish (Technical University of Nova Scotia, Canada) and Dr. Harris (P.M.L., U.K.) for providing auxenic microalgae samples. Dr. O. Bahzenova (University of Moscow, Russia) for providing samples of Siberian oils (under especially difficult circumstances). Dr. A.G. Douglas (N.R.G., University of Newcastle-upon-Tyne) for loan of micro- ozoniser and CDS pyroprobe. Prof. G. Eglinton and Prof. J.R. Maxwell (O.G.U., University of Bristol) for allowing me access to Finnegan TSQ 70 mass spectrometer, and Jim Carter for technical assistance with the NERC-supported MS-MS facility. Dr. W. Prowse and Prof. J.R. Maxwell (O.G.U.. University of Bristol) for obtaining 400 MHz »H NMR spectra. Dr. K. Oliver (BP Research, U.K.) for GC-FTIR analyses. Dr. G. Wolff (EOCGG, Liverpool University) for elemental analyses. Dr. B. Mycke (FINA Research, Belgium) for GC-IRMS analyses. Mr. Alan Aldridge (Database, U.K.) for IRMS analyses. Dr. A. Rees and Dr. R. Evens (University of Plymouth) for technical assistance with PYGC-MS, and Ag* HPLC and 270 MHz *H NMR, respectively. Ms. B. Wharton and Dr. P. O'Sullivan (University of Plymouth) for epipelic diatom identifications. Dr. T. Peakman who originally suggested the isomerisation reaction. Dr. J. SinningheDamst^and Dr. M. Kohnen (O.G.U., Technical University of Delft, Holland), Dr. F. Kenig (I.F.P., France), Dr. J. Grimalt (CID-CSIC, Spain) and Dr. J. Robson (SOAFD, Aberdeen) for valuable discussions. The technical staff of the Department of Environmental Sciences, University of Plymouth, most notably Roger Srodzinski, for maintenance and assistance with the Kratos mass spectrometer (under difficult operating conditions), but also Mr. I. Doidge (for his advice on my car!), Mr. A. Tonkin, Mr. A. Arnold and the late Mr. K. Pearson. Friends and collegues (past and present) in Plymouth and at Silwood Park, particularly Dr. Anthony Lewis (for the accomodation). Dr. Andy Revill (for leaving me!), Dr. Andy Rees, Dr. Mark Gough, all in P.E.G.G and Research Rovers ("No surrender"), and Dr. R. Large and Dr. P. Tibbetts (M-Scan Ltd; for their patience awaiting completion of this thesis). To whoever built "The Spaniard Inn" at Cargreen, many thanks. So convenient! I would also like to thank Dr. Murphy (St. Boniface's College, Plymouth) for initially motivating my interest in chemistry and Dr. M. Rhead (University of Plymouth) for giving me the opportunity to experience the "joys" of research prior to starting my Ph.D. A special "thank you" to Hez for use of her Mac, cooking lots of chick pea curries and correcting my grammar (amongst other things). PREFACE This thesis is presented in eight chapters. Each chapter is divided into subsections {e.g. LI, 1.2 ... etc.). Further subdivisions are similarly numbered sequentially and, where necessary, by the use of italics. Compound structures are assigned unique numbers (e.g. 1, 2 ... etc.), generally in chronological order of appearance, in the text and are presented at the end of each chapter. Chapter 1 provides an introduction and general background to the research described herein. Chapters 2-6 describe the research into the characterisation, distribution and fate of highly branched isoprenoid (HBI) hydrocarbons. Suggestions for further work are given in Chapter 7, whereas Chapter 8 covers the experimental and analytical procedures used. ABBREVIATIONS USED IN THE TEXT HBI highly branched isoprenoid DCM dichloromethane Na2S04 sodium sulphate LiAlH4 lithium aluminium hydride Mg magnesium CeClj.THaO cerium chloride heptahydrate EijO diethyl ether THF letrahydrofuran NaOH sodium hydroxide AgNOj silver nitrate KCl potassium chloride Na^PjO, sodium pyrophosphate DMF dimethylformamide CS2 carbon disulphide BuLi butyllithium TsOH toluene-p-sulphonic acid HOAc acetic acid NaHCOa sodium hydrogen carbonate PtOz.HzO platinum (TV) oxide monohydrate TOC total organic carbon ODP Ocean Drilling Project TLC thin layer chromatography HPLC high performance liquid chromatography GC gas chromatography MS mass spectrometry PY pyrolysis NMR nuclear magnetic resonance FTIR fourier transform infra red spectroscopy RI retention index LRMS low resolution mass spectrometry IRMS isotope ratio mass spectrometry McL McLafferty rearrangement TABLE OF CONTENTS SECTION PAGE NUMBER CHAPTER ONE INTRODUCTION 1.1 INTRODUCTION 1 1.2 THE OCCURRENCE