Note
Novel polyfunctionalised geohopanoids in a recent lacustrine
sediment (Priest Pot, UK)
D.F. Watson *, P. Farrimond
Fossil Fuels and Environmental Geochemistry (Postgraduate Institute), Drummond Building, University of Newcastle upon Tyne, NE1 7RU, UK
Received 25 November 1999; accepted 2 October 2000 (returned to author for revision 16 December 1999)
Abstract
The sediment of a highly productive lake (Priest Pot, UK) contains the biosynthesised hopanoid bacteriohopanete-trol (BHT) and a series of hopanediols and triols. These polyols represent intermediate stages in the conversion of biologically produced hopanoids to the diagenetic products, hopanes. Two of these geohopanoids, assigned as trisho-mohopane-32,33-diol and bishomohopane-30,31,32-triol have not been previously reported in recent sediments, and their identi®cation, in addition to that of tetrakishomohopane-32,33,34-triol and a bishomohopanediol gives valuable information on the original composition and subsequent diagenesis of bacteriohopanepolyols within recent sediments.
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Keywords:Geohopanoids; Priest Pot (UK); Bacteriohopanepolyols; Hopanoid diagenesis
1. Introduction
Hopanoids are a complex group of biomarkers which occur ubiquitously in sediments (Rohmer et al., 1984). The biosynthesised precursor biohopanoids such as bacteriohopanetetrol (X; see Appendix), are synthesised as membrane constituents by a diverse range of bacteria including cyanobacteria, methylotrophs and purple non-sulphur bacteria, although they have not yet been isolated from obligately anaerobic bacteria (Rohmer et al., 1984; Rohmer, 1988; 1993). Biosynthesised C35
hopanoids occur as polyfunctionalised compounds with either four, ®ve or six functional groups on the side chain and are herein termed tetra-, penta-, and hexa-functionalised biohopanoids respectively (Appendix). In addition to the well known tetrafunctionalised bioho-panoid, bacteriohopanetetrol (BHT), varying complex groups such as glycosidic groups can be attached to the terminal carbon of the tetrafunctionalised biohopanoid
side chain resulting in a diverse group of molecules (Rohmer, 1988, 1993).
After death of the organism, diagenetic processes modify the initial side chain composition of the bioho-panoids, leading to the formation of geohopanoids including hopanoic acids, hopanols, hopenes and hopa-noidal aldehydes and ketones (Innes et al., 1997 and Refs. therein). Compounds such as these represent intermediate stages in the diagenesis of the poly-functionalised biohopanoid precursors to the hopanes which are routinely analysed in ancient sedimentary rocks and petroleums. An improved understanding of these diagenetic products and the reactions involved in the preservation and degradation of hopanoids in environmental samples is essential for optimising the application of hopanoid biomarkers. A recent publica-tion identi®ed the novel diagenetic intermediates tetra-kishomohopane-32,33,34-triol and bishomohopane-31,32-diol within recent lacustrine environments (Rodier et al., 1999); compounds which preserve information about both the precursor biohopanoids and the diage-netic processes involved. Here, we con®rm the presence of tetrakishomohopane-32,33,34-triol and a
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hopanediol in another European lake and report two additional novel geohopanoids, assigned as trishomo-hopane-32,33-diol and bishomohopane-30,31,32-triol, both of which comprise early diagenetic products of bacteriohopanepolyols.
2. Experimental
2.1. Sampling
The sediment (TOC=15.7%) was taken from the depth interval of 21±22 cm within an anoxic sediment core from the small highly productive lake, Priest Pot (54220N, 3000W; Robinson et al., 1984; Innes et al.,
1997).
2.2. Analysis of hopanols
Freeze-dried sediment was extracted with chloroform/ methanol (2:1 v/v) and the extract divided into aliquots for derivatisation targeted at speci®c hopanoid groups (see Innes et al., 1997). Hopanols were analysed as their acetate derivatives, formed by heating the extract with acetic anhydride and pyridine (4 ml; 1:1 v/v) at 50C for
1 h and leaving at room temperature overnight. The derivatised extract was rotary evaporated to dryness and treated with N,O bis(trimethylsilyl)tri¯uoroacetamide (BSTFA; heated at 50C for 1 h) prior to analysis.
2.3. Gas chromatography±mass spectrometry (electron impact)
GC±MS analysis was conducted on a Hewlett-Pack-ard 5890 II GC (split/splitless injector; 350C) linked to
a Hewlett-Packard 5972MSD (electron voltage 70 eV; ®lament current 220 mA; source temperature 270C;
multiplier voltage 2000 V; interface temperature 350C).
A DB5-HT column (15 m0.25 mm i.d.; 0.1 mm ®lm
thickness) was used with helium as the carrier gas. The oven temperature was programmed from 50±200C at
15C/min (held for 1 min), 200±250C at 10C/min (held
for 1 min) and 250±350C at 5C/min (held for 8 min).
The mass spectrometer was operated in full scan mode (m/z50±700).
2.4. Gas chromatography±mass spectrometry (chemical ionisation)
GC±MS (CI) was conducted on a Hewlett-Packard 6890 GC (split/splitless injector; 350C) linked to a
Hewlett-Packard 5973MSD. The same column, inter-face temperature and oven temperature program was used as described for GC±MS (EI). Ammonia was used as the reagent gas with a ¯ow of 15%. The mass spec-trometer was operated in selected ion monitoring mode
(SIM) targeting the molecular ions of the acetylated polyols [M+(CI)=m/zM+(EI)+18].
2.5. Identi®cation and quanti®cation of hopanoids
Hopanoids were primarily identi®ed as acetates by diagnostic molecular and fragmentation ions (CI & EI), where possible con®rmed by comparison with published mass spectra and in the case of compound VII by co-injection with an authentic standard. Hopanoids were quanti®ed using peak areas from them/z191 chroma-togram (EI) relative to an internal standard, 5a -andro-stan-3b-ol (m/z243).
GC±MS (EI; normalised to 100%)VIas diacetatem/z
556 (M+, 1%), 541 (M+- CH
3, 2%), 496 (M+- AcOH,
1%), 369 (side-chain cleavage, 18%), 335 (ring C clea-vage, 85%), 275 (335- AcOH, 9%), 215 (335- 2AcOH, 33%), 191 (ring C cleavage, 100%).VIIas diacetatem/z
570 (M+, 1%), 555 (M+- CH
3, 2%), 510 (M+- AcOH,
1%), 369 (side-chain cleavage, 22%), 349 (ring C clea-vage, 100%), 289 (349- AcOH, 4%), 229 (349- 2AcOH, 16%), 191 (ring C cleavage, 90%).VIIIas triacetatem/z
614 (M+, 1%), 554 (M+-AcOH, 1%), 494 (M+
-2AcOH, 5%), 393 (ring C cleavage, 14%), 333 (393-AcOH, 12%), 369 (side-chain cleavage, 6%), 368 (369-H, 1%), 213 (393- 3AcO(369-H, 18%), 191 (ring C cleavage, 100%).IXas triacetatem/z642 (M+, 1%), 627 (M+
-CH3, 2%), 421 (ring C cleavage, 87%), 369 (side-chain
cleavage, 21%), 361 AcOH, 4%), 301 (421-2AcOH, 5%), 241 (421- 3AcOH, 9%), 191 (ring C clea-vage, 100%).Xas tetraacetatem/z699 (M+- CH
3, 1%),
493 (ring C cleavage, 69%), 433 (493- AcOH, 6%), 373 (493- 2AcOH, 2%), 369 (side-chain cleavage, 28%), 368 (369- H, 4%), 313 3AcOH, 3%), 253 (493-4AcOH, 8%), 191 (ring C cleavage, 100%).
3. Results and discussion
Them/z191 chromatogram (Fig. 1) shows a series of hopanols present within the sediment sample. Tetra-hymanol (III) is also present in high abundance, asso-ciated with water column strati®cation (Sinninghe Damste et al., 1995), a situation which arises during the summer months in Priest Pot.
and two triols (Figs. 1 and 2; Table 1). Tetra-kishomohopane-32,33,34-triol (IX) has been previously identi®ed in low concentrations (ppb level) in several European lakes (Rodier et al., 1999) although it occurs in higher abundance in the Priest Pot sediment (2.0mg/g dry sediment). In addition, trishomohopane-32,33-diol (VII) and bishomohopane-30,31,32-triol (VIII) have been identi®ed (by EI and CI mass spectrometry) for the ®rst time in a sediment (although ®ve unsaturated trishomohopane-32,33-diols have been isolated from
Acetobacter aceti ssp. xylinumby Peiseler and Rohmer [1991]). For the trishomohopanediol the positions of the hydroxyl groups are assigned as C-32 and 33 by com-parison of the mass spectrum of compound VII with data published for a trishomohopane-32,33-diol synthe-sised by Peiseler and Rohmer (1991). Furthermore, a synthesised standard of trishomohopane-32,33-diol (M. Rohmer) co-eluted with compoundVII.
A bishomohopanediol (VI) has also been identi®ed in this sediment sample although the positions of the hydroxyl groups on the side chain cannot be determined conclusively. Rodier et al. (1999) identi®ed a bishomo-hopanediol peak detected in an European lake sediment as bishomohopane-31,32-diol (VIb). The reason for this identi®cation was not given. The bishomohopanediol
peak reported here is also present after treatment of the sample with periodic acid followed by sodium borohy-dride. This procedure cleaves vicinal 1,2-diols to pro-duce aldehydes which are subsequently repro-duced to terminal alcohols by sodium borohydride (see Rohmer et al., 1984). The presence of this compound after the treatment suggests that the correct assignment is that of a non-vicinal arrangement of the two alcohol groups, i.e. bishomohopane-30,32-diol (VIa). However, a sec-ond, slightly later-eluting bishomohopanediol is formed during the procedure and it is this peak that would be expected to be the C30,32-diol. On this evidence we cannot con®rm the side-chain structure of the bisho-mohopanediol reported here. It is plausible that both the C-30,32 and the C-31,32 arrangements of bacter-iohopanediol could be present in recent sediments given that both bacteriohopane-30,32,33,34,35-pentol and bacteriohopane-31,32,33,34,35-pentol have been identi-®ed in cyanobacteria (e.g. Ourisson and Rohmer, 1992; Zhao et al., 1996) as possible precursors (see Appen-dix).
The four hopanediols and triols reported in this sedi-ment are interpreted to have been formed from poly-functionalised biohopanoid precursors by natural oxidative cleavage and subsequent degradation of the side chain during early diagenesis, a process similar to the 1,2-diol cleavage that occurs during laboratory H5IO6 / NaBH4 degradation (Zundel and Rohmer,
1985). The stereochemistry is assumed to be 17b(H), 21b(H), but has not been determined.
Identi®cation of the speci®c biological precursors of these individual compounds can be constrained from our knowledge of those compounds produced by bac-teria (Rohmer, 1988; 1993), thus providing information about the bacterial contributions to sedimentary organic matter. Both trishomohopane-32,33-diol and kishomohopane-32,33,34-triol can be assigned to tetra-functionalised biohopanoid precursors (such as BHT, or composite tetrafunctionalised compounds which are abundant in Priest Pot sediments; Innes et al., 1997; Farrimond et al., 1998), although these cannot be
ascri-Table 1
Compound identi®cations and concentrations
Peak number Compound Concentration (mg/g dry sediment)
I Diploptene (Hop-22(29)-ene) 9.8
II Hop-21-ene 3.0
III Tetrahymanol 23.7
IV 17a(H),21b(H) bishomohopanol 17.6
V 17b(H),21b(H) bishomohopanol 24.9
VI Bishomohopanediol (VIa or b) 3.4
VII Trishomohopane-32,33-diol 0.7
VIII Bishomohopane-30,31,32-triol 3.7
IX Tetrakishomohopane-32,33,34-triol 2.0
X Pentakishomohopane-32,33,34,35-tetrol (bacteriohopanetetrol) 12.8 Fig. 1. M/z 191 chromatogram of the acetylated extract of a
bed to particular bacterial sources (Rodier et al., 1999). Finally, bishomohopane-30,31,32-triol can be ascribed with certainty to a derivation from hexafunctionalised biohopanoid precursors which are known to be abun-dant in Priest Pot sediments (Innes et al., 1997). These compounds (of which only aminobacteriohopanepentol has been positively identi®ed in Methylococcus luteus,
Methylococcus capsulatusandMethylomonas methanica; Neunlist and Rohmer, 1985; Zhou et al., 1991) are thought to be characteristic of Type I methylotrophic
bacteria (Neunlist & Rohmer, 1985) which are sig-ni®cant primary producers in Priest Pot.
It is evident that hopanoid distributions within Recent sediments have the potential to reveal valuable informa-tion regarding the nature of the early diagenetic reacinforma-tions of the biohopanoids. The C32±C34hopanediols and triols
reported here apparently represent incomplete side chain degradation products which may be intermediates in the formation of the expected C30±C32 hopanol products.
However, it cannot be excluded that these compounds
arise through C30+C3 and C30+C4 parallel synthetic
pathways to that leading to the formation of C35
bac-teriohopanepolyols (Peiseler and Rohmer, 1991). Whe-ther incomplete or not, degradation products they may also go on to be preserved as C32±C34hopanoid
diage-netic products if their side chains are preserved through defunctionalisation or via incorporation into macro-molecular organic matter. Further diagenetically pro-duced hopanoids with multiple side chain functionalities probably remain to be identi®ed in Recent sediments, and should shed further light on the diagenetic fate and speci®c biomarker potential of the hopanoids.
Acknowledgements
The Natural Environment Research Council (NERC) are thanked for funding of a PhD studentship (DW). We also thank P. Donohoe (NRG) for help with GC± MS analyses, and H. Innes and P. Fox for discussion on hopanoid analysis. The authors wish to thank an anon-ymous reviewer for their constructive comments and advice and M. Rohmer for providing a standard of trishomohopane-32,33-diol.
Associate EditorÐA.G. Douglas
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Structures of compounds referred to in the text; X bacteriohopanetetrol; VIa bishomohopane-30,32-diol; VIb
