ALKANE BIOMARKERS. GEOCHEMICAL SIGNIFICANCE AND APPLlCA TION IN OIL SHALE GEOCHEMISTRY

F.J. GONZALEZ-VILA Instituto de Recursos Naturales y Agrobiologia, C.S.I.e. P.O. Box 1052 41080-Sevilla Spain

ABSTRACT. The application of alkane biomarkers for source, palaeoenvironmental reconstruction, maturity and biodegradation of sedimentary organic matter is briefly reviewed. Data on the extractable biomarker composition of a Spanish oil shale, as well as some considerations on future work in biomarker geochemistry are also included.

1. Biomarkers. Concept and Applications

la spite of the progress made in the last decades, understanding the origin and fate of organic compounds in Nature still remains an intriguing and difficult task. However, the concept of using organic compounds as measures of events and processes in the natural worId is now widely accepted and applied in a wide range of disciplines.

The organic substances present in recent and ancient sediments are thought to reflect the source inputs and the environmental conditions, including microbial activity, at the time of deposition. A restricted number of these organic substances, the so-called molecular fossils or biological markers (biomarkers for short), are molecules directly inherited from the organisms living at the time of deposition which have been preserved without subsequent alteration, or with only minor changes, so that their basic carbon skeleton remains intact (1,2)

The term biomarker stems therefore from the similarity of certain chemical structures in the sedimentary record to precursor molecules which occur in unique biosynthetic products of specific groups of organisms in the natural environment. Compounds whose biological origin is uncertain but whose structural specificity makes them useful as correlation parameters in source diagnosis are also referred to as biomarkers. Small changes in the structures of these biomarkers are due to microbial transformations in the water column and in recent sediments or chemical reactions during diagenesis in ancient sediments. It should be emphasized that, although the inputs contributing to the

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C. Snape (ed,). Composítion. Geoc~mistry ami Conversion af Oil Shales, 51 ~8, © 1995 Kluwer Academic Publishers, Printed in t~ Nether/ands.

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sedimentary organic matter are deduced from the presence of specific biomarkers, their absence does not necessarily imply that the biosynthetic precursors and the corresponding biota were not present in the depositional environment.

The biomarker approacb, i.e. isolation and recognition of biomarkers from different environmeots, constitutes a powerfu) too) in many areas of geocbemistry. Tbe use of biomarkers has moved from an academic pursuit (assessement of the nature of the different organic inputs ioto a sedimeot, reconstruction of paleo-environments, study of the processes of early diagenesis and tbermal maturation, etc) to a highly applied analytical technique in technological fields sucb as petro)eum and gas exploration, based mainly on the determination of source, maturity, migration and biodegradation of bydrocarbons. In all cases, conclusions based on biomarker distribution data are stronger than those based on any single compound or class of compounds.

2. ExtractioD,IsolatioD and AnaJysis of Biomarkers

Biomarker determinations provide a chal)enge for the analytical cbemist. In general. the biomarkers of greatest interest are present in relatively low concentrations in crude oils and rock extracts, whicb are complex mixtures of organic compounds.

Extraction of biomarkers from sediments and oils is. normalIy accomplisbed by conventional solid-liquid extraction procedures using different solvents or mixtures ot tbem in Soxhlet apparatus. Over recent years new extraction techniques based on the use of ultrasonic agitation ()) and supercritical fluids lO) have been developed. The supercritical fluid extraction (SFE) seems to bave a promising future, since it offer~ considerable advantages regarding extraction time and elimination of environmentai bazards.

Afier removal of asphaltenes, insoluble in n-pentane, the extracts are usuall~ fractionated into saturated, aromatic and polar components by adsorptior. chromatograpby on column or on modern solid phase extraction cartridges (~) .

Then-alkanes are usually tbe most abundant compounds in the saturated fractions L);

crude oils and rock extracts, and sometimes it is necessary to remove them as ar analytical step to effectively enhance the concentration of other more specifi, biomarkers, such as branched and cyclic alkanes. Many concentration techniques hay, been used quite effectively. Amongst tbem tbe clatration with urea and the use o molecular sieves seem to be the most useful to separate n-alkanes from thei: brancbed/ cyclic congeners.

The most common methods for detection and identification of biomarkers in comple: mix~es are the computerised Gas Chromatography-Mass Spectrometry (GC-MS systems, which became readily available and are today indispensable for routine analysi of crude oils and sediment extracts (6). Relatively different new techniques, such as GC MS-MS and Supercritical Fluid Chromatography (SFC)-MS have introduced a numbe of alternative methods for tbe analysis of stereoisomers and high molecular weigt biomarkers.

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3.Geochemical Significance of tbe Various Molecular Classes of Hydrocarbons Found in Sediment Extracts and Petroleum

A coroprehensive review of all the areas of biomarker geochemistry which have undergone a number of important reevaluations in the past decade (2.1-10) is beyond the scope of this reporto Rather this chapter will essentially comprise an overview of existing data (taken froro the aboye monograpbs, and only recent references to illustrate trends and developments are included) on those aspects of alkane biomarker distribution which relate lO the type of organic matter, the depositional environment, the thermal maturity and indications for biodegradation. Another useful application of biomarkers in oil exploration are the oil-oil and oil-source rocks correlations, as well as the determination of relative migration distances of expelled hydrocarbons. Structures marked throughout the text with Roman numbers are presented in Appendix l.

3.1. NORMAL ALKANES

N-alkanes are trace constituents of biologicallipids. However, these compounds have excellent preservation characteristics and may therefore be regarded as biomarkers. The most common application of n-alkanes is to infer the source of the organic materiaL Input of terrigenous source material is indicated by a strong predominance of high molecular weight odd-numbered alkanes (n-Cn , n-C29 and n-Cll ), which are associated with leaf waxes. Marine algae are known to produce lower molecular weight n-alkanes with maximum at n-CI1 and without any odd or even chain length preference. Freshwater algae synthesize longer chain length n-alkanes, with a predomiilance similar to that in higher land plants. Studies of bacteria have shown them to produce n-alkanes in the n-CI, to n-Cn region with no marked odd/even preference.

The distribution of n-alkanes have been used to ínter specific depositional environment of source rocks. For example, the occurrence of n-alkane distributions with an even-number preference is usually assumed to reflect evaporitic facies or marine carbonate environments (10.11). This is probably a function of the highIy reducing nature of the depositional environment which causes reduction rather than decarboxylation of the biologically-4erived even carbon numbered fatty acids (12). The observation of even/odd predominance in well-oxygenated systems has prompted other authors to suggest a possible bacterial autochthonous origin for these alkanes (13.14).

Maturity and biodegradation can also influence the distribution of n-alkanes. Thus, as maturity increases the odd/even or evenlodd predominance decreases. Bacterial degradation of crude oils causes progressive removal of n-aIkanes beginning from the low molecular weight homologues.

3.2. BRANCHED ALKANES

3.2.1. Monomethyl branched alkanes. These compounds occur mainIy as the iso- (2-methyl) and anteiso- (3-methyl) alkanes, and derive from plant waxes (C2,-C31 with an

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odd chain predominance) or bacteria (Cn-C,. iso, Cn and C lI anteiso). Mid-chain branched alkanes, in particular methyl heptadecanes (C,s), with the meth:

substituent at C7 7 ·or Cs have been proposed as biomarkers of cyanobacteria and henl possibly of hypersaline environments (15,16)

3.2.2. Acyclic lsoprenoids. These compounds are known constituents of plants, animo tissues and bacterial cell walls and have been reported in all classes of geologic, samples. The phytol side chain of chlorophyl1-a accounts for the widespread occurrenc of acyclic isoprenoids in the biosphere. They are chemically formed from variOl combinations of C, isoprene units (1) through three main types of linkages : head-to-ta (the most conunon, and include compounds such as pristane C,,(ll), phytane C2Q(III) 3D

homologues up to C.,), tail-to-tail (squalane, perhydro-.6-carotane, lycopane, etc), an head-to-head (Cn-C«) typical of thermophiJic and other archaebacteria).

Isoprenoids, and in particular the ratio of pristane to pbytane, have been widely use to obtain information on depositional enviroomeots 00 the premise that both pristar and phytane are derived from phytol. Diagenesis of CM phytol produces by reductic and dehydratioo in C2Q phytane io highIy reducing conditions, whereas in less anox conditions oxidation and decarboxylation prevail yielding C,9 pristane. However, sorr important limitations 00 the use of the pristane/phytane ratio as ao indicator for oxyge levels have arisen (,,17), and should be taken into consideration before using th;

parameter. Since most Recent hypersaline eovirooments are characterised by reducing condition~

the pristane/phytane ratio has also beeo suggested as an iodicator for salinity (', Specific acyclic isoprenoids associated with particular eovirooments, such as the regulé. C25 isoprenoid and botryococcane (IY), have been reported bydifferent authors (9).

3.3. MONOCYCLlC ALKANES

It is assumed that n-alkylcyclohexanes and n-alkyl,methylcyc1ohexanes arise fror cyclization of biologically-derived fatty acids (19). Tbeir distribution in crude oils an sediment extracts follows normally the n-alkane distribution with a large odd over eve carbon number predominance.

Alkyl cyclohexanes with isoprenoid side chains are probably derived frol carotenoids, which are abundant in algae and some higher plants, or are generated fror. bacterial reworking of previously sedimented acyclic isoprenoids (10). The distributi0 and catagenesis of cycloalkanes in oils has been recently reviewed (21).

3.4. BICYCLlC ALKANES (SESQUITERPENOIDS)

These compounds have been identified in fossil resins, sediments and petroleum (22). Th, two main types of bicycloalkanes identified are the drimanes (y) and the eudesmane (Yl), The latter are believed to be derived from eudesmanol, an alcohol found preser in higher plants, whereas drimanes are though to be derived from the degradation (" polycyc1ic alkane precursors in bacteria and appear to be ubiquitous. The larg

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number of isomers present in the samples a1so raises the possibility of stereochemical interconversion as a result of diagenesis and maturation.

3.5. TRICYCLIC ALKANES.DITERPENOIDS AND TRICYCLYC TERPANES

The presence of diterpenoid hydrocarbons in fossil fuels and sediments is a very strong indication of a terrigenous input lo tbe oils or source rocks, since tbese structures are predominant constituents of higher plant resins and supportive tissues, especially in conifers. The skeletal structure types found in fossil fuels consist primarily of derivatives of the abietane (YII), pimarane (VIII) and Iabdane (!X) series.

Another series of tricyclic aIkanes commonly found in marine sediments and erude oils is the homologous series of extended tricyelic terpanes (X) in the range from C,H}4 to C4,H86, and possibly higher. Tbey are not reported to be found in terrestrial souree rocles or oils derived from terrigenous source material. Different natural precursors, such as the tricycIobexaprenol, formed anaerobically from an ubiquitous eell eonstituent, hexapreool, have been proposed for this series (23).

The trieycIie terpane distribution can be related to the nature of the depositional environment. Thus, oils source from carbonate souree rocks usually show a dominant C23 homologue (24). 00 the otber hand, several autbors have proposed the potential use of trieyclic terpanes as a maturity indicator in the lower maturity levels on the basis of the variation in the relative amount of trieycIie terpanes to hopanes (23), and ehanges observed in tbe isomers stereochemistry (26). Tricyclie terpanes are especially useful io correlating nondegraded to biodegraded crude oils since they survive even severe biodegradation processes (24).

3.6. TETRACYCLIC ALKANES

3.6.1. Non-Suroidal Tetracycüc Terpanes. Tbe main types oftetracyelic alkanes found io sediment and crude oils originate from different sources: a) kaurane (XI), beyerane (XlI) and pbyIlocladane (Xill) are produced by diagenesis of tetracyclic alkenes characteristics of conifer resios; b) tbe series of 4,5-secotriterpanes (XIV) is thought to be formed from plant derived pentacycIic triterpenoids by thermal cracking, and e) the 17,21-secohopanes. (XY) and 8,14-secohopanes (XVI) derived from pentaeycIic hopanoids by cleavage oftbe 17(21) and 8(14) bonds respectively, taking place either by microbial activity during early diagenesis or by tbermocatalytic degradation of hopanes (21).

Tetracyclic terpanes are usually present in a series from C2• to C21 • Differences in probable precursors of tetracyclic and tricyclic terpanes malees the ratio of ~4 tetracyclic terpane over ~ tricyclic terpane a source parameter (l).

3.6.2. Steroidal Tetracyclanes. Sleranes. Steranes cornmoniy found in mature sediments and crude oils are derived via diagenesis from sterols (XYID which are widely dispersed in plants and mierorganisms, with the en and C2B sterols most abundant in marine organisms and the ~ sterols in higher plants (2B). ClO 4-desmethylsterols oceur

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in marine invectebrates and marine algae (11), and 4-methylsterols are significantl present in dinoflagellates (29).

Generally specific information on the organisms which relies on the position L

double bonds and functional groups will be lost, prior to oil generation. However, mo' of the diagnostic information available from steranes in fossil fuel studies is mainJ based on the stereochemical complexity of their basic skeleton, which is ofien retaine and is observed in sediment extracts and oils.

Biomarker maturity determinations are generally based upon changes in steranes an triterpanes (7). Sterols generally contain a 5,6 double bond and occur naturally as tt. 20R epimer with the 14a(H), 17a(H) configuration. As diagenesis progresses the doub bond is hydrogenated and a new epimeric center with a mixture of 5a(H) and 58(H) formed with a predominance of the 5a(H) epimer due to its greater thermal stabilir; As the level of maturity increases the 148(H),178(H) isomers are formed as a mixtur of the 20R and 20S epimers.

Sorne low molecular weight steranes, such as ~I pregnane and en homopregnar. (XYill) have been used as indicators of hypersaline and carbonate environments (JO 1

Many parameters involving the complex sterane distribution for source, deposition environment, maturation, migration and biodegradation indicators have been proposc (2.1.32). However, some cautions concerning the use of some indicators has be.: enphasized by different authors (33.3<). In Table 1 the most widely used parameters al

summarized (2) Diasteranes (XIX) present in oils and source rocks in relatively low abundances éL

formed by acid-clay-catalyzed rearrangement of regular steranes, and therefore can t used for depositional environment determinations.

3.7. PENTAeyeLIe ALKANES

3.7.1. Hopane-type triterpanes (XX). These constitute the group of triterpanes me extensively used in biomarker studies. Their naturally occurring precursors, diplopte; and e" tetrahydroxyhopane, are widely distributed among bacteria, blue-green alga­sorne grasses and lichens and several feros (2).

Diagenesis and maturation of organic material containing the precursors leads defunCtionalization and change ofthe biological178(H),216(H) stereochemistry to t

thermodynamically more stable 17a(H),216(H) configuration. At the same tim formation of another series of hopane analogs, known as moretanes (XXI), with te 17.6(H),21a(H) configuration occurs.

The e31 and higher homologues can occur as either 22S and 22R epimers due lO t chical center at position en. Increasing maturation leads to mixtures of 22S and 2: (naturally occurring) epimers with an equilibrium ratio of approximately 3:2. The rat of the ~7 17a(H)-hopane (Tm), over the ~ 18a(H) (Ts), more resistant to therrr. maturation than Tm, is used as a maturity parameter of oil and source rocks samplt

Most of the samples show the presence of regular hopanes from e27 to e", with t

absence of the en homologue. The detection of this compound in a few notar exceptions have been associated with the specific environment of deposition, such

fable 1.- SlJIIDary of biomarkers parameters based on steranes and triterpanes that have been used in petrole\lll stmies, (Philp, 1985).

* tricyclic terpanes (%) * change in paraffin ooncentration plus increase in sterane and triterpane ooncentration * C 9 + C30 primary terpanes I C

2l + C 8 seoondary terpanes

* 1~a(H)-22,29,30-trisnorbopane rm)' 18a(H)-22,29,30 trlsnorhopane (Ts) * 56(H) steranes I 17a(H) hopanes * 176(H),216(H) / 176(H),216{H) + 178(H),21a(H) + 17a(H),216(H) hopanes * 22R / 22R + 22S 17a(H),216(H)-homohopanes * 208 / 208 + 20R 136(H),17a(H)-diacholestane * 248 / 248 + 24R (20R)-24-methyl-aaa-cholestane * 20R / 20R + 208 -24-ethyl-aaa-cholestane * a86 / a86 + aaa (20R + 20S) 24-ethylcholestane * aaa-c28 steranes / aaa-C29 steranes * aaa-C steranes / aaa-C29 steranes * Sao (~bR) + a86 (20R) / aaa (20R) C29 steranes * 136(H),17a(H)(208) / aaa (20R) C27 + C 8 + C29 steranes * Boa (20R)-C28 + a86 (20R+20S)-C28 + ~ (20R+20S)-C29 / aaa (20R)-C28 steranes * C27 trlaromatic steranes I C27 triaromatic + C28 monoaromatic steranes * C21 monoaraoatic steranes I C2l triaromatic + C28 monoaromatic steranes * C26 triaromatic steranes I C20 triaromatic t C27 triaromatic steranes * preferential removal of C27-208 diasterane over 20R eplmer * preferential removal of regular steranes over diasteranes * 17a(8)-C30 bopanes / 178(8)-(C29 + ClO) IOOretanes * 17a(8)/178(H)-trisnorhopanes

~lliill2n

Source Migratlon/Maturation Source/Maturation Source IMa tura tiOll Migration in comon sources Maturation Maturation Maturation Maturation IIaturation Maturation Source Source Hlgration Hlqration Source Maturatlon/aromatlzatlon Maturation/bond breaking IIaturation/bond breaking Blodegradation Blodegradation Source rocks by pyrol ysis Source rocles by pyrol ysis

aaa, aB6, B68, Sao refer to the stereochetnlstry of the H atOlll at the 5, 14 and 17 positions, respectively, of tbe regular steranes

Ul -.1

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mat structures formed by sulphide oxidizing bacteria in upwelling areas (35,36). In evaporite facies the enhanced reducing character of the environmeots leads h

greater preservation of the hopane side chain during diageoesis. In hypersaline settin the occurrence of e,. andlor el5 hopanes lS characteristic and are found in higt concentrations than the other homohopanes (30). A very strong predominance of ( hopanes has been reported 10 occur in samples derived from peats and coals. This r. been attributed to decarboxilation of en hopanoic acids wruch are abundant in su samples. High cooceotrations of moretanes have also been noted in them and res from the direct input of mor etanol and moretaoone into the sediment from land pi¿ sources.

Hopanes having an additionaI methyI substituent in ring A have been reported . numerous authors 10 occur in crude oils and sediments, most of them deposited carbonate and/or hypersaline environments. Summons and Jahnke (37) have recem shown that the most commonly reported methylhopanes are 2-methylhopanes. The ml significant parameters involving the hopanes are also iocluded in Table 1 (2).

3.7.2. Non-hopane type triterpanes. Lupanes (XXII), oleananes (XXlll), feman (XXlY) and ursanes (XXV) biomarkers characteristic of plant triterpenoids have be:c frecuently found in coals and oils derived from terrigenous source materials. Tht precursors are widely distributed in higher plants. e3Q bicadinanes (XXVI) found in 01

have been shown to be characteristic cracking products of angiosperm fossil resins. Tr 18a(H)-0Ieanane has been observed in a number of deltaic sediments and crude oi related to them (2-4,31,31). The biogenic precursors of oleanane are presumed to be oleaner triterpeooids such as 6-amyrin, which are associated with high1y specialized terrigenOl plants.

The origin of gammacerane (XXYlJ), peotacyclic triterpane, ofien used as é

indicator for salinity, is still unclear. TetrahymanoI, the only pentacyclic triterpeno fouod in protozoa and fungus, is the only knowo poteotiaI biological precursor , ganunacerane (39). As Gammacerane is preseot in many sampIes from varioL enviroomeots, its use as a biomarker for salinity is oot based on its occurrence, b rather 00 its relative abundance (31).

3.8. HEXAeyeLIe HOPANOID ALKANES

e31 -C35 bexacyclic hopanoid alkanes (XXVIII) have beeo used as an indicator for anOXl paIeoeovironmeots (40,41). These authors have suggested that hexacyclic alkanes wer derived from bacteriohopanetetrol (e35) precursors through cyclisatioo of the side chai ~ of extended hopanoids during earIy diagenesis.

The hexacyclic hopanes are very resistant to bacterial attack and may be very useft: for defining palaeoenvironments and biodegradation degree of source rocles and oils

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4. Use of Biomarkers in Oil Shale Geochemistry. Case study of the Puertollano (Spain) Oil Shale

For many years organic-geochemical studies were largely limited to tbe most economically important organic materials found buried in tbe Eartb, namely coals, oil shales and petroleum. Oil shales have a special geochemical interest botb as potential source rocles and as an energy resource in tbemselves, altbough most oil shale mining projects are currently non-viable commercially.

OH shales are a complex group of rocles which were deposited in a wide range of sedimentary environments and have a wide range of largely alginite-dominated maceral assemblages. According to a recent classification of oil shales as regards to tbeir relative abundance of macerals (42), tbey overlap witb botb coals and petroleum source rocles so tbat any classification of oil shales has to take tbese otber types into account. In tbis context tbe use of biomarker distribution as an alternative to tbe precise classification of tbe different types of oil shales might be taken into consideration.

Bitumens are present in some oH sbales such as tbe Green River Formation of tbe U .S.A. and tbe Irati Formation of Brazil. The bitumens can impregnate and may partially replace some macerals (43). Past work on tbe biomarkers in oil shale has involved characterization of tbe biomarkers in bitumen and determination of tbe mannec in which tbeir quantity varied witb deptb and otber characteristics of tbe sbale witbin a given coreo Many outstanding worles on tbe application of biomarkers foc source rocles-oil correlations have been carried out on tbese sedimentary rocles (44).

To show tbe utility for oil shale geochemistry of tbe biomarker approach, we have taken as specific case study the Puertollano deposit, tbe most important in tbe lberian Península, which until now has been only partially characterized from tbe geochemical viewpoint (45).

Table 3 shows tbe composition of tbe alipbatic extractable hydrocarbon fraction of tbis deposito Experimental metbods were similar to tbose recently used for characterization of peat and cOO deposits (46).

The hydrocarbon distribution in tbe Puertollano OH Shale is dominated by n-alkanes, n-alkylcyclohexanes, n-aIkyl,metbylcyclohexanes and mid-chain monomethyl alkanes. The cyclohexanes showed a distribution very similar to tbat of n-alkanes suggesting a common origino Triterpenoid distributions showed a major contribution of bacterially derived hopanoids. The compounds identified comprised Cn -C3, hopanes (no Cu), ~7-

C,2 8, 14-secohopanes, ~-C,2 ring-D aromatized 8,14-secohopanes, C,2-C3' benzohopanes and a C,I hexacyclic saturated hopane. The conifer and angiosperm derived material is expressed in two different groups of diagenetic compounds.

Diterpenoid compounds as abietane, norabietane, simonellite, retene and tbe sesquiterpenoid cadalene are characteristics of tbe conifer resios, whereas tbe terpanes with oleanane and ursane skeleton are characteristics of angiosperms. It is concluded from tbese results tbat tbe Puertollano oH shale has been formed in a lacustrine environment witb significant input of higher plant derived organic material.

Some parameters, 50ch as a) tbe abundance of tbe ~ and C30 primary terpanes relative to tbe otber members of the series, b) tbe ratio of ~ 176(H),21a(H)-moretane

Table 2.-Hain series of compounds detected in the saturated fraction of the Puertollano Oil Shale extracts.

Series Range Hain 0/001 Structure2

canponent

n-alkanes ClJ-C39 C23 420 iso-alkanes C16-C32 20 anteiso-alkanes C16-C 32 20 mid-chain methyl alkanes (16-C32 40 acyclic isoprenoids C15-C20 C19 70 I,Ir n-alkyl-cyclohexanes C13-C28 80 methyl-n-alkyl-cyclohexanes C18-C28 80 bicyclic terpanes CH-CI6 C16 40 V,VI diterpanes C19-C 20 35 VII, VIII, IX tricyclic triterpanes C19-C 28 C23 40 XI -XVII tetracyclic terpanes C24 C24 10 tetracyclic 17,21 secohopanes C2~ C24 10 XV 8,14 secohopanes C27- 30 5 XVI id. ring-D aromatized C27-C31 C29 5 steranes C27-C 29 10 XVII-XIX pentacyclic hopanes C27-C35 C30 100 XX benzohopanes C3(35 C32 10 hexacyclic hopanes 31 C31 5 XXVIII

(1) relative abundan ces (2) Roman numbers refers to the structures shown in Appendix 1

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lO 17a(H),21B(H)-hopane (0.37), c) the predominance of Tm over Ts, and d) the predominance ofthe biolo~ica1ly occuring aaR isomers over the maturation induced BB­isomers in \be mixture of regular and rearranged steranes are indicative of the low maturity level of this oil shale and the absence of biodegradation.

5. Future Work 00 Biomarkers

The promising future seen for biomarkers by Eglinton two decades ago (47) in basis lO the development of a oumber of applications has been largely realized. Today, oow that many of the molecular methods and ftndings have been adsorbed by the industry (common use of the so-called "biomarker strategy" in exploration studies), the interest in biomarkers has again moved to new applications and extensions of the basic molecular organic geochemical approach. Thus, biomarkers have been applied in the study of recent sediments, to marine sciences, archaeologica1 chemistry, biogeochemical ecology, biomolecular paleontology, molecular stratigraphy, etc.

Important progress in biomarkers geocbemistry is expected from future investigations on nonhydrocarbon polar fractions and in particular suIphur biomarkers having well established structural relationships with known precursors (41). It has been noted, however, that the "natural sulphurization" during the early stages of diagenesis may lead lO selective removal ofthe specific hydrocarbon biomarker precursor, and therefore to biased palaeoenvironmental reconstruction (~."'l

Of especial interest at present is the use of biomarkers in environmental and pollution studies, since these compounds provide information on the chemica1 and biologica1 degradation of organic molecules in modero environments. Improved abilities to monitor and control waste disposal from modero demands, ranging from the disposal of sewage to improved understanding of landfill waste disposal should be included. Biomarkers have also been useful for frngerprinting the source of oil spills and other organic wastes and tracing the source of this pollution.

A particular goal is lO link the patterns of the geologica1 record to the behaviour of the atmosphere, increasing our understanding of the fundamental causes of climate change. An outstanding precedent of these investigations is the use of biomarkers from certain marine aIgae for paleoclimatic reconstructions based on their ability to synthesize long chain unsaturated ketones (~7-C39) with the degree of unsaturation depending on the ocean water temperature (").

The interest in biomarkers depends on the distribution of biocbemical precursors in presentIy living organisms. It should be emphasized, however, that knowledge of the distribution of different precursors in contemporaneous organisms is still insufficient. Progress in this field and consequentIy in biomarkers geochemistry are dependent 00

the development and improvement of new technologies and novel analytica1 schemes to study the complex mixtures of chemica1s in geologica1 samples.

Good examples foc the oew frontiers in biomarker geochemistry are the recent identification of biomarkers of high molecular weigbts ( > C40) by using higb­temperature capillar gas chromalOgraphy ('2), and the development of reliable

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instruments for determination of carbon isotopie eompositions of individual eomponents of oils and rock extracts (53), which seem to be a useful para meter in the eorrelation of oil to oil or oil to source rock and as indicator to identify the precursor of biomarkers.

5. Acknowledgement

I thanks the Spanish Direccion General de Investigacion Científica y Tecnica (Project PB91-0074) for subsidizing this work. Dr .1. C. Del Rio gave invaluable encouragement and advice.

6. References

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and references therein. 3. C.G.BLANCO, J.G.PRADO, M.D.GUILLEN and A.G. BORREGO,Q[g,.

Geochem., 1992, 18,313. 4 . J.c. MONIN, D. BARTH, M. PERRUT, M. ESPITALIE and B. DURAND,

Or~.Geochem., 1988, 13, 1079. 5 . K.H. ALTGELT and T.H. GOUW (eds) , Chromaro¡:raphy in Perroleum

Analysis, Marcel Dekker (1979). 6 . C.M. WHITE, lntroduction ro mass spectrometric techniques Jor Jossil fuel

analysis in this volume 7 . A.S. MACKENZIE, Advances in Perro/eum Geochemistry, J.Brooks and D.H.

Welte (eds), Academic Press (1984). 8 . R.B. JOHNS, Biolo¡:ical Markers in lhe Sedimentary Record, Elsevier (1986). 9 . R.P. PHILP and C.A. LEWIS, Ann. Rev. Earth Planer Sci.,1987, 15,363. 10. J.M. MOLDOWAN, P. ALBRECHT and R.P. PHILP, Biolo¡:ical Markers i1

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