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Baccharane (18,19-secolupane): A rare triterpenoid skeleton widespread in Triassic sediments and petroleum from the Adriatic Basin
Geochimica e! Cosmochimica Acta Vol. 57, pp. 3201-3205 Copyright © 1993 Pergamon Press Ltd. Printed in U.S.A.
0016-7037/93/$6.00 + .00
LETI'ER
Baccharane (18,19-secolupane): A rare triterpenoid skeleton widespread in Triassic sediments and petroleum from the Adriatic Basin JACQUES POINSOT,I PIERRE ADAM,l JEAN M. TRENDEL,1 PIERRE ALBRECHT,L* and ANGELO RIVA2 qnstitut de Chimie, Universit6 Louis Pasteur, l rue Blaise Pascal, 67000 Strasbourg, France 2AGIP S.p.A., 20097 San Donato Milanese, Milano, Italy
(Received March 1, 1993; accepted in revisedform April 29, 1993) Abstract--Baccharane (18,19-secolupane) 1, a triterpenoid skeleton seldom encountered in living organisms, has been characterized together with 17/~H-des-E-lupane 2, one of its minor degradation products, in Italian Triassic sediment and crude oil samples from the Adriatic Basin by synthesis of standards. Reports of baecharane-derived triterpenoids in the field of natural products are scant and up to now restricted to terrestrial plants (angiosperms and ferns). In this regard, sedimentary baccharane derivatives may indicate a continental contribution (ferns?) to fossil organic matter; but they may also represent a contribution of as yet undetermined marine microorganisms, as has been proposed for dammarenes and dammaranes. Alternatively, they could be the bio/geochemical transformation products of biosynthetically closely related precursors such as lupane- or dammarane-type triterpenoids. The apparently restricted occurrence of baccharane derivatives in geological samples (up to now detected only in Triassic sediments of the Adriatic Basin) confers upon these compounds great value as correlation parameters and possibly as source indicators. INTRODUCTION
structural identification of novel biomarkers is essential to assess not only the origin of organic matter, but also transformations taking place in the subsurface. In this note, we report the occurrence ofbaccharane (l 8,19secolupane) 1 and of 17flH-des-E-lupane 2, a probable sidechain degradation product of baccharane, in Italian Triassic sediment and crude oil samples from the Adriatic Basin (MATTAVELL!and NOVELLI, 1990). They were identified by comparison with synthetic standards.
MANY KINDS of polycyclic triterpenoid hydrocarbons have been reported from sediments and petroleum, and these biological markers are now widely used in geochemical studies, either to assess the origin of organic matter and the palaeoenvironment of deposition, or as maturity and correlation parameters (PETERS and MOLDOWAN, 1993). Beside the ubiquitous hopanoid series of bacterial origin, they often appear as structurally well-individualized Ca0 compounds with occasional skeletal modifications. Some of these biomarkers have biological precursors and producing organisms known for a long time. For example, oleananes are the molecular fossils of higher plant triterpenes oxygenated at C-3, and their presence in geological samples would therefore indicate a terrestrial contribution to the sediments (WHITEHEAD,1974; EKWEOZORet al., 1979a, b; RIVA et al., 1987). On the other hand, others have their origin only recently better understood, or still remain orphan. This is the case of gammacerane (HILLS et al., 1966), the most likely precursor of which, tetrahymanol, has been lately shown to be widespread in marine sediments and in marine ciliates (TEN HAVENet al., 1989; VENKATESAN, 1989; HARVEY and MCMANUS, 1991). Dammarenes (and dammaranes) are a good illustration of compounds which are widely distributed in marine sediments, but the origin of which still is a matter of speculation, although a microbial source appears likely (MEUNIER-CHRISTMANNet al., 1991 ). Furthermore, the precise geological fate of dammarenes remains somewhat unclear. In this respect, the unequivocal
EXPERIMENTAL Sample Extraction and Fractionation
One carbonate source rock (central Italy) and several crude oils from the Adriatic Basin were analyzed. Organic extract (toluenemethanol 3:1 and then chloroform-methanol 3:1, twice at 50°C for 1 h) or crude oils were fractionated over silica gel (columns or plates). Hexane was used to elute the less-polar fraction containing the nonaromatic hydrocarbons. Molecular sieving (5 /~) of the latter afforded the branched and cyclichydrocarbonssubsequentlyanalyzed by GC-MS. Physical Measurements
NMR spectroscopy NMR spectra were taken on a Bruker AM-400 spectrometer operating at observation frequencies of 400 MHz and 100 MHz for ~H and a3Cnuclei,respectively,and data were recorded at 27°C. Chemical shifts (6) are reported in ppm from tetramethylsilane,usingthe solvent (C~Dr:6~H 7.16, 613C 128.04) as internal reference. Additional NMR experiments ('3C broad band ~H decoupled and DEPT spectra, homonuclear ~H-~HCOSY and NOESY, heteronuclear inverse onebond and long-range 2'3j~H-~3Ccorrelation spectra) were performed on the final hydrocarbon standards in order to ensure their stereochemistry (CrD6 was used as deuterated solvent instead of
* Author to whom correspondence should be addressed. 3201
3202
J. Poinsot et al.
CDC13, as it gave a considerably better splitting of methyl group signals).
100-
Gas chromatography-mass spectrometry
80-
Mass spectra of synthetic compounds and of sedimentary hydrocarbons were recorded on a Finnigan MAT TSQ 70 spectrometer operating at 70 eV and connected to a Varian 3400 gas chromatograph equipped with an on-column injector, a J & W DB-5 column (60 m × 0.25 mm, 0.1 um film thickness), and using He as carrier gas. Coinjections of synthetic standards and natural hydrocarbons were performed on the apolar DB-5 column and on a polar SUPELCOWAX column (60 m × 0.25 mm, 0.1 t~m film thickness).
60-
Synthesis of Reference Hydrocarbons
4055 20-
81 [ 109 137
329
i ,I .IL,[[ ~,I, ~ 11!4.9,L 205233t.t. 1oo
100
Baccharane was prepared according to a previously described synthetic route (SUOKASand HASE, 1971) slightly modified and using, in our case, lup-20(29)-ene 3 as starting material (Fig. I). Dehydrogenation of lup-20(29)-ene 3 with mercuric acetate (ALUSON et al., 1961) afforded lupa-18,20(29)-diene 4, which was selectively hydrogenated into lup-I 8-ene 5. Treatment of the latter with ruthenium tetra-oxide led to the diketone 6, the bis-ethylene thioketal 7 of which gave baccharane I after Raney-Ni desulfurization. 17flHDes-E-lupane 2 was obtained starting from the diketone 6, which was submitted to a vapour-phase retro-Michael reaction (AQUINO NETO et al., 1986), affording a mixture (25:75) of the two ketones 8 and 9 readily separable by silica gel TLC (CH2C12). Raney-Ni desulfurization of the ethylene thioketal 10 furnished the 17BH-des-Elupane 2. All synthetic intermediates gave satisfactory and/or expected analytical data (m.p., tH-NMR, MS, UV, TLC-Rf), and the mass spectrum ofbaccharane I (Fig. 2b) is virtually identical to that previously published (ANTHONSENet al., 1970). Baccharane 1: ~H-NMR 6 1.75 (IH, dddd, J = 12.2, 12.2, 3.7, 3.7 Hz, H-13), 1.047 (3H, s, 26-CH3), 0.951 (3H, s, 28-CH3), 0.939 (6H, 2xd, J = 6.7 Hz, 29-CH3 and 30CH3), 0.915 (6H, 2xs, 27-CH3 and 23-CH3), 0.897 (3H, s, 25-CH3), 0.871 (3H, s, 24-CH3). t3C-NMR 6 33.62 (C-23), 33.21 (C-13), 23.24 (C-28), 22.92 (C-29 and C-30), 21.82 (C-24), 16.69 (C-25), 16.01 (C26), 13.49 (C-27). 17~H-des-E-lupane 2: IH-NMR ~ 1.47 (IH, dddd, J = 12.0, 12.0, 3.5, 3.5 Hz, H-13), 1.001 (3H, s, 26-CH3), 0.928 (3H, d, J = 6.3 Hz, 28-CH3), 0.908 (3H, s, 23-CH3), 0.898 (3H, d, J = 0.8 Hz, 25-CH3), 0.879 (3H, d, J = 0.6 Hz, 27-CH3), 0.864 (3H, s, 24CH3). ~3C-NMR ~ 38.26 (C-13), 33.61 (C-23), 22.91 (C-28), 21.82 (C-24), 16.71 (C-25), 16.01 (C-26), 14.18 (C-27).
191
200
b]
191
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400
m/z
400
.220
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lOO C_~
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191
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I
136
9° 100
200
° 300
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FIG. 2. Mass spectra (70 eV) of synthetic references: (a) 17,21secohopane 11; (b) baccharane 1; (c) 17BH-des-E-lupane 2.
RESULTS AND DISCUSSION
29
Identification of Baccharane 1 and 17~H-des-E-lupane 2 a,b
.
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FIG. 1. Synthesis of baccharane 1 and of 17flH-des-E-lupane 2 starting from lup-20(29)-ene 3: (a) Hg(OAc)2/AcOH; (b) HES/pyridine; (c) H2, PtO2/hexane; (d) RuO4/CCI4; (e) ethane- 1,2-dithiol, BFa-ethcrate; (t) Raney-Ni, H2/toluene-ethanol; (g) NaOH, cotton wool, 300°C.
A C30 tetracyclic terpane, frequently detected in crude oils of the Adriatic Basin, was first tentatively identified, on the basis of mass spectral data, as being 17,21-secohopane 11 (RIVA et al., 1989). Indeed, its mass spectrum (Fig. 2b) displays a prominent base peak at m/z 191, typical of the polycyclic terpane ring A/B moiety, and a characteristic fragment at m/z 329, usually diagnostic of the loss of the side chain of 17,21-secohopanes (TRENDEL et al., 1982; AQUINO NETO et al., 1983). More precise studies, however, showed small but significant differences between its mass spectrum and that of synthetic 17,21-secohopane 11 (Fig. 2a,b), and also completely different G C retention times for the two compounds which often co-occur in the samples from the Adriatic Basin, as illustrated by the partial m/z 191 mass fragmentogram of a typical petroleum from the Adriatic Sea (Fig. 3). Further investigations revealed that the unknown C30 product could have the baccharane (18,19-secolupane) skeleton 1, as both display quite similar mass spectra (ANTHONSEN et al., 1970). This c o m p o u n d is often accompanied by a very minor C24
Baccharane in petroleum from the Adriatic Basin
m/z
3203
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FIG. 3. Typical m/z 191 mass fragmentogram of the branched and cyclic alkane fraction of a petroleum from the Adriatic Basin. tetracyclic hydrocarbon of GC retention time slightly shorter than that of the C24 17,21-secohopane 12 (Fig. 3), and with a mass spectum (Fig. 2c) similar to the latter (AQuINO NETO et al., 1983). As this unknown C24 hydrocarbon occurs in samples only when the C3o hydrocarbon is present, it appeared reasonable that it could derive from the latter by removal of the side chain. These hypotheses were confirmed by comparison of the synthetic standards I and 2 with the sedimentary structures. Natural and synthetic compounds show quasi superimposable mass spectra and coeluted on apolar and polar GC columns, thus demonstrating their identity (see Experimental section).
baccharane skeleton could originate directly from marine microorganisms as a result, for instance, of a proton-induced cyclization ofsqualene leading, through a dammarenyl cation, to a baccharadiene subsequently hydrogenated in sediments. However, there is also evidence for a terrigenous input to our samples, as indicated by optical investigations on the kerogen of the rock, and therefore a continental contribution of organisms such as ferns may also be taken into consideration, although no other structures (except hopanes !) usually present in extant ferns could be detected. Alternatively, the baccharane skeleton could stem from a secondary transformation process undergone by
Origin of Baccharane and 17/3H-des-E-lupane
Baccharane-type triterpenes have been rarely reported in the field of natural products, and such a skeleton has, until about ten years ago, only been postulated as an intermediate leading to partial (baccharis oxide; ANTHONSEN et al., 1970) or fully rearranged (shionone; TAKAHASHIet al., 1967) structures. Hosenkol-A is the first example of a natural baccharane triterpenoid (SHOJI et al., 1983), and since then, reports of the natural occurrence of this skeleton in angiosperms are scant (e.g., FUJIOKA et al., 1988). However, during the last decade, unsaturated baccharane-type hydrocarbons have been identified in some genera of ferns (e.g., Lemmaphyllum, MASUDA et al., 1983; Microsorium, AGETA et al., 1992). They occur together with a large variety of triterpenoid structures (ferneries, dammarenes, etc.) quite frequently encountered in sediments. Various hypotheses can be put forward to explain the occurrence ofbaccharane in geological samples. The latter could either have a direct biological origin or result from diagenetic processes (Fig. 4). Indeed, as it has been proposed for dammarenes (and dammaranes) (MEUNIER-CHRISTMANN et al., 1991), the
continentalorganisms1 /
,ern ? ,
e
~diagenesis
marine 1 organisms
~
~agenesis ? ~
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1
-
Cupane
derivative
esis "
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Dammarenes
FIG. 4. Possible origins of baccharane 1 and 17/3H-des-E-lupane2.
3204
J. Poinsot et al.
biosynthetically closely related products. Thus, on one hand, b a c c h a r a n e - - w h i c h biosynthetically is a prelupane (RuzICKA, 1959)--could find its origin in the (bio)degradation of a lupane skeleton adequately functionalized, by cleavage of the C(18)-C(19) bond. Lupane derivatives are normally characteristic of angiosperms (e.g., MAHATO et al., 1992), and, as our samples (Late Trias) predate their evolution (Cretaceous), such a hypothesis would appear unlikely. Nevertheless, it should be kept in mind that triterpenoid structures usually considered as typical of angiosperms (i.e., lupane, oleanane) have also been found in more primitive groups of terrestrial plants such as ferns (AGETA and ARAI, 1983; AGETA et al., 1992) and consequently are not so strictly characteristic of flowering plants as generally presumed. It follows that it may be theoretically possible to find lupane and oleanane skeletons and their derivatives in sediments older than Cretaceous. On the other hand, the baccharane skeleton could be formed by ring D enlargement, via a C-20 dammaranyl carbonium intermediate, of d a m m a r a n e derivatives such as dammaran-20-ols or even dammar-13(17)-enes. Although laboratory experiments aiming at the chemically induced rearrangement of the C-20 cation of the dammarane-type into baccharane-type skeleton have failed (TORI et al., 1977), clay-catalyzed reactions may still lead to such a rearrangement since it has been shown (SIESKIND et al., 1979) that diaster13( 17)-enes could undergo D-homoannulation when treated with montmorillonite K10 clay. In this respect, the baccharane backbone could be viewed as a result of the diagenetic transformation of dammarenes. Several observations, however, will argue against this hypothesis. At first, to our knowledge, ring D homologated diasteranes have not yet been conclusively identified in sediments, showing that ring D enlargement is a process which does not currently take place under natural conditions. Furthermore, the occurrence of baccharane seems strictly restricted to the Adriatic Basin and therefore cannot be considered as a usual transformation product of the relatively ubiquitous sedimentary dammarenes. Moreover, the often pure carbonate depositional environment of our samples would not appear especially propitious to promote such a transposition (usually only very low quantities of diasteranes are present), and no further rearranged backbones (e.g., shionane) could be detected. Lastly, no correlation seems to exist between the occurrence in sediments of d a m m a r e n e s / d a m m a r a n e s and that of baccharane. F r o m all these considerations, it appears that the presence of baccharane in the Adriatic Basin would most likely reflect a specific source rather than diagenetic transformations of structurally related compounds. Whether this source is of marine or continental nature remains, of course, to be established. CONCLUSIONS 1) The structures of two tetracyclic hydrocarbons widespread in Italian Triassic sediments and petroleum of the Adriatic basin, baccharane 1 and 17/3H-des-E-lupane 2, have been conclusively identified by comparison with synthetic references. This constitutes the first report ofbaccharane derivatives from a geological source.
2) From their restricted sedimentary occurrence and the nature of the depositional environment, these molecular fossils most likely derive from biological precursors bearing the baccharane skeleton rather than being diagenetic transformation products of biosynthetically closely related products such as dammarenes. A contribution of either marine or continental organisms can be envisaged. 3) Due to its restricted distribution in geological samples baccharane could turn out to be a highly specific biomarker in terms of source and correlation parameter. Acknowledgments--We thank S. Marc, M. Edmond, and M. C. Schweigert for their much appreciated technical assistance, and AGIP S.p.A. for financial support and permission to publish. We also thank R. P. Philp, J. Rullkrtter, R. Summons, and P. Sundararaman for their helpful comments in reviewing the manuscript. Editorial handling. G. Faure
REFERENCES AGETA H. and ARA1Y. (1983) Fern constituents: Pentacyclic triterpenoids isolated from Polypodium niponicum and P. formosanum. Phytochem. 22, 180 l- 1808. AGETA H., ARAI Y., and MASUDAK. (1992) Chemical and chemotaxonomical studies on fern triterpenoids in Polypodiaceae. Abstracts of the 18th IUPAC Symposium on the Chemistry of Natural Products, Strasbourg (France), 30.8-4.9.1992, p. 591. ALLISONJ. M., LAWRIEW., MCLEANJ., and BEATONJ. M. (1961) Dehydrogenation with mercuric acetate in the lupane series: Part II. Lupeol and its derivatives. J. Chem. Soc., 5224-5230. ANTHONSEN T., BRUUN T., HEMMER E., HOLME D., LAMVIK A., SUNDE E., and S~RENSEN N. A. (1970) Baccharis oxide, a new triterpenoid from Baccharis halim~/blia L. Acta Chem. Scand. 24, 2479-2488. AQUINO NETO F. R., TRENDELJ. M., RESTLEA., CONNANJ., and ALBRECHTP. A. (1983) Occurrence and formation oftricyclic and tetracyclic terpanes in sediments and petroleum. In Advances in Organic Geochemistry 1981 (ed. M. BJOP~OYet al.), pp. 659-667. J. Wiley & Sons. AQUINO NETO F. R., TRENDEL J. M., and ALBRECHT P. (1986) Synthetic intermediates derived from triterpenoids by the retroMichael reaction in the vapour phase. Tetrahedron 42, 5621-5626. EKWEOZOR C. M., OKOGUN J. I., EKONG D. E. U., and MAXWELL J. R. (1979a) Preliminary organic geochemical studies of samples from the Niger delta (Nigeria): I. Analyses of crude oils for triterpanes. Chem. Geol. 27, 11-28. EKWEOZORC. M., OKOGUNJ. I., EKONG D. E. U., and MAXWELL J. R. (! 979b) Preliminary organic geochemical studies of samples from the Niger delta (Nigeria): If. Analyses of shale for triterpenoid derivatives. Chem. Geol. 27, 29-37. FUJIOKA T., IWAMOTO M., IWASE Y., OKABE H., MIHASHI K., and YAMAUCHIT. (1988) Studies on the constituents of Actinostemma lobatum Maxim: Ill. Structures of Actinostemmosides E and F, new baccharane-type triterpene glycosides isolated from the herb. Chem. Pharm. Bull. 36, 2772-2777. HARVEY H. R. and MCMANUS G. B. (1991) Marine ciliates as a widespread source of tetrahymanol and hopan-3/3-ol in sediments. Geochim. Cosmochim. Acta 55, 3387-3390. HILLS I. R., WHITEHEADE. V., ANDERSD. E., CUMMINSJ. J., and ROBINSON W. E. (1966) An optically active triterpane, gammacerane in Green River, Colorado, oil shale bitumen. J. Chem. Soe., Chem. Commun., 752-754. MAHATO S. B., NANDY A. K., and RoY G. (1992) Tfiterpenoids. Phytochem. 31, 2199-2249. MASUDAK., SHIOJIMAK., and AGETAH. (1983) Fern constituents: Six tetracyclic triterpenoid hydrocarbons having different carbon skeletons, isolated from Lemmaphyllum microphyllum var. obovatum. Chem. Pharm. Bull. 31, 2530-2533. MATTAVELLIL. and NOVELLI L. (1990) Geochemistry and habitat of the oils in Italy. AAPG Bull. 69, 1623-1639.
Baccharane in petroleum from the Adriatic Basin MEUNIER-CHRISTMANN C., ALBRECHT P., BRASSELL S. C., TEN HAVEN H. L., VAN DER LINDEN B., RULLKOTTERJ., and TRENDEL J. M. ( 1991 ) Occurrence of dammar- 13(17)-enes in sediments: Indications for a yet unrecognized microbial constituent? Geochim. Cosmochim. Acta 55, 3475-3483. PETERS K. E. and MOLDOWANJ. M. (1993) The Biomarker Guide. Prentice Hall. RlVA A., CACCIALANZAP. G., and QUAGLIAROLIF. (1987) Recognition of 18/3(H) oleanane in several crudes and Tertiary-Upper Cretaceous sediments. Definition of a new maturity parameter. Org. Geochem. 13, 671-675. RIVA A., RIOLO J., MYCKE B., OCAMPO R., CALLOT H. J., ALBRECHT P., and NALI R. (1989) Molecular parameters in Italian carbonate oils: Reconstruction of past depositional environments. 14th International Meeting on Organic Geochemistry, Paris, 18 22.9.1989, Abstr. 335. RUZICKA L. (1959) History of the isoprene rule. Proc. Chem. Soc., 341-360. SHOJ1 N., UMEYAMAA., TAIRA Z., TAKEMOTO T., NOMOTO K., MIZUKAWA K., and OHIZUMI Y. (1983) Chemical structure of hosenkol-A, the first example of the natural baccharane triterpenoid of the missing intermediate to shionane and lupane. J. Chem. Soc., Chem. Commun.. 871-873. SIESK1ND O., JOLY G., and ALBRECHT P. (1979) Simulation of the
3205
geochemical transformations of sterols: Superacid effect of clay minerals. Geochim. Cosmochim. Acta 43, 1675-1679. SUOKASE. and HASE T. (1971) The stereochemistry at C-17 of baccharis oxide. The synthesis of baccharan-3(3-ol and baccharan-3one. Acta Chem. Scand. 25, 2359-2360. TAKAHASHI T., MORIYAMA Y., TANAHASHIY., and OURISSON G. (1967) The structure of shionone. Tetrahedron Lett., 2991-2996. TEN HAVEN H. L., ROHMER M., RULLKOTTERJ., and BISSERETP. (1989) Tetrahymanol, the most likely precursor of gammacerane, occurs ubiquitously in marine sediments. Geoehim. Cosmochim. dcta 53, 3073-3079. TORI M., TSUYUKI T., and TAKAHASHI T. (1977) The reaction of dammarane derivatives: Reactions involving C-20 earbonium ions. Bull. Chem. Soc. Japan 50, 3349-3359. TRENDEL J. M., RESTLE A., CONNAN J., and ALBRECHT P. (1982) Identification of a novel series of tetracyclic terpene hydrocarbons (C24 - C27) in sediments and petroleum. Y. Chem. Soc., Chem. Commun., 304-306. VENKATESANM. I. (1989) Tetrahymanol: Its widespread occurrence and geochemical significance. Geochim. Cosmoehim. Acta 53, 3095-3101. WH ITEHEAD E. V. ( 1974) The structure of petroleum pentacyclanes. In Advances in Organic Geochemistry 1973 (ed. B. TISSOT and F. BIENNER), pp. 225-243. Editions Technip.
0016-7037/93/$6.00 + .00
LETI'ER
Baccharane (18,19-secolupane): A rare triterpenoid skeleton widespread in Triassic sediments and petroleum from the Adriatic Basin JACQUES POINSOT,I PIERRE ADAM,l JEAN M. TRENDEL,1 PIERRE ALBRECHT,L* and ANGELO RIVA2 qnstitut de Chimie, Universit6 Louis Pasteur, l rue Blaise Pascal, 67000 Strasbourg, France 2AGIP S.p.A., 20097 San Donato Milanese, Milano, Italy
(Received March 1, 1993; accepted in revisedform April 29, 1993) Abstract--Baccharane (18,19-secolupane) 1, a triterpenoid skeleton seldom encountered in living organisms, has been characterized together with 17/~H-des-E-lupane 2, one of its minor degradation products, in Italian Triassic sediment and crude oil samples from the Adriatic Basin by synthesis of standards. Reports of baecharane-derived triterpenoids in the field of natural products are scant and up to now restricted to terrestrial plants (angiosperms and ferns). In this regard, sedimentary baccharane derivatives may indicate a continental contribution (ferns?) to fossil organic matter; but they may also represent a contribution of as yet undetermined marine microorganisms, as has been proposed for dammarenes and dammaranes. Alternatively, they could be the bio/geochemical transformation products of biosynthetically closely related precursors such as lupane- or dammarane-type triterpenoids. The apparently restricted occurrence of baccharane derivatives in geological samples (up to now detected only in Triassic sediments of the Adriatic Basin) confers upon these compounds great value as correlation parameters and possibly as source indicators. INTRODUCTION
structural identification of novel biomarkers is essential to assess not only the origin of organic matter, but also transformations taking place in the subsurface. In this note, we report the occurrence ofbaccharane (l 8,19secolupane) 1 and of 17flH-des-E-lupane 2, a probable sidechain degradation product of baccharane, in Italian Triassic sediment and crude oil samples from the Adriatic Basin (MATTAVELL!and NOVELLI, 1990). They were identified by comparison with synthetic standards.
MANY KINDS of polycyclic triterpenoid hydrocarbons have been reported from sediments and petroleum, and these biological markers are now widely used in geochemical studies, either to assess the origin of organic matter and the palaeoenvironment of deposition, or as maturity and correlation parameters (PETERS and MOLDOWAN, 1993). Beside the ubiquitous hopanoid series of bacterial origin, they often appear as structurally well-individualized Ca0 compounds with occasional skeletal modifications. Some of these biomarkers have biological precursors and producing organisms known for a long time. For example, oleananes are the molecular fossils of higher plant triterpenes oxygenated at C-3, and their presence in geological samples would therefore indicate a terrestrial contribution to the sediments (WHITEHEAD,1974; EKWEOZORet al., 1979a, b; RIVA et al., 1987). On the other hand, others have their origin only recently better understood, or still remain orphan. This is the case of gammacerane (HILLS et al., 1966), the most likely precursor of which, tetrahymanol, has been lately shown to be widespread in marine sediments and in marine ciliates (TEN HAVENet al., 1989; VENKATESAN, 1989; HARVEY and MCMANUS, 1991). Dammarenes (and dammaranes) are a good illustration of compounds which are widely distributed in marine sediments, but the origin of which still is a matter of speculation, although a microbial source appears likely (MEUNIER-CHRISTMANNet al., 1991 ). Furthermore, the precise geological fate of dammarenes remains somewhat unclear. In this respect, the unequivocal
EXPERIMENTAL Sample Extraction and Fractionation
One carbonate source rock (central Italy) and several crude oils from the Adriatic Basin were analyzed. Organic extract (toluenemethanol 3:1 and then chloroform-methanol 3:1, twice at 50°C for 1 h) or crude oils were fractionated over silica gel (columns or plates). Hexane was used to elute the less-polar fraction containing the nonaromatic hydrocarbons. Molecular sieving (5 /~) of the latter afforded the branched and cyclichydrocarbonssubsequentlyanalyzed by GC-MS. Physical Measurements
NMR spectroscopy NMR spectra were taken on a Bruker AM-400 spectrometer operating at observation frequencies of 400 MHz and 100 MHz for ~H and a3Cnuclei,respectively,and data were recorded at 27°C. Chemical shifts (6) are reported in ppm from tetramethylsilane,usingthe solvent (C~Dr:6~H 7.16, 613C 128.04) as internal reference. Additional NMR experiments ('3C broad band ~H decoupled and DEPT spectra, homonuclear ~H-~HCOSY and NOESY, heteronuclear inverse onebond and long-range 2'3j~H-~3Ccorrelation spectra) were performed on the final hydrocarbon standards in order to ensure their stereochemistry (CrD6 was used as deuterated solvent instead of
* Author to whom correspondence should be addressed. 3201
3202
J. Poinsot et al.
CDC13, as it gave a considerably better splitting of methyl group signals).
100-
Gas chromatography-mass spectrometry
80-
Mass spectra of synthetic compounds and of sedimentary hydrocarbons were recorded on a Finnigan MAT TSQ 70 spectrometer operating at 70 eV and connected to a Varian 3400 gas chromatograph equipped with an on-column injector, a J & W DB-5 column (60 m × 0.25 mm, 0.1 um film thickness), and using He as carrier gas. Coinjections of synthetic standards and natural hydrocarbons were performed on the apolar DB-5 column and on a polar SUPELCOWAX column (60 m × 0.25 mm, 0.1 t~m film thickness).
60-
Synthesis of Reference Hydrocarbons
4055 20-
81 [ 109 137
329
i ,I .IL,[[ ~,I, ~ 11!4.9,L 205233t.t. 1oo
100
Baccharane was prepared according to a previously described synthetic route (SUOKASand HASE, 1971) slightly modified and using, in our case, lup-20(29)-ene 3 as starting material (Fig. I). Dehydrogenation of lup-20(29)-ene 3 with mercuric acetate (ALUSON et al., 1961) afforded lupa-18,20(29)-diene 4, which was selectively hydrogenated into lup-I 8-ene 5. Treatment of the latter with ruthenium tetra-oxide led to the diketone 6, the bis-ethylene thioketal 7 of which gave baccharane I after Raney-Ni desulfurization. 17flHDes-E-lupane 2 was obtained starting from the diketone 6, which was submitted to a vapour-phase retro-Michael reaction (AQUINO NETO et al., 1986), affording a mixture (25:75) of the two ketones 8 and 9 readily separable by silica gel TLC (CH2C12). Raney-Ni desulfurization of the ethylene thioketal 10 furnished the 17BH-des-Elupane 2. All synthetic intermediates gave satisfactory and/or expected analytical data (m.p., tH-NMR, MS, UV, TLC-Rf), and the mass spectrum ofbaccharane I (Fig. 2b) is virtually identical to that previously published (ANTHONSENet al., 1970). Baccharane 1: ~H-NMR 6 1.75 (IH, dddd, J = 12.2, 12.2, 3.7, 3.7 Hz, H-13), 1.047 (3H, s, 26-CH3), 0.951 (3H, s, 28-CH3), 0.939 (6H, 2xd, J = 6.7 Hz, 29-CH3 and 30CH3), 0.915 (6H, 2xs, 27-CH3 and 23-CH3), 0.897 (3H, s, 25-CH3), 0.871 (3H, s, 24-CH3). t3C-NMR 6 33.62 (C-23), 33.21 (C-13), 23.24 (C-28), 22.92 (C-29 and C-30), 21.82 (C-24), 16.69 (C-25), 16.01 (C26), 13.49 (C-27). 17~H-des-E-lupane 2: IH-NMR ~ 1.47 (IH, dddd, J = 12.0, 12.0, 3.5, 3.5 Hz, H-13), 1.001 (3H, s, 26-CH3), 0.928 (3H, d, J = 6.3 Hz, 28-CH3), 0.908 (3H, s, 23-CH3), 0.898 (3H, d, J = 0.8 Hz, 25-CH3), 0.879 (3H, d, J = 0.6 Hz, 27-CH3), 0.864 (3H, s, 24CH3). ~3C-NMR ~ 38.26 (C-13), 33.61 (C-23), 22.91 (C-28), 21.82 (C-24), 16.71 (C-25), 16.01 (C-26), 14.18 (C-27).
191
200
b]
191
[ 300
399414t m/z
400
m/z
400
.220
806040-
81 5~
/ 109135
1O0
200
lOO C_~
300
191
8081 60-
I
136
9° 100
200
° 300
m/z
400
FIG. 2. Mass spectra (70 eV) of synthetic references: (a) 17,21secohopane 11; (b) baccharane 1; (c) 17BH-des-E-lupane 2.
RESULTS AND DISCUSSION
29
Identification of Baccharane 1 and 17~H-des-E-lupane 2 a,b
.
c
•
,
4
28
("s s/'~ 29
.'~'J
~
5
f
O
,
1
o
1°o
,
7
6
23 24
lg ,r'~.s
o
o
FIG. 1. Synthesis of baccharane 1 and of 17flH-des-E-lupane 2 starting from lup-20(29)-ene 3: (a) Hg(OAc)2/AcOH; (b) HES/pyridine; (c) H2, PtO2/hexane; (d) RuO4/CCI4; (e) ethane- 1,2-dithiol, BFa-ethcrate; (t) Raney-Ni, H2/toluene-ethanol; (g) NaOH, cotton wool, 300°C.
A C30 tetracyclic terpane, frequently detected in crude oils of the Adriatic Basin, was first tentatively identified, on the basis of mass spectral data, as being 17,21-secohopane 11 (RIVA et al., 1989). Indeed, its mass spectrum (Fig. 2b) displays a prominent base peak at m/z 191, typical of the polycyclic terpane ring A/B moiety, and a characteristic fragment at m/z 329, usually diagnostic of the loss of the side chain of 17,21-secohopanes (TRENDEL et al., 1982; AQUINO NETO et al., 1983). More precise studies, however, showed small but significant differences between its mass spectrum and that of synthetic 17,21-secohopane 11 (Fig. 2a,b), and also completely different G C retention times for the two compounds which often co-occur in the samples from the Adriatic Basin, as illustrated by the partial m/z 191 mass fragmentogram of a typical petroleum from the Adriatic Sea (Fig. 3). Further investigations revealed that the unknown C30 product could have the baccharane (18,19-secolupane) skeleton 1, as both display quite similar mass spectra (ANTHONSEN et al., 1970). This c o m p o u n d is often accompanied by a very minor C24
Baccharane in petroleum from the Adriatic Basin
m/z
3203
191
100% .
Q
C29Ts ~J~
(~
[
[[[
~1~
C26 oo
o 2 o
z4o
ITll 1 = /IJo I• C30
•
•
26o
c~[3-Hopanes
o• •o
Hexahydrobenzohopanes 30-Norhopanes Di aZhopanes C33 17, 1-Secohopanes Tricyclopolyprenanes
c;,/ Gammacerane
Cz7
?
•
V I~ot-Hopanes
c,,
r
•
J
•
•
•
C35
V
28o
oc
360
FIG. 3. Typical m/z 191 mass fragmentogram of the branched and cyclic alkane fraction of a petroleum from the Adriatic Basin. tetracyclic hydrocarbon of GC retention time slightly shorter than that of the C24 17,21-secohopane 12 (Fig. 3), and with a mass spectum (Fig. 2c) similar to the latter (AQuINO NETO et al., 1983). As this unknown C24 hydrocarbon occurs in samples only when the C3o hydrocarbon is present, it appeared reasonable that it could derive from the latter by removal of the side chain. These hypotheses were confirmed by comparison of the synthetic standards I and 2 with the sedimentary structures. Natural and synthetic compounds show quasi superimposable mass spectra and coeluted on apolar and polar GC columns, thus demonstrating their identity (see Experimental section).
baccharane skeleton could originate directly from marine microorganisms as a result, for instance, of a proton-induced cyclization ofsqualene leading, through a dammarenyl cation, to a baccharadiene subsequently hydrogenated in sediments. However, there is also evidence for a terrigenous input to our samples, as indicated by optical investigations on the kerogen of the rock, and therefore a continental contribution of organisms such as ferns may also be taken into consideration, although no other structures (except hopanes !) usually present in extant ferns could be detected. Alternatively, the baccharane skeleton could stem from a secondary transformation process undergone by
Origin of Baccharane and 17/3H-des-E-lupane
Baccharane-type triterpenes have been rarely reported in the field of natural products, and such a skeleton has, until about ten years ago, only been postulated as an intermediate leading to partial (baccharis oxide; ANTHONSEN et al., 1970) or fully rearranged (shionone; TAKAHASHIet al., 1967) structures. Hosenkol-A is the first example of a natural baccharane triterpenoid (SHOJI et al., 1983), and since then, reports of the natural occurrence of this skeleton in angiosperms are scant (e.g., FUJIOKA et al., 1988). However, during the last decade, unsaturated baccharane-type hydrocarbons have been identified in some genera of ferns (e.g., Lemmaphyllum, MASUDA et al., 1983; Microsorium, AGETA et al., 1992). They occur together with a large variety of triterpenoid structures (ferneries, dammarenes, etc.) quite frequently encountered in sediments. Various hypotheses can be put forward to explain the occurrence ofbaccharane in geological samples. The latter could either have a direct biological origin or result from diagenetic processes (Fig. 4). Indeed, as it has been proposed for dammarenes (and dammaranes) (MEUNIER-CHRISTMANN et al., 1991), the
continentalorganisms1 /
,ern ? ,
e
~diagenesis
marine 1 organisms
~
~agenesis ? ~
o.o
l ..... "
1
-
Cupane
derivative
esis "
~ , 1~ L9 ~
Dammarenes
FIG. 4. Possible origins of baccharane 1 and 17/3H-des-E-lupane2.
3204
J. Poinsot et al.
biosynthetically closely related products. Thus, on one hand, b a c c h a r a n e - - w h i c h biosynthetically is a prelupane (RuzICKA, 1959)--could find its origin in the (bio)degradation of a lupane skeleton adequately functionalized, by cleavage of the C(18)-C(19) bond. Lupane derivatives are normally characteristic of angiosperms (e.g., MAHATO et al., 1992), and, as our samples (Late Trias) predate their evolution (Cretaceous), such a hypothesis would appear unlikely. Nevertheless, it should be kept in mind that triterpenoid structures usually considered as typical of angiosperms (i.e., lupane, oleanane) have also been found in more primitive groups of terrestrial plants such as ferns (AGETA and ARAI, 1983; AGETA et al., 1992) and consequently are not so strictly characteristic of flowering plants as generally presumed. It follows that it may be theoretically possible to find lupane and oleanane skeletons and their derivatives in sediments older than Cretaceous. On the other hand, the baccharane skeleton could be formed by ring D enlargement, via a C-20 dammaranyl carbonium intermediate, of d a m m a r a n e derivatives such as dammaran-20-ols or even dammar-13(17)-enes. Although laboratory experiments aiming at the chemically induced rearrangement of the C-20 cation of the dammarane-type into baccharane-type skeleton have failed (TORI et al., 1977), clay-catalyzed reactions may still lead to such a rearrangement since it has been shown (SIESKIND et al., 1979) that diaster13( 17)-enes could undergo D-homoannulation when treated with montmorillonite K10 clay. In this respect, the baccharane backbone could be viewed as a result of the diagenetic transformation of dammarenes. Several observations, however, will argue against this hypothesis. At first, to our knowledge, ring D homologated diasteranes have not yet been conclusively identified in sediments, showing that ring D enlargement is a process which does not currently take place under natural conditions. Furthermore, the occurrence of baccharane seems strictly restricted to the Adriatic Basin and therefore cannot be considered as a usual transformation product of the relatively ubiquitous sedimentary dammarenes. Moreover, the often pure carbonate depositional environment of our samples would not appear especially propitious to promote such a transposition (usually only very low quantities of diasteranes are present), and no further rearranged backbones (e.g., shionane) could be detected. Lastly, no correlation seems to exist between the occurrence in sediments of d a m m a r e n e s / d a m m a r a n e s and that of baccharane. F r o m all these considerations, it appears that the presence of baccharane in the Adriatic Basin would most likely reflect a specific source rather than diagenetic transformations of structurally related compounds. Whether this source is of marine or continental nature remains, of course, to be established. CONCLUSIONS 1) The structures of two tetracyclic hydrocarbons widespread in Italian Triassic sediments and petroleum of the Adriatic basin, baccharane 1 and 17/3H-des-E-lupane 2, have been conclusively identified by comparison with synthetic references. This constitutes the first report ofbaccharane derivatives from a geological source.
2) From their restricted sedimentary occurrence and the nature of the depositional environment, these molecular fossils most likely derive from biological precursors bearing the baccharane skeleton rather than being diagenetic transformation products of biosynthetically closely related products such as dammarenes. A contribution of either marine or continental organisms can be envisaged. 3) Due to its restricted distribution in geological samples baccharane could turn out to be a highly specific biomarker in terms of source and correlation parameter. Acknowledgments--We thank S. Marc, M. Edmond, and M. C. Schweigert for their much appreciated technical assistance, and AGIP S.p.A. for financial support and permission to publish. We also thank R. P. Philp, J. Rullkrtter, R. Summons, and P. Sundararaman for their helpful comments in reviewing the manuscript. Editorial handling. G. Faure
REFERENCES AGETA H. and ARA1Y. (1983) Fern constituents: Pentacyclic triterpenoids isolated from Polypodium niponicum and P. formosanum. Phytochem. 22, 180 l- 1808. AGETA H., ARAI Y., and MASUDAK. (1992) Chemical and chemotaxonomical studies on fern triterpenoids in Polypodiaceae. Abstracts of the 18th IUPAC Symposium on the Chemistry of Natural Products, Strasbourg (France), 30.8-4.9.1992, p. 591. ALLISONJ. M., LAWRIEW., MCLEANJ., and BEATONJ. M. (1961) Dehydrogenation with mercuric acetate in the lupane series: Part II. Lupeol and its derivatives. J. Chem. Soc., 5224-5230. ANTHONSEN T., BRUUN T., HEMMER E., HOLME D., LAMVIK A., SUNDE E., and S~RENSEN N. A. (1970) Baccharis oxide, a new triterpenoid from Baccharis halim~/blia L. Acta Chem. Scand. 24, 2479-2488. AQUINO NETO F. R., TRENDELJ. M., RESTLEA., CONNANJ., and ALBRECHTP. A. (1983) Occurrence and formation oftricyclic and tetracyclic terpanes in sediments and petroleum. In Advances in Organic Geochemistry 1981 (ed. M. BJOP~OYet al.), pp. 659-667. J. Wiley & Sons. AQUINO NETO F. R., TRENDEL J. M., and ALBRECHT P. (1986) Synthetic intermediates derived from triterpenoids by the retroMichael reaction in the vapour phase. Tetrahedron 42, 5621-5626. EKWEOZOR C. M., OKOGUN J. I., EKONG D. E. U., and MAXWELL J. R. (1979a) Preliminary organic geochemical studies of samples from the Niger delta (Nigeria): I. Analyses of crude oils for triterpanes. Chem. Geol. 27, 11-28. EKWEOZORC. M., OKOGUNJ. I., EKONG D. E. U., and MAXWELL J. R. (! 979b) Preliminary organic geochemical studies of samples from the Niger delta (Nigeria): If. Analyses of shale for triterpenoid derivatives. Chem. Geol. 27, 29-37. FUJIOKA T., IWAMOTO M., IWASE Y., OKABE H., MIHASHI K., and YAMAUCHIT. (1988) Studies on the constituents of Actinostemma lobatum Maxim: Ill. Structures of Actinostemmosides E and F, new baccharane-type triterpene glycosides isolated from the herb. Chem. Pharm. Bull. 36, 2772-2777. HARVEY H. R. and MCMANUS G. B. (1991) Marine ciliates as a widespread source of tetrahymanol and hopan-3/3-ol in sediments. Geochim. Cosmochim. Acta 55, 3387-3390. HILLS I. R., WHITEHEADE. V., ANDERSD. E., CUMMINSJ. J., and ROBINSON W. E. (1966) An optically active triterpane, gammacerane in Green River, Colorado, oil shale bitumen. J. Chem. Soe., Chem. Commun., 752-754. MAHATO S. B., NANDY A. K., and RoY G. (1992) Tfiterpenoids. Phytochem. 31, 2199-2249. MASUDAK., SHIOJIMAK., and AGETAH. (1983) Fern constituents: Six tetracyclic triterpenoid hydrocarbons having different carbon skeletons, isolated from Lemmaphyllum microphyllum var. obovatum. Chem. Pharm. Bull. 31, 2530-2533. MATTAVELLIL. and NOVELLI L. (1990) Geochemistry and habitat of the oils in Italy. AAPG Bull. 69, 1623-1639.
Baccharane in petroleum from the Adriatic Basin MEUNIER-CHRISTMANN C., ALBRECHT P., BRASSELL S. C., TEN HAVEN H. L., VAN DER LINDEN B., RULLKOTTERJ., and TRENDEL J. M. ( 1991 ) Occurrence of dammar- 13(17)-enes in sediments: Indications for a yet unrecognized microbial constituent? Geochim. Cosmochim. Acta 55, 3475-3483. PETERS K. E. and MOLDOWANJ. M. (1993) The Biomarker Guide. Prentice Hall. RlVA A., CACCIALANZAP. G., and QUAGLIAROLIF. (1987) Recognition of 18/3(H) oleanane in several crudes and Tertiary-Upper Cretaceous sediments. Definition of a new maturity parameter. Org. Geochem. 13, 671-675. RIVA A., RIOLO J., MYCKE B., OCAMPO R., CALLOT H. J., ALBRECHT P., and NALI R. (1989) Molecular parameters in Italian carbonate oils: Reconstruction of past depositional environments. 14th International Meeting on Organic Geochemistry, Paris, 18 22.9.1989, Abstr. 335. RUZICKA L. (1959) History of the isoprene rule. Proc. Chem. Soc., 341-360. SHOJ1 N., UMEYAMAA., TAIRA Z., TAKEMOTO T., NOMOTO K., MIZUKAWA K., and OHIZUMI Y. (1983) Chemical structure of hosenkol-A, the first example of the natural baccharane triterpenoid of the missing intermediate to shionane and lupane. J. Chem. Soc., Chem. Commun.. 871-873. SIESK1ND O., JOLY G., and ALBRECHT P. (1979) Simulation of the
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