Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Arau- Discussions ` es-Nancy cedex, Biogeosciences , with the exception 10514 10513 genera are observed. Both are characterized by some high shows some similarities to that of Araucaria and genus shows a high relative abundance of bicyclic sesquiterpenoids, particularly This discussion paper is/has been under review for the journal Biogeosciences (BG). Please refer to the corresponding final paper in BG if available. ing trace levels inpenoids the of case ofof saturated one abietanes. species, Distribution in(essentially of which derived the sesqui- from tetracyclic and abietanoic compounds acids) diter- Wollemia are predominant. absent High and similarities the betweenrelative abietane-type the abundance of tetracyclicditerpenoids. compounds with no predominance of other specific compounds of the cadalane-typetype, compounds bisabolane-type accompanied by aspenoids those well are of of as eudesmane- the labdane-type, chamazulene,abietanoic isopimarane, acids) pentamethyl-dihydroindenes. abietane-type (essentially as Diter- derivedtetracyclic well from diterpenoids, as these isohexyl compounds alkylaromatic show a hydrocarbons. relatively Compared lower to abundance, reach- the (common, inter- and infra-genericbotanical characteristics) allow palaeochemotaxonomy. Such to complete knowledgepalaeoflora our is knowledge relevant and in to palaeoclimatic the reconstruction,ies. reconstitution archaeology In of and this environmental work,organic solvent stud- major extracts of carbon fresh skeleton andspectrometry. pyrolyzed types The using of results gas show chromatography-mass terized are that by detected a all predominance in speciescaria of the of saturated Araucariaceae tetracyclic are diterpenoids. firstly Moreover, the charac- Several extant species of the Araucariaceaeinvested family (one for of the the families experimental of artificialalize ) were the maturation transformation by of confined biomolecules pyrolysis,experimental to in study geomolecules of order in diagenetized to laboratory molecular re- conditions. signatures The of the Araucariaceae species Abstract Biogeosciences Discuss., 9, 10513–10550, 2012 www.biogeosciences-discuss.net/9/10513/2012/ doi:10.5194/bgd-9-10513-2012 © Author(s) 2012. CC Attribution 3.0 License. France Received: 11 July 2012 – Accepted: 25Correspondence July to: 2012 Y. – Lu Published: ([email protected], [email protected]) 8 August 2012 Published by Copernicus Publications on behalf of the European Geosciences Union. Determination of the molecular signature of fossil conifers by experimental palaeochemotaxonomy – Part 1: The Araucariaceae family Y. Lu, Y. Hautevelle, and R. Michels University of Lorraine, UMR7566, CNRS G2R, BP 239, 54506 Vandœuvre-l 5 25 15 20 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | cult to perform ffi culties are related to (1) avail- ffi erences within the family, (3) to highlight ff 10516 10515 In order to fill these gaps, within the 7 extant families (Fig. 1), we investi- Most of palaeochemotaxonomic investigations are derived from published chemo- In addition to be useful to palaeofloristic and palaeoclimatic studies, botanical Botanical palaeochemotaxonomy has some specific attributes compared to palaeob- palaeochemotaxonomic (diagenetized) signature of(2) all to extant evaluate Araucariaceae the species, inter-the and molecular characteristics infra-generic which di should allowilies their distinction in from ancient other sediment conifer fam- samples.investigations on This palaeoflora, contribution palaeoclimatic could reconstruction, serve archaeology,tal environmen- research. as database to future gated the palaeochemotaxonomy of severalily extant (Table species 1) of using the anpyrolysis Araucariaceae (Hautevelle experimental fam- et method al., based 2006b).of on biomolecules This artificial into procedure maturation their allows by2000; corresponding to confined Gupta diagenetic simulate et geomolecules the al., (Stankiewicz conversion et 2006, al., 2007). Aims of this study are (1) to determine the common very scare. As pointed byable Hautevelle chemotaxonomic et data al. (generally (2006b),substances, focus di like on resin or specific essential biomolecules oil);may or (2) significantly degradation on and modify particular diagenesis the reactions,a which initial direct molecular chemotaxonomic fingerprint, relationshippart; making between (3) it an the di extant scarcity plantplants of containing and reference organic its collections molecules. fossil of counter- well preserved and identifiable fossils products in craft and funeral rites (e.g. Colombini et al.,taxonomic 2005; data Romanus et (Otto al.,plants and 2008). found Wilde, in 2001) sedimentaryet and rocks al., to (Otto 2011). some and Unfortunately, our Simoneit, part knowledge 2001; on from Otto botanical the palaeochemotaxonomy et is studies al., still of 2005; Dutta fossil palaeochemotaxonomy is used to trace diet habits, to understand the use of natural Mesozoic palynomorphs, they canbiomarkers be could directly linked be to readilytotal specific analyzed rock taxa samples. by of usual plants organic and geochemistry (3) procedurespalaeochemotaxonomy on has alsoronmental been science, recently it appliedrecent proved in anthropic to other impacts be toet instances. helpful soil In in al., (e.g, the envi- 2011; Farellapaper appreciation et Lavrieux mills of al., et past (e.g. 2001; land-use al., Leeming Heim and et and 2011) al., Nichols, as 2010; 1998; well Huang Wang as at to al., trace 2007). river In pollution archaeology, caused by otany and palynology in the reconstructionthrough of geological palaeoflora and times palaeoclimatic (e.g. evolutions et Vliex et al., al., 2006a). 1994; Indeed,record van (1) than Aarssen well et biomarkers al., preserved are 2000; plant more Hautevelle macrofossils, widespread (2) in on the the stratigraphic contrary to Palaeozoic and taxa and can be used astheir specific atomic markers. constituents While most recycled bioterpenoidsis in are incorporated the degraded into and surface sediments, processesthe thus of bioterpenoids joining the are the earth, transformed geological aproducts by cycle. minor called diagenesis, During geoterpenoids. part leading this Their to process, totally initial the retained chemotaxonomic formation (e.g. value of Simoneit, can reaction cient 1986; be sediments Otto partially may thus and or yield Simoneit, palaeochemotaxonomicbiological 2001). information precursors. inherited Geoterpenoids from of their an- Numerous studies of the molecular compositionout of the extant chemotaxonomic terrestrial values plants of have pointed manyular biological (e.g. compounds Aplin bioterpenoids et inThis partic- al., means 1963; that Smith, 1976; these Castro biomolecules et are al., synthesized 1996; by Mongrand a et restricted al., number 2001). of plant 1 Introduction 5 5 25 15 20 10 25 15 20 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Agathis , 2001). species ff sections, was Agathis Araucaria, Bunya, Bunya fossils appeared, but (Jussieu, 1978) repre- and the , 2001; Kunzmann, 2007b). Its Wollemia ff Araucaria and is by far the most endemic genus and Eutacta ord, 2005). These two genera, extend- remain somewhat unclear; ff Agathis (Jones et al., 1995) represented by the solo 10518 10517 Wollemia gene sequences: Agathis ective ff rbcL , especially the Wollemia . This sub-classification is in agreement with their mor- . . The latter, previously thought to be extinct, was rediscovered Araucaria Agathis Intermedia and could be infragenerically classified into four sections: is the most basal extant taxon in the family, sharing morphologies with , and species develop in the subtropical regions and extend into the marginal trop- , 2001). The monotypic genus Wollemia nobilis ff Wollemia Araucaria Eutacta phological characteristics (Stockey, 1982); phylogenetic relationships within the three genera areancestor); all monophyletic (i.e. all speciesAraucaria descend from an unique – – – – During the Jurassic, These phylogenitic relationships of Araucariaceae species were also confirmed by During the Cretaceous, theremained first restricted e to Australia and New Zealand (Hill and Bigwood, 1987; Stockey, maximum worldwide distribution wastaceous. achieved Their in distribution bothSouthern was hemispheres Hemisphere, then during with reduced thethe an Cre- Southeast during extension Asia). the to Middle the Northern Palaeocene to Hemispherethe (restricted the first genus to to appear andrange diversify. Their macrofossils of and pollens habitats broadened in aclimate wide the condition was two a hemispheres subtropical (Gondwana to warm-temperate and (Kershaw Laurasia), and where Wagsta Jurassic Appearance and evolution ofpollen Araucariaceae in are sedimentary series recordedEarly all by Traissic around both of the macrofossils Australiathe world. and unequivocal (as Araucariaceae The ancestor) representatives first were just foundMiddle pollen in Jurassic after record North (Stockey, the 1982; Yorkshire is from Kershaw from the Permo-Triassic and extinction the Wagsta and occupy the tropical islandsing (Dettmann and within Cli the equatorialgrow region under of a New mesothermalWagsta Guinea climate and limited Southeast to Asiaits the at few lower lower wild montane latitude, species zoneWollemi (about (Kershaw National 40 Park and adult in New plants) South live Wales deep (Australia). in2.1.2 a wilderness rainforest Fossil of Araucariaceae the record The geographic distribution of majorAraucaria Araucariaceae species is givenical in Table regions 1. Extant where there is a lower climatic variability, while most of in 1994 in Australia, cloistered deep in the New SouthSetoguchi Wales et (Jones al. et (1998) al., based 1995). on extending from subtropics to tropicsspecies (Enright are and described Hill, and 1995).(Salisbury, Nowadays, consist more 1807), of than three 40 represented well-definedsented by genera by which 21 19 are species, (1) speciesspecies (2) and (3) of several species during their evolution (Axsmith et al.,2.1 2004; Kunzmann, 2007a). , geographical distribution and living environment 2.1.1 Extant Araucariaceae Today, Araucariaceae species representsphere a forests. dominant The major component nativeica, of Araucariaceae Southwest the species Asia are South restricted and Hemi- to the South Western Amer- Pacific region and cover a large rainfall region Araucariaceae is considered as a Southernthe Hemisphere conifer first family, and conifer was among Triassic families (Stockey, 1994). to Diversity and be abundance ofup individualised Araucariaceae to are from nowadays well despite their preserved of some “Voltziales” reduction ancestors of geographic during distribution and the extinction 2 Some generalities concerning the Araucariaceae family 5 5 25 15 20 10 25 20 15 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | , ) 15 Agathis ¨ onig, 2000; but provided wood Agathis dammara became more promi- genus. At the end of the ), sesquiterpenoids (C Agathis 10 Agathis to be more competitive (Kershaw has been intensively used in the Wollemia Agathis , 2001). ff and especially 10520 10519 (kauri copal, kauri gum or kauri resin) was also Agathis australis ´ Agathis e, 1995). In parallel, there was a reduction of Wollemia r ff living in Indonesia and Philippines, as genus began to leave the Northern Hemisphere. As pointed species began to colonize and diversify in New Caledonia and is highly considered as a source of attractive, straight-grained, ) are abundantly synthesized by Araucariaceae. Compounds, , 2001). ff ) are here summarised. 10 Agathis 20 Araucaria Agathis Agathis , 2001), and it seems also be that for ff species, they regenerated from Early Miocene under subtropical to warm- may not have an equivalent economical value as Sesquiterpenoids are composed by monocyclic (e.g. elemane-, germacrane- and Regarding diterpenoids, they are represented by bicyclic, tricyclic and tetracyclic Monoterpenoids (C Furthermore, many Araucariaceae produce resins having archaeological and eco- Later, during the Cenozoic, sclarenes and biformenes), alcoholscommunic (like sclareols) acids) as and well isomers as may acids occur (like (Caputo agathic and and Mangoni, 1974; Caputo et al., types) and tricyclicpounds. These (e.g. chemicals are aromadendrane-, mainlyhols represented cubebane- and by acids, unsaturated and constituents hydrocarbons,Brophy of alco- gurjunane-types) et essential al., oil com- 2000). and However,like resins they angiosperms (e.g. are and Pietsch common bryophytes and (Otto in K and conifers Wilde, as 2001). well ascompounds. in Bicyclic other diterpenoids plants mainlycommon composed by in labdane-type all compounds are conifers. They are synthesized as unsaturated hydrocarbons (like species (Brophy etessential al., oils 2000). (Brophy They etlower represent al., molecular the 2000; weight, major Staniek thesepreserved et components chemicals in al., of are sediments. 2009). generally araucarian Furthermore, highly owing volatile and to not their humulane-types), readily bicyclic (e.g. cadinane-, copaane-, muurolane- and caryophyllane- for shipbuilding during several centuries. 2.2 Chemotaxonomy Available data on theand diterpenoids composition (C of monoterpenoids (C like pinene, thujene, limonene and cubebene are widespread in most of Araucariaceae widely used for the manufactureXX of centuries. paints, Other varnishes andproduce linoleum resin during having the XIX anCopal and has important also economic beenAraucaria value used (Manilla for a resin long or time Manilla in copal). jewellery and related arts. Furthermore, interest nowadays. The resin of past for the constructionof of New Zealand. canoes, Some houses of these and objects cult were objects well preserved by and native show Maori archaeological people The worldwide patchyposits, and as sparse inferred by distribution palaeobotanicpalaeoclimatic of and information. palynological Araucariaceae Indeed, data, in they canditions, thus preferentially sedimentary mainly provide growunder in de- valuable warm equatorial, and tropical(Kershaw or and wet subtropical Wagsta con- rainforests as well as peat swamps nomical values. easily worked timber. The wood of from South-Eastern Australia due toAraucaria the cooler conditions attemperate high conditions latitudes. (Kershaw Regarding and Wagsta 2.1.3 Araucariaceae interests major events in EarthNorth history: America; (1) (2) major rapid changes environmental changes in across forest the composition K-T innent boundary. from Europe the and Early Eocene towith the Early the Oligocene. diversification This development of wasfavourable synchronous angiosperms. conditions, such Owing as to themate their in ultramafic adaptation islands, soils ability on to peridotitesother many and Asia-Pacific un- the islands unstable (Ja cli- ofloristic exchanges between Gondwanagiosperms and during Laurasia. the Cretaceous Moreover, also appearance forced and of Wagsta an- Cretaceous, the out by Kunzmann (2007b), their retreat from Northern Hemisphere coincides with two 1994). This palaeogeographic limitation could have been due to the broken-up of palae- 5 5 25 15 20 10 25 15 20 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | A. and 2 Cl 2 97%, powder ≥ ) on the silica column. The v/v C and 700 bar during 24 h. These ◦ (Caputo and Mangoni, 1974) and (50/50 2 (Fluka No 62420, purity Cl ¨ onig, 2000; Otto and Wilde, 2001). Other 2 4 with an Accelerated Solvent Extractor (ASE 10522 10521 A. bidwillii species). 2 (Table 1). This sampling is believed to be well OH/CH Cl 3 2 species while these compounds are not reported by Agathis species: Wollemia ) on the alumina column, then with pentane, followed by ) and CH favors the generation of saturated over aromatic terpanes Agathis 4 v/v v/v and 1 C before being crushed. The plants to be pyrolyzed were intro- Araucaria ◦ (65/35 (50/50 2 2 Cl Cl 2 2 Araucaria C and 100 bar using CH (Otto and Wilde, 2001). ◦ , 8 OH/CH 3 The soluble organic compounds of fresh plants and their pyrolysates were extracted Distribution of bioterpenoids varies from species to another in fresh plants (Thomas, Tricyclic diterpenoids reported in Araucariaceae are composed essentially by the ¨ onig (2000), both tri- and tetracyclic diterpenoids are synthesized as unsaturated hy- CH pentane/CH ers (aliphatic, aromatic and polar).each Biomolecules representative of were the also studied. starting fresh plant samples of under 80 350, Dionex). The totallar extract fractions was using then liquid fractionatedpolar chromatography into fraction) on aliphatic, and alumina aromatic (separation silicadrocarbon and of fraction) (separation po- hydrocarbon columns. of The and aliphatic fractions and were successively aromatic eluted fractions with from CH the hy- duced into gold tubes withform). and The without LiAlH use ofduring LiAlH pyrolysis. The filedand gold introduced tubes into were autoclavesparameters and then of heated pyrolysis sealed were at determined under 280 to an obtain the argon broadest atmosphere distribution of biomark- representative of the intrinsic variability of the Araucariaceae family. 3.2 Experimental and analytical procedures This study focuses on the diagenetizedmation molecular of signature fresh of material Araucariaceae. Transfor- to itsartificial diagenetized maturation counterpart by was confined carried pyrolysis out developedand by by experimental Hautevelle et of al. the (2006b).a Twigs selected desiccator plants were at finely 45 cut and dried under vacuum for 24 h in 3.1 Samples 12 Araucariaceae speciesAgathis were selected for this study and are represented by 3 3 Samples and experimental procedures Brophy et al. (2000).former Such studied kind the resins of whileonomic contradictions the data is provides last valuable certainly studied knowledge essential due to oil. palaeochemotaxonomy. to Nevertheless, the chemotax- fact that the one or eventually tworetically compounds. specific Their to each moleculara araucarian composition compilation species could of and/or thus genus. severalThomas be These studies (1969) data theo- and insists originate some on from maric the inconsistencies and presence abietic may acids of occur. in compounds For like instance, agathic, communic, isopi- kaurane-, phyllocladane-, trachylobane- andand atisane-types Wilde, (Brophy 2001). et AccordingK al., to 2000; Thomas Otto (1969), Brophydrocarbons et (like al. pimaradienes, (2000) abietadienes,isopimaric and kaurenes, acids) Pietsh and etc.), and more acidswhich rarely (like are as only alcohols abietic reported (like and in phyllocladanol, few phenolic abietanes 1969; Brophy et al., 2000). For instance, each species seems to be dominated by bicyclic compounds, belonging to thebe clerodane-type, restricted like to clerodadienic two acidshunsteinii seem to abietane- and pimarane-types. Tetracyclic diterpenoids belong to the beyerane-, 1974a,b, 1976; Thomas, 1969, Pietsh and K 5 5 20 25 15 10 15 20 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | 1 1 − − ; Cmin ◦ -15 ion at + , completely genus 18 ) and dihydro- H 22 sesquiterpenoids 15 A. bidwillii H 15 and species are also com- C, and then 3 ◦ ) detected in the fresh Araucaria Araucaria cis- 109 (corresponding to a frag- 165 (corresponding to the loss to 130 ) are identified like cadinanes, 1 and Araucaria − A. bernieri 28 for aliphatic fraction and 8 mgml m/z m/z -curcumene (C H 1 − 15 ar Cmin trans- ), ◦ is showed in Fig. 2. Di- and tri- methyl- 30 H species. Bisabolane-type compounds, like C for 15 min. Helium was the carrier gas ◦ 0.25mm i.d., with 0.1 µm film thickness). The 15 10524 10523 × C at 15 ◦ Araucaria Araucaria 208, a low abundance (even absent) of the M ), are observed in the aliphatic and the aromatic frac- , both partially aromatized), cadalene (C -43) and a base peak at 24 22 + -curcumene is often found in trace amounts for all species. H ) and bisabolane-type compounds. Acyclic farnesane, prob- H m/z 15 Ar 32 15 H 15 flow rate). The MS operated in the electron impact mode (EI) at 70 eV ion- 193, a relatively high abundance of ion at 1 − -curcumene (C compounds isomers of saturated bisabolanear (C tion respectively. Bisabolane-type compounds are generally derivedfresh from plants. bisabolenes They represent found one in of(especially the the in major the class aliphatic of fractions), except for cadalane-type compounds. They are mainlywhich derived from are cadinol ubiquitous andof cadinenes saturated in cadalane-type vascular compounds plants. (C In the aliphatic fractions, a variety mentation of A/B-ring moietyaromatic fraction, after the the cadalane-type compounds lossand are of calamene represented (C precedent by calamenene units)aromatized). (Fig. One norcadalene 3a). (probably the In 1-isopropyl-7-methylnaphthalene ac- cording the to the spectrum from SinghIts et formation al., 1994) is is the detected result in all of species the (Fig. demethylation 3b). of cadalane-type compounds, and Farnesane (C ably derived fromplants, isomers is of identified farnesol in ( all muurolanes and amorphanes.a molecular Their ion spectra at m/z are commonlyof characterized a isopropyl by unit, M – – monly characterized by: 4.1.1 Sesquiterpenoids Distribution of sesquiterpenoids of naphthalenes are ubiquitous in each representative. 4.1 Molecular characteristics of artificially diagenetized Aliphatic, aromatic and polar fractionstative, were including studied the for extracts eachmainly of Araucariaceae focuses represen- fresh on plants palaeochemotaxonomy, the anderentially extracts studied. of of their pyrolyzed pyrolysates. plants Since were pref- our study detector. Data were acquired andCompounds processed were using identified the by Agilentdatabase comparison ChemStation (Wiley275) of or software. by mass interpretation spectra of mass with spectrometric literature fragmentation and patterns. library 4 Results and discussion Aliphatic and aromatic fractionsmatograph were coupled analysed with an using HP an 5971silylated mass HP spectrometer using 5890 (GC-MS). N,O-bis-(trimethylsilyl)trifluoroacetamide Polar Series fractions (BSTFA) were II beforecapillary gas analysis. column was The chro- a DB-5temperature J&W (60m programme was 70–315 followed by an(1 isothermal mlmin stage atization energy 315 and mass spectra were scanned from 50 to 500 Da using a quadrupole obtained fractions were diluted in hexanefor (4 the mgml others) before analysis by GC-MS. 3.3 Gas Chromatography-Mass Spectrometry (GC-MS) and identification of 5 5 10 15 20 25 20 15 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | ). 38 173 159 H 20 species . m/z m/z species. It A. cunning- and Araucaria ) with a molecular 169 (Fig. 3e). Due 24 and at trace level in Araucaria (H)-eudesmane are H β A. angustifolia 15 m/z A. araucana species are characterized by ) and a base peak at species, although they do not 20 A. bernieri ) is detected in the aromatic frac- H . According to its mass spectrum, 174 and a base peak at 28 14 . Moreover, eudesmanes-type com- H 18 m/z Araucaria Araucaria 133 was detected in all 188 (C 244 (C (H)-eudesmane and 4 A. nemorosa m/z 10526 10525 m/z α and m/z A. cunninghamii -carotene like in sporopollenin (which derives from some sesquiterpenoids, except for ) and other compounds identified as pentametyl-2,3- β 16 H 14 159 matches very well to fragment composed of two rings (in- A. laubenfelsii as well as in ), with a molecular ion at species. It shows a significant abundance in most species, but in , 18 184) more intense than its base peak at Araucaria m/z H 204 and a base peak at 13 m/z m/z ; Araucaria A. bidwillii -15) (Fig. 3f). Pentametyl-2,3-dihydroindene and chamazulene show a higher + furthermore, 1,3,4-trimetyl-2-(4-methylpentyl)benzene (C ion at but with a low abundance. According to Ellis et al. (1996), it belongs to the fam- (M abundance than other aromaticcadalene; bicyclic sesquiterpenoids with the exception of ily of isohexyl alkylaromatic(Fig. hydrocarbons 4a). (see Its Sect.pounds occurrence 4.1.2) (like could drimane) with during opened be the A-ring due pyrolysis. to the alteration of some bicyclic com- eudesmane-type compounds.identified 4 according to Wiley275 database and the spectrum published by Alexan- pounds are widely distributedHowever, they in were fossil not vascular reportedand plants in Otto and the (Alexander Wilde, previous et (2001), studies al., which of focus 1983). Brophy onionene fresh et (C material; al., (2000) according to Achari et al.could (1973). be It derived was from detected forpollen most exines) of (Achari etprobably directly al., derived 1973; from ionone Simoneit, that 1986). is abundant Inchamazulene in our the (C case, fresh plants; the ionene is to its carbonand skeleton a composed of pentenicskeleton, a ring, like ring aromadendrenes, it alloaromadendrene composedwidespread is in and of the spathulenol, fresh probably seven species which (Brophy derived2,3-dihydroindene, carbon et are identified al., according atoms from 2000; to Olawore, Wiley275 2005). precursor database, Pentametyl- a is spectrum of characterized with by a similar molecular ion at carbon no more precision to itspounds structure show was a proposed. high Inone abundance addition, of in cadalane-type major com- most ofhamii species samples and represent also der et al. (1983)vary (Fig. 3c, from d). one Their specie relativeA. to abundances heterophylla of another. these It two is compounds higher in dihydroindenes according tofraction. their Chamazulene spectra elutes are afterWiley275 also cadalene database, remarkable and is in itslar spectrum, the quite according ion aromatic similar to ( to the that of norcadalene but with a molecu- – – – – A peak having a molecular ion at cluding an aromatic ring) and twothe methylene loss units. This of fragment a is branched-chain probably formed with via 6 carbon atoms (Fig. 6a). This kind of fragmentation show a high abundancegenerally in derived each from representative.in labdadiene, As phenolic the shown fresh labdanes inand plants and the Wilde, (Caputo communic literature, 2001). Moreover, they and acids according are agathic Mangoni, to present acid de 1974; could Paiva Campello also Caputo and be et Fonseca the (1975), origin al., the of 1974a,b, labdanes 1976;tion in the in Otto case all of traces for its base peak at Bicyclic diterpenoids Bicyclic diterpenoids are essentially composedSeveral by isomers labdane-type of compounds labdane (C are identified in 4.1.2 Diterpenoids Diterpenoids in the extractsa of diversity pyrolysates of of bicyclic, tricyclic and tetracyclic compounds (Fig. 5). 5 5 25 20 15 10 15 10 25 20 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | , . These . Its mass . They are A. heterophylla , A. bidwillii Araucaria ) and a base peak at (H)-phyllocladanes and A. launbenfelsii 274 and a base peak at 38 β H A. bernieri 20 m/z 193 matches to a configuration (Caputo and Mangoni, 1974; Cox ), is detected in the aliphatic frac- 278 (C 36 (H)- and 16 m/z H α 20 m/z A. bidwillii 10528 10527 -isopimadinol, that are detected in some of the β -clerodadienes (Otto and Wilde, 2001) and is re- -beyerane, 16 ent . This kind of tricyclic aliphatic diterpenoids is proba- ent species (Fig. 6c). Compared to Alexander et al. (1995), (H)-kauranes, show relative high abundance in each repre- species including β -16 Araucaria A. nemorosa ent ). Distribution and abundance of these compounds for each represen- 34 and Araucaria H 20 -hydroxyisopimarene and 3 β (H)- and erent by the fragment intensities. They may rather be consistent with a trimethyl α 109) contains only two methylene units. The “third” methylene unit could thus ff 109 is detected with a low intensity in the aliphatic fraction of 123 (C -16 These abietane-type compounds are the major diterpenoids of A peak having a similar spectrum to those of methylretenes is observed in the aro- Other tricyclic diterpenoids, like abietane-type compounds, show a higher diversity, 2,6-dimetyl-1-(4-methylpentyl)naphthalene and 6-ethyl-2-dimetyl-1-(4-methylpentyl)- Another peak heaving a molecular ion at m/z m/z tative are given in Fig. 5a and Table 2. They are probably derived mainly from beyerene, are di phenanthrene. Tetracyclic diterpenoids Tetracyclic diterpenoids, mainly ent sentative. They are identified according toTheir the spectra spectra are published all by Noble characterized et by al. a (1985). molecular ion at Araucariaceae as well, showmostly degraded a during low pyrolysis. relative abundance in thematic fresh fractions plants of all andthe were spectra of these compounds have a same molecular ion and mass fragments, but bietane, 18- andsimonellite, 19-norabieta-8,11,13-trienes, 2- and tetrahydroretene, 9-methylretenes. While simonellite, theabietanes saturated bisnor- and abietane-type fichetellite compounds are like detected at trace level. mostly derived from dehydroabieticrolysates acid and more that generally isphenolic from observed abietanes abietanoic including in acid sugiol, hinokiol the inand and the polar ferruginol Simoneit, fresh (Otto fraction and plants. 2001), of Wilde, As 2001; which py- regards Otto are the known as abietane-type compounds precursors for bly derived from isopimara-7,15-dienes, 13-isopimardiene,well isopimara-8(9),15-diene as as 8 fresh plants. especially in the aromatic fraction. Related aromatic compounds are retene, dehydroa- A. laubenfelsii Pimarane-type compound, like isopimarane (C tion with a significant abundance for numerous species, like cal reactions, like aromatisation,the maturation ring-opening (Ellis processes et and al.,methylpentyl)naphthalene 1996). rearrangement, and Furthermore, during the 6-ethyl-2-dimetyl-1-(4-methylpentyl)-naphthalene co-occurrencegests sug- of precursors 2,6-dimetyl-1-(4- like tricyclicproducts diterpenoids are (Ellis structurally et quite al., dissimilar 1996), to for their which precursors. theTricyclic diterpenoids diagenetic Few aliphatic but a high diversity of aromatic tricyclic diterpenoids are observed. naphthalene are abundant fortwo most compounds species are identified andin from in Fig. spectra 4b, traces c. published in They bythe belong Ellis 1,3,4-trimetyl-2-(4-methylpentyl)benzene to et the previously al. family described.cific of (1996) Indeed, isohexyl and carbon alkylaromatic their given hydrocarbons skeleton spe- just with like an opened A-ring could be due to some specific chemi- including two rings (A( and B) andonly three be methylene bound at units, the whiletent carbon-9 with its position a (Fig. A/B-ring 6b). structure fragment Thisis similar obtained limited to configuration that to is of consis- astricted clerodane. few to However, derivatives two its of reportet in al., extant 2007). conifers described in the literature until now. m/z spectrum indicates that itbon skeleton could to also labdane. be Indeed, a its bicyclic fragment at compound and has a similar car- seems to be consistent with a monoaromatic labdane. No aromatic labdane has been 5 5 25 15 20 10 25 20 10 15 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | A. 238 ) with and 32 . Yet, no m/z (Fig. 6e). H erence of 19 ff 231 is rela- ) and a base 32 genus pyrolysates and m/z H in both their fresh 260 (C species, except of A. bernieri 21 , 168 and 184 respec- A. nemorosa m/z A. laubenfelsii Agathis and 288 (C m/z Araucaria and Araucaria species are observed. Far- . This spectrum matches very m/z A. angustifolia A. cunninghamii respectively. representative. 1,3,4-trimetyl-2-(4- ) species is given Fig. 8a. Distribu- A. araucana and A. bernieri Araucaria Ag. ( , 259; (2) Agathis -beyerane). Co-occurrence of phyllocladane- A. angustifolia 10530 10529 m/z ent A. nemorosa Agathis A. bidwillii , ; Fig. 4e), a base peak at 26 . Only phyllocladane-type or kaurane-type compounds A. angustifolia and in H 19 , as only phyllocladanes are observed, the C-19 tetracyclic A. araucana 123 is detected in - group. This configuration is in correspondence to a C-21 tetra- A. nemorosa 2 m/z A. angustifolia and genus among all conifers. Unfortunately, the diagenetic products of these 123 is detected in A. angustifolia all show the presence of phyllocladanes. This C-21 tetracyclic diterpane is m/z ; Fig. 4d) and 254 (C 22 H -kaurene, isokaurene and phyllocladene, as well as kauran-16-ol, phyllocladanol Araucaria 18 It is also interesting to point out that, according to Otto and Wilde (2001), tetracyclic A peak with a spectrum characterized by a molecular ion at Another peak, characterized by a molecular ion at methylpentyl)benzene is detected as well, but at trace level. 4.2.1 Sesquiterpenoids Composition of sesquiterpenoids in tion and relativesimilarities abundance of to major thenesane, sesquiterpenoids molecular cadalane-type are composition given compoundscadalene in of (saturated Table and 2. cadalanes, norcadalene),some Some calamene, high pentamethyl-dihydroindenes calamenene, relative and abundances chamazulene in show each However, these unidentified compoundsnomic may values have and some special relevant attentionies. palaeochemotaxo- should be paid to these biomarkers in4.2 future stud- Molecular characteristics of artificially diagenetized biomarkers. Nevertheless, many of themstance, are it quite was abundant observed in that several two(C species. peaks For having in- spectra with atively molecular co-appear ion with at 2,6-dimetyl-1-(4-methylpentyl)naphthalene and 6-ethyl-2-dimetyl- 1-(4-methylpentyl)-naphthalene. These two unidentifiedsimilar peaks abundance show to also 2,6-dimetyl-1-(4-methylpentyl)naphthalene and respectively 6-ethyl-2-dimetyl- 1-(4-methylpentyl)-naphthalene. The relationship of thetheir compounds co-occurrence is and not similar clear; fragmentation while tend each couple to a similar structure. Numerous compounds remain unidentified insome of the them, extract their of masstifications spectra as are given proposed in in the Fig.They Figs. 7a have, 4d, to suggests, e our respectively and knowledge, saturated never 7a–g. been tricyclic Tentative reported iden- diterpenoid. in previous studies on Unidentified compounds possible corresponding pyrolysis products could be identified. cyclic diterpane. Moreover, (1) accordingtively to more the abundant than spectrum,laubenfelsii the the ions ion at at probably a 16(H)-homo-phyllocladane. diterpenoids like trachylobane-type and atiserane-type compoundsfor are those exclusive two compounds are stillatiserene/isoatiserene are unknown identified to in the our fresh knowledge. In our study, trachylobane and peak at It is likelymolecular a weight methylated of tetracyclic 14an Da diterpane, additional to because -CH the of well known the tetracyclic positive diterpenoids di corresponds to are observed in a base peak at well to that ofFormation 17-nortetracyclic of diterpane this described C-19cyclic by diterpane, diterpane while Noble its is et precisethe likely al. molecular case structure (1986) due remains of (Fig. to unclear. Furthermore, 6d). diterpane the in could demethylation rather of be a a 17-norphyllocladane. C-20 tetra- and kaur-16-en-19-ol, observed in thepounds are fresh detected plants. in Furthermore,plant beyerane-type com- (beyerene) and the pyrolysatestype ( and kaurane-type compounds isA. observed angustifolia in most of ent 5 5 20 25 15 10 25 20 10 15 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | . W. Ag. and m/z . Satu- species Ag. moorei -curcumene -curcumene), ar , but in higher genera among ar Agathis and Ag. australis (H)-phyllocladane, Ag. australis α . The methylretenes Wollemia nobilis Agathis species (Fig. 8b) show , and their spectra are ; 16 34 Ag. moorei H and . The isohexyl alkylaromatic 20 and Ag. australis Agathis Ag. australis 264 and 274. A base peak at and C Ag. australis Araucaria species could be interesting for future 24 species. Regarding to the unidentified . Distribution and abundance of major m/z than in H Ag. robusta , the detected abietane-type compounds 20 species. Concerning the eudesmanes, they Ag. australis and (Fig. 4b, c). Abietane-type compounds show 10532 10531 genus, are observed only in Agathis . is characterized by the presence of eudesmane- Agathis Araucaria Araucaria Agathis Ag. robusta and Araucaria genera, is here absent. And do likewise 1,3,4-trimetyl-2- Ag. robusta W. nobilis Ag. australis genus are given in Table 2. shows a wide diversity of diterpenoids. On one hand, as Ag. australis . As for (H)- kauranes, are detected in high abundance except for β (H)- eudesmanes), bisabolane-type (bisabolane, -beyerane is identified only in -16 Agathis β Agathis Ag. moorei ent ent -curcumene), cadalane-type (saturated cadalanes, norcadalene, W. nobilis and Ag. robusta ar shows several similarities to other Araucariaceae genera (Figs. 2, 6, 9). (H)- and 4 species. (H)- and . Despite of the presence and absence, these tetracyclic compounds represent α is not as abundant as those observed in α Araucaria -16 Furthermore, For the sesquiterpenoids, Furthermore, some peaks show relatively high abundance and their spectra are re- Bicyclic diterpenoids are characterized by the presence of labdanes in the aliphatic Furthermore, bisabolane-type compounds (bisabolanes and dihydro- abietane-type compounds8,11,13-trienes, (like tetrahydroretene, retene, 2- and dehydroabietane, 9- 18- methylretenes as and well as 19- dehydroabietic norabieta- nobilis the sesquiterpenoids. Chamazulene,both which presents a(4-methylpentyl)benzene. relatively high abundance in the other Araucariaceaetype compounds species, (like labdanes), it isohexyl alkylaromatic1-(4-methylpentyl)naphthalene is hydrocarbons and (like 6-ethyl-2-dimetyl-1-(4-methylpentyl)-naphthalene), 2,6-dimetyl- characterized by the presence of labdane- Distribution and abundance of major sesqui-are and di- given terpenoids in of Table 2. this monotypic genus type (4 and dyhydro- calamene, calamenene and cadalene) compounds. Pentametyl-dihydroindene in given in Fig. 7h,spectral i. database, their Since molecular these structuressignificant spectra are but are not also yet neither unique clear. referenced Howeverbiomarker presence their in research. in frequent, literature nor in our 4.3 Molecular characteristics of artificially diagenetizedW. of nobilis Agathis spectively characterized by a molecular109 ion and at 259 areTheir observed chemical in formulas the are aliphatic respectively fraction C of compounds, like those of Fig. 7a–e, are also detected with a significant abundance in robusta as the major components within the and norabieta-8,11,13-trienes are detectedrated only abietanes at are trace generallyabundance level in in in trace in are in majority derivedas from from dehydroabietic other acid abietanoic (detectedin acids in the precursors fresh the plants). and pyrolysates) Onlyditerpenoids, (cetono)phenoilic as dehydroabietic the abietanes well acid (detected is observedent in the pyrolysats. Tetracyclic hydrocarbons (2,6-dimetyl-1-(4-methylpentyl)naphthalene andmethylpentyl)-naphthalene) 6-ethyl-2-dimetyl-1-(4- show high relativeand abundance in are most found of asa traces high for abundance in alland species. 19-norabieta-8,11,13-trienes, Aromatic tetrahydroretene, abietanes, 2- like and retene, 9-more dehydroabietane, methylretenes, abundant 18- are in much As sesquiterpenoids, compositions ofalso diterpenoids several of similarities toditerpenoids of those the of fraction and monoaromatic labdaneessentially in originate from the agathic aromaticisopimarane acid fractions is (Thomas, (cf. observed 1969). Sect. in The 4.1.2), tricyclic which diterpenoids like which are widespread inshow the a low abundance in thewere other only detected only in 4.2.2 Diterpenoids 5 5 25 15 20 10 25 15 20 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | - ` es- ent (H)- kau- β Ag. autralis . Moreover, species, by -16 genus. Presence ent -beyerane and W. nobilis Araucaria ent (H)- and Araucaria α -16 ent section species. Eutecta 10534 10533 ´ -beyerane, emy (Botanical Garden of Montet, Vandoeuvre-l ent genera, like labdane and isopimarane, are at trace species. Two types of molecular signature could be . Moreover, within the two types of diterpenoid signatures, Agathis . The tetracyclic compounds, like Araucaria and , presents high similarities to the . For the second type, the isohexyl alkylaromatic hydrocarbons genus is mainly characterized by the sesquiterpenoids, like eu- We thank Alain Rouiller for technical assistance. We thank Fred- genus is characterized by sesquiterpenoids, like cadalane-type Ag. robusta genus, the bisabolanes and eudesmanes are occasionally present in is, on the other hand, demonstrated by a great variety of other unknown Araucaria Ag. moorei Agathis Araucaria Wollemia nobilis ´ evenard (UMR 5125 PEPS, University Claude Bernard, Lyon I), Anne Allaria (Botanical the aromatic abietanes show always a high relativeWollemia abundance. nobilis of cadalane-types, bisabolanes, eudesmanes, penatamethyl-dihydroindenesalso and the absenceof of chamazulene characterizekauranes, the are sesquiterpenoids predominant among signature thein diterpenoids. the Other recurrent diterpenoids amount or absent.aromatic The abietanes recurrent do abietane-type not seemgenera. compounds as Moreover, are abundant there as identified. those isand The observed a are in great the of two variety same other ers, of relative like abundance unidentified as the aromatic much aromatic diterpenoidshydrocarbons. as abietane-type the compounds other and well the known biomark- isohexyl alkylaromatic Araucaria only certain species.similar Concerning to the those of diterpenoids,otherwise most distinguished. of One is speciesa characterized, are high as abundance highly most of of bons, tetracyclic but diterpanes a low and abundance isohexyl ofand saturated alkylaromatic abietanes. hydrocar- This is theshow case for a relatively lower abundancesaturated and abietanes the represent tetracyclic thus diterpane athe case are significant for absent. fraction The of the aliphatics like it is the compounds, chamazulene and penatamethyl-dihydroindenes. Compared to the the desmanes, bisabolanes, chamazulene, penatamethyl-dihydroindenesparticularly and by more cadalane-type compounds.cyclic For compounds the are diterpenoids,penoids, saturated largely other tetra- diterpenoids, predominant. like Compared labdane,hexyl isopimarane, alkylaromatic to abietanes hydrocarbons, the show as a wellisopimarane tetracyclic relatively as is diter- lower iso- only abundance. detected Moreover, in the – – – W. nobilis Nancy) for the help in the providing of samples. Acknowledgements. eric Th Garden of the Villaturelle, Paris), Thuret, Romaric INRA, Pierrel Antibes), and Marc Denis R Larpin (Museum Nationale d’Histoire Na- penoids allow to trace theers). transformation The of Araucariaceae biomolecules family into is geomoleculesurated characterized (biomark- tetracyclic by a diterpenoids. remarkable However, predominanceabietanes a of derived sat- high could abundance also be ofcariaceae pointed could sesquiterpenoids be out. and summarized The as palaeochemotaxonomy follows: of the Arau- of aromatic diterpenoids. The particularities ofilar these abundance compounds with are no that predominance they of are of any sim- specific compounds (Fig. 9d). 5 Conclusion Palaeochemotaxonomy of the Araucariaceaeuration family by is confined evaluated pyrolysis of using extant artificial species. mat- Major structures of sesqui- and diter- ranes). Phyllocladane-type compounds, being largelyaceae widespread species, in are most neither of detecteddue in Araucari- fresh to nor in the pyrolysatespresent high of at abundance trace of levelcited in tetracyclic in the Sect. diterpenoids, 4.1.2 aliphatic the (Fig. fraction. 7) saturated The show a unidentified abietanes significant compounds are abundance. like The diversity those of diterpenoids acid), tetracyclic diterpenoids (like 5 5 25 15 20 10 25 20 15 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | , Phy- , Phy- Agathis , Phytochem- , J. Paleontol., Araucaria cooki Araucaria cunninghami , Phytochemistry, 13, 467–470, Cheirolepidiaceae Araucaria angustifolia Araucaria bidwilli 10536 10535 , Phytochemistry, 41, 995–1011, 1996. coniferae ord, H. T.: Biogeography of Araucariaceae, Australia and New Zealand ff , Phytochemistry, 15, 1401–1402, 1976. ), Biochem. Syst. Ecol., 28, 563–578, 2000. nity of Cretaceous resins from India and Myanmar, Int. J. Coal Geol., 85, 49–55, ffi Araucaria 2006. Experimental evidence for theGeochem., formation 38, of 28–36, 2007. geomacromolecules from plant lipids,for Org. palaeoflora and palaeoclimatic changes(France), at Org. the Geochem., Dogger/Malm 37, transition 610–625, of 2006a. the Paris Basin 2001. preservation of fossil arthropods: an experimental study, Proc. Roy. Soc. B, 273, 2777–2783, Pacific, J. Veg. Sci., 10, 793–804, 1999. organic transport in the Rio Tapajos, Brazilian Amazon, Org. Geochem., 32, 1443–1458, 2011. carbons from aromatization-rearrangement ofa new terpenoids class in of biomarker, the Geochim. sedimentary Cosmochim. Acta, environment: 60, 4747–4763, 1996. Forest Histories: No. 2 Araucarian Forests, edited by: Dargavel, J.,botanical 2, a 2005. penoids of Southern Hemisphere conifers, Biochem. Syst. Ecol., 35,istry, 342–362, 14, 2007. 2299–2300, 1975. residues in pottery vessels90, of 2005. the Roman age from Antinoe (Egypt), Microchem. J., 79, 83– lignanoids in the order and 1974. tochemistry, 13, 475–478, 1974a. tochemistry, 13, 471–474, 1974b. Araucaria bidwilli Arkansas: implications for diversity and78, reproduction 402–409, in 2004. the oil of Wollemia nobilis and its comparison with other members of the Araucariaceae ( i I. methylation of4259–4266, phenanthrene 1995. and alkylphenanthrenes, Geochim. Cosmochim. Acta, 59, hydrocarbons, Phytochemistry, 2, 205–214, 1963. eudesmane in petroleum, J. Chem. Soc. Chem. Comm., 5, 226–228, 1983. degradation products from the pyrolysisa of spore coal sporopollenins and derived the from Green some River pollen shale, exines, Chem. Geol., 12, 229–234, 1973. Gupta, N. S., Michels, R., Briggs, D. E. G., Collinson, M. E., Evershed, R. P.,and Pancost, R.Hautevelle, D.: Y., Michels, R., Malartre, F., and Trouiller, A.: Vascular plant biomarkers as proxies Gupta, N. S., Michels, R., Briggs, D. E. G., Evershed, R. P., and Pancost, R. D.: The organic Farella, N., Lucotte, M., Louchouarn, P., and Roulet, M.: Deforestation modifying terrestrial Enright, N. J., Ogden, J., and Rigg, L. S.: Dynamics of forests with Araucariaceae in the Western Ellis, L., Singh, R. K., Alexander, R., and Kagi, R. I.: Formation of isohexyl alkylaromatic hydro- Dutta, S., Mallick, M., Kumar, K., Mann, U., and Greenwood, P. F.: Terpenoid composition and Dettmann, M. E. and Cli de Paiva Campello, J. and Fonseca, S. F.: Diterpenes from Cox, R. E., Yamamoto, S., Otto, A., and Simoneit, B. R. T.: Oxygenated di- and tricyclic diter- Colombini, M. P., Giachi, G., Modugno, F., and Ribechini, E.: Characterisation of organic Castro, M. A., Gordaliza, M., Miguel Del Corral, J. M., and Feliciano, A. S.: The distribution of Caputo, R., Dovinola, V., and Mangoni, L.: New labdane diterpenes from Caputo, R. and Mangoni, L.: Diterpenes from Caputo, R., Dovinola, V., and Mangoni, L.: New diterpenes from Caputo, R., Mangoni, L., Monaco, P., Pelosi, L., and Previtera, L.: Neutral diterpenes from Brophy, J. J., Goldsack, R. J., Wu, M. Z., Fookes, C. J. R., and Forster, P. I.: The steam volatile Axsmith, B. J., Krings, M., and Waselkov, K.: Conifer pollen cones from the Cretaceous of Aplin, T. T., Cambie, R. C., and Rutledge, P. S.:The taxonomic distribution of some diterpene Alexander, R., Bastow, T. P., Fisher, S. J., and Kagi, R. I.: Geosynthesis of organic compounds: Alexander, R., Kagi, R., and Noble, R.: Identification of the bicyclic sesquiterpenes drimane and Achari, R. G., Shaw, G., and Holleyhead, R.: Identification of ionene and other carotenoid The publication of this article is financed by CNRS-INSU. References 5 5 30 25 20 15 10 25 20 15 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | : with coniferae ¨ aischen Kreide, ´ eheret, J.-G., and Dis- sweet, grown in Nigeria, J. aus der europ seed oil identified as illuminant in , a new living Australian genus and re, T., and Veillon, J.-M.: Phylogenetic ff orestation of grassland with three species, ff Brassicaceae Araucaria Jussieu 10538 10537 Araucaria cunninghamii Wollemia nobilis , Org. Geochem., 42, 1315–1323, 2011. ¨ onig, W. A.: Enantiomers of sesquiterpene and diterpene hydrocarbons in Asteraceae species, Phytochem. Anal., 11, 99–105, 2000. ´ e, T.: Distribution and ecology of the conifers of New Caledonia, in: Ecology of the Southern r Cyclic terpenoids of contemporarycoals, resinous Organic plant Geochem., 10, detritus 877–889, and 1986. of fossil woods,related ambers compounds and in crude oils and sediments, Organic Geochem., 21, 249–256, 1994. relationships within Araucariaceae based1516, on 1998. rbcL gene sequences, Am. J. Bot., 85, 1507– remarks on their stigmata, and cotyledons, Trans. Linn. Soc. Lond., 8, 308–317, 1807. Jacobs, P., De Vos, D.,Nilotic and shells Waelkens, from M.: a first390, millennium 783–793, AD 2008. Coptic church in Bawit, Egypt, Anal. Bioanal. Chem., conifers – a review, Bot. Rev., 67, 141–238, 2001. iments and fossil plants36, from 907–922, the 2005. Miocene Clarkia Formation, Idaho, USA, Org.Araucaria Geochem., species and sediment frommochim. the Acta, Eocene 65, 3505–3527, Zeitz 2001. formation, Saxony, Germany, Geochim. Cos- markers of pulp-fibre-derived pollution in the DerwentFreshwater Res., Estuary, Tasmania, 49, using 7–17, sterol 1998. biomarkers, Mar. Bessoule, J.-J.: Taxonomy ofposition, gymnospermae: Phytochemistry, multivariate 58, 101–115, analyses 2001. of leaf fatty acid com- Essent. Oil Res., 17, 459–461, 2005. sity in the Mesozoic, Zool. Anz., 246, 257–277, 2007b. nar, J.-R.: Occurrence of triterpenyl acetates in soil and their potential as chemotaxonomical carbons in petroleum, Org. Geochem., 10, 825–829, 1986. vironmental reconstruction, Annu. Rev. Ecol. Syst., 32, 397–414, 2001. Palaeontogr. Abt. B, 276, 97–131, 2007a. some Australian coals, sediments2147, and 1985. crude oils, Geochim. Cosmochim. Acta, 49, 2141– Conifers, edited by: Enright,171–196, N. 1995. J. and Hill, R. S., Cambridgespecies University in Press, the Cambridge, Araucariaceae, Telopea, 6, 173–176, 1995. in Horto Regio Parisiensi Exaratam Anno 1774, L’Herissant, Paris, 1978. Alcheringa, 11, 325–335, 1987. ers and mean carbon residenceSoil time Biol. after Biochem., a 43, 1341–1349, 2011. oxisol decrease substantially2010. after 90 years of pasture, Org. Geochem., 41, 1219–1224, extant land plants: a contribution to2006b. palaeochemotaxonomy, Org. Geochem., 37, 1546–1561, ff Smith, P.: The chemotaxonomy of plants, Arnold, London, 1976. Singh, R. K., Alexander, R., and Kagi, R. I.: Identification and occurrence of norcadalenes and Simoneit, B. R. T., Grimalt, J. O., Wang, T. G., Cox, R. E., Hatcher, P. G., and Nissenbaum, A.: Setoguchi, H., Asakawa Osawa, T., Pintaud, J.-C., Ja Salisbury, R. A.: XI V. The characters of several genera in the natural order of Romanus, K., Van Neer, W., Marinova, E., Verbeke, K., Luypaerts, A., Accardo, S., Hermans, I., Otto, A., Simoneit, B. R. T., and Rember, W. C.: Conifer and angiosperm biomarkers in clay sed- Pietsch, M. and K Otto, A. and Wilde, V.: Sesqui-, di-, and triterpenoids as chemosystematic markers in extant Otto, A. and Simoneit, B. R. T.: Chemosystematics and diagenesis of terpenoids in fossil conifer Leeming, R. and Nichols, P. D.: Determination of the sources and distributionMongrand, of S., sewage and Badoc, A., Patouille, B., Lacomblez, C., Chavent, M., Cassagne, C., and Lavrieux, M., Jacob, J., LeMilbeau, C., Zocatelli, R., Masuda, K., Br Olawore, N. O.: Analysis of the leaf oil of Kunzmann, L.: Araucariaceae (Pinopsida): aspects in palaeobiogeography and palaeobiodiver- Noble, R. A., Alexander, R., Kagi, R. I., and Nox, J. K.: Identification of some diterpenoid hydro- Kunzmann, L.: Neue Untersuchungen zu Kershaw, P.: The southern conifer family Araucariaceae: history, status, and value for paleoen- Noble, R. A., Alexander, R., Kagi, R. I., and Knox, J.: Tetracyclic diterpenoid hydrocarbons in Jussieu, A. L. D.: Genera Plantarum Secundum Ordines Naturales Disposita, Iuxta Methodum Ja Jones, W. G., Hill, K. D., and Allen, J. M.: Huang, Z. Q., Davis, M. R., Condron, L. M., and Clinton, P.W.: Soil carbon pools, plant biomark- Hill, R. S. and Bigwood, A. J.: Tertiary gymnosperms from Tasmania: Araucariaceae, Heim, A., Hofmann, A., and Schmidt, M. W. I.: Forest-derived lignin biomarkers in an Australian Hautevelle, Y., Michels, R., Lannuzel, F., Malartre, F., and Trouiller, A.: Confined pyrolysis of 5 5 30 25 20 15 10 30 25 20 15 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | ´ eum National of Lyon (France) (Brazil, North Argentina(S. Chile and of Lyon (France) Montet (Nancy, France) (Northland and Coromandel d’Histoire Naturelle , 2007. 10539 10540 , , 1994. Podocarpus zamiifolius ¨ , uttmann, W.: Aromatized arborane/fernane hydrocarbons Dammara lanceolata , Dombeya excelsa , Aiton ex Lambert, 1837 Australia (Queensland and Botanical Garden of (Salisb.) Franco, 1952 Norfolk Island found in the trade Corbasson, 1968 South of New Caledonia Botanical Garden (Bertol.) Kuntze, 1898 South America Botanical Garden de Laubenfels, 1969 South of New Caledonia Botanical Garden (Molina) K. Koch, 1873 South America Botanical Garden of (D. Don) Loudon, 1829 New Zealand Mus Buchholz, 1949 New Caledonia Botanical Garden (C. Moore ex F. Muell.) Bailey, 1883 Australia Botanical Garden Hooker, 1843 Australia Botanical Garden of the (Lindl.) Mast. 1892 New Caledonia Botanical Garden Johns, Hill and Allen, 1994 Australia Wollemi Pine France A. imbricata A. excelsa A. braziliana, Columbea angustifolia Dammara moorei Dammara australis Com. : bunya, bunya pineCom. : bernier’s araucariacunninghamii (Queensland) Villa Thuret (France) of Lyon (France) Com. : parana pine, candelabraaraucana treeSyn. : Com. : monkeypuzzle tree and Paraguay) Com. : hoop pine, Moretonheterophylla Bay pineSyn. : Eutassa heterophylla Com. : SW. norfolk Argentina) Island pine laubenfelsii Com. : de Laubenfels araucarianemorosa North of NewCom. South : Wales) boise araucaria– Montet (Nancy, France) Com. : wollemi pine (Wollemi National Park) of Lyon (France) New Guinea of Lyon (France) – Syn. : Com. : moore kauri robusta Com. : queensland kauri, smooth-barkkauri, kauri pine (Queensland) of Lyon (France) Com. : kauri, New Zealandmoorei kauriSyn. : peninsulas) (Paris, France) Syn. : erent organs of a relictual conifer Wollemia nobilis, Biochemical Systematics doi:10.1007/bf02344070 ff doi:10.1007/s10661-006-9514-0 BunyaEutecta bidwillii bernieri Indermedia Phylogentic relationships, geographical range and provenance of selected Araucari- as molecular indicators ofthe Saar-Nahe floral Basin, changes southwestern Germany, in1994. Geochim. Upper Cosmochim. Acta, Carboniferous/Lower 58, Permian 4689–4702, strata of implications of petroleum hydrocarbonoutfalls contaminants along in the the coastal sediments229–237, of near Laizhou major Bay, drainage Bohai Sea, China, Environ. Monit. Assess., 125, sults, edited by: Eglinton, G. and Murphy, M. T. J.,etation Springer, changes Berlin, during 599–618, Jurassic 1969. times, Geochim. Cosmochim. Acta, 64, 1417–1424, 2000. 107, 493–502, Palynology, 37, 133–154, 1982. derived from di and Ecology, 38, 131–135, 2010. Wollemia nobilis Araucaria Araucaria angustifolia GenraAgathis Section Species australis Geographical range Samples provenance Table 1. Vliex, M., Hagemann, H. W., and P Wang, L., Lee, F. S. C., Wang, X. R., Yin, Y. F., and Li, J. R.: Chemical characteristics and source aceae species. van Aarssen, B. G. K., Alexander, R., and Kagi, R. I.: Higher plant biomarkers reflect palaeoveg- Thomas, B. R.: Kauri resins-modern and fossil, in: Organic Geochemistry – Methods and Re- Stockey, R.: Mesozoic Araucariaceae: Morphology and systematic relationships, J. Plant Res., Stockey, R. A.: The Araucariaceae: An evolutionary perspective, Review of Palaeobotany and Staniek, A., Muntendam, R., Woerdenbag, H. J., and Kayser, O.: Essential oil constituents 5 15 10 Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | – – + –– ++ ++ – tr tr tr + + ++ ––– + ++ + ++ ++ ++ + ++ ++ ++ + + – tr tr tr tr tr + + + + +++ + ++ + + ++ ++ + + + + + ++ + + + + + ++ + + + ++ – tr tr + + ++ + + ++ + + + + + + + + + + + + + + ++ 10542 10541 tr ++ Cupressaceae Pinaceae Podocarpaceae Taxaceae Taxodiaceae Sciadopityaceae + + + + + + + ++ Araucariaceae + + ++ + ++ + + + + + + ++ + + + + ++ ++ –– – ––– ––––– trtr tr tr tr tr + + + + ++ ++ ++ + + ++ ++

Agathis Araucaria Wollemia + ++ + ++ ++ + ++ ++ ++ + ++ ++ + + ++++++++ + +++ ++ + + + ++ +++ ++ + ++ + + + + + + + + ++ ++ + + + + + + + ++ + + + + + + + + + ++ ++ ++ + + + ++ ++ + + + ++ ++ ++ + + + + + + + + + + + + + + + ++ + + ++ ++ + + + + + + + + ++ + + + + + + ++ + + + + + + + + ++ + + + + + + + ++ ++ + + ++ + + + + + ++ + + ++ ++ + + ++ ++ + + ++ + + ++ ++ + + + + + + + + + + + ++ australis moorei robusta angustifolia araucana bidwillii bernieri cunninghamia heterophylla laubenfelsii nemorosa nobilis (Conifer) Major detected sesqui- and di- terpenoids types in Araucariaceae species. Pinophyta Phylogenetic classification of conifers. (252 Da) (252 Da) tr 24 22 H H -kaurane -beyerane normal abietane (dehydroabietic acid) phenolic abietane (ferru-ginol, sugiol and hinokiol) diagenetic products tr tr tr tr tr tr tr tr tr tr tr tr 19 18 Ent ?C ?C Phyllocladane-type hydrocarbons Abietane-type Ent Pimarane-type Isohexyl alkylaromatic Chamazulene Pentametyl-dihydroindene Diterpenoids Labdane-type Eudesmane-type Cadalane-type Ionene Class of structure Sesquiterpenoids Farnesane-type Bisabolane-type Fig. 1. Table 2. Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | (f) (c) and 3e (a) Saturated 3f 190 184 188 208 180 180 200 3b (a) 170 193 retention time 170 173 169 H 161 160 3f 180 160 177 154 158 4β(H)-eudesmane 150 150 165 160 140 141 145 Araucaria laubenfelsii Araucaria cunninghamii Araucaria nemorosa Araucaria heterophylia 140 tr 149 130 128 140 130 127 137 120 3e 3e 120 115 110 120 115 123 m/z 110 105 (H)-eudesmane (identified from 100 105 3f3f 109 100 β 100 90 91 4 Norcadalene 90 91 95 84 3b 3b 80 Pentametyl-2,3-dihydroindene 84 80 77 80 81 (d) retention time 77 species. The mass spectra of Fig. 3a–f 70 69 (1-isopropyl-7-metylnaphthalene ) 3f3f 70 69 63 60 60 65 60 55 50 51 50 51 Araucaria bernieri Araucaria bidwillii Araucaria angustifolia 40 Araucaria araucana f 41 b d tr b 40 Araucaria 15 208 10544 10543 180 n-C 184 200 170 * H * * 3c chamazulene (identified using Wiley275 database); * 169 H * * * 160 * 3a 180 * * * * 153 * * (e) 4α(H)-eudesmane * 150 * * 165 tr 160 141 m/z 165 140 tr retention time 149 130 norcadalene (identified from spectrum in Singh et al., 1994); x x x 128 x H 140 14 135 120 tr n-C (b) 115 123 110 tr 120 Araucaria laubenfelsii Araucaria cunninghamii Araucaria nemorosa Araucaria heterophylia m/z 102 100 15 92 109 90 100 Chamazulene n-C 95 H 80 3c * * * 77 * 80 * * * 81 70 * 70 * 3a * * * 63 * 69 60 * * * * * * 60 * * fractions of pyrolysis products of 51 50 55 * 41 40 40 * retention time 41 c e a (b) Mass spectra of some identified Araucariaceae sesquiterpenoids. 14 H x Partial chromatograms (sesquiterpenoids retention time window) of aliphatic n-C Araucaria bernieri Araucaria bidwillii Araucaria angustifolia Araucaria araucana (H)-eudesmane (identified using Wiley275 database); a α pentametyl-2,3-dihydroindene (identified using Wiley275 database). spectrum in Alexander et al., 1983); Fig. 3. cadalane-type compounds; 4 Fig. 2. aromatic are given in Fig. 3. x: absence of compound; tr: at trace level ; *: cadalane-type compounds. Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | 6c 6c (b) 6c 6c to 7g 7g 7f 7f 7e 7e 7e (d) 254 260 7d 7d 7d 7d 254 4c 240 and aromatic 238 4e 4d 240 220 (a) 220 retention time 220 7c 220 7c 7c 7b 7b 7b 207 1,3,4-trimetyl-2-(4- tr 4b 205 6-ethyl-2-dimetyl-1- 200 tr 200 197 195 (a) (c) 180 180 184 183 x 169 169 160 6a Araucaria laubenfelsii Araucaria cunninghamii 160 Araucaria nemorosa Araucaria heterophylia 153 153 6c 6c 6c m/z 140 141 140 141 7f 204 7g 128 128 7f 200 120 7f 120 7f 7f 7f 7e 7e 7e 7e 115 115 187 105 7d 7d 7d 100 100 180 4c 98 91 4e 89 4d 80 170 77 80 79 m/z 183 160 65 159 69 7c 7c 60 7b retention time 7b 7b 55 60 55 145 c e 4b 41 40 140 species. The mass spectra of Fig. 4b–e are given 133 120 240 240 119 10546 10545 240 238 Araucaria bernieri Araucaria bidwillii Araucaria angustifolia Araucaria araucana 6a b 224 223 105 220 220 100 211 209 91 6e H H H H 201 200 200 84 H 193 m/z 133 80 Araucaria H 77 181 6a 180 180 x 178 x x x 63 60 169 x x 160 x H 55 160 168 H 153 153 a H H H retention time H 141 141 140 2,6-dimetyl-1-(4-methylpentyl)- naphthalene; H H 140 H 128 128 m/z 120 (b) 120 115 115 Araucaria laubenfelsii Araucaria cunninghamii Araucaria nemorosa Araucaria heterophylia 105 100 101 100 6e H H 89 6e H 89 H H 80 80 H 77 76 x x m/z 169 x x 60 . 60 x x 55 55 x H H (c) 41 40 d b 41 40 H H H H 6d H retention time to Partial chromatograms (diterpenoids retention time window) of aliphatic Mass spectra of identified isohexyl alkylaromatic hydrocarbons in Araucariaceae species H H (e) Araucaria bernieri Araucaria bidwillii Araucaria angustifolia Araucaria araucana fractions of pyrolysis products of a in Fig. 4. The massgiven spectra in of Fig. Fig. 7. 6a–e *: are labdanes; given tr: in in Fig. trace; 6. x: The absence mass of spectra compound. of Fig. 7a–g are (b) Fig. 5. and methylpentyl)benzene; (4-methylpentyl)-naphthalenemethylretene. Mass spectra ofditerpenoids some but unidentified similar to Araucariaceae some of the identified isohexyl alkylaromatic hydrocarbons: and mass spectral cleavage patterns (identified after Ellis et al., 1996). Fig. 4. Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | (b, 17- (Only clero- (d) (a) (b) 288 280 248 273 240 233 260 259 220 phyllocladane 218 245 as a 16(H)-homo- 240 Tentatively identified identified Tentatively 231 260 240 203 258 250 264 200 260 252 220 238 217 7 m/z 273 243 237 223 240 189 H 220 245 9 3 230 240 H 180 203 4 m/z 231 - C 200 223 227 209 178 3 - C 2 220 200 210 220 189 195 208 165 209 180 160 177 m/z 1 200 195 180 199 200 179 190 193 164 168 160 179 m/z 203 180 m/z 187 140 180 160 170 179 : 16(H)-homo-phyllocladane. 173 150 155 244 165 Agathis Only for 240 160 140 165 145 160 157 152 140 135 150 120 119 (e) 229 151 Monoaromatic labdane; m/z 141 143 128 140 120 140 108 220 120 131 137 130 123 115 128 100 109 213 119 120 123 (a) 120 105 100 95 115 95 100 110 Tentatively identified as identified Tentatively 109 a trimethyl-phenanthrene 95 . , in Fig. 8a): in the aliphatic fraction; 200 105 80 109 100 195 100 81 95 : ? : ? : ? 83 80 95 80 91 90 30 22 24 : ? 36 H H H 69 81 183 69 80 81 80 19 18 19 H 77 60 180 19 C C C 60 60 70 69 69 C 55 67 55 173 Norlabdane 60 60 e b d f h 55 55 55 c 41 41 40 40 50 40 160 Agathis 159 240 260 280 143 238 276 280 (a, b ,d ,e) 250 140 274 280 260 252 10548 10547 278 260 261 252 7 223 237 260 240 220 H 260 128 237 3 230 247 259 7 8 - C 245 260 11 221 120 245 240 223 240 H 220 240 9 115 5 6 209 200 m/z 193 209 210 231 (only for - C 231 193 10 208 5 220 220 105 m/z 231 240 220 200 217 220 194 100 180 1 4 195 3 179 217 190 2 (i) 91 2 3 203 200 Agathis 200 Only for 182 205 179 m/z 193 180 220 191 165 203 200 m/z 109 189 160 170 m/z 1 80 180 165 77 177 180 177 165 207 152 160 189 200 152 a monoaromatic labdane Tentatively identified as Tentatively and m/z 159 163 67 141 150 140 152 180 163 160 193 160 m/z 175 141 Only for 151 60 140 175 140 150 m/z 128 A. nemorosa 55 180 179 130 128 140 137 140 120 a 128 119 (h) 160 163 135 : ? 120 115 36 165 121 115 m/z 120 121 105 120 110 160 H 105 149 100 a clerodane 20 109 103 140 151 100 C 109 : ? 89 100 100 90 : ? : ? : ? 137 95 89 34 95 89 24 24 22 140 80 138 H 77 H H H 80 81 20 Tricyclic diterpane Tricyclic 77 Tentatively identified as identified Tentatively 81 80 80 77 19 18 19 120 C 70 67 C C C 123 123 69 67 67 120 60 60 60 60 55 109 55 55 55 55 i c e 50 g a 100 109 Tetracyclic diterpane Tetracyclic 95 100 95 81 80 , in Fig. 5a), 81 80 69 69 60 in the aromatic fraction (Figs. 5b, 8b). 60 55 55 a 17-norphyllocladane methylretene (tentatively identified according to Alexander et al., 1995); Tentatively identified as identified Tentatively d 41 40 b 41 40 Mass spectra of some tentatively identified Araucariaceae diterpenoids and sug- Mass spectra of some unidentified but recurrent Araucariaceae diterpenoids. (c) A. nemorosa Fig. 7. for dane; norphyllocladane (identified according to Noble et al., 1986); c, d, e, f, g) Fig. 6. gested mass spectral cleavage patterns for Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | Discussion Paper | 6c 6c X X 7f 7f 7f 7f 7e 7e 7e species. Wollemia 6c 7d 7d 4c 4e 4d retention time 7c retention time 7f Agathis 7b H 7b x 4b H Aromatic diterpenoids 7f retention time win- x 7e (c, d) H (b) H 7d ? Agathis moorei Agathis robusta 6a Agathis australis H 4c H 4e x retention time 4d H H H H H x H Diterpenoids H H H H H X 7c x X H H 7b 7i H and diterpenoids 4b H H retention time H and diterpenoids Aliphatic diterpenoids c fractions of pyrolysis products of 7h (a) (a, b) Agathis moorei Agathis australis Agathis robusta b 3e 10550 10549 (b, d) d 3f 15 n-C H 3b * 3c retention time 3f * * * 3a and aromatic 3f * Aromatic sesquiterpenoids * * Agathis moorei Agathis australis Agathis robusta * (a, c) 3b 15 H * n-C retention time 3c * * * * 3f * H Sesquiterpenoids 3a * * * * * * 14 * * * x x n-C * x x * * * * a b ? retention time Partial chromatograms (sesquiterpenoids Partial chromatograms (sesquiterpenoids 14 . The mass spectra of Fig. 3a–c, f are given in Fig. 3. The mass spectra of Fig. 4b–e are n-C Aliphatic sesquiterpenoids Agathis moorei Agathis australis Agathis robusta a given in Fig. 4. Theare given mass in spectra Fig. of 7. Fig. *: 6c cadalane-type are compounds. given in Fig. 6. The mass spectra of Fig. 7b–f Fig. 9. nobilis windows) of aliphatic Fig. 8. The mass spectra ofFig. Fig. 4 3a–f by are b to given e.are in The given Fig. mass in spectra 3. Fig. of The 7. Fig. mass 6a, x: c spectra absence are of of given compound; Fig. in *: Fig. 4b–e 6. cadalane-type are The compounds. given mass spectra in of Fig. 7b–i dows) of aliphatic (left) and aromatic (right) fractions of pyrolysis products of