Org. Geochera. Vol. 22, No. 6, pp.

991-1004, 1994
Pergamon Copyright© 1994 ElsevierScienceLtd
0146-6380(94)00078-6 Printed in Great Britain.All rights reserved
0146-6380/94 $7.00+ 0.00

Extended saturated and monoaromatic tricyclic terpenoid carboxylic

acids found in Tasmanian tasmanite
D. A. AZEVEDO,l F. R. AQUINONETOl and B. R. T. SIMONEIT 2 .
Instituto de Quimica, Centro de Tccnologia, Bloco A, Sala 607, Universidade Federal do Rio de Janeiro,
Ilba do Fund~.o, Rio de Janeiro, 21949-900, Brazil and 2petroleum and Environmental Geochemistry
Group, College of Oceanic and Atmospheric Sciences, Oregon State University, Corvallis, OR 97331,
U.S.A.

(Received 23 November 1993; returned for revision 25 January 1994; accepted 1 June 1994)

Abstract--A series of tricyclic terpenoid carboxylic acids (C20--C40)was found in the acidic fraction of
Tasmanian tasmanite bitumen, occurring as a mixture of stereoisomers with mainly the 13~(H),14a(H)-
and 13~(H),14a(H)-configurations. These dominant acidic tricyclic consituents have the same carbon
skeleton as the ubiquitous tricyclic terpane biomarkers. A novel series of ring-C monoaromatic tricyclic
terpenoid carboxylic acids was also characterized. The series ranges from Ct9 to C39 and is the acidic
counterpart of the recently described series of monoaromatic tricyclic terpanes.

Key words--tricyclic terpenoid carboxylic acids, aromatic tricyclic terpenoid carboxylic acids, tasmanite,
biomarkers

INTRODUCTION their aromatic or unsaturated analogs. They were first

reported by Simoneit and Burlingame (1973) in the
Tasmanian tasmanite (Permian, 220-275 x 10 6 yr) is chemical oxidation products of tasmanite kerogen.
composed of compressed disks of tasmanites or tas- This was followed by Seifert (1975) with the identifi-
manitids (Newton, 1875). These unicellular organ- cation of C2~- and C24-tricyclic terpenoid carboxylic
isms are thought to be green algae (zoosporangia of acids in a California petroleum, based on their re-
Prasinophycean algae, Tappan, 1980; Sarjeant, 1989). duction to the corresponding hydrocarbons and
The tasmanitids are unique in that they are derived analysis of the mass spectra. However, these struc-
from a single species and hence yield simple bio- tures were not fully characterized. This was done by
marker mixtures that can be related to the precursor Cyr and Strausz (1983, 1984). They determined the
organisms (Burlingame et al., 1969; Simoneit and structures of the carboxylic acids from C2~ to C26 by
Burlingame, 1973; Simoneit et al., 1990, 1993). their transformation into the corresponding hydro-
The Tasmanian tasmanite is of particular interest carbons and comparison with the natural terpanes,
because its bitumen is composed mainly of com- isolated from Alberta oil sand deposits. The hydro-
pounds with the same tricyclic terpenoid skeleton in carbon obtained from the methyl ester of the C20
all fractions studied (saturated, unsaturated and aro- carboxylic acid, however, did not correspond to the
matic hydrocarbons). The saturated plus monounsat- tricyclic terpane, furthermore, the natural ester did
urated hydrocarbon fraction comprises primarily the not match any of the four possible isomers at C-13
ubiquitous extended tricyclic terpane series (Aquino and C-14. On the basis of this evidence and the
Neto et al., 1992; Simoneit et al., 1990) and two less information obtained from the mass spectral frag-
abundant series of A 9{11) and A ~3t~4}-unsaturated tri- mentation pattern of the Cz0 ester and the corre-
cyclic terpenes (Azevedo et al., 1991, 1994). The sponding alkane, a new structure was proposed,
aromatic hydrocarbon fraction is characterized by a which was confirmed recently by synthesis of the
series of extended ring-C monoaromatic tricyclic standards (Kaufman et al., 1988).
terpanes (Azevedo et al., 1990, 1992). We now report the identification of a family of
Tricyclic terpane hydrocarbons have been studied extended tricyclic terpenoid carboxylic acids found in
extensively and are found in many petroleums and the acidic fraction of the bitumen from Tasmanian
sediments. One of their potential geochemical precur- tasmanite. A less abundant series of extended ring-C
sors could be the corresponding tricyclic carboxylic monoaromatic tricyclic terpenoid carboxylic acids is
acids. There are few citations of saturated tricyclic also reported. They are related to the already known
carboxylic acids in geological samples and none of tricyclic terpane hydrocarbon series, as well as to the
recently discovered ring-C monoaromatic tricyclic
*Author for correspondence terpanes.

991
992 D.A. AZEVEDOe/ al.

conditions were used for the HRGC-MS-C analysis

I , ~ t60 g (8) I on a Hewlett-Packard Model 5987A instrument, with
linear scanning (50-500 Da, 1.87 s/decade), electron
i (0H2Cl2/ CHoOH) impact ionization (70eV) and also SIM analysis
(single ion monitoring) for detection of the higher
anam,,n (a) I members of the series (C30-C~6) (Condition A). The
I 74&3 ,mg (S : 1.24%) I
SIM analysis was also done by high-temperature-
I HRGC on an Ultra 2 fused silica column (Hewlett-
Aliquot I Packard--5.5 m x 0.20 mm, df = 0.33/~m) and tem-
101,4 rag,
perature programming from 90-370°C at 20°C/min
I (McCarthy& Duthie Chrom.) (Condition B) for better visualization of the higher
members of the series (C36-C40).
J , I
Acids (/It)
I 70.1mg I 21.5 rng
I (s : o.86%)1 (S : 0.26%) nC[2 nCi6
I ~B:,.,%I.I (B : 21.2%)
i I I ~i nC e
I llFE!l ; li§ZT c., TIC

I I
Hono¢lrboxyllc Acids ! otn~ Aa=
( as methyl esters) i ( as methyl esters )
3,4 mg 17.0 mg n¢ 2
(S 0.042%) (S 0.21%) a ! .c.nC,. ioc~6 m/z 74
(B 3.3%) (B t6.8%)
(A t5.8%) (A 79.1%) ! !, J j ~c8

Fig. 1. Yields of various fractions from sediment and

bitumen of Tasmanian tasmanite.
~,
C
llti}dJ ~..
iC~4 'Ci~
,(J,i~ ,..,
m/z 88
!,c, 5
LU
E X P E R I M E N T A L
Z
The Tasmanian tasmanite (Permian) was collected
by B.R.T.S. and T. C. Hoering in an abandoned shale
CO
== L
mine on the Mersey River, 3 miles south of Latrobe, D [ !cJ5 iCz° m/z I01
Tasmania (latitude 41°17'S, longitude 146°27'E and
elevation 35 m). A sample (60 g, Fig. 1), was washed
with dichloromethane, powdered in an agate mortar,
and extracted four times with dichloromethane/ h i, -- i , i • i, t
1
methanol (9:1) in an ultrasonic bath. The solvent was C
~T ~i ,j.~6r I

removed under vacuum on a rotary evaporator and E ~! m/z 191

C~4T
an aliquot of the bitumen (101.4mg, Fig. 1) was
subjected to McCarthy and Duthie chromatography
on a SiO2/KOH column with elution of the acids by
, r • I , i , r , i , i , r , [ • i , r ' i , J, I
a solution of 10% formic acid in diethyl ether, after
C,gA m/z 1 4 5 + 2 0 5 +
removal of the neutrals with diethyl ether (Loureiro F , IC2~
et al., 1986; McCarthy and Duthie, 1962). The acid I ,C~6A
fraction was concentrated and treated with dia-
zomethane in diethyl ether to esterify the acids. The
monocarboxylic acid methyl esters were purified by
silica gel (Merck) thin-layer chromatography (TLC)
4 6 8 I0 ~e 14 ~6 18 20 22 24 26 28 30
using hexane/dichloromethane (7:3) as eluent and lR (min)
methyl abietate as elution standard. The band corre-
sponding to the methyl esters was scraped off the
TLC plate after visualization under UV-light Fig. 2. (A) Total ion current (TIC) trace of the acid fraction
(254 nm) and eluted with diethyl ether. The yields of (as methyl esters) of Tasmanian tasmanite bitumen; (B) m/'_
the various fractions are shown in Fig. 1. 74 fragmentogram, key ion characteristic for normal car-
boxylic acid methyl esters; (C) rn/z 88 fragmentogram, key
The monocarboxylic acid methyl esters were ana- ion characteristic for 2-methyl-isoprenoid carboxytic acid
lyzed by high resolution gas chromatography methyl esters; (D) m/z !01 fragmentogram, key ion charac-
(HRGC) on a glass capillary column (SE-54, teristic for 3-methyl-is0prenoidcarboxylic acid methyl es-
25 m × 0.30 mm, dr = 0.25/~m) using hydrogen as ters; (E) m/z 191 fragmentogram, key ion characteristic for
tricyclic terpenoid earboxylic acid methyl esters; (F) com-
carrier gas and temperature programming from posite fragmentogram (m/z 145,203,217,231,245,273, 287,
80-30@C at 10£/min. The same chromatographic 329, 343, plus 357) for the aromatic tricyclic terpenoid
carboxylic acid methyl esters.
 Tricyclic terpenoid carboxylic acids 993

Standard methyl 13~(H),I4fl(H)- and 13fl(H), reduced on Pd/C 5% in hexane (Cyr and Strausz,
14fl(H)-dihydroisocopalate (C' and D' in Fig. 6) were 1983) or on PtO2 in methanol/ethyl acetate (5:1)
obtained by cyclization of methyl copalate in formic (Imamura and Rfiveda, 1980) to obtain only t h e / / p
acid (Heissler and Ladenburger, 1988). They are the isomer. The fl//standard was characterized by proton
enantiomers of the C20 tricyclic terpane acid methyl nuclear magnetic resonance and mass spectrometry.
esters. (Enantiomers have the same retention time in Veadeiroic acid methyl ester (14-ethylpodocarpa-
GC analysis, i.e. they coelute. Note: Usually geo- 8,11,13-triene-13-carboxy methyl ester) standard was
chemical literature refers to the stereochemistry of the isolated from Velloziaflavicans and characterized by
enantiomers, in this case fl~ and ~t~t, and this will be Pinchin and coworkers (1978). These standards were
used in the discussion of our results). Copalic acid coinjected with the monocarboxylic acid methyl ester
was isolated from copaiba oil, cyclized and then fraction on a SE-54 column (see above) and an OV-17

J A ~T 1 1114 22 27 32 C02~'[ 3

, ,~'; . .~ . . . ." . . . . . ',--, 3 , ",- 474

C C32T
~ m/z
488
c).
(/} I r T I t i I I I I K I I I I i I I I
OC

~ I I l l l l I I I I I I I I I I I I
m~

~Z

C~T
m/z
516

m/z
530
I I I i i i i I i ~ i ; i i i i i i i w

C 3 S T G m/z

16 20 24 28 32 36 40
t~ (min)

Fig. 3. Single ion monitoring (SIM) of the methyl ester fraction of Tasmanian tasmanite bitumen
(Condition A): (A) m/z 191 fragmentogram, characteristic for the saturated tricyclic terpenoid carboxylic
acid methyl esters; (B-G) mass fragmentograms of the molecular ions of the higher members of the series
(C31-C36).
994 D.A. AZEVEDOet al.

fused silica capillary column (Perkin-Elmer. canoic acid: iC~5 = 3.7.1 l-trimethyldodecanoic acid:
25m x 0.32mm, dr=0.25 pm) and analyzed by iC~9 = 2,6,10,14-tetramethylpentadecanoic acid; and
HRGC-MS-C. iC20 = 3.7.11.15-tetramethylhexadecanoic acid tall as
methyl esters. Fig. 2(C,D)].
RESULTS AND DISCUSSION
The total ion current (TIC) trace of the monocar- Extended tricyclic terpenoid carboxylic acids
boxylic acid fraction (as methyl esters J, the m z 74 Extended tricyclic terpenoid carboxylic acids are the
fragmentogram (key ion for linear carboxylic acid most abundant cyclic acids in Tasmanian tasmanite
methyl esters), m/z 88 and m / z 101 fragmentograms bitumen and have the same carbon skeleton as the
(key ions for isoprenoid carboxylic acid methyl es- tricyclic terpanes. They are of particular interest be-
ters), m/z 191 fragmentogram (key ion for tricyclic cause they could be the precursors of tricyclic terpane
terpenoid carboxylic acid methyl esters}, and the hydrocarbons. This hypothesis is reinforced by the fact
composite fragmentogram (m z 145. 203. 217. 231. that other biomarkers which might serve as precursors
245, 273, 287, 329, 343 plus 357) for the aromatic for tricyclic terpanes te.g. hopanes, steranes) are essen-
tricyclic carboxylic acid methyl esters are shown in tially absent (Simoneit et al.. 1990). SIM analysis of
Fig. 2. The monocarboxylic acids are composed the same fraction shows that the extended tricyclic
mainly of n-alkanoic acids ranging from C,, to C3,, terpenoid carbox31ic acids range at least up to C40
with a carbon number maximum at C,o, and a series (Figs 3 and 4). The tricyclic terpenoid carboxylic acid
of extended tricyclic terpenoid carboxylic acids dom- series also comprises a mixture of stereoisomers for all
inated by the C2~ homolog. A minor and novel series homologs, consistent with the low maturity of the
of monoaromatic tricyclic terpane carboxylic acids sample ( R o - 0.3-0.4%) (Aquino Neto et al.. 1992)
was also present. Four isoprenoid carboxylic acids The stereoisomeric distribution for the tricyclic
were observed, namely iCt4 = 2.6.10-trimethylunde- terpenoid carboxylic acids is shown in Fig. 5 The

( r A C'40

g:
._~
"~ 8 C2e

I I I I I 1 1 I [ I ~ r I I I I
C
C39
1~.. .

I [ I I r I I 9 [ [ I i ] I i i
6.0 7.0 BO .0 I0.0 1i.o 12.o 13.0
tR ( rnin )

Fig. 4. Single ion monitoring (SIM) of the methyl ester fraction of Tasmanian tasmanite (Condition B):
(A) m/z 191 fragmentogram, characteristic for the saturated tricyclic terpenoid carboxylie acid methyl
esters; (B) m/z 432 fragmentogram, molecular ion of the Cz8 homolog; (C) m/z 586 fragmentogram,
molecular ion of the C3~ homolog~
 - tO0
60OOO-
40000- Cz 0
nJll - 80

-6o

-40 m/z 191

zoo(X)o
"~~~I~ -20
.

i i I I -0

-I I'a=c=° 80

40 m/z 320
AJ~ I
I 1 ,0

4O001 40
m/z 334
O, I i
iI
I I
0

Z0001 B~',,Il.,,a~CZ2
10001
O, ~T- I ~ I I I IIt I'° 40
0
m/z 340

{,- lOOO Czs

I:
,, ..o.~

0 m/z 362
0.
¢/) s°° 1
o, ~bO

rr"
e,-....(i...o.o Cz4

~°°°t R/S R/S

I: m/z 376
n-- ~¢. I,c~C=5
R/S'" "
mh 390
400
O, , 0 I'° 40

RIS 6~.,I~Czs
2o0t 40 m/z 404
Oi I I l ' l ~ l ~ I "I I I
I i , 0

,o:t [."!~ c,I 40 m/z 418

"!1 c..
i '

}oo
! 0

40
mh 432
II I I I | | l | i I | ~ 0

i R/S!8o
.i~,, 400 m/z 446
; i i I I i I I : I

16 19 22 25 28
tR ( min)
Fig. 5. Ion map of tricyclic terpenoid carboxylic acid methyl esters for C204229. Key to stereoisomers (see
also Fig. 7): fl~t-C20 = 13fl(H), 14~t(H)-tricyclic terpenoid carboxylic acid methyl ester with 20 carbon atoms
(see Fig. 6).
996 D.A. AZEVEDOet al.

.•"
/\ I~ (A) '
'ICO2H
/ "" cto. (A')
CO2H
observed for the tic(-C:5 tricyclic terpane, which is the
first pseudohomolog of the saturate hydrocarbon
series with a chiral carbon in the side chain.
These results differ from those of Cyr and Strausz
(1983, 1984). Their tricyclic carboxylic acid concen-
trate of the Alberta oil sands consisted primarily of
the C20 and Cz3 and the diastereomeric C24 to C2~
homologs. The C2~ and the pair of C24 diastereomeric
acids comprised about 75% of the series. They did not
H ' "O2 observe coelution of the [3c~and ~tct-C:0standards with
CO2H
their sample and thus proposed another structure lbr
the C20 compound, which was confirmed recently by
oq6 (B) Kaufman et al. (1988). They retained the tricyclic
13o.(B')
terpenoid structure for the C2rC26 homologs.

~ I~ (c)
• .,,x

CO2H

a13(C')
CO2H
The mass spectra o f the C20~$29 members of the
tricyclic terpenoid carboxylic acids (as methyl esters.
no C2v) are shown in Fig. 8. The mass spectrometric
fragmentation pattern (Fig. 9) consists mainly of M +.
[M 15]+, m / z 191 (base peak), m / z 261 (from the
loss of the side chain at C-14), m / z 313 (from cleavage
at C-22 of the isoprenoidal side chain, for homologs
>C24), and smaller fragment ions (m/z 109, m / z 123)
probably derived from fragmentation of rings A and
CO2H CO2H
B, as was also observed by Cyr and Strausz (1983,
1984).

Monoaromatic tricvclic terpenoid carboxylic acidL~

cte{ (D) I~ (O') A novel series ol ring-C monoaromatic tricyclic
Fig. 6. Structures of the four stereolsomerlc C20 terpenoid carboxylic acids was found in the acid
saturated tricyclic terpenoid carboxylic acids: A fraction of this bitumen. The SIM analysis shows that
13fl(H),I4/~(H); B 13ct(H),t4f(H); C 13#(H),I4~t(H); the series ranges from C~9 to at least C39 (Fig. 10),
D--t 3~t(H),14a(H), and their enantiomers: A" with the C~9 homolog being the most abundant.
13c~(H),14ct(H); B' 13fl(H),140t(H); C' 13~(H),I4~(H);
D' 13fl(H),14fl(H).The pair ofenantiomers are not separ- The mass spectrum of the C t9 ring-C monoaro-
ated by the achiral GC conditions employed. matic tricyclic terpenoid carboxylic acid (as methyl
ester) is shown in Fig. 11 Inspection of the mass
spectra of the homologous senes shows the same
presence of 13fl(H),14et(H)-C20 and 13ct(H),14ct[H)- fragmentation pattern as the recently described rmg-
C20 was confirmed by coinjection of the synthetic C monoaromatic tricyclic terpane series, differing
standards (see Experimental) with the acid fraction only by 58 mass units (CO2Me group attached at the
on SE-54 and OV-17 glass capillary columns. All four C-14 side chain). The principal fragments include the
isomers of the C2~ homolog are observed: Bfl, ~fl, fla, aromatic C-ring plus the side chain with the CO 2Me
act (Fig. 6). The stereoisomeric distribution was group, which shift by 14 mass units up to C24 from
suggested by comparison with that observed for the one homolog to the next. For compounds greater
tricyclic terpane series by Chicarelli et aL (1988) and than C24, the fragmentation is dominated by cleavage
the observed elution order of the tiff and ~tfl-C20 of the side chain resulting in a base peak at m z 145.
standards. For C22 and C2s only the flct and ct~tisomers analogous to the monoaromatic tricyclic terpanes
were observed (see Experimental for clarification of (Azevedo et al.. 1990).
the conflicting stereoisomer nomenclature for the A suitable representation of the series up to C~0 is
standards and the geological series). obtained by plotting the composite fragmentogram of
Resolution of the R/S fl~tisomers for the C24 to C2e ions m/z 145. 203. 217. 231. 245, 273. 287. 329. 343.
pseudohomologs increases with higher carbon num- plus 357 (key ions of the series. Fig. 12). It should be
bers (Fig. 7). It is interesting to note that in the noted that the surprisingly intense C26 homolog
tricyclic terpane hydrocarbon series the C24 homolog (suggested by its relative retention time vs C25 and
does not show resolved R/S stereoisomers because C28, molecular ion and key fragments) may be due to
the carbon at position 22 is achiral by being linked to coelution as the composite fragmentogram involves
two methyl groups. In the C24 tricyclic terpenoid 10 different ions. Also the C~9 homolog at 16.8 min
carboxylic acid the carbon at position 22 is chiral (Fig. 12) has an identical mass spectrum as the other
because it is linked to a methyl and a carboxy group. isomer, which points to a probable C-13 carboxy
In the GC trace it shows a broadened peak for the methyl structure. By analogy with the interpretation
two resolved isomers. This is the same behaviour as of the mass spectra of the ring-C monoaromatic
 Tricyclic terpenoid carboxylic acids 997

'*'~ I c2, R/s

'"'1 "1
'"'1
1"~I
I I
• il

19.0 19.5 20.0 2@.5 21.0 21.5 ::=.g 22.5 23.0 23.5 24.e

.o,t l~/~ l%s R/s rrVz 3 ~

8e~]

>~

19.0 19.5 2 8 .el 2@.5 21.@ 21.5 22.0 22.5 23.0 23.5 24.0
|ml

: 3 0 0 -' c~a HIS I mlz

.. ~i\~1 404
150"

5~" I '

19.0 19.5 2g.e ::'@.5 21.g 21.5 22,e 22.5 23.e ~3.5 24.e

tR ( min )
Fig. 7. Expanded mass fragmentograms displaying the isomers of C24-C26. Note the greater resolution
of the #~ R/S isomers when going from C24 to C26.

tricyclic terpane series (Azevedo et aL, 1990), we can (vi) Group 6: fragments ofm/z 145 and 159 due
also classify these fragments into 7 groups: to loss of the side-chain from Group 4 frag-
ments, Scheme 4;
(i)Group h molecular ion M+'; (vii) Group 7: fragments of m/z 255 and 241 due
(ii)Group 2: [M - 15] +, Scheme 1; to the loss of the side chain from M ÷. and
(iii)Group 3: m/z 69, Scheme 1; [M - 15] + ions respectively, Scheme 5.
(iv) Group 4: [ M - 1 1 1 ] ÷ and [ M - 9 7 ] + pair,
The CO: Me group is interpreted to be attached to
ranging from m/z 189 and m/z 203, + 14n
the side chain at the C-14 position. There is consider-
(n = 0, 1, 2, 3. . . . ) respectively, Scheme 2;
able evidence for this conclusion:
(v) Group 5: four homologous ions of decreasing
intensity, ranging over m/z 215, 229, 243, 257 (a) A standard compound, veadeiroic acid (the
plus 14n; [ M - 8 5 ] + to [ M - 4 3 ] +, respect- CO2Me group is attached at C-13) was coin-
ively, Scheme 3; jected with the acid fraction. Its mass spectrum
 191

191 i i~ 1{11 "9 026

69 I C20 (C21H3602) : 69
\ ~ [ 23 :
\1 81 95 )Z3
I" ~,o,
I 109 123 I

• / M-t-
137
z,3 z,3 z~o ~3 ~
M*
~,zo
- c I" I//I,
IJJJli
• 1631
/ I ,.
\
I,o,,,3__ "',o,
M + r 4o
~o
I.~ ~/ \ ~, / l a_. ~ll.l,IJ, J k. / II \ \ ~,,- 3.5= / /
(...,,, o , , ,-, , , ,--.-;...~._,.,__;%,_;.,.,.=,L,L,., f0
A
gO lJO I I I ! I I t I ! I ! 1 T I I l I I t
160 ZOO 240 280 320 80 120 160 200 240 280 3ZO 360 IlO 120 100 200 240 2BO 3tO 340 400 440
V
'" 191 ' Jgl

¢g) / C21 24 ao
69 95 lOg I'

\, , ( ,~3,3, M +4c, , M; ,-, -' ,rl

/ \ ", z z°
L11J l i • -
, _e
-- . ~ .',..,: "{ ' ", ,"t o - ~ ./. -. . . i . . . . .x. . . . X \
J,
" [ L J JI - . . . . L J . . . . , N.
,~, _-- , I i ; "', O I
' go ' ,~o ' ,~o' =~o' 2~o' . o 32o 2~o'2~o'=;o'3~o'3;o',;o 80 120 !tiO LRO0 Z40 280 3ZO 340 400 440

rr"
' '[' C .~o
I;' / 29 .co
' .o
163 zs9
" / I M+ M+ \ M+ -,to
,, , ~ . , °. ~,, . . . . ~ 333, ,o,. ,,, ~, ? ' ~;~ ~ ' ',~/3'3"3'°'z°'r I , ,,, ~," ~3, ,;, ) - -20
. I ~ [ t J , JLJ .,L ,. .\_.. : J ":,'/ ~. i':.. _l J , . , . . . . . . ,I , . - ~ . , ; ~,~, }o . ,-"~ ,=,',.*,l, I, t , , !
r o
l
'' ! ,
' ~) ' , ; o ' , k , ' ~ ' ~ o ' z ; o ' fio'~$o' ' ;o'~;o'~;o'=~o'~,;o'~o s~ ~,o ,oo 100 150 200 2mJO 300 3S0 400 450 5(X)

niiz . . . . .
Fig. 8. Mass spectra of the tricyclic terpenoid carboxylic acids (as methyl esters) for C20-.C, 6, and C2~ and C2~.
 Tricyclic terpenoid carboxylic acids 999

m/z
261 , ~
C40 (C41H7602,M . W . 600)

" /I I

CO2CH 3

% I ; ,/)

) J,
, m/z
191 m/z
m/z 313
123
Fig. 9. Proposed fragmentation pattern for the saturated tricyclic terpenoid carboxylic acid methyl esters.

m/z 145

6 0 0 80 90 10.0 11.0 12'.0 13.0

t- B
lc2o
O
¢1)
n-
o I I i i i i i 1. i i i i i i I

D cse
m/z 551

i i i i i i , i i , i i

E C39 ,

i !
' 6'.o ' ~:o ' ~o ' ~.o ' ,&o ' ,,:o ,2'.o ,3.0
tR ( min )
Fig. 10. Single ion monitoring (SIM) of the methyl ester fraction (Condition B): (A) m/z 145
fragmentogram, characteristic of the aromatic tricyclic terpenoid carboxylic acid methyl esters (>C24);
(B) m/z 411 fragmentogram, (M - 15) + ion for the C2s homolog; (C) m/z 481 fragmentogram, (M - 15)+
ion for the C~3 homolog; (D) m/z 551 fragmentogram, (M - 15)+ ion for the C3s homolog; (E) m/z 565
fragmentogram, (M - 15) + ion for the C 3 9 homolog.

OG 22/6~H
1000 D.A. AZEVEDOet al.

~:e3

:'/
k
v 1.119 % . ~ CO2CH3
7711
t- Z
O \ 0~9 ~e
r"

.> ~a

0 69 "41
rr"

• .. al.dJ,_LJl£1,++ ... ' + It+.++

...... + . . . . . . + + : i + i +'+. . . . ~ ' ~ .......... "~"" "" ++i ...... +++ ..... +k+ ......

m/z
Fig. 11. Mass spectrum of the C~9 ring-C monoaromatic tricyclic terpenoid carboxylic acid methyl ester.

matches with that of one member of the series aromatic C-ring or in the side chain attached at
the C20 compound. However, they do not C-14.
coelute (methyl veadeiroate has a shorter reten- (c) For higher members of the series, >C24, the
tion time), excluding C-13 as a site for the fragmentation is dominated by cleavage of the
CO2 Me. side chain resulting in a peak at m / z 145 (same
(b) The key ions of the series are explained by the fragment that occurs in the monoaromatic
well proven rearrangements of rings A and B tricyclic terpane series).
to produce indanyl cations. They differ by 58 (d) The presence of a fragment at m / z 255 in the
mass units (CO 2Me group) from the key ions mass spectra of the series (loss of the side
of the ring-C monoaromatic tricyclic terpane chain) excludes the possibility of a COzMe
series (Azevedo et al., 1990). This indicates that group attached to the tings.
the CO2Me group must be present in the (e) Comparison with the saturated tricyclic ter-
indanyl cations, attached either directly to the penoid carboxylic acids, which are the main

i/z203145 I c 19 CO2CH3
217
231
24s (I) C3o
t'w 273 ~;t9
287 C26
I c c28

J+71.l.j, , J o i
I i~ e =1. e i4.e i~' e mac e ++e"
?.'e" ~ e a4"'.~e Id.i" e+ e :;,

tR (min)
Fig. 12. Composite fragmentogram (m/z 145, 203, 217, 231, 245, 273, 287, 329, 343, plus 357) for the
aromatic tricyclic terpenoid carboxylic acid methyl esters of Tasmanian tasmanite bitumen.
 Tricyclic terpenoid carboxylic acids 1001

GROUP 2
• CH3
_1'

GROUP 3
nCO2Me ~~~----~C02Me
( M - 15] + m/zS9

Scheme 1. Fragments [M - 15]+ (Group 2) and m/z 69 (Group 3).

cyclic constitutents of the acid fraction and tasmanite bitumen and was characterized as a series
whose CO2Me group was determined to be of 13- methyl-14-carboxy alkylpodocarpa-8,11,13-
attached to the side chain at the C-14 position trienes. The series ranges from C~9 to C39, with a
(see above). maximum at C~9, and is the acidic counterpart of
(f) Other positions usually found to be functional- the recently described series of monoaromatic tri-
ized in natural terpenoid compounds are at cyclic terpanes. The extended tricyclic terpenoid
C-18, C-19 and C-20. Nevertheless, those com- carboxylic acids of this bitumen are related to the
pounds have completely different mass spectra ubiquitous tricyclic terpane series. The main cyclic
(Pinto et al., 1987; Simoneit, 1977; Patitucci, constituents of the monocarboxylic acid fraction are
1983). the C20-C4o homologs of the series of tricyclic ter-
Therefore, structure I (Fig. 12) was tentatively assig penoid carboxylic acids. A pair of stereoisomers
ned for this monoaromatic tricyclic terpenoid car- (flct and a~) was observed for all pseudohomologs,
boxylic acid series based on the previous interpret- C20-C26, except for C2I, which consisted of all
ation. isomers, i.e. tiff, otfl, fl~t and ~tct. The diastereomers
were also resolved for the R/S flct homologs t> C24.
CONCLUSION Normal alkanoic acids ranging from C9 to
A novel series of monoaromatic tricyclic car- C30, with a carbon number maximum at C~6, and
boxylic acids was found in immature Tasmanian Cj4, C~5, C]9 and C20 isoprenoid acids are

GROUP 4

•[• .~nCOaMe +(~

~ ~ C ~
r"l n m/z 189 (n = O)
[M - 111] +

m/z 203 ( n . o)
[M - 97]+

Scheme 2. Ring A/B rearrangements give rise to a pair of fragments derived from m/z 189 and 203, plus
14n (Group 4).
1002 D. A. AZEVEDO et al.
GROUP 5

/% n

O=Me , L O2Me m/z21S(n=0)

• [M - 8 5 ] +

[M - 71] ÷

nCO=M e

- 57]+

[M - 4 3 ] *

Scheme 3. Ring A/B rearrangements give rise to four fragments derived from m,z 215, 229, 243, 257, plus
14n (Group 5).
GROUP6

nCO=Me
J
+~~~nCO~ 4e
+~ m / z 145

nCO=Me

m/z 159

Scheme 4. Fragments due to loss of the side chain give rise to ions m/z 145 and 159 (Group 6).
 Tricyclic terpenoid carboxylic acids 1003

GROUP7

~ CO2Me
..~ nCO2Me

~~~ 255
J ~,- .CH3
J ~,~ CH3
.

~ C02Me
...~ nCO2Me

Scheme 5. Fragments due to loss of the side chain from M +' or [M - 15]+ give rise to ions m/z 255 and
241 (Group 7).

also significant c o m p o n e n t s of the fraction a n d could Chicarelli M. I., Aquino Neto F. R. and Albrecht P. (1988)
be indicative o f the m a r i n e input. Ocurrence of four stereoisomeric tricyclic terpane series in
immature Brazilian shales. Geochim. Cosmochim. Acta 52,
T h e extended tricyclic terpenoid carboxylic acids 1955-1959.
are interpreted to be b i o m a r k e r s of tasmanitids, Cyr T. D. and Strausz O. P. (1983) The structures of tricyclic
which are ubiquitous kerogen macerals in petroleum terpenoid carboxylic acids and their parent alkanes in the
source rocks ( T a p p a n , 1980; Teichmiiller, 1986). Alberta oil sands. J. Chem. Soc., Chem. Commun.
1028-1030.
Thus, such b i o m a r k e r s can c o n t r i b u t e to the overall
Cyr T. D. and Strausz O. P. (1984) Bound carboxylic acids
b i t u m e n mixture a n d m a y prove to be useful as in the Alberta oil sands. Org. Geochem. 7, 127-140.
source indicators for certain m a r i n e organic m a t t e r De Grande S. M. B., Aquino Neto F. R. and Mello M. R.
(De G r a n d e et al., 1991, 1993). (1991) Tandem mass spectrometry analysis of tricyclic
terpanes. Proc. 12th Int. Mass Spectrom. Conf., p. 405.
Amsterdam.
Associate E d i t o r - - K . PETERS De Grande S. M. B., Aquino Neto F. R. and MeUo M. R.
(1993) Extended tricyclic terpanes in sediments and pet-
Acknowledgements--We thank FINEP and CNPq for finan- roleums. Org. Geochem. 20, 1039-1047.
cial support to conduct this research and Dr P. Albrecht and Heissler D. and Ladenburger C. (1988) Synthesis of (+)-
Dr K. E. Peters for their reviews. tricyclohexaprenol, a possible precursor of a family of
tricyclic geoterpanes, and synthesis of an isomer. Tetra-
hedron 44, 2513-2521.
REFERENCES lmamura P. M. and Rfiveda E. A. (1980) The C-13 configur-
ation of the bromine-containing diterpene isoaplysin-20.
Aquino Neto F. R., Trigiiis J., Azevedo D. A., Rodrigues Synthesis of debromoisoaplysin-20 and its C-13 epimer.
R. and Simoneit B. R. T. (1992) Organic geochemistry of J. Org. Chem. 45, 510-515.
geographically unrelated Tasmanites. Org. Geochem. 18, Kaufman T. S., Gonzalez-Sierra M. and Rrveda E. A.
791--808. (1988) Synthesis of an Alberta oil sand bitumen C20
Azevedo D. A., Aquino Neto F. R. and Simoneit B. R. T. tricyclic carboxylic acid beating a novel diterpenoid skel-
(1990) Mass spectrometric characteristics of a novel series eton. J. Chem. Soc. Perkin Trans. I, 2323-2327.
of ring-C monoaromatic tricyclic terpanes found in Tas- Loureiro M. R. B., Aquino Neto F. R. and Cardoso J. N.
martian tasmanite. Org. Mass Spectrom. 25, 475-480. (1986) Anfilise da matrria orgfinica de origem sedimentar.
Azevedo D. A., Aquino Neto F. R. and Simoneit B. R. T. I. Isolamento de ~icidos carboxilicos e hidrocarbonetos
(1991) Double bond location in a series of monounsatu- alif~iticos e aromfiticos. Quim. Nova 9, 15-19.
rated tricyclic terpanes. 12th Int. Mass Spectrom. Conf., McCarthy R. D. and Duthie A. H. (1962) A rapid quanti-
Amsterdam. tative method for the separation of free fatty acids from
Azevedo D. A., Aquino Neto F. R., Simoneit B. R. T. and other lipids. J. Lipid Res. 3, 117-119.
Pinto A. C. (1992) Novel series of tricyclic aromatic Newton E. T. (1875) On "'Tasmanite" and Australian
terpanes characterized in Tasmanian tasmanite. Org. "White Coal". Geol. Mag. 12, 337-342.
Geochem. 18, 9-16. Patitucci M. L. (1983) Constituintes quimicos de Vello:ia
Azevedo D. A., Aquino Neto F. R. and Simoneit B. R. T. leptopetala. M.Sc. Thesis, Universidade Federal do Rio de
(1994) Mass spectrometric characteristics of two novel Janeiro.
series of ring-C monounsaturated tricyclic terpenes found Pinchin R., Mayer L. M. U, and Pinto A. C. (1978)
in Tasmanian tasmanite. Org. Mass Spectrom In press. Veadeirol and veadeiroic acid, two novel diterpenes from
Burlingame A. L., Wszolek P. C. and Simoneit B. R. T. Vellozia flavicans. Phytochemistry 17, 1671-1672.
(1969) The fatty acid content of tasmanites. In Advances Pinto A. C., Zocker D. H. T., Queiroz P. P. S. and Kelecom
in Organic Geochemistry 1968 (Edited by Schenck P. A. A. (1987) Diterpenoids from Vellozia flavicans. Phyto-
and Havenaar I.), pp. 131-155. Vieweg, Braunschweig. chemistry 26, 2409-2411.
1004 D.A. AZEVEDOet al.

Sarjeant A. S. (1989) Personal communication. Simoneit B. R. T., Leif R. N., Aquino Neto F. R., Azevedo
Seifert W. K. (1975) Carboxylic acids in petroleum D. A., Pinto A. C. and Albrecht P. (1990) On the presence
and sediments. In Progress in the Chemistry of Organic of tricyclic terpane hydrocarbons in Permian tasmanites
Natural Products (Edited by Herz W., Grisebach H. algae. Naturwissenschaften 77, 380-383.
and Kisby G. W.), Vol. 32, pp. 2-49. Springer, New Simoneit B. R. T., Schoell M;, Dias R. F. and Aquino Neto
York. F. R. (1993) Unusual carbon isotope compositions of
Simoneit B. R. T. (1977) Diterpenoid compounds and biomarker hydrocarbons in a Permian tasmanite.
other lipids in deep-sea sediments and their geo- Geochim. Cosmochim. Acta 57, 4205-4211.
chemical significance. Geochim. Cosmochim. Acta 41. Tappan H. N. (1980) The Paleobiology of Plant Protists.
463--476. Freeman, San Francisco.
Simoneit B. R. T. and Burlingame A. L. (1973) Carboxylic Teichm/iller M. (1986) Organic petrology of source rocks,
acids derived from Tasmanian tasmanite by extractions history and state of the art. In Advances in Organic
and kerogen oxidations. Geochim. Cosmochim. Acta 37, Geochemistry 1985 (Edited by Leythaeuser D. and Rull-
595-610. k6tter J.), pp. 581--599~ Pergamon, Oxford.