Directory UMM :Data Elmu:jurnal:O:Organic Geochemistry:Vol31.Issue6.2000:

Organic Geochemistry 31 (2000) 497±507
www.elsevier.nl/locate/orggeochem

Petroleum tricyclic terpanes: predicted physicochemical
behavior from molecular mechanics calculations
Kenneth E. Peters
ExxonMobil Upstream Research Company, P.O. Box 2189, Houston, TX 77252-2189, USA
Received 31 July 1999; accepted 9 March 2000
(returned to author for revision 10 December 1999)

Abstract
Petroleum contains a diastereomeric doublet for each of the C25 to C29 tricyclic terpanes due to stereoisomerization at
C-22, where the elution order of the 22S and 22R epimers is unknown. Geometry-optimized molecular mechanics models
for each pair of epimers show similar calculated total energies, indicating similar thermal stability. Similar stability
explains the nearly equivalent size of the 22S and 22R chromatographic peaks for each doublet in nonbiodegraded petroleum. Molecular mechanics MM+ and COMPASS force-®eld calculations indicate an abrupt conformational change
between the C28 and C29 tricyclic terpanes, corresponding to a discontinuity on plots of molecular mass versus log of gas
chromatographic retention time. The second-eluting peak in each C26 to C29 doublet is more readily biodegraded, with
(Alberdi, M., Moldowan, J.M., Peters, K.E., Dahl, J.E., 2000. Stereoselective biodegradation of tricyclic terpanes in
heavy oils from Bolivar Coastal Fields, Venezuela. Submitted to Organic Geochemistry) or without microbial demethylation to form 17-nor-tricyclic terpanes. Factors controlling the chromatographic elution order of epimers are not
fully understood. However, elution order can be inferred if one assumes that epimers with greater calculated surface
areas are more susceptible to microbial attack, as for the extended hopanes where C-22 epimer elution order is known.

Surface areas of 22R epimers exceed 22S for C25 to C29 tricyclic terpanes, suggesting that the 22R epimers elute after
22S. Proof of elution order will require co-injection of authentic standards. Four epimers are possible for each of the
C30 and C31 tricyclic terpanes. Molecular mechanics and high-resolution chromatography suggest that all four peaks
occur in petroleum, but only two are normally observed due to co-elution. Complete resolution of these epimers will
require improved chromatographic methods. # 2000 Elsevier Science Ltd. All rights reserved.
Keywords: Tricyclic terpanes; Cheilanthane; Molecular mechanics; QSAR; Biodegradation

1. Introduction
Tricyclic terpanes or cheilanthanes were ®rst observed
in extracts from the Green River Formation (Anders and
Robinson, 1971; Gallegos, 1971). The most prominent
tricyclic terpanes (14-alkyl, 13-methylpodocarpanes, Fig.
1) range from C19 to at least C54 and are important
components in the saturated hydrocarbon fractions of
petroleum (Moldowan et al., 1983; De Grande et al.,
1993). Tricyclic terpanes are used to correlate crude oils
E-mail address: ken_peters@email.mobil.com

and source-rock extracts, predict source-rock characteristics, and evaluate the extent of thermal maturity and
biodegradation (Seifert et al., 1980; Seifert and Moldowan, 1981; Zumberge, 1987; Peters and Moldowan, 1993).

Extended tricyclic terpanes (>C24) contain a regular
isoprenoid side chain at C-14 (Aquino Neto et al., 1982)
as evidenced by lower abundance of the C22, C27, C32,
C37, and C42 homologs (Moldowan et al., 1983), which
require cleavage of two carbon±carbon bonds to form
from higher homologs (Fig. 1). Based on their results
for the C20 and C21 tricyclic terpanes, Chicarelli et al.,
(1988) infer that higher homologs in thermally mature

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498

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

Fig. 1. Mass chromatogram (m/z 191, solid trace) shows C19±C30 tricyclic terpanes in the saturated hydrocarbon fraction of heavily
biodegraded crude oil from Rubiales Field, Well 12 (2603±2608 feet), Llanos Basin, Colombia. C30 tricyclohexaprenane (inset) is a
widespread higher homolog in petroleun. Doublets eluting after C24 are stereoisomers resulting from the asymmetric center at C-22.
In-reservoir biodegradation resulted in C-10 demethylation of tricyclic terpanes and formation of the corresponding 17-nor-tricyclic

terpanes (m/z 177, dotted trace), where examples are indicated by arrows. Demethylation slightly favors the second-eluting tricyclic
terpane peak, especially for the C26, C28, and C29 homologs (e.g., Alberdi et al., 2000). Samples in Figs. 1 and 2 were treated with
molecular sieves to remove n-parans. C22DM TC=C22 13b,14a(H)-17-nor-tricyclic terpane, C27DM=l7a,21b(H)-25,28,30-trinorhopane, C28DM=17a,21b(H)-25,30-dinorhopane (inferred from retention times of m/z 177 peaks compared to probable precursor
on m/z 191), Ts=18a(H)-22,29,30-trisnorneohopane, Tm=17(H)-22,29,30-trisnorhopane.

petroleum show 13b,14(H)-stereochemistry. The C25 to
C29 tricyclic terpanes occur as diasteromeric doublet
peaks on m/z 191 mass chromatograms, which represent
epimers resulting from stereochemical di€erences at C-22
in the side chain (C25 is poorly resolved in Fig. 1). Splitting of these two peaks into four peaks should occur for
the C30 to C34 homologs because of the asymmetric center at C-27 (however, see below). Under the chromatographic conditions used by Moldowan et al. (1983)
splitting of the ®rst- and second-eluting peaks in each
doublet was not observed until C35 and C38, respectively.
Regular polyisoprenols, such as C30 tricyclohexaprenol
in bacterial membranes or malabaricatrienes from algae
or bacteria, likely account for many tricyclic terpanes in
petroleum (Ourisson et al., 1982; Aquino Neto et al., 1983;
Heissler et al., 1984; Behrens et al., 1999). Higher homologs may originate from C40 tricyclooctaprenol (Azevedo
et al., 1998) or larger precursors. High concentrations of
tricyclic terpanes and their aromatic analogs commonly

correlate with high paleolatitude Tasmanite-rich rocks,
suggesting an origin from these algae (Aquino Neto et
al., 1989, Azevedo et al., 1992; Peters et al., 1997).
However, stable carbon isotopic data suggest that other

algal or bacterial sources are possible (Revill et al.,
1994). Kruge et al. (1990) and De Grande et al. (1993)
note prominent tricyclic terpanes in saline lacustrine and
marine carbonate environments, suggesting to them that
the precursor organisms lived in moderate salinity conditions. However, caution must be exercised in these
interpretations because tricyclic terpanes are thermally
more stable than many other terpanes. Thus, highly
mature petroleum commonly contains abundant tricyclic
terpanes, regardless of source-rock organic matter input
(Peters and Moldowan, 1993).
The purpose of this work is to investigate four unexplained aspects of tricyclic terpane distributions in petroleum (e.g., Figs. 1 and 2) and to promote further research
to better de®ne their occurrence, stereochemistry, and
chromatographic properties.
1. The 22S and 22R epimer peaks in each C25 to C29
13b,14(H)-tricyclic terpane chromatographic

doublet show similar abundance in nonbiodegraded
oils and source-rock extracts (Fig. 2, top), unlike the
22S and 22R peaks for the extended 17,21b(H)hopanes (C31 to C35 hopanes, Peters et al., 1996).

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

499

2. Methods

Fig 2. Mass chromatogram (m/z 191, solid trace) shows C19±
C30 tricyclic terpanes in the saturated hydrocarbon fraction of
nonbiodegraded (top) and heavily biodegraded (bottom) crude
oils from Jusepin Field, Well 110 and Quiriquire Field, Well
449 (unknown depths), Maturin Basin, Venezuela. Relative
peak-heights show that biodegradation favors the second-eluting peak for the C28, C29, and C30 tricyclic terpanes (top versus
bottom) without formation of 17-nor-tricyclic terpanes (DM,
m/z 177, dotted trace). Although four C30 tricyclic terpane
peaks are possible due to the asymmetric carbon at C-27, only
two peaks occur due to co-elution, as discussed in text.

2. Chromatographic elution order for the 22S and 22R
epimers is unknown and plots of homolog number
or mass versus log retention time are non-linear
(Fig. 3).
3. Heavy biodegradation of crude oils in some reservoirs favors C-10 demethylation of the second-eluting epimer in each doublet to form the corresponding
17-nor-tricyclic terpane (e.g., Alberdi et al., 2000).
4. Routine gas chromatography±mass spectrometry
(GC±MS) resolves only two peaks for the C30 and
C31 tricyclic terpanes, whereas asymmetric centers
at C-22 and C-27 allow four peaks for each
homolog (22S27S, 22S27R, 22R27S, 22R27R).

Molecular mechanics calculations were completed on
structural models of the C21 to C31 tricyclic terpanes
using the MM+ force ®eld and default options (bond
dipoles, no cuto€s) in HyperChem 5.1/ChemPlus 2.01
(Professional Version, Molecular Visualization and
Simulation Program Package, Hypercube Inc., Gainesville,
Florida, 1998). Stereochemistry was assumed to be 5(H),

8b(CH3), 9b(H), 10b(CH3), l3b(H), l4b(H) (Aquino Neto
et al., 1982, Chicarelli et al., 1988). Geometry optimization
was completed using the Polak-Ribiere conjugate gradient in vacuo with a termination root mean square
(RMS) gradient of 0.01 kcal/AÊmol.
The conformational search option in HyperChem was
applied to each geometry-optimized structure to
improve the optimized energy (likely local minimum)
and approach the global energy minimum. Calculations
on large molecules, such as hopanes (Peters et al., 1996),
diahopanes (Dasgupta et al., 1995) and tricyclic terpanes, require long computing times. Conformational
search was completed in usage directed mode (Monte
Carlo multiple minimum approach; Chang et al., 1989)
by varying the torsion angles for key dihedral bonds in
the side chain of each compound, leaving the ring system in the conformation established by the stereochemistry and original geometry optimization. Dihedral
angles in the side chain were varied about C-13-21 for
the C21 to C24 tricyclic terpanes, C-13-21 and C-21-25
for the C25 to C28 tricyclic terpanes, and C-13-21, C-2l25, and C-25-29 for the C29 to C31 tricyclic terpanes. In
some cases for epimers A and B, the local energy minimum for epimer A was further reduced by inverting the
stereochemistry of the key asymmetric carbon atom in
the geometry-optimized conformation of epimer B and

repeating the conformational search.
Quantitative structure±activity relationships (QSAR)
were calculated using the minimum-energy structures
determined by conformational search in HyperChem.
QSAR is widely used to predict the physicochemical
behavior of compounds, including molecular volume
and surface area (e.g. Famini and Wilson, 1996 and
references therein). For example, solvent-accessible or
Van der Waals grid surface areas for geometry-optimized tricyclic terpanes were computed in ChemPlus 2.0
using the method of Bodor et al. (1989) and atomic radii
of Gavezotti (1983). The solvent probe radius was 1.4 AÊ.
The method to determine grid surface area requires
more computation time, but is more accurate than other
methods for a given set of atomic radii (Hasel et al.,
1988; Still et al., 1990).
Conformations of the tricyclic terpanes were also
determined by COMPASS force-®eld calculations using
Cerius21 software (Molecular Simulations Inc., San
Diego, California, 1999). COMPASS (Condensed-phase
Optimized Molecular Potentials for Atomistic Simulation

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K.E. Peters / Organic Geochemistry 31 (2000) 497±507

Fig. 3. Molecular mass versus log of chromatographic retention time for tricyclic terpanes in a nonbiodegraded West African oil (30.5
API) shows at least two distinct breaks in the linear trend at C24 and C29. The C24 and C29 tricyclic terpanes contain terminal isopropyl
groups where addition of a methyl group to form the next homolog activates asymmetric carbon atoms at C-22 and C-27, respectively.
Data points for the 22S and 22R diastereomers of the C25±C28 tricyclic terpanes plot in nearly identical locations due to the log retention
time scale. The HP-5 Ultra chromatographic column was slow-programmed to 325 C at 0.1 C/min as described in methods (e.g. Fig. 7).
A HP Ultra 2 column programmed to 325 C at 2 C/min. generates comparable breaks in the linear trend (Table 2).

Studies) is a force ®eld calibrated empirically from ab
initio data to yield structural, conformational, vibrational, and thermophysical properties in good agreement with experimental data (Sun, 1998). As a further
test, the lowest energy C28 and C29 22S and 22R conformers were optimized by the rigorous semi-empirical
AM1 method (Dewar et al., 1985) in Cerius2 to con®rm
the molecular mechanics conformational results.
Saturated hydrocarbon fractions from crude oils were
analyzed for tricyclic terpanes using multiple ion detection-gas chromatography±mass spectrometry (MIDGC±MS). The Hewlett Packard (HP) 5890 Series H gas
chromatograph was equipped with a 50 m HP Ultra 2

crosslinked 5% phenylmethylsilicone column (0.2 mm
I.D., 0.25 mm ®lm thickness) coupled to a HP 5970 mass
spectrometer with helium as carrier gas at 40 psi. After
injection at 270 C, volatile compounds were refocused
at 50 C for 4 min. The oven temperature was increased
to 150 C at 4 C/min, held for 5 min, and then increased
to 325 C at 2 C/min and held for 5 min. The mass
spectrometer was run in electron impact mode (70 eV)
and data were acquired using dwell times of 100 ms/
ion for processing on the HP ChemStation.
For the 6 h slow-programmed high-resolution experiment, a HP 5890 Series II gas chromatograph was coupled to a Finnigan TSQ-70 mass spectrometer used in
MID mode.
The 60 m HP-5 Ultra column (0.2 mm I.D., 0.1 mm
®lm thickness) was heated to 50 C for 4 min and then to
325 C at 0.1 C/min with split mode injection and 50 psi

helium carrier gas pressure. The injector and transfer
line temperatures were 270 C.

3. Results and discussion

Calculated total energies using the MM+ force ®eld
decrease with carbon number for geometry-optimized
conformations of the C25 to C28 tricyclic terpanes, but
increase for the C29 homolog (Fig. 4, Table 1). The 22S
and 22R epimers show similar energies, indicating similar
thermal stability throughout the range from C25 to C29.
For simplicity, Figs. 4 and 5 include data for only the C25
to C29 tricyclic terpanes, although complete data for the
range C21±C31 are in Table 1. The C24 tricyclic terpane
lacks an asymmetric center at C-22 and thus shows no
22S or 22R epimers, while the C30 and C31 tricyclic terpanes have asymmetric centers at both C-22 and C-27,
resulting in four possible epimers (Fig. 1, inset).
The calculated energies explain the similar size of the
®rst- and second-eluting chromatographic peaks for
each tricyclic terpane doublet in the range C25±C29 in
saturate fractions of oils and source-rock extracts (top
Fig. 2, Table 1). Although the C25±C28 22S tricyclic terpanes show slightly higher energies than the corresponding 22R epimers (Fig. 4), these energy di€erences
are less than those for the 22S and 22R epimers of the
extended 17a,2lb(H)-hopanes. The 22S and 22R epimer
energies di€er by less than 0.3 kcal/mol for C25±C29 tricyclic terpanes (Table 1), but range from 0.5 to 2.4 kcal/

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

501

Fig. 4. Relative energies of geometry-optimized C25±C29 22S and 22R tricyclic terpanes from molecular mechanics force-®eld calculations suggest that each 22S shows similar thermal stability to the corresponding 22R epimer. Inset shows abbreviated structure of the
C29 tricyclic terpane, with an asymmetric carbon at C-22, but not at C-27. C-27 becomes asymmetric for the C30 tricyclic terpane (Fig.
1). Table 1 includes data for the C21±C31 tricyclic terpanes.

mol for C31±C35 hopanes (Peters et al., 1996). The larger
energy di€erences for the hopane epimers result in
measured 22S/22R ratios near 1.2 to 1.5 in nonbiodegraded petroleum (Peters et al., 1996), while the
tricyclic terpane epimers generally show 22S/22R ratios
near 1.0 (e.g., Fig. 2, top). Most chromatographic
separations of C25 tricyclic terpanes into 22S and 22R
doublets are incomplete and C27 tricyclic terpanes are
low in crude oils or source-rock extracts (Figs. 1 and 2),
as discussed above.
Using a modi®ed Gibbs free energy equation and
assumptions described in Peters et al. (1996), the theoretical 22S/22R ratios for the C25±C29 tricyclic terpanes
in mature nonbiodegraded petroleum are 1.0, 1.0, 1.0,
0.9, and 0.8, respectively. Nearly all oil samples have
measured tricyclic terpane 22S/22R ratios near 1.0 when
the saturate fraction is treated with molecular sieves to
remove n-parans prior to analysis (Fig. 2, top). Exceptions include certain heavily biodegraded oils (Fig. 2,
bottom), highly mature oils where tricyclic terpanes are
low compared to contaminant peaks (e.g., Fig. 4 in Seifert and Moldowan, 1981), or where they were concentrated and possibly fractionated using an aluminum
oxide column (Fig. 1 in Moldowan et al., 1983).
The isoprene rule for the C29 tricyclic terpane requires
a terminal isopropyl group at C-26, which breaks the
conformational trend established by the lower homologs
from C25 to C28 (Fig. 1, inset). For this reason, the geometry-optimized shapes of the C29 and higher tricyclic
terpanes di€er from the lower homologs (Fig. 6) and they
show di€erent optimized dihedral angles, calculated

energies, volumes, and surface areas than might be
expected from the data for the lower homologs (Figs. 4
and 5, Table 1).
Chromatographic retention time increases discontinuously with mass for the tricyclic terpanes (Table
2, Fig. 3). For example, because the C25±C28 tricyclic
terpanes show a similar conformational style (Fig. 6)
with an asymmetric center at C-22, log retention time
versus mass is highly linear (R2=0.997). However, this
linear trend is broken at C24 and C29, where the terminal
carbons consist of an isopropyl group as discussed
above (Fig. 3, insets). One additional carbon attached to
these isopropyl groups activates either C-22 or C-27 as
an asymmetric center for C25 and C29 tricyclic terpanes,
respectively, which initiates a new conformational style
for higher homologs. For example, the transition from
C28 to C29 tricyclic terpanes results in a more compact
molecular shape. For geometry-optimized C28 tricyclic
terpanes, the side-chain is directed away from the ring
system, while the side-chain for the C29 tricyclic terpanes
curves back toward the ring system in a boomerang
shape (Fig. 6). The boomerang shape allows greater
mass without a corresponding increase in retention time
(di€erence between solid and dashed line above C28 in
Fig. 3).
The abrupt change in conformational style between
the C28 and the C29 tricyclic terpanes is surprising
because the side-chain simply increases from 9 to 10
carbons (Fig. 6). To test whether this remarkable conformational shift as determined by molecular mechanics
(MM+) is reproducible, more rigorous calculations

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K.E. Peters / Organic Geochemistry 31 (2000) 497±507

Table 1
Properties of geometry-optimized tricyclic terpanes from molecular mechanics and QSAR
Energyb

Volumec

Aread

Torsion Angle ( )

Carbona

C-22a

C-27a

Mass

(kcal/mol)

(AÊ3)

(AÊ2)

13±21

21±25

25ÿ29

21
22
23
24
25
25
26
26
27
27
28
28
29
29
30
30
30
30
31
31
31
31

m
m
m
m
R
S
R
S
R
S
R
S
R
S
R
S
R
S
R
S
R
S

m
m
m
m
m
m
m
m
m
m
m
m
m
m
S
S
R
R
S
S
R
R

290.53
304.56
318.59
332.61
346.64
346.64
360.67
360.67
374.69
374.69
388.72
388.72
402.75
402.75
416.77
416.77
416.77
416.77
430.80
430.80
430.80
430.80

ÿ49.35
ÿ49.81
ÿ50.36
ÿ51.56
ÿ53.41
ÿ53.38
ÿ54.06
ÿ54.03
ÿ54.65
ÿ54.61
ÿ55.27
ÿ55.23
ÿ55.05
ÿ54.82
ÿ56.33
ÿ56.09
ÿ56.59
ÿ56.36
ÿ57.64
ÿ57.70
ÿ57.47
ÿ57.52

939.47
991.97
1045.98
1086.41
1136.43
1136.38
1191.79
1190.12
1245.56
1245.89
1300.28
1300.09
1304.76
1298.58
1335.46
1332.09
1334.19
1331.28
1360.36
1363.32
1365.72
1367.11

516.06
544.06
577.47
594.73
619.29
611.64
648.09
644.22
671.26
671.26
706.37
701.28
680.67
674.57
685.01
677.63
685.12
680.40
692.09
693.00
696.68
695.93

ÿ98.93
ÿ99.64
ÿ99.72
ÿ99.64
ÿ100.07
ÿ99.18
ÿ99.95
ÿ99.18
ÿ99.11
ÿ99.11
ÿ100.29
ÿ98.97
ÿ101.90
ÿ101.28
ÿ101.14
ÿ100.09
ÿ101.14
ÿ100.67
51.13
50.67
51.18
51.07

M
M
M
M
ÿ64.44
64.05
ÿ64.85
64.02
ÿ64.65
64.39
ÿ64.97
63.83
53.68
53.74
53.01
52.97
53.14
53.81
ÿ53.90
ÿ52.93
ÿ53.29
ÿ52.77

M
M
M
M
M
M
M
M
M
M
M
M
ÿ65.20
ÿ64.83
ÿ65.28
ÿ65.26
ÿ65.50
ÿ65.20
64.74
65.46
65.71
66.70

a
Tricyclic terpane carbon number is followed by stereochemistry at C-22 and C-27, e.g. 31SR=C31 13b, 14(H)-tricyclic terpane
22S27R; M=missing.
b
Starting structures based on molecular mechanics energy minimization were subjected to a Monte Carlo conformational-search
routine in torsional coordinates to determine total energy as described in the text.
c
Molecular volume and grid surface area determined using quantitative structure±activity relationships (QSAR).
d
Only two peaks each are resolved for the C30 and C31 tricyclic terpanes using our chromatographic conditions.

were completed using the COMPASS force ®eld and
Cerius2 software (see methods). The conformational
shapes for the C24±C29 tricyclic terpanes determined
using the MM+ and COMPASS force ®elds are essentially the same, including the abrupt conformational
change from C28 to C29 (Fig. 6). Table 3 compares the
global-energy minima for the C28 and C29 22S tricyclic
terpanes with the energies of the nine nearest localenergy minimum conformations determined using the
COMPASS force ®eld. Di€erences in absolute energies
of conformers calculated by di€erent methods (Tables 1
and 3) are common and result from the methods of calculation. The critical values for interpretation are the
relative di€erences in energies between conformations
calculated by the same method. Table 3 shows that the
di€erences in energy between the global energy minimum and the next higher energy conformation are
about 0.04 and 1.34 kcal/mol for the C28 and C29 22S
tricyclic terpanes, respectively. The small energy di€erence for the two lowest energy C28 22S conformers is
consistent with a ¯exible side chain directed away from
the ring system. The energy di€erence between the two

lowest energy C29 22S conformers is signi®cant and
appears to be caused by increased intramolecular Van
der Waals interaction between the side chain and ring
system for the lowest energy conformer. This conclusion
is based on calculated intramolecular hydrogen±hydrogen distances in the range of 2.2±2.9 AÊ, which are similar
to intermolecular hydrogen±hydrogen distances among
hydrocarbons.
The lowest energy C28S and C29S conformers were also
optimized using the semi-empirical (AM1) method,
resulting in the same stable conformations. Semi-empirical calculations account for electronic interactions and
yield a more rigorous solution to conformation than
molecular mechanics. The calculated total energies for
22S and 22R epimers were similar for each tricyclic terpane homolog, supporting the results from the MM+
force-®eld calculations.
The stereochemistry of the C30±C32 tricyclic terpanes
is more complex than lower homologs due to an asymmetric center at C-27, which results in 22S27S, 22R27R,
22S27R, and 22R22S con®gurations (Table 1). However, petroleum with high tricyclic terpanes commonly

503

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

Fig. 6. The C25±C28 tricyclic terpanes show similar conformational styles based on molecular mechanics MM+ force-®eld
calculations (only the C25 and C28 22S and 22R geometryoptimized conformations are shown). An abrupt change in
confomational style occurs from the C28±C29 and higher tricyclic terpanes, resulting in a more compact molecular shape.
These MM+ force-®eld conformations are the same as those
determined by COMPASS force-®eld and semi-empirical
(AM1) calculations as discussed in the text.
Fig. 5. Molecular volumes and surface areas of geometry-optimized tricyclic terpanes from quantitative structure±activity
relationships (QSAR, bottom) increase from C25±C28, but the
trend is broken at C29, consistent with a change in conformational style (Fig. 6). The 22R epimers for C25±C29 homologs
show greater volumes and surface areas than the corresponding
22S epimers. Biodegradation of hopanes by C-10 demethylation favors epimers with greater surface areas (arrows at top,
Peters et al., 1996). The 22S hopane epimers elute prior to the
corresponding 22R epimers (Armanios, 1995). Assuming that
biodegradation also favors tricyclic terpane epimers with
greater surface areas, the C26 and C29 22R epimers will be preferentially biodegraded (bottom). Because the second-eluting
peak in each the C26±C29 tricyclic terpane doublet undergoes
preferential biodegradation (Figs. 1 and 2), it is inferred be the
22R epimer.

shows only two of the four expected chromatographic
peaks for each of the C30 and C31 tricyclic terpanes (e.g.
C30 in Fig. 1). These peaks are small and dicult to
identify on the m/z 191 mass chromatograms of many
petroleum samples. As discussed above, the C32 tricyclic
terpanes are low in all samples because they require
cleavage of two carbon±carbon bonds to form from
higher homologs.
Molecular modeling shows that the four epimers for
each of the C30 tricyclic terpanes have similar energies
(Table 1) and are thus likely to occur in similar abundance with no relatively high-energy, unfavorable con®gurations. This suggests that co-elution rather than
stability di€erences accounts for two rather than four

Table 2
Masses and chromatographic retention times of tricyclic terpane homologs in a nonbiodegraded oil from West Africa
Carbon

Mass
(amu)

RT(1)a
(min)

21
22
23
24
25
25
26
26
27
27
28
28
29
29
30
30
31
31
31
31

290.53
304.56
318.59
332.61
346.64
346.64
360.67
360.67
374.69
374.69
388.72
388.72
402.75
402.75
416.77
416.77
430.80
430.80
430.80
430.80

53.23
56.99
61.33
63.60
68.16
68.28
71.57
71.83
75.64
75.90
79.00
79.49
81.07
81.65
85.03
85.70
87.97
88.70
88.85
89.49

logRT(1)

RT(2)a
(min)

LogRT(2)

1.726
1.756
1.788
1.803
1.834
1.834
1.855
1.856
1.879
1.880
1.898
1.900
1.909
1.912
1.933
1.930
1.944
1.948
1.949
1.952

230.75
244.60
261.92
271.12
289.42
289.54
302.66
303.63
317.88
318.85
332.07
333.93
340.22
342.57
355.74
358.36
367.37
367.48
370.31
370.31

2.363
2.388
2.418
2.433
2.462
2.462
2.481
2.482
2.502
2.504
2.521
2.524
2.532
2.535
2.551
2.554
2.565
2.565
2.569
2.569

a
RT(1)=retention time using 50-m HP Ultra 2 column and
2 C/min heating rate, RT(2) = retention time using 60-m HP-5
Ultra column and 0.1 C heating rate (see Methods).

504

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

Table 3
COMPASS force-®eld conformer search results for ten lowest
energy conformations of the C28 and C29 22S tricyclic terpanes

Conformer

C28 22S
energy
(kcal/mol)

C29 22S
energy
(kcal/mol)

1
2
3
4
5
6
7
8
9
10

ÿ67.21
ÿ67.17
ÿ67.11
ÿ66.75
ÿ66.28
ÿ65.05
ÿ64.92
ÿ64.59
ÿ64.59
ÿ63.61

ÿ73.18
ÿ71.84
ÿ71.16
ÿ69.50
ÿ67.41
ÿ67.41
ÿ67.33
ÿ67.33
ÿ67.25
ÿ66.73

chromatographic peaks, where each peak represents two
epimers. Slow-ramp temperature-programmed chromatography supports these modeling results. A 6 h highresolution chromatography experiment failed to resolve
the two C30 tricyclic terpane peaks. However, the ®rst of
the two C31 tricyclic terpane peaks to elute shows
broadening, reduced peak height, and initial separation
into two poorly resolved components (Fig. 7). Routine

GC±MS on the same sample yields two C30 tricyclic
terpane peaks of approximately equal size. Chiral-phase
chromatography (Schurig, 1994; Tang et al., 1994) using
a 10 m CP Chirasil-Dex CB fused silica column did not
improve resolution of the C30 and C31 tricyclic terpanes
because the bonded phase became unstable at the
required temperatures.
Geometry-optimized C25±C28 tricyclic terpane epimers show systematic increases in molecular volume and
surface area based on QSAR (Fig. 5, bottom). However,
the C29 epimers do not conform to this trend. For
example, calculated surface areas for the C29 epimers are
less than the corresponding C28 epimers due to the
change in conformational style discussed above (Fig. 6).
Despite having identical masses, each 22R tricyclic terpane epimer has greater surface area than 22S in the
range from C25 to C29.
QSAR properties are widely used to predict the physicochemical behavior of compounds. For example, chromatographic elution times for di€erent compounds can
be described using various QSAR parameters, including
molecular volume, polarizability, covalent basicity or
acidity, and electrostatic basicity or acidity (Famini and
Wilson, 1996 and references therein). However, the
author is not aware of publications that use QSAR to
predict elution times for epimers. Correlation of absolute con®guration with elution order among epimers in

Fig. 7. Slow-programmed high-resolution chromatography of a nonbiodegraded West African oil (same as in Fig. 3) resolves two peaks
for C30, but suggests at least three peaks for C31 tricyclic terpanes. The ¯rst-eluting C31 peak is broader and shorter than that from routine
GC±MS (where it is slightly taller than the second peak) and initial separation of two co-eluting peaks is evident. Tricyclic terpanes were
enriched by high-performance liquid chromatography to minimize hopanes. Geometry-optimized models show that the four epimers for
each of the C30 and C31 tricyclic terpane epimers have similar energies, suggesting similar stability and abundance (Table 1).

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

homologous series of compounds is problematic
because many phenomena can a€ect their separation,
including hydrogen bonding, dipole-dipole interactions,
and other forces (Schurig, 1994). Unequivocal proof of
absolute con®guration requires co-injection of authentic
standards with known stereochemistry.
The extended hopanes may serve as analogs to predict
the elution order of the tricyclic terpane epimers. Like the
tricyclic terpanes, the C31±C35 hopanes are characterized
by 22S and 22R epimers (Fig. 5, top). However, unlike
the tricyclic terpanes where elution order of the epimers
is unknown, the 22S epimer for each hopane homolog
elutes prior to 22R, as con®rmed by co-elution experiments (Armanios, 1995). Hopane homologs of increasing
mass, volume, and surface area elute later during gas
chromatography. However, elution orders of hopane
epimers do not depend directly on mass (epimers have
the same mass), volume, or area, but rather on shape.
The hopane 22S epimers show a ``scorpion'' conformation and elute prior to their corresponding 22R epimers,
which show a ``rail'' conformation (Peters et al., 1996).
Data for the extended hopanes suggest that epimers
with larger volumes and surface areas are more susceptible to microbial attack. For each 22S and 22R hopane
doublet in the range C31±C35, the epimer with greater
volume and surface area is preferentially demethylated
at C-l0 to form 25-norhopanes during biodegradation
(Fig. 5, top, Peters et al., 1996). The volumes and surface areas of the 22S hopanes exceed the 22R epimers
for the C31 and C32 homologs, but are less than the C34
and C35 22R epimers. The 22S epimers of the C31 and
C32 hopanes show preferential conversion to 25-norhopanes compared to 22R, while the opposite applies to
the C34 and C35 epimers. The C33 hopane 22S and 22R
epimers show similar susceptibilities to biodegradation.
A study of Venezuelan crude oils shows that the second-eluting tricyclic terpane peak for each homolog in
the range C26±C29 undergoes preferential demethylation
at C-10 during biodegradation in some reservoirs
(Alberdi et al., 2000). The microbial demethylation of
tricyclic terpanes at C-10 to form 17-nor-tricyclic terpanes is analogous to C-10 demethylation of hopanes to
form 25-norhopanes in petroleum reservoirs (Peters et
al., 1996). Fig. 2 shows that the second-eluting tricyclic
terpane peak is also preferentially attacked in heavily
biodegraded oils that lack C-10 demethylation.
The QSAR results show that the C25±C29 22R tricyclic terpanes have larger surface areas and similar or
larger volumes than the corresponding 22S epimers. If
the analogy with the hopanes is correct, the 22R tricyclic terpane epimers are predicted to be preferentially
biodegraded compared to the corresponding 22S epimers. Because GC±MS shows that the second-eluting
peak for these compounds is preferentially biodegraded
(Figs. 1 and 2), it is inferred that 22S elutes prior to 22R
for the C25±C29 tricyclic terpanes.

505

4. Conclusions
Molecular mechanics and observations of tricyclic
terpanes in petroleum using routine and slow-programmed, high-resolution chromatography are used to
conclude the following:
1. 22S/22R epimer ratios for the C25±C29 tricyclic
terpanes approach 1.0 for nonbiodegraded petroleum because total energies of the favored conformations are similar (Fig. 4).
2. The second-eluting peak for each of the C26±C29
tricyclic terpanes is more readily biodegraded,
with or without microbial demethylation to form
l7-nor-tricyclic terpanes (Figs. 1 and 2).
3. A change in the preferred conformational style
occurs between geometry-optimized C28 and C29
tricyclic terpanes due to the branch point at C-27.
This change in conformational style results in discontinuous linear trends on plots of tricyclic terpane
mass versus log retention time (Fig. 3).
4. All four possible epimers for each of the C30 and
C31 tricyclic terpanes show optimized conformations with similar energies, suggesting that they
occur in similar abundance, but only two peaks
are observed for each by routine GC±MS. A 6
hour high-resolution chromatography experiment
indicates that at least one of the two peaks in the
C31 doublet consists of two co-eluting epimers
(Fig. 7). Improved chromatographic methods will
be required to separate these peaks.
5. 22S epimers are inferred to elute prior to 22R for
C25±C29 tricyclic terpanes. This prediction is based
on (1) the observation that the second-eluting peak
in each tricyclic terpane doublet undergoes preferential biodegradation and (2) the assumption
that epimers with larger exposed surface areas are
more readily biodegraded, as supported by analogy
with the 17(H)-hopanes (Fig. 5).
It is hoped that this study will stimulate further
research on tricyclic terpanes, especially in regard to
items 4 and 5 above. Proof of the chromatographic elution order for tricyclic terpane 22S and 22R epimers will
require co-elution experiments using synthesized
authentic standards. However, the predicted elution
order for tricyclic terpane epimers is consistent with
data for the hopanes, which contain an analogous
asymmetric center at C-22.
Acknowledgements
The author thanks Rajiv Bendale, Cli€ord Walters, J.
Michael Moldowan, Margarita Alberdi, James Kubicki,
and Jim Stinnett for technical discussions and ExxonMobil Upstream Research Company for permission to

506

K.E. Peters / Organic Geochemistry 31 (2000) 497±507

publish this work. John Zumberge (Geomark Research,
Inc.) kindly provided the chromatograms for Figs. 1 and
2. Robert Carlson, Francisco de Aquino Neto, Kirk
Schmitt, and Simon George provided useful review comments. Special thanks are due Yitian Xiao and John
Longo (ExxonMobil Upstream Research Company) for
assistance with the COMPASS force-®eld and semiempirical AM1 calculations and related discussions.
Associate EditorÐS. George

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