Appli cat i on N ote 3 0 2 6 3

Molecular Characterisation of Petroleum

M. Ahmed,1 S. Gong,1 D. Fuentes,1 S. Sestak,1 F. Theobald,2 and K. D’Silva3;
1
CSIRO Earth Science and Resource Engineering, North Ryde, Australia
2
Environmental Consulting, Cologne, Germany
3
Thermo Fisher Scientific, Bremen, Germany

Key Words
Analysis of aliphatic, aromatic and branched/cyclic
hydrocarbon fractions, Thermo Scientific DFS
High Resolution Mass Spectrometer

Introduction Located in Sydney Australia, CSIRO Organic

Comprehensive molecular characterisation of petroleum Geochemistry Team have conducted genetic
is used to solve issues related to exploration and characterisation of oils, fluid inclusion oils, natural gases
production of oil and gas. It can help identify processes and potential source rock extracts for various research
that occurred in reservoirs and along migration pathways. projects on behalf of international energy companies
It provides the ability to understand the key geological including Chevron, PETRONAS, Woodside, Oil Search,
information encoded in the geochemistry of gas and Exxon-Mobil, Petrobras, Interoil, Origin Energy, Roc Oil,
liquid hydrocarbons and can help oil companies deduce and BP Exploration & Production Inc.
the origin and maturity of petroleum hydrocarbons
CSIRO Organic Geochemistry Team currently use a
(natural gas, coal seam gas or oils). Geochemical fossils
Thermo Scientific™ DFS™ high resolution GC-MS
or biological markers, popularly called biomarkers,
for the molecular analyses of sedimentary organic
frequently convey genetic information about the types
matter, e.g., crude oils, fluid inclusion oils, condensates,
of organisms contributing to the organic matter of
potential source rock extracts, solid bitumen, pyrolyzates
sediments. They are used for correlations (oil-oil and
of ashpaltene/kerogen, etc. The DFS instrument provides
oil-source rock), for reconstitution of depositional
a range of capabilities required for this application,
environments, and also as indicators for diagenesis
including; high resolution, full scan, selected ion
and catagenesis.1
monitoring (also known as multiple ion detection, MID)
CSIRO Organic Geochemistry Team is a world leading and multiple reaction monitoring (MRM) with stable and
provider of research services to the petroleum industries predictable metastable product ion formation. All of these
and state and federal agencies. capabilities are combined by CSIRO for comprehensive
analyses of petroleum hydrocarbons.
2 This application note presents a brief
Mass Spectrometer Data Acquisition Methods
description of the methods and
chromatographic conditions, using Full scan of analysis of aliphatic and m/z 50-550, scan rate 0.5 sdec-1.
aromatic hydrocarbons GC program A.
the DFS high resolution GC-MS for
the analyses of: Multiple ion detection (MID) analysis m/z 128, 134, 142, 154, 156, 166,
of aromatic hydrocarbon fraction, 168, 170, 178, 183. GC program A.
• Aliphatic and aromatic hydrocarbon Method-A
fractions of the North Sea Oil-1, a
Multiple ion detection (MID) analysis m/z 178, 180, 182, 183, 184.03,
geochemical standard sample from of aromatic hydrocarbon fraction, 184.12, 192, 197, 198.05, 198.14,
the Norwegian Petroleum Directorate Method-B 202, 206, 212, 216, 220, 234.
• Branched /cyclic hydrocarbon fraction GC program A.
of a standard mixture of oils called Multiple ion detection (MID) analysis m/z 177, 183, 191, 205, 217,218,
AGSO STD obtained from Geoscience of aliphatic hydrocarbon or its 231.11, 231.21,253, 259. GC program B.
Australia (formerly, the Australian branched chain and cyclic fraction
Geological Survey Organization) Multiple Reaction Monitoring (MRM) m/z 370>191, 384>191, 398>191,
Results of these analyses for the identification/ analysis for hopanes 412>191, 426>191, 440>191, 454>191,
quantification of some selected hydrocarbon 468>191, 482>191. GC program B.
biomarkers are presented in the following Multiple Reaction Monitoring m/z 358>217, 372>217, 386>217,
section. (MRM) analysis for steranes 400>217, 414>217, 414>231.
GC program B.
Methods

Gas Chromatographic Methods Results

GC Thermo Scientific Trace Ultra GC
™ Crude oils and source rock extracts are complex mixtures
of many classes of compounds including source and
Sampler Thermo Scientific TriPlus XT™
maturity related biomarkers such as n-alkanes,
Carrier gas Helium isoprenoids, terpanes, steranes and diasteranes. Some
Carrier gas flow rate Constant (1.5 mL/min) biomarkers are often present in very low concentrations
Injector Split/Splitless requiring different modes of GC-MS analyses for their
identification and/or quantification; full scan, multiple ion
Injection temperature 260°C
detection and multiple reaction monitoring (MRM).
Injection volume 1 µL
Full scan analyses provide mass spectra for structural
Sample concentration 100 - 500 µg/mL in dichloromethane
elucidation and chromatograms of all ions. Multiple ion
GC column equivalent TraceGOLD™ TG-1MS 60 m × 0.25 mm, detection (MID) methods acquire only selected ions, e.g.,
0.25 µm film P/N 26099-1540 and m/z 123, 191, 217, etc. and provide results of better
TraceGOLD TG-5MS 60 m × 0.25 mm, sensitivity than full scan, providing information about
0.25 µm film P/N 26098-1540 trace components. This method can be used to determine
Temperature programs A) 40 °C ( 2 min) @ 4 °C min-1 to 310 °C the fingerprints for the selected compound types (e.g.,
(40 min) bicyclic sesquiterpanes, hopanes, steranes and diasteranes).
B) 40 °C (2 min) @ 20 °C min-1 to 200 °C Multiple reaction monitoring analyses monitor a selected
(0 min) 2 °C min-1 to 310 °C (40 min) daughter ion (e.g., m/z 217) formed from molecular ions
that decompose in the first field-free region of a double-
Mass Spectrometer Parameters focussing mass spectrometer. This method provides the
necessary selectivity required to analyze these specific
MS Thermo Scientific DFS
compounds in such a complex matrix; resulting in better
Electron energy 70 eV sensitivity and signal-to-noise ratios.
Resolution 1000 at 5% peak height
Injection temperature 260 °C
Transfer line temperature 280 °C
Source temperature 280 °C
Full Scan Analyses 3

RT: 0.00 - 99.01

Contaminant NL:2.25E9
n ‐C 14
26 TIC MS
[a] 32.87
NSO1_ALI_01A_M
24 29.49 36.06 39.08
39 08 Squalane (IS)
22 25.91

20 47.25
22.11 63.47
18
Relative Abundance

49.72
16
14 52.08
18.14
12
Pr 67.90
10 69.56
8 n ‐C 8 14.10 Ph n ‐C37
73.26
6 75.50
81.22 84.89
4 8.08 89.29
2 7.98
0 14
0.14
0
0 10 20 30 40 50 60 70 80 90
Time (min)

RT: 0.00 - 99.01 n ‐C 14 NL:7.92E7

32.87 m/z= 84.60- 85.60 MS
100
[b] 36.06 NSO1_ALI _01A_M
90 29.49
39.08 41.95
80
25.91
70
47.25
bbundance

60 22.11 Squalane (IS)

49.72
50
52.08
63.47
Relative A

40 n ‐C 8 18.14 58.59
Pr
Ph
30 14.10
64.38
20 10.34 67.89
n ‐C 37
69.56
10 73 .26
26 75.51
9 44
9.44
7.99 81.21 89.28
0
0 10 20 30 40 50 60 70 80 90
Time (min)
Figure 1. Full scan (a) Total ion chromatogram and (b) m/z 85 extracted ion chromatogram of the aliphatic hydrocarbon fraction showing the distributions of n-alkanes
and isoprenoids. Pr = Pristane, Ph = Phytane.
4 MID Analyses

RT: 20.00 - 82.00 SM: 5G

C30 αβ
100

90

80 C30 βα
Relative Abundance
70
Gammacerane
60 C30*
C31 βα
50 C31
αβ
C29 αβ S
40 C29*
C29Ts
30 20/3 R C αβ
S 32
26/3 Ts R S C33 αβ
20 Tm C34 αβ
19/3 R C35 αβ
21/3 23/3 S S
10 24/3 25/3 24/4 R R
0
20 25 30 35 40 45 50 55 60 65 70 75 80
Time (min)
Figure 2. MID mass chromatogram m/z 191 of the aliphatic hydrocarbon fraction showing the distribution of terpanes. Peak assignments define stereochemistry at
C-22 (S and R); αβ and βα denote 17α(H), 21β(H) -hopane and 17β(H), 21α(H)-moretane, respectively. 19/3 – 26/3 = C19 – C26 tricyclic terpanes, 24/4 = C24
tetracyclic terpane, Ts = C27 18α(H), 22,29,30-trisnorneohopane, Tm = C27 17α(H), 22,29,30-trisnorhopane.

RT: 44.00 - 64.00 SM: 5G

100 C 27αββ 20R + C29 βα 20S NL: 5.92E4
C 27 βα 20S m/z = 216.95 - 217.45 MS
C 27αββ 20S + C28 αβ 20R* NSO1_110
90
C 27ααα 20S 923_ALI_D

80

C 28 αβ 20S C 29ααα 20S

70 C 27ααα 20R C 29αββ 20R
C 28αββ 20R
Relative Abundance

C 27 βα 20R
C 29 βα 20R C 29αββ 20S
60 C 28 βα 20S*
C 28ααα 20S* C 29ααα 20R
C 28αββ 20S
50 C 27 αβ 20R
C 28βα20R* C 30ααα 20S
C 29 αβ
40
C 27 20S
C 28 C 30αββ 20S
αβ 20S ααα C 30αββ 20R
30 20R
C 30ααα 20R
20

10

44 46 48 50 52 54 56 58 60 62 64
Time (min)
Figure 3. MID mass chromatogram m/z 217 of the aliphatic hydrocarbon fraction showing C27–C29 steranes and diasteranes. Peak assignments define
stereochemistry at C-20 (S and R); βα, αβ, ααα and αββ denote 13β(H),17α(H)-diasteranes, 13α(H),17β(H)-diasteranes, 5α(H),14α(H),17α(H)-steranes and
5α(H),14β(H),17β(H)-steranes, respectively. * = isomeric peaks (24S and 24R).
 RT: 36.8 - 39.8 SM:5G RT: 47.00 - 49.00 SM:5G 5
100 100 9‐MP
[a] 1,3,6‐TMN NL: 1.41E6
[b]
NL: 1.54E6
m/z = 169.61- 170.61 MS m/z = 191.59 - 192.59 MS
90 1,3,5‐ + 1,4,6‐TMN NSO1_ARO _24JUL_B 90 nso1_aro _24jul_C
1,3,7‐ 1‐MP
TMN 2,3,6‐TMN
80 80
Relative Abundance

Relative Abundance
1,2,7‐TMN 2‐MP
70 70
3 MP
3‐MP
60 1,6,7‐TMN 60
50 50
1,2,5‐TMN
40 40
1,2,6‐TMN
30 30
20 1,2,4‐ 20
TMN
10 1,2,3‐TMN 10
0 0
37.0 37.5 38.0 38.5 39.0 39.5 47.0 47.5 48.0 48.5 49.0
Time (min) Time (min)
RT: 49.50 - 52.50 SM:5G RT: 52.00 - 60.00 SM:5G
100 1,3‐ + 3,9‐ + 3,10‐ + 2,10‐DMP
[c] NL: 9.86E5 100 [d] Retene NL: 9.86E5
m/z = 205.61- 206.61 MS m/z = 205.61- 206.61 MS
90 NSO1_aro _24jul_c 90 NSO1_aro _24jul_c

80 1,6‐ + 2,9‐ + 2,5‐DMP 80

70 70
Relative Abundance

Relative Abundance
60 2,7‐DMP
60
1,7‐DMP
50 3,5‐
, + 2,6‐DMP
, 50
40 2,3‐ + 1,9‐ + 4,9‐ + 4,10 -DMP 40
30 9‐ + 2‐ + 1‐EP + 30
3,6‐DMP
20 20
1,8‐DMP
10 3‐EP 1,2‐DMP 10
0 0
49.5 50.0 50.5 51.0 51.5 52.0 52.5 52 53 54 55 56 57 58 59 60
Time (min) Time (min)

Figure 4. (a) m/z 170.11, (b) m/z 192.09, (c) m/z 206.11 and (d) m/z 234.14 mass chromatograms of the aromatic hydrocarbon fraction showing the distributions
of C3-alkylnaphthalenes, methylphenanthrenes, C2-alkylphenanthrenes and retene, respectively. TMN = Trimethylnaphthalene, MP= Methylphenanthrene,
EP = Ethylphenanthrene, DMP = Dimethylphenanthrene.
6 MRM Analyses
RT: 50.00 - 80.00 SM: 5G RT: 58.59 - 88.59 SM: 5G

100 100 S
NL: 1.53E4
Ts C 31αβ
NL: 1.90E4

Relative Abundance
Bicadinane T   m/z 370 →191 m/z 426 →191 m/z= 189.70-192.70 F:
Tm
m/z= 189.70-192.70 F: 80 R + c EI SRM ms2
80 C27 Hopanes C31 Hopanes

Relative Abundance
+ c EI SRM ms2 [email protected]
[email protected]
[189.70-192.70] MS
60 [189.70-192.70] MS
60 AGSO_25JUL_HOPS AGSO_25JUL_HOPS
40
Bicadinane T1
40 20 C 31*
Bicadinane R
20 C 27 β 0
100 S NL: 8.87E3
C 32 αβ

Relative Abundance Relative Abundance

TIC F: + c EI SRM ms2
0 80 R m/z 440 →191 [email protected]
100 XT NL: 1.08E4
C32 Hopanes
[189.70-192.70] MS
m/z 384 →191 m/z = 189.70-192.70 F: 60 AGSO_25JUL_HOPS
Relative Abundance

25,30-BNH
+ c EI SRM ms2
80
29,30-BNH
C28 Hopanes [email protected] 40

28,30-BNH
[189.70-192.70] MS C 32 *
60 AGSO_ 25JUL _HOPS 20
0
TNH

40 100 NL: 4.49E3

S m/z 454 →191 m/z= 189.70-192.70 F:
20 80 C 33 αβ + c EI SRM ms2
[email protected]
R C33 Hopanes [189.70-192.70] MS
0 60 AGSO_25JUL_HOPS

100 C 29 αβ NL: 4.32E4 40 C 33 *

m/z = 189.70-192.70 F:
m/z 398 →191 200
Relative Abundance

80 + c EI SRM ms2
C29 Hopanes .
398 40@cid0 00
0
[189.70-192.70] MS
60 AGSO_25JUL_HOPS 100 S NL: 2.08E3
C 29 Ts

Relative Abundance Relative Abundance

C 34 αβ
m/z= 189.70-192.70 F:
40 80 + c EI SRM ms2
[email protected]
C 29 * C 29 25 ‐nor R m/z 468 →191
60 [189.70-192.70] MS
20 XT C34 Hopanes AGSO_25JUL_HOPS
C29 βα 40
0
20
100 C 30 αβ m/z 412 →191 NL: 3.97E4
0
Relative Abundance

m/z = 189.70-192.70 F:
80 C30 Hopanes + c EI SRM ms2 100 NL: 1.69E3
S
C 35 αβ
[email protected] m/z= 189.70-192.70 F:
Oleanane±Lupane [189.70-192.70] MS 80 + c EI SRM ms2
60 AGSO_25JUL_HOPS m/z 482 →191 [email protected]
60 R [189.70-192.70] MS
40 C 30* C 30 30 ‐nor C35 Hopanes AGSO_25JUL_HOPS
Gammacerane 40
20 20
0 0
50 55 60 65 70 75 80 60 65 70 75 80 85
Time (min) Time (min)
Figure 5. MRM chromatograms showing m/z M >191 metastable transitions of C27, C28, C29, C30, C31, C32, C33, C34 and C35 hopanes.
+

XT = cross talk, BNH = Bisnorhopane, TNH = 17 α(H), 18 α(H), 21 β(H)-trisnorhopane.

RT: 42.00 - 58.00 SM: 5G

100 C 27 βα 20S m/z 372 → 217 NL: 1.15E4
C 27ααα 20S
C 27 βα 20R C 27 Steranes and diasteranes
 
Relative Abundance

80 C 27αββ 20R m/z= 215.70-232.70 F: + c

C 27αβ 20 S C 27αββ 20S EI SRM ms2

60 C 27 αβ 20 R [email protected]

40
C 27ααα 20R XT [215.70-218.70] MS
_AGSO 06AUG12
_ STER
20 XT

0
C 28 βα 20S* C αββ20R
100 C 28 C 28ααα 20S* 28 NL: 4.02E3
C 28αββ 20S
βα 20R*
Relative Abundance

m/z= 215.70-232.70 F: + c
80
C 28ααα 20R
C 28αβ 20 S
EI SRM ms2
60 m/z 386 →217 [email protected]
C 28αβ 20R* XT
C Steranes and .[215 70 218 70] MS
40 XT 28
AGSO_06AUG12_STER
diasteranes
20
0
100 C 29 βα 20S NL: 9.22E3
C 29αβ 20 R C 29αββ 20S
m/z 400 → 217 C 29αββ 20R
Relative Abundance

m/z= 215.70-232.70 F: + c
80
C 29 Steranes and diasteranes
  EI SRM ms2
60 C 29βα20R
C 29 αβ C 29ααα 20R
[email protected]
C 29ααα 20S [215.70-218.70] MS
40 20S
AGSO_06AUG12_STER
20

0 C 30αββ 20S
100 C 30βα20S C 30 αββ 20R
C 30
ααα
NL: 8.80E2

80
m/z 414 → 217 C 30βα 20R 20R m/z= 215.70-232.70 F: + c
Relative Abundance

C 30 Steranes and diasteranes

  C 30ααα 20S EI SRM ms2
60 [email protected]
[215.70-218.70] MS
40
AGSO_06AUG12_STER
20
0
42 44 46 48 50 52 54 56 58
Time (min)
Figure 6. MRM chromatograms showing m/z M+>217 metastable transitions of C27, C28, C29 and C30 steranes and diasternaes. Peak assignments define
stereochemistry at C-20 (S and R); βα, αβ, ααα and αββ denote 13β(H),17α(H)-diasteranes, 13α(H),17β(H)-diasteranes, 5α(H),14α(H),17α(H)-steranes
and 5α(H),14β(H),17β(H)-steranes, respectively. XT = cross talks, * = isomeric peaks (24S and 24R).
Conclusions References
The capabilities of the Thermo Scientific DFS high 1. Tissot, B.P., Welte, D.H., 1984. Petroleum formation

Appli cat i on N ote 3 0 2 6 3

resolution mass spectrometer have enabled the and occurrence, pp. 699. Springer, Berlin.
continuation and development of oil biomarker research
2. Seifert, W.K., Moldowan, J.M., Jones, R.W., 1979.
at CSIRO Organic Geochemistry Laboratories.
Applications of biological marker chemistry to
The results generated using the DFS instruments allow petroleum exploration, 2, pp. 425-438. Heyden,
CSIRO to elucidate and quantify the molecular distribu- London.
tions of the aliphatic and aromatic hydrocarbon
3. Mackenzie, A.S., 1984. Applications of biological
biomarkers, such as, n-alkanes, regular isoprenoids,
markers in petroleum geochemistry. In: J.M. Brooks,
steranes and diasteranes, tri- and tetracyclic terpanes,
D.H. Welte (Eds.), Advances in Petroleum Geochemistry 1
hopanes, alkylnapthalenes, alkylphenanthrenes, retene,
(Ed. by J.M. Brooks, D.H. Welte), pp. 115-212. Academic
etc. (Figs 1 – 6). These results provide useful tools for the
Press, London.
assessment of:
4. Peters, K.E., Walters, C., Moldowan, J.M., 2005. The
• Secondary alteration processes
Biomarker Guide, pp. 1155 pp. Cambridge University
- biodegradation, evaporative fractionation,
Press, Cambridge.
water washing
• Source or origin of organic matter 5. Alexander, R., Larcher, A.V., Kagi, R.I., Price, P.L.,
- higher plants, algae, prokayriotic, eukaryotic 1988. The use of plant derived biomarkers for
correlation of oils with source rocks in the Cooper/
• Thermal maturity Eromanga Basin System, Australia. APEA Journal,
- immature, mature, over mature 28(1), 310-324.
• Palaeo-environmental condition
6. George, S.C., Volk, H., Dutkiewicz, A., Ridley, J.,
- marine, deltaic, lacustrine, terrigeneous
Buick, R., 2008. Preservation of hydrocarbons and
• Lithology biomarkers in oil trapped inside fluid inclusions for
- clay rich, carbonates >2 billion years. Geochimica et Cosmochimica Acta,
• Geological age 72, 844-870.

• Oil-oil and/or oil-source correlations 7. Gong, S., George, S.C., Volk, H., Liu, K., Peng, P.,
2007. Petroleum charge history in the Lunnan Low
Applications of such results in petroleum exploration
Uplift, Tarim Basin, China - Evidence from oil-bearing
geochemistry are described in numerous papers2–5 and
fluid inclusions. Organic Geochemistry, 38,
also in CSIRO publications.6–9 1341-1355.
Acknowledgements 8. Volk, H., Kempton, R.H., Ahmed, M., Gong, S.,
We would like to thank the team at CSIRO Sydney for George, S.C., Boreham, C.J., 2009. Tracking petroleum
the analysis of all samples and preparation of this systems in the Perth Basin by integrating microscopic,
application note. molecular and isotopic information on petroleum fluid
inclusions. Journal of Geochemical Exploration, 101,
109.

9. Ahmed, M., Volk, H., Allan, T., Holland, D., 2012.

Origin of oils in the Eastern Papuan Basin, Papua New
Guinea. Organic Geochemistry, 53, 137-152.

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