The first experimental demonstration of side chain extension of geoporphyrins in sediments

Article abstract of DOI:10.1246/cl.2010.1267

To investigate the formation process of high carbon number (>C ₃₂) sedimentary porphyrins, heating experiments of several porphyrins were performed. Chromic acid oxidation of the heating products of protoporphyrin IX dimethyl ester afforded 2-methyl-3-npropylmaleimide as the predominant product among the side-chain extension products formed. On the other hand, saturated substituents of etioporphyrin were also extended on heating to slowly form normal and branched homologs. These results may suggest that the transalkylation of porphyrin side chains proceeds mainly by a regioselective mechanism involving alkyl radical addition to a vinyl group of chlorophylls or their diagenetic products.

Full text of DOI:10.1246/cl.2010.1267

doi:10.1246/cl.2010.1267
Published on the web November 6, 2010
1267
The First Experimental Demonstration of Side Chain Extension of Geoporphyrins in Sediments
Kenta Asahina,1 Junya Asano,1 Gen Kumagai,1 Mitsuru Satou,1 Kouichi Nomoto,1
Yuichiro Kashiyama,1 Hajime Mita,2 and Shinya Nomoto*1
1Department of Chemistry, University of Tsukuba, Tsukuba, Ibaraki 305-8571
2Department of Life, Environment and Material Science, Fukuoka Institute of Technology, Fukuoka 811-0295
(Received September 3, 2010; CL-100756; E-mail: nomoto@chem.tsukuba.ac.jp)
To investigate the formation process of high carbon number
(>C32) sedimentary porphyrins, heating experiments of several
porphyrins were performed. Chromic acid oxidation of the heating
products of protoporphyrin IX dimethyl ester afforded 2-methyl-3-n-
propylmaleimide as the predominant product among the side-chain
extension products formed. On the other hand, saturated substituents
of etioporphyrin were also extended on heating to slowly form
normal and branched homologs. These results may suggest that the
transalkylation of porphyrin side chains proceeds mainly by a
regioselective mechanism involving alkyl radical addition to a vinyl
group of chlorophylls or their diagenetic products.
chains extending to C11 were obtained by chromic acid oxidation
of geoporphyrins from Cretaceous crude oil, together with
branched alkylmaleimides as minor components (Scheme 1).5
Similar results have been reported, in which the maleimides
containing n-alkyl side chains up to C5 predominated in the
oxidation products of geoporphyrins from Permian sedimentary
rocks, and 2-methyl-3-n-propyl- and 2-isobutyl-3-methylmale-
imide were appreciated as a biomarker of photosynthetic bacteria
utilizing bacteriochlorophylls, and further as an indicator
molecule of photic-zone euxinia (i.e., a seawater containing free
H2S) in connection with the massive extinction of organisms.6
In order to assess the origins of these high carbon number
geoporphyrins as well as the maleimides that possess long side
chains or specific alkyl groups related to bacteriochlorophylls, it
is important to elucidate the extension modes of porphyrin side
chains by transalkylation in sediments. For this purpose, we
performed heating experiments of protoporphyrin IX dimethyl
ester (2) and etioporphyrin I (3, Chart 2). Porphyrin 2 was
employed to elucidate side chain extension from a vinyl group at
the early stage of organic maturation in sediments and porphyrin
3, which possesses saturated alkyl substituents, was used as a
model substrate of thermally matured geoporphyrins.
Porphyrins 2 and 3 (ca. 0.2 mg) were heated at 300 and
350 °C for up to 360 h in a degassed sealed tube. The products
were oxidized with 10% CrO3 in 25% H2SO4 (1 mL) at 0 °C for
1 h and at room temperature for 1 h to afford a mixture of
maleimides and phthalimides that originated from benzoporphy-
rins formed during the heating.7 These products were then
extracted with benzene and analyzed by GC-MS equipped with
an FFAP capillary column (60 m © 0.25 mm i.d.). Identification
and quantification of the products were performed by comparison
of the GC-MS data with those of authentic samples.
Alkyl porphyrins in sediments and petroleum occur as
complex mixtures1 (Chart 1). They are suggested to be mostly
derived from defunctionalization of chlorophylls and hence have
the potential to provide information about the original chloro-
phylls, depositional conditions, and geochemical transforma-
tions.2 During the past 30 years, the molecular structures of more
than 80 porphyrins obtained from sedimentary organic matter
have been determined using HPLC, MS, and NMR techniques.
Oxidative degradation of porphyrins into maleimides and
subsequent analysis by GC-MS have also been applied to obtain
information regarding the nature of the side chains of geo-
porphyrins.3 Geoporphyrins with a skeleton of 32 carbons are
accounted for by a mechanism involving decarboxylation of
chlorophylls, but the presence has been shown of geoporphyrins
with carbon numbers greater than 32.4 It was proposed that these
high carbon number porphyrins were derived from bacterio-
chlorophylls (Chart 1) or from a random transalkylation at the
porphyrin side chains in sediments. On the other hand, it has been
reported that 2-n-alkyl-3-methylmaleimides (1) with n-alkyl side
Figure 1 shows typical gas chromatograms obtained by the
heating experiment. Porphyrin 2 afforded insoluble polymeric
OH
R1
NH
N
N
NH
N
R
R
N
N
N
N
N
Oxidation
Mg
HN
HN
O
O
N
N
R2
H
2-n-Alkyl-3-methylmaleimides (1)
O
Typical sedimentary porphyrins
CO2Farnesyl
Scheme 1.
Bacteriochlorophylls d
R1: Et, n-Pro, or i-Bu
R2: Me or Et
R1
N
N
N
N
Protoporphyrin IX dimethyl ester (2)
R2
Mg
R = R = vinyl
1
2
NH
N
N
R = R = CH CH CO Me
3
4
2
2
2
R = Me
5
HN
Etioporphyrin I (3)
R = R = R = R = Et
O
MeO2C
R5
CO2Phytyl
1
2
3
5
R = Me
R4
4
R3
Chlorophyll a
Chart 1.
Chart 2.
Chem. Lett. 2010, 39, 1267-1269
© 2010 The Chemical Society of Japan
www.csj.jp/journals/chem-lett/

1268
(a)
(b)
0.05
0.04
0.03
0.02
0.01
0
0.05
0.04
0.03
0.02
0.01
0
m
n
b
c
a
f
m/z
o
m/z
147, 161
p
l
111
k
125
139
153
167
165
175
e
i
s
0.2
0
0.1
0
0.1
DMMi / (DMMi + EMMi)
0.2
d
g
DMMi / (DMMi + EMMi + MceMMi)
r
v
q
t
u
h
j
Figure 3. Plots of the molar fractions of C8 maleimides formed from
heated protoporphyrin IX dimethyl ester (a) and etioporphyrin I (b)
against the maturity index. : MnPMi, : DEMi, : MiPMi.
20
35 40
Retention time / min
65
R
R
Figure 1. Mass fragmentograms of maleimides and phthalimides
obtained by oxidation of protoporphyrin IX dimethyl ester heated at
300 °C for 360 h. a: MMMi, b: DMMi, c: EMMi, d: MiPMi, e: DEMi,
f: MnPMi, g: EiPMi, h: MsecBMi, i: EnPMi, j: MiBMi, k: MnBMi,
l: MnPenMi, m: Pi, n: 3-MPi, o: MceMMi, p: 4-MPi, q: 3,6-DMPi,
r: 3-EPi, s: 3,5-DMPi, t: 4-EPi, u: 3,4-DMPi, v: 4,5-DMPi.
H
CH2-R
N
N
N
Scheme 2.
(a)
(b)
R
1
1
0.8
0.6
0.4
0.2
0
0.8
0.6
0.4
0.2
0
N
N
+
CH2-R
+
R
N
0.2
0
0.1
DMMi / (DMMi + EMMi)
0.2
0
0.1
DMMi / (DMMi + EMMi + MceMMi)
N
N
Figure 2. Plots of the molar fractions of C5 to C7 maleimides and
MceMMi formed from heated protoporphyrin IX dimethyl ester (a)
Scheme 3.
and etioporphyrin I (b) against the maturity index. : MceMMi,
EMMi, : DMMi, : MMMi.
:
Heating of 2 and 3 respectively afforded a characteristic
profile of C8 maleimide formation, as shown in Figure 3. MnPMi
obtained during the heating of 2 rapidly increased in the region of
the maturity index up to ca. 0.08, while all the C8 maleimides
from 3 gradually increased with increasing maturity. The amount
of MnPMi formed from 2 was about 4% of all the products at its
maximum. It is possible to consider that a methyl radical
generated in the reaction system selectively added itself to a
terminal carbon of a vinyl group of 2 to form a methyl-n-
propylpyrrole structure (Scheme 2). The rapid formation of
MnPMi may be attributed to the high reactivity of a vinyl group
to radicals, resulting in complete consumption of the vinyl group
until the maturity index of ca. 0.08.
Another remarkable difference was observed in the isomer
distribution of the C8 maleimides formed. Thus, the major
product from 3 was DEMi, which can be formed by one-carbon
extension at a methyl group of an ethylmethylpyrrole unit.
Formation of MnPMi exceeded that of MiPMi to some extent
(Scheme 3). The tendency for a saturated side chain to extend
into a straight chain was confirmed by separate heating experi-
ments using octaethylporphyrin, which afforded EnPMi and
EiPMi in the ratio of 4 to 1 in the range of the maturity index
(EMMi/(EMMi + DEMi)) up to 0.4.
products by heating, but maleimides and phthalimides were
detected after oxidation of the products. 2-Methyl-3-vinylmale-
imide that should have originated from the starting material was
not observed, possibly due to decomposition of the vinyl group
during the chromic acid oxidation. The determined alkyl
maleimides possessing extended alkyl side chains are three C8
(DEMi, MnPMi, and MiPMi), five C9 maleimides (EiPMi,
EnPMi, MsecBMi, MiBMi, and MnBMi), and a C10 maleimide
(MnPenMi). MnPenMi was not formed from 3.8
In Figures 2 and 3, the ratios of each maleimide to all the
maleimides and phthalimides obtained during the heating of
porphyrins 2 and 3 are plotted against DMMi/(DMMi +
EMMi + MceMMi) and DMMi/(DMMi + EMMi), respective-
ly, which represent thermal maturity of the porphyrins.3b
MceMMi that originates from the starting porphyrin rapidly
decreased at the initial stage of the heating by demethoxycarbon-
ylation, producing EMMi as shown in Figure 2a. The main
products formed from an ethylmethylpyrrole unit by the heating
of 2 and 3 were DMMi and MMMi, which increased with
increasing thermal maturation (Figure 2). When these porphyrins
were heated with naphthalene at 400 °C for 24 h and the products
were extracted and analyzed, the alkyl reactive species released
from the porphyrins were successfully captured as alkylnaph-
thalenes; the ratios of methyl-, ethyl-, and propylnaphthalenes
from 2 and 3 were 85:13:2 and 76:22:2, respectively. These
results may indicate that these alkyl reactive species affected the
extension of the porphyrins.
MnBMi was formed as the major product among the C9
maleimides during the heating of 2, and the amount was about a
tenth of that of MnPMi. Since an ethyl reactive species can be
generated from the porphyrin as stated above, MnBMi obtained
was likely formed by addition of an ethyl radical to a vinyl group
of a methylvinylpyrrole unit. The superior production of MnBMi
Chem. Lett. 2010, 39, 1267-1269
© 2010 The Chemical Society of Japan
www.csj.jp/journals/chem-lett/

1269
(a)
(b)
0.02
0.01
0.008
0.006
0.004
0.002
0
MnBMi
MnHeptylMi
MnPMi
0.01
0
MnOctylMi
MnNonylMi
MnPenMi
MnHexylMi
0
0.1
DMMi / (DMMi + EMMi)
0.2
0
0.1
DMMi / (DMMi + EMMi)
0.2
m/z 125
25
30
35
Retention time / min
40
45
Figure 5. Plots of the molar fractions of C8 (a) and, C9 and C10
maleimides (b) extracted from the stratigraphic sequence of sediments
at Soumaoki against the maturity index. : MnPMi, : DEMi,
:
Figure 4. Mass fragmentograms of maleimides obtained by oxida-
tion of protoporphyrin IX dimethyl ester heated at 350 °C for 6 h in the
presence of n-decane.
MiPMi, : MnBMi, : MnPenMi, : MiBMi, : MsecBMi, Ã:
EnPMi.
and EnPMi to branched C9 maleimides was observed in the
heating of 3, indicating a straight chain-forming tendency for the
side-chain extension of alkylporphyrins. MiBMi, which can
potentially be used as a biological marker for bacteriochloro-
phylls, was formed from both 2 and 3, though in small amounts.
The 2-n-alkyl-3-methylmaleimides generated by the heating
were MnPMi, MnBMi, and MnPenMi from porphyrin 2 and
MnPMi and MnBMi from 3, but those with even longer
substituents have been reported to exist in sediments.5 We then
performed heating experiments of the porphyrins in the presence
of n-alkane to explore the possibility of transalkylation from
coexisting carbon sources to porphyrin side chains. Porphyrins 2
and 3 were heated at 350 °C for 6 h with a large excess of n-
decane and the products were oxidized and analyzed. A major
homologous series of methylmaleimides with n-alkyl side chains
extending to C9 were observed in the heating products of 2
(Figure 4), showing that fragments of n-decane were introduced
to the porphyrin side chains without suffering any rearrangement.
Compared with the heating of 2, the superiority of 2-n-alkyl-3-
methylmaleimides was rather ambiguous in the heating products
of porphyrin 3, possibly because 2-n-alkyl-3-ethylmaleimides
were also produced.
for the formation of MnBMi and MnPenMi, both of which were
found in the sediments in larger amounts than the other C9
maleimides (Figure 5b). It is worth noting that the oxidative
extracts of the sediments contained MnPMi and MiBMi in the
amounts comparable to those produced during the heating of
protoporphyrin, showing that the major portions of these
maleimides are likely not of bacteriochlorophyll origin but
produced by geochemical transformation in sediments.
This study has established the chemical background for
interpreting the formation of high carbon number porphyrins in
sediments for the first time. Thus, the transalkylation of porphy-
rin side chains has been demonstrated to proceed mainly by a
regioselective mechanism involving alkyl radical addition to a
vinyl group of chlorophylls or their transformation products,
which could explain the molecular distribution of 2-n-alkyl-3-
methylmaleimides in oxidative extracts from sedimentary rocks.
This process was accompanied by reactions that extend saturated
substituents of etioporphyrin to produce its normal and branched
homologs, with normal ones being major products. The results of
this study also implicate the role of diagenesis as an important
control on the occurrence of structurally altered biomarker
molecules in matured sediments.
Maleimides were then identified in the oxidative extracts of
natural sediments, such as the Cretaceous/Tertiary stratigraphic
sequence of sediments at Soumaoki, Japan. Thus, after washing
of powdered sediment (ca. 5 g) with a mixture (10 mL) of
benzene and methanol (5:1, v/v) by sonication over five cycles,
the sample was oxidized with 10% CrO3 in 25% H2SO4 (10 mL)
at 0 °C for 2 h and at room temperature for 2 h. The suspension
was extracted five times with benzene (10 mL), and the extracts
were combined and concentrated carefully to 100 ¯L under a
nitrogen flow and analyzed as stated above. The total concen-
trations of all the maleimides and phthalimides in the sediments
were 3-70 nmol/g-rock and those of the C8 and the C9 to C10
maleimides were 40-580 and 40-220 pmol/g-rock, respectively.
The molar fractions of maleimides possessing extended side
chains in all the maleimides and phthalimides determined in the
sediments were comparable to those obtained during the heating
of the porphyrins. The isomer compositions of the C8 maleimides
were in accord with the results of the heating experiments of
protoporphyrin throughout the sequence, with MnPMi being the
predominant isomer (Figure 5a). This fact may indicate that a
methyl-n-propylpyrrole unit of sedimentary porphyrins had been
produced mainly from a methylvinylpyrrole unit of tetrapyrrole
pigments in the early stage of geochemical transformation and
preserved in the sediments. Alkyl radical addition could account
References and Notes
1
2
3
a) A. Treibs, Angew. Chem. 1936, 49, 682. b) E. W. Baker, J. Am.
Chem. Soc. 1966, 88, 2311.
B. Huseby, R. Ocampo, C. Bauder, H. J. Callot, K. Rist, T. Barth, Org.
Geochem. 1996, 24, 691.
a) A. Shimoyama, K. Kozono, H. Mita, S. Nomoto, Geochem. J. 2001,
35, 365. b) M. Kozono, S. Nomoto, A. Shimoyama, Chem. Lett. 2002,
470. c) S. Nomoto, H. Kigoshi, Res. Org. Geochem 2005, 20, 31.
a) R. Ocampo, H. J. Callot, P. Albrecht, J. Chem. Soc., Chem.
Commun. 1985, 200. b) J. Verne-Mismer, R. Ocampo, H. J. Callot, P.
Albrecht, Tetrahedron Lett. 1986, 27, 5257. c) R. Gibbison, T. M.
Peakman, J. R. Maxwell, Tetrahedron Lett. 1995, 36, 9057. d) D. H.
Mawson, J. S. Walker, B. J. Keely, Org. Geochem. 2004, 35, 1229.
J. Martin, E. Quirke, G. R. Shaw, P. D. Soper, J. R. Maxwell,
Tetrahedron 1980, 36, 3261.
4
5
6
7
8
R. D. Pancost, N. Crawford, J. R. Maxwell, Chem. Geol. 2002, 188,
217.
S. Nomoto, M. Satou, T. Yoshida, H. Mita, G. Kumagai, K. Nomoto,
H. Kigoshi, Y. Kashiyama, Chem. Lett. 2008, 37, 490.
Abbreviations of maleimides (Mis) and phthalimides (Pis) are as
follows; MMMi: 3-methylMi, DMMi: 2,3-dimethylMi, EMMi: 2-
ethyl-3-methylMi, DEMi: 2,3-diethylMi, MnPMi: 2-methyl-3-n-pro-
pylMi, MiPMi: 2-methyl-3-isopropylMi, EnPMi: 2-ethyl-3-n-propyl-
Mi, EiPMi: 2-ethyl-3-isopropylMi, MnBMi: 2-n-butyl-3-methylMi,
MiBMi: 2-isobutyl-3-methylMi, MsecBMi: 2-sec-butyl-3-methylMi,
MnPenMi: 2-methyl-3-n-pentylMi, MceMMi: 2-methoxycarbonyleth-
yl-3-methylMi, MPi: methylPi, EPi: ethylPi, DMPi: dimethylPi.
Chem. Lett. 2010, 39, 1267-1269
© 2010 The Chemical Society of Japan
www.csj.jp/journals/chem-lett/