Synthesis and characterization of novel N-para-ferrocenyl benzoyl dipeptide esters

Article abstract of DOI:10.1016/j.inoche.2006.06.013

N-para-ferrocenyl benzoyl dipeptide esters 2-5 were prepared by coupling para-ferrocenyl benzoic acid 1 to the dipeptide ethyl esters GlyGly(OEt) 2, GlyAla(OEt) 3, GlyLeu(OEt) 4 and GlyPhe(OEt) 5. The compounds were fully characterized by a range of NMR s

Full text of DOI:10.1016/j.inoche.2006.06.013

Inorganic Chemistry Communications 9 (2006) 1267–1270
www.elsevier.com/locate/inoche
Synthesis and characterization of novel N-para-ferrocenyl
benzoyl dipeptide esters
a
a
c
David Savage , Steven R. Alley a,b, Alok Goel a,b, Tara Hogan , Yoshiteru Ida ,
Paula N. Kelly a,b, Laurent Lehmann , Peter T.M. Kenny
a
a,b,
*
a
School of Chemical Sciences, Dublin City University, Dublin 9, Ireland
National Institute for Cellular Biotechnology, Dublin City University, Dublin 9, Ireland
School of Pharmaceutical Sciences, Showa University, Hatanodai, Shinagawa-ku, Tokyo 142-8555, Japan
b
c
Received 29 May 2006; accepted 10 June 2006
Available online 27 June 2006
Abstract
N-para-ferrocenyl benzoyl dipeptide esters 2–5 were prepared by coupling para-ferrocenyl benzoic acid 1 to the dipeptide ethyl esters
GlyGly(OEt) 2, GlyAla(OEt) 3, GlyLeu(OEt) 4 and GlyPhe(OEt) 5. The compounds were fully characterized by a range of NMR spec-
troscopic techniques, mass spectrometry (ESI-MS, FABMS, MS/MS) and cyclic voltammetry (CV). The a1 and b1 ions appeared at one
mass unit less than the expected mass in the tandem mass spectra. The CV curves of compounds 2–5 exhibited quasi-reversible redox
behavior similar to the Fc/Fc+ redox couple and displayed oxidation potential values in the 50–55 mV range, against the Fc/Fc+ redox
couple.
Ó 2006 Elsevier B.V. All rights reserved.
Keywords: Ferrocene; Bioorganometallic chemistry; Tandem mass spectrometry; Dipeptides; CV
The organometallic compound ferrocene has several
novel applications due to its stability, spectroscopic proper-
ties, electrochemical properties and ease of use. Research in
the area of ferrocenyl derivatives has focused on the prep-
aration of novel sensor compounds, peptide mimetic mod-
els and unnatural drugs [1–7]. As amino acids and peptides
play such diverse roles in biological systems, incorporation
of a redox active group onto these compounds is of partic-
ular interest. For example, the incorporation of a ferrocene
group onto proteins has shown the mediation of electron
transfer between electrodes and the protein redox site [8].
Also the synthesis and structural characterization of
N-ferrocenoyl and N-ferrocenyl amino acid and peptide
derivatives has been reported [9–26]. In addition ferrocene
has been incorporated in drugs such as antibiotics, aspirin,
anti-malarials and anti-cancer drugs such as tamoxifen
[27–30]. A review on the bioorganometallic chemistry of
ferrocene has been published [31]. Here, we now present
the synthesis and structural characterization of a series of
novel N-para-ferrocenyl benzoyl dipeptide ester deriva-
tives. The aim of this research is to incorporate three key
moieties in the synthesis of unusual biological materials,
namely, (i) an electroactive core, (ii) a conjugated linker
that can act as a chromophore and (iii) an amino acid or
dipeptide derivative that can interact with other molecules
via hydrogen bonding. The ferrocene moiety is linked to
the dipeptide ester via a para-disubstituted benzoyl group.
We recently reported the synthesis and structural charac-
terization of a series of N-para, N-meta, and N-ortho-ferr-
ocenyl benzoyl amino acid and N-meta-ferrocenyl
benzoyl dipeptide ester derivatives. [32–36].
*
Corresponding author. Address: School of Chemical Sciences, Dublin
City University, Dublin 9, Ireland. Tel.: +353 1 7005689; fax: +353 1
7005503.
para-Ferrocenyl benzoic acid 1 was prepared as previ-
ously reported [32]. Conventional peptide chemistry was
E-mail address: peter.kenny@dcu.ie (P.T.M. Kenny).
1387-7003/$ - see front matter Ó 2006 Elsevier B.V. All rights reserved.
doi:10.1016/j.inoche.2006.06.013

1268
D. Savage et al. / Inorganic Chemistry Communications 9 (2006) 1267–1270
employed in the preparation of the dipeptides. Thus equi-
molar quantities of the N-Boc protected amino acids were
reacted with the amino acid ethyl ester hydrochloride salts,
1,3-dicyclohexylcarbodiimide (DCC), catalytic amounts of
1-hydroxybenzotriazole (HOBt) and triethylamine in DCM
at 0 °C. Deprotection of the amino terminal was achieved
using TFA. The deprotected dipeptides were coupled to
para-ferrocenyl benzoic acid using equimolar amounts of
DCC and catalytic amounts of HOBt under basic condi-
tions (Scheme 1). Purification by silica gel column chroma-
tography furnished the pure products in yields of 52–67%
and all gave analytical and spectroscopic data in accor-
dance with the proposed structures [37]. The N-para-ferr-
ocenyl benzoyl derivatives 2–5 were characterized by a
combination of 1H NMR, 13C NMR, DEPT-135 and
1HA13C COSY (HMQC) spectroscopy, ESI-MS, FABMS,
MS/MS and CV. All the proton and carbon chemical shifts
for compounds 2–5 were unambiguously assigned by a
the meta and ortho protons on the (g5-C5H4) ring are pres-
ent at d 4.31 and d 4.63, respectively. The 13C NMR spectra
of compounds 2–5 show signals in the region d 67.2 to d
83.9 indicative of a monosubstituted ferrocene subunit.
The ipso carbon of the (g5-C5H4) ring appears in a very
narrow range of d 83.7 to d 83.9. This signal is absent in
the DEPT 135 spectra. The carbon atoms of the aromatic
ring are visible in the region d 126.2–144.5. The two quater-
nary carbons of the aromatic ring and the methylene
carbon atoms of derivatives 2–5 were identified by
DEPT-135. For example, in the 13C NMR spectrum of
N-{para-(ferrocenyl)benzoyl}-glycine-glycine ethyl ester 2,
as is typical for a para disubstituted compound, there are
only four aromatic signals. The two phenyl ipso carbons
appear at d 144.5 and d 130.8 and the two other aromatic
signals appear at d 127.7 and d 126.3, respectively. The sig-
nal at d 83.7 is absent in the DEPT 135 spectrum and there-
fore it is assigned as the ipso ferrocenyl carbon. The (g5-
C5H5) ring appears as an intense signal at d 70.2 and the
meta carbons on the (g5-C5H4) ring appear at d 70.1. The
ortho carbons on the (g5-C5H4) ring appear at d 67.2. A
1
combination of DEPT-135 and H-13C-COSY (HMQC).
1
The H and 13C NMR spectra for compounds 2–5 showed
peaks in the ferrocene region characteristic of a ferrocenyl
benzoyl moiety [32–36]. The protons in the ortho position
of the (g5-C5H4) ring appear in the region d 4.61 to d
4.64 whereas the protons in the meta position occur in
the range d 4.3 to d 4.32. The (g5-C5H5) ring appears in
the narrow region d 3.95 to d 3.98. The protons of the
para-disubstituted benzoyl group appear as two doublets
from d 7.43 to d 7.70. For example, in the case of N-
{para-(ferrocenyl)benzoyl}-glycine-glycine ethyl ester 2,
the aromatic protons are present as two doublets at d
7.45 and d 7.69, respectively. The (g5-C5H5) ring appears
1
complete assignment of the H and 13C NMR spectra of
compound 2 is presented in Table 1.
Since the introduction of soft ionization techniques such
as electrospray ionization (ESI) and fast atom bombard-
ment mass spectrometry (FABMS) a wide range of thermo-
labile and non-volatile compounds can be subjected to
mass spectrometric analysis [38,39]. FAB and ESI were
employed in the analysis of compounds 2–5 and confirmed
the correct relative molecular mass. Examination of the
mass spectra revealed the presence of intense radical-cat-
ions, with cation adducts due to sodium and potassium
1
as a singlet in the H NMR spectrum at d 3.96 whereas
O
O
i
OMe
+
H2N
Fe
Fe
OMe
ii
O
O
NH
Fe
OH
O
iii
Fe
HN
R
1
2-5
O
OEt
Scheme 1. Synthesis of N-para-ferrocenyl benzoyl dipeptide esters 2–5; GlyGly(OEt) 2, GlyAla(OEt) 3, GlyLeu(OEt) 4, GlyPhe(OEt) 5. (i) NaNO2, HCl,
5 °C, (ii) NaOH/MeOH, H2O, (iii) DCC, HOBt, Et3N, Dipeptide ethyl ester.

D. Savage et al. / Inorganic Chemistry Communications 9 (2006) 1267–1270
1269
Table 1
amide nitrogen. This is unusual as these a1 and b1 fragment
ions are usually produced without loss of a hydrogen atom
[40].
1H and 13C spectroscopic data for 2
13
14
O
O
2
3
H
N
11
18
16
The CV curves of compounds 2–5 exhibit quasi-revers-
ible redox behavior, similar to the Fc/Fc+ redox couple,
and display oxidation potential values ca. 51–55 mV higher
than ferrocene. For example, the Eo0 value for compound 2
is calculated to be 55 mV with respect to ferrocene. The
slight increase in the oxidation potential with respect to
unsubstituted ferrocene is explicable in terms of substituent
effects. Electron withdrawing amide groups on phenyl ring
make the ferrocene moiety electron deficient and more dif-
ficult to oxidize.
22
4
19
1
17
21
N
H
O
23
F5e
15
20
12
6
O
10
7
9
8
Site
1H NMR
13C NMR
83.7
HMQC
1
2, 5
3, 4
4.63
4.31
3.96
67.2
70.1
70.2
6–10
11
144.5
In conclusion, the novel N-para-ferrocenyl benzoyl
dipeptide esters 2–5 were prepared in good yields using
organic peptide synthetic protocols. The compounds were
characterized by NMR spectroscopic technique, tandem
mass spectrometry and CV. The tandem mass spectra
revealed the loss of a hydrogen atom in the production
of the a1 and b1 type product ions. In addition the com-
pounds are electroactive, with oxidation potentials compa-
rable to ferrocene and have the ability to interact with
other molecules via hydrogen bonding, which make these
compounds suitable as potential receptors for sensor
devices with electrochemical detection.
12, 13
14, 15
16
7.45
7.69
126.3
127.7
130.8
17
18
168.1a
4.12–15
4
44
19
20
169.8a
170.1
41.8
21
22
4.12–15
1.21
62.1
14.5
23
a
signals may be reversed.
also present. Fragment ions were not observed or were of
very low intensity. Therefore tandem mass spectrometry
was employed to confirm the integrity of the structures.
As the radical-cation species were of greater intensity than
the [M + H]+ species these ions were selected as the precur-
sor ions. Sequence specific ions were observer in all the
MS/MS spectra for compounds 2–5 confirming that the
glycine residue was linked to the benzoyl spacer group.
Important product ions were present at m/z 261, m/z 289,
m/z 317 and m/z 345 (Fig. 1). The ions at m/z 261 and
m/z 289 are due to the ferrocenylphenyl and ferro-
cenylbenzoyl subunits respectively. However, the expected
a1 and b1 product ions at m/z 318 and m/z 346 were not
observed, instead a1-1 and b1-1 product ions were observed
at m/z 317 and m/z 345 respectively. Accurate mass mea-
surement of the MS/MS spectrum for compound 4 gave
an elemental composition of C18H15N1O1Fe1 and a molec-
ular mass of m/z 317.0497 for the ion at nominal mass m/z
317. This corresponds to a ppm error of 0.1420. Obviously
a hydrogen atom has also been lost during the fragmenta-
tion process. The hydrogen atom is either one of the glycine
methylene hydrogen atoms or the hydrogen on the glycine
Acknowledgements
D.S. thank the Irish American Partnership and Dublin
City University for the funding of a studentship award
1999–2002. This research was partly supported by the Na-
tional Institute for Cellular Biotechnology under the Pro-
gramme for Research in Third Level Institutions (PRTLI,
round 3, 2001–2006).
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ethyl acetate furnished the title compound as orange needles (0.253 g,
56%).
m.p. 165–167 °C, Eo0 ¼ 55 mV (vs Fc/Fc+).
Analysis: found: C, 61.46; H, 5.56; N, 6.36.
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Mass spectrum: found: [M]+ 448
C23H24N2O4Fe requires: 448
I.R. mmax (KBr): 3321, 2924, 1742, 1622, 1574, 1501, 1311 cmꢀ1
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.
1H NMR (400 MHz) d (CDCl3): 7.69 (2H, d, J = 8 Hz, ArH), 7.45
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(2H, d, J = 5.2 Hz, ANHCH2COA), 3.96 {5H, s, (g5-C5H5)}, 1.21
(3H, t, J = 7.6 Hz, -OCH2CH3). 13C NMR (100 MHz) d (CDCl3):
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