Efficient synthesis of unsymmetrically disubstituted ferrocenes: Towards electrochemical dipeptide-Fc-biosensors

Article abstract of DOI:10.1016/S0040-4039(01)00251-9

Unsymmetrically disubstituted ferrocenes (1a, 1b), electrochemical dipeptide-Fc-biosensors, were synthesized by a straightforward synthetic method from 1,1′-ferrocenedicarboxylic acid. In addition, an efficient solution synthesis of pentapeptide (2) was reported.

Full text of DOI:10.1016/S0040-4039(01)00251-9

TETRAHEDRON
LETTERS
Pergamon
Tetrahedron Letters 42 (2001) 2601–2603
Efficient synthesis of unsymmetrically disubstituted ferrocenes:
towards electrochemical dipeptide-Fc-biosensors
Yiming Xu and Heinz-Bernhard Kraatz*
Department of Chemistry, 110 Science Place, University of Saskatchewan, Saskatoon, SK, Canada S7N 5C9
Received 25 January 2001; revised 7 February 2001; accepted 8 February 2001
Abstract—Unsymmetrically disubstituted ferrocenes (1a, 1b), electrochemical dipeptide-Fc-biosensors, were synthesized by a
straightforward synthetic method from 1,1%-ferrocenedicarboxylic acid. In addition, an efficient solution synthesis of pentapeptide
(2) was reported. © 2001 Elsevier Science Ltd. All rights reserved.
In recent years, electrochemical biosensors have been
developed for the analytical determination of biological
and organic analytes.1 The research of DNA biosensors
has been particularly successful,2 providing new meth-
ods for mutation detection. This technology offers the
benefits of being fast, easy-to-use, and inexpensive, as
well as having a wide range of potential applications,
such as the detection of diseases, monitoring air quality,
and detection of bacterial contamination in food items.
Thus, the development of new biosensors is presently
receiving a significant amount of attention.1,2
oligopeptide chain.4 We decided to prepare two Fc-
oligoproline constructs with a second podant peptide
chain able to interact strongly with aspartic proteases,
such as HIV-1 protease. We designed two 1,1%-bispep-
tide-Fc compounds (1a, 1b), in which the podant pen-
tapeptide (2) is attached to the 1%-Cp ring of the Fc
group. 2 has the identical peptide sequence to that
found in pepstatin,
a potent naturally occurring
inhibitor of aspartic protease, including pepsin, HIV-1,
and HIV-2 proteases,5 but has not been prepared
before. Structurally, 1a and 1b are unsymmetrically
1,1%-disubstituted ferrocenes. Importantly, known syn-
thetic methodologies for the preparation of unsymmet-
rically 1,1%-disubstituted ferrocene derivatives involve
complex transformations.6 Here we have overcome this
synthetic challenge and report the straightforward but
selective introduction of two different peptides into the
ferrocene framework to produce 1a and 1b. Further-
more, we report the efficient synthesis of pentapeptide
(2) in solution.
Our group has been interested in the electrochemistry
of ferrocenoyl (Fc) peptides and its potential applica-
tion for the development of Fc-based electrochemical
biosensors.3 We discovered that the redox properties of
the Fc moiety in Fc-peptides is sensitive even to small
variations in peptide structure.3 Furthermore, the oligo-
proline chain allows for electronic communication
between the Fc group and the C-terminus of the
O
Val-Val-Sta-Ala-Sta-OH
Fe
N
1a: n = 3
1b: n = 4
HO
O
O
n
R1HN
H
N
H
N
pepstatin: R1=iso-valeryl R2=H
O
O
2:
COOR2
R1=Boc
R2=Et
N
H
N
H
O
O
OH
OH
Keywords: ferrocene; synthesis; pepstatin; disubstitued ferrocenes; biosensors.
* Corresponding author.
0040-4039/01/$ - see front matter © 2001 Elsevier Science Ltd. All rights reserved.
PII: S0040-4039(01)00251-9

2602
Y. Xu, H.-B. Kraatz / Tetrahedron Letters 42 (2001) 2601–2603
O
OH
b
a
COOEt
N
H
COOEt
NHBoc
OH
NHBoc
3
4
NHBoc
OH
O
H
N
c
2
N
H
COOEt
O
OH
5
Scheme 1. Reagents and conditions. (a) i. TFA, ii. Boc-Ala-OSu, CH2Cl2, DMF, 84%; (b) i. TFA, ii. Boc-Sta-OSu, CH2Cl2,
DMF, 54%; (c) i. TFA, ii. Boc-Val2-OSu, HOBt, EDC, CH2Cl2, 48%.
The synthesis of pentapeptide (2) is shown in Scheme 1.
Statine (3) was prepared from -lucine in four steps
on silica gel. Since the active OBt-ester can be readily
exchanged for another peptide using the established
procedures,10 both 8a and 8b are very useful synthons
for ferrocene derivatives having two different peptide
chain and are readily prepared from 1,1%-ferrocenedi-
carboxylic acid. Compounds 8a and 8b were obtained
by treatment of 1.5 molar equivalents of oligoprolines
with 1 mol of 1,1%-ferrocenedicarboxylic acid in the
presence of N-hydroxybenzotriazolw (HOBt) and 1-
ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC)
in CH2Cl2 at room temperature (yield: 55% for 8a, 45%
for 8b) (Scheme 2).
L
with an overall yield of 27% {[h]2D0 −39 (c 1.1, MeOH);
lit.7 [h]2D0 −37.9 (c 0.84, MeOH)}.7 Commercially avail-
able Boc-Ala-OH was transformed to Boc-Ala-OSu via
a reported method.8 Treatment of Boc-Sta-OEt with
trifluoroacetic acid gave H-Sta-OEt, which was coupled
with Boc-Ala-OSu in a mixed solvent (CH2Cl2:DMF,
5:1) at room temperature to afford Boc-Ala-Sta-OEt
(4). After deprotection of the C-terminus by base
hydrolysis,7 3 was transformed into Boc-Sta-OSu and
then coupled with dipeptide 4 to form Boc-Sta-Ala-Sta-
OEt (5).
As outlined in Scheme 2, the N-terminal of pentapep-
tide (2) was deprotected to give H-Val2-Sta-Ala-Sta-
OEt, which was then successfully coupled with 8a and
8b in CH2Cl2 at room temperature to give 9a and 9b in
yields of 59 and 55%, respectively.12 Finally after
deprotection of the benzyl and ethyl groups in a basic
dioxane–water mixture, the target compounds 1a and
1b were obtained.13
Employing the reported method,9 Boc-Val2-OMe was
obtained from Boc-Val-OH and H-Val-OMe. Depro-
tection of the methyl group with NaOH in dioxane/
H2O afforded Boc-Val2-OH. In the last step of the
peptide coupling sequence, Boc-Val2-OH and tripeptide
5 were coupled together by the standard carbodiimide
protocol10 in CH2Cl2 at room temperature for 20 h to
give pentapeptide (2) {mp 188–190°C, [h]2D0 −60.1 (c 1.0,
CHCl3)}. The product was purified by column chro-
matography on silica gel (CH2Cl2–MeOH, 98:2).
In summary, we report a solution synthesis of penta-
peptide (2) which has the identical peptide sequence
to that in pepstatin. Furthermore, we report a straight-
forward and useful synthetic method for the prepara-
tion of unsymmetrically 1,1%-disubstituted ferrocenes
from 1,1%-ferrocenedicarboxylic acid. In this work, we
synthesized two novel 1,1%-dipeptide-ferrocenes (1a, 1b).
Currently work is in progress to evaluate the electro-
In our previous research on syntheses of bisoligoproline
ferrocenes, we isolated 1-oligoproline-1%-OBt-ferrocene
derivatives (8a, 8b).11 These complexes are air and
moisture stable and easily separated from the symmet-
ric 1,1%-bisoligoproline-ferrocenes by chromatography
O
N
N
N
COOH
O
a
Fe
H-Pron-OBzl
Fe
+
N
HOOC
BzlO
O
n=3
7b n=4
O
7a
n
8a n=3
8b n=4
6
O
Val-Val-Sta-Ala-Sta-OEt
b
c
Fe
1
N
BzlO
O
1a n=3
1b n=4
O
9a n=3
9b n=4
n
Scheme 2. Reagents and conditions. (a) HOBt, EDC, CH2Cl2; (b) H-Val2-Sta-Ala-Sta-OEt, CH2Cl2; (c) NaOH, dioxane/H2O.

Y. Xu, H.-B. Kraatz / Tetrahedron Letters 42 (2001) 2601–2603
2603
chemistry of 1a and 1b and their ability to bind to
aspartate proteases.
5. (a) Umezawa, H.; Aoyagi, T.; Morishima, H.; Matsuzaki,
M.; Hamada, M.; Takeuchi, T. J. Antibiot. 1970, 23, 259;
(b) Babine, R. E.; Bender, L. E. Chem. Rev. 1997, 97,
1359.
Acknowledgements
6. (a) Lai, L.-L.; Dong, T.-Y. J. Chem. Soc., Chem. Com-
mun. 1994, 2347; (b) Butler, I. R.; Quayle, S. C. J.
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This work was funded by a grant from the Health
Services Utilization and Research Commission
(HSURC) of the Province of Saskatchewan.
7. Rick, D. H.; Sun, E. T.; Boparai, A. S. J. Org. Chem.
1978, 43, 3624.
8. Anderson, G. W.; Zimmerman, J. E.; Callahan, F. M. J.
Am. Chem. Soc. 1964, 86, 1839.
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12. Compounds 9a and 9b were purified by column chro-
matography on silica gel (CH2Cl2–MeOH, 96:4). Selected
data for 9a and 9b. 9a: mp 129–130°C, [h]2D0 −46 (c 0.91,
CHCl3), HRMS (FAB) m/z calcd for C65H95FeN8O14
(M++1) 1267.6317, found 1267.6324. 9b: mp 131–132°C,
[h]2D0 −41 (c 1.3, CHCl3), HRMS (FAB) m/z calcd for
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respectively, by recrystallization from MeOH–Et2O.
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(c 1.0, MeOH), HRMS (FAB) m/z calcd for
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