The synthesis, structural characterization and in vitro anticancer activity of novel 1-alkyl-1′-N-meta-(ferrocenyl) benzoyl dipeptide esters and novel 1-alkyl-1′-N-ortho-(ferrocenyl) benzoyl dipeptide esters

Article abstract of DOI:10.1016/j.jorganchem.2014.04.027

1-Alkyl-1′-N-meta-(ferrocenyl) benzoyl dipeptide esters 7-18 and 1-alkyl-1′-N-ortho-(ferrocenyl) benzoyl dipeptide esters 19-30 were prepared by coupling the alkyl ferrocenyl benzoic acids 1-6 to the dipeptide ethyl esters Gly-Gly(OEt), Gly-L-Ala(OEt), Gly-L-Leu(OEt) and Gly-L-Phe(OEt) using the standard N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC), 1-hydroxybenzotriazole (HOBt) protocol. The alkyl groups employed in the synthesis were methyl, ethyl and propyl. The compounds were characterized using a combination of ¹H NMR, ¹³C NMR, DEPT-135 and ¹H-¹³C COSY (HMQC) spectroscopy and electrospray ionization mass spectrometry (ESI-MS). Selected compounds showed micromolar activity in the H1299 NSCLC cell line, with IC₅₀ values in the range of 2.6-20.1 μM.

Full text of DOI:10.1016/j.jorganchem.2014.04.027

Accepted Manuscript
The synthesis, structural characterization and in vitro anticancer activity of novel 1-
alkyl-1’-N-meta-(ferrocenyl) benzoyl dipeptide esters and novel 1-alkyl-1’-N-ortho-
(ferrocenyl) benzoyl dipeptide esters
Andy G. Harry , William E. Butler , Jennifer C. Manton , Mary T. Pryce , Norma
O'Donovan , John Crown , Dilip K. Rai , Peter T. M. Kenny
PII:
S0022-328X(14)00209-5
10.1016/j.jorganchem.2014.04.027
JOM 18562
DOI:
Reference:
To appear in: Journal of Organometallic Chemistry
Received Date: 26 March 2014
Revised Date: 25 April 2014
Accepted Date: 27 April 2014
Please cite this article as: A.G. Harry, W.E. Butler, J.C. Manton , M.T. Pryce, N. O'Donovan, J. Crown,
D.K. Rai, P.T.M. Kenny, The synthesis, structural characterization and in vitro anticancer activity of novel
1-alkyl-1’-N-meta-(ferrocenyl) benzoyl dipeptide esters and novel 1-alkyl-1’-N-ortho-(ferrocenyl) benzoyl
dipeptide esters, Journal of Organometallic Chemistry (2014), doi: 10.1016/j.jorganchem.2014.04.027.
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ACCEPTED MANUSCRIPT
1-Alkyl-1’-N-meta and ortho-(ferrocenyl) benzoyl dipeptide ethyl esters 7-30 were
prepared by conventional peptide chemistry using the EDC/HOBT protocol.
Compounds 7-30 exhibited cytotoxic effects on the human lung carcinoma cell line
H1299. 1-Methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 7 had
an IC50 value of 2.6 µM.
O
O
Fe
NH
HN
O
O
7

ACCEPTED MANUSCRIPT
The synthesis, structural characterization and in vitro anticancer activity of novel 1-alkyl-
1’-N-meta-(ferrocenyl) benzoyl dipeptide esters and novel 1-alkyl-1’-N-ortho-(ferrocenyl)
benzoyl dipeptide esters.
Andy G. Harrya, William E. Butlera,b, Jennifer C. Mantona , Mary T. Prycea, Norma O'Donovanb,
John Crownc, Dilip K. Raid, Peter T. M. Kennya,b*
a School of Chemical Sciences, Dublin City University, Dublin 9, Ireland
b National Institute for Cellular Biotechnology, Dublin City University, Dublin 9, Ireland
c Dept. Of Medical Oncology, St. Vincent’s University Hospital, Dublin 4, Ireland.
d Dept. of Food Biosciences, Teagasc Food Research Centre, Ashtown, Dublin 15,
Ireland.
Abstract
1-Alkyl-1’-N-meta-(ferrocenyl) benzoyl dipeptide esters 7-18
and 1-alkyl-1’-N-ortho-
(ferrocenyl) benzoyl dipeptide esters 19-30 were prepared by coupling the alkyl ferrocenyl
benzoic acids 1-6 to the dipeptide ethyl esters Gly-Gly(OEt), Gly-L-Ala(OEt), Gly-L-Leu(OEt)
and Gly-L-Phe(OEt) using the standard N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide
hydrochloride (EDC), 1-hydroxybenzotriazole (HOBt) protocol. The alkyl groups employed in
the synthesis were methyl, ethyl and propyl. The compounds were characterized using a
1
13
1
combination of H NMR, C NMR, DEPT-135 and H-13C COSY (HMQC) spectroscopy and
electrospray ionization mass spectrometry (ESI-MS). Selected compounds showed micromolar
activity in the H1299 NSCLC cell line, with IC50 values in the range of 2.6 to 20.1 ꢀM.
Keywords : Ferrocene; Bioorganometallic chemistry; Dipeptides; Cytotoxicity; Lung cancer.
* Corresponding Author
Dr. Peter T. M. Kenny,
School of Chemical Sciences,
Dublin City University,
Dublin 9, Ireland.
Fax: +353 1 7005503
Phone: +353 1 7005689
e-mail: peter.kenny@dcu.ie

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Introduction
Interest in metal complexes with biological applications is currently an active area of research [1-
3]. Some of the most promising metal anticancer agents are emerging from the field of
bioorganometallic chemistry. A recent review article has categorized the diverse range of metal
anticancer compounds according to their mode of action [4]. Also a perspective article on
organometallic anticancer compounds has been published [5]. Ferrocene is a particularly
attractive candidate for incorporation into biologically active compounds due to its aromaticity,
stability, redox properties and low toxicity [6, 7]. The reversible redox properties of ferrocene
have been strongly associated with its biological activity [8]. One key feature of ferrocene is the
ease by which it undergoes oxidation to form the ferricenium cation (Fc → Fc+). This occurs in a
reversible manner and is accommodated readily by the loss/gain of an electron from a non-
bonding high energy molecular orbital. Ferricenium salts that are known to inhibit tumor growth,
have been shown to produce hydroxyl (HO•) radicals under physiological conditions, leading to
oxidatively damaged DNA [9]. The catalytic generation of intracellular reactive oxygen species
(ROS) such as the HO• radical offers an attractive and alternative method to target and kill cancer
cells [10]. To date, the most promising ferrocene-based drug candidate consist of a [3]-
ferrocenophane motif and have a potent in vitro anti-proliferative effect in breast and prostate
cancer cells lines [11-13]. This research group have prepared N-ferrocenoyl and N-ferrocenyl
amino acid and dipeptide derivatives and investigated their potential applications [14-18]. It was
shown that N-(ferrocenyl)benzoyl and N-(ferrocenyl)naphthoyl amino acid and dipeptide
derivatives were potential anticancer agents [19-26]. In the case of our ferrocenyl-peptide
bioconjugates, the presence of the conjugating aromatic linker between the ferrocene and peptide
units lowers the redox potential to within the range of biologically accessible potentials, allowing
for the interconversion between the ferrocene and ferricenium species [24-25, 27]. The
anticancer activity of the ferrocenyl bioconjugates is possibly due at least in part to their low
redox potentials, which are within the range of biologically accessible potentials. Therefore in
order to explore this hypothesis further, alkyl groups were introduced on the previously
unsubstituted cyclopentadiene ring to further modify the redox potentials.
Recently, we reported a study of 1-alkyl-1’-N-para-(ferrocenyl)benzoyl dipeptide esters which
found that alkylation of the the previously unsubstituted cyclopentadiene ring resulted in an
increase in cytotoxicity [28]. We herein report the synthesis, structural characterization and

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biological evaluation of novel 1-alkyl-1’-N-meta-(ferrocenyl)benzoyl dipeptide esters and 1-
alkyl-1’-N-ortho-(ferrocenyl)benzoyl dipeptide esters. The bioconjugates consist of four key
moieties, namely (i) an electroactive core (ii) a conjugate linker (iii) an alkyl group and (iv) a
dipeptide ester. These novel derivatives differ from the previously reported N-meta and N-ortho-
(ferrocenyl)benzoyl dipeptide esters by having an alkyl group on the previously unsubstituted
cyclopentadiene ring [19,20]. The 1-alkyl-1’-N-meta and N-ortho-(ferrocenyl)benzoyl dipeptide
esters were prepared by coupling the methyl, ethyl and propyl ferrocenyl benzoic acids 1-6 to the
dipeptide ethyl esters using the conventional N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide
(EDC), 1-hydroxybenzotriazole (HOBt) protocol. The dipeptide esters employed in the synthesis
were Gly-Gly(OEt), Gly-L-Ala(OEt), Gly-L-Leu(OEt) and Gly-L-Phe(OEt). All the compounds
gave spectroscopic data in accordance with the proposed structures. In addition, the in vitro anti-
cancer activity of the bioconjugates against the human lung carcinoma cell line H1299 was
determined.
2. Results and Discussion
2.1. Synthesis
2.1.1. Synthesis of 1-alkyl-1’-N-meta-(ferrocenyl)benzoyl dipeptide esters 7-18 and 1-alkyl-1’-N-
ortho-(ferrocenyl)benzoyl dipeptide esters 19-30.
1-Alkyl-1’-N-meta-(ferrocenyl) benzoic acids 1-3 were prepared by coupling either methyl, ethyl
or propyl ferrocene to 3-amino benzoic acid via diazonium coupling whereas the 1-alkyl-1’-N-
ortho-(ferrocenyl) benzoic acids 4-6 were prepared by coupling either methyl, ethyl or propyl
ferrocene to 2-amino benzoic acid via diazonium coupling. The free N-terminal dipeptide ethyl
esters Gly-Gly(OEt), Gly-L-Ala(OEt), Gly-L-Leu(OEt) and Gly-L-Phe(OEt), were coupled to the
alkyl ferrocenyl benzoic acids 1-6 using EDC and HOBt in the presence of excess triethylamine
in dichloromethane (Scheme 1). EDC was used in preference to the less expensive coupling
reagent N, N'-dicyclohexylcarbodiimide (DCC) as its reaction by-products are easier to remove
compared to those of DCC, namely dicyclohexylurea (DCU). Purification by column
chromatography furnished the pure products. The 1-alkyl-1’-N-meta-(ferrocenyl)benzoyl
dipeptide esters 7-18 were obtained in yields of 14-25% and the 1-alkyl-1’-N-ortho-
(ferrocenyl)benzoyl dipeptide esters 19-30 in yields of 12-23%. The chemical structures of
compouds 7-18 are presented in table 1 and compounds 19-30 in table 2.

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The relatively low yields are partly due to the coupling procedure. The first step in amide bond
formation in the coupling protocol is formation of the O-acylisourea ester intermediate. This
intermediate is highly reactive, therefore side-reactions can pose a serious problem and can result
in extensive racemisation resulting in low yields. The addition of HOBt stabilizes the O-
acylisourea ester intermediate thus supressing side reactions, however the addition does not
result in 100 % suppression. As the reaction proceeds, upon addition of the dipeptide ethyl esters
the HOBt is displaced resulting in the formation of compounds 7-30. Due to the complexity of
the reaction which is associated with the competing reactions, low yields for compounds 7-30
were obtained. All of the compounds gave spectroscopic data in accordance with the proposed
structures.
2.2. 1H and 13C Spectroscopic analysis
All the proton and carbon chemical shifts for compounds 7-30 were unambiguously assigned by
1
1
a combination of DEPT-135 and H-13C COSY (HMQC). The H and 13C NMR spectra for
compounds 7-30 showed peaks in the ferrocene region characteristic of a disubstituted ferrocene
moiety [29-31]. For the cyclopentadiene ring attached to the benzoyl spacer moiety (η5C5H4-
benzoyl), the ortho protons appear as either singlets or triplets between δ 4.85 and δ 4.68, whilst
the meta protons appear as either singlets or triplets between δ 4.38 and δ 4.2. The protons on
the alkylated cyclopentadiene ring (η5-C5H4-alkyl) overlap with the signals of the methylene
groups of the glycine moiety, resulting in a multiplet being observed between δ 3.98 to δ 3.79.
For the alkyl groups attached to the cyclopentadiene ring, the methyl (-CH3) group proton signals
are present between δ 1.69 to δ 1.45. The methylene and methyl protons of the ethyl group
appear as a quartet between δ 2.07 to δ 1.96 and as a triplet between δ 1.03 to δ 0.7 respectively.
For the propyl group the protons on the methylene groups (-CH2CH2CH3) appear as a triplet
between δ 2.08 to δ 1.88 and as a multiplet between δ 1.4 to δ 1.23 whilst, the protons on the
methyl group (-CH2CH2CH3) appear as a triplet between δ 0.76 to δ 0.7.
13
The methyl group on the cyclopentadiene ring appears in the range δ 14.5 to δ 13.0 in the C
NMR spectra of compounds 7-10 and 19-22. The methylene and methyl carbon atoms of the
ethyl group appear in the range δ 25.8 to δ 20.8 and δ 14.2 to δ 13.0 respectively for compounds
11-14 and 23-26. For the propyl derivatives 15-18 and 27-30 the methylene group attached to
the cyclopentadiene ring appears in the range δ 33.8 to δ 31.0. The other methylene group
appears in the range δ 25.0 to δ 20.6 whereas the methyl group is present between δ 14.1 and δ

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13
13.1 The C NMR spectra of compounds 7-30 show signals in the region between δ 93.3 to δ
66.1 indicative of a disubstituted ferrocene unit. The ipso carbon atom on the (η5-C5H4-alkyl)
ring appears between δ 93.3 to δ 86.4 whilst the ipso on the (η5-C5H4-benzoyl) ring appears in
the range δ 85.6 to δ 80.0. Both of these signals are absent in the DEPT-135 spectra. The carbon
atoms on the benzoyl linker for both the meta and ortho series are non-equivalent therefore six
signals are observed. These appear between δ 142.5 and δ 123.2 for compounds 7-30. The
quaternary carbon atoms of the aromatic rings and the methylene carbon atoms of derivatives 7-
30 were identified by DEPT-135. Complete spectroscopic data for all the compounds is
presented in the experimental section.
2.3. In vitro anti-cancer activity of 7-30
The in vitro anti-proliferative effect of compounds 7-30 was studied in the H1299 non-small cell
lung cancer (NSCLC) cell line. Proliferation was measured using the acid phosphatase assay as
previously described [32]. In a preliminary screen the cells were treated with the 1-alkyl-1’-N-
meta and N-ortho-(ferrocenyl) benzoyl dipeptide esters 7-30 at a concentration of 10 µM and
were incubated for 5-6 days until cell confluency reached 80-90%. Cell survival was established
through determination of the acid phosphatase activity of surviving cells and growth inhibition
calculated relative to controls (untreated cells). The results for the preliminary screen are
depicted in table 3 for compounds 7-18 and in table 4 for compounds 19-30. The 1-alkyl-1’-N-
meta-(ferrocenyl) benzoyl dipeptide esters 7-18 displayed % growth inhibition in the range 20-
84%, whereas the 1-alkyl-1’-N-ortho-(ferrocenyl) benzoyl dipeptide esters 19-30 displayed %
growth inhibition in the range 14-48%. Because the 1-alkyl-1’-N-ortho-(ferrocenyl) benzoyl
dipeptide esters 19-30 displayed such poor % growth inhibition values at a concentration of 10
µM, they were not selected for further evaluation. We previously reported that N-{ortho-
(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester displayed an IC50 value of 5.3 µM [20].
Therefore alkylation of the the previously unsubstituted cyclopentadiene ring results in a
substantial decrease in the cytotoxicity of the 1-alkyl-1’-N-ortho-(ferrocenyl) benzoyl dipeptide
esters 19-30. The six most active compounds of the 1-alkyl-1’-N-meta-(ferrocenyl) benzoyl
dipeptide esters 7-18 which showed % growth inhibition values ≥ 70 % were selected for further
evaluation. A general trend was also observed in the 1-alkyl-1’-N-meta-(ferrocenyl) benzoyl
dipeptide derivatives, that is, the Gly-Gly(OEt) and the Gly-L-Ala(OEt) derivatives are more
active than the Gly-L-Leu and Gly-L-Phe ethyl esters which display % growth inhibition values

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≤ 55 %. For instance 1-methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 7
displayed a % growth inhibition value of 84 ± 3.8% whereas the 1-methyl-1’-N-{meta-
(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl ester 9 and the 1-methyl-1’-N-{meta-(ferrocenyl)-
benzoyl}-glycine-L-phenylalanine ethyl ester 10 displayed % growth inhibition values of 41 ±
1.0% and 20 ± 2.9% respectively. Therefore the 1-methyl, 1-ethyl and 1-propyl-1’-N- {meta-
(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl esters and the 1-methyl, 1-ethyl and 1-propyl-1’-
N-{meta-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester derivatives were not
investigated further. A loss of anti-proliferative activity is observed when chiral α-amino acids
with bulky side chains are used as the second amino acid in the dipeptide moiety. The IC50
values were determined for compounds 7, 8, 11, 12, 15 and 16. To determine the IC50 values of
the six compounds, individual 96-well plates containing H1299 cells were treated with the test
compound at concentrations ranging from 0.1 ꢀM to 100 ꢀM. The cells were then incubated for
5-6 days, until cell confluency was reached. Cell survival was determined by performing the acid
phosphatase assay. The IC50 value for each compound was calculated using Calcusyn software,
and standard deviations were calculated using data obtained from three independent experiments.
The IC50 values obtained are listed in table 5. From the IC50 values of the 1-alkyl-1’-N-{meta-
(ferrocenyl)-benzoyl}-glycine-glycine ethyl esters 7, 11 and 15 and the 1-alkyl-1’-N-{meta-
(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl esters 8, 12 and 16 it can be seen that the
cytotoxicity of the derivatives decreases with an increase of the size of the alkyl group
incorporated (propyl < ethyl < methyl). 1-Methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-
alanine ethyl ester 8 has an IC50 of 6.7 µM. We previously reported that N-{meta-(ferrocenyl)-
benzoyl}-glycine-L-alanine ethyl ester displayed an IC50 value of 4.0 µM [19]. Therefore
alkylation of the the previously unsubstituted cyclopentadiene ring results in a slight decrease in
the cytotoxicity of the 1-alkyl-1’-N-meta-(ferrocenyl) benzoyl dipeptide esters.
The in vitro cytotoxicity of the platinum(II)-based anti-cancer drug carboplatin 31 was also
evaluated against the H1299 cell line, and was found to have an IC50 value of 10.0 ± 1.60 µM
(Table 5). Thus compounds 7, 8 and 11 are more cytotoxic in vitro than the clinically employed
anti-cancer drug carboplatin. The most cytotoxic compound was 1-methyl-1’-N-{meta-
(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 7 which displayed an IC50 value of 2.6 µM
(Figure 1). Although compound 7 is the most active derivative of the 1-alkyl-1’-N-meta-
(ferrocenyl) benzoyl dipeptide esters it is less active compared to cisplatin 32 which displays an

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IC50 value of 1.5 µM against human H1299 lung carcinoma cells. These novel organometallic
anticancer compounds may induce DNA damage by the catalytic generation of reactive
oxygenated species (ROS) via a Fenton-type reaction, in which HO• radicals are generated from
the superoxide dismutation product, hydrogen peroxide (H2O2). In a previous study it was
shown that N-(6-ferrocenyl-2-naphthoyl)-glycine-glycine ethyl ester was capable of causing
oxidative damage to guanine via a ROS mediated mechanism. Cell cycle analysis in the H1299
cell line also suggested that these compounds induced apoptosis [25].
3. Conclusion
In conclusion, the novel 1-alkyl-1’-N-meta-(ferrocenyl)benzoyl dipeptide esters and 1-alkyl-1’-
N-ortho-(ferrocenyl)benzoyl dipeptide esters were synthesized. All the compounds gave
spectroscopic data in accordance with the proposed structures. The 1-alkyl-1’-N-meta-
(ferrocenyl)benzoyl dipeptide esters 7, 8, 11, 12, 15 and 16 showed micromolar activity in the
H1299 NSCLC cell line, with IC50 values I the range 2.6 to 20.1 ꢀM. The cytotoxicity of the
derivatives decreases with an increase of the size of the alkyl group on the previously
unsubstituted cyclopentadiene ring.
Alkylation of the the previously unsubstituted
cyclopentadiene ring results in a decrease in the cytotoxicity of both the 1-alkyl-1’-N-meta-
(ferrocenyl)benzoyl dipeptide esters and the 1-alkyl-1’-N-ortho-(ferrocenyl) benzoyl dipeptide
esters. This is in contrast to the 1-alkyl-1’-N-para-(ferrocenyl)benzoyl dipeptide esters whereby
alkylation of the the previously unsubstituted cyclopentadiene ring resulted in an increase in
cytotoxicity [26].
4. Experimental
4.1. General Procedures
All chemicals were purchased from Sigma-Aldrich, Lennox Chemicals, Fluorochem Limited or
Tokyo Chemical Industry UK Limited; and used as received. Commercial grade reagents were
used without further purification. When necessary, all solvents were purified and dried prior to
use. Riedel-Haën silica gel was used for thin layer and column chromatography. Melting points
were determined using a Stuart melting point (SMP3) apparatus and are uncorrected. Optical
rotation measurements were made on a Perkin Elmer 343 Polarimeter and are quoted in units of
10-1 deg cm2 g-1. Infrared spectra were recorded on a Perkin Elmer Spectrum 100 FT-IR with

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ATR. UV-Vis spectra were recorded on a Hewlett Packard 8452 A diode array UV-Vis
1
13
spectrophotometer. H and C NMR spectra were recorded in deuterated solvents on a Bruker
Avance 400 NMR. The 1H and 13C NMR chemical shifts are reported in ppm (parts per million).
Tetramethylsilane (TMS) or the residual solvent peaks have been used as an internal reference.
All coupling constants (J) are in Hertz. Electrospray ionisation mass spectra were performed on a
Micromass LCT mass spectrometer.
Cyclic voltammograms were recorded in anhydrous acetonitrile (Sigma-Aldrich), with 0.1 M
tetrabutylammonium hexafluorophosphate (TBAPF6) as a supporting electrolyte, using a CH
Instruments 600a electrochemical analyzer (Pico-Amp Booster and Faraday Cage). The
experiments were carried out at room temperature. A three-electrode cell consisting of a glassy
carbon working-electrode, a platinum wire counter-electrode and an Ag|AgCl reference electrode
was used. The glassy carbon electrode was polished with 0.3 ꢀm alumina followed by 0.05 ꢀm
alumina, between each experiment to remove any surface contaminants. The scan rate was 0.1 V
s-1. The concentration range of the ferrocene compounds was 1.0 mM in acetonitrile. The E0´
values obtained for the test samples were referenced relative to the ferrocene/ferricenium redox
couple.
4.2. General Procedure for the synthesis of the starting materials
4.2.1. 1-Methyl-1’-meta-ferrocenyl benzoic acid 1.
3-Aminobenzoic acid (3.43 g, 25.00 mmol) and methylferrocene (5.00 g, 25.00 mmol) were used
as starting materials. The title product was isolated by filtration and recrystallisation from
chloroform furnished the desired product as an orange solid (1.46 g, 18 %).
1H NMR (400MHz) δ (DMSO- d6): 12.85 (1H, s, -COOH), 8.00 (1H, s, ArH), 7.69 (2H, m,
ArH), 7.36 (1H, t, J = 7.2 Hz, ArH), 4.87 (2H, s, ortho on η5-C5H4-benzoyl), 4.41 (2H, s, meta
on η5-C5H4-benzoyl), 3.96 (4H, dd, η5-C5H4-alkyl), 1.62 ( 3H, s, -CH3).
13C NMR (100MHz) δ (DMSO- d6): 170.3 (C=O), 136.5 (Cq), 135.8 (Cq), 129.4, 128.2, 127.3,
125.1, 90.0 (Cipso η5-C5H4-alkyl), 83.2 (Cipso η5-C5H4-benzoyl), 72.2 (Cmeta η5-C5H4-benzoyl),
68.8 (Cmeta η5-C5H4-alkyl), 68.0 (Cortho η5-C5H4-alkyl), 67.3 (Cortho η5-C5H4-benzoyl), 13.9 (-
CH3).

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4.2.2. 1-Ethyl-1’-meta-ferrocenyl benzoic acid 2.
3-Aminobenzoic acid (3.20 g, 23.36 mmol) and ethylferrocene (5.00 g, 23.36 mmol)
were used as starting materials. The title product was isolated by filtration and recrystallisation
from chloroform furnished the desired product as an orange solid (1.48 g, 19 %).
1H NMR (400MHz) δ (DMSO- d6): 12.85 (1H, s, -COOH), 8.05 (1H, s, ArH), 7.70 - 7.66 (2H,
m, ArH), 7.39 (1H, t, J = 7.2 Hz, ArH), 4.89 (2H, t, J =1.6 Hz, ortho on η5-C5H4-benzoyl), 4.41
(2H, t, J =1.6 Hz, meta on η5-C5H4-benzoyl), 3.95 (4H, dd, η5-C5H4-alkyl), 2.31 (2H, q, J =7.6
Hz, -CH2CH3), 1.14 (3H, t, J = 7.6 Hz, -CH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 169.3 (C=O), 139.5 (Cq), 135.8 (Cq), 132.4, 128.7, 127.7,
125.9, 91.6 (Cipso η5-C5H4-alkyl), 81.7 (Cipso η5-C5H4-benzoyl), 71.2 (Cmeta η5-C5H4-benzoyl),
68.8 (Cmeta η5-C5H4-alkyl), 68.6 (Cortho η5-C5H4-alkyl), 66.9 (Cortho η5-C5H4-benzoyl), 23.0 (-
CH2CH3,-ve DEPT), 13.5 (-CH2CH3).
4.2.3. 1-Propyl-1’-meta-ferrocenyl benzoic acid 3.
3-Aminobenzoic acid (3.01 g, 21.93 mmol) and propyl ferrocene (5.00 g, 21.93 mmol)
were used as starting materials. The title product was isolated by filtration and recrystallisation
from chloroform furnished the desired product as an orange solid (1.48 g, 19 %).
1H NMR (400MHz) δ ( DMSO- d6 ): 12.85 (1H, s, -COOH), 8.00 (1H, s, ArH), 7.65 (2H, m,
ArH), 7.30 (1H, t, J = 7.2 Hz, ArH), 4.89 (2H, s, ortho on η5-C5H4-benzoyl), 4.41 (2H, s, meta
on η5-C5H4-benzoyl), 3.95 (4H, dd, η5 C5H4-alkyl), 2.31 (2H, t, J = 7.6 Hz, -CH2CH2CH3), 1.54 -
1.45 (2H, m, -CH2CH2CH3), 0.92 (3H, t, J = 7.2 Hz, -CH2CH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 168.3 (C=O), 139.5 (Cq), 136.8 (Cq), 132.4, 128.9, 127.5,
125.7, 89.6 (Cipso η5-C5H4-alkyl), 82.7 (Cipso η5-C5H4-benzoyl), 72.2 (Cmeta η5-C5H4-benzoyl),
68.9 (Cmeta η5-C5H4-alkyl), 68.6 (Cortho η5-C5H4-alkyl), 67.0 (Cortho η5-C5H4-benzoyl), 31.3 (-
CH2CH2CH3, -ve DEPT), 22.7 (-CH2CH2CH3, -ve DEPT), 14.4 (-CH2CH2CH3).
4.2.4. 1-Methyl-1’-ortho-ferrocenyl benzoic acid 4.
2-Aminobenzoic acid (3.43 g, 25.00 mmol) and methylferrocene (5.00 g, 25.00 mmol) were used
as starting materials. The title product was isolated by filtration and recrystallisation from
chloroform furnished the desired product as an orange solid (1.26 g, 16%).

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1H NMR (400 MHz) δ (DMSO- d6): 12.85 (1H, s, -COOH), 7.95 (1H, d, J = 7.6 Hz, ArH), 7.77
(1H, t, J = 7.2 Hz, ArH), 7.65 (1H, t, J = 6.4 Hz, ArH), 7.40 (1H, d, J = 7.6 Hz, ArH), 4.89 (2H,
s, ortho on η5-C5H4-benzoyl), 4.41 (2H, s, meta on η5-C5H4-benzoyl), 3.95 (4H, dd, η5-C5H4-
alkyl), 1.62 ( 3H, s, -CH3).
13C NMR (100 MHz) δ (DMSO- d6): 171.3 (C=O), 140.3 (Cq), 137.5 (Cq), 130.4, 128.7, 126.3,
125.9, 92.6 (Cipso η5-C5H4-alkyl), 80.7 (Cipso η5-C5H4-benzoyl), 73.9 (Cmeta η5-C5H4-benzoyl),
68.2 (Cmeta η5-C5H4-alkyl), 67.9 (Cortho η5-C5H4-alkyl), 67.0 (Cortho η5-C5H4-benzoyl), 13.5 (-
CH3).
4.2.5.1-Ethyl-1’-ortho-ferrocenyl benzoic acid 5.
2-Aminobenzoic acid (3.20 g, 23.36 mmol) and ethylferrocene (5.00 g, 23.36 mmol)
were used as starting materials. The title product was isolated by filtration and recrystallisation
from chloroform furnished the desired product as an orange solid (1.29 g, 17%).
1H NMR (400 MHz) δ (DMSO- d6): 12.85 (1H, s, -COOH), 7.97 (1H, d, J = 7.6 Hz, ArH), 7.74
(1H, t, J = 7.2 Hz, ArH), 7.62 (1H, t, J = 6.4 Hz, ArH), 7.44 (1H, d, J = 7.6 Hz, ArH), 4.89 (2H,
t, J =1.6 Hz, ortho on η5-C5H4-benzoyl), 4.41 (2H, t, J =1.6 Hz, meta on η5-C5H4-benzoyl), 3.95
(4H, dd, η5-C5H4-alkyl), 2.32 (2H, q, J =7.6 Hz, -CH2CH3), 1.13 (3H, t, J = 7.6 Hz, -CH2CH3);
13C NMR (100 MHz) δ (DMSO- d6):169.3 (C=O), 140.3 (Cq), 135.8 (Cq), 131.4, 127.2, 126.3,
125.1, 89.6 (Cipso η5-C5H4-alkyl), 80.7 (Cipso η5-C5H4-benzoyl), 70.2 (Cmeta η5-C5H4-benzoyl),
68.8 (Cmeta η5-C5H4-alkyl), 68.6 (Cortho η5-C5H4-alkyl), 67.0 (Cortho η5-C5H4-benzoyl), 22.7 (-
CH2CH3, -ve DEPT), 14.0 (-CH2CH3).
4.2.6. 1-Propyl-1’-ortho-ferrocenyl benzoic acid 6.
2-Aminobenzoic acid (3.01 g, 21.93 mmol) and propyl ferrocene (5.00 g, 21.93 mmol)
were used as starting materials. The title product was isolated by filtration and recrystallisation
from chloroform furnished the desired product as an orange solid (1.26 g, 17%).
1H NMR (400 MHz) δ (DMSO- d6): 12.85 (1H, s, -COOH), 7.95 (1H, d, J = 7.6 Hz, ArH), 7.77
(1H, t, J = 7.2 Hz, ArH), 7.65 (1H, t, J = 6.4 Hz, ArH), 7.40 (1H, d, J = 7.6 Hz, ArH), 4.89 (2H,
s, ortho on η5-C5H4-benzoyl), 4.41 (2H, s, meta on η5-C5H4-benzoyl), 3.95 (4H, dd, η5-C5H4-

ACCEPTED MANUSCRIPT
alkyl), 2.31 (2H, t, J = 7.6 Hz, -CH2CH2CH3), 1.54 - 1.45 (2H, m, -CH2CH2CH3), 0.92 (3H, t, J
= 7.2 Hz, -CH2CH2CH3);
13C NMR (100 MHz) δ (DMSO- d6): 169.3 (C=O), 138.5 (Cq), 135.8 (Cq), 130.4, 128.2, 126.3,
124.1, 90.6 (Cipso η5-C5H4-alkyl), 82.7 (Cipso η5-C5H4-benzoyl), 71.2 (Cmeta η5-C5H4-benzoyl),
68.7 (Cmeta η5-C5H4-alkyl), 68.2 (Cortho η5-C5H4-alkyl), 67.1 (Cortho η5-C5H4-benzoyl), 34.3 (-
CH2CH2CH3, -ve DEPT), 22.0 (-CH2CH2CH3, -ve DEPT), 14.0 (-CH2CH2CH3).
4.3. General procedure for the synthesis of 1-alkyl-1’-N-ferrocenyl benzoyl dipeptide ethyl esters
4.3.1. 1-Methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 7.
1-Methyl-1’-N-meta-ferrocenyl benzoic acid (0.25 g, 0.78 mmol) was dissolved in
dichloromethane (100 ml) at
0
oC. N-(3-dimethylaminopropyl)-N’-ethylcarbodiimide
hydrochloride (0.15 g, 0.78 mmol), 1-hydroxybenotriazole (0.11 g, 0.78 mmol), glycine glycine
ethyl ester hydrochloride (0.13 g, 0.78 mmol) and triethylamine (3 ml) were added and the
reaction mixture was allowed to stir at 0 oC for 45 min. The reaction mixture was then allowed to
stir at room temperature for 48 h. The reaction mixture was then washed with water and brine.
The organic layer was dried over MgSO4. The solvent was removed in vacuo to yield the crude
product which was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the the desired product as a red oil (0.09 g, 25 %), Eº´ = 10 mV (vs Fc/Fc+).
Mass spectrum: [M+Na]+ found: 499.1286
C24H26N2O4FeNa requires:
485.1291.
I.R. υmax (KBr): 3350 (NH), 1751 (C=Oester), 1627 (C=Oamide), 1602 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 332, 440 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.65 (1H, t, J = 6 Hz, -CONH-), 8.30 (1H, t, J = 7.6 Hz, -
CONH-), 8.00 (1H, s, ArH), 7.69 - 7.67 (2H, m, ArH), 7.39 (1H, t, J = 7.2 Hz, ArH), 4.80 (2H,
s, ortho on η5-C5H4-benzoyl), 4.31 (2H, s, meta on η5-C5H4-benzoyl), 4.10 (2H, q, J = 7.2 Hz, -
OCH2CH3) , 3.91 - 3.82 {8H, m, (η5-C5H4-alkyl), (-NHCH2CHO-), (-NHCH2CHO-)}, 1.61 (3H,
s, -CH3), 1.18 (3H, t, J = 7.2 Hz, -OCH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 170.5 (C=O), 167.2 (C=O), 166.5 (C=O), 136.5 (Cq), 135.8
(Cq), 129.4, 128.2, 127.3, 125.1, 92.3 (Cipso η5-C5H4-alkyl), 82.0 (Cipso η5-C5H4-benzoyl), 70.4
(Cmeta η5-C5H4-benzoyl), 69.1 (Cmeta η5-C5H4-alkyl), 68.4 (Cortho η5-C5H4-alkyl), 66.2 (Cortho η5-

ACCEPTED MANUSCRIPT
C5H4-benzoyl), 61.5 (-OCH2CH3, -ve DEPT), 42.6 (-NHCH2CO-, -ve DEPT), 40.7 (-
NHCH2CO-, -ve DEPT), 14.5 (-OCH2CH3), 14.2 (-CH3).
4.3.2. 1-Methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester 8.
Glycine-L-alanine ethyl ester hydrochloride (0.14 g, 0.78 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
yielding the title product as a red oil (0.08 g, 22 %), Eº´ = -30 mV (vs Fc/Fc+), [α]D20 = -13o (c
0.1, EtOH).
Mass spectrum: [M+Na]+ found: 499.1391
C25H28N2O4FeNa requires:
499.1396.
I.R. υmax (KBr): 3311 (NH), 1751 (C=Oester), 1620 (C=Oamide), 1601 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 333, 447 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.65 (1H, t, J = 6 Hz, -CONH-), 8.30 (1H, d, J = 7.6 Hz, -
CONH-), 8.05 (1H, s, ArH), 7.70 - 7.69 (2H, m, ArH), 7.39 (1H, t, J = 7.2 Hz, ArH), 4.81 (2H,
s, ortho on η5-C5H4-benzoyl), 4.34 (2H, s, meta on η5-C5H4-benzoyl), 4.30 - 4.24 (1H, m, -
CHCH3), 4.00 (2H, q, J = 6.6 Hz, -OCH2CH3) , 3.92 - 3.84 {6H, m, (η5-C5H4-alkyl), (-
NHCH2CO-)}, 1.59 (3H, s, -CH3), 1.31 (3H, d, J = 6.2 Hz, -CHCH3), 1.15 (3H, t, J = 6.6 Hz, -
OCH2CH3);
13C NMR (100MHz) δ (DMSO- d6): 170.5 (C=O), 167.0 (C=O), 166.9 (C=O), 136.5 (Cq), 135.8
(Cq), 129.0, 128.2, 126.3, 125.1, 90.3 (Cipso η5-C5H4-alkyl), 81.0 (Cipso η5-C5H4-benzoyl), 70.9
(Cmeta η5-C5H4-benzoyl), 68.4 (Cmeta η5-C5H4-alkyl), 67.5 (Cortho η5-C5H4-alkyl), 66.4 (Cortho η5-
C5H4-benzoyl), 60.8 (-OCH2CH3, -ve DEPT), 48.5 (-CHCH3), 42.2 (-NHCH2CO-, -ve DEPT),
17.0 (-CHCH3), 14.9 (-OCH2CH3), 14.5 (-CH3).
4.3.3. 1-Methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl ester 9.
Glycine-L-leucine ethyl ester hydrochloride (0.17 g, 0.78 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
20
yielding the title product as a red oil (0.07 g, 17 %), Eº´ = 30 mV (vs Fc/Fc+), [α]D = -19o (c
0.1, EtOH).
Mass spectrum: [M+Na]+ found: 541.1883
C28H34N2O4FeNa requires:
541.1888.

ACCEPTED MANUSCRIPT
I.R. υmax (KBr): 3287 (NH), 1742 (C=Oester), 1649 (C=Oamide), 1610 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 330, 445 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.65 (1H, t, J = 6 Hz, -CONH-), 8.30 (1H, d, J = 7.6 Hz, -
CONH-), 8.00 (1H, s, ArH), 7.69 - 7.66 (2H, m, ArH), 7.39 (1H, t, J = 7.0 Hz, ArH) 4.83 (2H,
s, ortho on η5-C5H4-benzoyl), 4.36 (2H, s, meta on η5-C5H4-benzoyl), 4.35 - 4.27 {1H, m, -
CH(CH2CH(CH3)2)}, 4.15 (2H, q, J = 7.2 Hz, -OCH2CH3) , 3.92 - 3.86 {6H, m, (η5-C5H4-
alkyl), (-NHCH2CO-)}, 1.69 - 1.45 {6H, m, ( -CH3), (-CH(CH2CH(CH3)2)}, 1.15 (3H, t, J = 7.2
Hz, -OCH2CH3), 0.90 - 0.85 {6H, m, -CH(CH2CH(CH3)2}.
13C NMR (100MHz) δ (DMSO- d6): 171.7 (C=O), 169.2 (C=O), 166.8 (C=O), 136.5 (Cq), 135.8
(Cq), 129.4, 128.2, 127.3, 126.1, 86.4 (Cipso η5-C5H4-alkyl), 82.1 (Cipso η5-C5H4-benzoyl), 71.4
(Cmeta η5-C5H4-benzoyl), 70.1 (Cmeta η5-C5H4-alkyl), 68.1 (Cortho η5-C5H4-alkyl), 66.5 (Cortho η5-
C5H4-benzoyl), 61.0 (-OCH2CH3, -ve DEPT), 51.0 {-CH(CH2CH(CH3)2)}, 42.7 (-NHCH2CO-, -
ve DEPT), 39.5 {-CH(CH2CH(CH3)2), -ve DEPT}, 25.0{-CH(CH2CH(CH3)2)}, 22.7 {-
CH(CH2CH(CH3)2)}, 21.4 {-CH(CH2CH(CH3)2)}, 14.9 (-OCH2CH3), 13.0 (-CH3).
4.3.4. 1-Methyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester 10.
Glycine-L-phenyalanine ethyl ester hydrochloride (0.20 g, 0.78 mmol) was used as a starting
material. The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl
20
acetate} yielding the title product as a red oil (0.06 g, 14 %), Eº´ = -10 mV (vs Fc/Fc+), [α]D
=
+8 o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 575.1774
C31H32N2O4FeNa requires:
575.1766.
I.R. υmax (KBr): 3273 (NH), 1729 (C=Oester), 1653 (C=Oamide), 1599 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 333, 449 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.68 (1H, t, J = 6.0 Hz, -CONH-), 8.32 (1H, d, J = 7.6 Hz,
-CONH-), 7.99 (1H, s, ArH), 7.71 - 7.69 (2H, m, ArH), 7.40 (1H, t, J = 7.0 Hz, ArH), 7.20 -
7.17 {5H, m, -CH(CH2Ph)}, 4.83 (2H, s, ortho on η5-C5H4-benzoyl), 4.45 - 4.40 {1H, m, -
CH(CH2Ph)}, 4.35 (2H, s, meta on η5-C5H4-benzoyl), 4.04 (2H, q, J = 7.2 Hz, -OCH2CH3) ,
3.95 - 3.87 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 3.06 - 2.93 {2H, m, -CH(CH2Ph)}, 1.55
(3H, s, -CH3), 1.12 ( 3H, t, J = 7.2 Hz, -OCH2CH3).

ACCEPTED MANUSCRIPT
13C NMR (100MHz) δ (DMSO- d6): 170.4 (C=O), 169.2 (C=O), 166.3 (C=O), 137.2 (Cq), 134.9
(Cq), 130.3 (Cq), 129.1, 128.4, 127.3, 126.1, 125.5, 124.1, 122.7, 91.2 (Cipso η5-C5H4-alkyl), 80.0
(Cipso η5-C5H4-benzoyl), 70.4 (Cmeta η5-C5H4-benzoyl), 70.0 (Cmeta η5-C5H4-alkyl), 68.4 (Cortho η5-
C5H4-alkyl), 67.9 (Cortho η5-C5H4-benzoyl), 60.8 (-OCH2CH3, -ve DEPT), 53.0 {-CH(CH2Ph)},
42.7 (-NHCH2CO-, -ve DEPT), 37.7 {-CH(CH2Ph), -ve DEPT}, 14.1 (-OCH2CH3), 13.0 (-CH3).
4.3.5. 1-Ethyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 11.
Glycine-glycine ethyl ester hydrochloride (0.12 g, 0.75 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.09 g, 25 %), Eº´ = -40 mV (vs Fc/Fc+).
Mass spectrum: [M+Na]+ found: 499.1387
C25H28N2O4FeNa requires:
499.1396.
I.R. υmax (KBr): 3311 (NH), 1741 (C=Oester), 1622 (C=Oamide), 1601 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 332, 440 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.49 (1H, t, J = 6 Hz, -CONH-), 8.23 (1H, t, J = 6 Hz, -
CONH-), 7.87 (1H, s, ArH), 7.67 - 7.63 (2H, m, ArH), 7.36 (1H, t, J = 7.2 Hz, ArH), 4.85 (2H,
t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.20 (2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl),
4.11 (2H, q, J = 7.2 Hz, - OCH2CH3), 3.98 - 3.87 {8H, m, (η5-C5H4-alkyl), (-NHCH2CO-), (-
NHCH2CO-)}, 1.96 (2H, q, J = 7.6 Hz, -CH2CH3), 1.26 (3H, t, J = 7.2 Hz, -OCH2CH3), 1.03
(3H, t, J = 7.6 Hz, -CH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 169.8 (C=O), 168.7 (C=O), 166.3 (C=O), 139.6 (Cq), 134.1
(Cq), 128.8, 128.5, 124.2, 124.1, 91.2 (Cipso η5-C5H4-alkyl), 83.1 (Cipso η5-C5H4-benzoyl), 70.0
(Cmeta η5-C5H4-benzoyl), 68.8 (Cmeta η5-C5H4-alkyl), 68.5 (Cortho η5-C5H4-alkyl), 66.2 (Cortho η5-
C5H4-benzoyl), 60.3 (-OCH2CH3, -ve DEPT), 42.4 (-NHCH2CO-, -ve DEPT), 40.6 (-
NHCH2CO-, -ve DEPT), 20.8 (-CH2CH3, -ve DEPT), 14.4 (-OCH2CH3), 14.0 (-CH2CH3).
4.3.6. 1-Ethyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester 12.
Glycine-L-alanine ethyl ester hydrochloride (0.13 g, 0.75 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
20
yielding the title product as a red oil (0.08 g, 22 %), Eº´ = 10 mV (vs Fc/Fc+), [α]D = -18o (c
0.1, EtOH).

ACCEPTED MANUSCRIPT
Mass spectrum: [M+Na]+ found: 513.1458
C25H28N2O4FeNa requires: 513.1453.
I.R. υmax (KBr): 3290 (NH), 1739 (C=Oester), 1635 (C=Oamide), 1605 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 333, 444 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.60 (1H, t, J = 6 Hz, -CONH-), 8.40 (1H, d, J = 7.0 Hz, -
CONH-), 8.00 (1H, s, ArH), 7.68 - 7.64 (2H, m, ArH), 7.40 (1H, t, J = 7.2 Hz, ArH), 4.68 (2H,
t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.34 (2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl),
4.30 - 4.24 (1H, m, - CHCH3), 4.00 (2H, q, J = 6.8 Hz, - OCH2CH3), 3.96 - 3.82 {6H, m, (η5-
C5H4-alkyl), (-NHCH2CO-)}, 2.05 (2H, q, J = 7.6 Hz, -CH2CH3), 1.28 (3H, d, J = 6.4 Hz, -
CHCH3), 1.20 (3H, t, J = 6.8 Hz, -OCH2CH3), 0.96 (3H, t, J = 7.6 Hz, -CH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 172.5 (C=O), 169.0 (C=O), 166.2 (C=O), 137.5 (Cq), 135.8
(Cq), 130.4, 129.2, 127.0, 125.2, 91.0 (Cipso η5-C5H4-alkyl), 84.0 (Cipso η5-C5H4-benzoyl), 71.5
(Cmeta η5-C5H4-benzoyl), 69.3 (Cmeta η5-C5H4-alkyl), 68.1 (Cortho η5-C5H4-alkyl), 66.5 (Cortho η5-
C5H4-benzoyl), 62.4 (-OCH2CH3, -ve DEPT), 47.6 (CHCH3), 42.8 (-NHCH2CO-, -ve DEPT),
21.8 (-CH2CH3, -ve DEPT), 19.0 (-CHCH3), 14.1 (-OCH2CH3), 13.5 (-CH2CH3).
4.3.7. 1-Ethyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl ester 13.
Glycine-L-leucine ethyl ester hydrochloride (0.16 g, 0.75 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.07 g, 18 %), Eº´ = -10 mV (vs Fc/Fc+), [α]D20 = -13o
(c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 555.1933
C29H36N2O4FeNa requires:
555.1922.
I.R. υmax (KBr): 3260 (NH), 1737 (C=Oester), 1643 (C=Oamide), 1615 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 333, 445 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.68 (1H, t, J = 6 Hz, -CONH-), 8.34 (1H, d, J = 6.8 Hz, -
CONH-), 8.01 (1H, s, ArH), 7.65 - 7.62 (2H, m, ArH), 7.45 (1H, t, J = 7.0 Hz, ArH), 4.78 (2H,
t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.30 (2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl),
4.34 - 4.26 {1H, m, -CH(CH2CH(CH3)2)}, 4.09 (2H, q, J = 7.0 Hz, -OCH2CH3), 3.97 - 3.86 {6H,
m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 2.04 (2H, q, J = 7.2 Hz, -CH2CH3), 1.60 - 1.40 {3H, m, -

ACCEPTED MANUSCRIPT
CH(CH2CH(CH3)2)}, 1.10 (3H, t, J = 7.0 Hz, -OCH2CH3), 0.88 - 0.84 {6H, m, -
CH(CH2CH(CH3)2)}, 0.74 (3H, t, J = 7.2 Hz, - CH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 170.9 (C=O), 168.9 (C=O), 166.4 (C=O), 138.5 (Cq), 135.8
(Cq), 130.1, 128.2, 127.3, 125.1, 89.9 (Cipso η5-C5H4-alkyl), 83.1 (Cipso η5-C5H4-benzoyl), 71.9
(Cmeta η5-C5H4-benzoyl), 69.3 (Cmeta η5-C5H4-alkyl), 68.0 (Cortho η5-C5H4-alkyl), 66.5 (Cortho η5-
C5H4-benzoyl), 62.5 (-OCH2CH3, -ve DEPT), 50.3 {-CH(CH2CH(CH3)2)}, 43.0 (-NHCH2CO-, -
ve DEPT), 39.9 {-CH(CH2CH(CH3)2), -ve DEPT}, 24.4 (-CH2CH3, -ve DEPT), 23.5 {-
CH(CH2CH(CH3)2)}, 23.0 {-CH(CH2CH(CH3)2)}, 22.4 {-CH(CH2CH(CH3)2)}, 14.4 (-
OCH2CH3), 13.7 (-CH2CH3).
4.3.8. 1-Ethyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester 14.
Glycine-L-phenylalanine ethyl ester hydrochloride (0.19 g, 0.75 mmol) was used as a starting
material. The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl
20
acetate} yielding the title product as a red oil (0.07 g, 18 %), Eº´ = 0.0 mV (vs Fc/Fc+), [α]D
=
+16o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 589.1768
C32H34N2O4FeNa requires:
589.1769.
I.R. υmax (KBr): 3300 (NH), 1739 (C=Oester), 1633 (C=Oamide), 1601 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 332, 444 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.54 (1H, t, J = 6.0 Hz, -CONH-), 8.30 (1H, d, J = 7.4 Hz,
-CONH-), 8.07 (1H, s, ArH), 7.69 - 7.67 (2H, m, ArH), 7.35 (1H, t, J = 7.0 Hz, ArH), 7.22 -
7.19 {5H, m, -CH(CH2Ph)}, 4.83 (2H, t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.43 - 4.40
{1H, m, -CH(CH2Ph)}, 4.37 (2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl), 4.08 (2H, q, J = 7.2
Hz, - OCH2CH3), 3.95 - 3.88 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 3.00 - 2.90 {2H, m, -
CH(CH2Ph)}, 2.05 (2H, q, J = 7.6 Hz, -CH2CH3), 1.10 (3H, t, J = 7.2 Hz, -OCH2CH3), 0.90
(3H, t, J = 7.6 Hz, -CH2CH3).
13C NMR (100MHz) δ (DMSO- d6): 172.4 (C=O), 169.7 (C=O), 167.0 (C=O), 135.0 (Cq), 131.7
(Cq), 130.7 (Cq), 129.7, 128.2, 127.4, 126.0, 124.2, 123.2, 122.7, 89.1 (Cipso η5-C5H4-alkyl), 83.0
(Cipso η5-C5H4-benzoyl), 73.2 (Cmeta η5-C5H4-benzoyl), 69.9 (Cmeta η5-C5H4-alkyl), 67.7 (Cortho η5-
C5H4-alkyl), 66.1 (Cortho η5-C5H4-benzoyl), 60.1 (-OCH2CH3, -ve DEPT), 52.8 {-CH(CH2Ph)},

ACCEPTED MANUSCRIPT
42.9 (-NHCH2CO-, -ve DEPT), 37.8 (-CH(CH2Ph), -ve DEPT), 24.0 (-CH2CH3, -ve DEPT), 14.9
(-OCH2CH3), 14.2 (-CH2CH3).
4.3.9. 1-Propyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 15.
Glycine-glycine ethyl ester hydrochloride (0.12 g, 0.72 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.08 g, 23 %), Eº´ = 20 mV (vs Fc/Fc+).
Mass spectrum: [M+Na]+ found: 513.1454
C26H30N2O4FeNa requires:
513.1453.
I.R. υmax (KBr): 3295 (NH), 1737 (C=Oester), 1631 (C=Oamide), 1609 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 334, 445 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.74 (1H, t, J = 6 Hz, -CONH-), 8.30 (1H, t, J = 6 Hz, -
CONH-), 8.08 (1H, s, ArH), 7.69 - 7.66 (2H, m, ArH), 7.40 (1H, t, J = 7.0 Hz, ArH), 4.80 (2H,
s, ortho on η5-C5H4-benzoyl), 4.34 (2H, s, meta on η5-C5H4-benzoyl), 4.01 (2H, q, J = 6.8 Hz, -
OCH2CH3), 3.95 - 3.85 {8H, m, (η5-C5H4-alkyl), (-NHCH2CO-), (-NHCH2CO-)}, 2.00 (2H, t, J
= 7.6 Hz, -CH2CH2CH3), 1.40 - 1.30 (2H, m, -CH2CH2CH3), 1.12 (3H, t, J = 7.2 Hz, -
OCH2CH3), 0.70 (3H, t, J = 7.2 Hz, - CH2CH2CH3);
13C NMR (100MHz) δ (DMSO- d6): 170.8 (C=O), 169.1 (C=O), 165.5 (C=O), 138.5 (Cq), 135.8
(Cq), 128.2, 127.9, 127.0, 123.2, 92.5 (Cipso η5-C5H4-alkyl), 83.0 (Cipso η5-C5H4-benzoyl), 71.7
(Cmeta η5-C5H4-benzoyl), 69.8 (Cmeta η5-C5H4-alkyl), 68.4 (Cortho η5-C5H4-alkyl), 66.7 (Cortho η5-
C5H4-benzoyl), 62.4 (-OCH2CH3, -ve DEPT), 42.8 (-NHCH2CO-, -ve DEPT), 41.9 (-
NHCH2CO-, -ve DEPT), 32.8 (-CH2CH2CH3, -ve DEPT), 25.0 (-CH2CH2CH3, -ve DEPT), 14.5
(-OCH2CH3), 14.0 (-CH2CH2CH3).
4.3.10. 1-Propyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester 16.
Glycine-L-alanine ethyl ester hydrochloride (0.13 g, 0.72 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
yielding the required product as a red oil (0.08 g, 22 %), Eº´ = 20 mV (vs Fc/Fc+), [α]D20 = -16o
(c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 527.1621
C27H32N2O4FeNa requires:
527.1609.

ACCEPTED MANUSCRIPT
I.R. υmax (KBr): 3290 (NH), 1739 (C=Oester), 1635 (C=Oamide), 1605 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 330, 449 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.65 (1H, t, J = 6 Hz, -CONH-), 8.41 (1H, d, J = 7.2 Hz, -
CONH-), 8.10 (1H, s, ArH), 7.71 - 7.69 (2H, m, ArH), 7.42 (1H, t, J = 7.0 Hz, ArH), 4.80 (2H,
s, ortho on η5-C5H4-benzoyl), 4.35 (2H, s, meta on η5-C5H4-benzoyl), 4.30 - 4.25 (1H, m, -
CHCH3), 4.10 (2H, q, J = 6.8 Hz, -OCH2CH3), 3.97 - 3.85 {6H, m, (η5-C5H4-alkyl), (-
NHCH2CO-)}, 2.00 (2H, t, J = 7.6 Hz, -CH2CH2CH3), 1.35 - 1.25 {5H, m, (-CH2CH2CH3), (-
CHCH3)}, 1.10 (3H, t, J = 6.8 Hz, -OCH2CH3), 0.72 (3H, t, J = 7.2 Hz, -CH2CH2CH3).
13C NMR (100MHz) δ ( DMSO- d6 ): 170.0 (C=O), 169.5 (C=O), 168.2 (C=O), 135.5 (Cq), 132.8
(Cq), 130.4, 129.2, 126.4, 123.9, 92.3 (Cipso η5-C5H4-alkyl), 82.6 (Cipso η5-C5H4-benzoyl), 71.6
(Cmeta η5-C5H4-benzoyl), 68.9 (Cmeta η5-C5H4-alkyl), 68.4 (Cortho η5-C5H4-alkyl), 67.9 (Cortho η5-
C5H4-benzoyl), 61.4 (-OCH2CH3, -ve DEPT), 48.6 (-CHCH3), 41.0 (-NHCH2CO-, -ve DEPT),
31.9 (-CH2CH2CH3, -ve DEPT), 21.0 (-CH2CH2CH3, -ve DEPT), 18.0 (-CHCH3), 14.2 (-
OCH2CH3), 13.9 (-CH2CH2CH3).
4.3.11. 1-Propyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl ester 17.
Glycine-L-leucine ethyl ester hydrochloride (0.16 g, 0.72 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.07 g, 18 %), [α]D20 = -18o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 569.2193
C30H38N2O4FeNa requires:
569.2181.
I.R. υmax (KBr): 3260 (NH), 1736 (C=Oester), 1643 (C=Oamide), 1612 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 331, 448 nm.
1H NMR (400MHz) δ (DMSO- d6): 8.65 (1H, t, J = 6 Hz, -CONH-), 8.30 (1H, d, J = 7.6 Hz, -
CONH-), 8.00 (1H, s, ArH), 7.72 - 7.70 (2H, m, ArH), 7.39 (1H, t, J = 7.0 Hz, ArH), 4.83 (2H,
s, ortho on η5-C5H4-benzoyl), 4.35 (2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl), 4.30 - 4.28
{1H, m, -CH(CH2CH(CH3)2)}, 4.19 (2H, q, J = 7.2 Hz, - OCH2CH3) , 3.95 - 3.88 {6H, m, (η5-
C5H4-alkyl), (-NHCH2CO-)}, 2.08 (2H, t, J = 7.6 Hz, -CH2CH2CH3), 1.69 - 1.45 {3H, m, -
CH(CH2CH(CH3)2)}, 1.35 - 1.25 (2H, m, -CH2CH2CH3), 1.19 (3H, t, J = 7.2 Hz, -OCH2CH3),
0.89 - 0.83 {6H, m, -CH(CH2CH(CH3)2)}, 0.70 (3H, t, J = 7.2 Hz, - CH2CH2CH3).

ACCEPTED MANUSCRIPT
13C NMR (100MHz) δ (DMSO- d6): 171.9 (C=O), 169.6 (C=O), 166.1 (C=O), 139.1 (Cq), 137.8
(Cq), 132.4, 129.7, 127.2, 125.7, 89.4 (Cipso η5-C5H4-alkyl), 83.3 (Cipso η5-C5H4-benzoyl), 71.0
(Cmeta η5-C5H4-benzoyl), 70.4 (Cmeta η5-C5H4-alkyl), 68.4 (Cortho η5-C5H4-alkyl), 66.1 (Cortho η5-
C5H4-benzoyl), 61.4 (-OCH2CH3, -ve DEPT), 51.3 (-CH(CH2CH(CH3)2), 42.9 (-NHCH2CO-, -ve
DEPT), 40.0 {-CH(CH2CH(CH3)2), -ve DEPT}, 31.0 (-CH2CH2CH3, -ve DEPT), 27.4 (-
CH(CH2CH(CH3)2) 24.7 (-CH2CH2CH3, -ve DEPT), 23.2 {-CH(CH2CH(CH3)2)}, 22.4 -
CH(CH2CH(CH3)2), 14.6 (-OCH2CH3), 13.6 (-CH2CH2CH3).
4.3.12. 1-Propyl-1’-N-{meta-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester 18.
Glycine-L-phenylalanine ethyl ester hydrochloride (0.18 g, 0.72 mmol) was used as a starting
material. The crude product was purified by column chromatography {eluant 1:1 hexane: ethyl
acetate} yielding the title product as a red oil (0.07 g, 17 %), Eº´ = 0.0 mV (vs Fc/Fc+), [α]D20 = -
13o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 603.2032
C33H36N2O4FeNa requires:
603.2034.
I.R. υmax (KBr): 3300 (NH), 1745 (C=Oester), 1633 (C=Oamide), 1612 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 330, 447 nm.
1H NMR (400MHz) δ ( DMSO- d6 ): 8.68 (1H, t, J = 6.0 Hz, -CONH-), 8.30 (1H, d, J = 7.6 Hz,
-CONH-), 8.00 (1H, s, ArH), 7.69 - 7.68 (2H, m, ArH), 7.40 (1H, t, ArH), 7.25 - 7.21 (5H, m, -
CH(CH2Ph)}, 4.80 (2H, s, ortho on η5-C5H4-benzoyl), 4.49 - 4.47 {1H, m, -CH(CH2Ph)}, 4.29
(2H, s, meta on η5-C5H4-benzoyl), 4.00 (2H, q, J = 7.2 Hz, -OCH2CH3), 3.95 - 3.83 {6H, m, (η5-
C5H4-alkyl), (-NHCH2CO-)}, 3.06 - 2.90 {2H, m, -CH(CH2Ph)}, 1.99 (2H, t, J = 7.6 Hz, -
CH2CH2CH3), 1.40 - 1.27 (2H, m, -CH2CH2CH3), 1.11 (3H, t, J = 7.2 Hz, -OCH2CH3), 0.70
(3H, t, J = 7.2 Hz, - CH2CH2CH3);
13C NMR (100MHz) δ (DMSO- d6): 171.9 (C=O), 169.2 (C=O), 166.9 (C=O), 137.0 (Cq), 132.7
(Cq), 130.1 (Cq), 128.7, 127.2, 126.8, 126.1, 125.2, 124.9, 123.7, 89.1 (Cipso η5-C5H4-alkyl), 84.0
(Cipso η5-C5H4-benzoyl), 71.4 (Cmeta η5-C5H4-benzoyl), 69.4 (Cmeta η5-C5H4-alkyl), 67.5 (Cortho η5-
C5H4-alkyl), 66.3 (Cortho η5-C5H4-benzoyl), 62.4 (-OCH2CH3, -ve DEPT), 53.7 (-CH(CH2Ph)),
43.2 (-NHCH2CO-, -ve DEPT), 39.2 {-CH(CH2Ph), -ve DEPT}, 33.8 (-CH2CH2CH3, -ve
DEPT), 20.6 (-CH2CH2CH3, -ve DEPT), 14.9 (-OCH2CH3), 13.1 (-CH2CH2CH3).

ACCEPTED MANUSCRIPT
4.3.13. 1-Methyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 19
Glycine glycine ethyl ester hydrochloride (0.13 g, 0.78 mmol) and
1-methyl-1’-N-ortho-ferrocenyl benzoic acid (0.25 g, 0.78 mmol) were used as starting
materials. The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl
acetate} yielding the desired product as a red oil (0.08 g, 22%).
Mass spectrum: [M+Na]+ found: 485.1289
C24H26N2O4FeNa requires:
485.1291.
I.R. υmax (KBr): 3310 (NH), 1741 (C=Oester), 1627 (C=Oamide), 1618 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 330, 445 nm.
1H NMR (400 MHz) δ (d6 DMSO): 8.74 (1H, t, J = 6.0 Hz, -CONH-), 8.33 (1H, t, J = 6 Hz, -
CONH-), 7.99 (1H, d, J = 7.0 Hz, ArH), 7.80 (1H, t, J = 7.2 Hz, ArH), 7.69 (1H, t, J = 6.4 Hz,
ArH), 7.40 (1H, d, J = 7.6 Hz, ArH), 4.83 (2H, s, ortho on η5-C5H4-benzoyl), 4.36 (2H, s, meta
on η5-C5H4-benzoyl), 4.11 (2H, q, J = 7.2 Hz, -OCH2CH3), 3.96 - 3.85 {8H, m, (η5-C5H4-alkyl),
(-NHCH2CO-), (-NHCH2CO-)}, 1.60 (3H, s, -CH3), 1.19 (3H, t, J = 7.2 Hz, -OCH2CH3).
13C NMR (100 MHz) δ (d6 DMSO): 170.5 (C=O), 169.0 (C=O), 168.3 (C=O), 137.5 (Cq), 135.8
(Cq), 130.4, 128.2, 127.3, 126.1, 93.3 (Cipso η5-C5H4-alkyl), 83.6 (Cipso η5-C5H4-benzoyl), 71.7
(Cmeta η5-C5H4-benzoyl), 70.5 (Cmeta η5-C5H4-alkyl), 68.9 (Cortho η5-C5H4-alkyl), 66.2 (Cortho η5-
C5H4-benzoyl), 61.5 (-OCH2CH3, -ve DEPT), 43.5 (-NHCH2CO-, -ve DEPT), 40.7 (-
NHCH2CO-, -ve DEPT), 14.6 (-OCH2CH3), 14.0 (-CH3).
4.3.14. 1-Methyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester 20
Glycine-L-alanine ethyl ester hydrochloride (0.14 g, 0.78 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the title compound as a red oil (0.07 g, 19%), [α]D20 = -15o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 499.1395
C25H28N2O4FeNa requires:
499.1396.
I.R. υmax (KBr): 3300 (NH), 1743 (C=Oester), 1630 (C=Oamide), 1600 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 324, 448 nm.
1H NMR (400 MHz) δ (DMSO- d6): 8.74 (1H, t, J = 6.0 Hz, -CONH-), 8.33 (1H, d, J = 6 Hz, -
CONH-), 7.99 (1H, d, J = 7.2 Hz, ArH), 7.81 (1H, t, J = 7.0 Hz, ArH), 7.69 (1H, t, J = 6.4 Hz,
ArH), 7.40 (1H, d, J = 7.6 Hz, ArH), 4.80 (2H, s, ortho on η5-C5H4-benzoyl), 4.35 (2H, s, meta

ACCEPTED MANUSCRIPT
on η5-C5H4-benzoyl), 4.31- 4.24 (1H, m, -CHCH3), 4.11 (2H, q, J = 6.8 Hz, -OCH2CH3), 3.93 -
3.84 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 1.58 (3H, s, -CH3), 1.30 (3H, d, J = 6.2 Hz, -
CHCH3), 1.17 (3H, t, J = 6.8 Hz, -OCH2CH3);
13C NMR (100 MHz) δ (DMSO- d6): 171.9 (C=O), 169.0 (C=O), 168.2 (C=O), 137.5 (Cq), 136.8
(Cq), 130.4, 128.2, 127.3, 126.1, 92.6 (Cipso η5-C5H4-alkyl), 84.0 (Cipso η5-C5H4-benzoyl), 71.0
(Cmeta η5-C5H4-benzoyl), 68.3 (Cmeta η5-C5H4-alkyl), 68.0 (Cortho η5-C5H4-alkyl), 66.1 (Cortho η5-
C5H4-benzoyl), 62.4 (-OCH2CH3, -ve DEPT), 47.3 (-CHCH3), 42.0 (-NHCH2CO-, -ve DEPT),
17.1 (-CHCH3), 14.9 (-OCH2CH3), 13.4 (-CH3).
4.3.15. 1-Methyl-1’-N-{ortho-(ferrocenyl)-benzoyl} glycine-L-leucine ethyl ester 21
Glycine-L-leucine ethyl ester hydrochloride (0.17 g, 0.78 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.06 g, 15%), [α]D20 = -21o (c 0.1, EtOH).
I.R. υmax (KBr): 3260 (NH), 1742 (C=Oester), 1639 (C=Oamide), 1611 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 322, 449 nm.
1H NMR (400 MHz) δ (DMSO- d6): 8.43 (1H, t, J = 7.6 Hz, -CONH-), 8.23 (1H, d, J = 7.6 Hz,
-CONH-), 7.83 (1H, d, J = 7.2 Hz, ArH), 7.50 (1H, t, J = 6.8 Hz, ArH), 7.27 (1H, t, J = 6.8
Hz, ArH), 7.15 (1H, d, J = 7.2 Hz, ArH), 4.82 (2H, s, ortho on η5-C5H4-benzoyl), 4.37 - 4.20
{3H, m, (meta on η5-C5H4-benzoyl), (-CH(CH2CH(CH3)2)}, 4.07 (2H, q, J = 7.0 Hz, -
OCH2CH3), 3.96 - 3.85 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 1.63 - 1.49 {6H, m, (-CH3), (-
CH(CH2CH(CH3)2)}, 1.18 (3H, t, J = 7.0 Hz, -OCH2CH3), 0.91 - 0.89 {6H, m, -
CH(CH2CH(CH3)2)}.
13C NMR (100 MHz) δ ( DMSO- d6 ): 172.9 (C=O), 169.4 (C=O), 167.8 (C=O), 136.2 (Cq),
134.8 (Cq), 133.1, 128.8, 127.4, 126.8, 90.1 (Cipso η5-C5H4-alkyl), 85.6 (Cipso η5-C5H4-benzoyl),
71.1 (Cmeta η5-C5H4-benzoyl), 70.0 (Cmeta η5-C5H4-alkyl), 69.8 (Cortho η5-C5H4-alkyl), 67.3 (Cortho
η5-C5H4-benzoyl), 61.4 (-OCH2CH3, -ve DEPT), 51.2 {-CH(CH2CH(CH3)2)}, 43.8 (-
NHCH2CO-, -ve DEPT), 41.2 {-CH(CH2CH(CH3)2), -ve DEPT}, 24.9 {-CH(CH2CH(CH3)2)},
22.9 {-CH(CH2CH(CH3)2)}, 21.9 {-CH(CH2CH(CH3)2)}, 14.2 (-OCH2CH3), 13.1 (-CH3).
4.3.16. 1-Methyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester 22

ACCEPTED MANUSCRIPT
Glycine-L-phenylalanine ethyl ester hydrochloride (0.20 g, 0.78 mmol) was used as a starting
material. The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl
acetate} yielding the desired product as a red oil (0.05 g, 12%), [α]D20 = +10o (c 0.1, EtOH).
I.R. υmax (KBr): 3253 (NH), 1755 (C=Oester), 1663 (C=Oamide), 1601 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 323, 449 nm.
1H NMR (400 MHz) δ (DMSO- d6): 8.58 (1H, t, J = 6.0 Hz, -CONH-), 8.32 (1H, d, J = 7.6 Hz,
-CONH-), 8.07 (1H, d, J = 7.0 Hz, ArH), 7.81 (1H, t, J = 7.2 Hz, ArH), 7.35 (1H, t, J = 7.0 Hz,
ArH ), 7.21 - 7.18 {6H, m, -CH(CH2Ph), ArH}, 4.83 (2H, s, ortho on η5-C5H4-benzoyl), 4.45 -
4.43 {1H, m, -CH(CH2Ph)}, 4.35 (2H, s, meta on η5-C5H4-benzoyl), 4.00 (2H, q, J = 7.2 Hz, -
OCH2CH3), 3.97 - 3.85 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-) }, 3.01 - 2.93 {2H, m, -
CH(CH2Ph)}, 1.54 (3H, s, -CH3), 1.10 (3H, t, J = 7.2 Hz, -OCH2CH3).
13C NMR (100 MHz) δ (DMSO- d6): 172.1 (C=O), 169.2 (C=O), 167.3 (C=O), 137.0 (Cq), 131.0
(Cq), 129.7 (Cq), 128.7, 128.0, 127.4, 126.4, 125.9, 124.7, 123.7, 93.2 (Cipso η5-C5H4-alkyl), 83.9
(Cipso η5-C5H4-benzoyl), 70.4 (Cmeta η5-C5H4-benzoyl), 70.0 (Cmeta η5-C5H4-alkyl), 68.3 (Cortho η5-
C5H4-benzoyl), 67.9 (Cortho η5-C5H4-alkyl), 62.4 (-OCH2CH3, -ve DEPT), 51.6 {-CH(CH2Ph)},
42.7 (-NHCH2CO-, -ve DEPT), 35.7 {-CH(CH2Ph), -ve DEPT}, 13.9 (-OCH2CH3), 13.0 (-CH3).
4.3.17. 1-Ethyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 23
Glycine-glycine ethyl ester hydrochloride (0.12 g, 0.75 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.08 g, 22%).
Mass spectrum: [M+Na]+ found: 499.1392
C25H28N2O4FeNa requires:
499.1396.
I.R. υmax (KBr): 3310 (NH), 1739 (C=Oester), 1631 (C=Oamide), 1617 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 325, 444 nm.
1H NMR (400 MHz) δ (DMSO- d6): 8.72 (1H, t, J = 6.0 Hz, -CONH-), 8.34 (1H, t, J = 6 Hz, -
CONH-), 8.00 (1H, d, J = 7.4 Hz, ArH), 7.81 (1H, t, J = 7.2 Hz, ArH), 7.70 (1H, t, J = 6.4 Hz,
ArH), 7.40 (1H, d, J = 7.6 Hz, ArH), 4.82 (2H, t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.31
(2H, t, J = 2 Hz, meta on η5-C5H4-benzoyl), 4.10 (2H, q, J = 6.8 Hz, - OCH2CH3), 3.97 - 3.85
{8H, m, (η5-C5H4-alkyl), (-NHCH2CO-), (-NHCH2CO-)}, 2.07 (2H, q, J = 7.6 Hz, -CH2CH3),
1.18 (3H, t, J = 6.8 Hz, -OCH2CH3), 0.98 (3H, t, J = 7.6 Hz, -CH2CH3).

ACCEPTED MANUSCRIPT
13C NMR (100 MHz) δ (DMSO- d6): 169.8 (C=O), 168.3 (C=O), 165.2 (C=O), 135.6 (Cq), 133.8
(Cq), 128.5, 127.2, 126.0, 124.0, 92.3 (Cipso η5-C5H4-alkyl), 81.0 (Cipso η5-C5H4-benzoyl), 71.0
(Cmeta η5-C5H4-benzoyl), 68.7 (Cmeta η5-C5H4-alkyl), 68.0 (Cortho η5-C5H4-alkyl), 66.1 (Cortho η5-
C5H4-benzoyl), 62.4 (-OCH2CH3, -ve DEPT), 43.4 (-NHCH2CO-, -ve DEPT), 41.7 (-
NHCH2CO-, -ve DEPT), 22.8 (-CH2CH3, -ve DEPT), 14.2 (-OCH2CH3), 13.0 (-CH2CH3).
4.3.18. 1-Ethyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester 24
Glycine-L-alanine ethyl ester hydrochloride (0.13 g, 0.75 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the title compound as a red oil (0.07 g, 19%), [α]D20 = -19o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 513.1447
C25H28N2O4FeNa requires:
513.1453.
I.R. υmax (KBr): 3289 (NH), 1740 (C=Oester), 1632 (C=Oamide), 1608 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 325, 443 nm.
1H NMR (400 MHz) δ (DMSO- d6): 8.48 (1H, t, J = 6.0 Hz, -CONH-), 8.34 (1H, d, J = 7.2 Hz,
-CONH-), 8.01 (1H, d, J = 6.8 Hz, ArH), 7.81 (1H, t, J = 7.2 Hz, ArH), 7.70 (1H, t, J = 6.4 Hz,
ArH), 7.41 (1H, d, J = 7.4 Hz, ArH), 4.72 (2H, t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.38
(2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl), 4.30 - 4.26 ( 1H, m, - CHCH3), 4.10 (2H, q, J =
6.8 Hz, - OCH2CH3), 3.92 - 3.80 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 2.00 (2H, q, J = 7.6
Hz, -CH2CH3), 1.30 (3H, d, J = 6.4 Hz, - CHCH3), 1.15 (3H, t, J = 6.8 Hz, -OCH2CH3), 0.90
(3H, t, J = 7.6 Hz, -CH2CH3);
13C NMR (100 MHz) δ (DMSO- d6): 172.5 (C=O), 168.0 (C=O), 167.2 (C=O), 138.5 (Cq), 134.0
(Cq), 129.0, 128.8, 127.5, 124.1, 92.7 (Cipso η5-C5H4-alkyl), 81.2 (Cipso η5-C5H4-benzoyl), 70.9
(Cmeta η5-C5H4-benzoyl), 69.8 (Cmeta η5-C5H4-alkyl), 68.5 (Cortho η5-C5H4-alkyl), 66.2 (Cortho η5-
C5H4-benzoyl), 60.8 (-OCH2CH3, -ve DEPT), 49.6 (-CHCH3), 42.2 (-NHCH2CO-, -ve DEPT),
25.8 (-CH2CH3, -ve DEPT), 17.6 (-CHCH3), 14.8 (-OCH2CH3), 14.0 (-CH2CH3).
4.3.19. 1-Ethyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl ester 25
Glycine-L-leucine ethyl ester hydrochloride (0.16 g, 0.75 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.06 g, 15%), [α]D20 = -11o (c 0.1, EtOH).

ACCEPTED MANUSCRIPT
I.R. υmax (KBr): 3275 (NH), 1747 (C=Oester), 1649 (C=Oamide), 1614 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 360, 451 nm.
1H NMR (400 MHz) δ(DMSO- d6): 8.68 (1H, t, J = 6.0 Hz, -CONH-), 8.34 (1H, d, J = 7.6 Hz,
-CONH-), 8.11 (1H, d, J = 6.6 Hz, ArH), 7.80 (1H, t, J = 7.2 Hz, ArH), 7.70 (1H, t, J = 6.4 Hz,
ArH), 7.40 (1H, d, J = 7.6 Hz, ArH), 4.83 (2H, t, J = 1.6 Hz, ortho on η5-C5H4-benzoyl), 4.33
(2H, t, J = 1.6 Hz, meta on η5-C5H4-benzoyl), 4.32 - 4.28 {1H, m, -CH(CH2CH(CH3)2)}, 4.20
(2H, q, J = 7.0 Hz, - OCH2CH3), 3.96 - 3.87 {6H m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 2.02 (2H,
q, J = 7.2 Hz, -CH2CH3), 1.65 - 1.40 {3H, m, -CH(CH2CH(CH3)2)}, 1.07 (3H, t, J = 7.0 Hz, -
OCH2CH3), 0.92 - 0.88 {6H, m, -CH(CH2CH(CH3)2)}, 0.70 (3H, t, J = 7.2 Hz, - CH2CH3);
13C NMR (100 MHz) δ (DMSO- d6): 170.4 (C=O), 169.6 (C=O), 166.4 (C=O), 136.6 (Cq), 135.8
(Cq), 131.4, 128.2, 127.7, 124.1, 92.4 (Cipso η5-C5H4-alkyl), 82.1 (Cipso η5-C5H4-benzoyl), 71.0
(Cmeta η5-C5H4-benzoyl), 70.4 (Cmeta η5-C5H4-alkyl), 68.5 (Cortho η5-C5H4-alkyl), 66.9 (Cortho η5-
C5H4-benzoyl), 61.4 (-OCH2CH3, -ve DEPT), 52.3 {-CH(CH2CH(CH3)2)}, 43.1 (-NHCH2CO-, -
ve DEPT), 40.1 {-CH(CH2CH(CH3)2), -ve DEPT}, 27.9 {-CH(CH2CH(CH3)2)}, 24.4 (-CH2CH3,
-ve DEPT), 21.7 {-CH(CH2CH(CH3)2)}, 20.4 {-CH(CH2CH(CH3)2)}, 14.7 (-OCH2CH3), 13.0 (-
CH2CH3).
4.3.20. 1-Ethyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester 26.
Glycine-L-phenylalanine ethyl ester hydrochloride (0.19 g, 0.75 mmol) was used as a starting
material. The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl
acetate} yielding the desired product as a red oil (0.06 g, 14%), [α]D20 = +14o (c 0.1, EtOH).
I.R. υmax (KBr): 3303 (NH), 1744 (C=Oester), 1629 (C=Oamide), 1608 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 325, 444 nm.
1H NMR (400 MHz) δ (DMSO- d6): 8.68 (1H, t, J = 6.0 Hz, -CONH-), 8.32 (1H, d, J = 7.6 Hz,
-CONH-), 8.03 (1H, d, J = 7.0 Hz, ArH), 7.81 (1H, t, J = 7.2 Hz, ArH), 7.79 (1H, t, J = 6.4 Hz,
ArH), 7.38 (1H, d, J = 6.8 Hz, ArH), 7.23 - 7.19 (5H, m, -CH(CH2Ph)), 4.83 (2H, t, J = 1.6 Hz,
ortho on η5-C5H4-benzoyl), 4.49 - 4.47 {1H, m, -CH(CH2Ph)}, 4.37 (2H, t, J = 1.6 Hz, meta on
η5-C5H4-benzoyl), 4.04 (2H, q, J = 6.4 Hz, -OCH2CH3), 3.89 - 3.80 {6H, m, (η5-C5H4-alkyl), (-
NHCH2CO-)}, 3.08 - 2.95 {2H, m, -CH(CH2Ph)}, 2.02 {2H, q, J = 7.6 Hz, -CH2CH3), 1.16 (3H,
t, J = 6.4 Hz, -OCH2CH3), 0.94 (3H, t, J = 7.6 Hz, -CH2CH3).

ACCEPTED MANUSCRIPT
13C NMR (100 MHz) δ (DMSO- d6): 171.5 (C=O), 169.1 (C=O), 168.2 (C=O), 139.0 (Cq), 130.9
(Cq), 129.1 (Cq), 128.5, 128.4, 127.1, 126.6, 125.2, 124.2, 121.7, 92.1 (Cipso η5-C5H4-alkyl), 81.1
(Cipso η5-C5H4-benzoyl), 71.4 (Cmeta η5-C5H4-benzoyl), 69.9 (Cmeta η5-C5H4-alkyl), 68.9 (Cortho η5-
C5H4-alkyl), 67.9 (Cortho η5-C5H4-benzoyl), 62.8 (-OCH2CH3, -ve DEPT), 53.6 {-CH(CH2Ph)},
42.2 (-NHCH2CO-, -ve DEPT), 39.8 {-CH(CH2Ph), -ve DEPT}, 23.0 (-CH2CH3, -ve DEPT),
15.0 (-OCH2CH3), 14.1 (-CH2CH3).
4.3.21. 1-Propyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-glycine ethyl ester 27.
Glycine-glycine ethyl ester hydrochloride (0.12 g, 0.72 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the title compound as a red oil (0.08 g, 23%).
Mass spectrum: [M+Na]+ found: 513.1464
C26H30N2O4FeNa requires:
513.1453.
I.R. υmax (KBr): 3387 (NH), 1735 (C=Oester), 1629 (C=Oamide), 1609 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 325, 443 nm.
1H NMR (400 MHz) δ( DMSO- d6): 8.76 (1H, t, J = 5.6 Hz, -CONH-), 8.34 (1H, t, J = 5.6 Hz, -
CONH-), 7.98 (1H, d, J = 7.4 Hz, ArH), 7.86 (1H, t, J = 7.2 Hz, ArH), 7.70 (1H, t, J = 6.8 Hz,
ArH), 7.39 (1H, d, J = 7.6 Hz, ArH), 4.83 (2H, s, ortho on η5-C5H4-benzoyl), 4.36 (2H, s, meta
on η5-C5H4-benzoyl), 4.12 (2H, q, J = 6.8 Hz, - OCH2CH3) , 3.96 - 3.85 {8H, m, (η5-C5H4-
alkyl), (-NHCH2CO-), (-NHCH2CO-)}, 2.05 (2H, t, J = 7.6 Hz, -CH2CH2CH3), 1.35 - 1.27 (2H,
m, -CH2CH2CH3), 1.10 (3H, t, J = 6.8 Hz, -OCH2CH3), 0.72 (3H, t, J = 7.2 Hz, - CH2CH2CH3);
13C NMR (100 MHz) δ (DMSO- d6): 168.8 (C=O), 168.3 (C=O), 167.9 (C=O), 142.5 (Cq), 137.8
(Cq), 130.2, 129.5, 128.9, 126.7, 91.8 (Cipso η5-C5H4-alkyl), 81.9 (Cipso η5-C5H4-benzoyl), 71.7
(Cmeta η5-C5H4-benzoyl), 69.1 (Cmeta η5-C5H4-alkyl), 67.0 (Cortho η5-C5H4-alkyl), 66.2 (Cortho η5-
C5H4-benzoyl), 61.9 (-OCH2CH3, -ve DEPT), 44.4 (-NHCH2CO-, -ve DEPT), 42.8 (-
NHCH2CO-, -ve DEPT), 32.8 (-CH2CH2CH3, -ve DEPT), 24.5 (-CH2CH2CH3, -ve DEPT), 14.3
(-OCH2CH3), 13.1 (-CH2CH2CH3).
4.3.22. 1-Propyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-alanine ethyl ester 28

ACCEPTED MANUSCRIPT
Glycine-L-alanine ethyl ester hydrochloride (0.13 g, 0.72 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.07 g, 19%), [α]D20 = -19o (c 0.1, EtOH).
Mass spectrum: [M+Na]+ found: 527.1617
C27H32N2O4FeNa requires:
527.1609.
I.R. υmax (KBr): 3291 (NH), 1740 (C=Oester), 1639 (C=Oamide), 1618 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 325, 446 nm.
1H NMR (400 MHz) δ ( DMSO- d6): 8.68 (1H, t, J = 6.0 Hz, -CONH-), 8.41 (1H, d, J = 7.0 Hz,
-CONH-), 8.01 (1H, d, J = 7.6 Hz, ArH), 7.80 (1H, t, J = 7.2 Hz, ArH), 7.79 (1H, t, J = 6.0 Hz,
ArH), 7.38 (1H, d, J = 7.6 Hz, ArH), 4.85 (2H, s, ortho on η5-C5H4-benzoyl), 4.37 (2H, s, meta
on η5-C5H4-benzoyl), 4.33 - 4.28 (1H, m, - CHCH3), 4.11 (2H, q, J = 6.4 Hz, - OCH2CH3) ,
3.92 - 3.85 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 2.05 (2H, t, J = 7.6 Hz, -CH2CH2CH3),
1.38 - 1.29 {5H, m, (-CH2CH2CH3), (-CHCH3)}, 1.14 (3H, t, J = 6.4 Hz, -OCH2CH3), 0.70 (3H,
t, J = 7.2 Hz, - CH2CH2CH3).
13C NMR (100 MHz) δ (DMSO- d6): 172.9 (C=O), 169.0 (C=O), 166.2 (C=O), 137.2 (Cq), 135.8
(Cq), 130.4, 128.6, 127.4, 124.8, 91.3 (Cipso η5-C5H4-alkyl), 81.2 (Cipso η5-C5H4-benzoyl), 72.0
(Cmeta η5-C5H4-benzoyl), 68.8 (Cmeta η5-C5H4-alkyl), 68.5 (Cortho η5-C5H4-alkyl), 67.9 (Cortho η5-
C5H4-benzoyl), 60.4 (-OCH2CH3, -ve DEPT), 47.6 (-CHCH3), 42.0 (-NHCH2CO-, -ve DEPT),
31.0 (-CH2CH2CH3, -ve DEPT), 22.8 (-CH2CH2CH3, -ve DEPT), 17.4 (-CHCH3), 15.0 (-
OCH2CH3), 14.0 (-CH2CH2CH3).
4.3.23. 1-Propyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-leucine ethyl ester 29
Glycine-L-leucine ethyl ester hydrochloride (0.12 g, 0.72 mmol) was used as a starting material.
The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl acetate}
yielding the desired product as a red oil (0.05 g, 14%), [α]D20 = -17o (c 0.1, EtOH).
I.R. υmax (KBr): 3272 (NH), 1751 (C=Oester), 1633 (C=Oamide), 1610 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 324, 446 nm.
1H NMR (400 MHz) δ( DMSO- d6): 8.67 (1H, t, J = 6.0 Hz, -CONH-), 8.33 (1H, d, J = 7.6 Hz,
-CONH-), 8.03 (1H, d, J = 7.2 Hz, ArH), 7.81 (1H, t, J = 7.0 Hz, ArH), 7.79 (1H, t, J = 6.4 Hz,
ArH), 7.31 (1H, d, J = 7.6 Hz, ArH), 4.79 (2H, s, ortho on η5-C5H4-benzoyl), 4.35 (2H, s, meta
on η5-C5H4-benzoyl), 4.30 - 4.27 {1H, m, -CH(CH2CH(CH3)2)}, 4.17 (2H, q, J = 7.2 Hz, -

ACCEPTED MANUSCRIPT
OCH2CH3), 3.95 - 3.84 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 2.07 (2H, t, J = 7.6 Hz, -
CH2CH2CH3), 1.69 - 1.45 {3H, m, -CH(CH2CH(CH3)2)}, 1.38 - 1.23 (2H, m, -CH2CH2CH3),
1.17 (3H, t, J = 7.2 Hz, -OCH2CH3), 0.90 - 0.85 {6H, m, -CH(CH2CH(CH3)2)}, 0.74 (3H, t, J =
7.2 Hz, - CH2CH2CH3).
13C NMR (100 MHz) δ (DMSO- d6): 170.9 (C=O), 168.5 (C=O), 167.1 (C=O), 139.5 (Cq), 135.8
(Cq), 129.4, 127.9, 125.2, 124.9, 91.4 (Cipso η5-C5H4-alkyl), 82.9 (Cipso η5-C5H4-benzoyl), 71.0
(Cmeta η5-C5H4-benzoyl), 70.4 (Cmeta η5-C5H4-alkyl), 69.2 (Cortho η5-C5H4-alkyl), 66.8 (Cortho η5-
C5H4-benzoyl), 61.4 (-OCH2CH3, -ve DEPT), 51.3 {-CH(CH2CH(CH3)2)}, 43.9 (-NHCH2CO-, -
ve DEPT), 41.0 {-CH(CH2CH(CH3)2), -ve DEPT}, 31.0 (-CH2CH2CH3, -ve DEPT), 27.7 {-
CH(CH2CH(CH3)2)}, 24.4 (-CH2CH2CH3, -ve DEPT), 22.7 {-CH(CH2CH(CH3)2)}, 21.4 {-
CH(CH2CH(CH3)2)}, 14.6 (-OCH2CH3), 13.6 (-CH2CH2CH3).
4.3.24. 1-Propyl-1’-N-{ortho-(ferrocenyl)-benzoyl}-glycine-L-phenylalanine ethyl ester 30
Glycine-L-phenylalanine ethyl ester hydrochloride (0.18 g, 0.72 mmol) was used as a starting
material. The crude product was purified by column chromatography {eluent 1:1 hexane: ethyl
acetate} yielding the title compound as a red oil (0.06 g, 13%), [α]D20 = +17o (c 0.1, EtOH).
I.R. υmax (KBr): 3288 (NH), 1730 (C=Oester), 1629 (C=Oamide), 1615 (C=Oamide) cm-1.
UV-Vis λmax EtOH: 324, 445 nm.
1H NMR (400 MHz) δ( DMSO- d6 ): 8.50 (1H, t, J = 6.0 Hz, -CONH-), 8.26 (1H, d, J = 7.0 Hz,
-CONH-), 7.91 (1H, d, J = 7.0 Hz, ArH), 7.78 (3H, t, J = 7.2 Hz, ArH), 7.49 (1H, t, J = 7.2 Hz,
ArH), 7.30 (1H, d, J = 7.0 Hz, ArH) 7.16 - 7.10 {5H, m, -CH(CH2Ph)}, 4.73 (2H, s, ortho on η5-
C5H4-benzoyl), 4.40 - 4.38 {1H, m, -CH(CH2Ph)}, 4.27 (2H, s, meta on η5-C5H4-benzoyl), 3.96
(2H, q, J = 7.2 Hz, - OCH2CH3), 3.83 - 3.79 {6H, m, (η5-C5H4-alkyl), (-NHCH2CO-)}, 2.84 -
2.80 {3H, m, -CH(CH2Ph)}, 1.88 (2H, t, J = 7.6 Hz, -CH2CH2CH3), 1.39 - 1.33 (2H, m, -
CH2CH2CH3), 1.14 (3H, t, J = 7.2 Hz, -OCH2CH3), 0.76 (3H, t, J = 7.2 Hz, - CH2CH2CH3);
13C NMR (100 MHz) δ (DMSO- d6): 169.6 (C=O), 168.6 (C=O), 166.3 (C=O), 136.0 (Cq), 134.1
(Cq), 128.8 (Cq), 128.5, 126.2, 125.1, 124.2, 123.1, 122.2, 120.8, 91.2 (Cipso η5-C5H4-alkyl), 83.1
(Cipso η5-C5H4-benzoyl), 69.9 (Cmeta η5-C5H4-benzoyl), 68.8 (Cmeta η5-C5H4-alkyl), 68.5 (Cortho η5-
C5H4-alkyl), 66.2 (Cortho η5-C5H4-benzoyl), 60.3 (-OCH2CH3, -ve DEPT), 50.0 {-CH(CH2Ph)},
42.4 (-NHCH2CO-, -ve DEPT), 36.2 (-CH(CH2Ph), -ve DEPT), 31.1 (-CH2CH2CH3, -ve DEPT),
20.8 (-CH2CH2CH3, -ve DEPT), 14.3 (-OCH2CH3), 14.1 (-CH2CH2CH3).

ACCEPTED MANUSCRIPT
4.4. General procedure for in vitro cytotoxicity assays
4.4.1. Biological Assays. Cell Line.
H1299 was obtained from the American Tissue Culture Centre (ATCC). The cell line was grown
o
in RPMI-1640 supplemented with 10% fetal calf serum (FCS) at 37 C in a 5% CO2 humidified
chamber.
4.4.2. In Vitro Proliferation Assays.
Cells in the exponential phase of growth were harvested by trypsinisation and a cell suspension
of 1 x 104 cells per ml was prepared in fresh culture medium. The cell suspension (100 µL) was
added to a flat bottom 96-well plate (Costar, 3599), plates were agitated gently in order to ensure
even dispersion of cells over the surface of the wells, and then cells were incubated for an initial
24 hours in a 37 oC, 5% CO2 incubator, to allow cell attachment to the wells. A 10 mM stock
solution of a test sample was prepared in dimethyl sulfoxide; dilute solutions of the test sample
were prepared at 2X final concentration by spiking the cell culture medium with a calculated
amount of the stock solution. 100 µL aliquot of each dilute solution was added to each well of
the plate, the plate was gently agitated, and then incubated at 37 oC, 5% CO2 for 5-6 days, until
cell confluency reached 80-90%. Assessment of cell survival in the presence of the compounds
7-30 was determined by the acid phosphatase assay [32]. The percentage cell growth in the
presence of each compound was determined relative to the control cells. The concentration of
compounds causing a 50% growth inhibition (IC50 of the compound) was determined using
Calcusyn (Biosoft, UK).
Acknowledgements
The authors thank James Murphy for assistance with the biological testing.
References
[1] C. G. Hartinger, N. Metzler-Nolte, P. J. Dyson, Organometallics, 31 (2012), p.5677.
[2] N. P. E. Barry, P. J. Sadler, Chem. Comm., 49 (2013), p. 5106
[3] C. G. Hartinger, P. J. Dyson, Chem. Soc. Rev., 38 (2009), p. 391.
[4] G. Gasser, I. Ott, N. Metzler-Nolte, J. Med. Chem., 54 (2011), p. 3.