1.

Introduction

1.1. Carbenes

Carbenes are compounds containing a divalent carbon atom with only six
[16]
valence-shell electrons. Introduced by Doering to organic chemistry in the 1950s
and by Fischer to organometallic chemistry in 1964, [17] these fascinating species have
sparked the interest of organic, [18] inorganic,[19] and theoretical chemists[20] like no
other single class of molecules.
In a prototypical carbene molecule –C–, the carbon atom can be either linear
or bent. A linear geometry gives rise to a sp-hybridized carbene center with two
degenerate nonbonding orbitals (p x and py). The molecule bending removes the
degeneracy and the carbene atom is sp 2-hybridized. As a result, the former px orbital
is stabilized by acquiring s character (it is therefore called σ), while the p y orbital
remains almost intact and is often called pπ (Figure 1.5).[18b]

Figure 1.5. Frontier orbitals of prototypical linear and bent carbenes.

Most carbenes are bent and their frontier orbitals can be systematically
labeled as σ and pπ. The ground-state spin multiplicity is a fundamental property of
carbenes. Four electronic configurations can be envisioned (Figure 1.6).[18b] A triplet
carbene contains two non-bonding electrons in two different orbitals with parallel
spins (σ1pπ1). A singlet carbene, on the other hand, may have two non-bonding
electrons paired in the same σ or p π orbitals (σ2 or pπ2), or paired in two different
orbitals (σ1pπ1); the σ2state is generally more stable. The ground state spin multiplicity

1
dictates the reactivity of carbenes. Singlet carbenes have a filled and an empty
orbital, and thus are generally ambiphilic; triplet carbenes have two half-occupied
orbitals and behave like diradicals.[21]

Figure 1.6. Four possible electronic configurations of carbene.

A carbene’s ground-state electronic configuration is related to the energies of

its σ and pπ orbitals.[22] The σ-pπ separation is largely influenced by the substituents
on the carbenes. Infact steric and electronic effects of the substituents at the carbene
carbon atom control the multiplicity of the ground state. It is generally accepted that
the singlet ground state is stabilized by σ-electron withdrawing, generally more
electronegative substituents.[23]
This negative inductive effect causes a lowering of the relative energy of the
non-bonding σ orbital, while the relative energy of the p π orbital remains essentially
unchanged. Substituents with σ-electron donating properties decrease the energy
gap between the σ and the p π orbital stabilizing the triplet ground state. In addition to
the inductive effects, mesomeric effects play a crucial role. [24]
The substituents at the carbene carbon atom are classified into the three
categories (X), (Z), (C): substituents (X) which provide π electrons for the carbene
carbon atom, substituents (Z) which withdraw π electrons from the carbene center,
and substituents (C) which are carbon atoms being part of a conjugated system. [25]
The majority of the π acceptor-substituted carbene centers (Z 2C: where for example
Z= Li, BH2 or BeH) are in the singlet ground state although they are linear or almost
linear.[26] The combination of a π acceptor with a σ donor at the carbene carbon atom
XZC: leads to compounds that are almost linear about the carbon atom. Bertrand’s
almost linear phosphanyl(silyl)carbene, [27] (phosphanyl)(phosphonio)carbenes, [28] and
trifluoroethylidynesulfurtrifluoride[29] are among these derivatives.
Some surprisingly persistent triplet carbenes (half-lives of several ms up to 9
min) have been obtained by substitution at the carbene center by two carbon atoms
which are part of a conjugated system (alkenes, alkynes, aryl groups). Crucial
parameters for stability are the type and steric demand of the substituents (to prevent

2
dimerization to olefins) and their ability to prevent the reaction with elemental oxygen.
[30]

Singlet carbenes which are substituted with two π donors of type X 2C: are
strongly bent at the carbene carbon atom (Figure 1.7). The interaction of the π
electron pairs at the substituents with the p π orbital at the carbene carbon atom
raises the relative energy of the p π orbital. Important members of this class of
compounds are the dimethoxy carbenes and the dihalocarbenes. [31]

Figure 1.7. Singlet carbenes substituted with two π donors of type X2C:.

The N-heterocyclic carbenes (NHCs) exclusively contain X 2C: singlet carbene

centers for geometric and electronic reasons.
There are essentially three different types of transition metal carbene
complexes featuring different types of carbene ligands. They have all been named
after their discoverers: Fischer carbenes[17, 32]
Schrock carbenes[33] and Wanzlick-
Arduengo[34] (Figure 1.8).

Figure 1.8. Bonding in Fischer, Wanzlick-Arduengo and Schrock carbene transition metal
complexes.

Fischer carbenes complexes have an electrophilic carbene atom [35] that can be
attacked by a Lewis base. The Schrock carbenes complexes have a reversed
reactivity: they are usually employed in olefin metathesis (Grubbs’ catalyst) or as an
alternative to phosphorus ylides in the Wittig reaction. The NHCs are different from
both other types. They form transition metal complexes that are essentially inert,
although exceptions are known.
3
1.1.2 N-Heterocyclic Carbenes

During the last 15 years, carbene chemistry has advanced in a dramatic

fashion with the preparation of persistent triplet diarylcarbenes, [30c] and the isolation of
heteroatom-substituted singlet carbenes.[36]
In 1991, Arduengo et al. reported a ground-breaking work on the synthesis of
“A Stable Crystalline Carbene”.[37] The diadamantyl-substituted imidazol-2-ylidene
was prepared by deprotonation of the corresponding imidazolium salt with sodium or
potassium hydride in the presence of catalytic amounts of tBuOK or DMSO (Figure
1.9).

Figure 1.9. Preparation of the first N-Heterocyclic Carbene.

The free carbene was isolated and found to be remarkably stable. Soon after,
it was realized that the bulky adamantyl groups in diadamantyl-substituted imidazol-
2-ylidene are not required for the stabilization of the free carbene, and a whole family
of imadazol-2-ylidenes bearing different substituents were prepared following similar
methods.[38]
The idea of synthesizing imidazole-based NHCs was to improve the stability of
the carbene center by two adjacent nitrogen atoms. Two electronegative nitrogen
atoms (σ-acceptor) stabilize the unpaired electrons in the valence shell of the
carbene carbon by the inductive effect. [39] The carbene center is also stabilized by the
lone pair electrons of the nitrogen atoms (π-donor) by reducing the electron
deficiency of the carbene carbon, called mesomeric effect (Figure 1.10).

Figure 1.10. Orbital diagram of NHC showing electronic effects.

4
 The ease of synthesis, functionalization, isolation of NHCs, and the success in
metalation with a large variety of hard/soft metals make NHCs excellent ligands with
similarities to phosphines.[18b, 36b, 40]
NHCs have the ability to strongly coordinate as both Lewis acids and Lewis
bases to a wide variety of metals throughout the periodic table with their strong sigma
donor ability.[36b, 40] This property also makes NHCs able to bind to both low- and high-
oxidation state transition metals. Although NHCs were reported as pure σ-donors in
numerous metal complexes, recent structural and theoretical studies showed that
various degree of π back-donation may occur (Figure 1.11).

Figure 1.11. Orbitals diagram of NHC.

The molecular and electronic structure of N-Heterocyclic Carbenes have been

intensively investigated.
The molecular structure of diadamantyl-substituted imidazol-2-ylidene exhibits
features characteristic for an unsaturated NHC (Figure 1.12).[37]

Figure 1.12. Solid-state molecular structure of diadamantyl-substituted imidazol-2-ylidene.

The small N—Ccarbene—N angle of 102.2o in this structure is particularly notable.

This angle is significantly reduced from the 109.7 o value found in the corresponding
imidazolium salt. Other significant structural differences between an NHC and its
imidazolium precursor include the C 2—N1/3, N1—C5, and N3—C4 distances. They are
increased from those of the imidazolium salt, while the C 4—C5 bond lengths are
reduced slightly. These structural changes indicate an increase of p character in the

5
C2—N bonds and a decrease of π-delocalization in the imidazole ring upon
deprotonation of the imidazolium ion.[41]
The stability of an NHC is mainly due to electronic effects, although steric
hindrance may also play an important role. The interaction between the carbene
centers with the π-donating, σ-accepting amino substitutes provides most of the
[42]
stabilizing energy and the aromaticity of the imidazole ring brings an additional
stabilization of about 25 kcal/mol.[42b]

1.2.2. The synthesis of NHCs

A variety of routes to prepare NHCs is available. The most convenient way for
the synthesis of NHCs is the use of imidazole containing precursors. Numerous NHC
precursors can be obtained by following various efficient routes. The synthetic
pathways include orthoformate route (Figure 1.13),[40] desulfurization of the cyclic
thiourea derivatives (Figure 1.14),[40] vacuum thermolysis of methoxy derivatives
(Figure 1.15),[40] and condensation of an amine with glyoxal (Figure 1.16).[43]

Figure 1.13. Orthoformiate route.

Figure 1.14. Desulfurization of the cyclic thiourea derivatives.

Figure 1.15. Vacuum thermolysis of methoxy derivatives.

6
 Figure 1.16. Condensation of an amine with glyoxal.

Alternatively, NHCs can be derived from a large variety of natural molecules

containing an imidazole moiety (Figure 1.17).

Figure 1.17. Examples of natural molecules containing imidazoles.

1.2.3. Reactivity and ligand properties of N-Heterocyclic Carbenes

Both saturated and unsaturated NHCs display nucleophilic reactivity. Unlike

transient carbenes, a stable NHC does not undergo cyclopropanation with normal
olefins and C—H insertion into hydrocarbons. A saturated NHC is prone to
dimerization, but an unsaturated NHC hardly dimerizes due to a large singlet-triplet
gap. Because of its high Lewis basicity, the NHC behaves like a very good donor
ligand. Reports on the use of NHCs as ligands appeared as early as the 1960s when
Őfele et al. and Wanzlick et al. prepared the first metal NHC complexes by reacting
imidazolium salts with basic metal precursors. [44] Unfortunately, the limitation of this
method hampered the potential of NHC ligands. Arduengo’s [38a] isolation of stable
carbenes immediately led to a renaissance of carbene chemistry, including its
coordination chemistry. It has been shown that NHCs can coordinate to virtually
every metal in the periodic table, in both low and high oxidation states.
7
 Surprisingly, it is not trivial to classify NHCs as either “soft” or “hard” ligands.
At first glance, given the neutral carbon donor, an NHC should be a soft ligand, and it
does exhibit many features similar to those of basic tertiary phosphine ligands. On
the other hand, NHCs are also known to coordinate to “hard” metal fragments such
as high-valent early transition metals, lanthanide, and actinide ions. This mixed
character indeed makes NHCs potentially useful for supporting various redox events
at a coordinated metal center.

1.3. Metal-NHC complexes

Many metal-NHC complexes have been synthesized until now, and most of
them, especially those from coinage metal, are suitable for application in organic
chemistry as well for biochemical studies. There are several synthesis pathways,
which usually involve the reaction between a metal oxide like copper(I) and silver(I)
oxides and imidazolium salts, both in organic solvents or water [ Dalton trans., 2010,
39, 4489-4491].

In case of gold-NHC, synthesis can involve direct reaction (with deprotonation

of the imidazolium salt by LiHDMS, method A) and later an Au salt, but usually this
leads to low yields. Other approaches (method B) which leads to better yields, is the
transmetalation of a silver-precursor. [M.V. Baker et al. / Journal of Organometallic
Chemistry 690 (2005) 5625–5635]:

8
Through direct reaction of Ag2O with an imidazolium salt , M. Pellei et al. successfully
synthesized novel bis-carbene silver complexes functionalized with amides in 2012:

This confirming the Ag(I) bases (Ag2O, AgOAc, and Ag2CO3) as good reagents of
choice in the general synthesis of silver-NHC .

1.3.1 Anti-cancer properties

The discovery of the antitumor properties of cisplatin (Figure 1.30) by

Rosenberg has proven that not only drugs composed of the basic elements of the
periodic table (carbon, oxygen, nitrogen) but also those containing heavy elements
may be toxic for tumor cells. Cisplatin, along with its second generation analogues,
represents the most widely used chemotherapeutic agents.

Figure 1.30. Worldwide approved anti-cancer platinum based drugs.

However, cisplatin displays several limitations that restrict its utility to a great
extent. For example, cisplatin is effective only for a narrow spectrum of tumor cells
and additionally, because of its poor aqueous solubility (1 mg/mL), is administered
intravenously. Moreover, the various toxicity in not target tissues like nephrotoxicity,
neurotoxicity, hematogenesis, etc., associated with cisplatin, further complicates its

9
usage. Therefore discovering new complexes selectively active on cancerous cells
life cycle, in an anti-proliferative and/or pro-apoptotic manner, remains a challenge.
Several NHC complexes of group 11 metals have been studied as an
alternative to cisplatin. Before the evaluation of N-heterocyclic carbene metal
complexes as metallopharmaceuticals, it is necessary to underline the imidazolium
salts properties as antibacterial and antifungal. [74] Pernak and co-workers[75] examined
a number of 3-alkylthiomethyl-1-ethylimidazolium chlorides (Figure 1.31). The
antimicrobial activity was evaluated against Staphylococcus aureus,
Gaffkyatetragena, Sarcinalutea, Klebsiellapneumoniae, Serratia, marcescens,
Rhodothorulaglutinis, and Bacillus subtilis. It was found that the activity was related
to the length of the alkyl chain. Pernak investigated a series of 3-alkoxymethyl-1-
methylimidazolium salts with the effects of three different anions, Cl −, BF4− and PF6−.
[76]

Figure 1.31. 3-Alkylthiomethyl-1-ethylimidazolium chlorides and 3-alkoxymethyl-1-

methylimidazolium salts studied by Pernak and co-workers.

The minimum inhibitory concentration (MIC) and the minimum bactericidal or

fungicidal concentration (MBC) values were determined against the bacterial strains
M. luteus, S. epidermidis, S. aureus, methicillin resistant S. aureus, E. hirae, E. coli,
P. vulgaris, K. pneumoniae, and P. aeruginosa, and the fungal strains C. albicans
and R. rubra.
Although it was shown that the salts did possess antibacterial and antifungal
activities, their efficacy depended greatly on the length of the alkyl chain. The type of
anion did not exert any notable effect on the activity of the salts. The most active
compound against most bacterial strains was the 3-dodecyloxymethyl-1-
methylimidazolium salt.
Additional studies done by Cetinkayaet al.verified the same trends as
observed earlier.[77] They explored the effects of lipophilic substituents, anions, and
backbone substituents on the antimicrobial activity associated with a series of 1,3-

10
diazolidinium salts and the effect of ring size of pyrimidinium salts as a comparison
(Figure 1.32).

Figure 1.32. Structures of 1,3-diazolidinium and pyrimidinium salts bearing different alkyl
groups.

The complexes were tested against a number of bacterial organisms including

E. coli, S. epidermidis, S. aureus, E. faecalis, E. cloacae, P. aeruginosa, and C.
albicans and were compared to ampicillin, a β-lactam antibiotic used to treat general
bacterial infections. The most notable feature observed among 30 derivatives
synthesized is the remarkable enhancement of the antimicrobial activity against
Gram-negative and Gram-positive bacteria of three derivatives of 1,3-diazolidinium
salts by the addition of two mesityl or two mesitylmethyl substituents on both nitrogen
atoms (bulky and lipophilic groups). Variation of the counter-ion of the 1,3-
diazolidinium salts, with R1 and R3 as mesitylmethyl, greatly affected their
antimicrobial activity.[78]
Furthermore, variations to the backbone such as substitutions from hydrogens
to methyl, cyclohexyl or benzene groups were determined to decrease the activity of
the NHC salts. Lastly, the antimicrobial activity of several derivatives of 1,3-
diazolidinium salts was directly compared to that of pyrimidinium salts with identical
substituents and it was established that the ring size plays a crucial role. The
imidazolidinium salts were much more active than the pyrimidinium salts. [78]

1.3.2 Copper(I)-NHCs complexes

Biological and bioinorganic studies about copper(I)-NHCs compounds are

mainly dealing with phosphine containing complexes. However, copper(I)-NHCs have
anti-cancer activity and high stability in numerous conditions. [79] This relatively high
stability would allow them to reach biological targets inside the cell. Cu(I)-NHC
complexes could react with intracellular oxygen producing Reactive Oxygen Species
(ROS) which would lead to oxidative attack of DNA as reported for bleomycin.

11
 Figure 1.33. A copper(I)-NHC complex having anti-cancer activity.

Gasser and coworkers[79] reported that the copper NHC complex (Figure 1.33)
was more cytotoxic than cisplatin. This complex induced apoptosis and, unlike
cisplatin, arrested the cell cycle progression in the G1 phase.

1.3.3. Ag(I)-NHCs complexes

Silver has been recognized as an effective antimicrobial agent, specifically in

the form of silver nitrate since the 17th and 18th centuries.[80]
Although the cytotoxic effects of silver against gram-positive and gram-
negative bacteria have long been established, the mechanisms of action are not
completely understood. Sporadic studies of the cell toxicity mechanisms suggest that
silver ions kill organisms through a variety of ways. One of these could be the
interfering effect of Ag ions once penetrated through the lipid membrane of bacteria,
where can affect the cell metabolism by enzymatic interaction [Z. Anorg. Allg. Chem.
2011, 637, p.389]. This effect is mostly showed when the complex has a relatively
high lipophilicity, such has for the compound:

which is highly active against S. aureus [Z. Anorg. Allg. Chem. 2011, 637,
p.389].
12
 Other kind of silver-NHC complex which was shown to be a promising
antitumor molecule is the double carbene complex bearing two amide-functionalized
imidazole rings. During in-vitro trials, it showed an activity similar to that of cisplatin in
HCT-15 colon and A549 lung cancer cells.

Although silver is generally nontoxic to humans, it is widely known that a

prolonged and excessive exposure to silver causes the development of a rare and
irreversible pigmentation of the skin (argyria) and/or the eyes (argyrosis). This is a
characteristic blue-black discoloration formed by the interaction of silver ions with
melanin and proteins in the wound exudates to give silver sulfide. [81] Besides argyria,
other toxic effects of silver may include upper and lower respiratory tract irritation.

1.3.4 N-Heterocyclic Carbenes in Homogeneous Catalysis

Current interests in metal complexes of NHC ligands arise from reports of their
remarkable activity in homogeneous catalysis. In 1995, Hermann et al. found that
palladium(II) bis-carbene complexes reported in Figure 1.35 are very efficient
catalysts for Heck-coupling reactions.[90] The carbene ligands in these complexes
appear to stabilize both Pd(II) and Pd(0) species resulting in very high catalytic
turnover numbers. This study provided, as stated in the title of Hermann’s paper, “a
new structural principle” for catalyst design. Subsequently, numerous metal
complexes bearing NHC ligands have been synthesized and show improved
activities in various catalytic reactions including C—N coupling, [91] hydrosilation,[92]
ethylene/carbon monoxide copolymerization,[93] C—H activation,[94] hydrogenation,[95]
hydroformylation,[96] furan synthesis,[97] dehalogenation,[98] atom-transfer radical
polymerization,[99] and particularly C—C bond formation (Suzuki, [100] Heck,[101]
13
Sonogashira,[102] Stille,[103] and Grignard[104] cross coupling reactions) and olefin
metathesis[43, 105] reactions.

Figure 1.35. Palladium bis-carbene complexes that catalize Heck reactions.

Recent efforts are directed toward functionalizing the carbene fragment of

NHC-metal complexes of coinage metals in order to improve catalytic activity in
several organic reaction. On this side, good results were obtained by Ambroz
Almassy et al. [Organometallics 2010, 29, 2484-2490], where a sulfonated
imidazolylidene complex of gold(I) (in zwitterion form) showed high activity in the
catalysis of alkyne selective hydration in aqueous media even in absence of an acid
co-catalyst. [Organometallics 2010, 29, 2488]

The superior performance of the NHC catalysts in these reactions results from
the pronounced donor properties of NHC ligands. Experiments have confirmed that

14
typical NHCs are more electron-releasing than even the most basic phosphines. [106]
NHC complexes have high thermal and hydrolytic durability and reactions can often
be carried out in the air. Unlike phosphines, NHCs hardly dissociate from the metal
ions thus there is no need for excess of ligands. NHCs stabilize both low and high
valent species formed in catalysis resulting in higher turnover numbers and longer
catalyst lifetimes. In light of the vast numbers of applications and their distinctive
advantages, NHCs have now become a guiding “concept” in organometallic
catalysis.[40]

1.2 Zwitterionic ligands

The term “zwitterion” is known back in 1894, used for the first time by the
German chemist Friedrich Wilhelm Küster. Its proper meaning is “hermaphrodite ion”
, relating to the double-charged nature of these kind of molecules. As a matter of fact,
a typical zwitterion is a molecule with a positive and a negative charge coexisting at
same time with zero net charge. This feature gives several properties to the
molecule, such as water solubility which could be controlled through pH, hence
increased bio-availability giving the possibility to better deliver a chemical drug in
biologic environment.
The most famous zwitterionic molecule is the amino acid, the “building block”
of human body, which consist of a common skeleton bearing a positive charged
amino- moiety and a negative charged carboxylic acid moiety.
As zwitterions are a broad class of compounds, they can be associated also
with functionalised carbenes. Lars-Arne Schaper and co-workers [ Angew. Chem. Int.
Ed. 2012, 51, 2–22] discussed these compounds in particular with CO 2- ion:

15
And SO3- :

The final aim adding these functionality to a molecule (thus leading a proper
zwitterion) is to achive a good water solubility.
Making this in NHC-ligand design through sulfonates is extremely common, used in
detergents and surfactants for more than 70 years, sulfonate groups are known for
excellent water solubility arising from their high acidity. The zwitterions formed from
sufonates are used sometimes as ionic liquids [M. Yoshizawa, M. Hirao, K. Ito-Akita,
H. Ohno, J. Mater. Chem. 2001, 11, 1057 – 1062], but also in inorganic and
organometallic synthesis, taking advantage of their non-coordinating behevior.
[ Angew. Chem. Int. Ed. 2012, 51, 5 ].
The general route to prepare N-functionalised carbene with sulfonates is the use of
propane- or butanesultone, which were already used in the past to generate mono-
and bis(imidazolium), bis(triazolium) and benzimidazolium salts . [ Angew. Chem. Int.
Ed. 2012, 51, 5].

1.1. Nitrogen mustards

Mustard in organic chemistry is a common name for molecules derived from the well-
known mustard gases, with common structure of bis(2-chloroethil) sulphide:

Cl Cl
S

They became sadly known because of their extreme toxic properties, being used as
a blistering agent during World Wars I and II. However, lots of studies were lately
16
conducted in order to better understand their biological role, the action mechanism
and so the possible use in medicine. As early as 1919 it was known that mustard gas
was a suppressor of hematopoiesis [Krumbhaar EB (1919). "role of the blood and the
bone marrow in certain forms of gas poisoning". JAMA 72: 39–41 ]. Further studies
suggested the development of mustards with other atoms in place of sulphur:

The nitrogen mustard was one of the most promising compound, its toxicity is lower
than sulphur analogues, and it displays high reactivity toward nucleophiles, being
useful intermediate in organic chemistry [Org. Biomol. Chem., 2012, 10, 8786–8793 |
8789].
Also coordination compounds of nitrogen mustards are known, with some of them
having interesting biological properties. For example, carbonato and oxalato
cobalt(III) nitrogen mustard complexes show hypoxia-selective cytotoxicity in vitro.

17
The coordination of the nitrogen lone pairs to cobalt(III) suppresses the toxicity of the
nitrogen mustard because the electron pair is no longer available to act as an internal
nucleophile. This prevents the formation of the reactive aziridinium ion intermediate
in normal tissue, but not in hypoxic tissues (tumor environment) where cobalt can be
reduced to Co(II) with formation of highly cytotoxyc species [2 0 0 4 D a l t o n T r a n
s . , 2 0 0 4 , 6 1 1 – 6 1 8].

In human cells, free nitrogen mustard was found able to bind to the N7 nitrogen on
the DNA base guanine, crosslinking two strands and preventing cell duplication.
Because of this, nitrogen mustards form a class of versatile anticancer drugs used in
the treatment of various cancers including Hodgkin’s disease, non-Hodgkin’s
lymphoma, chronic leukemia, and lung, ovarian, and breast cancer.
As underlined by computational studies, two subsequent electrophilic attacks on two
guanine moieties can afford bifunctional adducts with both inter- and intrastrand
cross-links in DNA that are reminiscent of the binding mode of cisplatin. These cross-
links and the resulting distortions of the double helix of DNA are believed to be
responsible for the cytotoxic action, as they are expected to block or at least interfere
with the passage of polymerases, thereby inhibiting DNA transcription, which
ultimately leads to cell apoptosis [J. Org. Chem. 2012, 77, 5914−5921].

One of the problems to overcome is that these agents exhibit severe host
toxicity due to their poor selectivity toward cancer cells. One approach to reduce the
toxicity of cross-linking agents for normal cells is to use a prodrug and trigger it in
tumor cells by activation with oxidation, reduction, or photolysis. Also linking the
active moiety of the molecule to a particular receptor overexpressed in tumor cells,
for example the estrogen receptor in breast and ovarian tumors, led to good result in

18
this field [(a)Shawn,M.H.;Marquis, J.C.; Zayas, B.;Wishnok, J.S.;Liberman,R. G.;
Skipper, P. L.; Tannenbaum, S. R.; Essigmann, J. M.; Croy, R. G. Mol. Cancer Ther.
2006, 5, 977]. Another possibility, studied by Yunyan Kuang et al. [J. Am. Chem.
Soc. 2011, 133, 19278–19281], is to exploit the bigger amount of oxidative stress
found in tumor cells, due to bigger concentration of ROS (Reactive Oxygen Species),
to activate the drug once inside the body. They used boronic esters to provide good
and non-toxic leaving group for the following reaction, as boric acid and derivatives
are highly and safely metabolized in the body:

Nitrogen mustards also afford variable in-vivo efficiency and toxicity in

function of pH values. George S. Tarnowskia et al. analyzed the influence of different
pH values for the compound HN2 (Bis(2-chloroethyl)methylamine) in mice bearing 1-
day-old Ehrlich ascites tumor. This led to a greater prolongation of median survival
time and increased to a larger extent the number of survivors after 30 days when
HN2 was dissolved in vehicles with initial pH values of 5.6, 7.5 or 10.5 than with other
initial pH values. Moreover, when a toxic dose of HN2 were administered, it was
better tolerated (with prolonged survival time) if it administered in vehicles with the
initial pH values of 2 or 4. [Biochemical Pharmacology ,Volume 17, Issue 6, June
1968, Pages 989–1002].

19
 To conclude, the versatility, cheapness and well-understood action
mechanisms of nitrogen mustards and related compounds are surely good reasons
to study them in the field of antitumor drugs, as well as their coordination chemisty.

2. Experimental Section

2.1. Methods and materials

All reagents were purchased from Aldrich and used without further purification.
The bis(imidazol-1-yl)methane,[107] [CH2(Im)2] was prepared in accordance with the
literature method. Some syntheses and handling were carried out under an
atmosphere of dry oxygen-free dinitrogen, using standard Schlenk techniques. All
solvents were dried, degassed and distilled prior to use. Elemental analyses (C, H, N,
S) were performed in house with a Fisons Instruments 1108 CHNS-O Elemental
Analyser. Melting points were taken on an SMP3 Stuart Scientific Instrument. IR
spectra were recorded as neat from 4000 to 600 cm -1 with a Perkin-Elmer Spectrum
One System. IR annotations used: br = broad, m = medium, s = strong, sbr = strong
broad, sh = shoulder, w = weak.1H- and 13
C-NMR spectra were recorded on a
Oxford-400 Varian spectrometer (400.4 MHz for 1H and 100.1 MHz for 13C). Chemical
shifts () are quoted relative to internal SiMe 4. NMR annotations used: s = singlet, d=
doublet, t = triplet, q=quartet. Electrospray mass spectra (ESI-MS) were obtained in
positive- or negative-ion mode on a Series 1100 MSD detector HP spectrometer,
using an acetonitrile or water mobile phase. The compounds were added to reagent
grade methanol to give solutions of approximate concentration 0.1 mM. These
solutions were injected (1 L) into the spectrometer via a HPLC HP 1090 Series II
fitted with an auto sampler. The pump delivered the solutions to the mass
spectrometer source at a flow rate of 300 L min-1, and nitrogen was employed both
as a drying and nebulizing gas. Capillary voltages were typically 4000 V and 3500 V
for the positive- and negative-ion mode, respectively. Confirmation of all major

20
species in this ESI-MS study was aided by comparison of the observed and predicted
isotope distribution patterns, the latter calculated using the IsoPro 3.0 computer
program.

21