6,6′-Dimethyl-2,2′-bipyridine-4-ester: A pivotal synthon for building tethered bipyridine ligands

Article abstract of DOI:10.1016/j.tet.2009.06.111

We describe an efficient and scalable synthesis of 4-carbomethoxy-6,6′-dimethyl-2,2′-bipyridine starting from easily available substituted 2-halopyridines and based on the application of modified Negishi cross-coupling conditions. This compound is a versatile starting material for the synthesis of 4-functionalized 2,2′-bipyridines bearing halide, alcohol, amine, and other functionalities, suitable for conjugation to biological material (2a-c, 3a-g). The utility of this compound in the construction of more complex architectures was further demonstrated by the synthesis of two bifunctional lanthanide chelators; an open chain ligand based on one 2,2′-bipyridine unit and a cryptand based on three 2,2′-bipyridine units [N₂(bpy)₃COOMe]. In the field of luminophoric biolabels, the photophysical properties of the corresponding Eu(III) cryptate are reported.

Full text of DOI:10.1016/j.tet.2009.06.111

Tetrahedron 65 (2009) 7673–7686
Contents lists available at ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
6,60-Dimethyl-2,20-bipyridine-4-ester: A pivotal synthon for building tethered
bipyridine ligands
,
,
,
b
a
,
,b
Fabien Havas a b, Nadine Leygue a b, Mathieu Danel a , Beatrice Mestre b, Chantal Galaup a
,
´
Claude Picard a
,b,
*
a
`
´
´ ˆ
CNRS, Laboratoire de Synthese et Physico-Chimie de Molecules d’Interet Biologique, SPCMIB, UMR-5068, 118 Route de Narbonne, F-31062 Toulouse cedex 9, France
b Universite´ de Toulouse, UPS, Laboratoire de Synthe`se et Physico-Chimie de Mole´cules d’Inte´rˆet Biologique, SPCMIB, 118 route de Narbonne, F-31062 Toulouse cedex 9, France
a r t i c l e i n f o
a b s t r a c t
We describe an efficient and scalable synthesis of 4-carbomethoxy-6,60-dimethyl-2,20-bipyridine starting
from easily available substituted 2-halopyridines and based on the application of modified Negishi cross-
coupling conditions. This compound is a versatile starting material for the synthesis of 4-functionalized
2,20-bipyridines bearing halide, alcohol, amine, and other functionalities, suitable for conjugation to
biological material (2a–c, 3a–g). The utility of this compound in the construction of more complex ar-
chitectures was further demonstrated by the synthesis of two bifunctional lanthanide chelators; an open
chain ligand based on one 2,20-bipyridine unit and a cryptand based on three 2,20-bipyridine units
[N2(bpy)3COOMe]. In the field of luminophoric biolabels, the photophysical properties of the corre-
sponding Eu(III) cryptate are reported.
Article history:
Received 4 March 2009
Received in revised form 21 June 2009
Accepted 25 June 2009
Available online 2 July 2009
Keywords:
Cross-coupling
Negishi reaction
2,20-Bipyridines
Bifunctional ligand
Lanthanide
Ó 2009 Elsevier Ltd. All rights reserved.
Luminescence
1. Introduction
[N2(bpy)3] are well established16 while bioassays using a such
probe have considerable commercial application. It has been ap-
Much attention is given to 2,20-bipyridine (bpy) systems be-
cause of their central role in the fields of inorganic, organometallic,
and coordinations chemistries. Bpy ligands generally afford effi-
cient binding affinity for most elements in the periodic table due to
plied as a luminescent label in direct homogeneous immunoassays,
in in situ hybridization and in PCR detection, and finds wide ap-
plications in diagnostic and high throughput screening for drug
discovery (HTRFÒ technology).17 Functionalized bpy derivatives
have commonly been required for all the purposes described above.
Especially, the covalent attachment of lanthanide organocomplexes
to biological materials is an important step in the development of
luminescent bioprobes. In this direction, 2,20-bipyridine is an ideal
core for introduction of a single conjugation group opposite to the
complexing sites of the molecule. The introduction of a single re-
the chelation effect and the p
-accepting ability of these moieties.1 It
was shown that the transition metal–bpy complexes found various
applications in many areas of chemistry including catalysis,2 elec-
trochemistry,3 artificial photosynthesis,4 luminescent sensor mol-
ecules,5 nonlinear optical (NLO) materials6 and organic light
emitting diodes (OLEDs).7 Besides, bpy derivatives are attractive
building blocks for supramolecular chemistry and macromolecular
chemistry.8 Lanthanide complexes of bpy-based ligands have been
also the focus of much attention owing to their important appli-
cations as time-resolved luminescence labels in bioanalyses and
medical diagnostic.9–11 As a matter of fact, bpy fragments act as
effective sensitizer for causing visible luminescence from Eu(III),
Tb(III), Sm(III), Dy(III) centers and near-infrared (NIR) luminescence
from Yb(III) and Nd(III) centers.12–15 In particular, the luminescence
properties of the europium complex of the Lehn cryptand
active group precludes undesirable intra- and intermolecular cross-
18–20
linkings in conjugation reactions with biomolecules.
As
a matter of fact, cross-linking may have dramatic impact on the
biological and immunogenic properties of the ligand-biomolecule
conjugate. The already described procedures of Mukkala et al. af-
ford access to 4-substituted 2,20-bipyridine derivatives that allow
bioconjugation of luminescent lanthanide chelates.21 In this work,
the introduction of a single active functionality at the 4-position
was performed via a multistep procedure including the mono-N-
oxide formation of 6,60-dimethyl-2,20-bipyridine, the subsequent
introduction of a nitro group at the 4-position of bpy and further
derivatizations of the nitro functionality to create derivatives suit-
able for bioconjugation. Nevertheless, the synthesis of 4-
* Corresponding author. Tel.: þ33 5 61 55 62 96; fax: þ33 5 61 55 60 11.
E-mail address: picard@chimie.ups-tlse.fr (C. Picard).
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.06.111

7674
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
substituted-bpy moieties has been much less explored that ones
that are symmetrically disubstituted at the 4,40-position.22 As
a consequence, limited accessibility to unsymmetrically function-
alized bpy derivatives has restricted their potential use in the
construction of more sophisticated architectures.
properties of 6a$Eu3þ cryptate are presented and compared to
those of the corresponding bifunctionalized [N2(bpy)3(CO2Me)2
-
$Eu] cryptate (6b$Eu3þ), commercially available for HTRFÒ assays.
2. Results and discussion
As part of a research program aimed at the design and synthesis
of lanthanide luminescence bioprobes,12,13,23 we were interested in
the synthesis of ligands based on a bpy moiety containing an ap-
propriate monofunctionalization to be covalently attached to
bioactive molecules. In this direction, 4-carbomethoxy-6,60-di-
methyl-2,20-bipyridine (1, Scheme 1) is an useful synthon.
2.1. Synthesis of 4-carbomethoxy-6,60-dimethyl-2,20-
bipyridine (1)
In principle, the synthesis of the targeted bpy compound (1), 4-
carbomethoxy-6,60-dimethyl-2,20-bipyridine, can mainly be
achieved through three different routes: a) introduction of the
functional ester group into symmetrical bpy derivative (6,60-di-
methyl-2,20-bipyridine), b) ring-assembly methodology like the
Kro¨hnke procedure or its various procedures, c) preliminary func-
tionalization of two different pyridine components followed by
coupling methodology generally employed for the formation of
aryl–aryl bonds. The two former approaches are limited by not
easily accessible starting compounds, multi-step procedures with
low yields and (or) require harsch conditions.27,28 For the latter
approach, palladium-catalyzed cross-coupling procedures, such as
Stille-type and Negishi-type allow a more flexible preparation of
unsymmetrically substituted and functionalized bipyridines.29–32
On this basis, the synthesis of 1 was approached via a metal-cata-
lyzed coupling of the two pyridine fragments 6 and 9, which are
amenable to large laboratory scale preparations (Scheme 2). The
synthetic strategy for preparing these two starting materials is
depicted in Scheme 2.
R1
COOMe
1
2
N
N
N
N
R1 = COOH, CH2OH,
CONH-(CH2)2-NH2
4
3
CH2O(CH2)6R2
R3
N
N
N
N
X
X
R
3 = COOMe, X = Br
R2 = OH, Br, N3, NH2, C CH,
NHCOCH2Br, maleimide
R3 = CH2OTBDPS, X = Cl
COOMe
R6
R5
a
5
6
6
N
O
NH2
OEt
N
Br
N
N
N
N
N
N
N
N
COOEt
CN
N
N
N
N
R4O2C
CO2R4
CO2R4
b
R4O2C
EtO
N
H
O
7
O
R4 = tBu, R5= COOMe, CH2OH,
CH2OCH2C CH
R4 = H, R5 = COOH
6a R6 = H
6b R6 = COOMe
c
Scheme 1. Structures of 2,20-bipyridine derivatives.
COOH
COOMe
d
9
8
N
H
O
N
Cl
In this compound, the aromatic ester group opens avenues to
the introduction of various functional groups suitable for biological
labeling. The methyl groups are available for different functionali-
zation reactions allowing the introduction of bpy unit in multi-
functional chelating agents. Despite their prevalence, the synthesis
of 6,60-dimethyl-2,20-bipyridine compound bearing an ester func-
tion at the 4-position was reported only recently. The methyl ester
derivative 1 was prepared from a Stille-type cross-coupling re-
action requiring a pyridyltriflate derivative as coupling component
to provide satisfying yield.24 The corresponding ethyl ester de-
rivative was prepared by using a combination of Stille and Negishi
cross-coupling methodologies with a moderate yield.25 In this re-
port we describe a full account of our efforts to develop a scalable
and friendly preparation of 1 and the achievement of this goal via
a Negishi’s zinc coupling.26 We have also investigated the utility of
the methyl ester function and methyl groups of 1 by elaboration of
a number of 6,60-dimethyl-2,20-bipyridine derivatives asymmetri-
cally substituted at the 4-position with various moieties (com-
pounds 2 and 3, Scheme 1) and by the conversion of the methyl
groups into bromomethyl and chloromethyl functions (compounds
4, Scheme 1). As examples of practical applications in the direction
of time-resolved luminescent Ln(III) probes, we describe the fur-
ther elaboration of the monofunctionalized open chain ligands (5)
and the [N2(bpy)3(CO2Me)] cryptand (6a). The photophysical
Scheme 2. Preparation of starting materials. Reagents and conditions; (a) HBr, Br2,
NaNO2, H2O, ꢀ20 ꢁC (75%); (b) i) EtONa, rt, 12 h; ii) cyanoacetamide, 80 ꢁC, 6 h; iii)
acetic acid (68%); (c) HCl 6 N, reflux, 24 h (91%); (d) i) POCl3, reflux, 18 h; ii) MeOH, rt,
24 h (83%).
The reagent 2-bromo-6-methylpyridine 6 can be obtained
commercially or synthesized from cheap 2-amino-5-methylpyr-
idine by a Sandmeyer/halogenation sequence with HBr, bromine
and sodium nitrite.33 This reaction could be adapted to a 150 g scale
using a literature procedure.34 The trifunctionalized pyridine 9,
bearing an ester group was obtained in three steps starting from
diethyl oxalate by improvement of literature procedures.35–37 In
a first step, the sodium salt of ethyl acetopyruvate was generated in
situ by the treatment of diethyl oxalate with acetone in the pres-
ence of sodium ethoxide and this was reacted with cyanoacetamide
to give the 2-pyridone derivative 7 in 68% yield. Upon hydrolysis of
the nitrile group and concomitant loss of carbon dioxide of 7 in
aqueous 6 N HCl, acide 8 was produced in 91% yield. Using this
route, 8 can be readily prepared in a multi-gram scale using
a simple precipitation procedure at each step for purification.
Treatment of 8 with phosphorous oxychloride following by meth-
anol then afforded the targeted pyridyl derivative 9 in 83% yield
after a short column chromatography on silica gel. Compound 9

F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
7675
was thus obtained with an overall yield of 51% starting from diethyl
oxalate.
With compounds 6 and 9 in hands, we began our synthetic
studies by finding the optimal methodology for the desired cross-
coupling of these two partners (Scheme 3).
chloroquinoxaline and 3-pyridylmagnesium bromide inspired us to
attempt a similar coupling through Grignard reagent derived from
6. Disappointingly, no coupling product was isolated.
2.2. Compound 1: derivatization of the aromatic ester group
O
10
B
N Ph
The pivotal building block 1 with an ester group can be used as
a synthetic handle to introduce other functional groups such as
carboxylic acid, alcohol, amine (Scheme 4).
N
O
9
d
COOH
9, f
9, e
SiMe3
1
12
11
Bu3Sn
N
N
N
N
N
N
g
9
2a
b
a
a
c
6
6
ClZn
N
13
COOMe
CH2OH
b
Scheme 3. Syntheses of 1 used in this study. Reagents and conditions: (a) i) nBuLi, THF,
ꢀ78 ꢁC; ii) Me3SiCl, rt, 18 h (55%); (b) i) nBuLi, THF, ꢀ78 ꢁC; ii) Bu3SnCl, rt, overnight
(96%); (c) i) nBuLi, THF, ꢀ78 ꢁC; ii) ZnCl2, rt, 0.5 h; (d) PdCl2(PPh3)2, K3PO4, CuI, DMF,
100 ꢁC, 8 h (45%); (e) PdCl2(PPh3)2, PPh3, CuI, DMF, 120 ꢁC, 48 h according to Ref 40; (f)
Pd(PPh3)4, toluene, reflux, 48 h (30–66%); (g) PdCl2(PPh3)2, DIBALH, THF, reflux, 4 h
(83%).
N
N
N
1
2b
c
CONH(CH2)2NH2
Firstly, in order to carry out cross-coupling reactions under
Suzuki–Miyaura conditions, we used boronic ester 10 which has
been shown to be stable and is now commercially available.38
According to the procedure of Jones et al.,39 the coupling of
1.2 equiv of boronic ester 10 relative to chloropyridine 9 was per-
formed in the presence of 5 mol % PdCl2(PPh3)2 and CuI. In these
conditions, the desired bpy product 1 was obtained in modest
isolated yield (45%). The next attempts were made using the
Hiyama cross-coupling, between 9 and pyridylsilane 11, prepared
from 6 via halogen–lithium exchange and subsequent reaction with
chlorotrimethylsilane. By using the catalyst system recently
proposed by Pierrat et al. (PdCl2(PPh3)2/PPh3/CuI) for the palla-
dium-catalysed Hiyama cross-coupling of 2-bromopyridine and
chloropyridyltrimethyl silanes,40 these attempts were unsuccessful
and no cross-coupling product was isolated. These results high-
lighted again that the lack of electron-withdrawing substituent on
the pyridine ring of 2-trimethylsilylpyridines prevents the forma-
tion of the final aryl–aryl bond.40 We have also used the 2-tribu-
tylstannylpicoline 12, prepared as previously reported,41 as reactive
partner in a Stille-type reaction. Cross-coupling reaction was car-
ried out in degassed toluene in the presence of a catalytic amount of
Pd(PPh3)4 (5 mol %), and afforded the desired bipyridine 1, but
yields tend to be variable (30–60%). Although a classical acidic
aqueous workup was employed for removing tin byproducts,42 the
use of stannane compound give rises to difficulties in product pu-
rification (this may in part be responsible for the variation in yield).
The preparation of 1 by using Negishi-type chemistry and orga-
nozinc reagent was more convenient, being more reliable and
providing a better yield. The pyridylzinc chloride compound 13 was
satisfactorily prepared from bromide 6 by halogen–metal exchange
with n-butyllithium, followed by transmetallation with ZnCl2.
Heterocoupling reaction of 13 with 9 in the presence of 5 mol % of
a catalyst43 prepared from PdCl2(PPh3)2 and DIBALH (2 equiv) in
refluxing THF afforded compound 1 in 83% yield after column
chromatography. This process also benefits from the fact that it is
conveniently carried out in a single pot and allows the extension to
large-scale preparations (up to 10 g) of the versatile bipyridine 1,
with no significant loss of yield or efficiency. Finally, it is worth
mentioning that we also tried to prepare 1 by using a simple iron
salt as environmentally friendly catalyst. A precedent44 from
Fuernstner’s group for the Fe(acac)3 catalyzed coupling of
N
2c
Scheme 4. Derivatization of 1. Reagents and conditions. (a) K2CO3, MeOH/H2O, 80 ꢁC,
0.5 h (100%); (b) NaBH4, EtOH, rt, 16 h (82%); (c) NH2CH2CH2NH2, rt, 24 h (100%).
The ester function was hydrolysed in mild basic conditions to
the corresponding acid 2a and was easily reduced to hydroxy-
methyl group (2b) with NaBH4 in methanol. Moreover, the reaction
of the ester compound 1 with a large excess (30 equiv) of neat
ethylenediamine produces the aminoethylamide 2c cleanly and
quantitatively after 24 h at room temperature. It is worth noting
that the amido-amino compound 2c could be further coupled for
vectorisation to various biomolecules using commercial hetero-
bifunctional cross-linking reagents.17,45
The hydroxymethyl function of 2b offers another potential
source of further derivatization, through a biologically stable ether
linkage. In this direction, a representative series of bpy derivatives
bearing alcohol, bromide, amine and other functionalities, and
a classical alkyl tether of six atoms spacer was obtained (Scheme 5).
Access to the hydroxyl-terminated bpy derivative 3a required
deprotonation of 2b with NaH and reaction of the resultant alco-
holate with commercial (6-bromohexyloxy)-tert-butyldimethylsi-
lane. The resulting ether 14 was cleanly deprotected with TBAF to
yield target alcohol in 46% yield over the two steps from 2b. Sim-
ilarly, after deprotonation of 2b, the direct preparation of bromo-
terminated derivative 3b was possible utilizing a 2.6-fold excess of
1,6-dibromohexane. In this way compound 3b can be prepared in
50% yield. According to the classical Gabriel synthesis methodology,
amino-terminated bpy derivative 3c was prepared from 3b in a two
step procedure via its phthalimido precursor 15. The purified yield
of 15 was 78%, and this was converted into 3c in 96% yield by
treatment with hydrazine hydrate.
Derivatizing bpy agents containing haloacetamidyl (3d) or
maleimide (3e) moiety were also prepared. These electrophilic
groups are commonly used in biological setting and react selec-
tively with thiol groups either extant or introduced into bio-
molecules.46 The bromoacetamide derivative was made from the
amine 3c using the bifunctional bromoacetyl chloride reagent
(1.2 equiv) in the presence of N-methyl morpholine (2.5 equiv) in
72% yield. Two different routes for the preparation of the

7676
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
CH2O(CH2)6OTBDMS
CH2O(CH2)6- C CH
CH2OH
i
a
N
N
N
N
N
N
N
N
N
N
N
14
2b
3g
b
c
O
CH2O(CH2)6OH
CH2O(CH2)6Br
CH2O(CH2)6
N
d
O
N
g
N
N
N
3a
3b
15
h
e
O
CH2O(CH2)6N3
CH2O(CH2)6NH2
CH2O(CH2)6
N
O
N
N
f
N
3f
3c
3e
CH2O(CH2)6NHCOCH2Br
N
N
3d
Scheme 5. Derivatization of 2b. Reagents and conditions. (a) i) NaH, DMF, 60 ꢁC, 1 h; ii) Br(CH2)6OTBDMS, 60 ꢁC, 24 h (57%); (b) TBAF, THF, rt, 3 h (80%); (c) i) NaH, DMF, 60 ꢁC, 1 h;
ii) Br(CH2)6Br (2.6 equiv), 60 ꢁC, 20 h (50%); (d) potassium phthalimide, DMF, 100 ꢁC, 5 h (78%); (e) NH2NH2, MeOH, rt, 3 h (96%); (f) ClCOCH2Br, N-methyl morpholine, CH2Cl2, rt,
18 h (72%); (g) maleimide, PPh3, DEAD, THF, 0 ꢁC, 12 h (54%); (h) NaN3, DMF, rt, overnight (99%); (i) i)NaH, THF, 60 ꢁC, 1 h; ii) 8-bromo-1-octyne, reflux, 16 h (36%).
maleimide derivative 3e were investigated. The classical method
requires the reaction of amines with maleic anhydride, followed by
dehydration/cyclisation of the maleimic acid intermediates, usually
promoted by acid.47 An alternative method described by Walker
using alcohols as starting materials involves the direct N-alkylation
of maleimide under mild Mitsunobu conditions.48 Attempts to
prepare 3e by treating amine 3c with maleic anhydride followed by
sodium acetate in acetic acid were disappointing in terms of both
yield (<20%) and reproducibility. The desired maleimide derivative
can be conveniently prepared by exploiting the Mitsunobu re-
action. Thus, treatment of alcohol 3a with maleimide in the pres-
ence of DEAD and PPh3 furnished 3e in 54% yield.
COOCH3
CH2OH
N
N
N
N
N
N
N
1
2b
a
c
CH2OTBDPS
COOMe
N
17
16
O
To provide additional bpy reagents of general utility, we have
also investigated the introduction of azide and alkyne moieties.
These functional groups are inertness towards physiological con-
ditions, which allows the use of the Huisgen 1,3-dipolar cycload-
dition under copper (I) catalysis for in vitro and in vivo
bioconjugation.49 The bromo derivative 3b was easily transformed
into the respective azido derivative 3f in a nearly quantitative yield
by treatment with sodium azide at room temperature. The syn-
thesis of acetylene compound 3g began with the preparation of 8-
bromo-1-octyne in a two steps procedure. Firstly, 2-octyn-1-ol was
subjected to a Zipper reaction (triple bond isomerization) with
mixed alkali metal amide50 to afford terminal isomer 7-octyn-1-ol.
In this latter compound, the hydroxyl group was then replaced by
bromine group by reaction with NBS/PPh3 using standard meth-
odology, yielding 8-bromo-1-octyne in 60% yield. Reaction of this
bromo acetylene compound with sodium alcoholate of 2b afforded
3g in moderate yield (36%).
O
d
b
CH2OTBDPS
COOMe
N
N
N
N
18
O
O
4a
Br
Br
e
CH2OTBDPS
N
N
19
OCOCH3
OCOCH3
f
CH2OTBDPS
CH2OTBDPS
g
N
N
N
N
2.3. Compounds 1 and 2b: activation of the methyl groups at
the 6,60-positions
4b
20
Cl
Cl
OH
OH
Scheme 6. Activation of methyl groups of 1 and 2b. Reagents and conditions: (a) m-
CPBA, CHCl3, rt, 19 h (95%); (b) i) Ac2O, 120 ꢁC, 16 h; ii) HBr, AcOH, 70 ꢁC, 7 h (63%); (c)
TBDPSCl, imidazole, DMF, rt, 48 h (94%); (d) m-CPBA, CHCl3, rt, 24 h (78%); (e) Ac2O,
100 ꢁC, 24 h (68%); (f) K2CO3, MeOH/H2O, rt, 3 h (100%); (g) SOCl2, rt, 1.5 h (64%).
In pivotal compounds 1 and 2b, the methyl groups at the 6,60-
positions are available for different functionalization reactions
which are depicted in Scheme 6.

F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
7677
Initial attempts to obtain dibromide 4a directly from 1 by a free-
SO2NHNa
+
radical bromation afforded product in low yield. Specifically, 1 was
refluxed in benzene under irradiation in the presence of N-bromo-
succinimide and a catalytic amount of AIBN. All trials produced
a mixture of brominated products from which the targeted com-
pound was isolated in only 20% yield, at best, after careful purifica-
tions by column chromatography and recrystallisation. Moreover,
carrying out a selective debromination of polybrominated by-
products with diisobutylaluminium hydride51 or diethylphosphite
and N,N0-diisopropylethylamine,52 only slight improvements in
overall yield were obtained. Therefore, a more circuitous procedure
was employed by using the well-known Boekelheide rearrangement
followed by a pseudohalogen exchange.27 The 1,10-dioxide com-
pound 16 was readily prepared (95%) by oxidation of 1 with
3-chloroperoxybenzoic acid (m-CPBA) in chloroform at room tem-
perature. After treatment of 16 with acetic anhydride at reflux, the
resulting rearranged diacetate was displaced with hydrobromic acid,
33% in acetic acid, to give dibromide 4a in 63% yield. Stepwise con-
version of1 to 4awas thusmadein two steps with a totalyield of 60%.
The monoprotected triol 20 was obtained in four steps starting
from alcohol 2b and following the methodology developed for the
1/4a conversion. After protection of the hydroxyl group of 2b with
tert-butyldiphenylsilyl chloride in the presence of imidazole, sub-
sequent treatment with m-CPBA resulted in the formation of bis N-
oxide 18 (94 and 78% yield respectively for these two steps). When
bis N-oxide 18 was refluxed with acetic anhydride for 24 h, bis ester
19 was generated as the major product and was isolated after col-
umn chromatography in 68% yield. After attempting various base-
hydrolysis procedures on 19, it was found that the use of potassium
carbonate in a MeOH/H2O mixture effected a quantitative trans-
esterification to give 20. Finally, diol 20 was converted to the
dichloride 4b by treatment with thionyl chloride (64% yield).
Stepwise conversion of 2b to 4b was thus made in an overall yield
of 32%. It can be noticed, that 20 may be obtained more directly
from bis N-oxide 18 using the same methodology, without isolating
the intermediate bis ester 19 in 63% yield.
N
N
N
N
(a)
R N
N R
N
N
Br
Br
22, R = Ts
23, R = H
21
(b)
(c)
COOMe
COOMe
N
N
N
N
N
N
N
(d)
N
N
N
N
N
N
+
Ln3+
Na
N
N
N
6a.Eu3+, 6a.Gd3+
6a.Na+
Scheme 8. Synthesis of cryptates 6a$Naþ and 6a$Ln3þ. Reagents and conditions: (a)
EtOH, reflux 24 h (65%), according to ref 53; (b) H2SO4, 120 ꢁC, 2 h (88%) according to
ref 53; (c) 4a, Na2CO3, CH3CN, reflux, 48 h, [reactants]¼1.5.10ꢀ3 M (47%); (d) LnCl3. 6H2O
(1.1 equiv), MeOH, reflux, 24 h.
(BSA) as a model protein. Activation of the noncoordinated aro-
matic carboxylate function of 5b$Eu3þ was achieved with EDCI$HCl
in DMF and N-hydroxysuccinimide (NHS). In our experimental
conditions, a labeling ratio (number of 5b$Eu3þ per BSA) of 1.6 was
established by MALDI-TOF mass spectrometry (See Experimental
part).
Scheme 7 shows also the synthesis of acetylenic compound 5d,
suitable for ‘click-chemistry’. Starting from 5a, it was obtained in
a two step sequence: i) selective reduction of the methyl ester
group with NaBH4 and ii) alkylation of the hydroxyl group with
propargyl bromide.
The Lehn cryptand [N2(bpy)3] monofunctionalized by a methyl
ester moiety 6a is accessible by a stepwise procedure involving the
synthesis of the intermediate macrocyclic bis(bipyridinediyl)di-
amine 23 and its subsequent bridging (Scheme 8).
2.4. Application: synthesis of multidentate ligands
In the last part of this work, we used dibromide 4a in the con-
struction of architectures suitable for complexation of lanthanide
ions (Schemes 7 and 8).
Alkylation of the secondary amine of di-tert-butyl iminodiace-
tate with 4a was carried out in the presence of a mineral base to
reach compound 5a in excellent yield. Deprotection of the ester
groups in aqueous 6 N HCl gave quantitatively the penta-acid 5b. In
preliminary experiments, the suitability of the 5b$Eu3þ complex for
protein labeling was investigated using Bovine Serum Albumin
The macrocycle 23 was obtained as previously described.53
Condensation of the dibromo bipyridine fragment 21 with tosyla-
mide monosodium salt in EtOH at reflux afforded the ditosylated
compound 22 (65% yield) which was deprotected in concentrated
H2SO4 by heating giving 23 in a 88% yield. Treatment of macrocyclic
diamine 23 with dibromide 4a was carried out in acetonitrile
(reflux, 48 h) and in the presence of sodium carbonate as a base and
COOMe
COOH
COOH
5a
5b
5b.Eu3+
4a
N
N
N
N
N
N
a
b
c
+
-
-
O2C
CO2
N
Eu3+
-
N
N
N
N
N
COOtBu
COOtBu
HOOC
tBuOOC
-
COOH
COOH
COOtBu
COOtBu
O2C
CO2
HN
HOOC
tBuOOC
d
CH2OCH2C CH
CH2OH
5c
5d
e
N
N
N
N
N
N
N
N
tBuOOC
tBuOOC
tBuOOC
COOtBu
COOtBu
COOtBu
tBuOOC
COOtBu
Scheme 7. Synthesis of open chain ligands 5. Reagents and conditions: (a) Na2CO3, CH3CN, reflux, 17 h (91%); (b) aqueous 6 N HCl, 90 ꢁC, 72 h (100%); (c) EuCl3$6H2O, 1.1 equiv, H2O,
rt, 24 h; (d) NaBH4, MeOH, rt, 2 h (93%); (e) i) NaH, DMF, rt, ii) propargyl bromide, rt, 48 h (41%).

7678
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
without using high-dilution techniques (reactant conditions
1.5ꢂ10ꢀ3 M). Cryptand 6a was isolated as its NaBr complex after
purification by column chromatography (47% yield).
complex. By using the well-established isotope effect and the
Horrocks equation54 the hydration number (i.e., the number of
coordinated water molecules) is determined to be 2.3. After cor-
rection for the weaker effect of outer-sphere water molecules,55
this value is reduced to 2.1. We can also notice that, in D2O solution,
the lifetime of the 5D0 level is temperature independent, indicating
that thermally activated radiationless decay paths play any role in
the decay of the metal-centered emitting state. The modest value
measured for the emission quantum yield (1.8%) is attributed to
nonradiative deactivation via the O–H vibrations of the two inner
sphere water molecules and to the presence of low energy ligand-
to-metal charge-transfer excited state (LMCT), as commonly ob-
served for europium complexes with bpy-type cryptands.15,56
However, as reported in the TRACEÒ technology,57 the addition of
fluoride ions may be used for enhancing the luminescence effi-
2.5. Photophysical properties of the tethered trisbipyridine
Eu(III) cryptate 6a$Eu3D
The Ln3þ complexes (Ln¼Eu, Gd) of the ligand 6a were prepared
by ion exchange from 6a$Naþ. Sodium ion was displaced by simply
refluxing for 24 h the sodium cryptate in the presence of a slight
excess of LnCl3$6H2O in methanol solution. The resulting 6a$Ln3þ
complexes were isolated by precipitation with diethyl ether.
At room temperature 6a$Eu3þ cryptate is luminescent in the red
domain when excited into the lowest-energy LC absorption band
(306 nm), indicating that the energy is absorbed by the surround-
ing ligand and efficiently transferred to the chelated Eu3þ ion.
Sensitized Eu3þ emission is observed corresponding to 5D0/7FJ
(J¼0–4) transitions, the dominant band in the emission spectrum is
the transition 5D0/7F2 at 617 nm (Fig. 1).
ciency of europium cryptate. When 6a$Eu3þ (10
mM) is incubated at
room temperature in the presence of a saturating anion concen-
tration (0.4 M, Fꢀ), a 4-fold increase of the total emission intensity
is observed.
For 6a$Gd3þcryptate, ligand excitation at 77 K and time-re-
solved measurement at 77 K reveal
a broad emission band
extending from 400 to 750 nm, with a maximum at 487 nm, which
is assigned to the phosphorescence of the coordinated cryptand
(Fig. 1). The lowest excited state (6P7/2) of the Gd3þ ion lies above
(c)
31,000 cmꢀ1 58
and thus cannot accept energy from the ligand; as
,
a result, 6a$Gd3þ cryptate displays ligand-centered photophysical
process only. The position of the lowest cryptand-centered excited
triplet state was located at 461 nm (21,700 cmꢀ1), determined from
the highest energy phosphorescence feature of the spectrum
(Fig. 1) and the corresponding lifetime is 4.30 ms. This lifetime is
substantially shorter than that measured for parent sodium
cryptate (second timescale), indicating a great perturbation of the
electron density of the macrobicycle by the metal ion and sug-
gesting a strong interaction Ln3þ-ligand.
J = 2
(a)
(b)
As one can see from data gathered in Table 1, the luminescence
properties (
6a$Eu3þ are similar to those reported for the corresponding di-
substituted europium cryptate 6b$Eu3þ 59
The small differences
s, F) and the estimated hydration state of the metal in
J = 1
.
observed are close to the experimental error. The modification in
the triplet state energy is too small to alter the efficiency of the
energy transfer from the ligand to the europium ion. Clearly, like its
disubstituted analogue, 6a$Eu3þ may be used as a luminescent
marker for HTRF assays but without the potential drawback of the
occurrence of undesirable intra- and intermolecular cross-linking
in bioconjugation reactions.
J = 4
J = 0
J = 3
200
300
400
500
600
700
800
λ/nm
Figure 1. (a) excitation (250–450 nm, lem¼617 nm) and (b) emission (lexc¼306 nm)
spectra of the 6a$Eu3þ cryptate in water at 298 K. The emission bands arise from the
5D0/7FJ transitions; the J values are shown on the spectrum. (c) emission spectrum
(
lexc¼306 nm) of the 6a$Gd3þ cryptate in water/glycerol (7/3) at 77 K.
3. Conclusion
The luminescence lifetimes (s) and quantum yields (F) were
After attempting several methodologies generally employed for
the synthesis of unsymmetrical 2,20-bpy derivatives, it was found
that a modified Negishi cross-coupling reaction was the most ef-
fective for the multigram preparation of 4-carbomethoxy-6,60-di-
methyl-2,20-bipyridine. This bpy derivative is a versatile building
block for a wealth of further transformations, as demonstrated by
some examples. Subsequent transformations of the aromatic ester
measured under various experimental conditions. The results are
summarized in Table 1 where some pertinent data previously
obtained for the corresponding methyl ester disubstituted de-
rivative [N2(bpy)3(CO2Me)2. Eu] (6b$Eu3þ) are also shown for com-
parison purposes. The value of the luminescence lifetime in H2O
solution at 298 K (0.36 ms) is diagnostic of an aquo europium
Table 1
Absorption maxima (lmax in nm), absorption coefficients (emax in Mꢀ1 cmꢀ1), luminescence lifetimes (
energies (ET in cmꢀ1) for 6a$Eu3þ and its corresponding disubstituted analogue 6b$Eu3þa
s in ms), quantum yields (F%), hydration state (q) and triplet state
298 K
lmax
emax
s2H98 K
s
s2D98 K
s7D7 K
F2H98 K
F
qb
ET
c
298 K
H;Fꢀ
H;Fꢀ
6a$Eu3þ
306
306
27,500
26,400
0.36
0.34
1.17
d
1.70
1.50
1.70
1.50
1.8
2
7.0
d
2.1
2.2
21,700
21,800
6b$Eu3þd
a
Data obtained in aerated water, H2O (H) or D2O (D) solutions; in the absence or in the presence of 0.4 M KF (Fꢀ).
Number of coordinated H2O molecules calculated using the equation q¼1.11 (1/sHꢀ1/sDꢀ0.31).55
From phosphorescence spectra at 77 K of 6a$Gd3þ and 6b$Gd3þ complexes.
Data from ref 59.
b
c
d

F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
7679
group to various functionalities were successfully carried out,
4.2. 3-Cyano-6-methyl-2-oxo-1,2-dihydro-pyridine-4-
carboxylic acid ethyl ester (7)
opening avenues for further grafting of biological material by
classical methods or ‘click chemistry’. Using the methods described
here, it will be possible to synthesize 4-subsituted 2,20-bipyridine
derivatives with spacers of any desired length and functionalized
tails. We have also shown that the methyl groups at the 6,60-posi-
tions can be simply converted into bromomethyl and chloromethyl
groups, which are common moieties for the development of pre-
organized bpy-containing architectures. As first examples in the
direction of luminescent lanthanide complexes, an open chain li-
gand based on one 2,20-bipyridine unit and a cryptand based on
three 2,20-bipyridine units were synthesized. The preliminary la-
beling experiments and photophysical studies of their corre-
sponding Eu(III) complexes nominate these ligands as candidates
for luminescent labeling of biological material.
Absolute ethyl alcohol (300 mL) was placed in a round-bot-
tomed flask and purged with argon. Sodium (13.5 g, 0.59 mol) was
added per portions over a period of 1 h and the solution was stirred
overnight. A mixture of diethyl oxalate (68 mL, 0.50 mol) and ace-
tone (37 mL, 0.50 mol) was added dropwise, observing the pre-
cipitation of a solid turning from white to yellow. The resulting
slurry was stirred for 3 h at rt. After this time, cyanoacetamide
(42.0 g, 0.50 mol) was added and the mixture was heated to 80 ꢁC
for 6 h. Ethanol was removed and the solid residue was dried under
vacuum. The resulting solid was dissolved in boiling water
(135 mL), and then acetic acid (10 mL) was added, whereupon
a precipitate was formed. The orange-coloured precipitate was
collected by filtration and dried under vacuum to give 7 (70.3 g,
0.34 mol, yield 68%). Mp: 214–215 ꢁC. (litt63 mp 213 ꢁC). Rf (Silica
gel, 6% MeOH in CH2Cl2): 0.50. IR ymax: 3444, 2840, 2228 (C^N),
1733 (C]O ester), 1646 (C]O amide), 1251. 1H NMR (250 MHz,
4. Experimental
4.1. General methods
DMSO-d6)
d
: 1.31 (t, 3H, J¼7.1), 2.32 (s, 3H), 4.35 (q, 2H, J¼7.1), 6.55
Reactions requiring an inert atmosphere were run under Argon.
THF and Et2O were freshly distilled from Na/benzophenone, ace-
tonitrile was freshly distilled from P2O5, and CH2Cl2 was freshly
distilled from CaH2. Butyllithium was titrated before use with
menthol, using 1,10-phenanthrolin as indicator. Thin-layer chro-
matography was performed on Merck silica or alumina plates with
a fluorescence indicator. TLC spots were visualised by irradiation
with UV light or by exposure to iodine vapours (Rf values refer to
relative mobilities on TLC plates). Column chromatography was
(s, 1H). 13C NMR (75 MHz, DMSO-d6)
d
: 14.1 (CH3), 19.7 (CH3), 63.0
(CH2), 99.4 (Cq), 104.8 (CH), 115.1 (Cq), 147.6 (Cq), 154.6 (Cq), 161.5
(Cq), 163.2 (Cq). MS (ESIþ): m/z (%) 207 (15) [MþH]þ, 229 (100)
[MþNa]þ, 245 (43) [MþK]þ.
4.3. 6-Methyl-2-oxo-1,2-dihydropyridine-4-carboxylic acid (8)
Cyanoester 7 (15.0 g, 72.8 mmol) was added to 140 mL of 6 M aq
HCl, and the resulting mixture was heated to a gentle reflux with
stirring for 24 h. The hot reaction mixture was poured onto approx.
250 g of ice. A fine beige solid precipitated, which was collected by
vacuum filtration, washed with 30 mL of ice water, and dried under
vacuum, to give acid 8 (10.1 g, 66.0 mmol, yield 91%) as a pale beige
solid. Mp: >260 ꢁC dec. Rf (Silica gel, CH2Cl2/MeOH 1:1): 0.40. IR
ymax: 3424, 2930, 1718 (C]O acid), 1641 (C]O amide), 1444, 1234.
carried out on silica gel (Merck, 60–200
mm, porosity 60 Å) and on
alumina (Macherey-Nagel, activity IV, 50–200
mm).
Melting points were taken with a Bu¨chi mel-temp apparatus.
Infrared spectra were recorded on a Perkin–Elmer 883 spectro-
photometer. Samples were prepared as KBr pellets (solid sample) or
applied to NaCl plates (liquid sample). Selected characteristic ab-
sorbances are reported in cmꢀ1. Proton magnetic resonance spectra
(1H NMR) were recorded at 250 or 300 MHz on a Bruker AC
250 MHz or Bruker Avance 300 MHz spectrometer and are reported
1H NMR (250 MHz, DMSO-d6)
changeable protons), 6.37 (s, 1H), 6.61 (s, 1H). 13C NMR (75 MHz,
d: 2.23 (s, 3H), 3.59 (br s, ex-
DMSO-d6) d: 19.2 (CH3),103.2 (CH),118.0 (CH),143.1 (Cq),147.2 (Cq),
as follows: chemical shift
d
in parts per million (multiplicity, num-
163.7 (Cq), 166.6 (Cq). MS (ESIꢀ): m/z (%) 152 (100) [MꢀH]ꢀ.
ber of protons, coupling constant J in hertz). Residual protic solvent
was used as the internal reference, setting chloroform to
d
7.26.
4.4. 2-Chloro-6-methylpyridine-4-carboxylic acid methyl
ester (9)
Carbon magnetic resonance spectra (13C NMR) were recorded at
75 MHz on a Bruker Avance 300 MHz spectrometer. Chemicals
shifts are quoted in parts per million, referenced to the appropriate
To a stirred solution of POCl3 (10 mL, 107 mmol) was added acid
8 (2.37 g, 15.5 mmol), and the resulting solution was heated to
reflux for 18 h. The excess POCl3 was distilled off under reduced
pressure, the residue was cooled to 0 ꢁC, methanol (20 mL) was
added dropwise, and the resulting mixture was stirred for 24 h at rt.
The reaction mixture was neutralized with solid NaHCO3, and
partitioned between H2O (30 mL) and EtOAc (30 mL). The aqueous
layer was extracted with EtOAc, (2ꢂ30 mL) and the combined or-
ganic layers were treated with active carbon, then filtered through
Celite, dried (Na2SO4), and the solvent was removed under reduced
pressure. The crude product was purified by flash chromatography
over silica gel (CH2Cl2), to give ester 9 (2.40 g, 12.9 mmol, yield 83%)
solvent peak, taking chloroform as
d 77.0. Electrospray (ES) mass
spectra were obtained on a Perkin–Elmer SCIEX API 100 and Waters
Q-TOF spectrometers. Elemental analyses were carried out by the
‘Service d0Analyse’, Laboratoire de Chimie de Coordination (Tou-
louse). Absorption measurements were done with a Hewlett Pack-
ard 8453 temperature controlled spectrophotometer.
The Eu(III) and Gd(III) luminescence emission and excitation
spectra were acquired using a LS-50B Perkin–Elmer spectrofluo-
rimeter equipped with a Hamamatsu R928 photomultiplier tube
and with the low-temperature accessory No. L2250136. Lifetimes
s
(uncertaintyꢃ5%) are the average values from at least ten separate
measurements covering two or more lifetimes. The phosphores-
cence decay curves were fitted by an equation of the form I(t)¼I(0)
as a white solid. Mp: 61 ꢁC. Rf (Silica gel, CH2Cl2): 0.66. IR ymax
3092, 2961, 1736 (C]O), 1557, 1429, 1303, 1226, 1166. 1H NMR
(CDCl3, 300 MHz) : 2.60 (s, 3H), 3.95 (s, 3H), 7.63 (s, 1H), 7.68 (s,
:
exp (ꢀt/
s
) using a curve-fitting program. The luminescence quan-
d
tum yields (uncertaintyꢄ15%) were determined by the method
1H). 13C NMR (75 MHz, CDCl3)
d: 24.1 (CH3), 52.8 (CH3), 120.8 (Cq),
described by Haas and Stein,60 using as standard [Ru(bpy)3]2þ in
aerated water (
the solvent.
121.2 (Cq), 140.3 (Cq), 151.3 (Cq), 160.5 (Cq), 164.5 (Cq). MS (ESIþ):
F
¼0.028)61 and corrected for the refractive index of
m/z (%) 186 (100) [MþH]þ, 208 (35) [MþNa]þ.
The following compounds were prepared as described in the
literature: 2-Bromo-6-methylpyridine 6,33 2-tributylstannyl-6-
methylpyridine 12,41 6,6-Bis (bromomethyl)-2,20-bipyridine 20,62
macrocyclic diamines 21 and 22.53
4.5. 2-Methyl-6-(trimethylsilyl)-pyridine (11)
To a stirred solution of bromopyridine 6 (1.00 g, 5.81 mmol) in
anhydrous THF (10 mL), at ꢀ78 ꢁC, was added, dropwise over

7680
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
15 min, nBuLi (1.3 M in hexanes, 4.9 mL, 6.4 mmol). The resulting
red solution was stirred at ꢀ78 ꢁC for 3 h, followed by the dropwise
concentrated under pressure to give a residue, which was purified
by flash chromatography over alumina (petroleum ether/Et2O
90:10), to give bipyridine 1 (7.60 g, 31.4 mmol, yield 83%) as a white
solid.
addition over 10 min. of TMSCl (900 mL, 7.00 mmol). The resulting
mixture was allowed to warm up to rt, and stirring was continued
for 18 h. The reaction mixture was quenched by addition of H2O
(12 mL), and extracted with Et2O (3ꢂ15 mL). The combined organic
layers were dried (Na2SO4), and the solvent was removed under
reduced pressure. The crude product was purified by flash chro-
matography over silica gel (petroleum ether/Et2O 90:10 then
80:20), to give pyridylsilane 11 (530 mg, 3.21 mmol, yield 55%) as
a pale yellow liquid. IR ymax: 3053, 2957, 1574, 1444, 1248, 1141. 1H
4.6.4. Compound 1
Mp: 122ꢀ123 ꢁC (litt.24 116–117 ꢁC). Rf (alumina, petroleum
ether/Et2O 90:10): 0.50. IR ymax: 2958, 1733 (C]O), 1571, 1429,
1340, 1255. 1H NMR (300 MHz, CDCl3)
d: 2.64 (s, 3H), 2.69 (s, 3H),
3.97 (s, 3H), 7.18 (d, 1H, J¼7.8), 7.69 (t,1H, J¼7.8), 7.71 (s, 1H), 8.19 (d,
1H, J¼7.8), 8.71 (s, 1H). 13C NMR (75 MHz, CDCl3)
d: 24.6 (CH3), 52.5
NMR (300 MHz, CDCl3)
d
: 0.33 (s, 9H), 2.59 (s, 3H), 7.04 (dd, 1H,
(CH3),117.7 (CH),118.3 (CH),122.2 (CH),123.5 (CH),137.0 (CH),138.6
(Cq), 155.1 (Cq), 157.1 (Cq), 158.1 (Cq), 159.0 (Cq), 166.2 (Cq). MS
(DCI/NH3): m/z (%) 243 (100) [MþH]þ. Anal. Calcd for C14H14N2O2:
C, 69.41; H, 5.82; N, 11.56. Found: C, 69.25; H, 5.75; N, 11.44.
J¼7.8, 0.75), 7.31 (d, 1H, J¼7.2), 7.46 (t, 1H, J¼7.5). 13C NMR (75 MHz,
CDCl3)
d
: ꢀ1.7 (CH3), 24.9 (CH3), 122.3 (CH), 125.7 (CH), 133.9 (CH),
158.3 (Cq), 167.6 (Cq). MS (DCI/NH3): m/z (%) 166 (31) [MþH]þ.
4.6. 4-Carbomethoxy-6,60-dimethyl-2,20-bipyridine (1)
4.7. 6,60-Dimethyl-2,20-bipyridine-4-acetic acid (2a)
4.6.1. Via Suzuki–Miyaura cross-coupling reaction
A solution of K2CO3 (28.4 mg, 0.21 mmol) in water (2.5 mL) was
added to a stirred solution of ester 1 (50 mg, 0.21 mmol) in meth-
anol (5.5 mL). The solution was heated to 80 ꢁC for 0.5 h. The sol-
vent was removed under reduced pressure, and the aqueous layer
was acidified with 0.5 N HCl to pH 3.5–4.0. A white precipitate
separated upon cooling. The solid was collected by centrifugation
and dried under reduced pressure to give 2a (47 mg, 0.21 mmol,
yield 100%). Mp: >200 ꢁC dec. IR ymax: 2928, 1711 (C]O), 1592,
A mixture of pyridyl chloride 9 (0.63 g, 3.4 mmol), 2-pyr-
idylboronic ester 10 (1.15 g, 4.08 mmol), PdCl2(PPh3)2 (119 mg,
0.17 mmol), anhydrous K3PO4 (1.5 g, 7 mmol), and CuI (325 mg,
1.7 mmol) in anhydrous DMF (40 mL) was heated to 100 ꢁC under
Argon for 6 h. The reaction mixture was warmed up to rt, diluted
with diethyl ether (50 mL), washed with water (2ꢂ20 mL), dried
over MgSO4 and the solvent was removed under reduced pressure.
The solid residue was purified by flash chromatography (alumina,
1571. 1H NMR (300 MHz, CDCl3)
d: 2.62 (s, 3H), 2.66 (s, 3H), 7.31 (d,
petroleum ether/Et2O 90:10) to give bipyridine
1.53 mmol, yield 45%) as a white solid.
1
(370 mg,
1H, J¼7.6), 7.77 (d, 1H, J¼0.9), 7.81 (t, 1H, J¼7.7), 8.13 (d, 1H, J¼7.8),
8.58 (d, 1H, J¼0.9). 13C NMR (75 MHz, CDCl3)
d: 24.2 (CH3), 24.35
(CH3), 119.2 (CH), 120.0 (CH), 123.8 (CH), 125.1 (CH), 139.05 (CH),
141.5 (Cq),156.0 (Cq),157.7 (Cq),159.5 (Cq),160.55 (Cq),168.35 (Cq).
MS (DCIþ/CH4): m/z (%) 229.1 (100) [MþH]þ, 257.1 (17) [MþC2H5]þ.
4.6.2. Via Stille cross-coupling reaction
To a stirred degassed solution of pyridyl chloride 9 (0.63 g,
3.4 mmol) and of pyridylstannane 12 (1.53 g, 4.0 mmol) in toluene
(20 mL) was added Pd(PPh3)4 (200 mg, 0.17 mmol), and the
resulting mixture was heated to reflux for 24 h. The reaction mix-
ture was filtered over Celite and concentrated aqueous HCl (20 mL)
was added to the residue, and the resulting mixture was washed
with CH2Cl2 (3ꢂ20 mL). The aqueous layer was neutralized with
solid NaHCO3, and extracted with CH2Cl2 (3ꢂ20 mL). The latter
organic layers were dried (Na2SO4) and the solvent was removed
under reduced pressure. The crude product was purified by flash
chromatography (alumina, petroleum ether/Et2O 90:10) to give
bipyridine 1 (540 mg, 2.23 mmol, yield 66%) as a white solid.
4.8. 6,60-Dimethyl-4-hydroxymethyl-2,20-bipyridine (2b)
To a solution of ester 1 (4.84 g, 20.0 mmol) in anhydrous ethanol
(40 mL) was added NaBH4 as a solid (1.90 g, 50.2 mmol) with stir-
ring at rt under Argon. The resulting mixture was further stirred for
16 h at rt. A solution of satd aq NH4Cl (20 mL) was added, and the
resulting mixture was partitioned between CH2Cl2 (50 mL) and H2O
(50 mL). The aqueous layer was extracted with CH2Cl2 (2ꢂ50 mL),
the combined organic layers were dried (Na2SO4) and the solvent
was removed under reduced pressure. The crude product was pu-
rified by flash chromatography over silica gel (CH2Cl2/MeOH 98:2
then 90:10), to give alcohol 2b (3.51 g, 16.4 mmol, yield 82%) as
a white solid. Mp: 72–74 ꢁC. IR ymax: 3268, 2919, 1589, 1569, 1460,
4.6.3. Via Negishi cross-coupling reaction
A
solution of n-butyllithium (1.5 M in hexanes, 45 mL,
67.5 mmol) was slowly added to a stirred solution of bromopicoline
6 (10.08 g, 58.6 mmol) in anhydrous THF (50 mL) at ꢀ78 ꢁC under
Ar in a 500-mL glass reactor. During the addition the temperature
was kept at ꢀ60 ꢁC maximum. The resulting red solution was
stirred for 30 min at ꢀ78 ꢁC, followed by the dropwise addition of
anhydrous ZnCl2 (it was used after fusion by flame-drying under
reduced pressure) (9.14 g, 67.1 mmol) in THF (150 mL). The result-
ing mixture was warmed up to rt, and stirring was continued for
1 h. In a separate 1-L glass reactor, DIBALH (1.0 M in hexanes, 4 mL,
4 mmol) was added to a stirred solution of PdCl2(PPh3)2 (1.46 g,
2.08 mmol) in anhydrous THF (150 mL), at rt under Argon. The
yellow solution turning black was stirred for 30 min before the
addition of a solution of pyridyl chloride 9 (7.04 g, 37.9 mmol) in
THF (50 ml). The resulting solution was stirred at rt for 10 min. The
pyridylzinc chloride (13) solution prepared above was then added
via canula, and the resulting yellow mixture was heated at reflux
until complete consumption of compound 9 was observed by TLC
monitoring (4 h). The reaction mixture was cooled to rt, then satd
aq NaHCO3 solution (400 mL) was added. The aqueous layer was
extracted with Et2O (3ꢂ250 mL) and the organic extracts were
1417, 1069, 803. 1H NMR (250 MHz, CDCl3)
d: 2.64 (s, 3H), 2.67 (s,
3H), 4.72 (s, 2H), 7.13 (s, 1H), 7.17 (d, 1H, J¼7.6), 7.69 (t, 1H, J¼7.8),
8.08 (s, 1H), 8.13 (d, 1H, J¼7.6). 13C NMR (75 MHz, CDCl3)
d: 24.52
(CH3), 24.54 (CH3), 63.6 (CH2), 115.8 (CH), 118.5 (CH), 120.5 (CH),
123.2 (CH), 137.1 (CH), 151.1 (Cq), 155.8 (Cq), 156.0 (Cq), 157.9 (Cq),
158.2 (Cq). MS (ESIþ): m/z (%) 215.3 (100) [MþH]þ, 237.2 (29)
[MþNa]þ. Anal. Calcd for C13H14N2O: C, 72.87; H, 6.59; N, 13.07.
Found: C, 72.73; H, 6.30; N, 12.89.
4.9. N-(2-Aminoethyl)-6,60-dimethyl-2,20-bipyridine-4-
carboxamide (2c)
Ester 1 (100 mg, 0.41 mmol) was added per portions to ethyl-
enediamine (759 mg, 12.6 mmol) at rt. The mixture was stirred at rt
for 24 h, observing the consumption of the starting material on TLC.
Excess of ethylenediamine was eliminated under high vacuum to
give 2c as a grey solid (111 mg, 0.41 mmol, yield 100%). Mp¼150–
151 ꢁC. IR ymax: 3363, 3278, 3046, 2908, 2860, 1651 (C]O), 1563. 1H
NMR (300 MHz, CDCl3)
d: 2.63 (s, 3H), 2.67 (s, 3H), 2.97 (t, 2H,
J¼5.8), 3.55 (q, 2H, J¼5.8), 7.18 (d, 1H, J¼7.8), 7.60 (s, 1H), 7.70 (t, 1H,

F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
7681
J¼7.8), 8.21 (d, 1H, J¼7.8), 8.44 (s, 1H). 13C NMR (75 MHz, CDCl3)
d
:
EtOAc (50 mL), washed with H2O (3ꢂ15 mL), dried (Na2SO4), and
the solvent was removed under reduced pressure. The crude
product was purified by flash chromatography over silica gel
(CH2Cl2/MeOH 97:3), to give bromide 3b (361 mg, 0.96 mmol, yield
50%) as a yellow oil. Rf (alumina, CH2Cl2/petroleum ether 60:40):
24.55 (CH3), 24.6 (CH3), 41.2 (CH2), 42.6 (CH2), 114.7 (CH), 118.4
(CH), 121.0 (CH), 123.6 (CH), 137.2 (CH), 142.8 (Cq), 155.1 (Cq), 156.6
(Cq), 157.9 (Cq), 159.2 (Cq), 166.35 (Cq). MS (ESIþ): m/z (%) 271.3
(100) [MþH]þ.
0.48. 1H NMR (300 MHz, CDCl3)
d: 1.41–1.45 (m, 4H), 1.62–1.67 (m,
4.10. 4-[6-(tert-Butyldimethylsilanyloxy)hexyloxymethyl]-
2H), 1.83–1.88 (m, 2H), 2.57 (s, 6H), 3.34 (t, 2H, J¼6.8), 3.46 (t, 2H,
6,60-dimethyl-2,20-bipyridine (14)
J¼6.4), 4.50 (s, 2H), 7.12 (d, 1H, J¼7.6), 7.16 (s, 1H), 7.65 (t, 1H, J¼7.7),
8.01 (s, 1H), 8.17 (d, 1H, J¼7.8). 13C NMR (75 MHz, CDCl3)
d: 24.7
To a stirred solution of alcohol 2b (146 mg, 0.68 mmol) in an-
hydrous DMF (5 mL) was added sodium hydride (35.2 mg,
1.47 mmol) in one portion under Argon. The resulting mixture was
heated to 60 ꢁC for 1 h, followed by the addition of a solution of (6-
bromohexyloxy)-tert-butyldimethylsilane (419 mg, 1.42 mmol) in
DMF (3 mL). The resulting mixture was stirred at 60 ꢁC for 24 h.
After cooling down to rt, the reaction mixture was partitioned be-
tween H2O (10 mL) and EtOAc (50 mL). The organic layer was
washed with H2O (3ꢂ10 mL), dried (MgSO4), and the solvent was
removed under reduced pressure. The crude product was purified
by column chromatography on silica gel (petroleum ether/EtOAc
90:10), to give ether 14 (166 mg, 0.39 mmol, yield 57%) as a color-
(CH3), 25.4 (CH2), 28.0 (CH2), 29.5 (CH2), 32.7 (CH2), 33.8 (CH2), 70.8
(CH2), 71.5 (CH2), 116.5 (CH), 118.3 (CH), 121.2 (CH), 123.1 (CH), 137.0
(CH), 148.8 (Cq), 155.9 (Cq), 156.0 (Cq), 157.8 (Cq), 158.2 (Cq). MS
(ESIþ): m/z (%) 377.3 (100) [MþH]þ, 379.3 (94) [MþH]þ. Anal. Calcd
for C19H25N2OBr: C, 60.48; H, 6.68; N, 7.42. Found: C, 60.47; H, 7.23;
N, 7.81.
4.13. 2-[6-(6,60-Dimethyl-[2,20]bipyridinyl-4-ylmethoxy)-
hexyl]-isoindole-1,3-dione (15)
To a stirred solution of bromide 3b (172 mg, 0.46 mmol) in DMF
(5 mL) was added potassium phthalimide (168 mg, 0.91 mmol),
and the resulting mixture was heated to 100 ꢁC for 5 h. The reaction
mixture was diluted with 50 mL of EtOAc, washed with H2O
(4ꢂ10 mL), dried (Na2SO4), and the solvent was removed under
reduced pressure. The crude product was purified by flash chro-
matography over alumina (petroleum ether/Et2O 50:50) to give
phthalimide 15 (160 mg, 0.36 mmol, yield 78%) as a white solid.
Mp: 155 ꢁC. IR ymax: 2935, 2859, 1772 (C]O), 1713 (C]O), 1585,
less oil. Rf (silica gel, petroleum ether/EtOAc 90:10): 0.16. IR ymax
2930, 2857, 1586, 1570, 1462, 1419, 1254, 1098, 836. 1H NMR
(250 MHz, CDCl3) : 0.04 (s, 6H), 0.89 (s, 9H),1.38–1.67 (m, 8H), 2.64
:
d
(s, 6H), 3.52 (t, 2H, J¼6.6), 3.61 (t, 2H, J¼6.6), 4.57 (s, 2H), 7.16 (d, 1H,
J¼7.3), 7.20 (s, 1H), 7.69 (t, 1H, J¼7.6), 8.09 (s, 1H), 8.17 (d, 1H, J¼7.6).
13C NMR (75 MHz, CDCl3)
d
: ꢀ5.3 (CH3), 18.3 (Cq), 24.6 (CH3), 25.6
(CH2), 25.92 (CH3), 25.94 (CH2), 29.7 (CH2), 32.75 (CH2), 63.1 (CH2),
71.0 (CH2), 71.5 (CH2), 116.4 (CH), 118.2 (CH), 121.1 (CH), 123.0 (CH),
136.9 (CH), 148.8 (Cq), 155.85 (Cq), 155.9 (Cq), 157.7 (Cq), 158.1 (Cq).
MS (FABþ, MNBA): m/z (%) 429.2 (100) [MþH]þ.
1573, 1396, 1370, 1115. 1H NMR (300 MHz, CDCl3)
d: 1.38–1.46 (m,
4H), 1.60–1.75 (m, 4H), 2.63 (s, 6H), 3.51 (t, 2H, J¼6.6), 3.69 (t, 2H,
J¼7.2), 4.55 (s, 2H), 7.14 (d, 1H, J¼7.5), 7.18 (s, 1H), 7.65–7.71 (m, 3H),
7.82–7.85 (m, 2H), 8.08 (s, 1H), 8.17 (d, 1H, J¼7.8). 13C NMR (75 MHz,
4.11. 6-(6,60-Dimethyl-[2,20]bipyridinyl-4-ylmethoxy)-hexan-
1-ol (3a)
CDCl3,) d: 24.6 (CH3), 25.7 (CH2), 26.7 (CH2), 28.5 (CH2), 29.5 (CH2),
37.9 (CH2), 70.9 (CH2), 71.4 (CH2), 116.5 (CH), 118.3 (CH), 121.1 (CH),
123.0 (CH), 123.1 (CH), 132.1 (Cq), 133.8 (CH), 136.9 (CH), 148.8 (Cq),
155.8 (Cq), 155.9 (Cq), 157.8 (Cq), 158.1 (Cq), 168.4 (Cq). MS (ESIþ):
m/z (%) 444.2 (100) [MþH]þ.
To a stirred solution of silyl ether 14 (118.6 mg, 0.277 mmol) in
anhydrous THF (5 mL), at 0 ꢁC under Argon, was added dropwise
tetra-n-butylammonium fluoride (1 M in THF, 3 mL, 3 mmol). The
resulting solution was stirred at 0 ꢁC for 20 min, then warmed up to
rt, and stirring was continued for 3 h. The reaction mixture was
partitioned between H2O (10 mL) and CH2Cl2 (10 mL), and the
aqueous layer was extracted with CH2Cl2 (3ꢂ10 mL). The combined
organic layers were dried (Na2SO4) and the solvent was removed
under reduced pressure. The crude product was purified by flash
chromatography over alumina (CH2Cl2/MeOH 95:5), to give alcohol
3a (70 mg, 0.222 mmol, yield 80%) as a colorless oil. IR ymax: 3368,
4.14. 6-(6,60-Dimethyl-[2,20]bipyridinyl-4-ylmethoxy)-
hexylamine (3c)
To a stirred solution of phthalimide 15 (120.2 mg, 0.27 mmol) in
methanol (3 mL) was added hydrazine monohydrate (50 ml,
1.0 mmol), and the resulting mixture was stirred at rt for 3 h. The
reaction mixture was partitioned between H2O (5 mL) and CH2Cl2
(10 mL), and the aqueous layer was extracted with CH2Cl2
(3ꢂ5 mL). The combined organic layers were dried (Na2SO4) and
the solvent was removed under reduced pressure. The crude
product was purified by flash chromatography over alumina
(CH2Cl2/MeOH 98:2), to give amine 3c (82.2 mg, 0.26 mmol, yield
96%) as a yellow oil. IR ymax: 3368, 2931, 2858, 1609, 1585, 1573,
2932, 2858, 1587, 1569, 1420, 1115. 1H NMR (300 MHz, CDCl3)
d:
1.32–1.63 (m, 8H), 2.56 (s, 6H,), 3.40 (s, 1H), 3.46 (t, 2H, J¼6.5), 3.57
(t, 2H, J¼6.5), 4.50 (s, 2H), 7.08 (d, 1H, J¼7.5), 7.11 (s, 1H), 7.62 (t, 1H,
J¼7.8), 8.03 (s, 1H), 8.10 (d, 1H, J¼7.8). 13C NMR (75 MHz, CDCl3)
d:
24.6 (CH3), 25.5 (CH2), 25.9 (CH2), 29.6 (CH2), 32.6 (CH2), 62.8 (CH2),
70.9 (CH2), 71.5 (CH2), 116.5 (CH), 118.3 (CH), 121.2 (CH), 123.0 (CH),
137.0 (CH), 148.8 (Cq), 155.8 (Cq), 156.0 (Cq), 157.8 (Cq), 158.1 (Cq).
MS (ESIþ): m/z (%) 315.2 (100) [MþH]þ, 337.35 (36) [MþNa]þ. Anal.
Calcd for C19H26N2O2: C, 72.58; H, 8.33; N, 8.91. Found: C, 72.35; H,
8.23; N, 8.77.
1462, 1420, 1115. 1H NMR (300 MHz, CDCl3)
d: 1.30–1.77 (m, 8H),
2.63 (s, 6H), 2.70 (t, 2H, J¼6.75), 3.52 (t, 2H, J¼6.5), 4.57 (s, 2H), 7.15
(d, 1H, J¼7.8), 7.19 (s, 1H), 7.68 (t, 1H, J¼7.7), 8.10 (s, 1H), 8.18 (d, 1H,
J¼7.8). 13C NMR (75 MHz, CDCl3,)
d: 24.7 (CH3), 26.1 (CH2), 26.7
(CH2), 29.7 (CH2), 33.5 (CH2), 42.1 (CH2), 71.0 (CH2), 71.5 (CH2),116.5
(CH), 118.3 (CH), 121.2 (CH), 123.1 (CH), 137.0 (CH), 148.9 (Cq), 155.9
(Cq),156.0 (Cq),157.8 (Cq),158.2 (Cq). MS (ESIþ): m/z (%) 314.4 (100)
[MþH]þ. Anal. Calcd for C19H27N3O$1H2O: C, 68.85; H, 8.82; N,
12.68. Found: C, 68.58; H, 9.05; N, 12.50.
4.12. 4-(6-Bromohexyloxymethyl)-6,60-dimethyl-2,20-
bipyridine (3b)
To a stirred solution of alcohol 2b (411 mg, 1.92 mmol) in an-
hydrous THF (5 mL) was added under Argon sodium hydride
(75 mg, 3.1 mmol). The resulting mixture was heated to reflux for
1.5 h, followed by addition of 1,6-dibromohexane (1.22 g,
5.0 mmol). The resulting solution was heated to reflux under Argon
for 20 h. After cooling to rt, the reaction mixture was diluted with
4.15. 2-Bromo-N-[6-(6,60-dimethyl-[2,20]bipyridinyl-4-
ylmethoxy)-hexyl]-acetamide (3d)
To a stirred solution of amine 3c (141 mg, 0.45 mmol) in anhy-
drous CH2Cl2 (5 mL) was added under Argon N-methylmorpholine

7682
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
(132
m
L, 1.2 mmol). The resulting solution was cooled down to 0 ꢁC,
L,
4.18. 6,60-Dimethyl-4-(oct-7-ynyloxymethyl)-2,20-
bipyridine (3g)
followed by the addition of bromoacetyl chloride (45
m
0.54 mmol). The resulting solution was stirred at rt under Argon for
18 h. The reaction mixture was partitioned between H2O (10 mL)
and CH2Cl2 (10 mL), and the aqueous layer was extracted with
CH2Cl2 (3ꢂ5 mL). The combined organic layers were dried (Na2SO4)
and the solvent was removed under reduced pressure. The crude
product was purified by flash chromatography over alumina
(CH2Cl2/MeOH 99:1), to give compound 3d (140 mg, 0.322 mmol,
yield 72%) as a yellow oil. IR ymax: 2931, 2858, 1661, 1567, 1420, 1113.
4.18.1. 7-Octyn-1-ol
Under Argon, lithium (140 mg, 20 mmol) was added to dry 1,3-
diaminopropane (10 mL), and the mixture was heated at 70 ꢁC for
3 h. At that time, the starting dark blue color was faded and a green
suspension of the lithium salt was obtained. The reaction was
cooled to rt and potassium tert-butoxide (1.34 g, 12 mmol) was
added, affording a yellow solution. After stirring for 30 min at rt, 2-
1H NMR (300 MHz, CDCl3)
d
: 1.37–1.48 (m, 4H), 1.57 (p, 2H, J¼7.1),
octyn-1-ol (425 mL, 3 mmol) was added via a syringe. The orange-
1.67 (p, 2H, J¼7.1), 2.64 (s, 6H), 3.29 (q, 2H, J¼6.7), 3.53 (t, 2H,
J¼6.4), 3.87 (s, 2H), 4.57 (s, 2H), 7.17 (d, 1H, J¼7.6), 7.19 (s, 1H), 7.70
(t, 1H, J¼7.7), 8.11 (s, 1H), 8.19 (d, 1H, J¼7.9). 13C NMR (75 MHz,
brown solution was stirred at rt for a further 2 h and the resulting
beige mixture was poured into water (50 mL). The mixture was
extracted with CH2Cl2 (3ꢂ20 mL) and the combined organic ex-
tracts were washed successively with 10% HCl and saturated NaCl
solution. After drying over MgSO4, the organic layer was evaporated
under reduced pressure to give the title compound (380 mg,
3 mmol, yield 100%) as an oil. The title compound was used without
purification for the next reaction step. Rf (silica gel, CH2Cl2): 0.17. 1H
CDCl3) d: 24.0 (CH3), 24.3 (CH3), 25.8 (CH2), 26.6 (CH2), 29.2 (CH2),
29.4 (CH2), 29.5 (CH2), 40.1 (CH2), 71.0 (CH2), 71.2 (CH2), 117.4 (CH),
119.3 (CH), 121.9 (CH), 123.9 (CH), 137.8 (CH), 150.6 (Cq), 153.9 (Cq),
154.3 (Cq),157.8 (Cq),157.9 (Cq),165.2 (Cq). MS (ESIþ): m/z (%) 434.3
(97) [MþH]þ, 436.3 (100) [MþH]þ. Anal. Calcd for C21H28N3O2Br: C,
58.07; H, 6.50; N, 9.67. Found: C, 58.34; H, 6.55; N, 9.52.
NMR (300 MHz, CDCl3)
d
: 1.3–1.6 (m, 8H), 1.90 (t, 1H, J¼2.6), 2.20
(td, 2H, J¼6.8, 2.6), 3.68 (t, 2H, J¼6.5). 1H NMR data are in good
4.16. 1-[6-(6,60-Dimethyl-[2,20]bipyridinyl-4-ylmethoxy)-
hexyl]-pyrrole-2,5-dione (3e)
agreement with the literature.64
4.18.2. 8-Bromo-1-octyne
DEAD (37
m
L, 0.235 mmol) was added dropwise to a stirred so-
To a stirred solution of 7-octyn-1-ol (1 g, 7.93 mmol) and PPh3
(2.70 g, 10.3 mmol) in CH2Cl2 (10 mL), at 0 ꢁC under Argon, was
slowly added solid N-bromosuccinimide (1.83 g, 10.3 mmol), at
such a rate that the temperature did not rise above 10 ꢁC. The
resulting orange mixture was warmed up to rt, and stirring was
continued for 2 h. The solvent was removed under reduced pres-
sure and the solid orange residue was extracted with petroleum
ether. The extract was filtered, and the solvent was removed under
reduced pressure. The crude product was purified by column
chromatography over silica gel (petroleum ether) to give the title
compound (0.9 g, 4.76 mmol, yield 60%) as a colorless oil. Rf (silica
lution of alcohol 3a (60 mg, 0.191 mmol), maleimide (22.6 mg,
0.233 mmol) and PPh3 (61 mg, 0.233 mmol) in THF (2 mL) at 0 ꢁC
under Argon atmosphere. The reaction mixture was stirred at 0 ꢁC
for 12 h and then was quenched by addition of H2O (3 mL). The
reaction mixture was evaporated to dryness and the residue was
subjected to column chromatography on silicagel (CH2Cl2/petro-
leum ether 80:20) to give maleimide 3e (41 mg, 0.104 mmol, yield
54%) as a white solid. Mp: 98 ꢁC. Rf (alumina, CH2Cl2/petroleum
ether 80:20): 0.48. IR ymax: 2933, 2853, 1701 (C]O), 1562, 1407,
1369. 1H NMR (300 MHz, CDCl3)
d: 1.3–1.7 (m, 8H), 2.62 (s, 6H), 3.50
(t, 2H, J¼6.5), 3.51 (t, 2H, J¼7.4), 4.55 (s, 2H), 6.66 (s, 2H), 7.14 (d, 1H,
gel, petroleum ether): 0.22. 1H NMR (300 MHz, CDCl3)
d: 1.3–1.6 (m,
J¼7.5), 7.17 (s, 1H), 7.67 (t, 1H, J¼7.8), 8.07 (s, 1H), 8.16 (d, 1H, J¼7.8).
6H), 1.90 (m, 2H), 2.0 (t, 1H, J¼2.6), 2.20 (td, 2H, J¼6.8, 2.6), 3.40 (t,
2H, J¼6.8). 1H NMR data are in good agreement with the
literature.64,65
13C NMR (75 MHz, CDCl3)
d: 24.7 (CH3), 25.75 (CH2), 26.6 (CH2),
28.5 (CH2), 29.6 (CH2), 37.8 (CH2), 70.9 (CH2), 71.5 (CH2), 116.6 (CH),
118.4 (CH), 121.2 (CH), 123.1 (CH), 134.1 (CH), 137.0 (CH), 148.9 (Cq),
155.9 (Cq), 156.0 (Cq), 157.9 (Cq), 158.2 (Cq), 170.9 (Cq). MS (ESIþ):
m/z (%) 394.2 (100) [MþH]þ, 416.2 (9) [MþNa]þ. Anal. Calcd for
C23H27N3O3: C, 70.21; H, 6.92; N, 10.68. Found: C, 70.47; H, 7.23; N,
10.32.
4.18.3. Compound 3g
To a stirred solution of alcohol 2b (100 mg, 0.467 mmol) in an-
hydrous THF (1.5 mL) was added under Argon sodium hydride
(22.4 mg, 0.933 mmol). The resulting mixture was heated to reflux
for 1 h, followed by addition of 8-bromo-1-octyne (265 mg,
1.4 mmol). The resulting solution was heated to reflux under Argon
for 16 h. After cooling to rt, the reaction mixture was evaporated to
dryness and the residue was subjected to column chromatography
over alumina (petroleum ether then petroleum ether/CH2Cl2 60:40)
to give acetylenic compound 3g (54 mg, 0.168 mmol, yield 36%) as
a waxy white solid. Rf (alumina, petroleum ether/CH2Cl2 60:40):
0.38. IR ymax: 3300 (CH), 2936, 2859, 2110 (C^C), 1586, 1570, 1419,
4.17. 4-(6-Azido-hexyloxymethyl)-6,60-dimethyl-2,20-
bipyridine (3f)
To a stirred solution of bromide 3b (90 mg, 0.239 mmol) in an-
hydrous DMF (1 mL) was added under Argon sodium azide (90 mg,
1.38 mmol). The reaction mixture was stirred at rt for 14 h and
a saturated NH4Cl solution (5 mL) was added. The mixture was
extracted with CH2Cl2 and the organic layer was dried over MgSO4.
Evaporation of the solvent afforded the title compound 3f (80 mg,
0.236 mmol, yield 99%) as a yellow oil. Rf (alumina, CH2Cl2/petro-
leum ether 60:40): 0.48. IR ymax: 2936, 2860, 2095 (N3), 1586, 1570,
1117. 1H NMR (300 MHz, CDCl3)
d: 1.39–1.69 (m, 8H), 1.93 (t, 1H,
J¼2.6), 2.18 (td, 2H, J¼6.8, 2.6), 2.61 (s, 6H), 3.51 (t, 2H, J¼6.5), 4.55
(s, 2H), 7.13 (d, 1H, J¼7.5), 7.17 (s, 1H), 7.66 (t, 1H, J¼7.7), 8.09 (s, 1H),
8.16 (d, 1H, J¼7.8). 13C NMR (75 MHz, CDCl3)
d: 18.3 (CH2), 24.7
1418, 1260. 1H NMR (300 MHz, CDCl3)
d: 1.40–1.43 (m, 4H), 1.62–
(CH3), 25.7 (CH2), 28.4 (CH2), 28.5 (CH2), 29.6 (CH2), 68.2 (CH), 70.8
(CH2), 71.5 (CH2), 84.6 (Cq), 116.5 (CH), 118.3 (CH), 121.2 (CH), 123.1
(CH), 137.0 (CH), 148.8 (Cq), 155.9 (Cq), 156.0 (Cq), 157.8 (Cq), 158.2
(Cq). MS (ESIþ): m/z (%) 323.1 (100) [MþH]þ, 345.3 (23) [MþNa]þ.
1.67 (m, 4H), 2.63 (s, 6H), 3.26 (t, 2H, J¼6.9), 3.52 (t, 2H, J¼6.5), 4.56
(s, 2H), 7.15 (d, 1H, J¼7.8), 7.17 (s, 1H), 7.68 (t, 1H, J¼7.8), 8.10 (s, 1H),
8.17 (d, 1H, J¼7.8). 13C NMR (75 MHz, CDCl3)
d: 24.5 (CH3), 25.6
(CH2), 26.4 (CH2), 28.7 (CH2), 29.4 (CH2), 51.2 (CH2), 70.7 (CH2), 71.4
(CH2), 116.4 (CH), 118.2 (CH), 121.0 (CH), 123.0 (CH), 136.9 (CH),
148.7 (Cq), 155.7 (Cq), 155.9 (Cq), 157.7 (Cq), 158.0 (Cq). MS (ESIþ):
m/z (%) 340.3 (100) [MþH]þ, 362.4 (54) [MþNa]þ. Anal. Calcd for
C19H25N5O: C, 67.23; H, 7.42; N, 20.63. Found: C, 66.79; H, 7.74; N,
20.78.
0
0
0
4.19. 4-Carbomethoxy-6,6 -dimethyl-2,2 -bipyridine N1,N1 -
dioxide (16)
To a stirred solution of m-CPBA (3.26 g, 18.89 mmol) in chloro-
form (27 mL) was added a solution of bipyridine 1 (1.20 g,

F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
7683
4.96 mmol) in chloroform (18 mL). The resulting solution was
stirred at rt for 19 h, then neutralized with satd aq NaHCO3, fol-
lowed by extraction with CH2Cl2 (3ꢂ40 mL). The combined organic
layers were dried (Na2SO4) and concentrated, to yield a yellow
paste. The crude product was purified by column chromatography
over silica gel (CH2Cl2/MeOH 97:3) to give dioxide 16 (1.29 g,
4.70 mmol, yield 95%) as a pale yellow solid. Mp: 222–224 ꢁC. Rf
(silica gel, CH2Cl2/MeOH 95:5): 0.10. IR ymax: 3075, 1721 (C]O),
4.202. 4-(tert-Butyldiphenylsilanyloxymethyl)-6,60-dimethyl-
0
2,2 -bipyridine N1,N1 -dioxide (18)
To a stirred solution of bipyridine 17 (1.00 g, 2.21 mmol) in
CHCl3 (75 mL) was added dropwise a solution of m-CPBA (1.50 g,
8.69 mmol) in CHCl3 (55 mL), and the resulting solution was stirred
at rt for 24 h. The reaction mixture was washed with satd aq
NaHCO3 (2ꢂ30 mL). The organic layer was dried (Na2SO4), and the
solvent was removed under reduced pressure. The crude product
was purified by flash chromatography over silica gel (CH2Cl2/MeOH
95:5, then 80:20), to give dioxide 18 (844 mg, 1.74 mmol, yield 78%)
as a tan solid. Mp: 67 ꢁC. IR ymax: 3071, 2931, 2858, 1471, 1446, 1428,
1426,1329,1261, 998. 1H NMR (300 MHz, CDCl3)
d: 2.56 (s, 3H), 2.57
(s, 3H), 3.91 (s, 3H), 7.23 (t, 1H, J¼7.8), 7.33 (dd, 1H, J¼7.8, 2.1), 7.38
(dd, 1H, J¼7.5, 2.1), 7.94 (d, 1H, J¼2.7), 7.96 (d, 1H, J¼2.4). 13C NMR
(75 MHz, CDCl3) d: 17.77 (CH3), 17.82 (CH3), 52.6 (CH3), 124.1 (CH),
124.5 (Cq), 125.3 (CH), 125.5 (CH), 126.6 (CH), 127.0 (CH), 142.9 (Cq),
143.8 (Cq), 149.7 (Cq), 149.8 (Cq), 164.1 (Cq). MS (DCIþ, NH3): m/z
(%) 275 (73) [MþH]þ, 259 (75) [MꢀOþH]þ, 243 (100) [Mꢀ2OþH]þ.
1370, 1253, 1113. 1H NMR (300 MHz, CDCl3)
d: 1.10 (s, 9H), 2.57 (s,
6H), 4.71 (s, 2H), 7.21–7.45 (m, 11H), 7.66 (dd, 4H, J¼7.5, 1.5). 13C
NMR (75 MHz, CDCl3) d: 17.8 (CH3), 18.0 (CH3), 19.2 (Cq), 26.7 (CH3),
63.5 (CH2),122.3 (CH),123.8 (CH),124.0 (CH),125.1 (CH),126.6 (CH),
129.9 (CH), 132.6 (Cq), 135.7 (CH), 138.1 (Cq), 142.9 (Cq), 143.7 (Cq),
149.0 (Cq), 149.5 (Cq). MS (FABþ, MNBA): m/z (%) 485 (100)
[MþH]þ, 507 (7) [MþNa]þ.
4.20. 4-Carbomethoxy-6,60-bis(bromomethyl)-2,20-
bipyridine (4a)
Bipyridine dioxide 16 (758 mg, 2.76 mmol) was dissolved in
acetic anhydride (10 mL), and the resulting solution was heated to
120 ꢁC under Argon for 16 h with stirring. The reaction mixture was
evaporated to dryness, followed by addition of a 33% solution of HBr
in glacial acetic acid (19 mL). The resulting solution was heated to
70 ꢁC under Argon for 7 h. The reaction mixture was cooled to rt
and diluted with H2O (190 mL), and the resulting solution was
neutralized with satd aq NaHCO3 and extracted with CH2Cl2
(6ꢂ50 mL). The combined organic layers were dried (Na2SO4), and
the solvent was removed under reduced pressure. The crude
product was purified by flash chromatography over silica gel
(CH2Cl2) to give dibromide 4a (695 mg, 1.74 mmol, yield 63%) as
a pale yellow solid. Mp: 127–128 ꢁC. Rf (silica gel, CH2Cl2): 0.31. IR
ymax: 3077, 2949, 1730 (C]O), 1566, 1401, 1345, 1269, 1234, 1202,
4.23. 4-(tert-Butyldiphenylsilanyloxymethyl)-6,60-
diacetoxymethyl-2,20-bipyridine (19)
A solution of 450 mg (0.93 mmol) of dioxide 18 in acetic anhy-
dride (7 mL) was heated to 100 ꢁC with stirring for 24 h. The re-
action mixture was concentrated, and the residue was purified by
flash chromatography over alumina (Et2O/petroleum ether 45:55),
to give diacetate 19 (360 mg, 0.63 mmol, yield 68%) as a pale yellow
solid. Mp 126–128 ꢁC. IR ymax: 3071, 2932, 2857, 1746 (C]O), 1588,
1427, 1224, 1113. 1H NMR (300 MHz, CDCl3)
d: 1.18 (s, 9H), 2.18 (s,
3H), 2.20 (s, 3H), 4.90 (s, 2H), 5.29 (s, 2H), 5.35 (s, 2H), 7.36–7.50 (m,
8H), 7.73 (dd, 4H, J¼7.5, 1.5), 7.83 (t, 1H, J¼7.8), 8.33 (s, 1H), 8.38 (d,
1H, J¼7.8). 13C NMR (75 MHz, CDCl3)
d: 19.3 (Cq), 20.9 (CH3), 26.5
771. 1H NMR (300 MHz, CDCl3)
d: 4.01 (s, 3H), 4.65 (s, 2H), 4.67 (s,
(CH3), 26.8 (CH3), 64.3 (CH2), 66.9 (CH2), 67.0 (CH2), 117.1 (CH), 118.2
(CH), 120.3 (CH), 121.5 (CH), 127.8 (CH), 129.9 (CH), 132.9 (Cq), 135.4
(CH), 137.5 (CH), 152.1 (Cq), 155.15 (Cq), 155.2 (Cq), 155.5 (Cq), 170.6
(Cq). MS (ESIþ): m/z (%) 569.7 (100) [MþH]þ, 591.3 (77) [MþNa]þ.
Anal. Calcd for C33H36N2O5Si: C, 69.69; H, 6.38; N, 4.93. Found: C,
69.68; H, 6.56; N, 5.38.
2H), 7.51 (dd, 1H, J¼7.8, 0.9), 7.84 (t, 1H, J¼7.8), 8.01 (d, 1H, J¼1.5),
8.40 (dd, 1H, J¼7.8, 0.9), 8.89 (d, 1H, J¼1.5). 13C NMR (75 MHz,
CDCl3) d: 33.4 (CH2), 33.9 (CH2), 52.9 (CH3), 119.9 (CH), 120.6 (CH),
122.8 (CH), 124.1 (CH), 138.1 (CH), 139.6 (Cq), 154.5 (Cq), 156.5 (Cq),
156.7 (Cq), 157.3 (Cq), 165.4 (Cq). MS (ESIþ): m/z (%) 401.3 (100)
[MþH]þ, 423.0 (95) [MþNa]þ. Anal. Calcd for C14H12N2O2Br2: C,
42.03; H, 3.02; N, 7.00. Found: C, 42.10; H, 3.15; N, 6.78.
4.24. 4-(tert-Butyldiphenylsilanyloxymethyl)-6,60-
bis(hydroxymethyl)-2,20-bipyridine (20)
4.21. 4-(tert-Butyldiphenylsilanyloxymethyl)-6,60-dimethyl-
2,20-bipyridine (17)
To a stirred suspension of diacetate 19 (360 mg, 0.63 mmol) in
MeOH (6.5 mL) and H2O (0.6 mL) was added solid potassium car-
bonate (350 mg, 2.5 mmol). The resulting mixture was vigorously
stirred at rt for 3 h. The reaction mixture was diluted with CH2Cl2
(30 mL), washed sequentially with H2O (2ꢂ10 mL) and brine
(10 mL). The organic layer was dried (Na2SO4), and the solvent was
removed under reduced pressure to give diol 20 (305 mg,
0.63 mmol, yield 100%) as a glassy pale yellow oil. IR ymax: 3391,
3070, 2931, 2857, 1611, 1588, 1568, 1471, 1427, 1157, 1112. 1H NMR
To a stirred solution of alcohol 2b (1.40 g, 6.53 mmol) in anhy-
drous DMF (65 mL) was added, at rt under Argon, imidazole (1.34 g,
19.7 mmol), followed by the dropwise addition of tert-butyldi-
phenylchlorosilane (2.7 mL, 10.5 mmol). The resulting solution was
stirred at rt for 48 h. The reaction mixture was quenched with satd
aq NaHCO3 (60 mL), diluted with EtOAc (400 mL), sequentially
washed with H2O (2ꢂ200 mL) and brine (200 mL). The organic
layer was dried (Na2SO4), and the solvent was removed under re-
duced pressure. The crude product was purified by flash chroma-
tography over alumina (Et2O/petroleum ether 10:90), to give silyl
ether 17 (2.77 g, 6.12 mmol, yield 94%) as a white solid. Mp: 89–
91 ꢁC. IR ymax: 3070, 2958, 2930, 2857, 1585, 1570, 1427, 1418, 1113.
(300 MHz, CDCl3) d: 1.18 (s, 9H), 3.75–4.15 (s, br, 2H), 4.83 (s, 2H),
4.85 (s, 2H), 4.89 (s, 2H), 7.26 (d, 1H, J¼7.5), 7.31 (s, 1H), 7.36–7.46
(m, 6H), 7.72 (dd, 4H, J¼7.5, 1.5), 7.81 (t, 1H, J¼7.8), 8.31 (s, 1H), 8.33
(d,1H, J¼7.8). 13C NMR (75 MHz, CDCl3)
d: 19.3 (Cq), 26.8 (CH3), 63.9
(CH2), 64.3 (CH2), 116.7 (CH), 117.5 (CH),119.7 (CH), 120.6 (CH),127.8
(CH), 129.9 (CH), 132.8 (Cq), 135.4 (CH), 137.6 (CH), 152.4 (Cq), 153.9
(Cq), 154.0 (Cq), 158.3 (Cq), 158.4 (Cq). MS (ESIþ): m/z (%) 485.3
(100) [MþH]þ, 507.3 (12) [MþNa]þ.
1H NMR (300 MHz, CDCl3)
d: 1.18 (s, 9H), 2.63 (s, 3H), 2.67 (s, 3H),
4.86 (s, 2H), 7.15 (d,1H, J¼7.8), 7.28 (s,1H), 7.38–7.46 (m, 6H), 7.69 (t,
1H, J¼7.8), 7.72–7.75 (m, 4H), 8.15 (s, 1H), 8.21 (d, 1H, J¼7.8). 13C
NMR (75 MHz, CDCl3) d: 19.3 (Cq), 24.6 (CH3), 24.8 (CH3), 26.8
(CH3), 64.4 (CH2),115.3 (CH),118.1 (CH),119.9 (CH), 122.9 (CH), 127.8
(CH), 129.8 (CH), 133.1 (Cq), 135.5 (CH), 136.9 (CH), 151.1 (Cq), 155.7
(Cq), 155.9 (Cq), 157.7 (Cq), 158.0 (Cq). MS (FABþ, MNBA): m/z (%)
453 (100) [MþH]þ. Anal. Calcd for C29H32N2OSi: C, 76.95; H, 7.13; N,
6.19. Found: C, 76.76; H, 7.21; N, 6.01.
4.25. 4-(tert-Butyldiphenylsilanyloxymethyl)-6,60-
bis(chloromethyl)-2,20-bipyridine (4b)
Diol 20 (160 mg, 0.33 mmol) was dissolved in thionyl chloride
(3 mL) and stirred at rt for 1.5 h. The solution was evaporated in

7684
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
vacuo and the residue was dissolved in CH2Cl2 and washed with
a satd aq NaHCO3 solution. The organic layer was dried (MgSO4),
evaporated in vacuo and the crude product was purified by column
chromatography over alumina (petroleum ether/CH2Cl2 70:30) to
give dichloro derivative 4b (110 mg, 0.21 mmol, yield 64%) as
a colorless oil. Rf (alumina, petroleum ether/CH2Cl2 70:30): 0.69. IR
ymax: 3062, 2954, 2930, 2857, 1606, 1587, 1566, 1462, 1427, 1413,
5b$Eu3þ and the labeling procedure are described in the
following.
4.28.1. Synthesis of 5b$Eu3þ
A solution of 5b (2.95 mg, 6.0 mmol) with EuCl3$6H2O (1.1 equiv)
in 2 mL of H2O at pH 3 was stirred at room temperature for 24 h. The
excess of Eu3þ was precipitated by adjusting the pH to 8 using NaOH
solution. The solvent was removed, the resulting solid was dissolved
in a minimum of methanol, and diethylether was added to pre-
cipitate the compound 5b$Eu3þ. MS (ESIꢀ, H2O): m/z (%) 637.37 (94)
[MꢀH]ꢀ, 639.37 (100) [MꢀH]ꢀ, 661.38 (18) [MþNaꢀ2H]ꢀ, 661.38
(20) [MþNaꢀ2H]ꢀ. Luminescence (H2O, lexc¼317 nm): lem (relative
intensity, corrected spectrum), 580 (2.0), 592 (31.6), 618 (100), 651
1262. 1H NMR (300 MHz, CDCl3)
d: 1.16 (s, 9H), 4.72 (s, 2H), 4.76 (s,
2H), 4.88 (s, 2H), 7.36–7.46 (m, 6H), 7.50 (dd, 1H, J¼7.7, 1.0), 7.58 (d,
1H, J¼1.0), 7.69–7.72 (m, 4H), 7.84 (t, 1H, J¼7.8), 8.36 (d, 1H, J¼1.0),
8.39 (dd, 1H, J¼7.8, 1.0). 13C NMR (75 MHz, CDCl3)
d: 19.4 (Cq), 26.8
(CH3), 46.8 (CH2), 64.3 (CH2), 117.6 (CH), 119.9 (CH), 120.6 (CH),
122.8 (CH), 127.9 (CH), 129.9 (CH), 132.9 (Cq), 135.5 (CH), 137.9 (CH),
152.7 (Cq), 155.1 (Cq), 155.2 (Cq), 156.0 (Cq), 156.1 (Cq). MS (ESIþ):
m/z (%) 521.1 (9) [MþH]þ, 523.1 (7) [MþH]þ, 543.1 (100) [MþNa]þ,
545.1 (67) [MþNa]þ.
(5.6), 697 (74.2) nm.
s
(H2O)¼0.59 ms.
4.28.2. Labeling of BSA with NHS-5b$Eu3þ
5b$Eu3þ (1.38 mg, 2.2
mol) was dissolved in 100
drous DMF. Stocks solutions of EDCI$HCl (44 L, 2.2 mol) and N-
hydroxysuccinimide (11.4 L, 2.2 mol) in anhydrous DMF were
added, and the solution was stirred at rt for 24 h. Bovine serum
albumin (4.78 mg, 72 nmol) dissolved in 480 L of phosphate
m
mL of anhy-
4.26. Tetra (tert-butyl) 2,20,200,2000-{[4-(methoxycarbonyl)-
2,20-bipyridine-6,60-diyl]bis(methylenenitrilo)}-tetrakis
(acetate) (5a)
m
m
m
m
m
To a stirred solution of dibromide 4a (800 mg, 2.0 mmol) in
anhydrous acetonitrile (160 mL) was added Na2CO3 (2.12 g,
20.0 mmol) and di-tert-butyl iminodiacetate (1.03 g, 4.2 mmol).
The resulting mixture was stirred at reflux for 17 h, then filtered,
and the filtrate was concentrated in vacuo. The residue was taken
up in CH2Cl2 (100 mL) and washed with H2O (75 mL). The aqueous
layer was extracted with CH2Cl2 (50 mL), the combined organic
layers were dried (MgSO4) and the solvent was removed under
reduced pressure. The crude product was purified by column
chromatography on silicagel (CH2Cl2/MeOH 100:0, then 98:02) to
give compound 5a (1.33 g, 1.82 mmol, yield 91%) as a pale yellow
oil. Rf (Silica gel, CH2Cl2/MeOH 95:05): 0.29. IR ymax: 2978, 2932,
1734 (C]O), 1567, 1368, 1256, 1220, 1144, 991, 773. 1H NMR
buffer (50 mM, pH¼8) was added to achieve a 30:1 label/protein
reaction ratio, and the mixture was allowed to stir for 48 h at
room temperature. The reaction mixture was concentrated to
a low volume using a Microcon YM-10 centrifugal filter (Amicon,
Millipore, Bedford, MA), and then 400 mL of H2O was added and
the solution was concentrated again. This operation was repeated
once again to remove totally the unbound label. The extent of
conjugation was determined by MALDI-TOF-MS. The matrix was
prepared from a solution of sinapinic acid in acetonitrile. The
sample and the matrix solution were mixed in equal amounts
(1 mL) and added on the stainless steel probe. 1 mL of aqueous 0.1%
TFA was added to each spot and the sample was allowed to dry at
room temperature. The data were acquired in the linear mode.
The comparison of the molecular mass of the Europium labeled
BSA (67,418.2 Da) with that of the starting protein (66,401.1 Da)
allowed establishing that the mean number of europium com-
plexes labels per BSA molecule is 1.6. 5b$Eu3þ-BSA: Luminescence
(H2O, lexc¼317 nm): lem (relative intensity, corrected spectrum),
580 (6.0), 592 (29.8), 618 (100), 651 (5.5), 697 (65.9) nm.
(300 MHz, CDCl3) d: 1.46 (s, 36H), 3.51 (s, 4H), 3.54 (s, 4H), 3.96 (s,
3H), 4.13 (s, 2H), 4.18 (s, 2H), 7.69 (dd, 1H, J¼7.7, 1.1), 7.78 (t, 1H,
J¼7.7), 8.11 (d, 1H, J¼1.4), 8.31 (dd, 1H, J¼7.7, 1.1), 8.81 (d, 1H, J¼1.4).
13C NMR (75 MHz, CDCl3)
d: 28.1 (CH3), 52.4 (CH3), 55.7 (CH2), 55.8
(CH2), 59.4 (CH2), 59.9 (CH2), 80.97 (Cq), 81.02 (Cq),118.8 (CH),119.5
(CH), 122.0 (CH), 123.4 (CH), 137.3 (CH), 139.0 (Cq), 154.5 (Cq), 156.6
(Cq), 159.0 (Cq), 159.8 (Cq), 166.0 (Cq), 170.4 (Cq), 170.5 (Cq). MS
(FABþ/MNBA): m/z (%) 729 (100) [MþH]þ, 751 (30) [MþNa]þ. Anal.
Calcd for C38H56N4O10: C, 62.62; H, 7.74; N, 7.69. Found: C, 62.47; H,
7.65; N, 7.47.
s(H2O)¼0.67 ms.
4.29. Tetra (tert-butyl) 2,20,200,2000-{[4-(hydroxymethyl)-
2,20-bipyridine-6,60-diyl]bis(methylenenitrilo)}-tetrakis
(acetate) (5c)
4.27. 2,20,200,2000-[(4-Carboxy-2,20-bipyridine-6,60-
diyl)bis(methylenenitrilo)]-tetrakis (acetic acid) (5b)
To a solution of ester 5a (729 mg, 1.0 mmol) in anhydrous
methanol (50 mL) was added NaBH4 as a solid (1.13 g, 30.0 mmol)
in five portions (2 h) with stirring at rt under Argon. The resulting
mixture was further stirred for 2 h at rt and the solvent removed
under reduced pressure. A solution of satd aq NH4Cl (15 mL) was
added, and the resulting mixture was partitioned between CH2Cl2
(50 mL) and H2O (50 mL). The aqueous layer was extracted with
CH2Cl2 (30 mL), the combined organic layers were dried (MgSO4)
and the solvent was removed under reduced pressure. The crude
product was purified by column chromatography over alumina
(CH2Cl2/MeOH 99:01) to give alcohol 5c (652 mg, 0.93 mmol, yield
93%) as a colourless oil. Rf (Alumina, CH2Cl2/MeOH 99:01): 0.25. IR
ymax: 3357, 2980, 2933, 1734, 1639, 1588, 1457, 1394, 1369, 1249,
The ester 5a (77 mg, 0.106 mmol) in a 6 N HCL solution
(5 mL) was stirred at 90 ꢁC for 3 days. The solvent was removed
under reduced pressure to give compound 5b in the form of its
hydrochloride salt as a white solid (64 mg, 0.106 mmol, yield
100%). IR ymax: 3385, 3185, 3009, 1723 (C]O), 1591, 1572, 1510.
1H NMR (300 MHz, DMSO-d6)
d: 3.67 (s, 8H), 4.20 (s, 2H), 4.23
(s, 2H), 7.66 (d, 1H, J¼7.7), 7.99 (t, 1H, J¼7.7), 8.06 (d, 1H, J¼1.3),
8.31 (d, 1H, J¼7.7), 8.69 (d, 1H, J¼1.3). 13C NMR (75 MHz, DMSO-
d6) d: 54.2 (CH2), 54.3 (CH2), 58.9 (CH2), 59.2 (CH2), 118.3 (CH),
119.6 (CH), 122.6 (CH), 124.1 (CH), 138.2 (CH), 140.1 (Cq), 153.7
(Cq), 155.4 (Cq), 157.3 (Cq), 158.8 (Cq), 166.2 (Cq), 171.3 (Cq),
171.5 (Cq). MS (ESIþ): m/z (%) 491.3 (100) [MþH]þ, 592.2 (95)
[MþK]þ.
1155. 1H NMR (300 MHz, CDCl3)
d: 1.49 (s, 36H), 3.54 (s, 4H), 3.55
(s, 4H), 4.14 (s, 4H), 4.73 (s, 2H), 7.65 (s, 1H), 7.66 (d, 1H, J¼7.7),
4.28. Labeling of BSA with 5b$Eu3D
7.79 (t, 1H, J¼7.7), 8.33 (d, 1H, J¼6.7), 8.34 (s, 1H). 13C NMR
(75 MHz, CDCl3) d: 28.1 (CH3), 55.7 (CH2), 59.6 (CH2), 59.8 (CH2),
BSA was labeled with 5b$Eu3þ by using succinimidyl
63.9 (CH2), 81.0 (Cq), 117.0 (CH), 119.6 (CH), 120.4 (CH), 123.0 (CH),
137.3 (CH), 151.6 (Cq), 155.3 (Cq), 155.6 (Cq), 158.5 (Cq), 158.7 (Cq),
) . The preparation of
monoester (NHS-5b$Eu3þ of 5b$Eu3þ

F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
7685
170.5 (Cq). MS (CI/NH3): m/z (%) 701.6 (55) [MþH]þ, 718.6 (100)
[MþNH4]þ.
cooled to 4 ꢁC, and the resulting precipitate was isolated after
centrifugation.
Complex 6a$EuCl3: MS (FABþ, MNBA): m/z (%) 855 (50) [MꢀCl]þ,
819 (100) [MꢀClꢀHCl]þ. Luminescence (H2O lexc¼306 nm): lem
(relative intensity, corrected spectrum), 581 (3.3), 592 (42.0), 617
(81.8), 651 (6.8), 702 (100) nm.
4.30. Tetra (tert-butyl) 2,20,200,2000-{[4-(prop-2-ynyloxy-
methyl)-2,20-bipyridine-6,60-diyl]bis(methyl-
enenitrilo)}-tetrakis (acetate) (5d)
Complex 6a$GdCl3: MS (FABþ, MNBA): m/z (%) 860 (53)
[MꢀCl]þ, 824 (100) [MꢀClꢀHCl]þ.
To a stirred solution of alcohol 5c (100 mg, 0.14 mmol) in an-
hydrous DMF (2 mL) was added at rt under Argon sodium hydride
(6.9 mg, 0.28 mmol) in one portion, followed by the addition of
Acknowledgements
a solution of propargyl bromide in toluene (80 wt%, 50 mL,
This work was supported by the CNRS, the Ministry of Research
0.45 mmol). The resulting mixture was stirred at rt for 48 h, then
a solution of satd aq NaCl (5 mL) was added. The mixture was
extracted with CH2Cl2 (3ꢂ5 mL), the combined organic layers were
dried (MgSO4) and the solvent was removed under reduced pres-
sure. The crude product was purified by column chromatography
over alumina (CH2Cl2/MeOH 98:02) to give acetylenic compound
5d (42 mg, 0.057 mmol, yield 41%) as a colourless oil. Rf (Alumina,
CH2Cl2/MeOH 98:02): 0.29. IR ymax: 3270, 2978, 2931, 2116, 1733,
1609, 1585, 1566, 1456, 1417, 1393, 1368, 1254, 1221, 1147. 1H NMR
´
´ ´
of France and the Region Midi-Pyrenees. F.H. thanks the CNRS for
a postdoctoral fellowship.
References and notes
1. Reedijk, J. In Comprehensive Coordination Chemistry; Wilkinson, G., Sir, Gillard,
R. D., McCleverty, J. A., Eds.; Pergamon: Oxford, NY, 1987; Vol. 2, pp 73–98.
2. Chelucci, G.; Thummel, R. P. Chem. Rev. 2002, 102, 3129–3170.
3. Balzani, V.; Juris, A.; Venturi, M.; Campagna, S.; Serroni, S. Chem. Rev. 1996, 96,
759–833.
4. Sun, L.; Hammarstro¨m, L.; Åkermark, B.; Styring, S. Chem. Soc. Rev. 2001, 30,
36–49.
5. Keefe, M. H.; Benkstein, K. D.; Hupp, J. T. Coord. Chem. Rev. 2000, 205, 201–228.
6. Maury, O.; Le Bozec, H. Acc. Chem. Res. 2005, 38, 691–704.
7. Carlson, B.; Phelan, G. D.; Kaminsky, W.; Dalton, L.; Jiang, X.; Liu, S.; Jen, A. K.-Y.
J. Am. Chem. Soc. 2002, 124, 14162–14172.
8. Schubert, U. S.; Eschbaumer, C. Angew. Chem., Int. Ed. 2002, 41, 2892–2926.
9. Hemmila¨, I.; Laitala, V. J. Fluoresc. 2005, 15, 529–542.
10. Faulkner, S.; Pope, S. J. A.; Burton-Pye, B. P. Appl. Spectrosc. Rev. 2005, 40, 1–31.
11. Bu¨ nzli, J.-C. G. In Metal Ions in Biological Systems; Sigel, A., Sigel, H., Eds.; Marcel
Dekker: New York, NY, 2004; Vol. 42, pp 39–75.
12. Brunet, E.; Juanes, O.; Rodriguez-Ubis, J.-C. Curr. Chem. Biol. 2007, 1, 11–39.
13. Nasso, I.; Galaup, C.; Havas, F.; Tisne`s, P.; Picard, C.; Laurent, S.; Vander Elst, L.;
Muller, R. N. Inorg. Chem. 2005, 44, 8293–8305.
(300 MHz, CDCl3)
d
: 1.49 (s, 36H), 2.51 (t, 1H, J¼2.4), 3.55 (s, 4H),
3.56 (s, 4H), 4.15 (s, 4H), 4.29 (d, 2H, J¼2.4), 4.72 (s, 2H), 7.61 (s, 1H),
7.67 (d, 1H, J¼7.7), 7.80 (t, 1H, J¼7.7), 8.30 (s, 1H), 8.32 (d, 1H, J¼7.7).
13C NMR (75 MHz, CDCl3)
d: 28.2 (CH3), 55.74 (CH2), 55.76 (CH2),
57.9 (CH2), 59.6 (CH2), 59.9 (CH2), 70.3 (CH2), 75.1 (Cq), 79.3 (CH),
81.0 (Cq), 117.9 (CH), 119.6 (CH), 121.4 (CH), 123.1 (CH), 137.4 (CH),
148.0 (Cq), 155.3 (Cq), 155.8 (Cq), 158.8 (Cq), 158.9 (Cq), 170.61(Cq),
170.65 (Cq). MS (ESIþ): m/z (%) 739.7 (94) [MþH]þ, 761.6 (100)
[MþNa]þ.
4.31. NaBr complex of 6,60-{N,N0N,N0-[Bis (2,20-bipyridine-6,60-
dimethyl)]bis (aminomethyl)}-2,20-bipyridine-4-carboxylic
14. Faulkner, S.; Beeby, A.; Carrie´, M.-C.; Dadabhoy, A.; Kenwright, A. M.; Sammes,
P. G. Inorg. Chem. Commun. 2001, 4, 187–190.
acid methyl ester (6a$NaD
)
15. Sabbatini, N.; Guardigli, M.; Lehn, J.-M. Coord. Chem. Rev. 1993, 123, 201–228.
16. Alpha, B.; Ballardini, R.; Balzani, V.; Lehn, J.-M.; Perathoner, S.; Sabbatini, N.
Photochem. Photobiol. 1990, 52, 299–306.
17. Bazin, H.; Trinquet, E.; Mathis, G. Rev. Mol. Biotechnol. 2002, 82, 233–250.
18. Arano, Y.; Uezono, T.; Akizawa, H.; Ono, M.; Wakisaka, K.; Nakayama, M.;
Sakahara, H.; Konishi, J.; Yokoyama, A. J. Med. Chem. 1996, 39, 3451–3460.
19. Maisano, F.; Gozzini, L.; De Haen, C. Bioconjugate Chem. 1992, 3, 212–217.
20. Reilly, R.; Lee, N.; Houle, S.; Law, J.; Marks, A. Appl. Radiat. Isot. 1992, 43,
961–967.
21. Mukkala, V. M.; Sund, C.; Kwiatkowski, M. U.S. Patent 5,216,134, 1993; Chem.
Abstr. 1993, 119, 244992.
22. Newkome, G. R.; Patri, A. K.; Holder, E.; Schubert, U. S. Eur. J. Org. Chem. 2004,
235–254.
23. Nasso, I.; Bedel, S.; Galaup, C.; Picard, C. Eur. J. Inorg. Chem. 2008, 2064–2074
and references cited therein.
24. Mathieu, J.; Marsura, A. Synth. Commun. 2003, 33, 409–414.
25. Bedel, S.; Ulrich, G.; Picard, C.; Tisne`s, P. Synthesis 2002, 1564–1570.
26. Havas, F.; Danel, M.; Galaup, C.; Tisnes, P.; Picard, C. Tetrahedron Lett. 2007, 48,
999–1002.
27. Regnouf de Vains, J.-B.; Papet, A. L.; Marsura, A. J. Heterocycl. Chem. 1994, 31,
1069–1077.
28. Kro¨hnke, F. Synthesis 1976, 1–24.
To a stirred suspension of macrocyclic diamine 23 (120 mg,
0.3 mmol) and Na2CO3 (318 mg, 3 mmol) in anhydrous CH3CN
(150 mL), at reflux under Argon, was added, dropwise over 20 min,
a solution of dibromide 4a (120 mg, 0.3 mmol) in anhydrous CH3CN
(50 mL). The resulting mixture was stirred at reflux for 48 h. The
reaction mixture was filtered and concentrated in vacuo, to yield
a pale yellow solid. The crude product was purified by flash chro-
matography over silica gel (CH2Cl2/MeOH 93:7), to give sodium
cryptate 6a$Naþ (109 mg, 0.14 mmol, yield 47%) as a yellow solid.
Mp>250 ꢁC (dec). IR ymax: 3421, 2921, 1728 (C]O), 1590, 1577, 1433,
1267, 1171, 996. 1H NMR (300 MHz, CDCl3):
(br s, 2H), 3.95 (s, 3H), 7.27–7.36 (m, 4H), 7.74–7.94 (m, 12H), 8.35 (s,
1H). 13C NMR (75 MHz, CDCl3)
: 52.9 (CH3), 59.4 (CH2), 119.3 (CH),
d 3.81 (br s, 10H), 3.91
d
120.2 (CH), 120.3 (CH), 120.4 (CH), 122.8 (CH), 124.0 (CH), 124.5
(CH), 138.10 (CH), 138.15 (CH), 138.2 (CH), 139.2 (Cq), 154.3 (Cq),
155.05 (Cq), 155.15 (Cq), 156.4 (Cq), 158.1 (Cq), 158.3 (Cq), 158.7
(Cq), 159.8 (Cq), 164.9 (Cq). MS (FABþ, MNBA): m/z (%) 655 (100)
[MþNa]þ, 671 (9) [MþK]þ. HRMS-FABþ calcd for [MþNa]þ
(C38H32N8O2Na) 655.25459, found 655.25382. Anal. Calcd for
C38H32N8O2NaBr$2H2O: C, 59.15; H, 4.70; N, 14.52. Found: C, 58.82;
H, 4.89; N, 14.28.
29. Panetta, C. A.; Kumpaty, H. J.; Heimer, N. E.; Leavy, M. C.; Hussey, C. L. J. Org.
Chem. 1999, 64, 1015–1021.
30. Heller, M.; Schubert, U. S. J. Org. Chem. 2002, 67, 8269–8272.
31. Fang, Y.-Q.; Hanan, G. S. Synlett 2003, 852–854.
32. Hapke, M.; Brandt, L.; Lutzen, A. Chem. Soc. Rev. 2008, 37, 2782–2797.
33. Adams, R.; Miyano, S. J. Am. Chem. Soc. 1954, 76, 3168–3171.
34. Schubert, U. S.; Eschbaumer, C.; Heller, M. Org. Lett. 2000, 2, 3373–3376.
35. Marvel, C. S.; Dreger, E. E. In Organic Syntheses; Blatt, A. H., Ed.; Wiley: New
York, NY, 1941; Collect. Vol. 1, pp 238–240.
¨
36. Libermann, D.; Rist, N.; Grumbach, F.; Cals, S.; Moyeux, M.; Rouaix, A. Bull. Soc.
Chim. Fr. 1958, 687–694.
37. Cuiban, F.; Cilianu-Bibian, S.; Zaharescu, L. Fr Patent 1,362,967, 1964.
38. Hodgson, P. B.; Salingue, F. H. Tetrahedron Lett. 2004, 45, 685–687.
39. Jones, N. A.; Antoon, J. W.; Bowie, A. L., Jr.; Borak, J. B.; Stevens, E. P. J. Heterocycl.
Chem. 2007, 44, 363–367.
4.32. General procedure for the preparation of lanthanide
complexes of Cryptand 6a
To
a stirred solution of the lanthanide salt LnCl3$6H2O
(1.1 equiv) in methanol (20 ml) was added sodium cryptate 6a$Naþ
(46 mg, 0.06 mmol, 1 equiv) in chloroform (10 mL). After 24 h of
reflux, the solvents were evaporated, and the residue dissolved in
the minimum of methanol. Anhydre diethyl ether was added
carefully until the apparition of a slight trouble. The mixture was
40. Pierrat, P.; Gros, P.; Fort, Y. Org. Lett. 2005, 7, 697–700.
`
41. Hanan, G. S.; Schubert, U. S.; Volkmer, D.; Riviere, E.; Lehn, J.-M.; Kyritsakas, N.;
Fischer, J. Can. J. Chem. 1997, 75, 169–182.
42. Fallahpour, R. A. Synthesis 2000, 1138–1142.
43. Amat, M.; Hadida, S.; Pshenichnyi, G.; Bosch, J. J. Org. Chem. 1997, 62,
3158–3175.

7686
F. Havas et al. / Tetrahedron 65 (2009) 7673–7686
44. Fu¨rstner, A.; Leitner, A.; Me´ndez, M.; Krause, H. J. Am. Chem. Soc. 2002, 124,
13856–13863.
45. Lehn, J.-M.; Mathis, G.; Alpha, B.; Deschenaux, R.; Jolu, E. Eur. Patent 321,353,
1989; Chem. Abstr. 1990, 112, 179041.
54. Horrocks, W. D. W., Jr.; Sudnick, D. R. Acc. Chem. Res. 1981, 14, 384–392.
55. Supkowski, R. M.; Horrocks, W. D. W., Jr. Inorg. Chim. Acta 2002, 340, 44–48.
56. Longo, R.; Gonçalves e Silva, F. R.; Malta, O. L. Chem. Phys. Lett. 2000, 328,
67–74.
46. Brinkley, M. Bioconjugate Chem. 1992, 3, 2–13.
47. Mehta, N. B.; Phillips, A. P.; Lui, F. F.; Brooks, R. E. J. Org. Chem. 1960, 25,
1012–1015.
48. Walker, M. A. Tetrahedron Lett. 1994, 35, 665–668.
49. Kolb, H. C.; Sharpless, K. B. Drug Discovery Today 2003, 8, 1128–1137.
50. Abrams, S. R. Can. J. Chem. 1984, 62, 1333–1334.
51. Uenishi, J.; Tanaka, T.; Nishiwaki, K.; Wakabayashi, S.; Oae, S.; Tsukube, H. J. Org.
Chem. 1993, 58, 4382–4388.
57. Mathis, G.; Dumont, C.; Jolu, E. J. -P. WO Patent 92:01224, 1990; Chem. Abstr.
1992, 116, 147509.
58. Stein, G.; Wu¨rzberg, E. J. Chem. Phys. 1975, 62, 208–213.
59. Guillaumont, D.; Bazin, H.; Benech, J.-M.; Boyer, M.; Mathis, G. ChemPhysChem
2007, 8, 480–488.
60. Haas, Y.; Stein, G. J. Phys. Chem. 1971, 75, 3668–3677.
61. Nakamaru, K. Bull. Chem. Soc. Jpn. 1982, 55, 2697–2705.
62. Galaup, C.; Carrie´, M.-C.; Tisne`s, P.; Picard, C. Eur. J. Org. Chem. 2001, 2165–2175.
63. Henecka, H. Chem. Ber. 1949, 82, 36–41.
64. Patwardhan, A. P.; Thompson, D. H. Langmuir 2000, 16, 10340–10350.
65. Bauer, E. B.; Hampel, F.; Gladysz, J. A. Adv. Synth. Catal. 2004, 346, 812–822.
52. Liu, P.; Chen, Y.; Deng, J.; Tu, Y. Synthesis 2001, 2078–2080.
53. Rodriguez-Ubis, J.-C.; Alpha, B.; Plancherel, D.; Lehn, J.-M. Helv. Chim. Acta 1984,
67, 2264–2269.