Thionated coumarins and quinolones in the light triggered release of a model amino acid: Synthesis and photolysis studies

Article abstract of DOI:10.1016/j.tet.2012.07.021

Model amino acid ester conjugates bearing heterocycles with a thiocarbonyl group (thiocoumarins and thioquinolones) were prepared by a thionation reaction of the corresponding carbonyl precursors, with the aim of obtaining conjugates with higher photosensitivity at longer wavelengths. Photolysis studies were carried out under irradiation at different wavelengths in a photochemical reactor (250, 300, 350 and 419 nm), and it was found that the ester bond between the heterocycle and the model amino acid in the thionated analogues cleaved readily with shorter irradiation times than those of the corresponding precursors. Moreover, fast photolysis at 419 nm for thioquinolone ester conjugates (within minutes) was particularly interesting and the fact that some of the reported heterocycles displayed very different irradiation times, may provide the possibility for selective removal of one group in the presence of another photocleavable group. The (quinolin-2-thione-4-yl) methyl group may be considered as a new and very effective addition to the family of photocleavable protecting groups for carboxylic acids.

Full text of DOI:10.1016/j.tet.2012.07.021

Tetrahedron 68 (2012) 7892e7900
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Thionated coumarins and quinolones in the light triggered release of a model
amino acid: synthesis and photolysis studies
*
Andrea S.C. Fonseca, Ana M.S. Soares, M. Sameiro T. Gonc¸ alves, Susana P.G. Costa
Centre of Chemistry, University of Minho, Campus of Gualtar, 4710-057 Braga, Portugal
a r t i c l e i n f o
a b s t r a c t
Article history:
Model amino acid ester conjugates bearing heterocycles with a thiocarbonyl group (thiocoumarins and
thioquinolones) were prepared by a thionation reaction of the corresponding carbonyl precursors, with
the aim of obtaining conjugates with higher photosensitivity at longer wavelengths. Photolysis studies
were carried out under irradiation at different wavelengths in a photochemical reactor (250, 300, 350
and 419 nm), and it was found that the ester bond between the heterocycle and the model amino acid in
the thionated analogues cleaved readily with shorter irradiation times than those of the corresponding
precursors. Moreover, fast photolysis at 419 nm for thioquinolone ester conjugates (within minutes) was
particularly interesting and the fact that some of the reported heterocycles displayed very different ir-
radiation times, may provide the possibility for selective removal of one group in the presence of another
photocleavable group. The (quinolin-2-thione-4-yl) methyl group may be considered as a new and very
effective addition to the family of photocleavable protecting groups for carboxylic acids.
Ó 2012 Elsevier Ltd. All rights reserved.
Received 19 January 2012
Received in revised form 8 July 2012
Accepted 9 July 2012
Available online 15 July 2012
Keywords:
Thionation
Coumarins
Quinolones
Photocleavable protecting groups
Amino acid conjugates
Photolysis
1. Introduction
functional groups (such as alcohols, amines, phosphates, aldehydes,
ketones and carboxylic acids),4 whereas quinolone derivatives were
The use of light of appropriate wavelength can give access to
synthetic and biochemically relevant molecules from their corre-
sponding inactive conjugate precursors, provided that they possess
suitable light-sensitive moieties covalently bonded to the func-
tional groups of the molecule of interest. In recent years, the need
for new light-sensitive protecting groups has grown considering
their application in organic synthesis, as an alternative to classical
acid- and base-labile protecting groups, as well as in biochemical
and biophysical studies (for example, in molecular caging and the
mapping of the binding sites of peptide-binding receptors). The aim
of the research in this field is the design of more efficient groups
that allow orthogonal deprotection/cleavage for application in
biomolecules at wavelengths not detrimental to biological systems,
providing spatial and temporal resolution, and the development of
photoresponsive drugs.1e3 The use of longer wavelengths can
minimise side reactions and/or cell damage. Nevertheless, among
the considerable number of light-sensitive groups that have been
reported, only few cleave in a practical time by irradiation at
wavelengths superior to 400 nm.
recently reported by us for the first time as carboxyl protecting
groups cleavable by light.5 As for quinolines, they were first
reported some years ago as photolabile protecting groups for
amines and alcohols6 and more recently as protecting groups with
sensitivity to multiphoton excitation.7
Our current research interests include the design of new oxy-
gen and nitrogen heterocycles and their application as photo-
cleavable protecting groups.8 Now we present the synthesis of
thionated analogues of coumarin and quinolone in an attempt to
improve our previously reported promising findings, which
revealed that quinolone conjugates displayed shorter irradiation
times than the corresponding coumarin conjugates, at various
wavelengths of irradiation.5 It was expected that the replacement
of the heterocycle’s carbonyl by a thiocarbonyl group would result
in a bathochromic shift of the molecule’s absorption wavelength
and consequently on the photoresponsive properties of the com-
pounds at higher wavelengths, more appropriate for
bioapplications.
The thiocarbonyl group has well documented photochemistry
that includes hydrogen abstraction from primary alcohol, photo-
Coumarin derivatives are well established photolabile protect-
ing groups that have been used in the protection of various
reduction, (2
p
þ2
p) photocycloaddition and CeH bond insertion
processes.9 Applications of thiocarbonyl compounds have been
reported such as amino acid derivatives containing side-chain
thioamides for the synthesis of photoactivable peptides, and thio-
carbonyls used to photo-link peptides to modified RNA molecules.10
* Corresponding author. Fax: þ351 253 604382; e-mail address: spc@quimi-
ca.uminho.pt (S.P.G. Costa).
0040-4020/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2012.07.021

A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
7893
2. Results and discussion
methoxy group at position-6 was coupled in the same conditions to
N-benzyloxycarbonylalanine and glutamic acid, in order to obtain
a set of three quinolone conjugates 3feh with representative amino
acids (nonpolar aromaticdphenylalanine; nonpolardalanine;
acidicdglutamic acid) (Scheme 2, Table 1).
The IR spectra of conjugates 3b,c,geh showed bands due to
stretching vibrations of the ester carbonyl group of the hetero-
cycleeamino acid link between 1729 and 1740 cmꢀ1, respectively.
1H NMR spectra showed signals of the heterocycle aromatic pro-
2.1. Synthesis
4-Chloromethyl-7-methoxy-1-methylquinolone 1c was pre-
pared by
a
modified Knorr synthesis11 between ethyl 3-
oxobutanoate and 3-methoxy-N-methylaniline, by a multistep
synthesis involving oxidation of the methyl group at position-4 to
the aldehyde, by reaction with selenium dioxide, followed by re-
duction with sodium borohydride, and conversion to the chlor-
omethylated derivative by treatment with thionyl chloride.5
The synthesis of thionated coumarin derivatives 2 was accom-
plished by reaction of the corresponding heterocycle 1 with Law-
esson’s reagent (LR), by refluxing in dry toluene.12 The desired
thiocoumarins 2a,b were obtained in fair yields, after purification
by silica gel column chromatography. The same transformation was
attempted for quinolone derivative 1c but the desired product 2c
could not be isolated, with a thiocarbonylated derivative that suf-
fered loss of the chlorine being detected that prevented further
application (Scheme 1, Table 1). In the 13C NMR spectra of thio-
coumarins 2a,b, signals of the thiocarbonyl group were found at
tons, for example, for 3b and 3c H-3 as a singlet ( 6.18 and
d
6.59 ppm, respectively), the phenylalanine -CH (d 4.72 to
a
4.79 ppm), as well as the characteristic protons of the heterocycle
methylene group (at about
d 5.22 ppm). The confirmation of the
presence of the newly formed ester bond was also supported by 13C
NMR spectra signals of the carbonyl group, which were found at
d
171.03 and 171.13 ppm, respectively. The synthesis and charac-
terization data of conjugates 3a,def had been previously reported
by us and are presented in Table 1 for easier comparison.5
For thiocoumarins 2a,b the same coupling reaction was envis-
aged to yield the corresponding ester conjugates 4 but difficulties
were encountered. In these condition reactions, the characteriza-
tion data of the isolated product suggested that the chloromethyl
group suffered intramolecular nucleophilic attack by the sulfur
d
197.11 and 197.05 ppm, respectively, with a shift of about 25 ppm
from the carbonyl precursors.
Cl
Cl
2a-b
X
O
O
NCH3
R
R
R
O
N
S
S
Lawesson's reagent
toluene, reflux
a
b
c
CH3 (36%)
OCH3 (30%)
OCH3
Cl
R
X
O
1
2c
Scheme 1. Thionation of precursors 1 for the synthesis of thiocoumarins and thioquinolone 2.
atom, yielding a constrained sulfur-bridged product and conse-
quently, coupling to the amino acid did not take place.
As a result, thiocoumarin and thioquinolone conjugates 4 had to
be synthesised from the carbonyl precursors 3, by reaction with
Lawesson’s reagent in dry toluene in moderate to good yields
(50e80%) (Scheme 2, Table 1).
Table 1
Synthesis, UV/vis absorption and emission data for coumarin and quinolone con-
jugates 3 and their thionated analogues 4, in absolute ethanol
Compound
Yield (%)
Absorption
Emission
labs (nm)
log
3
lem (nm)
fF
Stokes’
The synthesised thiocompounds 4 were chosen in order to
evaluate the influence of the nature of the heterocycle and the
type and position of the substituents at the heterocyclic moiety on
the photocleavage properties of the conjugates. The structure of
the compounds was confirmed by the usual spectroscopic
techniques.
The replacement of the carbonyl at the heterocycle by a thio-
carbonyl (leaving the remaining carbonyl groups in the conjugate
unchanged) was supported by IR spectroscopy (absorption bands
between 1245 and 1290 cmꢀ1). In the 1H NMR spectra, the expected
signals of the heterocycle and the amino acid residue were visible,
with the main difference being the chemical shift of proton H-3 that
shift (nm)
3a5
3b
3c
84
67
62
91
56
62
96
59
70
73
50
80
55
78
30
52
317
322
328
330
339
354
355
354
377
395
385
384
388
401
395
393
3.92
4.18
3.95
3.68
3.71
3.70
3.78
3.69
4.23
4.16
4.31
4.19
4.14
4.13
3.69
3.49
397
397
377
385
396
418
417
414
425
428
433
433
429
428
469
444
0.026
0.154
0.059
0.100
0.074
0.049
0.052
0.049
0.009
0.007
0.008
0.007
0.010
0.005
0.014
0.012
80
75
49
55
57
64
62
60
48
33
48
49
41
27
74
51
3d5
3e5
3f5
3g
3h
4a
4b
4c
4d
4e
4f
4g
4h
was displaced to higher
jacent thiocarbonyl group, from
coumarins, and from 7.55 to 7.74 ppm, for thioquinolones, when
compared to the carbonyl precursors ( 6.27 to 6.37 ppm, for cou-
marins, and
6.64 to 6.83 ppm, for quinolones).5 The trans-
d
due to the deshielding effect of the ad-
d
7.06 to 7.15 ppm, for thio-
d
d
d
formation of the carbonyl into the thiocarbonyl group at position-2
Compounds 1b,c were used in the synthesis of ester conjugates
3b,c, by a potassium fluoride coupling reaction to the carboxylic
acid group of N-benzyloxycarbonyl-phenylalanine, in N,N-dime-
thylformamide at room temperature (Scheme 2, Table 1).13 The
corresponding phenylalanine conjugates 3b,c were obtained in
good yields (62 and 67%, respectively). Quinolone 1f bearing a
of the heterocyclic ring was also supported by the 13C NMR spectra
signals for the C]S carbon from
d 196.86 to 196.99 ppm, for thio-
coumarins, and from 181.11 to 184.93 ppm, for thioquinolones
d
(the C]O carbon signal in the precursors appearing from
d 158.54
to 161.51 ppm).5

7894
A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
R
O
Cl
O
O
R
N
H
O
Ph
R1
R2
R1
R2
O
O
CO2H
Ph
O
N
H
KF, DMF, rt
X
O
X
3
1
Lawesson's reagent
toluene, reflux
X
O
O
R1
H
H
H
H
R2
CH3
OCH3 CH2Ph
OCH3 CH2Ph
H
H
H
H
H
R
a
b
c
d
e
f
CH2Ph
R
O
NCH3
NCH3
NCH3 CH3
NCH3 OCH3
NCH3 OCH3
NCH3 OCH3
O
CH2Ph
CH2Ph
CH2Ph
CH3
(CH2)2CO2CH3
N
H
O
Ph
R1
R2
O
S
4
X
g
h
Scheme 2. Synthesis of thionated amino acid ester conjugates 4.
The photophysical properties (absorption and emission max-
ima, molar absorption coefficients and fluorescence quantum
yields, fF) of the newly synthesised compounds bearing a cou-
marin (2a,b, 3a,b and 4a,b) or a quinolone (3c and 4ceh) were
evaluated by tracing the UV/vis absorption and emission spectra
of degassed 10ꢀ5 M solutions in absolute ethanol (Table 1). Rela-
tive fluorescence quantum yields were calculated using 9,10-
diphenylanthracene as standard (fF¼0.95 in ethanol),14 for cou-
marin conjugates 3b,c, or a 0.05 M solution of quinine in sulfuric
acid as standard (fF¼0.546)15 in case of thiocoumarin 2a,b and
thioquinolone 4aeh conjugates. For the fF determination, the
fluorescence standard was excited at the wavelengths of maxi-
mum absorption found for each of the compounds to be tested
and in all fluorimetric measurements, the absorbance of the so-
lution did not exceed 0.1.
By comparison of the data of thiocoumarins 2a,b with the cor-
responding precursors 1a,b, it was found that the fluorescence
quantum yield was lower for the thionated derivatives. The same
observation was made when comparing thiocoumarin conjugates
4a,b with the corresponding precursors 3a,b, and the thio-
quinolone conjugates 4ceh with the corresponding precursors
3ceh, probably due to the heavy induced spineorbit coupling by
the sulfur atom giving rise to an efficient intersystem crossing
mechanism that lowers the fluorescence emission.16 Upon in-
troduction of the sulfur atom, a considerable bathochromic shift in
absorption (between 47 and 73 nm, being larger for coumarins) and
emission (between 10 and 60 nm) was observed, thus confirming
our expectations in terms of the shift to longer wavelengths of
absorption and emission maxima. In terms of the influence of the
nature of heterocycle, the quinolones displayed longer wavelengths
of absorption and emission, both for carbonyl and thiocarbonyl
compounds. In terms of the influence of the nature of the sub-
stituent at the heterocycle, for example, by comparing quinolones
3def, it was found that the compounds with a methyl 3e and
a methoxy group 3f absorbed and emitted at longer wavelengths,
when compared to the unsubstituted quinolone 3d. As for the in-
fluence of the position of the substituent at the heterocycle, com-
parison of methoxyquinolones 3c and 3f revealed that substitution
at position-7, in 3f, shifted both absorption and emission bands, by
26 and 41 nm, respectively, when compared to 3c. Considering the
Stokes’ shifts, it was found that thionated compounds had smaller
Stokes’ shifts. The amino acid structure in compounds 3feh and
4feh did not have a marked influence in the photophysical prop-
erties, as the obtained data were identical for the three amino acid
representatives.
2.2. Photolysis studies of ester conjugates 3b, c, g, h and 4aeh
The purpose of this work was to assess the photostability of the
link between the thionated analogues of coumarin or quinolone
and the model amino acid, in ester conjugates, having in mind the
development of photocleavable protecting groups with improved
properties, namely the lability at longer wavelengths of irradiation
that could be useful for biological studies. Therefore, photolysis
studies were carried out by irradiating 1ꢁ10ꢀ4 M solutions of
coumarin and quinolone conjugates 3b, c, g, h and 4aeh in
methanol/HEPES buffer solution (80:20) in a Rayonet RPR-100
reactor, at 254, 300, 350 and 419 nm. The solvent system was
chosen based on previous studies regarding different organic
solvents with varying aqueous content, which revealed that
photocleavage was faster in a mixture of methanol and HEPES
buffer solution.17 The course of the photocleavage reaction was
followed by reverse phase HPLC with UV detection and 1H NMR.
The determined irradiation time represents the time necessary for
the consumption of the starting materials until less than 5% of the
initial peak area (A) was detected. Based on HPLC data, the plot of
ln A versus irradiation time showed a linear correlation for the
disappearance of the starting material, which suggested a first
order reaction, obtained by the linear least squares methodology
for a straight line, with high correlation coefficients (Table 2). The
photolysis data of conjugates 3a,def had been previously reported
by us and are presented in Table 2 for easier comparison.5
It was intended to assess the potential for the applicability of
thiocoumarin and thioquinolones as photocleavable protecting
groups, when compared to the earlier reported non-thionated
precursors, and at longer wavelengths (350 and 419 nm) indeed the
thionated compounds cleaved much faster. Taking into consider-
ation the influence of the structure of the heterocycle on the pho-
tocleavage rates, it was found that the thioquinolone conjugates
4cef had shorter irradiation times, at all wavelengths of irradiation
studied, when compared to the thiocoumarin conjugates 4a,b.
Among the thioquinolone conjugates 4cef, the derivatives bearing
a methoxy group were the fastest to cleave (less than 1 min at all
wavelengths of irradiation). For these compounds, some consider-
ations on the type of substituent and its position can be drawn: the
position of the methoxy group does not influence the cleavage rates
(compare 4c and 4f) and a higher electron donor character of the
substituent seems to be beneficial for the cleavage (compare 4d, 4e
and 4f).
As for the effect of the amino acid structure in compounds 3feh
and 4feh, the phenylalanine conjugate cleaved faster than the

A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
7895
Table 2
Irradiation times (tirr, in min) and photochemical quantum yields (fP) for the photolysis of conjugates 3b, c, g, h and 4aeh at different wavelengths in methanol/HEPES buffer
(80:20) solution
Compound
254 nm
300 nm
350 nm
419 nm
tirr
fP
tirr
fP
tirr
fP
tirr
fP
3a5
3b
3c
34
32
0.9
9.3
2.3
0.8
d
1.11ꢁ10ꢀ3
1.74ꢁ10ꢀ3
1.94ꢁ10ꢀ2
3.42ꢁ10ꢀ3
1.13ꢁ10ꢀ2
1.58ꢁ10ꢀ2
d
129
34
0.5
7.6
1.7
1.2
d
6.35ꢁ10ꢀ5
2.35ꢁ10ꢀ4
2.28ꢁ10ꢀ2
5.30ꢁ10ꢀ3
3.89ꢁ10ꢀ2
5.52ꢁ10ꢀ2
d
3293
253
12
4.89ꢁ10ꢀ5
5.39ꢁ10ꢀ5
1.10ꢁ10ꢀ3
3.12ꢁ10ꢀ4
3.67ꢁ10ꢀ3
3.48ꢁ10ꢀ2
1.81ꢁ10ꢀ2
2.22ꢁ10ꢀ2
1.65ꢁ10ꢀ4
1.06ꢁ10ꢀ4
1.24ꢁ10ꢀ2
8.41ꢁ10ꢀ4
1.08ꢁ10ꢀ2
3.77ꢁ10ꢀ2
1.05ꢁ10ꢀ3
5.91ꢁ10ꢀ4
d
d
d
d
5.40ꢁ10ꢀ3
304
d
3d5
3e5
3f5
3g
3h
4a
69
d
3.9
0.6
3.4
3.1
32
46
0.6
10
1.1
0.5
8.5
10
193
34
452
780
33
2.70ꢁ10ꢀ2
1.61ꢁ10ꢀ1
1.12ꢁ10ꢀ2
7.02ꢁ10ꢀ3
2.36ꢁ10ꢀ4
7.06ꢁ10ꢀ5
1.90ꢁ10ꢀ2
1.32ꢁ10ꢀ3
1.43ꢁ10ꢀ2
2.13ꢁ10ꢀ2
5.92ꢁ10ꢀ4
6.96ꢁ10ꢀ4
d
d
d
d
13
30
0.5
8.3
2.7
0.5
d
6.62ꢁ10ꢀ4
1.51ꢁ10ꢀ4
2.51ꢁ10ꢀ2
2.34ꢁ10ꢀ3
4.94ꢁ10ꢀ3
7.34ꢁ10ꢀ2
d
61
2.31ꢁ10ꢀ4
2.72ꢁ10ꢀ4
2.24ꢁ10ꢀ2
9.60ꢁ10ꢀ4
4.68ꢁ10ꢀ3
8.72ꢁ10ꢀ3
d
4b
4c
4d
4e
4f
4g
4h
101
0.7
10
2.0
0.7
d
37
0.6
9.8
1.1
0.4
104
133
d
d
d
d
alanine and glutamic acid conjugates and the thiocarbonyl de-
rivatives had practicable irradiation times at 419 nm.
The photochemical quantum yields (fP) were calculated based
on half-lives (t1/2), molar absorption coefficients at the irradiation
Signals due to by-products related to the heterocycle were also
visible in the 1H NMR spectra. The same experiment was done for
the N-benzyloxycarbonylalanine conjugate 3g and the results for
irradiation at 350 nm can be seen in Fig. 1c.
wavelength ( ) and the incident photon flux (I0), which was de-
3
termined by potassium ferrioxalate actinometry.18 The photo-
cleavage process was not as efficient as desirable (see fP in Table 2),
possibly by deactivation via fluorescence pathways that compete
with the photochemical reaction, as well as the type of reactor used
(open chamber reactor).
3. Conclusions
Thionated coumarin and quinolone amino acid conjugates 4
were readily obtained in fair to good yields through a reaction of
the corresponding precursor coumarin and quinolone conjugates 3.
Upon photolysis, it was found that the thionated conjugates 4
cleaved significantly faster than the parent conjugates 3 at 419 nm
and cleavage kinetics parameters were obtained. The cleavage of
the ester bond and the release of the amino acid in short irradiation
times at all considered wavelengths of irradiation suggests that
thionated quinolone derivatives 4 may be considered as promising
alternatives as photocleavable protecting groups for carboxylic
acids. Fast photolysis at 419 nm for thioquinolone conjugates 4cef
is particularly interesting, since this wavelength is the most suit-
able for practical caging strategies as it avoids cell damage due to
short wavelengths. In addition, the fact that some of the proposed
heterocycles display very different photocleavage rates at the same
wavelength, may provide the possibility of selective removal of one
group in the presence of the other group.
Regarding the irradiation times obtained for thionated com-
pounds 4 and their precursors 3, at all wavelengths, one might
suggest the feasibility of selective photodeprotection. If thionated
and non-thionated groups of the same type were to be considered
for the protection of a bifunctional molecule through ester bonds, in
the case of thioquinolones, this situation could be possible espe-
cially at 419 nm, whereas at 254 nm and 300 nm thioquinolones and
their precursors present similar irradiation times. At 419 nm, this
observation is also true for thiocoumarins and their precursors. Also,
considering the possibility of selective photodeprotection between
the two heterocycles, among the thionated compounds it should be
possible to use thiocoumarins and thioquinolones simultaneously
as protecting groups for a bifunctional molecule, to cleave selec-
tively at the same wavelength, for example, the pairs 4b/4c or 4b/4f.
Within the same type of heterocycle, for the thioquinolones some
selectivity could be possible between the unsubstituted heterocycle
(in 4d) and the methoxylated heterocycles (in 4c and 4f), that cleave
ca. 15e20 times faster than the former, at the same wavelength of
irradiation. However, this type of selectivity can only be achieved if
no energy transfer occurs between the two heterocycles.19
Considering the short irradiation times obtained for thionated
compounds 4aef at 419 nm, the presented thiocoumarins and
thioquinolones are very promising as photocleavable protecting
groups for the long-wavelength photolytic release of biomolecules,
for example, in caging strategies.
Additionally to monitoring the photolysis process by HPLC/UV
detection, the release of N-benzyloxycarbonyl-phenylalanine, as
the expected product of the conjugate’s photolysis, was also fol-
lowed by 1H NMR in a methanol-d4/D2O (80:20) solution. Upon
irradiation at 419 nm of solutions of thiocoumarin 4b (Fig. 1a) and
thioquinolone 4c (Fig. 1b), the signals due to the benzylic-type CH2
4. Experimental section
4.1. General
All melting points were measured on a Stuart SMP3 melting
point apparatus. TLC analyses were carried out on 0.25 mm thick
precoated silica plates (Merck Fertigplatten Kieselgel 60F254) and
spots were visualised under UV light. Chromatography on silica gel
was carried out on Merck Kieselgel (230e240 mesh). IR spectra
were determined on a BOMEM MB 104 spectrophotometer. UV/vis
absorption spectra (200e700 nm) were obtained using a Shimadzu
UV/2501PC spectrophotometer. NMR spectra were obtained on
a Varian Unity Plus Spectrometer at an operating frequency of
300 MHz for 1H and 75.4 MHz for 13C or a Bruker Avance III 400 at
an operating frequency of 400 MHz for 1H and 100.6 MHz for 13C
using the solvent peak as internal reference at 25 ꢂC. All chemical
shifts are given in parts per million using dH Me4Si¼0 ppm as ref-
erence and J values are given in hertz. Assignments were made by
comparison of chemical shifts, peak multiplicities and J values and
were supported by spin decoupling-double resonance and
at position-4 of the heterocycles, visible at about
disappeared completely and the CH and CH2 signals of phenyl-
alanine in the conjugate form gradually decreased, and a close set
of signals appeared at about 4.50 and 3.20 ppm, respectively.
d 5.30e5.40 ppm,
a
b
d

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A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
Fig. 1. Partial 1H NMR spectra in methanol-d4/D2O (80:20) of the photolysis at 419 nm of (a) thionated coumarin conjugate 4b: (a) before irradiation; (b) after irradiation for 60 min;
(c) after irradiation for 165 min; (b) thionated quinolone conjugate 4c: (a) before irradiation; (b) after irradiation for 2 min; (c) after irradiation for 15 min. (c) Photolysis at 350 nm
of quinolone conjugate 3g: (a) before irradiation; (b) after irradiation for 5 min; (c) after irradiation for 10 min.

A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
7897
bidimensional heteronuclear correlation techniques. Low and high
resolution mass spectrometry analyses were performed at the
‘C.A.C.T.I.dUnidad de Espectrometria de Masas’, at University of
Vigo, Spain. Fluorescence spectra were collected using a Fluo-
roMax-4 spectrofluorometer. Photolyses were carried out using
a Rayonet RPR-100 chamber reactor equipped with 10 lamps of 254
(35 W), 300 (21 W), 350 (24 W) and 419 (14 W) nm. HPLC analyses
1.2 Hz, H3), 6.87(1H, dd, J 8.8 and 2.4 Hz, H6), 6.94(1H, d, J 2.8 Hz, H8),
7.65 (1H, d, J 8.4 Hz, H5). 13C NMR (100.6 MHz, DMSO-d6):
d¼28.90
(NCH3), 55.53 (OCH3), 59.51 (CH2), 99.05 (C8), 109.45 (C6), 112.25
(C4a), 113.86 (C3), 125.71 (C5), 141.30 (C8a), 149.67 (C4), 161.47 (C7),
161.47 (C2). FTIR (KBr 1%, cmꢀ1):
1572, 1458, 1433, 1353, 1336, 1299, 1268, 1233, 1203, 1185, 1127, 1095,
n
¼3430, 2388, 2293, 2226, 1641,
1075, 1048, 947, 867, 842, 791, 666, 511. UV/vis (ethanol, nm): lmax
were performed using a Licrospher 100 RP18 (5
mm) column in an
(log
3
)¼325 (3.76). MS: m/z (EI) 219 (Mþ, 100). HRMS: m/z (ESI) calcd
HPLC system composed by a Jasco PU-980 pump, a Shimadzu SPD-
GAV UV/vis detector and a Shimadzu C-RGA Chromatopac register.
All commercial reagents were used as received. The synthesis of
compounds 1a,5 1b,8b and 3a,def5 was previously reported by us.
for C12H13NO3 219.0900, found 219.0895.
The alcohol (1 equiv, 0.050 g, 2.28ꢁ10ꢀ1 mmol) was stirred with
thionyl chloride (20 equiv, 0.33 mL, 4.56 mmol) in dichloromethane
(10 mL) for 1 day at room temperature. The mixture was filtered and
after removal of the solvent under reduced pressure, 4-chloromethyl-
7-methoxy-1-methylquinolin-2(1H)-one 1c was obtained as beige
solid (0.030 g, 0.13 mmol, 55%). Mp¼108e110 ꢂC. 1H NMR (400 MHz,
4.2. Synthesisof 4-chloromethyl-7-methoxy-1-methylquinolin-
2(1H)-one, 1c
CDCl3):
(1H, s, H3), 6.83 (1H, d, J 2.4 Hz, H8), 6.90 (1H, dd, J 8.8 and 2.4 Hz, H6),
7.73 (1H, d, J 9.2 Hz, H5). 13C NMR (100.6 MHz, CDCl3):
d¼3.69 (3H, s, NCH3), 3.94 (3H, s, OCH3), 4.69 (2H, s, CH2), 6.67
1,4-Dimethyl-7-methoxy-quinolin-2(2H)-one was prepared by
reaction of the 3-methoxy-N-methylaniline (1 equiv, 0.250 g,
1.82 mmol)withethyl3-oxobutanoate(2 equiv, 0.46 mL, 3.65 mmol),
by stirring at 180 ꢂC for 45 min. After evaporation of the excess ethyl
3-oxobutanoate under vacuum, the residue was stirred with aqueous
70% sulfuric acid, at 95 ꢂC for 45 min. After cooling to room temper-
ature, water(10 mL)wasadded tothemixture, followed byextraction
with ethyl acetate (3ꢁ15 mL). The organic layer was dried with an-
hydrous magnesium sulfate, filtered and evaporated under reduced
pressure. The resulting solid was purified by silica gel column chro-
matography using chloroform/methanol (100:1) as eluent. Fractions
containing the product were combined and evaporated, yielding
a light yellow solid (0.267 g,1.31 mmol, 72%). Mp¼60e62 ꢂC.1H NMR
d¼29.54
(NCH3), 42.41 (CH2), 55.61 (OCH3), 99.34 (C8), 109.59 (C6), 112.71
(C4a), 119.00 (C3), 126.37 (C5), 142.10 (C8a), 144.10 (C4), 161.87 (C7),
162.23 (C2). FTIR (KBr 1%, cmꢀ1):
1670, 1625, 1595, 1561, 1523, 1451, 1435, 1415, 1395, 1327, 1305, 1284,
1267, 1231, 1178, 1079, 1059, 1036, 969, 925, 886, 846, 809, 799, 736,
n
¼3443, 3088, 2965, 2935, 2840,
703, 698, 682, 665, 635, 613. UV/vis (ethanol, nm): lmax (log
3
)¼327
(4.24). MS: m/z (EI) 239 (Mþ 37Cl, 30), 237 (Mþ 35Cl, 100). HRMS: m/z
(EI) calcd for C12H12NO237Cl [Mþ] 239.0535, found 239.0527; calcd for
C12H12NO235Cl [Mþ] 237.0546, found 237.0557.
4.3. General procedure for the synthesis of coumarin and
quinolone conjugates 3b,c, 3geh
(300 MHz, CDCl3):
d
¼2.41 (3H, d, J 0.9 Hz, CH3), 3.66 (3H, s, NCH3),
3.92 (3H, s, OCH3), 6.44 (1H, d, J 9.0 Hz, H5), 6.78 (1H, d, J 2.7 Hz, H8),
6.83 (1H, dd, J 9.0 and 2.4 Hz, H6), 7.60 (1H, d, J 0.9 Hz, H3). 13C NMR
4-Chloromethyl-7-methoxycoumarin 1b, 4-chloromethyl-7-
methoxy-1-methylquinolone 1c or 4-chloromethyl-6-methoxy-1-
methylquinolone 1f (1 equiv) were dissolved in dry N,N-dime-
(75.4 MHz, CDCl3):
(C8), 109.05 (C6), 115.50 (C4a), 118.14 (C5), 126.53 (C3), 141.40 (C4),
146.29 (C8a), 161.44 (C7), 162.57 (C2). FTIR (KBr 1%, cmꢀ1):
d¼18.92 (CH3), 29.22 (NCH3), 55.50 (OCH3), 98.83
n¼3426,
thylformamide (5 mL). N-Benzyloxycarbonyl
L-phenylalanine, L-al-
2973, 2935, 1647, 1620, 1588, 1519, 1490, 1467, 1442, 1415, 1392, 1370,
1357, 1343, 1319, 1297, 1264, 1231, 1217, 1184, 1164, 1147, 1110, 1066,
1033, 972, 862, 819, 789, 751, 741, 707, 673, 635, 606. UV/vis (ethanol,
anine or -glutamic acid methyl ester (1 equiv) and potassium
L
fluoride (3 equiv) were added to the mixture. The reaction mixture
was stirred at room temperature for 2 days. The mixture was fil-
tered and the solvent was removed by rotary evaporation. The
resulting solid was purified by column chromatography dichloro-
methane/methanol (100:1) and mixtures of increasing polarity.
nm): lmax (log
(EI) calcd for C12H13NO2 203.0946, found 203.0946.
3
)¼325 (4.09). MS: m/z (EI) 203 (Mþ, 100). HRMS: m/z
This compound (1 equiv, 0.170 g, 8.37ꢁ10ꢀ1 mmol) was reacted
with selenium dioxide (4 equiv, 0.376 g, 3.35 mmol) in chloroben-
zene (30 mL), by heating at reflux for 2 days. The mixture was fil-
tered hot and the solvent was removed by rotary evaporation. The
resulting solid was purified by silica gel column chromatography
using chloroform/methanol (100:1) as eluent. Fractions containing
the product were combined and evaporated, yielding 7-methoxy-
4.3.1. N-Benzyloxycarbonyl-L-phenylalanine (7-methoxycoumarin-4-
yl) methyl ester, 3b. The product was obtained as beige solid
(0.217 g, 4.45 mmol, 67%). Mp¼114.0e116.0 ꢂC. 1H NMR (300 MHz,
CDCl3):
(1H, m,
d
¼3.14 (2H, d, J 6.3 Hz,
bCH2), 3.86 (3H, s, OCH3), 4.72e4.79
a
H), 5.10 (2H, s, CH2 Z), 5.22 (2H, s, CH2), 5.42 (1H, d, J 7.8 Hz,
1-methylquinolin-2(2H)-one-4-carbaldehyde as
a
yellow solid
NH), 6.18 (1H, s, H3), 6.82 (1H, br s, H8), 6.83 (1H, dd, J 7.2 and
1.5 Hz, H6), 7.12 (2H, dd, J 7.8 and 2.7 Hz, H20 and H60), 7.24e7.32
(9H, m, H5, 3ꢁPheH Phe and 5ꢁPheH Z). 13C NMR (75.4 MHz,
(0.161 g, 0.74 mmol, 89%). Mp¼173e175 ꢂC. 1H NMR (400 MHz,
CDCl3):
d
¼3.73 (3H, s, NCH3), 3.94 (3H, s, OCH3), 6.84 (1H, d, J
2.4 Hz, H8), 6.92 (1H, dd, J 8.8 and 2.4 Hz, H6), 7.27 (1H, s, H3), 7.76
CDCl3):
d
¼38.03 (
bCH2), 55.02 (aC), 55.608 (OCH3), 61.99 (CH2),
(1H, d, J 9.2 Hz, H5), 10.09 (1H, s, CHO). 13C NMR (100.6 MHz,
67.03 (CH2 Z), 101.05 (C8), 110.36 (C4a), 110.48 (C3), 112.53 (C6),
124.43 (C5), 127.28 (C40), 128.01 (C200 and C600), 128.12 (C400), 128.40
(C300 and C500), 128.65 (C30 and C50), 128.99 (C20 and C60), 135.18
(C10), 135.92 (C100), 147.98 (C4), 155.39 (C8a), 155.61 (C]O ure-
thane), 160.51 (C7), 162.76 (C2), 171.03 (C]O ester). FTIR (KBr 1%,
CDCl3):
d 29.96 (NCH3), 55.59 (OCH3), 99.16 (C8), 110.41 (C6), 110.43
(C4a),128.17 (C5),128.58 (C3),139.86 (C4),142.48 (C8a),162.10 (C7),
162.24 (C2), 193.19 (CHO). UV/vis (ethanol, nm): lmax (log
3 )¼328
(4.28). MS: m/z (ESI) 218 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for
C12H12NO3 218.08117, found 218.08103.
cmꢀ1):
n
¼3324, 3065, 3031, 2937, 1732, 1697, 1615, 1538, 1497, 1454,
This aldehyde (1 equiv, 0.161 g, 7.69ꢁ10ꢀ1 mmol) was reacted
with sodium borohydride (0.7 equiv, 0.020 g, 5.39ꢁ10ꢀ1 mmol) in
ethanol (10 mL) for 1 day at room temperature. The solvent was re-
moved under reduced pressure and after purification by silica gel
column chromatography using chloroform/methanol (100:1) as elu-
ent, 4-hydroxymethyl-7-methoxy-1-methylquinolin-2(1H)-one was
obtained as white solid (0.067 g, 0.31 mmol, 40%). Mp¼106e108 ꢂC.
1409, 1343, 1268, 1210, 1150, 1138, 1069, 1027, 985, 909, 875, 854,
795, 774, 450, 666, 750, 699, 557. UV/vis (ethanol, nm): lmax
(log
3
)¼322 (4.18). MS m/z (ESI, %): 488 ([MþH]þ, 100). HRMS: m/z
(ESI) calcd for C28H26NO7 488.17038, found 488.17032.
4.3.2. N-Benzyloxycarbonyl-
L-phenylalanine (7-methoxy-1-
methylquinolin-2(1H)-one-4-yl) methyl ester, 3c. The product was
1H NMR (400 MHz, DMSO-d6):
OCH3), 4.71 (2H, d, J 4.0 Hz, CH2), 5.44 (1H, t, J 5.2 Hz, OH), 6.49 (1H, t, J
d
¼3.58 (3H, s, NCH3), 3.88 (3H, s,
obtained as
a
white solid (0.018 g, 0.04 mmol, 62%).
Mp¼138.1e139.8 ꢂC. 1H NMR (400 MHz, CDCl3):
¼3.14 (2H, t, J
d

7898
A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
6.8 Hz,
b
CH2), 3.69 (3H, s, NCH3), 3.93 (3H, s, OCH3), 4.72e4.77 (1H,
4.4.1. 4-Chloromethyl-7-methylbenzopyran-2(1H)-thione, 2a. The
solid crude was purified by column chromatography using chlo-
roform/hexane (1:2) as eluent. The product was isolated as yellow
solid (0.160 g, 0.71 mmol, 36%). Mp¼173.2e175.1 ꢂC. 1H NMR
m, aH), 5.10 (2H, d, J 3.2 Hz, CH2 Z), 5.25e5.34 (3H, m, CH2 and NH),
6.59 (1H, s, H3), 6.83 (2H, m, H8 and H6), 7.09 (2H, dd, J 5.6 and
1.6 Hz, H20 and H60), 7.23e7.35 (8H, m, 3ꢁPheH Phe and 5ꢁPheH Z),
7.46 (1H, d, J 5.2 Hz, H5). 13C NMR (100.6 MHz, CDCl3):
d
¼29.50
(300 MHz, CDCl3):
d
¼2.48 (3H, s, CH3), 4.60 (2H, d, J 0.6 Hz, CH2),
(NCH3), 38.16 (
b
CH2), 54.98 (
a
C), 55.58 (OCH3), 63.27 (CH2), 67.08
7.20 (1H, dd, J 8.4 and 1.2 Hz, H6), 7.28 (1H, s, H3), 7.32 (1H, br s, H8),
(CH2 Z), 99.27 (C8), 109.62 (C6), 112.72 (C4a), 117.65 (C3), 125.74 (C5),
127.24 (C40), 128.10 (C200), 128.18 (C600), 128.50 (C300, C400 and C500),
128.66 (C30 and C50), 129.15 (C20 and C60), 135.32 (C10), 136.09 (C100),
141.84 (C8a), 142.20 (C4), 155.63 (C]O urethane), 161.81 (C7), 162.15
7.60 (1H, d, J 8.4 Hz, H5). 13C NMR (75.4 MHz, CDCl3):
d
¼21.76 (CH3),
41.13 (CH2), 116.50 (C4a), 117.36 (C8), 123.76 (C5), 126.79 (C6),
128.00 (C3), 140.58 (C4), 143.97 (C7), 156.69 (C8a), 197.11 (C2). FTIR
(KBr 1%, cmꢀ1):
n
¼3054, 3021, 2915, 1622, 1606, 1544, 1436, 1409,
(C2), 171.13 (C]O ester). FTIR (KBr 1%, cmꢀ1):
n¼3360, 3064, 3031,
1301, 1277, 1263, 1245, 1205, 1177, 1154, 1112, 1054, 953, 923, 894,
2934,1738,1699,1662,1630,1604,1561,1527,1498, 1454,1320,1035,
1257, 1228, 1185, 1154, 1085, 1068, 1018, 974, 872, 843, 809, 755, 735,
881, 819, 711, 699, 666, 651, 643. UV/vis (ethanol, nm): lmax (log
3
)¼
380 (4.26). MS m/z (ESI, %): 227 ([MþH]þ 37Cl, 34), 225 ([MþH]þ
35Cl, 100). HRMS m/z (ESI) calcd for C11H1037ClOS 227.01059, found
227.01328; calcd for C11H1035ClOS 225.01354, found 225.01328.
703, 665, 508. UV/vis (ethanol, nm): lmax (log
3 )¼328 (3.95). MS m/z
(ESI, %): 501 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for C29H29N2O6
501.20065, found 501.20201.
4.4.2. 4-Chloromethyl-7-methoxybenzopyran-2(1H)-thione, 2b. The
solid crude was purified by column chromatography using chlo-
roform/hexane (1:3) and mixtures of increasing polarity as eluent.
The product was isolated as yellow solid (0.056 g, 0.23 mmol, 30%).
4.3.3. N-Benzyloxycarbonyl-L-alanine (6-methoxy-1-methylquinolin-
2(1H)-one-4-yl) methyl ester, 3g. The product was obtained as
a light yellow solid (0.130 g, 0.31 mmol, 96%). Mp¼144.0e145.0 ꢂC.
1H NMR (400 MHz, CDCl3):
s, NCH3), 3.87 (3H, s, OCH3), 4.50 (1H, br s,
d
¼1.48 (3H, d, J 6.8 Hz,
b
CH3), 3.72 (3H,
Mp¼169.3e171.0 ꢂC. 1H NMR (300 MHz, CDCl3):
d
¼3.91 (3H, s,
a
H), 5.11 (2H, d, J 4.2 Hz,
OCH3), 4.58 (2H, s, CH2), 6.97 (1H, dd, J 8.7 and 2.7 Hz, H6), 7.01 (1H,
CH2 Z), 5.38 (3H, m, CH2 and NH), 6.81 (1H, s, H3), 7.03 (1H, d, J
2.4 Hz, H5), 7.23 (1H, dd, J 8.0 and 2.4 Hz, H7), 7.34e7.37 (6H, m,
d, J 2.7 Hz, H8), 7.22 (1H, s, H3), 7.63 (1H, d, J 8.7 Hz, H5). 13C NMR
(75.4 MHz, CDCl3):
(C4a), 114.30 (C6), 125.12 (C5),126.33 (C3), 141.01 (C4), 158.50 (C8a),
163.26 (C7), 197.05 (C2). FTIR (KBr 1%):
d¼41.18 (CH2), 55.94 (OCH3), 100.76 (C8), 112.58
5ꢁPheH Z and H8). 13C NMR (100.6 Hz, CDCl3):
d
¼18.48 (
bCH3),
29.60 (NCH3), 49.77 (
a
C), 55.75 (OCH3), 63.10 (CH2), 67.05 (CH2 Z),
n
¼3064, 2969, 2935, 1728,
106.63 (C5), 116.04 (C8), 119.13 (C7), 119.33 (C4a), 120.91 (C3),
128.11 (C20), 128.19 (C60), 128.51 (C30, C40 and C50), 134.58 (C8a),
136.12 (C10), 141.91 (C4), 154.82 (C6),155.61 (C]O urethane),161.22
1625, 1600, 1544, 1518, 1464, 1443, 1426, 1408, 1398, 1351, 1301,
1267, 1251, 1243, 1213, 1171, 1102, 1057, 1017, 985, 969, 158, 701, 657,
643, 621 cmꢀ1. UV/vis (ethanol, nm): lmax (log
3 )¼391 (4.17). MS m/
(C2), 172.52 (C]O ester). FTIR (KBr 1%, cmꢀ1):
1746,1717,1656,1622,1592,1572,1510,1462,1433,1383,1333,1281,
1247, 1204, 1179, 1116, 1071, 1041, 909, 880, 813, 732, 699, 646. UV/
n
¼3292, 2849, 2250,
z (ESI, %): 243 ([MþH]þ 37Cl, 34), 241 ([MþH]þ 35Cl, 100). HRMS m/z
(ESI) calcd for C11H1037ClO2S 243.00550, found 243.00537; calcd for
C11H1035ClO2S 241.00845, found 241.00836.
vis (ethanol, nm): lmax (log
3
)¼355 (3.78). MS m/z (ESI, %): 425
([MþH]þ, 100). HRMS: m/z (ESI) calcd for C23H25N2O6 425.17135,
4.4.3. N-Benzyloxycarbonyl-L-phenylalanine (7-methylbenzopyran-
found 425.17049.
2(1H)-thione-4-yl) methyl ester, 4a. The solid crude was purified by
column chromatography using chloroform as eluent. The product
was isolated as yellow solid (0.072 g, 0.15 mmol, 70%).
4.3.4. 1-(6-Methoxy-1-methylquinolin-2(1H)-one-4-yl)methyl-5-
methyl-2-(N-benzyloxycarbonyl)aminopentanedioate, 3h. The prod-
uct was obtained as a light yellow oil (0.087 g, 0.18 mmol, 59%). 1H
Mp¼149.0e150.8 ꢂC. 1H NMR (400 MHz, CDCl3):
d
¼2.48 (3H, s,
CH3), 3.14 (2H, d, J 6.4 Hz, bCH2), 4.72e4.77 (1H, m, aH), 5.12 (2H, s,
NMR (400 MHz, CDCl3):
(2H, m, CH2), 3.63 (3H, s, OCH3), 3.67 (3H, s, OCH3), 3.85 (3H, s,
NCH3), 4.49 (1H, br s, H), 5.08 (2H, d, J 4.2 Hz, CH2 Z), 5.33 (2H, s,
d
¼2.04e2.27 (2H, m,
b
CH2), 2.42e2.44
CH2 Z), 5.19e5.21 (3H, m, NH and CH2), 7.09e7.15 (4H, m, H3, H6,
H20 and H60 Phe), 7.24e7.27 (4H, m, 3ꢁPheH Phe and H400),
7.35e7.36 (6H, m, H5, H8 and 4ꢁPheH Z). 13C NMR (100.6 MHz,
g
a
CH2), 5.71 (1H, d, J 8.0 Hz, NH), 6.78 (1H, s, H3), 7.01 (1H, d, J 2.0 Hz,
H5), 7.19 (1H, dd, J 8.0 and 2.0 Hz, H7), 7.28e7.32 (6H, m, 5ꢁPheH Z
CDCl3):
d¼21.76 (CH3), 38.22 (
bCH2), 55.04 (aC), 61.89 (CH2), 67.25
(CH2 Z), 116.29 (C4a), 117.33 (C8), 123.20 (C5), 126.38 (C3), 126.79
(C6), 127.47 (C40), 128.19 (C200), 128.28 (C600), 128.55 (C300 and C500),
128.81 (C30 and C50), 129.07 (C400, C20, C60), 135.11 (C10), 136.00
(C100), 138.95 (C4), 143.86 (C7), 155.64 (C]O urethane), 156.43
and H8). 13C NMR (100.6 Hz, CDCl3):
29.85 (gCH2), 51.76 (OCH3), 53.49 (aC), 55.66 (OCH3), 63.21 (CH2),
d
¼27.11 (
bCH2), 29.46 (NCH3),
67.02 (CH2 Z), 106.51 (C5), 115.94 (C8), 119.10 (C7), 119.23 (C4a),
120.95 (C3), 127.99 (C20), 128.08 (C60), 128.40 (C30, C40 and C50),
134.44 (C8a), 136.02 (C10), 141.72 (C4), 154.74 (C6), 155.95 (C]O
urethane), 161.09 (C2), 172.94 (C]O ester). FTIR (KBr 1%, cmꢀ1):
(C8a), 171.11 (C]O ester), 196.99 (C2). FTIR (KBr 1%, cmꢀ1):
3060, 3030, 2921, 1731, 1692, 1623, 1606, 1546, 1495, 1454, 1380,
1346, 1289, 1276, 1225, 1187, 1168, 1152, 1112, 1070, 1022, 982, 885,
n
¼3307,
n
¼3361, 3016, 2953, 1729, 1655, 1622, 1590, 1572, 1510, 1463, 1434,
815, 781, 751, 697, 666. UV/vis (ethanol, nm): lmax (log
3
)¼377
1247, 1213, 1181, 1043, 878, 813, 754, 698, 666. UV/vis (ethanol,
nm): lmax (log
HRMS: m/z (ESI) calcd for C26H29N2O8 497.19247, found 497.19198.
(4.23). MS m/z (ESI, %): 488 ([MþH]þ, 100). HRMS: m/z (ESI) calcd
3
)¼354 (3.69). MS m/z (ESI, %): 497 ([MþH]þ, 100).
for C28H26NO5S 488.15262, found 488.15142.
4.4.4. N-Benzyloxycarbonyl-L-phenylalanine (7-methoxybenzopyran-
4.4. General procedure for the synthesis of thionated cou-
marins and quinolones 2a,b and 4aeh
2(1H)-thione-4-yl) methyl ester, 4b. The solid crude was purified by
column chromatography using chloroform/methanol (100:1) as
eluent. The product was isolated as a yellow solid (0.098 g,
0.19 mmol, 73%). Mp¼122.1e123.7 ꢂC. 1H NMR (400 MHz, CDCl3):
Compounds 2a,b and 4aeh were prepared by reaction of the
corresponding chloromethylated coumarins 1a,b (1 equiv), cou-
marin conjugates 3a,b (1 equiv) or quinolone conjugates 3ceh
(1 equiv) with Lawesson’s reagent (0.8 equiv, in case of coumarins
and 0.6 equiv, in case of quinolones) in dry toluene (10 mL) by
heating at reflux for 1e2 days. The mixture was filtered hot and the
solvent was removed under reduced pressure in a rotary evapora-
tor. The solid crude was purified by column chromatography.
d¼3.15 (2H, d, J 6.4,
bCH2 Phe), 3.91 (3H, s, OCH3), 4.73e4.78 (1H, m,
aH), 5.13 (2H, s, CH2 Z), 5.19 (2H, d, J 6.4 Hz, CH2), 5.25 (1H, d, J
8.0 Hz, NH), 6.91 (1H, dd, J 8.8 and 2.4 Hz, H6), 7.00 (1H, d, J 2.4 Hz,
H8), 7.06 (1H, s, H3), 7.12 (2H, dd, J 8.0 and 2.0 Hz, H20 and H60),
7.24e7.39 (9H, m, H5, 3ꢁPheH Phe and 5ꢁPheH Z). 13C NMR
(100.6 MHz, CDCl3):
(CH3), 61.94 (CH2), 67.20 (CH2 Z), 100.71 (C8), 112.34 (C4a), 114.25
d
¼38.18 (
bCH2 Phe), 55.02 (aC Phe), 55.89

A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
7899
(C6), 124.54 (C5), 124.72 (C3), 127.43 (C40), 128.15 (C200), 128.24
(C600), 128.51 (C300, C400 and C500), 128.77 (C30 and C50), 129.06 (C20
and C60), 135.11 (C10), 135.99 (C100), 139.36 (C4), 155.61 (C¼O ure-
thane), 158.18 (C8a), 163.17 (C7), 171.08 (C¼O ester), 196.86 (C2).
(NCH3), 54.94 (
a
C), 62.86 (CH2), 67.09 (CH2 Z), 116.10 (C8), 122.21
(C4a), 124.18 (C5), 127.25 (C40), 128.10 (C200), 128.18 (C600), 128.49
(C300 and C500), 128.66 (C30 and C50), 129.11 (C20, C60 and C400), 132.47
(C3),132.69 (C7),134.27 (C6),134.56 (C4), 135.22 (C10), 136.08 (C100),
139.13 (C8a), 155.63 (C]O urethane), 171.18 (C¼O ester), 183.23
FTIR (KBr 1%, cmꢀ1):
n
¼3324, 3063, 3030, 2935, 1731, 1696, 1615,
1602, 1546, 1519, 1497, 1454, 1427, 1409, 1382, 1346, 1288, 1256,
1212, 1185, 1151, 1103, 1072, 1023, 881, 838, 815, 780, 750, 697, 666,
(C2). FTIR (KBr 1%, cmꢀ1):
n
¼3323, 3060, 3029, 2925, 1741, 1727,
1684, 1564, 1524, 1496, 1452, 1386, 1341, 1294, 1276, 1235, 1209,
1183, 1156, 1101, 1066, 1053, 996, 957, 891, 840, 813, 789, 741, 697,
642, 620. UV/vis (ethanol, nm): lmax (log
3
)¼397 (4.16). MS m/z (ESI,
%): 504 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for C28H26NO6S
665, 637. UV/vis (ethanol, nm): lmax (log
3 )¼388 (4.14). MS m/z (ESI,
504.14753, found 504.14629.
%): 501 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for C29H29N2O4S
501.18425, found 501.18243.
4.4.5. N-Benzyloxycarbonyl-L-phenylalanine (7-methoxy-1-
methylquinolin-2(1H)-thione-4-yl) methyl ester, 4c. The solid
crude was purified by column chromatography using chloroform/
methanol (100:1) as eluent. The product was isolated as yellow
solid (0.110 g, 0.19 mmol, 50%). Mp¼106.3e107.8 ꢂC. 1H NMR
4.4.8. N-Benzyloxycarbonyl-L-phenylalanine (6-methoxy-1-
methylquinolin-2(1H)-thione-4-yl) methyl ester, 4f. The solid crude
was purified by column chromatography using chloroform/meth-
anol (100:1) as eluent. The product was isolated as yellow solid
(0.060 g, 0.12 mmol, 78%). Mp¼158.2e160.0 ꢂC. 1H NMR (400 MHz,
(400 MHz, CDCl3):
d
¼3.12 (2H, t, J 7.2 Hz,
bCH2), 3.94 (3H, s,
OCH3), 4.27 (3H, s, NCH3), 4.70e4.75 (1H, m,
aH), 5.09 (2H, d, J
CDCl3):
NCH3), 4.70e4.75 (1H, m,
d
¼3.13 (2H, t, J 6.8 Hz,
b
CH2), 3.86 (3H, s, OCH3), 4.32 (3H, s,
2.4 Hz, CH2 Z), 5.24 (2H, d, J 12.4 Hz, NH), 5.29 (1H, s, CH2), 6.93
(1H, dd, J 8.8 and 2.0 Hz, H6), 6.97 (1H, d, J 2.0 Hz, H8), 7.08 (2H,
dd, J 8.0 and 2.0 Hz, H20 and H60 Phe), 7.15e7.36 (9H, m, 4ꢁPheH
Phe and 5ꢁPheH Z), 7.50 (1H, d, J 9.2 Hz, H5), 7.55 (1H, s, H3). 13C
a
H), 5.08 (2H, d, J 4.0 Hz, CH2 Z), 5.24 (1H,
d, J 8.0 Hz, NH), 5.30 (2H, s, CH2), 7.03 (1H, d, J 2.4 Hz, H5), 7.06 (2H,
dd, J 8.0 and 2.4 Hz, H20 and H60), 7.20e7.35 (9H, m, H7, 3ꢁPheH
Phe and 5ꢁPheH Z), 7.55 (1H, d, J 9.2 Hz, H8), 7.70 (1H, s, H3). 13C
NMR (100.6 MHz, CDCl3):
d
¼37.98 (NCH3), 38.02 (
bCH2), 54.89
NMR (100.6 MHz, CDCl3):
(aC), 55.72 (OCH3), 62.94 (CH2), 67.06 (CH2 Z), 105.92 (C5), 117.67
d
¼38.02 (
bCH2), 38.04 (NCH3), 54.95
(aC), 55.68 (OCH3), 62.96 (CH2), 66.97 (CH2 Z), 100.06 (C8), 112.23
(C6), 116.35 (C4a), 125.95 (C5), 127.12 (C40), 127.97 (C200), 128.07
(C600), 128.24 (C300, C400 and C-500), 128.57 (C30 and C50), 129.06 (C20
and C60), 130.12 (C3), 134.81 (C4), 135.24 (C10), 136.01 (C100),
142.42 (C8a), 155.55 (C]O urethane), 161.94 (C7), 171.05 (C]O
(C8), 120.22 (C7), 123.39 (C4a), 127.23 (C40), 128.05 (C200), 128.16
(C600), 128.47 (C300, C400 and C500), 128.63 (C30 and C50), 129.07 (C20
and C60), 133.32 (C3), 134.24 (C4), 135.17 (C10), 135.90 (C8a), 136.04
(C100), 155.60 (C]O urethane), 156.07 (C6), 171.19 (C]O ester),
ester), 184.11 (C2). FTIR (KBr 1%, cmꢀ1):
n
¼3360, 3060, 3029, 2934,
181.88 (C2). FTIR (KBr 1%, cmꢀ1):
1615, 1565, 1521, 1471, 1454, 1342, 1328, 1255, 1212, 1187, 1170, 1156,
1102, 1066, 1028, 875, 842, 813, 787, 750, 733, 699, 666. UV/vis
n
¼3346, 3032, 2944, 1761, 1687,
1739, 1699, 1615, 1552, 1454, 1415, 1348, 1324, 1290, 1231, 1108,
1060, 891, 750, 697. UV/vis (ethanol, nm): lmax (log
3
)¼385 (4.31).
MS m/z (ESI, %): 517 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for
(ethanol, nm): lmax (log
3
)¼401 (4.13). MS m/z (ESI, %): 517
([MþH]þ, 100). HRMS: m/z (ESI) calcd for C29H29N2O5S 517.17917,
C29H29N2O5S 517.17917, found 517.17747.
found 517.17775.
4.4.6. N-Benzyloxycarbonyl-
L
-phenylalanine
(1-methylquinolin-
2(1H)-thione-4-yl) methyl ester, 4d. The solid crude was purified by
column chromatography using chloroform/methanol (100:1) as
eluent. The product was isolated as yellow solid (0.103 g,
0.21 mmol, 80%). Mp¼169.5e171.2 ꢂC. 1H NMR (400 MHz, CDCl3):
4.4.9. N-Benzyloxycarbonyl-L-alanine (6-methoxy-1-methylquinolin-
2(1H)-thione-4-yl) methyl ester, 4g. The solid crude was purified by
column chromatography using chloroform/methanol (100:1) as
eluent. The product was isolated as
0.09 mmol, 30%). 1H NMR (300 MHz, CDCl3):
CH3), 3.89 (3H, s, OCH3), 4.32 (3H, s, NCH3), 4.48 (1H, br s,
a
yellow oil (0.042 g,
¼1.47 (3H, d, J 6.9 Hz,
H), 5.11
d¼3.12 (2H, t, J 5.6 Hz,
bCH2), 4.29 (3H, s, NCH3), 4.74 (1H, m, aH),
d
5.10 (2H, s, CH2 Z), 5.29 (3H, m, NH and CH2), 7.08 (2H, dd, J 8.0 and
1.6 Hz, H20 and H60), 7.28 (9H, m, H6, 3ꢁPheH Phe and 5ꢁPheH Z),
7.59 (2H, d, J 8 Hz, H8 and H5), 7.66 (2H, m, H3 and H7). 13C NMR
b
a
(2H, d, J 3.9 Hz, CH2 Z), 5.35 (3H, m, CH2 and NH), 7.03 (1H, d, J
2.4 Hz, H5), 7.27 (1H, dd, J 7.5 and 2.4 Hz, H7), 7.34e7.38 (5H, m,
5ꢁPheH Z), 7.57 (1H, d, J 8.2 Hz, H8), 7.74 (1H, s, H3). 13C NMR
(100.6 MHz, CDCl3):
d
¼37.90 (CH3), 38.03 (
bCH2), 53.37 (aC), 54.89
(CH2), 66.97 (CH2 Z), 116.09 (C8), 122.10 (C4a), 124.17 (C6), 124.47
(C5), 127.12 (C40), 127.98 (C200), 128.07 (C600), 128.39 (C300, C400 and
C500), 128.56 (C30 and C50), 129.04 (C20 and C60), 131.22 (C7), 132.60
(C3), 134.57 (C4), 135.21 (C10), 136.00 (C100), 140.75 (C8a), 155.54
(C]O urethane), 171.06 (C]O ester), 183.97 (C2). FTIR (KBr 1%,
(75.4 MHz, CDCl3):
d
¼18.42 (
bCH3), 38.27 (NCH3), 49.74 (aC), 55.75
(OCH3), 62.93 (CH2), 66.92 (CH2 Z), 105.77 (C5), 117.77 (C8), 120.25
(C7), 123.36 (C4a), 128.08 (C20), 128.11 (C60), 128.17 (C30, C40 and
C50), 132.88 (C3), 134.54 (C4), 135.90 (C8a), 136.09 (C10), 155.58 (C]
O urethane), 156.11 (C6), 172.53 (C]O ester) 181.86 (C2). FTIR (KBr
cmꢀ1):
n
¼3320, 3058, 3035, 3020, 2936, 1743, 1696, 1622, 1605,
1%, cmꢀ1):
n
¼3311, 2938, 1714, 1650, 1620, 1572, 1534, 1476, 1434,
1561, 1532, 1495, 1453, 1363, 1344, 1301, 1267, 1236, 1211, 1175, 1161,
1092, 1075, 1057, 908, 883, 790, 751, 743, 704, 666, 512. UV/vis
1381, 1336, 1248, 1212, 1174, 1117, 1070, 1039, 916, 822, 752, 699,
664. UV/vis (ethanol, nm): lmax (log
3
)¼395 (3.69). MS m/z (ESI, %):
(ethanol, nm): lmax (log
3
)¼384 (4.19). MS m/z (ESI, %): 487
441 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for C23H25N2O5S
([MþH]þ, 100). HRMS: m/z (ESI) calcd for C28H27N2O4S 487.16860,
441.14855, found 441.14823.
found 487.16693.
4.4.10. 1-(6-Methoxy-1-methylquinolin-2(1H)-thione-4-yl)methyl-5-
methyl-2-(N-benzyloxycarbonyl)aminopentanedioate, 4h. The solid
crude was purified by column chromatography using chloroform/
methanol (100:1) as eluent. The product was isolated as a yellow
oil (0.045 g, 0.09 mmol, 52%). 1H NMR (400 MHz, CDCl3):
4.4.7. N-Benzyloxycarbonyl-L-phenylalanine (1,6-dimethylquinolin-
2(1H)-thione-4-yl) methyl ester, 4e. The solid crude was purified by
column chromatography using chloroform/methanol (100:1) as
eluent. The product was isolated as yellow solid (0.095 g,
0.19 mmol, 55%). Mp¼160.3e162.1 ꢂC. 1H NMR (400 MHz, CDCl3):
d
¼2.00e2.29 (2H, m,
b
CH2), 2.37e2.49 (2H, m,
g
CH2), 3.66 (3H, s,
d
¼2.46 (3H, s, CH3), 3.14 (2H, t, J 6.4 Hz,
bCH2), 4.32 (3H, s, NCH3),
OCH3), 3.70 (3H, s, NCH3), 3.88 (3H, s, OCH3), 4.49 (1H, br s,
aH),
4.72e4.77 (1H, m, aH), 5.10 (2H, d, J 4.0 HZ, CH2), 5.30e5.33 (3H, m,
NH and CH2), 7.08 (2H, dd, J 8.0 and 1.6 Hz, H20 and H60), 7.18e7.53
(11H, m, H5, H7, H8, 3ꢁPheH Phe and 5ꢁPheH Z), 7.68 (1H, s, H3).
5.10 (2H, d, J 4.2 Hz, CH2 Z), 5.34 (2H, s, CH2), 5.46 (1H, d, J 7.9 Hz,
NH), 7.08 (1H, d, J 2.4 Hz, H5), 7.18 (1H, dd, J 7.9 and 2.1 Hz, H7),
7.25e7.35 (5H, m, 5ꢁPheH Z), 7.56 (1H, d, J 7.9 Hz, H8), 7.74 (1H, s,
13C NMR (100.6 MHz, CDCl3):
d
¼20.93 (CH3), 38.04 (
bCH2), 38.10

7900
A.S.C. Fonseca et al. / Tetrahedron 68 (2012) 7892e7900
H3). 13C NMR (100.6 Hz, CDCl3):
29.86 (gCH2), 51.89 (OCH3), 53.54 (aC), 55.76 (OCH3), 63.25 (CH2),
d
¼27.16 (
b
CH2), 29.56 (NCH3),
References and notes
1. (a) Mayer, G.; Heckel, A. Angew. Chem., Int. Ed. 2006, 45, 4900e4921; (b) Bochet,
C. G. J. Chem. Soc., Perkin Trans. 1 2002, 125e142.
2. (a) Bochet, C. G. Angew. Chem., Int. Ed. 2001, 40, 2071e2073; (b) Blanc, A.; Bo-
67.17 (CH2 Z), 105.86 (C5), 118.74 (C8), 120.33 (C7), 122.86 (C4a),
128.21 (C20), 128.52 (C60), 128.64 (C30, C40 and C50), 133.29 (C3),
134.32 (C4), 135.97 (C8a), 136.03 (C10), 155.91 (C]O urethane),
156.16 (C6), 173.02 (C]O(Flu)) 181.93 (C2). FTIR (KBr 1%, cmꢀ1):
chet, C. G. J. Org. Chem. 2002, 67, 5567e5577.
3. (a) Yu, H.; Li, J.; Wu, D.; Qiu, Z.; Zhang, Y. Chem. Soc. Rev. 2010, 39, 464e473; (b)
Lee, H.-M.; Larson, D. R.; Lawrence, D. S. ACS Chem. Biol. 2009, 4, 409e427; (c)
Deiters, A. Curr. Opin. Chem. Biol. 2009, 13, 678e686; (d) Ellis-Davies, G. C. R.
Chem. Rev. 2008, 108, 1603e1613; (e) Young, D. D.; Deiters, A. Org. Biomol. Chem.
2007, 5, 999e1005; (f) Loudwig, S.; Specht, A.; Goeldner, M. Actual. Chim. 2002,
7e12; (g) Shigeri, Y.; Tatsu, Y.; Yumoto, N. Pharmacol. Ther. 2001, 91, 85e92; (h)
Dorman, G.; Prestwich, G. D. Trends Biotechnol. 2000, 18, 64e77; (i) Curley, K.;
Lawrence, D. S. Pharmacol. Ther. 1999, 82, 347e354.
n
¼3349, 2928, 1730, 1649, 1621, 1572, 1510, 1459, 1433, 1380, 1245,
1178, 1040, 813, 699. UV/vis (ethanol, nm): lmax (log
3
)¼393 (3.49).
MS m/z (ESI, %): 513 ([MþH]þ, 100). HRMS: m/z (ESI) calcd for
C26H29N2O7S 513.16967, found 513.17725.
4. (a) Hagen, V.; Dekowski, B.; Nache, V.; Schmidt, R.; Geissler, D.; Lorenz, D.;
Eichhorst, I.; Keller, S.; Kaneko, H.; Benndorf, K.; Wiesner, B. Angew. Chem., Int.
Ed. 2005, 44, 7887e7891; (b) Geissler, D.; Antonenko, Y. N.; Schmidt, R.; Keller,
S.; Krylova, O. O.; Wiesner, B.; Bending, J.; Pohl, P.; Hagen, V. Angew. Chem., Int.
Ed. 2005, 44, 1195e1198; (c) Furuta, T.; Takeuchi, H.; Isozaki, M.; Takahashi, Y.;
Kanehara, M.; Sugimoto, M.; Watanabe, T.; Noguchi, K.; Dore, T. M.; Kurahashi,
T.; Iwamura, M.; Tsien, R. Y. ChemBioChem 2004, 5, 1119e1128; (d) Shembekar,
V. R.; Chen, Y.; Carpenter, B. K.; Hess, G. P. Biochemistry 2007, 46, 5479e5484;
(e) Gilbert, D.; Funk, K.; Dekowski, B.; Lechler, R.; Keller, S.; Mohrlen, F.; Frings,
S.; Hagen, V. ChemBioChem 2007, 8, 89e97.
5. Fonseca, A. S. C.; Gonc¸ alves, M. S. T.; Costa, S. P. G. Amino Acids 2010, 39,
699e712.
6. (a) Epling, G. A.; Walker, M. E. Tetrahedron Lett. 1982, 23, 3843e3846; (b) Epling,
G. A.; Provatas, A. A. Chem. Commun. 2002, 1036e1037.
4.5. General photolysis procedure
A 1ꢁ10ꢀ4 M methanol/HEPES buffer (80:20) solution of conju-
gates 3b,c, 3g,h and 4 (5 mL) was placed in a quartz tube and ir-
radiated in a Rayonet RPR-100 reactor at the desired wavelength.
The lamps used for irradiation were of 254, 300, 350 and
419ꢃ10 nm. HEPES buffer solution was prepared in distilled water
with HEPES (4-(2-hydroxyethyl)-1-piperazine ethanesulfonic acid)
(10 mM), sodium chloride (120 mM), potassium chloride (3 mM),
calcium chloride (1 mM) and magnesium chloride (1 mM) and the
pH adjusted to 7.2 with 1 M sodium hydroxide.
€
7. (a) Fedoryak, O. D.; Dore, T. M. Org. Lett. 2002, 4, 3419e3422; (b) Davis, M. J.;
Kragor, C. H.; Reddie, K. G.; Wilson, H. C.; Zhu, Y.; Dore, T. M. J. Org. Chem. 2009,
74, 1721e1729.
Aliquots of 100 mL were taken at regular intervals and analysed
8. (a) Piloto, A. M.; Fonseca, A. S. C.; Costa, S. P. G.; Gonc¸ alves, M. S. T. Tetrahedron
2006, 62, 9258e9267; (b) Fonseca, A. S. C.; Gonc¸ alves, M. S. T.; Costa, S. P. G.
Tetrahedron 2007, 63, 1353e1359; (c) Fernandes, M. J. G.; Gonc¸ alves, M. S. T.;
Costa, S. P. G. Tetrahedron 2008, 64, 3032e3038; (d) Soares, A. M. S.; Costa, S. P.
G.; Gonc¸ alves, M. S. T. Amino Acids 2010, 39, 121e133; (e) Soares, A. M. S.; Costa,
S. P. G.; Gonc¸ alves, M. S. T. Tetrahedron 2010, 66, 8189e8195; (f) Piloto, A. M.;
Soares, A. M. S.; Costa, S. P. G.; Gonc¸ alves, M. S. T. Amino Acids 2012, 42,
2275e2282; (g) Soares, A. M. S.; Piloto, A. M.; Hungerford, G.; Costa, S. P. G.;
Gonc¸ alves, M. S. T. Eur. J. Org. Chem. 2012, 922e930.
by RP-HPLC. The eluent was acetonitrile/water (3:1) at a flow rate of
0.8 mL/min, for all compounds, except for compounds 4a,b (1.0 mL/
min) and 3g,h, 4g,h (0.6 mL/min), previously filtered through
a Millipore, type HN 0.45 mm filter and degassed by ultra-sound for
30 min. The chromatograms were traced by detecting UV absorp-
tion at the wavelength of maximum absorption for each conjugate
(retention time: 3b, 6.5; 3c, 5.2; 3g, 4.7; 3h, 4.8; 4a, 9.0; 4b, 7.3; 4c,
7.0; 4d, 6.8; 4e, 8.0; 4f, 6.7; 4g, 5.6; 4h, 5.9 min).
9. Coyle, J. D. Tetrahedron 1985, 41, 5393e5425.
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12. Jesberger, M.; Davis, T. P.; Barner, L. Synthesis 2003, 13, 1929e1958.
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Acknowledgements
ˇ
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14. Morris, J. V.; Mahaney, M. A.; Huber, J. R. J. Phys. Chem. 1976, 80, 969e974.
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Taylor and Francis: Boca Raton, FL, 2006.
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Thanks are due to the Fundac¸ ao para a Ciencia e Tecnologia (FCT,
Portugal) for financial support to the NMR Portuguese Network
(PTNMR, Bruker Avance III 400-Univ. Minho), FCT and FEDER (Eu-
ropean Fund for Regional Development)-COMPETE-QREN-EU for
financial support to the Research Centre, CQ/UM [PEst-C/QUI/
UI0686/2011 (FCOMP-01-0124-FEDER-022716)] and project PTDC/
QUI/69607/2006 (FCOMP-01-0124-FEDER-007449). A Ph.D. grant
to A.S.C.F. (SFRH/BD/32664/2006) is also acknowledged.
ꢀ
17. Fernandes, M. J. G.; Gonc¸ alves, M. S. T.; Costa, S. P. G. Tetrahedron 2007, 63,
10133e10139.
ꢀ
18. Muller, C.; Even, P.; Viriot, M.-L.; Carre, M.-C. Helv. Chim. Acta 2001, 84,
3735e3741.
19. Bochet, C. G. Tetrahedron Lett. 2000, 41, 6341e6346.