A new synthesis of dehydroluciferin [2-(6′-hydroxy-2′- benzothiazolyl)-thiazole-4-carboxylic acid] from 1,4-benzoquinone

Article abstract of DOI:10.1016/j.tet.2013.05.019

A new synthesis of dehydroluciferin [2-(6′-hydroxy-2′- benzothiazolyl)-thiazole-4-carboxylic acid], the oxidative product of luciferin, has been realized starting from 1,4-benzoquinone. Reaction of this compound with l-cysteine ethyl ester, followed by an oxidation-cyclization step afforded 2-carbethoxy-6-hydroxybenzothiazole that was in situ hydrolyzed and decarboxylated to 6-hydroxybenzothiazole. The tert-butyl(dimethyl)silyl ether of this key intermediate was subjected to α-lithiation followed by formylation with DMF, and the resulting aldehyde condensed with l-cysteine ethyl ester. Dehydrogenation of the intermediate thiazolidine followed by deprotection afforded dehydroluciferin in 35% overall yield from 1,4-benzoquinone (69% from 6-hydroxybenzothiazole).

Full text of DOI:10.1016/j.tet.2013.05.019

Tetrahedron 69 (2013) 5893e5897
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Tetrahedron
journal homepage: www.elsevier.com/locate/tet
A new synthesis of dehydroluciferin [2-(60-hydroxy-20-
benzothiazolyl)-thiazole-4-carboxylic acid] from 1,4-benzoquinone
,
,
Pierangela Ciuffreda a, Silvana Casati a, Giuseppe Meroni b y, Enzo Santaniello b
*
a
ꢀ
Dipartimento di Scienze Biomediche e Cliniche “L. Sacco”, Universita degli Studi di Milano, Via G. B. Grassi, 74, 20157 Milano, Italy
Dipartimento di Scienze della Salute, Universita degli Studi di Milano, Via A. di Rudinì, 8, 20142 Milano, Italy
b
ꢀ
a r t i c l e i n f o
a b s t r a c t
A new synthesis of dehydroluciferin [2-(60-hydroxy-20-benzothiazolyl)-thiazole-4-carboxylic acid], the
oxidative product of luciferin, has been realized starting from 1,4-benzoquinone. Reaction of this com-
Article history:
Received 28 February 2013
Received in revised form 24 April 2013
Accepted 9 May 2013
pound with
hydroxybenzothiazole that was in situ hydrolyzed and decarboxylated to 6-hydroxybenzothiazole. The
tert-butyl(dimethyl)silyl ether of this key intermediate was subjected to -lithiation followed by for-
mylation with DMF, and the resulting aldehyde condensed with -cysteine ethyl ester. Dehydrogenation
L-cysteine ethyl ester, followed by an oxidationecyclization step afforded 2-carbethoxy-6-
Available online 14 May 2013
a
L
Keywords:
of the intermediate thiazolidine followed by deprotection afforded dehydroluciferin in 35% overall yield
from 1,4-benzoquinone (69% from 6-hydroxybenzothiazole).
Dehydroluciferin
Firefly luciferase
Natural products
Heterocycles
Ó 2013 Elsevier Ltd. All rights reserved.
Total synthesis
1. Introduction
is at present the preferred enzyme for in vivo optical imaging of
small animals.6 A few limits such as light absorption and scattering
The luciferase from the North American firefly Photinus pyralis
(PpyLuc; EC 1.13.12.7) catalyzes a bioluminescent reaction from
luciferin
{(S)-2-[60-hydroxy-20-benzothiazolyl]-2-thiazoline-4-
carboxylic acid; 1}, ATP, and oxygen. This reaction involves the
formation of an enzyme-bound adenylyl intermediate ( -luciferyl
by organs or the high dose of the reporter probe required for
a sufficient emission7 could be overcome by other imaging mo-
dalities, such as single photon emission computed tomography
(SPECT) and positron emission tomography (PET), that use radio-
nuclides for imaging of reporter genes.8 In this respect, only two
D-
D
adenylate; 2) that is oxidized by molecular oxygen with release of
pyrophosphate (PPi) and CO2. The intermediate, unstable dioxeta-
none (3) liberates oxyluciferin {2-[60-hydroxy-20-benzothiazolyl]-
4-hydroxythiazole, 4} in an excited state that is stabilized by the
emission of a photon (Scheme 1).1,2 About 80% of the enzyme-
examples of radioactive isotopomers of
D
-luciferin (1) are available,
i.e., 60-[11C-methyl]- -luciferin9 and 70-[123I]iodo-
D
D
-luciferin,10 but
these compounds were unable to locate the tumour and showed
poor cell uptake.9,11
We have recently undertaken a project12 aimed at developing
compounds suitably labelled with radioisotopes for positron
emission tomography (PET) in vivo imaging of a transgenic mouse
that expresses a luciferase reporter gene under the control of ac-
tivated oestrogen receptors.13 We have considered the structure of
PpyLuc inhibitors as possible targets for the introduction of the
positron emitting isotope 18F.14 Within this framework, we have
recently prepared a few 2,6-disubstituted benzothiazoles15 that
bound intermediate -luciferyl adenylate (2) is oxidized to oxy-
D
luciferin 4, but another enzyme pathway gives rise to a side re-
action in which the adenylate 2 is oxidized to the dehydroluciferyl
adenylate (5) with production of H2O2. Pyrophosphorolysis of the
adenylate 5 may occur with formation of dehydroluciferin [2-(60-
hydroxy-20-benzothiazolyl)-thiazole-4-carboxylic acid, 6].3
PpyLuc is a well characterized enzyme that finds a large number
of biotechnological applications4 and has been used, for example, as
an indicator of cell proliferation, gene delivery or gene expression
in cell culture and in living animals as a transgenic marker.5 PpyLuc
have shown values of IC50 (8.8e45.2
other compounds of similar structure proposed as competitive in-
hibitors of PpyLuc (Ki ranging from 25 to 58
M).16
In this context, it is noteworthy that dehydroluciferin (6) itself is
mM) that are in the range of
m
a good inhibitor of PpyLuc (Ki 1 m
M)17 and that the most potent
* Corresponding author. Tel.: þ39 (0)2 50323270; fax: þ39 (0)2 50323245;
inhibitor of luciferase, DLSA [50-O-[(N-dehydroluciferyl)-sulfa-
moyl]-adenosine, 7a (Ki¼340 nM)],18 is the sulfamoyl analogue of
dehydroluciferyl adenylate (5) (Fig. 1).
e-mail address: enzo.santaniello@unimi.it (E. Santaniello).
y
Present address: Drug Discovery Unit, Department of Experimental Oncology,
European Institute of Oncology (IEO), Via Adamello, 16, 20139 Milan, Italy.
0040-4020/$ e see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2013.05.019

5894
P. Ciuffreda et al. / Tetrahedron 69 (2013) 5893e5897
ATP/Mg2+
PPi/Mg2+
O2
N
S
S
N
N
S
S
N
N
S
S
N
O
O
O
O
O P Ade
HO
HO
HO
HO
CO2H
CO2H
AMP
O
O
3
D
-luciferin (1)
D
-luciferyl-AMP (2)
O2
CO2
H2O2
PPi/Mg2+
ATP/Mg2+
N
S
S
N
O
N
S
S
N
N
S
S
N
photon +
HO
O P Ade
OH
HO
O
O
oxyluciferin (4)
dehydroluciferin (6)
dehydroluciferyl-AMP (5)
Scheme 1. PpyLuc-catalyzed reactions.
compounds.1,22 The original synthetic approach is reported in
Scheme 3 and has been discussed in a recent review that summa-
NH2
N
N
X
N
N
O O
N
S
S
N
rizes syntheses of
-luciferin (1), derivatives and analogues.23 The
D
S
NH
O
HO
O
above preparations were carried out only on milligram scale and
generally proceeded with low yields of the final products, since the
intermediates 2-cyano-6-hydroxybenzothiazole (8a) and the 6-
methoxy analogue 8b were synthesized from para-anisidine with
overall yields of 6 and 10%, respectively. The reaction of the nitrile
O
OH OH
b. X = group containing 18
7a. X=H, DLSA
F
Fig. 1. 50-O-[(N-Dehydroluciferyl)-sulfamoyl]-adenosine (DLSA, 7a).
8a with D-cysteine affords D-luciferin (1) that can be oxidized to
dehydroluciferin (6) in a basic solution with either potassium fer-
ricyanide or oxygen (61% from 8a). Alternatively, dehydroluciferin
(6) was prepared from 2-cyano-6-methoxybenzothiazole (8b) (16%
yield).
With the aim of preparing a 18F-labelled DLSA (compound 7b),
a group X should be placed in DLSA 7a or in dehydroluciferin (6), its
synthetic precursor. The group X should be transformed into
a leaving group suitable to a nucleophilic substitution by 18F-fluo-
ride, thus allowing the introduction of the 18F positron emitting
isotope (compound 7b). In a radiosynthesis, this step usually is
accomplished following the experimental procedure traditionally
N
S
S
N
HO
CO2H
1
a
established for the production of 2-[18F]-fluoro-2-deoxy-
18F-FDG).19
For this project, two synthetic approaches to derivatives of
D-glucose
N
S
CN
b
c, d, e
(
RO
N
S
S
N
8a R=H
b R=CH3
dehydroluciferin substituted by a group X in the phenyl ring
(compound 11, Scheme 2) were considered. The group X could be
introduced by an electrophilic substitution at the phenol moiety of
a suitable 2,6-disubstituted benzothiazole (8) or dehydroluciferin
(6) itself. Alternatively, the group X or a related moiety should be
present in a synthetic precursor such as the substituted 1,4-
benzoquinone 10.
HO
CO2H
6
Scheme 3. Synthesis of dehydroluciferin (6) from 2-cyano-6-hydroxybenzothiazole
(8a) or from 2-cyano-6-methoxybenzotiazole (8b). Reagents and conditions: (a) D-
cystine/NH3, Na, H2O/MeOH 94%; (b) NaOH, O2, 65%; (c) H2S/Et3N, Py, rt, 3 h, 95%; (d)
methylbromopyruvate, MeOH, rt, 22 h and reflux 17 h, 88%; (e) HBr reflux, 1.5 h, 20%.
The initial steps of the synthesis of dehydroluciferin (6) from
1,4-benzoquinone (12) were carried out essentially as de-
scribed.20,21,24 Specifically, 1,4-benzoquinone 12 was converted into
2-carbethoxy-6-hydroxybenzothiazole 13a that was not purified,
but directly hydrolyzed in a basic medium. Acidification and de-
N
R
S
HO
8
X
X
carboxylation of the unstable acid 13b proceeded according to
N
S
N
S
S
N
24
€
CO2Et
Lowik et al. Purified 6-hydroxybenzothiazole 14 has been ob-
HO
O
HO
CO2H
tained by this simplified one-pot reaction in 50% yield and consti-
tutes the starting material for our synthesis of dehydroluciferin (6)
(Scheme 4).
9
11
X
The hydroxyl group of 6-hydroxybenzothiazole 14 was pro-
tected as the corresponding silyl ether and compound 15 was
converted into the aldehyde 16 by reaction with butyllithium, fol-
lowed by formylation with DMF. This procedure has been used to
prepare heteroaromatic aldehydes regioselectively and in good
yields.25 For instance, the reaction has been recently applied to the
preparation of 6-dimethylaminobenzothiazole-2-carbaldehyde.26
The aldehyde 16 was obtained in 89% yield from compound 15
as confirmed by the 1H NMR spectrum of the crude material,27 and
O
10
Scheme 2. Synthetic approaches to a dehydroluciferin derivative 11.
In connection with our works on the synthesis of 2-substituted-
6-hydroxy (or 6-methoxy) benzothiazoles20 and
D
-luciferin (1)
starting from 1,4-benzoquinone,21 we have now developed a new
synthesis of dehydroluciferin (6) starting from 1,4-benzoquinone.
was directly reacted with L-cysteine methyl ester in refluxing
2. Results and discussion
hexane, smoothly affording the thiazolidine derivative 17 (84%).
This compound has been obtained as an inseparable 3:2 mixture of
C-2 epimers that show well-resolved signals in the 500 MHz 1H
NMR spectrum.
The synthesis of dehydroluciferin (6) has been described in the
studies on the structure of -luciferin (1) and structurally related
D

P. Ciuffreda et al. / Tetrahedron 69 (2013) 5893e5897
5895
broadband probes and deuterium lock. Chemical shifts (
d
) are given
O
N
S
N
S
N
S
a, b
HO
d
c
in parts per million (ppm) and were referenced using residual
signals of the solvent as internal standard (1H: CHCl3, 7.26 ppm;
DMSO, 2.49 ppm. 13C: CDCl3, 77.0 ppm; DMSO-d6, 39.5 ppm).
Coupling constants (J) are in hertz (Hz) and the experimental error
in the measured 1He1H coupling constants is ꢁ0.5 Hz. The splitting
pattern abbreviations are as follows: s, singlet; d, doublet; t, triplet;
q, quartet; m, multiplet, and br s, broad peak. For two-dimensional
experiments, Bruker microprograms using gradient selection (gs)
were applied. Infrared spectra were recorded using a Jasco FTIR
4100 spectrometer as a thin film on NaCl plates.
CO2R
TDMSO
HO
O
14
15
13a R=Et
b R=H
12
N
S
S
N
N
S
f
g
e
CHO
H
TDMSO
CO2Et
TDMSO
H
17
16
h, i
N
S
S
N
6
TDMSO
CO2Et
18
4.2. 6-Hydroxybenzothiazole (14)
Scheme 4. Synthesis of dehydroluciferin (6) from 1,4-benzoquinone (12). Reagents
and conditions: (a) CysCO2Et/MeOH; (b) K3Fe(CN)6/NaOH/i-PrOH; (c) NaOH (overall
yield of steps aec 50%); t-BuDSiCl/imidazole/THF, 98%; (e) BuLi/DMF/THF (not iso-
lated); (f) CysCO2Et/TEA/hexane (steps e and f 84%); (g) MnO2/toluene, 95%; (h) TBAF/
THF; (i) NaOH (steps h and i 88%).
The previously described procedures21,24 have been adapted to
a one-pot preparation that is described here in detail. A solution of
1,4-benzoquinone (12) (1.0 g, 9.2 mmol) in methanol (20 mL) was
slowly added to a solution of L-cysteine ethyl ester hydrochloride
Oxidation of the thiazolidine 17 to compound 18 was most
conveniently performed with activated manganese dioxide MnO2
in toluene in nearly quantitative yield. The doubly protected
dehydroluciferin (compound 18) was desilylated with tetrabuty-
lammonium fluoride (TBAF) and then hydrolyzed with 4 M NaOH to
give dehydroluciferin (6) (88% yield after the two deprotection
steps). Yields of dehydroluciferin (6) were 35% from 1,4-
benzoquinone (12) and 69% from 6-hydroxybenzothiazole (14).
(1.7 g, 9.2 mmol) in methanol (10 mL) with stirring and cooling in
an ice bath under nitrogen. After stirring at room temperature for
1.5 h, the reaction was monitored by TLC (petroleum ether/ethyl
acetate 6:4) and the solvent was evaporated under vacuum. Iso-
propanol was added (60 mL) and the solution was cooled in an ice
bath under nitrogen and stirring. An aqueous solution of 1 M
K3Fe(CN)6 (36 mL) and 4 M NaOH (2.5 mL) was slowly added and
stirring was continued at room temperature. At the end of the re-
action (1 h) after TLC (petroleum ether/ethyl acetate, 8:2), the solid
was removed by filtration under vacuum and the solution extracted
with ethyl acetate. A small part of this solution was evaporated to
dryness to get the crude product verifying the presence of desired
compound 2-carbethoxy-6-hydroxybenzothiazole 13a: 1H NMR
3. Conclusions
In summary, starting from easily available 1,4-benzoquinone
(12), a simple and convenient procedure has been developed to
synthesize dehydroluciferin (6) in good yields (35% overall yield
from benzoquinone). The new synthetic procedure here estab-
lished allows for the first time the preparation of grams of dehy-
droluciferin (6), and is versatile enough to prepare substituted
dehydroluciferins (compounds 11). In fact, depending on the
substituted 1,4-benzoquinone derivatives 10 (Scheme 3) either
commercially available or ad hoc synthesized, a wide variety of
groups X in the phenyl ring of dehydroluciferin (6) could be
introduced.
(CD3OD):
d
¼7.97 (d, J¼9.0 Hz, 1H, 4-H), 7.37 (d, J¼2.0 Hz, 1H, 7-H),
7.13 (dd, J¼9.0 and 2.0 Hz, 1H, 5-H), 4.50 (q, J¼6.9 Hz, 2H, CH2CH3),
1.46 (t, J¼6.9 Hz, 3H, CH2CH3) ppm.
The ethyl acetate extract was then washed with 1 M NaOH
(50 mL), the aqueous phase was acidified with 3 M HCl, and
extracted with ethyl acetate. Evaporation of the solvent at 60 ꢂC
caused complete decarboxylation of the unstable acid 13b. Purifi-
cation by column chromatography (DCM/acetone 95:5) gave title
compound 14 as a colourless solid that was recrystallized from
aqueous ethanol (0.7 g, 4.6 mmol, 50%). Mp 190e192 ꢂC (lit.24
184e185 ꢂC). Rf¼0.35 (petroleum ether/AcOEt 6:4). 1H NMR
This would open the access to 18F-labelled DLSA 7b for PET
in vivo imaging of the transgenic mouse that expresses a luciferase
reporter gene under the control of activated oestrogen receptors.13
(CD3OD):
d
¼8.98 (s, 1H, 2-H), 7.88 (d, J¼8.3 Hz, 1H, 4-H), 7.38 (d,
J¼2.1 Hz, 1H, 7-H), 7.06 (dd, J¼8.3 and 2.1 Hz, 1H, 5-H) ppm. 13C
NMR (CD3OD):
122.8 (4-CH), 115.9 (5-CH), 106.1 (7-CH) ppm.
d
¼164.8 (6-C), 155.9 (9-C), 152.1 (2-CH), 134.9 (8-C),
4. Experimental section
4.1. General
4.3. 6-{[tert-Butyl(dimethyl)silyl]oxy}-1,3-benzothiazole (15)
All of the reagents and solvents, analytically pure, were obtained
from SigmaeAldrich and used as such without further purification.
Column chromatography was performed on Silica Gel 60 (70e230
mesh) using the specified eluents. The progress of the reactions was
monitored by analytical thin-layer chromatography (TLC) on pre-
coated glass plates (silica gel 60 F254-plate-Merck, Darmstadt,
Germany) and the products were visualized by UV light.
Purity of all products (ꢀ99%) was verified by thin-layer chro-
matography and NMR measurements. Elemental analyses were
obtained for all intermediates and are within ꢁ0.4% of theoretical
values.
Imidazole (400 mg) and tert-butyldimethylsilyl chloride
(800 mg) were added to a solution of 14 (0.7 g, 4.6 mmol) in dry
tetrahydrofuran (15 mL) under stirring and nitrogen at room tem-
perature. The reaction was monitored by TLC (petroleum ether/
ethyl acetate, 8:2) until all the starting material was consumed
(approximately 18 h). A solution of ammonium chloride was added
and the mixture was extracted with ethyl acetate. Purification by
column chromatography (petroleum ether/ethyl acetate 8:2) gave
title compound 15 as a colourless oil (1.2 g, 4.5 mmol, 98%). Rf¼0.38
(petroleum ether/AcOEt 9:1). 1H NMR (CDCl3):
d¼8.85 (s, 1H, 2-H),
Melting points were determined with a Stuart Scientific SMP3
melting point apparatus.
7.99 (d, J¼9.0 Hz, 1H, 8-H), 7.38 (d, J¼2.0 Hz, 1H, 5-H), 7.05 (dd,
J¼9.0 and 2.0 Hz, 1H, 7-H), 1.03 (s, 9H, SiC(CH3)3), 0.32 (s, 6H,
1H NMR spectra were recorded at 298 K in CDCl3 (isotopic en-
richment 99.95%) or DMSO-d6 (isotopic enrichment 99.98%) solu-
tions at 300 K using a Bruker AVANCE 500 instrument (500.13 MHz
for 1H, 125.76 MHz for 13C) using 5 mm inverse detection
2ꢃSi(CH3)2) ppm. 13C NMR (CDCl3):
¼153.9 (6-C), 151.8 (2-CH),
d
148.3 (9-C), 134.9 (4-C), 123.9 (8-CH), 120.2 (7-CH), 111.7 (5-CH),
25.7 (3ꢃSieCH3),18.3 (SieC), ꢄ4.4 (2ꢃSieCH3) ppm; IR (NaCl, neat)
3063, 2930, 2857, 2711, 2360, 1898, 1553, 1445, 1390, 1268, 1186,

5896
P. Ciuffreda et al. / Tetrahedron 69 (2013) 5893e5897
1046, 947, 854, 815, 738, 686, 613 cmꢄ1; C13H19NOSSi (265.45): C
58.82, H 7.21, N 5.28, S 12.08; found C 58.89, H 7.16, N 5.20, S 12.14.
NMR (CDCl3):
d
¼8.28 (s, 1H, 5-H), 7.94 (d, J¼8.8 Hz, 1H, 40-H), 7.35
(d, J¼2.5 Hz, 1H, 70-H), 7.04 (dd, J¼8.8 and 2.5 Hz, 1H, 50-H), 4.46 (q,
J¼7.0 Hz, 2H, OCH2CH3), 1.44 (t, J¼7.0 Hz, 3H, OCH2CH3), 1.02 (s, 9H,
4.4. Ethyl 2-{60-[tert-butyl(dimethyl)silyl]oxy-20-benzothia-
zolyl}-thiazolidine-4-carboxylate (17)
SiC(CH3)3), 0.26 (s, 6H, 2ꢃSi(CH3)2) ppm. 13C NMR (CDCl3):
¼162.5
d
(2-C) 160.9 (CO2Et), 158.2 (60-C), 154.9 (20-C), 148.4 (90-C), 148.2 (4-
C), 137.2 (80-C), 129.4 (5-CH), 124.1 (40-CH), 121.0 (50-CH), 111.8 (70-
CH), 61.8 (CO2CH2CH3), 25.7 (3ꢃSieC(CH3)3), 18.3 (SieC), 14.4
(CO2CH2CH3), ꢄ4.3 (2ꢃSieCH3) ppm; IR (NaCl, neat) 3053, 2930,
2957, 2896, 2360, 1730, 1551, 1494, 1391, 1336, 1265, 1212, 1101,
1022, 905, 822, 782, 704, 646 cmꢄ1; C19H24N2O3S2Si (420.62): C
54.25, H 5.75, N 6.66, S 15.25; found C 54.32, H 5.66, N 6.69, S 15.18.
A solution of n-butyllithium (3 mL 1.6 M in hexane, 4.8 mmol)
was added dropwise to a stirred solution of 15 (1.2 g, 4.5 mmol) in
THF (27 mL) at ꢄ78 ꢂC at a rate that kept the reaction temperature
below ꢄ70 ꢂC. On completion of addition, the dark-red reaction
mixture was stirred for 30 min at ꢄ78 ꢂC and a solution of DMF
(0.35 mL, 4.5 mmol) in tetrahydrofuran (1 mL) was added. After
1.5 h, the reaction mixture was allowed to warm to room temper-
ature and was quenched with saturated aqueous NH4Cl (10 mL).
The layers were separated, and the aqueous layer was extracted
with Et2O. The combined organic extracts were dried (Na2SO4) and
concentrated in vacuo to give 6-{[tert-butyl(dimethyl)silyl]oxy}-
1,3-benzothiazol-2-carbaldehyde (16) as an oil, immediately used
for the next reaction. Rf¼0.62 (petroleum ether/AcOEt 9:1). 1H NMR
4.6. 2-(6’-Hydroxy-20-benzothiazolyl)-thiazole-4-carboxylic
acid (6)
To a solution of compound 18 (1.5 g, 3.6 mmol) in THF (30 mL)
a 1 M solution in THF of tetrabutylammonium fluoride (7 mL,
7 mmol) was added. After 15 min the reaction mixture was washed
with a solution of sodium bicarbonate, extracted with DCM, and
concentrated to a small volume. To the remained THF solution 4 M
sodium hydroxide was added (2 mL, 8 mmol) and the mixture was
stirred at room temperature for 3 h. At the completion of the re-
action, 3 M HCl (3 mL, 9 mmol) was added. After filtration and
washing with diethyl ether the product was recovered as a solid
(0.89 g, 3.2 mmol, 88%). Mp 315 ꢂC (lit.22 315e321 ꢂC dec). Rf¼0.13
(CDCl3):
d
¼10.1 (s, 1H, CHO), 8.10 (d, J¼8.3 Hz, 1H, 4-H), 7.37 (d,
J¼2.1 Hz, 1H, 7-H), 7.15 (dd, J¼8.3 and 2.1 Hz, 1H, 5-H), 1.03 (s, 9H,
SiC(CH3)3), 0.28 (s, 6H, 2ꢃSi(CH3)2) ppm. 13C NMR (CDCl3):
¼185.2
d
(CHO), 162.7 (2-CH), 156.7 (6-C), 148.7 (9-C), 138.3 (8-C), 126.7 (4-
CH), 122.1 (5-CH), 111.8 (7-CH), 25.7 (3ꢃSieC(CH3)3), 18.3 (SieC),
ꢄ4.34 (2ꢃSieCH3) ppm.
(DCM/MeOH 8:2). 1H NMR (DMSO-d6):
d
¼10.16 (s, 1H, CO2H), 8.64
A mixture of aldehyde 16, L-cysteine ethyl ester hydrochloride
(s, 1H, 5-H), 7.92 (d, J¼8.8 Hz, 1H, 40-H), 7.47 (d, J¼2.4 Hz, 1H, 70-H),
(920 mg, 5 mmol), and triethylamine (1.2 mL, 8.6 mmol) in hexane
(30 mL) was refluxed under stirring for 2 h. At the end of the re-
action after TLC (petroleum ether/ethyl acetate 8:2), the solid was
removed by filtration under vacuum and the solution evaporated to
provide 17 (3:2 mixture of C-2 epimers) as an oil (1.6 g, 3.8 mmol,
84%). Rf¼0.48 (petroleum ether/AcOEt 8:2). 1H NMR (CDCl3):
7.05 (dd, J¼8.8 and 2.4 Hz, 1H, 50-H), 10.16 (s, 1H, OH) ppm. 13C NMR
(DMSO-d6):
d
¼162.1 (CO2H), 161.8 (2-C) 157.5 (60-C), 157.0 (20-C),
148.8 (90-C), 146.8 (4-C), 137.2 (80-C), 131.6 (5-CH), 124.7 (40-CH),
117.6 (50-CH), 107.5 (70-CH) ppm.
d
¼(major epimer) 7.86 (d, J¼8.5 Hz, 1H, 40-H), 7.28 (d, J¼2.1 Hz, 1H,
Acknowledgements
70-H), 6.99 (dd, J¼8.5 and 2.1 Hz, 1H, 50-H), 5.87 (s, 1H, 5-H),
4.31e4.25 (m, 3H, 4-H and OCH2CH3), 3.44 (dd, J¼10.5 and 6.6 Hz,
1H, 5a-H), 3.17 ( dd, J¼10.5 and 9.4 Hz, 1H, 5b-H), 1.33 (t, J¼7.0 Hz,
3H, OCH2CH3), 1.00 (s, 9H, SiC(CH3)3), 0.22 (s, 6H, 2ꢃSi(CH3)2) ppm.
13C NMR (CDCl3): 171.3 (CO2Et), 170.4 (20-C), 153.6 (60-C), 148.7 (90-
C),136.4 (80-C),123.6 (40-CH),120.0 (50-CH),111.7 (70-CH), 67.0 (2-C),
64.7 (4-C), 62.0 (CO2CH2CH3), 38.0 (5-CH), 25.7 (3ꢃSieC(CH3)3),
18.3 (SieC), 14,2 (CO2CH2CH3), ꢄ4.3 (2ꢃSieCH3) ppm.
The research has been funded by the European Project EMIL
(European Molecular Imaging Laboratories, LSHC-Ct-2004-503569)
and the University of Milan.
References and notes
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ChemBioChem 2006, 7, 929.
d
¼(minor epimer) 7.82 (d, J¼8.5 Hz, 1H, 40-H), 7.25 (d, J¼2.1 Hz,
1H, 70-H), 6.97 (dd, J¼8.5 and 2.1 Hz, 1H, 50-H), 6.08 (s, 1H, 5-H),
4.31e4.25 (m, overlapped, 2H, OCH2CH3), 4.05 (dd, J¼9.4 and
6.6 Hz, 1H, 4-H), 3.45 (dd, J¼10.5 and 6.6 Hz, 1H, 5a-H), 3.15 (dd,
J¼10.5 and 9.4 Hz,1H, 5b-H),1.32 (t, J¼7.0 Hz, 3H, OCH2CH3), 0.99 (s,
9H, SiC(CH3)3), 0.21 (s, 6H, 2ꢃSi(CH3)2) ppm. 13C NMR (CDCl3):
172.8 (CO2Et), 166.8 (20-C), 153.9 (60-C), 147.8 (90-C), 136.3 (80-C),
123.9 (40-CH), 120.3 (50-CH), 111.7 (70-CH), 68.0 (2-C), 66.1 (4-C),
61.8 (CO2CH2CH3), 38.7 (5-CH), 25.7 (3ꢃSieC(CH3)3), 18.3 (SieC),
14,2 (CO2CH2CH3), ꢄ4.3 (2ꢃSieCH3) ppm; IR (NaCl, neat) 3942,
3330, 2930, 2983, 2931, 2858, 2341, 1737, 1554, 1454, 1391, 1265,
1096, 1030, 947, 896, 857, 825, 782, 704 cmꢄ1; C19H28N2O3S2Si
(424.13): C 53.74, H 6.65, N 6.60, S 15.10; found C 53.80, H 6,66, N
6.72, S 15.18.
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To a solution of compound 17 (1.6 g, 3.8 mmol) in toluene
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tration through a Celite pad under vacuum and the solvent evap-
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95%). Mp 138e140 ꢂC. Rf¼0.52 (petroleum ether/AcOEt 8:2). 1H
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ꢀ
ꢀ
26. Hrobarik, P.; Sigmundova, I.; Zahradnìk, P. Synthesis 2005, 600.
27. The aldehyde 16 could be obtained also by a traditional reductioneoxidation of
the ester 13a, but overall yields were lower than the ones described in this
paper.