Synthesis of functionalized analogs of pyochelin, a siderophore of Pseudomonas aeruginosa and Burkholderia cepacia

Article abstract of DOI:10.1016/j.tet.2005.12.012

Using an improved synthesis of pyochelin, a siderophore common to several pathogenic Pseudomonas species, three functionalized pyochelin analogs were efficiently synthesized starting from appropriate 2-hydroxybenzonitriles.

Full text of DOI:10.1016/j.tet.2005.12.012

Tetrahedron 62 (2006) 2247–2254
Synthesis of functionalized analogs of pyochelin, a siderophore
of Pseudomonas aeruginosa and Burkholderia cepacia
†
Freddy Rivault, Viviane Schons, Clemence Liebert, Alain Burger, Elias Sakr,
´
Mohamed A. Abdallah, Isabelle J. Schalk and Gaetan L. A. Mislin
´
*
¨
´ ´ ´ ´
Metaux et Microorganismes: Chimie, Biologie et Applications, Departement des Recepteurs et Proteines Membranaires, UMR7175 Institut
´
Gilbert Laustriat, Ecole Superieure de Biotechnologie de Strasbourg, Boulevard Sebastien Brant F-67400 Illkirch, France
´
´
Received 10 November 2005; accepted 5 December 2005
Available online 27 December 2005
Abstract—Using an improved synthesis of pyochelin, a siderophore common to several pathogenic Pseudomonas species, three
functionalized pyochelin analogs were efficiently synthesized starting from appropriate 2-hydroxybenzonitriles.
q 2005 Elsevier Ltd. All rights reserved.
1. Introduction
in biological fluids is usually estimated to be only 10K18
M
and P. aeruginosa and B. cepacia use siderophores to
compete for iron with the host.
Iron is a crucial element for aerobic life, unfortunately its
bioavailability is limited by the low solubility of iron(III) at
physiological pHs. To overcome this problem, microorgan-
isms have developed very efficient iron uptake systems
mediated by low molecular weight molecules called
siderophores.1 Under iron limited conditions, microorgan-
isms synthesize and excrete these molecules into the
extracellular medium in order to chelate iron(III). In
Gram-negative bacteria, the siderophore–iron(III) complex
is recognized and transported by a specific outer membrane
receptor. The uptake process is driven by the proton motive
force of the inner membrane via an inner membrane
complex composed of TonB, ExbB and ExbD proteins.2,3
Pseudomonas aeruginosa and Burkholderia cepacia, are
two opportunistic Gram-negative bacteria, causes of severe
and often lethal lung infections especially for cystic fibrosis
and aids affected patients.4 The low permeability of the
outer membrane of these bacteria and the increasing
antibiotic mediated selective pressure raised emerging
multiresistant strains. During infection, these bacteria are
in the host in an iron limited environment: higher eucaryotes
contain substantial amount of this metal but tightly
associated with transport and storage proteins and not freely
available for pathogens. Consequently, the level of free iron
One way to increase the efficiency of antibiotics targeting
these bacteria is the Trojan horse strategy where antibiotics
are coupled to siderophores and transported across the
bacterial membranes via the iron uptake pathways.5 Such an
approach has been already developed giving promising
results with the pyoverdine mediated iron uptake system.6 A
disavantage of the pyoverdine iron uptake pathway is that
each P. aeruginosa strain produces its own pyoverdine
7
along with a corresponding specific transporter. Very few
crossfeeding have been observed between these different
pyoverdines and their corresponding transporters. For the
pyochelin 1 pathway, this siderophore is produced by all
P. aeruginosa and B. cepacia strains.8 Therefore, pyochelin
is a more interesting candidate for such a prodrug strategy
and the bactericidial activity of pyochelin–antibiotic
conjugate should be more extended compared with
pyoverdine based prodrugs (Fig. 1).
OH
COOH
4"
2'
5
N
N
4' 2"
H
S
S
H
H
Pyochelin 1
Figure 1. Structure of pyochelin 1.
Keywords: Pseudomonas; Burkholderia; Cystic fibrosis; Siderophore;
Pyochelin; Antibiotic; Trojan horse conjugate; Iran uptake.
*
Corresponding author. Tel.: C33 3 9024 4727; fax: C33 3 9024 4829;
e-mail: mislin@esbs.u-strasbg.fr
In view of its use as a versatile antibiotic vector, pyochelin 1
was functionalized in order to be further connected to the
selected antibiotics. Taking into account the distance
† Present address: Laboratoire de Chimie Bioorganique, UMR 6001 CNRS,
´
Universite de Nice Sophia Antipolis, Parc Valrose, F-06108 Nice Cedex
2, France.
0040–4020/$ - see front matter q 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2005.12.012

2248
F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
between the heteroatoms involved in iron chelation and the
easiness of synthetic access, the fact that bulky C5-
substituted analogs of pyochelin were shown to transport
iron at reasonable rates,9 position 5 on the aromatic ring was
selected in a first approach to host an amine function. The
synthesis of such pyochelin analogs was thus planned using
the improved protocol adapted from a total synthesis of
pyochelin 1 previously published.10 According to this
procedure, pyochelin analogs 2, 3 and 4 were thus
synthesized starting, respectively, from amino substituted
2-hydroxybenzonitriles 5, 6 and 7 (Fig. 2).
the selective deprotection of phenol and gave carbamate 5
isolated in an overall 51% yield over the three last steps
(Scheme 1).
OH
CN
OH
CN
O
N
i,ii
iii
O2N
H2N
8
9 (83%)
10
iv, v
OH
TeocHN
CN
OH
OH
CN
5
(51% over the three last steps)
COOH
H
N
N
TeocHN
TeocHN
S
S
5
2
3
4
Scheme 1. Synthesis of functionalized 2-hydroxybenzonitrile 5. (i) HNO3,
H2SO4, 0 8C. (ii) NaOH, H2O/EtOH, 20 8C. (iii) H2, Pd/C 10%, EtOH,
20 8C. (iv) p-NO2(C6H4)OCOCH2CH2Si(CH3)3, pyridine, DMAP, CH2Cl2
reflux. (v) Na2CO3, H2O/acetone reflux.
OH
OH
CN
COOH
H
N
N
S
S
6
7
NHTeoc
NHTeoc
Cyanophenols 6 and 7 were both prepared starting from
commercially available 2-hydroxybenzonitrile 11. In a first
step, compound 11 was converted in excellent yield into the
iodinated derivative 12 using N-iodosuccinimide in
presence of HBF4 at low temperature.16 Iodo compound
12 was then coupled to Teoc protected propargylamine 1317
via a Sonogashira coupling reaction, using a catalytic
amount of Pd(PPh3)4, copper(I)iodide and DIPEA in
DMF.18 Thus, the expected substituted 2-hydroxybenzo-
nitrile 6 was isolated in excellent yield and catalytically
reduced in compound 7 with hydrogen over palladium
adsorbed on charcoal. In this step, an extended hydrogen-
ation cause the reduction of both the alkyne triple bond and
the nitrile group. However, when the course of the reaction
was carefully checked, the expected carbamate 7 was
isolated usually in 90% yield (Scheme 2).
OH
CN
OH
COOH
H
N
N
S
S
NHTeoc
NHTeoc
Figure 2. Retrosynthetic routes to functionalized pyochelins 2, 3 and 4.
In analog 2 the functionality is directly grafted on pyochelin
whereas analogs 3 and 4 offer some alternative in terms of
distance and flexibility towards the siderophore. All along
the synthesis, the amine was protected with a trimethyl-
silylethoxycarbonyl (Teoc) group,11 in order to facilitate
purifications and to avoid side reactions. This protecting
group was chosen for its good stability during the synthetic
process and for the easiness and mildness of the
deprotection conditions. In this paper, we describe the
synthesis of functionalized 2-hydroxybenzonitriles 5, 6 and
7 and their efficient conversions into the three unprece-
dented pyochelin analogs 2, 3 and 4.
OH
CN
OH
CN
OH
i
ii
I
CN
11
12 (91%)
6 (95%)
2. Results and discussion
NHTeoc
iii
The 2-hydroxybenzonitrile derivative 5 was prepared
starting from 1,2-benzisoxazole 8. The regioselective
nitration of 8 with a sulfonitric mixture,12 and the
subsequent hydrolysis in basic conditions of the oxazole
moiety led to the expected 5-nitro-2-benzonitrile 9 isolated
in 83% overall yield.13 The nitro function was then reduced
into an amine. Using TiCl3 in MeOH the expected amine 10
was isolated in an average yield after a tedious purification
protocol.14 A quite quantitative yield was obtained by
catalytic hydrogenation using palladium on charcoal: the
resulting amine 10 was quite unstable and had either to be
stored at K20 8C in the dark and under argon or
immediately used as such for the next step. Since, the
phenol function of compound 10 appeared to be more
reactive than the amine function, amine 10 was treated with
more than a two fold excess of p-nitrophenyl-trimethyl-
silylethyl-carbonate15 and yielded a Teoc bisprotected
compound. Further treatment of the crude mixture with
sodium carbonate in refluxing wet acetone induced
OH
CN
7 (90%)
NHTeoc
Scheme 2. Synthesis of functionalized 2-hydroxybenzonitriles 6 and 7.
(i) NIS, HBF4$ Et2O, CH3CN, K10 8C. (ii) 13, Pd(PPh3)4, CuI, DIPEA,
DMF, 20 8C. (iii) H2, Pd/C 10%, EtOH, 20 8C.
Having in hand the different 2-hydroxybenzonitriles, the
synthesis of the pyochelin analogs was then investigated. In
natural pyochelin, the absolute configurations are 40R and
400R while configuration at C200 is wobble and C200:can
readily be epimerized. During the synthesis of pyochelin,
partial racemization occurs at C40 while the absolute
configuration at C400 remains unaffected thus affording a
mixture of pyochelin a (40R,200R,400R), b (40R,200S,400R), and

F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
2249
neopyochelin c (40S,200R,400R) and d (40S,200S,400R)
The Weinreb amides 17, 18 and 19 were then reduced to the
corresponding aldehydes 20, 21 and 22 using LiAlH4.20 The
resulting aldehydes were very sensitive and thus were used
straightforward without further purification. Condensation
with N-methylcysteine,21 in presence of potassium acetate
in hydroethanolic medium furnished the expected functio-
nalized pyochelins 2, 3 and 4 in 65–75% yield over two
steps (Scheme 4).
(Fig. 3).19,10a,d
OH
OH
COOH
H
COOH
H
4"
4"
N
N
N
N
4' 2"
4' 2"
S
S
S
S
H
H
H
H
a (4'R,2"R,4"R)
b (4'R,2"S,4"R)
OH
OH
OH
OH
COOH
H
COOH
H
O
O
4"
4"
N
i
N
N
N
R
N
R
N
4' 2"
4' 2"
OMe
N
S
H
S
H
H
S
S
S
S
17 : R = R1
18 : R = R2
19 : R = R3
20 : R = R1
21 : R = R2
22 : R = R3
H
H
H
H
d (4'S,2"S,4"R)
c (4'S,2"R,4"R)
Figure 3. The four diastereoisomers of synthetic pyochelin.
ii
OH
COOH
H
On the other hand, pyochelin and neopyochelin were tested
in iron uptake experiments and proved to transport iron with
almost the same rate.9 Our data indicate clearly that the
configuration at C40 is not determining in the recognition by
the receptor FptA and in the iron transport process. We thus
chose to use the inexpensive (R)-cysteine, as starting
material for the condensation with the functionalized
2-hydroxybenzonitriles. The three hydroxybenzonitriles 5,
6 and 7 were converted into the thiazolines 14, 15 and 16
when treated with (R)-cysteine in a refluxing mixture of
MeOH and phosphate buffer (0.1 M, pH 6.4). Thiazolines
14, 15 and 16 were used without further purification to
synthesize the corresponding Weinreb amides 17, 18 and
19. For this purpose, we modified the procedure we
described previously by reacting 14, 15 and 16 with N,O-
dimethylhydroxylamine and N,N-dimethylaminopropyl-
ethylcarbodiimide hydrochloride (EDCI). Under these
conditions, hydroxamic esters 17, 18 and 19 were easily
isolated in good to excellent yield after two steps using a
very simple work-up. (Scheme 3).
N
N
R
R1
R2
R3
=
=
=
-NHTeoc
S
S
NHTeoc
NHTeoc
2 (72% over two steps) : R = R1
3 (75% over two steps) : R = R2
4 (65% over two steps) : R = R3
Scheme 4. Conversion of the hydroxamic esters 17, 18 and 19 into the
pyochelin analogs 2, 3 and 4. (i) LiAlH4, THF, K40 to K10 8C. (ii) (R)-N-
Methylcysteine$HCl, AcOK, EtOH/H2O, 20 8C.
The ratios between the four stereoisomers a, b, c and d of
each analog were the following: 2 (5/10/60/25), 3 (15/15/45/
25) and 4 (20/10/50/20). These diastereoisomeric ratios
were determined using 1H NMR and comparing integration
of the characteristic N-methyl signals. In addition, using
COSY and 1H–13C correlation it was then possible to assign
the chemical shifts of each the four diastereoisomers. The
relative configurations of the stereocenters were unambigu0-0
ously assigned by NOESY experiments. When pro0t0on H2
was saturated, a marked Overhauser effect with 0H4 00 proton
was observed for a (40R,200R,400R) and c (4 S,2 R,400R)
isomers0whereas no nuclear Overhauser effect was observed
for b (4 R,200S,400R) and d (40S,200S,400R).
OH
CN
OH
3. Conclusion
i
N
R
R
COOH
In summary, we report in this article very efficient synthetic
ways to three unprecedented functionalized pyochelins:
analog 2 was obtained in nine steps (30% overall yield)
starting from the commercially available 1,2-benzisoxazole
whereas analogs 3 and 4 were prepared starting from
2-hydroxybenzonitrile in six steps (40% overall yield) and
seven steps (45% overall yield), respectively. These analogs
will be used to build Trojan horse type antibiotic conjugates,
which will be tested on different pathogenic clinical strains
of Pseudomonas aeruginosa and Burkholderia cepacia.
S
H
5 : R = R1
14 : R = R1
15 : R = R2
16 : R = R3
6 : R = R2
7 : R = R3
ii
OH
N
O
R1 =
R2 =
-NHTeoc
R
OMe
N
S
H
17 (81% over two steps) : R = R1
18 (62% over two steps) : R = R2
19 (89% over two steps) : R = R3
NHTeoc
NHTeoc
4. Experimental
R3 =
4.1. General procedures
Scheme 3. Conversion of 2-hydroxybenzonitriles 5, 6 and 7 into the
hydroxamic esters 17, 18 and 19. (i) (R)-cysteine, phosphate buffer 0.1 M,
pH 6.4, MeOH, 60 8C. (ii) CH3NHOCH3$HCl, DIPEA, EDCI, CH2Cl2,
0–20 8C.
All the reactions were carried out in glassware under inert
argon atmosphere. Solvents used were of analytical grade

2250
F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
purity. DMF was distilled prior to use and was stored over
˚
activated 4 A molecular sieves. Pyridine and N,N-diisopropyl-
4.2.2. 2-Hydroxy-5-iodobenzonitrile (12). To a solution of
2-hydroxybenzonitrile 11 (1.00 g, 8.30 mmol) in dry
acetonitrile (7.5 mL), HBF4 (1.8 mL of 54% solution in
Et2O) was added dropwise at K20 8C. N-Iodosuccinimide
(2.00 g, 9.13 mmol) was then added portionwise as such a
rate that the reaction temperature does not exceed K10 8C.
The mixture was then warmed up to room temperature,
vigorously stirred during 4 h, treated with an aqueous
solution of NaHSO3 38% (20 mL) and extracted with
t-butylmethylether (2!30 mL). The collected organic
layers were then washed with brine (30 mL) and dried
over Na2SO4. After filtration, the orange-brown solid was
adsorbed on silica before being filtered on a silica gel
column (30 g of SiO2, CH2Cl2) leading to compound 12
(1.85 g, 7.55 mmol, yield: 91%) isolated as a white powder.
Mp 169–172 8C; IR (neat) 3165, 2245, 1589, 1489, 1400,
1354, 1294, 1263, 1231, 1170, 1118, 1072, 950, 892, 873,
852, 815, 755, 730, 682. 1H NMR (CD3COCD3, 300 MHz)
d 6.92 (d, JZ9.0 Hz, 1H), 7.79 (dd, JZ2.0, 9.0 Hz, 1H),
7.90 (d, JZ2.0 Hz, 1H), 10.08 (br s, 1H); 13C NMR
(CD3COCD3, 75 MHz) d 80.4, 103.2, 115.5, 119.3, 141.8,
143.8, 160.4. SM-EI m/z 245. Anal. Calcd for C7H4INO: C,
34.31; H, 1.65; N, 5.72. Found C, 34.35; H, 1.70; N, 5.68.
ethylamine (DIPEA) were distilled and stored over KOH.
Reactions were monitored by thin-layer chromatography
(TLC) using Merck precoated silica gel 60F254 (0.25 mm).
Column chromatographies were performed using Merck
kieselgel 60 (63–200 mm). Melting points were determined
with a Stuart Scientific Bibby SMP3 apparatus and were
uncorrected. IR spectra were scanned neat using a Perkin
Elmer Spectrum One spectrophotometer. NMR spectra were
1
recorded either on a Bruker Avance 300 (300 MHz for H
and 75 MHz for 13C), a Bruker Avance 400 (400 MHz for
1H and 100 MHz for 13C) or a Bruker Avance 500
(500 MHz for 1H and 125 MHz for 13C). Elemental analyses
were performed in the Service d’Analyses de l’Institut de
´
Chimie at the Universite Louis Pasteur of Strasbourg. Mass
spectra were recorded in the Service de Spectrometrie de
´
´
Masse de l’Institut de Chimie at the Universite Louis
Pasteur of Strasbourg and were measured after calibration in
ES-TOF experiments performed on a Bruker Daltonic
MicroTOF mass spectrometer.
4.2. Protocols, analytical and spectral data
4.2.3. Prop-2-ynyl-carbamic acid 2-trimethylsilylethyl-
ester (13). Solid p-nitrophenyl-trimethylsilylethylcarbo-
nate15 (1150 mg, 4.07 mmol) was added to a solution of
propargylamine (515 mL, 7.51 mmol), DIPEA (800 mL,
4.59 mmol), DMAP (10 mg) in CH2Cl2 (14 mL), and the
solution was refluxed for 48 h. The reaction mixture was
then cooled to room temperature, diluted with CH2Cl2 and
washed twice with saturated aqueous NaHCO3. The organic
layer was dried over Na2SO4, solvent was removed under
reduced pressure, and the residue was purified by flash
chromatography on silica gel (hexane/Et2O: 9:1 then
hexane/Et2O: 8:2), to give carbamic ester 13 as a colorless
oil (740 mg, 3.71 mmol, yield: 81%);17 IR (neat) 3312,
2954, 1697, 1517, 1424, 1349, 1324, 1248, 1178, 1139,
1043, 973, 946, 857, 834, 767, 694. 1H NMR (CDCl3,
300 MHz) d 0.00 (s, 9H), 0.96 (t, JZ8.6 Hz, 2H), 2.21 (t,
JZ2.3 Hz, 1H), 3.94 (dd, JZ2.3, 5.6 Hz, 2H), 4.16 (t, JZ
8.6 Hz, 2H), 4.86 (br s, 1H); 13C NMR (CD3COCD3,
75 MHz) d K1.5 (3C), 17.7, 30.6, 53.4, 71.3, 79.9, 156.3.
4.2.1. (3-Cyano-4-hydroxy-phenyl)-carbamic acid 2-tri-
methylsilyl-ethyl ester (5). To a solution of nitro compound
911,12 (500 mg, 3.05 mmol) in EtOH (100 mL) was added
Pd/C 10% (158 mg, 0.15 mmol). The suspension was
degassed under reduced pressure and then stirred under a
hydrogen atmosphere. When all the starting material had
been consumed, the mixture was degassed with argon before
being filtered over Celite 545. The filtrate was evaporated
under reduced pressure and the dark solid residue consisting
predominantly of amine 10 was used without further
purification for the next synthetic step. The crude amine
10 was dissolved in pyridine (7 mL) then p-nitrophenyl-
trimethylsilylethylcarbonate (2.39 g, 8.44 mmol) and
DMAP (233 mg, 1.91 mmol) were added. This solution
was kept at 50 8C during 48 h before being evaporated under
reduced pressure. The crude oil was dissolved in a mixture
of acetone (20 mL) and water (20 mL) and Na2CO3 (2.00 g,
18.87 mmol) was added. This suspension was refluxed
(60 8C) during 24 h before being cooled down to room
temperature. The mixture was diluted with water (30 mL)
and extracted with Et2O (3!20 mL). The collected organic
layers were washed with brine, dried over Na2SO4, filtered
and evaporated under reduced pressure. The yellow
deliquescent crude mixture was chromatographed on a
silica gel column (30 g of SiO2, CH2Cl2/MeOH: 95:5)
leading to the expected substituted carbamate 5 (432 mg,
1.56 mmol, overall yield starting from nitro compound 9:
51%) isolated as a pale yellow solid. Mp 133–136 8C; IR
(neat) 3351, 3216, 2958, 2901, 2245, 1765, 1703, 1605,
1548, 1506, 1427, 1365, 1251, 1227, 1205, 1169, 1115,
4.2.4. [3-(3-Cyano-4-hydroxy-phenyl)-prop-2-ynyl]-
carbamic acid 2-trimethylsilylethyl ester (6). A solution
of iodinated compound 12 (707 mg, 2.88 mmol), carbamic
ester 13 (748 mg, 3.75 mmol) and DIPEA (2.40 mL,
1780 mg, 13.80 mmol) in freshly distilled DMF (13 mL)
was cooled down to 0 8C and degassed under reduced
pressure. Pd(PPh3)4 (172 mg, 0.15 mmol) and CuI (116 mg,
0.61 mmol) were then successively added under argon. The
mixture was degassed again at 0 8C and then warmed up at
room temperature and stirred 2 h. DMF was then removed
by distillation under reduced pressure and the crude mixture
was adsorbed on silica before being chromatographed on a
silica gel column (40 g of SiO2, hexane/Et2O: 2:1 then
hexane/Et2O: 1:1). The acetylenic compound 6 (869 mg,
2.74 mmol, yield: 95%) was isolated as a yellow foamy
solid. Mp 80–82 8C; IR (neat) 3400, 3130, 2954, 2230,
1668, 1605, 1537, 1508, 1412, 1376, 1355, 1305, 1261,
1250, 1217, 1176, 1142, 1111, 1069, 1043, 1006, 942, 890,
855, 836, 769, 726, 693, 662. 1H NMR (CDCl3, 300 MHz) d
1
1060, 1041, 975, 931, 885, 825, 766, 730, 697. H NMR
CDCl3, 300 MHz) d 0.09 (s, 9H), 1.05 (m, 2H), 4.27 (m,
2H), 6.53 (br s, 1H), 6.91 (d, JZ8.8 Hz, 1H), 7.41 (dd, JZ
2.5, 8.8 Hz, 1H), 7.56 (d, JZ2.5 Hz, 1H); 13C NMR
(CD3COCD3, 75 MHz) d 0.0 (3C), 19.7, 64.9, 101.9, 118.5,
118.9, 124.0, 127.5, 134.6, 156.2, 157.5. ESI-TOF m/z 579
(2MCNaC), 857 (3MCNaC). Anal. Calcd for
C13H18N2O3Si: C, 56.09; H, 6.52; N, 10.06. Found C,
55.83; H, 6.55; N, 9.79.

F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
2251
0.06 (s, 9H), 1.02 (t, JZ8.4 Hz, 2H), 4.16 (d, JZ5.9 Hz,
2H), 4.24 (t, JZ8.4 Hz, 2H), 5.01 (br s, 1H), 6.93 (d, JZ
8.7 Hz, 1H), 7.27 (d, JZ8.7 Hz, 1H), 7.50 (s, 1H); 13C NMR
(CDCl3, 75 MHz) d K1.5 (3C), 17.7, 31.4, 64.3, 84.9, 91.1,
100.0, 114.6, 115.9, 116.8, 136.2, 137.6, 157.2, 159.6; ESI-
TOF m/z 655 (2MCNaC), 971 (3MCNaC). Anal. Calcd
for C16H20N2O3Si: C, 60.73; H, 6.37; N, 8.85. Found C,
60.87; H, 6.25; N, 8.34.
4.3.1. {4-Hydroxy-3-[4-(methoxy-methyl-carbamoyl)-
4,5-dihydro-thiazol-2-yl]-phenyl}-carbamic acid 2-tri-
methylsilyl-ethylester (17). Isolated as a white powder
(overall yield over two steps from 5: 81%). Mp 119–122 8C;
IR (neat) 3300, 2953, 1716, 1645, 1561, 1486, 1459, 1433,
1394, 1344, 1293, 1230, 1193, 1179, 1062, 994, 964, 947,
860, 834, 810, 767, 693, 664. 1H NMR (CDCl3, 300 MHz) d
0.00 (s, 9H), 0.98 (m, 2H), 3.24 (s, 3H), 3.42 (dd, JZ9.3,
10.9 Hz, 1H), 3.70 (br t, JZ10.9 Hz, 1H), 3.77 (s, 3H), 4.18
(m, 2H), 5.62 (br t, JZ9.3 Hz, 1H), 6.65 (br s, 1H), 6.85 (d,
JZ8.9 Hz, 1H), 7.26 (br d, JZ8.9 Hz, 1H), 7.44 (br s, 1H),
12.5 (br s, 1H); 13C NMR (CDCl3, 75 MHz) d 0.0 (3C),
19.2, 34.1, 34.5, 63.3, 65.0, 76.2, 117.2, 118.9, 122.6, 126.7,
131.0, 155.6, 156.7, 171.3, 175.4. ESI-TOF m/z 426 (MC
HC), 448 (MCNaC), 851 (2MCHC), 873 (2MCNaC).
Anal. Calcd for C18H27N3O5SSi: C, 50.80; H, 6.39; N, 9.87.
Found C, 50.86; H, 6.13; N, 9.83.
4.2.5. [3-(3-Cyano-4-hydroxy-phenyl)-propyl]-carbamic
acid 2-trimethylsilylethyl ester (7). To a solution of
acetylenic carbamate 6 (700 mg, 2.22 mmol) in EtOH
(40 mL) was added Pd/C 10% (260 mg, 0.22 mmol). The
suspension was degassed under reduced pressure and then
stirred under hydrogen atmosphere. When all the starting
material had been consumed, the mixture was degassed with
argon before being filtered on Celite. The filtrate was
evaporated under reduced pressure and the oily residue was
chromatographed on a silica gel column (30 g of SiO2,
hexane/Et2O: 1:1) leading to the expected substituted
2-hydroxybenzonitrile 7 (640 mg, 2.00 mmol, yield: 90%)
isolated as a thick colorless oil, which slowly crystallized
upon storage in the refrigerator. Mp 89–91 8C; IR (neat)
3389, 3135, 2956, 2928, 2910, 2861, 2236, 1668, 1611,
1598, 1532, 1515, 1461, 1428, 1374, 1307, 1275, 1260,
1251, 1210, 1174, 1143, 1115, 1064, 1020, 980, 951, 934,
887, 837, 825, 782, 762, 694, 677. 1H NMR (CDCl3,
300 MHz) d 0.06 (s, 9H), 0.98 (t, JZ8.6 Hz, 2H), 1.78 (p,
JZ7.5 Hz, 2H), 2.56 (t, JZ7.5 Hz, 2H), 3.17 (q, JZ6.6 Hz,
2H), 4.16 (t, JZ8.6 Hz, 2H), 4.80 (br s, 1H), 6.90 (d, JZ
8.3 Hz, 1H), 7.21 (d, JZ8.3 Hz, 1H), 7.26 (s, 1H); 13C NMR
(CDCl3, 75 MHz) d K1.5 (3C), 17.7, 31.3, 31.5, 40.3, 63.6,
99.3, 116.5, 117.0, 132.2, 133.0, 134.1, 157.6, 158.0. ESI-
TOF m/z 321 (MCHC), 343 (MCNaC), 359 (MCKC),
663 (2MCNaC). Anal. Calcd for C16H24N2O3Si: C, 59.97;
H, 7.55; N, 8.74. Found C, 59.68; H, 7.70; N, 8.24.
4.3.2. (3-{4-Hydroxy-3-[4-(methoxy-methyl-carbamoyl)-
4,5-dihydro-thiazol-2-yl]-phenyl}-prop-2-ynyl)-carbamic
acid 2-trimethylsilylethylester (18). Isolated as a yellow
solid (overall yield over two steps from 6: 62%). Mp
160–162 8C; IR (neat) 3316, 2952, 1711, 1668, 1619, 1561,
1520, 1486, 1425, 1391, 1355, 1323, 1292, 1248, 1201,
1178, 1130, 1044, 1002, 970, 943, 891, 858, 832, 768, 735,
695, 662. 1H NMR (CDCl3, 300 MHz) d 0.05 (s, 9H), 1.01
(t, JZ8.3 Hz, 2H), 3.30 (s, 3H), 3.50 (dd, JZ9.4, 10.5 Hz,
2H), 3.79 (t, JZ10.5 Hz, 2H), 4.09 (d, JZ5.9 Hz, 2H), 4.12
(t, JZ8.3 Hz, 2H), 4.78 (br s, 1H), 5.69 (t, JZ9.4 Hz, 1H),
6.92 (d, JZ8.5 Hz, 1H), 7.39 (dd, JZ2.0, 8.5 Hz, 1H), 7.50
(d, JZ2.0 Hz, 1H), 12.5 (br s, 1H); 13C NMR (CDCl3,
75 MHz) d K1.5 (3C), 17.7, 31.5, 32.6, 33.0, 61.8, 63.5,
74.6, 82.2, 84.0, 113.2, 116.0, 117.4, 134.2, 136.5, 156.3,
159.2, 169.5, 173.6; ESI-TOF m/z 464 (MCHC), 927
(2MCHC). Anal. Calcd for C21H29N3O5SSi: C, 54.40; H,
6.30; N, 9.06. Found C, 54.76; H, 6.46; N, 8.82.
4.3.3. (3-{4-Hydroxy-3-[4-(methoxy-methyl-carbamoyl)-
4,5-dihydro-thiazol-2-yl]-phenyl}-propyl)-carbamic acid
2-trimethylsilanylethylester (19). Isolated as a white solid
(overall yield over two steps from 7: 89%). Mp 160–162 8C;
IR (neat) 3347, 2926, 2854, 1715, 1672, 1628, 1597, 1567,
1521, 1491, 1388, 1248, 1176, 1138, 1043, 1007, 988, 966,
858, 834, 767, 735, 694. 1H NMR (CDCl3, 300 MHz) d 0.00
(s, 9H), 0.94 (t, JZ8.6 Hz, 2H), 1.74 (p, JZ7.5 Hz, 2H),
2.54 (t, JZ7.5 Hz, 2H), 3.15 (m, 2H), 3.25 (s, 3H), 3.43 (dd,
JZ9.5, 11.0 Hz, 2H), 3.73 (t, JZ9.5 Hz, 1H), 3.79 (s, 3H),
4.41 (t, JZ8.6 Hz, 2H), 4.71 (br s, 1H), 5.64 (t, JZ8.9 Hz,
1H), 6.86 (d, JZ8.2 Hz, 1H), 7.15 (m, 2H); 12.11 (br s, 1H);
13C NMR (CDCl3, 75 MHz) d K1.5 (3C), 17.7, 29.7, 31.9,
32.6, 32.9, 40.4, 61.8, 62.9, 76.6, 115.9, 117.7, 130.1, 131.9,
133.5, 156.8, 157.2, 169.7, 173.9. ESI-TOF m/z 468 (MC
HC); HRMS calcd for C21H34N3O5SSi 468.1983, found
468.1981. Anal. Calcd for C21H34N3O5SSi: C, 53.93; H,
7.11; N, 8.99. Found C, 53.71; H, 7.25; N, 8.27.
4.3. General procedure for Weinreb amides (17), (18)
and (19)
Substituted 2-hydroxybenzonitriles 5, 6 or 7 (1.00 equiv) and
(R)-cysteine (2.00 equiv) were dissolved in methanol (8 mL/
mmol of substituted 2-hydroxybenzonitrile). The suspension
was warmed up to 50 8C then 0.1 M phosphate buffer pH 6.4
(8 mL/mmol of substituted 2-hydroxybenzonitrile) was
added. The mixture was vigorously stirred at 60 8C during
36 h. The resulting yellow solution was cooled down to room
temperature before being diluted with water and acidified to
pH 2.0 by addition of solid citric acid. After extraction with
CH2Cl2, the collected organic layers were dried over Na2SO4,
filtered and evaporated under reduced pressure. The crude
thiazolines14, 15and16isolatedasyellow powderswereused
without further purification for the next step. Thiazoline was
dissolved in CH2Cl2 (25 mL/mmol of thiazoline), the solution
was cooled down to 0 8C. EDCI (1.10 equiv) was then added,
immediately followed by a solution of N,O-dimethylhydroxyl-
amine hydrochloride (1.20 equiv) and DIPEA (1.20 equiv) in
CH2Cl2 (25 mL/mmol of N,O-dimethylhydroxylamine
hydrochloride). The mixture was allowed to warm up to
room temperature and was stirred during 2 h. The mixture
was directly adsorbed on silica before being chromato-
graphed on a silica gel column eluted with hexane/Et2O: 1:1,
leading to the expected Weinreb amides 17, 18 or 19.
4.4. General procedure for functionalized pyochelin
derivatives (2), (3) and (4)
To a solution of Weinreb amide 17, 18 or 19 in dry THF
(16 mL/mmol of Weinreb amide), cooled down to K40 8C,
LiAlH4 (1.30 equiv of a 1 M solution in THF) was injected
dropwise. The reaction temperature was allowed to rise

2252
F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
to K20 8C in 30 min. The reaction mixture was then
hydrolyzed by successive additions of a saturated aqueous
solution of NH4Cl (12 mL/mmol of LiAlH4) then a 1 M
aqueous solution of KHSO4 (5 mL/mmol of LiAlH4). The
mixture was warmed up to room temperature and
vigourously stirring was applied until two phases were
formed. After partition and extraction with Et2O, the
organic layers were collected, dried over Na2SO4, filtered
before being evaporated. The crude aldehyde, very sensitive
to oxidation, was used directly for subsequent reaction. It
was dissolved into a mixture of ethanol (20 mL/mmol of
Weinreb amide) and water (6 mL/mmol of Weinreb amide)
and to this solution were successively added, potassium
acetate (6.65 equiv) and (R)-N-methylcysteine hydrochloride
(2.12 equiv). The mixture was then gently stirred in the dark
during 14 h (overnight) before being diluted with water
(30 mL) then washed with hexane (30 ml). The aqueous
layer was then acidified to pH 4.0 by addition of solid citric
acid before being extracted with CH2Cl2. The organic layers
were collected, dried over Na2SO4, filtered and evaporated
under reduced pressure. The expected functionalized
pyochelins 2, 3 and 4 are isolated first as yellow foams
then were dissolved in cyclohexane containing a minimun
of CH2Cl2. After evaporation of the solvent under reduced
pressure, the expected pyochelins were isolated as yellow
powders.
(C4), 131.9 (C5), 154.8 (OCONH), 155.6 (C2), 172.3 (C20),
172.9 (COOH).
4.4.4. (2a) (40R,200R,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 1.03 (t, JZ8.5 Hz, 2H,
CH2–Si), 2.64 (s, 3H, N–CH3), 3.18–3.27 (m, 2H, H500),
3.49 (d, JZ9.0 Hz, 2H, H50), 3.73 (dd, JZ7.0–7.9 Hz, 1H,
H400), 4.21 (t, JZ8.5 Hz, 2H, CH2–O), 4.61 (d, JZ5.4 Hz,
1H, H200), 5.21 (td, JZ5.4–9.0 Hz, 1H, H40), 6.87 (dd,
JZ1.4–6.4 Hz, 1H, H3), 7.50 (d, JZ1.4 Hz, 1H, H4), 7.77
(s, 1H, H6), 8.49 (br s, 1H, NH). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.2 (CH2–Si), 32.8 (C50),
33.1 (C500), 41.3 (N–CH3), 63.2 (CH2–O), 73.3 (C400), 77.1
(C200), 80.3 (C40), 116.5 (C1), 117.8 (C3), 120.6 (C6), 1250.1
(C4), 131.9 (C5), 154.8 (OCONH), 155.6 (C2), 172.3 (C2 ),
172.9 (COOH).
4.4.5. (2b) (40R,200S,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 1.03 (t, JZ8.5 Hz, 2H,
CH2–Si), 2.49 (s, 3H, N–CH3), 3.18–3.27 (m, 2H, H500),
3.43–3.49 (m, 1H, H50), 3.63–3.68 (m, 1H, H50), 4.18–4.28
(m, 1H, H400), 4.21 (t, JZ8.5 Hz, 2H, CH2–O), 4.55 (d, JZ
8.2 Hz, 1H, H200), 5.01 (q, JZ8.2 Hz, 1H, H40), 6.87 (dd,
JZ1.4–6.4 Hz, 1H, H3), 7.50 (d, JZ1.4 Hz, 1H, H4), 7.77
(s, 1H, H6), 8.49 (br s, 1H, NH). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.2 (CH2–Si), 32.1 (C500),
35.1 (C50), 37.8 (N–CH3), 63.2 (CH2–O), 71.0 (C400), 77.9
(C200), 81.2 (C40), 116.5 (C1), 117.8 (C3), 120.6 (C6), 1250.1
(C4), 131.9 (C5), 154.8 (OCONH), 155.6 (C2), 172.3 (C2 ),
172.9 (COOH).
4.4.1. 20-[2-Hydroxy-5-(2-trimethylsilyl-ethoxycarbonyl-
amino)-phenyl]-3-methyl-2,3,4,5,40,50-hexahydro-[2,40]-
bisthiazolyl-4-carboxylic acid (2). Isolated as a pale yellow
powder (overall yield over two steps from 17: 72%),
proportions: 2a/2b/2c/2d: 5/10/60/25. IR (neat) 3301, 2952,
2891, 1698, 1628, 1568, 1544, 1494, 1431, 1394, 1299, 1224,
1191, 1061, 996, 954, 928, 857, 834, 766, 693; HRMS calcd
for C20H28N3O5S2Si 482.1245, found 482.1240.
4.4.6. 20-{2-Hydroxy-5-[3-(2-trimethylsilyl-ethoxy-
carbonylamino)-prop-1-ynyl]-phenyl}-3-methyl-2,3,4,
5,40,50-hexahydro-[2,40]bisthiazolyl-4-carboxylic acid
(3). Isolated as a pale yellow powder (overall yield over
two steps from 18: 75%), proportions: 3a/3b/3c/3d: 15/15/
45/25. IR (neat) 3320, 2953, 1709, 1619, 1569, 1525, 1487,
1395, 1324, 1247, 1196, 1128, 1041, 939, 857, 833, 775,
694; HRMS calcd for C23H30N3O5S2Si 520.1402, found
520.1408.
4.4.2. (2c) (40S,200R,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 1.03 (t, JZ8.5 Hz, 2H,
CH2–Si), 2.63 (s, 3H, N–CH3), 3.31 (dd, JZ7.0–11.2 Hz,
1H, H500), 3.45 (dd, JZ7.0–11.2 Hz, 1H, H500), 3.53 (dd, JZ
8.4–11.3 Hz, 1H, H50), 3.63 (dd, JZ8.4–11.3 Hz, 1H, H50),
4.01 (dd, JZ5.4–7.0 Hz, 1H, H400), 4.21 (t, JZ8.5 Hz, 2H,
CH2–O), 4.34 (d, JZ8.4 Hz, 1H, H200), 4.83 (q, JZ8.4 Hz,
1H, H40), 6.87 (dd, JZ1.4–6.4 Hz, 1H, H3), 7.50 (d, JZ
1.4 Hz, 1H, H4), 7.77 (s, 1H, H6), 8.49 (br s, 1H, NH). 13C
NMR (125 MHz, CD3COCD3) d K1.5 ((CH3)3Si), 18.2
(CH2–Si), 33.5 (C500), 34.9 (C50), 44.8 (N–CH3), 63.2
(CH2–O), 73.9 (C400), 79.4 (C200), 83.4 (C40), 116.5 (C1),
117.8 (C3), 120.6 (C6), 125.1 (C4), 131.9 (C5), 154.8
(OCONH), 155.6 (C2), 172.3 (C20), 172.9 (COOH).
4.4.7. (3c) (40S,200R,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.86 (t, JZ8.1 Hz, 2H,
CH2–Si), 2.63 (s, 3H, N–CH3), 3.35 (dd, JZ6.9–11.0 Hz,
1H, H500), 3.44 (dd, JZ5.2–11.0 Hz, 1H, H500), 3.55 (dd, JZ
8.4–11.5 Hz, 1H, H50), 3.66 (dd, 0J0 Z8.4–11.5 Hz, 1H, H50),
4.01 (dd, JZ5.2–6.9 Hz, 1H, H4 ), 4.12 (d, JZ5.4 Hz, 2H,
CH2–N), 4.14 (t, JZ8.1 Hz, 2H, CH2–O), 4.34 (d, JZ
8.4 Hz, 1H, H200), 4.84 (q, JZ8.4 Hz, 1H, H40), 6.59 (br s,
1H, NH), 6.91–6.95 (m, 1H, H3), 7.40–7.44 (m, 2H, H6C
H4). 13C NMR (125 MHz, CD3COCD3) d K1.5 ((CH3)3Si),
18.2 (CH2–Si), 31.5 (CH2–N), 33.5 (C500), 35.0 (C50), 44.7
(N–CH3), 63.0 (CH2–O), 73.7 (C400), 79.2 (C200), 81.5
(C^C–Ar), 83.1 (C40), 86.2 (C^C–Ar), 114.3 (C5), 116.9
(C1), 118.2 (C3), 134.1 (C6), 136.9 (C4), 157.1 (OCONH),
159.9 (C2), 171.6 (C20), 174.9 (COOH).
4.4.3. (2d) (40S, 200S, 400R). 1H NMR (300 MHz, CD3COCD3)
d 0.00(s, 9H, (CH3)3Si), 1.03 (t, JZ8.5 Hz, 2H, CH2–Si), 2.70
(s, 3H, N–CH3), 3.06 (dd, JZ2.0–10.4 Hz, 1H, H500), 3.18–
3.27(m, 1H, H500), 3.38–3.50 (m, 2H, H50), 4.21 (t, JZ8.5 Hz,
2H, CH2–O), 4.24 (dd, JZ2.0–6.6 Hz, 1H, H400), 5.05 (d, JZ
4.7 Hz, 1H, H200), 5.28 (td, JZ4.7–9.0 Hz, 1H, H40), 6.87 (dd,
JZ1.4–6.4 Hz, 1H, H3), 7.50 (d, JZ1.4 Hz, 1H, H4), 7.77 (s,
1H, H6), 8.49 (br s, 1H, NH). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.2 (CH2–Si), 32.2 (C50),
32.3 (C500), 36.2 (N–CH3), 63.2 (CH2–O), 70.5 (C400), 73.6
(C200), 79.3 (C40), 116.5 (C1), 117.8 (C3), 120.6 (C6), 125.1
4.4.8. (3d) (40S,200S,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.86 (t, JZ8.1 Hz, 2H,
CH2–Si), 2.70 (s, 3H, N–CH3), 3.05 (dd, JZ2.4–10.4 Hz,
1H, H500), 3.20–3.26 (m, 1H, H500), 3.42–3.56 (m, 2H, H50),
4.12 (d, JZ5.4 Hz, 2H, CH2–N), 4.14 (t, JZ8.1 Hz, 2H,
CH2–O), 4.24 (dd, JZ2.4–6.6 Hz, 1H, H400), 5.05 (d, JZ

F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
2253
4.7 Hz, 1H, H200), 5.30 (td, JZ4.7–9.3 Hz, 1H, H40), 6.59
(br s, 1H, NH), 6.91–6.95 (m, 1H, H3), 7.40–7.44 (m, 2H,
H6CH4). 13C NMR (125 MHz, CD3COCD3) d K1.5
((CH3)3Si), 18.2 (CH2–Si), 31.5 (CH2–N), 32.2 (C50), 32.3
(C500), 36.0 (N–CH3), 63.0 (CH2–O), 70.3 (C400), 73.3
(C200), 79.0 (C40), 81.5 (C^C–Ar), 86.2 (C^C–Ar), 114.3
(C5), 116.9 (C1), 118.2 (C3), 134.1 (C6), 136.9 (C4), 157.1
(OCONH), 159.9 (C2), 171.6 (C20), 174.9 (COOH).
(C5), 134.2 (C4), 157.0 (OCONH), 158.1 (C2), 172.0 (C20),
173.1 (COOH).
4.4.13. (4d) (40S,200S,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.96 (t, JZ8.2 Hz, 2H,
CH2–Si), 1.79 (p, JZ7.2 Hz, 2H, C–CH2–C), 2.62 (t, JZ
7.9 Hz, 2H, CH2–Ar), 2.71 (s, 3H, N–CH3), 3.06 (dd, JZ
2.4–9.7 Hz, 1H, H500), 3.11–3.17 (m, 2H, CH2–N), 3.18–
3.24 (m, 1H, H500), 3.40 (dd, JZ9.1–11.0 Hz, 1H, H50), 3.46
(dd, JZ9.1–11.0 Hz, 1H, H50), 4.10 (t, JZ8.2 Hz, 2H,
CH2–O), 4.25 (dd, JZ2.4–6.2 Hz, 1H, H400), 5.05 (d, JZ
4.6 Hz, 1H, H200), 5.27 (td, JZ4.6–9.1 Hz, 1H, H40), 6.18
(br s, 1H, NH), 6.87 (d, JZ8.2 Hz, 1H, H3), 7.26 (s, 1H,
H6), 7.28 (d, JZ2.0 Hz, 1H, H4). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.3 (CH2–Si), 32.1 (C50),
32.2 (C500), 32.5 (CH2–Ar), 32.7 (C–CH2–C), 36.1 (N–
CH3), 40.7 (CH2–N), 62.4 (CH2–O), 70.4 (C400), 73.6 (C200),
79.2 (C40), 116.6 (C1), 117.6 (C3), 130.6 (C6), 133.3 (C5),
134.2 (C4), 157.0 (OCONH), 158.1 (C2), 172.0 (C20), 173.1
(COOH).
4.4.9. (3a) (40R,200R,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.86 (t, JZ8.1 Hz, 2H,
CH2–Si), 2.63 (s, 3H, N–CH3), 3.20–3.24 (m, 2H, H500),
3.52 (d, JZ9.3 Hz, 2H, H50), 3.71 (dd, JZ6.8–8.0 Hz, 1H,
H400), 4.12 (d, JZ5.4 Hz, 2H, CH2–N), 4.14 (t, JZ8.1 Hz,
2H, CH2–O), 4.61 (d, JZ5.5 Hz, 1H, H200), 5.21 (td, JZ
5.5–9.3 Hz, 1H, H40), 6.59 (br s, 1H, NH), 6.91–6.95 (m,
1H, H3), 7.40–7.44 (m, 2H, H6CH4). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.2 (CH2–Si), 31.5
(CH2–N), 32.8 (C50), 33.1 (C500), 41.3 (N–CH3), 63.0
(CH2–O), 73.2 (C400), 76.9 (C200), 80.1 (C40), 81.5
(C^C–Ar), 86.2 (C^C–Ar), 114.3 (C5), 116.9 (C1),
118.2 (C3), 134.1 (C6), 136.9 (C4), 157.1 (OCONH),
159.9 (C2), 171.6 (C20), 174.9 (COOH).
4.4.14. (4a) (40R,200R,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.96 (t, JZ8.2 Hz, 2H,
CH2–Si), 1.79 (p, JZ7.2 Hz, 2H, C–CH2–C), 2.62 (t, JZ
7.9 Hz, 2H, CH2–Ar), 2.64 (s, 3H, N–CH3), 3.11–3.17 (m,
2H, CH2–N), 3.18–3.20 (m, 2H, H500), 3.48 (d, JZ9.1 Hz,
2H, H50), 3.71 (dd, JZ6.7–8.2 Hz, 1H, H400), 4.10 (t, JZ
8.2 Hz, 2H, CH2–O), 4.62 (d, JZ5.4 Hz, 1H, H200), 5.19 (td,
JZ5.4–9.1 Hz, 1H, H40), 6.18 (br s, 1H, NH), 6.87 (d, JZ
8.2 Hz, 1H, H3), 7.26 (s, 1H, H6), 7.28 (d, JZ2.0 Hz, 1H,
H4). 13C NMR (125 MHz, CD3COCD3) d K1.5 ((CH3)3Si),
18.3 (CH2–Si), 32.5 (CH2–Ar), 32.7 (C–CH2–C), 32.7
(C50), 33.0 (C500), 40.7 (CH2–N), 41.4 (N–CH3), 62.4 (CH2–
O), 73.2 (C400), 77.1 (C200), 80.2 (C40), 116.6 (C1), 117.6
(C3), 130.6 (C6), 133.3 (C5), 134.2 (C4), 157.0 (OCONH),
158.1 (C2), 172.0 (C20), 173.1 (COOH).
4.4.10. (3b) (40R,200S,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.86 (t, JZ8.1 Hz, 2H,
CH2–Si), 2.50 (s, 3H, N–CH3), 3.20–3.26 (m, 2H, H500),
3.47–3.52 (m, 1H, H50), 3.68–3.72 (m, 1H, H50), 4.12 (d,
JZ5.4 Hz, 2H, CH2–N), 4.14 (t, JZ8.1 Hz, 2H, CH2–O),
4.23 (t, JZ6.8 Hz, 1H, H4000), 4.55 (d, JZ8.1 Hz, 1H, H200),
5.02 (q, JZ8.1 Hz, 1H, H4 ), 6.59 (br s, 1H, NH), 6.91–6.95
(m, 1H, H3), 7.40–7.44 (m, 2H, H6CH4). 13C NMR
(125 MHz, CD3COCD3) d K1.5 ((CH3)3Si), 18.2 (CH2–Si),
31.5 (CH2–N), 32.0 (C500), 35.0 (C50), 37.7 (N–CH3), 63.0
(CH2–O), 70.8 (C400), 77.7 (C200), 80.9 (C40), 81.5
(C^C–Ar), 86.2 (C^C–Ar), 114.3 (C5), 116.9 (C1),
118.2 (C3), 134.1 (C6), 136.9 (C4), 157.1 (OCONH),
159.9 (C2), 171.6 (C20), 174.9 (COOH).
4.4.15. (4b) (40R,200S,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.96 (t, JZ8.2 Hz, 2H,
CH2–Si), 1.79 (p, JZ7.2 Hz, 2H, C–CH2–C), 2.50 (s, 3H,
N–CH3), 2.62 (t, JZ7.9 Hz, 2H, CH2–Ar), 3.11–3.17 (m,
2H, CH2–N), 3.21–3.24 (m, 2H, H500), 3.45 (dd, JZ9.0–
11.5 Hz, 1H, H50), 3.65 (dd, JZ9.0–11.5 Hz, 1H, H50), 4.10
(t, JZ8.2 Hz, 2H, CH2–O), 4.23 (t, JZ6.8 Hz, 1H, H400),
4.55 (d, JZ8.2 Hz, 1H, H200), 5.00 (q, JZ8.2 Hz, 1H, H40),
6.18 (br s, 1H, NH), 6.87 (d, JZ8.2 Hz, 1H, H3), 7.26 (s,
1H, H6), 7.28 (d, JZ2.0 Hz, 1H, H4). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.3 (CH2–Si), 32.0 (C500),
32.5 (CH2–Ar), 32.7 (C–CH2–C), 34.8 (C50), 37.7 (N–CH3),
40.7 (CH2–N), 62.4 (CH2–O), 70.9 (C400), 77.8 (C200), 81.1
(C40), 116.6 (C1), 117.6 (C3), 130.6 (C6), 133.3 (C5), 134.2
(C4), 157.0 (OCONH), 158.1 (C2), 172.0 (C2), 173.1
(COOH).
4.4.11. 20-{2-Hydroxy-5-[3-(2-trimethylsilyl-ethoxy-
carbonylamino)-propyl]-phenyl}-3-methyl-2,3,4,5,40,50-
hexahydro-[2,40]bisthiazolyl-4-carboxylic acid (4). Iso-
lated as a yellow powder (overall yield over two steps from
19: 65%), proportions: 4a/4b/4c/4d: 20/10/50/20. IR (neat)
3326, 2950, 1695, 1626, 1569, 1528, 1492, 1248, 1100, 857,
834, 775, 693; HRMS calcd for C23H36N3O5S2Si 526.1860,
found 526.1851.
4.4.12. (4c) (40S,200R,400R). 1H NMR (300 MHz, CD3-
COCD3) d 0.00 (s, 9H, (CH3)3Si), 0.96 (t, JZ8.2 Hz, 2H,
CH2–Si), 1.79 (p, JZ7.2 Hz, 2H, C–CH2–C), 2.62 (t, JZ
7.9 Hz, 2H, CH2–Ar), 2.63 (s, 3H, N–CH3), 3.11–3.17 (m,
2H, CH2–N), 3.30 (dd, JZ6.8–11.0 Hz, 1H, H500), 3.44 (dd,
JZ5.2–11.0 Hz, 1H, H500), 3.55 (dd, J0Z8.4–11.4 Hz, 1H,
H50), 3.62 (dd, JZ8.4–11.4 Hz, 1H, H5 ), 4.01 (dd, JZ5.2–
6.8 Hz, 1H, H400), 4.10 (t, JZ8.2 Hz, 2H, CH2–O), 4.33 (d,
JZ8.4 Hz, 1H, H200), 4.83 (q, JZ8.4 Hz, 1H, H40), 6.18
(br s, 1H, NH), 6.87 (d, JZ8.2 Hz, 1H, H3), 7.26 (s, 1H,
H6), 7.28 (d, JZ2.0 Hz, 1H, H4). 13C NMR (125 MHz,
CD3COCD3) d K1.5 ((CH3)3Si), 18.3 (CH2–Si), 32.5
(CH2–Ar), 32.7 (C–CH2–C), 34.4 (C500), 34.7 (C50), 40.7
(CH2–N), 44.7 (N–CH3), 62.4 (CH2–O), 73.8 (C400), 79.3
(C200), 83.4 (C40), 116.6 (C1), 117.6 (C3), 130.6 (C6), 133.3
Acknowledgements
The authors thank the CNRS (Centre National de la
Recherche Scientifique) and the association Vaincre
la Mucoviscidose for financial support and for providing a
postdoctoral fellowship to F. R. We also thank Le Grand-
´
Duche du Luxembourg for providing a doctoral fellowship
to V. S.

2254
F. Rivault et al. / Tetrahedron 62 (2006) 2247–2254
2001, 57, 1897–1902. (d) Rinehart, K. L.; Staley, A. L.;
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