A synthesis of all stereoisomers of tenuecyclamide A employing a fluorous-Fmoc strategy

Article abstract of DOI:10.1021/jo401647k

A concise liquid-phase combinatorial synthesis of all stereoisomers of Tenuecyclamide A was achieved using a mixture of d-/l-alanine with each stereoisomer encoded by a different f-Fmoc tag. The synthetic strategy using f-Fmoc reagents as the protecting group for amino acids has been demonstrated to be a useful method for diverse polypeptide analogue synthesis.

Full text of DOI:10.1021/jo401647k

Article
pubs.acs.org/joc
A Synthesis of All Stereoisomers of Tenuecyclamide A Employing a
Fluorous-Fmoc Strategy
Yuya Sugiyama, Fumitaka Eguchi, Atsushi Miyazaki, Koichi Hayashi, Hiroaki Takahashi,
Hiromi Hamamoto, Takayuki Shioiri, and Masato Matsugi*
Department of Applied Biological Chemistry, Faculty of Agriculture, Meijo University, 1-501 Shiogamaguchi, Tempaku-ku, Nagoya
468-8502, Japan
S
* Supporting Information
ABSTRACT: A concise liquid-phase combinatorial synthesis
of all stereoisomers of Tenuecyclamide A was achieved using a
mixture of D-/L-alanine with each stereoisomer encoded by a
different f-Fmoc tag. The synthetic strategy using f-Fmoc
reagents as the protecting group for amino acids has been
demonstrated to be a useful method for diverse polypeptide
analogue synthesis.
each individual isomer yet in the end does provide the entire
family of desired stereoisomers isolated individually.
INTRODUCTION
■
Since its introduction in 1997 by Curran et al., fluorous solid-
phase extraction (FSPE)1 has proven to be a highly efficient
and widely applicable technique for separating molecules that
are tagged with a fluorous domain from other nonfluorous
organic molecules.2 In recent years, the technique has evolved
such that molecules can be separated purely on the basis of
respective fluorine content regardless of other functionality
present in the remainder of the molecule.3 This salient feature
of FSPE allows for liquid-phase combinatorial synthesis called
fluorous mixture synthesis (FMS). In this technique, the
fluorous tags containing differing amounts of fluorine are
installed onto the various starting materials, which are then
mixed for the remainder of the synthesis and separated based
purely on fluorine content at the end, thereby greatly reducing
the number of synthetic steps required to build up a compound
library.4
In peptide synthesis, the Fmoc reagent is one of the most
well-known nitrogen protecting groups for amino acids.6 We
have previously reported a synthetic route toward two kinds of
fluorous-Fmoc reagents (f-Fmoc)7 and demonstrated that
fluorous tagged dipeptides can be separated due to the
remarkable differences in their retention times on a fluorous
HPLC (f-HPLC) column.8 Based on the success of this strategy
for dipeptides, we planned to use different f-Fmoc reagents with
varying amounts of fluorine as both a nitrogen protecting group
and a fluorous encoding tag for a liquid-phase combinatorial
peptide synthesis. Herein, we report the first example of a FMS
for all of the stereoisomers of the biologically active peptide
tenuecyclamide A9c (1) showcasing the f-Fmoc reagents as
tools for effective peptide isomer library synthesis.
A particularly novel application of FMS resides in
determining the stereochemical requirements of a molecule
for it to impart its biological activity. More specifically, the
biological activity of peptides is governed by the configuration
at the asymmetric center.5 From a drug discovery perspective, it
is interesting to study the biological activity of the non-natural
configuration to ascertain whether or not this modification
enhances or diminishes the biological activity or whether or not
it elicits an entirely different response. Synthetically, this
becomes a daunting task especially in the case of poly peptides
where for each additional center there are 2n possible
stereoisomers. FMS, however, is uniquely suited to address
this challenge because the various isomers can be individually
labeled with different length fluorous tags then mixed together.
The mixture can then be carried through the synthesis and then
separated at the very end purely based on the fluorine content,
thereby deconvoluting the entire mixture. This strategy saves a
significant amount of effort because the synthetic steps between
the mixing and the demixing do not have to be carried out on
RESULTS AND DISCUSSION
■
Tenuecyclamides A (1a) and B (1b) are cyclic hexapeptides
that differ only in configuration at a single stereocenter, and
they were both isolated from the cyanobacterium Nostoc
spongiaeforme var. tenue in 1998 (Figure 1).9 Although the exact
stereochemistry was not elucidated at the time of isolation and
structural identification, the assignment of the configuration
was subsequently achieved by Kelly’s group in the work that
employed solid-phase peptide synthesis.10 Tenuecyclamide A
(1a) was found to inhibit the division of sea urchin embryos
with an ED100 of 10.8 μM. To date, the biological activity of
tenuecyclamide B (1b) has not been explored in detail.9c
The first step in the synthesis of the tenuecyclamide family
was to protect L-alanine (2a) and D-alanine (2b) using two
different f-Fmoc reagents bearing C4F9 tags and C6F13 tags,
Received: July 29, 2013
Published: September 16, 2013
© 2013 American Chemical Society
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column,12 and the isolated individual compounds were
subsequently deprotected to give the enantiomerically pure
thiazoles 6a and 6b (Scheme 3).
a
Scheme 3. Preparation of 6a and 6b
Figure 1. Structure of tenuecyclamide A (1a) and B (1b).
a
Preparative f-HPLC conditions: 0 to 30 min 80% MeCN/H2O up to
100% MeCN. Flow rate: 7.0 mL/min; 254 nm; 6a, 80%; 6b, 80%.
respectively (Scheme 1). The f4-Fmoc reagent means bis-C4F9
tagged Fmoc, and the f6-Fmoc reagent means bis-C6F13 tagged
Similarly, a mixture of the oxazole-ring unit (7-mix) was
prepared from the tagged mixture of L- and D-alanine
synthesized previously. Scheme 4 shows the synthetic route
Scheme 1. N-Terminal Protection of Both L- and D-Alanine
by f-Fmoc Reagents
a
Scheme 4. Preparation of 7-mix
a
Reagents and conditions: (a) 2.2 equiv of L-threonine-OBn, 4.7 equiv
of HOBt, 4.7 equiv of HBTU, 4.2 equiv of DIEA, DMF, rt, 24 h,
quant; (b) 8.0 equiv of Dess−Martin periodinane, CH2Cl2, reflux, 2 h,
98%; (c) 18.0 equiv of Ph3PO, 9.0 equiv of Tf2O, CH2Cl2, 0 °C, 2 h,
quant; FSPE, 55% aq THF on fluorous silica gel; (d) H2, 20 mol %
Pd/C, MeOH/THF, rt, 2 h, 94%.
Fmoc reagent in all of the schemes. In this process, 1,4-dioxane
was employed for the f6-Fmoc protection simply because it
afforded greater solubility for the fluorous reagent in
comparison to the more commonly used MeCN.
Subsequently, using the synthesis reported by Kelly,10 we
prepared the two isomers of the thiazole-ring unit (5a and 5b)
as a mixture, where the two isomers are encoded by a different
fluorous tag (Scheme 2). To rapidly purify the reaction
to access 7-mix as a mixture of two compounds bearing either a
C4F9 or a C6F13 chain. In this case, FSPE was conducted after
step “c” to readily obtain the desired oxazole mixture 7-mix.
The synthesis then proceeded by splitting the 7-mix oxazole
mixture into two parts, with one part being condensed with 6a
and the other half with 6b, to give 8a-mix and 8b-mix,
respectively, each as a mixture of two compounds that are
orthogonally tagged (Scheme 5). 8a-mix and 8b-mix were then
each divided again into two parts to give a total of four
compound mixtures (Scheme 6). The allyl protecting groups in
all four mixtures were then deprotected separately to reveal the
corresponding acid and each mixture was subsequently
condensed again with either 6a or 6b, respectively. The
mixtures were purified by FSPE, and as a result, we obtained
a
Scheme 2. Preparation of 5a and 5b
a
Reagents and conditions: (a) 2.0 equiv of allyl alcohol, 1.1 equiv of
HOBt, 1.1 equiv of HBTU, 2.1 equiv of DIEA, DMF, rt, 24 h, quant;
(b) excess DEA, MeCN; (c) 1.0 equiv each of f4-Fmoc-L- and f6-Fmoc-
D-alanine, 2.2 equiv of HOBt, 2.2 equiv of HBTU, 4.2 equiv of DIEA,
DMF, rt, 12 h, 95% (two steps); FSPE, 55% aq THF on fluorous silica
gel; (d) 18.0 equiv of Ph3PO, 9.0 equiv of Tf2O, CH2Cl2, 0 °C, 2 h,
90%; FSPE, 70% aq MeOH on fluorous silica gel; (e) activated MnO2,
(10 w/w), CHCl3, reflux, 30 min, 75%.
a
Scheme 5. Preparation of 8a-mix and 8b-mix
mixtures and remove organic reagents and byproducts from the
desired fluorous tagged thiazoles after the reaction, FSPE was
conducted in steps “c” and “d”. In practice, the condensation
reaction in step “c” was accomplished by using HBTU and
HOBt. At the end of the reaction, the crude mixture was loaded
onto fluorous silica gel11 with MeOH, and this was followed by
an initial elution with aqueous MeOH which removed the
nontagged organic compounds (reagents and reagent by-
products) from the reaction mixture while the fluorous tagged
components were retained. A second elution with THF washed
the fluorous fraction off the column and as a result, the f-Fmoc
compounds were isolated in high yields without a tedious
purification procedure. The two components of 5a and 5b were
then separated by preparative f-HPLC8 using a fluorous HPLC
a
Reagents and conditions: (a) 2.4 equiv of 6a, 4.7 equiv of HOBt, 4.7
equiv of HBTU, 4.2 equiv of DIEA, DMF, rt, 12 h, 91%; FSPE, 55% aq
THF on fluorous silica gel; (b) 2.4 equiv of 6b, 4.7 equiv of HOBt, 4.7
equiv of HBTU, 4.2 equiv of DIEA, DMF, rt, 12 h, 98%; FSPE, 55% aq
THF on fluorous silica gel.
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Scheme 6. Preparation of 9bc-mix, 9ad-mix, 9ef-mix, and
a,b
9gh-mix and Separation by f-HPLC
Figure 2. f-HPLC analysis of 9bc-mix. HPLC conditions: FluoroFlash
HPLC column, 4.6 mm i.d., 150 mm length; 0 to 7.5 min 80% MeCN/
H2O up to 100% MeCN. Flow rate: 1.0 mL/min; 254 nm.
remarkable differences in retention time, each component of
the four two component mixtures in Scheme 6, 9bc-mix, 9ad-
mix, 9ef-mix and 9gh-mix, were separated to the correspond-
ing individual compounds 9a−h in almost quantitative yield
using preparative f-HPLC (Scheme 6).
After exploring a variety of conditions to effect the key
macrolactamization, using fully deprotected 9b as the test
substrate,13 we identified that employing a fluorous tagged
Mukaiyama reagent14 1015 was superior to other plausible
coupling reagents. Further reaction optimization revealed that
highly dilute reaction conditions, and syringe-pump addition of
the substrate were required to obtain superior yields. The
highest yield of 71% (three-step yield) for the macro-
lactamization was obtained when a DMF solution of 10 was
added dropwise over 24 h to a 0.5 mM DMF solution of 9b. At
the end of the cyclization reaction, the byproduct fluorous-
pyridone from 10 and other reagents used in the deprotection
process were effectively separated from the desired material by
precipitation. To achieve this, water was added to the reaction
mixture after the coupling, and the resulting suspension was
filtered. After a short column chromatography, further
purification of the products was not necessary as they were
found to be pure by 1H NMR. Having optimized the
lactamization conditions, we were poised to complete the
synthesis of all eight stereoisomers of tenuecyclamide A. This
was accomplished by treating compounds 9a−h individually
with the fluorous coupling reagent 10 after the f-Fmoc groups
had been removed with DEA and the allyl groups had been
removed using palladium catalysis. In this manner we
synthesized all eight target lactams (1a−g) with yields ranging
from 41 to 71% over the three step lactamization and
a
Reagents and conditions: (a) 20 mol % Pd(PPh3)4, 2.0 equiv of
PhSiH3, CH2Cl2, rt, 30 min; (b) 2.4 equiv of 6a, 4.7 equiv of HOBt,
4.7 equiv of HBTU, 4.2 equiv of DIEA, DMF, rt, 12 h; FSPE, 50% aq
THF on fluorous silica gel; 9bc-mix, 85%; 9ef-mix, 87% (two steps);
(c) 2.4 equiv of 6b, 4.7 equiv of HOBt, 4.7 equiv of HBTU, 4.2 equiv
of DIEA, DMF, rt, 12 h; FSPE, 50% aq THF on fluorous silica gel;
1
deprotection sequence (Scheme 7). The H NMR of both
tenuecyclamide A (1a) and B (1b) obtained here were
completely consistent with the data reported by Kelly.10 In
addition, 1H NMR of both of 1e and 1g, were entirely
consistent with the report.
b
9ad-mix, 80%; 9gh-mix, 82% (two steps). Preparative f-HPLC
conditions: 0−7.5 min 80% MeCN/H2O up to 100% MeCN. Flow
rate: 7.0 mL/min.
CONCLUSION
■
In summary, we have conducted a concise synthesis of all eight
stereoisomers of tenuecyclamide A (1a) based on an encoding
strategy of the amino acid stereocenters using f-Fmoc reagents.
If all stereoisomers were synthesized individually by the same
linear synthetic route as outlined in this work, 136 synthetic
steps would have been required; however, we carried out the
syntheses in a mere 52 steps which includes the f-Fmoc
protections. We believe that the f-Fmoc strategy will be a useful
methods for divergent polypeptide synthesis. Additional studies
and the examination of the biological activity of the novel
four pairs of compounds, namely 9bc-mix, 9ad-mix, 9ef-mix,
and 9gh-mix, each containing two compounds with different,
and thus chromatographically separable, fluorine content.
As a demonstration, we analyzed the 9bc-mix by f-HPLC
and confirmed that the C4F9-Fmoc compound 9b appeared at
10.8 min, whereas the C6F13-Fmoc compound 9c appeared at
23.8 min, a separation of 13.0 min completely dictated by the
relative fluorine content of each molecule (Figure 2). With such
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a
Hz, 2H), 5.26 (d, J = 7.8 Hz, 1H), 4.48−4.37 (m, 3H), 4.19 (t, J = 6.6
Hz, 1H), 3.02−2.96 (m, 4H), 2.52−2.32 (m, 4H), 1.49 (d, J = 7.0 Hz,
3H); 13C NMR (67.8 MHz, CDCl3) δ 177.8, 155.9, 144.6, 144.3,
139.7 (2C), 138.1 (2C), 127.9 (2C), 125.0 (2C), 124.9, 124.8, 120.1−
106.6 (12C), 67.0, 49.5, 47.0, 26.6 (2C), 26.5 (2C), 18.1; 19F NMR
(466 MHz, CDCl3) δ −126.06 (4F), −123.35 (4F), −122.78 (4F),
−121.79 (4F), −114.46 (4F), −80.72 (6F); HRMS [FAB+] m/z calcd
for C34H23F26NO4 1003.1212, found 1003.1248.
Scheme 7. Intramolecular Cyclization of 9a−h
Fmoc-Cys-S-Tr-OAllyl.10b HOBt·H2O (288 mg, 1.88 mmol) and
HBTU (712 mg, 1.88 mmol) were added to a solution of N-Fmoc-S-
trityl-L-cysteine (1.00 g, 1.71 mmol) in DMF (10 mL) at room
temperature. After 15 min, allyl alcohol (232 μL, 3.41 mmol) and
DIEA (614 μL, 3.76 mmol) were added separately, and the resulting
mixture was stirred at the same temperature for 24 h. The reaction
mixture was diluted with ethyl acetate and washed with H2O, saturated
aq NaHCO3, and brine, and the organic layer was dried over Na2SO4
and then concentrated. The crude residue was purified by silica gel
chromatography (EtOAc/hexane =1:3) to give the title compound
a
Reagents and conditions: (a) excess DEA, MeCN, rt, 30 min; (b) 10
mol % Pd(OAc)2, 0.8 equiv of PS-PPh3, 2.0 equiv of PhSiH3, CH2Cl2,
rt, 30 min; (c) 5.0−20.0 equiv of fluorous tagged Mukaiyama reagent
10, DMAP, TEA, DMF, 50 °C, 24 h, high dilution conditions; 1a,
48%; 1b, 71%; 1c, 70%; 1d, 51%; 1e, 41%; 1f, 49%; 1g, 43%; 1h, 48%
(three steps).
1
(1.07 g, quant): white solid; mp 70−72 °C; H NMR (270 MHz,
CDCl3) δ 7.76 (d, J = 7.3 Hz, 2H), 7.60 (d, J = 7.0 Hz, 2H), 7.41−7.38
(m, 8H), 7.33−7.18 (m, 11H), 5.95−5.80 (m, 1H), 5.33−5.23 (m,
3H), 4.62 (d, J = 5.1 Hz, 2H), 4.38−4.35 (m, 3H), 4.23 (t, J = 7.0 Hz,
1H), 2.68- 2.64 (m, 2H).
unnatural isomers of tenuecyclamide A obtained by this work
(1c−h) will be reported in due course.
f4/f6-Fmoc-Ala-Cys-S-Tr-OAllyl. To a solution of N-Fmoc-S-trityl-L-
cysteine-OAllyl (171 mg, 0.274 mmol) in MeCN (4 mL) was added
DEA (4 mL) at room temperature. The mixture was stirred at the
same temperature for 30 min. After concentration in vacuo, the
reaction mixture was azeotroped to dryness with MeCN (3 × 3 mL).
In another flask, to a solution mixture of f4-Fmoc-L-alanine (100 mg,
0.124 mmol) and f6-Fmoc-D-alanine (125 mg, 0.124 mol) in DMF (4
mL) were added HOBt·H2O (42 mg, 0.274 mmol) and HBTU (104
mg, 0.274 mol) at room temperature. After 15 min, a solution of the
above residue in DMF (2 mL) and DIEA (88 μL, 0.523 mmol) was
added. The reaction mixture was stirred at room temperature for 12 h.
After the addition of 1.0 M HCl aq and then dilution with ethyl
acetate, the organic layer was washed with H2O, saturated aq NaHCO3
and brine, dried over Na2SO4, and concentrated. The crude residue
was purified by FSPE. A column was packed with fluorous silica gel
(4.98 g) using 55% aq THF as the solvent. The crude reaction mixture
was then loaded onto this column and eluted with 43 mL of 55% aq
THF followed by 65 mL of THF. Evaporation of the THF fraction by
vacuum centrifuge gave the title product (304 mg, 95%): pale yellow
EXPERIMENTAL SECTION
■
General Experimental Methods. Except as otherwise indicated,
all reactions were carried out under a positive pressure of nitrogen. All
the laboratory chemicals were purchased and used without
purification. Solvents were removed at a heating bath temperature of
40 °C and reduced pressure by rotary evaporation. Nonvolatile
compounds were dried in vacuo at 0.01 mbar. All reactions were
magnetically stirred and monitored by thin layer chromatography
(TLC) using silica gel plates. Purification by chromatography was
performed on silica gel (65−210 Å). NMR spectra were recorded at
270 MHz (1H), 67.8 MHz (13C), and 466 MHz (19F), respectively.
Chemical shifts δ are referred in terms of ppm, and J-coupling constants
are given in Hz. Abbreviations for multiplicity are as follows: s
(singlet), d (doublet), t (triplet), q (quadruplet), quin (quintet signal),
m (multiplet), br (broad signal). HPLC analysis and separations were
performed on FluoroFlash columns (20 mm i.d., 250 mm length and
4.6 mm i.d., 150 mm length), and the oligopeptide analogue were
purified by HPLC with UV/vis detector. High-resolution mass spectra
were measured by FAB and EI using a magnetic sector analyzer. High-
resolution FAB and EI mass spectra were calibrated with Ultramark
1621 and PFK, respectively, prior to data acquisition. Optical rotations
were measured at a Na D-line (λ = 589 nm) using a 100 mm cell.
f4-Fmoc-L-alanine 3a. A solution of f4-Fmoc-OSu (1.20 g, 1.45
mmol) in MeCN (20 mL) was added to a solution of L-alanine (193
mg, 2.17 mmol) in 10% aq Na2CO3 (5 mL) slowly at 0 °C. After 5
min, the reaction mixture was warmed to room temperature and
stirred for 3 h. The reaction acidified with 1.0 M HCl aq to pH 2.0.
The organic layer was extracted with diethyl ether, dried over Na2SO4,
and concentrated. The residue was purified by column chromatog-
raphy on silica gel (CHCl3/MeOH = 50:1) to give f4-Fmoc-L-alanine
(1.11 g, 95%): white solid; mp 75−76 °C; [α]2D5 = +1.0 (c = 0.10,
CHCl3); 1H NMR (270 MHz, CDCl3) δ 7.67 (d, J = 7.3 Hz, 2H), 7.42
(s, 2H), 7.25 (d, J = 7.8 Hz, 2H), 5.27 (d, J = 7.3 Hz, 1H), 4.44−4.39
(m, 3H), 4.18 (t, J = 6.9 Hz, 1H), 3.02−2.96 (m, 4H), 2.51−2.32 (m,
4H), 1.49 (d, J = 6.5 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ 177.9,
155.8, 144.6, 144.3, 139.7 (2C), 138.1 (2C), 127.9 (2C), 125.0 (2C),
124.9 (2C), 120.1−111.0 (8C), 67.0, 49.5, 47.0, 26.5 (4C), 18.5; 19F
NMR (466 MHz, CDCl3) δ −125.87 (4F), −124.27 (4F), −114.66
(4F), −80.91 (6F); HRMS [FAB+] m/z calcd for C30H23F18NO4
803.1340, found 803.1418.
1
solid; mp 68−74 °C; H NMR (270 MHz, CDCl3) δ 7.67 (d, J = 8.1
Hz, 4H), 7.42−7.37 (m, 16H), 7.28−7.15 (m, 22H), 6.45 (d, J = 5.4
Hz, 1H), 6.16 (d, J = 7.8 Hz, 1H), 5.94−5.21 (m, 2H), 5.44−5.21 (m,
6H), 4.59−4.51 (m, 6H), 4.41−4.35 (m, 6H), 4.16 (t, J = 6.5 Hz, 2H),
3.00−2.95 (m, 8H), 2.78−2.58 (m, 4H), 2.49−2.33 (m, 8H), 1.49−
1.26 (m, 6H); 19F NMR (466 MHz, CDCl3) δ −126.00 (4F), −125.85
(4F), −124.23 (4F), −123.29 (4F), −122.72 (4F), −121.75 (4F),
−114.65 (4F), −114.38 (4F), −80.92 (6F), −80.66 (6F).
f4/f6-Fmoc-thiazoline Ring Unit. To a solution of triphenylphos-
phine oxide (388 mg, 1.40 mmol) in dry CH2Cl2 (3 mL) was added
trifluoromethanesulfonic anhydride (0.11 mL, 0.698 mmol) slowly at 0
°C under N2. After the reaction mixture was stirred at 0 °C for 10 min,
a solution of f4/f6-Fmoc-alanine-S-trityl-L-cysteine-OAllyl (400 mg,
0.155 mmol) in dry CH2Cl2 (3 mL) was added. The reaction mixture
was stirred at 0 °C for 2 h. The reaction was quenched with saturated
aq NaHCO3. The organic layer was washed with H2O, dried over
Na2SO4, and concentrated. The crude residue was purified by FSPE. A
column was packed with fluorous silica gel (22.1 g) using 70% aq
MeOH as the solvent. The crude reaction mixture was then loaded
onto this column and eluted with 120 mL of 70% aq MeOH followed
by 80 mL of THF. The evaporation of the THF fraction by vacuum
centrifuge gave the title product (287 mg, 90%): white solid; mp 68−
72 °C; 1H NMR (270 MHz, CDCl3) δ 7.68 (d, J = 8.1 Hz, 4H), 7.44
(s, 4H), 4.32−7.23 (m, 4H), 5.99−5.85 (m, 2H), 5.64 (d, J = 6.8 Hz,
1H), 5.55 (d, J = 7.0 Hz, 1H), 5.39- 5.29 (m, 2H), 5.27−5.23 (m, 2H),
5.18−5.10 (m, 2H), 4.69−4.68 (m, 6H), 4.40 (m, 4H), 4.19 (t, J = 7.0
Hz, 2H), 3.63−3.55 (m, 4H), 3.02−2.96 (m, 8H), 2.52−2.35 (m, 8H),
1.49 (d, J = 4.6 Hz, 6H); 19F NMR (466 MHz, CDCl3) δ −125.99
f6-Fmoc-D-alanine 3b. f6-Fmoc-D-alanine was synthesized from f6-
Fmoc-OSu and D-alanine in 96% yield as a white solid by following the
procedure used for the synthesis of f4-Fmoc-L-alanine 3a: white solid;
mp 112−113 °C; [α]2D5 = −1.9 (c = 0.10, CHCl3); H NMR (270
1
MHz, CDCl3) δ 7.67 (d, J = 7.3 Hz, 2H), 7.42 (s, 2H), 7.25 (d, J = 7.3
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(4F), −125.83 (4F), −124.22 (4F), −123.29 (4F), −122.71 (4F),
−121.71 (4F), −114.66 (4F), −114.43 (4F), −80.89 (6F), −80.62
(6F).
by vacuum centrifuge gave 6a as colorless oil, which was used without
purification (2.3 mg, quant): colorless oil; [α]2D5 = −13.3 (c = 0.265,
CHCl3); 1H NMR (270 MHz, CDCl3) δ 8.12 (s, 1H), 6.11−5.97 (m,
1H), 5.41 (dd, J = 17.0 Hz, 1.4 Hz, 1H), 5.30 (dd, J = 10.5 Hz, 1.4 Hz,
1H), 4.86 (dd, J = 5.9 Hz, 1.4 Hz, 2H), 4.50 (q, J = 6.8 Hz, 1H), 1.86
(s, 2H), 1.55 (d, J = 6.5 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ
179.9, 161.1, 146.6, 131.9, 127.4, 118.8, 65.9, 49.7, 24.8; HRMS [FAB
+] m/z calcd for C9H13N2O2S 213.0698, found 213.0709.
Compound 5a and 5b. A solution of ring-closing product (198 mg,
0.0962 mmol) and activated MnO2 (1982 mg, 10 w/w) in dry CHCl3
(10 mL) was stirred at 60 °C for 30 min under N2. After MnO2 was
removed by filtration through a pad of Celite and washed with CHCl3,
the filtrate was concentrated. The crude residue was purified by silica
gel chromatography (EtOAc/hexane = 1/2) to give 5a and 5b (148
mg, 75%): white solid; mp 94−104 °C; 1H NMR (270 MHz, CDCl3)
δ 8.09 (s, 2H), 7.66 (d, J = 7.3 Hz, 4H), 7.43 (s, 4H), 7.23 (d, J = 7.8
Hz, 4H), 6.10−5.95 (m, 2H), 5.67 (d, J = 5.9 Hz, 2H), 5.40 (dd, J =
17.0, 1.4 Hz, 2H), 5.29 (dd, J = 10.5, 1.4 Hz, 2H), 5.19 (quin, J = 6.6
Hz, 2H), 4.84 (d, J = 5.9 Hz, 4H), 4.52−4.37 (m, 4H), 4.16 (t, J = 5.5
Hz, 2H), 3.01−2.95 (m, 8H), 2.51−2.32 (m, 8H), 1.65 (d, J = 5.9 Hz,
6H); 19F NMR (466 MHz, CDCl3) δ −126.00 (4F), −125.85 (4F),
−124.22 (4F), −123.26 (4F), −122.69 (4F), −122.0 (4F), −114.73
(4F), −114.47 (4F), −80.88 (6F), −80.63 (6F). Fluorous analytical
HPLC (flow rate; 1.0 mL/min; 0 to 30 min: 80% MeCN to 100%
MeCN; 30 to 60 min: 100% MeCN): tR = 22.0 min (f4-Fmoc-(S)-
thiazole), 41.9 min (f6-Fmoc-(R)-thiazole).
Preparative Separation of 5a and 5b. The preparative separation
of 5a and 5b was carried out on a SHIMADZU HPLC system.
Compounds 5a and 5b were dissolved in THF (2.0 mL) and filtered
through a syringe filter (0.45 μm pore size) prior to injection. The
separation was carried out on a FluoroFlash HPLC column, 20 mm
i.d., 250 mm length. The separation was achieved by gradient elution
with 80% MeCN/H2O up to 100% MeCN in 30.0 min, followed by
isocratic elution with 100% MeCN over 60 min with a constant flow
rate of 7.0 mL/min. A UV detector (254 nm) was used to manually
identify the peaks. 5a and 5b (40 mg) was injected chromatographic
run. The yield of the demixing was 80%, and the following two
compounds were isolated: f4-Fmoc-(S)-thiazole-ring unit 5a (14 mg)
and f6-Fmoc-(R)-thiazole-ring unit 5b (18 mg).
f4-Fmoc-(S)-thiazole-ring unit 5a: white solid; mp 110−111 °C;
[α]2D5= −5.64 (c = 0.13, CHCl3); 1H NMR (270 MHz, CDCl3) δ 8.10
(s, 1H), 7.68 (d, J = 7.8 Hz, 2H), 7.43 (s, 2H), 7.25 (d, J = 7.3 Hz,
2H), 6.11−5.96 (m, 1H), 5.55 (d, J = 5.1 Hz, 1H), 5.41 (dd, J = 17.3,
1.4 Hz, 1H), 5.30 (dd, J = 10.5, 1.4 Hz, 1H), 5.22−5.17 (m, 1H), 4.85
(d, J = 5.9 Hz, 2H), 4.52−4.39 (m, 2H), 4.18 (t, J = 6.2 Hz, 1H),
3.02−2.96 (m, 4H), 2.51−2.32 (m, 4H), 1.66 (d, J = 5.9 Hz, 3H); 13C
NMR (67.8 MHz, CDCl3), δ 173.7, 160.9, 155.8, 147.0 (2C), 144.6
(2C), 144.3 (2C), 139.8 (2C), 138.1, 131.9 (2C), 128.0, 127.5, 125.1,
125.0, 120.2−110.1 (9C), 67.0, 66.1, 49.4, 47.2, 26.6 (2C), 22.6 (3C);
19F NMR (466 MHz, CDCl3) δ −125.85 (4F), −124.21 (4F),
−114.59 (4F), −80.86 (6F); HRMS [FAB+] m/z calcd for
C36H29F18N2O4S 927.1562, found 927.1513.
Compound 6b. Compound 6b was prepared from f6-Fmoc-(R)-
thiazole-ring unit 5b in quantitative yield as colorless oil by following
the procedure used for prepare of 6a: colorless oil; [α]2D5 = +9.86 (c =
1
0.191, CHCl3); H NMR (270 MHz, CDCl3) δ 8.12 (s, 1H), 6.12−
5.97 (m, 1H), 5.41 (dd, J = 17.3 Hz, 1.4 Hz, 1H), 5.29 (dd, J = 10.5
Hz, 1.4 Hz, 1H), 4.85 (dd, J = 5.9 Hz, 1.4 Hz, 2H), 4.50 (q, J = 6.8 Hz,
1H), 1.86 (s, 2H), 1.55 (d, J = 6.5 Hz, 3H); 13C NMR (67.8 MHz,
CDCl3) δ 179.9, 161.1, 146.6, 131.9, 127.4, 118.8, 65.9, 49.7, 24.8;
HRMS [FAB+] m/z calcd for calcd for C9H13N2O2S 213.0698, found
213.0688.
f4/f6-Fmoc-Ala-Thr-OBn. A solution of f4-Fmoc-L-alanine (6.00 g,
7.47 mmol) and f6-Fmoc-D-alanine (7.47 g, 7.47 mmol) in DMF (120
mL) was treated with HOBt·H2O (5.37 g, 35.10 mmol) and HBTU
(13.31 g, 35.10 mmol) at room temperature. After 15 min, H-Thr-
OBn·(COOH)2 (4.92 g, 16.43 mmol) and DIEA (5.3 mL, 31.36
mmol) were added to the above reaction mixture. The reaction
mixture was stirred at room temperature for 24 h. After the addition of
1.0 M HCl aq and then dilution with ethyl acetate, the organic layer
was washed with H2O, saturated aq NaHCO3, and brine, dried over
Na2SO4, and concentrated. The crude residue was pass through a short
pad of silica gel (EtOAc/hexane =1:3) to obtain the title compounds
1
(15.80 g, 94%): pale yellow solid; mp 115−118 °C; H NMR (270
MHz, CDCl3) δ 7.67 (d, J = 7.3 Hz, 4H), 7.42 (s, 4H), 7.32−7.23 (m,
16H), 6.80−6.65 (m, 2H), 5.47−5.42 (m, 2H), 5.23−5.12 (m, 4H),
4.64 (dd, J = 8.4, 1.9 Hz, 2H), 4.40−4.38 (m, 6H), 4.16 (t, J = 6.9 Hz,
2H), 3.02−2.96 (m, 8H), 2.52−2.32 (m, 8H), 2.18−2.02 (m, 2H),
1.46−1.39 (m, 6H), 1.21−1.18 (m, 6H); 19F NMR (466 MHz,
CDCl3) δ −126.04 (4F), −125.89 (4F), −124.25 (4F), −123.30 (4F),
−122.74 (4F), −121.77 (4F), −114.69 (4F), −114.46 (4F), −80.89
(6F), −80.65 (6F).
f4/f6-Fmoc-Ala-Thr(CO)-OBn. Dess−Martin periodinane (155
mg, 0.0365 mol) was added to a solution of f4/f6-Fmoc-dipeptides
(100 mg, 0.0457 mmol) in CH2Cl2 (4 mL). The reaction mixture was
stirred at 40 °C for 2 h. The reaction was quenched with saturated aq
sodium hyposulfite/saturated aq NaHCO3 = 1/1 v/v. The organic
layer was washed with saturated aq NaHCO3, dried over Na2SO4, and
concentrated. The crude residue was passed through a short pad of
silica gel to obtain the title compounds (99 mg, 98%): pale yellow
1
solid; mp 86−88 °C; H NMR (270 MHz, CDCl3) δ 7.68 (d, J = 7.3
Hz, 4H), 7.42 (s, 4H), 7.34−7.33 (m, 12H), 7.25 (d, J = 7.6 Hz, 4H),
5.33−5.15 (m, 8H), 4.44−4.39 (m, 6H), 4.18 (t, J = 6.2 Hz, 2H),
3.03−2.96 (m, 8H), 2.52−2.30 (m, 14H), 1.41 (t, J = 7.3 Hz, 6H); 19F
NMR (466 MHz, CDCl3) δ −126.02 (4F), −125.88 (4F), −124.24
(4F), −123.29 (4F), −122.73 (4F), −121.74 (4F), −114.70 (4F),
−114.42 (4F), −80.87 (6F), −80.65 (6F).
f6-Fmoc-(R)-thiazole-ring unit 5b: white solid; mp 75−76 °C;
[α]2D5 = +8.24 (c = 0.12, CHCl3); 1H NMR (270 MHz, CDCl3) δ 8.10
(s, 1H), 7.68 (d, J = 8.1 Hz, 2H), 7.43 (s, 2H), 7.25 (d, J = 7.8 Hz,
2H), 6.10−5.96 (m, 1H), 5.57 (d, J = 6.5 Hz, 1H), 5.41 (dd, J = 17.0,
1.4 Hz, 1H), 5.30 (dd, J = 10.5, 1.4 Hz, 1H), 5.22−5.17 (m, 1H), 4.85
(d, J = 5.9 Hz, 2H), 4.52−4.39 (m, 2H), 4.17 (d, J = 6.6 Hz, 1H),
3.02−2.96 (m, 4H), 2.51−2.39 (m, 4H), 1.66 (d, J = 5.9 Hz, 3H); 13C
NMR (67.8 MHz, CDCl3), δ 173.6, 160.8, 155.5, 147.0 (2C), 144.5
(2C), 144.4 (2C), 139.7 (2C), 138.1, 131.8 (2C), 128.0, 127.4, 125.0,
124.9, 120.1−108.2 (13C), 66.9, 66.0, 49.3, 47.1, 26.6 (2C), 22.7
(2C), 21.1; 19F NMR (466 MHz, CDCl3) δ −125.99 (4F), −123.28
(4F), −122.71 (4F), −121.70 (4F), −114.38 (4F), −80.66 (6F);
HRMS [FAB+] m/z calcd for C40H29F26N2O4S 1127.1435, found
1127.1387.
Compound 6a. Diethylamine (2 mL) was added to a solution of f4-
Fmoc-(S)-thiazole-ring unit 5a (10 mg, 0.0108 mmol) in MeCN (2
mL), and the reaction mixture was stirred at room temperature for 30
min. After concentration in vacuo, the reaction mixture was
azeotroped to dryness with MeCN (3 × 3 mL). The crude residue
was purified by FSPE. A column was packed with fluorous silica gel
(0.32 g) using 50% aq THF as the solvent. The crude reaction mixture
was then loaded onto this column and eluted with 2 mL of 50% aq
THF followed by 4 mL of THF. The evaporation of the THF fraction
f4/f6-Fmoc-oxazole Ring Unit. To a solution of triphenylphosphine
oxide (18.33 g, 65.89 mmol) in dry CH2Cl2 (120 mL) was added
trifluoromethanesulfonic anhydride (5.1 mL, 32.95 mmol) slowly at 0
°C under N2. After the reaction mixture was stirred at same
temperature for 10 min, a solution of f4/f6-Fmoc-oxidative products
(8.00 g, 3.66 mmol) in dry CH2Cl2 (60 mL) was added to the above
reaction mixture. The reaction mixture was stirred at 0 °C for 2 h. The
reaction was quenched with saturated aq NaHCO3. The organic layer
was washed with H2O, dried over Na2SO4, and concentrated. The
crude residue was purified by FSPE. A column was packed with
fluorous silica gel (405.0 g) using 55% aq THF as the solvent. The
crude reaction mixture was then loaded onto this column and eluted
with 1020 mL of 55% aq THF followed by 1120 mL of THF. The
evaporation of the THF fraction by vacuum centrifuge gave the title
1
compounds (7.95 g, quant): white solid; mp 81−84 °C; H NMR
(270 MHz, CDCl3) δ 7.67 (d, J = 7.8 Hz, 4H), 7.44−7.23 (m, 18H),
5.53 (d, J = 7.8 Hz, 2H), 5.36 (s, 4H), 5.00 (quin, J = 6.4 Hz, 2H),
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4.42 (d, J = 6.5 Hz, 4H), 4.16 (t, J = 6.9 Hz, 2H), 3.02−2.96 (m, 8H),
2.58 (s, 6H), 2.51−2.32 (m, 8H), 1.56−1.55 (m, 6H); 19F NMR (466
MHz, CDCl3) δ −126.01 (4F), −125.87 (4F), −124.25 (4F), −123.32
(4F), −122.72 (4F), −121.75 (4F), −114.67 (4F), −114.47 (4F),
−80.87 (6F), −80.63 (6F).
FSPE. A short column was packed with fluorous silica gel (7.2 g) using
50% aq THF as the solvent. The crude reaction mixture was then
loaded onto this column and eluted with 120.0 mL of 50% aq THF
followed by 25.0 mL of THF. The evaporation of the THF fraction by
vacuum centrifuge gave product 9bc-mix as a colorless semisolid (202
mg, 85%). Fluorous analytical HPLC (flow rate; 1.0 mL/min; 0 to 7.5
min: 80% MeCN/H2O to 100% MeCN; 7.5 to 30 min: 100% MeCN):
tR =10.8 min (9b), 23.8 min (9c).
Compound 9ad-mix. Compound 9ad-mix was synthesized from
8a-mix and 6b in 80% yield as a colorless semisolid by following the
procedure used for the synthesis of 9bc-mix. Fluorous analytical
HPLC (flow rate; 1.0 mL/min; 0 to 7.5 min: 80% MeCN/H2O to
100% MeCN; 7.5 to 30 min: 100% MeCN): tR = 14.9 min (9a), 23.8
min (9d).
Compound 9ef-mix. Compound 9ef-mix was synthesized from 8b-
mix and 6a in 87% yield as a colorless semisolid by following the
procedure used for the synthesis of 9bc-mix. Fluorous analytical
HPLC (flow rate; 1.0 mL/min; 0 to 7.5 min: 80% MeCN/H2O to
100% MeCN; 7.5 to 30 min: 100% MeCN): tR = 15.5 min (9e), 23.1
min (9f).
Compound 9gh-mix. Compound 9gh-mix was synthesized from
8b-mix and 6b in 82% yield as a colorless semisolid by following the
procedure used for the synthesis of 9bc-mix. Fluorous analytical
HPLC (flow rate; 1.0 mL/min; 0 to 7.5 min: 80% MeCN/H2O to
100% MeCN; 7.5 to 30 min: 100% MeCN): tR = 15.1 min (9g), 23.1
min (9h).
Compound 7-mix. The ring closing product (f4/f6-Fmoc-oxazole-
ring unit) (1.00 g, 0.465 mmol) and Pd/C (11 mg, 0.0930 mmol) in
THF/MeOH (10 mL, 1/1 v/v) was stirred at room temperature for 2
h under H2 balloon. The catalyst was removed by filtration through a
pad of Celite and washed with ethyl acetate. The filtrate was
concentrated to give 7-mix (865 mg, 94%). The residue was
immediately used for the next step without further purification:
colorless oil; 1H NMR (270 MHz, CDCl3) δ 7.66 (d, J = 7.8 Hz, 4H),
7.44−7.41 (m, 4H), 7.23 (d, J = 7.0 Hz, 4H), 5.07−5.02 (m, 2H), 4.40
(d, J = 6.5 Hz, 4H), 4.15 (t, J = 6.6 Hz, 2H), 3.00−2.94 (m, 8H), 2.62
(s, 6H), 2.51−2.25 (m, 8H), 1.58 (d, J = 7.3 Hz, 6H); 19F NMR (466
MHz, CDCl3) δ −126.01 (4F), −125.86 (4F), −124.24 (4F), −123.29
(4F), −122.73 (4F), −121.74 (4F), −114.63 (4F), −114.41 (4F),
−80.91 (6F), −80.65 (6F).
Compound 8a-mix. A solution of 7-mix (250 mg, 0.127 mmol) in
DMF (4 mL) was treated with HOBt·H2O (91 mg, 0.597 mmol) and
HBTU (226 mg, 0.597 mmol) at room temperature. After 15 min, a
solution of 6a (67 mg, 0.305 mmol) in DMF (4 mL) and DIEA (91
μL, 0.55 mmol) were sequentially added to the above solution. The
reaction mixture was stirred at room temperature for 12 h. After the
addition of 1.0 M HCl aq and dilution with ethyl acetate, the organic
layer was washed with H2O, saturated aq NaHCO3, and brine, dried
over Na2SO4, and concentrated. The crude residue was purified by
FSPE. A short column was packed with fluorous silica gel (5.1 g) using
55% aq THF as the solvent. The crude reaction mixture was then
loaded onto this column and eluted with 62.9 mL of 55% aq THF
followed by 25.0 mL of THF. The evaporation of the THF fraction by
vacuum centrifuge gave product 8a-mix (272 mg, 91%): white solid;
mp 123−126 °C; 1H NMR (270 MHz, CDCl3) δ 8.10−8.07 (m, 2H),
7.70−7.67 (m, 4H), 7.47−7.43 (m, 6H), 7.26−7.24 (m, 4H), 6.09−
5.95 (m, 2H), 5.57−5.52 (m, 2H), 5.43−5.27 (m, 6H), 5.05−4.94 (m,
2H), 4.85−4.82 (m, 4H), 4.46−4.44 (m, 4H), 4.21 (t, J = 6.3 Hz, 2H),
3.01−2.96 (m, 8H), 2.62 (s, 6H), 2.51−2.32 (m, 8H), 1.75−1.71 (m,
6H), 1.62−1.53 (m, 6H); 19F NMR (466 MHz, CDCl3) δ −125.99
(4F), −125.86 (4F), −124.25 (4F), −123.32 (4F), −122.72 (4F),
−121.73 (4F), −114.64 (4F), −114.42 (4F), −80.87 (6F), −80.65
(6F).
Preparative Separation of 9bc-mix. Compound 9bc-mix was
dissolved in THF (1.5 mL) and filtered through a syringe filter (0.45
μm pore size) prior to injection. The separation was carried out on a
FluoroFlash HPLC column, 20 mm i.d., 250 mm length. The
separation was achieved by gradient elution with 80% MeCN/H2O up
to 100% MeCN in 7.5 min, followed by isocratic elution with 100%
MeCN over 60 min with a constant flow rate of 7.0 mL/min. A UV
detector (254 nm) was used to manually identify the peaks. 9bc-mix
(78 mg) was injected for the chromatographic run. The yield of the
demixing was quantitative yield and the following two compounds
were isolated: 9b (30 mg) and 9c (48 mg).
Compound 9b: colorless semisolid; [α]2D5 = −11.2 (c = 0.2,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.10 (s, 1H), 8.00 (s, 1H),
7.87 (d, J = 8.6 Hz, 1H), 7.67 (d, J = 8.1 Hz, 2H), 7.43 (d, J = 6.5 Hz,
2H), 7.32 (d, J = 7.8 Hz, 1H), 7.25 (d, J = 8.6 Hz, 2H), 6.09−5.94 (m,
1H), 5.59 (quin, J = 7.2 Hz, 1H), 5.50 (quin, J = 7.3 Hz, 1H), 5.42−
5.26 (m, 3H), 4.99 (quin, J = 5.9 Hz, 1H), 4.83 (d, J = 5.9 Hz, 2H),
4.45−4.43 (m, 2H), 4.18 (t, J = 6.5 Hz, 1H), 3.01−2.95 (m, 4H), 2.62
(s, 3H), 2.51−2.31 (m, 4H), 1.79 (d, J = 7.3 Hz, 3H), 1.70 (d, J = 6.8
Hz, 3H), 1.57 (d, J = 6.5 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ
173.2, 173.0, 161.3, 161.2, 161.1, 160.9, 160.8, 160.4 (2C), 155.4,
154.2, 149.0, 146.7 (2C), 144.6, 144.2, 139.6 (2C), 138.0 (2C), 131.8
(2C), 128.4 (2C), 127.9, 127.7 (2C), 124.9, 124.0, 120.1−110.4 (9C),
66.9, 65.9, 47.1 (2C), 46.7, 45.2, 26.5 (2C), 20.9 (3C), 11.6; 19F NMR
(466 MHz, CDCl3) δ −125.85 (4F), −124.27 (4F), −114.61 (4F),
−80.85 (6F); HRMS [FAB+] m/z calcd for C49H43F18N6O7S2
1233.2347, found 1233.2366.
Compound 8b-mix. Compound 8b-mix was synthesized from 7-
mix and 6b in 98% yield as a white solid by following the procedure
1
used for the synthesis of 8a-mix: white solid; mp 121−124 °C; H
NMR (270 MHz, CDCl3) δ 8.10−8.07 (m, 2H), 7.68 (d, J = 8.1 Hz,
4H), 7.46−7.43 (m, 6H), 7.25 (d, J = 6.8 Hz, 4H), 6.10−5.94 (m,
2H), 5.54 (quin, J = 7.1 Hz, 2H), 5.43−5.26 (m, 6H), 4.99 (quin, J =
6.4 Hz, 2H), 4.85−4.81 (m, 4H), 4.46−4.44 (m, 4H), 4.20 (t, J = 6.6
Hz, 2H), 3.02−2.96 (m, 8H), 2.61 (s, 6H), 2.50−2.31 (m, 8H), 1.73
(d, J = 6.8 Hz, 6H), 1.53 (d, J = 7.8 Hz, 6H); 19F NMR (466 MHz,
CDCl3) δ −126.04 (4F), −125.85 (4F), −124.24 (4F), −123.29 (4F),
−122.69 (4F), −121.71 (4F), −114.62 (4F), −114.39 (4F), −80.88
(6F), −80.64 (6F).
Compound 9c: colorless semisolid; [α]2D5 = +3.63 (c = 0.2, CHCl3);
1H NMR (270 MHz, CDCl3) δ 8.10 (s, 1H), 8.00 (s, 1H), 7.87 (d, J =
8.6 Hz, 1H), 7.67 (d, J = 8.1 Hz, 2H), 7.43 (d, J = 7.3 Hz, 2H), 7.32
(d, J = 8.6 Hz, 1H), 7.24 (d, J = 8.1 Hz, 2H), 6.09−5.94 (m, 1H), 5.59
(quin, J = 7.4 Hz, 1H), 5.50 (quin, J = 7.4 Hz, 1H), 5.42−5.26 (m,
3H), 4.99 (quin, J = 6.5 Hz, 1H), 4.83 (d, J = 5.9 Hz, 2H), 4.47−4.43
(m, 2H), 4.18 (t, J = 6.5 Hz, 1H), 3.01−2.95 (m, 4H), 2.62 (s, 3H),
2.50−2.31 (m, 4H), 1.79 (d, J = 7.3 Hz, 3H), 1.70 (d, J = 6.8 Hz, 3H),
1.57 (m, 3H); 13C NMR (67.8 MHz, CDCl3) δ 173.2, 173.0, 161.4,
161.3, 161.1, 160.9, 160.4 (2C), 155.4, 154.2, 149.0, 146.7 (2C), 144.6,
144.2, 139.7 (2C), 138.1 (2C), 131.8 (2C), 128.5 (2C), 127.9, 127.7
(2C), 124.9, 124.0, 120.2−108.2 (13C), 67.1, 65.9, 47.1, 46.7, 46.6,
45.2, 26.6 (2C), 21.0 (2C), 20.9, 11.6; 19F NMR (466 MHz, CDCl3) δ
−125.98 (4F), −123.31 (4F), −122.66 (4F), −121.68 (4F), −114.41
(4F), −80.61 (6F); HRMS [FAB+] m/z calcd for C53H43F26N6O7S2
1433.2221, found 1433.2236.
Compound 9bc-mix. A solution of 8a-mix (210 mg, 0.0891 mmol)
and Pd(PPh3)4 (21 mg, 0.0178 mmol) in dry CH2Cl2 (5 mL) was
stirred at room temperature for 10 min in N2. To the solution mixture
was added PhSiH3 (22 μL, 0.178 mmol), and then the mixture was
stirred at room temperature for 30 min. The mixture was through a
pad of Celite and washed CH2Cl2. The filtrate was concentrated. The
residue was immediately used for the next step without further
purification: A solution of the above residue in DMF (6 mL) was
treated with HOBt·H2O (64 mg, 0.419 mmol) and HBTU (159 mg,
0.419 mmol) at room temperature. After 15 min, a solution of 6a (45
mg, 0.214 mmol) in DMF (2 mL) and DIEA (63 μL, 0.374 mmol)
were sequentially added to the above solution at room temperature.
The mixture was stirred at the same temperature for 12 h. After the
addition of 1.0 M HCl aq and dilution with ethyl acetate, the organic
layer was washed with H2O, saturated aq NaHCO3, and brine, dried
over Na2SO4, and concentrated. The crude residue was purified by
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Preparative Separation of 9ad-mix. The preparative separation of
9ad-mix was carried out in the same manner as 9bc-mix. Compound
9ad-mix (223 mg) was injected for the chromatographic run. The
yield of the demixing was 96%, and the following two compounds
were isolated: 9a (102 mg) and 9d (113 mg).
47.0, 46.7, 46.6, 45.1, 26.6 (2C), 21.0 (2C), 20.9, 11.6; 19F NMR (466
MHz, CDCl3) δ −126.09 (4F), −123.35 (4F), −122.77 (4F), −121.78
(4F), −114.42 (4F), −80.75 (6F); HRMS [FAB+] m/z calcd for
C53H43F26N6O7S2 1433.2221, found 1433.2268.
Preparative Separation of 9gh-mix. The preparative separation of
9gh-mix was carried out in the same manner as for 9bc-mix.
Compound 9gh-mix (140 mg) was injected for the chromatographic
run. The yield of the demixing was 97%, and the following two
compounds were isolated: 9g (64 mg) and 9h (72 mg).
Compound 9a: colorless semisolid; [α]2D5 = −14.4 (c = 0.088,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.11 (s, 1H), 8.01 (s, 1H),
7.90 (d, J = 7.8 Hz, 1H), 7.67 (d, J = 7.8 Hz, 2H), 7.43 (d, J = 7.3 Hz,
2H), 7.35 (d, J = 7.8 Hz, 1H), 7.24 (d, J = 7.8 Hz, 2H), 6.10−5.94 (m,
1H), 5.59 (quin, J = 7.7 Hz, 1H), 5.50 (quin, J = 7.6 Hz, 1H), 5.43−
5.26 (m, 3H), 6.07−5.97 (m, 1H), 4.83 (d, J = 9.5 Hz, 2H), 4.51−4.39
(m, 2H), 4.18 (t, J = 6.2 Hz, 1H), 3.00−2.94 (m, 4H), 2.61 (s, 3H),
2.50−2.31 (m, 4H), 1.78 (d, J = 7.3 Hz, 3H), 1.70 (d, J = 6.5 Hz, 3H),
1.58 (d, J = 6.5 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ 173.1,
173.0, 161.3, 161.2, 161.1, 160.9, 160.4 (2C), 155.4, 154.2, 149.0,
146.7 (2C), 144.6, 144.3, 139.7, 139.6, 138.1 (2C), 131.8 (2C), 128.4
(2C), 127.9, 127.7 (2C), 124.9, 124.0, 120.0−110.9 (9C), 66.9, 65.9,
47.0, 46.7, 46.6, 45.2, 26.6 (2C), 26.5, 20.9 (2C), 11.6; 19F NMR (466
MHz, CDCl3) δ −125.85 (4F), −124.24 (4F), −114.63 (4F), −80.89
(6F); HRMS [FAB+] m/z calcd for C49H43F18N6O7S2 1233.2347,
found 1233.2344.
Compound 9g: colorless semisolid; [α]2D5 = −1.57 (c = 0.05,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.11 (s, 1H), 8.04 (s, 1H),
7.95 (d, J = 7.8 Hz, 1H), 7.68 (d, J = 7.3 Hz, 2H), 7.39−7.26 (m, 3H),
7.25 (d, J = 8.6 Hz, 2H), 6.09−5.93 (m, 1H), 5.60 (quin, J = 7.6 Hz,
1H), 5.50 (quin, J = 7.3 Hz, 1H), 5.43−5.26 (m, 3H), 4.99 (quin, J =
7.1 Hz, 1H), 4.83 (d, J = 5.4 Hz, 2H), 4.52−4.39 (m, 2H), 4.19 (t, J =
6.2 Hz, 1H), 3.00−2.95 (m, 4H), 2.62 (s, 3H), 2.50−2.31 (m, 4H),
1.76 (d, J = 6.4 Hz, 3H), 1.70 (d, J = 7.0 Hz, 3H), 1.56 (d, J = 6.5 Hz,
3H); 13C NMR (67.8 MHz, CDCl3) δ 173.2, 173.1, 161.3, 161.2,
161.0, 160.9, 160.4 (2C), 155.4, 154.2, 149.0, 146.7 (2C), 144.6, 144.3,
139.7 (2C), 138.1 (2C), 131.8 (2C), 128.4 (2C), 127.9, 127.7 (2C),
124.9, 124.0, 120.2−110.0 (9C), 66.9, 65.9, 47.1 (2C), 46.7, 45.2, 26.6,
26.5, 20.9 (3C), 11.6; 19F NMR (466 MHz, CDCl3) δ −125.84 (4F),
−124.24 (4F), −114.62 (4F), −80.86 (6F); HRMS [FAB+] m/z calcd
for C49H43F18N6O7S2 1233.2347, found 1233.2377.
Compound 9d: colorless semisolid; [α]2D5 = +6.92 (c = 0.095,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.12 (s, 1H), 8.01 (s, 1H),
7.86 (d, J = 8.1 Hz, 1H), 7.68 (d, J = 7.8 Hz, 2H), 7.43 (d, J = 6.8 Hz,
2H), 7.33 (d, 8.4 Hz, 1H), 7.25 (d, J = 8.1 Hz, 2H), 6.10−5.96 (m,
1H), 5.59 (quin, J = 7.3 Hz, 1H), 5.50 (quin, J = 7.3 Hz, 1H), 5.44−
5.27 (m, 3H), 5.00 (quin, J = 6.9 Hz, 1H), 4.85 (d, J = 5.9 Hz, 2H),
4.52−4.40 (m, 2H), 4.18 (t, J = 6.6 Hz, 1H), 3.01−2.95 (m, 4H), 2.63
(s, 3H), 2.51−2.30 (m, 4H), 1.79 (d, J = 7.3 Hz, 3H), 1.71 (d, J = 7.3
Hz, 3H), 1.57 (d, J = 7.3 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ
173.1, 173.0, 161.3, 161.2, 161.1, 160.9, 160.4 (2C), 155.4, 154.2,
149.0, 146.7 (2C), 144.6, 144.3, 139.7 (2C), 138.1 (2C), 131.8, 131.7,
128.5 (2C), 127.9, 127.7 (2C), 124.9, 124.0, 120.2−108.5 (13C), 66.9,
65.9, 47.1, 46.7, 46.6, 45.2, 26.6 (2C), 25.6, 20.9 (2C), 11.6; 19F NMR
(466 MHz, CDCl3) δ −125.98 (4F), −123.30 (4F), −122.70 (4F),
−121.72 (4F), −114.39 (4F), −80.64 (6F); HRMS [FAB+] m/z calcd
for C53H43F26N6O7S2 1433.2221, found 1433.2207.
Compound 9h: colorless semisolid; [α]2D5 = +12.4 (c = 0.087,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.10 (s, 1H), 8.00 (s, 1H),
7.88 (d, J = 8.6 Hz, 1H), 7.67 (d, J = 7.8 Hz, 2H), 7.43 (d, J = 6.8 Hz,
2H), 7.33 (d, J = 7.8 Hz, 1H), 7.25 (d, J = 7.8 Hz, 2H), 6.09−5.94 (m,
1H), 5.59 (quin, J = 7.3 Hz, 1H), 5.50 (quin, J = 7.3 Hz, 1H), 5.43−
5.26 (m, 3H), 5.00 (quin, J = 7.4 Hz, 1H), 4.83 (d, J = 6.5 Hz, 2H),
4.46−4.44 (m, 2H), 4.18 (t, J = 6.6 Hz, 1H), 3.01−2.95 (m, 4H), 2.62
(s, 3H), 2.51−2.32 (m, 4H), 1.79 (d, J = 7.3 Hz, 3H), 1.70 (d, J = 6.8
Hz, 3H), 1.57 (d, J = 7.0 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ
173.2, 173.1, 161.2 (2C), 161.1, 160.9, 160.4 (2C), 155.4, 154.2, 149.0,
146.7 (2C), 144.6, 1443, 139.7, 139.6, 138.1(2C), 131.8 (2C), 128.4
(2C), 127.9, 127.7 (2C), 124.9, 124.0, 120.2−107.1 (13C), 66.9, 65.9,
47.1, 46.7, 45.2, 43.4, 26.7 (2C), 21.0 (2C), 20.9, 11.6; 19F NMR (466
MHz, CDCl3) δ −125.97 (4F), −123.28 (4F), −122.71 (4F), −121.58
(4F), −114.45 (4F), −80.62 (6F); HRMS [FAB+] m/z calcd for
C53H43F26N6O7S2 1433.2221, found 1433.2218.
Preparative Separation of 9ef-mix. The preparative separation of
9ef-mix was carried out in the same manner as for 9bc-mix.
Compound 9ef-mix (90 mg) was injected for the chromatographic
run. The yield of the demixing was 98%, and the following two
compounds were isolated: 9e (42 mg) and 9f (46 mg).
Procedure for Cyclization Using 9a. Diethylamine (2 mL) was
added to a solution of 9a (28 mg, 0.0230 mmol) in MeCN (2 mL),
and the mixture was stirred at rt for 30 min. After concentration in
vacuo, the reaction mixture was azeotroped to dryness with MeCN (3
× 3 mL). Pd(OAc)2 (0.5 mg, 0.00230 mmol) and styrene polymer-
bound triphenylphosphene (11 mg, 1.7 mol/g, 0.0184 mmol) were
added to a flask containing dry CH2Cl2 (3 mL) under N2. After the
mixture was stirred for 15 min, a solution of the above residue in dry
CH2Cl2 (2 mL) and PhSiH3 (6 μL, 0.0461 mmol) were added
separately. The reaction progress was monitored by TLC, and the
reaction was complete in 30 min. The reaction mixture was passed
through a pad of Celite and concentrated. The residue was used in the
next step without further purification. The resulting amino acid
derivative was dissolved in DMF (16 mL) solution. This solution was
added to a flask containing fluorous tagged Mukaiyama reagent (373
mg, 0.461 mmol), TEA (192 μL, 1.38 mmol), and DMAP (23 mg,
0.184 mmol) in dry DMF (32 mL) at 50 °C over 8 h using a syringe
pump. After the addition was complete, the mixture was stirred at the
same temperature for 24 h. To the reaction mixture was added H2O
(12 mL) and then filtered. The filtrate was diluted with ethyl acetate
and washed with 1.0 M HCl aq, H2O, and brine. The organic layer was
dried over Na2SO4 and concentrated. The crude residue was purified
by short silica gel chromatography (EtOAc/hexane = 4/1) to obtain
1a as a colorless semisolid (5.1 mg, 48%).
Compound 9e: colorless semisolid; [α]2D5 = −14.9 (c = 0.06,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.12 (s, 1H), 8.04 (s, 1H),
7.88 (d, J = 7.8 Hz, 1H), 7.68 (d, J = 7.8 Hz, 2H), 7.43 (s, 2H), 7.32
(d, J = 7.8 Hz, 1H), 7.25 (d, J = 7.8 Hz, 2H), 6.08−5.97 (m, 1H), 5.59
(quin, J = 7.3 Hz, 1H), 5.50 (quin, J = 7.3 Hz, 1H), 5.43−5.26 (m,
3H), 4.99 (quin, J = 5.9 Hz, 1H), 4.84 (d, J = 4.6 Hz, 2H), 4.53−4.39
(m, 2H), 4.19 (t, J = 6.6 Hz, 1H), 3.00−2.95 (m, 4H), 2.62 (s, 3H),
2.50−2.34 (m, 4H), 1.78 (d, J = 6.5 Hz, 3H), 1.70 (d, J = 7.0 Hz, 3H),
1.56 (d, J = 6.5 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ 173.2,
173.1, 161.4, 161.2 (2C), 160.9, 160.4 (2C), 155.5, 154.3, 149.1, 146.9,
146.8, 144.6, 144.4, 139.7 (2C), 138.1 (2C), 131.9 (2C), 128.5 (2C),
128.0, 127.8 (2C), 125.0, 124.1, 120.2−110.4 (9C), 67.0, 66.0, 49.7,
46.8, 46.7, 45.2, 26.6 (2C), 21.0 (3C), 11.5; 19F NMR (466 MHz,
CDCl3) δ −125.88 (4F), −124.23 (4F), −114.58 (4F), −80.86 (6F);
HRMS [FAB+] m/z calcd for C49H43F18N6O7S2 1233.2347, found
1233.2397.
Compound 9f: colorless semisolid; [α]2D5 = +13.1 (c = 0.12,
1
CHCl3); H NMR (270 MHz, CDCl3) δ 8.12 (s, 1H), 8.01 (s, 1H),
7.87 (d, J = 7.8 Hz, 1H), 7.68 (d, J = 7.3 Hz, 2H), 7.43 (d, J = 6.8 Hz,
2H), 7.33 (d, J = 7.8 Hz, 1H), 7.25 (d, J = 7.3 Hz, 2H), 6.10−5.94 (m,
1H), 5.59 (quin, J = 7.2 Hz, 1H), 5.50 (quin, J = 7.3 Hz, 1H), 5.44−
5.27 (m, 3H), 5.00 (quin, J = 6.8 Hz, 1H), 4.84 (d, J = 5.1 Hz, 2H),
4.48−4.44 (m, 2H), 4.18 (t, J = 6.5 Hz, 1H), 3.01−2.95 (m, 4H), 2.62
(s, 3H), 2.51−2.32 (m, 4H), 1.79 (d, J = 6.5 Hz, 3H), 1.71 (d, J = 7.3
Hz, 3H), 1.58 (d, J = 8.1 Hz, 3H); 13C NMR (67.8 MHz, CDCl3) δ
173.2, 173.1, 161.4, 161.1, 161.0 (2C), 160.4 (2C), 155.9, 155.3, 149.0,
146.7 (2C), 144.6, 144.3, 139.7, 139.6, 138.1 (2C), 131.8 (2C), 128.4
(2C), 127.9, 127.7 (2C), 124.9, 124.0, 120.0−108.4 (13C), 66.9, 65.9,
Tenucyclamide A (1a):10a colorless semisolid; 1H NMR (270 MHz,
CDCl3) δ 8.69−8.68 (m, 2H), 8.61 (d, J = 6.8 Hz, 1H), 8.24 (s, 1H),
8.17 (s, 1H), 5.60 (quin, J = 6.6 Hz, 1 H), 5.37 (quin, J = 6.4 Hz, 1H),
5.14 (quin, J = 6.3 Hz, 1H), 2.70 (s, 3H), 1.80 (d, J = 6.8 Hz, 3H),
1.71−1.69 (m, 6H).
10270
dx.doi.org/10.1021/jo401647k | J. Org. Chem. 2013, 78, 10264−10272

The Journal of Organic Chemistry
Article
Tenuecyclamide B (1b).10a Compound 1b was synthesized from 9b
in 71% yield as a colorless semisolid by following the procedure used
for the synthesis of 1a: colorless semisolid; 1H NMR (270 MHz,
CDCl3) δ 8.70−8.66 (m, 2H), 8.60 (d, J = 7.8 Hz, 1H), 8.18 (s, 1H),
8.14 (s, 1H), 5.68 (quin, J = 7.0 Hz, 1H), 5.44 (quin, J = 6.6 Hz, 1H),
5.24 (quin, J = 6.9 Hz, 1H), 2.69 (s, 3H), 1.73−1.71 (m, 9H).
Compound 1c. Compound 1c was synthesized from 9c in 70%
yield as a colorless semisolid by following the procedure used for the
synthesis of 1a: colorless semisolid; [α]2D5 = −24.0 (c = 0.21, CHCl3);
1H NMR (270 MHz, CDCl3) δ 8.70 (d, J = 6.5 Hz, 1H), 8.65−8.60
(m, 2H), 8.20 (s, 1H), 8.17 (s, 1H), 5.58 (quin, J = 6.6 Hz, 1H), 5.42
(quin, J = 6.6 Hz, 1H), 5.21 (quin, J = 6.6 Hz, 1H), 2.69 (s, 3H), 1.79
(d, J = 6.8 Hz, 3H), 1.70−1.65 (m, 6H); 13C NMR (67.8 MHz,
CDCl3) δ 171.7, 170.6, 161.3, 160.7, 159.7, 159.6, 154.2, 149.2, 148.3,
128.6, 124.6, 123.1, 48.0, 47.9, 44.2, 24.7, 24.5, 19.9, 11.7; HRMS [EI
+] m/z calcd for C19H20N6O4S2 460.0987, found 460.0953.
AUTHOR INFORMATION
Corresponding Author
*E-mail: matsugi@meijo-u.ac.jp.
Notes
■
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
This work was partially supported by the Ministry of Education,
Science, Sports and Culture, Grant-in-aid for Scientific
Research (C) 23580154 and the Research Institute of Meijo
University. We thank Meiji Seika Pharma Co., Ltd. for funding
this work.
DEDICATION
■
Compound 1d. Compound 1d was synthesized from 9d in 51%
yield as a colorless semisolid by following the procedure used for the
synthesis of 1a: colorless semisolid; [α]2D5 = +40.3 (c = 0.29, CHCl3);
1H NMR (270 MHz, CDCl3) δ 8.68−8.66 (m, 2H), 8.56 (d, J = 6.8
Hz, 1H), 8.21 (s, 1H), 8.12 (s, 1H), 5.56 (quin, J = 6.5 Hz, 1H), 5.41
(quin, J = 6.6 Hz, 1H), 5.28 (quin, J = 6.8 Hz, 1H), 2.70 (s, 3H),
1.72−1.70 (m, 9H); 13C NMR (67.8 Hz, CDCl3) δ 171.7, 170.6,
161.3, 160.7, 159.7, 159.6, 154.2, 149.2, 148.3, 128.6, 124.6, 123.1,
48.0, 47.9, 44.2, 24.7, 24.5, 19.9, 11.7; HRMS [EI+] m/z calcd for
C19H20N6O4S2 460.0987, found 460.0953.
We dedicate this paper to Prof. Dennis P. Curran on the
occasion of his 60th birthday.
REFERENCES
■
(1) (a) Curran, D. P.; Hadida, S.; He, M. J. Org. Chem. 1997, 62,
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h, I., Curran, D. P.,
Compound 1e.10a Compound 1e was synthesized from 9e in 41%
yield as a colorless semisolid by following the procedure used for the
́
1
synthesis of 1a: colorless semisolid; H NMR (270 MHz, CDCl3) δ
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S. J.; Sefler, A. M. J. Am. Chem. Soc. 1998, 120, 4105. (b) Sakurai, K.;
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8.68−8.66 (m, 2H), 8.57 (d, J = 5.9 Hz, 1H), 8.21 (s, 1H), 8.12 (s,
1H), 5.56 (quin, J = 6.8 Hz, 1H), 5.41 (quin, J = 6.4 Hz, 1H), 5.28
(quin, J = 6.9 Hz, 1H), 2.70 (s, 3H), 1.72−1.68 (m, 9H).
Compound 1f. Compound 1f was synthesized from 9f in 49% yield
as a colorless semisolid by following the procedure used for the
synthesis of 1a: colorless semisolid; [α]2D5 = +2.4 (c = 0.144, CHCl3);
1H NMR (270 MHz, CDCl3) δ 8.69−8.67 (m, 2H), 8.61 (d, J = 6.5
Hz, 1H), 8.24 (s, 1H), 8.17 (s, 1H), 5.60 (quin, J = 6.8 Hz, 1H), 5.36
(quin, J = 6.3 Hz, 1H), 5.14 (quin, J = 6.1 Hz, 1H), 2.70 (s, 3H), 1.79
(d, J = 6.8 Hz, 3H), 1.73−1.64 (m, 6H); 13C NMR (67.8 Hz, CDCl3)
δ 172.2, 171.4, 161.9, 160.7, 160.0, 159.6, 153.6, 148.8, 148.6, 128.6,
124.9, 123.9, 48.3, 48.0, 44.9, 24.9, 23.7, 20.8, 11.6; HRMS [EI+] m/z
calcd for C19H20N6O4S2 460.0987, found 460.0980.
Compound 1g.10a Compound 1g was synthesized from 9g in 43%
yield as a colorless semisolid by following the procedure used for the
1
synthesis of 1a: colorless semisolid; H NMR (270 MHz, CDCl3) δ
8.70 (d, J = 5.9 Hz, 1H), 8.65−8.60 (m, 2H), 8.20 (s, 1H), 8.17 (s,
1H), 5.58 (quin, J = 6.6 Hz, 1H), 5.42 (quin, J = 6.6 Hz, 1H), 5.21
(quin, J = 6.6 Hz, 1H), 2.69 (s, 3H), 1.79 (d, J = 6.8 Hz, 3H), 1.70−
1.65 (m, 6H).
Compound 1h. Compound 1h was synthesized from 9h in 48%
yield as a colorless semisolid by following the procedure used for the
synthesis of 1a: colorless semisolid; [α]2D5 = +4.14 (c = 0.98, CHCl3);
1H NMR (270 MHz, CDCl3) δ 8.70−8.66 (m, 2H), 8.59 (d, J = 7.8
Hz, 1H), 8.18 (s, 1H), 8.13 (s, 1H), 5.67 (quin, J = 6.9 Hz, 1H), 5.44
(quin, J = 6.6 Hz, 1H), 5.24 (quin, J = 6.8 Hz, 1H), 2.69 (s, 3H),
1.73−1.68 (m, 9H); 13C NMR (67.8 Hz, CDCl3) δ 171.4, 170.9,
161.6, 160.5, 159.7, 159.4, 153.9, 148.9, 148.5, 128.4, 124.3, 123.5,
47.5, 47.3, 44.2, 25.0, 24.9, 20.8, 11.6; HRMS [EI+] m/z calcd for
C19H20N6O4S2 460.0987, found 460.0941.
́
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(b) Curran, D. P.; Dandapani, S.; Werner, S.; Matsugi, M. Synlett
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ASSOCIATED CONTENT
■
S
* Supporting Information
Proton, carbon, and fluorine NMR data for all new compounds
and fluorous HPLC chart of 5a, 5b, 9bc-mix, 9ad-mix, 9ef-mix,
and 9gh-mix. This material is available free of charge via the
Internet at http://pubs.acs.org.
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The Journal of Organic Chemistry
Article
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(11) Fluorous silica gel (FluoroFlash silica gel, 40 μm), which is
available from SIGMA-Aldrich, was used for all FSPE process.
(12) Preparative and analytical f-HPLC were conducted by
FluoroFlash columns (20 mm i.d., 250 mm length and 4.6 mm i.d.,
150 mm length) purchased from Fluorous Technologies, Inc. The
corporation is closing; however, fluorous column (Wakopak Fluofix-II
120E) having almost same separation ability is available from Wako
Pure Chemical Industries, Ltd.
(13) Although other condensation reagents such as PyBOP
((benzotriazol-1-yloxy) tripyrrolidinophosphonium hexafluorophos-
phate) or DPPA (diphenyl phosphorazidate) or DEPC (diethox-
yphosphoryl cyanide) or Yamaguchi reagent (2,4,6-trichlorobenzoyl
chloride) were examined under high dilution conditions (0.5 mM),
fluorous tagged Mukaiyama reagent 8 was the most effective for the
lactamization in terms of the yield and handling.
(14) Mukaiyama, T.; Usui, M.; Shimada, E.; Saigo, K. Chem. Lett.
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Kunda, Y.; Miyazaki, A.; Matsui, J.; Nakamura, S.; Hamamoto, H.;
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