HMDO-promoted peptide and protein synthesis in ionic liquids

Article abstract of DOI:10.1021/jo400797t

Hexamethyldisiloxane (HMDO) has been developed to efficiently promote the metal-free direct coupling of an amino function of one cysteine-free peptide or protein and a C-terminal thioester of the second peptide in ionic liquids. The amide-coupling reaction proceeds smoothly under mild conditions to afford the corresponding products in good to excellent yields (63-94%). Peptide couplings were also achieved using in-situ-generated thioesters by the thioesterification of oxo esters.

Full text of DOI:10.1021/jo400797t

Article
pubs.acs.org/joc
HMDO-Promoted Peptide and Protein Synthesis in Ionic Liquids
Jianli Duan,∥,† Yao Sun,∥,† Hao Chen,† Guofu Qiu,† Haibing Zhou,† Ting Tang,§ Zixin Deng,†
and Xuechuan Hong*,†,‡
†Key Laboratory of Combinatorial Biosynthesis and Drug Discovery (Wuhan University), Ministry of Education, and Wuhan
University School of Pharmaceutical Sciences, Wuhan, 430071, PR China
‡State Key Laboratory of Bioorganic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of
Sciences, Shanghai 200032, PR China
§College of Health Management, Hangzhou Normal University, Hangzhou, Zhejiang 310036, PR China
S
* Supporting Information
ABSTRACT: Hexamethyldisiloxane (HMDO) has been developed to efficiently promote the metal-free direct coupling of an
amino function of one cysteine-free peptide or protein and a C-terminal thioester of the second peptide in ionic liquids. The
amide-coupling reaction proceeds smoothly under mild conditions to afford the corresponding products in good to excellent
yields (63−94%). Peptide couplings were also achieved using in-situ-generated thioesters by the thioesterification of oxo esters.
associated with the need for a terminal cysteine. A more
general ligation technology to affect direct amidation of natural
or unnatural cysteine-free peptides under mild conditions
would greatly expand the utility of total protein synthesis.
Proteins containing a C-terminal thioester are very important
intermediates in protein modification. It is well-known that C-
INTRODUCTION
■
The development of methods for reliable, chemoselective
conjugation of synthetic molecules to peptides and proteins is
an important pursuit in chemical biology.1 These conjugation
methods have made possible the successful preparation of
semisynthetic proteins in large amounts for the study of the
protein−protein interactions, protein functionalities and time-
resolved localization in living cells.2 The formation of native
amide bonds plays a crucial role in the ligation and modification
of peptides and proteins,3 and many research groups have been
developing new strategies for building up amide bonds.4 Native
chemical ligation (NCL) was originally developed by Kent and
co-workers5 in 1994, and it is one of the most widely used
chemoselective ligation tools to join two peptide fragments
under mild reaction conditions and without the need for
protecting groups. However, a particular limitation of NCL is
its intrinsic reliance on the presence of a cysteine residue or a
Cys-mimicking auxiliary residue at the ligation juncture.
Cysteine is uncommon, comprising only 1.7% of all residues
in proteins (Scheme 1A).6 Additionally, the NCL technique
presents challenges for proteins with more than ∼100 amino
acids, and Cys residues can undergo disulfide-bond formation
and influence the overall folding pathway.7 Therefore, it
remains a challenging task to overcome the limitations
α
terminal thioesters can be selectively prepared by engineered
inteins/Sortase-mediated protein ligation, also referred to as
expressed protein ligation (EPL). This approach is an extension
of NCL, and hence any soluble protein could be converted to
the respective peptide thioester by the robust intein (Scheme
1B) or Sortase (Scheme 1C).8 Several peptide backbone
modifications using thioesters have been reported, but these
methods use either some toxic metals or organic solvents,9
which could lead to epimerization of amino acids. From the
perspective of biological chemistry, it is desirable to design
chemical reactions for peptides and proteins that proceed at
physiological pH, temperature and pressure.10 Inspired by the
above trans-thioesterification reaction between the peptide/
protein thioesters, we set out to develop a general ligation that
Received: April 18, 2013
Published: June 24, 2013
© 2013 American Chemical Society
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Scheme 1. Native Chemical Ligation and Protein Thioester Synthesis Using Express Engineered Inteins and Sortase
Article
α
RESULTS AND DISCUSSION
Scheme 2. A New Strategy for the Modification of Peptides
and Proteins: (A) Conceptual Depiction of Thioester-
Mediated Ligation and (B) Approach Discussed in the
Present Work
■
Our earlier method for the peptide synthesis involved EtOH as
the solvent,11 and this solvent is not a suitable choice in native
biological systems. Therefore, we investigated the modification
using ionic liquids instead of EtOH. Because of their unique
properties, ionic liquids are fascinating to chemists and
biochemists in various fields. Besides their potential for green
chemistry (mainly due to their catalytic ability and safety for the
biological system), many organic synthesis reactions have been
carried out in ionic liquids, and these include amino acid and
peptide synthesis.13 The catalyst BSA is an expensive and
unstable reagent, and we therefore pursued to find a new
catalytic system to modify peptides and proteins (Table 1). At
first, the modification of L-Phe-OEt with N-Cbz-L-Phe thioester
was evaluated at various reaction conditions (Table 1). It is
noteworthy that ionic liquids and HMDO can promote the
reactions individually in good yields in an extended time (Table
1, entries 4 and 5). Optimization studies revealed that the best
results were obtained by the combination of HMDO (1 equiv)
could be used as a new strategy for the modification of peptides
and proteins (Scheme 2A).
The thioester substrates for the new chemoselective peptide/
protein modification would be obtained through thiolysis of C-
terminally intein or Sortase A. The key feature of the new
methodology concerns the selective installation of the peptide
thioester into the peptide backbone without cysteine residue in
the second coupling component (Scheme 2B). Recently, our
group reported on the metal-free direct amidation of thioester
peptides with amino acid esters without cysteine residues in the
presence of bistrimethylsilylacetamide (BSA) in EtOH under
mild conditions.11 Liebeskind and co-workers published similar
results at the same time for peptide synthesis with thioacids as
the starting materials in toxic organic solvents.12 In our efforts
to develop a more efficient synthesis of peptides and proteins
via their thioesters or CoA esters or even oxo esters under such
mild conditions, we have discovered that hexamethyldisiloxane
(HMDO) promotes effectively the metal-free direct amide-
coupling reaction between a C-terminal thioester of one
peptide or protein and an amino function of the second
cysteine-free peptide under mild conditions in ionic liquids.
Herein, we report on such a method for the modification of
peptides and proteins.
Table 1. Optimization for the Synthesis of Peptide 14a
time
(h)
yield
a
entry
silyl compounds
solvents (2 mL)
EtOH
(%)
1
BSA (1 equiv)
HMDO (1 equiv)
HMDO (1 equiv)
HMDO (2 mL)
−
72
72
72
72
72
24
24
48
24
48
48
65
74
86
87
45
54
70
93
75
63
42
2
EtOH
3
i-PrOH
−
[BMIM]PF6
4
5
6
HMDO (0.5 equiv) [BMIM]PF6
7
HMDO (1 equiv)
HMDO (1 equiv)
HMDO (1 equiv)
HMDO (1 equiv)
HMDO (1 equiv)
[BMIM]PF6
8
[BMIM]PF6
9
[BMIM]BF4
10
11
[BMIM]BF4:H2O = 4:1
[BMIM]BF4:H2O = 2:1
a
Isolated yield based on the thioester.
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a
Table 2. Peptide Synthesis via HMDO-Mediated Ligation
b
entry
α-amino acid ester (11)
thioester (13)
dipeptide (14)
reaction time (h)
yield (%)
1
L-Phe-OEt (11a)
D-Phe-OMe (11b)
D-Phg-OEt (11c)
L-Leu-OMe (11d)
L-Ser-OEt (11e)
L-Leu-OMe (11d)
L-His-OMe (11f)
L-Phe-OEt (11a)
L-Trp-OMe (11g)
L-Ser-OEt (11e)
L-Leu-OMe (11d)
L-Trp-OMe (11g)
L-Phe-OEt (11a)
Gly-OEt (11h)
Cbz-L-Phe-S(p-tolyl) (13a)
Cbz-L-Phe-S(p-tolyl) (13a)
Cbz-L-Phe-S(p-tolyl) (13a)
Cbz-L-Phe-S(p-tolyl) (13a)
Cbz-L-Phe-S(p-tolyl) (13a)
Cbz-L-Trp-S(p-tolyl) (13b)
Cbz-L-Trp-S(p-tolyl) (13b)
Cbz-L-Trp-S(p-tolyl) (13b)
Cbz-L-Trp-S(p-tolyl) (13b)
Cbz-L-Trp-S(p-tolyl) (13b)
Cbz-L-Pro-S(p-tolyl) (13c)
Cbz-L-Pro-S(p-tolyl) (13c)
Cbz-L-Ala-S(p-tolyl) (13d)
Cbz-L-Met-S(p-tolyl) (13e)
Cbz-D-Phe-S(p-tolyl) (13f)
Ac-D-Phe-S(p-tolyl) (13g)
Cbz-Gly-S(p-tolyl) (13h)
Cbz-L-Phe-S(p-tolyl) (13a)
Cbz-L-Phe-L-Phe-OEt (14a)
Cbz-L-Phe-D-Phe-OMe (14b)
Cbz-L-Phe-D-Phg-OEt (14c)
Cbz-L-Phe-L-Leu-OMe (14d)
Cbz-L-Phe-L-Ser-OEt (14e)
Cbz-L-Trp-L-Leu-OMe (14f)
Cbz-L-Trp-L-His-OMe (14g)
Cbz-L-Trp-L-Phe-OEt (14h)
Cbz-L-Trp-L-Trp-OMe (14i)
Cbz-L-Trp-L-Ser-OEt (14j)
Cbz-L-Pro-L-Leu-OMe (14k)
Cbz-L-Pro-L-Trp-OMe (14l)
Cbz-L-Ala-L-Phe-OEt (14m)
Cbz-L-Met-Gly-OEt (14n)
Cbz-D-Phe-Gly-OEt (14o)
Ac-D-Phe-Gly-OEt (14p)
48
46
48
24
48
24
48
24
48
48
27
48
29
12
12
12
12
93
82
80
83
85
63
63
89
82
86
63
79
80
91
90
91
94
92
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
c
Gly-OEt (11h)
d
Gly-OEt (11h)
D-Phe-OMe (11i)
Gly-OH (11j)
Cbz-Gly-D-Phe-OMe (14q)
Cbz-L-Phe-Gly-OH (14r)
e
12
a
b
c
All reactions performed at 0.5 mmol in [BMIM]PF6 (2 mL) at 40 °C. Isolated yield based on the thioester. 13f (>99% ee); 14o (>99% ee);
d
HPLC conditions: Chiral Pak AD-H, 5 μm, 4.6 mm × 250 mm, n-hexane/2-propanol = 70:30, 210 nm, flow rate = 1.0 mL min−1. 13g (50.4% ee),
e
14p (49.4% ee), Chiral Pak AD-H, 5 μm, 4.6 mm × 250 mm, n-hexane/2-propanol = 70:30, 210 nm, flow rate = 1.0 mL min−1. Thioester (1.0
equiv), Gly-OH (2.00 equiv), NaOH (2.0 equiv).
Scheme 3. Synthesis of Tripeptide 17 and Tetrapeptide 19 via HMDO-Mediated Ligation
and ionic liquids such as 1-butyl-3-methylimidazolium hexa-
fluorophosphate ([BMIM]PF6) and 1-butyl-3-methylimidazo-
lium tetrafluoroborate ([BMIM]BF4) (Table 1, entries 8−9). It
was also found that the HMDO loading presents a critical
determinant of the reaction efficiency; increasing the amount of
HMDO from 0 to 1 equiv can significantly boost the yield from
45 to 70% (Table 1, entries 5−7). Peptide bond formation was
also accomplished in aqueous media ([BMIM]BF4:water >4:1
v/v), and the reactions become much slower with increasing
amounts of water (Table 1, entries 10−11).
Encouraged by this promising result, the scope of the
reaction was tested with a number of thioesters and α-amino
acid esters (Table 2). To our delight, every thioester substrate
tested produced the desired products in good to excellent yields
(63−94%) in [BMIM]PF6, though some α-substituted amino
acids required an extended reaction time (48 h) (Table 2,
entries 1 and 4−13). Treatment of the thioester 13a with
unnatural α-amino acid esters (11b, 11c) resulted in the
formation of unnatural dipeptides (14b, 14c) in excellent yields
(Table 2, entries 2−3). The amidation reaction of α-
unsubstituted amino acid esters such as Gly-OEt 11h proceeds
very well to provide the desired N-Cbz or N-Ac peptides 14n−
14p in 90−91% yields (Table 2, entries 14−16). When the N-
Cbz protected α-unsubstituted thioester such as 13h was
utilized as the coupling partner with the α-substituted amino
acid ester, the desired peptide 14q was obtained in 94% yield
(Table 2, entry 17). The amino acid can be directly used in the
coupling reaction to give the desired product 14r in 92% yield
(Table 2, entry 18). In order to examine the extent of
racemization, an enantiomeric mixture of 14o was synthesized
in the same manner, and HPLC data revealed that no
detectable racemization occurred for the synthesis of D-14o
in the HMDO-mediated coupling reaction. For the N-Ac
protected thioester 13g, the enantiomeric excess of the
corresponding peptide 14p is almost identical to that of the
thioester 13g. Moreover, tripeptide 17 and tetrapeptide 19
were successfully synthesized via this ligation (Scheme 3). It is
noteworthy that this modification reaction is compatible with a
free hydroxyl or thiol group, and hence, the method has
potential for applications to glycopeptide and polypeptide
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Scheme 4. Modification of Stearic Acid Thioester 20, Cinnamoyl-CoA 24, and Glycyrrhizic Acid Thioester 26 via HMDO-
Mediated Ligation
Scheme 5. Proposed Approach for Lysozyme Modification
synthesis. Though the reaction mechanism remains under
investigation, we speculate that HMDO not only can react with
amines to form N-silylamine intermediates but also activates
the carbonyl group of the thioester in the polar solvent system,
and that both of these effects synergistically facilitate peptide
amide bond formation.12,14
Further, the reaction was tested on stearic acid thioester 20
and cinnamoyl-CoA 24 to examine their potential use as probes
to study protein dynamics (Scheme 4). Very pleasingly, the
reactions proceeded smoothly and provided the desired
products 22, 23 and 25 in excellent yields. The reaction of
cinnamoyl-CoA 24 was monitored by HPLC analysis (see
Supporting Information). The α-amino acid esters or amines
could be used as the chemical chain terminator to intercept and
off-load truncated biosynthetic species from acyl carrier protein
(ACP) domains of polyketide synthase (PKSs).15 Interestingly,
modification of the glycyrrhizic acid α-trisubstituted thioester
26 via HMDO-mediated ligation afforded the desired product
27 in 65% yield.
In light of these results, the functionalization of proteins was
attempted. The lysine residue of proteins has attracted much
attention due to its importance in many post-translational
modifications in cell physiology and pathology. The amino
group of a lysine residue can be modified by activated esters,
sulfonyl chlorides and isocyanates.10a,16 We envisioned that the
thioesters could be used to modify lysine residues of proteins
via our HMDO-mediated ligation methodology in ionic liquids.
Lysozyme was chosen as the model to test this protein
modification because of its excellent chemical and thermal
stabilities (Scheme 5).
Using the optimized conditions, lysozyme was reacted with
N-Cbz-L-Phe-S(p-tolyl) (13a) at 37 °C and analyzed by LTQ
Orbitrap XL-MS. Analysis of the reaction mixture after 24 h
confirmed consumption of lysozyme, along with a single peak
(m/z 1621.6776) corresponding to the mass of N-Cbz-L-Phe-
lysozyme (Figure 1B). In order to test our methodology is
applicable for other small molecules modification lysozyme, we
also tested the ability of N-Cbz-L-Trp-S(p-tolyl) (13b) to
modify lysozyme (Figure 1C).
Finally, various oxo esters instead of thioesters were tested to
expand the general reaction scope of the substrates. The
coupling reactions of oxo esters are quite different and much
slower compared to that of the thioesters. This may in part be
attributed to the higher thermodynamic stability and lower
activity of oxo esters.17 However, the reaction was much faster
using a catalytic amount of p-tolylSH (0.1 equiv) (Table 3). We
expected the thioesters to be more reactive intermediates in the
direct addition of amino acid esters toward oxo esters (Scheme
6). In our mechanistic hypothesis, the thiol would play the part
of a catalytic mediator, and then the in situ-generated thioesters
by the thioesterification of oxo esters would be converted into
the peptide in the presence of HMDO in ionic liquids. Analysis
of the reaction mixture by LTQ Orbitrap XL-MS indicated the
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Figure 1. LTQ Orbitrap XL-MS spectrum of the lysozyme (A), N-Cbz-L-Phe-lysozyme (28) (B), and N-Cbz-L-Trp-lysozyme (29) (C).
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through thiolysis of C-terminally fused inteins or enzyme
Sortase A. We are currently exploring this approach for several
proteins without cysteine residue, and progress will be reported
in due course.
Table 3. Peptide Synthesis of Oxo Esters via HMDO-
Mediated Ligation
EXPERIMENTAL SECTION
■
General Information. 1H and 13C NMR spectra were recorded on
400 MHz spectrometers in deuteriochloroform (CDCl3) unless
otherwise stated. All 1H chemical shifts are reported in ppm (δ)
relative to TMS (0.00); 13C shifts are reported in ppm (δ) relative to
CDCl3 (77.0). Data are reported in the following order: chemical
shifts are given (δ); multiplicities are indicated as br (broadened), s
(singlet), d (doublet), t (triplet), q (quartet), m (multiplet), app
(apparent); coupling constants, J, are reported (Hz); integration is
provided. Optical rotation values were measured at 20 °C with
different solvents. Melting points were taken without calibration.
Analytical thin-layer chromatography (TLC) was performed on silica
gel aluminum sheets with F-254 indicator. Visualization was
accomplished by UV light or with solutions of K2CO3/KMnO4 in
water. Purification by chromatography was performed using 230−400
mesh SiO2 with compressed air as a source of positive pressure. HPLC
analyses were carried out with a binary pump on Symmetry C18 (4.6
× 150 mm, 5 μm) and CHIRAL PAK AD-H (4.6 mm I.D. × 250 mm,
5 μm). Solvents for reactions and chromatography were reagent grade
and used as received. “Brine” refers to a saturated aqueous solution of
NaCl. Solution of NaHCO3 refers to saturated aqueous solutions.
Solvents used as reaction media were purchased in >99% purity
without further purification, unless otherwise specified. High
Resolution MS was analyzed by Thermo LTQ XL Orbitrap.
p-tolylSH
reaction
time (h)
dipeptide
yield
a
entry
1
oxo ester (30)
(equiv)
(14)
(%)
Cbz-L-Phe-oxo
ester (30a)
0
72
62
55
72
60
48
14a
20
72
68
25
70
71
2
3
4
5
6
Cbz-L-Phe-oxo
ester (30a)
0.1
0.5
0
14a
Cbz-L-Phe-oxo
ester (30a)
14a
Cbz-L-Ala-oxo
ester (30a)
14m
14m
14m
Cbz-L-Ala-oxo
ester (30a)
0.1
0.5
Cbz-L-Ala-oxo
ester (30a)
a
Isolated yield based on oxo ester.
Scheme 6. Possible Pathway for the 1,2-Addition of Amino
Acid Esters to Oxo Esters by the Thioester Intermediates
Starting Materials. Cinnamoyl CoA was prepared according to
the literature.18 Thioesters were prepared from the corresponding N-
Cbz-L-amino acids.11
General Procedure A: Preparation of Thioesters Derived
from N-Cbz-L-Amino Acids. 4-Methylbenzenethiol (10.5 mmol)
and 1-hydroxybenzotriazole (15 mmol) were added to a solution of
the N-Cbz-L-amino acid (10 mmol) in ethyl acetate (10 mL) at 0 °C
followed by N,N′-diisopropylcarbodiimide (10 mmol). The mixture
was stirred for 24 h at room temperature, and the reaction progress
was monitored by TLC analysis. At the end of the reaction a few drops
of 50% acetic acid in ethyl acetate were added. The mixture was
filtered through Celite, and the organic phase was washed with
NaHCO3 solution and brine, dried over MgSO4, filtered, and
concentrated in vacuo. The crude product was purified by flash
chromatography affording the desired products.
formation of the thioester intermediate (m/z 406.1482, Figure
2).
CONCLUSIONS
■
In summary, we have developed an innovative strategy for the
modification of peptides and proteins that relies on HMDO-
mediated ligation in ionic liquids and proceeds smoothly under
mild conditions. The peptide coupling scope is expanded using
in situ-generated thioesters by the thioesterification of oxo
esters. This method is environmental friendly and does not
require a cysteine residue to form a native amide bond, and the
new method therefore presents a significant expansion to extant
NCL strategies. The approach will be particularly useful in
combination with the chemoselective thioester formation
N-Cbz-L-Phenylalanine p-toluene thioester (13a). Following the
general procedure A, the reaction performed with N-Cbz-L-phenyl-
alanine (2.990 g, 10 mmol) afforded the desired product as a white
solid (3.244 g, 80% yield): mp 114−116 °C (lit.11 116−117 °C);
20
1
[α]D −66.6 (c 0.42, CHCl3); H NMR (400 MHz, CDCl3) δ =
7.38−7.19 (m, 14H), 5.27 (d, J = 8.0 Hz, 1H), 5.16 (s, 2H), 4.89−4.84
(m, 1H), 3.24 (m, 2H), 2.41 (s, 3H); 13C NMR (101 MHz, CDCl3) δ
Figure 2. LTQ Orbitrap XL-MS spectrum of the thioesterification reaction mixture.
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= 199.1, 155.6, 139.9, 136.0, 135.3, 134.5, 130.1, 129.4, 128.7, 128.6,
128.3, 128.1, 127.3, 123.4, 67.3, 61.3, 38.5, 21.4; HRMS (ESI) Calcd
for C24H24NO3S ([M + H]+) 406.1471, found 406.1483.
N-Boc-Phenylalanine p-toluene thioester (16). Following the
general procedure A, the title compound was prepared as a white solid
(3.195 g, 86% yield): mp 166−167 °C; [α]D20 −38.4 (c 0.60, CHCl3);
1H NMR (400 MHz, CDCl3) δ = 7.27−7.11 (m, 9H), 4.92 (br, 1H),
4.70 (br, 1H), 3.11−3.00 (m, 2H), 2.30 (s, 3H), 1.36 (s, 9H); 13C
NMR (101 MHz, CDCl3) δ = 155.0, 139.8, 135.7, 134.6, 130.1, 129.5,
128.7, 127.1, 123.7, 80.5, 60.8, 42.3, 38.4, 23.4, 21.4; HRMS (ESI)
Calcd for C21H26NO3S ([M + H]+) 372.1628, found 372.1633.
N-Boc-Phenylalanine-glycine p-toluene thioester (18). Following
the general procedure A, the title compound was prepared as a white
N-Cbz-L-tryptophan p-toluene thioester (13b). Following the
general procedure A, the title compound was prepared as a pale yellow
solid (3.779 g, 85% yield): mp 50−52 °C (lit.11 51−52.5 °C); [α]D
20
1
−82.3 (c 0.50, CHCl3); H NMR (400 MHz, CDCl3) δ = 8.12 (s,
1H), 7.59 (d, J = 8.0 Hz, 1H), 7.41−7.34 (br, 6H), 7.26−7.12 (m,
6H), 7.06 (d, J = 4.0 Hz, 1H), 5.37 (d, J = 8.0 Hz, 1H), 5.16 (s, 2H),
4.93−4.88 (m, 1H), 3.46−3.31 (m, 2H), 2.40 (s, 3H); 13C NMR (101
MHz, CDCl3) δ = 139.8, 136.2, 134.6, 130.0, 128.5, 128.2, 128.1,
123.2, 122.4, 119.9, 111.2, 109.5, 67.2, 61.0, 28.3, 21.3; HRMS (ESI)
Calcd for C26H25N2O3S ([M + H]+) 445.1580, found 445.1589.
N-Cbz-L-Proline p-toluene thioester (13c). Following the general
procedure A, the title compound was prepared as a white crystal.
(2.318 g, 65% yield): mp 57−59 °C (lit.11 61−62 °C); [α]D20 −105 (c
1
solid (3.215 g, 75% yield): mp 186−188 °C; H NMR (400 MHz,
CDCl3) δ = 7.31−7.21 (m, 9H), 6.71 (br, 1H), 4.98 (br, 1H), 4.43 (br,
1H), 4.31−4.14 (m, 2H), 3.19−3.02 (m, 2H), 2.38 (s, 3H), 1.39 (s,
9H); 13C NMR (101 MHz, CDCl3) δ = 140.1, 136.5, 134.6, 130.2,
129.3, 128.7, 127.0, 122.6, 48.8, 29.7, 28.2, 21.3; HRMS (ESI) Calcd
for C23H29N2O4S ([M + H]+) 429.1843, found 429.1856.
1
Stearic acid p-toluene thioester (20). Following the general
0.50, CHCl3); H NMR (400 MHz, CDCl3) δ = 7.45−7.32 (m, 6H),
procedure A, the title compound was prepared as a white solid (3.315
7.25−7.14 (m, 3H), 5.34−5.12 (m, 2H), 4.69−4.57 (m, 1H), 3.74−
3.61(m, 2H), 2.40 (s, 3H), 2.31−2.15 (m, 3H), 2.10−1.98 (m, 1H);
13C NMR (101 MHz, CDCl3) δ = 200.6, 200.1, 155.1, 154.5, 139.7,
139.6, 136.6, 136.4, 134.6, 134.5, 130.0, 128.5, 128.4, 128.0, 127.9,
123.8, 123.6, 127.9, 123.8, 123.6, 67.3, 66.4, 65.9, 47.3, 46.9, 31.7, 30.7,
24.1, 23.4, 21.3; HRMS (ESI) Calcd for C20H22NO3S ([M + H]+)
356.1315, found 356.1321.
g, 85% yield): mp 92−94 °C (lit.19 46.5 °C); H NMR (400 MHz,
1
CDCl3) δ = 7.32−7.23 (m, 4H), 2.67 (t, J = 12.0 Hz, 2H), 2.39 (s,
3H), 1.76−1.68 (m, 2H), 1.28 (s, 28H), 0.92 (t, J = 12.0 Hz, 3H); 13C
NMR (101 MHz, CDCl3) δ = 198.2, 139.3, 134.2, 130.1, 124.3, 43.6,
31.9, 29.7, 29.6, 29.4, 29.3, 29.2, 29.0, 21.3, 14.0; IR (film) 2918,
2850,1710, 1534, 1469, 1400, 1043, 947, 806, 762 cm−1; HRMS (ESI)
Calcd for C25H43OS ([M + H]+) 391.3029, found 391.3029.
N-Cbz-L-Alanine p-toluene thioester (13d). Following the general
procedure A, the title compound was prepared as a white solid (2.505
Glycyrrhizic acid p-toluene thioester (26). Following the general
g, 76% yield): mp 61−63 °C (lit.11 58−62 °C); [α]D −28.6 (c 0.7,
20
procedure A, the title compound was prepared as a white solid (5.015
1
g, 85% yield): mp 260−262 °C; H NMR (400 MHz, CDCl3) δ =
CHCl3); 1H NMR (400 MHz, CDCl3) δ = 7.42−7.24 (m, 9H), 5.32−
5.19 (m, 1H), 5.16 (s, 2H), 4.66−4.59 (m, 1H), 2.40 (s, 3H), 1.50 (d,
J = 8.0 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ = 199.8, 155.5,
139.9, 134.6, 130.1, 128.6, 128.3, 128.2, 123.3, 67.3, 56.6, 21.4, 19.0;
HRMS (ESI) Calcd for C18H20NO3S ([M + H]+) 330.1158, found
330.1160.
7.30−7.22 (m, 4H), 7.13 (d, J = 8.0 Hz, 2H), 6.04 (d, J = 8.0 Hz, 2H),
5.63 (s, 1H), 4.92−4.87 (m, 1H), 4.25−4.20 (m, 2H), 3.24−3.05 (m,
4H), 2.80 (d, J = 16.0 Hz, 1H), 2.31 (s, 1H), 2.18−2.12 (m, 1H),
2.05−1.96 (m, 1H), 1.90−1.56 (m, 10H), 1.41−1.22 (m, 34H), 1.15−
1.11 (m, 10H), 1.05 (s, 3H), 1.00 (s, 4H), 0.88−0.80 (m, 16H), 0.75
(s, 3H); 13C NMR (101 MHz, CDCl3) δ = 171.7, 129.2, 128.6, 127.2,
78.8, 61.8, 61.7, 54.9, 52.6, 47.7, 45.3, 43.6, 43.1, 41.7, 39.1, 37.8, 37.2,
37.0, 32.7, 31.9, 31.8, 29.6, 29.4, 28.3, 28.1, 27.3, 26.3, 23.5, 23.4, 18.6,
17.5, 16.7, 16.4, 15.6, 14.2, 14.1; IR (film) 3310, 3034, 2951, 1732,
1660, 1529, 1447, 1344, 1254, 1048, 744, 700 cm−1; HRMS (ESI)
Calcd for C37H53O3S ([M + H]+) 577.3710, found 577.3710.
N-Cbz-L-Methionine p-toluene thioester (13e). Following the
general procedure A, the title compound was prepared as a white solid
(3.116 g, 80% yield): mp 141−142 °C (lit.11 143−144 °C); [α]D
20
1
−35.2 (c 0.50, CHCl3); H NMR (400 MHz, CDCl3) δ = 7.41−7.35
(m, 5H), 7.31−7.24 (m, 4H), 5.54 (d, J = 8.0 Hz, 1H), 5.20 (s, 2H),
4.77−4.71 (m, 1H), 2.67−2.57 (m, 2H), 2.40 (s, 3H), 2.31−2.30 (m,
1H), 2.27 (s, 3H), 2.25−1.99 (m, 1H); 13C NMR (101 MHz, CDCl3)
δ = 199.1, 155.8, 140.0, 136.0, 135.4, 134.5, 130.1, 128.6, 128.3, 128.2,
123.1, 67.4, 60.0, 32.0, 30.0, 21.4, 15.4; HRMS (ESI) Calcd for
C20H24NO3S2 ([M + H]+) 390.1192, found 390.1199.
N-Cbz-L-Phenylalanine oxo ester (30a). 4-Chlorophenol (10.5
mmol) and 1-hydroxybenzotriazole (15 mmol) were added to a
solution of the N-Cbz- L-amino acid (10 mmol) in ethyl acetate (10
mL) at rt followed by N,N′-diisopropylcarbodiimide (10 mmol). The
mixture was stirred for 24 h at room temperature, and the reaction
progress was monitored by TLC analysis. At the end of the reaction, a
few drops of 50% acetic acid in ethyl acetate were added. The mixture
was filtered through Celite, and the organic phase was washed with
NaHCO3 solution and brine, dried over MgSO4, filtered, and
concentrated in vacuo. The crude product was purified by flash
chromatography affording the desired products; the title compound
was prepared as a white solid (3.485 g, 85% yield): mp 136−138 °C;
1H NMR (400 MHz, CDCl3) δ = 7.35−7.31 (m, 12H), 7.21−7.19 (d,
J = 12.0 Hz, 2H), 6.92−6.89 (d, J = 12.0 Hz, 2H), 5.31 (br, 1H), 5.13
(s, 2H), 4.88 (br, 1H), 3.25−3.24 (d, J = 4.0 Hz, 2H); 13C NMR (101
MHz, CDCl3) δ = 170.1, 135.1, 131.6, 129.5, 129.4, 128.8, 128.6,
128.3, 128.2, 127.5, 122.6, 67.2, 55.0, 38.2; IR (film) 3310, 3034, 2951,
1732, 1660, 1529, 1447, 1344, 1254, 1048, 744, 700 cm−1; HRMS
(ESI) Calcd for C23H21ClNO4 ([M + H]+) 410.1154, found 410.1138.
N-Cbz-L-Alanine oxo ester (30b). Following the above general
procedure, the title compound was prepared as a white solid (2.939 g,
88% yield): mp 104−107 °C; 1H NMR (400 MHz, CDCl3) δ = 7.37−
7.33 (m, 8H), 7.05−7.03 (d, J = 8.0 Hz, 2H), 7.92−7.89 (d, J = 12.0
Hz, 2H), 5.34 (br, 1H), 5.14 (s, 2H), 4.63 (br, 1H), 1.61 (d, J = 8 Hz,
3H); 13C NMR (101 MHz, CDCl3) δ = 171.5, 155.6, 131.6, 129.6,
128.6, 128.2, 122.7, 67.1, 49.8, 18.4; IR (film) 3336, 1777, 1692, 1531,
1488, 1456, 1359, 1298, 1261, 1202, 1152, 1120, 1072 cm−1; HRMS
(ESI) Calcd for C17H17ClNO4 ([M + H]+) 334.0841, found 334.0852.
General Method B: Preparation of Amino Acid Ester Free
Amine (11a−11i).20 The amino acid ester hydrochloride salt (10
mmol) was mixed with tetrahydrofuran (THF) (20 mL). Triethyl-
N-Cbz-D-Phenylalanine p-toluene thioester (13f). Following the
general procedure A, the reaction performed with N-Cbz-D-phenyl-
alanine (2.990 g, 10 mmol) afforded the desired product as a white
20
solid (3.163 g, 78% yield): mp 110−112 °C; [α]D +18.6 (c 0.52,
CHCl3); 1H NMR (400 MHz, CDCl3) δ = 7.36−7.15 (m, 14H), 5.26
(d, J = 8.0 Hz, 1H), 5.12 (s, 2H), 4.85−4.80 (m, 1H), 3.20−3.10 (m,
2H), 2.37 (s, 3H); 13C NMR (101 MHz, CDCl3) δ = 199.2, 155.6,
139.9, 136.1, 135.4, 134.6, 130.2, 129.5, 128.8, 128.6, 128.3, 128.1,
127.3, 123.4, 110.0, 67.3, 61.3, 38.4, 21.4; HRMS (ESI) Calcd for
C24H24NO3S ([M + H]+) 406.1471, found 406.1488.
N-Ac-D-Phenylalanine p-toluene thioester (13g). Following the
general procedure A, the title compound was prepared as a white solid
(2.347 g, 75% yield): mp 158−159 °C; [α]D20 +56.4 (c 0.65, CHCl3);
1H NMR (400 MHz, CDCl3) δ = 7.74−7.65 (m, 1H), 7.36−7.10 (m,
8H), 5.92 (d, J = 8.0 Hz, 1H), 5.06−5.00 (m, 1H), 3.10 (d, J = 8.0 Hz,
2H), 2.30 (s, 3H), 1.93 (s, 3H); 13C NMR (101 MHz, CDCl3) δ =
198.7, 169.8, 140.0, 135.3, 134.6, 130.0, 129.3, 128.7, 127.3, 59.3, 38.3,
23.2, 21.4; HRMS (ESI) Calcd for C18H20NO2S ([M + H]+)
314.1209, found 314.1216.
N-Cbz-Glycine p-toluene thioester (13h). Following the general
procedure A, the title compound was prepared as a white solid (2.625
g, 83% yield): mp 93−95 °C; 1H NMR (400 MHz, CDCl3) δ = 7.37−
7.21 (m, 10H), 5.41 (br, 1H), 5.16 (s, 2H), 4.41 (d, J = 4.0 Hz, 2H),
2.37 (s, 3H), 1.93 (s, 3H); 13C NMR (101 MHz, CDCl3) δ = 196.5,
140.1, 134.7, 130.3, 128.6, 128.3, 128.2, 122.7, 67.4, 50.5, 21.3; HRMS
(ESI) Calcd for C17H18NO3S ([M + H]+) 316.1002, found 316.1015.
7019
dx.doi.org/10.1021/jo400797t | J. Org. Chem. 2013, 78, 7013−7022

The Journal of Organic Chemistry
Article
amine (200 mmol) was added dropwise and stirred overnight at room
temperature. The reaction mixture was filtered and washed with THF
(10 mL). The solution was concentrated under a vacuum to afford the
amino acid ester free amine.
yield): mp 70−72 °C (lit.11 73−75 °C); [α]D20 +46.4 (c 0.5, CHCl3)
1H NMR (400 MHz, CDCl3) δ = 8.16 (s, 1H), 7.73 (br, 1H), 7.39−
7.11 (m, 10H), 6.11 (br, 1H), 5.56 (br, 1H), 5.14 (s, 2H), 4.54−4.50
(m, 2H), 3.67 (s, 3H), 3.39 (br, 1H), 3.22 (br, 1H), 1.71 (br, 2H),
1.49 (br, 1H), 0.87 (t, J = 8.0 Hz, 6H); 13C NMR (101 MHz, CDCl3)
δ = 172.8, 171.0, 136.2, 128.5, 128.2, 128.1, 123.5, 122.3, 119.8, 118.9,
111.2, 52.2, 50.8, 41.5, 24.6, 22.6, 21.9; HRMS (ESI) Calcd for
C26H32N3O5 ([M + H]+) 466.2336, found 466.2344.
General Method C: Modification of Amines with Thiol Esters
in the Presence of HMDO. N-Cbz-L-amino acid thioester or other
thioester (0.5 mmol, 1.0 equiv) was dissolved in ionic liquids (2 mL).
HMDO (0.5 mmol, 1.0 equiv) and amino acid ester free amine (1.0
mmol, 2.0 equiv) were added stepwise. The reaction mixture was
stirred at 37 °C for 24−48 h, cooled to room temperature, and
concentrated under reduced pressure. The remaining residue was
dissolved in ethyl acetate (20 mL) and washed with 1 M aqueous HCl
(5 mL), 2 M aqueous NaOH (5 mL), and brine (5 mL). The organic
layer was dried over Na2SO4, filtered, and concentrated to dryness.
The crude product was purified by flash chromatography.
N-Cbz-L-Phe-L-Phe-OEt (14a). Following the general procedure C,
the title compound was prepared as a white solid (220 mg, 93% yield):
mp 142−144 °C (lit.21 138−139 °C); [α]D20 −9.9 (c 0.5, CHCl3); 1H
NMR (400 MHz, CDCl3) δ = 7.36−7.15 (m, 13H), 7.04−6.96 (m,
2H), 6.45 (br, 1H), 5.36 (br, 1H), 5.09 (s, 2H), 4.84−4.76 (m, 1H),
4.45 (br, 1H), 4.16−4.13 (m, 2H), 3.08−3.06 (m, 4H), 1.24−1.19 (m,
3H); 13C NMR (101 MHz, CDCl3) δ = 170.9, 170.4, 155.9, 136.2,
135.6, 129.4, 129.3, 128.7, 128.5, 128.2, 128.0, 127.1, 67.1, 61.5, 53.4,
53.1, 38.0, 29.0, 14.1; HRMS (ESI) Calcd for C28H31N2O5 ([M +
H]+) 475.2227, found 475.2228.
N-Cbz-L-Trp-L-His-OMe (14g). Following the general procedure C,
the title compound was prepared as a white solid (154.0 mg, 63%
20
1
yield): mp 163−165 °C; [α]D −22.8 (c 0.5, CHCl3) H NMR (400
MHz, CD3OD-d6) δ = 8.45 (s, 1H), 7.59 (d, J = 8.0 Hz, 1H), 7.34−
7.30 (m, 6H), 7.19 (s, 1H), 7.12 (t, J = 6.0 Hz, 2H), 7.03 (t, J = 8.0
Hz, 1H), 5.08−5.07 (m, 2H), 4.72−4.70 (m, 1H), 4.39 (t, J = 6.0 Hz,
1H), 3.65 (s, 3H), 3.25−3.19 (m, 2H), 3.10−3.02 (m, 2H); 13C NMR
(101 MHz, CD3OD-d6) δ = 173.3, 170.3, 156.9, 136.8, 136.6, 136.5,
133.7, 129.7, 128.0, 127.5, 127.2, 123.3, 120.9, 118.4, 117.8, 110.8,
109.2, 27.4, 26.6; HRMS (ESI) Calcd for C26H28N5O5 ([M + H]+)
490.2085, found 490.2080.
N-Cbz-L-Trp-L-Phe-OEt (14h). Following the general procedure C,
the title compound was prepared as a pale yellow solid (228.2 mg, 89%
20
1
yield): mp 158−160 °C; [α]D +9.2 (c 0.5, CHCl3) H NMR (400
MHz, CDCl3) δ = 8.06 (s, 1H), 7.71 (br, 1H), 7.38−7.34 (m, 6H),
7.24−7.14 (m, 5H), 7.02 (br, 1H), 6.68 (d, J = 8.0 Hz, 2H), 6.18 (d, J
= 8.0 Hz, 1H), 5.49 (br, 1H), 5.13 (s, 2H), 4.75−4.73 (m, 1H), 4.52
(br, 1H), 4.12−4.05 (m, 2H), 3.38 (br, 1H), 3.18 (q, 1H), 2.98 (d, J =
8.0 Hz, 2H), 1.69 (br, 1H), 1.22 (t, J = 8.0 Hz, 3H); 13C NMR (101
MHz, CDCl3) δ = 170.8, 170.7, 136.2, 135.6, 129.2, 128.5, 128.4,
128.2, 128.0, 127.0, 123.4, 122.3, 119.9, 118.9, 111.2, 61.4, 53.3, 37.8,
28.5, 14.0; HRMS (ESI) Calcd for C30H32N3O5 ([M + H]+) 514.2336,
found 514.2334.
N-Cbz-L-Phe-D-Phe-OMe (14b). Following the general procedure C,
the title compound was prepared as a white solid (188 mg, 82% yield):
20
1
mp 141−142 °C; [α]D −24.6 (c 0.5, CHCl3); H NMR (400 MHz,
CDCl3) δ = 7.36−7.13 (m, 13H), 6.91−6.89 (m, 2H), 6.31 (br, 1H),
5.28 (br, 1H), 5.06 (s, 2H), 4.86−4.81 (m, 1H), 4.45 (br, 1H), 4.16−
4.13 (m, 2H), 3.66 (s, 3H), 3.08−2.89 (m, 4H); 13C NMR (101 MHz,
CDCl3) δ = 171.4, 170.4, 136.2, 135.5, 129.3, 129.2, 128.7, 128.6,
128.5, 128.2, 128.0, 127.2, 127.1, 67.1, 56.2, 53.0, 52.3, 38.5, 37.8;
HRMS (ESI) Calcd for C27H29N2O5 ([M + H]+) 461.2071, found
461.2070.
N-Cbz-L-Trp-L-Trp-OMe (14i). Following the general procedure C,
the title compound was prepared as a pale yellow solid (231.2 mg, 82%
yield): mp 185−187 °C (lit.23 175 °C); [α]D +13.4 (c 0.5, CHCl3)
20
1H NMR (400 MHz, CDCl3) δ = 7.93 (d, J = 8.0 Hz, 1H), 7.26−6.96
(m, 15H), 6.47 (d, J = 8.0 Hz, 1H), 6.38 (d, J = 8.0 Hz, 1H), 5.16 (br,
1H), 4.98 (m, 2H), 4.81 (m, 1H), 4.38 (br, 1H), 3.56 (d, J = 12.0 Hz,
3H), 3.21 (m, 1H), 3.06 (m, 2H); 13C NMR (101 MHz, CDCl3) δ =
171.8, 170.4, 136.3, 129.4, 129.3, 128.7, 128.6, 128.2, 128.1, 128.0,
123.0, 122.2, 119.7, 118.4, 111.2, 109.4, 52.8, 52.5, 52.4, 27.5; HRMS
(ESI) Calcd for C31H31N4O5 ([M + H]+) 539.2289, found 539.2290.
N-Cbz-L-Trp-L-Ser-OEt (14j). Following the general procedure C,
the title compound was prepared as a white solid (195.7 mg, 86%
N-Cbz-L-Phe-D-Phg-OEt (14c). Following the general procedure C,
the title compound was prepared as a white solid (184 mg, 80% yield):
20
1
mp 146−148 °C; [α]D −32.8 (c 0.5, CHCl3); H NMR (400 MHz,
CDCl3) δ = 7.36−7.30 (m, 9H), 7.19−7.15 (m, 5H), 7.07 (br, 2H),
6.71 (br, 1H), 5.48 (d, J = 8.0 Hz, 1H), 5.08 (s, 2H), 4.49 (br, 1H),
4.21−4.08 (m, 2H), 3.05−3.00 (m, 2H), 1.20 (t, J = 8.0 Hz, 3H); 13C
NMR (101 MHz, CDCl3) δ = 170.4, 170.0, 136.0, 129.3, 128.9, 128.7,
128.5, 128.2, 128.0, 127.2, 110.0, 61.9, 56.0, 13.9; IR (film) 3734,
3006, 1732, 1695, 1651, 1536, 1451, 1377, 1317, 1258, 1214, 1136,
1041, 744, 698; HRMS (ESI) Calcd for C27H29N2O5 ([M + H]+)
461.2071, found 461.2071.
20
1
yield): mp 118−119 °C; [α]D +16.4 (c 0.5, CHCl3) H NMR (400
MHz, DMSO-d6) δ = 8.47 (d, J = 8.0 Hz, 1H), 7.53 (d, J = 8.0 Hz,
1H), 7.33−7.09 (m, 10H), 5.15 (br, 1H), 4.94 (s, 2H), 4.41 (br, 2H),
4.12−4.06 (m, 2H), 3.78 (br, 1H), 3.68 (br, 1H), 3.07−3.00 (m, 1H),
2.78−2.69 (m, 1H), 1.21 (t, J = 8.0 Hz, 3H); 13C NMR (101 MHz,
DMSO-d6) δ = 172.4, 170.9, 156.3, 138.6, 137.4, 129.7, 128.7, 128.4,
128.1, 127.8, 126.6, 65.6, 61.0, 56.3, 55.2, 37.9, 14.5; IR (film) 3737,
3401, 2922, 2856, 1703, 1671, 1525, 1457, 1342, 1258, 1136, 1023,
847 cm−1; HRMS (ESI) Calcd for C24H28N3O6 ([M + H]+) 454.1973,
found 454.1974.
N-Cbz-L-Phe-L-Leu-OMe (14d). Following the general procedure C,
the title compound was prepared as a white solid (176.7 mg, 83%
yield): mp 110−112 °C. (lit.22 109−110 °C); [α]D −23.4 (c 0.5,
20
1
CHCl3) H NMR (400 MHz, CDCl3) δ = 7.37−7.21 (m, 10H), 6.23
(br, 1H), 5.37 (br, 1H), 5.11 (s, 2H), 4.58−4.47 (m, 2H), 3.71 (s,
3H), 3.16−3.05 (m, 2H), 1.60−1.43 (m, 3H), 0.92−0.86 (m, 6H); 13C
NMR (101 MHz, CDCl3) δ = 172.7, 170.5, 156.0, 136.2, 129.4, 128.7,
128.5, 128.2, 128.0, 127.0, 67.1, 56.0, 52.3, 50.8, 41.5, 38.3, 24.7, 22.7;
HRMS (ESI) Calcd for C24H31N2O5 ([M + H]+) 427.2227, found
427.2212.
N-Cbz-L-Pro-L-Leu-OMe (14k). Following the general procedure C,
the title compound was prepared as a pale yellow solid (118 mg, 63%
yield): mp 73−75 °C (lit.24 78−80 °C); [α]D20 −56.7 (c 0.5, CHCl3)
1H NMR (400 MHz, CDCl3) δ = 7.37 (br, 5H), 7.12 (br, 0.5H), 6.34
(br, 0.33H), 5.23−5.14 (m, 2H), 4.55 (br, 1H), 4.39 (br, 1H), 3.73−
3.45 (m, 5H), 2.36−2.17 (m, 1H), 1.92 (s, 3H), 1.61 (br, 3H), 0.91 (d,
J = 4.0 Hz, 6H); 13C NMR (101 MHz, CDCl3) δ = 173.2, 171.4,
136.5, 128.6, 128.0, 127.8, 67.3, 60.7, 60.3, 52.2, 50.9, 47.5, 46.9, 41.2,
24.9, 24.6, 22.7, 21.9, 14.1; HRMS (ESI) Calcd for C20H29N2O5 ([M +
H]+) 377.2071, found 377.2070.
N-Cbz-L-Pro-L-Trp-OMe (14l). Following the general procedure C,
the title compound was prepared as a colorless oil (177.3 mg, 79%
yield): [α]D20 −32.4 (c 0.5, CHCl3) 1H NMR (400 MHz, CD3OD-d6)
δ = 7.55−7.49 (m, 1H), 7.38−7.31 (m, 3H), 7.23 (br, 3H), 7.10−7.08
(m, 1H), 7.04−7.0 (m, 1H), 5.15−5.08 (m, 1H), 5.00−4.95(m, 2H),
4.73−4.71 (m, 1H), 4.32−4.29 (m, 1H), 3.66 (d, J = 12.0 Hz, 3H),
3.50−3.43 (m, 2H), 3.28−3.10 (m, 2H), 2.22−2.12 (m, 1H), 1.96−
N-Cbz-L-Phe-L-Ser-OEt (14e). Following the general procedure C,
the title compound was prepared as a white solid (176.1 mg, 85%
yield): mp 97−99 °C (lit.11 100−103 °C); [α]D +12.3 (c 0.5,
20
1
CHCl3) H NMR (400 MHz, CD3OD-d6) δ = 7.62 (d, J = 8.0 Hz,
1H), 7.36−7.28 (m, 6H), 7.14−7.09 (m, 3H), 7.03−6.99 (m, 1H),
5.03 (s, 2H), 4.56−4.53 (m, 1H), 4.50−4.48 (m, 1H), 4.20−4.12 (m,
2H), 3.89−3.85 (m, 1H), 3.78−3.74 (m, 1H), 3.34−3.29 (m, 2H),
3.16−3.10 (m, 1H), 1.27 (t, J = 8.0 Hz, 3H); 13C NMR (101 MHz,
CD3OD-d6) δ = 173.2, 170.1, 156.9, 136.7, 136.6, 128.0, 127.5, 127.4,
127.3, 123.4, 121.0, 118.4, 117.9, 110.9, 109.4, 66.2, 61.5, 61.2, 55.9,
54.9, 27.8, 13.1; HRMS (ESI) Calcd for C22H27N2O6 ([M + H]+)
415.1864, found 415.1863.
N-Cbz-L-Trp-L-Leu-OMe (14f). Following the general procedure C,
the title compound was prepared as a pale yellow solid (147 mg, 63%
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1.79 (m, 3H); 13C NMR (101 MHz, CDCl3) δ = 170.8, 170.5, 136.2,
135.6, 128.5, 128.2, 128.0, 127.0, 123.4, 122.3, 119.8, 118.8, 111.2,
61.4, 53.3, 42.2, 37.8, 23.5, 14.1; HRMS (ESI) Calcd for C25H28N3O5
([M + H]+) 450.2023, found 450.2022.
N-Cbz-L-Ala-L-Phe-OEt (14m). Following the general procedure C,
the title compound was prepared as a white solid (159 mg, 80% yield):
mp 97−99 °C (lit.25 97−98 °C) [α]D20 +18.4 (c 0.5, CHCl3) 1H NMR
(400 MHz, CDCl3) δ = 7.37−7.24 (m, 8H), 7.13 (d, J = 8.0 Hz, 2H),
6.55 (br, 1H), 5.36 (br, 1H), 5.15 (q, 2H), 4.87−4.82 (m, 1H), 4.26−
4.16 (m, 3H), 3.18−3.07 (m, 2H), 1.36 (d, J = 4.0 Hz, 3H), 1.27 (t, J =
4.0 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ = 171.8, 171.2, 155.8,
136.1, 135.7, 129.3, 128.5, 128.2, 128.0, 127.1, 67.0, 61.6, 53.2, 50.4,
37.8, 18.5, 14.1; HRMS (ESI) Calcd for C22H27N2O5 ([M + H]+)
399.1914, found 399.1918.
CDCl3) δ = 171.7, 169.5, 136.7, 129.3, 128.6, 126.8, 61.5, 41.3, 28.2,
14.1; HRMS (ESI) Calcd for C18H27N2O5 ([M + H]+) 351.1914,
found 351.1918.
N-Boc-L-Phe-Gly-OEt. Title compound (350 mg, 1 mmol) was
dissolved in DCM (3 mL). The reaction mixture was cooled at 0 °C
and then added with TFA (1 mL). The reaction mixture was stirred at
0 °C for 40 min and then diluted with DCM, washing with NaHCO3
and brine. The organic layer was dried over MgSO4 and then
concentrated to get the crude product as yellow oil. The crude product
used directly in the next reaction without purification
N-Boc-L-Phe-L-Phe-Gly-OEt (17). Following the general procedure
C, the title compound was prepared as a white solid (190 mg, 76%
yield): mp 164−167 °C; 1H NMR (400 MHz, DMSO-d6) δ = 7.55 (t,
J = 8 Hz, 1H), 7.43 (t, J = 8 Hz, 1H), 7.27−7.17 (m, 10H), 6.89 (br,
1H), 4.60 (br, 1H), 4.13 (q, J = 8.0 Hz, 3H), 3.86 (br, 2H), 3.07−3.03
(m, 1H), 2.87 (d, J = 16.0 Hz, 2H), 2.69 (s, 2H), 1.28 (s, 9H), 1.21 (t,
J = 8 Hz, 3H), 13C NMR (101 MHz, DMSO-d6) δ = 171.3, 169.5,
155.0, 138.0, 137.5, 129.2, 129.1, 128.0, 127.9, 127.2, 126.2, 126.0,
124.4, 119.0, 109.6, 78.0, 60.4, 55.7, 53.3, 28.0, 14.0; HRMS (ESI)
Calcd for C27H36N3O6 ([M + H]+) 498.2599, found 498.2603.
N-Boc-L-Phe-Gly-L-Phe-Gly-OEt (19). Following the general proce-
dure C, the title compound was prepared as a white solid (193 mg,
70% yield): mp 178−180 °C; 1H NMR (400 MHz, MeOD) δ = 7.31−
7.21 (m, 10H), 4.70 (br, 1H), 4.60 (br, 3H), 4.31−4.27 (m, 1H),
4.22−4.17 (q, J = 8.0 Hz, 2H), 3.94−3.89 (m, 3H), 3.67 (d, J = 12.0
Hz, 1H), 3.28−3.11 (m, 2H), 2.98−2.81 (m, 2H), 1.38 (s, 9H), 1.30
(t, J = 8.0 Hz, 3H); 13C NMR (101 MHz, MeOD) δ = 175.0, 173.9,
171.1, 138.5, 130.4, 130.3, 129.5, 129.4, 127.8, 127.7, 62.3, 57.6, 56.0,
43.5, 42.1, 38.9, 38.7, 28.7, 14.5; HRMS (ESI) Calcd for C29H39N4O7
([M + H]+) 555.2813, found 555.2825.
N-Cbz-L-Met-Gly-OEt (14n). Following the general procedure C,
the title compound was prepared as a white solid (167 mg, 91% yield):
mp 74−76 °C (lit.11 74−76 °C); [α]D −19.5 (c 0.2, EtOH) H
NMR (400 MHz, CDCl3) δ = 7.61−7.27 (m, 5H), 6.70 (s, 1H), 5.57
(d, J = 7.9 Hz, 1H), 5.20−5.04 (m, 2H), 4.44 (dd, J = 14.1 Hz, 6.8,
1H), 4.21 (q, 2H), 4.03 (ddd, J = 23.1 Hz, 18.3 Hz, 5.4 Hz, 2H), 2.60
(t, J = 7.0 Hz, 2H), 2.34−1.85 (m, 5H), 1.28 (t, J = 7.0 Hz, 3H); 13C
NMR (101 MHz, CDCl3) δ = 171.3, 169.5, 156.1, 136.1, 128.6, 128.3,
128.1, 67.2, 61.7, 53.7, 41.3, 31.5, 29.9, 15.1, 14.1; HRMS (ESI) Calcd
for C17H25N2O5S ([M + H]+) 369.1479, found 369.1493.
20
1
N-Cbz-D-Phe-Gly-OEt (14o). Following the general procedure C,
the title compound was prepared as a white solid (173 mg, 90% yield):
20
1
mp 109−111 °C; [α]D −17.5 (c 0.57, EtOH) H NMR (400 MHz,
CDCl3) δ = 7.37−7.18 (m, 10H), 6.34 (br, 1H), 5.33 (br, 1H), 5.08 (s,
2H), 4.50 (br, 2H), 4.21 (q, J = 8.0 Hz, 2H), 3.11 (d, J = 12.0 Hz, 2H),
1.28 (t, J = 8.0 Hz, 3H); 13C NMR (101 MHz, CDCl3) δ = 171.0,
169.3, 136.2, 129.3, 128.7, 128.6, 128.2, 128.0, 127.1, 67.2, 61.6, 41.3,
14.1; HRMS (ESI) Calcd for C21H25N2O5 ([M + H]+) 385.1758,
found 385.1757.
Compound 22. Following the general procedure C, the reaction
performed with stearic acid thiol ester (185 mg, 0.50 mmol) afforded
the title compound as a white solid (135 mg, 80% yield): mp 123−124
1
N-Ac-D-Phe-Gly-OEt (14p). Following the general procedure C, the
°C; H NMR (400 MHz, CDCl3) δ = 5.41 (br, 1H), 3.30 (q, 2H),
title compound was prepared as a white solid (132 mg, 91% yield): mp
2.19 (t, J = 8.0 Hz, 2H), 1.66−1.62 (m, 4H), 1.27 (s, 30H), 0.96−0.88
(m, 6H); 13C NMR (101 MHz, CDCl3) δ = 173.0, 39.2, 36.9, 31.9,
31.7, 29.7, 29.6, 29.5, 29.4, 29.3, 25.8, 22.7, 20.0, 14.1, 13.7; IR (film)
3315, 2966, 2921, 2853, 1688, 1618, 1571, 1526, 1449, 1381, 1260,
1171, 1039 cm−1; HRMS (ESI) Calcd for C22H46NO ([M + H]+)
340.3574, found 340.3575.
Compound 23. Following the general procedure C, the title
compound was prepared as a white semisolid (188 mg, 82% yield): 1H
NMR (400 MHz, CDCl3) δ = 7.33−7.26 (m, 3H), 7.14 (d, J = 8.0 Hz,
2H), 5.91 (d, J = 8.0 Hz, 1H), 4.93 (q, 1H), 4.22 (q, 2H), 3.16 (t, J =
4.0 Hz, 2H), 2.39 (t, J = 8.0 Hz, 0.4H), 2.21 (t, J = 8.0 Hz, 1.6H),
1.68−1.59 (m, 2H), 1.28 (s, 31H), 0.92 (t, J = 8.0 Hz, 3H); 13C NMR
(101 MHz, CDCl3) δ = 172.6, 171.7, 135.9, 129.3, 128.5, 127.0, 61.5,
52.9, 37.9, 36.6, 31.9, 29.7, 29.6. 29.5, 29.4, 29.3, 29.2, 25.6, 22.7, 14.1;
HRMS (ESI) Calcd for C29H50NO3 ([M + H]+) 460.3785, found
460.3784.
Compound 25. Following the general procedure C, cinnamoyl-
CoA (10 mg, 1.1 μmol), HMDO (50 μL) and L-Phe-OEt (10.6 mg, 55
μmol) were added to [BMIM]BF4 (0.3 mL). The reaction mixture was
monitored by HPLC analysis (see Supporting Information): 1H NMR
(400 MHz, CDCl3) δ = 7.76 (d, J = 4.0 Hz, 2H), 7.55−7.52 (m, 1H),
7.47−7.43 (m, 2H), 7.33−7.16 (m, 6H), 7.18 (d, J = 8.0 Hz, 2H), 6.62
(br,1H), 5.91 (d, J = 8.0 Hz, 1H), 5.12 (q, J = 8.0 Hz, 1H), 5.91 (q, J =
8.0 Hz, 2H), 3.34−3.24 (m, 2H), 1.32 (t, J = 8.0 Hz, 3H).13C NMR
(101 MHz, CDCl3) δ = 171.6, 135.9, 131.7, 129.4, 128.6, 128.5, 127.1,
127.0, 61.6, 53.5, 37.9, 14.1; HRMS (ESI) Calcd for C20H22NO3 ([M
+ H]+) 324.1594, found 324.1595.
Compound 27. Following the general procedure C, the title
compound was prepared as a white solid (214.5 mg, 65% yield): mp
256−258 °C.1H NMR (400 MHz, CDCl3) δ = 7.31−7.19 (m, 6H),
7.12 (d, J = 4.0 Hz, 2H), 6.01 (d, J = 12.0 Hz, 1H), 5.63 (s, 2H), 4.92−
4.87 (m, 1H), 4.25−4.14 (m, 3H), 3.88−3.80 (m, 2H), 3.24−3.06 (m,
4H), 2.89−2.77 (m, 1H), 2.31 (s, 1H), 2.12−2.11 (m, 1H), 2.01−1.96
(m, 1H), 1.89−1.80 (m, 2H), 1.71−1.58 (m, 8H), 1.34−1.00 (m,
36H), 0.80 (s, 3H), 0.75 (s, 3H), 0.70 (d, J = 12.0 Hz, 1H); 13C NMR
(101 MHz, CDCl3) δ = 129.2, 128.6, 52.6, 47.4, 43.6, 43.1, 39.1, 37.0,
20
133−136 °C; [α]D +2.0 (c 0.50, EtOH) 1H NMR (400 MHz,
CDCl3) δ = 7.29−7.20 (m, 5H), 6.70 (br, 1H), 6.42 (br, 1H), 4.78 (d,
J = 8.0 Hz, 2H), 4.20−4.15 (m, 2H), 3.98−3.91 (m, 2H), 3.09−3.06
(m, 2H), 1.96 (s, 3H), 1.28 (t, J = 8.0 Hz, 2H); 13C NMR (101 MHz,
CDCl3) δ = 171.3, 170.2, 169.3, 136.5, 129.2, 128.6, 127.0, 61.5, 54.2,
41.3, 38.2, 23.1, 14.1; HRMS (ESI) Calcd for C15H21N2O4 ([M +
H]+) 293.1496, found 293.1495.
N-Cbz-Gly-D-Phe-OMe (14q). Following the general procedure C,
the title compound was prepared as a colorless oil (174 mg, 94%
20
1
yield): [α]D +14.8 (c 1.0, EtOH) H NMR (400 MHz, CDCl3) δ =
7.34−7.22 (m, 9H), 7.08 (d, J = 8.0 Hz, 2H), 6.57 (br, 1H), 5.48 (br,
1H), 5.10 (s, 2H), 4.90−4.85 (m, 1H), 3.90−3.79 (m, 2H), 3.70 (s,
3H), 3.13−3.06 (m, 2H); 13C NMR (101 MHz, CDCl3) δ = 171.7,
168.6, 136.1, 135.6, 129.2, 128.6, 128.5, 128.3, 128.1, 127.2, 67.2, 53.1,
52.4, 44.5, 37.8; HRMS (ESI) Calcd for C20H23N2O5 ([M + H]+)
371.1601, found 371.1626.
N-Cbz-L-Phe-Gly-OH (14r). N-Cbz-L-Phe p-toluene thiol ester (156
mg, 0.50 mmol) was dissolved in [BMIM]PF6 (2 mL), followed by
adding HMDO (81 mg, 0.50 mmol), Gly-OH (75 mg, 1.00 mmol)
and NaOH (40 mg). The mixture was heated at 37 °C for 12 h and
then cooled to room temperature. After workup, the title compound
was prepared as a white solid (163 mg, 92% yield): mp 118−120 °C;
[α]D20 −8.7 (c 1.0, EtOH) 1H NMR (400 MHz, DMSO-d6) δ = 8.45−
8.40 (m, 1H), 7.58−7.19 (m, 10H), 4.98−4.90 (m, 2H), 4.34 (t, J =
8.0 Hz, 1H), 3.83−3.77 (m, 2H), 3.07 (d, J = 12.0 Hz, 1H), 2.78 (t, J =
12.0 Hz, 1H); 13C NMR (101 MHz, DMSO-d6) δ = 171.9, 171.1,
155.8, 138.2, 136.9, 129.2, 128.2, 128.0, 127.7, 127.4, 126.2, 65.1, 56.0,
40.7, 37.5; HRMS (ESI) Calcd for C19H21N2O5 ([M + H]+) 357.1445,
found 375.1445.
L-Phe-Gly-OEt (15). Following the general procedure C, N-Boc-L-
Phe-Gly-OEt (15a) was prepared as a white solid (154 mg, 88%
20
1
yield): [α]D +7.2 (c 1.0, EtOH) H NMR (400 MHz, CDCl3) δ =
7.23−7.13 (m, 5H), 6.63 (br, 1H), 5.14 (d, J = 8.0 Hz, 1H), 4.38 (br,
1H), 4.13 (d, J = 8.0 Hz, 2H), 3.97−3.82 (m, 2H), 3.09−2.96 (m,
2H), 1.31 (s, 9H), 1.21 (t, J = 8.0 Hz, 3H); 13C NMR (101 MHz,
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31.8, 28.4, 28.1, 26.5, 23.5, 23.4, 23.3, 18.6, 15.6, 14.2; IR (film) 3504,
3395, 3340, 2965, 2927, 2870, 1738, 1715, 1654, 1618, 1574, 1520,
1456, 1385, 1326, 1248, 1206, 1034 cm−1; HRMS (ESI) Calcd for
C41H60NO5 ([M]+) 646.4466, found 646.4470.
REFERENCES
■
(1) (a) Bioconjugate Techniques, 2nd ed.; Hermanson, G. T., Ed.;
Academic Press: San Diego, 2008. (b) In The Wiley Encyclopedia of
Chemical Biology; Begley, T., Ed; Wiley-VCH: Weinheim, 2008. (c) Li,
K.; Luo, C.; Wang, D.; Zheng, Y. G. Med. Res. Rev. 2012, 32, 815.
(d) Wan, W.; Wang, Y.-S.; Liu, W. R. Org. Chem. Curr. Res. 2012, 1,
e111.
(2) (a) Hang, H. C.; Wilson, J. P.; Charron, G. Acc. Chem. Res. 2011,
44, 699. (b) BEST, M. D.; Rowland, M. M.; Bostic, H. E. Acc. Chem.
Res. 2011, 44, 686. (c) Debets, M. F.; Berkel, S. S. V.; Dommerholt, J.
A.; Dirks, J.; Rutjes, F. P. J. T.; Delft, F. L. V. Acc. Chem. Res. 2011, 44,
805. (d) Chen, Z.; Popp, B. V.; Bovet, C. L.; Ball, Z. T. ACS Chem.
Biol. 2011, 6, 920. (e) Romero, O.; Filice, M.; Rivas, B.; Carrasco-
Lopez, C.; Klett, J.; Morreale, A.; Hermoso, J. A.; Guisan, J. M.;
Abiane, O.; Palomo, J. M. Chem. Commun. 2012, 48, 9053. (f) Xu, G.;
Shin, S. B. Y.; Jaffrey, S. R. ACS Chem. Biol. 2011, 6, 1015. (g) Patra, C.
R.; Rupasinghe, C. N.; Dutta, S. K.; Bhattacharya, S.; Wang, E.; Spaller,
M. R.; Mukhopadhyay, D. ACS Chem. Biol. 2012, 7, 770.
(3) Kent, S. B. H. Chem. Soc. Rev. 2009, 38, 338.
(4) (a) Srinivas, K. V. N. S.; Das, B. J. Org. Chem. 2003, 68, 1165.
(b) Allen, C. L.; Davulcu, S.; Williams, J. M. Org. Lett. 2010, 12, 5096.
(c) Lin, Y. S.; Alper, H. Angew. Chem., Int. Ed. 2001, 40, 779.
(d) Damkaci, F.; DeShong, P. J. Am. Chem. Soc. 2003, 125, 4408.
(5) (a) Dawson, P. E.; Muir, T. W.; Clark-Lewis, I.; Kent, S. B. H.
Science 1994, 266, 776. (b) Dawson, P. E.; Churchill, M. J.; Ghadiri, M.
R.; Kent, S. B. H. J. Am. Chem. Soc. 1997, 119, 432.
(6) (a) McCaldon, P.; Argos, P. Proteins: Struct., Funct., Genet. 1988,
4, 99. (b) Hackenberger, C. P. R.; Schwarzer, D. Angew. Chem., Int. Ed.
2008, 47, 10030.
Modification of Lysozyme with Thioesters. Following the
general procedure C, thioester (0.10 mmol) was added to [BMIM]-
BF4/CH3CO2NH4 buffer solution (20 mM, pH = 7.0, 0.5 mL),
followed by addition of HMDO (20 μL) and lysozyme (20 mg). The
reaction mixture was completed at 37 °C for 24 h and then cooled to
room temperature. The desired product was analyzed by HRMS.
Reaction of Oxo Ester with Amines. The oxo ester and a
catalytic amount of thiol (0.1 equiv) were added to [BMIM]PF6 in a
round-bottom flask. The mixture was heated at 37 °C. After the
completion, the reaction mixture was cooled to room temperature and
concentrated under reduced pressure. The residue was diluted with
ethyl acetate (15 mL) and washed with 1 M aqueous HCl (4 mL), 2 M
aqueous NaOH (4 mL), and brine (4 mL), successively. The organic
layer was collected and dried over anhydrous Na2SO4, filtered, and
concentrated to dryness. The crude product was purified by flash
chromatography to provide the product (25% ethyl acetate in
petroleum ether and then 10% MeOH in DCM).
ASSOCIATED CONTENT
■
S
* Supporting Information
NMR spectra for all compounds. HPLC data for compounds
14o, 14p, 19, and 20. This information is available free of
charge via the Internet at http://pubs.acs.org.
(7) (a) Dawson, P. E.; Kent, S. B. Annu. Rev. Biochem. 2000, 69, 923.
(b) Dhall, A.; Chatterjee, C. ACS Chem. Biol. 2011, 6, 987.
(8) Muir, T. W.; Sondhi, D.; Cole, P. A. Proc. Natl. Acad. Sci. U. S. A.
1998, 95, 6705.
(9) (a) Zhang, L.; Tom, J. P. Tetrahedron Lett. 1997, 38, 4375.
(b) Kurosu, M. Tetrahedron Lett. 2000, 41, 591.
(10) (a) Sletten, E. M.; Bertozzi, C. R. Angew. Chem., Int. Ed. 2009,
48, 6974. (b) Chalker, J. M.; Bemardes, G. J. L.; Davies, B. G. Acc.
Chem. Res. 2011, 44, 730.
(11) Chen, H.; He, M.; Wang, Y.; Cui, Y.; Li, Y.; Zhou, H.; Hong, X.;
Deng, Z. Green. Chem. 2011, 13, 2723.
AUTHOR INFORMATION
■
Corresponding Author
*E-mail: xhy78@whu.edu.cn.
Author Contributions
∥J. Duan and Y. Sun contributed equally to this work.
Notes
The authors declare no competing financial interest.
(12) Wu, W.; Zhang, Z.; Liebeskind, L. J. Am. Chem. Soc. 2011, 133,
14256.
ACKNOWLEDGMENTS
■
We would like to thank Professor Rainer Glaser (University of
MissouriColumbia) for his insightful suggestions and his
contributions to the preparation of the manuscript. We also
thank Professor Lianrong Wang, Professor Yuhui Sun,
Professor Xudong Qu, Professor Changming Zhao, and
Professor Tiangang Liu for their helpful discussions. This
work was supported by the National Science and Technology
Major Project of the Ministry of Science and Technology of
China (No. 2012ZX10004801-003-011), Key Project of
Chinese Ministry of Education (No. 313040), the Specialized
Research Fund of the Doctoral Program of Higher Education
(No. 20110141120017), the Fundamental Research Funds for
the Central Universities (No.2012306020201), National Mega
Project on Major Drug Development (2011ZX09401-302), the
NSFC for several groups led by Professor Z. Deng (No.
31170049, 31130068, 31270119, 31200037, 81273411), the
Fund of State Key Laboratory of Phytochemistry and Plant
Resources in West China (P2010-KF10), and the Fund of State
Key Laboratory of Bioorganic and Natural Products Chemistry,
Shanghai Institute of Organic Chemistry.
(13) Plaquevent, J.-C.; Levillain, J.; Guillen, F.; Malhiac, C.;
Gaumont, A.-C. Chem. Rev. 2008, 108, 5035.
(14) (a) Wenschuh, H.; Beyenmann, M.; Winter, R.; Bienert, M.
Tetrahedron Lett. 1996, 37, 5483. (b) Kim, H.; Gardner, B.; Kahn, M.
Tetrahedron Lett. 1995, 36, 6013.
(15) (a) Tosin, M.; Smith, L.; Leadlay, P. F. Angew. Chem., Int. Ed.
2011, 50, 11930. (b) Tosin, M.; Betancor, L.; Stephens, E.; Li, W. M.
A.; Spencer, J. B.; Leadlay, P. F. ChemBioChem 2010, 11, 539.
(16) Lim, R. K. V.; Lim, Q. Chem. Commun. 2010, 46, 1589.
(17) (a) Yang, W.; Drueckhammer, D. G. J. Am. Chem. Soc. 2001,
123, 11004. (b) Idoux, J. P.; Hwang, P. T. R.; Hancock, C. K. J. Org.
Chem. 1973, 38, 4239. (c) Connors, D. K. A.; Bender, M. L. J. Org.
Chem. 1961, 26, 2498.
(18) Stoeckigt, J.; Zenk, M. H. Z. Naturforsch., C: J. Biosci. 1975, 30C,
352.
(19) Sasin, G. S.; Sasin, R.; Capron, N. J. Org. Chem. 1956, 21, 852.
(20) Jason, D. M. K.; Jasim, M. A. A.; Peter, B. Synth. Commun. 2010,
40, 1161.
(21) Inouye, K.; Voynick, I. M.; Delpierre, G. R.; Fruton, J. S.
Biochemistry 1966, 5, 2473.
(22) Tian, J.; Gao, W.; Zhou, D.; Zhang, C. Org. Lett. 2012, 14, 3020.
(23) Alexis, C.; Mathieu, T.; Karen, W.; Vanessa, R.; Francois, C.;
Gwilherm, M. Org. Lett. 2008, 10, 3841.
(24) Yu, K.; Johnson, R. L. J. Org. Chem. 1987, 52, 2051.
(25) Bergmann, M.; Fruton, J. S. J. Biol. Chem. 1942, 145, 247.
DEDICATION
■
Dedicated to the memory of Professor Richard Loeppky.
Deceased on April 21, 2012.
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