Gram-Scale Solution-Phase Synthesis of Heptapeptide Side Chain of Teixobactin 1

Article abstract of DOI:10.1055/s-0039-1690232

We report herein a scalable synthesis of linear heptapeptide side chain of the depsipeptide natural product teixobactin through solution phase. The synthesis of heptapeptide was achieved through an efficient coupling of suitably protected tripeptide and tetrapeptide comprising of three d-amino acids and four usual l-amino acid subunits.

Full text of DOI:10.1055/s-0039-1690232

SYNLETT
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2
1
4
1
4
3
7
-
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0
9
6
© Georg Thieme Verlag Stuttgart · New York
2019, 30, A–E
letter
A
en
S. Donikela et al.
Letter
Synlett
Gram-Scale Solution-Phase Synthesis of Heptapeptide Side Chain
of Teixobactin1
Sangeetha Donikelaa,b ◊
H2N
O
Kiranmai Nayania ◊
Vishnuvardhan Nomulaa
Prathama S. Mainkara,b
0
0
0
0-0
0
0
1-5
4
0
7-
3
7
9
9
O
O
OH
O
H
H
N
H
N
N
HN
N
H
N
H
linear heptapeptide side chain
0
0
0
0-0
0
0
2-
2
6
9
6-3
7
5
9
O
O
O
O
OH
Srivari Chandrasekhar*a,b
0
0
0
0-0
0
0
3-3
6
9
5-
4
3
4
3
HN
HN
O
linear side chain
a Department of Organic Synthesis and Process Chemistry,
CSIR-Indian Institute of Chemical Technology (IICT),
Hyderabad 500007, Telangana, India
srivaric@iict.res.in
b Academy of Scientific and Innovative Research (AcSIR),
Ghaziabad 201002, India
NH
O
O
H
N
CO2Me
OBn
BocHN
O
NH HN
O
cyclic peptide part
O
key dipeptide
HN
HN
NH
◊ These authors contributed equally to this work
teixobactin
Received: 03.09.2019
Accepted after revision: 14.10.2019
Published online: 25.10.2019
groups to take up total6 and analogues,6c,7–12 synthesis of
teixobactin and to elucidate its pharmacophore.10,12–14 Most
of these use solid-phase synthesis to achieve the desired
target.
DOI: 10.1055/s-0039-1690232; Art ID: st-2019-b0461-l
Abstract We report herein a scalable synthesis of linear heptapeptide
side chain of the depsipeptide natural product teixobactin through
solution phase. The synthesis of heptapeptide was achieved through an
efficient coupling of suitably protected tripeptide and tetrapeptide
comprising of three D-amino acids and four usual L-amino acid sub-
units.
D-Gln
H2N
Ile
Ile
Ser
O
O
O
OH
H
H
N
H
5
3
N
N
HN
N
N
H
4
2
6
7
H
1
O
O
O
Keywords peptides, amino acids, antibiotics, solution-phase synthe-
sis, heptapeptide, depsipeptide
O
OH
HN
O
8
O
HN
D-Thr
D-allo-Ile
Ala
NH
O
O
Ser
D-N-MePhe
9
11
The World Health Organization (WHO) recently re-
leased a list of 12 drug-resistant bacteria which are a threat
to the human population.2 These bacteria have developed
resistance to most of the known antibiotics available in the
market. This includes strains resistant to vancomycin,
which is considered last line of treatment due to its activity
of inhibiting lipid II. In the light of this grim situation, dis-
covery of teixobactin (1) from -proteobacteria, Eleftheria
terrae, is considered a groundbreaking achievement.3 Teixo-
bactin has inhibited growth of Gram-positive bacteria in-
cluding Mycobacterium tuberculosis, and resistance was not
detected under the laboratory conditions. Teixobactin (1)
has been shown to inhibit lipid II and lipid III resulting in
disruption of formation of bacterial cell wall. The accumu-
lation of cell-wall precursor UDP-MurNAc-pentapetide in
the teixobactin treated Staphylococcus aureous cells indicat-
ed that one of the steps in membrane formation were in-
hibited.3,4 Teixobactin comprises of 11 amino acids out of
which four are D-amino acids and is a cyclic depsipeptide
with linear heptamer side chain (1–7) and macrocycle (8–
11) (Figure 1). The presence of unusual amino acid, L-allo-
enduracididine5 (end), 13-membered macrocycle, and its
very important biological activity has prompted several
Ile
O
NH HN
10
O
HN
HN
L-allo-end
NH
Figure 1 Structure of teixobactin (1)
Our group has been working in the area of peptides and
peptidomimetics involving synthesis of unusual amino ac-
ids.15,16 With the global focus shifting to antimicrobial resis-
tance (AMR), we have initiated synthesis of some of the new
antibacterials. Peptide-containing antibacterials are of spe-
cial interest to us and in line with this, total synthesis of
teixobactin was commenced in our group with an objective
to find out new potent lead molecules.
Most of the reports involving synthesis of teixobactin
and analogues are based on solid-phase peptide synthesis.6–12
However, the solution-phase synthesis has its advantages
over solid phase in large-scale feasibility and economically
viable production of small-to-medium-length peptides for
industrial purposes.17 Two groups have reported the syn-
thesis of unusual amino acid, the L-allo-enduracididine part
© 2019. Thieme. All rights reserved. Synlett 2019, 30, A–E
Georg Thieme Verlag KG, Rüdigerstraße 14, 70469 Stuttgart, Germany

B
S. Donikela et al.
Letter
Synlett
of the macrocyle of 1.6b,18 Furthermore, several groups have
concentrated on the synthesis of teixobactin analogues by
changing the side-chain residues8a,9,10,13,19 and enduracidi-
dine residue of the macrocycle,7,8,12,19 whose biological
properties were compared with the natural product. The
studies reveal that changes in the side chain result in de-
creasing the activity, and replacement of enduracididine
residue (end) in the macrocycle does not affect potency of
the antibacterial activity. In addition, the X-ray crystallo-
graphic structure of a derivative of teixobactin reveals that
the unique pattern of hydrophobicity and stereochemistry
of side chain residues (1–7) results in formation of amyloid-
like fibrils, which is responsible for the activity against
Gram-positive bacteria.20 Hence, the conjugates of this hep-
tapeptide side chain with different active/antibiotic motifs
may result in the identification and development of new
potent analogues against antibiotic-resistant bacterial
strains. In a recent publication, Xu et al. completed the syn-
thesis21 of 1 and Reddy et al. achieved analogue synthesis22
using solution phase. Both these groups based their syn-
thetic strategy on hexapeptide (1–6) and macrolide (7–11)
as key intermediates. The approach used by Xu et al. focuses
on coupling of two tripeptides to obtain hexapeptide (1–6),
whereas Reddy et al. have added all six amino acids in a lin-
ear fashion. We observed that the dipeptide part (2–3) and
dipeptide part (5–6) of the heptapeptide side chain are
identical (–NH–Ile–Ser–CO–). Thus, we planned to prepare
this dipeptide in large quantity and couple with the re-
quired amino acids so that a concise (short) and scalable
synthesis could be achieved that will enable total as well as
analogue synthesis of teixobactin. We present herein a
solution-phase synthesis of the heptamer side chain of 1.
Retrosynthesis of teixobactin revealed the presence of a
linear heptamer 2 and cyclic depsipeptide 3. The linear
heptamer fragment 2 would be obtained via coupling of a
tripeptide 4 and tetrapeptide 5 which in turn could be ob-
tained from a key dipeptide 7 (Scheme 1).
The synthesis of both tri- and tetrapeptides began with
commercially available amino acids. The synthesis of
tripeptide 4 was started with a key dipeptide 7 intermedi-
ate. It was obtained from the coupling of commercially
available Boc-L-Ser(OBn)-OH (11) and Boc-L-Ile-OH (12).
The compound 11 was treated with acetyl chloride in
MeOH under reflux for simultaneous conversion of acid
into methyl ester and Boc deprotection to get H-Ser(OBn)-
OMe as a hydrochloride salt, which was coupled with the
acid 12 using N-(3-dimethylaminopropyl)-N′-ethylcarbo-
diimide hydrochloride (EDC·HCl) and hydroxybenzotriazole
teixobactin (1)
NH2
TrtHN
O
O
NH
O
O
O
O
OBn
H
H
N
H
N
N
+
HN
N
H
N
O
NH HN
H
O
O
O
CO2Me
O
OBn
O
BocN
cyclic
depsipeptide 3
HN
HN
linear heptamer 2
NH
TrtHN
O
H
N
CO2Me
OBn
HN
O
OBn
+
H
N
H
O
N
O
FmocHN
N
H
O
BocN
O
CO2Me
tripeptide 4
tetrapeptide 5
TrtHN
O
OH
O
OBn
H
H
N
BocHN
N
CO2Me
O
N
+
BocHN
+
H
BocN
O
CO2Me
OH
O
FmocHN
N-Fmoc-N'-trityl-D-glutamine (8)
OBn
key dipeptide 7
O
6
tripeptide 9
O
H
N
CO2Me
OBn
BocHN
BocHN
OH +
O
10
key dipeptide 7
Scheme 1 Retrosynthesis of teixobactin (1)
© 2019. Thieme. All rights reserved. Synlett 2019, 30, A–E

C
S. Donikela et al.
Letter
Synlett
(HOBt) as coupling reagents and DIPEA as base in dichloro-
methane to give dipeptide 7 in 90% yield over two steps.
N-Methylation23 on Boc-protected D-phenylalnine (13) us-
ing NaH and MeI in THF gave Boc-N-Me-D-phenylalanine
(6) in 95% yield. The dipeptide 7 upon Boc deprotection,24
with TFA/CH2Cl2 (1:1) to get free amine as a TFA salt 7a, fol-
lowed by coupling with the acid 6 using HOBt, EDC·HCl and
DIPEA in CH2Cl2 gave tripeptide 4 in 86% yield over two
steps (Scheme 2).
O
OBn
H
Boc-D-allo-isoleucine (10)
HOBt, EDC⋅HCl, DIPEA
H
N
BocHN
N
CO2Me
OBn
N
TFA⋅H2N
H
0 °C to rt, 18 h
82% (over two steps from 7)
O
CO2Me
O
7a
14
TrtHN
O
1. CH2Cl2/TFA (1:1),
0 ° to rt, 3 h
O
OBn
H
N
H
N
FmocHN
N
2. N-Fmoc-N'-trityl-D-glutamine (8)
HATU, DIPEA, 0 °C to rt, 24 h
86% (over two steps)
H
O
O
CO2Me
1. AcCl, MeOH
H
tetrapeptide 5
BocHN
CO2H
OBn
reflux, 3 h
N
CO2Me
OBn
BocHN
2. HOBt, EDC⋅HCl
Scheme 3 Synthesis of tetrapeptide 5
O
Boc-L-Ile-OH (12)
DIPEA, CH2Cl2, 0 °C to rt,
12 h, 90% (over two steps)
Boc-L-Ser(OBn)-OH (11)
7
O
H
O
H
O
Boc
N
N
CO2Me
OBn
N
HN
OH
OBn
BocHN
OH
HN
OH
LiOH⋅H2O
O
O
O
O
ref. 23
THF/H2O (7:3), 0 °C
1 h, rt, 2 h, 98%
BocN
BocN
4
15
Boc-D-phenylalanine (13)
Boc-N-Me-D-phenylalanine (6)
TrtHN
O
H
H
N
CO2Me
OBn
HN
N
CO2Me
1. diethylamine
CH2Cl2, rt, 3 h
RHN
6, HOBt, EDC⋅HCl
O
OBn
O
H
H
linear heptame
side chain 2
DIPEA
O
O
N
N
OBn
FmocHN
N
0 °C to rt, 18 h
86% (over two steps)
2. HATU, DIPEA, 15
CH2Cl2, 0 °C, 1 h, rt, 24 h
70% (over two steps)
BocN
H
O
O
CO2Me
7; R = Boc
CH2Cl2/TFA (1:1)
0 °C to rt, 2 h
tripeptide 4
7a; R = H
as TFA salt
5
Scheme 4 Synthesis of linear heptamer 2
Scheme 2 Synthesis of tripeptide 4
The dipeptide 7 was also used for the synthesis of tetra-
petide 5. The amine 7a (as TFA salt), obtained from Boc
deprotectction of dipeptide 7 using TFA in CH2Cl2, was cou-
pled with Boc-D-allo-isoleucine (10) using HOBt, EDC·HCl
and DIPEA in CH2Cl2 to give tripeptide 14 in 82% yield over
two steps. The Boc deprotection of tripeptide 14 gave the
corresponding amine which was further coupled with N-
Fmoc-N′-trityl-D-glutamine (8) using 1-[bis(dimethylami-
no)methylene]-1H-1,2,3-triazolo[4,5-b]pyridinium-3-ox-
ide hexafluorophosphate (HATU) and DIPEA to give tetra-
peptide 5 (Scheme 3).
Ester hydrolysis of tripeptide 4 using LiOH·H2O in
THF/H2O gave acid 15. The Fmoc group in tetrapeptide 5
was deprotected with diethylamine25 in CH2Cl2, and the ob-
tained free amine was coupled with acid 15 using HATU
and DIPEA in CH2Cl2 to accomplish heptamer side chain of
teixobactin 2 in 70% yield over two steps (Scheme 4). The
epimerization of amino acids is a major concern in peptide
synthesis. Here, the amount of epimerization during hep-
tamer synthesis was quantified by HPLC. The analysis
showed less than 2% epimerization (see Supporting Infor-
mation).
In conclusion, we have completed a solution-phase con-
vergent synthesis of heptapeptide side chain of the ‘super
antibiotic’, teixobactin on a gram scale in 37% overall yield.
The synthesis was achieved through an efficient coupling of
suitably protected tripeptide and tetrapeptide which were
prepared from a common dipeptide.26,27 This synthetic
scheme will allow us to synthesize analogues of the natural
product. The synthesis of analogues of 1 and their biological
evaluation will be presented in due course.
Funding Information
S. D. thanks University Grants Commission (UGC), New Delhi for re-
search fellowship. K. N. thanks Women Scientists Scheme (WOS-A,
Grant No. SR/WOS-A/CS-58/2017), Department of Science & Technol-
ogy (DST), Government of India for fellowship and research grant. P. S.
M. thanks Indian Council of Medical Research (ICMR), Government of
India for research grant (Grant No. AMR/IN/111/2017-ECD-II). S. C.
thanks the Department of Science and Technology (DST), Govern-
ment of India for a J C Bose fellowship (Grant No. SB/S2/JCB-
002/2015).()
© 2019. Thieme. All rights reserved. Synlett 2019, 30, A–E

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S. Donikela et al.
Letter
Synlett
Acknowledgment
(14) Yang, H.; Du Bois, D. R.; Ziller, J. W.; Nowick, J. S. Chem. Commun.
2017, 53, 2772.
The authors thank the Council of Scientific and Industrial Research
(CSIR) – Indian Institute of Chemical Technology (IICT), Ministry of
Science and Technology, Government of India for research facilities.
(15) For selected papers, see: (a) Chandrasekhar, S.; Babu, B. N.;
Prabhakar, A.; Sudhakar, A.; Reddy, M. S.; Kiran, M. U.;
Jagadeesh, B. Chem. Commun. 2006, 1548. (b) Chandrasekhar, S.;
Kiranmai, N.; Kiran, M. U.; Devi, A. S.; Reddy, G. P. K.; Idris, M.;
Jagadeesh, B. Chem. Commun. 2010, 46, 6962; and the references
cited therein.
Supporting Information
(16) Chandrasekhar, S.; Pendke, M.; Muththe, C.; Akondi, S. M.;
Mainkar, P. S. Tetrahedron Lett. 2012, 53, 1292.
(17) (a) Andersson, L.; Blomberg, L.; Flegel, M.; Lepsa, L.; Nilsson, B.;
Verlander, M. Biopolymers 2000, 55, 227. (b) Chandrudu, S.;
Simerska, P.; Toth, I. Molecules 2013, 18, 4373.
(18) Craig, W.; Chen, J.; Richardson, D.; Thorpe, R.; Yuan, Y. Org. Lett.
2015, 17, 4620.
(19) Parmar, A.; Iyer, A.; Prior, S. H.; Lloyd, D. G.; Goh, E. T. L.;
Vincent, C. S.; Palmai-Pallag, T.; Bachrati, C. Z.; Breukink, E.;
Madder, A.; Lakshminarayanan, R.; Tayler, E. J.; Singh, I. Chem.
Sci. 2017, 8, 8183.
(20) Yang, H.; Wierzbicki, M.; Du Bois, D. R.; Nowick, J. S. J. Am. Chem.
Soc. 2018, 140, 14028.
(21) Gao, B.; Chen, S.; Hou, Y. N.; Zhao, Y. J.; Ye, T.; Xu, Z. Org. Biomol.
Chem. 2019, 17, 1141.
(22) (a) Gunjal, V. B.; Reddy, D. S. Tetrahedron Lett. 2019, 60, 1909.
(b) Dhara, S.; Gunjal, V. B.; Handore, K. L.; Reddy, D. S. Eur. J. Org.
Chem. 2016, 4289.
(23) (a) Reddy, G. V.; Kumar, R. S. C.; Shankaraiah, G.; Babu, K. S.;
Rao, J. M. Helv. Chim. Acta 2013, 96, 1590. (b) Malkov, A.;
Varnkova, K.; Cerny, M.; Kocovsky, P. J. Org. Chem. 2009, 74,
8425. (c) Ferrie, L.; Reymond, S.; Capdevielle, P.; Cossy, J. Org.
Lett. 2006, 8, 3441.
Supporting information for this article is available online at
https://doi.org/10.1055/s-0039-1690232.
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p
p
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gInformati
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p
p
orti
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References and Notes
(1) CSIR-IICT communication number: IICT/Pubs./2019/316.
(2) (a) Willyard, C. Nature 2017, 543, 7643. (b) Von Nussbaum, F.;
Süssmuth, R. D. Angew. Chem. Int. Ed. 2015, 54, 6684; Angew.
Chem. 2015, 127, 6784. (c) Arias, A.; Murray, C. B. E. N. Engl. J.
Med. 2015, 372, 1168. (d) Tobin, K. C. Nature Rev. Microbiol.
2015, 13, 126. (e) Hunter, P. EMBO Rep. 2015, 16, 563. (f) Kostic,
M. Chem. Biol. 2015, 22, 159. (g) Piddock, L. J. V. J. Antimicrob.
Chemother. 2015, 70, 2679.
(3) Ling, L. L.; Schneider, T.; Peoples, A. J.; Spoering, A. L.; Engels, I.;
Conlon, B. P.; Mueller, A.; Schaeberle, T. F.; Hughes, D. E.;
Epstein, S.; Jones, M.; Lazarides, L.; Steadman, V. A.; Cohen, D.
R.; Felix, C. R.; Fetterman, K. A.; Millett, W. P.; Nitti, A. G.; Zullo,
A. M.; Chen, C.; Lewis, K. Nature 2015, 517, 455.
(4) (a) Ng, V.; Chan, W. C. Chem. Eur. J. 2016, 22, 12606. (b) Öster, C.;
Walkowiak, G. P.; Hughes, D. E.; Spoering, A. L.; Peoples, A. J.;
Catherwood, A. C.; Tod, J. A.; Lloyd, A. J.; Herrmann, T.; Lewis, K.;
Dowson, C. G.; Lewandowski, J. R. Chem. Sci. 2018, 9, 8850.
(5) Atkinson, D. J.; Naysmith, B. J.; Furkert, D. P.; Brimble, M. A. Beil-
stein J. Org. Chem. 2016, 12, 2325.
(6) (a) Jin, K.; Sam, L. H.; Laam, Po. K. H.; Lin, D.; Ghazvini Zadeh, E.
H.; Chen, S.; Yuan, Y.; Li, X. Nat. Commun. 2016, 7, 12394.
(b) Giltrap, A. M.; Dowman, L. J.; Nagalingam, G.; Ochoa, J. L.;
Linington, R. G.; Britton, W. J.; Payne, R. J. Org. Lett. 2016, 18,
2788. (c) Zong, Y.; Fang, F.; Meyer, K. J.; Wang, L.; Ni, Z.; Gao, H.;
Lewis, K.; Zhang, J.; Rao, Y. Nat. Commun. 2019, 10, 3268.
(7) Jad, Y. E.; Acosta, G. A.; Naicker, T.; Ramtahal, M.; El-Faham, A.;
Govender, T.; de la Torre, H. G.; Albericio, F. Org. Lett. 2015, 17,
6182.
(8) (a) Parmar, A.; Iyer, A.; Vincent, C. S.; Van Lysebetten, D.; Prior,
S. H.; Madder, A.; Taylor, E. J.; Singh, I. Chem. Commun. 2016, 52,
6060. (b) Parmar, A.; Iyer, A.; Lloyd, D. G.; Vincent, C. S.; Prior, S.
H.; Madder, A.; Taylor, E. J.; Singh, I. Chem. Commun. 2017, 53,
7788. (c) Parmar, A.; Lakshminarayanan, R.; Iyer, A.; Mayandi,
V.; Goh, E. T. L.; Lloyd, D. G.; Chalasani, M. L. S.; Verma, N. K.;
Prior, S. H.; Beuerman, R. W.; Madder, A.; Taylor, E. J.; Singh, I.
J. Med. Chem. 2018, 61, 2009.
(9) Yang, H.; Chen, K. H.; Nowick, J. S. ACS Chem. Biol. 2016, 11,
1823.
(10) Abdel Monaim, S. A. H.; Jad, Y. E.; Acosta, G. A.; Naicker, T.;
Ramchuran, E. J.; El-Faham, A.; Govender, T.; Kruger, H. G.; de la
Torre, B. G.; Albericio, F. RSC Adv. 2016, 6, 73827.
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9712.
(26) Synthetic Procedure for Key Dipeptide Intermediate, Methyl
O-Benzyl-N-[(tert-butoxycarbonyl)-L-isoleucyl]-L-serinate (7)
To a mixture of commercially available Boc-L-Ser(OBn)-OH (11,
5 g, 16.9 mmol, 1.0 equiv) in MeOH (50 mL) under nitrogen
atmosphere, was added MeCOCl (1.8 mL, 25.3 mmol, 1.5 equiv)
slowly at 0 °C. The reaction mixture was stirred at reflux for 3 h,
and the solvent was distilled off in vacuo to get L-Ser(OBn)-OMe
as a hydrochloride (4.1 g, 16.9 mmol). In another round-bot-
tomed flask, Boc-L-isoleucine (12, 3.9 g, 16.9 mmol, 1.0 equiv)
was dissolved in CH2Cl2 (50 mL) under nitrogen atmosphere,
and HOBt (2.5 g, 18.5 mmol, 1.1 equiv) and EDC·HCl (4.85 g,
25.3 mmol, 1.5 equiv) were added sequentially at 0 °C and
stirred for 15 min. To this reaction mixture was added dropwise
a solution of the above-obtained L-Ser(OBn)-OMe hydrochloride
salt (4.1 g, 16.9 mmol, 1.0 equiv) and DIPEA (11.5 mL, 67.6
mmol, 4.0 equiv) in dry CH2Cl2 (20 mL) at 0 °C and stirred for 1
h. Then reaction mixture was maintained at room temperature
for 12 h. The reaction mixture was diluted with CH2Cl2 (100 mL)
and washed with saturated aqueous NH4Cl solution (2 × 75 mL).
The organic layer was separated and washed with saturated
aqueous NaHCO3 solution (2 × 75 mL) followed by brine solution
(75 mL). The organic layer was separated, dried over anhydrous
Na2SO4, and concentrated under vacuum. The residue was puri-
fied by silica gel column chromatography (15% EtOAc in hex-
anes) to give key dipeptide 7 (6.39 g, 90%) as white solid. Rf = 0.5
(11) Abdel Monaim, S. A. H.; Jad, Y. E.; Ramchuran, E. J.; El-Faham, A.;
Govender, T.; Kruger, H. G.; de la Torre, B. G.; Albericio, F. ACS
Omega 2016, 1, 1262.
(12) Wu, C.; Pan, Z.; Yao, G.; Wang, W.; Fang, L.; Su, W. RSC Adv. 2017,
7, 1923.
(13) Parmar, A.; Prior, S. H.; Iyer, A.; Vincent, C.; Van Lysebetten, D.;
Breukink, E.; Madder, A.; Taylor, E. J.; Singh, I. Chem. Commun.
2017, 53, 2016.
20
(20% EtOAc in hexanes); mp 160–162 °C; []D +26.30 (c 1,
CHCl3). 1H NMR (400 MHz, CDCl3):  = 7.36–7.31 (m, 2 H), 7.31–
7.23 (m, 3 H), 6.67 (d, J = 6.9 Hz, 1 H), 5.10 (d, J = 7.4 Hz, 1 H),
4.73 (dt, J = 7.9, 3.1 Hz, 1 H), 4.50 (q, J = 12.2 Hz, 2 H), 4.08–3.98
© 2019. Thieme. All rights reserved. Synlett 2019, 30, A–E

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S. Donikela et al.
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(m, 1 H), 3.89 (dd, J = 9.5, 3.2 Hz, 1 H), 3.72 (s, 3 H), 3.66 (dd, J =
9.5, 3.2 Hz, 1 H), 1.95–1.79 (m, 1 H), 1.51–1.45 (m, 1 H), 1.44 (s,
9 H), 1.20–1.09 (m, 1 H), 0.95 (d, J = 6.8 Hz, 3 H), 0.90 (t, J = 7.4
Hz, 3 H) ppm. 13C NMR (100 MHz, CDCl3):  = 171.4, 170.5,
155.7, 137.5, 128.6, 128.0, 127.8, 79.9, 73.4, 69.6, 59.2, 52.6,
37.8, 28.4, 24.8, 15.6, 11.7 ppm. IR (thin film): max = 3285,
wise to the above reaction mixture at 0 °C and stirred for 30
min. Then reaction mixture was maintained at room tempera-
ture for 18 h. After completion of reaction, the reaction mixture
was diluted with CH2Cl2 (100 mL) and washed with saturated
aqueous NH4Cl solution (2 × 75 mL). The organic layer was sepa-
rated and washed with saturated aqueous NaHCO3 solution (2 ×
75 mL) followed by brine solution (75 mL). The organic layer
was separated, dried over anhydrous Na2SO4, and concentrated
under reduced pressure. The residue was purified by silica gel
column chromatography (25% EtOAc in hexanes) to give tripep-
tide 14 (5.16 g, 82%, over 2 steps) as white solid. Rf = 0.5 (30%
EtOAc in hexanes); mp 134–136 °C; []D20 +13.80 (c 1, CHCl3). 1H
NMR (500 MHz, CDCl3):  = 7.37–7.24 (m, 5 H), 6.75–6.64 (m, 2
H), 5.02 (d, J = 6.6 Hz, 1 H), 4.73 (dt, J = 8.0, 3.2 Hz, 1 H), 4.57–
4.43 (m, 2 H), 4.41 (dd, J = 8.6, 6.3 Hz, 1 H), 4.24–4.10 (m, 1 H),
3.89 (dd, J = 9.5, 3.3 Hz, 1 H), 3.73 (s, 3 H), 3.64 (dd, J = 9.5, 3.0
Hz, 1 H), 2.07–1.95 (m, 1 H), 1.95–1.85 (m, 1 H), 1.58–1.48 (m, 1
H), 1.44 (s, 9 H), 1.42–1.34 (m, 1 H), 1.24–1.12 (m, 2 H), 0.98–
0.87 (m, 9 H), 0.82 (d, J = 6.8 Hz, 3 H) ppm. 13C NMR (125 MHz,
CDCl3):  = 171.8, 170.8, 170.4, 155.8, 137.5, 128.6, 128.1, 127.8,
80.1, 73.4, 69.5, 58.3, 57.5, 52.7, 37.7, 37.3, 28.4, 26.6, 24.9, 15.4,
14.2, 11.8, 11.5 ppm. IR (thin film): max = 3323, 2966, 2930,
2964, 2928, 1707, 1642, 1512, 1367, 1165, 863, 749, 666 cm–1
.
HRMS (ESI): m/z calcd for [M + Na]+ C22H34N2O6Na: 445.2315;
found: 445.2313.
(27) Synthetic Procedure for Tripeptide, Methyl O-Benzyl-N-(tert-
butoxycarbonyl)-D-alloisoleucyl-L-isoleucyl-L-serinate (14)
To a solution of Boc-Ile-Ser(OBn)-OMe (7, 5.0 g, 11.83 mmol, 1.0
equiv) in dry CH2Cl2 (5.0 mL) under nitrogen atmosphere was
added TFA (5.0 mL, 65.2 mmol, 5.53 equiv) at 0 °C and stirred
for 30 min at this temperature. Then reaction was maintained at
room temperature for 2 h. After completion, the volatiles were
removed in vacuo. The residue was dried under high vacuum for
1 h to get the corresponding amine salt (5.2 g, crude) as a pale
brown thick liquid which was used for the next step directly.
Boc-D-allo-isoleucine (10, 2.73 g, 11.80 mmol, 1.0 equiv) was
dissolved in CH2Cl2 (30 mL) under nitrogen atmosphere. HOBt
(1.75 g, 12.98 mmol, 1.1 equiv) and EDC·HCl (3.4 g, 17.7 mmol,
1.5 equiv) were added sequentially at 0 °C. A mixture of amine
salt (5.2 g, crude, ca. 11.9 mmol, 1.0 equiv) and DIPEA (8.08 mL,
47.32 mmol, 4.0 equiv) in dry CH2Cl2 (50 mL) was added drop-
1747, 1658, 1501, 1365, 1211, 1163, 1020, 867, 749, 665 cm–1
.
HRMS (ESI): m/z calcd for [M + Na]+ C28H45N3O7Na: 558.3155;
found: 558.3156.
© 2019. Thieme. All rights reserved. Synlett 2019, 30, A–E