Method to build 2,4-substituted selenazole from β-azido diselenide and carboxylic acid: A formal synthesis of selenazofurin

Article abstract of DOI:10.1016/j.tet.2018.05.010

A novel approach for the synthesis of 2,4-disubstituted selenazoles using carboxylic acids (or anhydrides) and β-azido diselenide was achieved via a one-pot cascade formation of selenazoline (Staudinger reduction/diselenide cleavage/selenocarbonylation/az

Full text of DOI:10.1016/j.tet.2018.05.010

Accepted Manuscript
Method to build 2,4-substituted selenazole from β-azido diselenide and carboxylic
acid: A formal synthesis of selenazofurin
Junfei Qiao, Yi Liu, Yuguo Du
PII:
S0040-4020(18)30518-0
10.1016/j.tet.2018.05.010
TET 29517
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 30 March 2018
Revised Date: 27 April 2018
Accepted Date: 3 May 2018
Please cite this article as: Qiao J, Liu Y, Du Y, Method to build 2,4-substituted selenazole from β-azido
diselenide and carboxylic acid: A formal synthesis of selenazofurin, Tetrahedron (2018), doi: 10.1016/
j.tet.2018.05.010.
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to
our customers we are providing this early version of the manuscript. The manuscript will undergo
copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please
note that during the production process errors may be discovered which could affect the content, and all
legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT
Method to Build 2,4-Substituted Selenazole from β-Azido Diselenide
and Carboxylic Acid: A Formal Synthesis of Selenazofurin
Junfei Qiao, Yi Liu and Yuguo Du*
O
NH
2
Se
N
Se
N
N
3
FDP / Et
CN, 70 oC, 5 h
CH
N / Ph
P
MnO2 / DCE
reflux, 8 h
3
3
R
R
Se
N
MeO
Se
+
RCOOH
OMe
OMe HO
O
3
O
2
O
O
More than 15 examples are given
HO
OH
Selenazofurin is formally synthesized
5

ACCEPTED MANUSCRIPT
Method to Build 2,4-Substituted Selenazole from β-Azido Diselenide and
Carboxylic Acid: A Formal Synthesis of Selenazofurin
1, 2
3
1, 2
Junfei Qiao, Yi Liu and Yuguo Du*
1
State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for
2
Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China; School of
Chemistry and Chemical Engineering, University of Chinese Academy of Sciences, Beijing
3
1
00049, China; School of Chemistry and Chemical Engineering, Yantai University, Yantai
2
64005, China.
Corresponding Author: *E-mail: duyuguo@rcees.ac.cn
ABSTRACT
A novel approach for the synthesis of 2,4-disubstituted selenazoles using carboxylic
acids (or anhydrides) and β-azido diselenide was achieved via a one-pot cascade
formation of selenazoline (Staudinger reduction / diselenide cleavage /
selenocarbonylation / aza-Wittig reaction) and a following MnO -promoted oxidation.
2
This method offers excellent substrate flexibility, and its valuable application is
exemplified by an efficient total synthesis of Selenazofurin which exhibited good
antitumor activities against K562 and A549 cells.
1
. Introduction
The physiological roles of selenium are usually ascribed to selenium-containing
1
proteins that have been identified in the human proteome and in other species.
Biological incorporation of selenium proceeds through selenocysteine which is
considered as the twenty-first proteinogenic amino acid being coded by the UGA
2
codon. Studies also reveal that selenium deficiency is associated with Kashin-Beck
disease, cardiovascular disease, thyroid disease, cognitive decline, male infertility and
3
-5
cancers. There has been an increased attention on selenium-containing compounds
due to their abilities to act as glutathione peroxidase (GPX) mimics, catalytic
6
antioxidants, and cancer-chemopreventive compounds. Accordingly, diverse
selenium derivatives have been prepared and their anticancer activities were
investigated both in vivo and in vitro, suggesting that their main action modes were
carried out by inducing apoptosis of tumor cells and reducing cancerous metastasis in
7
-9
animals.
Other activities were also disclosed with antihypertensive, antiviral,
1
0
antibacterial, and antifungal properties.
In particular, 1,3-selenazoline and 1,3-selenzole, the interesting class of
five-membered heterocycles containing selenium and nitrogen atoms at 1- and
3
-positions respectively, have long been under investigations because of their
1

ACCEPTED MANUSCRIPT
5, 11
outstanding biological and medicinal potentials.
For examples (Fig. 1), a ribose
modified selenazole derivative (selenazofurin 1) exhibited amazing anti-tumor
properties, as well as a broad spectrum of antiviral activities in cell culture
12
experiments. Selenazole 2 showed potent antiproliferative activity against MCF-7
1
3
(
Human breast cancer) cells. More interestingly, when the thiazole structure of
febuxostat was replaced by selenazole, the generated analog 3 provides an excellent
inhibition activity on xanthine oxidase with IC at nM level, which is three times
5
0
1
4
more potent than the marketed reference drug.
Due to the importance of 1,3-selenazoles in drug development, many procedures
6
have been reported to allow the synthesis of more diversified derivatives. Among
them, Hantzch condensation of α-haloketone with selenoureas or selenoamides
5
remains the most widely explored method. However, selenourea analogs are not
1
5
convenient to use because of the high cost and low stability to the air and light. Thus,
an operational convenient and environmental benign method for the synthesis of
1
,3-selenazole is still interesting. Herein, we would like to report our new method for
the preparation of 1,3-selenazoles with β-azido diselenides and carboxylic acids or
anhydrides, as well as the efficient synthesis of selenazofurin 1 using this protocol.
Figure 1. Structures of bioactive 1,3-selenazole compounds
2
. Results and discussion
1
6
Based on our previous research on the phosphine triggered thiazole formation, we
envisaged that its structural analog selenazole could also be made through the
sequential diselenide cleavage / selenocarbonylation / intramolecular Staudinger
Reduction / Wittig reactions using β-azido diselenide and carboxylic compound in
one-pot. To verify this hypothesis, the substrate β-azido diselenide 6 was prepared
from commercial available N-Boc-L-serine methyl ester (4) in two steps (Scheme 1).
O
O
O
a, b
BocHN
MeO
c, d
N3
Se
5
OMe
NHBoc
Se
6
OMe
HO
OMe
Se
MeO
Se
N3
NHBoc
4
O
O
Scheme 1. Synthesis of the β-azido diselenide 6.
2

ACCEPTED MANUSCRIPT
Reagents and conditions: (a) compound 4 (1.0 equiv), Br (2.0 equiv), Ph P (2.0 equiv), imidazole
2
3
(2.0 equiv) in DCM, rt, 2h, 86%; (b) N H •H O/Se/NaOH/DMF, rt, 3h, 70%; (c) TFA/DCM, rt, 3h;
2 4 2
(
d) NaN /Tf O, ZnSO , Et N, CH OH/H O/DCM, 0 ºC, overnight, 44% over two steps.
3
2
4
3
3
2
Thus, compound 4 was treated with triphenylphosphine, bromine, and imidazole
in dry dichloromethane (DCM) to convert hydroxyl group into the corresponding
bromide in 86% yield. The bromide was added to a DMF solution containing a
mixture of elemental selenium, hydrazine monohydrate, and NaOH. The reaction
between selenium and hydrazine in the presence of NaOH gave Na Se , which
2
2
condensed with the bromide to give the N-Boc-protected homoselenocystine 5. It is
worth to note that the molar ratio between selenium and NaOH had better be kept at
1
:1 to ensure the predominant product diselenide 5. Treatment of 5 with 10% TFA in
DCM removed Boc group, followed by ZnSO promoted azide transfer using freshly
4
1
7
prepared TfN , obtained the expected β-azido diselenide 6 in 44% overall yield.
3
a
Table 1. Screening of the reaction conditions for selenazoline synthesis
Entry
Coupling
reagent
EDCI
FDP
Base
Time
Temperature
Solvent
Yield
(%)
23
o
(h)
5
( C)
1
2
3
4
5
6
7
8
9
DIPEA
reflux
reflux
reflux
40
DCM
DCM
DCM
Et N
3
5
29
FDP
NMP
5
10
EDCI
FDP
DIPEA
5
CH CN
70
3
Et N
3
5
40
CH CN
73
3
FDP
Et N
3
5
70
CH CN
84
3
FDP
Et N
3
5
70
DMF
49
FDP
Et N
3
5
70
dioxane 30
FDP
Et N
3
5
70
toluene
40
45
1
0
FDP
Et N
3
3
70
CH CN
3
a
Unless otherwise noted, 6 (1.0 equiv), acid (4.0 equiv), coupling reagent (4.0 equiv), base (8.0
equiv), Ph P (8.0 equiv).
3
With substrate 6 in hand, we next investigated the possibility of making
selenazoline, the precursor of selenazole preparation. Following our previous
16
synthesis of 2,4-disubstituted thiazoline, β-azido diselenide 6 was condensed with
benzoic acid in the presence of Ph P, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide
3
3

ACCEPTED MANUSCRIPT
hydrochloride (EDCI) and 1,1'-Dimethyltriethylamine (DIPEA) in DCM under reflux.
However, the desired selenazoline product 7 was obtained in an unacceptable low
yield (23%, Table 1, entry 1). A series of experimental results suggested that the
variation of coupling agent and base did not improve the reaction outcome much
(
Table 1, entries 1-3), until we realized that the stability of the Staudinger
1
8
intermediate during selenazoline formation might be crucial. We found that the
solvent had a great effect on the reaction in terms of the yield, and the relative polar
solvent CH CN was found to be the best in selenazoline synthesis (entries 4-9). This
3
is quite different from that of in thiazoline synthesis wherein the nonpolar solvent
DCM was favorable to the target formation. The reaction time and temperature were
also important factors impacting on the reaction (entries 5, 6, 10). Thus, an optimized
typical process for selenazoline synthesis was exemplified as follows: condensation of
β-azido diselenide 6 with benzoic acid in the presence of pentafluorophenyl
o
diphenylphosphate (FDP), Ph P, and Et N in CH CN at 70 C for 5h afforded methyl
3
3
3
2
-pheny1-1,3-selenazoline-4-carboxylate (7) in good yield (84%). The transformation
of 6 into 7 could be explained as shown in the following proposed pathway (Figure 2).
PPh3
O
N3
Ph P
N
Se Se
N PPh3
Ph P
3
N
Se
Se
OMe
PPh3
3
MeO
Se N3
O
O
O
Staudinger reaction
OMe
OMe
MeO
O
6
O
F
F
O
O
F
O
P
OPh
P
O
OPh
OPh
O
P
+
COOH
F
F
OPh
F
F
OPh
F
F
O
O
OPh
Selenocarbonylation
F
Ph
Se
N
O
aza-Wittig reaction
Ph
OMe
3
Ph P
N
Se
O
O
7
OMe
Figure 2. Proposed pathway for the transformation of compound 6 into 7.
Table 2. Exploration of conditions to convert selenazoline to the corresponding
selenazole
Se
N
Se
N
oxidation
Ph
Ph
OMe
OMe
O
O
7
8
Entry Oxidation reagent
Temperature
Time
Yield (%)
4

ACCEPTED MANUSCRIPT
a
1
2
3
4
5
I2 / DCM
rt
24h
24h
8h
0
a
I2 / THF
rt or reflux
0
b
o
IBX / DMSO
45 C
0
c
DBU / BrCCl / DCM
rt
8h
45-71
94
3
d
o
Fresh MnO / DCE
70 C
8h
2
a
b
c
selenazoline 7 (1.0 equiv), I (2.0 equiv). IBX (1.0 equiv). BrCCl (1.0-4.0 equiv), DBU
2
3
d
(
2.0-8.0 equiv). MnO (10 equiv)/DCE, 70 ºC, 8h.
2
1
9,20
There are very few literatures
about the direct oxidation of selenazoline to
selenazole. Our exploration indicated that the activated MnO in 1,2-dichlorethane
2
(DCE) was promising in case of the substrate 7 (Table 2), and the followed scope
research results are summarized in Table 3. The current method is applicable to a wide
variety of carboxylic substrates (acids and anhydrides, entries 1-12), except for the
relatively strong organic acids which might cause intermediate decomposition (entries
3
and 7). Curious about the chiral stability at selenazoline C-4, compound 9b was
o
subjected to Et N in CD OD at 60 C for 4 h. A clear hydrogen–deuterium exchange
3
3
was observed at C-4 in proton NMR experiment, presumably through base-catalyzed
ester-enol epimerization (see ESI Fig. S1). Attempts to prepare the selenazole via
1
6
one-pot five-step reaction following an established procedure gave low yields of
product, presumably due to the easy decomposition of -bromoester selenazoline
α
intermediate under BrCCl oxidation. We thus prepared the desired selenazole in two
3
steps, i.e., to prepare selenazoline via four-reaction in one-pot and followed by a
further MnO oxidation. It was reported that racemization on the α-stereogenic
2
21,22
position of thiazole was encountered with Hantzsch method.
To check this, the
optically pure carboxylic acid 11 and Boc-D-Phenylalanine 12 were subjected to the
selenazole formation under our reaction conditions (entries 14, 15), respectively. The
experiments turned out that both compounds were good substrates for this reaction
and no detectable racemization was observed in the corresponding selenazoles (see
ESI Fig. S2-S5). This finding certainly enlarges the potential applications of our
1
9, 23
method comparing to the reported methods.
a, b
Table 3. Synthesis of 1,3-selenazolines and selenazoles
O
Se
Se
N
RCOOH
MnO2 / DCE
70 oC, 8 h
FDP / Et N / Ph P
3
3
R
R
N3 Se
Se N3
OMe +
OMe
OMe
or
RCO) O
N
CH CN, 70 oC, 5 h
3
MeO
(
2
O
O
O
6
9a-l
1
0a-l
Entry
Substrate
Product Yield (%) Product
Yield (%)
5

ACCEPTED MANUSCRIPT
1
2
3
4
5
6
7
8
9
1
2-I-PhCOOH
9a
9b
9c
9d
9e
9f
9g
9h
9i
60
65
0
10a
10b
10c
10d
10e
10f
10g
10h
10i
10j
10h
10i
8
89
92
/
4-Cl-PhCOOH
4-NO -PhCOOH
2
3-Br-PhCOOH
75
65
71
0
90
93
92
/
2-OAc-PhCOOH
3-CH -PhCOOH
3
CF COOH
3
n-BuCOOH
60
71
65
54
66
76
95
93
90
95
93
94
n-C H COOH
5
11
0
n-C H COOH
9j
9h
9i
7
15
c
c
c
11
n-butyric anhydride
n-caproic anhydride
benzoic anhydride
1
2
3
1
7
1
4
5
9k
9l
77
62
10k
10l
90
91
(11)
1
(12)
a
Unless otherwise noted, β-azido diselenide 6 (1.0 equiv), acid (4.0 equiv), FDP (4.0 equiv), Et N
3
b
c
(
8.0 equiv), Ph P (8.0 equiv), 70 ºC in CH CN, 5h. MnO (10 equiv)/DCE, 70 ºC, 8h. coupling
3 3 2
reagent FDP was not added.
As the methodology has been established, we then focused on the application of
our method to a formal synthesis of 2- -D-ribofuranosylselenazole-4-carboxamide
selenazofurin 1). Being an anti-tumor drug candidate under clinical evaluation,
β
2
4
(
2
5
selenazofurin is 10-fold more dose potent than its sulfur counterpart in several in
vitro and in vivo antitumor screenings acting as an inhibitor of inosine monophosphate
dehydrogenase (IMPD) via selenazofurin adenosine dinucleotide (SAD).
6

ACCEPTED MANUSCRIPT
Scheme 2. Formal synthesis of Selenazofurin 1.
Reagents and conditions: (a) β-azido diselenide 6 (1.0 equiv), 13 (4.0 equiv), FDP (4.0 equiv),
Et N (8.0 equiv), Ph P (8.0 equiv), 70 ºC in CH CN, 5h, 70%; (b) MnO (10 equiv)/DCE, 70 ºC,
3
3
3
2
8
h, 85%; (c) MsOH/CH Cl, rt, 8h, 90%; (d) CH OH/NH , rt, 5h, in quantitative.
3 3 3
Our strategy towards the total synthesis of selenazofurin is depicted in Scheme 2.
The precursor, 2,3,5-tri-O-benzyl- -D-ribofuranosyl formic acid (13), was
synthesized from commercially available D-Ribose following a combination of two
β
2
3, 26
literature procedures.
FDP, Ph P, and Et N in CH CN afforded ribofuranosyl selenazoline 14 (Scheme 2) in
Treatment of 13 with β-azido diselenide 6 in the presence of
3
3
3
7
0% yield as an enantiomeric mixture at C-4 of selenazoline (R/S = 1.00:1.38, ESI Fig.
S6). The crude mixture was further oxidized with active MnO in DCE affording the
2
expected ribosyl selenazole 15 in 85% isolated yield. Coupling constant of J Hz at 2.8
1
ppm in H NMR spectrum clearly indicated that ribosyl selenazole 15 was in β
2
6
configuration. No
α product was observed in our studies. This was quite
encouraging as the anomeric mixture of ribofuranosyl residue was obtained from
2
5
other reported method. Hydrogenolysis of 15 with various Pd catalysts on carbon
did not give a complete removal of benzyl ethers, possibly due to the poisoning of Pd
2
7
catalyst by selenium in the substrate. To our delight, the clean cleavage of benzyl
ethers from 15 was carried out smoothly with methanesulfonic acid in CHCl to
3
afford the desired 16 in a yield of 90%. Finally, treatment of 16 with NH in methanol
3
furnished selenazofurin 1 quantitatively. The physical data of the synthetic
2
8
selenazofurin 1 is in consistent with that of the reported previously.
The antitumor activity of the synthetic 1 was evaluated against three tumor cell
2
9
lines (K562, A549 and HCT-116) according to a literature procedure, and the results
are outlined in Table 4. Compound 1 exhibited good antitumor activities against K562
and A549 cells with IC at 0.29 and 0.85 µM, respectively.
5
0
Table 4. In Vitro antitumor activity of the synthetic compound 1
7

ACCEPTED MANUSCRIPT
IC (µM)
5
0
Compound
K562
0.29
A549
0.85
HCT-116
>10
1
Taxol
0.0026
0.0026
0.0025
3
. Conclusions
We have successfully demonstrated a facile transformation of dialkyl diselenides into
selenazoles in good yields via a cascade Staudinger reduction / diselenide cleavage /
selenocarbonylation / aza-Wittig reaction and a following oxidation. The method
provides a broad substrate applicability and its potentials are also exemplified by an
efficient total synthesis of Selenazofurin which exhibited good antitumor activities
against K562 and A549 cells. Further applications of this strategy to the design and
synthesis of other related selenazole-containing bioactive compounds are currently
under investigation in our laboratory.
4
4
. Experimental section
.1 General Experimental
All solvents were dried according to the established procedures before use. All
reagents were purchased from commercial corporations. Flash chromatography (FC)
was performed using silica gel (200−300 meshes) according to the protocol of Still,
3
0
Kahn, and Mitra. All reactions under standard conditions were monitored by
thin-layer chromatography (TLC) on gel F254 plates. Optical rotations were measured
using a polarimeter. High-resolution mass spectrometry data (HRMS) were acquired
1
13
using a Q-TOF analyzer. H NMR and C NMR were measured on 400 MHz and 100
MHz spectrometers (NMR in CDCl with TMS as an internal standard). Chemical
3
shifts (δ) are given in parts per million and coupling constants (J) in Hertz.
4
.2. Experimental Procedures
Synthesis of Boc-L-selenocystine methyl ester (5)
To a pre-cooled mixture of triphenylphosphine (4.40 g, 16.8 mmol) in dry DCM (50
mL) was dropwisely added a solution of Br (0.86 mL, 16.8 mmol) in DCM (20 mL)
2
at 0 °C under nitrogen protection. After 20 min, a mixture of N-Boc-L-serine methyl
ester (1.84 g, 8.4 mmol) and imidazole (1.14 g, 16.8 mmol) in DCM (50 mL) was
slowly added to the above pale yellow solution and the mixture was stirred for another
two hours at 0 °C. Subsequently, the solids were filtered off and the solvent was
evaporated. The resulting solid was re-dissolved in DCM (10 mL) and purified by
column chromatography (hexanes /EtOAc, 6:1) to give the bromide intermediate in
8
6% yield (2.03 g, 7.2 mmol).
To a suspension of Se (3.945 g, 47.6 mmol) and pulverized NaOH (1.998 g, 47.6
8

ACCEPTED MANUSCRIPT
mmol) in DMF was added N H ·H O (0.4 mL). The mixture was stirred under argon
2
4
2
for 3h at 60 °C, then the above bromide (1.9 g, 6.8 mmol) was added dropwise and
stirred for 3h further. TLC indicated complete disappearance of the starting material,
and the mixture was poured into aqueous HCl (1 M, 50 mL) and extracted with DCM
(
3 x 50 mL). The combined organic solvent was dried over Na SO and evaporated in
2 4
vacuo. The crude mixture was purified by FC (silica gel, hexanes/EtOAc 2:1) to give
25
D
compound 5 in 70% yield as a yellow solid (1.33 g). m.p. 36-37 ºC; [α] 68.0 (c 1.0,
1
CHCl ); H NMR (400 MHz, MeOD), δ (ppm) 5.39 (d, J = 7.6 Hz, 1H, NH), 4.61-
3
13
4
.62 (m, 1H, CH), 3.75 (s, 3H, CH ), 3.32-3.43 (m, 2H, CH ); C NMR (100 MHz,
3 2
MeOD), δ (ppm) 28.5, 32.5, 52.8, 53.8, 80.5, 155.2, 171.4; HRMS (ESI): calcd for
+
C H N O Se , [M+Na] 587.0387; found: 587.0369.
18
32
2
8
2
Synthesis of (2R, 2'R)-dimethyl 3,3'-diselanediylbis(2-azidopropanoate) (6)
In flask one, selenocystine methyl ester 5 (562 mg, 1 mmol) was dissolved in TFA
(1.5 mL) in dry DCM (15 mL) and the mixture was stirred for 3 h. When TLC
indicated complete reaction, the mixture was evaporated under reduced pressure to
give L-selenocystine methyl ester trifluoroacetic acid salt quantitatively as a yellow
2
5
solid. m.p. 148-149 ºC; [α] D -39.0 (c 1.0, CH3OH); 1H NMR (400 MHz, MeOD), δ
(
ppm) 3.44-3.49 (m, 2H, CH ), 3.56-3.60 (m, 2H, CH ), 3.88 (s, 6H, OCH ), 4.45 (m,
2 2 3
2
H, CH); 13C NMR (100 MHz, MeOD), δ (ppm) 28.4, 54.1, 54.4, 169.7; HRMS
+
(ESI): calcd for C H N O Se , [M+H] 364.9519; found: 364.9500.
8
16
2
4
2
In flask two, a solution of NaN (390 mg, 6 mmol) in H O (1 mL) and CH Cl (1 mL)
3
2
2
2
was added Tf O (846 mg, 3 mmol) and stirred vigorously for 2 h at 0 °C. The biphasic
2
mixture was neutralized with saturated aqueous NaHCO (1 mL), extracted with
3
CH Cl (2 x 1 mL), and the combined organic phase was washed with saturated
2
2
aqueous NaHCO solution (2 x 1 mL) again. The resulting TfN solution in CH Cl
3
3
2
2
was used directly without further purification.
To a pre-mixed mixture of L-selenocystine methyl ester TFA salt (662 mg, 1 mmol),
ZnSO (4 mg, 0.02 mmol) and Et N (253 mg, 2.5 mmol) in MeOH (6.7 mL) and
4
3
water (1.3 mL) was added dropwise the freshly prepared TfN solution at 0 °C. The
3
homogeneous mixture was stirred at 0 °C until the reaction completed (TLC
monitoring), then quenched by addition of phosphate buffer (pH 3). The pH of the
aqueous layer was carefully adjusted to pH 2 by addition of dilute aqueous HCl. The
water phase was separated and extracted with CH Cl (3 x 5 mL), and the combined
2
2
organic layer was washed with brine, dried over Na SO , and concentrated in vacuo.
2
4
The residue was purified by FC (silica gel, hexanes/EtOAc 3:1) to give compound 6
25
1
as an orangered oil (0.85 g, 44% for two steps). [α] D -35.6 (c 1.0, CHCl ); H NMR
3
(
400 MHz, CDCl ), δ (ppm) 3.22 (dd, J = 8.0 Hz, J = 12.8 Hz, 1H); 3.37 (dd, J = 5.6
3
1
3
Hz, J = 12.8 Hz, 1H); 3.84 (s, 3H); 4.24-4.27 (m, 1H); C NMR (100 MHz, CDCl3),
9

ACCEPTED MANUSCRIPT
+
δ (ppm) 30.2, 53.2, 62.6, 169.6; HRMS (ESI): calcd for C H N O Se , [M+Na]
8
12
6
4
2
4
38.9148; found: 438.9137.
General Procedure for Selenazoline Formation.
Typical procedure for the synthesis of methyl-2-phenylselenazoline-carboxylate 7. To
a solution of benzoic acid (122 mg, 1 mmol) in anhydrous acetonitrile (10 mL) were
added triethylamine (Et N) (202 mg,
2
mmol) and pentafluorophenyl
3
diphenylphosphate (FDP) (416 mg, 1 mmol) at room temperature. After stirring for 5
min, dialkyl β-azido diselenide 6 (104 mg, 0.25 mmol) and Ph P (524 mg, 2 mmol)
3
were added, and the mixture was stirred at 70 ℃ for another 5h, then concentrated
under reduced pressure. The residue was purified by FC (silica gel, hexanes/EtOAc
1
0:1) to give compound 7 as yellow oil (113 mg).
General Procedure to Oxide Selenazoline to Selenazole
Typical procedure for the formation of methyl-2-phenylselenazole-carboxylate 8. To a
solution of methyl-2-phenylselenazoline-carboxylate 7 (177 mg, 1 mmol) in
1
,2-dichloroethane (10 mL) was added activated MnO (860 mg, 10 mmol). The
2
o
mixture was then heated at 70 C for 8 h under a nitrogen atmosphere. After filtration,
the mixture was evaporated in vacuo. The residue was chromatographed on silica gel
(hexanes/EtOAc 8:1) to give compound 8 (105 mg).
Methyl 2-(2-iodophenyl)selenazoline-4-carboxylate (7)
1
The title compound was obtained in 84% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 3.71-3.78 (m, 2H), 3.81 (d, J = 4.0 Hz, 3H), 5.22 (t, J = 8.8 Hz, 1H),
3
1
3
7
.37−7.40 (m, 2H), 7.44−7.47 (m, 1H), 7.78 (d, J = 7.2 Hz, 2H); C NMR (100 MHz,
CDCl ), δ (ppm) 31.5, 52.9, 81.0, 128.7, 129.3. 131.8, 135.1, 169.7, 171.4; HRMS
3
+
(
ESI): calcd for C H NO Se, [M+Na] 291.9853; found: 291.9860.
11
11
2
Methyl 2-phenylselenazole-4-carboxylate (8)
The title compound was obtained in 94% yield according to the typical procedure as
1
white solid. m.p. 108-110 ºC; H NMR (400 MHz, CDCl ), δ (ppm) 3.96 (d, J = 1.6
3
13
Hz, 3H), 7.41−7.49 (m, 3H), 7.93-7.96 (m, 2H), 8.88 (s, 1H); C NMR (100 MHz,
CDCl ), δ (ppm) 52.7,127.6, 129.2, 131.0, 134.6, 135.7, 149.0, 162.4, 175.4; HRMS
3
+
(
ESI): calcd for C H NO Se, [M+Na] 289.9696; found: 289.9702.
11
9
2
Methyl 2-(2-iodophenyl)selenazoline-4-carboxylate (9a)
1
The title compound was obtained in 60% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 3.82-3.95 (m, 2H), 3.84 (s, 3H), 5.25 (t, J = 9.0 Hz, 1H), 7.04-7.09
3
(
m, 1H), 7.35 (td, J = 0.8 Hz, J = 7.6 Hz, 1H), 7.45 (dd, J = 1.2 Hz, J = 7.6 Hz,
1 2 1 2
1
3
1
3
H), 7.88 (dd, J = 0.8 Hz, J = 8.0 Hz, 1H); C NMR (100 MHz, CDCl ), δ (ppm)
1 2 3
3.3, 53.0, 80.7, 94.4, 128.2, 129.5, 131.3, 140.3, 140.4, 170.7, 170.9; HRMS (ESI):
+
calcd for C H INO Se, [M+Na] 417.8814; found: 417.8822.
11
10
2
1
0

ACCEPTED MANUSCRIPT
Methyl 2-(2-iodophenyl)selenazole-4-carboxylate (10a)
1
The title compound was obtained in 89% yield as a yellowish oil (89%): H NMR
(
400 MHz, CDCl ), δ (ppm) 3.96 (s, 3H), 7.11 (td, J = 1.6 Hz, J = 8.0 Hz, 1H),
3 1 2
7
9
1
.40-7.44 (m, 1H), 7.82 (dd, J = 1.6 Hz, J = 8.0 Hz, 1H), 7.98 (d, J = 8.0 Hz, 1H),
1 2
1
3
.06 (s, 1H); C NMR (100 MHz, CDCl ), δ (ppm) 52.7, 96.9, 128.5, 131.3, 131.3,
3
37.0, 140.1, 140.7, 147.8, 162.4, 174.5; HRMS (ESI): calcd for C H INO Se,
11
8
2
+
[
M+Na] 415.8663; found: 415.8664.
Methyl 2-(4-chlorophenyl)selenazoline-4-carboxylate (9b)
1
The title compound was obtained in 65% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 3.75-3.81 (m, 2H), 3.96 (s, 3H), 5.21 (t, J = 9.0 Hz, 1H), 7.72 (d, J =
3
1
3
8
5
.4 Hz, 2H), 7.38 (d, J = 8.8 Hz, 2H); C NMR (100 MHz, CDCl ), δ (ppm) 31.9,
3
3.0, 80.9, 129.0, 130.5, 133.6, 138.0, 168.6, 171.2; HRMS (ESI): calcd for
+
C H ClNO Se, [M+Na] 325.9463; found: 325.9439.
11
10
2
Methyl 2-(4-chlorophenyl)selenazole-4-carboxylate (10b)
1
The title compound was obtained in 92% yield as white solid. m.p. 136-138 ºC; H
NMR (400 MHz, CDCl ), δ (ppm) 3.96 (s, 3H), 7.41 (d, J = 8.4 Hz, 2H), 7.88 (d, J =
3
1
3
8
1
.4 Hz, 2H), 8.89 (s, 1H); C NMR (100 MHz, CDCl ), δ (ppm) 52.7, 128.7(2),
3
29.5(2), 134.2, 134.8, 137.1, 149.1, 162.2, 173.9; HRMS (ESI): calcd for
+
+
C H ClNO Se, [M+Na] 323.9298; found: 323.9292, [M+K] 339.9038; found:
11
8
2
3
39.9077.
Methyl 2-(3-bromophenyl)selenazoline-4-carboxylate (9d)
1
The title compound was obtained in 75% yield as colorless oil. H NMR (400 MHz,
CDCl ), δ (ppm) 3.75-3.82 (m, 2H), 3.84 (s, 3H), 5.23 (t, J = 8.8 Hz, 1H), 7.27 (t, J =
3
13
8
.0 Hz, 1H), 7.58-7.60 (m, 1H), 7.67 (d, J = 8.0 Hz, 1H), 7.98 (m, 1H); C NMR (100
MHz, CDCl ), δ (ppm) 33.2, 54.3, 82.2, 124.2, 129.5, 131.6, 133.0, 136.0, 138.3,
3
+
1
69.6, 172.4; HRMS (ESI): calcd for C H NO Se, [M+Na] 369.8949; found:
11
11
2
3
69.8973.
Methyl 2-(3-bromophenyl)selenazole-4-carboxylate (10d)
1
The title compound was obtained in 90% yield as white solid. m.p. 122-123 ºC; H
NMR (400 MHz, CDCl ), δ (ppm) 3.97 (s, 3H), 7.31 (t, J = 8.0 Hz, 1H), 7.60 (dt, J =
3
1
13
0
.8 Hz, J = 8.0 Hz, 1H), 7.84-7.85 (m, 1H), 8.13 (m, 1H), 8.92 (s, 1H); C NMR
2
(
100 MHz, CDCl ), δ (ppm) 52.8, 123.4, 126.3, 130.1, 130.7, 133.8, 135.2, 137.3,
3
+
1
3
49.1, 162.2, 173.4; HRMS (ESI): calcd for C H INO Se, [M+Na] 367.8792; found:
11
8
2
67.8795.
Methyl 2-(2-acetoxyphenyl)selenazoline-4-carboxylate (9e)
1
The title compound was obtained in 65% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 2.36 (s, 3H), 3.70 (d, J = 8.4 Hz, 2H), 3.83 (s, 3H), 5.16 (t, J = 9.2
3
Hz, 1H), 7.13 (d, J = 8.0 Hz, 1H), 7.27-7.31 (m, 1H), 7.47 (t, J = 7.6 Hz, 1H), 7.81 (d,
1
3
J = 7.6 Hz, 1H); C NMR (100 MHz, CDCl ), δ (ppm) 21.6, 31.1, 52.9, 80.6, 123.7,
3
1
1

ACCEPTED MANUSCRIPT
1
26.3, 127.8, 131.6, 132.2, 148.3, 164.3, 169.5, 171.2; HRMS (ESI): calcd for
+
C H NO Se, [M+Na] 349.9907; found: 349.9932.
1
1
11
2
Methyl 2-(2-acetoxyphenyl)selenazole-4-carboxylate (10e)
1
The title compound was obtained in 93% yield as white solid. m.p. 155-156 ºC; H
NMR (400 MHz, CDCl ), δ (ppm) 2.49 (s, 3H), 3.97 (s, 3H), 7.24-7.26 (m, 1H), 7.35
3
1
3
(
t, J = 7.6 Hz, 1H), 7.49 (t, J = 7.6 Hz, 1H), 8.28 (t, J = 7.6 Hz, 1H), 8.98 (s, 1H); C
NMR (100 MHz, CDCl ), δ (ppm) 22.0, 52.6, 123.7, 126.6, 127.6, 129.8, 131.4, 135.5,
3
+
1
47.8, 148.2, 162.4, 167.9, 168.9; HRMS (ESI): calcd for C H NO Se, [M+Na]
11 11 2
3
47.9751; found: 347.9779.
Methyl 2-(4-methylhenyl)selenazoline-4-carboxylate (9f)
1
The title compound was obtained in 71% yield as a pale yellow oil. H NMR (400
MHz, CDCl ), δ (ppm) 2.36 (s, 3H), 3.70-3.78 (m, 2H), 3.82 (s, 3H), 5.20 (t, J = 8.8
3
13
Hz, 1H), 7.19 (d, J = 8.0 Hz, 2H), 7.67 (d, J = 8.0 Hz, 2H); C NMR (100 MHz,
CDCl ), δ (ppm) 21.6, 31.3, 52.9, 80.8, 129.2(2), 129.4(2), 132.3, 142.3, 169.5, 171.5;
3
+
HRMS (ESI): calcd for C H NO Se, [M+Na] 306.0009; found: 306.0026.
1
2
13
2
Methyl 2-(4-methylphenyl)selenazole-4-carboxylate (10f)
1
The title compound was obtained in 92% yield as white solid. m.p. 115-117 ºC; H
NMR (400 MHz, CDCl ), δ (ppm) 2.35 (s, 3H), 3.94 (s, 3H), 7.21 (d, J = 8.0 Hz, 2H),
3
1
3
7
5
.81 (d, J = 7.6 Hz, 2H), 8.83 (s, 1H); C NMR (100 MHz, CDCl ), δ (ppm) 21.6,
3
2.6, 127.4, 129.8, 132.9, 134.1, 141.4, 148.7, 162.3, 175.5; HRMS (ESI): calcd for
+
C H NO Se, [M+H] 282.0033; found: 282.0038.
12
11
2
Methyl 2-propylselenazoline-4-carboxylate (9h)
1
The title compound was obtained in 60% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 0.96 (t, J = 7.4 Hz, 3H), 1.60−1.70 (m, 2H), 2.59 (m, 2H), 3.64 (dd,
3
1
3
J = 8.8 Hz, J = 1.2 Hz, 2H), 3.79 (s, 3H), 4.98 (t, J = 8.8 Hz, 1H); C NMR (100
1
2
MHz, CDCl ), δ (ppm) 13.7, 21.7, 31.4, 39.6, 52.9, 80.1, 171.4, 173.7; HRMS (ESI):
3
+
calcd for C H NO Se, [M+Na] 258.0009; found: 257.9987.
8
13
2
Methyl 2-propylselenazole-4-carboxylate (10h)
1
The title compound was obtained in 95% yield as colorless oil. H NMR (400 MHz,
CDCl ), δ (ppm) 1.02 (t, J = 7.4 Hz, 3H), 1.77−1.86 (m, 2H), 3.03 (t, J = 7.8 Hz, 2H),
3
1
3
3
5
2
.91 (s, 3H), 8.81 (s, 1H); C NMR (100 MHz, CDCl ), δ (ppm) 13.7, 24.4, 39.1,
3
+
2.6, 134.6, 147.2, 162.3, 179.7; HRMS (ESI): calcd for C H NO Se, [M+Na]
8
11
2
55.9853; found: 255.9835.
Methyl 2-pentylselenazoline-4-carboxylate (9i)
1
The title compound was obtained in 71% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 0.89 (t, J = 6.8 Hz, 3H), 1.31−1.35 (m, 4H), 1.59-1.65 (m, 2H), 2.61
3
1
3
(
t, J = 7.9 Hz, 2H), 3.60-3.67 (m, 2H), 3.80 (s, 3H), 4.98 (t, J =8.4 Hz, 1H); C NMR
(
100 MHz, CDCl ), δ (ppm) 14.1, 22.5, 28.0, 31.3, 31.3, 37.8, 52.8, 80.1, 171.4, 174.0;
3
+
HRMS (ESI): calcd for C H NO Se, [M+Na] 286.0322; found: 286.0294.
1
0
17
2
1
2

ACCEPTED MANUSCRIPT
Methyl 2-pentylselenazole-4-carboxylate (10i)
1
The title compound was obtained in 93% yield as colorless oil. H NMR (400 MHz,
CDCl ), δ (ppm) 0.88 (t, J = 6.8 Hz, 3H), 1.31−1.42 (m, 4H), 1.74-1.82 (m, 2H), 3.04
3
1
3
(
t, J = 7.8 Hz, 2H), 3.91 (s, 3H), 8.80 (s, 1H); C NMR (100 MHz, CDCl ), δ (ppm)
3
1
4.0, 22.5, 30.7, 31.3, 37.2, 52.6, 134.6, 147.2, 162.3, 179.9; HRMS (ESI): calcd for
+
+
C H NO Se, [M+Na] 284.0166; found: 284.0151, [M+K] 299.9905; found:
12
19
2
2
99.9899.
Methyl 2-heptylselenazoline-4-carboxylate (9j)
1
The title compound was obtained in 65% yield as pale yellow oil. H NMR (400 MHz,
CDCl ), δ (ppm) 0.87 (t, J = 9.0 Hz, 3H), 1.27−1.31 (m, 8H), 1.58-1.66 (m, 2H), 2.61
3
(
t, J = 7.6 Hz, 3H), 3.64 (d, J = 9.6 Hz, 2H), 3.80 (s, 3H), 4.98 (t, J = 12.4 Hz, 1H);
1
3
C NMR (100 MHz, CDCl ), δ (ppm) 14.2, 22.7, 28.3, 29.1, 29.1, 31.3, 31.8, 37.8,
3
+
5
2.8, 80.1, 171.4, 173.9; HRMS (ESI): calcd for C H NO Se, [M+H] 292.0816;
1
2
21
2
+
found: 292.0803, [M+Na] 314.0635; found: 314.0626.
Methyl 2-heptylselenazole-4-carboxylate (10j)
1
The title compound was obtained in 90% yield as colorless oil. H NMR (400 MHz,
CDCl ), δ (ppm) 0.86 (t, J = 6.4 Hz, 3H), 1.27−1.44 (m, 8H), 1.75-1.82 (m, 2H), 2.61
3
1
3
(
t, J = 7.6 Hz, 3H), 3.05 (t, J = 7.8 Hz, 2H), 3.92 (s, 3H), 8.81 (s, 1H); C NMR (100
MHz, CDCl ), δ (ppm) 14.2, 22.7, 29.1, 29.1, 31.0, 31.7, 37.3, 52.6, 134.6, 147.2,
3
+
1
62.3, 179.9; HRMS (ESI): calcd for C H NO Se, [M+Na] 312.0479; found:
1
2
19
2
+
3
12.0482, [M+K] 328.0218; found: 328.0203.
(
S)-Methyl 2-(1-(tert-butyldiphenylsilyloxy)ethyl)selenazoline-4-carboxylate (9k)
1
The title compound was obtained in 77% yield (R/S = 55:45) as pale yellow oil. H
NMR (400 MHz, CDCl ), δ (ppm) 1.11(s, 4.05H,), 1.12 (s, 4.95H), 1.23(d, J = 6.4
3
Hz,1.35H), 1.28 (d, J = 6.4 Hz, 1.65H), 3.47(d, J = 9.6 Hz, 1.1H), 3.53(dd, J = 8.8 Hz,
J = 2.0 Hz, 0.9H), 3.78(s, 1.35H), 3.79(s, 1.65H), 4.67−4.75 (m, 1H), 4.91(t, J = 9.6
Hz, 0.45H), 5.(dt, J = 9.6 Hz, J = 0.8 Hz, 0.55H), 7.32−7.48 (m, 6H), 7.34-7.44 (m,
1
3
6
2
1
H), 7.65-7.0 (m, 4H); C NMR (100 MHz, CDCl ), δ (ppm) 19.4, 23.4, 23.5, 27.0,
3
8.6, 28.7, 52.8, 72.1, 72.2, 80.5, 80.7, 127.8, 127.8, 127.8, 130.0,130.0, 130.0, 132.8,
32.9, 133.7, 133.8, 135.9, 136.0, 136.1, 136.1, 171.3, 171.4, 181.5, 182.1; HRMS
+
(
ESI): calcd for C H NO SeSi, [M+Na] 498.0980; found: 498.0962.
2
3
29
3
(
S)-Methyl 2-(1-(tert-butyldiphenylsilyloxy)ethyl)selenzole-4-carboxylate (10k)
25
The title compound was obtained in 90% yield as colorless oil. [α]
-31.0 (c 0.85,
D
1
CHCl ); H NMR (400 MHz, CDCl ), δ (ppm) 1.13 (s, 9H), 1.35 (d, J = 6.4 Hz, 3H),
3
3
3
7
2
1
.91(s, 3H), 5.12(q, J = 6.4 Hz, 1H), 7.32−7.48 (m, 6H), 7.62 (d, J = 7.2 Hz, 2H),
1
3
.70 (d, J = 6.8 Hz, 2H), 8.87 (s, 1H); C NMR (100 MHz, CDCl ), δ (ppm) 19.4,
3
5.5, 27.1(3), 52.6, 72.8, 127.9(2), 128.0(2), 130.2, 130.3, 132.5, 133.5, 134.5,
35.9(2), 136.0(2), 147.9, 162.5, 186.9; HRMS (ESI): calcd for C H NO SeSi,
2
3
27
3
+
+
[
M+Na] 496.0823; found: 496.0839, [M+K] 512.0562; found: 512.0577.
1
3

ACCEPTED MANUSCRIPT
Methyl
-((R)-1-((tert-butoxycarbonyl)amino)-2-phenylethyl)selenazoline-4-carboxylate
9l)
2
(
1
3
The title compound was obtained in 62% yield as a pale yellow oil (R/S = 32:68). C
NMR (100 MHz, CDCl ), δ (ppm) 28.4, 30.9, 31.2, 39.6, 39.7, 52.9, 56.5, 80.1, 80.2,
3
8
1
4
0.4, 127.0, 127.1, 128.5, 128.6, 129.7, 129.8, 136.1, 136.3, 155.0, 155.1, 170.9,
+
71.1, 175.3, 176.3; HRMS (ESI): calcd for C H N O Se, [M+H] 413.0980; found:
1
8
24
2
4
13.0973.
Methyl
-((R)-1-((tert-butoxycarbonyl)amino)-2-phenylethyl)selenazole-4-carboxylate
10l)
2
(
2
5
The title compound was obtained in 91% yield as pale yellow oil. [α] D 5.8 (c 1.0,
1
CHCl ); H NMR (400 MHz, CDCl ), δ (ppm) 1.38 (s, 9H), 3.29 (bs, 1H), 3.38-3.42
3
3
(
m, 1H), 3.95 (s, 3H), 5.25 (bs, 2H), 7.14-7.16 (m, 2H), 7.21-7.30 (m, 3H), 8.83 (s,
1
3
1
H); C NMR (100 MHz, CDCl ), δ (ppm) 128.4, 41.2, 52.6, 56.6, 80.6, 127.2, 128.8,
3
1
29.6, 135.2, 136.3, 148.0, 155.2, 162.3, 181.2; HRMS (ESI): calcd for
+
C H N O Se, [M+Na] 433.0642; found: 433.0654.
18
22
2
4
Methyl 2-(2,3,5-tri-O-benzyl-β-D-ribofuranosyl)selenazoline-4-carboxylate (14)
To a solution of acid 13 (434 mg, 1 mmol) in anhydrous acetonitrile (10 mL) were
added Et N (202 mg, 2 mmol) and pentafluorophenyl diphenylphosphate (FDP, 416
3
mg, 1 mmol) at room temperature. After stirring for 5 min, β-azido diselenide 6 (104
mg, 0.25 mmol) and Ph P (524 mg, 2 mmol) was added into the solution. The reaction
3
mixture was stirred at 70 ℃ for another 5h, and the solvent was removed under
diminished pressure. The residue was purified by FC (silica gel, hexanes/EtOAc 7:1)
1
3
to give compound 14 (70%, R/S = 58:42,) as yellow oil. C NMR (100 MHz, CDCl ),
3
δ (ppm) 29.3, 29.4, 52.7, 52.7, 70.1, 70.2, 72.1, 72.1, 72.3, 72.4, 77.6, 78.0, 78.0, 80.6,
8
1
0.7, 80.8, 80.9, 81.6, 81.9, 83.6, 83.6, 127.6, 127.7, 127.9, 127.9, 127.9, 128.0, 128.3,
28.4, 128.4, 137.6, 137.7, 137.9, 138.3, 138.3, 171.1, 171.1, 176.2, 176.6; HRMS
+
(ESI): calcd for C H NO Se, [M+H] 596.1551; found: 596.1543.
3
1
33
6
Methyl 2-(2,3,5-tri-O-benzyl-β-D-ribofuranosyl)selenazole-4-carboxylate (15)
To a solution of 14 (100 mg, 0.17 mmol) in 1, 2-dichloroethane (5 mL) was added
activated MnO (870 mg, 1 mmol). The mixture was refluxed for 8 h under a nitrogen
2
atmosphere, then filtered and the filtrate was evaporated in vacuo. The residue was
chromatographed on silica gel (EtOAc/hexane 2:1) to give compound 15 (85 mg, 85%)
2
5
1
as colorless oil. [α]
105.8 (c 0.5, CHCl ); H NMR (400 MHz, CDCl ), δ (ppm)
D
3
3
,
,
,
,
,
,
,
,
,
3
1
.67 (dd, J4
= 3.8 Hz, J
= 4.8 Hz, 1H, H ), 3.85 (dd, J
= 2.4 Hz, J
=
, 5a
4 , 5a
5a
4 , 5b
5a , 5b
,
,
,
,
,
0.8 Hz, 1H, H ), 4.00 (s, 3H, OCH ), 4.04 (dd, J
= 7.6 Hz, J
= 4.8 Hz, 1H,
5
b
3
3 , 2
3 , 4
1
4

ACCEPTED MANUSCRIPT
,
,
,
,
,
,
,
H ), 4.04 (dd, J
= 3.0 Hz, J
= 4.8 Hz, 1H, H ), 4.41-4.45 (m, 2H, CH , H ),
3
3 , 2
3 , 4 3 2 4
4
.68-4.56 (m, 4H, CH ), 4.79 (d, J = 11.6 Hz, 1H, CH ), 4.89 (d, J = 12.0 Hz, 1H,
2 2
,
,
,
CH ), 5.40 (d, J
= 2.8 Hz, 1H, H ), 7.28-7.39 (m, 13H, Ph), 7.47-7.48 (m, 2H, Ph),
1
2
1 , 2
13
,
8
7
1
6
.88 (s, 1H, H ); C NMR (100 MHz, CDCl ), δ (ppm) 52.5, 69.3, 72.2, 72.4, 73.5,
1
3
7.3, 81.3, 81.7, 84.5, 127.8, 127.9, 128.0, 128.5, 128.5, 135.5, 137.6, 137.8, 138.2,
+
62.4, 180.7; HRMS (ESI): calcd for C H NO Se, [M+K] 632.0954; found:
3
1
31
6
32.0981.
Methyl 2-β-D-Ribofuranosylselenazole-4-carboxylate (16)
To a solution of compound 15 (50 mg, 0.084mmol) in CHCl (1 mL) was added
3
methanesulfonic acid (0.06 mL, 0.963 mmol). The reaction mixture was stirred at
room temperature for 8 h, then neutralized with NaOH (0.13 mL aqueous solution,
0
.963 mmol) and concentrated. The solid was washed with a co-solvent
(
CHCl /MeOH, 3 mL, v/v 2:1) and filtered through Celite, and the residue was
3
purified by FC (silica gel, methanol/EtOAc 1:10) to give compound 16 (24 mg, 90%)
2
5
as colorless solid. m.p. 113-114 °C (lit. m.p. 113–115 °C); [α] D -10.0 (c 0.5,
1
CH OH); H NMR (400 MHz, CDCl ), δ (ppm) 3.71 (dd, J = 2.8 Hz, J = 12.0 Hz,
3
3
1
H), 3.82 (dd, J = 4.2 Hz, J = 12.0 Hz, 1H), 3.90 (s, 3H), 4.02−4.08 (m, 2H), 3.47 (d,
13
J = 4.8 Hz, 1H), 4.98 (d, J = 4.4 Hz, 1H), 9.05 (s, 1H); C NMR (100 MHz, CDCl ),
3
δ (ppm) 52.9, 63.4, 72.9, 78.6, 86.4, 86.4, 136.8, 148.8, 163.6, 183.1; HRMS (ESI):
+
calcd for C H NO Se, [M+Na] 345.9806; found: 345.9796.
1
0
13
6
2
-β-D-Ribofuranosylselenazole-4-carboxamide（Selenazofurin 1）
Compound 16 (20 mg) was treated with an ammonia saturated methanol (10 mL) at 0
ºC for 5 h. The mixture was evaporated off in vacuo, and the residue was washed with
chloroform (2 mL x 3), then purified by FC (silica gel, methanol/EtOAc 1:5) to give
selenazofurin 1 (19 mg, quantitative yield) as colorless solid. m.p. 133-134 ºC (lit.
2
5
1
m.p. 131–133 ºC); [α] D -4.6 (c 0.4, MeOH); H NMR (400 MHz, MeOD), δ (ppm)
,
,
,
,
5a
,
,
,
3
.71 (dd, J4 = 4.4 Hz, J
= 12 Hz, 1H, H ), 3.79 (dd, J
= 2.8 Hz, J
= 12
,5a
5a ,5b
4,5b
5a ,5b
,
,
,
,
Hz, 1H, H ), 4.16 (t, J = 4.8 Hz, 1H, H ), 4.09-4.05 (m, 2H, H , H ), 4.97 (d, J =
5
b
2
3
4
,
13
5
.2 Hz, 1H, H ), 8.86 (s, H , 1H); C NMR (100 MHz, MeOD), δ (ppm) 63.6, 73.1,
1 5
7
8.5, 86.1, 86.7, 132.4, 152.0, 166.3, 181.7; HRMS (ESI): calcd for C H N O Se,
9
12
2
5
+
[
M+Na] 330.9809; found: 330.9824.
.3 Cells and cytotoxicity assays
4
Three human cell lines, A549, K562 and HCT-116 were cultured in RPMI 1640
medium (GIBCO-Invitrogen, NY) with 10% fetal bovine serum (FBS) and
−
1
−1
supplemented with glutamine (2 mmol L ), penicillin G (100 µg mL ), and
−
1
streptomycin (100 µg mL ) at 37 °C under 5% CO . MTT assay was carried out on
2
K562 (chronic erythroleukaemia), A549 and HCT 116 cells to test the cytotoxicity of
1
5

ACCEPTED MANUSCRIPT
Selenazofurin. The cells under study were harvested, counted and seeded into a 96
well plate at a density of 3000–5000 cells per well depending on their doubling times.
The cells were allowed to adhere for 24 h at 37 °C and 5% CO . After 24 h incubation,
2
the culture media was removed, 100 µL of test chemical dissolved in dimethyl
sulphoxide (DMSO) and culture medium was added to the cells, culture medium
containing DMSO only (without test compounds) was used for the control cells. Then,
the cells were incubated with compounds for 48 h under the same conditions. After 48
h, the culture medium containing compounds was removed and 100 µL of culture
−
1
media containing 5 mg mL MTT (Thiazoyl blue tetrazolium bromide) was added to
each well and cells were further incubated for 4 hours. After 4 hours incubation the
media containing MTT was aspirated and 100 µL DMSO was added to each well to
solubilize the crystallized formazan product. The plates were read on a plate reader at
5
70 nm and a reference wavelength of 630 nm. The absorbance readings for 630 nm
were subtracted from the 570 nm readings and the results were adjusted by dividing
the average by the DMSO control to adjust for any toxicity that may have occurred in
this control treatment set. The percentage inhibition was calculated based on
1
00−[(mean OD of treated cell × 100)/mean OD of vehicle treated cells (DMSO)].
The IC values were calculated using Probit Software. All the tests were repeated in
5
0
at least three independent experiments.
Acknowledgments
This work was supported in part by the Strategic Priority Research Program of CAS
(XDB14040201), the MOST (project 2015CB931903), and NNSFC (projects
2
1672255 and 21621064). We thank Xuefeng Sun for high resolution mass
determination.
Supporting Information
Supplementary data related to this article can be found at
References
1
2
3
. M. P. Rayman, P. Nutr. Soc. 2007;64: 527-542.
. J. Lu and A. Holmgren, J. Biol. Chem. 2009;284: 723-727.
. V. Zaharia, A. Ignat, B. Ngameni, V. Kuete, M. L. Moungang, C. N. Fokunang, M.
Vasilescu, N. Palibroda, C. Cristea, L. Silaghi-Dumitrescu and B. T. Ngadjui, Med.
Chem. Res. 2013;22: 5670-5679.
4
. E. Dominguez-Alvarez, M. Gajdacs, G. Spengler, J. A. Palop, M. A. Marc, K.
Kiec-Kononowicz, L. Amaral, J. Molnar, C. Jacob, J. Handzlik and C. Sanmartin,
Bioorg. Med. Chem. Lett. 2016;26: 2821-2824.
5
. G. Mugesh, W.-W. du Mont and H. Sies, Chem. Rev. 2001;101: 2125-2180.
1
6

ACCEPTED MANUSCRIPT
6
. V. Facchinetti, A. C. S. Nery, M. M. Avellar, C. R. B. Gomes, M. V. N. de Souza
and T. R. A. Vasconcelos, Curr. Org. Synth. 2015;12: 140-149.
7
8
9
. N. Rajesh, Mini-Rev. Med. Chem. 2008;8: 657-668.
. M. Kogami and M. Koketsu, Org. Biomol. Chem. 2015;13: 9405-9417.
. L. Wang, Z. Yang, J. Fu, H. Yin, K. Xiong, Q. Tan, H. Jin, J. Li, T. Wang, W. Tang,
J. Yin, G. Cai, M. Liu, S. Kehr, K. Becker and H. Zeng, Free Radical Biol. Med.
2
012;52: 898-908.
1
0. M. Soriano-Garcia, Curr. Med. Chem. 2004;11: 1657-1669.
11. E. Cordeau, S. Cantel, D. Gagne, A. Lebrun, J. Martinez, G. Subra and C. Enjalbal,
Org. Biomol. Chem. 2016;14: 8101-8108.
1
2. M. Koketsu and H. Ishihara, Curr. Org. Chem. 2003;7: 175-185.
3. H. C. Zhao, Y. P. Shi, Y. M. Liu, C. W. Li, L. N. Xuan, P. Wang, K. Zhang and B.
Q. Chen, Bioorg. Med. Chem. Lett. 2013;23: 6577-6579.
1
1
4. Q. Guan, Z. Cheng, X. Ma, L. Wang, D. Feng, Y. Cui, K. Bao, L. Wu and W.
Zhang, Eur. J. Med. Chem. 2014;85: 508-516.
1
5. C. Pizzo and S. G. Mahler, J. Org. Chem. 2014;79: 1856-1860.
6. Y. Liu, X. Sun, X. Zhang, J. Liu and Y. Du, Org. Biomol. Chem. 2014;12:
1
8
453-8461.
7. P. Loos, C. Ronco, M. Riedrich and H.-D. Arndt, Eur. J. Org. Chem. 2013;
290-3315.
8. Y. Wang, Y. Liang, L. Jiao, D.-M. Du and J. Xu, J. Org. Chem. 2006;71:
983-6990.
9. T. Murai, K. Yamaguchi, F. Hori and T. Maruyama, J. Org. Chem. 2014;79:
930-4939.
1
1
1
2
2
2
3
6
4
0. F. Purseigle, D. Dubreuil, A. Marchand, J. P. Pradère, M. Goli and L. Toupet,
Tetrahedron 1998;54: 2545-2562.
1. J.-B. Xavier, A. Fernando and A. Mercedes, Curr. Top. Med. Chem. 2014;14:
1
244-1256.
2. Y. Liu, P. He, Y. Zhang, X. Zhang, J. Liu and Y. Du, J. Org. Chem. 2018;83:
897-3905.
3
23. P. C. Srivastava and R. K. Robins, J. Med. Chem. 1983;26: 445-448.
24. J. A. Yalowitz and H. N. Jayaram, Anticancer Res., 2000;20: 2329-2338.
25. P. Dan Cook and D. J. McNamara, J. Heterocycl. Chem. 1986;23: 155-160.
26. H. Togo, S. Ishigami, M. Fujii, T. Ikuma and M. Yokoyama, J. Chem. Soc., Perkin
Trans. 1, 1994; 2931-2942.
2
7. V. R. Choudhary and M. G. Sane, J. Chem. Technol. Biotechnol. 1998;73:
3
36-340.
2
8. W. J. Hennen, B. C. Hinshaw, T. A. Riley, S. G. Wood and R. K. Robins, J. Org.
Chem. 1985;50: 1741-1746.
17

ACCEPTED MANUSCRIPT
2
3
9. T. J. Boritzki, D. A. Berry, J. A. Besserer, P. D. Cook, D. W. Fry, W. R. Leopold
and R. C. Jackson, Biochem. Pharmacol. 1985;34: 1109-1114.
0. W. C. Still, M. Kahn and A. Mitra, J. Org. Chem. 1978;43: 2923-2925.
1
8