Gram scale production of 1-azido-β-d-glucose: Via enzyme catalysis for the synthesis of 1,2,3-triazole-glucosides

Article abstract of DOI:10.1039/c9ra00736a

The production of analytical amounts of azido sugars is used as a means of verifying catalytic acid/base mutations of retaining glycosidase, but application of this process to preparative synthesis has not been reported. The catalytic acid/base mutant of Thermoanaerobacterium xylanolyticus GH116 β-glucosidase, TxGH116D593A, catalyzed the gram scale production of 1-azido-β-d-glucose (1) from p-nitropheyl-β-d-glucopyranoside (pNPGlc) and azide via a transglucosylation reaction. Overnight reaction of the enzyme with pNPGlc and NaN₃ in aqueous MES buffer (pH 5.5) at 55 °C produced 1 (3.27 g), which was isolated as a white foamy solid in 96% yield. This 1 was successfully utilized for the synthesis of fifteen 1,2,3-triazole-β-d-glucosyl derivatives (2-16) containing a variety of functional groups, via click chemistry.

Full text of DOI:10.1039/c9ra00736a

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Gram scale production of 1-azido-b-D-glucose via
enzyme catalysis for the synthesis of 1,2,3-triazole-
glucosides†
Cite this: RSC Adv., 2019, 9, 6211
Jaggaiah N. Gorantla,a Salila Pengthaisong,a Sunaree Choknud,a
Teadkait Kaewpuang,b Tanaporn Manyum,a Vinich Promarakb and James R. Ketudat
a
*
Cairns
The production of analytical amounts of azido sugars is used as a means of verifying catalytic acid/base
mutations of retaining glycosidase, but application of this process to preparative synthesis has not been
reported. The catalytic acid/base mutant of Thermoanaerobacterium xylanolyticus GH116 b-glucosidase,
TxGH116D593A, catalyzed the gram scale production of 1-azido-b-D-glucose (1) from p-nitropheyl-b-D-
glucopyranoside (pNPGlc) and azide via a transglucosylation reaction. Overnight reaction of the enzyme
with pNPGlc and NaN3 in aqueous MES buffer (pH 5.5) at 55 ꢀC produced 1 (3.27 g), which was isolated
as a white foamy solid in 96% yield. This 1 was successfully utilized for the synthesis of fifteen 1,2,3-
triazole-b-D-glucosyl derivatives (2–16) containing a variety of functional groups, via click chemistry.
Received 28th January 2019
Accepted 14th February 2019
DOI: 10.1039/c9ra00736a
rsc.li/rsc-advances
early study with the acid/base mutant of Agrobacterium b-
glucosidase, the substrates 2,4-dinitrophenyl-b-D-glucopyrano-
Introduction
side (DNPGlc) and 4-nitrophenyl-b-D-glucopyranoside (pNPGlc)
served as donor substrates for the transglucosylation of various
nucleophiles, including formate, azide, acetate, cyanide and
benzoate.5 Similarly, the Thermoanaerobacterium xylanolyticum
TxGH116 b-glucosidase acid/base mutant, TxGH116D593A, was
previously produced and rescued by azide to verify that the D593
acts as a catalytic acid/base.6 Kinetic characterization of trans-
glucosylation reactions catalyzed by b-glucosidase catalytic
acid/base mutants to produce 1-azido-b-D-glucose (1) has also
been reported,5–7 but exploration of this reaction as a prepara-
tive method has not been reported.
Synthesis of 1-azido-b-D-glucose (1) has also been reported
from unprotected glucose using 1,3-dimethylimidazolinium
salts, base and NaN3 in deuterium oxide,8–10 and other
reagents.11,12 1-Azido-b-D-glucose (1) has been used in the
copper-catalyzed azide–alkyne cycloaddition (CuAAC) reaction
for the construction of triazole-glucosides.13–16 Triazole-
glucosides exhibit a wide range of glucosidase,17–19 glycogen
phosphorylase,20–22 and carbonic anhydrase23,24 inhibitory
activities (Fig. 1). The triazole-glucosides also exhibit binding
affinity towards various proteins,25–28 and were found to be
potential radiopharmaceuticals,29,30 pesticides,31 water soluble
prodrugs, and inhibitors against cancer associated carbonic
anhydrases,32,33 and to have antinociception34 activities (Fig. 1).
Triazole-glucosides and their various activities have been
reviewed.35,36
Enzymes are well known for their unique ability to catalyze
stereospecic reactions and produce stereoselective products.1–3
Retaining b-glycosidases are one group of enzymes displaying
this property via generation of products with the same b-
anomeric conguration as their substrates. The retaining b-
glycosidase mechanism involves the displacement of the agly-
cone leaving group from the anomeric carbon of the sugar by
the catalytic nucleophile with acid-assistance by the catalytic
acid/base, which protonates the leaving group.4,5 The a-glycosyl
enzyme intermediate, thus formed, is resolved by attack of
a water molecule or other nucleophile on the anomeric carbon
to displace the catalytic nucleophile residue and release a b-
sugar or b-glycoside, respectively. This second step is generally
facilitated by the deprotonation of the incoming nucleophile
(water) molecule by the catalytic acid/base residue.
Acid/base mutants of b-glycosidase enzymes, in which the
catalytic carboxylate is replaced with a nonionizable amino acid
sidechain, have been frequently reported to catalyze the
formation of b-glucosides without signicant hydrolysis.4,5 In an
aSchool of Chemistry, Institute of Science, & Center for Biomolecular Structure,
Function and Application, Suranaree University of Technology, Nakhon Ratchasima
30000, Thailand. E-mail: cairns@sut.ac.th; Fax: +66 44 224185; Tel: +66 44 224193
bDepartment of Materials Science and Engineering, School of Molecular Science and
Engineering, Vidyasirimedhi Institute of Science and Technology, Rayong 21210,
Thailand
† Electronic supplementary information (ESI) available: Supplementary material
contains Fig. S1–S6, including purication, 1H NMR spectra of 1–16, 13C NMR
spectra of 1–16, and HRMS spectrum of compound 16. See DOI:
10.1039/c9ra00736a
In the present study, a catalytic process for the gram scale
production of the 1-azido-b-D-glucose (1) by an enzyme
(TxGH116D593A) in water was developed. The puried enzyme
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Fig. 1 Reported bioactive 1,2,3-triazole-b-D-glucosides.
catalyzed the production of gram scale amounts of 1 from and sodium azide, while the MES buffer was maintained at pH
pNPGlc and sodium azide in aqueous MES buffer (pH ¼ 5.5), in 5.5, based on previous pH optimization. Following these initial
ꢀ
an overnight reaction at 55 C. Aer the overnight incubation, experiments, the production was screened with 1 : 3, 1 : 6, 1 : 9,
thin layer chromatography (TLC) indicated the nearly complete 1 : 12 and 1 : 15 (w/w) ratios of enzyme : pNPGlc substrate,
conversion of pNPGlc substrate into 1, by the enzyme via 100 mM sodium azide and 50 mM MES buffer, pH 5.5, in 1 mL
transglucosylation with azide as the acceptor nucleophile. The water, and yellow color was observed aer the 24 h incubation at
product (1) could be conveniently puried and reacted with 55 ꢀC. TLC (Fig. 2) showed the formation of a new polar
various alkynes to form a variety of 1,2,3-triazole-b-D-glucosyl compound, with the Rf of 1-azido-b-D-glucose and near complete
derivatives (2–16), without the need for further deprotection of conversion of substrate, except at the 1 : 15 ratio, where the
the sugar.
substrate conversion was clearly not completed.
Again, we ran the same experiment as explained above with
1 : 13 and 1 : 14 (w/w) ratios of enzyme : substrate, the substrate
was almost completely converted into a product, as indicated by
TLC (Fig. 2). Taking advantage of this optimization of reaction
conditions, we used the 1 : 14 (w/w) ratio of enzyme : substrate
Results and discussion
Initially, we optimized the catalytic transglucosylation reaction
conditions by varying the concentration of enzyme, substrate,
Fig. 2 TLC profile of 1-azido-b-D-glucose (1) formation after 24 h from 3 mg/1 mg to 15 mg/1 mg and 5 g/370 mg of pNPGlc/enzyme
(TxGH116D593A). In each case, detection under UV light is shown on the left and carbohydrate staining with 10% sulphuric acid in ethanol and
charring is shown on the right. The numbers below the TLC lanes indicate ratios of pNPGlc to enzyme.
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Fig. 3 Structures of alkynes (1a–o) used in click chemistry.
Fig. 4 Structures of 1,2,3-triazole-b-D-glucosyl derivatives 2–16.
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for the large scale synthesis of 1-azido-b-D-glucose (1), from compound 1 represented 96% of the theoretical yield from the
pNPGlc and azide catalyzed by TxGH116D593A in a 370 mL 5 g of pNPGlc used in the reaction. This was similar to the yield
reaction volume (see ESI, Fig. S1†). Aer 24 hours of reaction at of 230 mg obtained from 330 mg of pNPGlc (98% yield) that we
55 ꢀC, the reaction mixture was lyophilized to dryness, redis- obtained in a preliminary small-scale reaction, suggesting
solved in methanol (MeOH) and puried by silica gel column reproducibly efficient conversion at milligram to gram scale.
chromatography. The pNP product eluted in 100% ethyl acetate The puried compound (1) was thus ready for the synthesis of b-
(EtOAc, 300 mL), and then the unreacted pNPGlc was isolated as D-glucosyl-1,2,3-triazole derivatives via a click chemistry.
a mixture with 1-azido-b-D-glucose (1) (2.645 g) (see ESI Fig. S1†),
Compound 1 was subsequently joined with different
with EtOAc/MeOH (95 : 5, v/v). The product 1 (800 mg) (see ESI compounds in which a propargyl group was attached to an
Fig. S1†), was isolated in pure form by elution with EtOAc/ alcohol, ethers, esters, amines, and amides (Fig. 3), as well as
MeOH (94 : 6, v/v). Finally, a trace of glucose byproduct (see phenyl acetylenes in a copper-catalyzed azide–alkyne cycload-
ESI Fig. S1†) was eluted with EtOAc/MeOH (1 : 1, v/v). The pure dition (CuAAC) reactions, giving rise to the 1,2,3-triazole-b-D-
1-azido-b-D-glucose (1) was dried under reduced pressure to give glucosyl derivatives 2–16 (Fig. 4, Table 1).
a white foamy solid, Rf ¼ 0.36 (CHCl3/MeOH/NH3, 7 : 2.8 : 0.2,
Initially, except for 1a, all the alkynes 1b–1k and 1n–o (Fig. 3)
1
v/v/v). Compound 1 was characterized with H, 13C NMR and were freshly prepared from their respective starting materials by
1
HRMS analysis and the H NMR spectrum resembles that re- EDC coupling, reductive amination and alkylation reactions.
ported in the literature (see ESI†).6 In compound 1, the The phenyl acetylenes (1l–m), were obtained from their
anomeric proton (H-1) shied to downeld and merged with respective aldehydes by the Seyferth–Gilbert homologation
water solvent residual peak at d 4.8 ppm in 600 MHz (see ESI reaction.37
page S4†), but the H-1 proton was observed at d 4.55 ppm (H-1,
d, J ¼ 9 Hz, 1H) in the 400 MHz spectrum (see ESI page S3†).
Pure 1-azido-b-D-glucose (1) was successfully coupled with
propargyl groups attached to hydroxyl, ether, amine, amide and
To increase the yield, the obtained mixture (2.645 g) was ester functional groups by copper-catalyzed azide–alkyne
redissolved in 6 mL of water/methanol (95 : 5, v/v) and sepa- cycloaddition (CuAAC) reactions at room temperature. In all the
rated over a Sephadex LH-20 column in the same solvent (ESI reactions, two equivalents each of CuI and DIPEA were used for
Fig. S2†). The product (1) eluted rst in pure form, followed by the coupling of 1 and alkynes 1a–k (Fig. 3) giving rise to most of
a minor amount of pNPGlc. Drying this portion of product the water soluble 1,2,3-triazole-b-D-glucosyl derivatives 2,20 3,38
yielded an additional 2.470 g of compound 1 as a foamy white 4, 5, 6,39 7, 8, 9, 10,38 11,38 12, in good to excellent yields (Fig. 4,
solid. Added with the 0.800 g from the rst step, the 3.270 g of Table 1). When the phenyl acetylenes were reacted with
Table 1 General reaction of the substrates used in click reactions
Entry
Alkyne
Cu salt (eq.)
Base (eq.)
Solvent system
Triazole-glucoside
Isolated yield
1
2
3
4
5
6
7
8
1a
1b
1c
1d
1e
1f
1g
1h
1i
1j
1k
1l
1m
1l
1m
1n
1o
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
CuI (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
DIPEA (2)
NaASc (0.2)
NaASc (0.2)
NaASc (0.4)
NaASc (0.4)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : CH3CN (1 : 2)
H2O : t-BuOH (1 : 5)
H2O : t-BuOH (1 : 5)
H2O : t-BuOH (1 : 1)
H2O : t-BuOH (1 : 2)
2
3
4
5
6
7
8
9
94%
60%
46.5%
43%
66%
51%
62%
78%
96%
80%
93%
—
9
10
11
12
10
11
12
13
14
15
16
17
No reaction
No reaction
13
14
15
16
CuI (2)
—
CuSO4$5H2O (0.1)
CuSO4$5H2O (0.1)
CuSO4$5H2O (0.2)
CuSO4$5H2O (0.2)
73%
75%
82%
79%
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compound 1 in the same solvent system, no triazole product analysis was performed on a Thermo Scientic Exactive Mass
was observed, prompting us to explore different click reaction Spectrometer with ions denoted in m/z.
conditions. Later, we found the phenyl acetylenes (1l–m)
successfully reacted with compound 1 when catalytic amounts
Process for the gram scale production of 1-azido-b-D-glucose
of CuSO4$5H2O (10%) and sodium ascorbate (20%) were used in
(1) from pNP-glucose using enzyme (TxGH116D593A) and
water and tert-butanol (1 : 5, v/v), leading to the formation of
purication of 1
triazole-glucoside derivatives 13 (ref. 40) and 14 (Fig. 4, Table 1).
The TxGH116D593A enzyme was produced and puried as
previously described,6 but without cleavage and removal of the
fusion tag. Compound 1 was produced in reactions with
In the above reaction conditions in water : tert-butanol (1 : 1
and 1 : 2, v/v), the compounds 15 and 16 (ref. 41) (Fig. 4, Table
1), were obtained by joining compound
1 with mono-
100 mM NaN3 (since higher concentrations did not produce
signicantly more product, data not shown), in 50 mM 4-mor-
pholineethanesulphonic acid (MES buffer), pH 5.5, and varying
concentrations of pNPGlc (Henan Tianfu Chemical Co. Ltd.,
Zhengzhou, China) substrate and TxGH116D593A enzyme. For
the large-scale reaction, 5 g pNPGlc (45 mM) was reacted with
370 mg TxGH115D593A (0.01 mM) as catalyst in a total volume
of 370 mL in a 1 L Duran bottle (see ESI Fig. S1†). Aer 24 hours
propargylated uorescein and di-propargylated hydroquinone
(1n and 1o), respectively.
It should be noted that compounds such as 4, 5 and 15
(Fig. 4), could be difficult to produce from click reactions with
tetra-acetyl-1-azido-b-D-glucose, since the internal ester and
lactone groups may be liable during deprotection reactions. For
instance, the deprotection of the glucose in generation of the
glucose ester of gibberellin 4 (GA4, the starting material for
compound 4) led to relatively low yield.7 In fact, for click
chemistry, the tetra-acetyl-1-azido-b-D-glucose could be depro-
tected before coupling and there are synthetic routes to 1-azido-
b-D-glucose directly.8–12 However, the enzymatic generation of
compound 1 reported here represents is a convenient method
for its generation. Compound 1 can then be coupled to such
labile groups without the need for deprotection.
ꢀ
of incubation in a water bath at 55 C, the yellow-colored reac-
tion mixture was frozen and subjected to lyophilization (ESI
Fig. S1†) for 48 h. The resulting yellow-colored dried mixture
was redissolved in methanol, the insoluble material was
removed by ltration through cotton and the soluble portion
was adsorbed on silica gel and loaded onto a silica gel column.
Purication of 1-azido-b-D-glucose (1) using silica gel column
chromatography
Conclusions
The silica gel with the adsorbed yellow-colored crude mixture
was loaded onto a silica gel (60–200 mesh) column (18.4 cm ꢁ 4
cm), in ethyl acetate (EtOAc, 100%). The column was washed
with 100% EtOAc (300 mL) to elute the p-nitrophenol, and then
with EtOAc/MeOH (95 : 5, v/v, 5 ꢁ 150 mL) to elute the
unreacted pNPGlc in a mixture with 1-azido-b-D-glucose (1).
Elution with EtOAc/MeOH (94 : 6, v/v, 5 ꢁ 150 mL) gave pure
product (see ESI Fig. S1†). Finally, the column was eluted with
EtOAc/MeOH (1 : 1, v/v) to elute the trace of glucose (see ESI
Fig. S1†). The compound was monitored by charring the TLC
plate aer spraying with 10% (v/v) sulfuric acid in ethanol. The
eluent fractions containing pure compound 1 and compound 1
mixed with pNPGlc were separately concentrated under reduced
pressure. The pure 1-azido-b-D-glucose (1) dried to a white
foamy solid (800 mg), Rf ¼ 0.36 (CHCl3/MeOH/NH3, 7 : 2.8 : 0.2,
v/v/v). The mixture of pNPGlc and compound 1 dried to yield
white powder (2.645 g) (ESI Fig. S1†).
The enzyme TxGH116D593A was successfully produced in
a multi-milligram scale and subsequently utilized for the gram
scale synthesis of 1-azido-b-D-glucose (1) in aqueous medium.
The compound 1 was further conjugated with different alkynes
via a 1,2,3-triazole as a linker. The complete process proceeds in
a safe and convenient way for the generation of novel 1,2,3-tri-
azole-b-D-glucosyl derivatives (2–16) in a short route by avoiding
deprotection steps. In addition, compound 1 was produced
from pNPGlc in an inexpensive way, when compared with the
commercial price of the product. This is the foremost method
developed for the gram scale synthesis of compound 1 using an
enzyme (TxGH116D593A), and its utilization for the synthesis of
glucoconjugates. Compound 1 is widely used in different elds
of synthesis and 2–16 are under screening for their inhibitory
activities.
Methods and experimental section
Purication of mixture of 1-azido-b-D-glucose (1) and pNP-b-D-
glucoside by Sephadex LH-20 column chromatography
Silica gel 60 F254 aluminum TLC plates were used to monitor the
reactions with short-wavelength ultraviolet light and by char-
ring the TLC plate aer spraying with 10% sulfuric acid in The obtained mixture (2.645 g, ESI Fig. S1†) was redissolved in
ethanol to visualize the spots. Column chromatography was 6 mL of water : methanol (95 : 5, v/v) and loaded onto an 18 cm
performed on silica gel 60–200 mesh and Sephadex LH-20 resin. ꢁ 3.5 cm Sephadex LH-20 column (see ESI Fig. S2†). The column
1H NMR and spectra were recorded at 400 MHz and 600 MHz was eluted with 2 ꢁ 100 mL of water/methanol (95 : 5, v/v). The
and 13C NMR spectra at 100 MHz, 125 MHz and 150 MHz on product (1) eluted in pure form, aer which the minor amount
a Bruker Avance III 600 MHz instrument with a Cryo probe of pNPGlc eluted. The product purity in the selected fractions
Prodigy. Either deuterated water (D2O), or deuterated DMSO was shown by TLC (ESI Fig. S2†), and these fractions were
(CD6SO) was used as the solvent. Chemical shis are given in pooled into a round bottom ask, and freeze-dried, resulting in
parts per million and coupling constants in hertz. HR-ESI-MS 2.470 g of compound 1 as a white foamy solid (ESI Fig. S2†).
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1-Azido-b-D-glucose (1)
Compound 4
1H NMR (D2O, 400 MHz): d 4.55 (H-1, d, J ¼ 9 Hz, 1H), 3.73 (H- To a solution of 1 (20 mg, 0.097 mmol, 1.2 eq.) and compound
6a, dd, J ¼ 2, 12.4 Hz, 1H), 3.55 (H-6b, dd, J ¼ 5.6, 12.8, Hz, 1H), 1c (30 mg, 0.081 mmol, 1 eq.) in water/acetonitrile (1 : 2, v/v, 1.5
3.37–3.30 (H-4, H-5, m, 2H), 3.25 (H-2, appt, J ¼ 9.4 Hz, 1H), and mL), were added copper iodide (31 mg, 0.162 mmol, 2 eq.) and
1
3.072 (H-3, appt, J ¼ 8.5 Hz, 1H); H NMR (D2O, 600 MHz). In DIPEA (28 mL, 0.162 mmol, 2 eq.), and the resulting yellow
the 600 MHz NMR analysis, the anomeric proton peak merged colored reaction mixture was stirred at room temperature 1 h.
with the solvent peak. The chemical shis of the observed peaks TLC indicated the almost complete conversion of the starting
were d 3.89 (H-6a, d, J ¼ 12 Hz, 1H), 3.72 (H-6b, d, J ¼ 12 Hz, 1H), material into a polar compound. The reaction mixture was
3.51–3.48 (H-4, H-5, m, 2H), 3.40 (H-2, appt, J ¼ 9 Hz, 1H), and ltered through a celite pad, washed with methanol (15 mL),
3.24 (H-3, appt, J ¼ 9 Hz, 1H); 13C NMR (D2O, 150 MHz) d 90.1, adsorbed on silica gel, and puried by chromatography using
77.9, 75.7, 72.8, 69.2 and 60.6; HR-ESI-MS m/z [M + Na]+ ethyl acetate/methanol (90 : 10, v/v) as solvent. Compound 4
C6H11N3O5Na calc'd for 228.05964, found 228.05975.
(26 mg, 46.5% yield) was obtained as a pale brown solid. It had
a silica gel TLC Rf ¼ 0.30 (EtOAc/MeOH 9 : 1, v/v, as solvent); 1H
NMR (D2O, 600 MHz): d 8.12 (s, 1H), 5.71 (d, J ¼ 8.7 Hz, 1H), 4.99
(s, 1H), 4.85 (s, 1H), 4.52–4.44 (m, 3H), 3.96 (appt, J ¼ 8.5 Hz,
1H), 3.90 (d, J ¼ 12 Hz, 1H), 3.80–3.75 (m, 2H), 3.71–3.68 (m,
2H), 3.62–3.59 (m, 1H), 3.07 (d, J ¼ 10 Hz, 1H), 2.62–2.60 (m,
2H), 2.06–2.04 (m, 2H), 1.91–1.85 (m, 2H), 1.80–1.56 (m, 6H),
1.35–1.29 (m, 2H), and 1.03 (s, 3H); 13C NMR (D2O, 150 MHz):
d 181.9, 174.1, 158.3, 123.4, 106.5, 97.1, 87.4, 78.9, 75.9, 72.2,
69.6, 69.0, 60.5, 55.0, 53.0, 52.5, 51.3, 51.1, 43.6, 38.8, 36.6, 34.3,
30.8, 27.2, 26.5, 15.6, and 13.7; HR-ESI-MS m/z [M + Na]+ calc'd
for C28H38N4NaO9 597.25365, found 597.25415.
Compound (2)
To a solution of 1 [26 mg, 0.126 mmol, 1 equivalent (eq.)] in
water/acetonitrile (1 : 2, 2 mL) was added propargyl alcohol 1a
(22 mL, 0.38 mmol, 3 eq.), followed by copper iodide (48 mg,
0.252 mmol, 2 eq.) and DIPEA (44 mL, 0.252 mmol, 2 eq.), and
the resulting yellow color reaction mixture was stirred at room
temperature overnight. TLC indicated the complete conversion
of the starting material into a polar compound. The reaction
mixture was diluted with 5 mL methanol and then passed
through a celite pad by washing with 10 mL methanol; the
ltrate was concentrated under reduced pressure. The resulting
crude product was redissolved in 4 mL methanol and adsorbed
Compound 5
on silica gel and subjected to purication by silica gel chro- To a solution of 1 (25 mg, 0.121 mmol, 1 eq.) and propargyl-
matography in solvent ethyl acetate/methanol (80 : 20, v/v), benzoate 1d (35 mg, 0.304 mmol, 1.8 eq.) in water/acetonitrile
which gave rise to pure compound 2 (31 mg, 94% yield) as (1 : 2, v/v,
2 mL), were added copper iodide (46 mg,
a white solid. Rf ¼ 0.13 (EtOAc/MeOH, 9 : 1, v/v); 1H NMR (D2O, 0.243 mmol, 2 eq.) and DIPEA (42 mL, 0.243 mmol, 2 eq.), and
600 MHz) d 8.39 (s, 1H), 5.94 (d, J ¼ 8 Hz, 1H), 4.94 (s, 2H), 4.24– the resulting yellow color reaction mixture was stirred at room
4.20 (m, 1H), 4.09 (d, J ¼ 11.7 Hz, 1H), 3.97–3.90 (m, 3H), and temperature overnight. TLC indicated the formation of a polar
3.82–3.81 (m, 1H); 13C NMR (D2O, 150 MHz) d 147.2, 123.4, 87.5, compound. The reaction mixture was ltered through celite
78.9, 76.0, 72.3, 69.0, 60.5 and 54.7; HR-ESI-MS m/z [M + Na]+ pad, washed with methanol (20 mL), and adsorbed on silica gel.
calc'd for C9H15N3O6Na 284.08585, found 284.08640.
Purication by silica gel chromatography using ethyl acetate/
methanol (90 : 10, v/v) as solvent gave rise to compound 5
(19 mg, 43% yield) as a white solid. Silica gel TLC gave an Rf ¼
0.26 (EtOAc/MeOH, 9 : 1, v/v as solvent); 1H NMR: (D2O, 600
MHz) d 8.37 (s, 1H), 8.01 (d, J ¼ 6.6 Hz, 2H), 7.67 (s, 1H), 7.51 (d,
J ¼ 6.1 Hz, 2H), 5.78 (d, J ¼ 8.8 Hz, 1H), 5.51 (s, 2H), 4.01 (appt, J
¼ 8.6 Hz, 1H), 3.91 (d, J ¼ 12 Hz, 1H), 3.79–3.71 (m, 3H), 3.63
(appt, J ¼ 8.9 Hz, 1H); 13C NMR (D2O, 150 MHz) d 168.2, 134.0,
129.5, 129.0, 128.8, 125.0, 87.6, 78.9, 75.9, 72.3, 69.0, 60.5, and
57.9; HR-ESI-MS m/z [M + Na]+ calc'd for C16H19N3NaO7
388.11207 amu, found 388.11189.
Compound 3
To a solution of 1 (25 mg, 0.121 mmol, 1 eq.) and prop-
argylcinnamide (1b) (45 mg, 0.243 mmol, 2 eq.) in water/
acetonitrile (1 : 2, v/v, 2 mL), were added copper iodide
(46 mg, 0.243 mmol, 2 eq.) and DIPEA (42 mL, 0.243 mmol, 2
eq.), and the resulting yellow colored reaction mixture was
stirred at room temperature for 2.5 h. TLC indicated the
formation of a polar compound. The reaction mixture was
ltered through a celite pad, washed with methanol (15 mL),
adsorbed on silica gel and puried by column chromatography
using ethyl acetate/methanol (90 : 10, v/v), which afforded
Compound 6
compound 3 (28 mg, 60%) as a white solid. Rf ¼ 0.2 (EtOAc/ To a solution of 1 (18 mg, 0.087 mmol, 1 eq.) and propargyl-
1
MeOH, 8.5 : 1.5, v/v); H NMR (D2O, 600 MHz) d 8.14 (s, 1H), benzyl ether 1e (63 mg, 0.435 mmol, 5 eq.) in water/
7.60 (bs, 2H), 7.51 (d, J ¼ 15.7 Hz, 1H), 7.51 (bs, 3H), 6.61 (d, J ¼ acetonitrile (1 : 2, v/v, 2 mL), were added copper iodide
15.7 Hz, 1H), 5.72 (d, J ¼ 8.7 Hz, 1H), 4.59 (s, 2H), 3.97 (appt, J ¼ (33 mg, 0.174 mmol, 2 eq.) and DIPEA (30 mL, 0.174 mmol, 2
8.5 Hz, 1H), 3.88 (d, J ¼ 12 Hz, 1H), 3.76–3.68 (m, 3H), and 3.60 eq.), and the resulting yellow colored reaction mixture was
(appt, J ¼ 9 Hz, 1H); 13C NMR (D2O, 150 MHz) d 168.8, 141.6, stirred at room temperature overnight. TLC indicated formation
134.3, 130.3, 129.0, 128.0, 123.1, 119.8, 87.4, 78.9, 75.9, 72.3, of a polar compound, just below to the starting material. The
68.9, 60.4, and 34.6; HR-ESI-MS m/z [M + Na]+ calc'd for reaction mixture was ltered through a celite pad, washed with
C
18H22N4NaO6 413.14370, found 413.14374.
methanol (15 mL), adsorbed on silica gel and puried by silica
6216 | RSC Adv., 2019, 9, 6211–6220
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gel chromatography using ethyl acetate/methanol (90 : 10, v/v) and 27.1; HR-ESI-MS m/z [M + Na]+ calc'd for C25H34N6NaO8
as solvent to afford compound 6 (20 mg, 66% yield) as a pale 569.23358, found 569.23474.
yellow viscous solid. Silica gel TLC showed an Rf ¼ 0.25 (EtOAc/
MeOH, 9 : 1, v/v, as solvent); 1H NMR: (D2O, 600 MHz); d 8.24 (s, Compound 9
1H), 7.44–7.42 (m, 5H), 5.76 (d, J ¼ 8.9 Hz, 1H), 4.75 (s, 2H,
To a solution of 1 (25 mg, 0.121 mmol, 1 eq.) and p-nitro-benzyl-
merged with solvent peak), 4.65 (s, 2H), 4.01 (appt, J ¼ 8.9 Hz,
propargylamine 1h (32 mg, 0.18 mmol, 1.5 eq.) in water/
1H), 3.92 (d, J ¼ 12 Hz, 1H), 3.80–3.71 (m, 3H), and 3.64 (appt, J
acetonitrile (1 : 2, v/v, 2 mL), were added copper iodide (46 mg,
0.242 mmol, 2 eq.) and DIPEA (42 mL, 0.242 mmol, 2 eq.), and the
¼ 9 Hz, 1H); 13C NMR: (D2O, 150 MHz): d 136.9, 128.8, 128.7,
128.4, 124.5, 87.4, 78.9, 75.9, 72.3, 72.3, 69.0, 62.2, and 60.4; HR-
resulting yellow colored reaction mixture was stirred at room
ESI-MS m/z [M + Na]+ calc'd for C16H21N3NaO6 374.13281, found
temperature for 6 h. TLC indicated the completion of starting
material into a polar compound. The reaction mixture was
374.13373.
ltered through celite pad, washed with methanol (20 mL), and
Compound 7
adsorbed on silica gel. Purication by silica gel chromatography
using ethyl acetate/methanol (70 : 30, v/v) as solvent afforded
compound 9 (35 mg, 74% yield) as white solid. Silica gel TLC gave
an Rf ¼ 0.29 (EtOAc/MeOH, 7 : 3, v/v, as solvent); 1H NMR (D2O,
600 MHz): d 8.23 (d, J ¼ 6.9 Hz, 1H), 8.1 (s, 1H), 7.56 (d, J ¼ 6.7 Hz,
1H), 5.75 (d, J ¼ 8.5 Hz, 1H), 4.94 (s, 2H), 4.06–4.02 (m, 3H), 3.93
(d, J ¼ 12 Hz, 1H), 3.81–3.71 (m, 3H), and 3.64 (appt, J ¼ 9 Hz,
1H); 13C NMR (D2O, 150 MHz): d 147.2, 129.7, 123.9, 87.5, 79.0,
76.0, 72.3, 69.0, 60.5, 54.5, and 50.9; HR-ESI-MS m/z [M + Na]+
calc'd forC16H21N5NaO7 418.13387, found 418.13445.
To a solution of 1 (27 mg, 0.131 mmol, 1 eq.) and propargyl-
indole-3-acetamide 1f (45 mg, 0.210 mmol, 1.6 eq.) in water/
acetonitrile (1 : 2, v/v, 2 mL), were added copper iodide
(50 mg, 0.263 mmol, 2 eq.) and DIPEA (46 mL, 0.263 mmol, 2
eq.), and the resulting yellow colored mixture was stirred at
room temperature for 4 h. Silica gel TLC indicated the almost
complete conversion of the starting material into a polar
compound. The reaction mixture was ltered through celite
pad, washed with methanol (20 mL), and adsorbed on silica gel.
Purication by silica gel chromatography using ethyl acetate/
Compound 10
methanol (90 : 10, v/v) as solvent afforded compound
7
(27.3 mg, 51% yield) as a pale brown solid. Silica gel TLC
showed an Rf ¼ 0.30 (with EtOAc/MeOH, 9 : 1, v/v, as solvent);
1H NMR (D2O, 600 MHz):d 7.68 (s, 1H), 7.50 (appt, J ¼ 8.4 Hz,
2H), 7.28 (s, 1H), 7.23 (appt, J ¼ 7.2 Hz, 1H), 7.12 (appt, J ¼
7.2 Hz, 1H), 5.58 (d, J ¼ 9 Hz, 1H), 4.42 (s, 2H), 3.86 (d, J ¼
11.4 Hz, 1H), 3.82–3.79 (m, 4H), 3.66.3.62 (m, 2H), and 3.55
(appt, J ¼ 9.6 Hz, 1H); 13C NMR (D2O, 150 MHz): d 175.4, 145.3,
136.3, 126.5, 125.1, 122.4, 122.2, 119.5, 118.3, 112.0, 107.5, 87.3,
78.8, 75.8, 72.2, 68.9, 60.4, 34.6, and 32.5; HR-ESI-MS m/z [M +
Na]+ calc'd for C19H23N5NaO6 440.15460, found 440.15543.
To a solution of 1 (26 mg, 0.126 mmol, 1 eq.) and Boc-
propargylamine 1i (59 mg, 0.378 mmol, 3 eq.) in water/
acetonitrile (1 : 2, v/v, 2 mL) were added copper iodide
(48 mg, 0.253 mmol, 2 eq.) and DIPEA (44 mL, 0.253 mmol, 2
eq.), and the resulting yellow colored reaction mixture was
stirred at room temperature for 3 h. TLC indicated the complete
conversion of the starting material into a polar compound. The
reaction mixture was ltered through celite pad, washed with
methanol (15 mL), and adsorbed on silica gel. Purication by
silica gel chromatography using ethyl acetate/methanol (96 : 4,
v/v) as solvent afforded compound 10 (44 mg, 96% yield) as
a white solid. Silica gel TLC demonstrated an Rf ¼ 0.35 (with
EtOAc/MeOH, 8.5 : 1.5, v/v, as solvent); 1H NMR (D2O, 600
MHz): d 8.11 (s, 1H), 5.72 (d, J ¼ 8.9 Hz, 1H), 4.36 (s, 2H), 3.99
(appt, J ¼ 8.5 Hz, 1H), 3.91 (d, J ¼ 12 Hz, 1H), 3.79–3.70 (m, 3H),
3.62 (appt, J ¼ 9 Hz, 1H), and 1.47 (s, 9H); 13C NMR (D2O, 150
MHz): d 158.0, 122.9, 87.4, 78.9, 75.9, 72.3, 69.0, 60.4, and 27.6;
HR-ESI-MS m/z [M + Na]+ calc'd for C14H24N4NaO7 383.15427,
found 383.15500.
Compound 8
To a solution of 1 (22 mg, 0.107 mmol, 1 eq.) and propargyl-
tryphtophanamide 1g (44 mg, 0.128 mmol, 1.2 eq.) in water/
acetonitrile (1 : 2, v/v, 2 mL), were added copper iodide
(41 mg, 0.214 mmol, 2 eq.) and DIPEA (37 mL, 0.214 mmol, 2
eq.), and the resulting reaction mixture was stirred at room
temperature for 30 min. TLC indicated the complete conversion
of starting material into a polar compound. The reaction
mixture was ltered through celite pad, washed with methanol
(25 mL), and adsorbed on silica gel. Purication by chroma-
Compound 11
tography using ethyl acetate/methanol (92 : 8, v/v) as solvent To a solution of 1 (20 mg, 0.097 mmol, 1 eq.) and propargyl-
afforded compound 8 (35.5 mg, 62% yield) as a white solid. benzamide 1j (31 mg, 0.195 mmol, 2 eq.) in water/acetonitrile
Silica gel TLC gave an Rf ¼ 0.2 (EtOAc/MeOH, 8.5 : 1.5, v/v, as (1 : 2, v/v,
2 mL), were added copper iodide (37 mg,
solvent); 1H NMR (D2O, 600 MHz): d 7.60 (d, J ¼ 7.8 Hz, 1H), 7.50 0.195 mmol, 2 eq.) and DIPEA (34 mL, 0.195 mmol, 2 eq.), and
(s, 1H), 7.46 (d, J ¼ 8.4 Hz, 1H), 7.22 (appt, J ¼ 7.2 Hz, 1H), 7.13 the resulting yellow colored reaction mixture was stirred at
(appt, J ¼ 7.2 Hz, 1H), 7.05 (s, 1H), 5.64 (d, J ¼ 9 Hz, 1H), 4.40– room temperature for 40 minutes. TLC indicated the complete
4.18 (m, 3H), 3.97–3.93 (m, 2H), 3.78 (dd, J ¼ 5.4, 12.6 Hz, 1H), conversion of the starting material into a polar compound. The
3.74–3.73 (m, 2H), 3.69 (appt, J ¼ 9 Hz, 1H), 3.61 (appt, J ¼ 9 Hz, reaction mixture was ltered through celite pad, washed with
1H), 3.14 (s, 2H), 1.34 (s, 6H) and 1.22 (s, 3H); 13C NMR (D2O, methanol (15 mL), and adsorbed on silica gel. Purication by
150 MHz): d 174.4, 157.2, 136.0, 126.7, 124.4, 121.8, 119.2, 118.3, silica gel chromatography using ethyl acetate/methanol (90 : 10,
111.8, 108.7, 87.3, 78.8, 75.9, 72.1, 68.9, 60.4, 55.6, 34.1, 27.5, v/v) as solvent afforded compound 11 (28 mg, 80% yield) as
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a white solid. Silica gel TLC demonstrated an Rf ¼ 0.27 (with
EtOAc/MeOH, 8.5 : 1.5, v/v, as solvent); 1H NMR (D2O, 600
MHz): d 8.15 (s, 1H), 7.76 (d, J ¼ 7.2 Hz, 2H), 7.59 (appt, J ¼
7.6 Hz, 1H), 7.50 (appt, J ¼ 8.4 Hz, 2H), 5.71 (d, J ¼ 9.6 Hz, 1H),
4.68 (s, 2H), 3.96 (appt, J ¼ 9 Hz, 1H), 3.87 (d, J ¼ 11.4 Hz, 1H),
3.76–3.66 (m, 3H), and 3.59 (appt, J ¼ 9.6 Hz, 1H); 13C NMR
(D2O, 150 MHz): d 170.9, 145.1, 133.2, 132.3, 128.8, 127.1, 123.1,
87.4, 78.8, 75.9, 72.2, 68.9, 60.4, and 34.9; HR-ESI-MS m/z [M +
Na]+ calc'd for C16H20N4NaO6 387.12805, found 387.12903.
Compound 14
A solution of 1 (25 mg, 0.0121 mmol, 1 eq.) and 4-cyano-
phenylacetylene 1m (23 mg, 0.182 mmol, 1.5 eq.) in water/tert-
butanol (1 : 5, v/v, 3 mL) was heated in a water bath until the
reaction mixture became homogeneous. Then, CuSO4$5H2O
(3 mg, 0.0121 mmol, 0.1 eq.) and sodium ascorbate (5 mg,
0.0242 mmol, 0.2 eq.) were added and the resulting colorless
reaction mixture was stirred at room temperature overnight.
TLC indicated the complete conversion of the starting material
into a polar compound. The reaction mixture was ltered
through celite pad, washed with methanol (20 mL), and
adsorbed on silica gel. Purication by silica gel chromatography
using ethyl acetate/methanol (94 : 6, v/v) as solvent afforded
compound 14 (30 mg, 75% yield) as a white solid. Silica gel TLC
(ran twice), demonstrated an Rf ¼ 0.20 (with EtOAc/MeOH, 9 : 1,
Compound 12
To a solution of 1 (50 mg, 0.243 mmol, 1 eq.) and PMB-
propargylether 1k (129 mg, 0.731 mmol, 3 eq.) in water/
acetonitrile (1 : 2, v/v, 4 mL) were added copper iodide
(93 mg, 0.486 mmol, 2 eq.) and DIPEA (85 mL, 0.486 mmol, 2
eq.), and the resulting yellow colored reaction mixture was
stirred at room temperature overnight. TLC indicated the
complete conversion of the starting material into a polar spot.
The reaction mixture was ltered through a celite pad, washed
with methanol (25 mL), and adsorbed on silica gel. Purication
by silica gel chromatography using ethyl acetate/methanol
(90 : 10, v/v) as solvent afforded compound 12 (87 mg, 93%
yield) as a pale yellow solid. Silica gel TLC demonstrated an Rf ¼
0.27 (with EtOAc/MeOH, 8.5 : 10, v/v, as solvent); 1H NMR (D2O,
600 MHz): d 8.17 (s, 1H), 7.32 (d, J ¼ 8.4 Hz, 2H), 6.98 (d, J ¼
7.8 Hz, 2H), 5.71 (d, J ¼ 9.6 Hz, 1H), 4.68 (s, 2H), 4.54 (s, 2H),
3.96 (appt, J ¼ 9 Hz, 1H), 3.88 (d, J ¼ 12.4 Hz, 1H), 3.80 (s, 3H),
3.76–3.66 (m, 3H), and 3.59 (appt, J ¼ 9 Hz, 1H); 13C NMR (D2O,
150 MHz): d 158.9, 144.4, 130.4, 129.5, 124.4, 87.4, 78.9, 75.9,
72.2, 71.9, 68.9, 61.9, 60.4, and 55.4; HR-ESI-MS m/z [M + Na]+
calc'd for C17H23N3NaO7 is 404.14337, found 404.14513.
1
v/v, as solvent); H NMR (DMSO-d6, 600 MHz): d 9.05 (s, 1H),
8.09 (d, J ¼ 7.8 Hz, 2H), 7.94 (d, J ¼ 7.8 Hz, 2H), 5.61 (d, J ¼ 9 Hz,
1H), 5.45 (d, J ¼ 5.4 Hz, 1H), 5.33 (d, J ¼ 2.4 Hz, 1H), 5.17 (d, J ¼
4.8 Hz, 1H), 4.63 (d, J ¼ 5.4 Hz, 1H), 3.81–3.77 (m, 1H), 3.72 (dd,
J ¼ 5.4, 10.8 Hz, 1H), 3.52–3.42 (m, 3H), and 3.29–3.25 (m, 1H);
13C NMR (DMSO-d6, 150 MHz): d 145.3, 135.5, 133.5, 126.2,
122.7, 119.2, 110.7, 88.2, 80.4, 77.2, 72.7, 70.0, and 61.2; HR-ESI-
MS m/z [M + Na]+ calc'd for C15H16N4NaO5 355.10184, found
355.10254.
Compound 15
To a solution of 1 (20 mg, 0.097 mmol, 1 eq.) and mono-
propargylated uorescein 1n (20 mg, 0.048 mmol, 0.5 eq.) in
water/tert-butanol (1 : 1, v/v, 2 mL) were added CuSO4$5H2O
(3 mg, 0.0097 mmol, 0.2 eq.) and sodium ascorbate (4 mg,
0.019 mmol, 0.4 eq.), and the resulting yellow colored reaction
mixture was stirred at room temperature overnight. TLC indi-
cated the complete conversion of the starting material into
a uorescent polar compound. The reaction mixture was
ltered through celite pad, washed with methanol (20 mL) and
adsorbed on silica gel. Purication by silica gel chromatography
using ethyl acetate/methanol (94 : 6, v/v) as solvent afforded
compound 15 (23 mg, 82% yield) as a yellow solid. Silica gel TLC
demonstrated an Rf ¼ 0.22 (with EtOAc/MeOH, 8.5 : 1.5, v/v, as
solvent); 1H NMR (DMSO-d6, 600 MHz): d 10.15 (s, 1H), 8.47 (s,
1H), 8.01 (d, J ¼ 7.8 Hz, 1H), 7.80 (appt, J ¼ 7.2 Hz, 1H), 7.73
(appt, J ¼ 7.2 Hz, 1H), 7.28 (d, J ¼ 7.8 Hz, 1H), 7.12 (s, 1H), 6.79
(d, J ¼ 8.4 Hz, 1H), 6.72 (s, 1H), 6.66 (d, J ¼ 8.4 Hz, 1H), 6.58 (s,
2H), 5.56 (d, J ¼ 9.6 Hz, 1H), 5.39 (d, J ¼ 6 Hz, 1H), 5.26 (d, J ¼
4.2 Hz, 1H), 5.23 (s, 2H), 4.62–4.61 (m, 1H), 3.80–3.76 (m, 1H),
3.69 (dd, J ¼ 5.4, 9 Hz, 1H), 3.46–3.37 (m, 4H), and 3.26–3.23 (m,
1H); 13C NMR (DMSO-d6, 150 MHz): d 169.1, 160.2, 160.0, 152.9,
152.3, 152.2, 142.6, 136.1, 130.6, 129.5, 129.4, 126.5, 125.1,
124.5, 124.4, 113.3, 112.8, 111.8, 109.9, 102.7, 102.0, 88.0, 83.1,
80.4, 77.4, 72.5, 70.0, 61.8, and 61.2; HR-ESI-MS m/z [M + H]+
calc'd for C29H26N3O10 576.16182, found 576.16227.
Compound 13
A solution of 1 (25 mg, 0.0121 mmol, 1 eq.) and 4-nitro-
phenylacetylene 1l (27 mg, 0.182 mmol, 1.5 eq.) in water/tert-
butanol (1 : 5, v/v, 3.5 mL) was heated in a 50 ꢀC water bath until
the reaction mixture became homogeneous and then CuSO4-
$5H2O (3 mg, 0.0121 mmol, 0.1 eq.) and sodium ascorbate
(5 mg, 0.0242 mmol, 0.2 eq.), were added and the resulting
colorless reaction mixture was stirred at room temperature
overnight. TLC indicated the complete conversion of the start-
ing material into a polar compound. The reaction mixture was
ltered through celite pad, washed with methanol (15 mL), and
adsorbed on silica gel. Purication by silica gel chromatography
using ethyl acetate/methanol (94 : 6, v/v) as solvent afforded
compound 13 (31 mg, 73% yield) as a white solid. Silica gel TLC
(ran twice), demonstrated an Rf ¼ 0.20 (with EtOAc/MeOH, 9 : 1,
1
v/v, as solvent); H NMR (DMSO-d6, 600 MHz): d 9.13 (s, 1H),
8.34 (d, J ¼ 8.4 Hz, 2H), 8.17 (d, J ¼ 7.8 Hz, 2H), 5.62 (d, J ¼
9.6 Hz, 1H), 5.46 (d, J ¼ 6 Hz, 1H), 5.33 (d, J ¼ 4.8 Hz, 1H), 5.18
(d, J ¼ 6 Hz, 1H), 4.62 (appt, J ¼ 6 Hz, 1H), 3.82–3.78 (m, 1H),
3.73 (dd, J ¼ 5.4, 11.4 Hz, 1H), 3.53–3.43 (m, 3H), and 3.28–3.25
(m, 1H); 13C NMR (DMSO-d6, 150 MHz): d 147.2, 144.9, 137.4,
126.4, 124.9, 123.1, 88.3, 80.4, 77.2, 72.8, 72.0, and 61.2; HR-ESI-
Compound 16
MS m/z [M + Na]+ calc'd for C14H16N4NaO7 375.0916, found To
a solution of 1 (44 mg, 0.214 mmol, 2 eq.) and
+
375.0919.
dipropargylated-hydroquinone 1o (20 mg, 0.107 mmol, 1 eq.) in
6218 | RSC Adv., 2019, 9, 6211–6220
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water/tert-butanol (1 : 2, v/v, 2 mL), were added CuSO4$5H2O 10 S. Shoda, A. Kobayashi, M. Noguchi, T. Tanaka, H. Nagai and
(5.3 mg, 0.0214 mmol, 0.2 eq.) and sodium ascorbate (8.5 mg, T. Matsumoto, JP2009292790A, 2009.
0.0428 mmol, 0.4 eq.), and the resulting reaction mixture was 11 M.-L. Larabi, C. Frechou and G. Demailly, Tetrahedron Lett.,
stirred at room temperature for 48 h. TLC indicated the 1994, 35, 2175–2178.
formation of a polar compound. The reaction mixture was 12 A. E. Meslouti, D. Beaupere, G. Demailly and R. Uzan,
ltered through a celite pad, which was washed with methanol Tetrahedron Lett., 1994, 35, 3913–3916.
(40 mL), and the solution adsorbed on silica gel. Purication by 13 R. Beniazza, N. Bayo, F. Molton, C. Duboc, S. Massip,
silica gel chromatography using ethyl acetate/methanol (80 : 20,
N. McClenaghan, D. Lastecoueres and J.-M. Vincent,
v/v) as solvent afforded compound 16 (51 mg, 79% yield) as
Beilstein J. Org. Chem., 2015, 11, 1950–1959.
a white solid. Silica gel TLC demonstrated an Rf ¼ 0.28 (with 14 R. Beniazza, R. Lambert, L. Harmand, F. Molton, C. Duboc,
1
EtOAc/MeOH, 6 : 4, v/v, as solvent); H NMR (D2O, 600 MHz):
d 8.25 (s, 2H), 6.99 (s, 4H), 5.71 (d, J ¼ 9 Hz, 2H), 5.20 (s, 4H),
3.96 (appt, J ¼ 9 Hz, 2H), 3.87 (d, J ¼ 12 Hz, 2H), 3.76–3.66 (m,
S. Denisov, G. Jonusauskas, N. D. McClenaghan,
D. Lastecoueres and J. M. Vincent, Chem.–Eur. J., 2014, 20,
13181–13187.
6H), and 3.59 (appt, J ¼ 9 Hz, 2H); 13C NMR (D2O, 150 MHz): 15 A. Pathigoolla, R. P. Pola and K. M. Sureshan, Appl. Catal., A,
152.3, 143.7, 124.5, 116.9, 87.4, 79.9, 75.9, 72.2, 68.9, 62.0, 60.4,
2013, 453, 151–158.
54.4, and 42.5; HR-ESI-MS m/z [M
+
Na]+ calc'd for 16 (a) B. L. Wilkinson, L. F. Bornaghi, S.-A. Poulsen and
T. A. Houston, Tetrahedron, 2006, 62, 8115–8125; (b)
G. C. Feast, O. E. Hutt, X. Mulet, C. E. Conn,
C. J. Drummond and G. P. Savage, Chem.–Eur. J., 2014, 20,
2783–2792; (c) S. Muthana, H. Yu, H. Cao, J. Cheng and
C
24H32N6NaO12 619.21975, found 619.19861.
Conflicts of interest
The authors have led a patent application for the process
X. Chen, J. Org. Chem., 2009, 74, 2928–2936.
described in this paper (TH1801005294), but otherwise have no 17 S. Dedola, D. L. Hughes, S. A. Nepogodiev, M. Rejzek and
conicts of interest to declare.
R. A. Field, Carbohydr. Res., 2010, 345, 1123–1134.
18 N. Anand, N. Jaiswal, S. K. Pandey, A. K. Srivastava and
R. P. Tripathi, Carbohydr. Res., 2011, 346, 16–25.
19 L. L. Rossi and A. Basu, Bioorg. Med. Chem. Lett., 2005, 15,
3596–3599.
Acknowledgements
The authors are thankful to Dr Ravi Shanker L. and Dr Yanling
Hua for assistance with HRMS and NMR, respectively. Financial 20 E. D. Chrysina, E. Bokor, K.-M. Alexacou, M.-D. Charavgi,
support from Suranaree University of Technology (SUT) and the
Thailand Research Fund (TRF), Thailand, grant BRG5980015,
and the National Research University project grant from the
G. N. Oikonomakos, S. E. Zographos, D. D. Leonidas,
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