Synthesis of purine nucleoside—amino acid conjugates and their photophysical properties

Article abstract of DOI:10.1016/j.tet.2016.05.043

2,6-Bis-(1,2,3-triazol-1H-1-yl)purine nucleosides react at C(6) with the N- and S-termini of various amino acids by expelling the corresponding 1,2,3-triazole as a leaving group in nucleophilic aromatic substitution reaction. This gives rise to a novel type of conjugate between purine nucleosides and amino acids. The obtained amino acid—derived 6-amino-2-(1,2,3-triazol-1H-1-yl)-purine derivatives showed useful levels of fluorescence with quantum yields up to 38% and Stokes shifts up to 91?nm. Glutathione and a cysteine-containing nonapeptide selectively reacted with their S-termini and produced the expected C(6) conjugates in good to excellent yields.

Full text of DOI:10.1016/j.tet.2016.05.043

Accepted Manuscript
Synthesis of purine nucleoside—amino acid conjugates and their photophysical
properties
Dace Cīrule, Kristers Ozols, Oskars Platnieks, Ērika Bizdēna, Ilze Māliņa, Māris Turks
PII:
S0040-4020(16)30442-2
10.1016/j.tet.2016.05.043
TET 27770
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 29 January 2016
Revised Date: 8 May 2016
Accepted Date: 16 May 2016
Please cite this article as: Cīrule D, Ozols K, Platnieks O, Bizdēna E, Māliņa I, Turks M, Synthesis of
purine nucleoside—amino acid conjugates and their photophysical properties, Tetrahedron (2016), doi:
10.1016/j.tet.2016.05.043.
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Synthesis of purine nucleoside – amino acid
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Dace Cīrule, Kristers Ozols, Oskars Platnieks, Ērika Bizdēna, Ilze Māliņa, and Māris Turks
Faculty of Materials Science and Applied Chemistry, Riga Technical University, P. Valdena 3/7, Riga, LV-
1048, Latvia

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Tetrahedron
journal homepage: www.elsevier.com
Synthesis of purine nucleoside – amino acid conjugates and their photophysical
properties
Dace Cīrule, Kristers Ozols, Oskars Platnieks, Ērika Bizdēna, Ilze Māliņa, and Māris Turks*
Faculty of Materials Science and Applied Chemistry, Riga Technical University, P. Valdena 3/7, Riga, LV-1048, Latvia
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
2,6-Bis-(1,2,3-triazol-1H-1-yl)purine nucleosides react at C(6) with the N- and S-termini of
various amino acids by expelling the corresponding 1,2,3-triazole as a leaving group in
nucleophilic aromatic substitution reaction. This gives rise to a novel type of conjugate
between purine nucleosides and amino acids. The obtained amino acid – derived 6-amino-2-
(1,2,3-triazol-1H-1-yl)-purine derivatives showed useful levels of fluorescence with quantum
yields up to 38% and Stokes shifts up to 91 nm. Glutathione and a cysteine-containing
nonapeptide selectively reacted with their S-termini and produced the expected C(6)
conjugates in good to excellent yields.
Available online
Keywords:
Purine nucleosides
Amino acids
Triazolyl purine derivatives
Fluorescence
2016 Elsevier Ltd. All rights reserved
.
SNAr reactions
1. Introduction
purine nucleoside “double conjugates” with both 1,2,3-triazoles
and amino acids 3 (Figure 1).
Azolylpurine nucleoside derivatives 1 (Figure 1) exhibit a
broad range of notable biological activities.1 Purine nucleoside
analogs possessing
a 1,2,3-triazolyl substituent in the
heterocyclic base form a distinct class among the purine-azole
conjugates that became popular during the past decade.2 The
members of 1,2,3-triazole-purine nucleoside family are known
to act as adenosine receptor antagonists,3 with a high affinity to
adenosine A3 receptors,4 inhibitors of aryl acid adenylating
enzymes,5 and potential anti-HCV agents.6 Some 1,2,3-triazole
derivatives showed activity against cytomegalovirus.7,8
Additionally, triazolylpurine nucleosides show useful levels of
fluorescent properties1,9,10 that can lead to their use as sensors in
biological chemistry.11,12
On the other hand, introduction of amino acid moieties into
biologically active compounds can lead to optimal
pharmacokinetic and pharmacodynamic characteristics.13 It is
also known that purine – amino acid conjugates 2 (Figure 1)
exhibit useful levels of biological activity that among others
include antibacterial14 and antiviral15,16 properties. The amino
acid residue can be introduced in the purine ring at its positions
C(2), C(6) and C(8).
Figure 1. Azolylpurine nucleosides 1, purine – amino acid conjugates 2
and purine nucleoside conjugates with both 1,2,3-triazole and amino acid
moieties 3.
Generally, amino acid moiety can be attached to the purine
core either by cross-coupling chemistry17,18,19,20,21 or by
nucleophilic aromatic substitution reactions.22,23,24,25,26,27
Regarding the attachment of the amino acid moiety by its N-
terminus to C(6) in the purine cycle, 6-halopurine derivatives
are used almost exclusively. Thus, the El-Bayouki group
This prompted us to develop a synthetic method leading to
1* Corresponding author. Tel.: +371 67089251
e-mail address: maris_turks@ktf.rtu.lv (M. Turks)

2
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triazolyl derivative 7 was obtained. The reaction sequence 4→7
developed
a
synthesis of N-(purin-6-yl)-amino acid
gave 59% overall yield.
derivatives through SNAr reaction between 9-(p-anisyl)-6-
chloropurine and the sodium salts of natural α-amino acids.28
Another approach involves the use of 9-benzyl-6-chloro-9H-
purine and amino acid esters in the presence of triethylamine in
ethanol at 120 ºC. It was shown that modifications at purine
C(2) and N(9) led to decreased yields.29 Also N2-protected 2-
amino-6-chloro-9H-purine undergoes SNAr reaction with
selected amino acid esters.13 It should be noted that most of the
described examples discuss the reactions between purine as
heterocyclic base and amino acid derivatives. Nevertheless,
Sheng et al.,30 and Qu&Guo et al.29 in 2012 have described
syntheses of purine nucleoside – amino acid conjugates.
Following the aforementioned approaches, also in these latter
cases 6-chloropurine riboside was used as a starting material.31
At this point we can conclude that to the best of our knowledge
no other leaving groups than chlorine have been used in SNAr
reactions between substituted purines and amino acid N-
terminus.
With the bis-triazolyl derivative 7 in hand we started to
elaborate the reaction conditions for the envisaged nucleophilic
aromatic substitution with protecting group free amino acids. It
was found that the reactivity of the starting material changed
dramatically depending on the solvent and base. The
preliminary screening of three parameters (amino acid, solvent,
and base) indicated that proline was the best reacting amino
acid which converted to the purine nucleoside – amino acid
conjugate in a time scale applicable for screening conditions.
Thus, in a parallel set of experiments, starting material 7 and
proline were combined with various bases in various solvents
to obtain the information on optimal reaction conditions
(Scheme 2, Figures 2,3).
We have recently reported the synthesis of 2,6-bis-
(triazolyl)purine nucleosides and their regioselective reactions
with primary and secondary amines9 and thiols.32 The obtained
N6-substituted 2-triazolyl adenosine analogs possessed good
fluorescent properties.9 Thus, we were keen to develop further
purine nucleoside SNAr chemistry which uses 1,2,3-triazole
moieties as leaving groups and to study potential photophysical
properties of the obtained amino acid conjugates with purine
nucleosides. So far only one report superficially describes the
fluorescent properties of some selected N-(purin-6-yl)amino
acid derivatives.29
Scheme 2. Reaction of 2,6-bis-triazolyl derivative 7 with proline
according to Figure 2 and 3.
2. Results and discussion
The key intermediate 7 was obtained in three steps from 2,6-
dichloropurine (4) (Scheme 1).9 First, compound 4 was reacted
with sodium azide,33 then 5 was further converted to 6 using
Vorbrüggen glycosylation.34 In the following reaction with
phenylacetylene in t-BuOH/water, bis-
Figure 2. Screening of reaction conditions I for transformation 7 +
proline → 8a: bases and solvents. The data represent the level of
conversion to 8a by HPLC-DAD. Experimental conditions: nucleoside 7
(10 mg, 0.015 mmol) + L-proline (3.5 mg, 0.03 mmol) + selected base
(0.03 mmol) in selected solvent (1 mL) at 40 °C.
It was found that the combination of DMSO and NEt3 can
be regarded as optimal. The use of K2CO3 as the base also
resulted in good conversion to product 8a. The latter result is
valid only in the case when strictly anhydrous conditions are
ensured. Deviation from this results in partial deacetylation
followed by formation of unidentified byproducts.
Additionally, the desired product is not very stable in the
reaction mixtures containing K2CO3. Diisopropylethylamine
(DIPEA) showed comparable results to those obtained by NEt3,
but the latter was chosen due to economic reasons.
Scheme 1. Synthesis of purine nucleoside 2,6-bis-triazolyl derivative 7.

3
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(Figure 4). To our surprise, the extra amount of water (0.1 mL
in 0.7 mL of DMSO) which was used in the NMR experiment
to treat the solubility issue of glutathione increased the rate of
reaction strikingly. Hence, 90% conversion of 7 was observed
in 3 min. In the NMR experiments the 2-bromomesitylene was
used as an internal standard. This allows us to conclude that
reaction 7+10 → 11+12 proceeds without formation of any
significant side-products. It is interesting to note that similar
reaction rate acceleration by addition of water has been
reported in SNAr reactions between chloropyridine systems and
thiols.35,36 It was proposed that RSHꢀꢀꢀOH2 hydrogen bond
formation increases the nucleophilicity of the sulfur center. On
the other hand, H-bond formation between purine ring nitrogen
and water might stabilize the intermediate Meisenheimer
adduct. In an analogous preparative-scale experiment, the
Figure 3. Screening of reaction conditions II for transformation 7 +
proline → 8a: various solvents in combination with NEt3. The data
represent the level of conversion to 8a by HPLC-DAD.Experimental
conditions: nucleoside 7 (20 mg, 0.03 mmol) + L-proline (7 mg, 0.06
mmol) + NEt3 (6 mg, 0.06 mmol) in selected solvent (2 mL) at 40 °C.
The elaborated reaction conditions were applied in the
synthesis of purine nucleoside – amino acid conjugates. Thus,
L-proline provided quantitative conversion to the expected
product 8a in 2 hours at room temperature also on a preparative
scale, however, the isolated yield reached 60%. For other
reagent combinations the bis-triazolyl derivative 7 was treated
with various natural α-amino acids in DMSO in the presence of
triethylamine at 50 °C for 15-24 h to give sugar moiety
protected nucleosides 8b-h in 48-63% yields (Scheme 3, Table
1). Under these conditions only trace amounts of partially
deprotected products were observed by LC-MS. Due to the
poor solubility of deprotected nucleoside 9, O-acetylated
nucleosides 8a-h were used for UV/VIS and fluorescence
measurements. If deprotected nucleosides are necessary,
cleavage of the protecting groups could be achieved using
MeNH2/H2O (Scheme 3). In this way the glycine adduct 7 was
obtained in 65% yield over 2 steps.
Scheme 3. 2,6-Bis-triazolylpurine nucleoside substitution with natural
α-amino acids at C-6 (see Table 1).
To investigate the scope of the nucleophilic agents suitable
for SNAr reactions, ligation between 7 and the tripeptide
glutathione (10) was performed (Scheme 4). Reaction went
smoothly in DMSO solution with Et3N additive to give a sole
product 11 in 1-2 h. The clean and rapid conversion was
examined closer by carrying out the reaction in an NMR tube
Scheme 4. S-Chemoselective ligation of 2,6-bis-triazolylpurine
nucleoside 7 with glutathione (10) and nonapeptide 13.

4
Tetrahedron
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clean and rapid conversion of the starting materials was
observed. In the first 15 minutes about half of peptide 13 had
transformed into product 14, releasing triazole 12 as a leaving
group. Furthermore, practically full conversion of 13 was
observed in 50 minutes. Also in this case exclusive formation
of the S-adduct was observed as seen by its UV spectrum
(Figure 6). The observed S-selectivity can be explained by the
fact that thiols are better nucleophiles than amines and among
other transformations have been previously reported to react
selectively in Michael additions37 and acylation reactions.38 In a
related recent example S-selective SNAr reactions were reported
between fluorinated aryl derivatives and cysteine containing
oligopeptides.39
Absorption and fluorescence spectra of the novel amino acid
derivatives 8a-h are shown in Figures 7-9. All compounds have
been investigated in methanol solutions. UV/Vis and emission
spectra of the three compounds with the brightest fluorescence
(8a, 8e, 8f) as well as the randomly selected 8d were further
examined in DMSO, MeCN, THF and chloroform (Figure 8
and the Supplementary data). The experimental data are
1
Figure 4. A part of H NMR spectra (300 MHz) of a mixture of bis-
triazolyl purine 7 and glutathione (10) in DMSO-d6/D2O. a) combination
of nucleoside 7 and peptide 10 in the absence of base; b) the previous
mixture in 3 min after addition of NEt3 (2.5 equiv).
6.0
11
14
A
5.0
4.0
3.0
2.0
1.0
0.0
B
C
210
230
250
Wavelength, nm
270
Figure 6. Absorption spectra of compounds 11, 14 in comparison with
those of previously reported 6-alkylthio-2-triazolyl-purine nucleoside
derivatives A-C.32 The structures of compounds A-C are given in the
Supplementary data.
Figure 5. HPLC-DAD traces of the reaction mixture 7 + 13 → 14 + 12:
a) combination of nucleoside 7 and peptide 13 in the absence of base; b) the
previous mixture in 15 min after addition of NEt3 (5.0 equiv); c) the
reaction mixture in 50 min after initial addition of NEt3.
8a
3.5
8b
3.0
8c
2.5
2.0
1.5
1.0
0.5
0.0
8d
8e
8f
DMSO-H2O solvent system was used to afford conjugate 11 in
82% isolated yield. Again, no starting material was detected
after 15 min of stirring at room temperature. The
chemoselectivity leading to the S-product was proved by
combination of NMR and UV spectra. In particular the UV
spectra of substituted 6-thio-2-triazolyl-purines show a very
specific pattern characterized by two pronounced maxima inthe
wavelength regions of 230 and 255 nm (Figure 6). On the other
hand, the N6-adducts 8a-h and other 6-amino-2-triazolyl-purine
derivatives9 are characterized by UV spectra with a very
different pattern (Figure 7). Encouraged by these results we
tested the reactivity of bistriazolyl derivative 7 towards
nonapeptide 13. The reaction was carried out in a rather diluted
solution at room temperature (1.5 mg of peptide 13 in 1 mL of
DMSO + 1.3 equiv. of nucleoside 7) and its progress was
monitored by HPLC (Figure 5). As in case with glutathione, a
8g
8h
250
275
300
325
350
Wavelength, nm
Figure 7. UV spectra of compounds 8a-h (c = 1×10-4 M, MeOH).
collected in Table 1. The absorption spectra of the obtained N6-
substituted 2-triazolyl-adenosine derivatives 8a-h show two
maxima at ca. 250 and 305 nm with the first one being about

5
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Table 1. Photophysical properties of compounds 8a-h
Products 8a-h
λmax
(exc),
nm
λmax
(emis),
nm
Stokes
shift, nm
Quantum
yield, %
Entry
Yield
Solvent
1
2
3
4
5
MeOH
DMSO
MeCN
THF
310
310
310
310
310
395
385
387
379
386
85
75
77
69
76
11
17
34
25
34
8a, 60%
CHCl3
6
7
8b, 50%
8c, 48%
MeOH
MeOH
300
300
387
387
87
87
4
2
8
MeOH
DMSO
MeCN
THF
300
305
305
295
300
297
300
295
295
300
300
300
295
295
298
386
381
389
378
390
388
384
386
377
386
386
380
386
376
386
86
76
84
83
90
91
84
91
82
86
86
80
91
81
88
3
9
6
10
11
12
13
14
15
16
17
18
19
20
21
22
8d, 50%
8e, 60%
8f, 63%
32
21
38
9
CHCl3
MeOH
DMSO
MeCN
THF
20
20
13
21
11
20
18
7
CHCl3
MeOH
DMSO
MeCN
THF
CHCl3
17
23
24
8g, 59%
8h, 55%
MeOH
MeOH
300
297
392
386
92
89
3
4

6
Tetrahedron
ACCEPTED MANUSCRIPT
twice as intensive as the second one (Figure 7). Also a shoulder
is present at 275 nm. These spectra are virtually unaffected by
solvent (see Supplementary data). Next we proceeded to the
emission measurements. UV irradiation at 295- 310 nm was
used to induce the fluorescence. Such wavelength could allow
excitation in biological systems, e.g. in the presence of nucleic
acids and most amino acid side-chains.40,41 All purine
nucleoside – amino acid conjugates with the N6-attachment
(products 8a-h) exhibit relatively pronounced fluorescent
properties. On the other hand, the thio-adduct 11 revealed very
weak fluorescence for which the fluorescence quantum yield
(QY) was not determined. The fluorescence maxima for
compounds 8a-h are positioned in the range of 375-395 nm and
show a small dependence on solvent (Figure 8 and 9, see also
3. Conclusion
In conclusion, we have successfully extended the scope of
nucleophilic aromatic substitution of bis-triazolyl-purine
nucleosides with functionalized N- and S-nucleophiles. It was
found that the N-linked triazolylpurine nucleoside – amino acid
conjugates possess fluorescent properties with quantum yields
up to 38%, which is comparable or on some occasions slightly
lower than that of simple N-adducts.1,9 We have also shown
that peptides possessing both N- and S-nucleophilic sites
exhibit excellent reactivity and complete S-selectivity in
nucleophilic aromatic substitution on the given substrate. This
can be regarded as a novel and useful entry for the synthesis of
various bioconjugates. One can expect that similar
transformations are also possible in pyrimidine and variously
annulated pyrimidine systems. It should be noted that the UV
spectra of the corresponding N- or S-adducts possess a
characteristic pattern and as such can be used in structure
elucidation. Both evaluation of biological activity of the
obtained products and application studies of the developed
method in biological chemistry are underway in our laboratory
and will be reported elsewhere.
Supplementary data). Still, it is worth pointing out that a
significant amount of fluorescence is observed in the visible
range. On the other hand, the QY are strongly influenced by the
solvent. Among the solvents studied, the highest QY usually
were observed in chloroform and acetonitrile solutions, while
in most cases the lowest value was attributed to samples
dissolved in methanol. For instance, QY experiences a 12-fold
increase upon solvent change from methanol to chloroform in
the case of 8d (Table 1).
4. Experimental Section
4.1. General
1.4
8a
8b
8c
8d
8e
8f
1.2
1.0
0.8
0.6
0.4
0.2
0.0
All non-aqueous reactions were carried out under an inert argon
atmosphere in oven-dried glassware (unless otherwise noted).
Commercial reagents were used without purification. Solvents
for synthetic preparations were distilled prior to use, and if
required, dried over standard drying agents (THF from metallic
sodium, MeOH from metallic magnesium and iodine, DMSO,
DMF, and Et3N from CaH2). Solvents of HPLC grade were
used for UV and fluorescence measurements. Column
chromatography was performed on silica gel (60 Ǻ, 40‒63 ꢁm,
Merck). Analytical thin layer chromatography (TLC) was
performed on E. Merck Kieselgel 60 F254. High-performance
liquid chromatography (HPLC) analyses were performed using
Agilent Technologies 1200 Series chromatograph. HPLC
column: X Bridge C18 4.6 × 150 mm (3.5 µm); eluent system:
A (0.01 M aqueous KH2PO4 / MeCN (94/6, V/V)), B (MeCN);
gradient regime: 0‒7‒10‒12 min, B eluent change 10‒95‒95‒
10%. IR spectra were recorded in KBr with FT-IR Perkin
8g
8h
11
320
370
420
470
520
570
Wavelength, nm
Figure 8. Fluorescence spectra of compounds 8a-h and 11 (c =
1×10-4 M, MeOH).
4.0
3.5
Elmer Spectrum BX (4000‒450 cm-1). H-NMR and 13C-NMR
1
THF
DMSO
MeCN
Chloroform
MeOH
3.0
2.5
2.0
1.5
1.0
spectra were recorded at 300 MHz and at 75.5 MHz,
respectively. The chemical shifts (δ) are given in parts per
million relative to the residual signals of non-deuterated solvent
1
DMSO-d6 for H-NMR (2.50 ppm) and 13C-NMR (39.5 ppm).
Multiplicity is indicated by: s (singlet), d (doublet), t (triplet),
m (multiplet), br. s (broad signal). The coupling constants (J)
are reported in Hz. HRMS (ESI) analysis were performed on
Agilent 1290 Infinity series (Agilent Technologies, Germany)
UPLC chromatographic system connected with Agilent 6230
TOF LC/MS (Agilent Technologies, Germany) mass selective
detector (ESI+). UPLC column: Extend C18 RRHD 2.1 × 50
mm (1,8 µm); eluent system: A (0.1% HCOOH in MeCN),
B (0.1% HCOOH in H2O). The UV-Vis absorption spectra
0.5
0.0
320
370
420
470
520
570
Wavelength, nm
Figure 9. Solvent effects on absorption and fluorescence spectra of
compound 8a (c = 1×10-4 M).

7
ACCEPTED MANUSCRIPT
were acquired using Perkin-Elmer 35 UV/Vis spectrometer
with a 1 cm path length quartz cell. Emission spectra and
absolute photoluminescence quantum yields were measured by
the Quanta Master 40 steady state spectrofluorometer using 1
cm and 0.5 cm path length quartz cuvettes, correspondingly. In
the quantum yield measurements, 6 inch integrating sphere by
LabSphere was employed.
kept at 0‒5 ºC for 24 h. The resulting precipitate was filtered,
washed on the filter with cold water (30 mL) and dried under
reduced pressure. The crude product was purified by silica gel
column chromatography (MeOH in CH2Cl2; gradient 10%→
20%) to give compound 8b (0.15 g, 50%) as a white foam. Rf =
0.25 (EtOAc/MeOH 2:1); IR (KBr) ν (cm-1): 3394, 1752, 1625,
1
o
1491, 1459, 1420, 1375, 1231; H-NMR (300 MHz, 70 C,
DMSO-d6) δ (ppm): 9.14 (s, 1H, H-C(triazole)), 8.34 (s, 1H,
H-C(8)), 8.01 (d, 2H, 3J = 7.6 Hz, Ar), 7.85 (br. s, 1H, (-NH-)),
7.49 (t, 2H, 3J = 7.6 Hz, Ar), 7.39 (t, 1H, 3J = 7.6 Hz, Ar), 6.28
4.2. N-(9-(2’,3’,5’-Tri-O-acetyl-β-D-ribofuranosyl)-2-(4-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-
L
-proline (8a)
3
3
(d, 1H, J1’-2’ = 4.3 Hz, H-C(1’)), 6.08 (dd, 1H, J1’-2’ = 4.3 Hz,
Triethylamine (0.13 mL, 1.00 mmol, 2 equiv.) was added to
a solution of bis-triazolyl derivative 7 (0.33 g, 0.50 mmol,
1 equiv.) and L-proline (0.11 g, 1.00 mmol, 2 equiv.) in DMSO
3J2’-3’ = 5.8 Hz, H-C(2’)), 5.85 (t, 1H, J2’-3’ = J3’-4’ = 5.8 Hz,
H-C(3’)), 4.79‒4.52 (m, 1H, H-C(2’’)), 4.45 (dd, 1H, J4’-5a’
3.8 Hz, J5a’-5b’
3
3
3
=
2
= 12.4 Hz, Ha-C(5’)), 4.44‒4.26 (m, 3H,
(10 mL). The resulting reaction mixture was stirred for 2‒3 h at
20‒25 ºC (HPLC control). Then crushed ice (40 mL) was added
to the reaction mixture followed by addition of diluted (1:1)
hydrochloric acid until the pH ~1. The resulting mixture was
kept at 0‒5 ºC for 24 h. The resulting precipitate was filtered,
washed on the filter with cold water (30 mL) and dried under
reduced pressure. The crude product was purified by silica gel
column chromatography (EtOH in EtOAc; gradient 0%→ 50%)
to give compound 8a (0.18 g, 60%) as a white powder. Product
8a exists as a 6:4 mixture of rotamers in a mixture of DMSO-d6
+ D2O. Rf = 0.5 (EtOAc/EtOH 1:1); IR (KBr) ν (cm-1): 3445,
2965, 1750, 1605, 1420, 1425, 1370, 1230, 1020. 1H-NMR
(300 MHz, DMSO-d6 + D2O) δ (ppm): 9.25 (s, 0.4H,
H-C(triazole)), 9.21 (s, 0.6H, H-C(triazole)), 8.37 (s, 0.6H,
H-C(8)), 8.29 (s, 0.4H, H-C(8)), 8.01 (d, 0.8H, 3J = 7.6 Hz, Ar),
H-C(4’), Hb-C(5’), (-OH)), 2.16, 2.09, 1.89 (3s, 9H,
3
H3CC(O)O-C (2’,3’,5’)), 1.54 (d, 3H, J = 6,9 Hz, H3C(3’’)).
13C-NMR (75.5 MHz, DMSO-d6+D2O) δ (ppm): 175.3, 170.4,
170.2, 169.8, 154.2, 149.2, 148.6, 146.8, 141.5, 130.1, 129.3,
128.7, 125.8, 120.0, 119.3, 86.7, 79.5, 72.7, 70.2, 62.7, 50.8,
20.6, 2.05, 20.4, 18.5. HRMS (ESI) calcd for C27H29N8O9
[M+H]+, 609.2052; found 609.2047 (0.82 ppm).
4.4. N-(9-(2’,3’,5’-Tri-O-acetyl-β-D-ribofuranosyl)-2-(4-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-L-
phenylalanine (8c)
Product 8c was obtained by the general synthetic procedure
using bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.),
L-
phenylalanine (0.17 g, 1.00 mmol, 2 equiv.) and triethylamine
(0.13 mL, 1.00 mmol, 2 equiv.). Silica gel chromatography
(EtOH in EtOAc; gradient 25%→50%) provided product 8c
(0.16 g, 48%) as a white powder. Rf = 0.61 (EtOAc/EtOH 3:1);
IR (KBr) ν (cm-1): 3373, 1753, 1615, 1495, 1455, 1435, 1415,
3
3
7.93 (d, 1.2H, J = 7.6 Hz, Ar), 7.49 (d, 0.8H, J = 7.6 Hz, Ar),
3
7.44‒7.27 (m, 2.2H, Ar), 6.27 (d, 0.6H, J1’-2’ = 5.0 Hz,
H-C(1’)), 6.24 (d, 0.4H, J1’-2’ = 4.5 Hz, H-C(1’)), 6.01 (dd,
3
3
3
0.4H, J1’-2’ = 4.5 Hz, J2’-3’ =5.9 Hz, H-C(2’)), 5.96 (dd, 0.6H,
3J1’-2’ = 5.0 Hz, 3J2’-3’ =6.0 Hz, H-C(2’)), 5.81 (dd, 0.4H, 3J2’-3’
=
1
o
5.9 Hz, 3J3’-4’ =5.4 Hz, H-C(3’)), 5.74 (dd, 0.6H, 3J2’-3’ = 6.0 Hz,
3J3’-4’ =4.7 Hz, H-C(3’)), 5.32‒5.25 (m, 0.4H, H-C(proline)),
4.70‒4.61 (m, 0.6H, H-C(proline)), 4.50‒4.22 (m, 3H, H-C(4’),
H2C(5’)), 4.19‒4.06 (m, 1.2H, H-C(proline)), 3.96‒3.84 (m,
0.4H, H-C(proline)), 3.82‒3.73 (m, 0.4H, H-C(proline)), 2.30‒
1374, 1234; H-NMR (300 MHz, 70 C, DMSO-d6 + D2O) δ
(ppm): 9.03 (s, 1H, H-C(triazole)), 8.29 (s, 1H, H-C(8)), 7.98
3
3
(d, 2H, J = 7.6 Hz, Ar), 7.49 (t, 2H, J = 7.6 Hz, Ar), 7.39 (t,
1H, 3J = 7.6 Hz, Ar), 7.24 (d, 2H, 3J = 7.2 Hz, Ar), 7.11 (t, 2H,
3J = 7.2 Hz, Ar), 7.04 (t, 1H, 3J = 7.4 Hz, Ar), 6.23 (d, 1H, 3J1’-
3
3
= 4.0 Hz, H-C(1’)), 6.01 (dd, 1H, J1’-2’ = 4.0 Hz, J2’-3’
=
1.84 (m,
4H,
H-C(proline)),
2.14,
2.05 (2s,
6H,
2’
3
3
H3CC(O)O-C(2’,3’,5’)), 1.94 (s, 1.8H, H3CC(O)O-C(2’,3’,5’)),
1.83 (s, 1.2H, H3CC(O)O-C(2’,3’,5’)). 13C-NMR (300 MHz,
DMSO-d6 + D2O) δ (ppm): 175.8/175.2, 170.6/170.5, 170.3/
170.1, 169.9/169.8, 153.5/152.4, 150.4/149.5, 149.0/148.6,
146.8/146.7, 140.1 (C(8) for both rotamers), 130.3/130.2,
129.3/129.2, 128.7/128.5, 125.9/125.7, 120.5/120.1, 119.9/
119.4, 86.3/86.1, 80.0/79.5, 72.8/72.6, 70.6/70.3, 63.2 (C(5’)
for both rotamers), 63.0/62.8, 49.4/48.1, 31.3/29.2, 24.8/22.0,
20.7 (CH3CO for both rotamers), 20.6 (CH3CO for both
rotamers), 20.5/20.4. HRMS (ESI): calcd for C29H31N8O9
[M+H]+, 635.2214; found 635.2210 (0.63 ppm).
5.7 Hz, H-C(2’)), 5.83 (t, 1H, J2’-3’ = J3’-4’ = 5.7 Hz, H-C(3’)),
4.90‒4.75 (m, 1H, H-C(2’’)), 4.45‒4.34 (m, 2H, H-C(4’), Ha-
3
2
C(5’)), 4.28 (dd, 1H, J4’-5b’ = 6.5 Hz, J5a’-5b’ = 13.2 Hz, Hb-
C(5’)), 3.50‒3.39 (m, 1H, Ha-C(3’’)), 3.19 (dd, 1H, 3J =
6.7 Hz, 2J= 13.8 Hz, Hb-C(3’’)), 2.13, 2.06, 1.84 (3s, 9H,
H3CC(O)O-C (2’,3’,5’)). 13C-NMR (75.5 MHz, DMSO-d6) δ
(ppm): 172.0, 170,5, 170.2, 169.8, 154.2, 149.0, 148.3, 146.4,
141.1, 138.8, 130.0, 129.5, 128.9, 128.3, 127.7, 125.8, 125.6,
119.7, 119.0, 86.1, 79.2, 72.4, 70.0, 65.6, 56.0, 37.1, 20.4, 20.3,
20.2. HRMS (ESI) calcd for C33H33N8O9 [M+H]+, 685.2365;
found 685.2356 (1.31 ppm).
4.3. General procedure for the preparation of purine
nucleoside - amino acid conjugates 8b-h: N-(9-(2’,3’,5’-Tri-
4.5. N-(9-(2’,3’,5’-Tri-O-acetyl-β-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-
D
-ribofuranosyl)-2-(4-
L
-valine (8d)
O-acetyl-β-D-ribofuranosyl)-2-(4-phenyl-1H-1,2,3-triazol-1-
Product 8d was obtained by the general synthetic procedure
using bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.),
yl)-9H-purin-6-yl)-
L
-alanine (8b)
L-
Triethylamine (0.13 mL, 1.00 mmol, 2 equiv.) was added to
a solution of bis-triazolyl derivative 7 (0.33 g, 0.50 mmol,
1 equiv.) and L-alanine (0.09 g, 1.00 mmol, 2 equiv.) in DMSO
(10 mL). The resulting reaction mixture was stirred for 15‒24 h
at 50 ºC (HPLC control). Then crushed ice (40 mL) was added
to the reaction mixture followed by addition of diluted (1:1)
hydrochloric acid until the pH ~1. The resulting mixture was
valine (0.12 g, 1.00 mmol, 2 equiv.) and triethylamine
(0.13 mL, 1.00 mmol, 2 equiv.). Silica gel chromatography
(MeOH in CH2Cl2; gradient 10%→20%) provided product 8d
(0.16 g, 50%) as a yellowish powder. Rf = 0.50 (CH2Cl2/MeOH
20:1); IR (KBr) ν (cm-1): 3389, 2965, 1752, 1620, 1493, 1434,
1
o
1373, 1233; H-NMR (300 MHz, 70 C, DMSO-d6 + D2O) δ
(ppm): 9.10 (s, 1H, H-C(triazole)), 8.26 (s, 1H, H-C(8)), 7.92

8
Tetrahedron
ACCEPTED MANUSCRIPT
3
3
3
2
(d, 2H, J = 7.5 Hz, Ar), 7.46 (t, 2H, J = 7.5 Hz, Ar), 7.36 (t,
4.30 (dd, 1H, J4’-5b’ = 6.6 Hz, J5a’-5b’ = 13.3 Hz, Hb-C(5’)),
3
3
1H, J = 7.5 Hz, Ar), 6.22 (d, 1H, J1’-2’ = 3.5 Hz, H-C(1’)),
6.03‒5.96 (m, 1H, H-C(2’)), 5.84 (t, 1H, 3J2’-3’ = 3J3’-4’ = 5.7 Hz,
H-C(3’)), 4.66‒4.55 (m, 1H, H-C(2’’)), 4.43‒4.32 (m, 2H, H-
2.14, 2.07, 1.85 (3s, 9H, H3CC(O)O-C(2’,3’,5’)), 2.11‒2.02 (m,
1H, H-C(3’’)), 1.72‒1.57 (m, 1H, Ha-C(4’’)), 1.31‒1.25 (m,
3
1H, Hb-C(4’’)), 0.99 (d, 3H, J = 6.8 Hz, H3C-C(3’’)), 0.93 (t,
3
2
3
C(4’), Ha-C(5’)), 4.26 (dd, 1H, J4’-5b’ = 5.2 Hz, J5a’-5b’
=
3H, J = 7.4 Hz, H3C(5’’)). 13C-NMR (75.5 MHz, DMSO-
12.7 Hz, Hb-C(5’)),2.34‒2.20 (m, 1H, H-C(3’’)), 2.10, 2.04,
1.76 (3s, 9H, H3CC(O)O-C(2’,3’,5’)), 0.94 (d, 6H, 3J = 6.6 Hz,
H3C(4’’)). 13C-NMR (75.5 MHz, DMSO-d6+D2O) δ (ppm):
175.8, 171.6, 171.3, 171.0, 155.0, 149.8, 149.0, 147.7, 142.3,
130.3, 130.0, 129.6, 126.4, 120.6, 119.6, 87.7, 79.9, 73.5, 70.6,
63.0, 61.3, 31.7, 21.1, 21.0, 20.9, 20.5, 18.9. HRMS (ESI)
calcd for C29H33N8O9 [M+H]+, 637.2365; found 637.2362
(0.47 ppm).
d6+D2O) δ (ppm): 173.6, 170.4, 170.2, 169.8, 154.8, 149.1,
148.8, 146.9, 141.6, 130.1, 129.3, 128.8, 125.8, 119.9, 119.2,
86.8, 79.6, 72.8, 70.2, 62.8, 59.0, 36.9, 25.4, 20.6, 20.5, 20.4,
16.1, 11.7. HRMS (ESI) calcd for C30H35N8O9 [M+H]+,
651.2522; found 651.2536 (2.15 ppm).
4.8. N-(9-(2’,3’,5’-Tri-O-acetyl-β-D-ribofuranosyl)-2-(4-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-glycine (8g)
Product 8g was obtained by the general synthetic procedure
using bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.),
glycine (0.08 g, 1.00 mmol, 2 equiv.) and triethylamine
(0.13 mL, 1.00 mmol, 2 equiv.). Silica gel chromatography
(CH2Cl2/EtOH 3:1) provided product 8g (0.17 g, 59%) as a
white powder. Rf = 0.25 (CH2Cl2/EtOH 3:1); IR (KBr) ν (cm-1):
4.6. N-(9-(2’,3’,5’-Tri-O-acetyl-β-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-
D
-ribofuranosyl)-2-(4-
L
-leucine (8e)
1
Product 8e was obtained by the general synthetic procedure
using bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.),
-leucine (0.13 g, 1.00 mmol, 2 equiv.) and triethylamine
3400, 1751, 1626, 1492, 1459, 1438, 1376, 1234; H-NMR
o
(300 MHz, 70 C, DMSO-d6 + D2O) δ (ppm): 9.12 (s, 1H, H-
C(triazole)), 8.31 (s, 1H, H-C(8)), 7.96 (d, 2H, 3J = 7.5 Hz, Ar),
7.48 (t, 2H, 3J = 7.5 Hz, Ar), 7.38 (t, 1H, 3J = 7.5 Hz, Ar), 6.25
L
(0.13 mL, 1.00 mmol, 2 equiv.). Silica gel chromatography
(MeOH in CH2Cl2; gradient 10%→20%) provided product 8e
(0.19 g, 60%) as a yellowish powder. Rf = 0.23 (CH2Cl2/MeOH
9:1); IR (KBr) ν (cm-1): 3392, 2960, 1752, 1625, 1491, 1459,
1439, 1419, 1375, 1232; 1H-NMR (300 MHz, 70 oC, DMSO-d6
+ D2O) δ (ppm): 9.09 (s, 1H, H-C(triazole)), 8.33 (s, 1H, H-
3
3
(d, 1H, J1’-2’ = 3.9 Hz, H-C(1’)), 6.04 (dd, 1H, J1’-2’ = 3.9 Hz,
3
3J2’-3’
=
5.9 Hz, H-C(2’)), 5.84 (t, 1H, J2’-3’ = =
3J3’-4’
5.9 Hz, H-C(3’)) 4.45‒4.36 (m, 2H, H-C(4’), Ha-C(5’)),
3
2
4.29 (dd, 1H, J4’-5b’ = 6.2 Hz, J5a’-5b’ = 13.2 Hz, Hb-C(5’)),
4.15 (br. s, 2H, H2C(2’’)), 2.12, 2.06, 1.82 (3s, 9H, H3CC(O)O-
C(2’,3’,5’)); 13C-NMR (75.5 MHz, DMSO-d6+D2O) δ (ppm):
172.5, 170.8, 170.6, 170.2, 155.1, 149.4, 148.8, 147.1, 141,9,
130.2, 129.6, 129.0, 126.0, 120.3, 119.6, 87.1, 79.6, 73.0, 70.3,
62.8, 44.0, 20.8, 20.7, 20.6. HRMS (ESI) calcd for C26H27N8O9
[M+H]+, 595.1896; found 595,1881 (2.52 ppm).
3
3
C(8)), 7.96 (d, 2H, J = 7.5 Hz, Ar), 7.49 (t, 2H, J = 7.5 Hz,
3
3
Ar), 7.38 (t, 1H, J = 7.5 Hz, Ar), 6.25 (d, 1H, J1’-2’ = 4.0 Hz,
3
3
H-C(1’)), 6.05 (dd, 1H, J1’-2’ = 4.0 Hz, J2’-3’
=
5.8 Hz,
3
3
H-C(2’)), 5.85 (t, 1H, J2’-3’ = J3’-4’ = 5.8 Hz, H-C(3’)), 4.87‒
4.62 (m, 1H, H-C(2’’)), 4.45‒4.37 (m, 2H, H-C(4’), Ha-C(5’)),
3
2
4.29 (dd, 1H, J4’-5b’ = 6.2 Hz, J5a’-5b’ = 13.3 Hz, Hb-C(5’)),
2.13, 2.07, 1.83 (3s, 9H, H3CC(O)O-C(2’,3’,5’)) 1.82‒1.74 (m,
3H, H2C(3’’), H-C(4’’)), 0.97‒0.89 (m, 6H, H3C(5’’)). 13C-
NMR (75.5 MHz, DMSO-d6+D2O) δ (ppm): 175.2, 170.6,
170.4, 170.0, 154.9, 149.2, 148.8, 147.0, 141.7, 130.2, 129.8,
129.4, 128.9, 125.9, 119.3, 87.0, 79.5, 72.9, 70.2, 62.8, 53.4,
41.1, 25.0, 23.5, 22.1, 20.7, 20.6, 20.5. HRMS (ESI) calcd for
C30H35N8O9 [M+H]+, 651.2522; found 651.2523 (0.15 ppm).
4.9. N-(9-(2’,3’,5’-Tri-O-acetyl-β-D-ribofuranosyl)-2-(4-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-L-methionine
(8h)
Product 8h was obtained by the general synthetic procedure
using bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.),
L-
methionine (0.15 g, 1.00 mmol, 2 equiv.) and triethylamine
(0.13 mL, 1.00 mmol, 2 equiv.). Silica gel chromatography
(MeOH in CH2Cl2; gradient 10%→50%) provided product 8h
(0.18 g, 55%) as a white powder. Rf = 0.30 (CH2Cl2/MeOH
9:1); IR (KBr) ν (cm-1): 3370, 2928, 1749, 1622, 1490, 1440,
4.7. N-(9-(2’,3’,5’-Tri-O-acetyl-β-D-ribofuranosyl)-2-(4-
phenyl-1H-1,2,3-triazol-1-yl)-9H-purin-6-yl)-L-isoleucine
(8f)
1
o
1418, 1374, 1231; H-NMR (300 MHz, 70 C, DMSO-d6 +
D2O) δ (ppm): 9.12 (s, 1H, H-C(triazole)), 8.35 (s, 1H, H-
3
3
C(8)), 7.97 (d, 2H, J = 7.5 Hz, Ar), 7.49 (t, 2H, J = 7.5 Hz,
Product 8f was obtained by the general synthetic procedure
using bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.),
3
3
Ar), 7.38 (t, 1H, J = 7.5 Hz, Ar), 6.27 (d, 1H, J1’-2’ = 4.1 Hz,
H-C(1’)), 6.05 (dd, 1H, J1’-2’ = 4.1 Hz, J2’-3’
H-C(2’)), 5.84 (t, 1H, J2’-3’ = J3’-4’ = 5.7 Hz, H-C(3’)), 4.84‒
4.63 (m, 1H, H-C(2’’)), 4.47‒4.37 (m, 2H, H-C(4’), Ha-C(5’)),
L-
3
3
=
5.7 Hz,
isoleucine (0.13 g, 1.00 mmol, 2 equiv.) and triethylamine
(0.13 mL, 1.00 mmol, 2 equiv.). Silica gel chromatography
(MeOH in CH2Cl2; gradient 10%→15%) provided product 8f
(0.20 g, 63%) as a yellowish powder. Rf = 0.23 (CH2Cl2/MeOH
9:1); IR (KBr) ν (cm-1): 3394, 2965, 1751, 1624, 1490, 1458,
1441, 1419, 1374, 1231. 1H-NMR (300 MHz, 70 oC, DMSO-d6
+ D2O) δ (ppm): 9.10 (s, 1H, H-C(triazole)), 8.36 (s, 1H, H-
3
3
3
2
4.30 (dd, 1H, J4’-5b’ = 6.5 Hz, J5a’-5b’ = 13.2 Hz, Hb-C(5’)),
2.62 (t, 2H, 3J = 7.6 Hz, H2C(4’’)), 2.31‒2.15 (m, 2H,
H2C(3’’)), 2.14, 2.07, 1.86 (3s, 9H, H3CC(O)O-C(2’,3’,5’)),
2.02 (s, 3H, H3C-S-C(4’’)). 13C-NMR (75.5 MHz, DMSO-
d6+D2O) δ (ppm): 173.9, 170.5, 170.2, 169.9, 154.7, 149.1,
148.7, 146.9, 141.6, 130.1, 129.3, 128.8, 125.8, 120.0, 119.4,
86.7, 79.5, 72.8, 70.2, 62.8, 54.2, 31.7, 30.3, 20.6, 20.5, 20.4,
14.9. HRMS (ESI) calcd for C29H33N8O9S [M+H]+, 669.2086;
found 669.2086 (0 ppm).
3
3
C(8)), 7.97 (d, 2H, J = 7.5 Hz, Ar), 7.49 (t, 2H, J = 7.5 Hz,
3
3
Ar), 7.38 (t, 1H, J = 7.5 Hz, Ar), 6.27 (d, 1H, J1’-2’ = 4.3 Hz,
H-C(1’)), 6.05 (dd, 1H, J1’-2’ = 4.3 Hz, J2’-3’
H-C(2’)), 5.84 (t, 1H, J2’-3’ = J3’-4’ = 5.7 Hz, H-C(3’)), 4.84‒
4.60 (m, 1H, H-C(2’’), 4.46‒4.37 (m, 2H, H-C(4’), Ha-C(5’)),
3
3
=
5.7 Hz,
3
3

9
ACCEPTED MANUSCRIPT
4.10.
N-(9-(β-
D
-ribofuranosyl)-2-(4-phenyl-1H-1,2,3-
144.8, 130.8, 129.9, 129.1, 128.6, 125.6, 120.8, 86.4, 80.0,
72.7, 70.3, 62.9, 53.2, 52.9, 43.6, 31.6, 31.1, 26.7, 20.6, 20.5,
20.4. HRMS (ESI): calcd for C34H39N10O13S [M+H]+,
827.2419; found 827.2420 (0.12 ppm).
triazol-1-yl)-9H-purin-6-yl)-glycine (9)
Bis-triazolyl derivative 7 (0.33 g, 0.50 mmol, 1 equiv.) and
glycine (0.08 g, 1.00 mmol, 2 equiv.) were dissolved in DMSO
(10 mL). Triethylamine (0.13 mL, 1.00 mmol, 2 equiv.) was
added and the mixture was stirred for 15‒24 h at 50 ºC (HPLC
control). Then the reaction mixture was diluted with
brine (20 mL) and filtered. The resulting solution was treated
with an aqueous solution of methylamine (25 w-%, 4 mL) and
stirred for 2‒3 h at ambient temperature (HPLC control).
Precipitate was filtered, washed on the filter with cold water
(5 mL), ethanol (3×5 mL) and dried under reduced pressure.
Product 9 (0.15 g, 65%) was obtained as a white powder.
IR (KBr) ν (cm-1): 3369, 1637, 1492, 1459, 1440, 1384, 1243;
1H-NMR (300 MHz, DMSO-d6 + D2O) δ (ppm): 9.24 (s, 1H,
4.12. Purine nucleoside – nonapeptide conjugate 14
Triethylamine (1 ꢁL, 7.2 ꢁmol, 5.0 equiv.) was added to a
solution of bis-triazolyl derivative 7 (1.2 mg, 1.8 ꢁmol,
1.3 equiv.) and nonapeptide 13 (1.5 mg, 1.4 ꢁmol, 1.0 equiv.)
(CASLO ApS; Denmark; lyophilized trifluoroacetate salt).
HPLC-DAD and UPLC-HRMS analysis revealed a clean
conversion into purine nucleoside – nonapeptide conjugate 14
(tR = 5.10 min; HRMS (ESI) calcd for C68H94N21O21S [M+H]+,
1572.6648; found 1572.6611 (2.35 ppm)) and 4-phenyl-1H-
1,2,3-triazole (12) (tR = 4.94 min; HRMS (ESI) calcd for
C8H8N3 [M+H]+, 146.0713; found 146.0705 (5.48 ppm)).
HPLC eluent system: C (0.1% TFA water solution / MeCN
(95/5, V/V)), B (MeCN); gradient regime: 0‒5‒8‒10‒12 min,
B eluent change 0‒40‒95‒95‒0%. HPLC traces are provided in
figure 5.
3
H-C(triazole)), 8.44 (s, 1H, H-C(8)), 8.00 (d, 2H, J = 7.4 Hz,
Ar), 7.48 (t, 2H, 3J = 7.4 Hz, Ar), 7.37 (t, 1H, 3J = 7.4 Hz, Ar),
3
3
5.96 (d, 1H, J1’-2’ = 5.9 Hz, H-C(1’)), 4.62 (dd, 1H, J1’-2’
=
3
3
5.9 Hz, J2’-3’ = 4.9 Hz, H-C(2’)), 4.20 (dd, 1H, J2’-3’ = 4.9 Hz,
3J3’-4’ = 2.7 Hz, H-C(3’)), 4.02‒3.95 (m, 1H, H-C(4’)), 3.92 (s,
2H, H2C(2’’)), 3.68 (dd, 1H, 3J4’-5a’ = 3.8 Hz, 2J5a’-5b’ = 12.0 Hz,
Ha-C(5’)), 3.57 (dd, 1H, 3J4’-5b’ = 4.0 Hz, 2J5a’-5b’ = 12.0 Hz, Hb-
C(5’)). 13C-NMR (75.5 MHz, DMSO-d6+D2O) δ (ppm): 172.4,
154.5, 149.3, 149.1, 146.8, 140.4, 130.2, 129.4, 128.7, 125.9,
120.4, 119.2, 87.8, 86.0, 74.0, 70.6, 61.6, 45.0. HRMS (ESI)
calcd for C20H21N8O6 [M+H]+,469.1579; found 469.1581 (0.42
ppm).
Acknowledgements
The authors thank Latvian-Lithuanian-Taiwanese co-project
W1935//LV-LT-TW/2015/2 for financial support. K.O. and
D.C. thank the Latvian Academy of Sciences and JSC
“Olainfarm” for scholarships. K.O. is also indebted to K.
Morbergs foundation for scholarship. The authors thank
Professor V. Kampars for the UV and fluorescence spectra.
4.11. Purine nucleoside – glutathione conjugate 11
Water (0.1 mL) and DMSO (0.6 mL) was added to a
mixture of bis-triazolyl derivative 7 (75 mg, 0.11 mmol, 1.0
equiv.) and glutathione (10) (38 mg, 0.12 mmol, 1.1 equiv.)
and stirred for 5 minutes. Triethylamine (40 ꢁL, 0.28 mmol, 2.5
equiv.) was then added and stirring was continued for an
additional 10 minutes (TLC control, DCM/MeOH 9:1) at
ambient temperature. The resulting solution was purified on
reverse-phase C(18) silica gel column (MeCN/Water 0→33%)
to obtain compound 11 (76 mg, 82% (93% purity by HPLC)) as
a white powder. HPLC eluent system: A (0.01 M aqueous
KH2PO4 / MeCN (94/6, V/V)), B (MeCN); gradient regime: 0‒
7‒10‒12 min, B eluent change 0‒45‒95‒0%.
Supplementary data
Supplementary data associated with this article can be found
in the online version, at http://
References
(1)
(2)
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IR (KBr) ν (cm-1): 3440, 1750, 1655, 1585, 1360, 1240, 1050.
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3
7.5 Hz, H-C(Ar)), 7.51 (t, 2H, J = 7.5 Hz, H-C(Ar)), 7.38 (d,
3
1H, 3J = 7.5 Hz, H-C(Ar)), 6.38 (d, 1H, J1’-2’ = 4.7 Hz,
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3
3
H-C(1’)), 6.04 (dd, 1H, J1’-2’ = 4.7 Hz, J2’-3’ = 5.7 Hz, H-
3
3
C(2’)), 5.78 (dd, 1H, J2’-3’ = 5.7 Hz, J3’-4’ = 5.3 Hz, H-C(3’)),
4.78 (dd, 1H, J2’’-3a’’ = 4.5 Hz, J2’’-3b’’ = 9.3 Hz, H-C(2’’)),
4.49−4.42 (m, 2H, H-C(4’), Ha-C(5’)), 4.36 (dd, 1H, J5a’-5b’
13.0 Hz, J4’-5b’ = 6.7 Hz, Hb-C(5’)), 4.16 (dd, 1H, J2’’-3a’’
3
3
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2
=
=
=
=
3
3
2
2
Zhang, H.; Zhou, L.; Coats, S. J.; McBrayer, T. R.; Tharnish, P.
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4.5 Hz, J3a’’-3b’’ = 13.8 Hz, Ha-C(3’’)), 3.59 (d, 1H, J1a’’-1b’’
16.6 Hz, Ha-C(1’’)), 3.57 (dd, 1H, 2J3a’’-3b’’ = 13.8 Hz, 3J2’’-3b’’
9.3 Hz, Hb-C(3’’)), 3.47 (d, 1H, 2J1a’’-1b’’ = 16.6 Hz, Hb-C(1’’)),
3.33 (t, 1H, 3J5’’-6’’ = 6.4 Hz, H-C(6’’)), 2.47 (dd, 1H, 2J4a’’-4b’’
=
=
3
2
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H3CC(O)O-C(2’, 3’, 5’)), 2.02‒1.93 (m, 2H, H2C(5’’)). 13C-
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