Synthesis and structure-activity relationships of pyridinyl-1 H -1,2,3-triazolyldihydroisoxazoles as potent inhibitors of tubulin polymerization

Article abstract of DOI:10.1016/j.ejmech.2014.11.063

Three series of compounds; pyridinyl-1H-1,2,3-triazoles, pyridinyl-1H-1,2,3-triazolylisoxazoles and pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles with TMP moiety were designed, synthesized and screened for their anti-cancer and anti-tubulin properties. By sequentially designing three series of compounds comprising of dihydroisoxazole in the linker, a small substituent like chlorine on one side (R¹) and aromatic group (R) on the pyridine ring, we have optimized the anti-cancer as well as anti-tubulin activity. Pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles 28b and 28c were found to be potent anti-cancer agents against all the cell lines tested with a concomitant accumulation of cells in the G2/M phase of the cell cycle. Molecular modeling suggests that the trimethoxyphenyl ring in 28b and 28c occupies the cholchicine binding domain of β-tubulin, whereas, the dihydroisoxazole extends towards the interface of α/β-tubulin.

Full text of DOI:10.1016/j.ejmech.2014.11.063

Accepted Manuscript
Synthesis and structure-activity relationships of pyridinyl-1H-1,2,3-
triazolyldihydroisoxazoles as potent inhibitors of tubulin polymerization
P. Suman , T.Ramalinga Murthy , K. Rajkumar , D. Srikanth , Ch. Dayakar , Chandan
Kishor , Anthony Addlagatta , Shasi V. Kalivendi , B.China Raju
PII:
S0223-5234(14)01105-2
10.1016/j.ejmech.2014.11.063
EJMECH 7556
DOI:
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 19 September 2014
Revised Date: 17 November 2014
Accepted Date: 30 November 2014
Please cite this article as: P. Suman, T.R. Murthy, K. Rajkumar, D. Srikanth, C. Dayakar, C. Kishor, A.
Addlagatta, S.V. Kalivendi, B.C. Raju, Synthesis and structure-activity relationships of pyridinyl-1H-1,2,3-
triazolyldihydroisoxazoles as potent inhibitors of tubulin polymerization, European Journal of Medicinal
Chemistry (2015), doi: 10.1016/j.ejmech.2014.11.063.
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ACCEPTED MANUSCRIPT
Graphical Abstract:
Synthesis
and
structure-activity
relationships
of
pyridinyl-1H-1,2,3-
triazolyldihydroisoxazoles as potent inhibitors of tubulin polymerization
P. Suman, T. Ramalinga Murthy, K. Rajkumar, D. Srikanth, CH. Dayakar, Chandan Kishor,
Anthony Addlagatta, Shasi V Kalivendi, B. China Raju
Three series of compounds were prepared and among them, pyridinyl-1H-1,2,3-
triazolyldihydroisoxazoles (28b and 28c) with TMP was found to be potent anti-cancer
agents and tubulin inhibitors.

ACCEPTED MANUSCRIPT
Synthesis and structure-activity relationships of pyridinyl-1H-1,2,3-
triazolyldihydroisoxazoles as potent inhibitors of tubulin polymerization
a
b
a
b
a
P. Suman, T. Ramalinga Murthy, K. Rajkumar, D. Srikanth, Ch. Dayakar, Chandan
b
b
b
a
Kishor, Anthony Addlagatta,* Shasi V Kalivendi,* B. China Raju*
a
b
Natural Product Chemistry Division and Centre for Chemical Biology,
CSIR-Indian Institute of Chemical Technology, Hyderabad 500 007, India.
Corresponding authors: Tel: +91-40-27191725; Fax: +91-40-27160512.
Email: chinaraju@iict.res.in (BCR), kalivendi@iict.res.in (SVK), anthony@iict.res.in (AA)
ABSTRACT: Three series of compounds; pyridinyl-1H-1,2,3-triazoles, pyridinyl-1H-1,2,3-
triazolylisoxazoles and pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles with TMP moiety were
designed, synthesized and screened for their anti-cancer and anti-tubulin properties. By
sequentially designing three series of compounds comprising of dihydroisoxazole in the
1
linker, a small substituent like chlorine on one side (R ) and aromatic group (R) on the
pyridine ring, we have optimized the anti-cancer as well as anti-tubulin activity. Pyridinyl-
1
H-1,2,3-triazolyldihydroisoxazoles 28b and 28c were found to be potent anti-cancer agents
against all the cell lines tested with a concomitant accumulation of cells in the G2/M phase of
the cell cycle. Molecular modeling suggests that the trimethoxyphenyl ring in 28b and 28c
occupies the cholchicine binding domain of β-tubulin, whereas, the dihydroisoxazole extends
towards the interface of α,β-tubulin.
Keywords: 2-Chloronicotinaldehyes, Pyridinyl-1H-1,2,3-triazoles, Triazolylisoxazoles,
Triazolyldihydroisoxazoles, Anti-cancer activity, Anti-Tubulin activity, Molecular modeling.
1

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1
. Introduction
Cancer is the second major cause of death in U.S. and rest of the world [1]. Hormones,
immune conditions, inherited genes, viruses, chemicals and radiation are some of the
responsible factors for the development of various cancers [2]. The uncontrolled growth and
spreading of abnormal cells result in the disruption of tissues that leads to death in cancer
patients. Tumor requires the development of vasculature to gather oxygen and nutrients;
therefore, inhibition of tumor growth by targeting vasculature could be one of the efficient
ways to fight against cancer [3]. Microtubules are involved in cellular process and
suppression of microtubule polymerization blocks cell division at mitosis, leading to cell
death and also the disruption of tumor vasculature. Therefore, microtubules are proven targets
for the development of anti-cancer agents. Drugs that disrupt microtubule/tubulin dynamics
are widely used in cancer therapy. The majority of these molecules act by binding to tubulin,
an α,β-heterodimer that forms the core of the microtubule. Microtubule targeting agents
(MTA) perturb the mitosis and arrest the cell cycle during interphase [4]. MTAs are known to
interact with tubulin in any of the four sites namely laulimalide, taxane/epothilone, vinca
alkaloid, and colchicines binding pockets [5]. Because of its effective inhibition of mitosis,
colchicine (1, Fig. 1) is identified as effective anti-cancer agent. Colchicine based molecules
disrupt the vasculature agents which are under investigation for cancer therapy [6].
Combretastatins are natural products that bind to tubulin and inhibit its polymerization [7].
Combretastatin A-4 (2, CA-4, Fig. 1) is the potent anti-cancer drug, binds to the colchicine
binding site of β-tubulin. CA-4 exhibits strong growth inhibition at nanomolar concentrations
against a wide variety of human cancers including multidrug resistant (MDR) cancer [8].
Low water solubility of CA-4 limits its efficacy in vivo. Hence, water soluble combretastatin
analogs have been developed [9]. Combretastatin A-4P (3, fosbretabulin disodium, Fig. 1),
and its amino acid derivative AVE-8062 (4, Ombrabulin, Fig. 1) act as vascular-disrupting
2

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agents (VDA) by rapidly depolymerizing microtubules. Structure-activity relationship (SAR)
studies have shown that 3,4,5-trimethoxyphenyl ring (TMP, 5-6, Fig. 1) [10] and 4-
methoxysubstituted phenyl ring by a double bond with cis geometry are important for
cytotoxic activity of the combretastatins [11]. Therefore, various five-membered heterocyclic
compounds such as thiazoles, imidazolones, pyrazoles, and triazoles (7, Fig. 1) with cis
restricted geometry were reported as tubulin inhibitors [12]. Based on these results, the TMP
moiety was found to be essential for anti-tubulin activity. In a recent study, Shetty et al
synthesized nicotinyl based 1H-indolyl-methoxyphenylsulfonamide derivatives as potent
antimitotic agents [13].
2
-Chloronicotinaldehydes based heterocyclic compounds, such as, Imidacloprid
analogues, 2-chloro-5-methylpyridine-3-olefin derivatives, 1,8-naphthyridines and Baylis-
Hillman (BH) adducts were synthesized. BH adducts displayed potent anti-malarial activity;
quinolines and olefins derivatives displayed good anti-microbial activity [14]. Further, 2-
chloronicotinaldehyde based Knoevenagel derivatives, (E)-α,β-unsaturated esters/ketones and
1
H-1,2,3-triazolylbenzohydrazides displayed promising anti-mycobacterial and anti-cancer
activities [15]. As part of our research interest in the design and synthesis of biologically
important heterocyclic compounds [16], we focused on 2-chloronicotinaldehydes in
combination with TMP. Based on our design, we presume that TMP will act as a
pharmacophore in directing the molecules into the colchicine binding pocket of tubulin while
the variable region provides affinity. In this study, we report synthesis of 2-
chloronicotinaldehydes based 1H-1,2,3-triazoles, 1H-1,2,3-triazolylisoxazoles and 1H-1,2,3-
triazolyldihydroisoxazoles with TMP and evaluation of their anti-tubulin and anti-
proliferative activity.
3

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2
. Chemistry
Three series of compounds such as pyridinyl-1H-1,2,3-triazoles 13a-j, pyridinyl-1H-1,2,3-
triazolylisoxazoles 25a-j and pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles 28a-j with
trimethoxyphenyl moiety were prepared starting from 2-chloronicotinaldehydes (Scheme 1-
6
). Vilsmeier reaction of enamides provided 2-chloronicotinaldehydes 8a-d [17]. Sodium
borohydride (NaBH ) reduction of 2-chloronicotinaldehydes 8a-d afforded corresponding
4
alcohols 9a-d and subsequent azidation with diphenylphosphoryl azide (DPPA) in presence
of DBU at room temperature [11a] provided azides 10a-d (Scheme 1). Cycloaddition reaction
(Click) of azide 10a with propargyl alcohol in presence of copper sulphate and sodium
ascorbate in aqueous tert-butanol at room temperature afforded pyridinyl-1H-1,2,3-
triazolylmethanol 11 [18]. Initially, we have chosen the Williamson reaction to prepare
pyridinyl-1H-1,2,3-triazole 13a. Triazolyl alcohol 11 and trimethoxy benzyl bromide 12 in
presence of K CO and KI in acetone under reflux conditions provided pyridinyl-1H-1,2,3-
2
3
triazole 13a in moderate yield (21%, Scheme 2). A parallel etherification reaction with NaH
in dry DMF also did not improve the yield. Hence, we have developed an alternate method to
prepare pyridinyl-1H-1,2,3-triazoles 13a-d with improved yields as depicted in Scheme 3.
The synthetic strategy for the preparation of pyridinyl-1H-1,2,3-triazoles 13a-d has been
accomplished by employing click reaction of 3-(azidomethyl)-2-chloropyridines 10a-d with
1
,2,3-trimethoxy-5-propynyloxy-methylbenzene 16. NaBH reduction of 3,4,5-trimethoxy-
4
benzaldehyde 14 provided alcohol 15 and subsequent propargylation of 15 with propargyl
bromide in presence of NaH in dry THF afforded trimethoxy-5-propynyloxy-methylbenzene
1
6 [19]. Click reaction of azides 10a-d with alkyne 16 in presence of copper sulphate and
sodium ascorbate in aqueous tert-butanol provided triazoles 13a-d (Scheme 3). Having
achieved the synthesis of pyridinyl-1H-1,2,3-triazoles 13a-d, in the next step, we have
prepared morpholine and thiomorpholine substituted pyridinyl compounds 13e-j.
4

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Nucleophilic
substitution
reaction
of
2-chloronicotinaldehydes
8a-c
with
morpholine/thiomorpholine 17a-b in presence of K CO in dry DMF at 110 °C furnished the
2
3
corresponding nicotinaldehydes 18a-f. Nicotinaldehydes 18a-f upon reduction, azidation and
click reaction provided corresponding pyridinyl-1H-1,2,3-triazoles 13e-j (Scheme 4). All
compounds 13a-j reported in this manuscript are new and hence are characterized by various
spectroscopic methods.
After successful preparation of 13a-j, the present protocol was extended to prepare
pyridinyl-1H-1,2,3-triazolylisoxazoles 25a-j and dihydroisoxazoles 28a-j. Oxime 22 was
prepared from 3,4,5-trimethoxybenzaldehyde 21 with hydroxylamine hydrochloride in
MeOH-H O. 1,3-Dipolar cycloaddition of oxime 22 with propargyl alcohol furnished
2
trimethoxyphenyl-isoxazolyl-methanol 23 [20]. Propargylation of 23 with propargyl bromide
provided corresponding alkyne derivative 24. Cycloaddition of substituted 3-
azidomethylpyridines 10a-d and 20a-f with alkyne 24 furnished pyridinyl-1H-1,2,3-
triazolylisoxazoles 25a-j (Scheme 5). Next, 1,3-dipolar cycloaddition of oxime 22 with allyl
alcohol produced trimethoxyphenyl-dihydroisoxazolyl-methanol 26 and upon propargylation
of 26 with propargyl bromide furnished corresponding alkyne 27. Cycloaddition of
substituted azides 10a-d and 20a-f with alkyne 27 provided dihydroisoxazoles 28a-j (Scheme
6
). 25a-j and 28a-j have been characterized by spectroscopic methods.
3
. Biology
Three set of compounds i) pyridinyl-1H-1,2,3-triazoles, ii) pyridinyl-1H-1,2,3-
triazolylisoxazoles
and
iii)
pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles
with
trimethoxyphenyl (TMP) were synthesized. Anti-cancer activity was performed on four
cancer cell lines. In addition, the ability of compounds in inhibiting tubulin polymerization
was studied. The interactions of inhibitor within the colchicine binding pocket of tubulin was
5

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also analyzed employing molecular modeling studies in order to identify the mode of
inhibition of the most active compounds.
4
. Results and discussion
The pyridinyl-1H-1,2,3-triazoles 13a-j, pyridinyl-1H-1,2,3-triazolylisoxazoles 25a-j and
pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles 28a-j were screened for anti-cancer activity on
four cancer cell lines (lung adenocarcinoma (A549), prostate cancer (DU-145), human
epithetical cervical cancer (HeLa) and human breast cancer (MCF7) by MTT assay [21]
followed by anti-tubulin activity and compared the efficacy with colchicine [22].
4
.1. Anti-cancer activity of trimethoxybenzyloxy-1H-1,2,3-triazolylpyridines
The IC values of the pyridinyl-1H-1,2,3-triazoles 13a-j were tabulated in Table 1; based
5
0
on the obtained IC values against the four cancer cell lines and percentage inhibition of
5
0
tubulin polymerization, moderate activity was observed. There was no specific pattern among
the compound structures and their inhibition potential suggesting that the variable region
(triazolylpyridine derivatives) is not able to dock in the enzyme pocket that also affects their
anti-cancer activity. It is important to note that the TMP in the combretastatin is linked to a
cis-stilbene which is a rigid juncture. Since a flexible linker in the 13a-j series connects the
TMP, to bring some rigidity near the TMP group we decided to add an isoxazole moiety.
Based on this we have synthesized next series of compounds 25a-j.
4
.2. Anti-cancer activity of 1H-1,2,3-triazolyl-3,4,5-trimethoxyphenylisoxazoles
The IC50 values of the pyridinyl-1H-1,2,3-triazolylisoxazoles 25a-j were presented in
Table 2. Based on the observed IC50 values against four cancer lines and extent of anti-
tubulin activity, some changes in the potency have been observed as compared to the 13a-j
series. Compounds 25c and 25i displayed improved activity against tubulin polymerization as
compared to their counterparts (13c and 13i respectively). However, an opposite trend is
6

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noticed in the case of 25e, 25h and 25j. We believe that our design from 13 to 25 series is in
the right direction. However, based on modeling studies we realized that converting the
isoxazole to dihydroisoxazole might provide increase in flexibility to the linker that may
bring better affinity for the molecules to bind to the tubulin. Therefore, we have designed and
synthesized dihydroisoxazoles 28a-j and their activity was measured against four cancer cell
lines and their anti-tubulin effects.
4
.3. Anti-cancer activity of 1H-1,2,3-triazolyl-3,4,5-trimethoxyphenyl-4,5-dihydroisoxazoles
The IC values of the pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles 28a-j are presented
5
0
in Table 3. As inferred from the modeling studies, there has been a drastic improvement in
the affinity of molecules to bind tubulin and thereby exhibited better anti-cancer activity. Five
compounds 28a-d and 28h displayed improved activity. There is clear correlation with
respect to anti-tubulin activity and cytotoxicity. For instance, 28b and 28c inhibited the
tubulin polymerization to about 70% at 10 ꢀM concentration and these compounds exhibited
cytotoxicity in low micromolar concentrations against all the four cell lines tested. Since
there is a direct correlation between tubulin polymerization and observed cytotoxicity, we
rationalized their behavior using structure-activity relationship. Among the 28a-j, the
compounds 28a-d, one of the substituents on the pyridine ring was common (chlorine atom at
nd
th
2
position) while substituent at 5 position is variable. The best activity was observed when
a phenyl 28b or p-tolyl 28c group occupied this position. However, when the p-flurophenyl
group 28d replaces this position, a slight decrease in the activity was observed. The
compound with a small substituent like methyl 28a also displayed better activity. On the
nd
other hand, when morpholine or thiomorpholine group replaces the chlorine atom at 2
position, invariably, almost all molecules except for 28h lost their potential against tubulin
polymerization as well as anti-cancer activity. To confirm that the anti-cancer activity by 28b
7

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and 28c is indeed through their anti-tubulin effects (Table 4), we have carried out series of
experiments described below.
4
.4. Anti-mitotic effects of compounds 28b-c
Previous reports suggest that the pharmacological inhibition of tubulin assembly leads to
cell cycle arrest at G2/M phase [22]. To examine the anti-mitotic effects of pyridinyl-1H-
,2,3-triazolyldihydroisoxazole derivatives, we performed cell cycle analysis of most active
compounds 28b and 28c by flow cytometry. Cell cycle analysis revealed that treatment with
1
2
2
4
8b and 28c exhibited 87% and 78% accumulation of cells in G2/M phase respectively (Fig.
).
.5. Effects of 28b and 28c on cellular microtubules network and nuclear morphology
To examine the anti-tubulin effects of compounds 28b and 28c at cellular level, we treated
HeLa cells at 10 µM concentrations for 24 h and analyzed the cellular microtubule network
by immunohistochemistry followed by nuclear staining with 4',6-diamidino-2-phenylindole
(DAPI). Results demonstrate that cells treated with 28b and 28c exhibited severely disrupted
microtubule organization, as compared to the controls (Fig. 3). The planar arrangement of
chromosomes in 28b and 28c treated cells in conjunction with distinct microtubule
organization centers clearly indicates mitotic-arrest, whereas, control cells exhibited a normal
microtubule organization (Fig. 3).
4
.6. Cell cycle related expression of Cyclin B1
Cyclin B1/CDc2 complex regulates the progression of cell cycle at G2/M phase. Maximum
expression of cyclin B1 expression will be seen during metaphase [23]. Immunoblot analysis
of cyclin B1 following treatment of cells with 28b and 28c (10 ꢀM for 24 h) resulted in an
increased expression of cyclin B1 as compared to DMSO treated controls [24]. Nocodazole, a
known anti-tubulin agent (2 µM) also increased cyclin B1 expression under similar
8

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experimental conditions (Fig. 4A). Therefore, the increased protein levels of cyclin B1 by
2
8b and 28c confirms that these compounds acts as anti-tubulin agents and block the cells at
mitotic phase.
4
.7. Analysis of soluble versus polymerized tubulin in cells
A dynamic equilibrium exists between the intracellular pool of α,β-tubulin heterodimers
and the microtubule polymer. Microtubule disrupting agents target this dynamic equilibrium
22]. Microtubule stabilizing (Paclitaxel) drugs promote polymerization, whereas, tubulin
[
depolymerization agents (Nocodazole and Colchicin) inhibit polymerization [25]. To further
analyze whether the block in cell cycle at G2/M by 28b and 28c can be reflected even in the
cellular levels of soluble and polymerized tubulin (microtubules), we treated HeLa cells with
2
8b and 28c at 10 µM concentration for 24 h and nocodazole (2 µM) was employed as a
positive control. Following treatments, intracellular levels of soluble (free tubulin) and
polymerized (tubulin from microtubules) fractions of tubulin were analyzed by
immunoblotting. Results indicate that while DMSO treated cells (controls) exhibited nearly
equal distribution of tubulin in soluble and insoluble fraction, cells treated with nocodazole
demonstrated complete shift into soluble fraction. Similar to nocodazole, cells treated with
2
8b and 28c showed higher amounts of tubulin in the soluble fraction as compared to the
polymerized fraction (Fig. 4B).
4
.8. Docking study
AutoDock version 4.2 was used to dock pyridinyl-1H-1,2,3-triazolylisoxazoles 25b-c and
pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles 28b-c into the colchicine-binding site of β-
tubulin [26]. Results suggest that the docking position of the trimethoxyphenyl ring binds
well in the colchicine-binding pocket with extensive hydrophobic contacts within the binding
pocket of the β-chain (Fig. 5). TMP binds in the pocket where the A ring of colchicine
normally binds and surrounded by amino acids Cys241, Leu242, Ala250, Leu255, Val318,
9

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and Ile378 of β-chain. Presence of isoxazole (25b and 25c) and 4,5-dihydroisoxazole (28b
and 28c) affects the activity. Due to the presence of 4,5-dihydroisoxazole in compounds 28b
and 28c, triazole moiety bends toward Asn101, Ser178 and Thr179 of α-chain and
participates in a series of hydrogen bonds. In the case of planar isoxazole (25b and 25c),
triazole moves away from these amino acids. Extensive interactions of 28b and 28c with the
tubulin core when compared to 25b and 25c may be responsible for their enhanced anti-
tubulin effects. A similar explanation can be provided for pyridinyl-1H-1,2,3-triazole series
1
3a-j that lack the 4,5-dihydroisoxazole moiety for their poor activity profile. Chlorine atom
of 2-chloro-5-phenylpyridine 28b is involved in hydrogen bond with side chain of Ser178.
This also suggests that the space in this region is limited and bulkier substituent’s like
morpholine and thiomorpholine 28e-j cannot occupy this position providing the basis for
poor activity of compounds with these substitutions.
5
. Conclusion
In conclusion, three series of compounds such as pyridinyl-1H-1,2,3-triazoles, pyridinyl-
H-1,2,3-triazolylisoxazoles and pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles were
1
prepared and screened for their anti-proliferative and anti-tubulin activity. By sequential
designing of three series of compounds comprising dihydroisoxazole in the linker, a small
1
substituent like chlorine on one side (R ) and aromatic group (R) on the other on the pyridine
ring, we have optimized the anti-tubulin as well as anti-cancer activity. Pyridinyl-1H-1,2,3-
triazolyldihydroisoxazoles 28b and 28c were found to be potent anti-cancer agents against all
the cell lines tested with accumulation of cells in the G2/M phase of the cell cycle. Molecular
modeling suggests that the trimethoxyphenyl ring in 28b and 28c occupy the cholchicine
binding domain of β-tubulin, whereas, the dihydroisoxazole extends towards the interface of
α,β-tubulin. Thus, these results suggest that pyridinyl-1H-1,2,3-triazolyldihydroisoxazoles,
1
0

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particularly 28b and 28c have potential to be developed as new a class of tubulin
polymerization inhibitors.
6
. Experimental section
6
.1. General
All chemicals and reagents were purchased from Aldrich (Sigma-Aldrich, USA), AVRA
Chemicals Pvt. Ltd (Hyderabad, India) and were used without further purification. Reactions
were monitored by thin layer chromatography (TLC) on pre-coated silica gel 60 F254 (mesh);
spots were visualized under UV light. Column chromatographic separations were carried out
on silica gel (60-120 mesh). Melting points were determined on a Mettler-Temp apparatus
and are uncorrected. An IR spectrum was recorded with a Thermo Nicolet Nexus 670
1
13
spectrometer. H NMR and C NMR spectra were recorded on Bruker Avance 300 and 500
MHz spectrometers. Chemical shifts (δ) are quoted in parts per million and are referenced to
tetramethylsilane (TMS) as internal standard. ESIMS obtained on 7070H spectrometer
operating at 70 eV using a direct inlet system. HRMS were carried out on Agilent 6510 Q-
TOF LC/MS instrument.
6
.2. General procedure for the synthesis of 2-chloronicotinaldehydes (8a-d)
The compounds 8a-d were prepared from its corresponding enamides as per our previous
reported method. Ref. [17]
.3. General procedure for the synthesis of 2-morpholine/thiomorpholinenicotinaldehydes
18a-f)
Morpholine 17a (1.2 mmol) was added to a stirred solution of 2-chloro-5-
methylnicotinaldehyde 8a (1 mmol) in N,N-dimethylformamide and K CO (1.5 mmol) at
6
(
2
3
room temperature. The reaction mixture was heated at 110 ºC for 5 h. After completion of the
reaction (TLC), the mixture was quenched with ice water and solid was filtered off. The solid
was recrystallized (EtOAc/hexane) gave 5-methyl-2-morpholinonicotinaldehyde 18a as
1
1

ACCEPTED MANUSCRIPT
yellow color solid. Similarly 18b-f was prepared from corresponding substituted 2-
chloronicotinaldehydes 8a-c.
6
.3.1. 5-Methyl-2-morpholinonicotinaldehyde (18a)
1
Yield: 86%; yellow solid; m.p: 90-92 ºC; H NMR (CDCl , 300 MHz): δ 2.32 (s, 3H), 3.38
3
13
(
t, J = 4.53 Hz, 4H), 3.88 (t, J = 4.53 Hz, 4H), 7.84 (s, 1H), 8.25 (s, 1H), 10.09 (s, 1H); C
NMR (CDCl , 75 MHz): δ 17.21, 51.75, 66.77, 119.60, 126.106, 140.33, 152.86, 160.42,
3
+
1
90.00; ESI-MS: m/z, 207 [M+H] .
6
.3.2. 5-Methyl-2-thiomorpholinonicotinaldehyde (18b)
1
Yield: 83%; yellow solid; m.p: 85-87 ºC; H NMR (CDCl , 300 MHz): δ 2.30 (s, 3H), 2.80
3
13
(
t, J = 5.03 Hz, 4H), 3.66 (t, J = 5.03 Hz, 4H), 7.84 (s, 1H), 8.23 (s, 1H), 10.04 (s, 1H); C
NMR (CDCl , 75 MHz): δ 17.22, 27.53, 53.97, 119.75, 126.08, 139.99, 152.79, 161.18,
3
+
1
89.99; ESI-MS: m/z, 223 [M+H] .
6
.3.3. 2-Morpholino-5-phenylnicotinaldehyde (18c)
1
Yield: 88%; yellow solid; m.p: 110-112 ºC; H NMR (CDCl , 300 MHz): δ 3.52 (t, J =
3
4
.14 Hz, 4H), 3.89 (t, J = 4.14 Hz, 4H), 7.38 (t, J = 6.77 Hz, 1H), 7.47 (t, J = 7.15 Hz, 2H),
1
3
7
.56 (d, J = 7.52 Hz, 2H), 8.22 (s, 1H), 8.64 (s, 1H), 10.10 (s, 1H); C NMR (CDCl , 75
3
MHz): δ 51.34, 66.78, 119.04, 126.29, 127.67, 129.03, 129.34, 136.54, 139.09, 150.64,
+
1
60.38, 189.62; ESI-MS: m/z, 269 [M+H] .
6
.3.4. 5-Phenyl-2-thiomorpholinonicotinaldehyde (18d)
1
Yield: 85%; yellow solid; m.p: 104-106 ºC; H NMR (CDCl , 300 MHz): δ 2.80-2.88 (m,
3
4
H), 3.75-3.83 (m, 4H), 7.37 (t, J = 7.17 Hz, 1H), 7.47 (t, J = 7.17 Hz, 2H), 7.56 (d, J = 8.12
1
3
Hz, 2H), 8.22 (s, 1H), 8.64 (s, 1H), 10.06 (s, 1H); C NMR (CDCl , 75 MHz): δ 27.51,
3
5
3.64, 119.24, 126.33, 127.70, 129.06, 129.37, 136.58, 138.60, 150.55, 161.11, 189.61; ESI-
+
MS: m/z, 285 [M+H] .
1
2

ACCEPTED MANUSCRIPT
6
.3.5. 2-Morpholino-5-p-tolylnicotinaldehyde (18e)
1
Yield: 81%; yellow solid; m.p: 114-116 ºC; H NMR (CDCl , 300 MHz): δ 2.39 (s, 3H),
3
3
7
5
1
6
.50 (t, J = 4.36 Hz, 4H), 3.86 (t, J = 4.36 Hz, 4H), 7.27 (d, J = 7.55 Hz, 2H), 7.46 (d, J =
1
3
.74 Hz, 2H), 8.19 (s, 1H), 8.63 (s, 1H), 10.10 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.43,
3
1.80, 67.17, 119.53, 126.52, 129.80, 130.14, 134.01, 137.96, 139.04, 150.89, 160.73,
+
90.11; ESI-MS: m/z, 283 [M+H] .
.3.6. 2-Thiomorpholino-5-p-tolylnicotinaldehyde (18f)
1
Yield: 85%; yellow solid; m.p: 134-136 ºC; H NMR (CDCl , 300 MHz): δ 2.41 (s, 3H),
3
2
8
1
.81-2.89 (m, 4H), 3.75-3.81 (m, 4H), 7.28 (d, J = 6.79 Hz, 2H), 7.45 (d, J = 8.30 Hz, 2H),
1
3
.19 (s, 1H), 8.62 (s, 1H), 10.07 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.08, 27.53, 53.71,
3
14.20, 119.35, 126.18, 129.78, 135.65, 137.63, 138.27, 150.43, 161.09, 189.75; ESI-MS:
+
m/z, 299 [M+H] .
6
.4. General procedure for the synthesis of pyridinylmethanols (9a-d & 19a-f)
Sodium borohydride (4.2 mmol) was added portion wise to a stirred solution of
nicotinaldehyde 8a (3.5 mmol) in methanol (10 mL) at 0 ºC. The reaction mixture was stirred
for 1 h at room temperature. Methanol was removed under reduced pressure; water (5 mL)
was added and extracted with ethyl acetate (2 x 10 mL). The combined organic extracts were
washed with brine, dried over Na SO , and solvent was removed under reduced pressure. The
2
4
residue was purified by column chromatography using silica gel (ethyl acetate/hexane)
afforded (2-chloro-5-methylpyridin-3-yl)methanol 9a as color less solid. Similarly
compounds 9b-d & 19a-f was synthesized from corresponding 8b-d & 18a-f.
6
.4.1. (2-Chloro-5-methylpyridin-3-yl)methanol (9a)
1
Yield: 89%; colorless solid; m.p: 84-86 ºC; H NMR (CDCl , 300 MHz): δ 2.31 (s, 3H),
3
1
3
4
.72 (s, 2H), 7.73 (s, 1H), 8.06 (s, 1H); C NMR (CDCl , 75 MHz): δ 17.64, 60.94, 132.69,
3
+
1
34.48, 137.63, 145.92, 147.74; ESI-MS: m/z, 158 [M+H] .
1
3

ACCEPTED MANUSCRIPT
6
.4.2. (2-Chloro-5-phenylpyridin-3-yl)methanol (9b)
1
Yield: 85%; colorless solid; m.p: 138-140 ºC; H NMR (CDCl , 300 MHz): δ 4.85 (s, 2H),
3
7
8
1
6
.42 (t, J = 7.32 Hz, 1H), 7.48 (t, J = 7.78 Hz, 2H), 7.52 (d, J =7.17 Hz, 2H), 8.10 (s, 1H),
1
3
.52 (s, 1H); C NMR (CDCl , 75 MHz): δ 61.32, 127.02, 128.40, 129.11, 134.48, 135.20,
3
+
36.13, 136.44, 146.04, 147.71; ESI-MS: m/z, 220 [M+H] .
.4.3. (2-Chloro-5-p-tolylpyridin-3-yl)methanol (9c)
1
Yield: 87%; colorless solid; m.p: 103-105 ºC; H NMR (CDCl , 300 MHz): δ 2.32 (t, J =
3
5
8
1
6
.28 Hz, 1H), 2.42 (s, 3H), 4.84 (d, J = 5.28 Hz, 2H), 7.27 (d, J = 7.55 Hz, 2H), 7.47 (d, J =
1
3
.30 Hz, 2H), 8.07 (s, 1H), 8.50 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.16, 61.62, 126.88,
3
+
29.86, 133.53, 134.48, 135.06, 135.19, 136.10, 138.47, 146.12; ESI-MS: m/z, 234 [M+H] .
.4.4. (2-Chloro-5-(4-fluorophenyl)pyridin-3-yl)methanol (9d)
1
Yield: 89%; colorless solid; m.p: 108-110 ºC; H NMR (CDCl , 300 MHz): δ 4.84 (s, 2H),
3
1
3
7
.16 (t, J = 8.54 Hz, 2H), 7.50-7.55 (dd, J = 5.16, 8.69 Hz, 2H), 8.06 (s, 1H), 8.45 (s, 1H); C
NMR (CDCl , 75 MHz): δ 61.32, 116.03, 116.31, 128.71, 128.82, 132.54, 134.48, 135.06,
3
+
1
45.88, 147.74, 161.38, 164.68; ESI-MS: m/z, 238 [M+H] .
6
.4.5. (5-Methyl-2-morpholinopyridin-3-yl)methanol (19a)
1
Yield: 84%; colorless solid; m.p: 102-104 ºC; H NMR (CDCl , 300 MHz): δ 2.28 (s, 3H),
3
3
.11 (t, J = 4.73 Hz, 4H), 3.85 (t, J = 4.73 Hz, 4H), 4.71 (s, 2H), 7.44 (s, 1H), 8.08 (s, 1H);
1
3
C NMR (CDCl , 75 MHz): 17.58, 50.72, 62.01, 67.10, 127.81, 128.84, 137.92, 146.86,
3
+
1
58.04; ESI-MS: m/z, 209 [M+H] .
6
.4.6. (5-Methyl-2-thiomorpholinopyridin-3-yl)methanol (19b)
1
Yield: 86%; colorless solid; m.p: 90-92 ºC; H NMR (CDCl , 300 MHz): δ 2.22 (s, 3H),
3
2
.75 (t, J = 4.53 Hz, 4H), 3.28 (t, J = 4.53 Hz, 4H), 4.63 (s, 2H), 7.41(s, 1H), 8.02 (s, 1H);
1
3
C NMR (CDCl , 75 MHz): δ 17.50, 28.03, 52.76, 61.61, 128.04, 128.79, 137.79, 146.66,
3
+
1
58.94; ESI-MS: m/z, 225 [M+H] .
1
4

ACCEPTED MANUSCRIPT
6
.4.7. (2-Morpholino-5-phenylpyridin-3-yl)methanol (19c)
1
Yield: 84%; colorless solid; m.p: 124-126 ºC; H NMR (CDCl , 300 MHz): δ 3.21 (brs,
3
4
H), 3.88 (brs, 4H), 4.79 (s, 2H), 7.39 (t, J = 6.98 Hz, 1H), 7.46 (t, J = 7.55 Hz, 2H), 7.54 (d,
1
3
J = 7.17 Hz, 2H), 7.84 (s, 1H), 8.49 (s, 1H); C NMR (CDCl , 75 MHz): δ 50.64, 62.18,
3
6
2
6
7.11, 126.72, 127.65, 127.78, 128.98, 132.22, 135.80, 137.43, 145.05, 159.20; ESI-MS: m/z,
+
71 [M+H] .
.4.8. (5-Phenyl-2-thiomorpholinopyridin-3-yl)methanol (19d)
1
Yield: 87%; colorless solid; m.p: 122-124 ºC; H NMR (CDCl , 300 MHz): δ 2.81-2.87
3
(m, 4H), 3.41-3.48 (m, 4H), 4.78 (s, 2H), 7.38 (t, J = 6.79 Hz, 1H), 7.46 (t, J = 7.55 Hz, 2H),
1
3
7
5
2
6
.54 (d, J = 6.79 Hz, 2H), 7.85 (s, 1H), 8.49 (s, 1H); C NMR (CDCl , 75 MHz): δ 28.09,
3
2.83, 62.12, 126.78, 127.71, 128.16, 129.05, 137.48, 144.92, 145.06, 160.18. ESI-MS: m/z,
+
87 [M+H] .
.4.9. (2-Morpholino-5-p-tolylpyridin-3-yl)methanol (19e)
1
Yield: 88%; colorless solid; m.p: 100-102 ºC; H NMR (CDCl , 300 MHz): δ 2.39 (s, 3H),
3
3
7
5
.19 (t, J = 4.57 Hz, 4H), 3.88 (t, J = 4.73 Hz, 4H), 4.79 (s, 2H), 7.25 (d, J = 7.78 Hz, 2H),
1
3
.43 (d, J = 8.08 Hz, 2H), 7.80 (s, 1H), 8.48 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.05,
3
0.71, 62.11, 67.11, 126.52, 127.81, 129.67, 132.17, 134.54, 135.58, 137.49, 144.86, 159.02;
+
ESI-MS: m/z, 285 [M+H] .
6
.4.10. (2-Thiomorpholino-5-p-tolylpyridin-3-yl)methanol (19f)
1
Yield: 86%; colorless solid; m.p: 140-142 ºC; H NMR (CDCl , 300 MHz): δ 2.32 (s, 3H),
3
2
7
2
.84 (t, J = 5.03 Hz, 4H), 3.44 (t, J = 5.03 Hz, 4H), 4.76 (s, 2H), 7.26 (d, J = 8.08 Hz, 2H),
1
3
.43 (d, J = 7.93 Hz, 2H), 7.81 (s, 1H), 8.47 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.15,
3
8.16, 52.86, 62.25, 126.63, 128.12, 129.77, 132.42, 134.57, 135.59, 137.63, 144.92, 160.03;
+
ESI-MS: m/z, 301 [M+H] .
1
5

ACCEPTED MANUSCRIPT
6
.5. General procedure for the synthesis of azidomethylpyridines (10a-d & 20a-f)
DBU (8.4 mmol) was added to a stirred solution of (2-chloro-5-methylpyridin-3-
yl)methanol 9a (5.6 mmol) in dry toluene (15 mL) at room temperature, and followed by
diphenyl phosphoryl azide (6.7 mmol). The reaction mixture was stirred for additional 3 h at
room temperature. Reaction was quenched with aqueous NH Cl solution (8 mL) and
4
extracted with ethyl acetate (2 x 10 mL). The combined organic extracts were washed with
brine, dried over Na SO , and solvent was removed under reduced pressure. The residue was
2
4
purified by column chromatography using silica gel (ethyl acetate/hexane) provided 3-
azidomethyl)-2-chloro-5-methylpyridine 10a as color less solid. Similarly, compounds 10b-
d & 20a-f was synthesized from corresponding 9b-d & 19a-f.
(
6
.5.1. 3-(Azidomethyl)-2-chloro-5-methylpyridine (10a)
1
Yield: 79%; colorless solid; m.p: 120-122 ºC; H NMR (CDCl , 300 MHz): δ 2.35 (s, 3H),
3
1
3
4
1
6
.50 (s, 2H), 7.57 (s, 1H), 8.18 (s, 1H); C NMR (CDCl , 75 MHz): δ 17.53, 51.31, 129.32,
3
+
32.76, 138.68, 14.19, 149.00; ESI-MS: m/z, 183 [M+H] .
.5.2. 3-(Azidomethyl)-2-chloro-5-phenylpyridine (10b)
1
Yield: 82%; colorless solid; m.p: 143-145 ºC; H NMR (CDCl , 300 MHz): δ 4.57 (s, 2H),
3
1
3
7
1
6
.38-7.58 (m, 5H), 7.92 (s, 1H), 8.54 (s, 1H); C NMR (CDCl , 75 MHz): δ 51.20, 126.75,
3
+
28.38, 128.96, 129.83, 135.65, 135.94, 146.70, 148.51, 146.70; ESI-MS: m/z, 245 [M+H] .
.5.3. 3-(Azidomethyl)-2-chloro-5-p-tolylpyridine (10c)
1
Yield: 84%; colorless solid; m.p: 114-116 ºC; H NMR (CDCl , 300 MHz): δ 2.40 (s, 3H),
3
4
.59 (s, 2H), 7.30 (d, J = 8.30 Hz, 2H), 7.47 (d, J = 7.55 Hz, 2H), 7.92 (s, 1H), 8.54 (s, 1H);
1
3
C NMR (CDCl , 75 MHz): δ 21.20, 51.54, 120.25, 125.08, 126.94, 129.66, 130.01, 133.03,
3
+
1
36.23, 138.84, 146.82, 148.39; ESI-MS: m/z, 259 [M+H] .
1
6

ACCEPTED MANUSCRIPT
6
.5.4. 3-(Azidomethyl)-2-chloro-5-(4-fluorophenyl)pyridine (10d)
1
Yield: 87%; colorless solid; m.p: 116-118 ºC; H NMR (CDCl , 300 MHz): δ 4.62 (s, 2H),
3
1
3
7
.19 (t, J = 8.30 Hz, 2H), 7.52-7.59 (dd, J = 5.28, 9.06 Hz, 2H), 7.90 (s, 1H), 8.53 (s, 1H); C
NMR (CDCl , 75 MHz): δ 51.49, 116.18, 116.48, 128.96, 130.21, 135.42, 136.11, 146.90,
3
+
1
61.54, 164.48; ESI-MS: m/z, 263 [M+H] .
6
.5.5. 4-(3-(Azidomethyl)-5-methylpyridin-2-yl)morpholine (20a)
1
Yield: 82%; colorless solid; m.p: 120-122 ºC; H NMR (CDCl , 300 MHz): δ 2.32 (Ss,
3
3
1
1
6
H), 3.08 (t, J = 4.70 Hz, 4H), 3.86 (t, J = 4.70 Hz, 4H), 4.40 (s, 2H), 7.46 (d, J = 1.81 Hz,
1
3
H), 8.14 (d, J = 1.81 Hz, 1H); C NMR (CDCl , 75 MHz): δ 17.45, 50.25, 51.10, 66.87,
3
+
22.98, 128.28, 139.36, 147.43, 158.84; ESI-MS: m/z, 234 [M+H] .
.5.6. 4-(3-(Azidomethyl)-5-methylpyridin-2-yl)thiomorpholine (20b)
1
Yield: 79%; colorless solid; m.p: 110-112 ºC; H NMR (CDCl , 300 MHz): δ 2.28 (s, 3H),
3
1
3
2
.77-2.88 (m, 4H), 3.32-3.56 (m, 4H), 4.37 (s, 2H), 7.46 (s, 1H), 8.14 (s, 1H); C NMR
(
CDCl , 75 MHz): δ 17.55, 27.90, 50.38, 53.20, 129.74, 129.93, 139.05, 147.80, 160.12; ESI-
3
+
MS: m/z, 250 [M+H] .
6
.5.7. 4-(3-(Azidomethyl)-5-phenylpyridin-2-yl)morpholine (20c)
1
Yield: 85%; colorless solid; m.p: 132-134 ºC; H NMR (CDCl , 300 MHz): δ 3.21 (t, J =
3
4
.53 Hz, 4H), 3.87 (t, J = 4.72 Hz, 4H), 4.48 (s, 2H), 7.38 (t, J = 7.17 Hz, 1H), 7.48 (t, J =
1
3
7
.74 Hz, 2H), 7.56 (d, J = 7.17 Hz, 2H), 7.84 (s, 1H), 8.54 (s, 1H); C NMR (CDCl , 75
3
MHz): δ 50.59, 51.01, 66.94, 122.82, 126.68, 127.66, 128.97, 131.72, 137.01, 137.19,
+
1
45.75, 160.02; ESI-MS: m/z, 296 [M+H] .
6
.5.8. 4-(3-(Azidomethyl)-5-phenylpyridin-2-yl)thiomorpholine (20d)
1
Yield: 84%; colorless solid; m.p: 122-124 ºC; H NMR (CDCl , 300 MHz): δ 2.77-2.89
3
(m, 4H), 3.40-3.52 (m, 4H), 4.46 (s, 2H), 7.38 (t, J = 7.17 Hz, 1H), 7.47 (t, J = 7.55 Hz, 2H),
1
3
7
.56 (d, J = 6.98 Hz, 2H), 7.86 (s, 1H), 8.52 (s, 1H); C NMR (CDCl , 75 MHz): δ 27.45,
3
1
7

ACCEPTED MANUSCRIPT
5
0.56, 52.99, 123.36, 126.65, 127.77, 128.97, 131.81, 136.85, 137.42, 145.07, 160.37; ESI-
+
MS: m/z, 312 [M+H] .
6
.5.9. 4-(3-(Azidomethyl)-5-p-tolylpyridin-2-yl)morpholine (20e)
1
Yield: 88%; colorless solid; m.p: 100-102 ºC; H NMR (CDCl , 300 MHz): δ 2.41 (s, 3H),
3
3
7
5
.20 (t, J = 4.51 Hz, 4H), 3.88 (t, J = 4.51 Hz, 4H), 4.47 (s, 2H), 7.27 (d, J = 7.90 Hz, 2H),
1
3
.45 (d, J = 7.90 Hz, 2H), 7.82 (s, 1H), 8.52 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.05,
3
0.61, 51.05, 66.95, 122.85, 126.51, 129.69, 131.75, 134.27, 136.83, 137.56, 145.61, 159.84;
+
ESI-MS: m/z, 310 [M+H] .
6
.5.10. 4-(3-(Azidomethyl)-5-p-tolylpyridin-2-yl)thiomorpholine (20f)
1
Yield: 85%; colorless solid; m.p: 128-130 ºC; H NMR (CDCl , 300 MHz): δ 2.40 (s, 3H),
3
2
8
5
.80-2.86 (m, 4H), 3.42-3.48 (m, 4H), 4.45 (s, 2H), 7.28 (d, J = 7.55 Hz, 2H), 7.48 (d, J =
1
3
.30 Hz, 2H), 7.82 (s, 1H), 8.52 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.06, 27.85, 50.67,
3
3.11, 120.153, 123.16, 126.54, 129.70, 129.96, 136.75, 137.58, 145.64, 160.80; ESI-MS:
+
m/z, 326 [M+H] .
6
.6. Synthesis of (1-((2-chloro-5-methylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl) methanol
11)
Propargyl alcohol (0.75 mmol) was added to a stirred solution of 3-(azidomethyl)-2-
(
chloro-5-methylpyridine 10a (0.83 mmol) in tert-butanol (0.7 mL) and H O (0.7 mL) at room
2
temperature, followed by CuSO ·5H O (0.04 mmol) and sodium ascorbate (0.11 mmol). The
4
2
reaction mixture was stirred for 12-16 h at room temperature and after completion of the
reaction (TLC), the reaction mixture was extracted with ethyl acetate (2 x 5 mL). The organic
extract was washed with H O and dried over anhydrous Na SO . The residue was purified by
2
2
4
column chromatography using silica gel (ethyl acetate/hexane) furnished (1-((2-chloro-5-
methylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl) methanol 11.
1
8

ACCEPTED MANUSCRIPT
6
.6.1. (1-((2-Chloro-5-methylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl) methanol (11)
1
Yield: 82%; colorless solid; m.p.: 120-122 ºC. H NMR (CDCl , 300 MHz): δ 2.29 (s, 3H),
3
1
3
4
.80 (s, 2H), 5.65 (s, 2H), 7.33 (s, 1H), 7.68 (s, 1H), 8.21 (s, 1H). C-NMR (CDCl , 75
3
MHz): 17.48, 49.34, 56.02, 122.50, 126.12, 132.69, 137.63, 145.92, 147.15, 147.74. ESI-MS:
+
m/z, 239 [M+H] .
6
.7. Synthesis of 2-chloro-5-methyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-
triazol-1-yl) methyl)pyridine (13a)
Potassium carbonate (1.5 mmol) and potassium iodide (0.2 mmol) were added to a stirred
solution of (1-((2-chloro-5-methylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl) methanol 11 (1
mmol) in acetone at room temperature and followed by 3,4,5-trimethoxybenzyl bromide (1.2
mmol). The reaction mixture was heated to reflux for 6 h and solvent was removed under
reduced pressure. The residue was dissolved in ethyl acetate (2 x 5 mL) and washed with cold
water. The organic extract was dried over Na SO , and solvent was removed under reduced
2
4
pressure. The residue was subjected to column chromatography purification to provide 2-
chloro-5-methyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-yl)
methyl)pyridine 13a.
6
.7.1.
2-Chloro-5-methyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-yl)
methyl)pyridine (13a)
-1
Yield: 21%; colorless solid; m.p.: 145-147 ºC; IR νmax (cm ): 3136, 2854, 2938, 1592,
1
1
459, 1428, 1332, 1236, 1127, 1078, 1048; H NMR (CDCl , 300 MHz): δ 2.29 (s, 3H), 3.83
3
(s, 3H), 3.85 (s, 6H), 4.54 (s, 2H), 4.69 (s, 2H), 5.62 (s, 2H), 6.58 (s, 2H), 7.35 (s, 1H), 7.67
1
3
(
s, 1H), 8.20 (s, 1H); C NMR (CDCl , 75 MHz): δ 17.51, 50.72, 55.99, 60.72, 63.37, 72.80,
3
1
04.75, 123.00, 128.43, 133.16, 133.34, 137.35, 139.52, 145.53, 147.13, 150.11, 153.17;
+
+
ESI-MS: m/z, 419 [M+H] ; ESI-HRMS: m/z calcd for C H ClN O [M+H] 419.1474,
2
0
23
4
4
found 419.1480.
1
9

ACCEPTED MANUSCRIPT
6
.8. Synthesis of (3,4,5-trimethoxyphenyl)methanol (15)
NaBH (6.1 mmol) was added portion wise to a stirred solution of 3,4,5-trimethoxy
4
benzaldehyde 14 (5.1 mmol) in methanol (10 mL) at 0 ºC. The reaction mixture was stirred
for 1 h at room temperature. Methanol was removed under reduced pressure; water (5 mL)
was added and extracted with ethyl acetate (2 x 10 mL). The organic extract was washed with
brine, dried over Na SO , and solvent was removed under reduced pressure. The residue was
2
4
purified by column chromatograph using silica gel (ethyl acetate/hexane 1:4) afforded 15 as
colorless liquid in 92% yield.
6
.9. Synthesis of 1,2,3-trimethoxy-5-((prop-2-yn-1-yloxy)methyl)benzene (16)
The compound (3,4,5-trimethoxyphenyl)-methanol 15 (4.5 mmol) was dissolved in THF
(15 mL) and added to a stirred suspension of NaH (60%, 5.4 mmol) in THF (5 mL) at 0 ºC.
The reaction mixture was stirred for 30 min at the same temperature. Propargyl bromide (4.9
mmol) was added to the reaction mixture at the same temperature and stirring was continued
for an additional 5 h at room temperature. After completion the reaction (TLC), the reaction
mixture was quenched with ice water and extract with ethyl acetate (2 x 20 mL). The organic
extract was washed with brine, dried over Na SO , and solvent removed under reduced
2
4
pressure. The residue was subjected to silica gel column chromatography purification (ethyl
acetate/hexane 1:10) to afford 1,2,3-trimethoxy-5-((prop-2-yn-1-yloxy)methyl)benzene 16.
6
.9.1. 1,2,3-Trimethoxy-5-((prop-2-yn-1-yloxy)methyl)benzene (16)
1
Yield: 85%; cream brown liquid; H NMR (CDCl , 300 MHz): δ 2.48 (s, J = 2.44 Hz, 1H),
3
1
3
3
.83 (s, 3H), 3.87 (s, 6H), 4.19 (d, J = 2.28 Hz, 2H), 4.54 (s, 2H), 6.59 (s, 2H); C NMR
(
CDCl , 75 MHz): δ 55.97, 57.01, 60.73, 71.65, 74.69, 79.47, 104.85, 132.82, 137.42,
3
+
1
53.16; ESI-MS: m/z, 237 [M+H] .
2
0

ACCEPTED MANUSCRIPT
6
. 10. Synthesis of 3,4,5-trimethoxybenzaldehyde oxime (22)
NaHCO (5.6 mmol) was added portion wise at 0 ºC to a stirred solution of hydroxylamine
3
hydrochloride (3.7 mmol) in water (7 mL) at room temperature and the mixture was stirred
for 30 min. The 3,4,5-trimethoxybenzaldehyde 14 (3.1 mmol) dissolved in methanol (5 mL)
and added to the above reaction mixture and stirring was continued for an additional 6 h.
Methanol was removed under reduced pressure and the residue was extracted with diethyl
ether. The organic extract was washed with brine, dried over Na SO and solvent removed
2
4
under reduced pressure gave 22 as colorless solid in 85% yield.
.11. (3-(3,4,5-Trimethoxyphenyl)isoxazol-5-yl)methanol (23)
N-Chlorosuccinimide (1.3 mmol) and pyridine (2 drops) was added to a stirred solution of
6
oxime 22 (1.3 mmol) in anhydrous CHCl (15 mL) at room temperature. The reaction
3
mixture was stirred for 1 h at 50-60 ºC. Propargyl alcohol (1.4 mmol) was added to the
reaction mixture and followed by triethylamine (1.95 mmol) in CHCl (5 mL). The reaction
3
mixture was stirred at 25 ºC for 2 h and water was added (10 mL). The organic layer was
separated, washed with 2.5% HCl (15 mL) and followed by water (15 mL) and with brine
solution (15 mL). The solvent was removed under reduced pressure and the residue was
purified by column chromatography using silica gel (ethyl acetate/hexane 1:3) afforded (3-
(3,4,5-Trimethoxyphenyl)isoxazol-5-yl)methanol 23.
6
.11.1. (3-(3,4,5-Trimethoxyphenyl)isoxazol-5-yl)methanol (23)
1
Yield: 75%; Colorless liquid; H NMR (CDCl , 300 MHz): δ 3.89 (s, 3H), 3.91 (s, 6H),
3
1
3
4
.81 (s, 2H), 6.50 (s, 1H), 6.99 (s, 2H); C NMR (CDCl , 75 MHz): δ 56.25, 56.60, 60.92,
3
+
9
9.90, 104.09, 124.26, 139.62, 153.56, 162.25, 172.03; ESI-MS: m/z, 266 [M+H] .
2
1

ACCEPTED MANUSCRIPT
6
.12. 5-((Prop-2-ynyloxy)methyl)-3-(3,4,5-trimethoxyphenyl)isoxazole (24)
The compound 24 was prepared according to the method described as per section 6.9 by
employing (3-(3,4,5-trimethoxyphenyl)isoxazol-5-yl)methanol 23 (3.4 mmol) with propargyl
bromide (3.8 mmol).
6
.12.1. 5-((Prop-2-ynyloxy)methyl)-3-(3,4,5-trimethoxyphenyl)isoxazole (24)
1
Yield: 84%; H NMR (CDCl , 300 MHz): δ 2.54 (t, J = 2.54 Hz, 1H), 3.89 (s, 3H), 3.92 (s,
3
13
6
H), 4.28 (d, J = 2.44 Hz, 2H), 4.75 (s, 2H), 6.58 (s, 1H), 7.02 (s, 2H); C NMR (CDCl , 75
3
MHz): δ 56.19, 57.94, 60.89, 62.03, 75.72, 78.39, 101.38, 103.92, 124.15, 139.54, 153.53,
+
1
62.24, 168.98; ESI-MS: m/z, 304 [M+H] .
6
.13. Synthesis of (3-(3,4,5-trimethoxyphenyl)-4,5-dihydroisoxazol-5-yl)methanol (26)
The compound 26 was prepared according to the method described as per section 6.11. by
employing oxime 22 (1.6 mmol) and allyl alcohol (1.6 mmol).
6
.13.1. (3-(3,4,5-trimethoxyphenyl)-4,5-dihydroisoxazol-5-yl)methanol (26)
1
Yield: 80%; H NMR (CDCl , 300 MHz): δ 3.24-3.32 (dd, J = 7.93, 16.42 Hz, 1H),
3
3
.37-3.43 (dd, J = 10.38, 16.42 Hz, 1H), 3.66-3.72 (dd, J = 4.53, 12.27 Hz, 1H), 3.84-3.86
(dd, J = 3.39, 10.95 Hz, 1H), 3.87 (s, 3H), 3.88 (s, 6H), 4.82-4.91 (m, 1H), 5.30 (s, 2H), 6.89
1
3
(
s, 2H); C NMR (CDCl , 75 MHz): δ 39.53, 40.67, 56.00, 62.91, 81.62, 101.49, 131.81,
3
+
1
46.00, 149.93, 151.19, 156.45; ESI-MS: m/z, 268 [M+H] .
6
.14. Synthesis of 5-((prop-2-ynyloxy)methyl)-3-(3,4,5-trimethoxyphenyl)-4,5-dihydro
isoxazole (27)
The compound 27 was prepared according to the method described as per section 6.9 by
employing compound 26 (3.4 mmol) and propargyl bromide (3.8 mmol).
6
.14.1. 5-((prop-2-ynyloxy)methyl)-3-(3,4,5-trimethoxyphenyl)-4,5-dihydro isoxazole (27)
1
Yield: 87%; H NMR (CDCl , 300 MHz): δ 2.46 (t, J = 2.44 Hz, 1H), 3.25-3.30 (dd, J =
3
7
.47, 16.63 Hz, 1H), 3.37-3.43 (dd, J = 10.83, 16.47 Hz, 1H), 3.68-3.70 (dd, J = 5.18, 10.37
2
2

ACCEPTED MANUSCRIPT
Hz, 1H), 3.74-3.77 (dd, J = 5.03, 10.37 Hz, 1H), 3.88 (s, 3H), 3.89 (s, 6H), 4.24 (d, J = 2.28
1
3
Hz, 2H), 4.91-4.97 (m, 1H), 6.91 (s, 2H); C NMR (CDCl , 75 MHz): δ 37.28, 56.13, 58.65,
3
6
0.80, 70.30, 74.93, 79.14, 79.49, 103.92, 124.75, 139.67, 153.16, 156.17; ESI-MS: m/z, 306
+
[
M+H] .
6
.15. General procedure for the synthesis of pyridinyl-1H-1,2,3-triazoles, triazolylisoxazoles,
triazolyldihydroisoxazoles (13a-j, 25a-j and 28a-j)
,2,3-Trimethoxy-5-((prop-2-yn-1-yloxy)methyl)benzene 16 (0.75 mmol) was added to a
1
stirred solution of 3-(azidomethyl)-2-chloro-5-methylpyridine 10a (0.83 mmol) in tert-
butanol (0.7 mL) and H O (0.7 mL) at room temperature, followed by CuSO ·5H O (0.04
2
4
2
mmol) and sodium ascorbate (0.11 mmol). The reaction mixture was stirred for 13 h and
extracted with ethyl acetate (2 x 5 mL), the organic extract was washed with H O, dried over
2
Na SO . The residue was purified by column chromatography using silica gel (ethyl
2
4
acetate/hexane) furnished 2-chloro-5-methyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-
1
,2,3-triazol-1-yl) methyl)pyridine 13a as colorless solid. Yield: 78%. Similarly compounds
3b-j, 25a-j and 28a-j were prepared from the corresponding azides 10b-d, 20a-f and
1
alkynes 16, 24 and 27.
.15.1. 2-Chloro-5-phenyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-
yl)methyl)pyridine (13b)
6
-1
Yield: 78%; colorless solid; m.p.: 141-143 ºC; IR νmax (cm ): 3135, 2938, 2841, 1591,
1
1
4
1
1
1
4
458, 1429, 1235, 1126, 1084, 764; H NMR (CDCl , 300 MHz): δ 3.82 (s, 3H), 3.84 (s, 6H),
3
.53 (s, 2H), 4.69 (s, 2H), 5.72 (s, 2H), 6.57(s, 2H), 7.41-7.47 (m, 5H), 7.72 (s, 2H), 8.60 (s,
1
3
H); C NMR (CDCl , 75 MHz): δ 50.86, 56.03, 60.76, 63.41, 72.80, 104.80, 123.05,
3
27.01, 128.83, 129.08, 129.24, 133.19, 135.42, 136.75, 137.27, 145.71, 148.05, 148.70,
+
+
53.21; ESI-MS: m/z, 481 [M+H] ; ESI-HRMS: m/z calcd for C H ClN O [M+H]
25
26
4
4
81.1630, found 481.1637.
2
3

ACCEPTED MANUSCRIPT
6
.15.2. 2-Chloro-5-(p-tolyl)-3-((4-(((3,4,5-trimethoxybenzyl)oxy)methyl)-1H-1,2,3-triazol-1-
yl)methyl)pyridine (13c)
-1
Yield: 75%; colorless solid; m.p.: 143-145 ºC; IR νmax (cm ): 2930, 2851, 1591, 1459,
1
1
430, 1331, 1227, 1126, 772; H NMR (CDCl , 300 MHz) δ 2.39 (s, 3H), 3.80 (s, 3H), 3.83
3
(s, 6H), 4.52 (s, 2H), 4.69 (s, 2H), 5.72 (s, 2H), 6.58 (s, 2H), 7.25 (d, J = 8.12 Hz, 2H), 7.38
13
(
d, J = 7.91 Hz, 2H), 7.70 (s, 1H), 7.72 (s, 1H), 8.58 (s, 1H); C NMR (CDCl , 75 MHz): δ
3
2
1
1.11, 50.87, 55.99, 60.75, 63.38, 72.77, 104.75, 123.03, 126.79, 128.96, 129.93, 132.44,
+
33.18, 136.65, 137.02, 138.94, 145.65, 147.87, 148.32, 153.18; ESI-MS: m/z, 495 [M+H] ;
+
ESI-HRMS: m/z calcd for C H O N Cl [M+H] 495.1782, found 495.1793.
2
6
28
4
4
6
.15.3. 2-Chloro-5-(4-fluorophenyl)-3-((4-(((3,4,5-trimethoxybenzyl)oxy)methyl)-1H-1,2,3-
triazol-1-yl)methyl)pyridine (13d)
-1
Yield: 79%; colorless solid; m.p.: 123-125 ºC; IR νmax (cm ): 2939, 2841, 1592, 1510,
1
1
459, 1435, 1422, 1231, 1126, 1084, 772; H NMR (CDCl , 300 MHz): δ 3.83 (s, 3H), 3.85
3
(s, 6H), 4.51 (s, 2H), 4.69 (s, 2H), 5.71 (s, 2H), 6.58 (s, 2H), 7.15 (t, J = 8.68 Hz, 2H), 7.42-
1
3
7
7
1
1
4
6
.49 (dd, J = 5.09, 8.68 Hz, 2H), 7.68 (s, 1H), 7.73 (s, 1H), 8.55 (s, 1H); C NMR (CDCl ,
3
5 MHz): δ 50.67, 55.88, 60.61, 63.28, 72.67, 104.67, 116.02, 116.30, 123.74, 128.06,
28.63, 129.08, 131.42, 133.09, 135.59, 137.00, 145.52, 147.66, 148.60, 153.08, 161.38,
+
+
64.68; ESI-MS: m/z, 499 [M+H] ; ESI-HRMS: m/z calcd for C H O N ClF [M+H]
2
5
25
4
4
99.1536, found 499.1542.
.15.4. 4-(5-Methyl-3-((4-(((3,4,5-trimethoxybenzyl)oxy)methyl)-1H-1,2,3-triazol-1-yl)
methyl)pyridin-2-yl)morpholine (13e)
-1
Yield: 81%; colorless solid; m.p.: 109-111 ºC; IR νmax (cm ): 3473, 3135, 2957, 2927,
1
2
3
7
853, 1592, 1506, 1457, 1331, 1238, 1123, 1048; H NMR (CDCl , 300 MHz): δ 2.24 (s,
3
H), 3.04 (m, 4H), 3.79-3.92 (m, 13H), 4.54 (s, 2H), 4.69 (s, 2H), 5.60 (s, 2H), 6.59 (s, 2H),
1
3
.15 (s, 1H), 7.56 (s, 1H), 8.15 (s, 1H); C NMR (CDCl , 75 MHz): δ 17.57, 49.34, 51.31,
3
2
4

ACCEPTED MANUSCRIPT
5
6.03, 60.74, 63.47, 66.97, 72.72, 104.48, 114.24, 122.76, 122.93, 129.11, 133.32, 138.36,
+
1
45.48, 148.45, 153.22, 158.75; ESI-MS: m/z, 470 [M+H] ; ESI-HRMS: m/z calcd for
+
C H O N [M+H] 470.2381, found 470.2398.
24
32
5
5
6
.15.5. 4-(5-Methyl-3-((4-(((3,4,5-trimethoxybenzyl)oxy)methyl)-1H-1,2,3-triazol-1-yl)
methyl)pyridin-2-yl)thiomorpholine (13f)
-1
Yield: 76%; colorless solid; m.p.: 131-133 ºC; IR νmax (cm ): 2924, 2845, 1591, 1456,
1
1
3
2
4
1
456, 1421, 1232, 1126, 773; H NMR (CDCl , 300 MHz): δ 2.21 (s, 3H), 2.79-2.81 (m, 4H),
3
.27-3.31 (m, 4H), 3.82 (s, 3H), 3.85 (s, 6H), 4.53 (s, 2H), 4.69 (s, 2H), 5.54 (s, 2H), 6.58 (s,
3
H), 7.13 (s, 1H), 7.54 (s, 1H), 8.14 (s, 1H); C NMR (CDCl , 75 MHz): δ 17.44, 27.82,
3
9.21, 53.21, 55.87, 60.60, 63.34, 72,61, 104.64, 122.69, 129.03, 133.11, 137.22, 138.22,
+
45.32, 148.27, 153.03, 159.58; ESI-MS: m/z, 486 [M+H] ; ESI-HRMS: m/z calcd for
+
C H O N S [M+H] 486.2153, found 486.2169.
24
32
4
5
6
.15.6. 4-(5-Phenyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-yl)methyl)
pyridin-2-yl)morpholine (13g)
-1
Yield: 73%; colorless solid; m.p.: 101-103 ºC; IR νmax (cm ): 2938, 2851, 1592, 1453,
1
1
238, 1123, 768; H NMR (CDCl , 300 MHz): δ 3.16 (t, J = 4.57 Hz, 4H), 3.82 (s, 9H), 3.88
3
(t, J = 4.57 Hz, 4H), 4.51 (s, 2H), 4.68 (s, 2H), 5.66 (s, 2H), 6.55 (s, 2H), 7.35-745 (m, 5H),
1
3
7
6
1
.48 (s, 1H), 7.56 (s, 1H), 8.55 (s, 1H); C NMR (CDCl , 75 MHz): δ 49.37, 51.02, 55.85,
3
0.59, 63.33, 66.75, 72.54, 104.61, 122.55, 122.74, 126.49, 127.72, 128.87, 132.25, 133.11,
+
35.96, 136.62, 145.51, 146.20, 153.03, 159.55; ESI-MS: m/z, 532 [M+H] ; ESI-HRMS: m/z
+
calcd for C H O N [M+H] 532.2541, found 532.2554.
2
9
34
5
5
6
.15.7. 4-(5-Phenyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-yl)methyl)
pyridin-2-yl)thiomorpholine (13h)
-1
Yield: 77%; colorless solid; m.p.: 131-133 ºC; IR νmax (cm ): 3135, 2931, 2841, 1593,
1
1
505, 1457, 1421, 1370, 1331, 1234, 1126, 819, 771; H NMR (CDCl , 300 MHz): δ 2.82-
3
2
5

ACCEPTED MANUSCRIPT
2
2
.85 (m, 4H), 3.39-3.43 (m, 4H), 3.82 (s, 9H), 4.51 (s, 2H), 4.68 (s, 2H), 5.62 (s, 2H), 6.55 (s,
1
3
H), 7.35-744 (m, 5H), 7.46 (s, 1H), 7.55 (s, 1H), 8.55 (s, 1H); C NMR (CDCl , 75 MHz):
3
δ 27.84, 49.53, 53.24, 56.01, 60.71, 63.49, 72.69, 104.85, 122.72, 122.85, 126.63, 127.85,
28.98, 132.48, 133.20, 136.13, 136.74, 137.51, 145.68, 146.36, 153.19, 160.66; ESI-MS:
1
+
+
m/z, 548 [M+H] ; ESI-HRMS: m/z calcd for C H N O S [M+H] , 549.1543, found
29
33
5
4
5
49.1549.
.15.8. 4-(5-p-Tolyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-yl)methyl)
6
pyridin-2-yl)morpholine (13i)
-1
Yield: 79%; colorless solid; m.p.: 116-118 ºC; IR νmax (cm ): 3136, 3006, 2959, 2852,
1
1
3
2
1
6
1
593, 1505, 1454, 1365, 1331, 1239, 1122, 819, 755; H NMR (CDCl , 300 MHz): δ 2.37 (s,
3
H), 3.14 (t, J = 4.57 Hz, 4H), 3.82 (s, 9H), 3.87 (t, J = 4.73 Hz, 4H), 4.52 (s, 2H), 4.67 (s,
H), 5.65 (s, 2H), 6.55 (s, 2H), 7.20 (d, J = 7.92 Hz, 2H), 7.33 (d, J = 8.08 Hz, 2H), 7.47 (s,
1
3
H), 7.55 (s, 1H), 8.54 (s, 1H); C NMR (CDCl , 75 MHz): δ 21.01, 49.52, 51.19, 56.00,
3
0.73, 63.46, 66.90, 72.68, 104.81, 122.65, 122.76, 126.46, 129.71, 132.45, 133.30, 133.81,
+
35.92, 137.45, 137.82, 145.66, 146.24, 153.18, 159.50; ESI-MS: m/z, 546 [M+H] ; ESI-
+
HRMS: m/z calcd for C H O N [M+H] 546.2703, found 546.2712.
3
0
36
5
5
6
.15.9. 4-(5-p-Tolyl-3-((4-((3,4,5-trimethoxybenzyloxy)methyl)-1H-1,2,3-triazol-1-yl)
methyl)pyridin-2-yl)thiomorpholine (13j)
-1
Yield: 76%; colorless solid; m.p: 135-137 ºC; IR νmax (cm ): 2923, 2851, 1592, 1505,
1
1
2
2
1
1
1
505, 1453, 1370, 1331, 1219, 1126, 772; H NMR (CDCl , 300 MHz): δ 2.37 (s, 3H), 2.81-
3
.85 (m, 4H), 3.38-3.42 (m, 4H), 3.82 (s, 9H), 4.51 (s, 2H), 4.68 (s, 2H), 5.61 (s, 2H), 6.55 (s,
H), 7.20 (d, J = 7.93 Hz, 2H), 7.33 (d, J = 8.12 Hz, 2H), 7.45 (s, 1H), 7.54 (s, 1H), 8.52 (s,
1
3
H); C NMR (CDCl , 75 MHz): δ 21.05, 27.89, 49.55, 53.29, 56.03, 60.76, 63.51, 72.72,
3
04.83, 122.73, 122.91, 126.58, 129.74, 132.52, 133.24, 133.81, 135.99, 137.48, 137.87,
+
45.71, 146.18, 153.21, 160.44; ESI-MS: m/z, 562 [M+H] .
2
6

ACCEPTED MANUSCRIPT
6
.15.10. 5-(((1-((2-Chloro-5-methylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)
methyl)-3-(3,4,5-trimethoxyphenyl)isoxazole (25a)
-1
Yield: 83%; colorless solid; m.p.: 80-82 ºC; IR νmax (cm ): 3132, 2935, 1585, 1466, 1424,
1
1
4
8
1
1
238, 1127, 1078; H NMR (CDCl , 300 MHz): δ 2.29 (s, 3H), 3.89 (s, 3H), 3.91 (s, 6H),
3
.73 (s, 2H), 4.80 (s, 2H), 5.62 (s, 2H), 6.60 (s, 1H), 7.00 (s, 2H), 7.35 (s, 1H), 7.73 (s, 1H),
1
3
.21(s, 1H); C NMR (CDCl , 75 MHz): δ 17.46, 50.75, 56.15, 60.80, 62.87, 63.95, 101.42,
3
03.93, 123.28, 124.05, 128.31, 133.31, 139.44, 139.53, 144.57, 147.12, 150.09, 153.46,
+
62.15, 169.08; ESI-MS: m/z, 486 [M+H] ; ESI-HRMS: m/z, calcd for C H ClN O
23
25
5
5
+
[
M+H] 486.1525, found 486.1538.
6
.15.11. 5-(((1-((2-Chloro-5-phenylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)
methyl)-3-(3,4,5-trimethoxyphenyl)isoxazole (25b)
-1
Yield: 85%; colorless solid; m.p.: 96-98 ºC; IR νmax (cm ): 2933, 2855, 1571, 1508, 1454,
1
1
4
1
1
1
431, 1371, 1237, 1127, 1084, 766; H NMR (CDCl , 300 MHz): δ 3.77 (s, 9H), 4.60 (s, 2H),
3
.64 (s, 2H), 5.58 (s, 2H), 6.48 (s, 1H), 6.87 (s, 2H), 7.19-7.39 (m, 5H), 7.58 (s, 1H), 7.71 (s,
1
3
H), 8.44 (s, 1H); C NMR (CDCl , 75 MHz): δ 50.68, 55.95, 60.64, 62.67, 63.75, 101.29,
3
03.69, 123.37, 123.90, 126.74, 128.58, 128.85, 129.00, 129.42, 135.47, 136.39, 137.00,
+
39.27, 144.41, 147.71, 148.49, 153.28, 161.98, 168.95; ESI-MS: m/z, 548 [M+H] ; ESI-
+
HRMS: m/z, calcd for C H ClN O [M+H] 548.1690, found 548.1695.
2
8
27
5
5
6
.15.12. 5-(((1-((2-Chloro-5-p-tolylpyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)
methyl)-3-(3,4,5-trimethoxyphenyl)isoxazole (25c)
-1
Yield: 83%; colorless solid; m.p.: 106-108 ºC; IR νmax (cm ): 3133, 3005, 2937, 1586,
1
1
3
7
467, 1424, 1238, 1127, 818, 756; H NMR (CDCl , 300 MHz): δ 2.36 (s, 3H), 3.89 (s, 3H),
3
.91 (s, 6H), 4.72 (s, 2H), 4.77 (s, 2H), 5.71 (s, 2H), 6.58 (s, 1H), 7.01 (s, 2H), 7.25 (d, J =
1
3
.78 Hz, 2H), 7.38 (d, J = 7.78 Hz, 2H), 7.70 (s, 1H), 7.76 (s, 1H), 8.57 (s, 1H); C NMR
(
CDCl , 75 MHz): δ 21.07, 50.91, 56.16, 60.84, 62.89, 63.96, 101.43, 103.89, 123.33,
3
2
7

ACCEPTED MANUSCRIPT
1
1
24.09, 126.76, 128.84, 129.09, 132.40, 136.60, 136.97,138.92, 139.49, 144.68, 147.85,
+
48.32, 153.48, 162.19, 169.10; ESI-MS: m/z, 562 [M+H] ; ESI-HRMS: m/z, calcd for
+
C H ClN O [M+H] 562.1845, found 562.1850.
29
29
5
5
6
.15.13. 5-(((1-((2-Chloro-5-(4-fluorophenyl)pyridin-3-yl)methyl)-1H-1,2,3-triazol-4-yl)
methoxy)methyl)-3-(3,4,5-trimethoxyphenyl)isoxazole (25d)
-1
Yield: 85%; colorless solid; m.p.: 111-113 ºC; IR νmax (cm ): 3516, 3138, 3007, 2940,
1
1
3
7
601, 1572, 1512, 1436, 1416, 1372, 1236, 1128, 1085, 838; H NMR (CDCl , 300 MHz): δ
3
.86 (s, 3H), 3.87 (s, 6H), 4.69 (s, 2H), 4.75 (s, 2H), 5.68 (s, 2H), 6.50 (s, 1H), 6.96 (s, 2H),
1
3
.11 (s, 2H), 7.41-7.43 (s, 2H), 7.64 (s, 1H), 7.75 (s, 1H), 8.51 (s, 1H); C NMR (CDCl , 75
3
MHz): δ 50.83, 56.16, 60.83, 62.91, 63.99, 101.44, 103.93, 116.18, 116.35, 123.36, 124.05,
1
1
28.73, 128.80, 129.01, 131.52, 135.71, 137.05, 139.56, 144.72, 147.82, 148.70, 153.49,
+
62.19, 164.13, 169.05; ESI-MS: m/z, 566 [M+H] ; ESI-HRMS: m/z, calcd for
+
C H ClFN O [M+H] 566.1602, found 566.1601.
28
26
5
5
6
1
.15.14. 4-(5-Methyl-3-((4-(((3-(3,4,5-trimethoxyphenyl)isoxazol-5-yl)methoxy)methyl)-1H-
,2,3-triazol-1-yl)methyl)pyridin-2-yl)morpholine (25e)
-1
Yield: 87%; colorless solid; m.p.: 118-120 ºC; IR νmax (cm ): 2929, 2852, 1570, 1454,
1
1
4
2
414, 1370, 1238, 1124, 770; H NMR (CDCl , 300 MHz): δ 2.22 (s, 3H), 3.03-3.06 (t, J =
3
.53 Hz, 4H), 3.83- 3.36 (t, J = 4.72 Hz, 4H), 3.89 (s, 3H), 3.91 (s, 6H), 4.70 (s, H), 4.77 (s,
1
3
H), 5.58 (s, 2H), 6.58 (s, 1H), 7.01 (s, 2H), 7.13 (s, 1H), 7.58 (s, 1H), 8.15 (s, 1H); C
NMR (CDCl , 75 MHz): δ 17.53, 49.34, 51.27, 56.14, 60.83, 62.80, 64.02, 66.91, 101.45,
3
1
1
5
03.86, 122.51, 123.05, 124.05, 129.10, 138.25, 139.49, 144.56, 148.49, 153.48, 158.69,
+
+
62.19, 169.08; ESI-MS: m/z, 537 [M+H] ; ESI-HRMS: m/z, calcd for C H N O [M+H]
2
7
33
6
6
37.2449, found 537.2456.
2
8