Synthesis and SAR of new isoxazole-triazole bis-heterocyclic compounds as analogues of natural lignans with antiparasitic activity

Synthesis and SAR of new isoxazole-triazole bis-heterocyclic compounds as

analogues of natural lignans with antiparasitic activity

†

⁑

Lara A. Zimmermann , Milene H. de Moraes , Rafael da Rosa , Eduardo B. de Melo ,

†

#

‖

†

⁑

†;*

Fávero R. Paula , Eloir P. Schenkel , Mario Steindel and Lílian S. C. Bernardes .

†

Department of Pharmaceutical Sciences, Federal University of Santa Catarina, 88040-900,

Florianópolis, 88040-900, SC, Brazil.

⁑

Department of Microbiology, Immunology and Parasitology, Federal University of Santa

Catarina, Florianópolis, 88040-900, SC, Brazil.

#

Department of Pharmacy, Western Paraná State University, Cascavel, 858119-110, PR,

Brazil.

Course of Pharmacy, Federal University of Pampa, Uruguaiana, 97500-970, RS, Brazil.

Corresponding author: l.bernardes@ufsc.br

‖

*

ABSTRACT Despite the impressive scientific and technological advances of recent decades, no

effective treatment is currently available for Chagas disease. Our research group has been

studying the design and synthesis of analogues of natural lignans aiming to identify compounds

with antiparasitic activity. This article reports the synthesis of 42 novel bis-heterocyclic

derivatives and the structure-activity relationship study conducted based on results of biological

assays against Trypanosoma cruzi amastigotes. Thirty-seven compounds were active, and

eight of them had GI₅₀values lower than 100 μM (GI₅₀88.4–12.2 M). A qualitative structure

activity relationship study using three dimensional descriptors was carried out and showed a

correlation between growth inhibitory potency and the presence of bulky hydrophobic groups

located at rings A and D of the compounds. Compound 3-(3,4-dimethoxyphenyl)-5-((4-(4-

pentylphenyl)-1H-1,2,3-triazol-1-yl)methyl)isoxazole (31) was the most active in the series (GI₅₀

12.2 μM), showing, in vitro, low toxicity and potency similar to benznidazole (GI₅₀10.2 μM).

These results suggest that this compound can be a promising scaffold for the design of new

trypanocidal compounds.

KEYWORDS isoxazole, triazole, anti-trypanosomatid agents, Trypanosoma cruzi, Leishmania

amazonensis.

ABBREVIATIONS %GI: percentage of parasite growth inhibition at 100 μM; GI : 50% growth

50

inhibition concentration; Bnz: benznidazole; MW: microwave; NA: not active at the tested

concentration; ND: not determined due to absence of trypanocidal activity; SI: selectivity index

(

CC₅₀

THP-1

cells/GI₅₀);

(r)TR:

(recombinant)

trypanothione

reductase.

S1

GRAPHICAL ABSTRACT

HIGHLIGHTS

42 bis-heterocyclic analogues of natural lignans were synthesized.

All compounds had their trypanocidal and leishmanicidal activities evaluated.

The most active compounds were evaluated against (r)TR.

SAR showed a correlation between growth inhibition and the presence of bulky hydrophobic

groups.

S2

1.

INTRODUCTION

Chagas disease and leishmaniasis caused by protozoan parasites Trypanosoma

cruzi and Leishmania spp., respectively, are tropical and subtropical diseases that

affect millions of people worldwide and are major causes of mortality and morbidity,

1,2

especially in Latin America . Current treatments have major drawbacks, such as

variable efficacy, long therapy duration, prolonged hospitalization, high costs, and the

potential development of drug resistance. Thus, there is an urgent need to discover

3–5

new drugs candidate for the treatment of these parasitic diseases

.

The study of natural products is a promising strategy in drug development for

obtaining lead compounds. Of all drugs approved between 1981 and 2014 for the

6

treatment of human diseases, about 56% are related to natural products

.

Furthermore, the search for plant-derived compounds with high antiparasitic activity

has received much attention because of the structural diversity of these secondary

7,8

metabolites

.

Our research group has been studying the design and synthesis of analogues of

the trypanocidal natural lignans veraguensin (1) and grandisin (2) using different

strategies of molecular modification, aiming to identify hit compounds with appropriate

9–11

chemical and biological properties

. Recently, we have described the synthesis of a

series of derivatives containing the isoxazole ring as a bioisosteric replacement of the

9

central tetrahydrofuran ring of natural lignans 1 and 2 . As a continuation of our

9

studies, considering promising molecules previously identified (4-6) , we synthesized a

series of bis-heterocyclic derivatives containing the isoxazole moiety and a triazole ring

as a spacer group between the aromatic units (Figure 1). These heterocyclic systems

are found in several drugs and show interesting properties because of their chemical

12–14

characteristics, biological and pharmacological applications

. The five-membered

heterocyclic ring triazole was selected because of its ability to act as both hydrogen

bond acceptor (HBA) and donor (HBD), structure rigidity and stability under oxidative

15

and reductive conditions . These aspects combined contribute to ongoing SAR

studies on trypanocidal lignan derivatives and for designing chemically more complex

compounds without considerable negative effects on druglikeness.

In this work, compounds were planned by modifying rings A and D of the

proposed scaffold (7) using the different substitution patterns seen in natural

derivatives (lignans 1–3), which contain methoxy and methylenedioxy groups, and by

varying the substituent groups on ring D regarding their electronic and structural

diversity. Presenting the synthesis of 42 new compounds, this article describes the

structure-activity relationship (SAR) studies that were carried out based on the results

of biological assays against T. cruzi amastigotes. Additionally, the most promising

synthesized compounds were evaluated against Trypanothione reductase enzyme,

considering that it is a trypanosomatid-specific target involved in the redox metabolism

16–21

of the parasite and is essential for its survival

.

S3

Figure 1. Natural neolignans veraguensin (1), grandisin (2), and 6,6'-((2R,3R,4R,5R)-3,4-

dimethyltetrahydrofuran-2,5-diyl)bis(4-methoxybenzo[d][1,3]dioxole) (3), 3,5-disubstituted

isoxazoles (4-6) and scaffold of the proposed bis-heterocyclic derivatives (7). TC_T_r_y_p_o: T. cruzi

trypomastigotes; TC_A_m_a: T. cruzi amastigotes.

2.

RESULTS AND DISCUSSION

2.1. SYNTHESIS

The synthetic route was designed to be economically feasible and reproducible

and to allow the rapid preparation of a library of structurally diverse compounds in good

yields (Scheme 1). Initially, an aldoxime (b) was synthesized in 98% yield by

microwave irradiation of a commercial aldehyde (a) and hydroxylamine hydrochloride

22

.

Subsequently, this aldoxime was reacted with N-chlorosuccinimide, generating in

situ the corresponding arylcarboximidoyl chloride, which was reacted with propargyl

alcohol in the presence of copper sulfate to form a 3,5-disubstituted isoxazole

23

derivative (d) . This compound was tosylated and reacted with sodium azide under

2

4,25

microwave irradiation, giving intermediate azide f in 85% yield

. Finally, the bis-

heterocyclic compounds (h) were obtained by copper(I)-catalyzed azide–alkyne

cycloadditions (CuAAC) under microwave irradiation in yields ranging from 38 to 99%.

1

13

The compounds 7–48 were characterized by H and C NMR and mass spectrometry.

Spectral data are available in the Supplementary Material. The compounds were

evaluated for drug-likeness using Lipinski’s Rule of Five, and most of them had the

properties necessary for good oral bioavailability (Table S1, Supplementary Material)

26,27

.

Scheme 1. Synthetic route of bis-heterocyclic compounds. (i) NH OH.HCl, DABCO, MW:

2

7

0 ºC, 100 W, 1 min (60-98%); (ii) NCS, DMF, sodium ascorbate, CuSO , NaHCO , t-

4 3

BuOH:H O, propargyl alcohol, r.t, 4 h (82%) or NCS, DMF, sodium ascorbate, CuSO4,

2

NaHCO ,DMF,propargyl alcohol, MW: 35 ºC, 150 W, 11 min (54-76%); (iii) TsCl, Et N,

3

K CO ,CH Cl :H O, 1 h (49-76%); (iv) NaN , DMF, MW: 70 ºC, 150 W, 10 min (89-80%); (v)

2

3

2

3

CuSO , sodium ascorbate, alkyne, DMF, 70 ºC, 100 W, 10 min (38-89%).

4

S4

2.2. BIOLOGIC INVESTIGATION AND STRUCTURE-ACTIVITY RELATIONSHIP STUDY

Compounds were tested in vitro on THP-1 cells (human monocytic leukaemia cell

line) infected with T. cruzi amastigotes. Cytotoxicity assays were performed on THP-1

cells. These results are shown in Tables 1-3. All compounds were additionally

evaluated against amastigotes of Leishmania amazonensis; however, no compound

showed promising activity at 100 μM (Table S2, Supplementary Material).

Table 1. Inhibitory activity of the first series of compounds at 100 μM against

Trypanosoma cruzi amastigotes.

Yield

b

Compound

R₃

a

%GI

GI (M)

CC (M)

SI

50

(

%)

76

38

88

76

84

49

92

88

99

89

77

68

87

47

42

97

7

8

9

H

33.7 ± 0.8

43.3 ± 6.3

12.4 ± 1.7

95.3 ± 0.7

45.2 ± 4.4

89.5 ± 0.9

18.6 ± 2.5

0.0 ± 0.0

43.1 ± 2.5

46.9 ± 4.5

28.7 ± 2.4

0.00 ± 0.0

32.6 ± 6.9

0.0 ± 0.0

>100

ND

-

2.6

-

2.3

-

4-OCH

2-OCH

3

1

2

0

1

2

3

4

5

6

7

8

9

0

1

2

3,5-OCH

4-CH

2,4,5-CH

2-CH ,4-OCH

2,5-CH

C(OCH

4-NO

3

54.5 ± 2.1

>100

88.4  4.8

>100

142.8 ±10.9

ND

200.5 60.0

ND

3

d

NA

ND

3

3-C H

2

)CH-4

>100

2

3

2

4-OCF

4-F

3

d

NA

4-N(CH

4-NH

4-CH OH

4-(CH CH

)

3

>100

2

d

NA_d

2

0.0 ± 0.0

NA

2

d

)

0.0 ± 0.0_c

ND

>500

2

4

3

B_anz

93.0 ± 0.6

10.2  0.1

>49.02

b

Yield of the CuAAC reaction. Results are expressed as mean ± SD of an experiment

performed in triplicate. Bnz: percentage of growth inhibition of T. cruzi amastigotes at 20 μM.

c

d

NA, not active at the tested concentration. SI: selectivity index = CC₅₀THP-1/GI₅₀_.

The first series of molecules synthesized had an unsubstituted A ring and

variations of substituent groups on ring D. As compound 7 had the most simplified

scaffold of the bis-heterocyclic derivatives, we considered its percentage of parasite

growth inhibition (33.7%) to compare and discuss the SAR data of other compounds.

Biological data suggest that the substitution of hydrogens for methoxy (OMe) groups

affects the inhibitory activity according to the position of the group. The presence of 4-

OMe contributed to a slight increase in activity (compound 8, GI 43.3%), whereas the

presence of 2-OMe led to a decrease in activity (compound 9, GI 12.4%). The

presence of 3,5-OMe (compound 10) led to an increase in activity (GI 95.3%, GI₅₀54.5

μM, SI 2.6). In addition to the influence of the position of the methoxy groups, we

investigated the trypanocidal activity of derivatives with a substituted methyl group on

ring D. The addition of a 4-methyl group also led to a slight increase in activity

(

compound 11, GI 45.2%), similar to the observed for the 4-OMe derivative (8).

Compound 12, which received a 2,4,5-trimethyl substitution, had an increase in activity

GI 89.5%, GI₅₀88.4 μM, SI 2.3). The combination of a methyl group at the 2-position

(

and a methoxy group at the 4-position (compound 13) decreased activity (GI 18.6%).

S5

These results demonstrate the contribution of methoxy and methyl groups to

compound activity, especially for derivatives with rings substituted at the 3,5 or 4,5

positions; these groups probably occupy and act on a hydrophobic region of a putative

molecular target. We also observed a small decrease in activity for derivatives with a 2-

methyl group, which may be related to the conformational restriction caused by the

presence of ortho substituents in bis-aromatic compounds, altering their conformation

and, consequently, influencing how they bind to a target. However, the presence of a

3,4-dimethyl substituent (compound 12) contributed to inhibitory activity. The activity of

compounds 9, 13, and, 14 suggests that a 2-substituent may decrease activity, but the

combination of hydrophobic substituents at positions 3,5 or 4,5 has a positive effect on

inhibitory activity. Furthermore, the 6-methoxy-β-naphthalene group, present in

compound 15 (GI 43.1%), also seemed to be unfavorable for biological activity;

because of its rigidity, this group might prevent the molecule from adopting an

appropriate conformation to interact with a molecular target, even though it can form

hydrophobic interactions. By analyzing the effects of electron-donor or electron-

withdrawing groups at the para position of ring D, we observed that, for compounds

16–18, the increase in biological activity was directly proportional to the increase in the

electron-withdrawing capacity of the substituent. Most compounds containing electron-

donor groups were able to inhibit parasite growth (8, 11 and 19), with the exception of

those that have ortho substituents (9; 14) and compounds 20 (4-NH ) and 21 (4-

2

CH OH), which have polar substituents that would hinder possible hydrophobic

2

interactions.

Based on the chemical structure of the most active compounds of the first series

(

10 and 12) and considering that the different active natural lignans (1–3) have

28,29

methoxy or methylenedioxy groups as substituents on ring A

, a second series of

molecules was synthesized and evaluated against amastigotes of T. cruzi (23–28)

Table 2).

(

Table 2. Inhibitory activity of the second series of compounds at 100 μM against

Trypanosoma cruzi amastigotes.

a

b

Compound

R₁

R₂

H

OCH

O

R₃

Yield (%)

%GI

GI (M)

CC (M)

SI

50

2

3

4

5

6

7

8

OCH

3

2,4,5-CH

3

89

82

94

96

88

60

33.4  2.8

3.6  0.7

89.6  0.3

74.8  3.7

13.4 ± 0.4

87.9 ± 1.1_c

93.0 ± 0.6

>100

ND

> 500

ND

-

3,5-OCH

2,4,5-CH

3,5-OCH

2,4,5-CH

3,5-OCH

3

> 9.0

> 16.5

-

4.2

>49.0

3

55.2  5.0

30.3  4.3

>100

49.9  8.9

10.2  0.1

3

OCH

2

3

2

3

207.6 9.3

>500

Bn_az

b

Yield of the CuAAC reaction. Results are expressed as mean ± SD of an experiment

performed in triplicate. Bnz: percentage of growth inhibition of T. cruzi amastigotes at 20 μM.

c

SI: selectivity index =CC₅₀THP-1/GI₅₀

Regarding analogues containing the 2,4,5-trimethylphenyl ring, we observed that

the 3,4-OMe substitution on ring A led to an increase both in trypanocidal potency and

in selectivity. Similar results were obtained with derivatives containing the 3,5-

S6

dimethoxyphenyl ring. Of these, compounds 26 and 28 were more potent and selective

than the compound 10. These results suggest that the presence of substituents at

positions meta and para of ring A are important for biological activity.

On the basis of these observations, a third series of compounds (29–42) was

synthesized (Table 3). In the new series, the 3,4-dimethoxy substitution pattern on ring

A was maintained and the effect of modifying substituent groups on ring D was

evaluated. Comparison of compounds 29, 30, and 31, which have the 4-OCH , 4-OCF ,

3

and 4-(CH ) CH substituents on the D ring, respectively, to their analogues (8, 17, and

2

4

3

22) (Table 1) showed that the insertion of a dimethoxylated substituent on ring A

favored trypanocidal activity. However, no significant change in activity was observed

for analogues 43–48 (Table 3), which contained the 4-methoxy or 3,4-methylenedioxy

substituents on ring A, corroborating the idea that, for the bis-heterocyclic derivatives

presented herein, the presence of the 3,4-dimethoxyphenyl A ring is important for

biological activity.

Table 3. Inhibitory activity of the third series of compounds at 100 μM against

Trypanosoma cruzi amastigotes

a

b

Compound

R

1

R

2

R

3

Yield (%)

%GI

GI (M)

CC (M)

SI

5

0

50

2

3

4

9

0

1

2

3

4

5

6

7

8

9

0

1

2

3

4

5

6

7

8

OCH

3

OCH

H

3

4-OCH

4-OCF

3

62

89

94

50

87

92

93

95

82

97

78

88

65

92

89

85

94

99

96

91

81.9  2.0

78.5  1.8

80.2  0.2

42.8  1.1

43.4  3.3

12.2  3.3

7.6  0.3

43.7  3.8

13.6  1.2

4.0  0.7

37.8  4.4

32.0  1.6

33.9  3.5

26.2  0.8

43.2  3.6

9.9 ± 0.0

22.7  3.5

40.1  3.5

12.2  1.9

>100

> 500

227.2  56.3

> 500

ND

> 22.0

5.7

3

4-(CH

4-NH

)

CH

> 41.0

2

4

3

-

2

4-CH

OH

2

>100

ND

-

H

4-CH

2-OCH

2-CH ,4-OCH

2,5-CH

>100

ND

-

3

>100

ND

-

3

>100

ND

-

3

>100

ND

-

3

4-NO

4-F

>100

ND

-

2

>100

ND

-

4-N(CH

C(OCH

)

>100

ND

-

3

2

3-C

H

)CH-4

>100

ND

-

2

3

OCH

>100

ND

-

3

OCH

O

>100

ND

-

2

3

OCH

H

OCF

26.0  2.8

26.8 ± 1.4

13.4  0.4

48.9 ± 0.9

>100

ND

-

3

OCH

2

3

OCH

(CH

)

CH

>100

ND

-

3

2

4

3

OCH

>100

ND

4

c

Bnz

93.0 ± 0.6

10.2  0.1

>500

>49.0

a

b

Yield of the CuAAC reaction. Results are expressed as mean ± SD of an experiment

c

performed in triplicate. Bnz: percentage of growth inhibition of T. cruzi amastigotes at 20 μM.

SI: selectivity index =CC₅₀THP-1/GI₅₀

Compounds 29, 30, and 31 showed similar percentages of parasite growth

inhibition to those of compounds 25 and 26. However, the comparison between their

GI₅₀values showed that derivatives 29 and 31 were the most active in the series and

S7

presented good selectivity indexes (29: GI₅₀22.7 M, SI >22.0; 31: GI₅₀12.2 M, SI

41.0). The experimental results showed that the activity of compound 31 was similar

>

to those of the commercially available drug benznidazole (GI₅₀10.2 M, SI >49.1). The

presence of the 4-pentyl substituent in the derivative 31 may have been determinant for

its increased activity; as this group is hydrophobic and highly flexible, it can adopt a

conformation that favors the interaction with the hydrophobic region of the molecular

target.

To investigate one of the possible mechanisms of action of our bis-heterocyclic

compounds, derivatives that showed GI < 100 µM (10, 12, 25, 26, 28, 29, 30, and 31)

50

were evaluated for their ability to inhibit T. cruzi trypanothione reductase. None

compound showed significant inhibition at 100 μM (Table S3, Supplementary Material),

suggesting that trypanocidal activity is not related to the inhibition of this enzyme.

Complementary studies must be carried out in order to clarify the mechanism of action

of this class of compounds.

2.3 STRUCTURE-ACTIVITY RELATIONSHIP STUDY

30

A qualitative study using three-dimensional descriptors was performed

to

complement the observed SAR data of the synthesized compounds. The objective of

this step was to increase the understanding of how the studied compounds could

present their activities against the studied strain. Using the fractional factorial design

(

FFD) approach for variable selection, we obtained a Partial Least Square (PLS)

regression model composed of 95 molecular interaction field (MIF) descriptors, which

2

originated two latent variables. The results obtained for the fit of the model (R = 0.74;

2

LOO

RMSEC = 0.50; F₂_.₃₉= 55.50; critical F = 3.24) and its internal predictive ability (Q

=

0

.52; RMSECV = 0.68) indicate that the model explains and predicts information at

2

31

levels recommended by the literature (R > 0.6; Q

> 0.5) .The model is

LOO

represented graphically in Figure 2, based on compounds 10 (active) and 18 (inactive).

Distances between probes that favor activity are presented as red lines, whereas

distances between probes that are unfavorable are shown as blue lines. Most of the

descriptors that favor activity correspond to distances that match exactly those of rings

A and D, in which the variations of substituent groups were carried out.

Figure 2. Graphical representation of SAR model obtained using GRIND descriptors. Green

spheres: TIP fields; yellow spheres: DRY fields; blue spheres: N1 fields; blue lines: distances

between fields (in angstroms) that have a negative impact on activity; red lines: distance

between fields that have a positive impact on activity. 10: compound classified as 1; 18:

compound classified as −1.

S8

The mechanistic interpretation was based on the six most relevant descriptors

Table 4 and Figure S1, Supplementary Material). Spatial characteristics and

(

hydrophobicity are present in four descriptors; the three descriptors that favor inhibitory

activity (40_DRY-DRY, 413_DRY-TIP, and 583_N1-TIP) correspond to hydrophobic

and steric groups located on rings A and D. The presence of these DRY and TIP field

descriptors in these regions is consistent with that observed for the modifications

carried out in compounds 29, 30 and 31. These results corroborate the experimental

data, showing that the occurrence of activity is highly dependent on hydrophobic and

bulky groups located at rings A and D of the compounds. In 413_DRY-TIP, for

example, the distance range of 18.8–19.2 Å is not present in compound 18. These

results are supported by other studies where good antiparasitic activity was related to

10,32,33

lipophilicity and the presence of bulky groups

Therefore, considering that the

model does not contradict the experimental results, and still presents positive

correlation with other published studies, it may assist in the development of new

derivatives active against the growth of T. cruzi.

Table 4. Characteristics of the six most relevant descriptors of the model.

Sign Descriptor

Field

Distance range (Å)

−

+

−

+

8

DRY-DRY

N1-N1

3.2–3.6

16.0–16.4

8.0–8.4

40

142

160

413

583

N1-N1

15.2–15.6

18.8–19.2

13.6–14.0

DRY-TIP

N1-TIP

3.

CONCLUSION

In conclusion, 42 new bis-heterocyclic compounds were synthesized and

evaluated for trypanocidal and leishmanicidal activity. The synthetic route established

involves four steps of reactions and allows obtaining the analogues quickly and in good

yields. On the synthesized series, eight compounds (10, 12, 25, 26, 28, 29, 30, and 31)

substantially inhibited the growth of the parasite, showing GI₅₀values in the of 12.2 to

88.4 µM. Moreover, it was possible to establish the relationship between chemical

structure and trypanocidal activity, which was supported by the model generated using

GRIND descriptors. Derivative 31 was the most active of the series showing, in vitro,

low toxicity and a potency in the low micromolar range, similar to that of the reference

drug benznidazole. These results suggest that compound 31 can be considered a

promising scaffold for the design of new trypanocidal compounds.

4.

EXPERIMENTAL SECTION

4.1 Synthesis

Uncorrected melting points were determined by using a MICROQUÍMICA MQAPF-301

1

13

apparatus. All H and C NMR spectra were obtained in Nuclear Brucker Advance

DPX 400 MHz and Varian Oxford AS-400 using TMS as internal standard (δ = 0.00

ppm), unless indicated otherwise. Dimethyl sulfoxide-d6 (DMSO‑ d6), Chloroform-d

(

CDCl ) and methanol-d4 (MeOD-d4) were used as the solvents. In the absence of

3

TMS as standard, chemical shift values (δ) are given in parts per million using residual

1

solvent peaks ( H NMR: δ = 7.26 ppm for CDCl , δ = 2.50 ppm for DMSO-d and δ =

H

3

H

6

H

S9

13

3

.31 ppm for MeOD-d4; C NMR: δ = 77.0 ppm for CDCl , δ = 39.4 ppm for DMSO-

C 3 C

d and δ = 49.0 ppm for MeOD-d4) as internal standard. The residual water peak can

6

C

1

be seen in the following chemical shifts: H NMR: δ = 1.56 ppm for CDCl , δ = 3.33

ppm for DMSO-d and δ = 4.87 ppm for MeOD-d4 . When peak multiplicities are

H

3

H

34

6

H

reported, the following abbreviations are used: s, singlet; d, doublet; t, triplet; m,

multiplet; dd, doublet of doublets. Mass spectra were performed in Agilent's 1100

Series LC linked a Bruker micrOTOF. Reactions under microwave irradiation were

conducted in Discovery - CEM Explorer microwave reactor with cooling, pressure and

gas addition systems. The solvents were purchased from Tedia® and reagents were

purchased from Sigma–Aldrich®, they were treated and purified when necessary

35

according to methodologies in literature . Thin layer chromatography was performed

on silica G60 gel layers SILICYCLE® with fluorescence indicator F-254 and column

chromatography was performed using silica gel with particle size: 40-63 and 63-200μm

(

Sigma Aldrich) and hexane: ethyl acetate (Tedia) as eluent.

4.1.1. General procedure for preparation of aldoxime b1 – b4.

A mixture of 4-methoxybenzaldehyde (3.7 mmol), hydroxylamine hydrochloride (7.4

mmol) and 1,4-diazabicyclo [2.2.2] octane (DABCO) (3.7 mmol) was irradiated in

microwave in a sealed tube during 1 min., at 70 ºC and 300 W. To the reaction mixture

was added HCl 5% (10.0 mL) and the crude product was extracted with CH Cl (2 x

2

1

0.0 mL). The organic phase was dried with anhydrous Na SO and then the solvent

2 4

was removed under reduced pressure. The mixture was purified by column

chromatography [hexane: ethyl acetate, 70-30% (v/v)] affording (E)-4-

methoxybenzaldehyde oxime (b1) (520.1 mg, 3.4 mmol, 93% yield) as a white solid

yielding the expected product.

4.1.1.1. (E)-3,4-dimethoxybenzaldehyde oxime (b2).

Compound b2 was prepared as described in general procedure 4.1.1. The product b2

(

468.8 mg; 2.6 mmol, 86% yield) was obtained as a white solid. The spectral data for

22

the title compound are in agreement with the reference already reported

.

4.1.1.2. (E)–benzaldehyde oxime (b3).

Compound b3 was prepared as described in general procedure 4.1.1. The product b3

22

(

2.3 mg; 2.3 mmol, 98% yield) was obtained as a white solid . The spectral data of

this compound was compared with the commercially available compound.

4.1.1.3. (E)-benzo [1,3] dioxole-5-carbaldehyde oxime (b4).

Compound b4 was prepared as described in general procedure 4.1.1. The product b4

338.5 mg, 2.0 mmol, 60% yield) was obtained as a white solid. The spectral data for

(

36

the title compound are in agreement with the reference already reported .

4.1.2. Synthesis of isoxazole (3-(4-methoxyphenyl)isoxazol-5-yl)methanol (d1).

To a solution of (E)-4-methoxybenzaldehyde oxime (b1) (790.0 mg; 5.3 mmol) in

DMF (15.8 mL), was slowly added a small quantity of N-chlorosuccinimide (NCS). The

reaction temperature increased (if this did not happen, add 50.0 µL de HCl 1N and the

temperature should increase). After, the other amount of NCS (774.2 mg, 5.8 mmol)

was added. The reaction was stirred for one hour at room temperature and controlled

by TLC. The reaction was quenched by addition of ethyl ether (20.0 mL) and washed

with NaCl (2 x 20.0 mL). The organic phase was dried with anhydrous Na SO and then

2

4

the solvent was rotaevaporatored. The carboxyimidoyl chloride obtained was diluted in

S10

t-BuOH:H O (1:1) (3.8 mL) and then propargylic alcohol (340.0 µL, 5.8 mmol), CuSO₄

2

(

7.5 mg; 0.05 mmol, diluted in 50.0 µL of water), sodium ascorbate (104.40 mg; 0.5

mmol) and NaHCO (1.3 g; 15.8 mmol) were added. The reaction was stirred for 4

3

hours, then diluted in ethyl acetate (20.0 mL) and washed with H O (2 x 20.0 mL). The

2

organic phase was dried with anhydrous Na SO and then the solvent was removed

2

4

under reduced pressure. The purified compound (d1) was obtained after column

chromatography [hexane: ethyl acetate, 8:2 (v/v)]. Light yellow solid (890.0 mg; 4.3

o

mmol, 82% yield). Mp.: 90-92 C. Rf.:0.40 [hexane: ethyl acetate, 60:40 % (v/v)]. IR

-1 1

(

KBr): 3352, 2837, 1610, 1527, 1431, 1259, 1082, 840, 802 cm . H NMR (400 MHz,

CDCl ) δ 7.74 (d, J= 8.5 Hz, 2H), 6.98 (d, J= 8.5 Hz, 2H), 6.52 (s, 1H), 4.81 (s, 2H),

3

13

3

.86 (s, 3H). C NMR (100 MHz, CDCl ) δ 171.5, 162.1, 161.0, 128.2, 121.4, 114.3,

3

9

99.8, 56.7, 55.4 .

4.1.3.General procedure for preparation of isoxazoles d2-d4.

Under microwave irradiation, to a solution of oxime (1.03 mmol) in DMF (0.3 mL),

was added NCS (1.1 mmol) and the mixture was irradiated in a sealed tube during 1

min., at 35 ºC and 150 W. After that, propargylic alcohol (1.1 mmol), CuSO (0.01

4

mmol) dilute in 50.0 µL of water, sodium ascorbate (0.10 mmol) and NaHCO (4.10

3

mmol) were added to the tube. The reaction was irradiated again during 10 min., in the

same conditions above. When reaction time was over, the mixture was diluted in

EtOAc (20.0 mL) and washed with NaCl (2 x 20.0 mL). The organic phase was dried

with anhydrous Na SO and the solvent was removed under reduced pressure yielding

2

4

the expected product.

4.1.3.1. (3-(3,4-dimethoxyphenyl)isoxazol-5-yl)methanol (d2).

Compound d2 was prepared as described in general procedure 4.1.3. The product d2

259.66 mg, 1.1 mmol, 55% yield) was obtained as a white solid. The spectral data for

(

9

the title compound are in agreement with the reference already reported .

4.1.3.2. (3-phenylisoxazol-5-yl)methanol (d3).

Compound d3 was prepared as described in general procedure 4.1.3. The product d3

27.36 mg, 0.2 mmol, 76% yield) was obtained as a light yellow solid. The spectral data

(

9

for the title compound are in agreement with the reference already reported .

4.1.3.3. (3-(benzo[d][1,3]dioxol-5-yl)isoxazol-5-yl)methanol (d4).

Compound d3 was prepared as described in general procedure 4.1.3. The product d3

144.6 mg, 0.7 mmol, 54% yield) was obtained as a light yellow solid. The spectral data

(

9

for the title compound are in agreement with the reference already reported .

4.1.4. General procedure for preparation of tosylates e1-e4.

To a solution of the isoxazol (d) (0.1 mmol) in CH Cl :H O (1:1) at molar

2

4

concentration 10 mol/L, were added: triethylamine (TEA) (0.01 mmol), K CO (0.2

2

3

mmol) and, finally, 4-toluenesulfonyl chloride (0.2 mmol) divided in 6 equal parts added

every 10 min. The pH was monitored and adjusted by addition of NaOH to 10-11. The

reaction was stirred for 8 hours at room temperature. At the end of the reaction, 50.0

µL of TEA was added and the reaction further stirred for 10 min. Then, the solution was

quenched with CH Cl (20.0 mL) and washed with saturated solution of NaCl (2 x 20.0

2

mL). The organic phase was dried with anhydrous Na SO and the solvent was

2

4

removed under reduced pressure. The mixture was purified by chromatography

Hexane:EtOAc 80:20 % (v/v) yielding the expected product.

[

S11

4.1.4.1. (3-(4-methoxyphenyl)isoxazol-5-yl)methyl 4-methylbenzenesulfonate (e1).

Compound e1 was prepared as described in general procedure 4.1.4. The product e1

27.3 mg, 0.1 mmol, 76 % yield) was obtained as a white solid. The spectral data for

(

9

the title compound are in agreement with the reference already reported .

4.1.4.2. (3-(3,4-dimethoxyphenyl)isoxazol-5-yl)methyl 4-methylbenzenesulfonate (e2).

o

White solid (26.3mg, 0.1 mmol, 63% yield). Mp.: 112- 113 C. Rf.: 0.24

[

9

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2837, 1583, 1525, 1469, 1355, 1174, 1018,

-1 1

64, 813, 779, 734 cm . H NMR (400 MHz, CDCl ) δ 7.82 (d, J= 8.4 Hz, 2H), 7.35 (d,

3

J= 2.0 Hz, 1H), 7.35 (d, J= 8.4 Hz, 2H), 7.22 (dd, J= 8.3, 2.0 Hz, 1H), 6.92 (d, J= 8.3

Hz, 1H), 6.54 (s, 1H), 5.17 (s, 2H), 3.94 (s, 3H), 3.93 (s, 3H), 2.42 (s, 3H).

4.1.4.3. (3-phenylisoxazol-5-yl)methyl 4-methylbenzenesulfonate (e3).

o

White solid (40.7mg, 1.2 mmol, 49% yield). Mp.: 93-94 C. Rf.: 0.50 [Hexane:EtOAc,

6

–

(

0:40 % (v/v)]. IR (KBr): 3115, 1957, 1469, 1436, 1357, 1172, 1095, 939, 817, 767,

-1 1

94cm . H NMR (300 MHz, CDCl ) δ: 7.81 (d, J= 8 Hz, 2H), 7.76 – 7.68 (m, 2H), 7.49

3

7.42 (m, 3H), 7.34 (d, J= 8 Hz, 2H), 6.55 (s,1H), 5.19 (s, 2H), 2.40 (s, 3H). HSQC

1

13

H: 300 MHz, C: 75 MHz, CDCl ) δ 130.0, 129.1, 128.1, 126.8, 103.1, 61.1, 21.6.

3

C

4.1.4.4. (3-(benzo[d][1,3]dioxol-5-yl)isoxazol-5-yl)methyl 4-methylbenzenesulfonate

(

e4).

o

Light yellow solid (51.7mg, 0.1 mg, 57% yield). Mp.: 84-85 C. Rf.: 0.6 [Hexane:EtOAc,

6

1

2

0:40 % (v/v)]. IR (KBr): 3126, 2781, 1597, 1516, 1465, 1423, 1359, 1244, 1180, 1095,

-1

1

035, 954, 929, 873, 812, 744cm . H NMR (400 MHz, CDCl ) δ 7.81 (d, J= 8.3 Hz,

3

H), 7.35 (d, J= 8.3 Hz, 2H), 7.25 (d, J= 2.0 Hz, 1H), 7.19 (dd, J=8.4, 2.0 Hz, 1H), 6.87

(

d, J= 8.4 Hz, 1H), 6.46 (s, 1H), 6.03 (s, 2H), 5.30 (s, 1H), 5.17 (s, 2H), 2.43 (s, 3H).

4.1.5. General procedure for preparation of azides f1-f4.

To a solution of the tosylate (e) (0.1 mmol) in 0.2 mL of DMF sodium azide (NaN₃)

(

0.6 mmol) was added. The reaction was irradiated in microwave in a sealed tube

during 10 min., at 70 ºC and 150 W. At the end, the reaction was quenched with EtOAc

20.0 mL) and washed with satured solution of NaCl (2 x 20.0 mL). The organic phase

was dried with anhydrous Na SO and was removed under reduced pressure. The

(

2

4

mixture was purified by chromatography [hexane: ethyl acetate, 90:10 % (v/v)] yielding

the expected product.

4.1.5.1. 5-(azidomethyl)-3-(4-methoxyphenyl)isoxazole (f1).

Compound f1 was prepared as described in general procedure 4.1.5. The product f1

11.4 mg, 0.05 mmol, 89% yield) was obtained as a white solid. The spectral data for

(

9

the title compound are in agreement with the reference already reported .

4.1.5.2. 5-(azidomethyl)-3-(3,4-dimethoxyphenyl)isoxazole (f2).

o

White solid (386.0 mg, 1.5 mmol, 80% yield). Mp.: 88–89 C. Rf.: 0.8 [Hexane:EtOAc,

6

8

2

3

0:40 % (v/v)]. IR (KBr): 2839, 2098, 1612, 1583, 1525, 1471, 1234, 1020, 912, 871,

-1 1

54, 765 cm . H NMR (300 MHz, CDCl ) δ 7.42 (d, J= 2.0 Hz, 1H), 7.30 (dd, J= 8.3,

3

.0 Hz, 1H), 6.94 (d, J= 8.3 Hz, 1H), 6.57 (s, 1H), 4.50 (s, 2H), 3.96 (s, 3H), 3.94 (s,

H).

4.1.5.3. 5-(azidomethyl)-3-phenylisoxazole (f3).

S12

o

White solid (209.6 mg, 1.0 mmol, 85% yield). Mp.: 51-52 C. Rf.: 0.9 [Hexane:EtOAc,

6

1

–

0:40 % (v/v)]. IR (KBr): 3120, 2999, 2106, 1604, 1577, 1469, 1444, 1404, 1290, 1251,

-1 1

080, 950, 883, 773, 694 cm . H NMR (300 MHz, CDCl ) δ 7.88 – 7.76 (m, 2H), 7.53

3

1

13

7.40 (m, 3H), 6.60 (s, 1H), 4.50 (s, 2H). HSQC ( H: 300 MHz, C: 75 MHz, CDCl₃)

δ_C126.9, 129.5, 101.2, 45.5.

.1.5.4. 5-(azidomethyl)-3-(benzo[d][1,3]dioxol-5-yl)isoxazole (f4).

Light yellow solid (230.6 mg, 0.94 mmol, 87% yield). Mp.: 55-56 C. Rf.: 0.7

4

o

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3126, 2781, 2137, 2102, 1597, 1516, 1465,

423, 1359, 1244, 1180, 1095, 1035, 954, 929, 873, 812, 744 cm . H NMR (400 MHz,

-1 1

CDCl ) δ 7.33 (d, J= 1.7 Hz, 1H), 7.27 (q, J= 8.1, 1.7 Hz, 1H), 6.89 (d, J= 8.1 Hz, 1H),

3

13

6

.52 (s, 1H), 6.04 (s, 2H), 4.49 (s, 2H). C NMR (100 MHz, CDCl ) δ 166.9, 162.7,

3

149.4, 148.3, 122.5, 121.3, 108.7, 106.9, 101.5, 101.1, 45.5.

4.1.6. General procedure for synthesis of triazole derivatives (h).

To a solution of the azide intermediate (0.1 mmol) in 0.2 mL of DMF in a

microwave tube, CuSO (0.05 mmol) diluted in 50.0 µL of water, sodium ascorbate

4

(

0.025 mmol) and the alkyne (0.2 mmol) were added. The tube was sealed and the

solution was irradiated (70 ºC, 150 W) for 10 min and, then, TLC analysis showed the

complete consumption of the starting material. The reaction was diluted with 15 mL of

brine, extracted with ethyl acetate (3 x 10 mL), dried by anhydrous Na SO and

2

4

concentrated under reduced pressure. The resulting crude product was purified by

column chromatography on silica gel [hexane: ethyl acetate, 60:40 % (v/v)] yielding the

expected product.

4.1.6.1. 3-phenyl-5-((4-phenyl-1H-1,2,3-triazol-1-yl)methyl)isoxazole (7).

o

White solid (28.8 mg, 0.09 mmol, 76% yield). M.P: 175-176 C. R.f: 0.59

[

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3116, 1612, 1579, 1462, 1442, 1406, 1218,

-1 1

1

7

5

1

+

082, 761, 690 cm . H NMR (400 MHz, CDCl ) δ 7.95 (s, 1H, H-triazole), 7.83 (d, J=

3

.4 Hz, 2H, Ar), 7.77-7.75 (m, 2H, Ar), 7.45-7.41 (m, 6H, Ar), 6.61 (s, 1H, H-isoxazole),

13

.77 (s, 2H, CH ). C NMR (100 MHz; CDCl ) δ 165.6, 163.1, 148.9, 130.6, 130.2,

2

3

29.2, 129.1, 128.7, 128.3, 127.0, 126.0, 120.0, 102.7, 45.5. HRMS (ESI-TOF) m/z: [M

+

H]+ Calcd for C H N O 303.1246, found 303.1301 [M + H] .

18

15

4

4.1.6.2. 5-((4-(4-methoxyphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole (8).

o

Light yellow solid (15.9 mg, 0.05 mmol, 38% yield). M.P: 210-211 C. R.f: 0.55

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3174, 2968, 2837, 1608, 1577, 1548, 1465,

-1 1

253, 1068, 758, 696 cm . H NMR (400 MHz, CDCl ) δ 8.36 (dd, J= 7.7, 1.7 Hz, 1H,

3

Ar), 8.23 (s, 1H, H-triazole), 7.79 – 7.74 (m, 2H, Ar), 7.47 – 7.43 (m, 3H, Ar), 7.34 (dd,

J= 7.5, 1.7 Hz, 1H, Ar), 7.09 (td, J= 7.6, 1.0 Hz, 1H, Ar), 6.99 (d, J= 8.4 Hz, 1H, Ar),

13

6

1

3

.56 (s, 1H, H-isoxazole), 3.95 (s, 3H, Ar-OCH₃). C NMR (125 MHz, DMSO‑ d6) δ

67.3, 162.2, 155.4, 142.1, 130.4, 129.1, 128.1, 126.7, 126.6, 124.5, 120.7, 118.7,

11.6, 102.0, 55.5, 44.5. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O

19

17

4

2

+

33.1352, found 333.1410 [M + H] .

4.1.6.3. 5-((4-(2-methoxyphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole (9).

o

Light yellow solid (36.0 mg, 0.10 mmol, 88% yield). M.P: 210-211 C; R.f: 0.36

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3172, 2968, 2837, 1658, 1583, 1548, 1442,

-1 1

253, 1068, 777, 758, 698 cm . H NMR (300 MHz, CDCl ) δ 8.36 (dd, J=7.7, 1.7 Hz,

3

1H, Ar), 8.22 (s, 1H, H-triazole), 7.79 – 7,75 (m, 2H, Ar), 7,48 – 7,43 (m, 3H, Ar), 7,34

S13

(

1

ddd, J=7,7, 8,5, 1.0 Hz, 1H, Ar), 7.10 (ddd, J=8.5, 8.0, 1.7 Hz, 1H, Ar), 6.99 (dd, J=8.0,

.0 Hz, 1H, Ar), 6.57 (s, 1H, H-isoxazole), 5.79 (s, 2H, CH ), 3.95 (s, 3H, Ar-OCH ).

2

3

1

13

HSQC ( H: 300 MHz, C: 75 MHz, CDCl ) δ 127.8, 123.4, 126.8, 130.3, 129.0, 129.1,

3

C

1

21.0, 110.9, 102.1, 45.3, 55.4. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O

19 17 4 2

+

333.1352, found 333.1371 [M + H] .

4.1.6.4.

5-((4-(3,5-dimethoxyphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole

(

10).

o

White solid (34.3 mg, 0.09 mmol, 76% yield). M.P: 119-120 C. R.f: 0.31

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3128, 2839, 1593, 1556, 1427, 1207, 1157,

-1

1

066, 833, 773, 696, 688 cm . H NMR (300 MHz, CDCl ) δ 7.93 (s, 1H, H-triazole),

3

7.78-7.75 (m, 2H, Ar), 7.49 – 7.42 (m, 3H, Ar), 7.00 (d, J= 2.3 Hz, 2H, Ar), 6.61 (s, 1H,

H-isoxazole), 6.46 (t, J=2.3 Hz, 1H, Ar), 5.77 (s, 2H, CH ), 3.84 (s, 6H, Ar-OCH ).

2

3

1

13

HSQC ( H: 300 MHz, C: 75 MHz, CDCl ) δ 129.1, 126.9, 120.2, 103.7, 102.3, 101.0,

3

C

5

5.5, 45.4. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 363.1457, found

20 18 4 3

+

.

363.1882 [M + H]

4.1.6.5. 3-phenyl-5-((4-(p-tolyl)-1H-1,2,3-triazol-1-yl)methyl)isoxazole (11).

o

White solid (33.0 mg, 0.10 mmol, 84% yield). M.P: 171-172 C. R.f: 0.57

[

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3113, 2920, 2852, 1610, 1581, 1498, 1444,

-1 1

1

7

2

228, 819, 769, 688 cm . H NMR (400 MHz, CDCl ) δ 7.90 (s, 1H, H-triazole), 7.77-

3

.74 (m, 2H, Ar), 7.72 (d, J= 8.0 Hz, 2H, Ar), 7.45-7.46 (m, 3H, Ar), 7.23 (d, J= 8.0 Hz,

13

H, Ar), 6.60 (s, 1H, H-isoxazole), 5.74 (s, 2H, CH ), 2.37 (s, 3H, AR-CH ). C NMR

2

3

(

100 MHz, CDCl ) δ 165.7, 163.1, 149.0, 138.5, 130.7, 129.7, 129.2, 128.3, 127.4,

3

1

27.0, 125.9, 119.6, 102.4, 45.5, 21.4. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for

+

C H N O 317.1402, found 317.1462 [M + H] .

19

17

4

4.1.6.6.

3-phenyl-5-((4-(2,4,5-trimethylphenyl)-1H-1,2,3-triazol-1-yl)methyl)isoxazole

(

12).

o

White solid (21.1 mg, 0.06 mmol, 49% yield). M.P: 200-201 C. R.f: 0.63

[

1

Hexane:EtOAc, 60:40 % (v/v)], IR (KBr): 3163, 2966, 2883, 1614, 1573, 1504, 1442,

-1

1

230, 945, 871, 771, 734, 698 cm . H NMR (300 MHz, CDCl ) δ 7.84 (s, 1H, H-

3

triazole), 7.80 – 7.75 (m, 2H, Ar), 7.53 (s, 1H, Ar), 7.48-7.45 (m, 3H, Ar), 7.30 (s, 1H,

Ar), 7.05 (s, 1H, Ar), 6.66 (s, 1H, H-isoxazol), 5.80 (s, 2H, CH ), 2.39 (s, 3H, AR-CH ),

2

3

1

13

2

.27 (s, 3H, AR-CH ). HSQC ( H: 300 MHz, C: 75 MHz, CDCl ) δ 132.2, 129.9,

3

C

1

29.1, 126.9, 121.9, 102.3, 45.2, 20.6, 19.2, HRMS (ESI-TOF) m/z: [M + H]+ Calcd

+

for C H N O 345.1715, found 345.1750 [M + H] .

21

4

4.1.6.7. 5-((4-(4-methoxy-2-methylphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-

phenylisoxazole (13).

o

White solid (31.2 mg, 0.09 mmol, 92% yield). M.P: 127-128 C. R.f: 0.46

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3140, 2960, 1606, 1577, 1492, 1444, 1406,

[

-

1 1

1

3

2

3

236, 1205, 1039, 848, 829, 770, 692 cm . H NMR (300 MHz, CDCl ) δ 7.79-7.75 (m,

3

H, Ar/H-triazol), 7.69 (d, J= 9.3 Hz, 1H, Ar), 7.50 – 7.43 (m, 2H, Ar), 6.82 (dd, J= 9.3,

.4 Hz, 1H, Ar), 6.81 (d, J= 2.4 Hz, 1H, Ar) 6.62 (s, 1H, H-isoxazol), 5.78 (s, 2H, CH ),

.83 (s, 3H, Ar-OCH3), 2.44 (s, 3H, Ar-CH3). HSQC ( H: 300 MHz, C: 75 MHz,

2

1

13

CDCl ) δ 130.2, 129.2, 126.9, 121.4, 116.3, 111.5, 102.3, 55.3, 45.2, 21.5. HRMS

3

C

+

(

ESI-TOF) m/z: [M + H]+ Calcd for C H N O 347.1508, found 347.1763 [M + H] .

20 19 4 2

4.1.6.8. 5-((4-(2,5-dimethylphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole (14).

S14

o

White solid (29.0 mg, 0.09 mmol, 88% yield). M.P: 136-137 C. R.f: 0.46

[

7

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3107, 1614, 1581, 1440, 1404, 1864, 815,

-1 1

69, 690 cm . H NMR (300 MHz, CDCl ) δ 7.83 (s, 1H, H-triazole), 7.81 – 7.74 (m 2H,

3

Ar), 7.62 (d, J= 1.4 Hz, 1H, Ar), 7.49 – 7.43 (m, 3H, Ar), 7.16 (d, J= 7.8 Hz, 1H, Ar),

7

3

1

.09 (dd, J=7.8, 1.4 Hz, 1H, Ar), 6.62 (s, 1H, H-isoxazol), 5.79 (s, 2H, CH ), 2.42 (s,

2

1

13

H, Ar-CH3), 2.36 (s, 3H, Ar-CH3). HSQC ( H: 300 MHz, C: 75 MHz, CDCl ) δ_C

3

30.8, 129.5, 129.2, 129.2, 126.9, 122.0, 102.2, 45.3, 20.9, 20.9, HRMS (ESI-TOF)

+

m/z: [M + H]+ Calcd for C H N O 331.1559, found 3331.1787 [M + H] .

20

19

4

4.1.6.9.

5-((4-(6-methoxynaphthalen-2-yl)-1H-1,2,3-triazol-1-yl)methyl)-3-

phenylisoxazole (15).

o

Light yellow solid (38.0 mg, 0.10 mmol, 99% yield). M.P: 197-198 C. R.f: 0.6

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3118, 2962, 1610, 1546, 1508, 1469, 1440,

-1 1

224, 1211, 1029, 902, 856, 821, 767, 686cm . H NMR (300 MHz, CDCl ) δ 8.28 (s,

3

1H, Ar), 8.02 (s, 1H, H-triazole), 7.89 (dd, J= 8.0, 1.0 Hz, 1H, Ar), 7.83 – 7.76 (m, 4H,

Ar), 7.51 – 7.43 (m, 3H, Ar), 7.19 (d, J= 2.0 Hz, 1H, Ar), 7.15 (s, 1H, Ar), 6.64 (s, 1H, H-

1

13

isoxazole), 5.81 (s, 2H, CH ), 3.94 (s, 3H, Ar-OCH ). HSQC ( H: 300 MHz, C: 75

2

3

MHz, CDCl ) δ 129.7, 129.1, 127.1, 124.6, 124.3, 119.7, 119.4, 105.8, 102.4, 55.3,

3

C

4

5.4. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 383.1508, found 383.1780

23 19 4 2

+

[

M + H] .

4.1.6.10. 5-((4-(4-nitrophenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole (16).

o

White solid (30.8 mg, 0.09 mmol, 89% yield). M.P: 131-132 C. R.f:

0

1

2

7

.43[Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3115, 2922, 850, 1612, 1581, 1467,

-1 1

408, 1226, 839, 808, 765, 688 cm . H NMR (300 MHz, CDCl ) δ 8.31 (d, J= 9.0 Hz,

3

H, Ar), 8.10 (s, 1H, H-triazole), 8.03 (d, J=9.0 Hz, 2H, Ar), 7.81 – 7.76 (m, 2H, Ar),

1

,50 – 7.45 (m, 3H, Ar), 6.69 (s, 1H, H-isoxazole), 5.83 (s, 2H, CH ). HSQC ( H: 400

2

13

MHz, C: 100 MHz, CDCl ) δ 130.6, 129.0, 126.9, 126.4, 124.6, 121.2, 102.8, 45.3.

3

C

HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 348.1097, found 348.1391 [M +

18

14

5

3

+

H] .

4.1.6.11.

3-phenyl-5-((4-(4-(trifluoromethoxy)phenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (17).

o

White solid (30.0 mg, 0.08 mmol, 77% yield). M.P: 182-183 C. R.f: 0.37

[

8

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3113, 3089, 1614, 1581, 1556, 1309, 1209,

-1 1

50, 810, 767, 694 cm . H NMR (300 MHz, CDCl ) δ 7.90 (s, 1H, H-triazole), 7.81 (d,

3

J= 8.5 Hz, 2H, Ar), 7.74 – 7.69 (m, 2H, Ar), 7.43-7.38 (m, 3H, Ar), 7.23 (d, J= 8.5 Hz,

1

3

2

1

H, Ar), 6.58 (s, 1H, H-isoxazole), 5.73 (s, 2H, CH ). C NMR (75 MHz, CDCl ) δ

2

3

2

65.3, 163.2, 149.4 (d, J = 1.5 Hz), 147.6, 130.7, 129.2, 129.0, 128.3, 127.4, 127.0,

21.6, 120.8 (d, J = 257.6 Hz), 120.2, 102.6, 45.5. HRMS (ESI-TOF) m/z: [M + H]+

Calcd for C H F N O 387.1069, found 387.1032 [M + H] .

CF

1

CF

+

19

14

3

4

2

4.1.6.12. 5-((4-(4-fluorophenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole (18).

o

White solid (21.9 mg, 0.07 mmol, 68% yield). M.P: 195-196 C. R.f: 0.57

[

8

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3101, 1612, 1560, 1496, 1406, 1228, 1082,

-1 1

33, 781, 767, 692 cm . H NMR (300 MHz, CDCl ) δ 7.91 (s, 1H, H-triazole), 7.80 (m,

3

4

H, Ar), 7.53 – 7.43 (m, 3H, Ar), 7.15-7.10 (m, 2H, Ar), 6.63 (s, 1H, H-isoxazole), 5.78

13

1

(

s, 2H, CH₂). C NMR (75 MHz, CDCl ) δ: 165.3, 163.0, 162.6 (d, J = 257.7 Hz),

3 CF

3

2

1

=

3

47.9, 130.5, 129.1, 128.1, 127.6 (d, J_C_F= 8.2 Hz), 126.8, 126.3, 119.6, 115.9 (d, J

CF

21.9 Hz), 102.4, 45.4. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H FN O

18

14

4

+

21.1152, found 321.1417 [M + H] .

S15

4.1.6.13. N,N-dimethyl-4-(1-((3-phenylisoxazol-5-yl)methyl)-1H-1,2,3-triazol-4-yl)aniline

(

19).

o

Light yellow solid (29.9 mg, 0.09 mmol, 87% yield). M.P: 160-161 C. R.f: 0,54

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3124, 2987, 2808, 1618, 1606, 1556, 1508,

-1 1

454, 1442, 1359, 1224, 948, 821, 773, 698 cm . H NMR (300 MHz, CDCl ) δ 7.80 (s,

3

1H, H-triazole), 7.78 – 7.74 (m, 2H, Ar), 7.70 (d, J= 8.9 Hz, 2H, Ar), 7.47 – 7.42 (m, 3H,

Ar), 6.76 (d, J= 8.9 Hz, 2H, Ar), 6.58 (s, 1H, H-isoxazole), 5.73 (s, 2H, CH ), 2.99 (s,

2

1

13

6

H, N-CH ). HSQC ( H: 300 MHz, C: 75 MHz, CDCl ) δ 129.1, 126.9, 118.4, 112.5,

3

C

1

02.2, 45.3, 40.5. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 346.1668,

20

5

+

found 346.1960 [M + H] .

4.1.6.14. 4-(1-((3-phenylisoxazol-5-yl)methyl)-1H-1,2,3-triazol-4-yl)aniline (20).

o

Light yellow solid (15.0 mg, 0.05 mmol, 47% yield). M.P: 220- 221 C. R.f: 0.15

[

Hexane:EtOAc, 60:40 % (v/v)], IR (KBr): 3460, 3360, 3223, 3122, 1730, 1631, 1614,

-1 1

1

7

3

566, 1502, 1442, 1404, 1296, 1045, 833, 771, 688 cm . H NMR (300 MHz, CDCl ) δ

3

.80 (s, 1H, H-triazole), 7.79 – 7.74 (m, 2H, Ar), 7.63 (d, J= 8.2 Hz, 2H, Ar), 7.46 (m,

13

H, Ar), 6.74 (d, J= 8.2 Hz, 2H, Ar), 6.59 (s, 1H, H-isoxazole), 5.75 (s, 2H, CH ).

C

2

NMR (125 MHz, MeOD) δ 164.2, 146.5, 131.3, 129.9, 127.7, 121.7, 120.7, 116.1,

1

03.1, 45.9. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 318.1355, found

18 16 5

+

318.1628 [M + H] .

4.1.6.15.

(4-(1-((3-phenylisoxazol-5-yl)methyl)-1H-1,2,3-triazol-4-yl)phenyl)methanol

(

21).

o

White solid (17.4 mg, 0.05 mmol, 42% yield). M.P: 170-171 C. R.f: 0.13

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3228, 3115, 3091, 2904, 1612, 1581, 1548,

444, 1404, 1203, 10493 1004, 763, 688 cm . H NMR (400 MHz, MEOD) δ 8.40 (s,

-1

1

H, H-triazole), 7.83 – 7.80 (m, 4H, Ar), 7.49 – 7.46 (m, 3H, Ar), 7.44 (d, J= 8.5 Hz, 2H,

13

Ar), 6.89 (s, 1H, H-isoxazole), 5.92 (s, 2H, CH ), 4.65 (s, 2H, CH ). C NMR (100 MHz,

2

MeOD) δ 165.5, 163.0, 148.4, 141.7, 130.5, 129.0, 128.7, 128.0, 127.4, 126.8, 125.8,

1

20.4, 102.5, 64.1, 45.3. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O

19 17 4 2

+

333.1352, found 333.1409 [M + H] .

4.1.6.16. 5-((4-(4-pentylphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-phenylisoxazole (22).

o

White solid (36.0 mg, 0.09 mmol, 97% yield). M.P: 201-202 C. R.f: 0.31

[

7

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3118, 1606, 1581, 1514, 1352, 1232, 860,

-1

1

79, 694 cm . NMR H (300 MHz, CDCl ) δ 7.90 (s, 1H, H-triazole), 7.78-7.73 (m, 2H,

3

Ar), 7.74 (d, J= 8.1 Hz, 2H, Ar), 7.49 – 7.42 (m, 3H, Ar), 7.24 (d, J= 8.1 Hz, 2H, Ar),

6

2

4

3

.60 (s, 1H, H-isoxazole), 5.76 (s, 2H, CH ), 2.68 – 2.57 (m, 2H, CH ), 1.67 – 1.57 (m,

2 2

1

H, CH ), 1.38 – 1.29 (m, 4H, CH -CH ), 0.89 (t, J= 6.8 Hz, 3H, CH -CH ). HSQC ( H:

2

3

13

00 MHz, C: 75 MHz, CDCl ) δ 128.9, 128.9, 126.0, 119.5, 102.2, 45.4, 35.7, 31.5,

3

C

1.0, 22.5, 14.0. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 373.2028,

23

25

4

+

found 373.2338 [M + H] .

4.1.6.17.

3-(4-methoxyphenyl)-5-((4-(2,4,5-trimethylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (23).

o

White solid (38.0 mg, 0,09, 97% yield). Mp: 127-128 C. IR (KBr): 2835, 1610, 1531,

-

1

7

6

427, 1255, 1176, 1033, 829 cm . H NMR (300 MHz, CDCl ) δ 7.80 (s,1H, triazole),

3

.71 (d, J=8.9 Hz, 2H, Ar), 7.58 (s, 1H, Ar), 7.04 (s, 1H, Ar), 6.96 (d, J= 8.9 Hz, 2H, Ar),

.56 (s,1H, Isoxazole), 5.77 (s, 2H, CH ), 3.85 (s, 3H, Ar-OCH ), 2.40 (s, 3H, CH ),

2

3

S16

1

3

2

1

3

.26 (s, 6H, CH ). C NMR (100 MHz, CDCl ) δ 165.5, 162.7, 161.5, 148.2, 137.0,

3

34.4, 132.8, 132.4, 130.1, 128.4, 126.9, 121.8, 120.9, 114.6, 102.1, 55.5, 45.5, 20.9,

9.5, 19.3. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 375.1816, found

22

23

4

2

+

.

75.1999 [M + H]

4.1.6.18.

5-((4-(3,5-dimethoxyphenyl)-1H-1,2,3-triazol-1-yl)methyl)-3-(4-

methoxyphenyl)isoxazole (24).

o

Light yellow solid (32,2mg, 0,08 mmol, 82% yield). Mp:142–143 C. IR (KBr): 2837,

-1

1

620, 1591, 1529, 1429, 1255, 1157, 1043, 860, 831, 730 cm . H NMR (300 MHz,

CDCl ) δ 7.92 (s,1H, H-triazole), 7.71 (d, J= 8.8 Hz, 2H, Ar), 7.00 (d, J= 2.3 Hz, 2H,

3

Ar), 6.96 (d, J= 8.8 Hz, 2H, Ar), 6.56 (s,1H, H-isoxazole), 6.46 (t, J= 2.3 Hz, 1H, Ar),

13

5

.75 (s, 2H, CH ), 3.85 (s, 3H, Ar-OCH ), 3.84 (s, 6H, Ar-OCH ). C NMR (100 MHz,

2 3 3

CDCl ) δ 165.2, 162.7, 161.5, 161.4, 148.7, 132.0, 128.4, 120.8, 120.3, 114.6, 103.9,

3

1

02.2, 101.2, 55.7, 55.5, 45.5. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O

21 21 4

+

393.1563, found 393.1803 [M + H] .

4.1.6.19. 3-(3,4-dimethoxyphenyl)-5-((4-(2,4,5-trimethylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (25).

o

White solid (38.0 mg, 0.09 mmol, 97 % yield). M.P: 98–99 C. R.f: 0,18[Hexane:

EtOAc, 60:40 % (v/v)], IR (KBr): 2831, 1618, 1589, 1525, 1475, 1435, 1259, 1234,

-

1 1

1

7

1

153, 1029, 893, 796, 742 cm , H NMR (400 MHz, CDCl ) δ 7.80 (s, 1H, H-triazole),

3

.58 (s, 1H, Ar), 7.37 (d, J= 1.9 Hz, 1H, Ar), 7.26 (dd, J= 8.4, 1.9 Hz, 2H, Ar), 7.04 (s,

H, Ar), 6.91 (d, J= 8.4 Hz, 1H, Ar), 6.57 (s, 1H, H-isoxazole), 5.77 (s, 2H, CH ), 3.93

2

(

s, 3H, CH ), 3.92 (s, 3H, Ar-OCH ), 2.40 (s, 3H, AR-CH ), 2.27 (s, 3H, AR-CH ) 2.26

3 3 3 3

13

s, 3H, AR-CH₃). C NMR (100 MHz, CDCl ) δ 165.6, 162.8, 151.1, 149.6, 148.2,

3

137.0, 134.4, 132.8, 132.5, 130.1, 126.8, 121.8, 121.1, 120.2, 111.3, 109.4, 102.2,

5

6.2, 56.1, 45.5, 21.0, 19.5, 19.3. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for

+

C H N O 405.1927, found 405.2223 [M + H] .

23

25

4

3

4.1.6.20.

3-(3,4-dimethoxyphenyl)-5-((4-(3,5-dimethoxyphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (26).

o

White solid (40.0 mg, 0.09 mmol, 96% yield). M.P: 145–146 C. R.f: 0.07

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2841, 1654, 1591, 1527, 1477, 1419, 1157,

-1 1

060, 842, 821 cm . H NMR (400 MHz, CDCl ) δ 7.92 (s, 1H, H-triazole), 7.36 (d, J=

3

1.9 Hz, 1H, Ar), 7.25 (dd, J= 8.3, 1.9 Hz, 1H, Ar), 7.00 (d, J= 2.3 Hz, 2H, Ar), 6.91 (d,

J= 8.3 Hz, 1H, Ar), 6.56 (s, 1H, H-isoxazole), 6.46 (t, J=2.3 Hz, 1H, Ar), 5.75 (s, 2H,

13

CH ), 3.93 (s, 3H, Ar-OCH ), 3.92 (s, 3H, Ar-OCH ), 3.84 (s, 6H, Ar-OCH ). C NMR

2

3

(

100 MHz, CDCl ) δ 165.3, 162.8, 161.4, 151.2, 149.6, 148.7, 132.0, 121.0, 120.3,

3

1

20.2, 111.3, 109.5, 103.9, 102.3, 101.1, 56.2, 56.1, 55.7, 45.6, HRMS (ESI-TOF) m/z:

+

[

M + H]+ Calcd for C H N O 423.1668, found 423.2027 [M + H] .

22 23 4

4.1.6.21.

3-(benzo[d][1,3]dioxol-5-yl)-5-((4-(2,4,5-trimethylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (27).

o

White solid (28.2 mg, 0.07 mmol, 88% yield). M.P: 170-171 C. R.f: 0.38

[

9

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3120, 2862, 1604, 1517, 1463, 1240, 1043,

-1 1

39, 869, 812 cm . H NMR (400 MHz, CDCl ) δ 7.72 (s, 1H, H-triazole), 7.50 (s, 1H,

3

Ar), 7.21 (d, J= 1.8 Hz, 1H, Ar), 7.15 (dd, J= 8.1, 1.8 Hz, 1H, Ar), 6.97 (s, 1H, Ar), 6.79

d, J= 8.1 Hz, 1H, Ar), 6.45 (s, 1H, H-isoxazole), 5.94 (s, 2H, O-CH -O), 5.68 (s, 2H,

(

2

13

CH ), 2.32 (s, 3H, Ar-CH ), 2.19 (s, 3H, Ar-CH ), 2.19 (s, 3H, Ar-CH ) C NMR (100

2

3

S17

MHz, CDCl ) δ 165.54, 162.62, 149.61, 148.45, 148.16, 137.04, 134.42, 132.74,

3

132.42, 130.05, 126.76, 122.23, 121.87, 121.49, 108.82, 107.00, 102.21, 101.70,

7

7.48, 77.16, 76.84, 45.41, 20.91, 19.54, 19.29. HRMS (ESI-TOF) m/z: [M + H]+

+

Calcd for C H N O 389.1614, found 389.1952 [M + H] .

22

21

4

3

4.1.6.22.

3-(benzo[d][1,3]dioxol-5-yl)-5-((4-(3,5-dimethoxyphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (28).

o

White solid (20.1 mg, 0.05 mmol, 60% yield). M.P: 100-101 C. R.f: 0.3

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3118, 2839, 1741, 1591, 1517, 1462, 1357,

-1 1

240, 1157, 1041, 939, 869, 808, 773, 682 cm . H NMR (500 MHz, DMSO‑ d ) δ 8.76

6

(

s, 1H, H-triazole), 7.44 – 7.41 (m, 3H, Ar), 7.09 (s, 1H, H-isoxazol), 7.07 (d, J= 2.3 Hz,

2

5

1

4

H, Ar), 7.05 (d, J= 8.5 Hz, 1H, Ar), 6.49 (t, J= 2.3 Hz, 1H, Ar), 6.11 (s, 2H, O-CH -O),

2

13

.97 (s, 2H, CH ), 3.81 (s, 6H, Ar-OCH ). C NMR (125 MHz, DMSO‑ d ) δ 166.6,

2

3

6

61.9, 160.9, 149.0, 148.0, 146.7, 132.2, 122.4, 121.9, 121.2, 108.8, 106.5, 103.2,

02.2, 101.6, 100.1, 55.3, 44.7. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O

07.1355, found 407.1348 [M + H] .

21

18

4

5

+

4.1.6.23. 3-(3,4-dimethoxyphenyl)-5-((4-(4-methoxyphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (29).

o

White solid (23.0 mg, 0.06mmol, 62% yield). M.P: 135–136 C. R.f: 0.13

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2833, 1608, 1579, 1529, 1465, 1247, 1139,

-1

1

031, 975, 858, 821, 763 cm . H NMR (400 MHz, CDCl ) δ 7.85 (s, 1H, H-triazole),

3

7.77 (d, J= 8.8 Hz, 2H, Ar), 7.37 (d, J= 1.7 Hz, 1H, Ar), 7.25 (dd, J= 8.4, 1.7 Hz, 1H,

Ar), 6.97 (d, J= 8.8 Hz, 2H, Ar), 6.92 (1H, d, J=8.4 Hz), 6.56 (s,1H, H-isoxazole), 5.76

13

(

s, 2H, CH ), 3.94 (s, 3H, Ar-OCH ), 3.93 (s, 3H, Ar-OCH ), 3.85 (s, 3H, Ar-OCH ). C

2 3 3 3

NMR (100 MHz, CDCl ) δ 165.5, 162.8, 160.0, 151.2, 149.6, 148.7, 127.3, 122.9,

3

1

21.0, 120.2, 119.2, 114.5, 111.3, 109.5, 102.2, 56.2, 56.1, 55.5, 45.5. HRMS (ESI-

+

TOF) m/z: [M + H]+ Calcd for C H N O 393.1563, found 393.1975 [M + H] .

21

4

4.1.6.24. 3-(3,4-dimethoxyphenyl)-5-((4-(4-(trifluoromethoxy)phenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (30).

o

White solid (38.0 mg, 0.08 mmol, 89% yield). M.P: 148–149 C. R.f: 0.15

[

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2837, 1606, 1587, 1529, 1473, 1276, 1159,

-1 1

1

8

1

026, 854, 806 cm . H NMR (400 MHz, CDCl ) δ 7.95 (s, 1H, H-triazole), 7.87 (d, J=

3

.8 Hz, 2H, Ar), 7.37 (d, J= 1.9 Hz, 1H, Ar), 7.28 (d, J= 8.8 Hz, 2H, Ar), 7.26 (dd, J=8.4,

.9 Hz, 1H, Ar), 6.92 (d, J= 8.4 Hz, 1H, Ar), 6.60 (s, 1H, H-isoxazole), 5.78 (s, 2H,

1

3

CH ), 3.94 (s, 3H, Ar-OCH ), 3.93 (s, 3H, Ar-OCH ). C NMR (100 MHz, CDCl ) δ

1

2

3

1

65.1, 162.9, 151.2, 149.6, 149.4, 147.6, 129.0, 127.4, 121.6, 120.9, 120.7 (d, J_C_F

=

2

57.4 Hz),120.3, 120.2, 111.3, 109.4, 102.4, 56.2, 56.1, 45.5. HRMS (ESI-TOF) m/z:

+

[

M + H]+ Calcd for C H F N O 447.1280, found 447.1659 [M + H] .

21 18 3 4 4

4.1.6.25.

3-(3,4-dimethoxyphenyl)-5-((4-(4-pentylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (31).

o

White solid (39.3 mg, 0.09 mmol, 94% yield). M.P: 78–79 C. R.f: 0.12 [Hexane:

EtOAc, 60:40 % (v/v)]. IR (KBr): 2852, 1747, 1604, 1585, 1529, 1471, 1433, 1230,

-1 1

1

147, 854, 820 cm . H NMR (300 MHz, CDCl ) δ 7.84 (s, 1H, H-triazole), 7.67 (d, J=

3

8.1 Hz, 2H, Ar), 7.29 (d, J= 1.9 Hz, 1H, Ar), 7.18 (dd, J= 8.3, 1.9 Hz, 1H, Ar), 7.16 (d,

J= 8.1 Hz, 1H, Ar), 6.84 (d, J= 8.3 Hz, 1H, Ar), 6.48 (s, 1H, H-isoxazole), 5.69 (s, 2H,

CH ), 3.86 (Ar-OCH ), 3.85 (Ar-OCH ), 2.60 – 2.51 (m, 2H, CH ), 1.61 – 1.50 (m, 2H,

2

3

2

13

CH ), 1.32 – 1.18 (m, 4H, CH ), 0.82 (t, J= 6.8 Hz, 3H, CH ). C NMR (100 MHz,

2

3

S18

CDCl ) δ 165.5, 162.8, 151.1, 149.6, 149.0, 143.7, 129.1, 127.6, 125.9, 121.0, 120.2,

3

1

19.7, 111.3, 109.5, 102.2, 56.2, 56.1, 45.6, 35.9, 31.6, 31.2, 22.7, 14.1. HRMS (ESI-

+

TOF) m/z: [M + H]+ Calcd for C H N O 433.2240, found 433.2650 [M + H] .

25

29

4

3

4.1.6.26.

4-(1-((3-(3,4-dimethoxyphenyl)isoxazol-5-yl)methyl)-1H-1,2,3-triazol-4-

yl)aniline (32).

o

White solid (18.0 mg, 0.05mmol, 50% yield). M.P:167–168 C. R.f: 0.12

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3325, 3062, 1651, 1585, 1438, 1359, 1280,

-1

1

178, 960, 837, 819 cm . H NMR (300 MHz, CDCl ) δ 7.79 (s, 1H, H-triazole), 7.63

3

(

2

1

d, J= 8.5 Hz, 2H, Ar), 7.36 (d, J= 1.8 Hz, 1H, Ar), 7.25 (dd, J=8.4, 1.8 Hz, 2H, Ar), 6.91

d, J= 8.4 Hz, 1H, Ar), 6.73 (d, J= 8.5 Hz, 2H, Ar), 6.53 (s, 1H, H-isoxazole), 5.73 (s,

13

H, CH ), 3.93 (s, 3H, Ar-OCH ), 3.92 (s, 3H, Ar-OCH ). C NMR (100 MHz, CDCl ) δ

2 3 3 3

65.5, 162.7, 151.0, 149.4, 149.1, 146.8, 127.1, 120.9, 120.5, 120.1, 118.5, 115.2,

11.2, 109.3, 102.0, 56.1, 56.0, 45.4, HRMS (ESI-TOF) m/z: [M + H]+ Calcd for

+

C H N O 378.1566, found 378.1904 [M + H] .

20

5

3

4.1.6.27.

(4-(1-((3-(3,4-dimethoxyphenyl)isoxazol-5-yl)methyl)-1H-1,2,3-triazol-4-

yl)phenyl)methanol (33).

o

White solid (32.6 mg, 0.08 mmol, 87% yield). M.P: 109–110 C. R.f:

0.10

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3560, 2853, 1614, 1587, 1529, 1458, 1433,

-1

1

230, 1155, 1022, 900, 839, 800 cm . H NMR (400 MHz, CDCl ) δ 7.94 (s, 1H, H-

3

triazole), 7.82 (d, J= 8.0 Hz, 2H, Ar), 7.43 (d, J= 8.0 Hz, 2H, Ar), 7.36 (d, J= 1.8 Hz, 1H,

Ar), 7.25 (dd, J= 8.3, 1.8 Hz, 1H, Ar), 6.91 (d, J= 8.3 Hz, 1H, Ar), 6.58 (s, 1H, H-

isoxazole), 5.76 (s, 2H, CH ), 4.73 (s, 2H, CH ), 3.93 (s, 3H, Ar-OCH ), 3.92 (s, 3H, Ar-

2

3

13

OCH ). C NMR (100 MHz, CDCl ) δ 165.3, 162.8, 151.1, 149.5, 148.5, 141.7, 129.0,

3

1

27.5, 125.9, 120.8, 120.2, 111.3, 109.4, 102.3, 64.5, 56.1, 56.0, 45.4. HRMS (ESI-

+

TOF) m/z: [M + H]+ Calcd for C H N O 393.1563, found 393.1993 [M + H] .

21

4

4.1.6.28. 3-(3,4-dimethoxyphenyl)-5-((4-phenyl-1H-1,2,3-triazol-1-yl)methyl)isoxazole

(

34).

o

Light yellow solid (32.0 mg, 0.09 mmol, 92% yield. M.P: 158–159 C. R.f: 0.15

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2839, 1602, 1587, 1529, 1421, 1230, 1138,

-1 1

018, 902, 850, 763 cm . H NMR (400 MHz, CDCl ) δ 7.94 (s, 1H, H-triazole), 7.84 (

3

dd, J= 8.0, 1.3 Hz, 2H, Ar), 7.44 (dd, J= 7.58, 1.3 Hz, 3H, Ar), 7.37 (d, J= 2.0 Hz, 1H,

Ar), 7.37 (dd, J= 8.3, 2.0 Hz, 2H, Ar), 6.92 (d, J= 8.3 Hz, 1H, Ar), 6.57 (s, 1H, H-

13

isoxazole), 5.78 (s, 2H, CH ), 3.94 (s, 3H, Ar-OCH ), 3.93 (s, 3H, Ar-OCH ). C NMR

2

3

(

100 MHz, CDCl ) δ 165.4, 162.83 151.2, 149.6, 148.9, 130.2, 129.1, 128.7, 126.0,

3

1

21.0, 120.2, 120.0, 111.3, 109.5, 102.3, 56.2, 56.1, 45.6. HRMS (ESI-TOF) m/z: [M +

+

H]+ Calcd for C H N O 363.1457, found 362.1828 [M + H] .

20

19

4

3

4.1.6.29. 3-(3,4-dimethoxyphenyl)-5-((4-(p-tolyl)-1H-1,2,3-triazol-1-yl)methyl)isoxazole

(

35).

o

Light yellow solid (33.8 mg, 0.09 mmol, 93% yield). M.P: 176–177 C. R.f: 0.18

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2841, 1604, 1585, 1527, 1421, 1265, 1228,

-1

1

139, 1018, 906, 854, 815, 765 cm . H NMR (400 MHz, CDCl ) δ 7.90 (s, 1H, H-

3

triazole), 7.72 (d, J= 8.0 Hz, 2H, Ar), 7.36 (d, J= 1.65 Hz, 1H, Ar), 7.27 – 7.21 (m, 3H,

Ar), 6.90 (d, J= 8.3 Hz, 1H, Ar), 6.56 (s, 1H, H-isoxazole), 3.93 (s, 3H, Ar-OCH ), 3.91

3

1

3

(

s, 3H, Ar-OCH ), 2.37 (s, 3H, Ar-CH ). C NMR (100 MHz, CDCl ) δ 165.5, 162.8,

3 3 3

1

3

51.1, 149.6, 148.9, 138.5, 129.7, 127.4, 125.9, 121.0, 120.2, 119.7, 111.3, 109.4,

02.2, 56.2, 56.1, 45.5, 21.4, HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O

21

4

3

+

77.1614, found 377.1981 [M + H] .

S19

4.1.6.30.

3-(3,4-dimethoxyphenyl)-5-((4-(2-methoxyphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (36).

o

Light yellow solid (36.0 mg, 0.09 mmol, 95% yield). M.P: 136–137 C. R.f: 0.32

[

1

Hexane: EtOAc, 60:40 % (v/v)]. IR (KBr): 2827, 1622, 1583, 1527, 1477, 1435, 1257,

-1

1

163, 1033, 833,796, 742 cm . H NMR (400 MHz, CDCl ) δ 8.26 (dd, J=7.8, 1.8

3

Hz,1H, Ar), 8.12 (s, 1H, H-triazole), 7.26 (d, J= 1.8 Hz, 1H, Ar), 7.22 (dd, J= 8.3, 1.6 H,

1

8

3

H, Ar), 7.15 (dd, J= 8.4, 1.6 Hz, 1H, Ar), 6.99 (1H, dd, J= 8.3, 7.4 Hz), 6.91 (1H, d, J=

.4 Hz), 6.83 (1H, d, J= 8.3 Hz), 6.44 (s, 1H, H-isoxazole), 5.69 (s, 2H, CH ), 3.87 (s,

2

13

H, Ar-OCH ), 3.86 (s, 3H, Ar-OCH ), 3.84 (s, 3H, Ar-OCH ). C NMR (100 MHz,

3

CDCl ) δ 166.0, 162.8, 155.9, 151.1, 149.6, 144.2, 129.4, 127.8, 123.5, 121.2, 121.1,

3

1

20.2, 119.1, 111.3, 111.0, 109.5, 101.9, 56.2, 56.1, 55.6, 45.5. HRMS (ESI-TOF) m/z:

+

[

M + H]+ Calcd for C H N O 393.1563, found 393.1855 [M + H] .

21 21 4 4

4.1.6.31. 3-(3,4-dimethoxyphenyl)-5-((4-(4-methoxy-2-methylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (37).

o

Light yellow solid (32.2 mg, 0.08 mmol, 82% yield). M.P.: 124–125 C. R.f: 0.10

[

1

Hexane: EtOAc, 60:40 % (v/v)]. IR (KBr): 2839, 1606, 1583, 1527, 1444, 1419, 1267,

-1

1

134, 1039, 1022, 850, 825, 796 cm . H NMR (400 MHz, CDCl ) δ 7.77 (s, 1H, H-

3

triazole), 7.70 (d, J= 7.4 Hz, 1H, Ar), 7.38 (d, J= 1.9 Hz, 1H, Ar), 7.26 (dd, J= 8.3, 1.9

Hz, 1H, Ar), 6.92 (d, J= 8.3 Hz, 1H, Ar), 6.84-6.81 (m, 2H, Ar), 6.58 (s, 1H, H-

isoxazole), 5.77 (s, 2H, CH ), 3.94 (s, 3H, Ar-OCH ), 3.93 (s, 3H, Ar-OCH ), 3.83 (s,

2

3

13

3

1

H, Ar-OCH ), 2.45 (s, 3H, Ar-CH ). C NMR (100 MHz, CDCl ) δ 165.5, 162.8, 159.8,

3 3 3

51.1, 149.6, 148.0, 137.4, 130.4, 122.3, 121.5, 121.0, 120.2, 116.5, 111.7, 111.3,

09.4, 102.2, 56.2, 56.1, 55.4, 45.5, 21.7. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for

+

C H N O 407.1719, found 407.2078 [M + H] .

22

23

4

4.1.6.32.

3-(3,4-dimethoxyphenyl)-5-((4-(2,5-dimethylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (38).

o

White solid (36.5 mg, 0.09 mmol, 97% yield). M.P.: 163-164 C. R.f: 0.17 [Hexane:

EtOAc, 60:40 % (v/v)]. IR (KBr): 2837, 1610, 1581, 1523, 1431, 1232, 1020, 902, 817,

-1 1

7

79 cm . H NMR (300 MHz, CDCl ) δ 7.83 (s, 1H, H-triazole), 7.63 (d, J= 1.2 Hz, 1H,

3

Ar), 7.38 (d, J= 2.0 Hz, 1H, Ar), 7.26 (dd, J= 8.3, 2.0 Hz, 1H, Ar), 7.17 (d, J= 7.8 Hz,1H,

Ar), 7.09 (dd, J= 7.8, 1.2 Hz,1H, Ar), 6.92 (d, J= 8.3 Hz, 1H, Ar), 6.58 (s, 1H, H-

isoxazole), 5.79 (s, 2H, CH ), 3.94 (s, 3H, Ar-OCH ), 3.93 (s, 3H, Ar-OCH ), 2.43 (s,

2

3

13

3

1

H, Ar-CH ), 2.36 (s, 3H, Ar-CH ). C NMR (100 MHz, CDCl ) δ 165.5, 162.8, 151.16,

3 3 3

49.6, 148.2, 135.8, 132.5, 131.0, 129.6, 129.3, 129.3, 122.1, 121.0, 120.2, 111.3,

09.4, 102.2, 56.2, 56.1, 45.5, 21.0, 21.0. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for

+

C H N O 391.1770, found 391.2101[M + H] .

22

23

4

3

4.1.6.33.

3-(3,4-dimethoxyphenyl)-5-((4-(4-nitrophenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (39).

o

Yellow solid (30.5 mg, 0.07 mmol, 78% yield). M.P: 167–168 C. R.f: 0.07 [Hexane:

EtOAc, 60:40 % (v/v)]. IR (KBr): 2839, 1604, 1590, 1527, 1514, 1431, 1336, 1267,

-

1 1

1

2

7

5

141, 1024, 908, 854, 804, 767 cm . H NMR (300 MHz, CDCl ) δ 8.24 (d, J= 8.9 Hz,

3

H, Ar), 8.03 (s, 1H, H-triazole), 7.95 (d, J= 8.9 Hz, 2H, Ar), 7.31 (d, J= 1.9 Hz, 1H, Ar),

.20 (dd, J= 8.3, 1.9 Hz, 2H, Ar), 6.86 (d, J= 8.3 Hz, 1H, Ar), 6.58 (s, 1H, H-isoxazol),

.74 (s, 2H, CH ), 3.87 (s, 3H, Ar-OCH ), 3.86 (s, 3H, Ar-OCH ), C NMR (100 MHz,

13

2

3

CDCl ) δ 164.9, 162.8, 151.0, 149.3, 147.5, 146.3, 136.2, 126.3, 124.3, 122.1, 120.6,

3

S20

1

20.2, 111.2, 109.3, 102.5, 55.9, 55.8, 45.3. HRMS (ESI-TOF) m/z: [M + H]+ Calcd

+

for C H N O 408.1308, found 408.1751 [M + H] ,

20

18

5

4.1.6.34.

3-(3,4-dimethoxyphenyl)-5-((4-(4-fluorophenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (40).

o

White solid (32.3 mg, 0.08 mmol, 88% yield). M.P: 173–174 C. R.f: 0.12

[

Hexane:EtOAc, 60:40 % (v/v)], IR (KBr): 2839, 1604, 1587, 1529, 1496, 1421, 1232,

-1 1

1

7

2

1

4

3

138, 1028, 898, 831, 767 cm . H NMR (300 MHz, CDCl ) δ 7.91 (s, 1H, H-triazole),

3

.83 – 7.78 (m, 2H, Ar), 7.37 (d, J= 1.8 Hz, 1H, Ar), 7.26 (dd, J= 8.3, 1.8 Hz, 1H, Ar),

.15 – 7.09 (m, 2H, Ar), 6.91 (d, J= 8.3 Hz, 1H, Ar), 6.59 (s, 1H, H-isoxazole), 5.76 (s,

1

3

H, CH ), 3.93 (s, 3H, Ar-OCH ), 3.92 (s, 3H, Ar-OCH ). C NMR (125 MHz, CDCl ) δ

2

3

1

3

65.2, 162.9 (d, J = 255.2 Hz), 162.8, 151.1, 149.5, 147.9, 127.7 (d, J = 8.2 Hz),

26.2, 120.8, 120.2, 120.0, 116.0 (d, J_C_F= 22.0 Hz), 111.3, 109.4, 102.4, 56.1, 56.0,

CF

2

5.4. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H FN O 381.1363, found

20

18

4

3

+

81.1761 [M + H] .

4.1.6.35. 4-(1-((3-(3,4-dimethoxyphenyl)isoxazol-5-yl)methyl)-1H-1,2,3-triazol-4-yl)-

N,N-dimethylaniline (41).

o

White solid (25.5 mg, 0.062 mmol, 65% yield). M.P: 164–165 C. R.f: 0.07

[

8

Hexane:EtOAc, 60:40 % (v/v)], IR (KBr): 2806, 1612, 1556, 1525, 1506, 1261, 1024,

-1 1

54, 813 cm . H NMR (300 MHz, CDCl ) δ 7.81 (s, 1H, H-triazole), 7.72 (d, J= 8.8 Hz,

3

2H, Ar), 7.36 (d, J= 1.6 Hz, 2H, Ar), 7.25 (dd, J= 8.3, 1.6 Hz, 2H, Ar), 6.91 (d, J= 8.8

Hz, 2H, Ar), 6.84 (d, J= 8.3 Hz, 2H, Ar), 6.54 (s, 1H, H-isoxazole), 5.73 (s, 2H, CH ),

2

13

3

.93 (s, 3H, Ar-OCH ), 3.92 (s, 3H, Ar-OCH ), 3.01 (s, 6H, N-CH ). C NMR (100 MHz,

3 3 3

CDCl ) δ 165.8, 162.8, 151.1, 150.8, 149.6, 149.4, 126.9, 121.1, 120.2, 118.5, 118.4,

3

1

12.6, 111.3, 109.5, 102.1, 56.2, 56.1, 45.5, 40.6. HRMS (ESI-TOF) m/z: [M + H]+

+

Calcd for C H N O 406.1879, found 406.2239 [M + H] .

22

24

5

3

4.1.6.36. 3-(3,4-dimethoxyphenyl)-5-((4-(6-methoxynaphthalen-2-yl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (42).

o

Light yellow solid (39.3 mg, 0.09 mmol, 92% yield). M.P: 185–186 C. R.f: 0.30

EtOAc: Hexane 60:40% (v/v)]. IR (KBr): 28843, 1604, 1587, 1527, 1467, 1232, 1147,

[

-1 1

1

020, 893, 864, 823 cm . H NMR (300 MHz, CDCl ) δ 8.26 (s, 1H, Ar), 8.00 (s, 1H, H-

3

triazole), 7.88 (dd, J=8.4, 1.6 Hz, 1H, Ar), 7.77 (dd, J= 8.7, 3.7 Hz, 2H, Ar), 7.37 (d, J=

1.8 Hz, 1H, Ar), 7.25 (dd, J= 8.3, 1.8 Hz, 1H, Ar), 7.16 (dd,J= 8.7, 2.5 Hz, 1H, Ar), 7.15

–

7.12 (m, 1H, Ar), 6.90 (d, J= 8.3 Hz, 1H, Ar), 6.58 (s, 1H, H-isoxazole), 5.77 (s, 2H,

13

CH ), 3.93 (s, 6H, Ar-OCH ), 3.91 (s, 3H, Ar-OCH ). C NMR (100 MHz, CDCl ) δ

2

3

1

4

65.5, 162.9, 158.3, 151.2, 149.6, 149.1, 134.7, 129.9, 129.1, 127.6, 125.4, 124.7,

24.4, 121.0, 120.3, 119.9, 119.6, 111.3, 109.5, 106.0, 102.3, 100.1, 56.2, 56.1, 55.5,

5.6. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 443.1719, found

25

23

4

+

43.2177 [M + H] .

4.1.6.37.

3-(4-methoxyphenyl)-5-((4-(4-methoxyphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (43).

o

White solid (89% yield). Mp: 196.0-198.0 C. IR (KBr): 2835, 1614, 1577, 1529, 1435,

-1 1

1

251,1180, 1029, 827 cm . H NMR (400 MHz, CDCl ): δ = 7.85 (s,1H, triazole), 7.76

3

(

d, J= 8.8 Hz, 2H, Ar), 7.71 (d, J= 8.8 Hz, 2H, Ar), 6.96 (d, J= 8.8 Hz, 4H, Ar), 6.55

13

s,1H, isoxazole), 5.75 (s, 2H, CH ), 3.85 (s, 3H, Ar-OCH ), 3.85 (s, 3H, Ar-OCH ). C

2

3

NMR (100 MHz, CDCl ) δ 165.4, 162.8, 161.5, 160.0, 148.6, 128.4, 127.3, 122.6,

3

1

20.6, 119.8, 114.6, 114.6, 102.4, 55.5, 55.4, 45.4. HRMS (ESI-TOF) m/z: [M + H]+

+

Calcd for C H N O 363.1457, found 363.1670 [M + H] .

20

19

4

3

S21

4.1.6.38.

3-(benzo[d][1,3]dioxol-5-yl)-5-((4-(4-methoxyphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (44).

o

White solid (19.3 mg, 0.05 mmol, 85% yield). M.P: 181-182 C. R.f: 0.28

[

9

Hexane:EtOAc, 60:40 % (v/v)], IR (KBr): 3124, 2835, 1604, 1560, 1516, 1454, 1247,

-1

1

35, 871, 831, 813, 781, 742 cm . H NMR (400 MHz, CDCl ) δ 7.84 (s, 1H, H-

3

triazole), 7.76 (d, J= 8.8 Hz, 2H, Ar), 7.29 (d, J= 1.6 Hz, 1H, Ar), 7.22 (dd, J= 8.1, 1.6

Hz, 1H, Ar), 6.96 (d, J= 8.8 Hz, 2H, Ar), 6.86 (d, J= 8.1 Hz, 1H, Ar), 6.52 (s, 1H, H-

13

isoxazole), 6.02 (s, 2H, O-CH -O), 5.74 (s, 2H, CH ), 3.84 (s, 3H, Ar-OCH ), C NMR

2

3

(

100 MHz, CDCl ) δ 165.5, 162.7, 160.0, 149.7, 148.7, 148.5, 127.3, 122.9, 122.3,

3

1

+

21.5, 119.2, 114.5, 108.8, 107.0, 102.2, 101.7, 55.5, 45.5, HRMS (ESI-TOF) m/z: [M

+

H]+ Calcd for C H N O 377.1250, found 377.1409 [M + H] .

20

17

4

4.1.6.39.

3-(4-methoxyphenyl)-5-((4-(4-(trifluoromethoxy)phenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (45).

o

White solid (42.6 mg, 0.10 mmol, 94% yield). M.P: 191–192 C. R.f: 0.32

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 2841, 1612, 1575, 1529, 1435, 1282, 1215,

-1 1

161, 1031, 833 cm . H NMR (300 MHz, CDCl ) δ 7.95 (s, 1H, H-triazole), 7.86 (d, J=

3

8

.8 Hz, 2H, Ar), 7.71 (d, J= 8.9 Hz, 2H, Ar), 7.29 (d, J= 8.8 Hz, 2H, Ar), 6.96 (d, J=8.9

13

Hz, 2H, Ar), 6.58 (s, 1H, H-isoxazole), 5.77 (s, 2H, CH ), 3.85 (s, 3H, Ar-OCH ).

NMR (125 MHz, CDCl ) δ 165.0, 162.7, 161.4, 149.25 (d, J = 1.5 Hz), 147.3, 128.7,

C

2

3

2

3

CF

1

128.3, 127.3, 121.4, 120.6, 120.5, 120.4 (d, J_C_F= 256.3 Hz), 114.5, 102.3, 55.3, 45.3.

HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H F N O 417.1175, found 417.1481 [M

20

16

3

4

3

+

H] .

4.1.6.40. 3-(benzo[d][1,3]dioxol-5-yl)-5-((4-(4-(trifluoromethoxy)phenyl)-1H-1,2,3-triazol-

1-yl)methyl)isoxazole (46).

o

White solid (26.0 mg, 0.06 mmol, 98% yield). M.P: 166-167 C. R.f: 0.43

[

8

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3115, 1608, 1517, 1350, 1255, 1039, 846,

-1 1

17, 756 cm . H NMR (300 MHz, CDCl ) δ 7.95 (s, 1H, H-triazol), 7.87 (d, J= 8.5 Hz,

3

2H, Ar), 7.29 (d, J= 1.2 Hz, 1H, Ar), 7.28 (d, J= 8.5 Hz, 2H, Ar), 7.23 (dd, J= 8.1, 1.2

Hz, 1H, Ar), 6.87 (d, J= 8.1 Hz,1H, Ar), 6.55 (s, 1H, H-isoxazole), 6.03 (s, 2H, O-CH -

2

13

O), 5.77 (s, 2H, CH ). C NMR (125 MHz, MeOD) δ 167.7, 164.1, 163.8, 151.0, 149.9,

30.9, 128.4, 123.2, 122.7, 122.6, 122.0 (d, J_C_F= 258.8 Hz), 109.8, 107.7, 103.3,

03.1, 46.1. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H F N O 431.0967, found

2

1

4

20

14

3

4

+

31.1271 [M + H] .

4.1.6.41.

3-(4-methoxyphenyl)-5-((4-(4-pentylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (47).

Solid white solid (42,3 mg, 0,11 mmol, 96% yield), F.M.: C H N O M.M.: 402,49

2

4

26

4

2,

o

g/mol, M.P:150–151 C, R.f: 0,46 [Hexane:EtOAc, 60:40 % (v/v)], IR (KBr): 2848,

614, 1577, 1529, 1440, 1251, 1031, 819, 532 cm , NMR H (300 MHz, CDCl ) δ 7.90

-1

1

3

(

8

s, 1H, H-triazole), 7.74 (d, J= 8.3 Hz, 2H, Ar), 7.70 (d, J= 8.9 Hz, 2H, Ar), 7.24 (d, J=

.3 Hz, 2H, Ar), 6.96 (d, J= 8.9 Hz, 2H, Ar), 6.54 (s, 1H, H-isoxazole), 5.75 (s, 2H,

CH ), 3.85 (s, 3H, Ar-OCH ), 2.67 – 2.57 (m, 2H, Ar-CH ), 1.70 – 1.57 (m, 2H, CH ),

2

3

2

13

1

.38 – 1.27 (m, 4H, CH -CH ), 0.89 (t, J=6.8 Hz, 3H, CH -CH ). NMR C (100 MHz,

2 2 2 3

CDCl ) δ 165.4, 162.7, 161.5, 149.0, 143.6, 129.1, 128.4, 127.6, 125.9, 120.8, 119.6,

3

1

14.6, 102.2, 55.5, 45.5, 35.9, 31.6, 31.2, 22.7, 14.2, HRMS (ESI-TOF) m/z: [M + H]+

+

Calcd for C H N O 403,2134, found 403,2473 [M + H] .

24

27

4

2

S22

4.1.6.42.

3-(benzo[d][1,3]dioxol-5-yl)-5-((4-(4-pentylphenyl)-1H-1,2,3-triazol-1-

yl)methyl)isoxazole (48).

o

White solid (31.0 mg, 0.07 mmol, 91% yield). M.P.: 165-166 C. R.f: 0.54

[

1

Hexane:EtOAc, 60:40 % (v/v)]. IR (KBr): 3124, 2854, 1602, 1517, 1488, 1469, 1340,

-1 1

242, 1043, 933, 871, 815, 781 cm . H NMR (500 MHz, DMSO‑ d ) δ 8.65 (s, 1H, H-

6

triazole), 7.77 (d, J= 8.1 Hz, 2H, Ar), 7.41 (d, J= 1.7Hz, 1H, Ar), 7.41 (dd, J= 8.5, 1.7

Hz, 1H, Ar), 7.27 (d, J= 8.1 Hz, 2H, Ar), 7.06 (s, 1H, H-isoxazole), 7.03 (d, J= 8.5 Hz,

1

H, Ar), 6.10 (s, 2H, O-CH -O), 5.95 (s, 2H, CH ), 2.62 – 2.57 (m, 2H, CH ), 1.62 –

2 2 2

13

1.55 (m, 2H, CH ), 1.34 – 1.23 (m, 4H, CH -CH ), 0.86 (t, J= 7.0 Hz, 3H, CH -CH ). C

2

3

NMR (125 MHz, DMSO‑ d ) δ 166.7, 161.8, 149.0, 148.0, 146.9, 142.3, 128.8, 127.9,

6

125.2, 121.9, 121.6, 121.2, 108.8, 106.5, 102.1, 101.6, 44.7, 34.8, 30.8, 30.5, 21.9,

1

3.9. HRMS (ESI-TOF) m/z: [M + H]+ Calcd for C H N O 417.1927, found 417.2254

24

25

4

3

+

[

M + H] .

4.2. Biological evaluation

4.2.1. Screening against T. cruzi amastigote

The in vitro tripanocidal activity in amastigote forms of T. cruzi was evaluated by

37

a colorimetric beta-galactosidase assay . The β-galactosidase T. cruzi, Tulahuen

strain was provided by the Laboratory of Cellular and Molecular Parasitology, Centro

de Pesquisas René Rachou, FIOCRUZ, Belo Horizonte. Culture-derived

trypomastigotes raised from infected L929 cell line were used to infect differentiated

6

human THP-1 derived macrophage (4.0x10 cells/well) in 96-well microplates in a

38

parasite/cell ratio of 2:1 and incubated overnight at 37ºC and 5% CO₂. The medium

containing non-internalized parasites was removed and replaced with 180 µl of fresh

complete medium. Infected cell layer were treated by addition of 20.00 μl of each

sample (2 µM of DMSO-diluted stock solution in 18 µM of RPMI-1640), in triplicate,

with the compounds solubilized in dimethyl sulfoxide (DMSO) diluted at 100 µM,

followed by incubation for 72h at 37°C and 5% CO . After treatment, cells were

2

carefully washed with PBS and incubated for 4h at 37°C with 250 µl of

Chlorophenolred-ß-D-galactopyranoside (Sigma-Aldrich Co., St. Louis, MO, USA)

(

CPRG) at 100 µM and Nonidet P-40 (Amresco Inc, Solon, Ohio, USA) (NP-40) 0,1%.

The absorbance was measured at 570 nm and reference at 630 nm in an automated

microplate reader. Benznidazole (Sigma Aldrich) was used as positive control. The

results are expressed as percentage of parasite growth inhibition.

4.2.2. Screening against Leishmania amazonensis amastigote

For the leishmanicidal screening against intracellular L. amazonensis

amastigotes was used colorimetric methodology. THP-1 (ATCC TIB202) cells

4

(

4.0 × 10 per well) were cultivated in 96 well plates in RPMI-1640 medium without

phenol red (Sigma-Aldrich, CO. St. Louis, MO, USA) supplemented with 10% FBS (Life

Technologies, USA), 12.5 mM HEPES, penicillin (100 U/ml), streptomycin (100 μg/ml),

sodium pyruvate (1 mM) (Gibco) and Glutamax (2 mM) (Gibco) and treated with

100 ng/ml of phorbol 12-myristate 13-acetate (PMA, Sigma-Aldrich Co., St. Louis, MO,

USA ) for 72 h at 34 °C in a 5% CO , to allow THP-1 cells differentiation into non-

2

38

dividing macrophages . L. amazonensis MHOM/BR/77/LTB0016 promastigotes,

expressing β-galactosidase, were grown at 26 °C in Schneider's insect medium (Sigma

Chemical Co., St. Louis, MO, USA) supplemented with 5% heat inactivated fetal bovine

serum FBS and 2% of human urine. Four days old culture promastigotes

6

(

40.0 × 10 parasites/ml) were washed with phosphate buffered saline, pH 7.4 (PBS)

S23

an incubated in RPMI-1640 supplemented with 10% human B+ serum heat-inactivated

for 1 h at 34 °C to parasite opsonization. THP-1 cells were incubated with a

parasite/cell ratio of 10:1 for 3 h at 34 °C and 5% CO . After this period, non-adherent

2

parasites were removed by one wash with PBS and infected cells were incubated with

180 μl of full supplemented RPMI-1640 medium for another 24 h to allow the

transformation of promastigotes into intracellular amastigotes. The infected cells were

treated with 20.00 µl of each compound (2 µM of DMSO-diluted stock solution in 18 µM

of RPMI-1640) in triplicate. After treatment, cells were carefully washed with PBS and

incubated for 4 h at 37 °C with 250 μl of chlorophenolred-ß-D-galactopyranoside

(

Sigma–Aldrich Co., St. Louis, MO, USA) (CPRG) at 100 μM and Nonidet P-40

Amresco Inc, Solon, Ohio, USA) (NP-40) 0.1%. Optical density was read at

570/630 nm in an Infinite M200 TECAN, Austria. Amphotericin B (2 µM) (Sigma) was

used as positive control and DMSO 1% as negative control.

4.2.3. Human THP-1-derived macrophages

THP-1 cells (human monocytic leukaemia cell line) were grown and cultivated in

6

9

6 well plates (4.0 × 10 cells/well), as described above, treated with the compounds

(

7

diluted in DMSO) in concentrations ranging from 15.6 M to 500 M and incubated for

2 h (37 °C, 5% CO ). Cell viability was assessed by the MTT assay, which consists in

2

the colorimetric measurement of the metabolization of 3-(4,5-dimethylthiazol-2-yl)-2,5-

diphenyltetrazolium bromide (MTT) to formazan by viable cells. DMSO 1% was used

as negative control and the optical density was read at 540 nm in an Infinite M200

TECAN microplate reader immediately after the dissolution of formazan crystals with

DMSO.

4.2.4. Trypanothione reductase enzyme assay

Trypanothione reductase from T. cruzi (TcTR), was expressed in Escherichia coli

BL21DE3 and purified by affinity chromatography. TryR (1 m-unit), HEPES (40 mM, pH

7.5), NADPH (0.15 mM), DTNB (25 M) and EDTA (1 mM) were incubated in 96 well

plates (final volume= 240 L) for 5 min (27 °C) before T(S) (1 M) and the tested

2

3

9

compound (diluted in DMSO) were added . Compounds and controls were pre-

o

incubated at 27 C for 30 min and 10 L of DTNB was added to the reaction mixture.

Following, absorbance at 412 nm was measured for 30 min to determine the enzymatic

activity. Clomipramine was used as positive control and DMSO 1% as negative control.

4.2. Structure-Activity Relationships Study

The structure-activity relationship (SAR) study was carried out using a three-

dimensional approach. For this to be possible, three-dimensional structures were built

in HyperChem 7 (Hyper Co.), and calculations of MM+ force field, followed by AM1

semi-empirical theory level, were carried out using the same software.

Field descriptors were obtained using the software Pentacle (Molecular

Discovery Ltd). This software uses the GRIND approach, that calculate 3D field

descriptors in an alignment-independent way. The descriptors were computed using an

30

combination of ALMOND and CLACC algorithms

Based in the approach used by Ermondi et al., a binary SAR study was carried

out. Compounds with inhibition less to 40%, or inactive (24), were set as -1, whereas

40

with inhibition greater or equal to 40% (18) were set as 1 . Pentacle generated DRY-

DRY (hydrophobic-hydrophobic groups), O-O (hydrogen bond acceptor-hydrogen bond

acceptor groups), N1-N1 (hydrogen bond donor-hydrogen bond donor groups), and

TIP-TIP (shape-shape groups) descriptors, and the combinations between these (DRY-

O, DRY-N1, DRY-TIP, O-N1, O-TIP, and N1-TIP). This initial set was reduced using

S24

30

the Fractional Factorial Design (FDD) variable selection method , and the model was

41,42

built by Partial Least Squares (PLS)

. In this step and a step subsequent,

descriptors were autoscaled, the most adequate data pre-processing approach for

43

QSAR studies . Thus, the quality of the model assessed based on its coefficient of

2

determination (R ), the root mean square error of calibration (RMSEC), the F-test, the

2

coefficient of determination of cross-validation (Q _L_O_O), and the root mean square error

31

of cross-validation (RMSECV) .

SUPPLEMENTARY MATERIAL

Supplementary data associated with this article can be found in

These data include experimental details and characterization data for the reported

compounds, NMR spectra, mass spectrometry and biological data (PDF)

AUTHOR INFORMATION

CORRESPONDING AUTHOR

E-mail address: l.bernardes@ufsc.br (L.S.C. Bernardes).

FUNDING SOURCES

This work was supported by the Brazilian agencies FAPESC/CNPq, (PRONEX, grant

number 2671/2012-9), CAPES, CNPq (Universal, grant number 407626/2016-6),

Fundação Araucária, and FAPERGS.

ACKNOWLEDGMENT

We gratefully acknowledge José Carlos Tomaz (FCFRP, USP, Brazil), Vinícius

Palaretti (FFLCH, USP, Brazil), Luis Otávio Zamoner (FCFRP, USP, Brazil), Louis

Pergaud Sandjo (CIF, UFSC, Brazil), and Ana Luísa Morotti for the spectral analyses.

REFERENCES

1.

Bhutta ZA, Sommerfeld J, Lassi ZS, Salam RA, Das JK. Global burden, distribution, and

interventions for infectious diseases of poverty. Infect Dis Poverty. 2014;3(1):21.

doi:10.1186/2049-9957-3-21.

2

.

WHO. Investing to Overcome the Global Impact of Neglected Tropical Diseases: Third

WHO Report on Neglected Diseases 2015. Geneva; 2015. doi:ISBN 978 92 4 156486 1.

Leal SM. In vitro Antileishmanial, Trypanocidal and Mammalian Cell Activities of Diverse

N,N’-Dihetaryl Substituted Diamines and Related Compounds. Sci Pharm.

3

2013;81(1):43-55. doi:10.3797/scipharm.1205-14.

4

.

No JH. Visceral leishmaniasis: Revisiting current treatments and approaches for future

discoveries. Acta Trop. 2016;155:113-123. doi:10.1016/j.actatropica.2015.12.016.

Scarim CB, Jornada DH, Chelucci RC, de Almeida L, dos Santos JL, Chung MC. Current

advances in drug discovery for Chagas disease. Eur J Med Chem. 2018;155:824-838.

doi:10.1016/j.ejmech.2018.06.040.

5

6

.

Newman DJ, Cragg GM. Natural Products as Sources of New Drugs from 1981 to 2014.

J Nat Prod. 2016;79(3):629-661. doi:10.1021/acs.jnatprod.5b01055.

Singh N, Mishra BB, Bajpai S, Singh RK, Tiwari VK. Natural product based leads to fight

7

against

leishmaniasis.

Bioorg

Med

Chem.

2014;22(1):18-45.

doi:10.1016/j.bmc.2013.11.048.

8

.

Izumi E, Ueda-Nakamura T, Dias Filho BP, Veiga Júnior VF, Nakamura CV. Natural

products and Chagas’ disease: a review of plant compounds studied for activity against

Trypanosoma cruzi. Nat Prod Rep. 2011;28(4):809. doi:10.1039/c0np00069h.

da Rosa R, de Moraes MH, Zimmermann LA, Schenkel EP, Steindel M, Bernardes LSC.

Design and synthesis of a new series of 3,5-disubstituted isoxazoles active against

Trypanosoma cruzi and Leishmania amazonensis. Eur J Med Chem. 2017;128:25-35.

doi:10.1016/j.ejmech.2017.01.029.

9

10.

Hartmann AP, de Carvalho MR, Bernardes LSC, et al. Synthesis and 2D-QSAR studies

S25

of neolignan-based diaryl-tetrahydrofuran and -furan analogues with remarkable activity

against Trypanosoma cruzi and assessment of the trypanothione reductase activity. Eur

J Med Chem. 2017;140:187-199. doi:10.1016/j.ejmech.2017.08.064.

11.

Bernardes LSC, Kato MJ, Albuquerque S, Carvalho I. Synthesis and trypanocidal activity

of 1,4-bis-(3,4,5-trimethoxy-phenyl)-1,4-butanediol and 1,4-bis-(3,4-dimethoxyphenyl)-

1,4-butanediol.

Bioorganic

Med

Chem.

2006;14(21):7075-7082.

doi:10.1016/j.bmc.2006.07.006.

1

2.

3.

Dua R, Shrivastava S, Sonwane SK, Srivastava SK. Pharmacological Significance of

Synthetic Heterocycles Scaffold: A Review. Adv Biol Res (Rennes). 2011;5(3):120-144.

Sysak A, Obmińska-Mrukowicz B. Isoxazole ring as a useful scaffold in a search for new

1

therapeutic

doi:10.1016/j.ejmech.2017.06.002.

Zhu J, Mo J, Lin H zhi, Chen Y, Sun H peng. The recent progress of isoxazole in

medicinal chemistry. Bioorganic Med Chem. 2018;26(12):3065-3075.

doi:10.1016/j.bmc.2018.05.013.

Bonandi E, Christodoulou MS, Fumagalli G, Perdicchia D, Rastelli G, Passarella D. The

agents.

Eur

J

Med

Chem.

2017;137:292-309.

14.

15.

16.

1,2,3-triazole ring as a bioisostere in medicinal chemistry. Drug Discov Today.

2017;22(10):1572-1581. doi:10.1016/j.drudis.2017.05.014.

Schmidt A, Krauth-Siegel RL. Enzymes of the trypanothione metabolism as targets for

antitrypanosomal drug development. Curr Top Med Chem. 2002;2(11):1239-1259.

doi:10.2174/1568026023393048.

1

7.

8.

Krauth-Siegel RL, Inhoff O. Parasite-specific trypanothione reductase as a drug target

molecule. Parasitol Res. 2003;90(SUPPL. 2):S77-S85. doi:10.1007/s00436-002-0771-8.

Krauth-Siegel RL, Bauer H, Schirmer RH. Dithiol Proteins as Guardians of the

Intracellular Redox Milieu in Parasites: Old and New Drug Targets in Trypanosomes and

1

Malaria-Causing Plasmodia.

doi:10.1002/anie.200300639.

Angew Chemie

Int Ed.

2005;44(5):690-715.

1

9.

0.

Cavalli A, Bolognesi ML. Neglected Tropical Diseases: Multi-Target-Directed Ligands in

the Search for Novel Lead Candidates against Trypanosoma and Leishmania. J Med

Chem. 2009;52(23):7339-7359. doi:10.1021/jm9004835.

Lo Presti MS, Bazán PC, Strauss M, Báez AL, Rivarola HW, Paglini-Oliva PA.

Trypanothione reductase inhibitors: Overview of the action of thioridazine in different

2

stages

doi:10.1016/j.actatropica.2015.02.012.

Field MC, Horn D, Fairlamb AH, et al. Anti-trypanosomatid drug discovery: an ongoing

challenge and continuing need. Nat Rev Microbiol. 2017;15(4):217-231.

of

Chagas

disease.

Acta

Trop.

2015;145:79-87.

2

1.

2.

a

doi:10.1038/nrmicro.2016.193.

2

Hajipour AR, Rafiee F, Ruoho AE. A rapid and convenient method for the synthesis of

aldoximes under microwave irradiation using in situ generated ionic liquids. J Iran Chem

Soc. 2010;7(1):114-118. doi:10.1007/BF03245867.

2

3.

4.

Hansen T V., Wu P, Fokin V V. One-pot copper(I)-catalyzed synthesis of 3,5-

disubstituted isoxazoles. J Org Chem. 2005;70(19):7761-7764. doi:10.1021/jo050163b.

Morita J-I, Nakatsuji H, Misaki T, Tanabe Y. Water-solvent method for tosylation and

mesylation of primary alcohols promoted by KOH and catalytic amines. Green Chem.

2

2005;7:711. doi:10.1039/b505345e.

2

5.

6.

Himo F, Lovell T, Hilgraf R, et al. Copper(I)-catalyzed synthesis of azoles. DFT study

predicts unprecedented reactivity and intermediates. J Am Chem Soc. 2005;127(1):210-

216. doi:10.1021/ja0471525.

2

Lipinski CA. Drug-like properties and the causes of poor solubility and poor permeability.

J Pharmacol Toxicol Methods. 2000;44(1):235-249. doi:10.1016/S1056-8719(00)00107-

6.

2

7.

8.

Lipinski CA. Lead- and drug-like compounds: The rule-of-five revolution. Drug Discov

Today Technol. 2004;1(4):337-341. doi:10.1016/j.ddtec.2004.11.007.

Da Silva Filho AA, Costa ES, Cunha WR, Silva MLA, Nanayakkara NPD, Bastos JK. In

vitro antileishmanial and antimalarial activities of tetrahydrofuran lignans isolated from

Nectandra megapotamica (Lauraceae). Phyther Res. 2008;22(10):1307-1310.

doi:10.1002/ptr.2486.

2

29.

Felippe LG, Baldoqui DC, Kato MJ, et al. Trypanocidal tetrahydrofuran lignans from

Peperomia

blanda.

Phytochemistry.

2008;69(2):445-450.

doi:10.1016/j.phytochem.2007.08.012.

S26

30.

31.

32.

33.

34.

35.

36.

Pastor M, Cruciani G, McLay I, Pickett S, Clementi S. GRid-INdependent Descriptors

GRIND): A Novel Class of Alignment-Independent Three-Dimensional Molecular

Descriptors. J Med Chem. 2000;43(17):3233-3243. doi:10.1021/jm000941m.

(

Roy PP, Roy K. On Some Aspects of Variable Selection for Partial Least Squares

Regression

Models.

QSAR

Comb

Sci.

2008;27(3):302-313.

doi:10.1002/qsar.200710043.

da Silva Santos J, de Melos JLR, Lima GS, et al. Synthesis, anti-Trypanosoma cruzi

activity and quantitative structure relationships of some fluorinated thiosemicarbazones.

J Fluor Chem. 2017;195:31-36. doi:10.1016/j.jfluchem.2017.01.013.

Pauli I, Ferreira LG, de Souza ML, et al. Molecular modeling and structure–activity

relationships for a series of benzimidazole derivatives as cruzain inhibitors. Future Med

Chem. 2017;9(7):641-657. doi:10.4155/fmc-2016-0236.

Gottlieb HE, Kotlyar V, Nudelman A. NMR Chemical Shifts of Common Laboratory

Solvents

as

Trace

Impurities.

J

Org

Chem.

1997;62(21):7512-7515.

doi:10.1021/jo971176v.

Armarego WLF, Chai CLL. Chemical Methods Used in Purification. In: Purification of

Laboratory Chemicals. Elsevier. Elsevier; 2009:61-79. doi:10.1016/B978-1-85617-567-

8.50010-X.

Arshad M, Bhat AR, Pokharel S, et al. Synthesis, characterization and anticancer

screening of some novel piperonyl-tetrazole derivatives. Eur J Med Chem. 2014;71:229-

236. doi:10.1016/j.ejmech.2013.11.008.

3

7.

8.

Buckner FS, Verlinde CLMJ, Flamme ACL a, Voorhis WCV a N. Efficient Technique for

Screening Drugs for Activity against. Microbiology. 1996;40(11):2592-2597.

Schwende H, Fitzke E, Ambs P, Dieter P. Differences in the state of differentiation of

THP-1 cells induced by phorbol ester and 1 ,25-dihydroxyvitamin D3. J Leukoc Biol.

3

1996;59.

3

9.

0.

Hamilton CJ, Saravanamuthu A, Eggleston IM, Fairlamb AH. Ellman’s-reagent-mediated

regeneration of trypanothione in situ: substrate-economical microplate and time-

dependent inhibition assays for trypanothione reductase. Biochem J. 2003;369:529-537.

doi:10.1042/BJ20021298.

Ermondi G, Caron G, Pintos IG, et al. An application of two MIFs-based tools (Volsurf+

and Pentacle) to binary QSAR: The case of a palinurin-related data set of non-ATP

competitive Glycogen Synthase Kinase 3β (GSK-3β) inhibitors. Eur J Med Chem.

4

2011;46(3):860-869. doi:10.1016/j.ejmech.2010.12.024.

4

1.

2.

Teófilo RF, Martins JPA, Ferreira MMC. Sorting variables by using informative vectors as

a strategy for feature selection in multivariate regression. J Chemom. 2009;23(1):32-48.

doi:10.1002/cem.1192.

Eriksson L, Jaworska J, Worth AP, Cronin MTD, McDowell RM, Gramatica P. Methods

for Reliability and Uncertainty Assessment and for Applicability Evaluations of

4

Classification-

and

Regression-Based

QSARs.

Environ

Health

Perspect.

2003;111(10):1361-1375. doi:10.1289/ehp.5758.

43.

Martins JP a, Ferreira MMC. QSAR modeling: um novo pacote computacional open

source para gerar e validar modelos QSAR. Quim Nova. 2013;36(4):554-560.

doi:10.1590/S0100-40422013000400013.

S27

Supporting Information

Synthesis and SAR of new isoxazole-triazole bis-heterocyclic compounds as

analogues of natural lignans with antiparasitic activity

†

⁑

†

#

Lara A. Zimmermann , Milene H. de Moraes , Rafael da Rosa , Eduardo B. de Melo ,

‖

†

⁑

†;*

Fávero R. Paula , Eloir P. Schenkel , Mario Steindel and Lílian S. C. Bernardes .

†

Department of Pharmaceutical Sciences, Federal University of Santa Catarina, 88040-900,

Florianópolis, 88040-900, SC, Brazil.

⁑

Department of Microbiology, Immunology and Parasitology, Federal University of Santa

Catarina, Florianópolis, 88040-900, SC, Brazil.

#

Department of Pharmacy, Western Paraná State University, Cascavel, 858119-110, PR,

Brazil.

Course of Pharmacy, Federal University of Pampa, Uruguaiana, 97500-970, RS, Brazil.

Corresponding author: l.bernardes@ufsc.br

‖

*

S28

Table of contents

Figure S1. Best GRIND descriptors associated with compounds 10 and 18 .............................................................30

Table S1. Molecular properties of (7-48) calculated using StarDrop 5 Program .......................................................31

Table S2: Leishmanicidal activity of compounds 7-48 against Leishmania amazonensis intracellular

amastigotes ..............................................................................................................................................................32

Table S3: Trypanosoma cruzi rTR inhibitory activity of compounds 10, 12, 25-26, 28-31.......................................33

R EPRESENTATIVE SPECTRA ...................................................................................................................................................34

S29

Article Doi

Synthesis and SAR of new isoxazole-triazole bis-heterocyclic compounds as analogues of natural lignans with antiparasitic activity

DOI: 10.1016/j.bmc.2018.08.025

Source and publish data:

Authors:

Article abstract of DOI:10.1016/j.bmc.2018.08.025

Full text of DOI:10.1016/j.bmc.2018.08.025

Products guided by the article

R&D Labs maybe for 105363-12-0

Relevant to this article

Hot Product