Experimental and theoretical study on the reaction of N3-phenyl- (pyridin-2-yl)carbohydrazonamide with itaconic anhydride

Article abstract of DOI:10.1016/j.molstruc.2012.04.087

Two new 1,2,4-triazole-containing alkenoic acid derivatives were obtained from the reaction of N-phenyl-(pyridin-2-yl)carbohydrazonamide with itaconic anhydride, depending on the reaction conditions. The structures of 2-((4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-yl)methyl)acrylic acid or (E)-2-methyl-3(4-phenyl-5-(pyridine-2-yl)-4H-1,2,4-triazol-3-yl)acrylic acid were confirmed by means of 1D and 2D NMR spectroscopic data as well as by single-crystal X-ray diffraction analysis. The experiential ¹H and ¹³C chemical shifts were compared with those calculated with B3LYP, EDF1, and EDF2 density functional theories. The theoretical study of the observed terminal-to-internal alkene isomerization was performed with density functional (DFT) B3LYP/6-31+G method using SM8 water and DMF solvation models. Antimicrobial activities of the newly prepared alkenoic acid derivatives were verified experimentally by a broth microdilution method.

Full text of DOI:10.1016/j.molstruc.2012.04.087

Journal of Molecular Structure 1022 (2012) 211–219
Contents lists available at SciVerse ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstruc
Experimental and theoretical study on the reaction
of N3-phenyl-(pyridin-2-yl)carbohydrazonamide with itaconic anhydride
a
a
b
c,
⇑
_
Bozena Modzelewska-Banachiewicz , Renata Paprocka , Liliana Mazur , Jarosław Saczewski
,
d
e
e
´
´
Jolanta Kutkowska , Dorota K. Ste˛pien , Michał Cyranski
a Department of Organic Chemistry, Faculty of Pharmacy, Nicolaus Copernicus University, Jurasza 2, 85-089 Bydgoszcz, Poland
b Department of General and Coordination Chemistry, Maria Curie-Sklodowska University, M. Curie-Sklodowska Sq. 2, 20-031 Lublin, Poland
c Department of Chemical Technology of Drugs, Medical University of Gdansk, Hallera 107, 80-416 Gdansk, Poland
d Department of Genetics and Microbiology, Maria Curie-Sklodowska University, Akademicka 19, 20-033 Lublin, Poland
e Department of Chemistry, University of Warsaw, Pasteura 1, 02-093 Warsaw, Poland
h i g h l i g h t s
"
"
"
"
New isomeric 1,2,4-triazole-containing alkenoic acids are obtained.
Mechanisms of base-catalyzed and neutral thermal 1,3-proton shifts are studied theoretically.
The structures are confirmed by 1D and 2D NMR and single crystal X-ray diffraction analysis.
Antimicrobial activity of the 1,2,4-triazole-containing alkenoic acids are investigated.
a r t i c l e i n f o
a b s t r a c t
Article history:
Two new 1,2,4-triazole-containing alkenoic acid derivatives were obtained from the reaction of N-phe-
nyl-(pyridin-2-yl)carbohydrazonamide with itaconic anhydride, depending on the reaction conditions.
The structures of 2-((4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-yl)methyl)acrylic acid or (E)-2-
methyl-3(4-phenyl-5-(pyridine-2-yl)-4H-1,2,4-triazol-3-yl)acrylic acid were confirmed by means of 1D
and 2D NMR spectroscopic data as well as by single-crystal X-ray diffraction analysis. The experiential
1H and 13C chemical shifts were compared with those calculated with B3LYP, EDF1, and EDF2 density
functional theories. The theoretical study of the observed terminal-to-internal alkene isomerization
was performed with density functional (DFT) B3LYP/6-31+G⁄ method using SM8 water and DMF solva-
tion models. Antimicrobial activities of the newly prepared alkenoic acid derivatives were verified exper-
imentally by a broth microdilution method.
Received 3 March 2012
Received in revised form 24 April 2012
Accepted 25 April 2012
Available online 8 May 2012
Keywords:
1,2,4-Triazole
Carbohydrazonamide
Itaconic anhydride
Alkenoic acids
Ó 2012 Elsevier B.V. All rights reserved.
1,3-Proton shift reaction
X-ray diffraction analysis
1. Introduction
tions. On the other hand, it is well known that derivatives of vari-
ous alkenoic acids [2–4] and cinnamic acid [5] exhibit
pronounced antibacterial and antifungal activities.
a
1,2,4-Triazole derivatives constitute an important class of het-
erocyclic compounds, which gained considerable attention in re-
cent years due to a broad spectrum of biological properties,
which include: antimicrobial, antiviral, anticancer, antiasthmatic,
anticonvulsant, antidepressant, antihypertensive, antiemetic, hyp-
notic, sedative, anxiolytic, antithyroid, hypoglycemic and antimi-
graine activity [1]. Among them fluconazole and itraconazole
belong to the most powerful antimycotic drugs used today in the
treatment and prevention of superficial and systemic fungal infec-
In view of the above, we expected that by combining the 1,2,4-
triazole ring system and the alkenoic acid moiety, a new class of
heterocyclic analogues of cinnamic acid with the desired antimi-
crobial properties could be obtained. Our research plane aimed at
achieving that goal was based on previous investigations of hydra-
zonamide chemistry [6–9] and chemical properties of itaconic
anhydride [10,11]. The synthetic pathways leading to 2-((4-phe-
nyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-yl)methyl)acrylic acid (3)
and (E)-2-methyl-3(4-phenyl-5-(pyridine-2-yl)-4H-1,2,4-triazol-
3-yl)acrylic acid (4) are presented in Scheme 1. The molecular
structures of alkenoic acid derivatives obtained were confirmed
⇑
Corresponding author. Tel.: +48 583491953; fax: +48 583491654.
E-mail address: js@gumed.edu.pl (J. Saczewski).
0022-2860/$ - see front matter Ó 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.molstruc.2012.04.087

212
B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
by 1D and 2D NMR spectral data and single crystal X-ray diffrac-
tion analysis. Moreover, a facile isomerization of the terminal al-
kene derivative into the more thermodynamically stable internal
alkene prompted us to investigate the mechanisms of both the
base-catalyzed and the thermal 1,3-proton shifts using DFT
B3LYP/6-31+G⁄ method combined with SM8 (H2O, DMF) solvation
models.
purified by crystallization from water to give 0.46 g (1.5 mmol,
15%) of 3.
M.p. 164–166 °C. 1H NMR (300 MHz, DMSO-d6): d = 3.56 (s, 2 H,
CH2), 5.58 (s, 1 H, CH), 6.13 (s, 1 H, CH), 7.30–7.37 (m, 3 H, CH),
7.45–7.48 (m, 3 H, CH), 7.89 (t, J = 6,0 Hz, 1 H, CH), 7.96 (d,
J = 6.0 Hz, 1H, CH), 8.29 (d, J = 6.0 Hz, 1 H, CH), 12.59 (bs, 1 H,
COOH) ppm. 13C NMR (100 MHz, DMSO-d6): d = 29.3, 125.5,
126.0, 128.7, 129.2, 130.9, 131.0, 136.9, 137.9, 139.0, 148.5,
150.7, 154.3, 155.8, 168.9. FT-IR (KBr, cmꢀ1): 3426m, 3078m,
1711s, 1635m, 1591m, 1500s, 1457s, 1417m, 1218m, 1164m,
1008m, 954w, 792m, 781m, 700m. ESI-MS m/z (%): 306 (57%),
261 (100%), 130 (28%), 78 (41%), 77 (50%). Anal. Calc. for
2. Experimental
2.1. Materials and methods
C
17H14N4O2: C, 66.66; H, 4.61; N, 18.29%; found: C, 66.46; H,
Chemicals were purchased from Aldrich and used without fur-
ther purification. Melting points were measured on a Mel TEMP
1002D apparatus and are uncorrected. Mass spectra data were ob-
tained on Finingan Trace DSQ GC/MS with electrospray ion source.
The FT-IR spectra were recorded from KBr pellets in the range of
4000–400 cmꢀ1 on Perkin Elmer Spectrum 2000 infrared spec-
trometer. The elemental analysis was made with a Vario Macro
11.45-0000 (Elementar Analysesysteme GmbH, Germany) operat-
ing with the software VARIOEL. 1H NMR and 13C NMR spectra were
recorded on a Bruker Avance spectrometer in DMSO-d6 with TMS
as an internal standard. N3-phenyl-(pyridine-2-yl)carbohydrazo-
namide (1) was synthesized according to the previously described
procedure [6].
4.62; N, 18.07%.
2.3. Isomerization of 3 to (E)-2-methyl-3-(4-phenyl-5-(pyridin-2-yl)-
4H-1,2,4-triazol-3-yl)acrylic acid (4)
Method I: Compound 3 (0.46 g, 1.5 mmol) was dissolved in 2%
aqueous solution of sodium hydroxide (10 mL) and the reaction
mixture was left at room temperature for 12 h. Then, the solution
was cooled to room temperature and neutralized with 1% hydro-
chloric acid. The precipitate thus obtained was collected by filtra-
tion, washed with water and dried in vacuum to give 0.44 g
(1.4 mmol, 95.6%) of 4.
Method II: Compound 3 (0.46 g, 1.5 mmol) was dissolved in
dimethylformamide (10 mL) and heated at 150 °C for 4 h. Upon
cooling to room temperature the reaction mixture was filtered
and the filtrate was diluted with water (10 mL). The product that
precipitated was collected by filtration, washed with water and
dried in vacuum to yield 0.27 g (0.88 mmol, 58.7%) of 4.
2.2. Synthesis of 2-((4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-
yl)methyl)acrylic acid (3)
The carbohydrazonamide 1 (2.12 g, 10 mmol) and itaconic
anhydride 2 (1.12 g, 10 mmol) were dissolved in anhydrous diethyl
ether (50 mL) and the reaction mixture was left at room tempera-
ture for 7 days. The solid product that precipitated was collected by
filtration and washed with diethyl ether. Then, the solid was dis-
solved in chloroform (30 mL), heated at reflux for 5 min. and fil-
tered. The filtrate was evaporated to dryness and the solid
residue was washed with anhydrous diethyl ether, dried and
M.p. 234–236 °C. 1H NMR (500 MHz, DMSO-d6): d = 2.42 (d, 3H,
CH3, J = 1.5 Hz); 6.85 (d, 1H, CH, J = 1.5 Hz); 7.36–7.41 (m, 3H, CH);
7.49–7.54 (m, 3H, CH); 7.92 (t, J = 7.9 Hz, 1H, CH); 8.04 (d, J = 7.9 Hz,
1H, CH); 8.30–8.32 (m, 1H, CH); 12.8 (s, 1H, COOH). 13C NMR
(125 MHz, DMSO-d6): d = 15.2, 119.0, 124.0, 124.5, 127.8, 129.4,
129.5, 134.5, 136.0, 137.2, 146.2, 149.1, 152.4, 152.5, 168.2. FT-IR
(KBr, cmꢀ1): 3428s, 3055m, 2923w, 2496w, 1938w, 1704s,
Table 1
Experimental and DFT B3LYP/6-31+G⁄ 1H and 13C NMR chemical shifts d of 3 and 4 with the root-mean-square deviation (RMSD).
H, or C atom atom
Chemical shift d experimental
B3LYP/6-31+G⁄ calculated
B3LYP/6-31+G⁄ correcteda
EDF1
EDF2
3
CA4H2
CA6H
CA6H0
RMSD
3.56
5.58
6.13
3.06
5.95
6.45
0.40
3.26
6.04
6.49
0.38
3.12
5.99
6.51
0.41
4
CA12H3
CA22H
CA3PH
CA4PH
CA5PH
CA6PH
RMSD
C-12
C-22
C-2
C-1
C-3
2.42
6.85
8.04
7.92
7.38
8.31
2.64
6.60
8.68
7.49
6.80
8.04
0.43
16.4
116.7
130.0
157.0
146.5
148.9
141.9
118.0
128.7
116.1
141.3
6.60
2.82
6.59
8.73
7.49
6.79
8.15
0.46
19.2
114.5
128.7
154.7
144.8
147.1
139.6
116.5
125.9
114.2
139.5
8.48
2.69
6.70
8.74
7.51
6.82
8.05
0.43
16.5
116.9
130.3
156.9
146.2
148.7
141.5
118.3
128.9
116.5
141.5
6.54
15.2
16.14
122.8
136.0
168.9
156.4
158.8
151.3
124.1
135.0
122.7
151.0
3.16
119.0
136.0
168.3
152.5
152.4
146.2
124.1
137.3
124.5
149.1
C-5
CA2P
CA3P
CA4P
CA5P
CA6P
RMSD
a
The corrected 13C chemical shifts account local environment in addition to directly calculated shifts.

B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
213
1644m, 1498s, 1440s, 1295s, 1241s, 992m, 793m, 774s, 696m,
586m. ESI-MS m/z (%): 306 (85%), 288 (24%), 261 (100%), 156
(32%), 130 (69%), 78 (67%), 77 (87%). Anal. Calc. for C17H16N4O3:
C, 62.95; H, 4.97; N, 17.27%; found: C, 62.86; H, 5.12; N, 16.89%.
F2 for all reflections except those with very negative F2. The
weighted R factors wR and all goodness-of-fit S values are based
on F2. The non-hydrogen atoms were refined anisotropically. In 3
the H atoms adjacent to O1 and C6 atoms were found in the differ-
ence-Fourier maps and refined with isotropic displacement param-
eters. All remaining
H atoms were positioned geometrically
2.4. Synthesis of (E)-2-methyl-3-(4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-
[CAH = 0.97 Å for CH2 and 0.93 Å for CH(ar)] and refined using the
riding model with Uiso(H) = 1.2Ueq(C). In 4 all hydrogen atoms were
located from a difference Fourier map; their positions and thermal
parameters were refined isotropically. The scattering factors were
taken from Tables 6.1.1.4 and 4.2.4.2 [16].
All figures presenting the results of X-ray diffraction determina-
tion were made using the MERCURY program [17]. Crystal data and
structural refinement for 3 and 4 are specified in Table 2. Selected
bond lengths, bond angles, major torsion angles and hydrogen
bonds are given in Tables 3 and 4. Displacement ellipsoids and
atom numbering are shown in Figs. 1 and 2, respectively. Packing
and hydrogen bonds are shown in Figs. 3–6.
triazol-3-yl)acrylic acid (4)
A solution of hydrazonamide 1 (2.12 g, 10 mmol) and itaconic
anhydride (2) (1.12 g, 10 mmol) in anhydrous diethyl ether
(50 mL) was stirred vigorously at room temperature for 2 h. The
solid that precipitated was collected by filtration, washed with
diethyl ether and dried under vacuum. Then, the dry solid thus ob-
tained was dissolved in 2% aqueous NaOH solution and the reac-
tion mixture was heated under reflux for 2 h. Upon cooling to
room temperature the solution was neutralized with 1% HCl. The
solid that precipitated was collected by filtration, washed with
water and dried in vacuum to give 2.32 g (7.57 mmol, 75%) of 4.
2.5. X-ray structure analysis
Single-crystals of 3 and 4 suitable for X-ray analysis, were ob-
tained by slow crystallization from ethanol. The XRD measure-
ments for 3 and 4 were performed at 100 K on Oxford Diffraction
Xcalibur CCD diffractometer with the graphite-monochromatized
Table 3
Selected geometric parameters.
3
4
Bond distances (Å)
C1AO1
C1AO2
C1AC2
C2AC6
C2AC4
C3AC4
N1AN2
N2AC3
C3AN4
Mo K
monochromated Enhance Cu K
tals were positioned at 55 mm from the CCD camera. Data sets
were collected using the scan technique, with an angular scan
a
radiation and Gemini
j-axis diffractometer with graphite-
1.326(2)
1.213(2)
1.493(2)
1.315(2)
1.505(2)
1.498(2)
1.383(2)
1.307(2)
1.374(2)
1.375(2)
1.315(2)
1.315(3)
1.220(2)
1.497(3)
1.494(3)
1.345(3)
1.454(3)
1.380(2)
1.318(3)
1.377(2)
1.373(2)
1.311(2)
a
radiation, respectively. The crys-
x
width of 0.75° and 1.0° for 3 and 4, respectively. In both cases
the data were corrected for Lorentz and polarization effects. Mul-
ti-scan absorption correction was also applied [12]. Data reduction
and analysis were carried out with the CrysAlis RED, Oxford Dif-
fraction Ltd. [13]. The structures were solved by direct methods
[14] and refined using SHELXL [15]. The refinement was based on
N4AC5
C5AN1
Bond angles (°)
O2AC1AO1
O2AC1AC2
O1AC1AC2
124.2(1)
123.1(1)
112.7(1)
123.5(2)
124.1(2)
112.4(2)
Table 2
Crystal data and structure refinement details for 3 and 4.
Torsion angles (°)
O1AC1AC2AC4
O1AC1AC2AC6
C1AC2AC4AC3
C6AC2AC4AC3
C2AC4AC3AN2
3
4
160.0(1)
ꢀ25.4(2)
ꢀ59.5(2)
126.1 (2)
ꢀ13.1(2)
ꢀ150.7(2)
28.4(2)
179.2(2)
0.2(3)
Formula
C
17H14N4O2
C17H16N4O3
324.34
100(2)
Formula weight (g molꢀ1
T (K)
)
306.32
100(2)
0.71073
19.9(3)
Wavelength (Å)
Crystal system, space group
1.54184
Monoclinic, Pn
5.8875(2)
8.5996(3)
15.7666(6)
96.743(4)
792.74(5)
2
Monoclinic, P21/n
9.1499(3)
10.0043(3)
16.2523(7)
104.686(4)
1439.10(9)
4
1.414
3.56–28.28
0.097
a (Å)
b (Å)
c (Å)
b (°)
V (Å3)
Z
Table 4
Geometry of proposed hydrogen bonds for 3 and 4.
Calculated density (g cmꢀ3
h Range (°)
)
1.359
5.14–67.08
0.793
DAH (Å)
Hꢂ ꢂ ꢂA (Å)
Dꢂ ꢂ ꢂA (Å)
<DAHꢂ ꢂ ꢂA (°)
3
Absorption coefficient
O1AH1oꢂ ꢂ ꢂN3(i)
C11AH11ꢂ ꢂ ꢂO2(ii)
C6AH6aꢂ ꢂ ꢂO1(iii)
C6AH6bꢂ ꢂ ꢂN1(iv)
C14AH14ꢂ ꢂ ꢂN1(v)
C8AH8ꢂ ꢂ ꢂN1(vi)
1.00(2)
0.93
1.03(2)
1.04(2)
0.93
1.74(3)
2.63
2.66(2)
2.69(2)
2.54
2.705(3)
3.342(3)
3.411(3)
3.590(2)
3.412(2)
3.318(3)
164(3)
134
127(3)
149(2)
157
(mmꢀ1
)
Crystal size (mm)
Crystal color and form
0.49 ꢁ 0.25 ꢁ 0.16
0.31 ꢁ 0.23 ꢁ 0.11
Colorless prism
8777/2706
[Rint = 0.041]
2551
Colorless prism
Reflections collected/unique 21461/3556
[Rint = 0.059]
2463
0.93
2.46
154
Observed reflections
(I > 2 (I))
Data/restraints/parameters
4
r
O1wAH1wꢂ ꢂ ꢂO2
O1wAH2wꢂ ꢂ ꢂN1(vii)
O1wAH2wꢂ ꢂ ꢂN2(vii)
O1AH1ꢂ ꢂ ꢂO1w(ii)
C8AH8ꢂ ꢂ ꢂO2(viii)
C6AH62ꢂ ꢂ ꢂN3(ix)
0.81(3)
0.88(3)
0.88(3)
0.94(3)
0.94(2)
0.98(3)
2.02(3)
1.90(3)
2.49(3)
1.59(3)
2.47(3)
2.60(3)
2.820(2)
2.780(2)
3.262(2)
2.515(2)
3.230(2)
3.503(3)
173(2)
178(3)
146(3)
167(3)
138(2)
155(2)
3556/0/220
R1 = 0.0403;
wR2 = 0.0914
R1 = 0.0664;
wR2 = 0.0984
0.923
2706/2/279
R1 = 0.0332;
wR2 = 0. 0803
R1 = 0.0359;
wR2 = 0.0816
1.026
Final R indices (I > 2
r(I))
R indices (all data)
Goodness-of-fit on F2
Completeness to h max. (%)
99.7
0.31/ꢀ0.21
99.9
0.13/ꢀ0.21
Symmetry codes: (i) x ꢀ 1, y, z; (ii) 1 + x, y, z; (iii) ꢀx, ꢀy, ꢀz; (iv) 1 ꢀ x, ꢀy, ꢀz; (v) 1/
2 + x, 1/2 ꢀ y, 1/2 + z; (vi) 1 ꢀ x, 1 ꢀ y, ꢀz; (vii) x, y ꢀ 1, z; (viii) x ꢀ 1, 1 + y, z; (ix) 1/
2 + x, 2 ꢀ y, z ꢀ 1/2.
Largest diff. peak and hole
(e Åꢀ3
)

214
B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
Fig. 1. The molecular structure of 3 showing the atom-numbering scheme. Displacement ellipsoids are drawn at the 50% probability level.
Fig. 2. Molecular structure and atomic numbering for 4. Displacement ellipsoids are drawn at 50% probability level.
Fig. 3. Part of the crystal structure of 3, showing hydrogen-bonded (dashed lines) molecular chains.

B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
215
Fig. 4. Part of the crystal structure of 4, showing hydrogen-bonded (dashed lines)
molecular chains.
Fig. 6. Packing view of 4 along [100] direction. Dashed lines indicate hydrogen
bonds.
cus faecalis ATCC 29212, Sarcina lutea; Mycobacterium smegmatis,
Nocardia spp, and the pathogenic fungus Candida albicans. Bacterial
cultures were grown on liquid Luria–Bertani (LB) medium at
temperature 37 °C. The minimum inhibitory concentration
(MIC50) was measured by replicating 105 CFU/mL onto microplates
Fig. 5. Packing view of 3 along [010] direction.
supplemented with 50, 75, 100, and 250 lg/mL of corresponding
compound dissolved in dimethylsulfoxide (DMSO). Tests were
performed in triplicate for each concentration, in all the tests,
DMSO was used as the control. After 18 h the growth of bacteria
was estimated spectrophotometrically (550 nm) with an ELISA
detector.
2.6. Theoretical calculations
All the quantum-chemical calculations were carried out with
use of the Spartan 08 program package provided by Wavefunction,
Inc. [18]. The geometries were fully optimized in vacuum with DFT
B3LYP method using 6-31+G⁄ basis set combined with water SM8
solvation model [19]. Frequency calculations were performed for
all structures to prove the energy minima. The Gibbs free energies
were obtained from the electronic energies corrected with the
zero-point energies (ZPE), thermal energies involving temperature
increase from 0 to 298.15 or 423.15 K and entropies. The relative
energies were obtained by subtracting the energy of the starting
structures from the energies of all the other geometries and con-
verting these differences into kcal/mol. Vibrational frequency anal-
yses and IRC calculations were performed for all found transition
states (TSs) to prove that they assuredly connect the reactants
and products.
3. Results and discussion
3.1. Synthesis
It is known that itaconic anhydride easily isomerizes to citra-
conic anhydride in the presence of organic bases such as pyridine
or N,N-dimethylaniline [10,11]. As shown in Scheme 2, this isomer-
ization proceeds via anionic intermediate which is stabilized in po-
lar solvents. Therefore, to secure the formation of the final product
with external C@C double bond, we have run the reaction of
anhydride 1 with carbohydrazonamide 2 in diethyl ether at room
temperature (Scheme 1). 2-((4-Phenyl-5-(pyridin-2-yl)-4H-1,2,4-
triazol-3-yl)methyl)acrylic acid (3) thus obtained was further
transformed into a more thermodynamically stable (E)-2-methyl-
3-(4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-yl)acrylic acid (4)
by the treatment with aqueous 2% NaOH solution at room temper-
ature for 12 h (base-catalyzed isomerization, 95% yield) or by heat-
ing in wet DMF at 150oC for 4 h (thermal isomerization, 58% yield).
It should be pointed out that the compound 3 incorporating ter-
minal C@C double bond was separated in pure form in rather low
yield (15%) and attempts in optimization of the synthetic procedure
2.7. Antibacterial in vitro activity of compounds 3 and 4
The susceptibility to the new synthesized compounds was
determined by a broth microdilution method in standard 96-well
sterile flat-bottom polystyrene plates (Kartell). The following
strains were tested: Gram-negative bacteria: Escherichia coli ATCC
25922, Pseudomonas aeruginosa ATCC 27853, Yersinia enterocolitica
O3; Gram-positive: Staphylococcus aureus ATCC 25923, Enterococ-

216
B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
Scheme 1. The reaction of N-phenyl(pyridine-2-yl)carbohydrazonamide (1) with itaconic anhydride (2).
O
O
R
R
NH
NH
O
O
B
RNH2
O
O
CH2
OH
B-H
B-H
2
O
O
O
RNH2
ion pair
O
O
stabilised
O
CH3
OH
in polar solvent
O
Scheme 2. Reactivity of itaconic anhydride.
by varying organic solvents and reaction temperature were not suc-
cessful due to formation of intractable mixtures of products. We
succeeded, however, in obtaining the compound 4 in 75% yield from
hydrazonamide 1 and itaconic anhydride 2 by the treatment of the
crude intermediate A, initially formed in diethyl ether solution,
with aqueous NaOH solution at reflux (Scheme 1).
The molecular structures of the newly prepared 1,2,4-triazole
derivatives 3 and 4 were confirmed unambiguously by 1H and
13C NMR spectroscopic data including 1HA13C HSQC and HMBC
correlation studies (Supplementary material) as well as by single
crystal X-ray diffraction analysis (Figs. 1–6).
The experimental NMR spectra of the compounds 3 and 4 were
compared with the calculated proton and carbon shifts obtained by
means of the density functional calculations (Table 1). The allyl
group of 3 was similarly described by the all tested models, how-
ever the Empirical Density Functional 1 (EDF1) method exhibited
the lowest root-mean-square deviation (RMSD) of 0.38 ppm. On
the contrary, in the case of the 1H and 13C NMR spectra of 4 the
B3LYP and EDF2 models gave considerably better results than
EDF1, especially for the carbon atoms. It is worth noting that the
most accurate 13C NMR prediction was obtained using the cor-
rected B3LYP method which accounts local environment in addi-
tion to the directly calculated shifts (RMSD = 3.16 ppm) [18].
3.2. X-ray crystallography
Compounds 3 and 4 crystallize in the monoclinic space group
P21/n and Pn, respectively. An independent part of the unit cell con-
sists of one molecule of 2-methylene-4-oxo-4-(2-((phenylamino)
(pyridin-2-yl)methylene)hydrazinyl)butanoic acid in the former
case, and 2-((4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-yl)methyl)
acrylic acid with water molecule (1:1), in the latter one.
The molecule of 3 is composed of the central 1,2,4-triazole ring,
substituted at the C3, N4 and C5 atoms by 2-methacrylic acid, phe-
nyl and 2-pyridyl groups, respectively (Fig. 1). The interatomic dis-
tances within the triazole ring are not equal (Table 3) but
comparable with those observed for the other closely related
1,2,4-triazole derivatives [20,21]. The N1AC5, N2AC3 bonds, with
the interatomic distances of 1.315(2) and 1.307(2) Å, display a
double-bond character, whereas those of C3AN4 and N4AC5 ones
[1.374(2) and 1.375(2) Å] are of intermediate character. The central
heterocyclic ring in 3 is planar but not coplanar with its substitu-
ents. The pyridyl and phenyl rings are twisted about the external
bond to the 1,2,4-triazole ring; the angles between the least-
squares planes through the pyridyl/phenyl and triazole rings are
18.55(7)° and 63.66(5)°, respectively. The planes through the pyr-
idyl and benzene rings form a dihedral angle of 56.20(5)°. As in the

B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
217
Scheme 3. The base promoted 1,3-proton shift of 2-((4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-yl)methyl)acrylic acid (3) with the relative electronic energy (
D
Ee, kcal/
mol) and Gibbs free energy (D
G°, kcal/mol, 298.15 K) profiles calculated with B3LYP 6-31+G⁄ method using SM8 water solvation model [19].
parent itaconic acid [22] the carboxyl O1 atom in 3 is cisoidally ori-
ented with respect to the terminal @CH2 group. It is worth noting,
that the same orientation was observed in the crystal structures of
most C3-unsubstituted 2-methacrylic acid derivatives, reported so
far [23]. However, contrary to the free acid, a significant deviation
from planarity is observed for the methacrylic system in 3 as indi-
cated from the torsion angles O1AC1AC2AC4 and O1AC1AC2AC6,
being of 160.0(1)° and ꢀ25.4(2)°, respectively. This might be
caused by steric hindrance between the carboxyl and triazole moi-
eties, which seems to be supported by relative widening of O1-C1-
C2 angle (by about 1.3°). The C1AO2 and C2AC6 distances
[1.213(2) and 1.315(2) Å] are in good agreement with the localized
double bond lengths, whereas the C1AC2 distance [1.493(2) Å]
confirms the presence of unconjugated Csp2ACOOH single bond
[23,24].
Geometry of 4 displays significant similarities to 3, except obvi-
ous hydrogen transfer from CH2 fragment (at C4 carbon atom) to
CH2 group (at C6 carbon atom), what involves appropriate changes
of hybridizations and methyl group formation (Fig. 2). Therefore,
for instance, in both systems the pyridine and benzene rings are
twisted in respect to the 1,2,4-triazole ring to similar extent: the
angles between the best planes of the pyridine or benzene rings
and triazole fragment are 23.90° and 66.86°, respectively. The bond
lengths in the methacrylic moiety are within normal ranges and
are comparable to those observed for closely related mesaconic
acid [25]. The relative orientation of carboxyl C1 and triazole C3
atoms indicate transoid conformer with the carboxyl group twisted
from the plane of C4@C2(C6)AC1 atoms by the angle of 28.75(5)°.
The most characteristic feature of crystal 3 is the presence of
catemers formed by translation related molecules propagated
along the a axis (Fig. 3). Within each chain ‘head-to-tail’ oriented
molecules are linked by the O1AH1oꢂ ꢂ ꢂN3(i) and C11AH11ꢂ ꢂ ꢂO2(ii)
hydrogen bonds (symmetry codes as in Table 4) giving R22(7) rings.
Furthermore, the molecules belonging to the adjacent antiparallel
chains are connected by an extended net of weak CAHꢂ ꢂ ꢂO/N
hydrogen bonds between the atoms of 2-methacrylic system as
well as the pyridyl/phenyl and triazole rings giving rise to complex
3D supramolecular architecture (Fig. 5). The pattern of interactions
in (E)-2-methyl-3-(4-phenyl-5-(pyridin-2-yl)-4H-1,2,4-triazol-3-
yl)acrylic acid (4) is totally different. The co-crystallizing water
molecule behaves as a structural gluing factor [26,27]. It links three
molecules together and serves as the hydrogen bond acceptor to
hydroxyl group at carboxylic fragment (O1ꢂ ꢂ ꢂO1w(ii) [2.515 Å]) of
one molecule and hydrogen bond donor to carbonyl group
(O1wꢂ ꢂ ꢂO2 [2.820 Å]) and nitrogen atom at triazole ring
(O1wꢂ ꢂ ꢂN1(viii) [2.780 Å]) of two other molecules, as shown in
Figs. 4 and 6.
3.3. Theoretical studies
The observed 3 ? 4 isomerization in aqueous NaOH solution
corresponds to numerous base-promoted 1,3-proton shifts de-
scribed in the literature [28–30] including isomerisation of itaconic
acid to mesaconic acid [31]. In order to elucidate the chemical
mechanism, which leads to rearranged isomeric triazole derivative
4, we have performed B3LYP/6-31+G⁄ DFT calculations with use of
SM8 water solvation model [19]. As depicted in Scheme 3, the pro-
posed reaction sequence is comprised of the initial exothermic
abstraction of the acidic methylene proton (
that leads to the formation of carbanion C and the subsequent
hydrolysis giving rise to the formation of salt D (
G° = ꢀ5.7 kcal/
D
G° = ꢀ5.3 kcal/mol)
D
mol). The following neutralization of carboxylate function with di-
luted hydrochloric acid provides the isomerized alkene derivative
4. It is worth noting that weaker bases such as triethylamine or
pyridine were ineffective in catalyzing the conversion of 3 to 4.
From a theoretical point of view, however, the most interesting
is the less frequently investigated [32] neutral thermal rearrange-
ment of 3 to 4 observed in DMF solution at 150oC. In this study we
have considered three possible mechanisms of 1,3-proton shifts
presented in Scheme 4.
First, the direct C ? C proton transfer via transition state TS1
leading to cis-olefin E. The activation energy of the rearrangement
3 ? E via direct intramolecular proton transfer is very high
(DEe = 73.2 kcal/mol), and therefore, it should not occur. In fact,
the cis-isomer E was not detected in NMR spectrum of the crude
reaction product.
Second, the two step mechanism which consists in the initial
C ? N 1,3-proton shift via transition state TS2 (
mol) furnishing the dearomatized triazole F, which is followed by
N ? C 1,5-proton shift via transition state TS3 ( Ee = 36.2 kcal/
DEe = 73.9 kcal/
D
mol) that leads to the final product 4. Although the activation
energy of C ? N 1,3 proton transfer is slightly lower than that

218
B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
Scheme 4. Thermal 1,3-proton shift reaction with the relative electronic energy (DEe, kcal/mol) and Gibbs free energy (DG°, kcal/mol, 423.15 K) profiles calculated with
B3LYP 6-31+G⁄ method using SM8 DMF solvation model [19].
calculated for direct C ? C transfer, it is still prohibitive, and there-
fore, the formation of F should be considered an unfavorable
process.
Third plausible mechanism, i.e. water-mediated double-proton
transfer reaction is based on interactions between the solute and
the solvent molecules. The thermal rearrangement of 3 to 4 is car-
ried out in the reagent grade DMF which contains a considerable
amount of water, hence, the ‘‘water-relay’’ mechanism cannot be
excluded. According to this model, the nucleophilic water can
accept a proton from the donor site of the solute molecule and
transfer a different proton to the acceptor site, which often results
in significant lowering of the energy barrier in proton transfer-re-
lated reactions, including various 1,3-proton shifts [33–40]. As
shown in Scheme 4, upon addition of water molecule the energy
barrier of the rearrangement of 3 ? F is lower by 34.1 kcal/mol
than that for the direct C ? N 1,3-proton shift. The transition state
TS4 is characterized by
DEe = 39.8 kcal/mol and a single imaginary
frequency (i1300 cmꢀ1) referring to CAH—O bond breakage and

B. Modzelewska-Banachiewicz et al. / Journal of Molecular Structure 1022 (2012) 211–219
219
N—HAO bond formation. The C—H and N—H atom distances are
1.428 and 1.055 Å, while the O—H atoms are separated by 1.241
and 1.689 Å, respectively.
Interestingly, the subsequent relayed N ? C 1,5-proton shift in
F leading to the final product 4 via transition state TS5 is energet-
ically more favorable than the alternative direct shift via TS3
Cambridge Crystallographic Data Centre as supplementary publi-
cations Nos. CCDC-861928 (3) and CCDC-830714 (4). Copies of
available materials can be obtained free of charge on application
to CCDC, 12 Union Road, Cambridge CB2 1EZ, UK (fax: +44 1223
336 033; e-mail: data_request@ccdc.cam.ac.uk or www: http://
www.ccdc.cam.ac.uk/data_request/cif). Supplementary data asso-
ciated with this article can be found, in the online version, at
http://dx.doi.org/10.1016/j.molstruc.2012.04.087.
(DEe = 29.3 vs 36.2 kcal/mol). However, the ‘‘catalytic’’ effect of
relay is significantly reduced due to entropic cost associated with
constructing the relay pathway with aid of an supramolecule com-
prising eight-membered ring. As a result water assisted F ? 4
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Appendix A. Supplementary material
Cartesian coordinates, computed electronic energies and Gibbs
free energies for structures 3, 4, B, C, D, E, F, TS1-5; HSQC and
HMBC NMR spectra of compound 4. Crystallographic data for the
structures reported in this paper have been deposited with the