Synthesis, X-ray structure, vibrational spectroscopy, DFT, biological evaluation and molecular docking studies of (E)-N’-(4-(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide

Author statement

Khalid Karrouchi: Conceptualization, Methodology, Original Draft, Writing - Review &

Editing.

Silvia A. Brandán : Theoretical, Calculations Writing - Original Draft, Writing - Review &

Editing.

Yusuf Sert: Theoretical, Calculations Writing - Original Draft.

Hakima El-marzouqi: Resources.

Smaail Radi: Supervision, Project administration.

Marilena Ferbinteanu: Software, Validation, Writing - Review.

My El Abbes Faouzi : Visualization, Investigation.

Yann Garcia: Supervision, Investigation

M’hammed Ansar: Supervision, Project administration.

Synthesis, X-ray structure, Vibrational spectroscopy, DFT, biological evaluation and

molecular docking studies of (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-1H-

pyrazole-3-carbohydrazide

a,*

b,*

c

d

Khalid Karrouchi , Silvia A. Brandán , Yusuf Sert , Hakima El-marzouqi , Smaail Radi

e

f

d

g

h

,

Marilena Ferbinteanu , My El Abbes Faouzi , Yann Garcia , M’hammed Ansar

a

b

c

Laboratory of Analytical Chemistry and Bromatology, Faculty of Medicine and Pharmacy,

Mohamed V University, Rabat, Morocco.

Cátedra de Química General, Instituto de Química Inorgánica, Facultad de Bioquímica, Química y

Farmacia, Universidad Nacional de Tucumán, Ayacucho 471, 4000, Tucumán, Argentina.

Sorgun Vocational School, Science and Art Faculty-Department of Physics, Yozgat Bozok University,

Yozgat, Turkey.

d

e

f

Laboratory of Pharmacology and Toxicology, Pharmacokinetic Research Team, Faculty of Medicine

and Pharmacy, University Mohammed V, Rabat, Morocco

Laboratory of Applied Chemistry and Environment (LCAE), Department of Chemistry, Faculty of

Sciences, University Mohamed Premier, Oujda 60000, Morocco.

Inorganic Chemistry Department, Faculty of Chemistry, University of Bucharest, Dumbrava Rosie

2

3, Bucharest 020462, Romania.

g

Institute of Condensed Matter and Nanosciences, Molecular Chemistry, Materials and Catalysis

IMCN/MOST), Université catholique de Louvain, Place L. Pasteur 1, 1348 Louvain-la-Neuve,

(

Belgium.

h

Laboratory of Medicinal Chemistry, Faculty of Medicine and Pharmacy, Mohamed V University,

Rabat, Morocco.

*

Corresponding authors:

Khalid.karrouchi@um5s.net.ma (K. Karrouchi)

brandansa@yahoo.com.ar (Silvia A. Brandán)

1

Abstract

New crystal, (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide

(3) has been synthesized and characterized by FT-IR, NMR, ESI-MS and single crystal X-ray

diffraction (XRD). The optimized molecular structures of free base and cationic species of (3)

in gas phase and aqueous solution, vibrational frequencies and, corresponding vibrational

assignments have been investigated experimentally and theoretically by using the B3LYP/6-

3

1G* and B3LYP/6-311++G** methods. High solvation energy values are observed for both

species of (3) in solution while the NBO and AIM studies support the higher stability of the

cationic species in solution. The high energy values E_σ_→_σ_*and E_σ_→_π_*transitions, due to the

∆

planarity of both CH groups linked to N atom, could support the high reactivities of its free

3

base and cationic species, as compared with naloxone, cocaine and scopolamine. Complete

vibrational assignments of 105 and 108 vibration modes expected for free base and cationic

species of (3) together with the corresponding harmonic force constants are here reported. In

vitro antidiabetic and antioxidant activities were revealed for (3). The molecular docking

studies of the title compound revealed that it may exhibit anti-diabetic activity via inhibition

of α-glucosidase PDB:3A4A enzyme.

Keywords: Pyrazole; Crystal structure; Vibrational spectroscopy; DFT; NBO; Antioxidant

activity; Antidiabetic activity; Molecular docking.

2

1

. Introduction

Hydrazones, carbohydrazides and similar derivatives represent an important class of organic

compounds of interest in the medicinal and pharmaceutical fields [1-11] because one of many

studies has reported that hydrazones improve the antitumor selectivity and toxicity profile of

antitumor agents by forming drug carrier systems employing suitable carrier proteins [5].

Other studies have evidenced that many pyrazole derivatives present therapeutic activity [12-

1

7], some are pesticides [18-20] and, other exhibit a wide gamma of biological properties,

such as anti-inflammatory, anti-cancer, antioxidants, antidepressants, antivirals, analgesics,

anti-parkinsons, anti-alzheimer, anti-glaucoma, anti-diabetic, anti-tubercular and anti-

leishmanial [21-35]. With this background set, the determinations of structural, electronic and

topological properties of these pyrazole derivatives are essential to know the influence of

different groups on the structures in order to understand the connections of these groups with

their biological properties. On the other hand, the vibrational analyses and, in particular the

vibrational assignments of all bands observed in the experimental infrared and Raman spectra

are of great aid to identify all species in any medium, especially when these studies are

combined with theoretical calculations derived from density functional theory (DFT) [36-39].

In the present work, a new crystalline derivative has been synthesized, (E)-N'-(4-

(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3) and, then it was

1

13

characterized by using FT-IR spectrum in the solid phase, H- and C-NMR spectra in

DMSO-d solution, ESI-MS and single crystal X-ray diffraction (XRD). These experimental

6

studies were accomplished with theoretical DFT calculations by using the functional hybrid

B3LYP together with the 6-311++G** basis set in order to predict the structural, electronic,

topological and vibrational properties in the gas phase [40,41]. Taking into account the wide

range of biological activities reported for pyrazole derivatives, this new derivative was here

evaluated by in vitro anti-diabetic and anti-oxidant activities. Hence, the reactivities of this

derivative in gas phase and in aqueous solution were predicted by using the frontier orbitals

with the B3LYP/6-311++G** method while their behaviours in both media were evaluated

calculating some descriptors useful for compound containing different type of rings [42-51].

All calculations in aqueous solution were performed with the self-consistent reaction field

(

SCRF) method by using the integral equation formalism variant polarised continuum

IEFPCM) and universal solvation models [52-54]. The bands observed in the experimental

(

infrared spectrum were assigned by using the harmonic force field calculated with the

SQMFF methodology and the Molvib program at the same level of theory [55-57]. Here, the

3

main scaled force constants by using the same level of theory were reported for this new

1

13

pyrazole derivative. Comparisons among experimental and predicted FT-IR, H- and C-

NMR spectra have showed reasonable concordance among them. In addition, the interactions

of the title molecule with α-Glucosidase PDB:3A4A and antioxidant peroxiredoxin 5

PDB:1HD2 receptors were investigated by molecular docking studies.

2

. Experimental section

.1. General methods

2

All chemical reactions were purchased Sigma-Aldrich. Reactions were checked with TLC

using aluminum sheets with silica gel 60 F254 from Merck. Melting points were measured

using a Buchi B-545 digital capillary melting point apparatus and used without correction.

The FT-IR spectrum was recorded with Perkin-Elmer VERTEX 70 FT-IR spectrometer

-1

1

13

covering ﬁeld 400–4.000 cm . H and C NMR spectra were recorded in solution in DMSO-

d₆, on Bruker spectrometer (300 MHz). The chemical shifts are expressed in parts per million

(ppm) by using tetramethylsilane (TMS) as internal reference. Mass spectra were collected

using API 3200 LC/MS/MS system, equipped with an ESI source.

2

.2. Synthesis

General procedure for the synthesis of (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-

H-pyrazole-3-carbohydrazide (3) :

1

Target compound (3) described here was facilely synthesized according to the literature

procedures [58-61] (Scheme 1). To a solution of 5-methyl-1H-pyrazole-3-carbohydrazide (1)

(1 mmol) in 10 mL of ethanol was added an equimolar amount of the 4-

dimethylaminobenzaldehyde (2) in the presence of two drops of acetic acid. The mixture was

maintained under reflux for 2 h, until TLC indicated the end of reaction. Then, the reaction

mixture was poured in cold water, and the precipitate formed was filtered out washed with

ethanol and recrystallized from ethanol.

-1

Yield 81 %, M.p. 259-261 °C; IR (ATR, ν(cm )) : 3233 (NH), 1648 (C=O), 1602 (N=CH);

1

H-NMR (300 MHz, DMSO-d , δ(ppm)): δ = 2.26 (s, 3H, CH ), 2.94 (s, 6H, N(CH ) ), 6.45

6

3

3 2

(s, 1H, H-pyrazole), 6.72 (d, J = 8.7 Hz, 2H, H-Ar), 7.46 (d, J = 8.7 Hz, 2H, H-Ar), 8.30 (s,

1

3

1

H, -NH), 11.24 (s, 1H, N=CH) 13.01 (s, 1H, NH-pyrazole) ; C NMR: (300MHz, DMSO-

d₆, δ (ppm)): 10.78, 40.28, 105.11, 112.26, 122.33, 128.76, 140.36, 146.60, 148.42, 151.84,

+

1

58.45. MS: m/z = 272.3 (M+H) .

4

Scheme 1. The synthetic route of compound 3.

.3. Single crystal X-ray diffraction

X-ray single crystal data were collected by using MoKα (

2

λ

= 0.71075 Å) radiation on a

Rigaku R-Axis Rapid II diffractometer equipped with an large-area curved imaging plate

detector (460.0 x 256.0 mm), 3-circle goniometer and high-frequency 5 kW sealed tube. All

calculations were performed using the CrystalStructure [62] crystallographic software

package except for refinement, which was performed using the software package Olex1.2

[

63]. The structure was solved by direct methods and refined in a routine manner (SHELXL)

64]. Non-hydrogen atoms were refined anisotropically. Hydrogen atoms were refined using

the riding model. Molecular graphics were generated by using MERCURY 3.9 [65] and POV-

Ray software [66]. The details of the X-ray crystal data and the structure solution as well as

the refinement are given in Table 1. A summary of selected bond lengths [Å] and angles [°]

are given in Table S1 (ESI). CCDC 1989792.

2

.4. Computational details

The initial theoretical structure of (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-1H-

pyrazole-3-carbohydrazide (3) was that experimental determined by X-ray crystal data taken

from the corresponding CIF file. The optimizations in gas phase and in aqueous solution were

performed with the Revision A.02 of Gaussian09 program [67] and the hybrid B3LYP/6-

3

11++G** method [40,41]. The cationic species of (3) was also optimized because it is

expected in solution. The IEFPCM and universal solvation methods were employed in the

optimization of (3) in solution because the solvent effects are considered with both models

[

52-54]. The Moldraw program was used to compute the volumes in both media [68]. Atomic

Merz-Kollman (MK) [69] and natural population analysis (NPA) charges, molecular

electrostatic potential (MEP) and stabilization energy were predicted with the version 3.1 of

NBO program [70,71] while the topological properties were computed by using the AIM2000

program [72]. The mapped MEP surface of (3) was generating with the version 5.0 of

GaussView program [73] while the normal internal coordinates together with the SQMFF

methodology and the version 7.0 of Molvib program [55-57] were employed in the vibrational

5

study. To perform the assignments, only potential energy distribution (PED) contributions ≥

0 % were considered. The predicted Raman spectrum in the gas phase in activities was

1

corrected to intensities with the equations suggested in the literature [74]. The Ultraviolet-

visible spectrum was also predicted in aqueous solution by using the Time-dependent DFT

calculations (TD-DFT) [67] by using NStates=100 and the B3LYP/6-311++G** method. The

1

13

predicted H- and C-NMR spectra in solution by using the GIAO method [75] with the

hybrid B3LYP/6-311++G** method were compared with those experimental obtained in

DMSO-d solution. Finally, the reactivities and behaviours of (3) in both media were

6

predicted with the frontier orbitals calculated with the B3LYP/6-311++G** level of theory.

From the difference between both orbitals were computed the gap values and the chemical

potential (μ), electronegativity (χ), global hardness (η), global softness (S), global

electrophilicity index (ω) and nucleophilicity indexes (Ε) descriptors [36-39,42-51].

2

.5. Antidiabetic activity

The α-glucosidase, β-galactosidase and α-amylase inhibition assays were conducted according

to previously reported protocols [31, 76].

2

.6. Antioxidant activities

The antioxidant activities of the title compound were determined in vitro by 2,2-diphenyl-1-

picrylhydrazyl (DPPH), 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid (ABTS) and

ferric reducing antioxidant powerꢀ(FRAP) methods according to the procedures described in

our previous work [31].

3

. Results and discussion

.1. X-ray crystal structure description

The compound 3 was analysed by single crystal X-ray diffraction. The summary of

crystallographic information is listed in Table 1. The compound 3 crystalized in the

monoclinic space group P2 /c. The molecule is almost planar and forms dihedral angles

1

between the central plane of the carbohydrazide moiety and the plane of the pyrazole ring

o

(

6.64 ) or the plane of the benzene ring (11.66 ). The unit cell contains four molecules. The

crystal packing (Figure 1) shows that two by two molecules are arranged in parallel plans

within a distance of ~10.2Å and there are no supramolecular interactions between them.

Crystal packing for 3 along a, b and c axes are shown in Figures S1-S3.

3

.2. Optimization geometry and properties in both media

6

Calculated total and corrected by ZPVE energies, dipole moments and volumes of (E)-N'-(4-

dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3) as free base and

(

cationic species in gas phase and in aqueous solution are presented in Table 2 by using the

B3LYP/6-311++G** and B3LYP/6-31G* methods while the theoretical optimized structure

for the free base can be seen in Figure 2 together with the atoms labelling. The cationic

structure is presented in Figure S4. The phenyl ring is identified as R1 in those two species of

(3) while R2 is designed to the pyrazole ring. The analyses of the results show a very

important contraction in the volume of (3) as free base in aqueous solution and a strong

increase in the dipole moment value is also observed with the B3LYP/6-311++G** method.

On the contrary, an expansion in the volume of free base of (3) in aqueous solution is

observed with the other method and a slight diminishing in the dipole moment value while the

cationic species with the B3LYP/6-31G* method reveals a decrease or contraction of volume

in aqueous solution. The expansion or contraction of volume observed for both species of (3)

in solution can be easily attributed to the presence of group donors of H bonds, such as the O

and N atoms and, to the presence of acceptors N-H groups in the structure of (3). Here, the

formations of H bonds in solution justify the variations of those two properties in solution

because (3) is clearly a weak base. Table S1 shows the values of Mulliken, Merz-Kollman and

NPA charges, molecular electrostatic potentials (MEP) and bond orders, expressed as Wiberg

indexes of free base of (3) in gas phase and in aqueous solution by using B3LYP/6-311++G**

level of theory. In general, we observed that the MK and NPA charges present practically the

same signs on the five N atoms and the only O atom but the values are different between them

and different from the Mulliken charges. However, the NPA charges practically do not

present changes in solution, as compared with the corresponding values in gas phase.

Regarding the atomic charges in solution, it is observed that the cationic species of (3) could

be formed by the protonation of N11 and N23 atoms but the higher Mulliken charges predict a

higher value on N11 atom while the MK charges predict a most negative value on N23 (-

0

.493 a.u.) than the corresponding to N11 (-0.245 a.u.) indicating, this way, that the

protonation could occur in the N23. Here, the calculations performed for both probable

cationic species have shown all positive frequencies when the optimized cation is protonated

in the N23 while two imaginary frequencies are obtained when the cation is protonated in the

N11. Consequently, the cationic species of (3) in solution indicate the protonation of N23

atom.

7

Other interesting property studied here for (3) are the molecular electrostatic potentials (MEP)

calculated from the MK charges [69] on the N and O atoms whose results are given in Table

S1. Analyzing the MEP values on those atoms obtained for the free base of (3) by using the

B3LYP/6-311++G** level of theory it is observed few changes in solution, but when the

mapped MEP surface is evaluated for (3) in gas phase with the other basis set from Figure S5

different colorations are clearly observed on its surface. Thus, the strong red colours on the

O16, N11 and N23 atoms indicate nucleophilic sites while on the H14 and H22 atoms that

belong to the N13-H14 and N19-H22 bonds are observed blue colours that clearly indicate

electrophilic sites. Hence, reactions with potential electrophiles or nucleophiles biological

reactive take places on those two important reaction sites.

If now, the bond order (BO), expressed as Wiberg index are analyzed for (3) from Table S1, it

is observed that N19 atom presents the higher value in both media because this atom is linked

to the most labile H22 atom and, for this reason, the strong blue coloration on the mapped

MEP surface it is observed on this atom. In general, the values for all atoms decrease in

solution with exception of BO for the N11 and N23 atoms which increase in this medium due

to the hydration with water molecules.

Taking into account the changes predicted for (3) in solution, corrected and uncorrected

solvation energies by the total non-electrostatic terms and by zero point vibrational energy

(

ZPVE) of free base and cationic species of (3) by using the B3LYP/6-311++G** and

B3LYP/6-31G* methods are compared in Table 3. Comparisons of corrected solvation energy

∆G ) for those two species of (3) with those predicted for the free base and cationic species

(

c

of naloxone [77], cocaine [78] and scopolamine [79] by using the B3LYP/6-31G* method are

shown in the same table because these three species show similar solvation energy values in

aqueous solution although they present different biological properties. On the other hand, the

structure of (3) has two CH groups in the N33 atom, as also is observed in the structures of

3

2

cocaine [78] and scopolamine [79] but, in (3) that N23 atom has sp hybridization, different

3

from the other compounds with sp hybridization and only one CH group. On the contrary,

3

naloxone also presents the N atom in sp hybridization but linked to an allyl chain and to two

CH of ring [77]. Structures of compared compounds can be seen in Figure S6. Probably, the

2

sp hybridization of N23 atom in (3) could justify the differences in the biological properties

observed for this species, in relation to the other compared species [77-79]. Despite the

calculations were performed with different methods, the values show that (3) as free base

presents a higher ∆G (-117.85 kJ/mol by using the 6-31G* basis set and -129.50 kJ/mol by

c

8

using the other basis set) than the other ones while the cationic species evidence higher

values, as expected because these species are charged and, for these reasons, they are hydrated

in solution. Note that the cationic species of (3) presents a ∆G value between the naloxone

c

and scopolamine species.

3

.4. Geometrical parameters

The optimized geometrical parameters of (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-

H-pyrazole-3-carbohydrazide (3) as free base in gas phase and aqueous solution by using the

1

B3LYP/6-311++G** method are presented in Table 4 together with the corresponding

experimental ones determined in this work. Root-mean-square deviation (RMSD) values are

also included in that table to evaluate the differences between theoretical and experimental

results. Hence, good correlations are observed for bond lengths and angles with RMSD values

between 0.018 and 0.012 for bond lengths and of 1.3 and 1.2 for bond angles. The higher

deviations are clearly observed for dihedral angles because the C1-C12-N11-N13 dihedral

angle in gas phase presents a negative value while change at positive in solution, in

accordance to experimental one. On the contrary, the C12-N11-N13-C15, N13-C15-C17-N23

and C15-C17-N23-N19 dihedral angles are predicted in both media with negative signs but

experimentally they have positive signs and different values. A contrary resulted is observed

for the other two N13-C15-C17-C18 and O16-C15-C17-N23 dihedral angles because they are

predicted in both media with positive signs and experimentally both present negative signs

and different values. Evidently, the changes in the geometrical parameters of (3) in solution

are related to its hydration and to high solvation energy observed in this medium. The good

correlations evidenced in the bond lengths and angles, despite the differences observed in

some dihedral angles, indicate that the optimized structures of (3) in both media by using the

B3LYP/6-311++G** method can be used to perform the vibrational analysis and the

determination of force fields in the two media.

3

.5. NBO and AIM studies

The study of stability of (3) in both media is a very important factor taking into account that

structurally this species presents acceptors and donors groups and, also reveals in vitro

antidiabetic and antioxidant activities. Both NBO and AIM calculations are tools useful to

investigate the presence of different interactions by using respectively the Second Order

Perturbation Theory Analysis of Fock Matrix in NBO Basis and the topological properties

according to the Bader’s theory of atoms in molecules (AIM) [70-72]. In the first study the

9

version 3.1 of NBO program was employed [70]. Hence, main delocalization energies for the

free base of (3) by using the B3LYP/6-311++G** method in gas phase and aqueous solution

are summarized in Table S2. The evaluation of results shows that for (3) in gas phase are

observed only four interactions:

energy values while in solution the

E_σ_→_σ_*and E_σ_→_π_*transitions with surprisingly high energy values are observed. In these

∆E_π_→_π_*,

∆E_L_P_→_σ_*, ∆E_L_P_→_π_*and ∆E_π_*_→_π_*interactions with low

∆E_π_→_σ_*interaction together with other two additional

∆

two latter interactions are involved transitions of both rings and of N33 atom with both CH₃

groups. Hence, in total six interactions are observed for (3) in aqueous solution. Therefore, the

total energy value highly favours to the free base of (3) in solution whose value justifies its

high solvation energy value and high stability in water. Probably, this resulted indicates that

the free base species of (3) is as a cationic species in solution because it is hydrated.

NBO analyses have shown high stability of (3) in solution, as compared with the values in gas

phase and, for this reason, the presences of other inter or intra-molecular interactions of free

base should be investigated in both media by using the topological properties, according to the

Bader’s theory [71]. To perform these calculations, the version 2000 of AIM program was

used [72]. Hence, in Table S3 is presented the analysis of the electron density, ρ(r), the

2

Laplacian values, ∇

ρ(r), the eigenvalues (λ1, λ2, λ3) of the Hessian matrix and, the |λ1|/λ3

ratio calculated in the Bond Critical Points (BCPs) and Ring critical point (RCPs) for the free

base of (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3)

in gas and aqueous solution phases by using the B3LYP/6-311++G** method. These results

show that in gas phase only the two expected RCP1 of ring 1 (R1) and RCP2 of ring 2 (R2)

are observed while two Bond Critical Points (C4-H5···H31 and C7-H8···H35) and two new

Ring critical points (RCPN1 and RCPN2) appear in solution. Obviously, those two BCPs

present the characteristics of an interaction ionic or highly polar covalent where λ1/λ3< 1 and

2

∇

ρ(r) > 0 (closed-shell interaction). In Figure 3 can be seen the molecular graphics of (3) in

aqueous solution showing the two BCPs C4-H5···H31 and C7-H8···H35, two RCP1 and

RCP2 and, the two RCPN1 and RCPN2. The presence of two BCPs in solution predict the

high stability of (3) in this medium, as also was predicted by NBO calculations.

3

.6. Vibrational study

Both B3LYP/6-31G* and B3LYP/6-311++G** methods have optimized the structures of free

base and cationic species of (3) in gas phase and aqueous solution with C symmetry and due

1

to the presence of 37 atoms in the structure of free base 105 vibration modes are expected for

1

0

this species while for the cationic one are expected 108 vibration modes. The vibration modes

of both species present activity in both spectra. The experimental FT-IR spectrum of the title

compound was recorded in a solid state using reflectance (ATR) mode. A comparison of

experimental ATR spectrum of free base of (3) in the solid phase with the corresponding

predicted for free base and cationic species in gas phase by using the B3LYP/6-311++G**

-1

method can be seen in Figure 4. The group of IR bands between 2867 and 2633 cm together

-1

with the decreasing in the intensity of IR band at 1742 cm attributed to C=O stretching

mode, predicted with high intensity by SQM calculations, could probably justify the presence

of cationic species of (3). According Figure 4, the predicted ATR spectrum of cationic species

shows higher and lower intensities of bands associated respectively to the stretching modes of

N-H group of pyrazine ring and C=O bond. On the other hand, the higher number of predicted

-1

IR bands in approximately the same 4000-2000 and 2000-0 cm regions could also support

the presence of cationic species in the solid phase. In the determination of force fields for the

two species of (3) the normal internal coordinates were employed together with the SQMFF

methodology and the version 7.0 of Molvib program [55-57]. In the assignments, only

potential energy distribution (PED) contributions ≥ 10 % were considered. The predicted

Raman spectra for those two species in the gas phase in activities were transformed to

intensities and both are observed in Figure S7. These transformations were performed with

the equations suggested in the literature [74]. In Table 5 are summarized observed and

calculated wavenumbers and assignments for the free base and cationic species of (3) in gas

phase by using the B3LYP/6-311++G** method. A brief discussion by regions is presented

below.

Band Assignments

-1

4

000-2000 cm region. This region is typical of N-H, C-H and CH stretching modes. Here,

3

-1

the free base presents two N-H stretching modes (predicted at 3494 and 3348 cm ) while for

-1

the cationic species are expected three N-H stretching modes at 3401, 3316 and 2889 cm

where this latter band is attributed to N19-H22 bond of pyrazine ring. In the cationic species,

the N19-H22 stretching mode is predicted very intense in gas phase by B3LYP/6-311++G**

calculations. In heterocyclic compounds, the N–H stretching vibrations appears strongly in the

−

1

region 3500–3000 cm [36,50,80]. Pillai et al. [81] have reported NH stretching bands at

-1

3

517 cm for pyrazole and at 3388 cm for carbohydrazide (-CONH-N=). In the present

−

1

study, the FT-IR band appears at 3232 cm is assigned to N–H stretching modes of vibrations

for pyrazole and carbohydrazide, respectively. The C-H stretching vibrations of aromatic

1

-1

rings give rise to bands in the region 3200–3000 cm in aromatic compounds [38,43-48,77-

0]. The C-H stretching modes for the two species are predicted by SQM calculations in

approximately the same regions and, for these reasons, the ATR bands between 3141 and

8

-1

2

633 cm are assigned to these vibration modes. Here, the band at 3116 cm is assigned to C-

H stretching vibrations of pyrazole ring, which is in good agreement with the values reported

for pyrazole derivatives [81]. The CH stretching modes for para-substituted benzenes are

-1

found in the region 3100-3000 cm [43-48,77-80]. The series of IR bands between 3026 and

-1

2

915 cm were assigned as CH stretching modes of the 4-dimethylaminobenzyl ring. The C-

-1

H stretching mode of azomethine appears as a weak band at 2867 cm in the infrared

-1

spectrum. Methyl stretching vibrations are observed in the region 2975-2765 cm [36,43,45-

4

7,49-51,77-80]. Note that in the cationic species the symmetric CH stretching modes of

3

-1

three groups are predicted at 2810, 2796 and 2776 cm while for the free base are not

predicted bands in this region, for these reasons, the presence of cationic species in the solid

-1

phase could be clearly justified with the experimental IR bands at 2806, 2723 and 2633 cm .

The presence of an adjacent group such as a N or O atom can result in a significant frequency

shift in the methyl (CH ) in-phase stretch to lower frequency. Due to the absence of

3

experimental Raman spectrum the symmetries of corresponding CH stretching modes were

3

not confirmed.

-1

2

000-1000 cm region. In this region are expected the C=O, C=C, C-C and C-N stretching

modes, deformation and rocking modes of NH, CH and CH groups and, some deformations

3

of pyrazole and 4-dimethylaminobenzyl rings can also be observed. The C=O stretching mode

is usually one of the most representative in an infrared spectrum, it appears in a wavenumber

-1

region relatively free of other vibrations (1800–1600 cm ) [45-47,77-79]. This mode was

-1

assigned at 1691 cm by Pillai et al. [81]. In our study, the SQM calculations predict that

-1

stretching mode in the free base as an intense band at 1711 cm while in the cationic species

-

1

this mode is also predicted intense at 1604 cm . On the other hand, the C7-H8 in-plane

deformation mode in free base is predicted with double intensity, as compared with the C=O

vibration, according Figure 4. However, such prediction is no experimentally observed

probably due to the inter-molecular hydrogen bonds between N1-H1···O1i, as determined in

the experimental structure. Here, it is necessary to clarify that the theoretical calculations were

performed in the gas phase where the packing forces were not considered, hence, the

differences among the predicted spectra and the corresponding experimental one. Then, the

-1

C=O stretching modes for both species can be assigned to the strong IR band at 1648 cm .

1

2

-1

The C=C stretching of both species are predicted between 1611 and 1502 cm , hence, these

-1

modes are assigned to group of intense IR bands between 1648 and 1522 cm . Pillai et al.

-1

[

81] have reported the stretching vibrations νC=N at 1553 cm and νN-N at 1108 cm in

pyrazole derivative while in the carbohydrazide the C=N and C-N stretching bands are

-1

expected in the range 1672-1566 cm and 1275 ± 55 cm , respectively [82]. Therefore, the,

-1

intense bands observed at 1611 and 1549 cm are assigned to C12=N11 stretching modes of

free base and cationic, respectively. In the free base, the C-N stretching mode is assigned at

-1

1

549 cm and C-N stretching mode is assigned at 1250 cm . The N-N stretching mode has

-1

been reported at 1118 cm by Sheeja et al. [82] at 1066 cm by Govindarasu et al. [83] and,

-1

at 1156 cm by Freitas et al. [84]. The N19-N23 and N11-N13 stretching modes in the free

-1

base and cationic species of (3) are assigned respectively at 1170 and 1062 cm and, at 1182

-1

and 1020 cm , as predicted by calculations. In the cationic species the N11-N13 stretching

-1

mode can be also assigned to the very strong IR band at 1128 cm because in that position

this mode is observed coupled with other vibration mode.

In para substituted benzene, the C-H in-plane bending or deformations (β) vibrations are

-1

observed in the region 1400-1000 cm and are usually of medium to weak intensity [36,49-

1,77-79,81]. Here, some bands due to C–H in-plane bending vibration in both species

interact somewhat with other vibrations and they can be assigned to the bands in the region

5

−

1

between 1475 and 1136 cm . The N13-H14 in-plane deformation corresponding to

carbohydrazide group in both species is predicted at higher wavenumbers than the other N19-

-1

H22 and N23-H38 ones. Hence, the IR bands at 1549, 1408 and 1255 cm are assigned to

those vibration modes.

The deformation and rocking modes of the CH groups in both species can be assigned

3

-1

respectively to the IR bands observed between 1461/1338 and 1158/961 cm , as predicted by

calculations and as observed in compounds with similar groups [45,47,50,78,79].

-1

1

000-10 cm region. In this region, the C-C and C-N stretching modes, C-H and N-H out-of-

plane deformations, deformations, wagging and twisting modes of C=O group, twisting CH₃

and deformations and torsions modes of both pyrazole and 4-dimethylaminobenzyl rings are

expected. These vibration modes were assigned taking into account the SQM calculations

performed here and by comparison with assignments for similar species [18-25,34-44]. A

detail is presented in Table 5. The C=O in-plane deformation and the out-of-plane

-1

deformation are expected in the regions 625 ± 70 and 540 ± 80 cm , respectively [81]. Here,

1

3

-1

the IR bands observed at 904 and 742 cm are assigned to these modes corresponding to free

-1

base while these modes for the cationic species are predicted at 667 and 362 cm , for which,

they can be assigned in this region.

-1

The out-of-plane CH deformations are observed between 900 and 600 cm [36,37,39,45,48-

1,77-80]. Generally the CH out-of-plane deformations typical for para substituted benzenes

5

-1

are assigned in the 840 ± 50 cm region [83]. In the present work, these vibration modes in

-1

the both species can be associated to the IR bands observed between 983 and 765 cm while

the corresponding N-H out-of-plane deformations are predicted and assigned to the bands

-1

observed between 734 and 501 cm . The remaining vibration modes are assigned within the

characteristic region and reported in Table 5.

3

.7. Force Fields

The character of different bonds can be also descript by the harmonic force constants and, for

these reasons, for free base and cationic species of (3) these parameters were calculated from

the corresponding harmonic force fields expressed in internal coordinates. Hence, the SQMFF

procedure and the Molvib program were employed [55-57]. In Table 6 those constants for

both species in gas phase by using the B3LYP/6-311++G** method were compared with the

reported for the free base and cationic species of naloxone and scopolamine in the same

medium by using the B3LYP/6-31G* method [77,79]. Comparing first both species of (3), in

the cationic species it is observed that due to additional N23-H38 bond the value of its force

-

1

-1

constant obviously decreases from 6.42 mdyn Å in the free base to 5.72 mdyn Å in the

cationic one. Also, the f( C=O) force constant in the cationic species decreases, as compared

ν

with the corresponding to the free base. This difference in the value can be clearly attributed

to the bond lengths between both involved atoms because in the free base the predicted C=O

distance is 1.209 Å while in the cationic ones is 1.248 Å. Comparisons with the other two

species show that the free base of (3) presents approximately the same value than that

observed in scopolamine. On the other hand, the f(

dimethylaminobenzyl ring in the two species of (3) have practically the same values than

those observed for naloxone and scopolamine while the f( C-H)_R₂force constants for the

pyrazole ring decrease slightly in the cationic species, in relation to the free base [77,79]. The

f( N-CH ) force constants have practically the same values in both species of (3) but in

naloxone and scopolamine the values change because its cationic species are formed in the N

atom that contain the CH groups [77,79]. Note that the f( C-N)_C_h_a_i_nforce constant of cationic

νC-H)_R₁force constant of 4-

ν

3

ν

3

1

4

species presents a lower value than that observed for the free base, indicating this way, that

the carbohydrazide group is influenced by the N23-H38 bond. In the same way, the f( C-N)_R,

f( N-N) , f( N-N)_C_h_a_i_nand f( C=C)_R₂force constants change in the cationic when they are

compared with the corresponding to free base. Evidently, the f( N-N) force constant (3.90

ν

R

ν

R

-1

mdyn Å ) in the cationic species is most influenced that f(

ν

N-N)_C_h_a_i_nforce constant of

carbohydrazide group (6.73 mdyn Å ), as expected because the N23-H38 belong to pyrazole

ring. Moreover, the cationic species generate a increase in the f( C=C) force constant of

-1

ν

R2

pyrazole ring, as compared with the corresponding to f(νC=C)_R₁force constant of 4-

dimethylaminobenzyl ring.

3

.8. Ultaviolet-visible spectra

The electronic ultraviolet-visible spectra of free base and cationic species of (3) in aqueous

solution were also predicted by using the B3LYP/6-311++G** method. The electronic spectra

predicted for those two species can be seen in Figure S8. The predicted spectrum for the free

base shows a maximum at 343.6 nm with a shoulder in c.a. 268.4 nm while for the cationic

form the maximum can be observed in c.a. 225 nm. The intense observed band in the

spectrum of free base and, expected for the cationic one, can be assigned to π→π* transitions

due to the presence of C=C double bonds of both pyrazole and 4-dimethylaminobenzyl rings

and to carbohydrazide group and, also can be assigned to n→π* transitions which are

predicted for the two species by NBO calculations (Figure S9). In the UV-visible spectrum of

cationic form the π→π* and n→π* transitions are predicted with low intensities as compared

with the intense band attributed to σ→σ* and σ→π* transitions by using NBO analyses.

Here, the high values of these latter two transitions predicted for the cationic species overlap

to the π→π* and n→π* transitions. On the other hand, the shoulder observed in the predicted

UV-Vis spectrum for the free base in aqueous solution could be attributed to the cationic form

because it species is also expected in solution. These assignments for both species are in

agreement with those reported for species containing similar rings [45-47,51].

3

.9. NMR studies

1

13

The H and C NMR spectra of the title compounds were obtained by using TMS as an

1

13

internal standard and DMSO-d as solvent. Both H and C NMR spectra can be seen in

6

Figures S10 and S11, respectively while the chemical shifts are summarized in Tables 7 and 8

compared with the corresponding theoretical ones for the free base and cationic species in

1

5

1

aqueous solution by using the B3LYP/6-311++G** and GIAO methods [75]. The H NMR

1

spectra corresponding to compound (3) are presented in Figure S10, respectively. The H

NMR spectra of the title molecule displayed a two singlets at δ 2.26 and 2.94 ppm due to CH₃

and N(CH ) protons, respectively. The chemical shifts of pyrazole proton (C4-H) appear as a

3

2

singlet at δ 6.45 ppm. The chemical shifts of the phenyl protons appeared as two doublets at δ

.72 and 7.46 ppm. The chemical shifts of the azomethine (N=CH) and amide (NHCO)

protons appear as a singlet at 8.30 and 11.24 ppm, respectively. The chemical shifts of NH

6

1

3

pyrazole proton appeared as singlet at δ 13.01 ppm. The C NMR spectra of the title

compound showed the chemical shifts of C=O are at 158.45 ppm. The signals at 148.42 ppm

are clearly assigned for azomethine group (N=CH) chemical shifts. The signals at 105.11,

1

46.60 and 151.84 ppm are assigned for pyrazole carbons, while that the aromatic carbon

chemical shifts of the compound occurred in the range of 112.26-140.36 ppm. In general,

good correlations with low RMSD values are found when the theoretical chemical shifts for

free base are compared with the corresponding experimental ones by using the root-mean-

square deviation (RMSD) values. However, for the cationic species are observed higher

RMSD values than those predicted for free base, thus, the low δ value predicted for δ of H22

(2.8 ppm) could be attributed to H bond formation in solution while the high RMSD value of

the chemical shifts of C atoms in the cationic species is due to δ of C17 (108.2 ppm) probably

because it atom is linked to N23 atom which could be protonated in aqueous solution.

3

.10. ESI-MS study

The ESI-MS spectra show molecular ion peaks with m/z values 272.2 correspond to the

+

molecular weight [M+H] . The m/z value 294.4 correspond to the sodiated molecular ion

+

peak [M+Na] (Figure S12). These values are in good agreement with the proposed

composition for the title molecule (C H N O).

1

4

17

5

3

.11. Hirshfeld surface analysis

The Hirshfeld surface analysis of the (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-1H-

pyrazole-3-carbohydrazide (3) was performed with Crystal Explorer 3.1 program [85] to

determine the possible molecular packing, intra and intermolecular hydrogen bond

interactions in the structure. Additionally, this analysis allows the visualization of

intermolecular interactions by such as red, blue and white different colours [86, 87]. As an

input file in the program, Crystallographic Information File (cif*) of the compound was used.

In Figure 5 was indicated d_n_o_r_mmapped on Hirshfeld surface for visualizing the intercontacts

1

6

of the title compound and d_n_o_r_mvalue was obtained as -0.2548 (red points) to 1.2144 a.u.

blue colors). As seen from the Figure 5, the dark red points near the C, O, N atoms result

(

from C=O…H interactions with 2.273 Å and 2.782 Å and N-H…N interactions with 2.337 Å

were observed and these regions have a significant role in the molecular packing.

Additionally, the two-dimensional fingerprint plots with their relative contributions or

percentage contributions to the Hirshfeld surface were shown in Figure 6. The most important

interactions were determined with H⋯H (51.2%), C⋯H/H⋯C (18%), N⋯H/H⋯N (12.4%),

O⋯H/H⋯O (11%), and N⋯C/C⋯N (5.6%) contributions. Lastly, the percentage

contributions of other intermolecular contacts are less than 2% in the Hirshfeld surface.

3

.12. Antidiabetic activity

Inhibition of α-glucosidase and α-amylase can retard carbohydrate digestion, thus causing a

reduction in the rate of glucose absorption into the blood. Therefore, inhibition of these

enzyme activities in digestive organs is considered to be a therapeutic approach for managing

diabetes (type 2). β-galactosidase is a tetrameric enzyme of historical and scientific

importance. Moreover, the hydrolysis of lactose gives galactose and glucose. Intramolecular

galactose transfer yields allolactose, the natural inducer of the lac operon. In the context, the

title compound was evaluated in vitro against α-glucosidase, α- amylase and β-galactosidase.

The result summarized in Table 9 and compared to the reference drug Acarbose and the

phenolic compound Quercetin.

From the observed results in Table 9, the in vitro α-glucosidase inhibition study shows that

the product 3 show a better activity for a concentration of 0.08 mM with percent inhibition of

7

9.83%, higher than that of the Acarbose as a reference (29%). The results of the β-

galactosidase test reveal that the title compound has a good inhibitory activity with a

percentage of 64.6%, comparable to that of Quercetin (68%) for a concentration of 3.30 mM.

For the alpha-amylase inhibition test, the results obtained show that the compound 3 have a

mean inhibitory activity (20.51%) relative to the Acarbose with a percentage of 36% for a

concentration 3.53 mM.

3

.13. Antioxidant activity

The antioxidant activity of the synthesized compound 3 has been systematically evaluated

using three different assays at 3.70 mM. The scavenger capacity is determined by measuring

the decrease in the absorption of the DPPH and ABTS radicals, and the results are expressed

as Trolox equivalents (TE) (μg TE / mg of compound). Meanwhile, we have also evaluated

1

7

3

+

2+

the ability of synthesized compounds to reduce Fe to Fe by using the Ferric Reducing

Antioxidant Power (FRAP) test. These three assays are mainly used to measure the direct

involvement of the compound in improving the primary antioxidant activity.

One of the most widely used methods for evaluating total antioxidant activity is the

determination of DPPH% trapping activity by its simple, rapid, sensitive and reproducible

procedure. The DPPH reagent allows us to determine the intrinsic capacity of a substance

having groups (RH) such as -NH to give a hydrogen atom or electrons. In our study, the

results are expressed as Trolox equivalents (ET) (μg ET / mg of compound), the tile

compound 3 present a promising free DPPH scavenging capacity with 3.65 trolox equivalent

(

Table 10).

The ABTS method is based on the ability of hydrogen or electrondonating antioxidants to

decolorize the performed radical monocation of

2,2′‑azino‑bis(3‑ethyl‑benzthiazoline‑6‑sulfonic acid) generated due to oxidation of ABTS

(

with potassium persulfate. The radical scavenging abilities showed by the tested compounds

towards this assay typically revealed that, the title molecule 3 exhibit moderate radical

scavenging ability, expressed as trolox equivalents (μg Trolox/mg of compound) with value

5

.02 trolox equivalents, respectively. Subsequently, we also evaluated the capacity of the

3

+

2+

synthesized compounds for reducing Fe to Fe by employing the ferric reducing antioxidant

power (FRAP) test. The compound 3 displays the higher effect in reducing ferric to ferrous

iron, expressed as ascorbic acid (AA) equivalents (μg AA/mg of compound) reaching more

than 7.03 ascorbic acid equivalents (Table 10).

3

.14. Frontier orbital and descriptors

The frontier orbitals studies of (3) are of great importance due to the antidiabetic and

antioxidant activities that this synthesized species has revealed. Hence, the gap energies in

both media were calculated for the free base from the differences between the highest

occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) by

using B3LYP/6-311++G** level of theory [42]. Then, the chemical potential (μ),

electronegativity (χ), global hardness (η), global softness (S), global electrophilicity index (ω)

and global nucleophilicity index (E) descriptors were also computed because these parameters

are useful to predict the chemical reactivity and kinetic stability of these species [36-39,43-

5

1]. Thus, a low gap value indicates that the species presents high chemical reactivity and low

kinetic stability [42]. In Table S4 are presented HOMO and LUMO orbitals, gap values and

descriptors for free base of (3) in gas and aqueous solution phases by using the B3LYP/6-

1

8

3

11++G** method. The values for the free base of (3) were compared with those reported in

gas phase for the same species of naloxone, cocaine and scopolamine in gas phase by using

the B3LYP/6-31G* method [77-79]. Regarding the results for (3) in both media we observed

a slight increases in the reactivity of free base of (3) in water and comparing with the values

for the other three species, clearly, (3) is most reactive than naloxone, cocaine and

scopolamine. The scopolamine species is the less reactive while the free base of (3) in

aqueous solution has higher reactivity. The formation of cationic species of (3) in solution

could justify the high reactivity due to its higher stability in solution, as suggested by NBO

and AIM calculations. Probably, the higher reactivity of (3) and its antioxidant and

antidiabetic activities can be attributed to N(CH ) group because both CH groups are planar

3

2

3

2

in (3) with the N33 atom in sp hybridization, different from the other compared compounds.

Hence, the higher reactivity of (3) in water is in agreement with the higher solvation energy

values of free base and cationic species in solution. In relation, to the descriptors it is

observed that both global electrophilicity (ω) and global nucleophilicity (E) indexes present

higher and lower values, respectively in (3), as compared with the naxolone, cocaine and

scopolamine [77-79]. These differences probably could be related to the dispositions planar of

two CH groups in (3) while in those three alkaloids are no planar.

3

.15. Molecular docking

The docking computations are very important techniques in structure-based drug design and

this method can estimate the binding-conformation mode within the target protein [88, 89]. In

this section, firstly having medical and pharmacological importance (E)-N'-(4-

(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3) molecule was

optimized with B3LYP/6-311++G(d,p) method/basis set and was recorded as Protein Data

Bank (PDB) format. In second step, the target proteins PDB:3A4A (α-Glucosidase) and

peroxiredoxin 5-PDB:1HD2 (antioxidant) were identified with the help of literature and

experimental activity study and they were obtained from the Protein Data Bank [90].

Additionally in the proteins water molecules and co-factors were cleaned and recorded PDB

format. Here the PDBQT formats of ligand and two proteins were prepared with Discover

Studio Visualizer 4.0 (DSV 4.0) software [91]. The AutoDock Vina program [92] was used

for molecular docking calculations. The docking results obtained during this research were

evaluated and were described as follows:

1

9

In the first instance, the active sites of α-Glucosidase protein-PDB:3A4A were determined as

ARG442, ASP352, HIS351, GLU277, VAL216, ASP215, ARG213, PHE178, PHE159,

HIS112, TYR72, ASP69, ASP38, TRP36, ASP34, ASN32 and ASP30 and according to these

active residues the grid boxes were determined as centre_x=33.212, centre_y=-9.817,

centre_z=22.508, size_x=88, size_y=46, size_z=50, spacing=0.375. The obtained docking

results for (3)-PDB: 3A4A were given in Table 11 and Figure 7. From the experimental

activity and theoretical docking results, the best results were observed in the interaction

between (E)-N'-(4-(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3)

molecule and α-Glucosidase protein-PDB: 3A4A. According to the theoretical affinity

binding energies, the best binding was determined with -7.9 (kcal/mol) energy and active two

hydrogen bonding. These two active hydrogen bondings were seen between ASP215 and H3

atom with 2.03 Å bond distance, between ARG442 and O1 atom with 3.11 Å bond distance.

Additionally van der Waals, attractive charge, carbon-hydrogen bond, π-cation, π-anion, π- π-

T shaped and π-alkyl interactions were determined from the Figure 7.

In the second instance, the active sites of antioxidant peroxiredoxin 5 protein PDB:1HD2

were determined as LEU149, THR147, GLN133, ARG127, SER118, SER115, ASP113,

LYS93, GLU91, ALA90, GLN68, GLY66, ALA64, LYS63, CYS47, GLY46, PRO45,

THR44, PRO40 and according to these active residues the grid boxes were determined as

centre_x=10.129, centre_y=38.905, centre_z=22.741, size_x=80, size_y=82, size_z=112,

spacing=0.375. Similarly, the obtained docking results for (3)-PDB: 1HD2 were presented in

Table 11 and Figure 8. According to the affinity binding energies, the best binding was

determined with -6.2 (kcal/mol) energy and 1 active and 1 non-active hydrogen bondings.

The active hydrogen bonding was observed between GLN68 and H3 atom with 2.07 Å bond

distance, and non-active hydrogen bonding was observed between GLN92 and N7 atom with

3

.11 Å bond distance. Additionally, van der Waals, carbon-hydrogen bond, π-cation and π-

anion, interactions were also determined from the Figure 8.

Finally, the inhibition constants for (3) and PDB:3A4A and (3)-PDB: 1HD2 interactions

were calculated as 1.61904 μM and 28.5343 μM, respectively by using Ki=exp(ΔG/RT)

-3

equation, where, ΔG, R and T are the docking binding energy, gas constant (1.9872036×10

kcal/mol) and room temperature (298.15 K), respectively. From the molecular docking results

of in silico antidiabetic and antioxidant, it is concluded that (E)-N'-(4-

2

0

(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3) molecule can be

designed as potential antidiabetic agent .

4

. Conclusions

Synthesis, crystal structure data, antioxidant and antidiabetic activities of (E)-N'-(4-

dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide (3) are reported. The

(

compound was experimentally characterized by using FT-IR, NMR, ESI-MS and single

crystal X-ray diffraction (XRD). The molecular structures of free base and cationic species of

(3) were determined theoretically in gas phase and aqueous solution by using B3LYP/6-31G*

and B3LYP/6-311++G** methods. High solvation energy values are observed for both

species of (3) in aqueous solution while the NBO and AIM studies support the higher stability

of the cationic species in solution. The high energy values ∆E_σ_→_σ_*and ∆E_σ_→_π_*interactions

that involve transitions of both rings and of N33 atom with both CH groups, due to the

3

planarity of both CH groups linked to N atom, could support the high reactivities of its free

3

base and cationic species, as compared with naloxone, cocaine and scopolamine. The

harmonic force fields for both species were reported together with the complete vibrational

assignments of 105 and 108 vibration modes expected respectively for free base and cationic

species of (3). The harmonic force constants of free base and cationic species of (3) were also

reported together with the Raman spectra for the two species and their electronic spectra in

aqueous solution. In addition, the frontier orbitals studies have evidenced high reactivity of

both species of (3) which justify its significant antioxidant activity, whilst showed the best

activity against α-glucosidase higher than that acarbose, and showed activity against α-

amylase similar to Quercetin. Finally, The molecular docking studies of the title compound

revealed that it may exhibit anti-diabetic activity via inhibition of α-glucosidase PDB:3A4A

enzyme.

Conflicts of interest

All authors declare that there are no conflicts of interest.

Acknowledgments

This work was supported by the WBI-COP22 Morocco project and UM5R as well as grants

from CIUNT Project Nº 26/D608 (Consejo de Investigaciones, Universidad Nacional de

Tucumán). The authors would like to thank Prof. Tom Sundius for his permission to use

MOLVIB.

2

1

Supporting Information Available: Tables S1-S5 and Figures S1-S12.

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Article Doi

Synthesis, X-ray structure, vibrational spectroscopy, DFT, biological evaluation and molecular docking studies of (E)-N’-(4-(dimethylamino)benzylidene)-5-methyl-1H-pyrazole-3-carbohydrazide

DOI: 10.1016/j.molstruc.2020.128541

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