FT-IR, FT-Raman, UV, NMR spectra and molecular structure investigation of (E)-2-(3-chloropyrazin-2-yl)-1-(3-ethyl-2, 6-diphenyl piperidin-4-ylidene) hydrazine: A combined experimental and theoretical study

Article abstract of DOI:10.1016/j.molstruc.2015.07.024

This work presents the characterization of (E)-2-(3-chloropyrazin-2-yl)-1-(3-ethyl-2, 6-diphenyl piperidin-4-ylidene) hydrazine (HDE) by quantum chemical calculations and spectral techniques. The structure was investigated by FT-IR, FT-Raman, UV-vis and NMR techniques. The geometrical parameters and energies have been obtained from Density functional theory (DFT) B3LYP (6-31G (d, p)) basis set calculations. The geometry of the molecule was fully optimized, vibrational spectra were calculated and fundamental vibrations were assigned on the basis of total energy distribution (TED) of the vibrational modes, calculated with scaled quantum mechanics (SQM) method. ¹H and ¹³C NMR chemical shifts of the molecule were calculated using Gauge-independent atomic orbital method (GIAO). The electronic properties such as excitation energies, wavelength, HOMO, LUMO energies performed by Time dependent density functional theory (TD-DFT) results complements with the experimental findings. NBO analysis has been performed for analyzing charge delocalization throughout the molecule. The calculation results were applied to simulate spectra of the title compound, which show excellent agreement with observed spectra. To provide information about the interactions between human cytochrome protein and the novel compound theoretically, docking studies were carried out using Schr?dinger software.

Full text of DOI:10.1016/j.molstruc.2015.07.024

Accepted Manuscript
FT-IR, FT-Raman, UV, NMR Spectra and Molecular Structure investigation of (E)-1-
(3-choloropyrazin-2-yl)-2-(3-ethyl-2, 6-diphenyl piperidin-4-ylidene) hydrazine: A
combined Experimental and Theoretical study
A. Therasa Alphonsa, C. Loganathan, S. Athavan, Dr. S. Kabilan
PII:
S0022-2860(15)30135-6
10.1016/j.molstruc.2015.07.024
MOLSTR 21679
DOI:
Reference:
To appear in: Journal of Molecular Structure
Received Date: 6 April 2015
Revised Date: 16 June 2015
Accepted Date: 13 July 2015
Please cite this article as: A. Therasa Alphonsa, C. Loganathan, S. Athavan, S. Kabilan, FT-IR, FT-
Raman, UV, NMR Spectra and Molecular Structure investigation of (E)-1-(3-choloropyrazin-2-yl)-2-(3-
ethyl-2, 6-diphenyl piperidin-4-ylidene) hydrazine: A combined Experimental and Theoretical study,
Journal of Molecular Structure (2015), doi: 10.1016/j.molstruc.2015.07.024.
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FT-IR, FT-Raman, UV, NMR Spectra and Molecular Structure investigation of (E)-1-
(3-choloropyrazin-2-yl)-2-(3-ethyl-2, 6-diphenyl piperidin-4-ylidene) hydrazine: A
combined Experimental and Theoretical study
A.Therasa Alphonsaa, C. Loganathana, S.Athavana, S. Kabilana*
aDepartment of Chemistry, Annamalai University, Annamalainagar - 608002, Tamilnadu, India.
Abstract
This work presents the characterization of (E)-1-(3-choloropyrazin-2-yl)-2-(3-ethyl-2,
6-diphenyl piperidin-4-ylidene) hydrazine (HDE) by quantum chemical calculations and
spectral techniques. The structure was investigated by FT-IR, FT-Raman, UV-Vis and NMR
techniques. The geometrical parameters and energies have been obtained from Density
functional theory (DFT) B3LYP (6-31G (d, p)) basis set calculations. The geometry of the
molecule was fully optimized, vibrational spectra were calculated and fundamental vibrations
were assigned on the basis of total energy distribution (TED) of the vibrational modes,
1
calculated with scaled quantum mechanics (SQM) method. H and 13C NMR chemical shifts
of the molecule were calculated using Gauge-independent atomic orbital method (GIAO).
The electronic properties such as excitation energies, wavelength, HOMO, LUMO energies
performed by Time dependent density functional theory (TD-DFT) results complements with
the experimental findings. NBO analysis has been performed for analyzing charge
delocalization throughout the molecule. The calculation results were applied to simulate
spectra of the title compound, which show excellent agreement with observed spectra. To
provide information about the interactions between human cytochrome protein and the novel
compound theoretically, docking studies were carried out using Schrödinger software.
*Corresponding author: Dr. S. Kabilan
Email: profdrskabilanaa@gmail.com, Tel: 9443924629

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Keyword: TED, Docking, HOMO, LUMO, NBO and Hyperpolarizability.
1. Introduction
In recent years there has been a growing interest pertaining to the synthesis of
heterocyclic systems owing to their varied reactivity, biological and physiological activity
[1-5]. Nitrogen containing heterocycles always signified a subject of great interest due to
their ubiquity in nature and massive presence as part of the skeletal backbone of many
therapeutic agents. Piperidones have an important role in the field of medicinal chemistry due
to their biological activities, including anticancer and cytotoxic properties [6]. Piperidones
are also reported to possess analgesic, anti-inflammatory, central nervous system (CNS)
stimulating, local anesthetic, anticancer and antimicrobial activity [7]. Particularly due to
their varied biological properties such as antiviral, antifungal [8] and anti tumor analgesic [9]
activities, 4-piperidones gaining a considerable importance. Also it plays an important role
which embraces the organometallic field in different aspects [10-13].
Prostakov and Gaivoronskaya have reviewed about the significance of
piperidin-4-one as intermediate in the synthesis of compounds of physiologically active [14].
The extensive studies undertaken in the past on 4-piperidones have their relation to the
synthesis of drug [15]. Noller & Baliah prepared substituted piperidones by heating a
mixture of ketone, aldehyde and ammonium acetate in acetic acid in the ratio of 1:2:1 [16].
Introduction of various substituents on the piperidone heterocycle improved the biological
efficiency. When 2- and/or 6-positions of 4-piperidones are occupied by aryl groups, their
biological activities are improved. The presence of alkyl substitution in 2 or 3 position is also
attributed to biological activities [17-18]. A series of 2, 6–diphenyl–4-piperidone are

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synthesized by different dialkyl ketone, aromatic aldehyde and ammonium acetate by
Mannich condensation reaction [16].
Literature survey reveals that to the best of our knowledge, inadequate experimental
and theoretical studies have been made on the vibrational, absorption spectra of piperidone
and its derivatives. Most recently Paulo R. Oliveto et al investigated the spectroscopic and
theoretical studies of 3-(4’ substituted phenyl sulfanyl)-1-methyl-2-piperidone [19].
Subashchandrabose et al analyzed the spectroscopic FT-IR, FT-Raman and energy
distribution of 3-pentyl-2, 6-diphenyl piperdin-4-one [20]. In addition to these studies,
number researches are also done about the antimicrobial activities of piperidone derivatives
[21-23]. To the best of our knowledge, neither experimental nor theoretical studies of the title
compound have not been available until now.
In our present work, the molecular structural parameters like bond length, bond
angle, dihedral angle, electronic, translational , rotational and vibrational frequencies, NBO
have been calculated for the compound, (E)-1-(3-choloropyrazin-2-yl)-2-(3-ethyl-2,6-
diphenyl piperidin-4-ylidene) hydrazine (HDE) (Fig.1). Optimized structural parameters are
obtained using the Density Functional Theory (DFT), performing B3LYP/ (6-31G, d, p) level
of calculations. The calculated molecular structural parameters and vibrational frequencies
have been analyzed and compared with the experimental values. HOMO, LUMO analysis
with UV spectroscopic studies is used to elucidate the information about the charge transfer
within the molecule.
2. Experimental
2.1. Synthesis
2, 3-dichloro pyrazine, ammonium acetate, Benzaldehyde, 2-pentanone are purchased
from Sigma-Aldrich chemical company with a stated purity and are used as such without
further purification. The title compound is synthesized according to scheme 1.

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2.2. Instruments
The FT-IR spectrum of the molecule is recorded in the region of 400- 4000 cm -1 on
Thermo Scientific Nicolet iS5 (iDl transmission) using KBr pellet. FT-Raman spectrum is
recorded using 1064 nm line of Nd: YAG laser as excitation source with wave length in the
region 3500-50 cm -1 on a Bruker RFS 100/S FT-Raman spectrophotometer. The detector is a
liquefied nitrogen cooled Ge detector. The Ultraviolet absorption spectra of HDE dissolved in
DMSO are examined in the range of 200 -800 nm using a Hitachi model U-2001
spectrophotometer. NMR spectrum is performed in Bruker DPX 400 MHz at 300K. The
compound is dissolved in DMSO-d6. Chemical shifts are reported in ppm relative to
1
tetramethylsilane (TMS) for protons and carbons. H and 13C NMR spectra are obtained at a
base frequency of 400 MHz and 100 MHz respectively.
2.3. Computational Techniques
All computational studies are carried out at Density Functional theory (DFT) level on
a Dell workstation equipped with V3 quardcore, Xeon processor E3–1240, 3.40 GHz
personal computer with 8 GB total RAM using Gaussian 09W program package [24]. DFT
calculations are less time consuming and include a significant part of the electron correlation
leading to good accuracy. The calculations are carried out with the Becke’s three parameter
exchange functional with the LYP correction (B3LYP) and the basis set, 6-31 G (d, p) are
used in appropriate calculations. The computational work has begun with the conformational
analysis of the compound. The selected geometric parameters of the optimized structure of
HDE are given in Table 1S.

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2.4. Docking Studies
The Docking studies reported in this work were performed on a Dell workstation
equipped with V2 quardcore, Xeon processor E3–1225, 3.20 GHz personal computer with 8
GB total RAM and with Schrödinger software suite, LLC, New York, 2012.
Molecular Docking study was performed by P450 2C19 proteins (human cytochrome
P450 monooxygenase) to find the binding and interaction properties of synthesized
compounds. The 3D crystallographic structure of human cytochrome protein complex [25]
(PDB ID: 4GQS) was downloaded from Protein Data Bank (www.rcsb.com). The protein
complex was prepared after pre–processing by Protein Preparation Wizard [26] in Maestro
9.3.5 version of Schrödinger software. The minimization of the protein complex was
continued using OPLS–2005 (Optimized Potential for Liquid Simulations) force field [27]
until the root mean square deviation (RMSD) reached the value of 0.3 Å.
The 2D structure of compound HDE was imported from the project table and the
structure was minimized and geometrically refined using Ligprep [28] module. Conformers
were generated using OPLS–2005 force field and torsional search method with distance
dependent dielectric solvation treatment. The extra precision (XP) mode of docking was used
to find the interactions between the active site of human cytochrome protein and the
synthesized molecule using Glide [29] application of the Schrödinger software suite.
3. Results and Discussion
3.1. Vibrational analysis
The synthesized HDE is subjected to IR and Raman spectral analysis and the
observed IR and Raman spectrum of HDE is given in Figure 1S. The theoretical vibrational
spectra of the compound HDE are analyzed using DFT/B3LYP 6-31G (d, p) Table 2S. The
non-negative vibrational frequency obtained from DFT calculations confirms that the
optimized geometry of compound HDE is located at the minima on the potential energy state.

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The theoretical vibrational frequencies are found to be in good agreement with the observed
vibrational frequencies. The theoretical vibrational frequencies obtained for compound HDE
is interpreted by means of Total energy distribution (TED %) calculations using SQM
method. The normal modes assignment of the theoretical IR frequencies is visualized and
substantiated with the help of the Gaussview 5.0 visualization program. The synthesized
HDE consists of 53 atoms and hence has 162 normal modes of vibrations which includes 115
stretching, 116 bending and 93 torsional modes of vibration. Scaling factor values[30]of 0.96
used for C-H,C-X stretching, bending,wagging,ring puckering and torsion vibrational [31]
frequencies, respectively. The molecule HDE belongs to C1 symmetry. The significant
normal modes with TED (10%) are given in Table 2S.
N-H vibrations
The experimental wave number of 3414cm-1 in the IR and 3412cm-1 (mode no: 153)
in the Raman spectrum of HDE is attributed for N-H stretching. The N-H stretching vibration
modes are visualized in the mode nos.153 and 152 with the scaled frequencies 3433cm-1 to
3357cm-1 respectively. Both the obtained theoretical and vibrational frequencies are pure
vibrations with TED % of 100 %.
Aromatic stretching group vibrations
In general, the aromatic C-H stretching frequencies appear in the range of 3100-3000
cm-1 [32] and the ring C-C stretching vibrations occur in the region 1650-1400 cm-1 [33]. In
the present work, the bands observed at 3102 cm-1 and 3103 cm-1 in both infrared and Raman
spectra for HDE have been assigned to the aromatic C-H stretching vibrations. In theoretical
calculation, mode nos.140-151 corresponds to the theoretical aromatic C-H stretching
vibrations.

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Methyl, methylene group vibrations
The methyl groups present in HDE undergoes vibrations like symmetric stretching,
asymmetric stretching, symmetric and asymmetric bending modes. The symmetric and
asymmetric stretching mode of the methyl group appears in the range of 2860-2935 cm-1 and
2925-2985 cm-1 [34]. The stretching in the methyl groups is well explained by the mode nos.
112 and 110 in the theoretical vibrational study of the compound HDE. The theoretical wave
numbers go well with the experimental both IR and Raman wave numbers. The bending
vibrations of the methyl group are found to appear in the region of 1465-1440 cm-1 and 1390-
1370 cm-1 [35]. The bending vibrations of the methyl groups are observed at 1411, 1412 cm-1
in the IR and Raman spectrum. The theoretical investigation well explains these bending
vibrations as asymmetric bending vibration of the methyl group with the help of the mode
no.112 in the scaled frequency of 1429 cm-1.
The asymmetric and symmetric C-H stretching in methylene group normally appears
in the region of 3100-2900 cm-1 [36]. In the present study, the observations of asymmetric
and symmetric stretching of the C-H in methylene group (C7) in compound HDE observed at
2970, 2932 and 2934 cm-1 both in IR and Raman spectrum are complemented by the
theoretical investigation which well explains the asymmetric and symmetric stretching in the
methylene group in the respective scaled frequencies 3011, 2990, 2949, 2928, and 2908 cm-1
(mode nos.138, 136-134, 132). The bending vibration of the methylene group is explained
theoretically in the frequency range 1457, 1452 cm-1(mode no.120, 119). The bending
vibration of the methylene group is observed at 1459 cm-1.
C=N Vibrations
The C=N vibration normally appears in the region 1471 – 1689 cm-1 [37]. The strong
band observed at 1719 cm-1 in the experimental IR spectra (1716 cm-1 in FT-Raman) of HDE

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is assigned to C=N vibration. The mode no 129 with the scaled frequency 1647 cm-1
visualizes the C=N stretching.
N-N Vibrations
The experimental wave number of 949 cm-1 in the IR spectra whereas in Raman at
951 cm-1 of HDE is attributed for N-N stretching. The N-N stretching vibration mode is
visualized in the mode no. 69 with the scaled frequency 948 cm-1.
3.2.1H and 13C NMR spectral analysis of HDE.
1
1
The H NMR spectrum of HDE has been given in Figure 2S. In the H NMR
spectrum of the compound HDE Table 1 in which H (7) signal appears as triplet at <delta>
1
0.721 ppm. The multiplet at <delta> 1.290 ppm in the H NMR indicates the presence of H
(6). The H (3) protons are exposed in the region around <delta> 4.741 to 4.968 ppm. The
1
doublet at <delta> 2.69 ppm in the H NMR spectrum of the compound HDE shows the
presence of H (8) protons. The 1H NMR spectrum of HDE reveals a multiplet at <delta> 3.73
ppm for H (2) attached to a phenyl ring with the integral value corresponding to four protons.
The 13C NMR spectrum of the compound HDE has been displayed in Figure 3S. The
13
carbon chemical shifts are presented in Table 1. The C NMR spectrum of the synthesized
compound HDE has been recorded in DMSO. The low field signal observed around <delta>
10 ppm in 13C NMR spectra of HDE is due to the carbon C (5). The C (8) carbon is identified
13
at <delta> 63.0 ppm in C NMR of HDE. The carbon in the azine linkage C (23) and C (4)
are shielded by the electron cloud and hence exposed by the up-field signals in the range
13
<delta> 158.1 ppm to <delta> 160.6 ppm. The signal at <delta> 59.2 ppm in the C NMR
spectrum of HDE is due to the carbon C (2).
The 13C NMR peaks of the C5 carbon atom appear to be the same in experimental and
theoretical values for which it is presented in the Table 1.

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NMR spectroscopy is the key to reveal the conformational analysis of organic
molecules. Good quality geometries must be taken into consideration for the quantum
calculation of the absolute isotropic magnetic shielding tensors to yield more reliable results.
There are many reports on NMR isotropic magnetic shielding tensor calculations employing
1
the GIAO method associated with the Density functional theory (DFT) [38]. The GIAO H
13
NMR and C NMR chemical shift calculations of the stable conformer is made in DMSO
[scrf= (solvent=DMSO)] by using B3LYP / 6-31G (d, p) basis set. The isotropic values in
the calculations are subtracted from a scaling factor of 182.4656 and 31.882 to obtain the
13
1
chemical shifts for C and H NMR respectively. The obtained chemical shift values were
compared with the experimentally observed values. The observed and the calculated carbon
1
chemical shift values are found to be in good agreement whereas the theoretical H NMR
chemical shifts are found to be lesser than the experimental values. The calculated values are
given in the Table 1 along with the observed values. The relationship between the
experimental chemical shift and computed chemical shift values are given in Figure 2. The
relation between the observed and computed values predicts that the conformation deducted
theoretically should be the favorable conformation of the synthesized HDE.
3.3. Natural Bond orbital analysis
The natural bond orbital analysis provides an efficient method for studying intra-and
intermolecular bonding and interaction among bonds, and also provides a convenient basis
for investigating charge transfer or conjugative interaction in molecular systems, it could
enhance the analysis of the delocalization of charge in the system.
The donor bonding orbitals, the acceptor antibonding orbitals, the donor lone pair
atoms are given in Table 2 along with the E (2) values which estimates the interaction
between the donor (filled) and acceptor (vacant) orbitals. The E (2) energy is the lowering
energy that occurs during the hyperconjugative electron transfer process and hence E (2) can

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be referred to as stabilization energy. Larger the E (2) values, greater is the stability of the
molecule. In the NBO analysis of the compound HDE, the E (2) values are greater for the
delocalization of the electrons between the bonds present in the phenyl ring. The lone pair of
electrons present in the Nitrogen atom N22 is delocalized to C3-N28 antibonding orbitals
with high delocalization energy which may be attributed to the presence of chloro substituent.
The delocalization of electrons from the lone pair of the Nitrogen atom N21 to C4 – C5
antibonding orbitals is not so effective because of the absence of electron withdrawing group
in the rings nearer to N21.
3.4. Charge Analysis
The charge quantifies the electronic structure changes under atomic displacement. It
is related directly to the chemical bonds present in the molecule, it influences dipole moment,
polarizability, electronic structure and more properties of molecular systems. Natural charge
analysis tends to predict larger charges than several other population analysis like Mulliken
charges. Natural charges considers the population in the orbitals that will overlap to form a
bond and those that are too near the core of an atom to be involved in bonding. Mulliken
charges are the partial charges confined to the atoms. The Mulliken and natural charge
distribution of the molecule calculated on B3LYP level with 6-31G (d, p) basis set are shown
in Table 3. The Mulliken charge distribution in the compound HDE is found to be greater
than the corresponding natural charges (Figure 3). The carbon atoms 4C, 23C and 24C
possess positive charges in Mulliken charge and natural charge analysis. The carbon atoms
2C, 15C, 27C, 26C and 9C possess positive Mulliken charge and negative natural charge. All
the hetero atoms in the compounds possess negative Mulliken and negative charges.
3.5. Molecular electrostatic potential (MEP) surfaces
The reactive behavior of the molecule is visualized with the help of three dimensional
MEP surface. MEP surface describes the charge distribution in the molecule and helps in

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predicting the sites for nucleophilic and electrophilic attack in the molecule. The MEP
surface has been plotted for the molecules HDE in Figure 4. Region of negative charge is
pictured out in red colour and it is found around the electronegative Cl and N in the azine
linkage in the molecule HDE. The red colour region is susceptible to electrophilic attack.
The blue colour region represents strong positive region and is prone to nucleophilic attack.
The green colour region corresponds to a potential half way between the two extremes red
and blue region.
3.6. HOMO-LUMO Pictures of HDE
HOMO-LUMO pictures of molecule HDE is presented in Figure 5. The pz Orbitals of
C15,C16,C17,C18,C19,C20,C7,C9,C10,C11,C12,C13 and C14 carbon atoms do not involve
in the formation of the HOMO orbitals in the molecule HDE. The pz orbitals of
C6,C7,N1,C5,C8 C9,C10,C11,C12,C13,C14,C20,C15,C16,C17,C18,C19 and C20 carbon
atoms alone do not involve in the formation of the LUMO orbital. The band gap between the
HOMO and LUMO orbital energy is found to be 4.485 eV.
3.7. Polarizability calculations of HDE
The calculated values of the dipole moment (µ), the polarizability (α0) and first
hyperpolarizability (βtot) by finite field approach are given in Table 4 alone with the
corresponding components. The field independent and field dependent dipole moment µ
values of are calculated to be 2.71 and 0.78 respectively. The highest value of dipole
moment is observed for the component µy for both the field independent and dependent
conditions. The compound HDE is found to be a polar molecule having non – zero dipole
moment components.
The calculated polarizabilities αij have non – zero and are dominated by the diagonal
components. The βtot value of HDE is found to 3.212 x 10-30 esu. Delocalization of charges
in particular directions is indicated by large values of those particular components of

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polarizability and hyperpolarizabilty, first order hyperpolarizability is dominated by βxzz and
βzzz values. This shows that the delocalization of the charges in the presence of an external
field is endorsed in those directions. The first hyperpolarizability of HDE (<beta>tot) is 9
times greater that of urea (0.3728 x 10-30 esu), hence this molecule can have a considerable
NLO activity and can find its application in the material science field too.
3.8. UV spectrum and electronic properties
The UV absorption spectrum of the title compound in DMSO solvent was recorded
within the 200-400 nm range and representative spectrum is shown in Figure 4S.
It is
observed that the recorded absorption bands centered at 278 nm DMSO. In order to support
our experimental observations TD-DFT calculations on electronic absorption spectra were
performed based on the B3LYP/6-31G (d, p) [39] level optimized structure in gas phase. The
TD-DFT method predicted the maximum absorption peak at 272 nm with strong oscillator
strength, while this peak was recorded at 278 nm in DMSO. Due to Frank-Condon principle
this peak (λmax) corresponding to vertical excitation. Calculations of the molecular orbital
geometry show that the visible absorption maxima of this molecule correspond to the electron
transition between frontier orbitals such as translation from HOMO to LUMO. The HOMO
energy characterizes the ability of electron giving, the LUMO characterizes the ability of
electron accepting, and the gap between HOMO and LUMO characterizes the molecular
chemical stability [40]. The energy gap between the HOMOs and LUMOs is a critical
parameter in determining molecular electrical transport properties because it is a measure of
electron conductivity. The absorption maximum values are observed 278 nm those show
agreement in theoretical value. The deviation between experiment and theory may be
resulted from solvent effects. Solvent makes the chemical environment of molecule in the
simulation become very complex. The maximum absorption wavelength corresponds to the
electronic transition from the highest occupied molecular orbital HOMO to lowest

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unoccupied molecular orbital LUMO. All the observed transitions are π-π* ones. All
assignments agree well with the values available in the literature and experimental values.
3.9. Docking studies
Docking studies were carried out to evaluate the binding affinity and the interactions
between the synthesized compound and the human cytochrome protein (PDB ID: 4GQS).
The docking score of the different conformers of compound HDE varies from -8.4 to -3.6.
The docking results clearly indicate that the tested compound display two types of
interactions with three amino acids present in the active site of 4GQS. The 2D and 3D view
of interactions between the compound and 4GQS is shown in Figure 6. The aromatic rings
present in the 2nd and the 6th positions of the piperidyl ring were showed π−π stacking
interactions with PHE114 and PHE476 residues. The –NH group of the piperidyl ring shows
hydrogen bond interaction with GLY296 residue.
4. Conclusions
A novel Unsymmetrical azine (E)-1-(3-chloropyrazin-2-yl)-2-(3-ethyl-2,6-diphenyl
piperidin-4-ylidene) hydrazine (HDE) was synthesized and characterized by FT-IR, FT-
Raman, UV-Vis, 1H and 13C techniques. The theoretical IR vibrational frequencies are found
to be in good agreement with the observed and theoretical IR vibrational frequencies of HDE
are analyzed by means of total energy distribution (TED%) calculations using scaled
quantum mechanics (SQM) method. The GIAO 1H and 13C NMR chemical shift calculations
of the stable conformer is made in DMSO by using B3LYP/6-31G (d,p) basis set. The
compound HDE is subjected to NBO analysis, the E(2) values are greater for the
delocalization of the electrons between the bonds present in the phenyl ring. In the Nitrogen
atom N22 antibonding orbitals with high delocalization energy which may be due to the
presence of chloro substituent. The first hyper polarizability of HDE is 9 times greater than
of urea (0.3728 x 10-30 esu), hence this molecule can have considerable NLO activity and can

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find its application in material science field too. The docking results clearly indicate that the
tested compound display two types of interactions with three amino acids present in the
active site of 4GQS.
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FIGURE and SCHEME CAPTIONS
Scheme 1.
Fig. 1. Numbering of HDE used in this study
Fig. 2. Comparison of observed and theoretical chemical shifts values of 1H and 13C spectrum
of HDE
Fig.3. Comparison of Mulliken charges and NBO charges of HDE
Fig. 4. MEP plot of HDE
Fig. 5. HOMO-LUMO pictures of HDE
Fig. 6. 2D and 3D interaction between HDE and 4GQS

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Scheme 1

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Fig. 1. Numbering of HDE used in this study

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Fig. 2. Comparison of Experimental and theoretical chemical shifts values of 1H and 13C
spectrum of HDE

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Mulliken Charges
NBO Charges
0.6
0.4
0.2
0.0
-0.2
-0.4
-0.6
Fig. 3. Comparison of Mulliken charges and NBO charges of HDE

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Fig. 4. MEP plot of HDE

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LUMO
HOMO
Fig. 5. HOMO-LUMO pictures of HDE

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2D view
3D view
Fig. 6. 2D and 3D interaction between HDE and 4GQS

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TABLE CAPTIONS
Table 1. Experimental and computational 1H and 13C NMR spectral analysis of HDE in
DMSO solution by DFT method
Table 2. Second order perturbation theory analysis of fock matrix in NBO basis for HDE
Table 3. Mulliken charges and NBO charges of HDE
Table 4. Dipole moment µ (D), mean polarizability α0 (x 10-24 esu) and first
hyperpolarizability βtot (x10-33 esu) of HDE by DFT method

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Table 1. Experimental and computational 1H and 13C NMR spectral analysis of HDE in
DMSO solution by DFT method
1H chemical shift values
Atom
H7
Expt.value Theo.value
0.721(t)
1.290(m)
2.695(d)
3.737(m)
4.968(t)
4.741(t)
0.501
H6
1.678
H8
3.633
H2
4.295
H3a
H3b
2.705
3.081
Aromatic protons 7.439(s)
7.342-7.698
8.102
H27
7.894(s)
7.821(s)
10.046(s)
11.056(s)
H26
7.839
H1--N1
H22--N22
0.633
7.105
13C chemical shift values
Atom
C7
Expt.value Theo.value
10.61
17.44
44.96
51.68
59.25
63.03
15.20
C6
30.15
C3
38.53
C5
51.64
C2
61.76
C8
67.32
Aromatic carbons 128.54
123.54
128.91
138.98
139.41
145.64
187.31
Pyrazine
Pyrazine
Pyrazine
Pyrazine
C4
129.28
134.51
135.44
158.13
160.61

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Table 2. Second order perturbation theory analysis of fock matrix in NBO basis for HDE
E2
Donor
(i)
Acceptor
(j)
ED
(j,e)
E(j)-E(i)
a.u
Type
ED (i,e)
F(i,j) a.u
(Kcal/Mol)
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
π-π*
σ-σ*
σ-π*
σ-σ*
σ-σ*
C15-C16 1.65250 C20-C19
C15-C16 1.65250 C17-C18
C20-C19 1.66506 C15-C16
C20-C19 1.66506 C17-C18
C17-C18 1.66362 C15-C16
C17-C18 1.66362 C20-C19
C23-N28 1.70876 C24-N25
C23-N28 1.70876 C27-C26
C24-N25 1.79094 C23-N28
C24-N25 1.79094 C27-C26
C27-C26 1.66809 C23-N28
C27-C26 1.66809 C24-N25
0.32736 20.15
0.33339 20.75
0.34552 20.55
0.33339 20.35
0.34552 20.02
0.32736 20.24
0.38584 14.26
0.28739 25.49
0.45173 15.82
0.28739 16.43
0.45173 15.04
0.38584 22.04
0.32914 19.57
0.33364 21.30
0.34568 20.85
0.33364 20.03
0.34568 19.64
0.32914 20.58
0.04346 12.09
0.45173 35.61
0.05890 10.75
0.05890 11.74
0.38584 13.81
0.28
0.28
0.28
0.28
0.28
0.28
0.30
0.33
0.32
0.35
0.26
0.26
0.28
0.28
0.29
0.28
0.29
0.28
0.81
0.30
0.84
0.84
0.31
0.067
0.068
0.068
0.068
0.068
0.068
0.060
0.082
0.068
0.069
0.058
0.068
0.066
0.069
0.069
0.067
0.067
0.068
0.089
0.098
0.086
0.090
0.063
C9-C14
C9-C14
1.64942 C10-C11
1.64942 C13-C12
C10-C11 1.66701 C9-C14
C10-C11 1.66701 C13-C12
C13-C12 1.66235 C9-C14
C13-C12 1.66235 C10-C11
N21
N22
N28
N25
1.92884 C4-C5
1.76998 C23-N28
1.90466 C23-C24
1.89411 C23-C24
1.92507 C24-N25
LP(3)-π* Cl29
a - energy of hyper conjugative interaction (stabilization energy), b - energy difference donor and acceptor i and j
NBO orbitals, c - F (i-j) is the fock matrix element between i and j NBO orbitals

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Table 3. Mulliken charges and NBO charges of HDE
Atom Mulliken charges NBO charges
N1
-0.49
0.01
-0.68
-0.06
-0.52
0.33
C2
C3
-0.25
0.33
C4
C5
-0.08
-0.18
-0.32
-0.02
0.14
-0.32
-0.45
-0.68
-0.07
-0.03
-0.22
-0.23
-0.23
-0.22
-0.23
-0.04
-0.23
-0.23
-0.23
-0.23
-0.21
-0.30
-0.48
0.34
C6
C7
C8
C9
C10
C11
C12
C13
C14
C15
C16
C17
C18
C19
C20
N21
N22
C23
C24
N25
C26
C27
N28
Cl29
-0.12
-0.08
-0.08
-0.09
-0.12
0.09
-0.12
-0.09
-0.08
-0.08
-0.10
-0.35
-0.45
0.54
0.05
0.14
-0.39
0.05
-0.40
-0.06
-0.01
-0.49
-0.01
0.08
-0.48
0.005