Synthesis, spectra, electronic structure, molecular docking and cytotoxicity investigation on 2-(piperidin-1-ylmethyl)-isoindoline-1,3-dione – A Mannich base system

Article abstract of DOI:10.1016/j.molstruc.2020.129151

A Mannich base molecule of 2-(piperidin-1-ylmethyl)isoindoline-1,3-dione (PPF) has been synthesized using Mannich base condensation reaction. The molecular geometry of PPF molecule was optimized by B3LYP/6-311G(d,p) basis set. The vibrational frequencies have been assigned with the aid of normal coordinate analysis. Total energy distributions (TED) were used to explain each vibrational wavenumber assignments of a molecule. The ¹H and ¹³C NMR spectroscopic analyses were predicted by using gauge independent atomic orbital (GIAO) method. The thermal stability was determined by applying the thermo gravimetric/differential scanning calorimetry (TG/DSC) studies. The HOMO-LUMO energies, population density of states (PDOS), overlap population density of states (OPDOS), reduced density gradient (RDG), Mulliken atomic charges and molecular electrostatic potential (MEP) surface were plotted to investigate the van der Waals, attraction, repulsion, non-covalent and covalent interactions of the compound. The natural bond orbital (NBO) charge analysis was used in explaining the intermolecular charge transfer and resonance stabilization energy of molecule. Besides, the molecular docking studies have been performed. The cytotoxic activity of the PPF molecule has been resolved.

Full text of DOI:10.1016/j.molstruc.2020.129151

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Journal of Molecular Structure 1224 (2021) 129151
Contents lists available at ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstr
Synthesis, spectra, electronic structure, molecular docking and
cytotoxicity investigation on 2-(piperidin-1-ylmethyl)-isoindoline-
1,3-dione – A Mannich base system
Krishnan Sarojini Devia, Palaniappan Subramanib,∗, Namadevan Sundaraganesanc,
Mani Jeevad, Susairaj Jone Pradeepac, Balakrishnan Karthikeyane
a Department of Chemistry, Sri-La-Sri Kasivasi Sawaminatha Swamigal Arts College, Thiruppanathal, Thanjavur 612504, Tamil Nadu, India
b Chemistry Section, Faculty of Engineering and Technology, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India
c Physics Section, Faculty of Engineering and Technology, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India
d Department of Chemistry, National Institute of Technology, Tiruchirappalli 620 015, Tamil Nadu, India
e Department of Chemistry, Annamalai University, Annamalainagar 608 002, Tamil Nadu, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A Mannich base molecule of 2-(piperidin-1-ylmethyl)isoindoline-1,3-dione (PPF) has been synthesized us-
ing Mannich base condensation reaction. The molecular geometry of PPF molecule was optimized by
B3LYP/6-311G(d,p) basis set. The vibrational frequencies have been assigned with the aid of normal co-
ordinate analysis. Total energy distributions (TED) were used to explain each vibrational wavenumber
assignments of a molecule. The 1H and 13 C NMR spectroscopic analyses were predicted by using gauge
independent atomic orbital (GIAO) method. The thermal stability was determined by applying the thermo
gravimetric/differential scanning calorimetry (TG/DSC) studies. The HOMO-LUMO energies, population
density of states (PDOS), overlap population density of states (OPDOS), reduced density gradient (RDG),
Mulliken atomic charges and molecular electrostatic potential (MEP) surface were plotted to investigate
the van der Waals, attraction, repulsion, non-covalent and covalent interactions of the compound. The
natural bond orbital (NBO) charge analysis was used in explaining the intermolecular charge transfer and
resonance stabilization energy of molecule. Besides, the molecular docking studies have been performed.
The cytotoxic activity of the PPF molecule has been resolved.
Received 15 November 2019
Revised 23 August 2020
Accepted 26 August 2020
Available online 27 August 2020
Keywords:
Mannich base
NMR
DFT
NBO
Thermal
cytotoxicity analysis
© 2020 Elsevier B.V. All rights reserved.
1. Introduction
piperidine derivatives have been used in pre-clinical and medicinal
studies [7]. Piperidine derivatives were proven for their pharmaco-
Generally, the Mannich condensation reaction plays signifi-
cant role for the preparation of organic compounds in the field
of organic and pharmaceutical chemistry. The N-Mannich reac-
tion is a three components one-pot reaction of primary or sec-
ondary amines containing electron-deficient nitrogen functional
group such as imide, amide, sulfonamide, etc., with the formation
of carbon bridge. The N-Mannich base compounds act as a pro-
drug form to design amine-containing antibiotic, which has been
previously investigated by Koch and coworkers [1]. The piperidine
heterocyclic system has many biological applications such as anal-
gesic, anti-hypersensitive, central nervous system depressant an-
tiviral, bactericidal etc. [2–6]. Watson et al. confirmed that, several
logical activity as they form a very significant element of molec-
ular structures on essential drug such as minoxidil and raloxifene
[8]. They are acting as intermediates for the preparation of antisep-
tics, cross-linking agents, crystalline adducting agents, dyes, poly-
mer and pesticides. Phthalimide structure has an isoindole moi-
ety. The famous examples comprise of auxin (hormone for plant
growth), indigo (plant colourant), serotonin (neurotransmitter) and
tryptophan 23 (aromatic side chain of amino acid). Phthalimides
among the heterocyclic scaffolds have specific class of function
on biological activity and that have properties like antipsychotics,
anti-inflammatory agents, herbicides and insecticides. Phthalimide
derivatives are used to identify the new class of reverse transcrip-
tase inhibitors for non-nucleoside HIV-1 virus [9]. Most of the
phthalimide moieties exhibit QSAR derived properties like elec-
tronic, steric and hydrophobic that suits for HIV-1 reverse tran-
scriptase as an inhibitor [10]. Similarly, the present title Mannich
∗
Corresponding author.
E-mail address: saro87che@gmail.com (P. Subramani).
https://doi.org/10.1016/j.molstruc.2020.129151
0022-2860/© 2020 Elsevier B.V. All rights reserved.

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K.S. Devi, P. Subramani and N. Sundaraganesan et al. / Journal of Molecular Structure 1224 (2021) 129151
base molecular system 1-[(pyridin-2-yl-amino)methyl]pyrrolidine-
2,5-dione (SFAP) which is under study, has its own biological mer-
its particularly on antioxidant activity.
A significant amount of the people are affected by lung can-
cer due to exposure towards tobacco smokes for long period and
few people who neither have certainty nor smoked affected as a
passive smoker. In some cases, the lung cancer is caused by ge-
netic factors and exposure of toxic gases and air pollutants. The
cytotoxic drugs play major role in chemotherapy to cure partic-
ularly this type cancers [11]. Many chemists have been devel-
oped to resolve the cytotoxic activity of numerous isoindoline-1,3-
dione derived compounds [12–14]. For instance, Collin et al. had
confirmed N-pyridinyl(alkyl)phthalimides connected to N-phenyl-
4,5,6,7-tetrafluorophthalimides could act as an inhibitor for the
treatment of tumour necrosis factor (TNF) production [15].
The N-pyridinyl and N-quinolinyl substituted compounds such
as phthalimides and succinimides have been synthesized and these
compounds recognized the cytotoxicity inhibit the action of growth
of a number of cultured cell lines. The main objective of this study
is to access the cytotoxic activity of the title molecule and poten-
tial of chosen commercially available activity against lung cancer
drugs in correlated with originate using lung cancer cell line on
A549 as MTT assay. The new Mannich base molecule 2-(piperidin-
1-ylmethyl)isoindoline-1,3-dione (PPF) which was synthesized con-
sists of molecular formula such as C14 H16 N2O2. PPF has colourless
substance, which has a melting point in the range of 113–115 °C.
The PPF molecule is based on coarse aqueous suspension, with
slight basicity level by having 8.6 pH value. Literature survey re-
veals that, the results based on quantum chemical calculations and
spectral analysis on PPF molecule have not been reported so far.
Herein, we report the complete interpretations of the vibrational
spectra based on the observed and computed results, which are
satisfactory and endurable to each other. In the present study, we
have attempted to investigate the bonding nature and thermody-
namic properties of the PPF molecule theoretically.
2. Experimental and computational details
Fig. 1. Experimental (a) FT-IR and (b) FT-Raman spectra of PPF molecule.
The PPF has been synthesized by using Mannich condensation
reaction. The raw chemicals employed in the synthesis were pur-
chased from Alpha and Aldrich Chemical Company with spectro-
scopic grade of purification. Hence, there were no need for any fur-
ther purification on reactants such as phthalimide, piperidine and
formaldehyde along with solvent used all the way through various
stages of the synthesis.
2.2. Spectroscopy measurements
The Fourier transform infrared spectrum of the molecule was
recorded in the range from 4000 to 400 cm–1 using the Perkin
Elmer FT-IR spectrophotometer by KBr pellet method as shown in
Fig. 1a. Fourier transform Raman spectrum of the molecule was
analyzed in the region from 4000 to 50 cm–1 by Bruker RFS 27
spectrophotometer utilizing 1064 nm excitation wavelength from a
Nd:YAG laser in the range 5000–50 cm–1 as shown in Fig. 1b. The
proton and carbon NMR were recorded using Bruker 500 MHz Ad-
vance NMR spectrometer operating at 500 MHz as shown in Fig. 2.
These measurements were completed with the help of highly pro-
tic solvent like dimethyl sulfoxide (DMSO-d6) solution. The solu-
tion was established by means of 0.2 mg for 1H NMR and 30 mg
for 13C NMR of the sample and dissolved in 5 mL of DMSO-d6 so-
lution. The chemical shifts of the spectrum were produced in ppm
corresponds to tetramethyl silane (TMS). Mass spectrum was done
2.1. Synthesis of 2-(piperidin-1-ylmethyl)-isoindoline-1,3-dione
The PPF was synthesized by adding phthalimide, piperidine and
formaldehyde in an equimolar ratio (Scheme 1) as one-pot. A mix-
ture of phthalimide (7.35 g, 0.05 mol), piperidine (5.0 mL, 0.05
mol) and 40% formaldehyde solution (3.8 mL, 0.05 mol) in ethanol
were mixed and refluxed for 12 h. The reaction mixture was al-
lowed to cool at room temperature and the solid mass obtained
was filtered, washed with distilled water. Then it was dried and re-
crystallized from absolute ethanol to yield a white crystalline prod-
uct (yield 92%).
Scheme 1. Synthesis of 2-(piperidin-1-ylmethyl)isoindoline-1,3-dione (PPF).

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K.S. Devi, P. Subramani and N. Sundaraganesan et al. / Journal of Molecular Structure 1224 (2021) 129151
3
Fig. 2. (a) 1H and (b) 13 C NMR spectra of PPF molecule.
using high resolution Q-Tof Mass, mass spectrometer as detector
(Perkin Elmer), Turbo Massver 5.4.2 software for data analysis and
the GC parameters were found as split 10:1 injection; column:
Elite-5MS (30 m × 0.25 mm I.D, 250 μm df); injection port tem-
perature: 250 °C; carrier gas: helium; flow rate: 1 mL/min; oven
temperature: initially 60 °C for 2 min, ramped to 300 °C at 10
°C/min, 300 °C for 6 min. The MS conditions were found as fol-
lows: transfer line heater: 240 °C; ion source temperature: 240 °C;
electron impact ionization (EI) mode; ionization energy: 70 eV. The
analysis was performed in scan mode, from 10 to 600 amu at a
speed of 1.5 scans/s. The solvent delay was set to 2.0 min as shown
in Fig. 3. The TG/DSC study was analyzed using thermo gravimet-
ric instrument TGA7 (Perkin Elmer) Q500 Hi-Res TGA from TA in-
struments, DSC7 (Perkin Elmer) Q200 Hi-Res TGA from TA instru-
ments as shown in Fig. 4. The UV-Vis transition spectrum of the
sample was produced in DMSO solution in the spectral range from
600 to 200 nm with Lambda-35 Perkin Elmer spectrophotometer
as shown in Fig. S1 (Supplementary information).
2.3. Theoretical calculations
In our current study, the density functional theory (DFT) sim-
ulations [16] were performed with Gaussian 09 package [17] by
means of Becke’s three parameters hybrid exchange functional
[18] and Lee-Yang-Parr correlation functional (B3LYP) [19,20]. The
basis level of 6-311G (d, p) was done selectively, on account of ad-
vantage of doing faster simulations along with relative fine preci-
sion and molecular structure [21,22]. The vibrational assignments
were described and scaled using the appropriate factor [23,24].
Molecular electrostatic potential was performed using the similar
set of theory and designed with GaussView 5 [25]. The vibrational
frequencies computed were scaled and TED has been derived us-

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K.S. Devi, P. Subramani and N. Sundaraganesan et al. / Journal of Molecular Structure 1224 (2021) 129151
ν0 = 9398.5 cm–1, which corresponds to the wavelength of 1064
nm of a Nd:YAG laser), νi represents the frequency of normal mode
(cm–1), while the Si is the Raman scattering activity of the normal
mode Qi. The IiR is assumed in arbitrary unit. In the above said
equation, the h, k, C and T denote the Planck constant, Boltzmann
constant, speed of light and temperature in Kelvin, respectively.
2.5. Docking studies
The molecular docking study was investigated by using lung
cancer and diabetic proteins to find the active binding sites and
interaction properties of amino acids with synthesized molecules.
The 3D crystal structure of protein complex (PDB format) was
downloaded from Protein Data Bank (www.rcsb.com). The protein
complex and ligand were organized after pre-processing by protein
preparation wizard and ligand preparation wizard [37] in Maestro
9.3.5 version of Schrodinger software.
Fig. 3. Mass spectrum of PPF molecule.
2.6. In vitro anticancer studies
2.6.1. Cell line
A549 (Lung Cancer) cells were initially procured from National
Centre for Cell Sciences (NCCS), Pune, India and maintained by
Dulbecco’s modified Eagles medium, DMEM (Sigma Aldrich, USA).
The cell line was cultured in 25 cm2 tissue culture flask with
DMEM supplemented with 10% FBS, L-glutamine, sodium bicar-
bonate and antibiotic solution containing: Penicillin (100 U/mL),
Streptomycin (100 μg/mL), and Amphotericin B (2.5 μg/mL). Cul-
tured cell lines were kept at 37 °C in a humidified 5% CO2 incu-
bator. The viability of cells were evaluated by direct observation
of cells by inverted phase contrast microscope and followed by 3-
(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT)
assay method.
Fig. 4. TG/DSC curves of PPF molecule.
ing VEDA 4 program [26]. The 1H and 13C NMR isotropic chem-
ical shielding were provided with gauge-invariant atomic orbital
(GIAO) method [27,28] for the mentioned optimized parameters
of B3LYP/6-311G(d, p) basis set. The order of GIAO technique is
one of the excellent prevalent approaches for simulating the nu-
clear magnetic shielding tensors and useful for consistent studies
of magnetic characters. The NBO analysis was performed with NBO
3.1 version [29] as employed in the Gaussian 09 program. The TD-
DFT [30,31] used to find out the electronic transitions, absorbance
wavelengths and vertical excitation and oscillator strengths of PPF
molecule in solvent phase and isolated gaseous phase was carried
out. The TD-DFT approach was carried out to determine the fron-
tier molecular orbitals (HOMO and LUMO) energies of a molecule.
GaussSum 3.0 [32] was used to draw a diagram for density of state
(DOS) spectrum of the PPF molecule. The Multiwfn [33] and VMD
program [34] are used to plot RDG graph, respectively. The thermo-
dynamic characters of the PPF molecule such as enthalpy change
(ꢀH), entropy (S) and specific heat capacity (CV) were simulated
with different temperature ranges from 100 to 1000 K.
2.6.2. Cell treatment procedure
Two days old confluent monolayer of cells was trypsinized and
the cells were suspended in 10% growth medium. 100 μL cell sus-
pension (5 × 104 cells/well) was seeded in 96 well tissue culture
plates and incubated at 37 °C in a humidified 5% CO2 incubator.
2.6.3. Preparation of compound stock
About
1 mg of sample was weighed and dissolved in 1
mL DMEM using a cyclomixer. The sample solution was filtered
through 0.22 μm Millipore syringe filter to ensure the sterility.
2.6.4. Anticancer evaluation
After 24 h, the growth medium was removed, each freshly pre-
pared compounds in 5% DMEM were serially diluted five times
by two fold dilution (100, 50, 25, 12.5, 6.25 μg in 500 μL of 5%
DMEM) and each concentration of 100 μL were added in triplicates
to the respective wells and incubated at 37 °C in a humidified 5%
CO2 incubator. Non-treated control cells were also maintained.
2.4. Raman intensities
The predicted Raman intensities (IiR), investigated by Raman ac-
tivities (Si), were displayed using the Gaussian 09 program. Raman
intensities can be expressed by the mathematical equation as ex-
plained in Eq. 1 [35,36].
2.6.5. Anticancer assay by direct microscopic observation
Entire plate was observed after 24 h of treatment in an in-
verted phase contrast tissue culture microscope (Olympus CKX41
with Optika Pro5 CCD camera) and microscopic observations were
recorded as images. Any detectable changes in the morphology of
the cells, such as rounding or shrinking of cells, granulation and
vacuolization in the cytoplasm of the cells were considered as in-
dicators of cytotoxicity.
ꢀ
ꢁ
4
IiR = C v − −v v −−1Bi−−1Si
(1)
(
)
0
i
i
where Bi is the temperature factor which interprets the intensity
contribution of excited vibrational states, denoted by the Boltz-
mann appropriation.
2.6.6. Anticancer assay by MTT method
B = 1 − − exp − −hv c/KT
(2)
In the IiR equation, the ν0 is the frequency of laser excita-
tion line (in this work, we have utilized the excitation frequency
(
)
i
i
Fifteen mg of MTT was reconstituted in 3 mL PBS until com-
pletely dissolved and sterilized by filter sterilization. After 24 h of
incubation period, the sample content in wells were removed and