Antiradical Properties of trans-2-(4-substituted-styryl)-thiophene

Article abstract of DOI:10.1007/s10895-020-02629-5

2-substituted thiophene compounds with electron donating and electron withdrawing p-phenyl substitution were synthesized and studied their radical scavenging properties using DPPH assay and DFT method. It is shown that p-hydroxy and p-amino phenyl substit

Full text of DOI:10.1007/s10895-020-02629-5

Journal of Fluorescence
https://doi.org/10.1007/s10895-020-02629-5
ORIGINAL ARTICLE
Antiradical Properties of trans-2-(4-substituted-styryl)-thiophene
Anamika Gusain1 & Naresh Kumar1 & Jagdeep Kumar1 & Gunjan Pandey1 & Prasanta Kumar Hota1
Received: 25 May 2020 /Accepted: 5 October 2020
#
Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract
2-substituted thiophene compounds with electron donating and electron withdrawing p-phenyl substitution were synthesized and
studied their radical scavenging properties using DPPH assay and DFT method. It is shown that p-hydroxy and p-amino phenyl
substituted compound exhibit radical scavenging activity. From DFT and radical scavenging studies, a correlation between IC50
with the bond dissociation enthalpy, proton affinity, ground state dipole moment and optical band gap of compound is found.
Compounds 1–3 with electron withdrawing substituent (NO2, CN, Cl) do not show any radical scavenging properties, whereas
compounds 6–7 with electron donating substituent (OH, NH2) show antiradical properties. Further, the antiradical activity is
reduced drastically by replacing the -OH and -NH2 with methoxy and -N-alkylating group respectively in 6 and 7. The compound
with p-hydroxy phenyl substitution, exhibits stronger antiradical activity as compared to the p-amino phenyl substitution due to
smaller O-H bond dissociation energy as compared to the N-H bond. From DPPH and DFT studies, it is suggested that the radical
scavenging activity in 2-substituted thiophene is occurred through proton transfer mechanism. The other possible SET, SPLET
mechanisms are also corroborated.
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Keywords Antioxidant ability Thiophene Phenol Stilbene Absorption Fluorescence Density functional theory Bond
.
.
dissociation energy Ionization energy Proton affinity
Introduction
petroleum products and in electronic device applications [1–4].
Some of the well known naturally occurring radical scavengers
are flavonoids [5–11], glutathione [12, 13], vitamin A, vitamin C
[14, 15], vitamin E [16], uric acid [17], caffeic and ferulic acids
[18, 19], β-carotene [10], curcumin [20–22], bilirubin [23].
However, some of these anti-radicals like β-carotene, vitamin
A, and vitamin E have no suppressing effect towards abnormal
biological activity. [24–26]. Thus, in recent years, there have
been growing interest in developing novel radical scavengers that
fulfill the need for industrial and pharmaceutical applications.
Many synthetic antioxidants were designed and synthesized to
improve the radical scavenging properties that can be used in
biological and industrial applications [27–34]. For example, an-
tiradicals based on stilbenoid [35–38], butylated hydroxy phenyl
compounds, BHA, BHT [1], dihydroquinoline ethoxyquin [39]
are useful for above applications. Recently, some of the thio-
phene based hybrid compounds, such as dithinyldene cyclohex-
anone (IC50: 4 μM) [40], thiophene based schiff base (IC50:
5 μM) [41] are known to exhibit remarkably higher radical scav-
enging properties and useful in wide range of applications
[42–46]. Thus, in order to grasp the basic fundamental of radical
scavenging activity, studies on the antiradical properties on var-
ious donor and acceptor substituted ethenyl thiophenes (1–8)
(Fig. 1) were carried out using DPPH assay and DFT methods.
Radical scavengers are active molecules, which play important
role in many area of chemistry, biology and material science such
as in food storage, cosmetic, pharmaceuticals, oil, rubber,
Anamika Gusain and Naresh Kumar contributed equally to this work.
Highlights
Thiophene compounds with p-hydroxy and p-amino phenyl substitutent,
exhibit antiradical activity with IC50 range from 45 μM to 165μM . The
activity is comparable to vitamin E (IC50 : 26 μM)
Correlation between the anti-radical activity with the ground state dipole
moment, bond dissociation enthalpy, ionization potential and proton af-
finity of thiophene compound is elucidated.
In thiophene compounds, the radical scavenging activity is predominantly
occurred through hydrogen atom transfer mechanism. The other possible
mechanisms such as SET, SPLET are also discussed.
Electronic supplementary material The online version of this article
(https://doi.org/10.1007/s10895-020-02629-5) contains supplementary
material, which is available to authorized users.
* Prasanta Kumar Hota
p.hota@hnbgu.ac.in
1
Department of Chemistry, School of Sciences, Hemvati Nandan
Bahuguna Garhwal University, Srinagar
(Garhwal), Uttarakhand 246174, India

J Fluoresc
Fig. 1 Structure of compounds 1–
8
It is shown that the thiophene compounds with donor substituent
(-OH and -NH2) exhibit antiradical properties, whereas the activ-
ity is reduced in presence of withdrawing substituent. In the
presence of thiophene compound, the DPPH radical is mostly
quenched through proton transfer mechanism. The results are
also supported by various thermodynamic parameters obtained
through TDDFT calculation.
in presence of pyridine-piperidine mixture as described earlier
[47–54], e.g. typical synthetic protocol for compound 1 is as
follows: 2-formyl thiophene (0.93 mL, 0.01 mol) was refluxed
with mixture of 10 mL of freshly distilled pyridine, 0.6 mL of
piperidine and 3.62 g, (0.02 mol) of p-nitrophenyl acetic acid
at 100 °C for eight hours and the progress of the reaction was
monitored by thin layer chromatography. The reaction mix-
ture was then cooled and poured in ice-cold water and treated
with 100 mL of diluted hydrochloric acid to remove excess of
pyridine from the reaction mixture. A yellow colored product
was extracted in chloroform and purified by column chroma-
tography using 2% ethyl acetate in petroleum ether as the
eluting solvent, when the desired compound was obtained in
30% yield. Compound 7 was obtained through reduction re-
action of 1 [52]. For this purpose, ethenyl thiophene 1 in
ethanol was refluxed in presence of aqueous ferrous sulfate
and ammonia solution at 100 °C for 3 h. All the products were
purified by column chromatography using 2–10% ethyl ace-
tate in petroleum ether (60–80 °C) as the eluting solvent.
Compound 8 was synthesized through alkylation of com-
pound 7 in presence of potassium-tert-butoxide. Compound
2 was prepared through condensation of 2-formyl thiophene
and corresponding phosphite using Wadsworth-Emmons re-
action [52, 55, 56]. All compounds show satisfactory physico-
chemical data (UV-Vis, FTIR, 1H and 13C NMR, GC-MS and
CHNS analysis).
Experiment
Material and Methods
Chemicals are purchased from M/s. Sisco Research Laboratory.
Radleys make, Carousel 6 plus reaction station was used for the
synthesis of compounds 1–8. Perkin Elmer Lambda 750 UV/
VIS/NIR spectrophotometer is used to record the absorption
spectra and Perkin Elmer LS-55 fluorescence spectrophotome-
ter is used to record the fluorescence spectra using a red PMT
detector system. FTIR spectra were recorded on a Impact
1
Nicolet-400 spectrophotometer using KBr discs. The H
NMR and 13C NMR spectra were recorded in CDCl3 on a
JEOL 500 MHz FTNMR instruments. GC-MS spectra were
recorded on a GCD 1800A Hewlett packard GC-mass spec-
trometer. CHNS analyses were carried out on a Theoquest CE
instrument 1112 series CHNS auto analyzer. Melting points
were determined on a Lab India make melting point apparatus.
For spectroscopic studies, UV grade solvents were used.
Radical Scavenging Activities
Synthesis of Compounds 1–8
For radical scavenging activity, 2,2-diphenyl-1-
picrylhydrazyl (DPPH) assay was carried out by following
the protocol described elsewhere [57]. In a typical experimen-
tal procedure, 100 μM concentration of DPPH solution was
prepared by adding 0.4 mL of 1 × 10−3 M methanolic solution
of DPPH. A varying concentration of testing compound (0 to
The synthetic scheme of all compounds are shown in Fig. 2.
The substituted p-phenyl ethenyl-E-thiophenes (1, 3–6) were
synthesized by the condensation of p-substituted phenyl acetic
acid with the corresponding 2-formylthiophene (2:1 M ratio)

J Fluoresc
Fig. 2 Synthetic scheme for
ethenyl thiophenes (1–8); (i) pyr-
idine-piperidine, 100 °C, 8 h; (ii)
(a) p-cyano benzyl bromide,
P(OEt)3, 150 °C, 3 h; (b) DMF,
NaH, 0 °C, 1 h; (iii) FeSO4, aqu.
NH3, ethanol, 100 °C, 2 h; (iv)
Potassium-tert-butoxide, tert-bu-
tyl alcohol, Butyl bromide, r.t. 3 h
400 μM) was then added to DPPH solution depending upon
the antiradical activity. The decrease in the absorbance of
DPPH radical at 517 nm was then measured at regular interval
of time (0–45 min duration). The DPPH solution with stan-
dard antioxidant, vitamin E is used as a positive control. All
the experiments were performed in triplicate and the average
of absorbance was taken for calculating the inhibition concen-
tration. The 50% inhibition concentration (IC50) is the concen-
tration of antioxidant at which the 50% of absorbance of
DPPH radical is quenched with respect to the control
(Ablank). The IC50 is calculated from the plot of % inhibition
vs. concentration of antioxidant, and using the eq. 1.
%Inhibition of DPPH free radical
hn
ꢀ
i
¼
Ablank−Asample =ðAblankÞ x 100
ð1Þ
where Ablank = Absorbance of DPPH radical in absence of
antioxidant; Asample = Absorbance of DPPH radical in pres-
ence of varying concentration of antioxidant.
Table 1 Ground state dipole moment (μg), absorption (λabs) and fluorescence wavelength maximum (λem), extinction coefficient (ε), optical band gap,
DPPH radical inhibition concentration (IC50) of ethenyls thiophenes in methanol
Com
μg (Debye)
λ
abs (nm)a
λ
em (nm)a
ε (M−1 cm−1
)
Band Gap(eV)b
Band Gap(eV)c
Band Gap(eV)d
IC50 (μM)
1 -NO2
2 -CN
3 -Cl
6.65
5.60
1.87
0.34
1.19
1.42
3.00
3.48
372
340
327
321
332
325
343
367
614
412
384
385
387
378
435
420
21,500
39,000
30,400
21,000
44,500
24,280
27,500
12,400
2.89
3.28
3.46
3.51
3.43
3.47
3.20
2.88
3.14
3.31
3.49
3.55
3.44
3.53
3.19
3.16
3.51
3.73
3.90
3.98
3.85
3.90
3.75
3.64
–
–
–
4 -H
–
5 -OCH3
6 -OH
7-NH2
8-NR2
–
45
165
322
a Experimentally obtained; Vitamin E (IC50, 26 μM, μg: 0.80); ascorbic acid (IC50 ~ 11 μM); 4-hydroxy stilbene (IC50 ~ 24 μM);
b Obtained through Tauc plot,
c Obtained through intersection of absorption and fluorescence spectra,
d Obtained through DFT

J Fluoresc
Fig. 5 Absorption spectra of compounds 1–8 in methanol
Fig. 3 A typical absorption spectra of 100 μM of DPPH radical alone and
in presence of 25 μM concentration of ethenyl thiophenes 6–8 and
vitamin E
predicted bond dissociation energy is most reliable and com-
parable with other computational methods [62, 63]. The DFT
method is validated using known compounds, vitamin E, 4-
hydroxy stilbene, 2,4,6-tri-tert-butylphenol, hydrogen radical,
whose results are well matched with the previously reported
experimental and theoretical data [64–66].
Time Dependent Density Functional Theory (TDDFT)
For the calculation of thermodynamic parameters (BDE, IP,
PA, PDE, ETE), the ab initio quantum chemical software
package ORCA is used [58]. The ground state dipole moment,
absorption and fluorescence wavelength maximum, the verti-
cal excitation energy and oscillator strength (ƒ) is computed
using time-dependent density functional theory (TDDFT) [59,
60]. The ground and excited state of the neutral, free radical,
cationic radical and anionic thiophenes are optimized through
B3LYP and BLYP functional using def2-SVP and aug-cc-
pVDZ basis set respectively [61]. The minimized geometry
is further confirmed by vibrational analysis, resulting in no
imaginary frequencies. This geometry is used as the input
for further calculations to obtain the frontier molecular orbitals
(FMOs), UV-Vis and fluorescence spectra. The TDDFT
Results and Discussion
Radical Scavenging Properties of Ethenyl Thiophene
The radical scavenging properties of thiophene compounds
(1–8) are carried out using DPPH assay in methanol. In gen-
eral, the DPPH free radical is quenched at 517 nm in presence
of an antioxidant. It is observed that the absorbance of DPPH
radical is quenched in the presence of thiophene compounds
6–8, whereas, the absorbance of DPPH radical is almost
Fig. 4 Plot of % of DPPH radical quenching vs. concentration of ethenyl
thiophenes 1–8
Fig. 6 Fluorescence spectra of compounds 1–8 in methanol

J Fluoresc
Correlation of Antiradical Activity with Optical
Properties
In order to understand the anti-radical activity in detail, the
optical properties of the molecule are studied. The absorption
and fluorescence spectra of 1–8 were recorded in solvent of
varying polarity (Fig. S1-S2). It is shown that the absorption
(λabs) and fluorescence wavelength (λem) of all these thio-
phenes are red shifted from non-polar solvent n-hexane to
polar solvent DMF (Table S1). The molar extinction coeffi-
cient (ε) of thiophene compounds lies in between
10,000 M−1 cm−1 to 30,000 M−1 cm−1. These indicate the
π → π* nature of transition in thiophene compounds 1–8
(Table 1, Figs. 5 and 6). The λabs is moderately red shifted
from non-polar solvent, n-hexane to polar solvent, DMF, by
15 nm, 3 nm, 4 nm, 4 nm, 5 nm, 3 nm, 17 nm and 17 nm,
whereas the λem is red shifted by 156 nm, 6 nm, 8 nm, 13 nm,
9 nm, 0 nm, 45 nm and 24 nm for 1–8 respectively. As com-
pared to 2-[phenyl ethenyl-E thiophene](4), the λabs and λem
of 2-[4-nitro phenyl ethenyl-E-thiophene](1) are red shifted by
51 nm and 229 nm respectively. Similarly, λabs and λem of 8
are red shifted by 45 nm and 35 nm, whereas a minimal
change of 0–4 nm is observed for thiophenes 3–6. The λem
is highly sensitive to solvent polarity and p-phenyl substituent.
The large red shift of λem in 1 suggest the involvement of
charge transfer excited state for 1, whereas, a moderate red
shift in 7 and 8 suggest a partial charge transfer in the excited
state of amine compounds 7 and 8. These type of charge
transfer phenomena are very common in nitro and amine com-
pounds [51–54]. The change in excited state dipole moment is
obtained for 1–8 as 13.85 Debye, 2.54 Debye, 2.47 Debye,
4.41 Debye, 1.91 Debye, 1.78 Debye, 6.52 Debye and 5.20
Debye respectively using McRay Plot (Fig. 7, Table S2).
These further indicate that compound 1 is highly dipolar and
exhibits charger transfer excited state, whereas amine
Fig. 7 McRay Plot, Stokes’ shift vs. solvent polarity parameter, F(ε,n) of
1–8
unaffected in presence of other thiophene compounds 1–5.
The % of quenching of DPPH radical is obtained from the
plot of % of inhibition vs. compound concentration (Fig. 3
and 4). The hydroxy (6) and amino (7) substituted thiophene
compounds exhibit radical scavenging properties with IC50:
45 μM and 165 μM respectively (Table 1). Upon O-alkylation
and N-alkylation of hydroxy and amino functional group re-
spectively, the radical quenching activity of 5 and 8 is drasti-
cally reduced (IC50 ~ 322 μM for 8 and no activity for 5). On
the other hand, thiophene compounds (1–5) do not show any
radical scavenging activity. Thus, the hydroxy and amino
compounds are capable of transferring hydrogen to the
DPPH radical and subsequently, the quenching of DPPH rad-
ical is occurring. These indicate that the thiophene compounds
quench the DPPH radical predominantly through the hydro-
gen atom transfer (HAT) mechanism, which leads to the for-
mation of neutral DPPHH molecule.
Fig. 8 TDDFT computed HOMO-LUMO energy of ethenyls 1–8

J Fluoresc
Fig. 11 Optical band gap of 1–8 and vitamin E (9)
lowest HOMO energy is observed (1: HOMO: −6.09 eV,
LUMO: −2.58 eV; 2: −5.92 eV, LUMO -2.18 eV; 3:
−5.60 eV, LUMO -1.69 eV; 4: −5.51 eV, LUMO -1.53 eV),
whereas a highest HOMO energy is computed for hydroxy
and amine compounds (6: HOMO: −5.21 eV, LUMO -
1.31 eV, 7:HOMO: −4.88 eV, LUMO -1.13 eV, 8: HOMO:
−4.77 eV, LUMO: −1.12 eV).
Similarly, the ground state dipole moment is computed for
1–8 using DFT method. It is shown that 1, 2, 7 and 8 exhibit
large ground state dipole moment (1: 6.65 Debye, 2: 5.60
Debye, 7: 3.00 Debye, 8: 3.48 Debye), whereas other thio-
phene compounds (3–6) show a small dipole moment
(Vitamin E: 0.80 Debye, 3: 1.87 Debye, 4: 0.34 Debye, 5:
1.19 Debye, 6: 1.42 Debye). As compared to 4, the dipole
moment is increased with increasing the electron withdrawing
capacity of p-pheny substituent (1-NO2: 6.65 Debye, 2-CN:
5.60 Debye, 3-Cl: 1.87 Debye, 4-H: 0.34 Debye) and also
with increasing the electron donating capacity of p-phenyl
substituent (8-NR2: 3.48 Debye, 7-NH2: 3.00 Debye, 6-OH:
1.42 Debye, 4-H: 0.34 Debye) (Fig. 12).
Fig. 9 TDDFT computed UV-Vis absorption spectra of compounds 1–8
compounds 7 and 8 undergo partial charge transfer and other
compounds have non-polar excited state.
The optical band gap of these compounds were obtained
using both experimental and theoretical methods (Table 1,
Table S3, Fig. S3). In all the methods, the optical band gap
follows the similar trend. The ground and excited state of thio-
phene compound are stabilized more in presence an electron
withdrawing p-phenyl substituent. However, as compared to
the unsubstituted thiophene compound 4, the optical band gap
of substituted compound is decreased either by an electron
withdrawing or electron donating p-phenyl substituent
(Table 1). The optical band gap obtained through DFT method
is little larger than the experimental method. The effect of sol-
vent is not taken in to account in DFT method and thus, there is
a less stabilization of ground and excited state in DFT method.
This leads to a shorter emission wavelength with little larger
optical band gap (Figs. 8, 9, 10 and 11, Fig. S3).
The HOMO and LUMO energy of ethenyl thiophene are
gradually increased in presence of an electron donating sub-
stituent. For nitro, cyano and chloro compounds (1–3), a
Thus, thiophene is acting as an electron donor or electron
acceptor depending upon the nature of p-phenyl substitution.
Interestingly, thiophenes with electron withdrawing substitu-
ent (NO2, CN, Cl) do not show any radical scavenging
Fig. 10 TDDFT computed fluorescence spectra of compounds 1–8
Fig. 12 Ground state dipole moment of 1–8 and vitamin E (9)

J Fluoresc
Table 2 TDDFT computed
hydrogen bond dissociation
(BDE), ionization potential (IP),
proton dissociation (PDE), proton
affinity (PA) and electron transfer
(ETE) energy (Kcal/mol) of
ethenyl thiophenes (1–8)
Compound
BDE ΔBDE IP
ΔIP
PDE
ΔPDE PA
ΔPA
ETE ΔETE
Vitamin E
1-NO2
2-CN
82.9
0
157.4
0
236.3
0
–
–
–
–
–
365.3
0
–
–
–
–
–
29.1
0
–
–
177.0 19.6
172.6 15.1
165.1 7.7
164.5 7.0
155.5 −1.9
156.6 −0.7
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
–
3-Cl
–
–
–
4-H
–
–
–
5-OCH3
6-OH
–
–
–
89.5
6.6
243.7 7.4
261.7 25.4
–
355.6 −9.68 45.5
16.4
IC50: 45 μM
7-NH2
99.1
16.1
148.3 −9.2
141.5 −15.8
379.5 14.2
31.1
2.0
IC50: 165 μM
8-NR2
–
–
–
–
–
IC50: 322 μM
ΔBDE = BDE-BDEvitamin E; ΔIP = IP-IPvitamin E; ΔPA = PA-PAvitamin E; ΔPDE = PDE-PDEvitamin E; ΔETE =
ETE-ETEvitamin E
•þ
IP ¼ EAr−X −EAr−XH
ð3Þ
ð4Þ
ð5Þ
ð6Þ
H
properties. On the other hand thiophene compounds (6–8)
with electron donating substituent such as amine and hydroxy,
exhibit anti-radical properties. The radical scavenging efficacy
of such compounds, however, is decreased further upon alkyl-
ation. The order of radical scavenging activity is: Vitamin E >
6-OH > 7-NH2 > 8-NR2. Thus, in these compounds, the radi-
cal scavenging activity is directly related to the optical band
gap (Fig. 11), whereas inversely related to the dipole moment
of the thiophene compounds (Fig. 12). The anti-radical mech-
anism is very complex and in order to understand the mecha-
nism in detail, the thermodynamic parameters such as BDE,
IP, PA, PDE, ETE of the molecules (1–8) are calculated using
the eq. 2–6 and the data is shown in Table 2.
−
PA ¼ EAr−X þ EHþ−EArXH
PDE ¼ EAr−X þ EHþ−EArX
•
•þ
H
•
−
−
ETE ¼ EAr−X þ Ee −EAr−X
where EAr-XH, EAr-X•, EAr-X H, EAr-X−, EH•, EH+, Ee− are the
enthalpies of Ar-XH, Ar-X• radical, Ar-X•+H cationic radical,
anion Ar-X−, H radical, H+ cation, electron respectively
[67–80].
Many radical scavenging mechanisms are well known in
the literature [67–80]. These include hydrogen atom transfer
(HAT) [67–70], single electron transfer (SET) [70–73], radi-
cal adduct formation (RAF) [74], sequential proton loss and
electron transfer (SPLET) [75, 76], sequential electron proton
transfer [SEPT] [77–79], sequential proton loss hydrogen
•+
BDE ¼ EAr−X• þ EH•−EAr−XH
ð2Þ
Scheme 1 Some of the plausible radical scavenging mechanisms, HAT, SET, SPLET, SEPT.

J Fluoresc
atom transfer (SPLHAT) [80] etc. (Scheme 1). The HAT
mechanism is associated with the hydrogen bond dissociation
enthalpy (BDE), whereas, SET is associated with ionization
potential (IP) of the antiradical. Similarly, SPLET is associat-
ed with proton affinity (PA) and electron transfer enthalpy
(ETE), whereas SPET is associated with both IP and PA of
the antiradical. Thus, thermodynamic parameters, such as
BDE, IP, PA, PDE, ETE provide most valuable information
in predicting the plausible mechanism.
From Table 2, the IP energy of 1–4 is found to be very large
(IP: 164.17–177.03 Kcal/mol), as compared to 5–8 (IP:
141.55–156.63 Kcal/mol). In 5–8 and vitamin E, the trend
of IP energy is vitamin E ~ 6 > 7 > 8. and the trend of radical
scavenging efficacy is: vitamin E > 6 > 7 > 8. It is known that
electron withdrawing substituent stabilize the neutral mole-
cule, and destabilize the radical and radical cation, which leads
to a higher IP energy [69, 70]. On the other hand, electron
donating substituent stabilized the radical, radical cation and
destabilized the neutral molecule, which leads to decrease in
the IP energy [70, 72]. Thus, compound with higher antirad-
ical activity should have a lower IP energy to act through the
SET mechanism [71, 72]. Similarly, the proton dissociation
energy (PDE) provides useful information for the later step of
SET-PT mechanisms [81] (Scheme 2). The PDE of cationic
radical of vitamin E, 6 and 7 is found as 236 kcal/mol,
243 kcal/mol, and 261 kcal/mol respectively. Thus PDE is
larger for thiophene with strong electron donating amine
substituent and it requires higher energy for the dissociation
of proton from the radical cation intermediate. Thus, it is sug-
gested that these compounds may not follow anti-radical ac-
tivity through SET or SET-PT mechanism.
To confirm the hydrogen atom transfer (HAT) mecha-
nism, the BDE is calculated for 6–7 and vitamin E. The O-
H and N-H bond dissociation energy (BDEO-H, BDEN-H) is
little larger for 6 and 7 (89 Kcal/mol for 6 and 99 Kcal/mol
for 7) as compared to the vitamin E (82.97 Kcal/mol). In 6
and 7, the O-H and N-H bond dissociation energy is in-
creased by 6 Kcal/mol (ΔBDEO-H) and 16 Kcal/mol
(ΔBDEN-H), as compared to vitamin E respectively. The
trend of BDE is: 7 > 6 > vitamin E and the radical scaveng-
ing activity is also reduced in the order Vitamin E > 6 > 7.
This trend of BDE and anti-radical activities is in accord to
the HAT mechanism. Further, replacing the OH and NH2
with methoxy (5: -OCH3) and di-alkyl (8: -N(C4H9)2]
group, results in the reduction of anti-radical activity.
These suggest that thiophene compounds exhibit antiradical
activities through hydrogen atom transfer (HAT) mecha-
nism (Scheme 2). In general, the antiradical activity of phe-
nol and amine compounds occur through H atom transfer
mechanism (HAT). Such compounds have smaller BDE
with more stabilized phenoxyl or imine radical [62, 67–69,
81–88]. In thiophene compounds, 6 and 7, the BDE of N-H
bond is 10 kcal/mol higher than O-H bond. This indicates
that the imine radical is less stable than the phenoxyl radical
Scheme 2 Possible antiradical mechanisms in ethenyl thiophenes

J Fluoresc
Acknowledgements PKH, AG, NK and JK are grateful to University
Grants Commission, New Delhi for research grant (No F.30-72/2014-
BSR) and research fellowship. GP is the recipient of national postdoctoral
research fellowship from Department of Science & Technology (DST),
New Delhi. Authors acknowledged AMRC, IIT Mandi for 1H NMR and
13C NMR facility.
and thus, compound 7 exhibits less anti-radical activity
compared to the hydroxy compound 6.
The other possible mechanism is the sequential proton loss
and electron transfer (SPLET) mechanism via the anionic in-
termediate (Scheme 2). In this mechanism the role of PA is
important. The PA for phenyl N-H and phenol O-H bond is
379 kcal/mol and 355 kcal/mol respectively. In the present
thiophene compounds, the trend of PA energy is: 7 > 6 and
the antiradical activity is in the order 6 > 7. If the anion gen-
erate through proton abstraction by following the SPLET
mechanism as shown in Scheme 2, the phenoxyl anion of
ethenyl thiophene could be stabilized more as compared to
the aminyl anion and consequently decrease in PA for 6 as
compared to 7 and increase in the antiradical activity of 6.
However, the ETE of phenoxyl anion (6) is 14 Kcal/mol is
larger compared to amine anion (7), which is the second step
of SPLET pathway (ETE: 45 kcal/mol for 6, 31 kcal/mol for
7). A more stabilized anion requires higher energy to transfer
electron to the free radical [69, 86]. Thus, SPLET mechanism
can be ruled out. Thus, in 6 and 7 as the BDE is smaller than IP
and PA energy, the antiradical activity could be through ther-
modynamically controlled HAT mechanism.
Author Contributions PKH, AG, NK synthesized and characterized the
1
compounds using H and 13C NMR, GC-MS, FTIR techniques. PKH,
AG, NK carried out the absorption, fluorescence measurement and ana-
lyzed the data. PKH, AG, NK and GP measured the antiradical activity.
PKH and JK designed and JK carried out the DFT calculation. PKH, AG
and NK wrote the paper.
References
1. Yehye WA, Rahman NA, Ariffin A, Hamid SBA, Alhadi AA,
Kadir FA, Yaeghoobi M (2015) Understanding the chemistry be-
hind the antioxidant activities of butylated hydroxytoluene (BHT):
A review. Eur J Med Chem 101:295–312
2. Jahnert T, Hager MD, Schubert US (2014) Application of phenolic
radicals for antioxidants, as active materials in batteries, magnetic
materials and ligands for metal-complexes. J Mater Chem A 2:
15234–15251
3. Carocho M, Ferreira ICFR (2013) A review on antioxidants,
prooxidants and related controversy: natural and synthetic com-
pounds, screening and analysis methodologies and future perspec-
tives. Food Chem Toxicol 51:15–25
4. Schmidt S, Pokorny J (2005) Potential application of oilseeds as
sources of antioxidants for food lipids - a review. Czech J Food Sci
23:93–102
5. Christen S, Peterhans E, Stocker R (1990) Antioxidant activities of
some tryptophan metabolites: possible implication for inflammato-
ry diseases. Proc Natl Acad Sci 87:2506–2510
6. Lunder TL (1992) Catechins of green tea: antioxidant activity. In
M. T. Huang, C. T. Ho, and C.Y. Lee. Eds., Phenolic compounds in
Food and their effects on health. II. Antioxidants and cancer pre-
vention. Am Chem Soc Sympos Ser. 114–120
Conclusion
In summary, the antiradical activities of p-phenyl substituted
ethenyl thiophenes were studied using DPPH assay and den-
sity functional theory. It is shown that ethenyls with strong
electron-donating substituent (NH2, OH) exhibit antiradical
activity, whereas ethenyls with electron withdrawing substit-
uent do not show antiradical activity. From the studies on the
optical properties, it is shown that ethenyl thiophene with
small ground state dipole moment and large optical band
gap, exhibits good antiradical properties.
7. Tournaire C, Croux S, Maurette MT, Beck I, Hocquaux M, Braun
AM, Oliveros E (1993) Antioxidant activities of flavonoids: effi-
ciency of singlet oxygen quenching. J Photochem Photobiol B 19:
205–215
From the studies on the thermodynamic parameters, it is
shown that amine and hydroxy substituted thiophenes have
smaller bond dissociation enthalpy (BDE) as compared to
ionization energy (IE) and proton affinity (PA). Thus, for
quenching of free radical, these compounds follow the ther-
modynamically controlled HAT mechanism. Ethenyls with
electron withdrawing substituent (NO2, CN, Cl) do not
show any antiradical properties owing to higher IP energy
and lack of loosely bound hydrogen atom. It is also note-
worthy to mention that the antiradical mechanism is very
complex and hence, kinetic and radical quenching studies
can provide more detail insights into the mechanism of ac-
tion. Overall, the present ethenyl thiophene exhibits substit-
uent dependent antiradical activity, which is interesting.
These result, however, provide a very useful information
in designing future molecules that exhibit efficient antirad-
ical activities.
8. Po-Geller B, Reiter RJ, Hardeland R, Tan DX, Barlow-Walden LR
(1996) Melatonin and structurally related endogenous indoles act as
potent electron donors and radical scavengers in vitro. Redox Rep
2:179–184
9. Kohen R, Jamamoto J, Cundi KC, Ames BN (1988) Antioxidant
activity of carnosine, homocarnosine, and anserine present in mus-
cle and brain. Proc. Nat. Acad. Sci. 81:3175–3179
10. Richard AL (1997) Naturally occurring antioxidants. Lewis pub-
lishers. New York, 1st edition
11. Cheynier V (2005) Polyphenols in foods are more complex than
often though. Am J Clin Nutr 81:223S–229S
12. Morse ML, Dahl RH (1978) Cellular glutathione is a key to the
oxygen effect in radiation damage. Nature 271:660–662
13. Hausladen A, Alscher RG (1993) Gluthanione antioxidants in
higher plants. CRC Press, Boca Raton, FL, pp 1–30
14. Kramer GF, Norman HA, Krizek DT, Mirecki RM (1991)
Influence of VV-B irradiation on polyamines, lipid peroxidation
and membrane lipids in cucumber. Phytochemistry 30:2101–2108
15. Aruoma OI (1993) Free radicals and food. Chem Brit:210–214

J Fluoresc
16. Osawa T, Kumazawa S, Kawakishi S (1991) Prunusols A and B,
novel antioxidative tocopherol derivatives isolated from the leaf
wax of Prunusgrayana maxim. Agric Biol Chem 55:1727–1731
17. Simic MG, Jovanovic SV (1989) Antioxidation mechanisms of uric
acid. J Am Chem Soc 111:5778–5882
38. Madadi NR, Zong H, Ketkar A, Zheng C, Penthala NR, Janganati
V, Bommagani S, Eoff RL, Guzman ML, Crooks PA (2015)
Synthesis and evaluation of a series of resveratrol analogues as
potent anti-cancer agents that target tubulin. Med Chem Comm 6:
788–794
18. Graf E (1992) Antioxidant potential of ferulic acid. Free Rad. Biol.
Med. 13:435–448
39. Kumar S, Engman L, Valgimigli L, Amorati R, Fumo MG, Pedulli
GF (2007) Antioxidant profile of ethoxyquin and some of its S, se
and Te analogues. J. Org. Chem. 72:6046–6055
40. Kar S, Ramamoorthy G, Sinha S, Ramanan M, Pola JK, Golakoti
NR, Nanubolu JB, Sahoo SK, Dandamudi RB, Doble M (2019)
Synthesis of diarylidenecyclohexanone derivatives as potential
anti-inflammatory leads against COX- 2/mPGES1 and 5-LOX.
New J Chem 43:9012–9020
19. Terao J, Karasawa H, Arai H, Nagao A, Suzuki T (1993) Peroxyl
radical scavenging efficiency of caffeic acid and its related phenolic
compounds in solution. Biosci Biotechnol Biochem 57:1204–1205
20. Tonnesen HH, Greenhill JV (1992) Studies on coumarine and
cucurminoids. XXII. Curcumin as a reducing agent and as a radical
scavenger. Int J Pharmaceut 87:79–87
21. Tonnesen HH, Smistad G, Agren T, Karlsen J (1993) Studies on
coumarine and cucurminoids. XXIII. Effects of Curcumin on lipo-
somal lipid peroxidation. Int J Pharmaceut 90:221–228
22. Bhattacharya M, Mandal P, Sen A (2009) In vitro detection of
antioxidants in different solvent fractions of ginger (Zingiber
officinale Rosc.), Indian. J Plant Physiol 14:23–27
23. Stocker R, Glazer AN, Ames BN (1987) Antioxidant activity of
albumin-bound bilirubin. Proc Nat Acad Sci 84:5918–5922
24. Jiang L (2010) Efficacy of antioxidant vitamins and selenium sup-
plement in prostate cancer prevention: a meta-analysis of random-
ized controlled trials. Nutr Cancer 62:719–727
41. Shanty AA, Mohanan PV (2018) Heterocyclic schiff bases as non
toxic antioxidants: solvent effect, structure activity relationship and
mechanism of action. Spectrochim Acta A 192:181–187
42. Tenti G, Egea J, Villarroya M, Leon R, Fernandez JC, Padin JF,
Sridharan V, Ramos MT, Menendez JC (2013) Identification of 4,
6-diaryl-1,4-dihydropyridines as a new class of neuroprotective
agents. Med Chem Commun 4:590–594
43. Aguiara ACV, Moura RO, Junior JFBM, Rocha HAO, Camara
RBG, Schiavona MSC (2016) Evaluation of the antiproliferative
activity of 2-amino thiophene derivatives against human cancer
cells lines. Biomed Pharmacother 84:403–414
44. Cardoso LNF, Nogueira TCM, Rodrigues FAR, Oliveira ACA, dos
Santos Luciano MC, Pessoa C, de Souza MVN (2017) N-
acylhydrazones containing thiophene nucleus: a new anticancer
class. Med Chem Res 26:1605–1608
45. Luo Y, Li X, Chen T, Wang Y, Zheng W (2012) Synthesis of a
novel thiophene derivative that induces cancer cell apoptosis
through modulation of AKT and MAPK pathways. Med Chem
Commun 3:1143–1146
25. Abner EL (2011) Vitamin E and all-cause mortality: a meta-analy-
sis. Current Aging Science 4:158–170
26. Bjelakovic G, Dimitrinka N, Christian G (2013) Meta-regression
analyses, meta-analyses, and trial sequential analyses of the effects
of supplementation with beta-carotene, vitamin A, and vitamin E
singly or in different combinations on all-cause mortality: do we
have evidence for lack of harm? PLoS One 8:74558–74755
27. Palozza P (2002) Design, synthesis, and antioxidant activity of
FeAOX-6, a novel agent deriving from a molecular combination
of the chromanyl and polyisoprenyl moieties. Free Rad Biol Med
33:1724–1735
28. Estevao MS, Carvalho LC, Ribeiro D, Couto D, Freitas M, Gomes
A, Marques MMB (2010) Antioxidant activity of unexplored in-
dole derivatives: synthesis and screening. Eur J Med Chem 45:
4869–4878
46. Zoubi WA, Mohamed SG, A-Hamdani AAS, Mahendradhany AP,
Ko YG (2018) Acyclic and cyclic imines and their metal com-
plexes: recent progress in biomaterial and corrosion applications.
RSC Adv 8:23294–23318
47. Singh AK, Hota PK (2003) Photoreactivity of donor-acceptor eth-
enes. Indian J Chem B 42:2048–2053
48. Singh AK, Hota PK (2005) Absorption and fluorescence spectral
properties of donor acceptor ethenes bearing indole and p-nitrophe-
nyl substituents. Res Chem Intermed 31:85–101
29. Mahajan P (2017) Synthesis, antioxidant, and anti-inflammatory
evaluation of novel thiophene-fused quinoline based β-diketones
and derivatives. J Heterocy Chem 54:1415–1422
30. Shen T, Wang X-N, Lou H-X (2009) Natural stilbenes: an over-
view. Nat Prod Rep 26:916–935
31. Jagtap UB, Bapat VA (2010) Artocarpus: A review of its traditional
uses, phytochemistry and pharmacology. J Ethnopharmacol 129:
142–166
49. Singh AK, Hota PK (2006) Fluorescence and photoisomerization
studies of p-nitrophenyl substituted indolic ethenes. J Phys Org
Chem 19:43–52
50. Singh AK, Hota PK (2007) Ethenyl indoles as neutral hydrophobic
fluorescence probes. J Phys Org Chem 20:624–629
32. Nopo-Olazabal C, Hubstenberger J, Nopo-Olazabal L, Medina-
Bolivar F (2013) Antioxidant activity of selected stilbenoids and
their bioproduction in hairy root cultures of muscadine grape (Vitis
rotundifolia michx.). J Agr Food Chem 61:11744–11758
33. Charles DJ (2013) Antioxidant properties of spices, herbs and other
sources. Springer, New York
34. Nimse SB, Pal D (2015) Free radicals, natural antioxidants, and
their reaction mechanism. RSC Adv 5:27986–28006
35. Fan G-J, Liu X-D, Qian Y-P, Shang Y-J, Li X-Z, Dai F, Fang J-G,
Jin X-L, Zhou B (2009) 4,4′-Dihydroxy-trans-stilbene, a resveratrol
analogue, exhibited enhanced antioxidant activity and cytotoxicity.
Biorg Med Chem 17:2360–2365
36. Tang J-J, Fan G-J, Dai F, Ding D-J, Wang Q, Lu D-L, Li R-R, Li X-
Z, Hu L-M, Jin X-L, Zhou B (2011) Finding more active antioxi-
dants and cancer chemoprevention agents by elongating the conju-
gated links of resveratrol. Free Radic Biol Med 50:1447–1457
37. Su D, Cheng Y, Liu M, Liu D, Cui H, Zhang B, Zhou S, Yang T,
Mei Q (2013) Comparison of piceid and resveratrol in antioxidation
and antiproliferation activities in vitro. PLoS One 8:e54505
51. Kumar N, Kumar J, Hota PK (2017) Substituent dependence charge
transfer and photochemical properties of donor-acceptor substituted
ethenyl thiophenes. J Fluoresc 27:1729–1738
52. Kumar N, Paramasivam M, Kumar J, Gusain A, Hota PK (2018)
Substituent dependent optical properties of p-phenyl substituted
ethenyl-E-thiophenes. J Fluoresc 28:1207–1216
53. Kumar N, Kumar J, Hota PK (2018) Substituent dependent
photoreactivity of donor- acceptor substituted phenyl ethenes.
Lett Org Chem 15:479–484
54. Kumar N, Paramasivam M, Kumar J, Gusain A, Hota PK (2019)
Tuning of optical properties of p-phenyl ethenyl -E-furans: A
solvatochromism and density functional theory. Spectrochim Acta
A 206:396–404
55. Wadsworth WS Jr, Emmons WD (1961) The utility of phosphonate
carbanions in olefin synthesis. J Am Chem Soc 83:1733–1738
56. Singh AK, Hota PK (2006) Substituent directed distal
photoisomerisation of indolic dienyl chromophores. Indian J.
Chem. B 45:2469–2473

J Fluoresc
57. Marinova G, Batchvarov V (2011) Evaluation of the methods for
determination of the free radical scavenging activity by DPPH.
Bulg J Agric Sci 17:11–24
74. Liebler DC, McClure TD (1996) Antioxidant reactions of β-caro-
tene: identification of carotenoid-radical adducts. Chem Res
Toxicol 9:8–11
75. Dorovic J, Dimitric Markovic JM, Stepanic V, Begovic N, Amic D,
Markovic Z (2014) Influence of different free radicals on scaveng-
ing potency of gallic acid. J Mol Model 20:2345
76. Alberto ME, Russo N, Grand A, Galano A (2013) A physicochem-
ical examination of the free radical scavenging activity of Trolox:
mechanism, kinetics and influence of the environment. Phys Chem
Chem Phys 15:4642–4650
77. Nakanishi I, Kawashima T, Ohkubo K, Kanazawa H, Inami K,
Mochizuki M, Fukuhara K, Okuda H, Ozawa T, Itoh S,
Fukuzumi S, Ikota N (2005) Electron-transfer mechanism in
radical-scavenging reactions by a vitamin E model in a protic me-
dium. Org Biomol Chem 3:626–629
78. Ouchi A, Nagaoka SI, Abe K, Mukai K (2009) Kinetic study of the
aroxyl radical-scavenging reaction of α- tocopherol in methanol
solution: notable effect of the alkali and alkaline earth metal salts
on the reaction rates. J Phys Chem B 113:13322–13331
79. Iuga C, Alvarez-Idaboy JR, Russo N (2012) Antioxidant activity of
trans-resveratrol toward hydroxyl and hydroperoxyl radicals: a
quantum chemical and computational kinetics study. J Org Chem
77:3868–3877
80. Leopoldini M, Chiodo SG, Russo N, Toscano M (2011) Detailed
investigation of the OH radical quenching by natural antioxidant
caffeic acid studied by quantum mechanical models. J Chem
Theory Comput 7:4218–4233 (ACS)
81. Klein E, Lukes V (2007) DFT/B3LYP study of the substituent
effect on the reaction enthalpies of the individual steps of sequential
proton loss electron transfer mechanism of phenols antioxidant ac-
tion: correlation with phenolic CO bond length. J Mol Struct
THEOCHEM 805:153–160
58. F. Neese, The ORCA program system, Wiley Interdiscip. Rev.:
Comput. Mol. Sci., 2 (2012) 73–78
59. Dreuw A, Head-Gordon M (2005) Single reference ab initio
methods for the calculation of excited states of large molecules.
Chem Rev 105:4009–4037
60. Kulhánek J, Bureš F, Wojciechowski A, Makowska-Janusik M,
Gondek E, Kityk IV (2010) Optical operation by chromophores
featuring 4,5-dicyanoimidazole embedded within poly (methyl
methacrylate) matrices. J Phys Chem A 114:9440–9446
61. Weigend F, Ahlrichs R (2005) Balance basis sets of split valence,
triple zeta valence and quadruple zeta valence quality for H to Rn:
design and assessment of accuracy. Phys Chem Chem Phys 7:
3297–3305
62. Leopoldini M, Russo N, Toscano M (2011) The molecular basis of
working mechanism of natural polyphenolic antioxidants. Food
Chem 125:288–306
63. DiLabio GA (1999) Using locally dense basis sets for the determi-
nation of molecular properties. J Phys Chem A 103:11414–11424
64. Wright JS, Johnson ER, DiLabio GA (2001) Predicting the activity
of phenolic antioxidants: theoretical method, analysis of substituent
effects, and application to major families of antioxidants. J Am
Chem Soc 123:1173–1183
65. Mahoney LR, Ferris FC, DaRooge MA (1969) Calorimetric study
of the 2,4,6-tri-tert- butylphenoxy radical in solution. J Am Chem
Soc 91:3883–3889
66. Denisov ET, Khudyakov IV (1987) Mechanisms of action and
reactivities of the free radicals of inhibitors. Chem Rev 87:1313–
1357
67. Markovic ZS, Mentus SV, Dimitric Markovic JM (2009)
Electrochemical and density functional theory study on the reactiv-
ity of fisetin and its radicals: implications on in vitro antioxidant
activity. J Phys Chem A 113:14170–14179
82. Burton GW, Ingold KU (1986) Vitamin E: application of the prin-
ciples of physical organic chemistry to the exploration of its struc-
ture and function. Acc Chem Res 19:194–201
83. King A, Young G (1999) Characteristics and occurrence of pheno-
lic phytochemicals. J Am Diet Assoc 99:213–218
84. Ban F, Lundqvist MJ, Boyd RJ, Eriksson LA (2002) Theoretical
studies of the cross-linking mechanisms between cytosine and ty-
rosine. J Am Chem Soc 124:2753–2761
85. Fang Y, Liu L, Feng Y, Li X-S, Guo Q-X (2002) Effects of hydro-
gen bonding to amines on the phenol/phenoxyl radical oxidation. J
Phys Chem A 106:4669–4678
86. Robbins RJ (2003) Phenolic acids in foods: an overview of analyt-
ical methodology. J Agr Food Chem 51:2866–2887
87. Singh N, Loader RJ, O’Malley PJ, Popelier PLA (2006)
Computation of relative bond dissociation enthalpies (ΔBDE) of
phenolic antioxidants from quantum topological molecular similar-
ity (QTMS). J Phys Chem A 110:6498–6503
88. Li MJ, Liu WX, Peng CR, Ren QH, Lu WC, Deng W (2013) A
DFT study on reaction of eupatilin with hydroxyl radical in solu-
tion. Int J Quantum Chem 113:966–974
68. Xue Y, Zheng Y, Zhang L, Wu W, Yu D, Liu Y (2013) Theoretical
study on the antioxidant properties of 2′-hydroxychalcones: H-atom
vs. electron transfer mechanism. J Mol Model 19:3851–3862
69. Farmanzadeh D, Najafi M (2013) On the antioxidant activity of the
tryptophan derivatives. Bull Chem Soc Jpn 86:1041–1050
70. Kumar J, Kumar N, Sati N, Hota PK (2020) Antioxidant properties
of ethenyl indole: DPPH assay and TDDFT studies. New J Chem
44:8960–8970
71. Kancheva VD, Saso L, Angelova SE, Foti MC, Slavova-Kasakova
A, Daquino C, Enchev V, Firuzi O, Nechev J (2012) Antiradical
and antioxidant activities of new bio-antioxidants. Biochimie. 94:
403–415
72. Nakanishi I, Shimada T, Ohkubo K, Manda S, Shimizu T, Urano S,
Okuda H, Miyata N, Ozawa T, Anzai K, Fukuzumi S, Ikota N,
Fukuhara K (2007) Involvement of electron transfer in the
radical-scavenging reaction of resveratrol. Chem Lett 36:1276–
1277
73. Mortensen A, Skibsted LH, Sampson J, Rice-Evans C, Everett SA
(1997) Comparative mechanisms and rates of free radical scaveng-
ing by carotenoid antioxidants. FEBS Lett 418:91–97
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