Experimental and Computational
methods for skin sensitization evaluation of isatin-hydrazone
Kamel Mokhnache*,
Ahlem Karbab, EL-Khamsa Soltani, Soraya Madoui, Hanane Khither, Noureddine Charef, Lekhmici Arrar
Laboratory of Applied Biochemistry, University Ferhat Abbas Setif 1, 19000, Algeria
*Corresponding Author E-mail: kamelmokhnache@yahoo.com
ABSTRACT:
A new isatin-hydrazone (IH) was selected
for theoretical and experimental studies. The HOMO and LUMO energies were investigated
using density functional theory (DFT) via B3LYP/6-31G (d,p), skin sensitization
prediction was carried out using Pred Skin software
program. The results demonstrate the reactivity of IH with Energy gap (Δ) of 0.0579 a.u,
low sensitizer effect towards human skin with probability of 60 %, and an
excellent topical anti-inflammatory effect against xylen-induced
ear odema in mice model with inhibition percentages
of 81.48%.
KEYWORDS: Hydrazone, skin sensitization, Topical,
Anti-inflammatory.
INTRODUCTION:
Indole or benzo [b]
pyrrole is a planar aromatic heterocyclic compound. It has a bicyclic
structure, consisting of a benzene ring fused to a pyrrole ring. It has, an
π-electronic system consisting of ten electrons from eight carbon atoms
and the non-binding doublet of the nitrogen atom [1]. A considerable number of
natural products are derived from this motif. In humans, serotonin or
5-hydroxytryptamine is a potent vasoconstrictor, mainly stored in blood
platelets, regulating gastric secretions and intestinal contractions [2]. This
substance also acts as a neurotransmitter, involved in the conduction of
impulses between nerve cells. In plants, heteroauxine
plays a role comparable to serotonin. 5,6-Dihydroxyindole is an essential
component of melanin, a brown pigment that stains the skin. Indigo is a
compound that is used as a colorant in the textile industry.
The indole ring is the most
heterocyclic unit in nature and one of the most attractive in the field of
organic chemistry concerning the development of new biologically active
structures and the discovery of new reactivities [3]. During the 1990s, the
discovery of a new class of drugs, the triptans, was a real advance in the
management of migraine attacks. For example, Zolmitriptan Zomig®
is marketed by Astrazeneca since 1998, Elitriptan (Zophren®), another
biologically active product with an indole ring, is marketed as part of the
suppression of nausea caused by chemotherapy or radiotherapy in the case of
anticancer treatments [4]. In this study, N'-[(E)-(5-bromo-1H-
indol-3-yl) methylidene] pyridine-4-carbohydrazide was selected for theoretical
and experimental studies. Molecular structure proprieties were investigated
using density functional theory (DFT) via B3LYP/6-31G (d,p), skin sensitization
prediction was carried out using Pred Skin software
program and was calculated using P value. In addition, topical
anti-inflammatory effect against xylen-induced
ear edema was investigated using mice model.
MATERIAL AND METHODS:
Quantum chemical calculation
Molecular structure
proprieties were investigated using density functional theory (DFT) via
B3LYP/6-31G (d,p), using the Gaussian 09 program [5],
where the following parameters: ionization potential (IP), electron affinities
(EA) [6], electronegativity (χ), chemical potential (μ) [7], global
hardness (η) [8], global softness (σ), and electrophilicity index
(ω) [9] were calculated by using the following equations [10]:
|
IP = – EHOMO |
(1) |
|
EA = –ELUMO |
(2) |
|
Ƞ = ELUMO – EHOMO |
(3) |
|
σ = 1 / Ƞ |
(4) |
|
χ = – (EHOMO – ELUMO) / 2 |
(5) |
|
µ = (EHOMO – ELUMO) / 2 |
(6) |
|
ω = µ2 / 2 Ƞ |
(7) |
Skin sensitization prediction
The online software program Pred Skin was used for skin sensitization prediction [11].
Skin sensitization
calculation:
Chemical-skin sensitizer
effect was calculated using the quantum chemical calculation according to the
following formulae: Predicted value (P) =15.3 x EHOMO + 5.08
When P is greater
than 0.50, the compound is predicted as a sensitizer, otherwise it is predicted
as a non-sensitizer [12]. Results compared with Ascorbic acid (AA) which
is skin whitening Agent [13] with no sensitizer effect.
Topical
anti-inflammatory effect:
Experimental
animals:
Experiments were
performed using mice, weighing 25-30g. They were obtained from Pasteur institut (Algeria) and housed in plastic cages under normal
laboratory conditions (12 h light/dark cycle, 23 ± 2°C) for an acclimatization
period of 7 days prior to the experiments. All the animals were given food and
water ad libitum.
Xylene-induced ear
edema in mice:
Adult albino female’s mice
(25-30g) were randomised into different groups of 6
mice each were used for the experiment. The investigated compounds (0.5mg/ear
edema), were topically applied to various groups. Inflammation was induced in
mice by topical application of 30μl of xylene and 30μl of different
synthetic product at the inner surface of the right ear. The thickness of the
ear is measured before and half hour after the induction of inflammation by a
digital caliper. The difference in thickness before and after the application
of xylene is calculated [14].
RESULTS AND DISCUSSION:
Quantum chemical
calculation
Geometry
optimization
Geometry optimization of
Isoniazid INH and its synthesized hydrazone analog IH was
performed in DFT/B3LYP/6-31G++(d,p)
level calculations. The optimized structure is shown in Fig. 1.
Fig. 1. Optimized structure of
IH
The analysis of the wave
function indicates that the electron absorption corresponds to the transition
between the ground state and the excited state, the electron donor distribution
in the busiest molecular orbital (HOMO) and the electron acceptor in the least
occupied molecular orbital (LUMO). Table 1 and Figure 2
illustrate the molecular energy, LUMO represents the ability to gain an
electron, so the HOMO represents an ability to lose an electron. The energy of
the HOMO is directly proportional to the ionization potential and the energy of
the LUMO is directly proportional to the electronic affinity.
The difference in orbital
energy between the HOMO and LUMO is called the HOMO-LUMO gap. The high HOMO
energy corresponds to a molecule more reactive with electrophiles in reactions,
low energy LUMO is reactive with nucleophiles.
According to the theory of
molecular orbitals, a high HOMO energy of one reagent molecule and a low LUMO
energy of another reagent are advantageous for the reaction between the two
molecules, because the electron transfers are easier from the HOMO of a LUMO
reagent on the other in the orbital interaction. The HOMO, LUMO and the energy
difference (HOMO-LUMO) of the monomer in the DFT with the 6-31G base (d, p)
were calculated. The HOMO-LUMO energy gap reveals that the difference in energy
reflects the chemical activity of the molecule.
Fig 2. Frontiers orbital
of the hydrazine
Table 1. Calculated global
scalar properties of IH
|
B3LYP/6-31G (d,p) |
||
|
Molecular energy (a.u) |
INH |
Hyd1 |
|
E(B3LYP) |
–472.2814 |
–3444.13 |
|
ELUMO |
–0.1045 |
–0.1228 |
|
EHOMO |
–0.2174 |
–0.1804 |
|
Energy gap (Δ) |
0.1129 |
0.0579 |
|
Ionization potential (I) |
0.2174 |
0.1804 |
|
Electron affinity (A) |
0.1045 |
0.1228 |
|
Global hardness (η) |
0.05645 |
0.0288 |
|
Global softness (S) |
0.02822 |
0.0144 |
|
Chemical potential (µ) |
–0.3219 |
–0.1516 |
|
Electophilicity (ω) |
0.9177 |
0.3975 |
|
Electronegativity (χ) |
0.1609 |
0.1516 |
Skin sensitization
calculation:
In first time, skin
sensitization was predicted by P value calculation using EHOMO
(Hartree), results shown in Table 2 show sensitizer effect of IH comparable
with INH and sensitizer effect comparable with AA with P values
of 23198, 1.75378 and 0.4301 respectively.
Table 2. P value and predicted
sensitization effect of INH, Hyd and AA
|
Compound |
EHOMO |
P values |
Skin sensitization |
|
INH |
-0.2174 |
1.75378 |
Sensitizer |
|
IH |
-0.1804 |
2.3198 |
Sensitizer |
|
AA |
-8.27 |
0.4301 |
Non-Sensitizer |
Skin sensitization prediction:
Results displayed
in Figures 3, and 4 explain the reasons of precedent results in Table
2, which indicate the atoms or the fragments contributed in the sensitizing
effect, green atom or fragment represent an increase in skin sensitization
potential; whereas, pink fragments represent a decrease in skin sensitization
potential, and gray fragments do not contribute to skin sensitization
potential.
Fig. 3. Effect of IH on a.
Human skin sensitization, b. Murine local lymph node assay (LLNA), c. Direct
peptide reactivity assay (DPRA), d. Human cell line activation test (h-CLAT),
e. KeratinoSensTM.
Topical anti-inflammatory:
Inflammation is a complex
biological response to harmful stimuli mainly mediated by two enzymes:
cyclooxygenase and lipoxygenase that generate prostaglandins and leukotrienes
respectively [15]. The clinical signs of this process are: The clinical signs of
this process are heat, redness, swelling and pain, and impaired functioning of
the affected organ may occur. At the tissue level, the inflammatory response is
characterized by increased vascular permeability, increased protein
denaturation and cell membrane alteration [16]. Topical anti-inflammatory
effect against xylen-induced ear edema was
investigated using mice model. The ability of IH to inhibit inflammation
induced by xylene was estimated. The obtained results show that IH has
an inhibitory effect against topical inflammation. This effect is significantly
(p≤0.001) lower than that of the positive control indomethacin (IND)
which expresses a percentage inhibition of 87.65 ± 0.13%. However, the
inhibition percentages obtained with IH is 81.48 ± 0.32%. Whereas, INH
exhibited similar effect to IND with inhibition percentage of 87.65±1.45
% (Figure 5).
Figure 5. Topical
anti-inflammatory effect of IH, INH and IND
CONCLUSION:
A new hydrazone IH,
derived from the isoniazid (anti-tubercular drug) was investigated for their
skin sensitization and anti-inflammatory effects. The predicted results
indicate the sensitizer effect of this hydrazone. On the other hand, results
revealed an excellent topical anti-inflammatory effect of IH.
REFERENCES:
1.
Sundberg,
R. J. The Chemistry of Indoles, Academic Press New York - London, 1970
2.
Rzeszotarska, B. 5-Hydroxytryptamine and Related Indolealkylamines, Springer-Verlag Berlin Heidelberg 1966,
19.
3.
Bartoli,
G.; Bencivenni, G.; Dalpozzo,
R. Chem. Soc. Rev. 2010, 39, 4449-4465.
4.
Generali,
J.A.; Cada, D.J. Hospital Pharmacy 2009, 44, 670-671
5. M.J. Frisch, G.W.
Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R.
Cheeseman, et al., Gaussian 09, Revision A.02, Gaussian, Inc., Wallingford CT,
2009.
6. J.B. Foresman, A.
Frisch, Exploring Chemistry with Electronic Structure Methods, Gaussian,
Pittsburg, Pa, USA, 1995.
7. L. Pauling, The
Nature of the Chemical Bond, Cornell University Press, Ithaca, NY, USA, 1960.
8. P. Senet, Chemical hardnesses of atoms and molecules from frontier orbitals, Chemical Physics Letters. 275
(1997) 527–532.
9.
R.G.
Parr, R.G. Pearson, Absolute hardness: companion parameter to absolute
electronegativity, Journal
of the American chemical society. 105 (1983) 7512–7516.
10.
Z. Bouanane, M. Bounekhel, M. Elkolli, F. Abrigach, M. Khoutoul, R. Bouyala, R. Touzani, A. Hellal, Synthesis,
structural, catecholase, tyrosinase and DFT studies
of pyrazoloquinoxaline derivatives, Journal of Molecular Structure.
1139 (2017) 238–246.
11.
R.C.
Braga, V.M. Alves, E.N. Muratov, J. Strickland, N. Kleinstreuer, A. Trospsha, C.H.
Andrade. Pred-Skin: A Fast and Reliable Web
Application to Assess Skin Sensitization Effect of Chemicals. Journal of Chemical Information and
Modeling. 57
(2017) 1013–1017.
12. Q. Ouyang, L. Wang,
Y. Mu, X.Q. Xie, Modeling skin sensitization
potential of mechanistically hard-to-be-classified aniline and phenol compounds
with quantum mechanistic properties, BMC Pharmacol. Toxicol. 76 (2014) 1-9.
13. C. Couteau, L. Coiffard, Overview of skin whitening agents: drugs and
cosmetic products, Cosmetics 27 (2016) 1-16.
14.
Kamel Mokhnache, Noureddine Charef, Soraya Madoui, Mohammad
S. Mubarak. Drug Classification and Acute Rodent Toxicity Predictions of
Bis-Phenolic Ligand: As A Topical Anti-Inflammatory and Antifungal Agent. GSJ.
6 (2018) 636-647
15. Nagi MN, Mansour MA
(2000). Protective Effect of Thymoquinone against Doxorubicininduced
Cardiotoxicity in Rats: A Possible Mechanism of Protection. Pharmacological
Research. 41: 283–289.
16. Dorward D.A., Lucas C.D.,
Rossi A.G., Haslett C. and Dhaliwal K. (2012). Imaging inflammation: mole-cular strategies to visualize key components of the
inflammatory cascade, from initiation to resolution. Pharmacology and
Therapeutics. 135(2), 182-199.
Received on 09.10.2019
Accepted on 30.11.2019
Accepted on 31.12.2019 ©A&V
Publications all right reserved
Research J. Topical
and Cosmetic Sci. 2020; 11(1):15-19.
DOI: 10.5958/2321-5844.2020.00004.7