Antimycobacterial, antimicrobial activity, experimental (FT-IR, FT-Raman, NMR, UV-Vis, DSC) and DFT (transition state, chemical reactivity, NBO, NLO) studies on pyrrole-isonicotinyl hydrazone

Accepted Manuscript Antimycobacterial, antimicrobial activity, experimental (FT-IR, FT-Raman, NMR, UV–Vis, DSC) and DFT (transition state, chemical reactivity, NBO, NLO) studies on pyrrole-isonicotinyl hydrazone

Poonam Rawat, R.N. Singh, Alok Ranjan, Sartaj Ahmad, Rajat Saxena PII: DOI: Reference:

S1386-1425(17)30109-9 doi: 10.1016/j.saa.2017.02.021 SAA 14943

To appear in:

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy

Received date: Revised date: Accepted date:

31 October 2016 5 February 2017 9 February 2017

Please cite this article as: Poonam Rawat, R.N. Singh, Alok Ranjan, Sartaj Ahmad, Rajat Saxena , Antimycobacterial, antimicrobial activity, experimental (FT-IR, FT-Raman, NMR, UV–Vis, DSC) and DFT (transition state, chemical reactivity, NBO, NLO) studies on pyrrole-isonicotinyl hydrazone. The address for the corresponding author was captured as affiliation for all authors. Please check if appropriate. Saa(2017), doi: 10.1016/ j.saa.2017.02.021

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ACCEPTED MANUSCRIPT Antimycobacterial, Antimicrobial activity, Experimental (FT-IR, FT-Raman, NMR, UV-Vis, DSC) and DFT (Transition State, Chemical Reactivity, NBO, NLO) Studies on Pyrrole-isonicotinyl hydrazone Poonam Rawat, R.N. Singh*, Alok Ranjan, Sartaj Ahmad and Rajat Saxena

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Department of Chemistry, University of Lucknow, Lucknow 226007, U.P., India

Abstract

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As part of a study of pyrrole hydrazone, we have investigated quantum chemical calculations, molecular geometry, relative energy, vibrational properties and antimycobacterial / antimicrobial

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activity of Pyrrole-2-carboxaldehyde isonicotinyl hydrazone (PCINH), by applying the density

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functional theory (DFT) and Hartree fock (HF). Good reproduction of experimental values is

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obtained and with small percentage error in majority of the cases in comparison to theoretical result (DFT). The experimental FT-IR and Raman wavenumbers were compared with the

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respective theoretical values obtained from DFT calculations and found to agree well. In crystal structure studies the hydrated PCINH (syn-syn conformer) shows different conformation than

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from anhydrous form (syn-anti conformer). The rotational barrier between syn-syn and syn-anti

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HNMR,

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conformers of PCINH is 12.7 kcal/mol in the gas phase. In this work, use of FT-IR, FT-Raman, 13

CNMR and UV-Vis spectroscopies have been made for full characterization of

PCINH. A detailed interpretation of the vibrational spectrum was carried out with the aid of normal coordinate analysis using single scaling factor. Our results support the hydrogen bonding pattern proposed by reported crystalline structure. The calculated nature of electronic transitions within molecule found to be π→π*. The electronic descriptors study indicates that PCINH can be used as robust synthon for synthesis of new heterocyclic compounds. The first static

ACCEPTED MANUSCRIPT hyperpolarizability (β0) of PCINH is calculated as 33.89 x 10–30 esu, (gas phase); 68.79 x 10–30 (CHCl3), esu; 76.76 x 10–30 esu (CH2Cl2), 85.16 x 10–30 esu (DMSO). The solvent induced effects on the first static hyperpolarizability were studied and found to increase as dielectric constants of the solvents increases. Investigated molecule shows better NLO value than Para nitroaniline

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(PNA). The compound PCINH shows good antifungal and antibacterial activity against

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Aspergillus Niger and gram-positive bacteria Bacillus subtilis, respectively. The compound also

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shows good antituberculosis activity against Mycobacterium tuberculosis H37Rv using the microplate alamar blue assay (MABA).

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Keywords: Solvent effect, NMR spectroscopy, TD-DFT calculations, Vibrational analysis,

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Reactivity descriptors.

* Corresponding author. e-mail: [email protected]

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Tel.: +91 9451308205

ACCEPTED MANUSCRIPT 1. Introduction Hydrazide-hydrazones have technical, commercial [1] and synthetic applications in their own right due to their physiological activity. They have been used in the field of drugs, photothermochemic compounds and precursors for organic synthesis [2] especially in indicators-

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chemistry [3]. Few pyrrole hydrazones were synthesized and showed to possess potential

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antituberculosis activity [4-6]. They also act as herbicides, insecticides, nematocides,

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rodenticides, plant growth regulators, sterilants for housefly and iron overload diseases [7]. In analytical chemistry hydrazones find applications as multi-dentate ligands for transition metals in

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colorimetric or fluorimetric determinations [8] and also used for the identification and isolation

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of carbonyl compounds. Their metal complexes have used for device applications such as telecommunications, optical computing, optical storage and optical information processing [9].

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The crystal structure of Pyrrole-2-carboxaldehyde isonicotinyl hydrazone and its hydrate was

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reported by Safoklov et.al. [10] based on single crystal X-ray diffraction data collected at ambient temperature. Christopher Glidewelld [11] re-determined the structure of Pyrrole-2-

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carboxaldehyde isonicotinyl hydrazone. The anhydrous crystal structure of PCINH is different

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from its hydrate structure. Gunay, [12] reported molecular structure and chemical shift assignment of pyrrole-2-carboxaldehyde isonicotinyl hydrazone monohydrate by DFT and HF on

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6-31g(d) basis set.

With increasing demand for number of pyrrole based compounds, in pharmaceutical, drugs, analytical and coordination chemistry attempts are being made in directions of simplifying the synthetic strategies of robust synthons as well as detailing out their spectral analysis, structural elucidation and chemical reactivity. Hydrogen bonds are of versatile importance in the fields of chemistry and bio-chemistry, which governs solvation, atmospheric chemistry, molecular

ACCEPTED MANUSCRIPT assemblies, molecular recognition, organic synthesis, supramolecular structures, biochemical phenomena and life processes [13]. Pyrrole and isonicotinyl hydrazide are two major heterocycles, whose compounds possess wide range of biological and material applications. Literature survey reveals that detail quantum chemical calculations and their correlation with

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experimental properties of Pyrrole-2-carboxaldehyde isonicotinyl hydrazone (PCINH) have not

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been reported yet. Therefore, our goal in this work has been to detail out the spectroscopies: FT-

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IR spectrum, FT-Raman; thermochemistry, HOMO-LUMO, NBO, molecular electrostatic potential surface, chemical reactivity, biological activity and NLO of PCINH and hope that the

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results of the present study would be helpful in future spectral analysis, biological activity and

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NLO as well as likely to contribute to the advancement of knowledge in this area. 2. Computational details

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All calculations were performed with the Gaussian 09 program package [14]. All calculations

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were carried out at the density functional level of theory employing the B3LYP functional and Hartree Fock (HF) theory with 6-311++G(d,p) basis set. The FT-IR and Raman spectra were

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calculated at the same level of theory and single scaling factor was applied to the calculated

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frequencies. Raman and IR spectra were simulated using line shape of Lorentzian curves type and the FWHM (full width at half maximum) of each peak was 18 cm-1. The normal mode

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analysis was performed with the help of calculated PED for each internal coordinates using localized symmetry defined using Pulay’s recommendations [15]. 1H and

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CNMR chemical

shifts of (PCINH) were calculated at the same level by employing the Gauge-Included Atomic Orbital (GIAO) [16] with PCM model and DMSO as solvent. A TD-DFT calculation was carried out to obtain the theoretical excitation/absorbance spectrum. The reaction energies ∆E°, standard enthalpies ∆H° and Gibbs free energies ∆G° of rotational conversions between the PCINH

ACCEPTED MANUSCRIPT conformers were derived from frequency calculations at the B3LYP/6-31G(d,p) level of theory. The reaction entropies (∆S°) of rotational conversions were evaluated using a thermodynamic equation ∆S° = (∆H°-∆G°)/T. The rate constant k(T) derived from transition state theory was computed from the Gibbs free energy of activation ∆‡G°, using k(T) = (kBT/hc°)exp(-∆‡G°/RT)

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where the factor cº is assigned to unity [17]. The equilibrium constant K at 298.15 K and 1 atm is

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computed using the equation ∆G°= - RT lnK. The Mullikan and Natural population analysis was

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performed at the UB3LYP/6-31G(d,p) level of theory. The global and local indexes (philicity, hardness, softness, chemical potential, electronegativity and condense Fukui functions etc.) of

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reactivity were calculated with Koopman´s approximation [18-21]. DSC thermogram of the

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powder sample of PCINH was recorded on Mettler Toledo DSC system model DSC 1:1 stare system with heating rate of 10/15oC/min, nitrogen flow at the rate of 30 ml/min and crucible was

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used of 40 μl Al.

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2.1 Experimental Details

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2.1.1 Preparation of Pyrrole-2-carboxaldehyde isonicotinyl hydrazone and General procedures A mixture of 2-Formyl-1H-pyrrole (1) (0.100g, 0.5122 mmol), Isoniazid (2) (0.1014g,

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0.5122 mmol) in 15 ml methanol was refluxed for 15 minutes. A shiny lemon colored precipitate

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was obtained on cooling. The precipitate was filtered off, washed with methanol and dried in air. Yield 76.74%, m.p-decomposed above 210oC. The 1H NMR and

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C NMR spectra were recorded in DMSO-d6 on Bruker DRX-300

spectrometer. The FT-IR-spectrum was recorded in KBr medium on a Bruker-spectrometer by averaging 64 scans with a resolution of 4 cm-1. The DART-Mass spectrum was recorded on JMS-T100LC, Accu TOF spectrometer. The UV-Vis absorption spectrum was recorded on

ACCEPTED MANUSCRIPT ELICO SL-164 spectrophotometer equipped with a 10-nm quartz cell. The measurement is done for a 10-5 M chloroform solution at 25ºC in the range 200-800 nm. 2.2.2 Biological Screening 2.2.2.1 Antimicrobial Screening

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The synthesized compounds were evaluated for their in vitro antifungal and antibacterial activity

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using Aspergillus Niger, Bacillus subtilis strain, respectively, in potato dextrose agar media

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(PDA). The antibacterial and antifungal activity was performed by Agar diffusion method at the concentration level of 200μg/ml and 100μg/ml. The solution (2.5 ml) of PCINH and Isoniazid

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(different concentrations) and 50 ml of molten sterile nutrient PDA media (5%) was poured into

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sterilized glass petridishes. A control PDA plate was also prepared for the test to compare the effect of test samples and to nullify the effect of solvent. All these plates were kept for 24 hours

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in UV chamber so that the samples had sufficient time to diffuse over a considerable area of the

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plate. After solidification of the medium, these plates were freshly seeded with small portion of the mycelium of fungus in form of 0.5 mm discs, carefully over the center of each PDA plate

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with the help of sterilized needle. The plates were kept for incubation at 25±1°C for 96 hours.

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Chloroform was used as solvent to prepare desired solution of PCINH and Isoniazid initially and also to maintain proper control. Inhibitory activity was measured (in mm) as the diameter of the

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observed inhibition zones.

2.2.2.2. Antimycobacterial Activity The PCINH was screened against Mycobacterium tuberculosis H37Rv using the microplate alamar blue assay (MABA). Primary screening was conducted at a single concentration of 10 µg/mL against Mycobacterium tuberculosis H37Rv, in BACTEC 12B medium using the Microplate Alamar Blue Assay (MABA) [22]. Compound first demonstrating at least 90%

ACCEPTED MANUSCRIPT inhibition in the primary screening (IC90 ≤ 10 µg/mL) was retested at lower concentrations by serial dilution against Mycobacterium tuberculosis H37Rv to determine the actual MIC, using the MABA method. The VERO cell cytotoxicity assay (50% inhibitory concentration IC50) was done in parallel with TB dose response assay to determine the selectivity index (SI), defined as

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the ratio of the measured IC50 (mammalian cell toxicity) to the IC90 (H37Rv) Mycobacterium

Results and discussion

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3.

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tuberculosis.

3.1 Molecular structure, energies and Topological analysis

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Initial geometry was taken from the single crystal X-ray diffraction data [10] and further

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geometry of the compound was optimized. The ground state optimized structure of the molecule is presented in Figure 1. The optimized and experimental single crystal X-ray diffraction

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structures of PCINH were compared by superimposing them using a least squares algorithm

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that minimizes the distances of the corresponding non hydrogen atoms (Figure 2). The agreement

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between the optimized and experimental crystal structure is quite good showing that the geometry optimization reproduces almost exact to experimental conformation. The PCINH

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crystallize in two forms hydrated (a) and anhydrous (b). Both forms have been crystallized in

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two different conformations. The form (a) crystallizes with water molecule in syn-syn conformation (Supplementary Figure S1) whereas forms (b) in syn-anti-conformation. The carbonyl group oxygen atom O(1) and azomethine framework nitrogen atoms N(5), and N(4) are in syn-syn positions due to the association with the water molecule whereas in anhydrous form (b) these atoms are in the syn-anti positions. The bond lengths of the azomethine framework of hydrate crystal (a) are identical with the anhydrous form (b) except N-N bond length which is shorter in (a) than (b) due to the association with the water molecule. In both forms the

ACCEPTED MANUSCRIPT endocyclic angles of the pyrrole ring are added up to exactly 540°, indicating that the heterocyclic pyrrole ring is perfectly planar. The relative energies of the molecule are calculated employing DFT functional (B3LYP). The energies for syn-syn and syn-anti-conformation for the optimized geometries of PCINH are -719.40259743 and -719.39507696 a.u. respectively. They

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have energy difference of 4.72 kcal/mol and in equilibrium syn-syn and syn-anti forms are

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supposed to be in ratio 99.97% and 0.03%, respectively.

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The Molecular structures of both forms possess C1 point group symmetry. The optimized structural parameters (bond lengths, bond angles, dihedral angle) of syn-syn and syn-anti-

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conformations for PCINH have been compared with those obtained experimentally from the

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single crystal X-ray diffraction data as shown in Supplementary Table S1. It has been seen that the DFT method give comparable geometries, which differ from each other by not more than

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0.037Å in bond length and 2.2º in bond angles. The elongated C-H bond is observed in DFT

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calculations. This is due to the fact that in X-ray crystal structures measurements positions of hydrogen atoms are not precise because they correspond to the electron density maxima of

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atoms. There are no differences between nuclei and maxima positions for heavier atoms, but

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there are differences for hydrogen atoms. The effect is that the length of the bond with the hydrogen atom determined from X-ray diffraction measurements is usually shorter than the

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distance between nuclei. Hence, the correction according to neutron diffraction results is often applied. The Bader’s theory of Atoms in molecules is useful tool to characterize the strength of hydrogen bond. QTAIM study the concept of chemical bond and bond strength in terms of topological features, electron density distribution function and energetic parameters. The nature and strength of weaker interactions are given in supplementary Table S2. The bond strength is determined by accumulation of electron density in the bond region and the screening of nuclei by

ACCEPTED MANUSCRIPT this density. The energy of syn-syn conformer further lowers in case of its hydration. The crystal structure of hydrated PCINH, pyrrolic NH interact with oxygen atom of water and >C=N and >C=O bond are hydrogen bonded in bifurcation with hydrogen atom of water. This kind of arrangement results due to packing forces. The hydrogen bonding interaction energy decreases

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on deviation from linearity in hydrogen bonded atoms. The molecular graph of PCINH.H2O is

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given in Supplementary Figure S2. Three types of intermolecular interaction are seen in the

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molecular graph. The intermolecular N5•••H28-O27 interaction is weaker than N4-H22•••O27 interaction. The hydrogen bond energy values of C11=O1•••H28-O27 and N4-H22•••O27 bond

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are found to be 5.26 and 10.86 kcal/mol, respectively. The N4-H22•••O27 bond is strongly

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bonded through intermolecular hydrogen bonding with oxygen atom of water. 3.2 Rotational energy barrier and reaction energies of PCINH

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In order to gain insight into the nature of the energetic of the rotational equilibrium

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between the two almost planer structure of PCINH we have performed computational calculations on both minima and the rotational transition state at same level of theory. Energy

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profiles for conversion of conformers are shown in Figure 3. Activation energies and

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thermodynamics quantities of PCINH are listed in Supplementary Table S3 and S4. The transition structure TS1 between the syn-syn and syn-anti forms was confirmed by imaginary

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frequencies at -129.21 cm-1. The rotational barrier between syn-syn and syn-anti forms is 12.70 kcal/mol in the gas phase. This rotational barrier is larger than the energy difference and thus the free rotation is quite a difficult task at room temperature. Forward rate coefficient of equilibrium conversion evaluated from Gibbs free energies of activation according to TS1 expressed as log kf is -19.2. The value of equilibrium constant between syn-syn and syn-anti forms is 1259.8. The value of equilibrium constant (k) is greater than 1 this means that the conversion would only be

ACCEPTED MANUSCRIPT possible when energy is provided to the syn-syn conformer. The enthalpy change (∆HReaction), Gibbs free energy change (∆GReaction) and entropy change (∆SReaction), of condensation reaction between 2-Formyl-1H-pyrrole and Isoniazid are also carried out at B3LYP/6-311++G(d,p) basis set. The calculated thermodynamic parameters of PCINH are given in Supplementary Table S4.

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The negative values of ∆GReaction and ∆HReaction show that the reaction is exothermic and

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spontaneous at room temperature.

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3.3 Thermal Analysis

The differential scanning calorimetry (DSC) study was made to check the purity of the

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sample as shown in Figure 4. On heating the PCINH sample in N2 atmosphere, in the range 30-

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250oC it produced a strong endothermic maximum at 211.01ºC and ΔHf = -150.78 J/g, without any glass transition and first cooling curve does not produce any crystallization peak. PCINH

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completely decompose at 220ºC as shown in the Figure 4. We have done the second heating till

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higher temperature more than the first heating to check the exact decomposition temperature. The absence of any glass transition and crystallization clearly demonstrates that the PCINH is

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thermally stable upto 180ºC confirming the purity of the PCINH sample.

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3.4 1H NMR Spectroscopy

The chemical shifts are calculated with GIAO approach using B3LYP method and 6-

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311++G(d,p) basis set. The experimental and calculated values of 1H and 13C chemical shifts of the title compound are given in Supplementary Table S5 and correlation graph of chemical shifts are shown in Supplementary Figure S3. Taking into account that the range of 13CNMR chemical shift of an analogous molecule is usually greater than 100ppm, this accuracy ensures the reliable interpretation of spectroscopic parameters. The carbon NMR spectrum shows only 9 peaks with different intensities, while 12 are present in the molecular formula. This suggests the presence of

ACCEPTED MANUSCRIPT symmetry which makes some of the carbon atom equivalent. Nine carbon peaks in the molecule are observed from 163.68 to 119. 83 ppm is calculated from 162.84 to 119.02 ppm. The calculated 1HNMR chemical shift show good agreement with the experimental values. 3.5 UV-Vis spectroscopy and Natural Bonding Orbital (NBO) analysis

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Absorption spectroscopy of organic compounds are based on transitions of n or π

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electrons to the π* excited state which takes place in the range 200–800 nm. The UV-Vis

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spectrum of syn-syn conformer has been recorded in chloroform. The UV-Vis spectrum has been calculated in gas and solvent phase at B3LYP functional and 6-311++G(d,p) basis set. The

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calculated solvent phase spectrum show good corroboration with experimental spectrum.

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Molecular orbitals are very useful in qualitative descriptions of bonding, biological activity and reactivity. So we calculated the highest occupied molecular orbital energy (HOMO), the lowest

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unoccupied molecular orbital energy (LUMO), and energy difference (H-L) between the frontier

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molecular orbitals (HOMO–LUMO) of the most stable syn-syn conformer. The observed and calculated electronic transitions of high oscillatory strength are listed in Supplementary Table S6

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and shown in Figure 5. The first electronic absorption corresponds to the transition from the

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ground to the first excited state and is mainly described by one electron excitation from the highest occupied molecular orbital (HOMO) to the lowest unoccupied molecular orbital

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(LUMO) with strong oscillatory strength of 0.3871 which is π-π* in nature. The HOMO is located over the pyrrole and hydrazone framework, the HOMO→LUMO transition implies an electron density transfer to more over pyridine ring. These atomic orbital compositions of the frontier molecular orbitals are sketch in Supplementary Figure S4. The second excitation originates from HOMO to LUMO+2 with oscillatory strength of 0.1158 which is ℼ-π* in nature. In HOMO→LUMO+2 transitions, electron density transfer to pyridine ring from HOMO orbital.

ACCEPTED MANUSCRIPT The calculated HOMO and LUMO energy gap explains the eventual charge transfer interactions taking place within the molecule. The max absorption peak observed at 352 nm can be assigned to π→π* transition and may be attributed to the excitation in the aromatic ring and C=O group. The Figure 5 also shows that the compound is transparent in the entire visible region and the

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absorption takes place in the UV range which is the key factor for studied molecule for NLO

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applications in the room temperature.

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The NBO analysis can be used to estimate delocalization of electron density between occupied Lewis-type orbitals and formally unoccupied non-Lewis NBOs, which corresponds to a

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stabilizing donor–acceptor interaction. Second order perturbation theory allows making

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conclusions about the strength of these interactions. The increased electron density at NH atoms leads to the elongation of N-H bond length and lowering of the N-H wavenumber. The electron

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density is transferred from n to antibonding σ* orbital of the C-C bond, explaining both the

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elongation and red shift. The C=O stretching mode can be used as good probe for evaluating the bonding configuration around the carbonyl oxygen atom and the electronic distribution between

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of the pyridine. The calculated second-order interaction energies (E2) using the NBO analyses

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have shown that the lone pairs localized on the N atoms donate the charge to carbon and hydrogen, and second order interaction energies are 23.10 and 21.36 kcal/mol. For donation of

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electron density from the lone pairs orbitals of the oxygen atoms to unoccupied carbon atom orbitals the second-order energies are equal to 36.41 and 35.87 kcal/mol. 3.6 Vibrational assignments In the study of the vibrational behavior of PCINH, infrared and Raman spectroscopy seem to be good method to decide whether PCINH forms dimer through H-bonding. Comparison of the wavenumbers calculated at B3LYP with experimental values is given in Supplementary

ACCEPTED MANUSCRIPT Table S7, reveals the over estimation of the calculated vibrational modes due to neglect of anharmonicity in real system. Ab initio harmonic vibrational frequencies (ω) are typically larger than the fundamentals (ν) observed experimentally. A major source of this disagreement is the neglect of anharmonicity effects in the theoretical treatment. Errors also arise because of

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incomplete incorporation of electron correlation and the use of finite basis sets. Thus, DFT and

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Hartree-Fock (HF) theory tend to overestimate vibrational frequencies because of improper

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dissociation behavior, a shortcoming that can be partially compensated for by the explicit inclusion of electron correlation. The overestimation of ab initio harmonic vibrational

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frequencies is, however, found to be relatively uniform, and as a result generic frequency scaling

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factors are often applied to obtain good overall agreement between the scaled theoretical harmonic frequencies and the anharmonic experimental frequencies. Experimental and calculated

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scaled infrared absorbance spectrum is shown in Figure 6 in the region 4000-400 cm-1 .

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The molecular conformation obtained from geometry optimization, exhibits no special symmetry and hence the molecule belongs to C1 point group. As a consequence, all the

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fundamental vibrations of free molecule are both IR and Raman active. DFT calculations yield

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Raman scattering amplitudes which cannot be taken directly to be the Raman intensities. The Raman scattering cross section, ∂σ/∂Ω, which are proportional to Raman intensity may be

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calculated from Raman scattering amplitude and predicted wavenumbers for each normal mode using the relationship [23, 24]

    4  j  2 4  4   0   j   h       45    hc j   8 2 c j  1  exp  kT     

 S j  

ACCEPTED MANUSCRIPT Where Sj and υj are the scattering activities and the predicted wavenumbers, respectively of the jth normal mode, υ0 is the wavenumber of Raman excitation line h, c and k are universal constants. The Raman intensities obtained using this relationship match quite nicely with the experimentally observed intensities. The simulated and experimental Raman spectrum for

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PCINH is shown in Figure 7.

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In the FT-IR spectrum of PCINH, pyrrole N–H stretching vibration (νNH) is observed at

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3286 cm-1, whereas it is calculated as 3319 cm-1. The observed νN–H at 3286 cm-1 is also in agreement with the reported strong absorption band at 3148 cm-1 for 2-formyl pyrrole [25]. The

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C–C stretching vibration in pyrrole ring is observed at 1535 cm-1 and is in good agreement with

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the calculated wavenumber at 1548 cm-1. The C=N stretching vibration is observed at 1547 cm-1, whereas it is calculated as 1622 cm-1. The observed stretching vibration of carbonyl group (νC=O)

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at 1662 cm-1 correlates well with the calculated wavenumber at 1690 cm -1. In FT-IR spectrum,

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the presence of -C=N band confirms the formation of hydrazone linkage. The νC=O stretching wavenumber observed at 1718 cm-1 in experimental FT-IR spectrum. The combination bands of

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the C–N deformations vibrations are observed at 1222, 1216 cm-1, whereas they are calculated as

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1228, 1220 cm-1, respectively. The C–H stretches in pyrrole and pyridine ring are observed in the range 3137-2938 cm-1 and correlates well with observed data as shown in Supplementary Table

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S7. The calculated N–N stretching vibration at 1134 cm-1 with 19% contribution in PED agrees well with observed wavenumber at 1100 cm-1. The observed pyrrole ring deformations (δR) at 831 and 668 cm-1 correspond to the calculated wavenumber at 843 and 665 cm-1 respectively.

ACCEPTED MANUSCRIPT 3.7 Chemical reactivity 3.7.1 Molecular electrostatic potential (MEP) and global and local reactivity descriptors Electrostatic potential correlates with dipole moment, electronegativity, partial charges and site of chemical reactivity of the molecule. A MEP surface for PCINH is given in Figure 8. The

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red colour around carbonyl group shows negative electrostatic potential and blue colour around

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the pyrrolic and amide NH shows most positive electrostatic potential, whereas the rest of the

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region of molecule is shown by green colour having moderate electrostatic potential. MEP shows two proton donor centers, viz., the amide NH group (NHamide) and the NH group of the pyrrole

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ring (NHpy). The NHamide group of hydrazone frame work linked with O=C group and NH

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group of pyrrole ring linked with nitrogen atom of pyridine ring through intermolecular hydrogen bonding.

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DFT [26] is computationally inexpensive quantum chemical tool that has been emerging

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nowadays for studying various chemical problems. The principle behind the DFT [26] is that the energy of a molecule can be determined from the electron density instead of a wavefunction. By

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using DFT it is possible to define and justify concepts of chemical reactivity. In this work an

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attempt has been made to explore the uses of DFT descriptors to investigate the reactive sites of 2-Formyl-1H-pyrrole, Isoniazid as well that of PCINH. Electronegativity (χ), chemical potential

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(μ), global hardness (η), global softness (S) electrophilicity index (ω) and polarizability are global reactivity descriptors, have been calculated on the basis of Koopman’s theorem [27] using the energies of frontier molecular orbitals εHOMO, εLUMO and given by equations (1-5) [28-34]. χ = -1/2(εLUMO +εHOMO)

(1)

μ = - χ = 1/2 (εLUMO +εHOMO)

(2)

η = 1/2 (εLUMO - εHOMO)

(3)

ACCEPTED MANUSCRIPT S = 1/2 η

(4)

ω = μ2/2η

(5)

All computed global descriptors are given in Table 1. The electrophilicity value of 2-Formyl-1Hpyrrole is higher than Isoniazid. Therefore, the 2-Formyl-1H-pyrrole behaves as electrophile and

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ECT = (∆Nmax)A - (∆Nmax)B

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interacting molecules has been calculated using equation (6)

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Isoniazid as nucleophile. Electrophilic charge transfer (ECT) [34] between the ∆Nmax values of

where, (∆Nmax)A = μA / ηA

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(∆Nmax)B = μB / ηB

(6)

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If (i) ECT > 0, charge flow from reactant I to another II (ii) ECT < 0, charge flow from II to I. ECT is calculated as 0.092 for reactant molecules 1 and 2, which indicates that charge flows

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from 2 to 1. Therefore, 1 acts as electron acceptor and 2 as electron donor. The specific site

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selectivity of the molecules has been calculated with the aid of local (condensed) Fukui function. The Fukui functions (fk+, fk-) are calculated using the equations (7-9). for nucleophilic attack

(7)

fk- = [q(N) – q(N-1)]

for electrophilic attack

(8)

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fk+ = [q(N+1) – q(N)]

where N, N-1, N+1 are total electrons present in neutral, cation and anion state of molecule

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respectively. The fk+ and fk- values for both reactant and products are listed in Supplementary Table S8. In 2-Formyl-1H-pyrrole fk+ maximum values at carbon C(6) of carbonyl group indicate that this site is prone to nucleophilic attack. The maximum value of fk- for Isoniozid, at N(9) of NH2 group indicate that this atom is more prone to electrophilc attack. Therefore, local reactivity descriptors for reactants 1 and 2 confirm the formation of product PCINH by nucleophilic attack of N(9) at C(6) site of reactant 1. The product PCINH undergoes both electrophilic and

ACCEPTED MANUSCRIPT nucleophilc attack. The maximum values of (fk+) at C(16) indicate that this site is more prone to nucleophilic attack. Therefore, local reactivity descriptors of PCINH favor the formation of new heterocyclic compounds such as thiadiazoline, thiazolidinones and azetidinones etc. by attack of nucleophilic part of the dipolar reagent on the C16 site of C16=N5 bond.

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3.8 Static dipole moment (μ0), mean polarizability (|α0|), anisotropy of polarizability (∆α) and

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first hyperpolarizability (β0)

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DFT based methods can be of great aid and adequate in searching for new nonlinear chemical compounds. Theoretical methods have been considered as useful techniques for

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prediction of polarizabilities and hyperpolarizabilities avoiding an expensive large amount of

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experimental synthetic work that precedes the measuring of NLO properties, what may not lead to a desired compound for practical applications. Computational calculation as an alternate

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choice provides extensive properties of materials, eg. hyperpolarizability which is difficult task

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to measure directly [35]. Polarizabilities and hyperpolarizabilities are described to response of a system in the presence of an applied electric field and determine the strength of molecular

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interactions, cross sections of different scattering and collision processes, as well as the non–

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linear optical (NLO) properties of the system [36]. DFT methods are the best in terms of accuracy and computational time [36]. In order to gain insight into the non-linear optical (NLO)

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property of the molecule, the first static hyperpolarizability (β) was calculated with double numerical differentiation of energies that is by the finite field perturbation method in vacuum as well as incorporating the solvent factors with increasing polarity and their calculated values are given in Table 2. It is noted from the Table 2 that there is gradual increase in calculated polarizability and hyperpolarizabilities with an increase in the dielectric constants of the solvents which may be explained due to the different stabilization of the frontier orbital’s of different

ACCEPTED MANUSCRIPT solvents. The first static hyperpolarizability (β0) of PCINH is calculated as 33.89 x 10–30, (gas phase), 68.79 x 10–30 (CHCl3), esu, 76.76 x 10–30 (CH2Cl2), 85.16 x 10–30 esu (DMSO). The p– Nitroaniline is chosen as a reference molecule having β0 value = 11.54 x 10–30 (gas phase), 28.20 x 10–30 (CHCl3), 38.51 x 10–30 (CH2Cl2), 40.51 x 10–30 (DMSO) esu.

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The polarizable continuum model (PCM) exploits a continuum solvation model (CSM)

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is a commonly used method in computational chemistry to model solvation effects. Modeling the

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solvent as a polarizable continuum, rather than individual molecules, makes ab initio computation feasible. The solute is accommodated inside a molecular cavity, built as a set of

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interlocking spheres centered on the atoms constituting the molecule under investigation.

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Mentioned solvents: CHCl3, CH2Cl2 and (CH3)2SO have been arranged with dielectric constant and dipole moment as CHCl3 Dielectric constant 4.8, Dipole moment 1.04 D, CH2Cl2 Dielectric

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constant 9.1, Dipole moment 1.60 D, (CH3)2SO Dielectric constant 47, Dipole moment 3. The

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values of the calculated properties (Dipole moment (µ0), Polarizability (|α0|), anisotropy of Polarizability (Δα), First Hyperpolarizability (β0)) of PCINH and its hydrated form in different

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solvents are arranged in Table 2. As dielectric constant and dipole moment of solvents increase

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value of above properties found to increase. The variation of NLO of azo-enaminone derivatives with variation of solvent properties (Dielectric constant and Dipole moment) has been reported

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in literature [37] where impact of solvent is very clear. PCM model considers local minimum of energy during the interaction in solvent than one might explicitly try to speculate in term of specific type of interaction. The variation of above calculated properties based on Polarizable Continuum Model (PCM) correlates well with variation of Dielectric constant and Dipole moment of solvents. The investigated molecule has higher hyperpolarizability value than p– Nitroaniline in gas as well as solvent phase.

ACCEPTED MANUSCRIPT 3.9 Evaluation of Antimicrobial activity Pyrrole and isonicotinyl hydrazide or Isoniazid derivatives are two major heterocycles, whose compounds possess wide range of biological activities. When pyrrole and Isoniazid moiety join together by azomethine framework (CH=NNHCO), formed PCINH compound show enhance

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antimicrobial activity has been observed than individual ones as shown in Table 3. Antifungal

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and antibacterial activity of study compound was studied against Aspergillus Niger and human

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pathogenic gram-positive bacteria Bacillus subtilis. PCINH and Isoniazid exhibited remarkable in vitro activity against test organism strain. The bar diagram of inhibition of zone for fungal and

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bacterial strain are shown in Figure 9. The systematic perusal of antimicrobial activity data

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reveals that, the antifungal and antibacterial effect is highly significant and pronounced for PCINH compound. A close inspection of screening data reveals presence of pharmacophore (-

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CH=NNHCO) highly influencing the antibacterial and antifungal activity PCINH than Isoniazid.

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PCINH and Isoniazid show comparatively low antimicrobial activity chloramphenicol (bacterial) and Nystatin (fungal), the reference drug.

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3.10 Evaluation of Antitubercular activity

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In this study, PCINH was evaluated against Mycobacterium tuberculosis. The PCINH displayed significant inhibitory effects (IC90 ≤ 10 µg/mL). PCINH showed an IC90 value of 2.68, IC50

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value of 3.11 and the selectivity index (SI) of 1.05 (Table 4). This type of effective derivative of isoniazid is ideally suited for further modification to obtain more active and less cytotoxic antimycobacterial compounds. Information about the geometry and structure of the molecule and its molecular electrostatic potential surfaces (MESP) are useful for understanding the relationship between molecular structure and biological activity. The EHOMO is a quantum chemical parameter which reflects the ability of molecule to electron donation. The higher the

ACCEPTED MANUSCRIPT energetic level of (HOMO), the lesser is the value of the ionization potential, so electrons from HOMO can easily be donated. The PCINH molecule has increased E HOMO value as well as increased biological activity than its precursor Isoniazid. Furthermore, the frontier molecular orbitals energy gap (the energy band gap (εL – εH) i.e. energy difference between the frontier

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molecular orbitals (LUMO - HOMO)) is another quantum chemical molecular descriptor that

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measures the molecular reactivity. The low energy gap (εL – εH) indicates higher reactivity of the

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compound. The low value of energy band gap (εL– εH) found for product (PCINH), therefore, is a good indication of the enhancement of the biological activity of the product PCINH. Similarly,

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lower value of global hardness (η) and global electrophilicity index (ω) clearly support the

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enhanced biological activities of product (PCINH) than that of reactant (INH). Higher values of chemical potential (μ) and global softness (S) clearly support the enhanced biological activities

Conclusions

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biological activities of the system.

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of product (PCINH) than that of reactant (INH). As per these data, theoretical results support the

A PCINH was prepared and characterized by a combined experimental and theoretical

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quantum chemical calculations. Vibrational spectroscopy and density functional calculation have

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been applied to the investigation of PCINH. The equilibrium geometries and harmonic vibrational wavenumbers of all the 26 normal modes of the molecule were determined and analyzed with DFT level of theory employing 6-311++G(d,p) basis set. The calculated 1H NMR and

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C NMR chemical shifts are in good agreement with the observed chemical shifts

experimentally. The calculated vibrational wavenumbers are in complement with the observed wavenumbers in FT-IR spectrum. The experimental FT-IR and Raman wavenumbers were compared with the respective theoretical values obtained from DFT calculations and found to

ACCEPTED MANUSCRIPT agree well with calculated frequencies. The rotational barrier between syn-syn and syn-anti forms is 12.70 kcal/mol in the gas phase. This rotational barrier is larger than the energy difference and thus the free rotation is quite a difficult task at room temperature. Forward rate coefficient of equilibrium conversion evaluated from Gibbs free energies of activation according to TS1

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expressed as log kf are -19.2. The value of equilibrium constant between syn-syn and syn-anti

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forms is 1259.8. The value of equilibrium constant (k) is greater than 1 this means that the

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conversion would only be possible when energy is provided to the syn-syn conformer. The local reactivity descriptors suggest that the investigated compound may be used as precursor for the

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syntheses of new heterocyclic compounds such as thiadiazoline, thiazolidinones and

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azetidinones. The first static hyperpolarizability (β0) of PCINH is calculated as 33.89 x 10–30, (gas phase), 68.79 x 10–30 (CHCl3), 76.76 x 10–30 (CH2Cl2), 85.16 x 10–30 esu (DMSO). The

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effects of solvent study revealed that there is a gradual increase in the calculated properties with

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increase in dielectric constant of the solvents. The investigated molecule has higher hyperpolarizability indicating that might be used as non–linear optical material. After systematic

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perusal of antimicrobial activity data reveals that, the antifungal and antibacterial effect is highly

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significant and pronounced for PCINH compound. The presence of pharmacophore (-

Isoniazid

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CH=NNHCO) highly influencing the antibacterial and antifungal activity of PCINH than

Acknowledgement

The one of the author is thankful to NCERT for providing fund for research work.

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References [1]. S.R. Sheeja, N. A. Mangalam, M.R. Prathapachandra Kurup, Y. S. Mary, H. T. Varghese, C. Y. Panicker, K. Raju, J. Mol. Struct. 973 (2010) 36–46 [2]. D. Ray, J. T. Foy, R. P. Hughes, I. Aprahamian, Nature Chem, 4(2012) 757-762 [3] A.V. Xavier (Ed.), Frontiers in Bioinorganic Chemistry, VCH, (1985). [4] A. Bijev, Drug Research 59(2009) 34-41 [5] A. Bijev, Arzneimittelforschung. 56(2006) 96-103. [6] Bijev, A.; Vladimirova, S.; Prodanova, P. Lett. Drug Des. Dis.; 6(2009) 508-517 [7] T.B. Chaston, D.R. Richardson, Am. J. Hematol. 73 (2003) 200-210. [8]. M. Fabian, G. Palmer, Biochem. 40 (2001) 1867-1874. [9] B.R. Bijini, S. Prasanna, M. Deepa, N. C. Mohanakumaran, K. Rajendrababu, Int. J. ChemTech Res. 4(2012) 739-74 [10] B. B. Safoklov, E. G. Atovmyan, L. A. Nikonova, V. V. Tkachev, S. M. Aldoshin, Russ. Chem. Bull. Internat. Edition 51(2002) 2224-2229. [11] M. S. V. Solange Wardell, V. N. de Souza Marcus, L. W. James, J. N. Low, C. Glidewell, Acta Crystal. Sec C, 62(2006) o47-o49 [12] N. Gunay, Atomic Mol. Phy. Book of Abstarct 40 (2008) [13]P. Rawat, R. N. Singh, J Mol. Struc. 1075(2014) 462-470 [14] M.J. Frisch, G.W. Trucks, H.B. Schlegel, G.E. Scuseria, M.A. Robb, J.R. Cheeseman, G. Scalmani, V. Barone, B. Mennucci, G.A. Petersson, H. Nakatsuji, M. Caricato, X. Li, H.P. Hratchian, A.F. Izmaylov, J. Bloino, G. Zheng, J.L. Sonnenberg, M. Hada, M. Ehara, K. Toyota, R. Fukuda, J. Hasegawa, M. Ishida, T. Nakajima, Y. Honda, O. Kitao, H. Nakai, T. Vreven, J.A. Montgomery, Jr., J.E. Peralta, F. Ogliaro, M. Bearpark, J.J. Heyd, E. Brothers, K.N. Kudin, V. N. Staroverov, T. Keith, R. Kobayashi, J. Normand, K. Raghavachari, A. Rendell, J.C. Burant, S. S. Iyengar, J. Tomasi, M. Cossi, N. Rega, J.M. Millam, M. Klene, J.E. Knox, J.B. Cross, V. Bakken, C. Adamo, J. Jaramillo, R. Gomperts, R.E. Stratmann, O. Yazyev, A.J. Austin, R. Cammi, C. Pomelli, J. W. Ochterski, R. L. Martin, K. Morokuma, V.G. Zakrzewski, G.A. Voth, P. Salvador, J.J. Dannenberg, S. Dapprich, A.D. Daniels, O. Farkas, J.B. Foresman, J.V. Ortiz, J. Cioslowski, and D. J. Fox, Gaussian 09, Revision B.01, Gaussian, Inc., Wallingford, CT, 2010. [15] P. Pulay, G. Fogarasi, F. Pang, J. Am. Chem. Soc. 101 (1979) 2550-2560. [16] Guang, Z.; Jianzhi, H.; Xiaodong, Z.; Lianfang, S.; Chaohui, Y.; Webb, G.A. Chem.l Phy. Lett. 266(19197) 533-536 [16] S. Chasvised, W. Rakrai, N. Morakot, B. Wanno, Intern. Trans. J. Engin. Man. Appl.Sci. Tec. 1(2011) 73-82 [17] R. N. Singh, A. Kumar. R. K. Tiwari, P. Rawat, Spectrochimica Acta Part A 112(2013) 182190. [18] P. Rawat, R. N. Singh, J. Mol. Struc. 1074(2014) 201-212. [19] P. Rawat, R. N. Singh, Arabian J. Chem. (2014) doi:10.1016/j.arabjc.2014.10.050 [20] R. N. Singh, P. Rawat, J. Mol. Struc. 1054-1055(2013) 65-67. [21] P. Rawat, R. N. Singh, Spectrochimica Acta Part A 140(2015) 344-355 [22] S. G. Franzblau, R. S. Witzig, J. C. Mclaughlin, P. Torres, G. Madico, A. Hernandez, M. T. Degnan, M. B. Coo, V. K. Quenzer, R. M. Freguson, R. H. Gilman. J. Clin. Microbiol, 36(1998) 362-366. [23] G.A. Guirgis, P. Klaboe, S. Shen, D.L. Powell, A. Gruodis, V. Aleksa, C.J. Nielsen, J. Tao, C. Zheng, J.R. Durig, J. Raman Spectrosc. 34 (2003) 322–336.

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[24] P.L. Polavarapu, J. Phys. Chem. 94 (1990) 8106–8112. [25] Y. Hoshina, T. Kobayashi, Engineering 4(2012) 139-145 [26] T. Van Mourik, M. Bühl, M. P. Gaigeot, Philos. Trans. A Math. Phys. Engin. Sci. 372(2012) 1-8 [27] T. Tsuneda, J. W. Song, S. Suzuki, K. Hirao, Chem. Phys. 133(2010) 174101 [28] P Rawat, RN Singh, J. Mol. Struc. 1084 (2015) 326-339 [29] P. Rawat, R. N.Singh J. Mol. Struc. 1082 (2015) 118–130 [30] P. Rawat, R. N.Singh J. Mol. Struc. 1100 (2015) 105-115 [31] P. Rawat, R. N.Singh J. Mol. Struc. 1097(2015) 214–225 [32] R.N. Singh, P. Rawat, S. Sahu J. Mol. Struc. 1065–1066(2014) 99–107 [33] P. Rawat, R. N. Singh, V. Baboo, P. Niranjan, H. Rani, R. Saxena, S. Ahmad J. Mol. Struc. 1129(2017) 37-49 [34] P. Rawat, R. N. Singh, S. Sahu, P. Niranjan, H. Rani, R. Saxena, S. Ahmad, ChemistrySelect 1(2016) 4008–4015 [35] Ana E. De A. Machado, L. A. De Souza, H. F. Dos Santos, Wagner B. De Almeida, J. Polymer Sci. Part B: Polymer Phys. 49(2011) 1410-1419 [36] O. Christiansen, J. Gauss, J. F. Stanton, Chem. Phys. Lett. 305(1999) 147-155 [37] F. S. Daniel, Machado, O.T. Lopes, I. T. Lima, D. A. Da Silva Filho, C. B. De Oliveira, J. Phys. Chem. C 2016, 120, 17660 – 17669

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Captions Figure Figure 1. Optimized geometry of ground state syn-syn, syn-anti conformers and Transition state Figure 2. Comparison of the experimentally determined structure (by single crystal x-ray diffraction) shown in green colour and optimized structure in gray colour of (a) syn-syn and (b) syn-anti conformer of PCINH (hydrogen atoms, not shown for clarity in presentation). Figure 3. Energy profiles for conversion of syn-syn (a), syn-anti (b) and transition state Figure 4. Differential scanning calorimetry (DSC) curve of PCINH for a heating rate of 15o C/min inN2 atmosphere. Figure 5. Experimental and calculated absorption spectrum of PCINH Figure 6. Experimental and calculated IR spectrum of PCINH along with correlation graph Figure 7. The simulated FT-Raman spectra of FCINH at (a) DFT and (b) HF level Figure 8. Molecular electrostatic potential (MEP) maps on the isodensity surface calculated at the B3LYP/6-311++G(d,p) level of theory: (a) from -6.301e-2 (red) to +6.301e-2 (blue) for syn conformer of PCINH. Figure 9. The bar diagram represent the antibacterial and antifungal zone of inhibition

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Tables Table 1. Calculated εHOMO, εLUMO, energy band gap (εL – εH), chemical potential (μ), electronegativity (χ), global hardness (η), global softness (S), global electrophilicity index (ω) for 2-Formyl-1H-pyrrole (Reactant I), Isoniazid (Reactant II) and Elecrophilicity based charge transfer (ECT) for reactant system [(I)↔(II)]. Table 2. Calculated Dipole moment (µ0), Polarizability (|α0|), anisotropy of Polarizability (Δα), First Hyperpolarizability (β0) and their components for PCINH with and without water molecule. Table 3. Antibacterial and antifungal screening data against studied compounds Table 4. Antitubercular activity screening data of the studied compounds

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TS (c) Figure 1. Optimized geometry of ground state syn-syn, syn-anti conformers and Transition state

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2 (a)

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Figure 2. Comparison of the experimentally determined structure (by single crystal x-ray diffraction) shown in green colour and optimized structure in gray colour of (a) syn-syn and (b) syn-anti conformer of PCINH (hydrogen atoms, not shown for clarity in presentation).

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Figure 3. Energy profiles for conversion of PCINH conformer syn-syn (a) to syn-anti (b) through transition state

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Figure 4. Differential scanning calorimetry (DSC) curve of PCINH at heating rate of 15° C/min in N2 atmosphere

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Figure 5. Experimental and calculated absorption spectrum of PCINH

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Figure 6. Experimental and calculated IR spectrum of PCINH along with correlation graph

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(a)

(b) Figure 7. The simulated FT-Raman spectra of FCINH at (a) DFT and (b) HF level

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Figure 8. Molecular electrostatic potential (MEP) maps on the isodensity surface, calculated at

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conformer of PCINH.

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Figure 9. The bar diagram plot of the zone of inhibition representing the antibacterial and

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ACCEPTED MANUSCRIPT Table 1. Calculated εHOMO, εLUMO, energy band gap (εL–εH), chemical potential (μ), electronegativity (χ), global hardness (η), global softness (S), global electrophilicity index (ω) for 2-Formyl-1H-pyrrole (Reactant I), Isoniazid (Reactant II) and Electrophilicity based charge transfer (ECT) for reactant system [(I)↔(II)]. εH εL εL– εH μ=-χ η S ω ECT 5.0055 -3.7613

2.5027

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-1.6337

5.4874 -4.3775

2.7437

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3.4920

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4.4270 -3.3316

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εH, εL, εL–εH, μ, χ, η, ω (in eV) and S (in eV–1)

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2.5072

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Table 2. Calculated Dipole moment (µ0), Polarizability (|α0|), anisotropy of Polarizability (Δα), First Hyperpolarizability (β0) and their components for PCINH with and without water molecule. Dipole Polariza- Anisotropy of First moment bility polarizability hyperpolarizability µ0 |α0| Δα β0 Gas phase PCINH 3.1103 21.1778 85.2062 33.8935 PCINH--H2O 2.3424 26.1546 97.4886 53.7731 PNA 7.1690 5.9870 25.8710 11.5480 CHCl3 PCINH 4.3469 28.4453 112.1796 68.7965 PCINH--H2O 2.5474 41.4905 111.0948 93.1154 PNA 8.6890 14.6410 53.2540 28.2031 CH2Cl2 PCINH 4.5817 29.5478 116.1235 76.7666 PCINH--H2O 2.7427 33.70166 122.1539 101.2658 PNA 9.266 16.107 59.177 38.5141 DMSO PCINH 4.8463 30.7024 120.1349 85.1679 PCINH--H2O 2.9671 35.0558 125.9909 109.6033 PNA 9.366 16.127 59.187 40.5141 HF Gas phase PCINH 4.4486 11.4788 56.4368 8.0484 PCINH--H2O 2.2166 19.63304 73.18562 10.14879 CHCl3 PCINH 4.3218 22.3506 81.6677 12.3589 PCINH--H2O 2.8512 23.2565 85.0910 14.4810 CH2Cl2 PCINH 4.1966 23.0968 83.8767 13.2204 PCINH--H2O 3.0250 24.0032 87.2889 15.1665 DMSO PCINH 4.4486 23.8822 86.1259 14.0484 PCINH--H2O 3.2272 24.8071 89.5187 15.9170 -24 -30 µ0 in Debye; |α0| and Δα in 10 esu; β0 in 10 esu,

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Table 3. Antibacterial and antifungal screening data against studied compounds and drugs Comp. / drugs Zone of inhibition in mm Bacillus subtilis Aspergillus niger 200 µg/ml 100 µg/ml 200 µg/ml 100 µg/ml 14 9 16 9 Isoniazid (INH) PCINH 24 12 20 14 32 24 Chloramphenicol 28 20 Nystatin 0 0 0 0 Control

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Table 4. Antitubercular activity screening data of the studied compounds and drugs Comp. IC90 µg/ml IC50 µg/ml SI Activity 7.57

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Isoniazid (INH) PCINH Rifampicin Control

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Graphical abstract

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Highlights

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►The intramolecular interactions have been studied with the help of NBO analysis.

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►Rotational barrier between syn and anti-structure of PCINH is 12.7 kcal/mol.

►PCINH shows good antimicrobial activity against fungal and bacterial strains.

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►The calculated β0 value for FPINH is 33.89 x 10-30 esu.

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