2.1. Computational details

All calculations were carried out using Gaussian 09 program package with the GaussView 05 molecular visualization program on personal computer ^[20,^21]. The molecular structure and vibrational computations of 5-acetyl-2,4-dimethylthiazole molecule were calculated by using Becke-3-Lee Yang Parr (B3LYP) and HSEH1PBE density functional theory methods with LanL2DZ basis set in ground state, respectively. Since the other well-known basis set lead us to imaginary vibrational frequency values, the mentioned basis set was used, and so the optimized molecular structure of title molecule in the ground state was obtained at the B3LYP/ and HSEH1PBE/ LanL2DZ levels. By considering the well-known systematic errors which come from the negligence of anharmonicity, electron correlation and basis set deficiencies, and furthermore, the computation in DFT method was performed in gas phase of isolated molecule while the experimental measurements were taken in solid phase for the title molecule ^[22,^23], the calculated vibrational wavenumbers were scaled as 0.961 for frequencies higher than 800 cm​-1 and 1.001 for frequencies less than 800 cm​-1 at the B3LYP and HSEH1PBE/LanL2DZ levels ^[24]. The assignments of fundamental vibrational modes of the title molecules were performed on the basis of total energy distribution (TED) analysis by using VEDA 4 program ^[25]. In the NMR calculations, the optimized molecular geometry of title molecule was first obtained at the B3LYP and HSEH1BPE/LanL2DZ levels for 5-acetyl-2,4- dimethylthiazole, by using the conductor-like polarizable continuum method (CPCM). Afterwards, the ​1H and ​13C NMR chemical shifts were calculated using the gauge- including atomic orbital (GIAO) method based on optimized at the mentioned levels in gas phase and in CHCl​3 solvent ^[26,^27]. The UV-vis spectra in gas phase and chloroform solvent were obtained using the time dependent DFT (TD-DFT) method. ^[28,^29] Furthermore, the FMO analyses of the title compound were performed by using DFT/HSEH1PBE method with 6-311++G(d,p) basis set and their 3D plots were verified.

2.2. Molecular Docking

Molecular docking studies of 5-acetyl-2,4-dimethylthiazole was performed by on SwissDock web server using EADock DSS algorithm ^[30]. High resolution crystal structure of vascular endothelial growth factor (VEGFR-2) (PDB ID: 2XIR) and β-ketoacyl-acyl carrier protein (ACP) synthase III (KAS III) (PDB ID: 1HNJ) were downloaded from a protein data bank website (https://www.rcsb.org/pdb/home/home.do). 5-Acetyl-2,4-dimethylthiazole was prepared for docking by energy minimized using the molecular mechanics and the semi-empirical AM1 methods with Gaussian 09 program package^[20]. All images in molecular docking studies were drawn with the UCSF Chimera package ^[31].

3.1. Molecular structure

The optimized molecular structures at the HSEH1PBE/Lan L2DZ levels basis set of the 5-acetyl-2, 4-dimethylthiazole compound are in Fig. 1. In Table I we only present some selected bond lengths, bond angles and dihedral angles which were calculated at the B3LYP and HSEH1BPE /LanL2DZ levels for the title molecule.

Figure 1 The optimized molecular geometry of 5-acetyl-2,4-dimethylthiazole at the B3LYP/LanL2DZ level.  

By considering the Table I and Fig. 1, in thiazole ring the double bond C1=N5 and the single bond C2-5N were calculated as 1.3203/1.3171 and 1.4025/1.3952 Å at the B3LYP and HSEH1BPE /LanL2DZ levels, respectively. Likewise, the bond lengths C1-S4 and C3-S4 were found as 1.8141/1.8001 and 1.8203/1.806 Å at the mentioned levels, respectively, which are the longest bond lengths of the title molecule. On the other hand, in CH​3CO- acetyl group of the 5-acetyl-2,4-dimethylthiazole compound, the double carbonyl C10=O19 bond length was calculated as 1.2588/1.2239 Å at the mentioned levels, respectively which are the shortest bond length of the title molecule. Furthermore, in the thiazole ring of the title compound the calculated S4-C1-N5, C1-S4-C3 and C1-N5-C2 bond angle values were found as 114.2158/114.3176, 87.3958/87.5172 and 113.4344/113.3176​∘ while the C3-C10-O19 bond angle in the CH​3CO- acetyl group were calculated as 119.9356/119.9199​∘ at the mentioned levels, respectively, as seen in Table I. Likewise, by considering some selected dihedral angles of the title compound exhibited in Table I, we should express that the compound under investigation is a non-planar molecule.

3.2. Vibrational analysis

The experimental and simulated IR spectra of the 5-acetyl-2,4-dimethylthiazole compound (C​7H​9NOS) are shown in Fig. 2. This molecule has 19 atoms, and its number of fundamental vibrational modes is 51. Furthermore, the observed and calculated at the B3LYP and HSEH1BPE/LanL2DZ levels vibrational frequencies and vibrational frequency assignments of the title compound are summarized in Table II. As seen in Table II, the assignments of vibrational frequencies were performed by using PED analysis.

Figure 2 The experimental and simulated IR spectra of 5-acetyl-2,4-dimethylthiazole. 

By considering Table II and Fig. 2, the ν(CH) symmetric stretching modes of the title molecule are observed at 2999.31, 2964.59, 2924.08 and 2961.56 cm^-1 while their calculated wavenumber values at the B3LYP and HSEH1PBE/LanL2DZ level were found as 3008.94/3042.80, 3004.72/3039.31, 2932.77/2957.62 and 2938.89/2961.81 cm^-1 which are the quite pure modes by the 100% and 79% PED contributions, respectively ^[32-^36]. On the other hand, the position of C=O stretching band at the interval 1870-1540 cm^-1 depends on properties such as the physical state, electronic and mass effects of neighboring substituents, intramolecular and intermolecular hydrogen bonding and conjugations ^[33,^34]. In this context, the C=O stretching band of the title molecule is observed at 1670.35 cm^-1 as a very strong band as seen in Fig. 2, and its calculated value at the mentioned levels were found as 1560.83/1595.54 cm^-1 which shows a fall of approximately 109.62/74.81 cm^-1 in carbonyl frequency for acetyl group.

As for the v(C=N) stretching modes of thiazole ring of the title molecule observed at 1510.28 cm^-1 was calculated as 1498.29/1528.25 cm^-1 at the B3LYP and HSEH1PBE /LanL2DZ levels respectively, which are coupled by v(C-C) stretching vibration. Furthermore, the v(C-N) stretching modes observed at 1446.61 and 1269.52 cm^-1 and calculated at the mentioned levels as 1444.87/1448.45 and 1223.73/1252.40 cm^-1, respectively. On the other hand, the v(S-C) stretching mode is observed at 667.37 cm^-1 as a strong band in Fig. 2 and calculated as 667.11/678.55 cm^-1 at the mentioned levels, respectively.

3.3. NMR analysis

The nuclear magnetic resonance (NMR) spectroscopy is a powerful tool for determining the composition, structure and function of complex molecules and to compute the reliable magnetic properties which provide the accurate predictions of molecular geometries and the isotropic chemical shift analysis ^[37,^38]. In this framework, the ¹H and ¹³C NMR chemical shift values were calculated at the mentioned levels by using GIAO model. The calculated ¹ H and ¹³ C NMR chemical shift values of the title molecule in gas phase and in the chloroform solvent at the B3LYP and HSEH1PBE /LanL2DZ levels are presented in Table III.

By considering Table III, the outstanding point for the calculated ​1H NMR chemical shift values in CHCl₃ solvent is that the all computed values vary at the interval 1.5944/1.6537-2.1332/2.2579 ppm at the B3LYP and HSEH1PBE /LanL2DZ levels since all protons of the 5-acetyl-2,4-dimethylthiazole compound (C₇H₉NOS) which has no aromatic ring proton belongs to the methyl groups.

On the other hand, as seen in Table III, the highest calculated carbon-13 NMR chemical shift value in CHCl₃ solvent of the title molecule at the B3LYP and HSEH1PBE /LanL2DZ levels was found as 194.838/190.306 ppm for the C10 atom which is connected to the O19 atom with high electronegativity while the lowest two calculated values in the chloroform solvent were found as 14.7945/10.49 ppm and 15.5823/11.2336 ppm for the C6 and C11 atoms, respectively.

3.4. UV-vis absorption and FMOs analysis

The experimental UV-vis in ethanol (EtOH), chloroform (CHCl₃) and N, N-dimethylformamide (DMF) solvents and the simulated at the B3LYP and HSEH1PBE/LanL2DZ levels in gas phase and chloroform solvent spectra of the 5-acetyl-2,4-dimethylthiazole molecule are presented in Fig. 3. Furthermore, the maximum absorption wavelengths (λ_max), excitation energies and oscillator strengths calculated by using TD-DFT/ B3LYP and HSEH1PBE/LanL2DZ levels of the title molecule in both gas and chloroform solvent are presented in Table IV.

Figure 3 The experimental (in CHCl₃, EtOH and DMF) solvents and simulated UV-vis spectra at the B3LYP and HSEH1PBE/LanL2DZ levels.  

The experimental maximum absorption wavelengths ( λmax) are observed at 243 nm, 265 nm, 278 nm, 295 nm and 315 nm in ethanol solvent, 259 nm, 295 nm, 309 nm, 318 nm and 337 nm in chloroform solvent and 295 nm and 316 nm in N, N-dimetylformamide solvent. By considering Table IV, the calculated maximum absorption wavelengths (?max) in chloroform solvent which correspond to the experimental 278, 265 and 243 nmλmax values in chloroform solvent were found as 277.46/269.74 nm, 251.02/242.69 nm and 214.33/207.75 nm at the B3LYP and HSEH1PBE/LanL2DZ levels, respectively. Therefore, the observed and calculated absorption bands can be attributed to theπ→π*, π→π* and n→π* transitions for the title molecule ^[39].

On the other hand, the highest occupied molecular orbital (HOMO) and the lowest unoccupied molecular orbital (LUMO) called the frontier molecule orbitals (FMOs) mean the outermost orbital filled by electrons and the first empty innermost orbital unfilled by electron, respectively. The HOMO is directly related to the ionization potential and behaves as an electron donor and the LUMO is directly related to the electron affinity and behaves as an electron acceptor. Therefore, the energy gap formed between HOMO and LUMO can be considered as a critical parameter and is an indicator in terms of the molecular chemical stability, and to identify the molecular electrical transport properties of any molecule. By using this energy gap, the molecular properties such as the chemical reactivity, polarizability, chemical hardness and softness, and electronegativity can be determined^[40,^41].

In this study, the 3-dimensional (3D) plots of the title molecule at the HSEH1PBE /LanL2DZ level are given in Fig. 4. The calculated energy gap value between the HOMO-LUMO for the title molecule is 4.42 eV at the mentioned level, as seen in Fig. 4. The large energy gap demonstrates the charge transfer occurs within of the title compound, and complex cannot be easily polarized. Besides, this value showing HOMO → LUMO contribution is originated from π→π* transition, as seen in Table IV and Fig. 4.

Figure 4 3D plots of HOMO-LUMO of 5-acetyl-2,4-dimethyl thiazole at the HSEH1PBE/LanL2DZ level.  

3.5. NBO analysis

The NBO analysis provides an explanation of the intramolecular and intermolecular bondings and interactions among bonds in molecular system. Furthermore, it allows to study on the hyperconjugation interactions or charge transfers (ICT) in the molecules.

In this framework, a stabilizing donor-acceptor interaction is corresponded to the delocalization of electron density between occupied Lewis type orbitals and formally unoccupied non-Lewis orbitals. Therefore, in order to make clear the intra- and inter-molecular interactions in the molecular systems, the stabilization energies of the molecules have been studied by using second-order perturbation theory. For each donor NBO (i) and acceptor NBO (j), the stabilization energy E ⁽²⁾ associated with electron delocalization between donor and acceptor is estimated as ^[42,^43]

where q _i is the donor orbital occupancy, ε _i and ε _j are diagonal elements (orbital energies), and F _ij is the off-diagonal NBO Fock matrix element (1). In this work, the results of second-order perturbation theory analysis of the Fock Matrix at the B3LYP and HSEH1PBE /LanL2DZ levels of theory for the 5-acetyl-2,4-dimethylthiazole molecule is presented in Table V.

By considering Table V, the stabilization energy values greater than 7.00 kcal mol^-1 are listed. Therefore, the strongest stabilization energy values calculated at the B3LYP and HSEH1PBE/LanL2DZ levels were found as 27.47/26.74 kcal/mol, respectively and corresponds to the transition between the lone pair n electrons of S4 electrons and π* antibonding electrons of the C1-N5 in thiazole ring. Furthermore, the transitions between n(N5)→σ*(C1-S4), n(O19) →σ* (C10-C15) and n(O19) →σ*(C3-C10) occurred in the title molecule. Likewise, the π→π* transition occurred between (C2-C3) and (C10-O19) bonds for the title molecule, as seen in Table V.

3.6. NLO parameters

The calculated results of the molecular polarizabilities at the B3LYP and HSEH1PBE /LanL2DZ levels of the finite-field approach for the 5-acetyl-2,4-dimethylthiazole molecule are given in Table VI. Therefore, total static dipole moment μ, the mean polarizability <α>, the anisotrotpy of the polarizability Δα and the mean first hyperpolarizability <β> which form the molecular polarizabilities of the title compound are shown in Table VI. As seen in Table VI, the μ in Debye, <α> and Δα in 10​-24 esu, and <β> in 10​-30 esu are presented as defined in cited^[44].

Therefore, the calculated values at the B3LYP and HSEH1PBE /LanL2DZ levels converted by using 1a.u=0.1482×10-24 electrostatic unit (esu) for α and 1a.u=8.6393×10-33 esu for β are reported as the polarizabilities and hyperpolarizability, respectively. On the other hand, it is well-known that the <β> value is a crucial factor for the

NLO parameters of the molecular systems and in this work, the calculated <β> values for the title molecule were found as 3.38/3.32×10-30 esu at the mentioned levels, as seen in Table VI.

3.7. Molecular Docking

The binding properties of 5-acetyl-2,4-dimethylthiazole to the VEGFR-2 kinase (PDB ID: 2XIR) and acetoacetyl-ACP synthase (PDB ID: 1HNJ) proteins were investigated, and molecular modelling studies were performed for ligand-protein interactions. Molecular docking studies provide the best representation of the association of the studied compound with the receptor, as well as hydrogen bonds that play an important role in ligand-protein interaction which is required for inhibition activity. As a result of the molecular docking studies, it was found that the optimized structure of the 5-acetyl-2,4-dimethylthiazole binds to the protein by hydrogen bonding with certain amino acids in the binding region of both proteins. The obtained energy and hydrogen bond location informations are given in Table VII.

According to the results, in the most stable docked pose between ligand-2XIR showed lowest interaction energy of -6.02 kcal/mol and was involved a hydrogen bonding interaction with Asp 1046 in the active site of 2XIR (Fig. 5). The amino acid Asp1046 had formed hydrogen bonding with nitrogen atom of 5-acetyl-2,4-dimethylthiazole (2.038 Å).

Figure 5 Binding model of 5-acetyl-2,4-dimethylthiazole with VEFGR-2 (PDB ID: 2XIR)  

On the other hand, ligand-1HNJ interaction exhibited the higher free energy at -5.96 kcal/mol than the ligand-2XIR couple. 5-Acetyl-2,4-dimethylthiazole and 1HNJ interaction also exhibited a hydrogen bonding between O atom of ligand and NH of Leu 191 with a 2.165 Å bond length (Fig. 6). Since the ligand forms a shorter hydrogen bonding with the 2XIR, the bond between the two is much stronger than ligand-1HNJ couple. Full fitness score value of ligand-2XIR (Fig. 7) and ligand-1HNJ (Fig. 8) interactions were found to be as and -1435.90 kcal/mol, respectively. These scores were in accordance with the order of the binding energy values. Docking assay of 5-acetyl-2, 4-dimethylthiazole with 2XIR showed a better full fitness score value and more preferred interaction.

Figure 6 Binding model of 5-acetyl-2,4-dimethylthiazole with KAS III (PDB ID: 1HNJ).  

Figure 7 The binding of 5-acetyl-2,4-dimethylthiazole in the active site of VEFGR-2 (PDB ID: 2XIR).  

Figure 8 The binding of 5-acetyl-2,4-dimethylthiazole in the active site of KAS III (PDB ID: 1HNJ).  

The docking study results clearly reveal that 5-acetyl-2,4-dimethylthiazole showed strong binding affinity towards the target proteins, achieving the best full fitness score and binding energy against 2XIR. However, full fitness score and binding energy for 1HNJ also appeared significant. On the basis of docking results, it was found that 5-acetyl-2,4-dimethylthiazole had potential to inhibit vascular endothelial growth factor receptor-2 kinase and β-Ketoacyl-acyl carrier protein synthase III.