Synthesis, Modeling Study and Antioxidants Activity of New Heterocycles Derived from 4-Antipyrinyl-2-Chloroacetamidothiazoles

: The present work reports the preparation of twelve new heterocyclic scaffolds containing an antipyrinyl-thiazole hybrid through the reaction of 4-antipyrinyl-2-chloroacetamido-thiazoles 1 and 6 with various types of nucleophiles, namely; ethyl thioglycolate, 2-mercaptobenzothiazole, 2-mercaptobenzoxazole, ammonium thiocyanate, malononitrile, and salicylaldehyde. The constructed compounds were characterized by conventional spectroscopic techniques (IR, 1 H NMR, 13 C NMR, and mass analysis). A DFT method (material studio package) was used to predict the geometry, bond lengths, bond angles, and dipole moments as well as other global chemical reactivities of the constructed antipyrinyl-thiazole compounds. Also, their semi-core pseudopods calculations (dspp) were carried out with DNP (double numerical basis sets plus polarization functional) to predict the properties of materials. In addition, the antioxidant activity of these antipyrinyl-thiazole scaffolds has been screened by the ABTS method. The results indicated that 2-(4-antipyrinylthiazolylamino)-5-substituitedbenzylidene-thiazol-4(5 H )-ones 10b and 10c exhibited the best antioxidant activity with a percentage inhibition of 85.74% and 83.51%, respectively. ), chemical potential ( µ ), electronegativity ( χ ), global softness (S), 17.13%. 58.62; 17.24%. C 24 18 5 2 N, C, 52.39;


Introduction
The antipyrine nucleus has been utilized as a useful precursor for the construction of many biologically important heterocycles [1,2]. Antipyrine derivatives are of great interest in medicine as a result of their broad range of pharmacological activity and clinical applications, including antibacterial [3], analgesic [4], and anti-inflammatory [4,5] as well as antitumor activity [6,7]. Antipyrine scaffolds are strong inhibitors of cyclooxygenase isoenzymes and platelet thromboxane and prostanoids synthesis [8,9]. In addition, thiazoles are valuable basic units in the field of medicinal science and are found in a wide assortment of bioactive scaffolds [10,11]. It has been well established that they possess antibacterial [12,13], anti-inflammatory [14,15], antitubercular [16,17], anticonvulsant [18,19], and anticancer [20][21][22][23] activities. Aminothiazole-containing medicines have been applied in clinical use for over thirty years, e.g., Famotidine is used to treat and prevent gastroesophageal reflux disease [24,25], Abafungin as an antifungal agent is used in the treatment of dermatomycoses [26] and Cefdinir is a well-known FDA-approved antibiotic and a third generation broad-spectrum cephalosporin [27]. Recently, some new thiazoles have been synthesized and proved to possess antioxidant power and DNA damage inhibition ability [28,29]. In light of these previous reports, we envisaged that integrating antipyrine and thiazole nuclei in one molecule could potentially produce new molecular hybrids with significant synergistic antioxidant activities. The reactivity of C-5 of thiazole ring in the key compound, 4-antipyrinyl-2chloroacetamidothiazole (1) towards electrophilic diazo-coupling reaction was investigated. Thus, the coupling reaction of compound 1 with diazotized sulphanilamide was carried out in ethanol containing sodium acetate at 0-5 °C to furnish 4-antipyrinyl-2-chloroacetamido-5-(4sulfamoylphenylazo)thiazole (6), see Scheme 2. The infrared spectrum of 6 showed three characteristic absorptions at 3351, 3264, and 1697 cm −1 to indicate the presence of -NH2, N-H, and C=O functional groups, respectively. The 1 H NMR spectrum displayed four singlet signals at δ 2.72 ppm for three protons of CH3-C, at δ 3.28 ppm for three protons of CH3-N, at δ 4.44 ppm for two protons of -CH2-group, and at δ 12.92 ppm for one proton of N-H. In addition, a multiplet signal for the aromatic protons and NH2 (δ 7.28-7.61 ppm), and a doublet signal for the two protons at δ 7.94-8.00 ppm (AA'-BB' system of the 1,4-disubstituted phenyl ring). The reactivity of C-5 of thiazole ring in the key compound, 4-antipyrinyl-2chloroacetamidothiazole (1) towards electrophilic diazo-coupling reaction was investigated. Thus, the coupling reaction of compound 1 with diazotized sulphanilamide was carried out in ethanol containing sodium acetate at 0-5 • C to furnish 4-antipyrinyl-2-chloroacetamido-5-(4-sulfamoylphenylazo)thiazole (6), see Scheme 2. The infrared spectrum of 6 showed three characteristic absorptions at 3351, 3264, and 1697 cm −1 to indicate the presence of -NH 2 , N-H, and C=O functional groups, respectively. The 1 H NMR spectrum displayed four singlet signals at δ 2.72 ppm for three protons of CH 3 -C, at δ 3.28 ppm for three protons of CH 3 -N, at δ 4.44 ppm for two protons of -CH 2 -group, and at δ 12.92 ppm for one proton of N-H. In addition, a multiplet signal The reaction of 2-chloroacetamido-thiazole derivative 6 with 2-mercaptobenzothiazole and/or 2mercaptobenzoxazole was achieved by refluxing in ethyl alcohol and sodium acetate for 2 h to furnish the corresponding sulfide derivatives 7a and 7b, respectively. The chemical structures of 7a and 7b were deduced from their compatible spectral and elemental analyses.
Treatment of the key compound 1 with ammonium thiocyanate in refluxing ethanol has been described to furnish 2-((4-antipyrinylthiazol-2-yl)imino)thiazolidin-4-one (9). A plausible mechanism for the reaction is indicated in scheme 3. The reaction proceeds via intramolecular cyclization of the thiocyanate intermediate 8 and the Dimroth-like rearrangements [31]. The product of this reaction was designed as the lactam derivative 9 and finds support from the literature of Vicini et al. [32]. The constructed thiazolidin-4-one scaffold 9 underwent condensation with three para-substituted benzaldehyde derivatives in glacial CH3COOH and anhydrous CH3COONa (Knoevenagel condensation reaction) furnishing the corresponding 2-(4-antipyrinylthiazol-2-ylamino)-5-(substituted-benzylidene)-thiazol-4(5H)-one derivatives 10a-c in good yields, see Scheme 3. The chemical structures of 10a-c were determined based on their compatible spectral analyses. Thus, the IR spectrum of 10a showed the characteristic absorptions at 3176 and 1698 cm −1 referring to the functional groups -NH-and carbonyl (C=O), respectively. The 1 H NMR spectrum of the same scaffold exhibited a singlet for three protons at δ 2.61 ppm (CH3-C), a singlet for three protons at δ 2.28 ppm (CH3-N), a singlet for three protons at δ 3.84 ppm (OCH3), and a doublet for two protons at δ 7.14 ppm (aromatic protons). The aromatic and thiazole C-5 protons resonated as a multiplet (δ 7.28-7.62 ppm), the olefinic proton resonated as a singlet at δ 7.73 ppm, while the proton on N-H function resonated as a singlet at δ 12.62 ppm. In compounds 10a-c, the Z conformation of the exocyclic C=C double bond was assigned on the basis of 1 H NMR spectroscopy and on the basis of literature data for analogous 4-thiazolidinones [33]. The 1 H NMR spectra of compounds 10a-c showed only one kind of methine proton that, deshielded by the adjacent C=O, was detected at 7.73-7.78 ppm, which are higher chemical shift values than those expected for E isomers.
The reaction of 2-chloroacetamido-thiazole derivative 6 with 2-mercaptobenzothiazole and/or 2-mercaptobenzoxazole was achieved by refluxing in ethyl alcohol and sodium acetate for 2 h to furnish the corresponding sulfide derivatives 7a and 7b, respectively. The chemical structures of 7a and 7b were deduced from their compatible spectral and elemental analyses.
Treatment of the key compound 1 with ammonium thiocyanate in refluxing ethanol has been described to furnish 2-((4-antipyrinylthiazol-2-yl)imino)thiazolidin-4-one (9). A plausible mechanism for the reaction is indicated in scheme 3. The reaction proceeds via intramolecular cyclization of the thiocyanate intermediate 8 and the Dimroth-like rearrangements [31]. The product of this reaction was designed as the lactam derivative 9 and finds support from the literature of Vicini et al. [32].
The constructed thiazolidin-4-one scaffold 9 underwent condensation with three para-substituted benzaldehyde derivatives in glacial CH 3 COOH and anhydrous CH 3 COONa (Knoevenagel condensation reaction) furnishing the corresponding 2-(4-antipyrinylthiazol-2ylamino)-5-(substituted-benzylidene)-thiazol-4(5H)-one derivatives 10a-c in good yields, see Scheme 3. The chemical structures of 10a-c were determined based on their compatible spectral analyses. Thus, the IR spectrum of 10a showed the characteristic absorptions at 3176 and 1698 cm −1 referring to the functional groups -NH-and carbonyl (C=O), respectively. The 1 H NMR spectrum of the same scaffold exhibited a singlet for three protons at δ 2.61 ppm (CH 3 -C), a singlet for three protons at δ 2.28 ppm (CH 3 -N), a singlet for three protons at δ 3.84 ppm (OCH 3 ), and a doublet for two protons at δ 7.14 ppm (aromatic protons). The aromatic and thiazole C-5 protons resonated as a multiplet (δ 7.28-7.62 ppm), the olefinic proton resonated as a singlet at δ 7.73 ppm, while the proton on N-H function resonated as a singlet at δ 12.62 ppm. In compounds 10a-c, the Z conformation of the exocyclic C=C double bond was assigned on the basis of 1 H NMR spectroscopy and on the basis of literature data for analogous 4-thiazolidinones [33]. The 1 H NMR spectra of compounds 10a-c showed only one kind of methine proton that, deshielded by the adjacent C=O, was detected at 7.73-7.78 ppm, which are higher chemical shift values than those expected for E isomers. Condensation reaction of equimolar amounts of chloroacetamido derivative 1 and malononitrile in absolute ethyl alcohol containing a few drops of triethylamine afforded the corresponding condensation product 2-amino-1-(4-antipyrinylthiazol-2-yl)-3-cyano-4,5-dihydro-5-oxo-1H-pyrrole (12) which was identified based on its compatible spectral analyses, see Scheme 4. Thus, the infrared spectrum of compound 12 exhibited the characteristic absorptions at 3293, 3191, 2216, and 1705 cm −1 related to the functional groups amino (NH2), nitrile (C≡N), and carbonyl (C=O), respectively. In the 1 H NMR spectrum, the protons of two methyl, methylene, thiazole-H5, and amino functions were secured by the presence of their characteristic signals at δ 2.65 ppm (singlet), δ 3.32 ppm (singlet), δ 4.36 ppm (singlet), δ 7.29 ppm (singlet), and δ 12.47 ppm (singlet), respectively. Treatment of chloroacetamide derivative 1 with salicylaldehyde in DMSO containing potassium carbonate was achieved by stirring the reaction mixture for 8 h followed by neutralization with dilute HCl to furnish N-(4-antipyrinylthiazol-2-yl) benzofuran-2-carboxamide (13). It was reasoned that the chlorine atom in acetamido derivative 1 could be substituted by nucleophiles, so the plausible Condensation reaction of equimolar amounts of chloroacetamido derivative 1 and malononitrile in absolute ethyl alcohol containing a few drops of triethylamine afforded the corresponding condensation product 2-amino-1-(4-antipyrinylthiazol-2-yl)-3-cyano-4,5-dihydro-5-oxo-1H-pyrrole (12) which was identified based on its compatible spectral analyses, see Scheme 4. Thus, the infrared spectrum of compound 12 exhibited the characteristic absorptions at 3293, 3191, 2216, and 1705 cm −1 related to the functional groups amino (NH 2 ), nitrile (C≡N), and carbonyl (C=O), respectively. In the 1 H NMR spectrum, the protons of two methyl, methylene, thiazole-H5, and amino functions were secured by the presence of their characteristic signals at δ 2.65 ppm (singlet), δ 3.32 ppm (singlet), δ 4.36 ppm (singlet), δ 7.29 ppm (singlet), and δ 12.47 ppm (singlet), respectively. Condensation reaction of equimolar amounts of chloroacetamido derivative 1 and malononitrile in absolute ethyl alcohol containing a few drops of triethylamine afforded the corresponding condensation product 2-amino-1-(4-antipyrinylthiazol-2-yl)-3-cyano-4,5-dihydro-5-oxo-1H-pyrrole (12) which was identified based on its compatible spectral analyses, see Scheme 4. Thus, the infrared spectrum of compound 12 exhibited the characteristic absorptions at 3293, 3191, 2216, and 1705 cm −1 related to the functional groups amino (NH2), nitrile (C≡N), and carbonyl (C=O), respectively. In the 1 H NMR spectrum, the protons of two methyl, methylene, thiazole-H5, and amino functions were secured by the presence of their characteristic signals at δ 2.65 ppm (singlet), δ 3.32 ppm (singlet), δ 4.36 ppm (singlet), δ 7.29 ppm (singlet), and δ 12.47 ppm (singlet), respectively. Treatment of chloroacetamide derivative 1 with salicylaldehyde in DMSO containing potassium carbonate was achieved by stirring the reaction mixture for 8 h followed by neutralization with dilute HCl to furnish N-(4-antipyrinylthiazol-2-yl) benzofuran-2-carboxamide (13). It was reasoned that the chlorine atom in acetamido derivative 1 could be substituted by nucleophiles, so the plausible Treatment of chloroacetamide derivative 1 with salicylaldehyde in DMSO containing potassium carbonate was achieved by stirring the reaction mixture for 8 h followed by neutralization with dilute HCl to furnish N-(4-antipyrinylthiazol-2-yl) benzofuran-2-carboxamide (13). It was reasoned that the chlorine atom in acetamido derivative 1 could be substituted by nucleophiles, so the plausible pathway for the formation of benzofuran derivative 14 could be via formation of intermediate 13, which subsequently underwent a condensation reaction between the formyl and active methylene group, see Scheme 5. The structural proof of compound 14 was determined based on its correct spectral analyses. Thus, the infrared spectrum of compound 14 showed absorptions at 3369 and 1689 cm −1 to secure the functional groups (N-H) and (C=O), respectively. The 1 H NMR spectrum clearly indicated the presence of a singlet for three protons at 2.62 (CH 3 -C), a singlet for three protons at 3.33 (CH 3 -N), a singlet for one proton at 7.28 ppm (thiazole-H5), a multiplet for ten protons in the region of δ 7.36-7.71 ppm (aromatic-H and furan-H3), and a singlet for one proton at δ 12.31 ppm (NH).

Computational Studies
DMOL3 module calculations were used to examine the cluster estimations [34] and DNP, the double numerical basis sets plus polarization functional (DNP) implemented in the Materials Studio bundle [35]. It is constructed to realize the large-scale density functional theory (DFT) calculations [36][37][38][39]. The geometric optimization is performed with no symmetry confinement.

Geometry Optimization
The molecular structures along with atomic numbering of the title 4-antipyrinylthiazole scaffolds (2, 4, 5, 6, 7, 9, 10, 12, and 14) are represented in Figures 1 and 2. The bond lengths and bond angles are included in the supplementary material, Tables S1-S24. The data obtained in these tables reveal that the bond lengths or bond angles are altered to some extent upon the formation of a new thiazole derivative, which is in turn dependent (or influenced) by the nature of the attacking electrophile or the experimental conditions.

Global Reactivity Descriptors
Density functional theory (DFT) was utilized to understand the chemical reactivity and site selectivity of the molecular systems. As highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) are the orbitals that are the most likely to be involved in chemical reactivity, where the HOMO (π donor) is the highest energy orbital that is still occupied; therefore, energetically, it is the orbital which is easiest to remove electrons from, thus acting as a Lewis base. On the other hand, the LUMO (π acceptor) is the lowest lying orbital that is empty; therefore, energetically, this orbital is the easiest to add more electrons to (acts as Lewis acid). The energy gap = EHOMO − ELUMO and clarifies the inevitable charge exchange interaction inside the molecule. The global electrophilicity index (ω), chemical potential (μ), electronegativity (χ), global softness (S), and global hardness (η), are determined by the well-known equations [40] and the data are listed in Table 1.

Computational Studies
DMOL3 module calculations were used to examine the cluster estimations [34] and DNP, the double numerical basis sets plus polarization functional (DNP) implemented in the Materials Studio bundle [35]. It is constructed to realize the large-scale density functional theory (DFT) calculations [36][37][38][39]. The geometric optimization is performed with no symmetry confinement.

Geometry Optimization
The molecular structures along with atomic numbering of the title 4-antipyrinylthiazole scaffolds (2, 4, 5, 6, 7, 9, 10, 12, and 14) are represented in Figures 1 and 2. The bond lengths and bond angles are included in the supplementary material, Tables S1-S24. The data obtained in these tables reveal that the bond lengths or bond angles are altered to some extent upon the formation of a new thiazole derivative, which is in turn dependent (or influenced) by the nature of the attacking electrophile or the experimental conditions.

Global Reactivity Descriptors
Density functional theory (DFT) was utilized to understand the chemical reactivity and site selectivity of the molecular systems. As highest occupied molecular orbital (HOMO) and lowest unoccupied molecular orbital (LUMO) are the orbitals that are the most likely to be involved in chemical reactivity, where the HOMO (π donor) is the highest energy orbital that is still occupied; therefore, energetically, it is the orbital which is easiest to remove electrons from, thus acting as a Lewis base. On the other hand, the LUMO (π acceptor) is the lowest lying orbital that is empty; therefore, energetically, this orbital is the easiest to add more electrons to (acts as Lewis acid). The energy gap = EHOMO − ELUMO and clarifies the inevitable charge exchange interaction inside the molecule. The global electrophilicity index (ω), chemical potential (µ), electronegativity (χ), global softness (S), and global hardness (η), are determined by the well-known equations [40] and the data are listed in Table 1.
The values of Frontier molecular orbitals energies (FMOs) namely E HOMO and E LUMO as well as their neighboring orbitals are negative, indicating the stability of the synthesized antipyrinyl-thiazole derivatives [41].

2.
Based on FMOs theory, the reaction occurs with maximum overlap between the HOMO on one molecule and the LUMO on the other and this is a controlling factor in many reactions. Therefore, orbitals of the derivative with the largest value of molecular orbital coefficients may be considered as the sites of electron donation. Thus, the HOMO level is mostly localized on N (2) It is well documented that the smaller the energy gap (E HOMO − E LUMO ) of a molecule the greater the reactivity, polarizability, and readiness to offer electrons to an acceptor and the molecule is considered to be "soft", which in turn affects its biological activity. Thus, the title compounds follow the order: 10b > 10c > 6 > 7a > 7b > 10a > 5 > 14 > 4 > 9 > 2 > 12. This means that compound 10b possesses the smallest energy gap and the highest electrophilicty index (ω = 30.864) among all newly synthesized thiazoles and, therefore, has the highest softness, polarizability, and reactivity [13].

Antioxidant Activity
The antioxidant activity assay was estimated using an ABTS free radical scavenging activity assay (2,2′-azinobis(3-ethylbenzothiazoline-6-sulfonic acid) [42]. Inhibition free radical ABTS in percent (I %) was calculated as per the equation: where Ablank is the absorbance of the control reaction (containing all reagents except the test compound), and Asample is the absorbance of the test sample.

Antioxidant Activity
The antioxidant activity assay was estimated using an ABTS free radical scavenging activity assay (2,2 -azinobis(3-ethylbenzothiazoline-6-sulfonic acid) [42]. Inhibition free radical ABTS in percent (I %) was calculated as per the equation: where A blank is the absorbance of the control reaction (containing all reagents except the test compound), and A sample is the absorbance of the test sample. The antioxidant activities of the newly synthesized 4-antipyrinylthiazole scaffolds 2, 4, 5, 6, 7, 9, 10, 12, and 14 were evaluated using the ABTS Radical Cation Decolorization Assay [43]. The results, shown in Table 2, indicated that compounds 10b, 10c, and 7a have excellent antioxidant properties compared to the reference of the test (L-Ascorbic acid, 88.88%). The derivatives of 2-(4-antipyrinylthiazolylamino)-5-substituitedbenzylidene-thiazol-4(5H)-one substituted with electron-withdrawing groups at the benzylidene moiety exhibited the best antioxidant activity. The percentage inhibition of scaffolds 10b and 10c substituted with nitro and bromide groups at the benzylidene moiety are 85.74% and 83.51%, respectively.
The replacement of the chlorine atom of the 2-chloroacetamido-thiazole compound 6 (percentage inhibition of 73.33%) by the benzothiazolyl moiety to afford the corresponding antipyrinyl-thiazole scaffold 7a enhanced the antioxidant activity with percentage inhibition 78.14%.

General Methods
Melting points have been determined on Gallenkamp electric apparatus (capillary method, Gallenkamp Co., London, UK). IR spectra (KBr discs) have been obtained on a Mattson 5000 FT-IR spectrometer (Shimadzu Co., Kyoto, Japan). The nuclear magnetic resonance spectra have been recorded using a WP 300 spectrometer (Bruker Co., Billerica, MA, USA) at 300 MHz for 1 H-NMR or 75.5 MHz for 13 C-NMR. The mass analysis (EI technique, 70 eV) was acquired by a Qp-2010 mass spectrometer (Shimadzu, Tokyo, Japan).