Rev. Colomb. Cienc. Quím. Farm., Vol. 45(2), 201-218, 2016 www.farmacia.unal.edu.co Scientific research article / http://dx.doi.org/10.15446/rcciquifa.v45n2.59936 Synthesis, characterization and antibacterial screening of some Schiff bases derived from pyrazole and 4-amino antipyrine Shipra Baluja1, Sumitra Chanda2 1Department of Chemistry, Saurashtra University, Rajkot-360 005, Gujarat, India E-mail: [email protected] 2Department of Biosciences, Saurashtra University, Rajkot-360 005, Gujarat, India Received: September 22, 2015 Accepted: April 7, 2016 Summary Some Schiff bases of pyrazole and 4-amino antipyrine have been synthesized. The antibacterial screening of these synthesized compounds was done in dimethyl forma- mide against four Gram positive bacteria viz. Bacillus cereus, Staphylococcus aureus, Staphylococcus epidermidids and Micrococcus luteus, and three Gram negative bacteria viz. Proteus mirabilis, Escherichia coli and Klebsiella aerogenes. It is observed that in comparison to Schiff bases of 4-amino antipyrine, pyrazole Schiff bases are better for inhibition for these selected Gram positive and Gram negative bacterial strains. Keywords: Pyrazole, 4-amino antipyrine, Schiff bases, dimethyl formamide, antibac- terial activity. Resumen Síntesis, caracterización y evaluación antibacteriana de algunas bases de Schiff derivadas de pirazol y 4-amino antipirina Se sintetizaron algunas bases de Schiff a partir de pirazol y 4-amino antipirina. La evaluación de la actividad antibacteriana de estos compuestos en dimetil formamida se realizó frente a cuatro bacterias Gram positivas, Bacillus cereus, Staphylococcus aureus, Staphylococcus epidermidids y Micrococcus luteus, y frente a tres bacterias Gram negativas, Proteus mirabilis, Escherichia coli y Klebsiella aerogenes. Se observó 201 Shipra Baluja, Sumitra Chanda mejor inhibición bacteriana frente a las diferentes cepas para las bases de Schiff basadas en pirazol comparadas con aquellas basadas en 4-amino antipirina. Palabras clave: Pirazol, 4-amino antipirina, bases de Schiff, dimetil formamida, acti- vidad antibacteriana. Introduction Schiff’s bases are an important class of organic compounds having a wide variety of applications in many fields such as analytical, biological and inorganic chemistry [1-5]. Some of these compounds act as corrosion inhibitors [6] and are used as catalysts in polymer [7-8] and dyes [9] industries. Further, Schiff bases have gained importance in medicinal and pharmaceutical fields due to a broad spectrum of biological activi- ties like antimicrobial [10-11], antifungal [12-13], antiviral [14], anti-inflammatory [15-16], analgesic [17-18], anticonvulsant [19], antitubercular [20], anticancer [21], antioxidant [22-23], anthelmintic [24], antimalarial [25-26] and so forth. In continuation of our previous research [27-28], in the present work, some new Schiff bases have been synthesized having pyrazole and 4-amino antipyrine moieties and their structure were confirmed by IR, NMR and mass spectral data. The screening of these compounds was also done to study their antibacterial properties in dimethyl formamide. Experimental Synthesis of pyrazole Schiff bases Synthesis of (1E)-1-(4-nitrophenyl)ethanone phenylhydrazone Equimolar solution of phenyl hydazine and p-nitro phenyl acetophenone in absolute ethanol was refluxed in water bath for 2 hours using glacial acetic acid as catalysis. The crude product was isolated and was crystallized from absolute alcohol. Synthesis of 3-(4-nitrophenyl)-1-phenyl-1H-pyrazole-4-carbaldehyde (1E)-1-(4-nitro phenyl) ethanone phenyl hydrazone (0.01M) was added to Vilsmayer- Haack reagent (prepared by drop wise addition of 3 ml POCl in ice cooled 25 ml 3 DMF) and was refluxed for 5 hours. The reaction mixture was poured into ice followed by neutralization using sodium bicarbonate. The crude product was isolated and crys- tallized from ethanol. 202 Synthesis and antibacterial screening of some Schiff bases Synthesis of Schiff bases In an ethanolic solution of 3-(4-nitro phenyl)-5-phenyl-4H-pyrazole-4-carbaldehyde and different aromatic amines, 2-3 drops of glacial acetic acid was added and the reac- tion mixture was refluxed for 10 hours. The resulting solution was cooled to room temperature and was poured in crushed ice with constant stirring. The product was filtered and washed with sodium bisulfate solution to remove the unreacted aldehyde. The crude product was crystallized from methanol and dried. The synthesized Schiff bases are: 1. SA-1: (E)-3-(4-nitrophenyl)-1-phenyl-4-((2-phenylhydrazono)methyl)-1H- pyrazole 2. SA-2: (E)-4-methyl-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline 3. SA-3: (E)-4-nitro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene) aniline 4. SA-4: (E)-4-methoxy-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline 5. SA-5: (E)-3-chloro-4-fluoro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline 6. SA-6: (E)-4-chloro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline 7. SA-7: (E)-4-fluoro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline 8. SA-8: (E)-2,5-dichloro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline 9. SA-9: (E)-2-(((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)amino) phenol. Synthesis of Schiff base from 4-amino antipyrine Equimolar amount of different aldehydes and 4-amino anti pyridine was dissolved in 30 ml methanol. 0.1 mole of 4-amino antipyrine and few drops of glacial acidic acid were added in this solution and the mixture was refluxed for 12-14 hours at 80 °C-85 °C in water bath. 203 Shipra Baluja, Sumitra Chanda The resulting solution was cooled to room temperature and then poured in crushed ice with constant stirring. The product was filtered and washed with sodium bisulfate solution to remove the unreacted aldehyde. The crude product was crystallized from methanol and dried. The following Schiff bases have been synthesized from 4-amino antipyrine: 1. SB-1: (E)-4-((4-(dimethylamino)benzylidene)amino)1,5-dimethyl-2-phenyl- 1H-pyrazol-3(2H)-one 2. SB-2: (E)-4-((4-chlorobenzylidene)amino)1,5-dimethyl-2-phenyl-1H-pyrazol- 3(2H)-one 3. SB-3: (E)-4-((4-fluorobenzylidene)amino)1,5-dimethyl-2-phenyl-1H-pyrazol- 3(2H)-one 4. SB-4: (E)-1,5-dimethyl-4-((2-nitrobenzylidene)amino)-2-phenyl-1H-pyrazol- 3(2H)-one 5. SB-5: (E)-1,5-dimethyl-4-((3-nitrobenzylidene)amino)-2-phenyl-1H-pyrazol- 3(2H)-one 6. SB-6: (E)-4-((4-hydroxybenzylidene)amino)1,5-dimethyl-2-phenyl-1H-pyra- zol-3(2H)-one 7. SB-7: (E)-4-((2-chlorobenzylidene)amino)1,5-dimethyl-2-phenyl-1H-pyrazol- 3(2H)-one. All these compounds were synthesized according to the reaction schemes given in fig- ures 1 and 2. The physical parameters such as molecular formula, molecular weight, melting point, percentage yields, and R values along with the solvent system of all these synthesized f compounds are given in tables 1 and 2 respectively. The IR spectra (KBr pellets) were scanned on IR (SHIMADZU-FTIR-8400) over the frequency range from 4000-400 cm-1. 1H NMR spectra were scanned on Bruker Spectrometer (400 MHz) by using deuterated DMSO as a solvent. The Mass spectra were scanned on GCMS-SHIMADZU-QP2010. 204 Synthesis and antibacterial screening of some Schiff bases Antibacterial activity Test microorganisms The synthesized compounds were tested for its antibacterial activity against four Gram positive bacteria viz. Bacillus cereus ATCC11778, Staphylococcus aureus ATCC 29737, Staphylococcus epidermidids ATCC 12228 and Micrococcus luteus ATCC10240, and three Gram negative bacteria viz. Proteus mirabilis NCIM2241, Escherichia coli ATCC25922 and Klebsiella aerogenes NCTC418. The microorganisms were obtained from National Chemical Laboratory (NCL), Pune, India. Microorganisms were main- tained at 4 °C on nutrient agar slants. Figure 1. Reaction scheme for pyrazole Schiff bases O + NO N 2 - Abs. Ethanol O + HC N 3 NH NH CH NH2 O 3 - VilsmeierHaack Formylation N N +O R-NH2 N N +O N N O- Abs. Ethanol O- N O R Figure 2. Reaction scheme for 4-amino antipyrine Schiff bases H3C NH2 H3C CH 3 N N H C R-CHO 3 N O N N R methanol O 205 Shipra Baluja, Sumitra Chanda Table 1. Physical constants of pyrazole Schiff bases. Serial Compound Mol. Wt. R* M.P. Yield R Molecular formula f No. code (g/mol) value (°C) (%) 1. SA-1 -NH-CH C H NO 383 0.67 266 65 6 5 22 17 5 2 2. SA-2 4-CH-CH4 C H NO 382 0.54 218 69 3 6 23 18 4 2 3 SA-3 4-NO- CH C H NO 413 0.62 246 65 2 6 4 22 15 5 4 3. SA-4 3-OCH-CH C H NO 398 0.54 258 78 3 6 4 23 18 4 3 4. SA-5 4-F,3-Cl-CH C H ClFNO 420 0.47 243 65 6 3 22 14 4 2 5. SA-6 4-Cl-CH C H ClNO 402 0.55 198 70 6 4 22 15 4 2 6. SA-7 4-F-CH C H FNO 386 0.60 268 72 6 4 22 15 4 2 7. SA-8 2,5-di Cl-CH C H ClNO 436 0.46 283 68 6 3 22 14 2 4 2 8. SA-9 2-OH-CH C H NO 384 0.47 256 76 6 4 22 16 4 3 *TLC solvent system: Hexane:Ethyl acetate- 7.0:3.0 Table 2. Physical constants of 4-amino antipyrine Schiff bases. Serial Compound Mol. Wt. R* M.P. Yield R Molecular formula f No. code (g/mol) value (°C) (%) 1. SB-1 4-N(CH)-CH C H NO 334 0.42 218 65 3 2 6 4 20 22 4 2. SB-2 4-Cl-CH C H ClNO 325 0.35 258 63 6 4 18 16 3 3 SB-3 4-F- CH C H FNO 309 0.48 230 64 6 4 18 16 3 3. SB-4 2-NO-CH C H NO 336 0.45 210 73 2 6 4 18 16 4 3 4. SB-5 3-NO-CH C H NO 336 0.28 205 68 2 6 4 18 16 4 3 5. SB-6 4-OH-CH C H NO 307 0.34 202 69 6 4 18 17 3 2 6. SB-7 2-Cl-CH C H ClNO 325 0.56 210 78 6 4 18 16 3 *TLC solvent system: Hexane:Ethyl acetate- 8.0:2.0 Preparation of the test compound For all the compounds, solutions were prepared at a concentration of 0.2 mg/ml in DMF. Preparation of the plates and microbiological assay The antibacterial evaluation was done by agar well diffusion method [29] using Muel- ler Hinton agar No. 2 as the nutrient medium. The agar well diffusion method was preferred to be used in this study since it was found to be better than the disc diffu- sion method as suggested by Essawi et al. [30]. The bacterial strains were activated by 206 Synthesis and antibacterial screening of some Schiff bases inoculating a loop full of test strain in 25 ml of N-broth and the same was incubated for 24 hours in an incubator at 37 oC. 0.2 ml of the activated strain was inoculated in Mueller Hinton agar. Mueller Hinton agar kept at 45 oC was then poured in the Petri dishes and allowed to solidify. After solidification of the media, 0.85 cm ditch was made in the plates using a sterile cork borer and these were completely filled with the test solution. The plates were incubated for 24 h at 37 oC. The mean value obtained for the three wells was used to calculate the zone of growth inhibition of each sample. The controls were maintained for each bacterial strain. The inhibition zone formed by these compounds against the particular test bacterial strain determined the antibacterial activities of the synthetic compounds. The observed activities are also compared with well known antibiotics. Results and discussion The physical parameters all the synthesized compounds are given in tables 1 and 2. Spectral data SA-1: (E)-3-(4-nitrophenyl)-1-phenyl-4-((2-phenylhydrazono)methyl)-1H-pyra- zole IR (cm-1, KBr): 1543 (C=C str.), 1598 (C=N str. (Schiff base), 1543 (C=N str., (pyra- zole moiety), 1256 (C-N str.), 961 (N-N str.), 1256 (N-O str.). 1H NMR (DMSO-d ) δ(ppm): 6.91- 7.02 (2H, doublet, Ar-H), 7.29- 7.34 (2H, doublet, 6 Ar-H), 7.61-7.62 (2H, multiplet, Ar-H), 7.42-7.59 (9 H, multiplet, Ar-H), 7.63 (1 H, singlet, Ar-H), 8.85 (1H, singlet, -N=CH). MS: (m/z) = 383 SA-2: (E)-4-methyl-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene) aniline IR (cm-1, KBr): 1537 (C=C str.), 1620 (C=N str. (Schiff base), 1598 (C=N str., (pyra- zole moiety), 1256 (C-N str.), 961 (N-N str.), 1288 (N-O str.), 2922 (C-H str. (alkane). 1H NMR (DMSO-d ) δ(ppm): 3.75 (3H, singlet, C-CH ), 6.86-6.92 (2H, doublet, 6 3 Ar-H), 7.32- 7.47 (2H, doublet, Ar-H), 7.51-7.63 (9 H, multiplet, Ar-H), 7.74 (1 H, singlet, Ar-H), 8.84 (1H, singlet, -N=CH). MS: (m/z) = 382 207 Shipra Baluja, Sumitra Chanda SA-3: (E)-4-nitro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene) aniline IR (cm-1, KBr): 1508 (C=C str.), 1635 (C=N str. (Schiff base), 1635 (C=N str., (pyra- zole moiety), 1242 (C-N str.), 965 (N-N str.), 1339 (N-O str.). 1H NMR (DMSO-d ) δ(ppm): 6.64-6.71 (2H, doublet, Ar-H), 7.26- 7.32 (2H, dou- 6 blet, Ar-H), 7.63-7.78 (9 H, multiplet, Ar-H), 7.95 (1 H, singlet, Ar-H), 8.67 (1H, singlet, -N=CH). MS: (m/z) = 413 SA-4: (E)-4-methoxy-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene) aniline IR (cm-1, KBr): 1504 (C=C str.), 1627 (C=N str. (Schiff base), 1596 (C=N str., (pyra- zole moiety), 1242 (C-N str.), 961 (N-N str.), 1338 (N-O str.), 1029 (C-O-C str. Ether). 1H NMR (DMSO-d ) δ(ppm): 3.85 (3H, singlet, C-CH ), 6.94-6.97 (2H, doublet, 6 3 Ar-H), 7.23- 7.28 (2H, doublet, Ar-H), 7.39-7.55 (9 H, multiplet, Ar-H), 7.56 (1 H, singlet, Ar-H), 8.74 (1H, singlet, -N=CH). MS: (m/z) = 398 SA-5: (E)-3-chloro-4-fluoro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl) methylene)aniline IR (cm-1, KBr): 1502 (C=C str.), 1598 (C=N str. (Schiff base), 1597 (C=N str., (pyra- zole moiety), 1226 (C-N str.), 961 (N-N str.), 1340 (N-O str.), 1049 (C-F str.), 754 (C-Cl str.). 1H NMR (DMSO-d ) δ(ppm): 6.62-6.66 (1H, doublet, Ar-H), 7.07- 7.12 (1 H, tri- 6 plet, Ar-H), 7.10-8.02 (10 H, multiplet, Ar-H), 8.37 (1 H, singlet, Ar-H), 8.70 (1H, singlet, -N=CH). MS: (m/z) = 420 SA-6: (E)-4-chloro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene) aniline IR (cm-1, KBr): 1540 (C=C str.), 1598 (C=N str. (Schiff base), 1635 (C=N str., (pyra- zole moiety), 1336 (C-N str.), 960 (N-N str.), 1387 (N-O str.), 754 (C-Cl str.). 208 Synthesis and antibacterial screening of some Schiff bases 1H NMR (DMSO-d ) δ(ppm): 6.52-6.62 (2H, doublet, Ar-H), 7.01- 7.12 (2H, dou- 6 blet, Ar-H), 7.33-7.52 (9 H, multiplet, Ar-H), 7.82 (1 H, singlet, Ar-H), 8.84 (1H, singlet, -N=CH). MS: (m/z) = 402 SA-7: (E)-4-fluoro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene) aniline IR (cm-1, KBr): 1500 (C=C str.), 1599 (C=N str. (Schiff base), 1637 (C=N str., (pyra- zole moiety), 1333 (C-N str.), 961 (N-N str.), 1230 (N-O str.), 1079 (C-F str.). 1H NMR (DMSO-d ) δ(ppm): 6.45-6.53 (2H, doublet, Ar-H), 7.14- 7.26 (2H, dou- 6 blet, Ar-H), 7.36-7.62 (9 H, multiplet, Ar-H), 7.96 (1 H, singlet, Ar-H), 8.85 (1H, singlet, -N=CH). MS: (m/z) = 386 SA-8: (E)-2,5-dichloro-N-((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methyl- ene)aniline IR (cm-1, KBr): 1521 (C=C str.), 1590 (C=N str. (Schiff base), 1628 (C=N str., (pyra- zole moiety), 1328 (C-N str.), 962 (N-N str.), 1248 (N-O str.), 751 (C-Cl str.). 1H NMR (DMSO-d ) δ(ppm): 6.43-6.51 (1H, doublet, Ar-H), 7.14- 7.20 (1 H, tri- 6 plet, Ar-H), 7.35-8.24 (10 H, multiplet, Ar-H), 8.28 (1 H, singlet, Ar-H), 8.96 (1H, singlet, -N=CH). MS: (m/z) = 436 SA-9: (E)-2-(((3-(4-nitrophenyl)-1-phenyl-1H-pyrazol-4-yl)methylene)amino) phenol IR (cm-1, KBr): 1515 (C=C str.), 1585 (C=N str. (Schiff base), 1634 (C=N str., (pyra- zole moiety), 1332 (C-N str.), 960 (N-N str.), 1241 (N-O str.). 1H NMR (DMSO-d ) δ(ppm): 6.42-6.56 (2H, doublet, Ar-H), 7.08- 7.14 (2H, dou- 6 blet, Ar-H), 7.25-7.63 (9 H, multiplet, Ar-H), 7.68 (1 H, singlet, Ar-H), 7.92 (1H, singlet, -OH), 8.89 (1H, singlet, -N=CH). MS: (m/z) = 384 SB-1: (E)-4-((4-(dimethylamino)benzylidene)amino)1,5-dimethyl-2-phenyl-1H- pyrazol-3(2H)-one 209 Shipra Baluja, Sumitra Chanda IR (cm-1, KBr): 2930 (-C-H str. (asym.)), 2804 (-C-H str. (sym.)), 1582 (C=C str.) 1648 (C=N str.), 1648 (C=O str.), 1582 (C=N str.), 1290 (C-N str.), 973 (N-N str.). 1H NMR (DMSO-d ) δ(ppm): 2.11-2.35 (6H, singlet, C-CH ), 2.56 (3H, singlet, 6 3 C-CH ), 3.25 (3H, singlet, N-CH ), 7.05- 7.10 (2H, triplet, Ar-H), 7.35-7.39 (1H, 3 3 triplet, Ar-H),7.47-7.94 (6H, multiplet, Ar-H), 9.85 (1H, singlet, N=CH-). MS: (m/z) = 334 SB-2: (E)-4-((4-chlorobenzylidene)amino)1,5-dimethyl-2-phenyl-1H-pyrazol- 3(2H)-one IR (cm-1, KBr): 2939 (-C-H str. (asym.)), 2850 (-C-H str. (sym.)), 1571 (C=C str.) 1571 (C=N str.), 1650 (C=O str.), 1593 (C=N str.), 1290 (C-N str.), 739 (C-Cl str.). 1H NMR (DMSO-d ) δ(ppm): 1H NMR (DMSO-d ) δ(ppm) : 2.32 (3H, singlet, 6 6 C-CH ), 3.22 (3H, singlet, N-CH ), 6.91- 6.96 (2H, triplet, Ar-H), 7.20-7.29 (1H, 3 3 triplet, Ar-H),7.35-7.55 (6H, multiplet, Ar-H), 9.74 (1H, singlet, N=CH-). MS: (m/z) = 325 SB-3: (E)-4-((4-fluorobenzylidene)amino)1,5-dimethyl-2-phenyl-1H-pyrazol- 3(2H)-one IR (cm-1, KBr): 2950 (-C-H str. (asym.)), 2830 (-C-H str. (sym.)), 1568 (C=C str.) 1597 (C=N str.), 1651 (C=O str.), 1597 (C=N str.), 1454 (C-H str.), 1221 (C-F str.). 1H NMR (DMSO-d ) δ(ppm): 2.47 (3H, singlet, C-CH ), 3.15 (3H, singlet, N-CH ), 6 3 3 7.07- 7.11 (2H, triplet, Ar-H), 7.30-7.34 (1H, triplet, Ar-H),7.38-7.85 (6H, multi- plet, Ar-H), 9.70 (1H, singlet, N=CH-). MS: (m/z) = 309 SB-4: (E)-1,5-dimethyl-4-((2-nitrobenzylidene)amino)-2-phenyl-1H-pyrazol- 3(2H)-one IR (cm-1, KBr): 2945 (-C-H str. (asym.)), 2835 (-C-H str. (sym.)), 1568 (C=C str.) 1591 (C=N str.), 1647 (C=O str.), 1591 (C=N str.), 1381 (N-O str.). 1H NMR (DMSO-d ) δ(ppm): 2.48 (3H, singlet, C-CH ), 3.21 (3H, singlet, N-CH ), 6 3 3 7.28- 7.32 (2H, triplet, Ar-H), 7.38-7.40 (1H, triplet, Ar-H),7.17-7.46 (6H, multi- plet, Ar-H), 9.90 (1H, singlet, N=CH-). MS: (m/z) = 336 210