Synthesis and characterization of new 4-aryl-2-(2-oxopropoxy)-6-(2,5-dichlorothiophene)nicotinonitrile and their furo[2,3-b]pyridine derivatives: Assessment of antioxidant and biological activity European Journal of Chemistry 9 (4) (2018) 375-381 European Journal of Chemistry View Journal Online View Article Online Synthesis and characterization of new 4-aryl-2-(2-oxopropoxy)-6-(2,5- dichlorothiophene)nicotinonitrile and their furo[2,3-b]pyridine derivatives: Assessment of antioxidant and biological activity Mahmoud Al-Refai 1,*, Mohammad Ibrahim 1, Abdullah Al-Fawwaz 2 and Armin Geyer 3 1 Department of Chemistry, Faculty of Science, Al Al-Bayt University, Al-Mafraq, 25113, Jordan mahmoud_alrefai@aabu.edu.jo (M.R.), mohammadibrahim@aabu.edu.jo (M.I.) 2 Department of Biological Sciences, Faculty of Science, Al Al-Bayt University, Al-Mafraq, 25113, Jordan al_fawwaz@aabu.edu.jo (A.F.) 3 Faculty of Chemistry, Philipps University Marburg, Hans-Meerwein-Straße 4, 35032 Marburg, Germany geyer@staff.uni-marburg.de (A.G.) * Corresponding author at: Department of Chemistry, Faculty of Science, Al Al-Bayt University, Al-Mafraq, 25113, Jordan. Tel: +962.2.6297000/2141 Fax: +962.2.6297021 e-mail: mahmoud_alrefai@aabu.edu.jo (M. Al-Refai). 10.5155/eurjchem.9.4.375-381.1792 Received: 27 September 2018 Received in revised form: 18 October 2018 Accepted: 20 October 2018 Published online: 31 December 2018 Printed: 31 December 2018 A new series of furo[2,3-b]pyridine derivatives bearing aryl substituents were synthesized in two steps, where, the cyano-(2H)-pyridones (1a-l) were converted to the corresponding nicotinonitriles (2a-l), followed by the Thorpe-Ziegler ring cyclization to the furo[2,3- b]pyridine derivatives (3a-l). All new compounds were characterized by 1D-NMR experiments (1H and 13C) and 2D-NMR experiments (COSY, HMBC and HSQC), as well as ESI- MS and HR-ESI-MS data. The new compounds were screened for their antioxidant activities by 2,2-diphenyl-1-picryl-hydrazylhydrate (DPPH) free radical assay. The highest radical scavenging effect was observed for nicotinonitriles 2d, 2h and 2l and furo[2,3-b]pyridines 3b, 3f and 3j by methanolic solvent at 4.0 mg/mL concentration. Remarkably, all nicotinonitriles and furo[2,3-b]pyridine exhibited a significant radical scavenging activity after 24 and 48 hours compared with 0.5 hour. Pyridine Thiophene Antioxidant Nicotinonitrile Cyanopyridone Furo[2,3-b]pyridine Cite this: Eur. J. Chem. 2018, 9(4), 375-381 Journal website: www.eurjchem.com 1. Introduction Furo[2,3-b]pyridine containing heterocycles are of interest because they show a wide range of pharmacological activities [1-6]. The furo[2,3-b]pyridine scaffold present in known many naturally occurring and biologically active compounds [7,8]. The synthesis of pyridine containing derivatives with antimic- robial and antioxidant activities with no toxic effects on health is of growing concern to combat the resistance of pathogenic microorganisms to one or several problems of antibiotics [9- 13]. The highly reactive chemical free radicals might form in different ways in the body and have the potential to damage cells, organelles and DNA, causing cancer, and cardiovascular and neurodegenerative diseases [14,15]. Therefore, numerous methods are used to test antioxidant activities of chemical compounds. Two free radicals that are commonly used to assess antioxidant activity in vitro are 2,2-diphenyl-1-picryl hydrazyl (DPPH) and 2,2'-azino-bis(3-ethylbenzothiazoline-6- sulphonic acid) (ABTS) [16]. Many investigators have studied the free radical scavenging activity based on a fixed endpoint (e.g. 30 min) [17]. Very recently, the crystal structure and fluorescence beha- vior of new furo[2,3-b]pyridine compound and a series of a nicotinonitrile and furo[2,3-b]pyridine derivatives bearing alkyl moieties have been investigated by our research group [18,19]. Herein, we report the synthesis and characterization of a new series of nicotinonitriles and furo[2,3-b]pyridines as potent antioxidant agents (Scheme 1). 2. Experimental 2.1. Materials 4-Aryl-3-cyano-(2H)-pyridones (1a-l) were prepared according to literature procedure [20,21] and used without further purification. Solvents were dried and distilled accor- ding to standard methods. ABSTRACT RESEARCH ARTICLE KEYWORDS European Journal of Chemistry ISSN 2153-2249 (Print) / ISSN 2153-2257 (Online) – Copyright © 2018 The Authors – Atlanta Publishing House LLC – Printed in the USA. This work is published and licensed by Atlanta Publishing House LLC – CC BY NC – Some Rights Reserved. http://dx.doi.org/10.5155/eurjchem.9.4.375-381.1792 http://dx.doi.org/10.5155/eurjchem.9.4.375-381.1792 https://crossmark.crossref.org/dialog/?doi=10.5155/eurjchem.9.4.375-381.1792&domain=pdf&date_stamp=2018-12-31 http://www.eurjchem.com/ http://dx.doi.org/10.5155/eurjchem.9.4.375-381.1792 mailto:mahmoud_alrefai@aabu.edu.jo mailto:mohammadibrahim@aabu.edu.jo mailto:al_fawwaz@aabu.edu.jo mailto:geyer@staff.uni-marburg.de mailto:mahmoud_alrefai@aabu.edu.jo http://www.eurjchem.com/ https://crossmark.crossref.org/dialog/?doi=10.5155/eurjchem.9.4.375-381.1792&domain=pdf&date_stamp=2018-12-31� 376 Al-Refai et al. / European Journal of Chemistry 9 (4) (2018) 375-381 i Br h gcb f Ar = F Cl Br lk e j O O O Br a d N H S Cl Cl Ar 1a-l CN O N S Cl Cl Ar 2a-l CN O O O NH2 ON S Cl Cl R 3a-l 1. K2CO3, DMF r.t., 1 h MeONa, MeOH reflux, 3-8 hO Cl2. ,1 h Scheme 1. Synthesis of nicotinonitriles (2a-l) and furo[2,3-b]pyridines (3a-l). 2.2. Instrumentation All purchased reagents were used as received. Solvents were dried and distilled according to standard protocols. NMR spectra, 1H (300 and 500 MHz) and 13C (75 and 125 MHz), were recorded in CDCl3 (used as an internal standard) at 300 K on Bruker spectrometers (Bruker, Germany). Chemical shifts, δ (ppm), were determined from the centre of the respective coupling pattern (s: singlet, d: doublet, dd: doublet of doublet, t: triplet). ESI-MS and HR-ESI-MS measurements were carried out on a LTQ-FT mass spectrometer (Thermo Fisher Scientific). The reactions were monitored using thin layer chromatography (TLC) using analytical TLC plates coated with silica gel (60F254, Merck). Preparative thin layer chromatography (PTLC) was performed at room temperature using CHCl3 as the mobile phase. 2.3. Synthesis 2.3.1. General procedure for the synthesis of nicotinonitriles (2a-l) The 4-aryl-3-cyano-(2H)-pyridone (1) (2.0 mmol), and K2CO3 (2.2 mmol, 1.1 equiv.) were dissolved in DMF (50 mL, dry) in a round-bottom flask equipped with a stir bar. The reaction mixture was stirred at the ambient temperature for 1 h. Chloroacetone (4.0 mmol, 2.0 equiv.) was added portion wise to the mixture and allowed to stir overnight. The reaction mixture was transferred to ice water. The precipitate formed was filtered off, washed with water, dried and then subjected to PTLC chromatography using chloroform as mobile phase to give the corresponding nicotinonitrile (2) (Scheme 1). 2-(2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-phenyl pyridine-3-carbonitrile (2a): Color: White solid. Yield: 97%. 1H NMR (300 MHz, CDCl3, δ, ppm): 2.33 (s, 3H, CH3COCH2O), 5.08 (s, 2H, CH3COCH2O), 7.21 (s, 1H, H-4'), 7.57 (m, 3H, H-3", 4", 5"), 7.58 (m, 2H, H-6", 2"), 7.66 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.4 (CH3COCH2O), 71.2 (CH3COCH2O), 93.8 (C- 3), 114.7 (CN-3), 116.8 (CH-5), 127.1 (CH-4'), 128.4 (CH-6", 2"), 129.2 (CH-3", 5"), 130.4 (CH-4"), 135.1 (Cq-, 3'), 135.7 (Cq- 1"), 151.5 (Cq-6), 157.1 (Cq-4), 163.2 (Cq-2), 203.0 (CO). MS (+ESI, m/z (%)): 403 ([M+H]+, 100), 405 ([M+H+2]+, 66), 407 ([M+H+4]+, 15), 425 ([M+Na]+, 84), 427 ([M+Na+2]+, 55), 429 ([M+Na+4]+, 13), 827 ([2M+Na]+, 15), 829 ([2M+Na+2]+, 22), 830 ([2M+Na+4]+, 14). HRMS (+ESI, m/z): 403.0070 [M+H]+, 405.0040 [M+H+2]+, 407.0012 [M+H+4]+, (calcd. for C19H13 Cl2N2O2S, 403.0069). HRMS (+ESI, m/z): 424.9889 [M+Na]+, 426.9860 [M+Na+2]+, 428.9831 [M+Na+4]+, (calcd. for C19H12 Cl2N2O2SNa, 424.9889). (2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-o-tolylpyri dine-3-carbonitrile (2b): Color: White solid. Yield: 93%. 1H NMR (300 MHz, CDCl3, δ, ppm): 2.29 (s, 3H, CH3-2"), 2.31 (s, 3H, CH3COCH2O), 5.06 (s, 2H, CH3COCH2O), 7.19 (s, 1H, H-4'), 7.33 (m, 4H, H-3", 4", 5", 6"), 7.50 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 19.9 (CH3-2"), 26.3 (CH3COCH2O), 71.2 (CH3COCH2O), 95.7 (Cq-3), 114.0 (CN-3), 117.5 (CH-5), 126.2, 127.1, 128.7, 129.8, 131.0 (CH-4', 3", 4", 5", 6"), 134.0, 135.1, 135.2, 135.6 (Cq-2', 3', 5', 1'', 2"), 151.3 (Cq-6), 158.3 (Cq-4), 162.7 (Cq-2), 202.8 (CO). MS (+ESI, m/z (%)): 417 ([M+H]+, 34), 419 ([M+H+2]+, 24), 421 ([M+H+24]+, 8), 439 ([M+Na]+, 100), 441 ([M+Na+2]+, 66), 443 ([M+Na+4]+, 15), 855 ([2M+ Na]+, 18), 857 ([2M+Na+2]+, 30), 859 ([2M+Na+4]+, 17). HRMS (+ESI, m/z): 439.0047 [M+Na]+, 441.0017 [2M+Na+4]+, (calcd. for C20H14Cl2N2O2SNa, 439.0045). 2-(2-Oxopropoxy)-4-(4-tert-butylphenyl)-6-(2, 5-dichloro thiophen-3-yl)pyridine-3-carbonitrile (2c): Color: White solid. Yield: 89%. 1H NMR (500 MHz, CDCl3, δ, ppm): 1.39 (s, 9H, C(CH3)3), 2.32 (s, 3H, COCH3), 5.02 (s, 2H, CH3COCH2O), 7.20 (s, 1H, H-4'), 7.58 (d, J = 8.7 Hz, 2H, H-3", 5"), 7.63 (d, J = 8.7, 5.4 Hz, 2H, H-2", 6"), 7.66 (s, 1H, H-5). 13C NMR (125 MHz, CDCl3, δ, ppm): 26.4 (COCH3), 31.2 (C(CH3)3), 34.9 (C(CH3)3), 71.2 (CH3COCH2O), 93.5 (Cq-3), 115.0 (CN), 116.7 (CH-5), 126.2 (CH-3", 5"), 126.5, 127.0 (Cq-2', 5'), 128.2 (CH-4', 2", 6"), 132.7 (Cq-1''), 135.1 (Cq-3'), 151.3 (Cq-6), 153.9 (Cq-4''), 157.0 (Cq-4), 162.6 (Cq-2), 203.1 (CO). MS (+ESI, m/z (%)): 460 ([M+H]+, 24), 462 ([M+H+2]+, 18), 482 ([M+Na]+, 63), 484 ([M+Na+2]+, 41), 941 ([2M+Na]+, 100), 943 ([2M+Na+2]+, 43). HRMS (+ESI, m/z): 481.0517 [M+Na]+, 483.0488 [M+Na+2]+, (calcd. for C23H20Cl2N2O2SNa, 481.0515). 2-(2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-(4-met- hoxyphenyl)pyridine-3-carbonitrile (2d): Color: White solid. Yield: 93%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.22 (s, 3H, CH3COCH2O), 3.82 (s, 3H, OCH3-4"), 4.97 (s, 2H, CH3COCH2O), 6.99 (d, J = 8.8 Hz, 2H, H-3", 5"), 7.11 (s, 1H, H-4'), 7.54 (s, 1H, H-5), 7.57 (d, J = 8.8 Hz, 2H, H-2", 6"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.5 (CH3COCH2O), 55.5 (OCH3-4"), 71.2 (CH3CO 2018 – European Journal of Chemistry – CC BY NC – DOI: 10.5155/eurjchem.9.4.375-381.1792 Al-Refai et al. / European Journal of Chemistry 9 (4) (2018) 375-381 377 CH2O), 93.2 (C-3), 114.6 (CH-3", 5"), 115.1 (CN-3), 116.5 (CH- 5), 127.1 (CH-4'), 130.0 (CH-2", 6"), 127.0, 127.9, 135.2, (Cq-2', 3', 5', 1''), 151.2 (Cq-6), 156.7 (Cq-4), 161.5 (Cq-4"), 163.4 (Cq- 2), 203.1 (CO). MS (+ESI, m/z (%)): 455 ([M+Na]+, 100), 457 ([M+Na+2]+, 66), 459 ([M+Na+4]+, 15), 887 ([2M+Na]+, 21), 889 ([2M+Na+2]+, 30), 891 ([2M+Na+4]+, 20). HRMS (+ESI, m/z): 454.9994 [M+Na]+, 456.9965 [M+Na+2]+, (calcd. for C20H14Cl2N2O3SNa, 454.9994). 2-(2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-(3-met- hoxyphenyl)pyridine-3-carbonitrile (2e): Color: White solid. Yield: 95%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.33 (s, 3H, CH3COCH2O), 3.92 (s, 3H, OCH3-3"), 5.09 (s, 2H, CH3COCH2O), 7.10 (ddd, J = 8.3, 2.4, 0.7 Hz, 1H, H-4"), 7.21 (t, J = 2.3 Hz, 1H, H-2"), 7.22 (s, 1H, H-4'), 7.25 (ddd, J = 7.6, 1.8, 1.1 Hz, 1H, H- 6"), 7.49 (t, J = 7.9 Hz, 1H, H-5"), 7.66 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.4 (CH3COCH2O), 55.5 (OCH3-3"), 71.2 (CH3COCH2O), 93.8 (Cq-3), 113.9 (CH-2"), 114.6 (CN-3), 116.1 (CH-5), 116.7 (CH-4"), 120.7 (CH-6"), 127.0 (CH-4'), 130.3 (CH-5"), 135.1 (Cq-1"), 136.9 (Cq-3'), 151.4 (Cq-6), 157.0 (Cq-4), 159.9 (Cq-3"), 163.2 (Cq-2), 202.9 (CO). MS (+ESI, m/z (%)): 455 ([M+Na]+, 100), 457 ([M+Na+2]+, 65), 459 ([M+Na+24]+, 15), 887 ([2M+Na]+, 20), 889 ([2M+Na+2]+, 33), 891 ([2M+ Na+4]+, 21). HRMS (+ESI, m/z): 454.9996 [M+Na]+, 456.9967 [M+Na+2]+, (calcd. for C20H14Cl2N2O3SNa, 454.9994). 2-(2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-(4-fluoro phenyl)pyridine-3-carbonitrile (2f): Color: White solid. Yield: 82%. 1H NMR (500 MHz, DMSO-d6, δ, ppm): 2.18 (s, 3H, COCH3), 5.30 (s, 2H, CH3COCH2O), 7.46 (t, J = 8.9 Hz, 2H, H-3", 5"), 7.71 (s, 1H, H-5), 7.78 (s, 1H, H-4'), 7.84 (dd, J = 8.9, 5.35 Hz, 2H, H-2", 6"). 13C NMR (125 MHz, DMSO-d6, δ, ppm): 26.4 (COCH3), 71.5 (CH3COCH2O), 115.3 (CN), 93.2 (Cq-3), 116.6, 116.4 (CH-3", 5"), 117.1 (CH-5), 127.0, 126.2 (Cq-2', 5'), 128.6 (CH-4'), 131.7 (CH-2", 6"), 132.2 (Cq-1''), 135.1(Cq-3'), 151.5 (Cq-6), 155.9 (Cq-4), 162.8, 163.3 (Cq-4''), 164.8 (Cq-2), 202.8 (CO). MS (+ESI, m/z (%)): 421 ([M+H]+, 68), 423 ([M+H+2]+, 41), 443 ([M+Na]+, 100), 445 ([M+Na+2]+, 67), 863 ([2M+Na]+, 30), 865 ([2M+Na+2]+, 46), 867 ([M+Na+4]+, 24). HRMS (+ESI, m/z): 442.9801 [M+Na]+, 444.9771 [M+Na+2]+, (calcd. for C19H11Cl2FN2O2SNa, 442.9795). 2-(2-Oxopropoxy)-4-(4-chlorophenyl)-6-(2, 5-dichlorothio phen-3-yl)pyridine-3-carbonitrile (2g): Color: White solid. Yield: 86%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.32 (s, 3H, CH3COCH2O), 5.08 (s, 2H, CH3COCH2O), 7.21 (s, 1H, H-4'), 7.56 (d, J = 8.7 Hz, 2H, H-3", 5"), 7.62 (d, J = 8.7 Hz, 2H, H-2", 6"), 7.61 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.3 (CH3COCH2O), 71.2 (CH3COCH2O), 93.7 (C-3), 114.4 (CN-3), 116.4 (CH-5), 127.0 (CH-4'), 129.5 (CH-3", 5"), 129.7 (CH-2", 6"), 126.7, 127.2, 134.0, 135.0 (Cq-2', 5', 1'', 4"), 136.9 (Cq-3'), 151.7 (Cq-6), 155.8 (Cq-4), 163.3 (Cq-2), 202.5 (CO). MS (+ESI, m/z (%)): 437 ([M+H]+, 23), 439 ([M+H+2]+, 21), 441 ([M+H +24]+, 8), 459 ([M+Na]+, 97), 461 ([M+Na+2]+, 100), 463 ([M+ Na+4]+, 36), 895 ([2M+Na]+, 21), 897 ([2M+Na+2]+, 39), 899 ([2M+Na+4]+, 34). HRMS (+ESI, m/z): 458.9501 [M+Na]+, 460.9471 [M+Na+2]+, 462.9442 [M+Na+4]+, (calcd. for C19H11 Cl3N2O2SNa, 458.9499). 2-(2-Oxopropoxy)-4-(2-bromophenyl)-6-(2, 5-dichlorothio phen-3-yl)pyridine-3-carbonitrile (2h): Color: White solid. Yield: 79%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.34 (s, 3H, CH3COCH2O), 5.08 (s, 2H, CH3COCH2O), 7.22 (s, 1H, H-4'), 7.40 (m, 2H, H-3", 4"), 7.50 (td, J = 7.9, 1.1 Hz, 1H, H-5"), 7.59 (s, 1H, H-5), 7.77 (d, J = 8.2 Hz, 1H, H-6''). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.4 (CH3COCH2O), 71.2 (CH3COCH2O), 95.6 (Cq-3), 113.8 (CN-3), 117.7 (CH-5), 121.7 (Cq-2"), 127.0 (CH-4'), 127.9, 130.4, 131.3, 133.6 (CH-3", 4", 5", 6"), 134.9 (Cq-1"), 136.6 (Cq- 3'), 151.3 (Cq-6), 156.5 (Cq-4), 162.5 (Cq-2), 202.9 (CO). MS (+ESI, m/z (%)): 481 ([M+H]+, 18), 483 ([M+H+2]+, 27), 485 ([M+H+24]+, 13), 503 ([M+Na]+, 68), 505 ([M+Na+2]+, 100), 507 ([M+Na+4]+, 52). HRMS (+ESI, m/z): 502.8994 [M+Na]+, 504.8970 [M+Na+2]+, 506.8943 [M+Na+4]+, (calcd. for C19H11 BrCl2N2O2SNa, 502.8994). 2-(2-Oxopropoxy)-4-(3-bromophenyl)-6-(2, 5-dichlorothio phen-3-yl)pyridine-3-carbonitrile (2i): Color: White solid. Yield: 81%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.32 (s, 3H, CH3COCH2O), 5.09 (s, 2H, CH3COCH2O), 7.20 (s, 1H, H-4'), 7.45 (t, J = 7.9 Hz, 1H, H-5"), 7.60 (1H, s, H-5), 7.63 (ddd, J = 7.8, 1.7, 1.0 Hz, 1H, H-6"), 7.70 (ddd, J = 8.04, 1.89, 1.0 Hz, 1H, H-4"), 7.77 (d, J = 1.8 Hz, 1H, H-2"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.4 (CH3COCH2O), 71.2 (CH3COCH2O), 93.8 (Cq-3), 114.3 (CN- 3), 116.5 (CH-5), 123.2 (Cq-3"), 126.9 (CH-6"), 127.0 (CH-4'), 130.7 (CH-5"), 131.3 (CH-2"), 133.4 (CH-4"), 134.9 (Cq-, 3'), 137.6 (Cq-1"), 151.8 (Cq-6), 155.4 (Cq-4), 163.2 (Cq-2), 202.6 (CO). MS (+ESI, m/z (%)): 503 ([M+Na]+, 67), 505 ([M+Na+2]+, 100), 507 ([M+Na+4]+, 46). HRMS (+ESI, m/z): 502.8994 [M+Na]+, 504.8970 [M+Na+2]+, 506.8943 [M+Na+4]+, (calcd. for C19H11BrCl2N2O2SNa, 502.8994). 2-(2-Oxopropoxy)-4-(3-bromo-4-methoxyphenyl)-6-(2, 5-di chlorothiophen-3-yl)pyridine-3-carbonitrile (2j): Color: White solid. Yield: 82%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.22 (s, 3H, CH3COCH2O), 3.91 (s, 3H, OCH3-4"), 4.98 (s, 2H, CH3CO CH2O), 6.99 (d, J = 8.9 Hz, 1H, H-5"), 7.11 (s, 1H, H-4'), 7.50 (s, 1H, H-5), 7.60 (dd, J = 8.6, 2.3 Hz, 1H, H-6"), 7.74 (d, J = 2.3 Hz, 1H, H-2"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.3 (CH3CO CH2O), 56.5 (OCH3-4"), 71.2 (CH3COCH2O), 93.4 (Cq-3), 112.0 (CH-5"), 112.5 (Cq-3"), 114.7 (CN-3), 116.3 (CH-5), 127.0 (CH- 4'), 129.0 (CH-6"), 129.1 (Cq-1"), 133.2 (CH-2"), 137.2 (Cq-3'), 151.5 (Cq-6), 155.2 (Cq-4), 157.6 (Cq-4"), 163.3 (Cq-2), 202.7 (CO). MS (+ESI, m/z (%)): 533 ([M+Na]+, 67), 535 ([M+Na+2]+, 100), 537 ([M+Na+24]+, 47). HRMS (+ESI, m/z): 532.9100 [M+Na]+, 534.9075 [M+Na+2]+, 536.9048 [M+Na+4]+, (calcd. for C20H13BrCl2N2O3SNa, 532.9100). 2-(2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-(naph thalen-1-yl)pyridine-3-carbonitrile (2k): Color: White solid. Yield: 84%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.33 (s, 3H, CH3COCH2O), 5.11 (s, 2H, CH3COCH2O), 7.14 (s, 1H, H-4'), 7.55- 7.98 (m, 8H, H-5, 2", 3", 4", 6", 7", 8", 9"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.4 (CH3COCH2O), 71.2 (CH3COCH2O), 96.2 (C- 3), 114.1 (CN-3), 118.5 (C-5), 126.4, 127.1, 128.1, 133.4, 133.8, 135.0 (Cq-2', 3', 5', 1", 5'', 10''), 124.6, 125.2, 126.6, 130.4, 127.1, 127.2, 127.3, 128.8 (CH-4', 2'', 3'', 4'', 6'', 7'', 8'', 9'''), 151.0 (Cq-6), 156.8 (Cq-4), 163.0 (Cq-2), 202.8 (CO). MS (+ESI, m/z (%)): 453 ([M+H]+, 20), 455 ([M+H+2]+, 13), 475([M+Na]+, 100), 477 ([M+Na+2]+, 64), 479 ([M+Na+24]+, 14), 927 ([2M+ Na]+, 32), 929 ([2M+Na+2]+, 45), 931 ([2M+Na+4]+, 26). HRMS (+ESI, m/z): 475.0047 [M+Na]+, 477.0017 [M+Na+2]+, (calcd. for C23H14Cl2N2O2SNa, 475.0045). 2-(2-Oxopropoxy)-6-(2, 5-dichlorothiophen-3-yl)-4-(naph thalen-6-yl)pyridine-3-carbonitrile (2l): Color: White solid. Yield: 76%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.25 (s, 3H, CH3COCH2O), 5.01 (s, 2H, CH3COCH2O), 7.15 (s, 1H, H-4'), 7.49- 7.96 (m, 7H, H- 5, 4", 5", 6", 7", 9", 10"), 8.09 (d, J =1.5 Hz, 1H, H-2"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.4 (CH3COCH2O), 71.2 (CH3COCH2O), 93.9 (C-3), 114.8 (CN-3), 117.0 (C-5), 123.9, 125.2, 133.0, 133.9, 135.1(Cq-2', 3', 5', 1", 3'', 8''), 127.0, 127.1, 127.7, 127.8, 128.6, 128.8, 129.1(CH-4', 2'', 4'', 5'', 6'', 7'', 9'', 10''), 151.4 (Cq-6), 157.1 (Cq-4), 163.3 (Cq-2), 202.9 (CO). MS (+ESI, m/z (%)): 453 ([M+H]+, 18), 455 ([M+H+2]+, 11), 475 ([M+Na]+, 100), 477 ([M+Na+2]+, 72), 479 ([M+Na+ 24]+, 14), 927 ([2M+Na]+, 26), 929 ([2M+Na+2]+, 40), 931 ([2M+Na+4]+, 25). HRMS (+ESI, m/z): 475.0047 [M+Na]+, 477.0017 [M+Na+2]+, (calcd. for C23H14Cl2N2O2SNa, 475.0045). 2.3.2. General procedure for the synthesis of furo[2, 3-b] pyridines (3a-l) The nicotinonitrile (2) (1.0 mmol), was dissolved in methanolic solution of a sodium methoxide, the resulting solution was heated under reflux for 3 h. After being cooled to 2018 – European Journal of Chemistry – CC BY NC – DOI: 10.5155/eurjchem.9.4.375-381.1792 378 Al-Refai et al. / European Journal of Chemistry 9 (4) (2018) 375-381 room temperature, the resulting solid was filtered off, washed with cold methanol and dried. The product was subjected to PTLC chromatography using the chloroform as mobile phase to afford the corresponding furo[2, 3-b]pyridine compound 3 (Scheme 1). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-phenylfuro[2, 3-b]pyridin-2-yl)ethanone (3a): Color: Yellow solid. Yield: 79%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.59 (s, 3H, CH3CO CH2O), 5.21 (bs, 2H, NH2), 7.52 (s, 1H, H-4'), 7.62 (m, 5H, H-2", 3", 4", 5", 6"), 7.81 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.2 (CH3CO), 109.7 (Cq-3a), 118.9 (CH-5), 127.9 (CH- 4'), 128.4 (CH-2", 6"), 125.0, 126.8, 137.6 (Cq-3, 2', 5', ), 129.1 (CH-3", 5"), 129.8 (CH-4"), 133.9 (Cq-2), 136.2 (Cq-1''), 136.5 (Cq-3'), 148.2 (Cq-4), 150.9 (Cq-6), 159.9 (Cq-7a), 190.1 (CO). MS (+ESI, m/z (%)): 403 ([M+H]+, 100), 405 ([M+H+2]+, 68), 407 ([M+H+4]+, 58), 425 ([M+Na]+, 23), 427 ([M+Na+2]+, 16), 429 ([M+Na+4]+, 11). HRMS (+ESI, m/z): 403.0071 [M+H]+, 405.0041 [M+H+2]+, 407.0021 [M+H+4]+ , (calcd. for C19H13Cl2 N2O2S, 403.0069). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-o-tolylfuro[2, 3-b]pyridin-2-yl)ethanone (3b): Color: Yellow solid. Yield: 81%. 1H NMR (300 MHz, CDCl3, δ, ppm): 2.55 (s, 6H, CH3CO, CH3-2"), 5.20 (bs, 2H, NH2), 7.43 (d, J = 7.1 Hz, 1H, H-3''), 7.50 (m, 3H, H- 4', 4'', 5''), 7.75 (s, 1H, H-5), 7.78 (d, J = 7.8 Hz, 1H, H-6''). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.2 (CH3CO, CH3-2"), 110.6 (Cq- 3a), 119.1 (CH-5), 128.0, 130.3, 131.1, 133.6 (CH-4', 3'', 4'', 5'', 6"), 122.2, 125.2, 126.7, 134.1, 136.5, 136.4 (Cq-2, 3, , 2', 5', 1'', 2''), 137.0 (Cq-3'), 146.1 (Cq-4), 150.8 (Cq-6), 159.5 (Cq-7a), 190.2 (CO). 1-(4-(4-Tert-butylphenyl)-3-amino-6-(2,5-dichlorothiophen -3-yl)furo[2, 3-b]pyridin-2-yl)ethanone (3c): Color: Yellow solid. Yield: 86%. 1H NMR (500 MHz, DMSO-d6, δ, ppm): 1.37 (s, 9H, C(CH3)3), 2.46 (s, 3H, COCH3), 5.87 (bs, 2H, NH2), 7.65 (s, 4H, H-2", 3", 5", 6"), 7.74 (s, 1H, H-4'), 7.75 (s, 1H, H-5). 13C NMR (125 MHz, DMSO-d6, δ, ppm): 26.7 (COCH3), 31.5 (C (CH3)3), 35.1 (C(CH3)3), 109.8 (Cq-3a), 119.6 (CH-5), 125.1, 125.9 (Cq-2', 5'), 126.7 (CH-3", 5"), 128.8 (CH-4', 2", 6"), 133.2 (Cq-1''), 133.4 (Cq-2), 136.7 (Cq-3'), 148.7 (Cq4), 150.4 (Cq-6), 152.8 (C-4''), 159.8 (Cq-7a), 188.8 (CO). MS (+ESI, m/z (%)): 464 ([M+H+4]+, 100), 941 ([2M+Na]+, 16). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-(4-methoxy phenyl)furo[2, 3-b]pyridin-2-yl)ethanone (3d): Color: Yellow solid. Yield: 88%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.55 (s, 3H, CH3CO), 3.91 (s, 3H, OCH3-4"), 5.59 (bs, 2H, NH2), 7.10 (d, J = 8.6Hz, 2H, H-3", 5"), 7.48 (s, 1H, H-4'), 7.54 (d, J = 8.6 Hz, 2H, H-2", 6"), 7.74 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.2 (CH3CO), 55.5 (OCH3-4"), 109.7 (Cq-3a), 114.8 (CH-3", 5"), 118.8 (CH-5), 128.0 (CH-4'), 129.8 (CH-2", 6"), 124.9, 126.7, 128.3, 133.8, 136.6 (Cq-2, 3, 2', 5', 1''), 137.6 (Cq-3'), 148.0 Cq- 4), 150.8 (Cq-6), 160.0 (Cq-4"), 160.9 (Cq-7a), 190.1 (CO). MS (+ESI, m/z (%)): 455 ([M+Na]+, 100), 457 ([M+Na+2]+, 67), 459 ([M+Na+24]+, 23), 887 ([2M+Na]+, 46), 889 ([2M+Na+2]+, 48), 891 ([2M+Na+4]+, 37). HRMS (+ESI, m/z): 454.9995 [M+Na]+, 456.9966 [M+Na+2]+, 458.9941 [M+Na+4]+, (calcd. for C20H14Cl2N2O3SNa, 454.9994). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-(3-methoxy phenyl)furo[2, 3-b]pyridin-2-yl)ethanone (3e): Color: Yellow solid. Yield: 90%. 1H NMR (300 MHz, CDCl3, δ, ppm): 2.55 (s, 3H, CH3CO), 3.89 (s, 3H, OCH3-3"), 5.60 (bs, 2H, NH2), 7.08 (d, J = 7.6 Hz, 2H, H-2", 4''), 7.09 (s, 1H, H-4'), 7.15 (d, J = 7.6, 1.8, 1.1 Hz, 1H, H-6"), 7.49 (t, J = 8.47 Hz, 1H, H-5"), 7.87 (s, 1H, H- 5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.2 (CH3CO), 55.5 (OCH3-3"), 109.7 (Cq-3a), 114.0 (CH-2"), 115.3 (CH-4"), 118.8 (CH-5), 120.5 (CH-6"), 127.9 (CH-4'), 130.5 (CH-5"), 126.8, 125.0, 136.5, 137.4 (Cq-2, 3, 2', 5', 1'' ), 137.5 (Cq-3'), 147.7 (Cq- 4), 150.8 (Cq-6), 159.5 (Cq-3"), 160.2 (Cq-7a), 190.1 (CO). MS (+ESI, m/z (%)): 433 ([M+H]+, 51), 435 ([M+H+2]+, 34), 437 ([M+H+4]+, 11), 455 ([M+Na]+, 100), 457 ([M+Na+2]+, 67), 459 ([M+Na+24]+, 15), 887 ([2M+Na]+, 37), 889 ([2M+Na+2]+, 48), 891 ([2M+Na+4]+, 30). HRMS (+ESI, m/z): 454.9996 [M+Na]+, 456.9966 [M+Na+2]+, (calcd. for C20H14Cl2N2O3SNa, 454.9994). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-(4-fluoro phen- yl)furo[2, 3-b]pyridin-2-yl)ethanone (3f): Color: Yellow solid. Yield: 71%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.45 (s, 3H, COCH3), 5.91 (bs , 2H, NH2), 7.46 (t, J = 8.9 Hz, 2H, H-3", 5"), 7.74 (s, 2H, H-4', 5), 7.75 (dd, J = 8.9, 5.4 Hz, 2H, H-2", 6"). 13C NMR (125 MHz, DMSO-d6, δ, ppm): 26.6 (COCH3), 110.0 (Cq- 3a), 116.7, 116.1 (CH-3", 5"), 119.7 (CH-5), 125.2, 126.0, 133.5, (Cq-2, 2', 5'), 128.9 (CH-4'), 131.5 (CH-2", 6"), 136.8 (Cq-1''), 137.9 (Cq-3'), 147.73 (Cq-4), 150.5 (Cq-6), 159.7 (Cq-7a), 162.5, 162.9, (Cq-4''), 189.0 (CO). MS (+ESI, m/z (%)): 421 ([M+H]+, 100), 423 ([M+H+2]+, 70), 425 ([M+H+4]+, 25), 443 ([M+Na]+, 82), 445 ([M+Na+2]+, 58), 863 ([2M+Na]+, 30), 865 ([2M+ Na+2]+, 50), 867 ([M+Na+4]+, 30). HRMS (+ESI, m/z): 442.9802 [M+Na]+, 444.9773 [M+Na+2]+, (calcd. for C19H11Cl2F N2O2SNa, 442.9795). 1-(3-Amino-4-(4-chlorophenyl)-6-(2, 5-dichlorothiophen-3- yl)furo[2, 3-b]pyridin-2-yl)ethanone (3g): Color: Yellow solid. Yield: 75%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.59 (s, 3H, CH3CO), 5.47 (bs, 2H, NH2), 7.52 (s, 1H, H-4'), 7.57 (d, J = 8.7 Hz, 2H, H-3", 5"), 7.60 (d, J = 8.7Hz, 2H, H-2", 6"), 7.78 (s, 1H, H- 5). 13C NMR (125 MHz, CDCl3, δ, ppm): 26.3 (CH3CO), 109.7 (Cq- 3a), 118.8 (CH-5), 129.7 (CH-3", 5"), 127.9 (CH-4'), 129.8 (CH- 2", 6"), 134.1, 134.6, 134.6, 136.3, 136.4 (Cq-2, 3, 2', 5', 1'', 4''), 137.0 (Cq-3'), 146.7 (Cq-4), 152.8 (Cq-6), 159.9 (Cq-7a), 190.3 (CO). MS (+ESI, m/z (%)): 437 ([M+H]+, 33), 439 ([M+H+2]+, 36), 441 ([M+H+24]+, 82). 1-(3-Amino-4-(2-bromophenyl)-6-(2, 5-dichlorothiophen-3- yl)furo[2, 3-b]pyridin-2-yl)ethanone (3h): Color: Yellow solid. Yield: 73%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.56 (s, 3H, CH3CO), 5.21 (bs, 2H, NH2), 7.45 (m, 2H, H-3", 4"), 7.53 (m, 2H, H-4', 5"), 7.77 (s, 1H, H-5), 7.82 (dd, J = 8.8 Hz, 1H, H-6''). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.3 (CH3CO), 110.6 (Cq-3a), 119.2 (CH-5), 122.2 (Cq-2"), 128.0, 130.4, 131.1, 133.6 (CH-4', 3", 4", 5", 6"), 126.8, 134.1, 136.4, 136.5, (Cq-2, 3, 2', 5', 1"), 137.0 (Cq-3'), 146.1 (Cq-4), 150.8 (Cq-6), 159.5 (Cq-7a), 190.2 (CO). MS (+ESI, m/z (%)): 481 ([M+H]+, 44), 483 ([M+H+2]+, 65), 485 ([M+H+24]+, 36), 503 ([M+Na]+, 66), 505 ([M+Na+2]+, 100), 507 ([M+Na+4]+, 52). HRMS (+ESI, m/z): 502.8995 [M+Na]+, 504.8971 [M+Na+2]+, 506.8943 [M+Na+4]+, (calcd. for C19H11BrCl2N2O2SNa, 502.8994). 1-(3-Amino-4-(3-bromophenyl)-6-(2, 5-dichlorothiophen-3- yl)furo[2, 3-b]pyridin-2-yl)ethanone (3i): Color: Yellow solid. Yield: 68%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.59 (s, 3H, CH3CO), 5.50 (bs, 2H, NH2), 7.49 (t, J = 7.8 Hz, 1H, H-5"), 7.51 (s, 1H, H-4'), 7.56 (ddd, J = 7.6, 1.6, 1.2 Hz, 1H, H-6"), 7.73 (ddd, J = 8.0, 1.9, 1.1 Hz, 1H, H-4"), 7.77 (t, J = 1.7 Hz, 1H, H-2"), 7.78 (s, 1H, H-5). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.3 (CH3CO), 109.7 (Cq-3a), 118.8 (CH-5), 123.5 (Cq-3"), 127.0 (CH-6"), 127.9 (CH-4'), 125.2, 126.9, 137.0 (Cq-3, 2', 5', ), 130.8 (CH-5"), 131.4 (CH-2"), 132.8 (CH-4"), 137.0 (Cq-3'), 138.1 (Cq-1''), 146.3 (Cq-4), 151.0 (Cq-6), 159.8 (Cq-7a), 190.3 (CO). MS (+ESI, m/z (%)): 481 ([M+H]+, 36), 483 ([M+H+2]+, 58), 503 ([M+Na]+, 60), 505 ([M+Na+2]+, 100), 507 ([M+Na+4]+, 66), 985 ([2M+Na]+, 47), 987 ([2M+Na+2]+, 63), 989 ([2M+Na+4]+, 53). 1-(3-Amino-4-(3-bromo-4-methoxyphenyl)-6-(2, 5-dichloro thiophen-3-yl)furo[2, 3-b]pyridin-2-yl)ethanone (3j): Color: Yellow solid. Yield: 66%. 1H NMR (300 MHz, CDCl3, δ, ppm): 2.22 (s, 3H, CH3CO), 3.92 (s, 3H, OCH3-4"), 4.98 (bs, 2H, NH2), 6.99 (d, J = 8.6 Hz, 1H, H-5"), 7.11 (s, 1H, H-4'), 7.50 (s, 1H, H- 5), 7.60 (dd, J = 8.6, 2.3 Hz, 1H, H-6"), 7.74 (d, J = 2.28 Hz, 1H, H-2"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.3 (CH3CO), 56.5 (OCH3-4"), 109.6 (Cq-3a), 112.3 (CH-5"), 112.8 (Cq-3"), 118.8 (CH-5), 127.9 (CH-4'), 128.7 (CH-6"), 133.3 (CH-2"), 126.8, 129.6, 134.0, 136.4 (Cq-2, 3, 2', 5', 1''), 137.2 (Cq-3'), 146.3 (Cq- 4), 150.9 (Cq-6), 157.2 (Cq-4"), 159.9 (Cq-7a), 190.2 (CO). MS (+ESI, m/z (%)): 533 ([M+Na]+, 70), 535 ([M+Na+2]+, 100), 2018 – European Journal of Chemistry – CC BY NC – DOI: 10.5155/eurjchem.9.4.375-381.1792 Al-Refai et al. / European Journal of Chemistry 9 (4) (2018) 375-381 379 537 ([M+Na+24]+, 65). HRMS (+ESI, m/z): 532.9101 [M+Na]+, 534.9077 [M+Na+2]+, 536.9050 [M+Na+4]+, 538.9026 [M+Na+ 6]+, (calcd. for C20H13BrCl2N2O3SNa, 532.9100). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-(naphthalen-1- yl)furo[2, 3-b]pyridin-2-yl)ethanone (3k): Color: Yellow solid. Yield: 63%. 1H NMR (500 MHz, CDCl3, δ, ppm): 2.55 (s, 3H, CH3CO), 5.03 (s, 2H, NH2), 7.54 (s, 1H, H-4'), 7.59-8.02 (m, 8H, H-5, 2", 3", 4", 6", 7", 8", 9"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.2 (CH3CO), 111.4 (Cq-3a), 120.0 (CH-5), 125.1, 125.3, 126.9, 127.0, 127.5, 128.0, 128.7, 130.0 (CH-4', 2'', 3'', 4'', 6'', 7'', 8'', 9'''), 126.8, 126.8, 130.7, 133.2, 133.7, 134.0, 136.5, 137.3 (Cq- 2, 3, 2', 3', 5', 1'', 3'', 8''), 146.3 (Cq-4), 150.8 (Cq-6), 159.7 (Cq- 7a), 189.7 (CO). MS (+ESI, m/z (%)): 453 ([M+H]+, 62), 455 ([M+H+2]+, 44), 457 ([M+H+4]+, 18), 475 ([M+Na]+, 100), 477 ([M+Na+2]+, 77), 479 ([M+Na+24]+, 18), 927 ([2M+Na]+, 44), 929 ([2M+Na+2]+, 55), 931 ([2M+Na+4]+, 41). HRMS (+ESI, m/z): 475.0048 [M+Na]+, 477.0018 [M+Na+2]+, (calcd. for C23H14Cl2N2O2SNa, 475.0045). 1-(3-Amino-6-(2, 5-dichlorothiophen-3-yl)-4-(naphthalen-3- yl)furo[2, 3-b]pyridin-2-yl)ethanone (3l): Color: Yellow solid. Yield: 67%. 1H NMR (300 MHz, CDCl3, δ, ppm): 2.57 (s, 3H, CH3CO), 5.55 (bs, 2H, NH2), 7.51 (s, 1H, H-4'), 7.65-8.01 (m, 8H, H-5, 2", 4", 5", 6", 7", 9", 10"). 13C NMR (75 MHz, CDCl3, δ, ppm): 26.3 (CH3CO), 109.7 (C-3a), 119.1 (C-5), 125.5, 127.4, 127.5, 127.9, 128.0, 128.1, 128.4, 129.4 (CH-4', 2'', 4'', 5'', 6'', 7'', 9'', 10''), 125.1, 126.8, 133.1, 133.4, 133.5, 133.9, 136.5, 137.4 (Cq- 2, 3, 2', 3', 5', 1'', 3'', 8''), 148.2 (Cq-4), 150.9 (Cq-6), 159.9 (Cq- 7a), 190.1 (CO). 2.4. Biological screening test 2.4.1. Antimicrobial activity Antimicrobial activity of the new nicotinonitriles (2a-l) and furo[2,3-b]pyridines (3a-l) was determined by the wells diffusion method, for these assays, cultures of the following microorganisms were used: two Gram-positive bacteria (Staphylococcus aureus and Bacillus subtilis), one Gram- negative bacteria (Escherichia coli), and fungal strains (Aspergillus niger and Penicillium sp.) [13]. 2.4.2. Antioxidant activity by DPPH method The antioxidant activity of methanolic solutions of compounds 2a-l and 3a-l was determined using the 2,2- diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging assay determined by a modified method described by Ayoola et al. [22]. Solutions of concentration 4.0 mg/mL were prepared for each solution of compounds 2a-l and 3a-l dissolved in methanol. Freshly prepared methanolic solution of DPPH (1.0 mL, 0.25 µM) was mixed with 2.0 mL of different samples of each solution of compounds 2a-l and 3a-l. The mixture was shaken by vortex and the absorbance was determined after 0.5, 24 and 48 hours by spectrophotometer at 517 nm. Ascorbic acid of same concentration (4.0) mg/mL was used as standard reference. Lower absorbance of the reaction mixture indicates higher free radical scavenging activity. The scavenging activity of the samples was calculated using the following equation: ( ) ( ) % 100AB ASPercentage of radical scavenging activity AA AB − = × (1) 3. Results and discussion 3.1. Synthesis and characterization of nicotinonitriles and furo[2,3-b]pyridines The nicotinonitrile derivatives 2a-l are readily obtained in very good to excellent yields by treatment of the 4-aryl-3- cyano-2(H)-pyridones (1a-l) with potassium carbonate and chloroacetone in DMF as a solvent at room temperature, Scheme 1. The synthesized derivatives and different substi- tuents are listed in Scheme 1. The reflux of nicotinenitriles (2a-l) in methanolic solution of sodium methoxide leads to the target furo[2,3-b]pyridines (3a-l) in good to high yields (Scheme 1). The structures of the new synthesized heterocycles were proposed using standard spectroscopic techniques including IR, 1D-NMR, 2D-NMR, and mass spectrometry. The exact chemical shifts of all protons and carbons were determined by a full analysis of 2D-NMR spectra. The complete data for all protons and carbons are listed in the experimental section. The ESI-MS and HR-ESI-MS data analyses revealed the correct molecular ion peaks for all compounds as suggested by their molecular formulas. The 1H NMR data of nicotinonitriles (2a-l) showed two aromatic resonances at δ 7.50-7.78, 7.11-7.77 ppm attributed to H-5 and H-4’, respectively, as well as two resonance signals in the up field region due to the oxygenated methylene protons –CH2– at δ 4.97-5.11 ppm while the methyl singlet of an acetate moiety in the region δ 2.18-2.34 ppm. Furo[2,3- b]pyridines (3a-l) showed similar 1H NMR signals to nicotine- nitriles (2a-l) except for the presence of an exchangeable protons at δ 4.98-5.91 ppm (NH2) instead of the oxygenated methylene. In the 13C NMR spectra, the –CH2– and CN carbons in the newly synthesized nicotinonitriles (2a-l) appeared at δ 71.2-71.5 ppm and δ 113.8-115.3 ppm, respectively. The absence of the –CH2– and CN carbon signals in these region confirms the formation of the furo[2,3-b]pyridine compounds 3a-l. The ESI/MS and HR-ESI/MS mass spectra confirmed the proposed molecular formulas of compounds 2a-l and 3a-l. 3.2. Biological screening 3.2.1. Antimicrobial activity The nicotinonitriles (2a-l) and furo[2,3-b]pyridine compounds 3a-l had no effect on any of bacterial nor fungal strains used in antimicrobial activity test. 3.2.2. Antioxidant activity An antioxidant is molecule that inhibit or quench free radical reactions and delay or inhibit the cellular damage [11,23,24]. A broader definition of antioxidant was suggested by Halliwell et al. 1995 as “any substance that when present at low concentrations, compared to those of an oxidizable substrate significantly delays or prevents oxidation of that substrate” [25]. Several compounds with the antioxidant activity have been used to slow down the radical associated oxidative reactions [11]. In the present study radical scavenging activity were evaluated to clarify the antioxidant properties of the nicotine- nitriles (2a-l) and furo[2,3-b]pyridine (3a-l) at various time intervals (0.5, 24 and 48 hours). The results of radical scavenging activity usually were determined based on a fixed endpoint [17]. Some investigators have proposed kinetic parameters that can provide more complete information about antioxidant behavior [26,27] and suggested that the kinetics could be more important than the total antioxidant capacities determined at a fixed point [11,28]. The DPPH reagent reacted very slowly with chemical samples, approaching, but not reaching, steady state after 8 h [16]. Some researchers reported similar slow reaction of most antioxidants which were tested when DPPH were used [16,29]. The scavenging percentage of ascorbic acid was higher than all other compounds. 2018 – European Journal of Chemistry – CC BY NC – DOI: 10.5155/eurjchem.9.4.375-381.1792 380 Al-Refai et al. / European Journal of Chemistry 9 (4) (2018) 375-381 0 10 20 30 40 50 60 70 80 90 100 2a 2b 2c 2d 2e 2f 2g 2h 2i 2j 2k 2l AA R ad ic al s ca ve ng in g ac tiv ity (A A % ) Compounds after 0.5 hour after 24 hours after 48 hours Figure 1. The DPPH radical scavenging activity of nicotinonitriles (2a-l) in methanol at 4.0 mg/mL concentration after 0.5, 24 and 48 hours. 0 10 20 30 40 50 60 70 80 90 100 3a 3b 3c 3d 3e 3f 3g 3h 3i 3j 3k 3l AA R ad ic al s ca ve ng in g ac tiv ity (A A % ) Compounds after 0.5 hour after 24 hours after 48 hours Figure 2. The DPPH radical scavenging activity of furo[2,3-b]pyridine (3a-l) in methanol at 4.0 mg/mL concentration after 0.5, 24 and 48 hours. Antioxidant activities of nicotinonitriles (2a-l) in methanol were measured using DPPH free radical scavenging assay monitored at various time intervals (0.5, 24 and 48 hours), and the results of DPPH scavenging effect are shown in Figure 1. Nicotinonitriles (2a-l) exhibited a significant antioxidant activity after 24 and 48 hours. The highest percentage (%) was shown by methanolic solvent of compounds 2d, 2h and 2l at 4.0 mg/mL concentration. The DPPH radical scavenging activity of furo[2,3- b]pyridine (3a-l) in methanol at 4.0 mg/mL concentration, monitored at various time intervals (0.5, 24 and 48 hours) are shown in Figure 2. Furo[2,3-b]pyridine (3a-l) exhibited a significant radical scavenging activity after 24 and 48 hours compared with 0.5 hour, all chemical compounds exhibited high radical scavenging activity after 24 and 48 hours compared with 0.5 hour. The highest radical scavenging activity percentage (%) was shown by compounds 3b, 3f and 3j after 24 hours with 86.1, 85.5 and 90.1%, respectively. 4. Conclusion New nicotinonitrile and furo[2,3-b]pyridine derivatives bearing aromatic substituents have been synthesized and fully characterized. The new heterocyclic compounds were studied for their antioxidant activity. Notably, both nicotinonitriles and furo[2,3-b]pyridine derivatives possess antioxidant activity after 24 and 48 hours compared with 0.5 hour. Acknowledgements We are grateful to Al Al-Bayt University (Mafraq, Jordan), and the Deutsche Forschungemeinschaft (DFG, Germany) for financial support. Disclosure statement Conflict of interests: The authors declare that they have no conflict of interest. Author contributions: All authors contributed equally to this work. Ethical approval: All ethical guidelines have been adhered. Sample availability: Samples of the compounds are available from the author. ORCID Mahmoud Al-Refai http://orcid.org/0000-0001-7154-7888 Mohammad Ibrahim http://orcid.org/0000-0003-0720-9889 Abdullah Al-Fawwaz http://orcid.org/0000-0003-0795-7537 Armin Geyer http://orcid.org/0000-0001-8096-2266 2018 – European Journal of Chemistry – CC BY NC – DOI: 10.5155/eurjchem.9.4.375-381.1792 http://orcid.org/0000-0001-7154-7888 http://orcid.org/0000-0003-0720-9889 http://orcid.org/0000-0003-0795-7537 http://orcid.org/0000-0001-8096-2266 Al-Refai et al. / European Journal of Chemistry 9 (4) (2018) 375-381 381 References [1]. Kumar, N. R.; Poornachandra, Y.; Nagender, P.; Mallareddy, G.; Kumar, R. N.; Ranjithreddy, P.; Kumar, G. C.; Narsaiah, B. Eur. J. Med. 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Introduction 2. Experimental 2.1. Materials 2.2. Instrumentation 2.3. Synthesis 2.3.1. General procedure for the synthesis of nicotinonitriles (2a-l) 2.3.2. General procedure for the synthesis of furo[2, 3-b] pyridines (3a-l) 2.4. Biological screening test 2.4.1. Antimicrobial activity 2.4.2. Antioxidant activity by DPPH method 3. Results and discussion 3.1. Synthesis and characterization of nicotinonitriles and furo[2,3-b]pyridines 3.2. Biological screening 3.2.1. Antimicrobial activity 3.2.2. Antioxidant activity 4. Conclusion Acknowledgements Disclosure statement ORCID References PrintField10: PrintField11: PrintField12: PrintField13: PrintField14: PrintField15: PrintField16: PrintField20: PrintField21: PrintField22: PrintField23: PrintField24: PrintField25: PrintField26: