# Synthesis, Characterization, and Cytotoxic Evaluation of Some Newly Substituted Diazene Candidates

Authors: Mohamed El-Naggar; Abdel-Nasser El-Shorbagi; Dina H. Elnaggar; Abd El-Galil E. Amr; Mohamed A. Al-Omar; Elsayed A. Elsayed
Journal: Journal of Chemistry
 (2018)

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## Abstract

A series of azocompounds containing methyl salicylate4a–k and 1-naphthyl moiety 6–8 was synthesized and tested as anticancer agents. Nitrosation of methyl 5-amino-2-hydroxybenzoate or 1-aminonaphthalene by using NaNO2 in the presence of HCl afforded diazonium salt derivatives 2 and 5, which were treated with substituted imino or substituted amino derivatives, to give the corresponding substituted amino-pent-2-en-3-yl-diazenylbenzoate 4a–k or 2-substituted-1-(naphthalen-1-yl)diazene derivatives 6a–h, 7a,b, and 8a,b. All the synthesized compounds were elucidated by elemental analysis and spectroscopic evidence.


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## Body

## 1. Introduction

In a previous work, the authors reported that certain synthesized substituted heterocyclic aromatic derivatives showed antiviral activities [1] and alpha reductase inhibition [2]. Also, some of new compounds containing the nitrogen atom have been synthesized and used as antihypertensive α-blocking [3], antiparkinsonian [4], antialzheimer [5], antimicrobial [6–8], and anti-inflammatory [9, 10] agents. There is an increasing and critical demand for more powerful anticancer agents, due to increased detection of cancer resistance [11]. Azocompounds are one of the largest types of organically synthesized compounds, which are effective both as drugs and cosmetics [12]. Also, they are extensively used in the field of dyeing textiles, biomedical studies, advanced applications in organic synthesis, and remarkable biological activities including antibacterial activity [13–20]. A number of azocompounds were synthesized comprising a drug moiety in their skeleton [21]. Salicylates are a class of chemicals with appreciative biological activities. Several biomedical studies have used methyl salicylate derivatives with promising results [22–24]. Aromatic amines such as benzidine, 1-naphthylamine, and 4-methylaniline have been recognized as potential carcinogens [25, 26]. From these observations, we decided to synthesize two series of azocompounds. One of these series contains methyl salicylate moiety, and the other involves 1-naphthyl moiety. All the synthesized compounds were screened as anticancer agents.


## 2. Experimental

### 2.1. Chemistry

All reagents and solvents were obtained from the commercial supplier and used without further purification. Melting point is uncorrected and was determined on an electrothermal melting point apparatus (Stuart Scientific, England, UK). Precoated silica gel plates (Kieselgel 0.25 mm, 60G F254, Merck, Germany) were used for thin layer chromatography (TLC) for reaction monitoring, and UV light was used for detection. The separation was done using column chromatography with silica gel 60 (Merck) eluted with appropriate solvent. Nuclear magnetic resonance spectra (NMR) were recorded using Bruker 500 MHz instrument. Chemical shifts were reported in ppm, and spectra were corrected using the residual solvent as a reference. Mass spectra were recorded on a Bruker Daltonics spectrometer.


#### 2.1.1. General Procedure for the Synthesis of Compounds4a–k

A mixture of methyl 5-aminosalicylate1 (0.56 g, 3.4 mmol) in concentrated hydrochloric acid (5 mL) was stirred with gentle heating until complete dissolving. Then, the mixture was cooled at a temperature from zero to −4°C, sodium nitrite (0.28 g, 4 mmol) in 2 mL of water was added portionwise at a temperature of −4°C to form the corresponding diazonium salt 2, and the diazonium salt formed was added to a mixture of 4-(substituted imino)pentan-2-one derivatives 3 (2 mmol) [27] in ethanol (15 mL) slowly under the same temperature allowing the mixture to stir at room temperature for an additional 3 hours. The solid formed was collected by filtration, washed with cold water several times, and then left to dry. The obtained product was recrystallized from ethanol to give target compounds 4a–k, respectively.(1) Methyl 5-(4-(benzylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4a). Yellow crystalline solid, m.p. 167-168°C, yield = 62%, and 1H-NMR (DMSO-d6) δ (ppm) 2.42 (s, 3H, CH3-CNH), 2.57 (s, 3H, CO-CH3), 3.93 (s, 3H, OCH3), 4.82 (s, 2H, Ar-CH2), 7.05 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.36–7.42 (m, 5H, ArH), 7.72 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.78 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.40 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.8, 29.0, 47.6, 53.0, 113.8, 118.9, 123.0, 125.6, 128.1, 128.2, 128.6, 129.4, 137.3, 144.5,159.1, 159.4, 169.3, and 196.4 (20C). ESI-MS: m/z, 368.16 (M+ + H). Anal. calcd for C20H21N3O4 (367.40): C, 65.38; H, 5.76; and N, 11.44. Found: C, 65.35; H, 5.78; and N, 11.40.(2) Methyl 5-(2-oxo-4-(phenethylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4b). Yellow crystalline solid, m.p. 131-132°C, yield = 60%, and 1H-NMR (DMSO-d6) δ (ppm) 2.39 (s, 3H, CH3-CNH), 2.52 (s, 3H, CO-CH3), 2.98 (t, 2H, J = 6.8 Hz, PhCH2CH2N), 3.85 (t, 2H, J = 6.7 Hz, ArCH2CH2N), 3.95 (s, 3H, OCH3), 7.03 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.34–7.30 (m, 5H, ArH), 7.59 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.76 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 13.84 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 29.1, 35.0, 44.6, 53.1, 113.8, 118.7, 122.4, 126.6, 127.0, 128.3, 129.0, 129.2, 138.7, 144.9, 159.1, 159.3, 169.5, and 196.3 (21C). ESI-MS: m/z, 382.18 (M+ + H). Anal. calcd for C21H23N3O4 (381.43): C, 66.13; H, 6.08; and N, 11.02. Found: C, 66.45; H, 6.02; and N, 11.00.(3) Methyl 5-(2-oxo-4-(1-phenylethylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4c). Yellow crystalline solid, m.p. 110-111°C, yield = 66%, and 1H-NMR (DMSO-d6) δ (ppm) 1.60 (d, 3H, J = 7.6 Hz, ArCH(CH3)N), 2.41 (s, 3H, CH3-C), 2.49 (s, 3H, CO-CH3), 3.94 (s, 3H, OCH3), 5.17 (q, 1H, J = 6.7 Hz, ArCH(CH3)N), 7.06 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.40–7.45 (m, 5H, ArH), 7.76 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.86 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.63 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.8, 24.0, 29.0, 53.1, 53.7, 114.0, 119.0, 122.2, 126.4, 126.6, 128.2, 128.5, 129.4, 143.0, 144.5, 159.2, 159.4, 169.3, and 196.4 (21C). ESI-MS: m/z, 382.18 (M+ + H). Anal. calcd for C21H23N3O4 (381.43): C, 66.13; H, 6.08; and N, 11.02. Found: C, 66.04; H, 6.03; and N, 10.98.(4) Methyl 5-(4-(2-hydroxyethylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4d). Yellow crystalline solid, m.p. 187-188°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.42 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.59 (t, 2H, J = 5.0 Hz, NCH2CH2OH), 3.66 (t, 2H, J = 4.2 Hz, NCH2CH2OH), 3.92 (s, 3H, OCH3), 5.13 (br s, 1H, NCH2CH2OH) 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.79 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 8.01 (s, 1H, Ar-H, benzoate ring), and 10.5 (br s, 1H, NH), 14.21 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.7, 29.1, 46.0, 53.1, 59.7, 113.9, 118.7, 122.1, 127.0, 128.4, 134.5, 154.1, 159.3, 169.5, and 196.4 (15C). ESI-MS: 322.27 (M+ + H). Anal. calcd for C15H19N3O5 (321.33): C, 56.07; H, 5.96; and N, 13.08. Found: C, 56.00; H, 5.92; and N, 13.00.(5) Methyl 5-(4-(isobutylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4e). Yellow crystalline solid, m.p. 94–95°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.00 (d, 6H, J = 6.7 Hz, NHCH2CH(CH3)2), 1.95–1.90 (m, 1H, NHCH2CH(CH3)2), 2.42 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.39 (dd, 2H, J = 5.4 Hz, NHCH2CH(CH3)2), 3.92 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.76 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.95 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.09 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.45, 2.36, 28.3, 29.1, 50.7, 53.1, 114.0, 118.9, 122.0, 126.8, 128.4, 134.5, 145.2, 159.3, 169.4, and 196.2 (17C). ESI-MS: m/z, 334.18 (M+ + H). Anal. calcd for C17H23N3O4 (333.38): C, 61.25; H, 6.95; and N, 12.60. Found: C, 61.12; H, 6.90; and N, 12.54.(6) Methyl 5-(4-(isopropylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4f). Yellow crystalline solid, m.p. 104-105°C, yield = 61%, and 1H-NMR (DMSO-d6) δ (ppm) 1.30 (d, 6H, J = 6.4 Hz, NHCH(CH3)2), 2.41 (s, 3H, CH3-C), 2.56 (s, 3H, COCH3), 3.92 (s, 3H, OCH3), 4.15–4.12 (m, 1H, J = 6.4 Hz, NHCH(CH3)2), 7.08 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.92 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.10 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 23.2, 29.1, 45.6, 53.1, 114.0, 118.8, 121.8, 127.1, 127.9, 134.5, 145.1, 159.3, 169.4, and 196.1 (16C). ESI-MS: m/z, 320.16 (M+ + H). Anal. calcd for C16H21N3O4 (319.36): C, 60.17; H, 6.63; and N, 13.16. Found: C, 60.10; H, 6.58; and N, 13.10.(7) Methyl 5-(4-(butylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4g). Yellow crystalline solid, m.p. 131-132°C, yield = 65%, and 1H-NMR (DMSO-d6) δ (ppm) 0.97 (t, 3H, J = 7.3 Hz, NHCH2CH2CH2CH3), 1.43-1.40 (m, 2H, J = 7.6 Hz, NHCH2CH2CH2CH3), 1.62–1.66 (m, 2H, J = 6.9 Hz, NHCH2CH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.53 (s, 3H, COCH3), 3.52 (t, 2H, J = 6.7 Hz, NHCH2CH2CH2CH3), 3.91 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.72 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.05 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.0, 16.4, 20.2, 29.0, 31.0, 43.1, 53.1, 113.9, 118.9, 122.6, 126.5, 128.2, 134.5, 145.1, 159.3, 169.4, and 196.2 (17C). ESI-MS: m/z, 334.18 (M+ + H). Anal. calcd for C17H23N3O4 (333.38): C, 61.25; H, 6.95; and N, 12.60. Found: C, 61.12; H, 6.86; and N, 12.52.(8) Methyl 5-(4-(3-hydroxypropylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4h). Yellow crystalline solid, m.p. 135-136°C, yield = 63%, and 1H-NMR (DMSO-d6) δ (ppm) 1.80–1.75 (m, 2H, J = 6.3 Hz, NHCH2CH2CH2OH), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.54 (br t, 2H, J = 5.7 Hz, NHCH2CH2CH2OH), 3.59 (br t, 2H, J = 6.7 Hz, NHCH2CH2CH2OH), 3.91 (s, 3H, OCH3), 4.70 (br s, 2H, NHCH2CH2CH2OH), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.77 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.98 (s, 1H, Ar-H, benzoate ring), and 10.50 (br s, 1H, NH), 13.95 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 29.1, 32.2, 40.0, 53.1, 58.4, 113.9, 118.7, 122.0, 127.1, 128.3, 134.5, 145.2, 159.3, 169.5, and 196.2 (16C). ESI-MS): m/z, 336.16 (M+ + H). Anal. calcd for C16H21N3O5 (335.36): C, 57.30; H, 6.31; and N, 12.53. Found: C, 57.10; H, 6.22; and N, 12.42.(9) Methyl 5-(2-oxo-4-(pentylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4i). Yellow crystalline solid, m.p. 127-128°C, yield = 72%, and 1H-NMR (DMSO-d6) δ (ppm) 0.92 (t, 3H, J = 6.9 Hz, NHCH2CH2CH2CH2CH3), 1.40–1.45 (m, 4H, NHCH2CH2CH2CH2CH3), 1.66–1.62 (m, 2H, NHCH2CH2CH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.53 (t, 2H, J = 6.7 Hz, NHCH2CH2CH2CH2CH3), 3.91 (s, 3H, OCH3), 7.06 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.05 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.3, 16.4, 22.3, 28.6, 29.2, 30.7, 43.4, 53.0, 113.8, 118.9, 122.7, 126.3, 128.2, 134.5, 145.1, 159.4, 169.4, and 196.2 (18C). ESI-MS: m/z, 348.19 (M+ + H). Anal. calcd for C18H25N3O4 (347.41): C, 62.23; H, 7.25; and N, 12.10. Found: C, 62.05; H, 7.16; and N, 12.05.(10) Methyl 5-(4-(ethylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4j). Yellow crystalline solid, m.p. 106-107°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.27 (t, 3H, J = 7.2 Hz, NHCH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.55 (q, 2H, NHCH2CH3), 3.91 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 13.88 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.8, 16.3, 29.1, 38.3, 53.1, 113.9, 118.8, 122.3, 126.9, 128.1, 134.5, 145.3, 159.4, 169.5, and 196.1 (15C). ESI-MS): m/z, 306.15 (M+ + H). Anal. calcd for C15H19N3O4 (305.33): C, 59.01; H, 6.27; and N, 13.76. Found: C, 58.86; H, 6.20; and N, 13.70.(11) Methyl 5-(2-oxo-4-(propylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4k). Yellow crystalline solid, m.p. 137-138°C, yield = 69%, and 1H-NMR (DMSO-d6) δ (ppm) 0.99 (t, 3H, J = 7.3 Hz, NHCH2CH2CH3), 1.68–1.64 (m, 3H, J = 7.2 Hz, NHCH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.50 (q, 2H, J = 6.9 Hz, NHCH2CH3), 3.92 (s, 3H, OCH3), 7.08 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.77 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.96 (s, 1H, Ar-H, benzoate ring), 10.52 (br s, 1H, NH), and 13.98 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 11.8, 16.4, 22.5, 29.1, 45.1, 53.1, 114, 118.8, 122.2, 126.9, 128.3, 134.5, 145.2, 159.3, 169.4, and 196.2 (16C). ESI-MS: m/z, 320.14 (M+ + H). Anal. calcd for C16H21N3O4 (319.36): C, 60.17; H, 6.63; and N, 13.16. Found: C, 60.00; H, 6.56; and N, 13.10.


#### 2.1.2. General Procedure for the Synthesis of Compounds6–8

A mixture of 1-aminonaphthalene (0.48 g, 3.4 mmol) and conc. HCl (5 mL) was stirred with gentle heating until complete dissolving. The reaction mixture was cooled at a temperature from zero to −4°C in an ice bath, and then sodium nitrite (0.28 g, 4 mmol) in water (2 mL) was added portionwise to the reaction mixture at the same temperature to form the diazonium salt5. To the diazonium salt 5, a solution of appropriate amide or iminoderivatives [27], namely, N-substituted 3-oxo-butanamide, ethyl 3-(substituted imino)butanoate, or N-alkyl-3-(alkyl imino)-butanamide in absolute ethanol (10 mL), was added while stirring at a temperature of −4°C, and the mixture was stirred at room temperature for an additional 3 hours. The solid formed was collected by filtration, washed with cold water several times, dried, and crystallized from dimethylformamide/ethanol to give the corresponding 6a–h, 7a,b, and 8a,b, respectively.(1) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-phenethylbut-2-enamide
(6a). Brownish crystalline solid, m.p. 141-142°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.50 (s, 3H, C-CH3), 2.88 (t, 2H, J = 7.2 Hz, ArCH2CH2NH), 3.62 (q, 2H, J = 7.0 Hz, ArCH2CH2NH), 7.23–7.20 (m, 1H, Ar), 7.33–7.30 (m, 4H, ArH), 7.63–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 6.7 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.04 (d, 1H, J = 7.9 Hz, naphthyl-H), 9.40 (br s, 1H, NHCO), and 15.67 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 26.4, 35.2, 40.7, 110.9, 119.8, 123.1, 125.1, 126.8, 126.9, 127.2, 127.5, 128.1, 128.9, 129.2, 129.3, 134.2, 136.9, 139.5, 164.9, and 198.7 (22C). ESI- MS: m/z, 360.17 (M+ + H). Anal. calcd for C22H21N3O2 (359.42): C, 73.52; H, 5.89; and N, 11.69. Found: C, 73.45; H, 5.80; and N, 11.60.(2) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-methylbut-2-enamide
(6b). Brownish crystalline solid, m.p. 175-176°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm) 2.52 (s, 3H, C-CH3), 2.88 (d, 3H, J = 4.9 Hz, CONHCH3), 7.63–7.60 (m, 2H, naphthyl-H), 7.72–7.68 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.2 Hz, naphthyl-H), 7.83 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.92 (d, 1H, J = 8.0 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.23 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 25.8, 26.4, 110.8, 119.8, 123.0, 124.9, 126.9, 127.1, 127.5, 128.3, 129.3, 134.2, 136.9, 165.5, and 198.5 (15C). ESI-MS: m/z, 270.12 (M+ + H). Anal. calcd for C15H15N3O2 (269.30): C, 66.90; H, 5.61; and N, 15.60. Found: C, 66.80; H, 5.50; and N, 15.52.(3) 2-(Naphthalen-1-yl-diazenyl)-N-tert-butyl-3-hydroxybut-2-enamide
(6c). Brownish crystalline solid, m.p. 126-127°C, yield = 59%, and 1H-NMR (DMSO-d6) δ (ppm) 1.43 (s, 9H, C(CH3)3), 2.52 (s, 3H, C-CH3), 7.63–7.60 (m, 2H, naphthyl-H), 7.74–7.40 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.90 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.42 (s, 1H, NHCO), and 15.69 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 26.5, 28.8, 51.3, 111.0, 119.7, 123.1, 125.1, 126.9, 127.1, 127.6, 128.4, 129.2, 134.2, 136.9, 164.7, and 199.3 (18C). ESI-MS: m/z, 312.17 (M+ + H). Anal. calcd for C18H21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.30; H, 6.75; and N, 13.40.(4) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-isopropylbut-2-enamide
(6d). Brownish crystalline solid, m.p. 124-125°C, yield = 62%, and 1H-NMR (DMSO-d6) δ (ppm) 1.23 (d, 6H, J = 6.6 Hz, CH(CH3)2), 2.52 (s, 3H, C-CH3), 4.11–4.00 (m, 1H, CH(CH3)2), 7.65–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 6.8 Hz, naphthyl-H), 7.90 (d, 1H, J = 8.3 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.26 (br s, 1H, NHCO), and 15.71 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 22.6, 40.8, 49.1, 110.9, 119.7, 123.1, 125.1, 127.0, 127.1, 127.5, 128.1, 129.3, 134.2, 136.9, 164.2, and 199.0 (17C). ESI-MS: m/z, 280.15 (M+ + H). Anal. calcd for C17H19N3O2 (297.35): C, 68.67; H, 6.44; and N, 14.13. Found: C, 68.60; H, 6.34; and N, 14.00.(5) 2-(Naphthalen-1-yl-diazenyl)-N-ethyl-3-hydroxybut-2-enamide
(6e). Brownish crystalline solid, m.p. 130–131°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.17 (t, 3H, J = 7.2 Hz, NHCH2CH3), 2.52 (s, 3H, C-CH3), 3.38–3.34 (m, 2H, NHCH2CH3), 7.64–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.33 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.9, 26.5, 33.60, 110.9, 119.8, 123.0, 125.0, 126.9, 127.1, 127.5, 128.2, 129.3, 134.2, 136.9, 164.8, and 198.7 (16C). ESI-MS: m/z, 284.14 (M+ + H). Anal. calcd for C16H17N3O2 (283.33): C, 67.83; H, 6.05; and N, 14.83. Found: C, 67.74; H, 6.00; and N, 14.78.(6) 2-(Naphthalen-1-yl-diazenyl)-N-butyl-3-hydroxybut-2-enamide
(6f). Brownish crystalline solid, m.p. 142-143°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm) 0.93 (t, 3H, J = 7.3 Hz, NHCH2CH2CH2CH3), 1.38–1.35 (m, 2H, J = 7.6 Hz, NHCH2CH2CH2CH3), 1.57–1.54 (m, 2H, J = 7.3 Hz, NHCH2CH2CH2CH3) 2.53 (s, 3H, C-CH3), 3.36 (q, 2H, J = 7.0 Hz, NHCH2CH2CH2CH3), 7.65–1.62 (m, 2H, naphthyl-H), 7.75–7.71 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.36 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.1, 20.2, 26.5, 31.3, 38.3, 110.9, 119.8, 123.1, 125.1, 126.9, 127.1, 127.5, 128.2, 129.3, 134.2, and 136.9, 164.9, 198.8 (18C). ESI-MS: m/z, 312.17 (M+ + H). Anal. calcd for C18H21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.35; H, 6.72; and N, 13.40.(7) 2-(Naphthalen-1-yl-diazenyl)-N-sec-butyl-3-hydroxybut-2-enamide
(6g). Yellow crystalline solid, m.p. 105-106°C, yield = 55%, and 1H-NMR (CDCl3) δ (ppm) 1.01 (t, 3H, J = 7.4 Hz, CH3CH2CH(NH)CH3), 1.28 (d, 3H, J = 6.6 Hz, CH3CH2CH(NH)CH3), 1.66–1.60 (m, 2H, CH3CH2CH(NH)CH3), 2.62 (s, 3H, C-CH3), 4.12–4.08 (m, 1H, CH3CH2CH(NH)CH3) 7.56–7.52 (m, 3H, naphthyl-H), 7.68 (d, 1H, J = 8.0 Hz, naphthyl-H), 7.82 (d, 1H, J = 8.0 Hz, naphthyl-H), 7.90 (d, 1H, J = 7.6 Hz naphthyl-H), 8.07 (d, 1H, J = 7.9 Hz naphthyl-H), 9.40 (br s, 1H, NHCO), and 15.81 (s, 1H, OH); 13C-NMR (CDCl3) δ (ppm). 10.4, 20.3, 26.2, 29.4, 46.0, 110.7, 120.2, 123.5, 124.8, 126.1, 126.4, 126.7, 127.6, 128.6, 134.1, 137.2, 164.7, and 199.4 (18C). ESI-MS: m/z, 312.16 (M+ + H). Anal. calcd for C18N21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.35; H, 6.72; and N, 13.40.(8) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-propylbut-2-enamide
(6h). Brownish crystalline solid, m.p. 157-158°C, yield = 65%, and 1H-NMR (CDCl3) δ (ppm) 1.04 (t, 3H, J = 7.4 Hz, CONHCH2CH2CH3), 1.68–1.63 (m, 2H, J = 7.3 Hz, CONHCH2CH2CH3), 2.62 (s, 3H, C-CH3), 3.42 (q, 2H, J = 7.0 Hz, CONHCH2CH2CH3), 7.57–7.53 (m, 3H, naphthyl-H), 7.68 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.90 (d, 1H, J = 7.5 Hz, naphthyl-H), 8.07 (br d, 1H, J = 8.5 Hz, naphthyl-H), 9.54 (br s, 1H, NHCO), and 15.79 (s, 1H, OH); 13C-NMR (CDCl3) δ (ppm) 11.6, 22.6, 26.2, 40.4, 110.7, 120.2, 123.5, 124.9, 126.1, 126.4, 126.7, 127.6, 128.6, 134.1, 137.1, 165.3, and 199.4 (17C). ESI-MS: m/z, 298.15 (M+ + H). Anal. calcd for C17H19N3O2 (297.35): C, 68.67; H, 6.44; and N, 14.13. Found: C, 68.55; H, 6.36; and N, 14.04.(9) Ethoxy 3-(benzylimino)-2-(naphthalen-1-yl-diazenyl)but-1-en-1-ol
(7a). Brownish crystalline solid, m.p. 110-111°C, yield = 77%, and 1H-NMR (DMSO-d6) δ (ppm) 1.32 (t, 3H, J = 7.1 Hz, OCH2CH3), 2.59 (s, 3H, C-CH3), 4.27 (q, 2H, J = 7.1 Hz, OCH2CH3), 4.83 (s, 2H, J = 3.4 Hz, ArCH2N), 7.50–4.46 (m, 9H, naphthyl-H and ArH), 7.65 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.72 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.88 (d, 1H, J = 8.2 Hz, naphthyl-H), and 14.90 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.9, 17.2, 50.2, 60.3, 111.2, 121.9, 124.1, 126.1, 126.4, 126.4, 126.6, 127.2, 128.1, 128.5, 129.3, 129.5, 134.3, 136.9, 137.9, 166.7, and 198.7 (23C). ESI-MS: m/z, 374.19 (M+ + H). Anal. calcd for C23H23N3O2 (373.45): C, 73.97; H, 6.21; and N, 11.25. Found: C, 73.85; H, 6.12; and N, 11.20.(10) Ethoxy 2-(naphthalen-1-yl-diazenyl)-3-(phenethylimino)but-1-en-1-ol
(7b). Brownish crystalline solid, m.p. 142-143°C, yield = 72%, and 1H-NMR (DMSO-d6) δ (ppm) 1.30 (t, 3H, J = 7.1 Hz, OCH2CH3), 2.48 (s, 3H, C-CH3), 3.08 (t, 2H, J = 7.1 Hz, ArCH2CH2N), 3.89 (t, 2H, J = 6.9 Hz, ArCH2CH2N), 4.23 (q, 3H, J = 7.1 Hz, OCH2CH3), 7.20–7.15 (m, 1H, naphthyl-H), 7.28–7.24 (m, 2H, naphthyl-H), 7.32–7.28 (m, 2H, naphthyl-H), 7.58–7.55 (m, 4H, ArH), 7.78–7.74 (m, 1H, naphthyl-H), 7.96–7.92 (m, 1H, naphthyl-H), and 14.59 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm). 14.9, 16.5, 35.9, 46.4, 60.1, 111.9, 122.6, 125.1, 126.3, 126.5, 126.7, 126.9, 127.5, 128.6, 128.9, 129.2, 129.2, 134.4, 136.9, 138.9, 166.7, and 198.7 (24C). ESI-MS: m/z, 388.19 (M+ + H). Anal. calcd for C24H25N3O2 (387.47): C, 74.39; H, 6.50; and N, 10.84. Found: C, 74.18; H, 6.42; and N, 10.76.(11) 1-(Butylamino)-3-(butylimino)-2-[(1-(naphthalen-1-yl)diazene]but-1-en-1-ol
(8a). Brownish crystalline solid, m.p. 91-92°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm). 0.93 (t, 3H, J = 6.4 Hz, NHCH2CH2CH2CH3), 0.96 (t, 3H, J = 7.3 Hz, NCH2CH2CH2CH3), 1.46–1.40 (m, 4H, J = 7.7 Hz, NHCH2CH2CH2CH3 and NCH2CH2CH2CH3), 1.61 (q, 2H, NHCH2CH2CH2CH3), 1.67–1.64 (m, 2H, NCH2CH2CH2CH3), 2.62 (s, 3H, C-CH3), 3.39 (q, 2H, J = 6.4 Hz, NHCH2CH2CH2CH3), 3.65 (q, 2H, J = 6.8 Hz, NCH2CH2CH2CH3), 7.56–7.52 (m, 4H, naphthyl-H), 7.76 (d, 1H, J = 8.2 Hz, naphthyl-H), 7.95–7.90 (m, 1H, naphthyl-H), 8.4 (d, 1H, J = 7.4 Hz, naphthyl-H), 11.3 (br s, 1H, NH), and 15.37 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.1, 14.2, 14.7, 20.0, 20.4, 31.4, 31.7, 37.9, 44.0, 111.3, 121.3, 122.8, 126.0, 126.3, 126.8, 126.8, 128.6, 129.0, 134.5, 148.7, 165.9, and 172.7 (22C). ESI-MS: m/z, 367.25 (M+ + H). Anal. calcd for C22H30N4O (366.50): C, 72.10; H, 8.25; and N, 15.29. Found: C, 72.00; H, 8.18; and N, 15.20.(12) N-(2-hydroxyethyl imino)-3-(2-hydroxyethylamino)-2-(naphthalen-1-yldiazenyl)but-2-en-2-ol
(8b). Brownish crystalline solid, m.p. 157-158°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.64 (s, 3H, C-CH3), 3.47 (q, 3H, J = 5.3 Hz, CONHCH2CH2OH), 3.62–3.66 (m, 6H, CONHCH2CH2OH and CNCH2CH2OH), 4.95 (t, 1H, J = 4.6 Hz, CONHCH2CH2OH), 5.08 (t, 1H, J = 4.4 Hz, CNCH2CH2OH), 7.54–7.50 (m, 4H, naphthyl-H), 7.74 (d, 1H, J = 7.8 Hz, naphthyl-H), 7.94–7.88 (m, 1H, naphthyl-H), 8.74–8.70 (m, 1H, naphthyl-H), 11.40 (br s, 1H, NH), and 15.37 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm).15.0, 41.1, 47.1, 60.0, 60.5, 111.1, 121.3, 123.9, 126.3, 126.4, 126.7, 126.8, 128.2, 129.2, 134.5, 148.8, 165.8, and 172.8 (18C). ESI-MS: m/z, 343.18 (M+ + H). Anal. calcd for C18H22N4O3 (342.39): C, 63.14; H, 6.48; and N, 16.36. Found: C, 63.00; H, 6.40; and N, 16.30.


### 2.2. Cytotoxicity Assay

Three cancer cell lines were used in the current study: MCF-7 (human breast adenocarcinoma), A549 (lung carcinoma), and HT-29 (human colorectal adenocarcinoma). Cancer cell lines were obtained from Sigma-Aldrich Chemical Company, USA, and were seeded in Dulbecco’s Modified Eagle Media (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS), 100 IU/mL penicillin, 100 mg/mL streptomycin, and 2 mM glutamine. The cells were maintained in a humidified atmosphere with 5% CO2 at 37°C.


#### 2.2.1. Antiproliferative Activity

The antiproliferative activity of the tested compounds was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as mentioned before [28–30]. In brief, cells were seeded in 96-well plates 24 h before different treatments. Test compounds were dissolved in dimethyl sulfoxide (DMSO) and further diluted with the cultivation medium. All target compounds (1–23) were preliminary evaluated in vitro at the single concentration of 5 μM against different cancer cell lines. Doxorubicin was used in parallel as a positive control drug at the single concentration of 2 μM. Different tested samples were added to the wells, and the cells were incubated at 37°C in a 5% CO2-humidified incubator for 48 h. After treatment, 10 μL of the MTT solution (5 mg/mL) was added to each well, and the plates were further incubated for 2 h. The media along with the MTT solution was removed and replaced by 100 μL of the DMSO solution to dissolve the precipitating formazan crystals. The intensity of the developed color was measured at 570 nm with an automatic multiwell plate reader (Varioskan™ LUX multimode microplate reader, Thermo Fisher Scientific, USA). Percentage inhibition of proliferation was calculated as a fraction of the vehicle control.


## 2.1. Chemistry

All reagents and solvents were obtained from the commercial supplier and used without further purification. Melting point is uncorrected and was determined on an electrothermal melting point apparatus (Stuart Scientific, England, UK). Precoated silica gel plates (Kieselgel 0.25 mm, 60G F254, Merck, Germany) were used for thin layer chromatography (TLC) for reaction monitoring, and UV light was used for detection. The separation was done using column chromatography with silica gel 60 (Merck) eluted with appropriate solvent. Nuclear magnetic resonance spectra (NMR) were recorded using Bruker 500 MHz instrument. Chemical shifts were reported in ppm, and spectra were corrected using the residual solvent as a reference. Mass spectra were recorded on a Bruker Daltonics spectrometer.


### 2.1.1. General Procedure for the Synthesis of Compounds4a–k

A mixture of methyl 5-aminosalicylate1 (0.56 g, 3.4 mmol) in concentrated hydrochloric acid (5 mL) was stirred with gentle heating until complete dissolving. Then, the mixture was cooled at a temperature from zero to −4°C, sodium nitrite (0.28 g, 4 mmol) in 2 mL of water was added portionwise at a temperature of −4°C to form the corresponding diazonium salt 2, and the diazonium salt formed was added to a mixture of 4-(substituted imino)pentan-2-one derivatives 3 (2 mmol) [27] in ethanol (15 mL) slowly under the same temperature allowing the mixture to stir at room temperature for an additional 3 hours. The solid formed was collected by filtration, washed with cold water several times, and then left to dry. The obtained product was recrystallized from ethanol to give target compounds 4a–k, respectively.(1) Methyl 5-(4-(benzylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4a). Yellow crystalline solid, m.p. 167-168°C, yield = 62%, and 1H-NMR (DMSO-d6) δ (ppm) 2.42 (s, 3H, CH3-CNH), 2.57 (s, 3H, CO-CH3), 3.93 (s, 3H, OCH3), 4.82 (s, 2H, Ar-CH2), 7.05 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.36–7.42 (m, 5H, ArH), 7.72 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.78 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.40 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.8, 29.0, 47.6, 53.0, 113.8, 118.9, 123.0, 125.6, 128.1, 128.2, 128.6, 129.4, 137.3, 144.5,159.1, 159.4, 169.3, and 196.4 (20C). ESI-MS: m/z, 368.16 (M+ + H). Anal. calcd for C20H21N3O4 (367.40): C, 65.38; H, 5.76; and N, 11.44. Found: C, 65.35; H, 5.78; and N, 11.40.(2) Methyl 5-(2-oxo-4-(phenethylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4b). Yellow crystalline solid, m.p. 131-132°C, yield = 60%, and 1H-NMR (DMSO-d6) δ (ppm) 2.39 (s, 3H, CH3-CNH), 2.52 (s, 3H, CO-CH3), 2.98 (t, 2H, J = 6.8 Hz, PhCH2CH2N), 3.85 (t, 2H, J = 6.7 Hz, ArCH2CH2N), 3.95 (s, 3H, OCH3), 7.03 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.34–7.30 (m, 5H, ArH), 7.59 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.76 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 13.84 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 29.1, 35.0, 44.6, 53.1, 113.8, 118.7, 122.4, 126.6, 127.0, 128.3, 129.0, 129.2, 138.7, 144.9, 159.1, 159.3, 169.5, and 196.3 (21C). ESI-MS: m/z, 382.18 (M+ + H). Anal. calcd for C21H23N3O4 (381.43): C, 66.13; H, 6.08; and N, 11.02. Found: C, 66.45; H, 6.02; and N, 11.00.(3) Methyl 5-(2-oxo-4-(1-phenylethylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4c). Yellow crystalline solid, m.p. 110-111°C, yield = 66%, and 1H-NMR (DMSO-d6) δ (ppm) 1.60 (d, 3H, J = 7.6 Hz, ArCH(CH3)N), 2.41 (s, 3H, CH3-C), 2.49 (s, 3H, CO-CH3), 3.94 (s, 3H, OCH3), 5.17 (q, 1H, J = 6.7 Hz, ArCH(CH3)N), 7.06 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.40–7.45 (m, 5H, ArH), 7.76 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.86 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.63 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.8, 24.0, 29.0, 53.1, 53.7, 114.0, 119.0, 122.2, 126.4, 126.6, 128.2, 128.5, 129.4, 143.0, 144.5, 159.2, 159.4, 169.3, and 196.4 (21C). ESI-MS: m/z, 382.18 (M+ + H). Anal. calcd for C21H23N3O4 (381.43): C, 66.13; H, 6.08; and N, 11.02. Found: C, 66.04; H, 6.03; and N, 10.98.(4) Methyl 5-(4-(2-hydroxyethylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4d). Yellow crystalline solid, m.p. 187-188°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.42 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.59 (t, 2H, J = 5.0 Hz, NCH2CH2OH), 3.66 (t, 2H, J = 4.2 Hz, NCH2CH2OH), 3.92 (s, 3H, OCH3), 5.13 (br s, 1H, NCH2CH2OH) 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.79 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 8.01 (s, 1H, Ar-H, benzoate ring), and 10.5 (br s, 1H, NH), 14.21 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.7, 29.1, 46.0, 53.1, 59.7, 113.9, 118.7, 122.1, 127.0, 128.4, 134.5, 154.1, 159.3, 169.5, and 196.4 (15C). ESI-MS: 322.27 (M+ + H). Anal. calcd for C15H19N3O5 (321.33): C, 56.07; H, 5.96; and N, 13.08. Found: C, 56.00; H, 5.92; and N, 13.00.(5) Methyl 5-(4-(isobutylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4e). Yellow crystalline solid, m.p. 94–95°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.00 (d, 6H, J = 6.7 Hz, NHCH2CH(CH3)2), 1.95–1.90 (m, 1H, NHCH2CH(CH3)2), 2.42 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.39 (dd, 2H, J = 5.4 Hz, NHCH2CH(CH3)2), 3.92 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.76 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.95 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.09 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.45, 2.36, 28.3, 29.1, 50.7, 53.1, 114.0, 118.9, 122.0, 126.8, 128.4, 134.5, 145.2, 159.3, 169.4, and 196.2 (17C). ESI-MS: m/z, 334.18 (M+ + H). Anal. calcd for C17H23N3O4 (333.38): C, 61.25; H, 6.95; and N, 12.60. Found: C, 61.12; H, 6.90; and N, 12.54.(6) Methyl 5-(4-(isopropylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4f). Yellow crystalline solid, m.p. 104-105°C, yield = 61%, and 1H-NMR (DMSO-d6) δ (ppm) 1.30 (d, 6H, J = 6.4 Hz, NHCH(CH3)2), 2.41 (s, 3H, CH3-C), 2.56 (s, 3H, COCH3), 3.92 (s, 3H, OCH3), 4.15–4.12 (m, 1H, J = 6.4 Hz, NHCH(CH3)2), 7.08 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.92 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.10 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 23.2, 29.1, 45.6, 53.1, 114.0, 118.8, 121.8, 127.1, 127.9, 134.5, 145.1, 159.3, 169.4, and 196.1 (16C). ESI-MS: m/z, 320.16 (M+ + H). Anal. calcd for C16H21N3O4 (319.36): C, 60.17; H, 6.63; and N, 13.16. Found: C, 60.10; H, 6.58; and N, 13.10.(7) Methyl 5-(4-(butylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4g). Yellow crystalline solid, m.p. 131-132°C, yield = 65%, and 1H-NMR (DMSO-d6) δ (ppm) 0.97 (t, 3H, J = 7.3 Hz, NHCH2CH2CH2CH3), 1.43-1.40 (m, 2H, J = 7.6 Hz, NHCH2CH2CH2CH3), 1.62–1.66 (m, 2H, J = 6.9 Hz, NHCH2CH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.53 (s, 3H, COCH3), 3.52 (t, 2H, J = 6.7 Hz, NHCH2CH2CH2CH3), 3.91 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.72 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.05 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.0, 16.4, 20.2, 29.0, 31.0, 43.1, 53.1, 113.9, 118.9, 122.6, 126.5, 128.2, 134.5, 145.1, 159.3, 169.4, and 196.2 (17C). ESI-MS: m/z, 334.18 (M+ + H). Anal. calcd for C17H23N3O4 (333.38): C, 61.25; H, 6.95; and N, 12.60. Found: C, 61.12; H, 6.86; and N, 12.52.(8) Methyl 5-(4-(3-hydroxypropylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4h). Yellow crystalline solid, m.p. 135-136°C, yield = 63%, and 1H-NMR (DMSO-d6) δ (ppm) 1.80–1.75 (m, 2H, J = 6.3 Hz, NHCH2CH2CH2OH), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.54 (br t, 2H, J = 5.7 Hz, NHCH2CH2CH2OH), 3.59 (br t, 2H, J = 6.7 Hz, NHCH2CH2CH2OH), 3.91 (s, 3H, OCH3), 4.70 (br s, 2H, NHCH2CH2CH2OH), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.77 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.98 (s, 1H, Ar-H, benzoate ring), and 10.50 (br s, 1H, NH), 13.95 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 29.1, 32.2, 40.0, 53.1, 58.4, 113.9, 118.7, 122.0, 127.1, 128.3, 134.5, 145.2, 159.3, 169.5, and 196.2 (16C). ESI-MS): m/z, 336.16 (M+ + H). Anal. calcd for C16H21N3O5 (335.36): C, 57.30; H, 6.31; and N, 12.53. Found: C, 57.10; H, 6.22; and N, 12.42.(9) Methyl 5-(2-oxo-4-(pentylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4i). Yellow crystalline solid, m.p. 127-128°C, yield = 72%, and 1H-NMR (DMSO-d6) δ (ppm) 0.92 (t, 3H, J = 6.9 Hz, NHCH2CH2CH2CH2CH3), 1.40–1.45 (m, 4H, NHCH2CH2CH2CH2CH3), 1.66–1.62 (m, 2H, NHCH2CH2CH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.53 (t, 2H, J = 6.7 Hz, NHCH2CH2CH2CH2CH3), 3.91 (s, 3H, OCH3), 7.06 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.05 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.3, 16.4, 22.3, 28.6, 29.2, 30.7, 43.4, 53.0, 113.8, 118.9, 122.7, 126.3, 128.2, 134.5, 145.1, 159.4, 169.4, and 196.2 (18C). ESI-MS: m/z, 348.19 (M+ + H). Anal. calcd for C18H25N3O4 (347.41): C, 62.23; H, 7.25; and N, 12.10. Found: C, 62.05; H, 7.16; and N, 12.05.(10) Methyl 5-(4-(ethylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4j). Yellow crystalline solid, m.p. 106-107°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.27 (t, 3H, J = 7.2 Hz, NHCH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.55 (q, 2H, NHCH2CH3), 3.91 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 13.88 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.8, 16.3, 29.1, 38.3, 53.1, 113.9, 118.8, 122.3, 126.9, 128.1, 134.5, 145.3, 159.4, 169.5, and 196.1 (15C). ESI-MS): m/z, 306.15 (M+ + H). Anal. calcd for C15H19N3O4 (305.33): C, 59.01; H, 6.27; and N, 13.76. Found: C, 58.86; H, 6.20; and N, 13.70.(11) Methyl 5-(2-oxo-4-(propylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4k). Yellow crystalline solid, m.p. 137-138°C, yield = 69%, and 1H-NMR (DMSO-d6) δ (ppm) 0.99 (t, 3H, J = 7.3 Hz, NHCH2CH2CH3), 1.68–1.64 (m, 3H, J = 7.2 Hz, NHCH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.50 (q, 2H, J = 6.9 Hz, NHCH2CH3), 3.92 (s, 3H, OCH3), 7.08 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.77 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.96 (s, 1H, Ar-H, benzoate ring), 10.52 (br s, 1H, NH), and 13.98 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 11.8, 16.4, 22.5, 29.1, 45.1, 53.1, 114, 118.8, 122.2, 126.9, 128.3, 134.5, 145.2, 159.3, 169.4, and 196.2 (16C). ESI-MS: m/z, 320.14 (M+ + H). Anal. calcd for C16H21N3O4 (319.36): C, 60.17; H, 6.63; and N, 13.16. Found: C, 60.00; H, 6.56; and N, 13.10.


### 2.1.2. General Procedure for the Synthesis of Compounds6–8

A mixture of 1-aminonaphthalene (0.48 g, 3.4 mmol) and conc. HCl (5 mL) was stirred with gentle heating until complete dissolving. The reaction mixture was cooled at a temperature from zero to −4°C in an ice bath, and then sodium nitrite (0.28 g, 4 mmol) in water (2 mL) was added portionwise to the reaction mixture at the same temperature to form the diazonium salt5. To the diazonium salt 5, a solution of appropriate amide or iminoderivatives [27], namely, N-substituted 3-oxo-butanamide, ethyl 3-(substituted imino)butanoate, or N-alkyl-3-(alkyl imino)-butanamide in absolute ethanol (10 mL), was added while stirring at a temperature of −4°C, and the mixture was stirred at room temperature for an additional 3 hours. The solid formed was collected by filtration, washed with cold water several times, dried, and crystallized from dimethylformamide/ethanol to give the corresponding 6a–h, 7a,b, and 8a,b, respectively.(1) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-phenethylbut-2-enamide
(6a). Brownish crystalline solid, m.p. 141-142°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.50 (s, 3H, C-CH3), 2.88 (t, 2H, J = 7.2 Hz, ArCH2CH2NH), 3.62 (q, 2H, J = 7.0 Hz, ArCH2CH2NH), 7.23–7.20 (m, 1H, Ar), 7.33–7.30 (m, 4H, ArH), 7.63–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 6.7 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.04 (d, 1H, J = 7.9 Hz, naphthyl-H), 9.40 (br s, 1H, NHCO), and 15.67 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 26.4, 35.2, 40.7, 110.9, 119.8, 123.1, 125.1, 126.8, 126.9, 127.2, 127.5, 128.1, 128.9, 129.2, 129.3, 134.2, 136.9, 139.5, 164.9, and 198.7 (22C). ESI- MS: m/z, 360.17 (M+ + H). Anal. calcd for C22H21N3O2 (359.42): C, 73.52; H, 5.89; and N, 11.69. Found: C, 73.45; H, 5.80; and N, 11.60.(2) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-methylbut-2-enamide
(6b). Brownish crystalline solid, m.p. 175-176°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm) 2.52 (s, 3H, C-CH3), 2.88 (d, 3H, J = 4.9 Hz, CONHCH3), 7.63–7.60 (m, 2H, naphthyl-H), 7.72–7.68 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.2 Hz, naphthyl-H), 7.83 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.92 (d, 1H, J = 8.0 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.23 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 25.8, 26.4, 110.8, 119.8, 123.0, 124.9, 126.9, 127.1, 127.5, 128.3, 129.3, 134.2, 136.9, 165.5, and 198.5 (15C). ESI-MS: m/z, 270.12 (M+ + H). Anal. calcd for C15H15N3O2 (269.30): C, 66.90; H, 5.61; and N, 15.60. Found: C, 66.80; H, 5.50; and N, 15.52.(3) 2-(Naphthalen-1-yl-diazenyl)-N-tert-butyl-3-hydroxybut-2-enamide
(6c). Brownish crystalline solid, m.p. 126-127°C, yield = 59%, and 1H-NMR (DMSO-d6) δ (ppm) 1.43 (s, 9H, C(CH3)3), 2.52 (s, 3H, C-CH3), 7.63–7.60 (m, 2H, naphthyl-H), 7.74–7.40 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.90 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.42 (s, 1H, NHCO), and 15.69 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 26.5, 28.8, 51.3, 111.0, 119.7, 123.1, 125.1, 126.9, 127.1, 127.6, 128.4, 129.2, 134.2, 136.9, 164.7, and 199.3 (18C). ESI-MS: m/z, 312.17 (M+ + H). Anal. calcd for C18H21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.30; H, 6.75; and N, 13.40.(4) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-isopropylbut-2-enamide
(6d). Brownish crystalline solid, m.p. 124-125°C, yield = 62%, and 1H-NMR (DMSO-d6) δ (ppm) 1.23 (d, 6H, J = 6.6 Hz, CH(CH3)2), 2.52 (s, 3H, C-CH3), 4.11–4.00 (m, 1H, CH(CH3)2), 7.65–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 6.8 Hz, naphthyl-H), 7.90 (d, 1H, J = 8.3 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.26 (br s, 1H, NHCO), and 15.71 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 22.6, 40.8, 49.1, 110.9, 119.7, 123.1, 125.1, 127.0, 127.1, 127.5, 128.1, 129.3, 134.2, 136.9, 164.2, and 199.0 (17C). ESI-MS: m/z, 280.15 (M+ + H). Anal. calcd for C17H19N3O2 (297.35): C, 68.67; H, 6.44; and N, 14.13. Found: C, 68.60; H, 6.34; and N, 14.00.(5) 2-(Naphthalen-1-yl-diazenyl)-N-ethyl-3-hydroxybut-2-enamide
(6e). Brownish crystalline solid, m.p. 130–131°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.17 (t, 3H, J = 7.2 Hz, NHCH2CH3), 2.52 (s, 3H, C-CH3), 3.38–3.34 (m, 2H, NHCH2CH3), 7.64–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.33 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.9, 26.5, 33.60, 110.9, 119.8, 123.0, 125.0, 126.9, 127.1, 127.5, 128.2, 129.3, 134.2, 136.9, 164.8, and 198.7 (16C). ESI-MS: m/z, 284.14 (M+ + H). Anal. calcd for C16H17N3O2 (283.33): C, 67.83; H, 6.05; and N, 14.83. Found: C, 67.74; H, 6.00; and N, 14.78.(6) 2-(Naphthalen-1-yl-diazenyl)-N-butyl-3-hydroxybut-2-enamide
(6f). Brownish crystalline solid, m.p. 142-143°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm) 0.93 (t, 3H, J = 7.3 Hz, NHCH2CH2CH2CH3), 1.38–1.35 (m, 2H, J = 7.6 Hz, NHCH2CH2CH2CH3), 1.57–1.54 (m, 2H, J = 7.3 Hz, NHCH2CH2CH2CH3) 2.53 (s, 3H, C-CH3), 3.36 (q, 2H, J = 7.0 Hz, NHCH2CH2CH2CH3), 7.65–1.62 (m, 2H, naphthyl-H), 7.75–7.71 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.36 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.1, 20.2, 26.5, 31.3, 38.3, 110.9, 119.8, 123.1, 125.1, 126.9, 127.1, 127.5, 128.2, 129.3, 134.2, and 136.9, 164.9, 198.8 (18C). ESI-MS: m/z, 312.17 (M+ + H). Anal. calcd for C18H21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.35; H, 6.72; and N, 13.40.(7) 2-(Naphthalen-1-yl-diazenyl)-N-sec-butyl-3-hydroxybut-2-enamide
(6g). Yellow crystalline solid, m.p. 105-106°C, yield = 55%, and 1H-NMR (CDCl3) δ (ppm) 1.01 (t, 3H, J = 7.4 Hz, CH3CH2CH(NH)CH3), 1.28 (d, 3H, J = 6.6 Hz, CH3CH2CH(NH)CH3), 1.66–1.60 (m, 2H, CH3CH2CH(NH)CH3), 2.62 (s, 3H, C-CH3), 4.12–4.08 (m, 1H, CH3CH2CH(NH)CH3) 7.56–7.52 (m, 3H, naphthyl-H), 7.68 (d, 1H, J = 8.0 Hz, naphthyl-H), 7.82 (d, 1H, J = 8.0 Hz, naphthyl-H), 7.90 (d, 1H, J = 7.6 Hz naphthyl-H), 8.07 (d, 1H, J = 7.9 Hz naphthyl-H), 9.40 (br s, 1H, NHCO), and 15.81 (s, 1H, OH); 13C-NMR (CDCl3) δ (ppm). 10.4, 20.3, 26.2, 29.4, 46.0, 110.7, 120.2, 123.5, 124.8, 126.1, 126.4, 126.7, 127.6, 128.6, 134.1, 137.2, 164.7, and 199.4 (18C). ESI-MS: m/z, 312.16 (M+ + H). Anal. calcd for C18N21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.35; H, 6.72; and N, 13.40.(8) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-propylbut-2-enamide
(6h). Brownish crystalline solid, m.p. 157-158°C, yield = 65%, and 1H-NMR (CDCl3) δ (ppm) 1.04 (t, 3H, J = 7.4 Hz, CONHCH2CH2CH3), 1.68–1.63 (m, 2H, J = 7.3 Hz, CONHCH2CH2CH3), 2.62 (s, 3H, C-CH3), 3.42 (q, 2H, J = 7.0 Hz, CONHCH2CH2CH3), 7.57–7.53 (m, 3H, naphthyl-H), 7.68 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.90 (d, 1H, J = 7.5 Hz, naphthyl-H), 8.07 (br d, 1H, J = 8.5 Hz, naphthyl-H), 9.54 (br s, 1H, NHCO), and 15.79 (s, 1H, OH); 13C-NMR (CDCl3) δ (ppm) 11.6, 22.6, 26.2, 40.4, 110.7, 120.2, 123.5, 124.9, 126.1, 126.4, 126.7, 127.6, 128.6, 134.1, 137.1, 165.3, and 199.4 (17C). ESI-MS: m/z, 298.15 (M+ + H). Anal. calcd for C17H19N3O2 (297.35): C, 68.67; H, 6.44; and N, 14.13. Found: C, 68.55; H, 6.36; and N, 14.04.(9) Ethoxy 3-(benzylimino)-2-(naphthalen-1-yl-diazenyl)but-1-en-1-ol
(7a). Brownish crystalline solid, m.p. 110-111°C, yield = 77%, and 1H-NMR (DMSO-d6) δ (ppm) 1.32 (t, 3H, J = 7.1 Hz, OCH2CH3), 2.59 (s, 3H, C-CH3), 4.27 (q, 2H, J = 7.1 Hz, OCH2CH3), 4.83 (s, 2H, J = 3.4 Hz, ArCH2N), 7.50–4.46 (m, 9H, naphthyl-H and ArH), 7.65 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.72 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.88 (d, 1H, J = 8.2 Hz, naphthyl-H), and 14.90 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.9, 17.2, 50.2, 60.3, 111.2, 121.9, 124.1, 126.1, 126.4, 126.4, 126.6, 127.2, 128.1, 128.5, 129.3, 129.5, 134.3, 136.9, 137.9, 166.7, and 198.7 (23C). ESI-MS: m/z, 374.19 (M+ + H). Anal. calcd for C23H23N3O2 (373.45): C, 73.97; H, 6.21; and N, 11.25. Found: C, 73.85; H, 6.12; and N, 11.20.(10) Ethoxy 2-(naphthalen-1-yl-diazenyl)-3-(phenethylimino)but-1-en-1-ol
(7b). Brownish crystalline solid, m.p. 142-143°C, yield = 72%, and 1H-NMR (DMSO-d6) δ (ppm) 1.30 (t, 3H, J = 7.1 Hz, OCH2CH3), 2.48 (s, 3H, C-CH3), 3.08 (t, 2H, J = 7.1 Hz, ArCH2CH2N), 3.89 (t, 2H, J = 6.9 Hz, ArCH2CH2N), 4.23 (q, 3H, J = 7.1 Hz, OCH2CH3), 7.20–7.15 (m, 1H, naphthyl-H), 7.28–7.24 (m, 2H, naphthyl-H), 7.32–7.28 (m, 2H, naphthyl-H), 7.58–7.55 (m, 4H, ArH), 7.78–7.74 (m, 1H, naphthyl-H), 7.96–7.92 (m, 1H, naphthyl-H), and 14.59 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm). 14.9, 16.5, 35.9, 46.4, 60.1, 111.9, 122.6, 125.1, 126.3, 126.5, 126.7, 126.9, 127.5, 128.6, 128.9, 129.2, 129.2, 134.4, 136.9, 138.9, 166.7, and 198.7 (24C). ESI-MS: m/z, 388.19 (M+ + H). Anal. calcd for C24H25N3O2 (387.47): C, 74.39; H, 6.50; and N, 10.84. Found: C, 74.18; H, 6.42; and N, 10.76.(11) 1-(Butylamino)-3-(butylimino)-2-[(1-(naphthalen-1-yl)diazene]but-1-en-1-ol
(8a). Brownish crystalline solid, m.p. 91-92°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm). 0.93 (t, 3H, J = 6.4 Hz, NHCH2CH2CH2CH3), 0.96 (t, 3H, J = 7.3 Hz, NCH2CH2CH2CH3), 1.46–1.40 (m, 4H, J = 7.7 Hz, NHCH2CH2CH2CH3 and NCH2CH2CH2CH3), 1.61 (q, 2H, NHCH2CH2CH2CH3), 1.67–1.64 (m, 2H, NCH2CH2CH2CH3), 2.62 (s, 3H, C-CH3), 3.39 (q, 2H, J = 6.4 Hz, NHCH2CH2CH2CH3), 3.65 (q, 2H, J = 6.8 Hz, NCH2CH2CH2CH3), 7.56–7.52 (m, 4H, naphthyl-H), 7.76 (d, 1H, J = 8.2 Hz, naphthyl-H), 7.95–7.90 (m, 1H, naphthyl-H), 8.4 (d, 1H, J = 7.4 Hz, naphthyl-H), 11.3 (br s, 1H, NH), and 15.37 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.1, 14.2, 14.7, 20.0, 20.4, 31.4, 31.7, 37.9, 44.0, 111.3, 121.3, 122.8, 126.0, 126.3, 126.8, 126.8, 128.6, 129.0, 134.5, 148.7, 165.9, and 172.7 (22C). ESI-MS: m/z, 367.25 (M+ + H). Anal. calcd for C22H30N4O (366.50): C, 72.10; H, 8.25; and N, 15.29. Found: C, 72.00; H, 8.18; and N, 15.20.(12) N-(2-hydroxyethyl imino)-3-(2-hydroxyethylamino)-2-(naphthalen-1-yldiazenyl)but-2-en-2-ol
(8b). Brownish crystalline solid, m.p. 157-158°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.64 (s, 3H, C-CH3), 3.47 (q, 3H, J = 5.3 Hz, CONHCH2CH2OH), 3.62–3.66 (m, 6H, CONHCH2CH2OH and CNCH2CH2OH), 4.95 (t, 1H, J = 4.6 Hz, CONHCH2CH2OH), 5.08 (t, 1H, J = 4.4 Hz, CNCH2CH2OH), 7.54–7.50 (m, 4H, naphthyl-H), 7.74 (d, 1H, J = 7.8 Hz, naphthyl-H), 7.94–7.88 (m, 1H, naphthyl-H), 8.74–8.70 (m, 1H, naphthyl-H), 11.40 (br s, 1H, NH), and 15.37 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm).15.0, 41.1, 47.1, 60.0, 60.5, 111.1, 121.3, 123.9, 126.3, 126.4, 126.7, 126.8, 128.2, 129.2, 134.5, 148.8, 165.8, and 172.8 (18C). ESI-MS: m/z, 343.18 (M+ + H). Anal. calcd for C18H22N4O3 (342.39): C, 63.14; H, 6.48; and N, 16.36. Found: C, 63.00; H, 6.40; and N, 16.30.


## 2.1.1. General Procedure for the Synthesis of Compounds4a–k

A mixture of methyl 5-aminosalicylate1 (0.56 g, 3.4 mmol) in concentrated hydrochloric acid (5 mL) was stirred with gentle heating until complete dissolving. Then, the mixture was cooled at a temperature from zero to −4°C, sodium nitrite (0.28 g, 4 mmol) in 2 mL of water was added portionwise at a temperature of −4°C to form the corresponding diazonium salt 2, and the diazonium salt formed was added to a mixture of 4-(substituted imino)pentan-2-one derivatives 3 (2 mmol) [27] in ethanol (15 mL) slowly under the same temperature allowing the mixture to stir at room temperature for an additional 3 hours. The solid formed was collected by filtration, washed with cold water several times, and then left to dry. The obtained product was recrystallized from ethanol to give target compounds 4a–k, respectively.(1) Methyl 5-(4-(benzylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4a). Yellow crystalline solid, m.p. 167-168°C, yield = 62%, and 1H-NMR (DMSO-d6) δ (ppm) 2.42 (s, 3H, CH3-CNH), 2.57 (s, 3H, CO-CH3), 3.93 (s, 3H, OCH3), 4.82 (s, 2H, Ar-CH2), 7.05 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.36–7.42 (m, 5H, ArH), 7.72 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.78 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.40 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.8, 29.0, 47.6, 53.0, 113.8, 118.9, 123.0, 125.6, 128.1, 128.2, 128.6, 129.4, 137.3, 144.5,159.1, 159.4, 169.3, and 196.4 (20C). ESI-MS: m/z, 368.16 (M+ + H). Anal. calcd for C20H21N3O4 (367.40): C, 65.38; H, 5.76; and N, 11.44. Found: C, 65.35; H, 5.78; and N, 11.40.(2) Methyl 5-(2-oxo-4-(phenethylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4b). Yellow crystalline solid, m.p. 131-132°C, yield = 60%, and 1H-NMR (DMSO-d6) δ (ppm) 2.39 (s, 3H, CH3-CNH), 2.52 (s, 3H, CO-CH3), 2.98 (t, 2H, J = 6.8 Hz, PhCH2CH2N), 3.85 (t, 2H, J = 6.7 Hz, ArCH2CH2N), 3.95 (s, 3H, OCH3), 7.03 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.34–7.30 (m, 5H, ArH), 7.59 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.76 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 13.84 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 29.1, 35.0, 44.6, 53.1, 113.8, 118.7, 122.4, 126.6, 127.0, 128.3, 129.0, 129.2, 138.7, 144.9, 159.1, 159.3, 169.5, and 196.3 (21C). ESI-MS: m/z, 382.18 (M+ + H). Anal. calcd for C21H23N3O4 (381.43): C, 66.13; H, 6.08; and N, 11.02. Found: C, 66.45; H, 6.02; and N, 11.00.(3) Methyl 5-(2-oxo-4-(1-phenylethylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4c). Yellow crystalline solid, m.p. 110-111°C, yield = 66%, and 1H-NMR (DMSO-d6) δ (ppm) 1.60 (d, 3H, J = 7.6 Hz, ArCH(CH3)N), 2.41 (s, 3H, CH3-C), 2.49 (s, 3H, CO-CH3), 3.94 (s, 3H, OCH3), 5.17 (q, 1H, J = 6.7 Hz, ArCH(CH3)N), 7.06 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.40–7.45 (m, 5H, ArH), 7.76 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.86 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.63 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.8, 24.0, 29.0, 53.1, 53.7, 114.0, 119.0, 122.2, 126.4, 126.6, 128.2, 128.5, 129.4, 143.0, 144.5, 159.2, 159.4, 169.3, and 196.4 (21C). ESI-MS: m/z, 382.18 (M+ + H). Anal. calcd for C21H23N3O4 (381.43): C, 66.13; H, 6.08; and N, 11.02. Found: C, 66.04; H, 6.03; and N, 10.98.(4) Methyl 5-(4-(2-hydroxyethylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4d). Yellow crystalline solid, m.p. 187-188°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.42 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.59 (t, 2H, J = 5.0 Hz, NCH2CH2OH), 3.66 (t, 2H, J = 4.2 Hz, NCH2CH2OH), 3.92 (s, 3H, OCH3), 5.13 (br s, 1H, NCH2CH2OH) 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.79 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 8.01 (s, 1H, Ar-H, benzoate ring), and 10.5 (br s, 1H, NH), 14.21 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.7, 29.1, 46.0, 53.1, 59.7, 113.9, 118.7, 122.1, 127.0, 128.4, 134.5, 154.1, 159.3, 169.5, and 196.4 (15C). ESI-MS: 322.27 (M+ + H). Anal. calcd for C15H19N3O5 (321.33): C, 56.07; H, 5.96; and N, 13.08. Found: C, 56.00; H, 5.92; and N, 13.00.(5) Methyl 5-(4-(isobutylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4e). Yellow crystalline solid, m.p. 94–95°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.00 (d, 6H, J = 6.7 Hz, NHCH2CH(CH3)2), 1.95–1.90 (m, 1H, NHCH2CH(CH3)2), 2.42 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.39 (dd, 2H, J = 5.4 Hz, NHCH2CH(CH3)2), 3.92 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.76 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.95 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.09 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.45, 2.36, 28.3, 29.1, 50.7, 53.1, 114.0, 118.9, 122.0, 126.8, 128.4, 134.5, 145.2, 159.3, 169.4, and 196.2 (17C). ESI-MS: m/z, 334.18 (M+ + H). Anal. calcd for C17H23N3O4 (333.38): C, 61.25; H, 6.95; and N, 12.60. Found: C, 61.12; H, 6.90; and N, 12.54.(6) Methyl 5-(4-(isopropylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4f). Yellow crystalline solid, m.p. 104-105°C, yield = 61%, and 1H-NMR (DMSO-d6) δ (ppm) 1.30 (d, 6H, J = 6.4 Hz, NHCH(CH3)2), 2.41 (s, 3H, CH3-C), 2.56 (s, 3H, COCH3), 3.92 (s, 3H, OCH3), 4.15–4.12 (m, 1H, J = 6.4 Hz, NHCH(CH3)2), 7.08 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.92 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.10 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 23.2, 29.1, 45.6, 53.1, 114.0, 118.8, 121.8, 127.1, 127.9, 134.5, 145.1, 159.3, 169.4, and 196.1 (16C). ESI-MS: m/z, 320.16 (M+ + H). Anal. calcd for C16H21N3O4 (319.36): C, 60.17; H, 6.63; and N, 13.16. Found: C, 60.10; H, 6.58; and N, 13.10.(7) Methyl 5-(4-(butylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4g). Yellow crystalline solid, m.p. 131-132°C, yield = 65%, and 1H-NMR (DMSO-d6) δ (ppm) 0.97 (t, 3H, J = 7.3 Hz, NHCH2CH2CH2CH3), 1.43-1.40 (m, 2H, J = 7.6 Hz, NHCH2CH2CH2CH3), 1.62–1.66 (m, 2H, J = 6.9 Hz, NHCH2CH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.53 (s, 3H, COCH3), 3.52 (t, 2H, J = 6.7 Hz, NHCH2CH2CH2CH3), 3.91 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.72 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.05 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.0, 16.4, 20.2, 29.0, 31.0, 43.1, 53.1, 113.9, 118.9, 122.6, 126.5, 128.2, 134.5, 145.1, 159.3, 169.4, and 196.2 (17C). ESI-MS: m/z, 334.18 (M+ + H). Anal. calcd for C17H23N3O4 (333.38): C, 61.25; H, 6.95; and N, 12.60. Found: C, 61.12; H, 6.86; and N, 12.52.(8) Methyl 5-(4-(3-hydroxypropylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4h). Yellow crystalline solid, m.p. 135-136°C, yield = 63%, and 1H-NMR (DMSO-d6) δ (ppm) 1.80–1.75 (m, 2H, J = 6.3 Hz, NHCH2CH2CH2OH), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.54 (br t, 2H, J = 5.7 Hz, NHCH2CH2CH2OH), 3.59 (br t, 2H, J = 6.7 Hz, NHCH2CH2CH2OH), 3.91 (s, 3H, OCH3), 4.70 (br s, 2H, NHCH2CH2CH2OH), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.77 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.98 (s, 1H, Ar-H, benzoate ring), and 10.50 (br s, 1H, NH), 13.95 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 16.3, 29.1, 32.2, 40.0, 53.1, 58.4, 113.9, 118.7, 122.0, 127.1, 128.3, 134.5, 145.2, 159.3, 169.5, and 196.2 (16C). ESI-MS): m/z, 336.16 (M+ + H). Anal. calcd for C16H21N3O5 (335.36): C, 57.30; H, 6.31; and N, 12.53. Found: C, 57.10; H, 6.22; and N, 12.42.(9) Methyl 5-(2-oxo-4-(pentylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4i). Yellow crystalline solid, m.p. 127-128°C, yield = 72%, and 1H-NMR (DMSO-d6) δ (ppm) 0.92 (t, 3H, J = 6.9 Hz, NHCH2CH2CH2CH2CH3), 1.40–1.45 (m, 4H, NHCH2CH2CH2CH2CH3), 1.66–1.62 (m, 2H, NHCH2CH2CH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.53 (t, 2H, J = 6.7 Hz, NHCH2CH2CH2CH2CH3), 3.91 (s, 3H, OCH3), 7.06 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 14.05 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.3, 16.4, 22.3, 28.6, 29.2, 30.7, 43.4, 53.0, 113.8, 118.9, 122.7, 126.3, 128.2, 134.5, 145.1, 159.4, 169.4, and 196.2 (18C). ESI-MS: m/z, 348.19 (M+ + H). Anal. calcd for C18H25N3O4 (347.41): C, 62.23; H, 7.25; and N, 12.10. Found: C, 62.05; H, 7.16; and N, 12.05.(10) Methyl 5-(4-(ethylamino)-2-oxopent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4j). Yellow crystalline solid, m.p. 106-107°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.27 (t, 3H, J = 7.2 Hz, NHCH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.55 (q, 2H, NHCH2CH3), 3.91 (s, 3H, OCH3), 7.07 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.78 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.93 (s, 1H, Ar-H, benzoate ring), 10.50 (br s, 1H, NH), and 13.88 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.8, 16.3, 29.1, 38.3, 53.1, 113.9, 118.8, 122.3, 126.9, 128.1, 134.5, 145.3, 159.4, 169.5, and 196.1 (15C). ESI-MS): m/z, 306.15 (M+ + H). Anal. calcd for C15H19N3O4 (305.33): C, 59.01; H, 6.27; and N, 13.76. Found: C, 58.86; H, 6.20; and N, 13.70.(11) Methyl 5-(2-oxo-4-(propylamino)pent-3-en-3-yl)diazenyl)-2-hydroxybenzoate
(4k). Yellow crystalline solid, m.p. 137-138°C, yield = 69%, and 1H-NMR (DMSO-d6) δ (ppm) 0.99 (t, 3H, J = 7.3 Hz, NHCH2CH2CH3), 1.68–1.64 (m, 3H, J = 7.2 Hz, NHCH2CH2CH3), 2.41 (s, 3H, CH3-C), 2.54 (s, 3H, CO-CH3), 3.50 (q, 2H, J = 6.9 Hz, NHCH2CH3), 3.92 (s, 3H, OCH3), 7.08 (d, 1H, J = 8.8 Hz, Ar-H, benzoate ring), 7.77 (d, 1H, J = 2.5 Hz, Ar-H, benzoate ring), 7.96 (s, 1H, Ar-H, benzoate ring), 10.52 (br s, 1H, NH), and 13.98 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 11.8, 16.4, 22.5, 29.1, 45.1, 53.1, 114, 118.8, 122.2, 126.9, 128.3, 134.5, 145.2, 159.3, 169.4, and 196.2 (16C). ESI-MS: m/z, 320.14 (M+ + H). Anal. calcd for C16H21N3O4 (319.36): C, 60.17; H, 6.63; and N, 13.16. Found: C, 60.00; H, 6.56; and N, 13.10.


## 2.1.2. General Procedure for the Synthesis of Compounds6–8

A mixture of 1-aminonaphthalene (0.48 g, 3.4 mmol) and conc. HCl (5 mL) was stirred with gentle heating until complete dissolving. The reaction mixture was cooled at a temperature from zero to −4°C in an ice bath, and then sodium nitrite (0.28 g, 4 mmol) in water (2 mL) was added portionwise to the reaction mixture at the same temperature to form the diazonium salt5. To the diazonium salt 5, a solution of appropriate amide or iminoderivatives [27], namely, N-substituted 3-oxo-butanamide, ethyl 3-(substituted imino)butanoate, or N-alkyl-3-(alkyl imino)-butanamide in absolute ethanol (10 mL), was added while stirring at a temperature of −4°C, and the mixture was stirred at room temperature for an additional 3 hours. The solid formed was collected by filtration, washed with cold water several times, dried, and crystallized from dimethylformamide/ethanol to give the corresponding 6a–h, 7a,b, and 8a,b, respectively.(1) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-phenethylbut-2-enamide
(6a). Brownish crystalline solid, m.p. 141-142°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.50 (s, 3H, C-CH3), 2.88 (t, 2H, J = 7.2 Hz, ArCH2CH2NH), 3.62 (q, 2H, J = 7.0 Hz, ArCH2CH2NH), 7.23–7.20 (m, 1H, Ar), 7.33–7.30 (m, 4H, ArH), 7.63–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 6.7 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.04 (d, 1H, J = 7.9 Hz, naphthyl-H), 9.40 (br s, 1H, NHCO), and 15.67 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 26.4, 35.2, 40.7, 110.9, 119.8, 123.1, 125.1, 126.8, 126.9, 127.2, 127.5, 128.1, 128.9, 129.2, 129.3, 134.2, 136.9, 139.5, 164.9, and 198.7 (22C). ESI- MS: m/z, 360.17 (M+ + H). Anal. calcd for C22H21N3O2 (359.42): C, 73.52; H, 5.89; and N, 11.69. Found: C, 73.45; H, 5.80; and N, 11.60.(2) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-methylbut-2-enamide
(6b). Brownish crystalline solid, m.p. 175-176°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm) 2.52 (s, 3H, C-CH3), 2.88 (d, 3H, J = 4.9 Hz, CONHCH3), 7.63–7.60 (m, 2H, naphthyl-H), 7.72–7.68 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.2 Hz, naphthyl-H), 7.83 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.92 (d, 1H, J = 8.0 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.23 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 25.8, 26.4, 110.8, 119.8, 123.0, 124.9, 126.9, 127.1, 127.5, 128.3, 129.3, 134.2, 136.9, 165.5, and 198.5 (15C). ESI-MS: m/z, 270.12 (M+ + H). Anal. calcd for C15H15N3O2 (269.30): C, 66.90; H, 5.61; and N, 15.60. Found: C, 66.80; H, 5.50; and N, 15.52.(3) 2-(Naphthalen-1-yl-diazenyl)-N-tert-butyl-3-hydroxybut-2-enamide
(6c). Brownish crystalline solid, m.p. 126-127°C, yield = 59%, and 1H-NMR (DMSO-d6) δ (ppm) 1.43 (s, 9H, C(CH3)3), 2.52 (s, 3H, C-CH3), 7.63–7.60 (m, 2H, naphthyl-H), 7.74–7.40 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.90 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.42 (s, 1H, NHCO), and 15.69 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 26.5, 28.8, 51.3, 111.0, 119.7, 123.1, 125.1, 126.9, 127.1, 127.6, 128.4, 129.2, 134.2, 136.9, 164.7, and 199.3 (18C). ESI-MS: m/z, 312.17 (M+ + H). Anal. calcd for C18H21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.30; H, 6.75; and N, 13.40.(4) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-isopropylbut-2-enamide
(6d). Brownish crystalline solid, m.p. 124-125°C, yield = 62%, and 1H-NMR (DMSO-d6) δ (ppm) 1.23 (d, 6H, J = 6.6 Hz, CH(CH3)2), 2.52 (s, 3H, C-CH3), 4.11–4.00 (m, 1H, CH(CH3)2), 7.65–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.78 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.83 (d, 1H, J = 6.8 Hz, naphthyl-H), 7.90 (d, 1H, J = 8.3 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.26 (br s, 1H, NHCO), and 15.71 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 22.6, 40.8, 49.1, 110.9, 119.7, 123.1, 125.1, 127.0, 127.1, 127.5, 128.1, 129.3, 134.2, 136.9, 164.2, and 199.0 (17C). ESI-MS: m/z, 280.15 (M+ + H). Anal. calcd for C17H19N3O2 (297.35): C, 68.67; H, 6.44; and N, 14.13. Found: C, 68.60; H, 6.34; and N, 14.00.(5) 2-(Naphthalen-1-yl-diazenyl)-N-ethyl-3-hydroxybut-2-enamide
(6e). Brownish crystalline solid, m.p. 130–131°C, yield = 75%, and 1H-NMR (DMSO-d6) δ (ppm) 1.17 (t, 3H, J = 7.2 Hz, NHCH2CH3), 2.52 (s, 3H, C-CH3), 3.38–3.34 (m, 2H, NHCH2CH3), 7.64–7.60 (m, 2H, naphthyl-H), 7.74–7.70 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.6 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.33 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.9, 26.5, 33.60, 110.9, 119.8, 123.0, 125.0, 126.9, 127.1, 127.5, 128.2, 129.3, 134.2, 136.9, 164.8, and 198.7 (16C). ESI-MS: m/z, 284.14 (M+ + H). Anal. calcd for C16H17N3O2 (283.33): C, 67.83; H, 6.05; and N, 14.83. Found: C, 67.74; H, 6.00; and N, 14.78.(6) 2-(Naphthalen-1-yl-diazenyl)-N-butyl-3-hydroxybut-2-enamide
(6f). Brownish crystalline solid, m.p. 142-143°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm) 0.93 (t, 3H, J = 7.3 Hz, NHCH2CH2CH2CH3), 1.38–1.35 (m, 2H, J = 7.6 Hz, NHCH2CH2CH2CH3), 1.57–1.54 (m, 2H, J = 7.3 Hz, NHCH2CH2CH2CH3) 2.53 (s, 3H, C-CH3), 3.36 (q, 2H, J = 7.0 Hz, NHCH2CH2CH2CH3), 7.65–1.62 (m, 2H, naphthyl-H), 7.75–7.71 (m, 1H, naphthyl-H), 7.77 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.91 (d, 1H, J = 8.4 Hz, naphthyl-H), 8.02 (d, 1H, J = 8.1 Hz, naphthyl-H), 9.36 (br s, 1H, NHCO), and 15.72 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.1, 20.2, 26.5, 31.3, 38.3, 110.9, 119.8, 123.1, 125.1, 126.9, 127.1, 127.5, 128.2, 129.3, 134.2, and 136.9, 164.9, 198.8 (18C). ESI-MS: m/z, 312.17 (M+ + H). Anal. calcd for C18H21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.35; H, 6.72; and N, 13.40.(7) 2-(Naphthalen-1-yl-diazenyl)-N-sec-butyl-3-hydroxybut-2-enamide
(6g). Yellow crystalline solid, m.p. 105-106°C, yield = 55%, and 1H-NMR (CDCl3) δ (ppm) 1.01 (t, 3H, J = 7.4 Hz, CH3CH2CH(NH)CH3), 1.28 (d, 3H, J = 6.6 Hz, CH3CH2CH(NH)CH3), 1.66–1.60 (m, 2H, CH3CH2CH(NH)CH3), 2.62 (s, 3H, C-CH3), 4.12–4.08 (m, 1H, CH3CH2CH(NH)CH3) 7.56–7.52 (m, 3H, naphthyl-H), 7.68 (d, 1H, J = 8.0 Hz, naphthyl-H), 7.82 (d, 1H, J = 8.0 Hz, naphthyl-H), 7.90 (d, 1H, J = 7.6 Hz naphthyl-H), 8.07 (d, 1H, J = 7.9 Hz naphthyl-H), 9.40 (br s, 1H, NHCO), and 15.81 (s, 1H, OH); 13C-NMR (CDCl3) δ (ppm). 10.4, 20.3, 26.2, 29.4, 46.0, 110.7, 120.2, 123.5, 124.8, 126.1, 126.4, 126.7, 127.6, 128.6, 134.1, 137.2, 164.7, and 199.4 (18C). ESI-MS: m/z, 312.16 (M+ + H). Anal. calcd for C18N21N3O2 (311.38): C, 69.43; H, 6.80; and N, 13.49. Found: C, 69.35; H, 6.72; and N, 13.40.(8) 2-(Naphthalen-1-yl-diazenyl)-3-hydroxy-N-propylbut-2-enamide
(6h). Brownish crystalline solid, m.p. 157-158°C, yield = 65%, and 1H-NMR (CDCl3) δ (ppm) 1.04 (t, 3H, J = 7.4 Hz, CONHCH2CH2CH3), 1.68–1.63 (m, 2H, J = 7.3 Hz, CONHCH2CH2CH3), 2.62 (s, 3H, C-CH3), 3.42 (q, 2H, J = 7.0 Hz, CONHCH2CH2CH3), 7.57–7.53 (m, 3H, naphthyl-H), 7.68 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.82 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.90 (d, 1H, J = 7.5 Hz, naphthyl-H), 8.07 (br d, 1H, J = 8.5 Hz, naphthyl-H), 9.54 (br s, 1H, NHCO), and 15.79 (s, 1H, OH); 13C-NMR (CDCl3) δ (ppm) 11.6, 22.6, 26.2, 40.4, 110.7, 120.2, 123.5, 124.9, 126.1, 126.4, 126.7, 127.6, 128.6, 134.1, 137.1, 165.3, and 199.4 (17C). ESI-MS: m/z, 298.15 (M+ + H). Anal. calcd for C17H19N3O2 (297.35): C, 68.67; H, 6.44; and N, 14.13. Found: C, 68.55; H, 6.36; and N, 14.04.(9) Ethoxy 3-(benzylimino)-2-(naphthalen-1-yl-diazenyl)but-1-en-1-ol
(7a). Brownish crystalline solid, m.p. 110-111°C, yield = 77%, and 1H-NMR (DMSO-d6) δ (ppm) 1.32 (t, 3H, J = 7.1 Hz, OCH2CH3), 2.59 (s, 3H, C-CH3), 4.27 (q, 2H, J = 7.1 Hz, OCH2CH3), 4.83 (s, 2H, J = 3.4 Hz, ArCH2N), 7.50–4.46 (m, 9H, naphthyl-H and ArH), 7.65 (d, 1H, J = 7.5 Hz, naphthyl-H), 7.72 (d, 1H, J = 8.1 Hz, naphthyl-H), 7.88 (d, 1H, J = 8.2 Hz, naphthyl-H), and 14.90 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.9, 17.2, 50.2, 60.3, 111.2, 121.9, 124.1, 126.1, 126.4, 126.4, 126.6, 127.2, 128.1, 128.5, 129.3, 129.5, 134.3, 136.9, 137.9, 166.7, and 198.7 (23C). ESI-MS: m/z, 374.19 (M+ + H). Anal. calcd for C23H23N3O2 (373.45): C, 73.97; H, 6.21; and N, 11.25. Found: C, 73.85; H, 6.12; and N, 11.20.(10) Ethoxy 2-(naphthalen-1-yl-diazenyl)-3-(phenethylimino)but-1-en-1-ol
(7b). Brownish crystalline solid, m.p. 142-143°C, yield = 72%, and 1H-NMR (DMSO-d6) δ (ppm) 1.30 (t, 3H, J = 7.1 Hz, OCH2CH3), 2.48 (s, 3H, C-CH3), 3.08 (t, 2H, J = 7.1 Hz, ArCH2CH2N), 3.89 (t, 2H, J = 6.9 Hz, ArCH2CH2N), 4.23 (q, 3H, J = 7.1 Hz, OCH2CH3), 7.20–7.15 (m, 1H, naphthyl-H), 7.28–7.24 (m, 2H, naphthyl-H), 7.32–7.28 (m, 2H, naphthyl-H), 7.58–7.55 (m, 4H, ArH), 7.78–7.74 (m, 1H, naphthyl-H), 7.96–7.92 (m, 1H, naphthyl-H), and 14.59 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm). 14.9, 16.5, 35.9, 46.4, 60.1, 111.9, 122.6, 125.1, 126.3, 126.5, 126.7, 126.9, 127.5, 128.6, 128.9, 129.2, 129.2, 134.4, 136.9, 138.9, 166.7, and 198.7 (24C). ESI-MS: m/z, 388.19 (M+ + H). Anal. calcd for C24H25N3O2 (387.47): C, 74.39; H, 6.50; and N, 10.84. Found: C, 74.18; H, 6.42; and N, 10.76.(11) 1-(Butylamino)-3-(butylimino)-2-[(1-(naphthalen-1-yl)diazene]but-1-en-1-ol
(8a). Brownish crystalline solid, m.p. 91-92°C, yield = 71%, and 1H-NMR (DMSO-d6) δ (ppm). 0.93 (t, 3H, J = 6.4 Hz, NHCH2CH2CH2CH3), 0.96 (t, 3H, J = 7.3 Hz, NCH2CH2CH2CH3), 1.46–1.40 (m, 4H, J = 7.7 Hz, NHCH2CH2CH2CH3 and NCH2CH2CH2CH3), 1.61 (q, 2H, NHCH2CH2CH2CH3), 1.67–1.64 (m, 2H, NCH2CH2CH2CH3), 2.62 (s, 3H, C-CH3), 3.39 (q, 2H, J = 6.4 Hz, NHCH2CH2CH2CH3), 3.65 (q, 2H, J = 6.8 Hz, NCH2CH2CH2CH3), 7.56–7.52 (m, 4H, naphthyl-H), 7.76 (d, 1H, J = 8.2 Hz, naphthyl-H), 7.95–7.90 (m, 1H, naphthyl-H), 8.4 (d, 1H, J = 7.4 Hz, naphthyl-H), 11.3 (br s, 1H, NH), and 15.37 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm) 14.1, 14.2, 14.7, 20.0, 20.4, 31.4, 31.7, 37.9, 44.0, 111.3, 121.3, 122.8, 126.0, 126.3, 126.8, 126.8, 128.6, 129.0, 134.5, 148.7, 165.9, and 172.7 (22C). ESI-MS: m/z, 367.25 (M+ + H). Anal. calcd for C22H30N4O (366.50): C, 72.10; H, 8.25; and N, 15.29. Found: C, 72.00; H, 8.18; and N, 15.20.(12) N-(2-hydroxyethyl imino)-3-(2-hydroxyethylamino)-2-(naphthalen-1-yldiazenyl)but-2-en-2-ol
(8b). Brownish crystalline solid, m.p. 157-158°C, yield = 70%, and 1H-NMR (DMSO-d6) δ (ppm) 2.64 (s, 3H, C-CH3), 3.47 (q, 3H, J = 5.3 Hz, CONHCH2CH2OH), 3.62–3.66 (m, 6H, CONHCH2CH2OH and CNCH2CH2OH), 4.95 (t, 1H, J = 4.6 Hz, CONHCH2CH2OH), 5.08 (t, 1H, J = 4.4 Hz, CNCH2CH2OH), 7.54–7.50 (m, 4H, naphthyl-H), 7.74 (d, 1H, J = 7.8 Hz, naphthyl-H), 7.94–7.88 (m, 1H, naphthyl-H), 8.74–8.70 (m, 1H, naphthyl-H), 11.40 (br s, 1H, NH), and 15.37 (s, 1H, OH); 13C-NMR (DMSO-d6) δ (ppm).15.0, 41.1, 47.1, 60.0, 60.5, 111.1, 121.3, 123.9, 126.3, 126.4, 126.7, 126.8, 128.2, 129.2, 134.5, 148.8, 165.8, and 172.8 (18C). ESI-MS: m/z, 343.18 (M+ + H). Anal. calcd for C18H22N4O3 (342.39): C, 63.14; H, 6.48; and N, 16.36. Found: C, 63.00; H, 6.40; and N, 16.30.


## 2.2. Cytotoxicity Assay

Three cancer cell lines were used in the current study: MCF-7 (human breast adenocarcinoma), A549 (lung carcinoma), and HT-29 (human colorectal adenocarcinoma). Cancer cell lines were obtained from Sigma-Aldrich Chemical Company, USA, and were seeded in Dulbecco’s Modified Eagle Media (DMEM) supplemented with 10% heat-inactivated fetal bovine serum (FBS), 100 IU/mL penicillin, 100 mg/mL streptomycin, and 2 mM glutamine. The cells were maintained in a humidified atmosphere with 5% CO2 at 37°C.


### 2.2.1. Antiproliferative Activity

The antiproliferative activity of the tested compounds was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as mentioned before [28–30]. In brief, cells were seeded in 96-well plates 24 h before different treatments. Test compounds were dissolved in dimethyl sulfoxide (DMSO) and further diluted with the cultivation medium. All target compounds (1–23) were preliminary evaluated in vitro at the single concentration of 5 μM against different cancer cell lines. Doxorubicin was used in parallel as a positive control drug at the single concentration of 2 μM. Different tested samples were added to the wells, and the cells were incubated at 37°C in a 5% CO2-humidified incubator for 48 h. After treatment, 10 μL of the MTT solution (5 mg/mL) was added to each well, and the plates were further incubated for 2 h. The media along with the MTT solution was removed and replaced by 100 μL of the DMSO solution to dissolve the precipitating formazan crystals. The intensity of the developed color was measured at 570 nm with an automatic multiwell plate reader (Varioskan™ LUX multimode microplate reader, Thermo Fisher Scientific, USA). Percentage inhibition of proliferation was calculated as a fraction of the vehicle control.


## 2.2.1. Antiproliferative Activity

The antiproliferative activity of the tested compounds was measured using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay as mentioned before [28–30]. In brief, cells were seeded in 96-well plates 24 h before different treatments. Test compounds were dissolved in dimethyl sulfoxide (DMSO) and further diluted with the cultivation medium. All target compounds (1–23) were preliminary evaluated in vitro at the single concentration of 5 μM against different cancer cell lines. Doxorubicin was used in parallel as a positive control drug at the single concentration of 2 μM. Different tested samples were added to the wells, and the cells were incubated at 37°C in a 5% CO2-humidified incubator for 48 h. After treatment, 10 μL of the MTT solution (5 mg/mL) was added to each well, and the plates were further incubated for 2 h. The media along with the MTT solution was removed and replaced by 100 μL of the DMSO solution to dissolve the precipitating formazan crystals. The intensity of the developed color was measured at 570 nm with an automatic multiwell plate reader (Varioskan™ LUX multimode microplate reader, Thermo Fisher Scientific, USA). Percentage inhibition of proliferation was calculated as a fraction of the vehicle control.


## 3. Results and Discussion

### 3.1. Chemistry

In the present work, a series of methyl 2-hydroxy-5-4-oxo-2-(substituted amino)pent-2-en-3-yl)diazenyl)benzoate4a–k were synthesized via the diazotization of methyl 5-amino-2-hydroxybenzoate 2 as the starting material. Nitrosation of methyl 5-amino-2-hydroxybenzoate 1 by using NaNO2 in the presence of HCl afforded the corresponding diazonium salt derivative 2. Treatment of 2 with 4-(substituted amino)pent-3-en-2-one derivatives 3 [31] in absolute ethanol with stirring at room temperature gave the corresponding methyl 2-hydroxy-5-(4-oxo-2-(substituted amino)pent-2-en-3-yl)diazenyl)benzoate 4a-k, respectively (Scheme 1).Scheme 1
Synthetic route of compounds4a–k.Additionally, nitrosation of 1-aminonaphthalene by using NaNO2 in the presence of HCl afforded the corresponding diazonium salt of naphthalen-1-amine 5. Treatment of diazonium compound 5 with N-substituted 3-oxo-butanamides (A) or ethyl 3-(substituted imino)butanoates (B) gave the corresponding azocompounds 6a–h and 7a,b, respectively. Finally, compound 5 was reacted with N-alkyl-3-(alkyl imino)butanamides (C) in the presence of sodium acetate to afford the corresponding compounds 8a,b, respectively (Scheme 2).Scheme 2
Synthetic route of compounds6–8.


### 3.2. Anticancer Activity

The results of the antiproliferative activity of the synthesized compounds (Table1, Figure 1) clearly showed that the synthesized compounds greatly varied in their effects on the tested cell lines. All the evaluated 23 compounds showed variable effects on MCF-7 cells. On the contrary, compounds 4f, 6f, 6g, 8a, and 8b were not effective against A549 and HT-29 cells. For MCF-7 cells, the most effective compounds are 4c, 4a, and 8a, whereas their toxicities were moderate on cells and reached 35, 23, and 22%, respectively. Compounds 6f, 4i, 6b, 4k, 6a, 4j, 6d, and 7a showed lower anticancer activities, and their toxicities obtained at the tested dose recorded 20, 16, 15, 13, 12, 11, 11, and 11%, respectively. The rest of the synthesized compounds showed weak anticancer activities, where they maximally reduced cell viabilities by 10%. Concerning A549 cells, compounds 4a and 6b showed a maximal potent effect, and their percentages of inhibition of cancer cells were obtained as 34 and 25%, respectively. On the contrary, compounds 7a, 4i, 4j, and 4h showed lower anticancer activities, and their maximal cytotoxicities reached 17, 15, 13, and 11%, respectively. For HT-29 cells, it can be seen that the most potent compound (compound 7b) against HT-29 cells reduced cell growth by about 39%. On the contrary, compounds 4f, 4d, and 7a showed weaker cytotoxic activities ranging only from 13 to 11%. It can be generally concluded that the most potent synthesized compounds are 7b against HT-29 cells, compound 4a against A549 cells, and compound 4c against MCF-7 cells.Table 1
Inhibition percentages of different synthesized compounds against MCF-7, A549, and HT-29 cell line.
Compound
Inhibition percentage ± SD (%)
MCF-7
A549
HT-29
4a
23 ± 3
34 ± 4
0.0 ± 8
4b
4 ± 5
6 ± 3
0.0 ± 8
4c
35 ± 3
10 ± 8
0.0 ± 3
4d
10 ± 5
9 ± 8
12 ± 8
4e
10 ± 2
3 ± 9
0.0 ± 4
4f
7 ± 3
0.0 ± 8
0.0 ± 6
4g
13 ± 7
0.0 ± 6
13 ± 7
4h
10 ± 5
11 ± 5
8 ± 6
4i
16 ± 5
15 ± 7
13 ± 4
4j
11 ± 4
13 ± 6
9 ± 6
4k
13 ± 7
10 ± 5
6 ± 6
6a
12 ± 6
7 ± 4
0.0 ± 7
6b
15 ± 3
15 ± 4
6 ± 7
6c
8 ± 4
5 ± 5
0.0 ± 4
6d
11 ± 5
3 ± 6
9 ± 7
6e
6 ± 5
2 ± 8
0.0 ± 7
6f
20 ± 2
0.0 ± 7
0.0 ± 8
6g
1 ± 6
0.0 ± 8
0.0 ± 8
6h
10 ± 5
0.0 ± 6
2 ± 8
7a
11 ± 4
17 ± 2
11 ± 6
7b
10 ± 3
0.0 ± 6
39 ± 7
8a
22 ± 5
0.0 ± 6
0.0 ± 8
8b
2 ± 4
0.0 ± 3
0.0 ± 8
Data are represented as mean ± SD.Figure 1
Effect of different compounds on the viability of MCF-7, A549 and HT-29 cell line. Data are represented as mean ± SD. (a) MCF-7; (b) A549; (c) HT-29.The abovementioned results showed that different cell lines varied greatly in their response against different synthesized compounds. This correlates well with previously reported results [32, 33], where this can be attributed to the inherent differences between different cells in their specific membrane structure and organization. For the synthesized derivatives, our results are in good agreement with those previously reported by Jakopec et al. [34], who reported on the cytotoxic activities of diazenes on several tumor cell lines. They attributed their results to both (1) the attachment of unsubstituted benzene on the amide nitrogen and (2) the extent of compound basicity. Moreover, the length of the conjugation plays a crucial role in the activity of the synthesized derivatives [34].Generally, it can be observed that the obtained anticancer activities of the newly synthesized diazene candidates are not promising as expected; therefore, the future work is planned to investigate further structural modifications of the prepared derivatives.


## 3.1. Chemistry

In the present work, a series of methyl 2-hydroxy-5-4-oxo-2-(substituted amino)pent-2-en-3-yl)diazenyl)benzoate4a–k were synthesized via the diazotization of methyl 5-amino-2-hydroxybenzoate 2 as the starting material. Nitrosation of methyl 5-amino-2-hydroxybenzoate 1 by using NaNO2 in the presence of HCl afforded the corresponding diazonium salt derivative 2. Treatment of 2 with 4-(substituted amino)pent-3-en-2-one derivatives 3 [31] in absolute ethanol with stirring at room temperature gave the corresponding methyl 2-hydroxy-5-(4-oxo-2-(substituted amino)pent-2-en-3-yl)diazenyl)benzoate 4a-k, respectively (Scheme 1).Scheme 1
Synthetic route of compounds4a–k.Additionally, nitrosation of 1-aminonaphthalene by using NaNO2 in the presence of HCl afforded the corresponding diazonium salt of naphthalen-1-amine 5. Treatment of diazonium compound 5 with N-substituted 3-oxo-butanamides (A) or ethyl 3-(substituted imino)butanoates (B) gave the corresponding azocompounds 6a–h and 7a,b, respectively. Finally, compound 5 was reacted with N-alkyl-3-(alkyl imino)butanamides (C) in the presence of sodium acetate to afford the corresponding compounds 8a,b, respectively (Scheme 2).Scheme 2
Synthetic route of compounds6–8.


## 3.2. Anticancer Activity

The results of the antiproliferative activity of the synthesized compounds (Table1, Figure 1) clearly showed that the synthesized compounds greatly varied in their effects on the tested cell lines. All the evaluated 23 compounds showed variable effects on MCF-7 cells. On the contrary, compounds 4f, 6f, 6g, 8a, and 8b were not effective against A549 and HT-29 cells. For MCF-7 cells, the most effective compounds are 4c, 4a, and 8a, whereas their toxicities were moderate on cells and reached 35, 23, and 22%, respectively. Compounds 6f, 4i, 6b, 4k, 6a, 4j, 6d, and 7a showed lower anticancer activities, and their toxicities obtained at the tested dose recorded 20, 16, 15, 13, 12, 11, 11, and 11%, respectively. The rest of the synthesized compounds showed weak anticancer activities, where they maximally reduced cell viabilities by 10%. Concerning A549 cells, compounds 4a and 6b showed a maximal potent effect, and their percentages of inhibition of cancer cells were obtained as 34 and 25%, respectively. On the contrary, compounds 7a, 4i, 4j, and 4h showed lower anticancer activities, and their maximal cytotoxicities reached 17, 15, 13, and 11%, respectively. For HT-29 cells, it can be seen that the most potent compound (compound 7b) against HT-29 cells reduced cell growth by about 39%. On the contrary, compounds 4f, 4d, and 7a showed weaker cytotoxic activities ranging only from 13 to 11%. It can be generally concluded that the most potent synthesized compounds are 7b against HT-29 cells, compound 4a against A549 cells, and compound 4c against MCF-7 cells.Table 1
Inhibition percentages of different synthesized compounds against MCF-7, A549, and HT-29 cell line.
Compound
Inhibition percentage ± SD (%)
MCF-7
A549
HT-29
4a
23 ± 3
34 ± 4
0.0 ± 8
4b
4 ± 5
6 ± 3
0.0 ± 8
4c
35 ± 3
10 ± 8
0.0 ± 3
4d
10 ± 5
9 ± 8
12 ± 8
4e
10 ± 2
3 ± 9
0.0 ± 4
4f
7 ± 3
0.0 ± 8
0.0 ± 6
4g
13 ± 7
0.0 ± 6
13 ± 7
4h
10 ± 5
11 ± 5
8 ± 6
4i
16 ± 5
15 ± 7
13 ± 4
4j
11 ± 4
13 ± 6
9 ± 6
4k
13 ± 7
10 ± 5
6 ± 6
6a
12 ± 6
7 ± 4
0.0 ± 7
6b
15 ± 3
15 ± 4
6 ± 7
6c
8 ± 4
5 ± 5
0.0 ± 4
6d
11 ± 5
3 ± 6
9 ± 7
6e
6 ± 5
2 ± 8
0.0 ± 7
6f
20 ± 2
0.0 ± 7
0.0 ± 8
6g
1 ± 6
0.0 ± 8
0.0 ± 8
6h
10 ± 5
0.0 ± 6
2 ± 8
7a
11 ± 4
17 ± 2
11 ± 6
7b
10 ± 3
0.0 ± 6
39 ± 7
8a
22 ± 5
0.0 ± 6
0.0 ± 8
8b
2 ± 4
0.0 ± 3
0.0 ± 8
Data are represented as mean ± SD.Figure 1
Effect of different compounds on the viability of MCF-7, A549 and HT-29 cell line. Data are represented as mean ± SD. (a) MCF-7; (b) A549; (c) HT-29.The abovementioned results showed that different cell lines varied greatly in their response against different synthesized compounds. This correlates well with previously reported results [32, 33], where this can be attributed to the inherent differences between different cells in their specific membrane structure and organization. For the synthesized derivatives, our results are in good agreement with those previously reported by Jakopec et al. [34], who reported on the cytotoxic activities of diazenes on several tumor cell lines. They attributed their results to both (1) the attachment of unsubstituted benzene on the amide nitrogen and (2) the extent of compound basicity. Moreover, the length of the conjugation plays a crucial role in the activity of the synthesized derivatives [34].Generally, it can be observed that the obtained anticancer activities of the newly synthesized diazene candidates are not promising as expected; therefore, the future work is planned to investigate further structural modifications of the prepared derivatives.



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*Source: 3626824-2018-11-01.xml*