Linear and Angular Heteroannulated Pyridines Tethered 6-Hydroxy-4,7-Dimethoxybenzofuran: Synthesis and Antimicrobial Activity

Abstract

2-Chloropyridine-3-carbonitrile derivative 1 was utilized as a key precursor to build a series of linear and angular annulated pyridines linked to a 6-hydroxy-4,7-dimethoxybenzofuran moiety. Reaction of substrate 1 with various hydrazines afforded pyrazolo[3,4-b]pyridines. Treatment of substrate 1 with 1,3-N,N-binucleophiles including 3-amino-1,2,4-triazole, 5-amino-1H-tetrazole, 3-amino-6-methyl-1,2,4-triazin-5(4H)-one and 2-aminobenzimidazole produced the novel angular pyrido[3,2-e][1,2,4]triazolo[4,3-a]pyrimidine, pyrido[3,2-e][1,2,4]tetrazolo[1,5-a]pyrimidine, pyrido[3′,2′:5,6] pyrimido[2,1-c][1,2,4]triazine and benzo[4,5]imidazo[1,2-a]pyrido[3,2-e]pyrimidine, respectively. Reaction of substrate 1 with 1,3-C,N-binucleophiles including cyanoacetamides and 1H-benzimidazol-2-ylacetonitrile furnished 1,8-naphthyridines and benzoimidazonaphthyridine. Moreover, reacting substrate 1 with 5-aminopyrazoles gave pyrazolo[3,4-b][1,8]naphthyridines. Finally, reaction of compound 1 with 6-aminouracils as cyclic enamines yielded pyrimido[4,5-b][1,8]naphthyridines. Some of the synthesized products showed noteworthy antimicrobial efficiency against all types of microbial strains. Structures of the produced compounds were established using analytical and spectroscopic tools.

1. Introduction

Benzofuran scaffolds, found in numerous natural products and medications, are of great therapeutic value [1,2,3]. Some drugs containing benzofurans with potential biological activities are approved by the USFDA or EMA [4]. Benzofurans constitute a valuable class in the field of drug discovery and development due to their interesting biological characteristics [5,6]. Noteworthy, substituted benzofurans exhibit significant efficiency against various tumors and cancer cell lines [7,8,9]. Also, benzofurans tethered different heterocyclic compounds displayed significant antimicrobial efficiency against a diversity of microbial strains [9,10,11,12]. Benzofuran derivatives also possess diverse biological properties, such as antioxidant, anti-inflammatory, antipyretic neuroprotective, analgesic as well treatment potential for Alzheimer’s diseases [13,14,15,16,17]. Theoretical studies and physical applications on benzofuran derivatives including HOMO-LUMO energy, MEP map, Mulliken atomic charges, dipole moments, solvatochromic, NBO and NLO, photophysical, photochemical and optoelectronic properties were also investigated [18,19,20,21,22]. A variety of synthetic strategies were developed to prepare heterocyclic compounds including benzofuran skeletons [23,24,25,26]. However, 6-substituted khellin represents an excellent building block to synthesize benzofuran-tethered heterocyclic systems due to the availability of electron-deficient γ-pyrone moiety [27,28,29,30,31]. On the other hand, a diversity of pyridines and their annulated heterocycles are widely synthesized using variable synthetic methodology [32,33,34,35]. Pyridine-based heterocycles exhibited promising pharmacological characteristics including antiviral, antiproliferative anticancer, antimicrobial, antimycobacterial, antifungal, anti-diabetic and anti-Alzheimer as well as inhibitors for acetylcholinesterase and butyrylcholinesterase [36,37,38,39,40,41,42]. Given the chemical and biological importance of benzopyrans and pyridines scaffolds, the current work aims to synthesize some new linear and angular annulated pyridines tethered to a 6-hydroxy-4,7-dimethoxybenzofuran moiety in one molecular frame utilizing o-choropyridinecarbonitrile 1 [43] as a building block, and to explore the biological activities of the prepared compounds.

2. Results and Discussion

2.1. Characterization of the Synthesized Compounds

It is known that compounds containing neighboring cyano and chloro functions are active building blocks for constructing nitrogen heterocyclic compounds [44,45]. Thus, chloropyridinecarbonitrile derivative 1 serves as an effective precursor for the synthesis of a variety of fused pyridines connected to a 6-hydroxy-4,7-dimethoxybenzofuranylcarbonyl moiety.

Reaction of substrate 1 with 3-hydrazino-5,6-diphenyl-1,2,4-triazine (2) [46] and 7-chloro-4-hydrazinoquinoline (3) [47], in refluxing DMF/TEA, afforded triazinyl/ quinolinylpyrazolo[3,4-b]pyridines 4 and 5, respectively (Scheme 1). Compounds 4 and 5 are formed via the nucleophilic addition of NH₂ group to the nitrile function in substrate 1, followed by pyrazole ring closure and elimination of an HCl molecule. The mass spectra of compounds 4 and 5 confirmed their molecular formulae C₃₂H₂₃N₇O₅ and C₂₆H₁₈ClN₅O₅ showing their parent ion peaks at m/z 585 and 515, respectively. The C≡N function, detected at ṽ 2227 cm⁻¹ in the spectrum of compound 1, disappeared in the IR spectra of products 4 and 5. The IR spectrum of products 4 and 5 presented distinctive absorption bands due to amino groups at ṽ 3368, 3293 and 3354, 3271 cm⁻¹, respectively. Also, characteristic absorption bands corresponding to C=O and C=N were recorded at ṽ 1652/1658 and 1610/1612 cm⁻¹. Further, the NH₂ groups were detected in the ¹H-NMR spectra of compounds 4 and 5 at δ 9.31 and 9.42 ppm, respectively, while the OH protons were seen at δ 12.29 and 12.52 ppm. In addition, two characteristic doublets attributable to H-3_furan and H-2_furan were seen in the ¹H NMR spectra of compounds 4 and 5 at δ 7.08/7.13 and 7.85/7.86 ppm, respectively.

Likewise, compound 1 was permitted to react with some 1,3-N,N-binucleophiles. Thus, the novel angular pyrido[3,2-e][1,2,4]triazolo[4,3-a]pyrimidine 6 and pyrido[3,2-e][1,2,4]tetrazolo[1,5-a]pyrimidine 7 were synthesized from reacting substrate 1 with 3-amino-1,2,4-triazole and 5-amino-1H-tetrazole, respectively (Scheme 2). The mass spectra of the compounds 6 and 7 displayed their molecular ion peaks at m/z 406 and 407, coinciding with the proposed molecular formulae C₁₉H₁₄N₆O₅ and C₁₈H₁₃N₇O₅, respectively. Their IR spectra showed distinctive absorption bands at ṽ 3348, 3265/3369, 3287 (NH₂) and 1649/1644 cm⁻¹ (C=O). The ¹H NMR spectra of compounds 6 and 7 revealed characteristic singlet signals due to H-4_pyridine and H-2_pyridine at δ 8.43/8.52 and 8.55/8.61, respectively. In addition, the amino protons were observed as D₂O exchangeable signals at δ 9.50 and 9.26 ppm. The spectrum of compound 6 displayed definite singlet signal assignable to H-3_triazole at δ 8.97. The carbonyl carbon in compounds 6 and 7 were observed in the downfield region in the ¹³C NMR spectra at δ 192.3 and 192.4 ppm, respectively, also the spectrum of compound 6 showed distinctive singlet due to C-3_triazole at δ 137.3 ppm.

Similarly, treatment of substrate 3 with 3-amino-6-methyl-1,2,4-triazin-5(4H)-one (8) [48] and 2-aminobenzimidazole, in boiling DMF/TEA, yielded the novel angular annulated pyrido[3′,2′:5,6]pyrimido[2,1-c][1,2,4]triazine 9 and benzo[4,5]imidazo[1,2-a] pyrido[3,2-e]pyrimidine 10, respectively (Scheme 3). The IR spectra of compounds 9 and 10 showed distinctive absorption bands at ṽ 3372,3296/3383,3268 (NH₂) and 1654/1648 cm⁻¹ (C=O). Also, the spectrum of compound 9 presented distinguish absorption band due to C=O_triazine at ṽ 1692 cm⁻¹. The ¹H NMR spectra of compounds 9 and 10 presented D₂O exchangeable signals due to amino protons at δ 9.32 and 9.52 ppm, respectively. The spectrum of compound 9 displayed an upfield signal at δ 2.18, corresponding to CH_{3 triazine}. The ¹³C NMR spectrum of compound 9 showed two specific signals attributed to CH_{3 triazine} and C=O_triazine at δ 17.3 and 166.2 ppm. The mass spectra of compounds 9 and 10 showed their molecular ion peaks at m/z 448 and 455, respectively, which coincided well with their proposed molecular formulae C₂₁H₁₆N₆O₆ and C₂₄H₁₇N₅O₅, respectively.

Next, compound 1 was permitted to react with some of 1,3-C,N-binucleophiles. Hence, reaction of compound 1 with cyanoacetamide, N-benzyl-2-cyanoacetamide and 1H-benzimidazol-2-ylacetonitrile, in boiling DMF/TEA, furnished 1,8-naphthyridine-3-carbonitriles 11, 12 and benzo[4,5]imidazo[1,2-a][1,8] naphthyridine-6-carbonitrile 13, respectively (Scheme 4). The IR spectra of compounds 11–13 showed characteristic absorption bands attributed to C≡N at ṽ 2224, 2221 and 2226 cm⁻¹, respectively. The spectra of compounds 11 and 12 showed characteristic absorption bands due to C=O_pyridine at 1681 and 1686 cm⁻¹. The ¹H NMR spectra of compounds 11–13 presented the NH₂ protons as exchangeable signals at δ 9.32, 9.28 and 9.41 ppm, respectively. The NH proton in compound 11 was seen at δ 11.04 ppm. Also, the CH₂ protons in compound 12 were recorded at δ 3.08 ppm. The ¹³C NMR spectra of compounds 11 and 12 showed characteristic signals attributed to C≡N, C=O_{naphthyridine} and C=O_ketone at δ 116.3/116.6, 169.5/169.1 and 193.2/194.1 ppm, respectively. The spectrum of compound 12 displayed the methylene carbon as definite signal at δ 29.4. The mass spectra of compounds 11–13 exhibited their parent ion peaks at m/z 406, 496 and 479 that agree well with the suggested molecular formulae C₂₀H₁₄N₄O₆ (406.35), C₂₇H₂₀N₄O₆ (496.47) and C₂₆H₁₇N₅O₅ (479.44), respectively.

Moreover, the reaction of substrate 1 with 5-amino-2,4-dihydro-3H-pyrazol-3-one and 5-amino-3-methyl-1H-pyrazole, in boiling DMF/TEA, gave linear annulated pyrazolo[3,4-b][1,8]naphthyridines 14 and 15, respectively (Scheme 5). The IR spectrum of compound 14 showed typical absorption bands at ṽ 3376, 3338, 3296 (NH₂, 2NH), 1667 (C=O_pyrazole) and 1646 cm⁻¹ (C=O). The ¹H NMR spectrum of compound 15 revealed characteristic singlet signals at δ 2.42, 8.52 and 8.64 ppm attributed to CH_{3 pyrazole}, H-4_pyridine and H-2_pyridine, in addition to three D₂O exchangeable signals at δ 9.32 (NH₂), 10.36 (NH) and 12.41 ppm (OH). The mass spectra of compounds 14 and 15 revealed their molecular ion peaks at m/z 421 and 319 that match well with the proposed molecular formulae C₂₀H₁₅N₅O₆ (421.36) and C₂₁H₁₇N₅O₅ (419.39), respectively. The carbon of C=O_pyrazole was seen in the ¹³C NMR spectrum of compound 14 in the downfield region at δ 165.5 ppm, while the spectrum of compound 15 presented distinctive signal due to CH_{3 pyrazole} at the upfield region δ 18.6 ppm (CH₃).

Finally, compound 1 was permitted to react with some cyclic enamines namely 6-aminouracil, 6-aminothiouracil and 6-amino-1,3-dimethyluracil, in DMF containing TEA, giving pyrimido[4,5-b][1,8]naphthyridines 16–18, respectively (Scheme 6). The mass spectra of compounds 16–18 presented their molecular ion peaks at m/z 449, 465 and 477 approving their suggested formula weights 449.37 (C₂₁H₁₅N₅O₇), 465.44 (C₂₁H₁₅N₅O₆S) and 477.43 (C₂₃H₁₉N₅O₇), respectively. The amino protons were observed in ¹H NMR spectra of compounds 16–18 at δ 9.58, 9.34 and 9.37 ppm, respectively. Two characteristic signals attributable to 2NCH₃ protons were seen in the ¹H NMR spectrum of compound 18 at δ 3.06 and 3.17 ppm. Further, the ¹³C NMR spectra of compounds 16 and 18 showed characteristic signals at δ 165.1/165.5 (C2 as C=O_pyrimidine) and 167.4/168.1 (C4 as C=O_pyrimidine), while The spectrum of compound 17 displayed specific signals due to C4 as C=O_pyrimidine and C2 as C=S_pyrimidine at δ 168.9 and 186.3 ppm, respectively. Also, the ¹³C NMR spectrum of compound 18 showed two characteristic signals at δ 28.9 and 30.0 corresponding to 2NCH₃ carbons.

2.2. Antimicrobial Estimation

The synthesized products were investigated for their antimicrobial assay, in vitro, against some Gram-positive bacteria (S. aureus and B. subtilis) and Gram-negative bacteria (S. typhimurium and E. coli), as well as yeast (C. albicans) and fungus (A. fumigatus).

To assess the antimicrobial efficacy of the synthesized products, the inhibitory zones, including the disc diameter (6 mm), were evaluated (Table 1) [49]. High inhibition action referred to zone diameter >2/3 zone diameter of control, while moderate activity means zone diameter ≤2/3 and >1/3 zone diameter of reference drug. Cycloheximide is the reference drug for fungus and yeast, while Chloramphencol for Gram-positive bacteria, and Cephalothin for Gram-negative bacteria.

According to the results in Table 1 (Chart 1 and Charts S1–S5), all examined compounds had a strong inhibitory impact on the tested strains of fungus and yeast; this may due to the presence of the 6-hydroxy-4,7-dimethoxy-1-benzofuran moiety which exists in all products. Meanwhile, the inhibitory effect against the microbial strains varies according to the effect of the synthesized heterocyclic rings. Compounds 4 and 5 presented high efficiency against both types of Gram-positive and Gram-negative bacteria and this may be attributed to the presence of triazinyl/quinolinyl-pyrazolopyridine moieties linked to the benzofuranylcarbonyl fragment. Also, building angular heterocyclic systems, namely pyridotetrazolopyrimidine 7 and pyridopyrimidotriazine 9, enhanced the inhibitory effects against all tested microorganisms. On the other hand, some linear heterocyclic systems such as 1,8-naphthyridines 11, 12 and pyrimidonaphthyridines 16–18 showed high inhibition actions towards all tested bacterial strains.

As illustrated above, due to the existence of the principal scaffold, 6-hydroxy-4,7-dimethoxy-1-benzofuran moiety, all of the examined products demonstrated valuable inhibitory effects towards yeast and fungus. Furthermore, the inhibitory action towards bacterial strains was improved by the inclusion of additional heterocyclic systems, such as pyrazolopyridine, pyridotetrazolopyrimidine, pyridopyrimidotriazine, 1,8-naphthyridine and pyrimidonaphthyridine. As a result, some of the produced compounds may have excellent antimicrobial properties.

3. Materials and Methods

3.1. General Information

General. Melting point determination was performed using a digital Stuart SMP3 device (Buchi, Flawil, Switzerland). The mass spectra were measured using Shimadzu (Tokyo, Japan) GC-2010 mass spectrometer (70 eV); in gas chromatography. The ¹H NMR (400 MHz) and ¹³C NMR (100 MHz) spectra were measured with the Mercury-400BB apparatus (vnmr1, Rheinstetten, Germany) using DMSO-d₆ as the solvent and TMS (δ) as the internal standard. Using KBr disks, an FTIR Nicolet (Green Bay, WI, USA) IS10 spectrophotometer (cm⁻¹) was used to record the infrared spectra. 2-Chloro-5-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]pyridine-3-carbonitrile (1) was prepared according to literature [43].

3.2. Biological Method

On medium potato dextrose agar (PDA), which comprised an infusion of 200 g potatoes, 6 g dextrose, and 15 g agar, the antimicrobial activity test was conducted. Filter paper disks of uniform size (6 mm in diameter, with three disks for each chemical) were carefully placed on an inoculated agar surface after being impregnated with an equivalent volume (10 µL) of dissolved compounds at concentrations of 500 and 1000 mg/mL in dimethylformamide (DMF). Following 36 h of incubation at 27 °C for bacteria and 48 h at 24 °C for fungi. The average diameter of the bacterial and fungal inhibitory zones surrounding the disks, measured in millimeters at concentrations of 500 and 1000 mg/mL, was recorded for each investigated compound [49].

3-Amino-1-(5,6-diphenyl-1,2,4-triazin-3-yl)-5-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-1H-pyrazolo[3,4-b]pyridne (4)

A mixture of compound 1 (0.72 g, 2 mmol) and 3-hydrazinyl-5,6-diphenyl-1,2,4-triazine (2) (0.58 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the pale-yellow crystals deposited were filtered and recrystallized from AcOH, mp > 300 °C, yield (0.88 g, 75%). IR (KBr, cm⁻¹): 3413 (OH), 3368, 3293 (NH₂), 1652 (C=O), 1610 (C=N), 1581 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.86 (s, 3H, OCH₃), 3.95 (s, 3H, OCH₃), 7.08 (d, 1H, J = 2.0 Hz, H-3_furan), 7.44–7.52 (m, 10H, Ar-H), 7.85 (d, 1H, J = 2.0 Hz, H-2_furan), 8.42 (s, 1H, H-4_pyridine), 8.48 (s, 1H, H-2_pyridine), 9.31 (s, 2H, NH₂ exchangeable with D₂O), 12.29 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (75 MHz, DMSO-d₆, δ): 58.7 (OMe), 59.8 (OMe), 102.3 (C3a′), 104.0 (C3′), 108.2 (C3a), 112.4 (C5′), 121.4 (Ar-C), 122.1 (Ar-C), 123.8 (C7′), 124.7 (Ar-C), 125.2 (Ar-C), 127.6 (Ar-C), 129.1 (C4′), 129.8 (Ar-C), 131.4 (Ar-C), 132.8 (C5), 135.7 (Ar-C), 138.2 (C-7a), 139.0 (C-5_triazine), 139.9 (C-6_triazine), 140.2 (C-3_triazine), 143.2 (C-3), 144.5 (C-4), 147.2 (C-2′), 148.1 (C-6), 149.1 (C6′), 151.6 (C7a′), 189.7 (C=O_ketone). Mass spectrum, m/z (I_r %): 585 (M⁺, 46), 555 (24), 452 (37), 353 (16), 324 (20), 220 (71), 178 (100), 159 (13), 133 (10), 117 (16), 93 (25), 77 (48), 64 (21). Anal. Calcd for C₃₂H₂₃N₇O₅ (585.57): C, 65.64; H, 3.96; N, 16.74%. Found: C, 65.37; H, 3.88; N, 16.64%.

3-Amino-1-(7-chloroquinolin-4-yl)-5-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-1H-pyrazolo[3,4-b]pyridne (5)

A mixture of compound 1 (0.72 g, 2 mmol) and 7-chloro-4-hydrazinylquinoline (3) (0.38 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the orange-yellow crystals so formed were filtered and recrystallized from AcOH, mp > 300 °C, yield (0.79 g, 78%). IR (KBr, cm⁻¹): 3408 (OH), 3354, 3271 (NH₂), 1658 (C=O), 1612 (C=N), 1588 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.87 (s, 3H, OCH₃), 3.97 (s, 3H, OCH₃), 7.13 (d, 1H, J = 2.0 Hz, H-3_furan), 7.52–7.56 (m, 3H, H-3_quinoline, H-5_quinoline and H-6_quinoline), 7.86 (d, 1H, J = 2.0 Hz, H-2_furan), 8.04 (s, 1H, H-8_quinoline), 8.18 (d, 1H, J = 7.6 Hz, H-2_quinoline), 8.53 (s, 1H, H-4_pyridine), 8.69 (s, 1H, H-2_pyridine), 9.42 (s, 2H, NH₂ exchangeable with D₂O), 12.52 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (75 MHz, DMSO-d₆, δ): 58.5 (OMe), 59.3 (OMe), 102.5 (C3a′), 106.4 (C3′), 109.3 (C3a), 112.5 (C5′), 122.0 (Ar-C), 122.6 (Ar-C), 123.3 (Ar-C), 123.7 (C7′), 124.6 (Ar-C), 125.1 (Ar-C), 126.4 (Ar-C), 127.8 (Ar-C), 128.6 (C4′), 134.5 (C8a_quinoline), 139.5 (C-7a), 140.5 (C4_quinoline), 143.4 (C2 _quinoline), 144.5 (C-4), 145.2 (C-3), 147.4 (C-2′), 148.2 (C-6), 150.5 (C6′), 152.6 (C7a′), 192.2 (C=O_ketone). Mass spectrum, m/z (I_r %): 515/517 (M⁺/M+2, 100/33), 354 (68), 312 (32), 221 (54), 162/164 (69/23), 134 (11), 117 (19), 94 (42), 77 (26), 64 (15). Anal. Calcd for C₂₆H₁₈ClN₅O₅ (515.90): C, 60.53; H, 3.52; N, 13.57%. Found: C, 60.41; H, 3.39; N, 13.52%.

5-Amino-3-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]pyrido[3,2-e] [1,2,4]triazolo[4,3-a]pyrimidine (6)

A mixture of compound 1 (0.72 g, 2 mmol) and 3-amino-1,2,4-triazole (0.16 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals deposited were filtered and recrystallized from iso-butanol, mp > 300 °C, yield (0.59 g, 72%). IR (KBr, cm⁻¹): 3406 (OH), 3348, 3265 (NH₂), 1649 (C=O), 1615 (C=N), 1594 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.84 (s, 3H, OCH₃), 3.92 (s, 3H, OCH₃), 7.11 (d, 1H, J = 2.0 Hz, H-3_furan), 7.92 (d, 1H, J = 2.0 Hz, H-2_furan), 8.43 (s, 1H, H-4_pyridine), 8.55 (s, 1H, H-2_pyridine), 8.97 (s, 1H, H-3_triazole), 9.50 (s, 2H, NH₂ exchangeable with D₂O), 12.33 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 58.0 (OCH₃), 59.2 (OCH₃), 102.3 (C5′), 104.3 (C4a), 106.7 (C3′), 109.3 (C3a′), 122.4 (C7′), 128.2 (C3), 129.6 (C4′), 137.3 (C9), 138.4 (C4), 139.3 (C2), 144.9 (C5), 145.2 (C2′), 148.5 (C10a), 149.7 (C6a), 151.6 (C6′), 152.8 (C7a′), 192.3 (C=O_ketone). Mass spectrum, m/z (I_r %): 406 (M⁺, 100), 350 (67), 320 (39), 288 (32), 221 (51), 185 (24), 148 (28), 134 (15), 118 (11), 94 (47), 77 (33), 64 (10). Anal. Calcd for C₁₉H₁₄N₆O₅ (406.35): C, 56.16; H, 3.47; N, 20.68%. Found: C, 55.96; H, 3.44; N, 20.39%.

5-Amino-7-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]pyrido[3,2-e] [1,2,4]tetrazolo[1,5-a]pyrimidine (7)

A mixture of compound 1 (0.72 g, 2 mmol) and 5-amino-1H-tetrazole (0.16 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals so formed were filtered and recrystallized from iso-butanol, mp > 300 °C, yield (0.57 g, 70%). IR (KBr, cm⁻¹): 3403 (OH), 3369, 3287 (NH₂), 1644 (C=O), 1611 (C=N), 1590 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.86 (s, 3H, OCH₃), 4.03 (s, 3H, OCH₃), 7.19 (d, 1H, J = 2.0 Hz, H-3_furan), 7.95 (d, 1H, J = 2.0 Hz, H-2_furan), 8.52 (s, 1H, H-4_pyridine), 8.61 (s, 1H, H-2_pyridine), 9.26 (s, 2H, NH₂ exchangeable with D₂O), 12.40 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 58.5 (OCH₃), 59.1 (OCH₃), 101.8 (C5′), 105.1 (C4a), 106.2 (C3′), 110.3 (C3a′), 123.1 (C7′), 128.5 (C3), 129.8 (C4′), 138.5 (C4), 138.8 (C2), 144.2 (C5), 145.1 (C2′), 148.1 (C10a), 149.2 (C6a), 151.1 (C6′), 152.7 (C7a′), 192.4 (C=O_ketone). Mass spectrum, m/z (I_r %): 407 (M⁺, 48), 349 (30), 304 (25), 274 (19), 221 (36), 192 (16), 171 (14), 159 (22), 133 (18), 117 (14), 93 (100), 77 (64), 64 (24). Anal. Calcd for C₁₈H₁₃N₇O₅ (407.34): C, 53.07; H, 3.22; N, 24.07%. Found: C, 52.83; H, 3.14; N, 23.95%.

5-Amino-3-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-9-methyl-10H-pyrido[3′,2′:5,6]pyrimido[2,1-c][1,2,4]triazin-10-one (9)

A mixture of compound 1 (0.72 g, 2 mmol) and 3-amino-6-methyl-1,2,4-triazin-5(4H)-one (8) (0.25 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the pale-brown crystals so formed were filtered and recrystallized from AcOH/H₂O, mp > 300 °C, yield (0.62 g, 69%). IR (KBr, cm⁻¹): 3405 (OH), 3372, 3296 (NH₂), 1692 (C=O_triazine), 1654 (C=O), 1604 (C=N), 1587 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 2.18 (s, 3H, CH_{3 triazine}), 3.82 (s, 3H, OCH₃), 3.91 (s, 3H, OCH₃), 7.25 (d, 1H, J = 2.4 Hz, H-3_furan), 7.85 (d, 1H, J = 2.4 Hz, H-2_furan), 8.49 (s, 1H, H-4_pyridine), 8.58 (s, 1H, H-2_pyridine), 9.32 (s, 2H, NH₂ exchangeable with D₂O), 12.40 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 17.3 (CH₃), 58.8 (OCH₃), 59.3 (OCH₃), 103.4 (C5′), 104.9 (C4a), 106.6 (C3′), 110.2 (C3a′), 123.0 (C7′), 129.3 (C4′), 130.0 (C3), 135.4 (C9), 138.5 (C4), 139.1 (C2), 143.6 (C5), 145.0 (C11a), 146.2 (C2′), 148.5 (C6a), 150.5 (C6′), 152.7 (C7a′), 166.2 (C=O_triazine), 193.6 (C=O_ketone). Mass spectrum, m/z (I_r %): 448 (M⁺, 68), 418 (100), 388 (32), 346 (29), 227 (17), 194 (15), 159 (17), 133 (22), 118 (13), 92 (36), 77 (32), 64 (14). Anal. Calcd for C₂₁H₁₆N₆O₆ (448.39): C, 56.25; H, 3.60; N, 18.74%. Found: C, 56.03; H, 3.47; N, 18.65%.

5-Amino-3-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]benzo[4,5] imidazo[1,2-a]pyrido[3,2-e]pyrimidine (10)

A mixture of compound 1 (0.72 g, 2 mmol) and 2-aminobenzimidazole (0.27 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals deposited were filtered and recrystallized from AcOH, mp > 300 °C, yield (0.65 g, 71%). IR (KBr, cm⁻¹): 3417 (OH), 3383, 3268 (NH₂), 1648 (C=O), 1607 (C=N), 1582 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.84 (s, 3H, OCH₃), 3.98 (s, 3H, OCH₃), 7.15 (d, 1H, J = 2.4 Hz, H-3_furan), 7.37–7.43 (m, 2H, Ar-H), 7.48–7.53 (m, 2H, Ar-H), 7.86 (d, 1H, J = 2.4 Hz, H-2_furan), 8.42 (s, 1H, H-4_pyridine), 8.53 (s, 1H, H-2_pyridine), 9.52 (s, 2H, NH₂ exchangeable with D₂O), 12.33 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 58.5 (OCH₃), 59.7 (OCH₃), 102.1 (C5′), 104.8 (C4a), 106.2 (C3′), 110.5 (C3a′), 120.3 (Ar-C), 121.1 (Ar-C), 122.8 (C7′), 124.1 (Ar-C), 125.2 (Ar-C), 126.1 (Ar-C), 129.6 (C4′), 130.3 (C3), 134.3 (Ar-C), 138.3 (C4), 139.7 (C2), 142.9 (C-13a), 144.7 (C5), 146.3 (C2′), 148.1 (C6a), 150.3 (C6′), 151.9 (C7a′), 192.3 (C=O_ketone). Mass spectrum, m/z (I_r %): 455 (M⁺, 100), 395 (59), 340 (46), 234 (21), 220 (35), 190 (24), 161 (32), 134 (26), 117 (10), 94 (56), 77 (42), 65 (17). Anal. Calcd for C₂₄H₁₇N₅O₅ (455.42): C, 63.29; H, 3.76; N, 15.38%. Found: C, 63.14; H, 3.52; N, 15.31%.

4-Amino-6-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carbonitrile (11)

A mixture of compound 1 (0.72 g, 2 mmol) and cyanoacetamide (0.16 g, 2 mmol) in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals so formed were filtered and recrystallized from DMF/H₂O, mp > 300 °C, yield (0.59 g, 73%). IR (KBr, cm⁻¹): 3411 (OH), 3385, 3316, 3274 (NH₂, NH), 2224 (C≡N), 1681 (C=O_pyridine), 1650 (C=O), 1608 (C=N), 1584 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.89 (s, 3H, OCH₃), 3.97 (s, 3H, OCH₃), 7.23 (d, 1H, J = 2.4 Hz, H-3_furan), 7.91 (d, 1H, J = 2.4 Hz, H-2_furan), 8.46 (s, 1H, H-4_pyridine), 8.52 (s, 1H, H-2_pyridine), 9.32 (s, 2H, NH₂ exchangeable with D₂O), 11.04 (s, 1H, NH exchangeable with D₂O), 12.64 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 59.0 (OCH₃), 60.2 (OCH₃), 87.1 (C3), 102.7 (C5′), 105.8 (C3′), 111.2 (C3a′), 116.3 (C≡N), 122.8 (C7′), 123.2 (C4a), 128.4 (C6), 129.8 (C4′), 138.2 (C5), 140.0 (C7), 144.7 (C4), 146.1 (C2′), 148.8 (C8a), 150.8 (C6′), 152.3 (C7a′), 169.5 (C2 as C=O_{naphthyridine}), 193.2 (C=O_ketone). Mass spectrum, m/z (I_r %): 406 (M⁺, 100), 340 (25), 256 (20), 221 (49), 172 (38), 133 (14), 117 (23), 93 (46), 77 (28), 64 (13). Anal. Calcd for C₂₀H₁₄N₄O₆ (406.35): C, 59.12; H, 3.47; N, 13.79%. Found: C, 59.06; H, 3.30; N, 13.58%.

4-Amino-1-benzyl-6-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-2-oxo-1,2-dihydro-1,8-naphthyridine-3-carbonitrile (12)

A mixture of compound 1 (0.72 g, 2 mmol) and N-benzylcyanoacetamide (0.32 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals deposited were filtered and recrystallized from AcOH, mp > 300 °C, yield (0.68 g, 68%). IR (KBr, cm⁻¹): 3415 (OH), 3371, 3288 (NH₂), 2221 (C≡N), 1686 (C=O_pyridine), 1657 (C=O), 1613 (C=N), 1579 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.08 (s, 2H, CH₂), 3.83 (s, 3H, OCH₃), 3.97 (s, 3H, OCH₃), 7.18 (d, 1H, J = 2.0 Hz, H-3_furan), 7.54–7.66 (m, 5H, Ar-H), 7.93 (d, 1H, J = 2.4 Hz, H-2_furan), 8.42 (s, 1H, H-4_pyridine), 8.50 (s, 1H, H-2_pyridine), 9.28 (s, 2H, NH₂ exchangeable with D₂O), 12.32 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 29.4 (NCH₂), 58.3 (OCH₃), 59.5 (OCH₃), 88.4 (C3), 103.5 (C5′), 106.2 (C3′), 111.4 (C3a′), 116.6 (C≡N), 123.1 (C7′), 125.3 (C4a), 125.6 (Ar-C), 127.0 (Ar-C), 128.6 (C6), 129.2 (C4′), 130.6 (Ar-C), 134.1 (Ar-C), 138.6 (C5), 139.2 (C7), 144.5 (C4), 146.6 (C2′), 149.2 (C8a), 150.8 (C6′), 152.5 (C7a′), 169.1 (C2 as C=O_{naphthyridine}), 194.1 (C=O_ketone). Mass spectrum, m/z (I_r %): 496 (M⁺, 59), 466 (52), 375 (46), 309 (47), 242 (32), 220 (64), 194 (26), 159 (21), 134 (15), 118 (16), 91 (100), 77 (57), 64 (23). Anal. Calcd for C₂₇H₂₀N₄O₆ (496.47): C, 65.32; H, 4.06; N, 11.29%. Found: C, 65.14; H, 4.01; N, 11.15%.

5-Amino-3-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-benzo[4,5]imidazo[1,2-a][1,8]naphthyridine-6-carbonitrile (13)

A mixture of compound 1 (0.72 g, 2 mmol) and 1H-benzimidazol-2-ylacetonitrile (0.31 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the pale-brown crystals deposited were filtered and recrystallized from DMF, mp > 300 °C, yield (0.71 g, 74%). IR (KBr, cm⁻¹): 3404 (OH), 3361, 3279 (NH₂), 2226 (C≡N), 1651 (C=O), 1612 (C=N), 1576 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.86 (s, 3H, OCH₃), 3.93 (s, 3H, OCH₃), 7.13 (d, 1H, J = 2.0 Hz, H-3_furan), 7.37–7.42 (m, 4H, Ar-H), 7.88 (d, 1H, J = 2.0 Hz, H-2_furan), 8.39 (s, 1H, H-4_pyridine), 8.57 (s, 1H, H-2_pyridine), 9.41 (s, 2H, NH₂ exchangeable with D₂O), 12.33 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (75 MHz, DMSO-d₆, δ): 59.1 (OMe), 60.2 (OMe), 87.3 (C-6), 102.9 (C3a′), 105.8 (C3′), 108.6 (C-9), 112.5 (C5′), 113.1 (C-7a), 117.2 (C≡N), 120.7 (Ar-C), 122.8 (C7′), 124.1 (Ar-C), 124.7 (Ar-C), 128.9 (Ar-C), 129.8 (C4′), 130.7 (Ar-C), 132.2 (Ar-C), 138.7 (C-11a), 142.3 (C-8), 143.2 (C-10), 145.1 (C-7), 147.2 (C2′), 148.0 (C-5a), 151.9 (C6′), 152.4 (C7a′), 191.3 (C=O_ketone). Mass spectrum, m/z (I_r %): 479 (M⁺, 100), 419 (64), 353 (47), 313 (43), 258 (24), 221 (36), 192 (18), 161 (15), 133 (12), 117 (19), 94 (68), 77 (44), 64 (19). Anal. Calcd for C₂₆H₁₇N₅O₅ (479.44): C, 65.13; H, 3.57; N, 14.61%. Found: C, 64.96; H, 3.40; N, 14.39%.

4-Amino-6-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-1,2-dihydro-3H-pyrazolo[3,4-b][1,8]naphthyridin-3-one (14)

A mixture of compound 1 (0.72 g, 2 mmol) and 5-amino-2,4-dihydro-3H-pyrazol-3-one (0.20 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the pale-brown crystals deposited were filtered and recrystallized from DMF, mp > 300 °C, yield (0.64 g, 76%). IR (KBr, cm⁻¹): 3407 (OH), 3376, 3338, 3296 (NH₂, 2NH), 1667 (C=O_pyrazole), 1646 (C=O), 1605 (C=N), 1582 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.87 (s, 3H, OCH₃), 4.03 (s, 3H, OCH₃), 7.16 (d, 1H, J = 2.0 Hz, H-3_furan), 7.93 (d, 1H, J = 2.0 Hz, H-2_furan), 8.50 (s, 1H, H-4_pyridine), 8.69 (s, 1H, H-2_pyridine), 9.39 (s, 2H, NH₂ exchangeable with D₂O), 10.44 (s, 1H, NH exchangeable with D₂O), 11.28 (s, 1H, NH exchangeable with D₂O), 12.48 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 59.4 (OCH₃), 59.9 (OCH₃), 103.4 (C5′), 105.5 (C3a), 106.7 (C3′), 111.6 (C3a′), 112.4 (C5a), 122.6 (C7′), 128.3 (C6), 129.2 (C4′), 137.4 (C5), 138.1 (C7), 143.0 (C9a), 145.2 (C4), 146.2 (C2′), 148.4 (C8a), 150.7 (C6′), 152.5 (C7a′), 165.5 (C=O_pyrazolone), 192.0 (C=O_ketone). Mass spectrum, m/z (I_r %): 421 (M⁺, 100), 378 (47), 318 (56), 278 (27), 221 (38), 201 (41), 172 (30), 157 (14), 133 (17), 118 (21), 93 (48), 77 (36), 65 (12). Anal. Calcd for C₂₀H₁₅N₅O₆ (421.36): C, 57.01; H, 3.59; N, 16.62%. Found: C, 56.86; H, 3.47; N, 16.50%.

4-Amino-6-(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-3-methyl-1H-pyrazolo[3,4-b][1,8]naphthyridine (15)

A mixture of compound 1 (0.72 g, 2 mmol) and 5-amino-3-methyl-1H-pyrazole (0.20 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals so formed were filtered and recrystallized from DMF/H₂O, mp > 300 °C, yield (0.66 g, 79%). IR (KBr, cm⁻¹): 3412 (OH), 3359, 3281 (NH₂), 1648 (C=O), 1617 (C=N), 1589 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 2.42 (s, 3H, CH_{3 pyrazole}), 3.92 (s, 3H, OCH₃), 4.00 (s, 3H, OCH₃), 7.22 (d, 1H, J = 2.4 Hz, H-3_furan), 7.96 (d, 1H, J = 2.4 Hz, H-2_furan), 8.52 (s, 1H, H-4_pyridine), 8.64 (s, 1H, H-2_pyridine), 9.32 (s, 2H, NH₂ exchangeable with D₂O), 10.36 (s, 1H, NH exchangeable with D₂O), 12.41 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 18.6 (CH₃), 59.2 (OCH₃), 60.0 (OCH₃), 102.8 (C5′), 105.7 (C3a), 106.1 (C3′), 111.5 (C3a′), 113.4 (C5a), 122.4 (C7′), 125.4 (C6), 129.6 (C4′), 135.3 (C3), 137.2 (C6), 138.1 (C8), 142.6 (C9a), 143.0 (C4), 145.9 (C2′), 148.5 (C8a), 150.8 (C6′), 153.1 (C7a′), 191.6 (C=O_ketone). Mass spectrum, m/z (I_r %): 419 (M⁺, 68), 389 (100), 319 (42), 278 (37), 220 (51), 198 (24), 157 (20), 133 (26), 118 (27), 92 (39), 77 (28), 64 (17). Anal. Calcd for C₂₁H₁₇N₅O₅ (419.39): C, 60.14; H, 4.09; N, 16.70%. Found: C, 59.93; H, 3.84; N, 16.59%.

5-Amino-7-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl] pyrimido[4,5-b][1,8]naphthyridine-2,4(1H,3H)-dione (16)

A mixture of compound 1 (0.72 g, 2 mmol) and 6-amino-2,3-dihydro pyrimidin-2,4(1H,3H)-dione (0.27 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the yellow crystals deposited were filtered and recrystallized from AcOH/H₂O, mp > 300 °C, yield (0.67 g, 75%). IR (KBr, cm⁻¹): 3407 (OH), 3384, 3297, 3227 (NH₂, 2NH), 1679 (2C=O_pyrimidine), 1656 (C=O), 1614 (C=N), 1583 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.92 (s, 3H, OCH₃), 3.98 (s, 3H, OCH₃), 7.13 (d, 1H, J = 2.4 Hz, H-3_furan), 7.88 (d, 1H, J = 2.4 Hz, H-2_furan), 8.34 (s, 1H, H-4_pyridine), 8.72 (s, 1H, H-2_pyridine), 9.58 (s, 2H, NH₂ exchangeable with D₂O), 10.33 (s, 1H, NH exchangeable with D₂O), 10.70 (s, 1H, NH exchangeable with D₂O), 12.45 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 58.7 (OCH₃), 59.6 (OCH₃), 102.5 (C5′), 104.0 (C4a), 107.4 (C3′), 109.2 (C3a′), 111.2 (C5a), 122.9 (C7′), 127.6 (C3), 129.3 (C4′), 137.8 (C6), 139.1 (C8), 144.3 (C10a), 144.9 (C5), 147.4 (C2′), 148.1 (C9a), 150.5 (C6′), 152.7 (C7a′), 165.1 (C2 as C=O_pyrimidine), 167.4 (C4 as C=O_pyrimidine), 192.8 (C=O_ketone). Mass spectrum, m/z (I_r %): 449 (M⁺, 100), 391 (68), 346 (39), 305 (43), 262 (52), 229 (41), 185 (38), 157 (19), 134 (25), 117 (16), 94 (31), 77 (28), 64 (10). Anal. Calcd for C₂₁H₁₅N₅O₇ (449.37): C, 56.13; H, 3.36; N, 15.58%. Found: C, 56.02; H, 3.21; N, 15.37%.

5-Amino-7-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]-2-thioxo-2,3-dihydropyrimido[4,5-b][1,8]naphthyridin-4(1H)-one (17)

A mixture of compound 1 (0.72 g, 2 mmol) and 6-amino-2-thioxo-2,3-dihydropyrimidin-4(1H)-one (0.29 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL), was heated under reflux for 4 h. After cooling, the canary yellow crystals so formed were filtered and recrystallized from AcOH, mp > 300 °C, yield (0.73 g, 78%). IR (KBr, cm⁻¹): 3401 (OH), 3370, 3284, 3216 (NH₂, 2NH), 1673 (C=O_pyrimidine), 1652 (C=O), 1619 (C=N), 1587 (C=C), 1226 (C=S). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.90 (s, 3H, OCH₃), 3.98 (s, 3H, OCH₃), 7.08 (d, 1H, J = 2.8 Hz, H-3_furan), 7.87 (d, 1H, J = 2.8 Hz, H-2_furan), 8.53 (s, 1H, H-4_pyridine), 8.67 (s, 1H, H-2_pyridine), 9.34 (s, 2H, NH₂ exchangeable with D₂O), 11.42 (s, 1H, NH exchangeable with D₂O), 11.78 (s, 1H, NH exchangeable with D₂O), 12.64 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 58.5 (OCH₃), 59.7 (OCH₃), 103.4 (C5′), 105.3 (C4a), 107.2 (C3′), 109.5 (C3a′), 112.4 (C5a), 123.1 (C7′), 126.9 (C3), 128.7 (C4′), 136.4 (C6), 137.6 (C8), 142.1 (C10a), 143.2 (C5), 146.3 (C2′), 147.8 (C9a), 151.2 (C6′), 152.5 (C7a′), 168.9 (C4 as C=O_pyrimidine), 186.3 (C2 as C=S_pyrimidine), 194.8 (C=O_ketone). Mass spectrum, m/z (I_r %): 465 (M⁺, 78), 407 (58), 348 (61), 318 (55), 263 (47), 221 (56), 173 (16), 159 (26), 133 (29), 118 (32), 92 (100), 77 (46), 64 (13). Anal. Calcd for C₂₁H₁₅N₅O₆S (465.44): C, 54.19; H, 3.25; N, 15.05; S, 6.89%. Found: C, 53.85; H, 3.17; N, 14.93; S, 6.81%.

5-Amino-7-[(6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl]1,3-dimethyl-pyrimido[4,5-b][1,8]naphthyridine-2,4(1H,3H)-dione (18)

A mixture of compound 1 (0.72 g, 2 mmol) and 6-amino-1,3-dimethylpyrimidine- 2,4(1H,3H)-dione (0.31 g, 2 mmol), in DMF (10 mL) containing TEA (0.1 mL) was heated under reflux for 4 h. After cooling, the pale-yellow crystals so formed were filtered and recrystallized from AcOH/H₂O, mp > 300 °C, yield (0.74 g, 77%). IR (KBr, cm⁻¹): 3402 (OH), 3376, 3283 (NH₂), 1676 (2C=O_pyrimidine), 1658 (C=O), 1610 (C=N), 1588 (C=C). ¹H NMR (400 MHz, DMSO-d₆, δ): 3.06 (s, 3H, NCH₃), 3.17 (s, 3H, NCH₃), 3.92 (s, 3H, OCH₃), 4.04 (s, 3H, OCH₃), 7.25 (d, 1H, J = 2.0 Hz, H-3_furan), 7.97 (d, 1H, J = 2.0 Hz, H-2_furan), 8.52 (s, 1H, H-4_pyridine), 8.68 (s, 1H, H-2_pyridine), 9.37 (s, 2H, NH₂ exchangeable with D₂O), 12.52 (s, 1H, OH exchangeable with D₂O). ¹³C NMR (100 MHz, DMSO-d₆, δ): 28.9 (NCH₃), 30.0 (NCH₃), 58.9 (OCH₃), 59.7 (OCH₃), 102.6 (C5′), 103.9 (C4a), 106.7 (C3′), 109.8 (C3a′), 111.1 (C5a), 123.2 (C7′), 126.8 (C3), 129.0 (C4′), 137.1 (C6), 139.3 (C8), 143.6 (C10a), 144.7 (C5), 146.2 (C2′), 148.3 (C9a), 151.2 (C6′), 151.8 (C7a′), 165.5 (C2 as C=O_pyrimidine), 168.1 (C4 as C=O_pyrimidine), 192.7 (C=O_ketone). Mass spectrum, m/z (I_r %): 477 (M⁺, 100), 416 (72), 375 (64), 333 (47), 256 (58), 221 (65), 194 (37), 173 (46), 158 (25), 133 (29), 118 (21), 94 (22), 77 (18), 64 (9). Anal. Calcd for C₂₃H₁₉N₅O₇ (477.43): C, 57.86; H, 4.01; N, 14.67%. Found: C, 57.64; H, 3.95; N, 14.48%.

4. Conclusions

In the current study, the recently synthesized 2-chloro-5-[(6-hydroxy-4,7- dimethoxy-1-benzofuran-5-yl)carbonyl]pyridine-3-carbonitrile (1) was efficiently utilized as a building block for the construction of various heterocyclic systems. Linear and angular annulated pyridines linked to the (6-hydroxy-4,7-dimethoxy-1-benzofuran-5-yl)carbonyl were efficiently synthesized through the reaction of starting precursor 1 with binucleophilic reagents. All the synthesized compounds showed a remarkable effect against yeast and fungus strains, while compounds 4, 5, 7, 9, 11, 12 and 16–18 exhibited significant inhibitory effects against all tested microorganisms.

Author Contributions

N.A.A. Investigation, Methodology, Formal analysis, Writing—review and editing; A.-S.B. Investigation, Methodology, Formal analysis, Writing—review and editing. M.A.I. Investigation, Methodology, Formal analysis, Writing—review and editing. All authors have read and agreed to the published version of the manuscript.

Institutional Review Board Statement

Not applicable.

Informed Consent Statement

Not applicable.

Data Availability Statement

Data are contained within the article and Supplementary Materials.

Acknowledgments

The authors thank the support from Princess Nourah bint Abdulrahman University Researchers Supporting Project number (PNURSP2024R403), Princess Nourah bint Abdulrahman University, Riyadh, Saudi Arabia.

Conflicts of Interest

The authors declare no conflict of interest.

Footnotes

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Schemes, Chart and Table

Scheme 1. Formation of pyrazolo[3,4-b]pyridines 4 and 5.

Scheme 2. Formation of pyridotriazolopyrimidine 6 and pyridotetrazolopyrimidine 7.

Scheme 3. Formation of angular pyridopyrimidotriazine 9 and benzoimidazopyrido-pyrimidine 10.

Scheme 4. Formation of naphthyridines 11, 12 and benzoimidazonaphthyridine 13.

Scheme 5. Formation of pyrazolo[3,4-b][1,8]naphthyridines 14 and 15.

Scheme 6. Formation of pyrimido[4,5-b][1,8]naphthyridines 16–18.

Chart 1. The antibacterial efficiency of synthesized compounds against S. aureus.

Table 1

Antimicrobial estimation, in vitro, for the prepared compounds 1–18 by disc diffusion measurement.

No.	Zone Diameter (mm) * (% with Respect to Reference Drug)
	Gram-Positive Bacteria				Gram-Negative Bacteria				Yeasts and Fungi
Compd. No.	S. aureus		B. subtilis		S. typhimurium		E. coli		C. albicans		A. fumigatus
	1000μg/mL	500μg/mL	1000μg/mL	500μg/mL	1000μg/mL	500μg/mL	1000μg/mL	500μg/mL	1000μg/mL	500μg/mL	1000μg/mL	500μg/mL
1	40%	38%	49%	48%	53%	50%	45%	44%	69%	71%	70%	73%
4	77%	77%	69%	72%	69%	68%	71%	74%	86%	75%	73%	77%
5	83%	81%	77%	76%	75%	68%	74%	70%	80%	75%	70%	69%
6	46%	50%	51%	52%	47%	46%	39%	41%	71%	64%	77%	73%
7	80%	77%	74%	76%	75%	71%	74%	70%	86%	79%	77%	81%
9	86%	88%	83%	84%	75%	71%	68%	74%	83%	75%	73%	73%
10	57%	58%	49%	48%	42%	39%	45%	52%	71%	68%	73%	77%
11	80%	81%	71%	72%	78%	75%	68%	70%	77%	71%	68%	69%
12	71%	69%	69%	72%	72%	71%	74%	78%	74%	68%	78%	77%
13	49%	42%	46%	40%	53%	43%	42%	41%	69%	71%	70%	69%
14	54%	54%	43%	44%	58%	50%	47%	44%	74%	71%	77%	81%
15	51%	50%	49%	48%	50%	46%	39%	41%	77%	75%	68%	73%
16	77%	77%	74%	72%	78%	68%	66%	67%	74%	71%	77%	81%
17	89%	85%	71%	72%	72%	64%	71%	70%	77%	75%	68%	73%
18	83%	83%	80%	76%	75%	71%	68%	71%	71%	68%	77%	77%
S	100%	100%	100%	100%	100%	100%	100%	100%	100%	100%	100%	100%

* Calculated from 3 times. S: Standard antibiotics, which are Cycloheximide for fungus and yeast, Chloramphencol for Gram-positive bacteria, and Cephalothin for Gram-negative bacteria.

Supplementary Materials

The following supporting information can be downloaded at: https://www.mdpi.com/article/10.3390/molecules29184496/s1. Supporting Materials: (A) Copies of 1H-NMR, 13C-NMR and mass spectral data for the synthesized compounds and (B) Antimicrobial efficiency of the synthesized compounds with respected to references drugs.

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