Water-Mediated Synthesis of (E)-3-(1-Methyl-1H-benzo[d]imidazol-5-yl)-N-phenethylacrylamide, a Caffeic Acid Phenethyl Amide Analogue

Abstract

Caffeic acid phenethyl ester (CAPE) is a phenolic natural product with diverse biological activities, notably anticancer properties. However, its ester group is metabolically unstable. The amide derivative, CAPA, offers improved metabolic stability to esterases but still possesses a metabolically liable catechol group. In this work, we describe the synthesis of a novel CAPA analogue in which the catechol is replaced with a benzimidazole bioisostere via a water-mediated Wittig reaction.

1. Introduction

Caffeic acid phenethyl ester (CAPE) (Figure 1) is a phenolic compound derived from bee propolis; it exhibits a wide range of biological activities such as antimicrobial, anti-inflammatory, antioxidant, and immunostimulatory effects [1,2,3,4,5]. This has led to CAPE’s application in chronic wound healing [6], protection from ischemia–reperfusion injury [7], and other applications [8]. However, CAPE’s metabolic instability has prompted efforts to develop more stable analogues by replacing CAPE’s metabolically unstable ester bond with an amide group yielding caffeic acid phenethyl amide (CAPA) (Figure 1). This substitution of an ester with an amide should lead to increased stability for carboxylesterases, which generally hydrolyze amides much slower than esters [9,10]. While this modification addresses the instability of CAPE towards esterases, both CAPE and CAPA contain catechol groups that can also serve as sites of metabolic instability [11] as well as toxophores due to redox cycling and covalent modification of the ortho-quinone form [12,13].

To address these limitations, we sought to design a CAPA analogue with a bioisostere for the catechol group. Benzimidazoles have been reported to serve as catechol bioisosteres in the context of catecholamines [14,15] and other biologically active phenols [16]. Thus, we report the preparation of one such benzimidazole CAPA analogue, (E)-3-(1-methyl-1H-benzo[d]imidazol-5-yl)-N-phenethyl acrylamide (1) (Figure 1).

Usually, CAPE and CAPA analogues are constructed using ester or amide coupling of caffeic acid precursors or the Wittig olefination reaction of the corresponding aldehyde to generate CAPE and CAPA [17,18,19]. For acid coupling approaches using organic solvents, high-purity reagents and specific activating agents are often required to achieve desired yields and selectivity. In contrast, we elected to employ the Wittig olefination route by employing an on-water reaction of the appropriate phosphonium salt with 1-methyl-1H-benzo[d]imidazole-5-carbaldehyde, following our recently reported procedure for the preparation of biologically active CAPA analogs [20]. Water has proven to be an effective medium for Wittig reactions involving a broad range of stabilized ylides and aldehydes. This type of Wittig reaction performs well when large hydrophobic groups, such as aromatic and heterocyclic aromatic carboxaldehydes, are paired with phosphonium salts. Although the reactants may sometimes exhibit poor solubility, the reactions typically result in high yields, with E-selectivity reaching up to 99% [21,22,23]. In this method, the aqueous Wittig reaction stabilized ylides are relatively straightforward, catalyst-free, and sustainable, achieve high E-selectivity, and minimize the environmental impact by reducing reliance on organic solvents.

2. Results and Discussion

The required aldehyde for the Wittig route to 1, 1-methyl-1H-benzo[d]imidazole-5-carbaldehyde 7 (Scheme 1), has previously been prepared via the reduction of 1-methyl-1H-benzo[d]imidazole-5-carbonitrile [24]; however, the yield for this reduction was not high (51% yield). Aldehyde 7 has also been prepared as a mixture of 5- and 6-carbaldehydes via the methylation of 1H-benzo[d]imidazole-5-carbaldehyde [16]. We elected to prepare 7 through Swern oxidation of the primary alcohol 6 which was prepared from the corresponding methyl ester 5 (Scheme 1).

The benzimidazole methyl ester 5 was synthesized using a modified literature procedure [25]. Starting with 4-fluoro-3-nitrobenzoic acid, Fisher esterification in methanol affords the methyl ester 2 in 95% yield (Scheme 1). The conversion of fluoro compound 2 to methylamine 3 was achieved in quantitative yield using 40% methylamine solution. Reduction of 3 with zinc metal affords methyl 3-amino-4-(methylamino) benzoate (4) in 91% yield. Subjecting 4 to microwave irradiation in the presence of formic acid at 150 °C for 15 min gives ester 5 (77% yield). LAH reduction of ester 5 affords the hydroxymethyl compound 6 in 81% yield. Swern oxidation of 6 affords the required aldehyde 7 in 85% yield.

The synthesis of benzimidazole CAPA analogue 1 proceeded through the Wittig reaction of 1-methyl-1H-benzo[d]imidazole-5-carbaldehyde (7) and phosphonium bromide (8) in water using sodium hydroxide as a base at 70 °C for 3 h (Scheme 2). After extraction with dichloromethane, purification via flash chromatography, and recrystallization from DCM in hexane, the CAPA analogue was obtained as a single isomer of (E)-3-(1-methyl-1H-benzo[d]imidazol-5-yl)-N-phenethyl acrylamide (1) in 64% yield (Scheme 2).

3. Materials and Methods

Unless otherwise noted, all materials were obtained from commercial suppliers and were used without further purification. (2-Oxo-2-(phenethyl amino) ethyl) triphenyl phosphonium bromide (8) was prepared using a method from the literature [26]. Flash chromatography was performed on a Teledyne Combi-Flash with RediSep Rf silica gel (230–400 mesh) using the mobile phase indicated. Melting points (open capillary) were determined on an Electrothermal series IA 9000 digital melting point apparatus and were uncorrected. Unless otherwise noted, ¹H and ¹³C NMR spectra were obtained on Bruker Advance Spectrometers operating at 400 MHz (100 MHz for ¹³C) or 500 MHz (126 MHz for ¹³C) and were determined in CDCl₃ and DMSO-d6. Chemical shifts are reported in ppm using solvent as an internal standard (7.26 ppm for ¹H and 77.0 ppm for ¹³C in CDCl₃ and 2.50 ppm for ¹H and 39.52 ppm for ¹³C in DMSO-d6). Low-resolution mass spectra were obtained on an Advion Expression L compact mass spectrometer with atmospheric pressure chemical ionization (APCI) or electrospray ionization (ESI). High-resolution mass spectra were obtained on a Waters Synapt XS spectrometer with ESI. Copies of all NMR, IR, and HRMS spectra are available in the Supplementary Materials.

Synthesis of methyl 4-fluoro-3-nitrobenzoate (2): To a solution of 4-fluoro-3-nitrobenzoic acid (1 g, 5.45 mmol) in methanol (10 mL), concentrated H₂SO₄ (0.5 mL) was added. The reaction mixture was heated to 60 °C for 8 h. After completion of the reaction (monitored using the TLC), the solution was concentrated under reduced pressure, diluted with water, and extracted with ethyl acetate (2 × 50 mL). The combined organic layers were washed with sodium bicarbonate solution (20 mL), dried over anhydrous sodium sulfate, and concentrated under vacuum to afford an off-white solid, 1.1 g, 95% yield; mp = 62–64 °C; IR 2952, 2885, 1726, 1268, 1072, 835 cm⁻¹; ¹H NMR (500 MHz, CDCl₃) δ 8.74 (dd, J = 7.2, 2.1 Hz, 1H), 8.33 (ddd, J = 8.7, 4.2, 2.2 Hz, 1H), 7.40 (dd, J = 10.1, 8.8 Hz, 1H), 3.99 (s, 3H); ¹³C NMR (126 MHz, CDCl₃) δ 164.1, 158.0 (d, J = 271 Hz), 136.5 (d, J = 10 Hz), 127.8, 127.8, 127.2 (d, J = 4 Hz), 118.8 (d, J = 21 Hz), 52.9; APCI MS m/z: 200.1 (MH⁺). Matches lit [27].

Synthesis of methyl 4-(methylamino)-3-nitrobenzoate (3): To a solution of 4-fluoro-3-nitro-benzoic acid (1 g, 5.02 mmol) in MeOH (10 mL) at 0 °C, 40% methylamine solution in water (5 mL) was added dropwise with vigorous stirring. The reaction mixture was stirred at ambient temperature for 4 h, during which time an orange precipitate formed. The reaction mixture was diluted with water (20 mL) and cooled to 5 °C in an ice bath. The reaction mixture was adjusted to pH 4 through the slow addition of conc. HCl. The precipitate that formed was collected by filtration and washed with 2 × 10 mL of distilled water. Drying in vacuo for 18 h gave 4-methylamino-3-nitrobenzoic acid as a yellow solid, 1.15 g, quantitative yield, mp = 138–140 °C; IR 2950, 1705, 1230, 1005, 972, 835 cm⁻¹; ¹H NMR (500 MHz, DMSO-d₆) δ 8.56 (d, J = 1.7 Hz, 2H), 7.94 (d, J = 9.1 Hz, 1H), 7.02 (d, J = 9.1 Hz, 1H), 3.81 (s, 3H), 2.99 (d, J = 5.0 Hz, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 165.4, 148.5, 136.1, 130.8, 128.7, 115.9, 115.0, 52.4, 30.3; APCI MS m/z: 211.1 (MH⁺). Matches lit [28].

Synthesis of methyl 3-amino-4-(methylamino) benzoate (4): To a solution of methyl 3-(methylamino)-4-nitrobenzoate (900 mg, 4.28 mmol) in THF/water/MeOH (1:1:1) (10 mL) at 0 °C, zinc dust (1.4 g, 21.41 mmol) and ammonium chloride (1.14 g, 21.41 mmol) were added. The reaction mixture was stirred at rt for 30 min. The reaction mixture was filtered on a bed of Celite and extracted with ethyl acetate (50 mL); the combined organic layers were dried over anhydrous sodium sulfate and concentrated under vacuum to afford a light brown solid, 700 mg, 91% yield; mp = 95–97 °C; IR 3403, 2948, 2525, 1692, 1599, 1296, 1166, 766 cm⁻¹; ¹H NMR (500 MHz, DMSO-d₆) δ 7.25 (dd, J = 8.2, 1.9 Hz, 1H), 7.18 (d, J = 1.9 Hz, 1H), 6.40 (d, J = 8.3 Hz, 1H), 5.39 (d, J = 3.4 Hz, 1H), 4.66 (s, 2H), 3.73 (s, 3H), 2.78 (d, J = 3.9 Hz, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 167.4, 142.1, 134.5, 121.3, 117.0, 114.3, 107.9, 51.6, 30.1; APCI MS m/z: 181.2 (MH⁺). Matches lit [29].

Synthesis of methyl 1-methyl-1H-benzo[d]imidazole-5-carboxylate (5): A solution of methyl 3-amino-4-(methyl amino) benzoate (500 mg, 2.77 mmol) in 3 mL of formic acid in a microwave vial was subjected to microwave heating to 150 °C for 15 min. The reaction mixture was concentrated under reduced pressure, diluted with water, and extracted with ethyl acetate (2 × 20 mL). The combined organic layers were washed with saturated sodium bicarbonate solution (20 mL) followed by brine solution (20 mL), dried over anhydrous sodium sulfate, and concentrated under vacuum. The crude product was triturated with a DCM/hexane mixture to obtain an orange solid, 400 mg, 77% yield; mp = 124–126 °C; IR 2984, 2908, 1735, 1478, 1283, 1234, 1043 cm⁻¹; ¹H NMR (400 MHz, DMSO-d₆) δ 8.34 (s, 1H), 8.26 (d, J = 1.1 Hz, 1H), 7.91 (d, J = 8.5 Hz, 1H), 7.68 (d, J = 8.5 Hz, 1H), 3.88 (s, 3H), 3.87 (s, 3H); ¹³C NMR (101 MHz, DMSO-d₆) δ 167.3, 147.3, 143.2, 138.3, 123.8, 123.6, 121.5, 110.9, 52.5, 31.4; APCI MS m/z: 191.1 (MH⁺). Matches lit [25].

Synthesis of (1-methyl-1H-benzo[d]imidazol-5-yl) methanol (6): To a solution of methyl 1-methyl-1H-benzo[d]imidazole-5-carboxylate (300 mg, 1.74 mmol) in 15 mL of THF under argon at 0 °C, LAH (38 mg, 2.55 mmol) was added. The reaction mixture was stirred at rt for 6 h. After complete consumption of the starting material (monitored using the TLC), the reaction mixture was carefully quenched with water (20 mL) and extracted with ethyl acetate (3 × 50 mL). The combined organic layers were dried over anhydrous sodium sulfate and concentrated under vacuum. The crude product was purified using Combi–Flash chromatography using 12% methanol in DCM as an eluant to obtain the product as a white solid, 163 mg, 83% yield; mp = 120–122 °C; IR 3379, 3256, 2949, 1758, 1246, 1043 cm⁻¹; ¹H NMR (500 MHz, DMSO-d₆) δ 8.13 (s, 1H), 7.58 (d, J = 0.5 Hz, 1H), 7.50 (d, J = 8.3 Hz, 1H), 7.24 (dd, J = 8.3, 0.9 Hz, 1H), 5.22 (s, 1H), 4.60 (d, J = 5.7 Hz, 2H), 3.82 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 145.1, 143.7, 136.4, 134.0, 122.1, 117.6, 110.1, 63.8, 31.1; APCI MS m/z: 163.4 (MH⁺); HR CIMS calcd. for C₉H₁₁N₂O (MH⁺) 163.0866, found 163.0883.

Synthesis of 1-methyl-1H-benzo[d]imidazole-5-carbaldehyde (7): To a solution of oxalyl chloride 165 mg in DCM (15 mL) at −78 °C under argon, DMSO (202 mg in DCM 10 mL) was added dropwise, followed by stirring at −78 °C for 30 min. Alcohol (150 mg in DCM 10 mL) was added dropwise to the reaction mixture and stirred for 30 min at −78 °C. Triethylamine (0.72 mL) was added to the reaction mixture and stirred at 0 °C for 30 min. The reaction mixture was allowed to warm to rt and stirred for an additional 1 h. The reaction mixture was quenched with water (50 mL) and extracted with DCM (2 × 25 mL), and the combined organic layers were dried over sodium sulfate and concentrated under vacuum to afford the crude product. The crude product was dissolved in 1 mL of DCM; then, 25 mL of hexane was added and the mixture was stirred for 20 min during which the product precipitated as a solid. The solids were filtered and dried under vacuum to afford a brown solid, 125 mg, 85% yield; mp = 119–121 °C; IR 3510, 3383, 1732, 1687, 1372, 1249 cm⁻¹; ¹H NMR (500 MHz, DMSO-d₆) δ 10.06 (s, 1H), 8.39 (s, 1H), 8.25 (s, 1H), 7.85 (dd, J = 8.4, 1.3 Hz, 1H), 7.76 (d, J = 8.4 Hz, 1H), 3.90 (s, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 193.2, 147.7, 143.4, 139.3, 131.5, 123.6, 123.1, 111.6, 31.5; APCI MS m/z: 161.1 (MH⁺); HR CIMS calcd. for C₉H₉N₂O (MH⁺) 161.0709, found 161.0720. Matches lit [16,24].

Synthesis of (E)-3-(1-methyl-1H-benzo[d]imidazol-5-yl)-N-phenethyl acrylamide (1): To a stirred solution of phosphonium salt 8 (113.3 mg, 0.22 mmol) in 2 mL of water at 0 °C, sodium hydroxide (30.0 mg, 0.75 mmol) was added portion-wise followed by the addition of 1-methyl-1H-benzo[d]imidazole-5-carbaldehyde 7 (30 mg, 0.18 mmol). The resulting mixture was stirred at 70 °C for 3 h. After the completion of the reaction (monitored using the TLC), the reaction mixture was diluted with water (10 mL) and extracted with dichloromethane (2 × 25 mL). The combined organic layers were washed with 20 mL of brine solution, dried over sodium sulfate, and concentrated under reduced pressure to afford the crude product. The crude product was purified via flash chromatography using (10% MeOH in DCM) is an eluant to afford the obtained compound, which was recrystallized from DCM (1 mL) in hexane (10 mL) to afford an off-white solid, 35 mg, 64% yield; mp = 151–153 °C; IR 3292, 3062, 2362, 1652, 1614, 1249 cm⁻¹; ¹H NMR (500 MHz, DMSO-d₆) δ 8.22 (s, 1H), 8.16 (d, J = 5.1 Hz, 1H), 7.84 (s, 1H), 7.60 (d, J = 8.3 Hz, 1H), 7.53 (t, J = 13.1 Hz, 2H), 7.36–7.18 (m, 5H), 6.62 (d, J = 15.7 Hz, 1H), 3.85 (s, 3H), 3.54–3.50 (m, 2H), 2.80 (t, J = 7.3 Hz, 3H); ¹³C NMR (126 MHz, DMSO-d₆) δ 165.8, 146.2, 144.0, 140.2, 139.9, 136.0, 129.2, 129.1, 128.8, 126.6, 122.3, 120.6, 119.5, 111.2, 40.8, 35.6, 31.3; HR CIMS calcd. for C₁₉H₂₀N₃O (MH⁺) 306.1601, found 306.1622.

4. Conclusions

In this study, we report a stereoselective on-water Wittig reaction of 1-methyl-1H-benzo[d]imidazole-5-carbaldehyde (7) with phosphonium salt (8) in the presence of a base to afford the novel benzimidazole CAPA analogue 1 in good yield. Biological studies of 1 are ongoing.