INTRODUCTION

Heterocycles constitute a wide class of organic compounds which contribute significantly in every facet of pure and applied chemistry [1]. Heterocyclic compounds have played an important role in bio­chemical industries as they are present in large propor­tion in biomolecules like vitamins, enzymes, biologi­cally active compounds, and natural products [2]. Indole derivatives have attracted considerable attention [3] due to the broad range of their biological properties such as antiviral [4], antibacterial [5], anticancer [6], antidepressant [7], and antifungal activities [8].

Derivatives of indole have a variety of medicinal uses as antihypertensive and antiparasitic agents, antidepressants, as well as in cardiology, neurology, and endocrinology [9]. One of the indole alkaloids is delavirdine which prevents human immunodeficiency virus (HIV) from proliferating in human body [10]. Arbidol is an antiviral agent which is used for the treatment of influenza viruses, and it also acts as an ef­ficient inhibitor of SARS-CoV-2 virus [11]. Rhopaladin is a marine indole alkaloid which showed repres­sive activity against c-ErbB-2 kinase and cyclin-depen­dent kinase 4 [12] (Fig. 1). Indole derivatives such as tadalafil, chaetoindolone A, eudistomin C, strepto­chlorin analogs [13], etc., have also been evaluated for the effective control of plant pathogens. Plant patho­gens cause damage to flora, fauna, and microorganisms [14]. Nowadays, the major threat to agriculture is the root-knot nematode of the Meloidogyne genus [15] out of which Meloidogyne incognita species majorly cause yield loss in different crops like tomato, brinjal, turmeric, melon, etc. [16] which damage crops worth $125 billion with a yield loss of 14% all over the world [17]. In India, annual loss in 30 crops caused by nematodes is in crores each year. Therefore, efforts have been made for the prevention of infectious nematodes.

Fig. 1.
figure 1

Structures of some indole-containing drugs.

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Various reagents have been used for the synthesis of indole dimers. The reagents commonly used for the substitution reaction of indole with aromatic aldehydes are mineral acids like HCl, H2SO4, and HF, but these are listed as hazardous catalysts [18]. Herein, we report a convenient one-step reaction for the synthesis of bis-indolyl methanes using glacial acetic acid under ultra­sonication conditions as it has emerged as a new lead in green organic synthesis [19]. Also, the use of glacial acetic acid offers many advantages such as cost effec­tive­ness, easy availability, lower toxicity, air stability, and easy separation of products by simple filtration, thereby eradicating the necessity of purification protocols such as chromatography and liquid–liquid extractions which are very time-consuming [20]. As far as nematicidal activity [21] is concerned, only limited information is available in this regard. To bridge this gap, bis(indolyl)methanes have been synthesized from indole and aromatic aldehydes and examined for their nematicidal activity against the root knot nematode M. incognita [22].

RESULTS AND DISCUSSION

Bis(indolyl)methane derivatives 3a–3g were syn­the­sized as shown in Scheme 1 by the reaction of indole (1) with aromatic aldehydes 2a–2g in water in the presence of acetic acid as catalyst at 40°C under ultrasonic irradiation. The crude products were recrys­tallized from ethanol to afford pure compounds 3a–3g in good yields. The structure of the synthesized com­pounds was confirmed by using various spectroscopic techniques, including 1H and 13C NMR, FT-IR spec­troscopy, and mass spectrometry. Compounds 3a–3g are colored crystalline solids that are stable in air and readily soluble in dimethyl sulfoxide, methylene chloride, chloroform, and other polar solvents.

Scheme
scheme 1

1.

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The proposed mechanism of indole dimerization with aromatic aldehydes involves electrophilic substi­tu­tion at the 3-position of indole by the aldehyde carbonyl carbon atom in acidic environment to give intermediate A which is converted to another interme­diate B via elimination of water molecule. The addition of the second indole molecule to B produces protonated bis(indolyl)methane C whose deprotonation affords final product 3 (Scheme 2).

Scheme
scheme 2

2.

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Indole (1) and its derivatives 3a–3g were evaluated for their nematicidal activity against Meloidogyne incognita at different concentrations (1500, 1000, 750, 500, 250, and 100 ppm) after exposure for 24, 48, 72, 96, and 120 h. The efficiency of compounds 1 and 3a–3g on the egg hatch inhibition of M. incognita at different concentrations and durations are shown in Figs. 2 and 3 (see also Supplementary Materials). The highest percent egg hatch inhibition was exhibited by the compounds at the maximum concentration (1500 ppm), and their efficiency declined as the con­cen­tration decreased (Fig. 2). Similarly, the maximum egg hatch inhibition was observed after 120 h of dura­tion, followed by 96, 72, 48, and 24 h (Fig. 3).

Fig. 2.
figure 2

Effect of indole (1) and its derivatives 3a–3g on percent egg hatch inhibition of M. incognita at different concentrations.

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Fig. 3.
figure 3

Effect of indole (1) and its derivatives 3a–3g on percent egg hatch inhibition of M. incognita at different durations.

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Figures 4 and 5 illustrate the effect of indole (1) and its derivatives 3a–3g on the percent mortality of second stage juveniles (J2) of M. incognita at different con­centrations and durations. The observed pattern was similar to that of percent egg hatch inhibition, i.e., the percent mortality decreased with decrease in the con­centration and exposure time (Supplementary Table 2).

Fig. 4.
figure 4

Effect of indole (1) and its derivatives 3a–3g on percent mortality of second stage juveniles of M. incognita at different concentrations.

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Fig. 5.
figure 5

Effect of indole (1) and its derivatives 3a–3g on percent mortality of second stage juveniles of M. incognita at different durations.

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The maximum percent mortality against second stage juvenile of M. incognita was seen after 120 h duration, followed by 96, 72, 48, and 24 h of exposure (Fig. 5). Thus, all the compounds showed both concen­tration and duration dependent manner for both percent egg hatch inhibition and percent mortality of root knot nematodes. 3,3′-[(4-Nitrophenyl)methyl]di(1H-indole) (3a) exhibited the highest percent egg hatch inhibition potential (96.83%) and maximum percent mortality potential (100.00%). For percent egg hatch inhibition, the order of efficiency was as follows: 3a > 3b > 3c > 3d > 3e > 3f > 1 > 3g which showed that 3a was the most effective and that 3g was the least effective. The same order was observed for percent mortality. Thus, the compounds having electron-withdrawing groups showed better nematicidal activity [23, 24].

EXPERIMENTAL

The melting points were measured in open-end capil­laries with a Nutronics digital melting point apparatus. The reactions were carried out using a Helix Biosciences Ultra Sonicator (220 V, 700 W) from the Central Instrumentation Laboratory (Punjab Agricul­tural University, Ludhiana). The 1H and 13C NMR spectra were recorded at 25°C on a Bruker Avance II spectrometer (500 and 125 MHz, respectively) using CDCl3 or DMSO-d6 as solvent and tetramethylsilane (TMS) as internal standard. The IR spectra (400– 4000 cm–1) were recorded on a Perkin Elmer Spectrum Two FT-IR spectrometer from samples prepared as KBr pellets. Elemental analyses were obtained on a Thermo Scientific instrument (Department of Chemistry, Guru Nanak Dev University, Amritsar).

General procedure for the synthesis of 3,3′-(aryl­methylene)di(1H-indoles) 3a–3g. A solution of indole (1, 0.138 mol, 0.162 g) in acetonitrile (5.0 mL) was added dropwise at room temperature to a solution of aromatic aldehyde 2a–2e (0.039 mol) in acetonitrile (5.0 mL). Water (20.0 mL) and a catalytic amount of acetic acid (0.5 mol %) were then added, and the mixture was irradiated at 40°C in an ultrasonicator. After completion of the reaction (2–8 h; TLC), the mixture was poured onto crushed ice, and the solid product was filtered off and recrystallized from hot methylene chloride.

3,3′-[(4-Nitrophenyl)methylene]di(1H-indole) (3a). Yield 78%, yellow crystalline solid, mp 220–222°C (from CH2Cl2), Rf 0.52 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1219 (C–N), 1344 (NO2), 1515 (NO2), 1554 (C=Carom), 2858 (C–H), 3023 (C–Harom), 3397 (N–H). 1H NMR spectrum (CDCl3), δ, ppm: 5.92 s (1H, CH), 6.62 d (2H, J = 1.7 Hz, Harom), 6.94–6.98 m (2H, Harom), 7.11–7.14 m (2H, Harom), 7.28 d (2H, J = 7.95 Hz, Harom), 7.31 d (2H, J = 8.15 Hz, Harom), 7.36–7.39 q (1H, J = 7.9 Hz, Harom), 7.62 d (1H, J = 7.65 Hz, Harom), 7.94 s (2H, NH, D2O exchange­able), 8.01 d.d (1H, J = 1.3, 1.45 Hz, Harom), 8.14 t (1H, J = 1.7 Hz, Harom). 13C NMR spectrum (CDCl3), δC, ppm: 111.26, 118.23, 119.55, 121.51, 122.34, 123.63, 123.67, 126.65, 129.13, 134.87, 136.75, 146.38, 148.51. Mass spectrum: m/z 368.09 [M + 1]+. Found, %: C 75.19; H 4.66; N 11.44; O 8.71. C23H17N3O2. Calculated, %: C 74.09; H 4.26; N 11.34; O 8.21. M 367.13.

3,3′-[(3-Nitrophenyl)methylene]di(1H-indole) (3b). Yield 82%, yellow crystalline solid, mp 218–220°C (from CH2Cl2), Rf 0.40 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1232 (C–N), 1338 (NO2), 1522 (NO2), 1551 (C=Carom), 2856 (C–H), 3019 (C–Harom), 3356 (N–H). 1H NMR spectrum (DMSO-d6), δ, ppm: 6.03 s (1H, Harom), 6.87–6.90 m (4H, Harom), 7.04– 7.07 m (2H, Harom), 7.29 d (2H, J = 7.95 Hz, Harom), 7.37 d (2H, J = 8.1 Hz, Harom), 7.61 d (2H, J = 8.7 Hz, Harom), 8.15 d.d (2H, J = 1.85, 1.32 Hz, Harom), 10.92 d (2H, J = 1.4 Hz, NH, D2O exchangeable). 13C NMR spec­trum (DMSO-d6), δC, ppm: 111.46, 116.55, 118.29, 118.79, 120.97, 123.30, 123.73, 126.24, 129.33, 136.47, 145.64, 153.02. Mass spectrum: m/z 368.23 [M + 1]+. Found, %: C 75.19; H 4.66; N 11.44; O 8.71. C23H17N3O2. Calculated, %: C 75.02; H 4.45; N 11.21; O 8.11. M 367.13.

3,3′-[(4-Fluorophenyl)methylene]di(1H-indole) (3c). Yield 75%, light brown crystalline solid, mp 78–80°C (from CH2Cl2), Rf 0.34 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1224 (C–N), 1332 (C–F), 1521 (C=Carom), 2860 (C–H), 3013 (C–Harom), 3448 (N–H). 1H NMR spectrum (CDCl3), δ, ppm: 5.93 s (1H, CH), 6.61 d (2H, J = 2.2 Hz, Harom), 6.93–6.98 m (2H, Harom), 7.11–7.15 m (2H, Harom), 7.27 d (2H, J = 5.90 Hz, Harom), 7.32 d (2H, J = 12.3 Hz, Harom), 7.37 q (1H, J = 12.9 Hz, Harom), 7.63 d (1H, J = 7.60 Hz, Harom), 7.94 s (2H, NH, D2O exchangeable), 8.02 d.d (1H, J = 3.8, 0.54 Hz, Harom), 8.15 t (1H, J = 3.15 Hz, Harom). 13C NMR spectrum (CDCl3), δC, ppm: 112.26, 118.35, 119.75, 119.91, 121.13, 122.34, 123.75, 124.93, 126.15, 129.56, 134.67, 136.65, 146.68, 148.15. Mass spectrum: m/z 341.23 [M + 1]+. Found, %: C 81.16; H 5.03; F 5.58; N 8.23. C23H17FN2. Calculated, %: C 80.86; H 4.91; F 5.28; N 8.13. M 340.39.

3,3′-[(3,4-Dimethoxyphenyl)methylene]di(1H-indole) (3d). Yield 90%, light pink crystalline solid, mp 198–200°C (from CH2Cl2), Rf 0.53 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1084 (C–O), 1198 (C–N) 1539 (C=Carom), 2831 (C–H), 3008 (C–Harom), 3240 (N–H). 1H NMR spectrum (CDCl3), δ, ppm: 3.74 s (3H, OMe), 3.83 s (3H, OMe), 5.82 s (1H, CH), 6.63 t (2H, J = 1.4 Hz, Harom), 6.75 d (1H, J = 8.25 Hz, Harom), 6.82 d.d (1H, J = 3.40 Hz, Harom), 6.92 d (1H, J = 1.95 Hz, Harom), 6.98–7.01 m (2H, Harom), 7.14–7.17 m (2H, Harom), 7.33 d (2H, J = 8.2 Hz, Harom), 7.39 d (2H, J = 7.95 Hz, Harom), 7.88 s (2H, NH, D2O exchangeable). 13C NMR spectrum (CDCl3), δC, ppm: 55.82, 55.85, 110.94, 111.04, 112.27, 119.23, 119.93, 119.97, 120.61, 121.93, 123.55, 127.10, 136.73, 136.75, 147.33, 148.71. Mass spectrum: m/z 383.29 [M + 1]+. Found, %: C 78.51; H 5.80; N 7.32; O 8.37. C25H22N2O2. Calculated, %: C 78.13; H 5.49; N 7.22; O 8.24. calculated: M 382.45.

3,3′-[(4-Methoxyphenyl)methylene]di(1H-indole) (3e). Yield 94%, light red crystalline solid, mp 180–182°C (from CH2Cl2), Rf 0.50 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1029 (C–O), 1147 (C–N), 1488 (C=Carom), 2689 (C–H), 3051 (C–Harom), 3197 (N–H). 1H NMR spectrum (CDCl3), δ, ppm: 3.83 s (3H, OMe), 5.82 s (1H, CH), 6.62 t (2H, J = 2.85 Hz, Harom), 6.75 d (1H, J = 8.30 Hz, Harom), 6.82 d.d (1H, J = 1.95, 0.76 Hz, Harom), 6.93 d (1H, J = 7.05 Hz, Harom), 6.93–7.04 m (2H, Harom), 7.14–7.17 m (2H, Harom), 7.32 d (2H, J = 9.95 Hz, Harom), 7.38 d (2H, J = 4.20 Hz, Harom), 7.88 s (2H, NH, D2O exchangeable). 13C NMR spectrum (CDCl3), δC, ppm: 55.84, 110.47, 111.49, 113.67, 119.22, 119.39, 119.91, 121.61, 122.13, 124.75, 127.90, 137.63, 137.65, 147.13, 148.53. Mass spec­trum: m/z 353.41 [M + 1]+. Found, %: C 81.79; H 5.72; N 7.95; O 4.54. C24H20N2O. Calculated, %: C 81.87; H 5.65; N 7.45; O 4.41. M 352.43.

4-[Di(1H-indol-3-yl)methyl]-2-methoxyphenol (3f). Yield 88%, grey crystalline solid, mp 111–113°C (from CH2Cl2), Rf 0.43 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1134 (C–O), 1214 (C–N), 1545 (C=Carom), 2899 (C–H), 3112 (C–Harom), 3342 (N–H), 3365 (O–H). 1H NMR spectrum (CDCl3), δ, ppm: 3.86 s (3H, OMe), 5.52 s (1H, CH), 5.79 s (1H, OH, D2O exchangeable), 6.68 d (2H, J = 1.50 Hz, Harom), 6.76 d (1H, J = 8.25 Hz, Harom), 6.84 d.d (1H, J = 2.05, 0.71 Hz, Harom), 6.92 d (1H, J = 2.05 Hz, Harom), 6.70 t (2H, J = 7.20 Hz, Harom), 7.14–7.17 m (2H, Harom), 7.34 d (2H, J = 8.15 Hz, Harom), 7.40 d (2H, J = 8.15 Hz, Harom), 7.90 s (2H, NH, D2O exchangeable). 13C NMR spectrum (CDCl3), δC, ppm: 55.93, 110.37, 110.96, 115.07, 119.18, 119.91, 119.96, 120.11, 121.87, 123.46, 127.10, 136.71, 144.93, 145.30. Mass spec­trum: m/z 369.19 [M + 1]+. Found, %: C 78.24; H 5.47; N 7.60; O 8.69. C24H20N2O2. Calculated, %: C 78.14; H 5.24; N 7.57; O 8.45. M 368.43.

4-[Di(1H-indol-3-yl)methyl]phenol (3f). Yield 78%, pink crystalline solid, mp 118–120°C (from CH2Cl2), Rf 0.47 (EtOAc–hexane, 20:80). IR spectrum, ν, cm–1: 1221 (C–N), 1322 (C–O), 1524 (C=Carom), 2864 (C–H), 3054 (C–Harom), 3247 (N–H), 3373 (O–H). 1H NMR spectrum (DMSO-d6), δ, ppm: 5.71 s (1H, CH), 6.66 d.d (2H, J = 8.55, 5.9 Hz, Harom), 6.78 d (2H, J = 1.80 Hz, Harom), 6.84–6.87 m (2H, Harom), 7.01–7.04 m (2H, Harom), 7.14 d (2H, J = 8.45 Hz, Harom), 7.26 d (2H, J = 8.11 Hz, Harom), 7.33 d (2H, J = 8.10 Hz, Harom), 9.11 s (1H, OH, D2O exchangeable), 10.75 d (2H, J = 1.65 Hz, NH, D2O exchangeable). 13C NMR spectrum (DMSO-d6), δC, ppm: 111.25, 114.62, 117.92, 118.54, 119.04, 120.63, 123.24, 126.54, 128.99, 135.07, 136.46, 155.13. Mass spec­trum: m/z 339.29 [M + 1]+. Found, %: C 81.63; H 5.36; N 8.28; O 4.73. C23H18N2O. Calculated, %: C 81.49; H 5.26; N 8.18; O 4.65. M 338.40.

Nematicidal activity. a. Egg hatch inhibition assay. A pure culture of root knot nematodes was raised on the crop of brinjal. Five egg masses were taken and placed in a solution of a tested compound in water (5 mL) with a concentration of 1500, 1000, 750, 500, 250, and 100 ppm. A small quantity of acetone was added to dissolve the compounds in distilled water for preparing stock solutions which were then diluted to a required concentration. Distilled water with the same amount of acetone was used as control in each treat­ment, and three replications of each treatment were made. Egg hatching was observed after 24, 48, 72, 96, and 120 h, maintaining the temperature at 27±2°C. Statistical analysis was performed, and critical differ­ences were calculated. The percent hatch inhibition was calculated as (C – T)/C×100, where C is the number of nematodes in the control sample, and T is the number of nematodes after treatment.

b. Second stage juvenile mortality assay. Freshly hatched second stage juveniles (J2) were taken for the mortality test. An average of 20 stage two juveniles were placed in 1 mL of distilled water, and 5 mL of a solution of each test compound prepared as in the hatching test was added with 4 replications each together with control. For each concentration, the results were recorded after 24, 48, 72, 96, and 120 h of exposure, and the percent ratio of the number of dead nematodes to the total number of nematodes was determined.

CONCLUSIONS

Arylbis(indolyl)methanes were synthesized by a one-step procedure from aromatic aldehydes and indole under ultrasonic irradiation. The proposed proce­dure is relatively easy, less time consuming, and greener than those reported previously. 3,3′-[(4-Nitro­phenyl)methylene]di(1H-indole) (3a) showed the best nematicidal activity against the root knot nematode M. incognita according to the egg hatch inhibition and second stage juvenile mortality assays. Arylbis­(indolyl)­methanes having electron-withdrawing groups were more effective.