Introduction

Isatin is an endogenous natural product found in a number of plants, including those of the genus Isatis [1] isolated in 1988 [2]. The reaction of isatin with various compounds have been under intensive studies by many authors [3–6]. Isatin derivatives have gained unique importance due to broad spectrum of pharmacological activities which are reflected by their use as antimicrobial agents. It has been reported that several compounds containing an isatin moiety possess antibacterial, antifungal, anticonvulsant, anti-inflammatory, and anti-HIV activities [7, 8]. Some of the heterocyclic compounds incorporated with isatin moiety such as pyrazoline, pyrimidine, and spiro derivatives shows significant application in different biological and industrial aspects [9, 10]. A number of indole derivatives are inhibitors of the Nor-A efflux pump in the human pathogenic bacterium Staphylococcus [11]. Different conventional methodologies have been used for the reaction of isatin with acetophenone. They have used hazardous chemicals like alkali KOH, piperidine, diethyleneamine, and basic alumina, solvents like DMF, acetic acid, and alcohols [12, 13]. Hence, to avoid such problems, it is necessary to go through clean, nonhazardous, and environmentally benign pathways for the synthesis.

Polyethylene glycol (PEG-400) prompted reactions have attracted the attention of organic chemists [14–16] due to their solvating ability and aptitude to act as a phase transfer catalyst, and their negligible vapour pressure, easy recyclability, ease of work-up, eco-friendly nature, and economical cost. PEG is non-toxic, non-halogenated, inexpensive, potentially recyclable and water soluble, which facilitates its removal from the reaction product.

In the present study, we have prepared 3-phenacylidene-2-indolinones by using bleaching earth clay (pH 12.5) as catalyst and PEG-400 as solvent.

Experimental

All the reagents and chemicals were used laboratory grade and purified prior to use. Melting points of all synthesized compounds were recorded in open capillary tubes and are uncorrected. IR spectra were recorded in KBr pellets on a FTIR Shimadzu spectrophotometer, and 1H NMR spectra in dimethylsulfoxide (DMSO) solvent were scanned on Aan VANCE 300 MHz spectrometer using TMS as an internal standard. The MS were recorded on a EI-Shimadzu-GC–MS spectrometer. Elemental analysis was carried out on a Carlo Ebra 106 Perkin-Elmer model 240 analyzer.

General procedure for the synthesis of 3-[2-(substituted-phenyl)-2-oxo-ethyl]-3-hydroxy-1,3-dihydro-indol-2-one (IIIa–e)

A mixture of isatin (0.01 mol), substituted acetophenone (IIa–e) (0.01 mol) and a catalytic amount of bleaching earth (10 wt% of pH 12.5) was placed in a PEG-400 (20 ml). The reaction mixture was stirred for 3–6 min at room temperature. The progress of the reaction was monitored by thin layer chromatography. The yellow solid that was separated was filtered and recrystallized from the alcohol–DMF mixture to afford 3-[2-(substituted-phenyl)-2-oxo-ethyl]-3-hydroxy-1,3-dihydro-indol-2-one (IIIa–e) in a significant yield.

Spectral data analysis of all compounds (IIIa–e)

3-[2-(4-Methoxy-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIa)

M.p. 190–195 °C; IR (KBr, ν, cm−1): 3,413 (–N–H), 2,930 (–C–H), 1,690 (–C=O), 1,665 (–C=O of lactam ring), 1,590 (C=C), 1155 (O–C); 1H NMR (400 MHz, DMSO-d 6) (δ, ppm): 8.7 (s, 1H, N–H), 8.5 (s, 1H conjugated –C–H), 3.7 (s, 3H, –OCH3), 6.95–7.90 (m, 8H, Ar–H); EIMS: m/z = 279 [M+1]+. Anal. calcd. for C17H13NO3: C, 73.11; H, 4.69; N, 5.02; O, 17.19 %. Found: C, 73.55; H, 4.45; N, 4.99; O, 17.10 %

3-(2-Oxo-2-p-tolyl-ethylidene)-1,3-dihydro-indol-2-one (IIIb)

M.p. 205–210 °C; IR (KBr, ν, cm−1): 3,400 (–N–H), 2,940 (–C–H), 1,695 (–C=O), 1,658 (–C=O of lactam ring), 1,585 (C=C), 1,140 (O–C); 1H NMR (400 MHz, DMSO-d 6) (δ, ppm): 8.5 (s, 1H, N–H), 8.2 (s, 1H conjugated –C–H), 1.8 (s, 3H, –CH3), 6.90–7.95 (m, 8H, Ar–H); EIMS: m/z = 263 [M+1]+. Anal. calcd. for C17H13NO2: C, 77.55; H, 4.98; N, 5.32; O, 12.15 %. Found: C, 76.72; H, 4.95; N, 5.11; O, 12.10 %

3-[2-(5-Chloro-2-hydroxy-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIc)

M.p. 235–240 °C; IR (KBr, ν, cm−1): 3,510 (–O–H), 3,390 (–N–H), 3,020 (=C–H), 1,680 (–C=O), 1,664 (–C=O of lactam ring), 1,580 (aromatic –C=C), 1,135 (O–C), 690 (C–Cl); 1H NMR (400 MHz, DMSO-d 6) (δ, ppm): 11.6 (s, 1H, O–H), 8.9 (s, 1H, N–H), 8.4 (S, 1H conjugated –C–H), 6.98–7.99 (m, 7H, Ar–H); EIMS: m/z = 299 [M+1]+. Anal. calcd. for C16H10ClNO3: C, 64.12; H, 3.36; Cl, 11.83; N, 4.67; O, 16.01 %. Found: C, 64.18 H, 3.385; N, 4.58; O, 16.05 %

3-[2-(3,5-Dichloro-2-hydroxy-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIId)

M.p. 246–250 °C; IR (KBr, ν, cm−1): 3,450 (–O–H), 3,270 (–N–H), 3,010 (=C–H), 1,670 (–C=O), 1,655 (–C=O of lactam ring), 1,580 (aromatic –C=C), 1,130 (O–C), 710 (C–Cl); 1H NMR (400 MHz, DMSO-d 6) (δ, ppm): 11.8 (s, 1H, O–H), 8.5 (s, 1H, N–H), 8.1 (S, 1H conjugated –C–H), 6.99–7.96 (m, 6H, Ar–H); EIMS: m/z = 333 [M+1]+. Anal. calcd. for C16H9Cl2NO3: C, 57.51; H, 2.71; Cl, 21.22; N, 4.19; O, 14.36 %. Found: C, 57.44; H, 2.80; Cl, 21.09; N, 4.21; O, 14.30 %

3-[2-(2-Hydroxy-5-methyl-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIe)

M.p. 225–230 °C; IR (KBr, ν, cm−1): 3,410 (–O–H), 3,320 (–N–H), 2,950 (–C–H), 1,683 (–C=O), 1,667 (–C=O of lactam ring), 1,588 (C=C), 1,130 (C–O); 1H NMR (400 MHz, DMSO-d 6) (δ, ppm): 10.8 (O–H), 8.6 (s, 1H, N–H), 8.3 (s, 1H conjugated –C–H), 1.9 (s, 3H, –CH3), 6.92–7.97 (m, 7H, Ar–H); EIMS: m/z = 279 [M+1]+. Anal. calcd. for C17H13NO3: C, 73.11; H, 4.69; N, 5.02; O, 17.19 %. Found: C, 73.01; H, 4.64; N, 4.9; O, 17.12 %

Results and discussion

It our efforts to develop a green, environmentally benign methodology, we are reporting here a green approach for the synthesis of 3-[2-(substituted-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIa–e). Because of the toxicological issues of hydrocarbon-based solvents on the environment, PEG-400 attracted our attention for use as a green reaction solvent with bleaching earth clay (pH 12.5) as catalyst. The cross-aldol condensation reaction between a mixture of isatin and substituted acetophenone catalyzed by bleaching clay earth and PEG 400 afforded corresponding α,β-unsaturated ketones (IIIa–e) within 5–6 min without formation of β-hydroxyl ketone [17]. Literature reviews revealed that, in conventional methodology, when toxic and expensive catalysts such as diethyl amine, piperidine and basic alumina are used in solvents like ethanol or DMF, the intermediate β-hydroxyl ketone was formed after 6–8 h. Further dehydration occurs during 1-2 h in the presence of acetic acid and concentrated HCl, affording the product 3-phenacylidine-2-indolinone (III). In some methodologies, the reaction between isatin and substituted acetophenone takes place by using basic alumina in microwave irradiation [18] to give the final product (III) without forming β-hydroxyl ketone (II) but here again, basic alumina is toxic and expensive. In our present methodology, we can get the desired product having better yields in a shorter time and in a greener way. It is well documented that PEG-400 and bleaching earth clay improves atom efficiency and yields of the product, as no side products are formed. Furthermore, the reaction using PEG-400 and bleaching earth clay has many advantages, such as their solvating ability and their acting as phase transfer catalyst, easy recyclability, ease of work-up, eco-friendly nature, and economical cost. PEG is non-toxic, non-halogenated, inexpensive, potentially recyclable and water soluble, which facilitates its removal from reaction products. Hence, we have developed new, simple, economical, safe, environmentally benign, alternative methodology for the synthesis of 3-[2-(substituted-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIa–e) (Scheme 1; Table 1).

Scheme 1
scheme 1

Synthesis of 3-[2-(substituted-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIa–e) using PEG-400

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Table 1 Synthesis of 3-[2-(substituted-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIa–e) using PEG-400
Full size table

Antimicrobial activity

The antibacterial activities of the synthesized compounds III a–e were determined by the agar diffusion method, as recommended by the National Committee for Clinical Laboratory Standards [26–28], against bacterial species Escherichia coli (MTCC 1650), Bacillus subtilis (MTCC 441), Staphylococcus aureus (MTCC 96), and Proteus vulgaris (MTCC 1771) and fungi like Aspergillus flavus (MTCC 2501), Candida albicans (MTCC 227), and Trichoderma viridae (MTCC 167). The drugs Ampicillin and Fluconazole are used as standard drugs for antibacterial and antifungal activities. The culture strains of the bacteria were maintained on nutrient agar slants at 37 ± 0.5 °C for 24 h.

The antibacterial activity was evaluated using nutrient agar plates seeded with 0.1 ml of the respective bacterial culture strain suspension prepared in sterile saline (0.85 %) at 105 CFU/ml dilutions The stock solutions were made by diluting compounds in DMSO to final concentrations ranging from 25 to 100 μg/ml. The wells, of 6 mm diameter, were filled with 0.1 ml of the compound solution separately for each bacterial strain. All the plates were incubated at 37 ± 0.5 °C for 24 h. Zones of inhibition of compounds in mm and MIC were noted.

For antifungal activity, all the culture strains of fungi were maintained on potato dextrose agar (PDA) slant at 27 ± 0.2 °C for 24–48 h until sporulation. Spores of strains were transferred into 5 ml of sterile distilled water containing 1 % Tween-80 (to properly suspend the spore). The spores were counted by a hemocytometer (106 CFU/ml). Sterile PDA plates were prepared containing 2 % agar, and 0.1 ml of each fungal spore suspension was spread on each plate and incubated at 27 ± 0.2 °C for 12 h. After incubation, wells were prepared using a sterile cork borer and each agar well was filled with 0.1 ml of the compound solution at the fixed concentration of 10 μg/ml. The plates were kept in a refrigerator for 20 min for diffusion and then incubated at 27 ± 0.2 °C for 24–28 h. After incubation, zones of inhibition of compounds were measured in mm, along with the standard.

The antibacterial results shown in Table 2 reveal that compounds IIIc, IIId, and IIIe show significant activity against all tested bacteria, such as E. coli (MTCC 1650), B. subtilis (MTCC 441), S. aureus (MTCC 96), and P. vulgaris (MTCC 1771) and possess significant zones of inhibition. The compounds IIIa and IIIb were less active against E. coli (MTCC 1650), and P. vulgaris (MTCC), and compound IIIb is inactive against B. subtilis (MTCC 441) and S. aureus (MTCC 96).

Table 2 Antimicrobial activity of synthesized 3-[2-(substituted-phenyl)-2-oxo-ethylidene]-1,3-dihydro-indol-2-one (IIIa–e)
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The results of the in vitro antifungal activities are summarized in compounds IIIa, IIIc, IIId, and IIIe which exhibited equal or stronger antifungal activities against all tested fungi, i.e.. A. flavus (MTCC 2501), C. albicans (MTCC 227), and T. viridae (MTCC 167) showing good zones of inhibition as compared to the standard drug, fluconazole. The antifungal activity of IIIb was shown to be lower due to a functional group effect. Significant antibacterial and antifungal activity was observed in compounds IIIa, IIIc, IIId, and IIIe due to the side chains R, R1, and R2 having hydroxyl and halo groups.

Conclusion

In summary, we have developed an efficient, one-pot and eco-friendly methodology for the synthesis of 3-phenacylidene-2-indolinones derivatives using recyclable bleaching earth clay (pH 12.5) as catalyst and PEG-400 as green solvent. The conventional methodology has two steps, and needs heating and has a longer reaction time. The synthesized compounds were confirmed by spectral analysis and further evaluated for their antimicrobial activity. The antibacterial and antifungal activity revealed that most of the compounds have moderate to good activity.