Eco-friendly Synthesis , Spectral Correlation Analysis , and Antimicrobial Activities of Substituted ( E )-1-benzylidene-2-( 3-nitrophenyl ) hydrazines

In the present study, a series of ten substituted (E)-1-benzylidene-2-(3-nitrophenyl)hydrazine compounds have been synthesized by condensation of 3-nitrophenylhydrazine with various substituted benzaldehydes using SiO2-H3PO4 catalyst under solvent free condition. This method involves shorter reaction time (5-10 minutes) with excellent yield (80-90%). The structures of the synthesized (E)-1-benzylidene-2-(3nitrophenyl)hydrazines have been characterized by their physical constants, UV, FT-IR and NMR spectral data. The characterized UV, FT-IR and NMR spectral data have been correlated with Hammett constants using single-linear and multi-linear regression analysis. From the results of single-linear and multi-linear regression analysis the effect of substituents on the spectral data have been discussed. The antimicrobial activities of all the (E)-1-benzylidene-2-(3-nitrophenyl)hydrazine compounds have been tested using BauerKirby method.


INTRODUCTION
Now-a-days solvent-free approach to the synthesis of molecules becomes an interesting one.Since the majority of solvents are either toxic or inflammable, they cause serious consequences during synthesis.In many cases, the solvent-free approach involves improves selectivity, reduces reaction time, simplifies separation, and purification of products where compared to the conventional methods [1][2][3][4][5].The development of greener methods is a major challenge in synthetic chemistry.Among the several advantages of green chemistry, the reduction or replacement of volatile organic solvents from the reaction medium is of utmost importance [6,7].Due to the increasing environmental and economical concern in the recent years, it is now essential for chemists to search for as many new environmentally benign methods as possible.The aims of green chemistry [8] are the prevention of wastes and the generation of substances with little or no toxicity to humans and the environment, in order to maximize atom economy.This can be achieved only if the final product is synthesized by avoiding the use of solvents whether they are harmful or not.
Green chemistry is used for designing chemical products and processes by reducing or eliminating the use of harmful chemicals and/or generating hazardous substances.Various green chemistry methods involving stirring, phase transfer catalyst, ionic liquid and many more techniques like microwave techniques include approaches for the creation of ''benign-by-design'' synthetic methods which are now accepted worldwide [9].Processes designed by green routes help in the promotion of resource and efficient utilization of energy.They involve low level of waste and hence the processes are becoming economically and environmentally beneficial.
Hydrazone-containing azomethine -NH─N=CH─ protons constitute an important class of compounds for development of new drugs.Many researchers had synthesized such type of compounds [10] as target molecules and evaluated their biological activities.Hydrazones was reported to possess antimicrobial [11], antitubercular [12,13], anticonvulsant [14], analgesic [15], anti-inflammatory [16], antiplatelet [17], anticancer [18,19], antifungal [20], antiviral [21], antitumoral [22,23], antibacterial [24] and antimalarial [25] activities.In recent years, environmentally benign synthetic methods have expected considerable attention and some solvent-free protocols have been developed [26,27].Schmeyers et al. reported the solid-state synthesis of various kinds of benzylidene aniline derivatives by grinding together solid anilines and solid benzaldehydes [28].Varma et al. reported the clay-catalyzed synthesis of imines and enamines under solvent-free conditions using microwave irradiation [29].There has been a large emphasis, both in the chemical industry and in academic research, on the development of environmentally benign solvents and reaction conditions.This is largely owed to the fact that traditional solvents, such as volatile organic solvents, have been implicated in a number of environmental problems [30].
In the present work, we report the synthesis of (E)-1-benzylidene-2-(3-nitrophenyl)hydrazine derivatives using grinding method.The reactions have been carried out by grinding together equivalent amounts of the appropriate substituted aldehyde and 3-nitrophenylhydrazine in the presence of SiO2-H3PO4 in a porcelain mortar, under solvent-free conditions.Grinding for about 5-10 minutes led to a colored solid mass of the crude product.The completion of the reaction was checked by TLC.Purification was carried out by simple Buchner filtration, washing with cold water, and crystallization from ethanol solvent to give the respective (E)-1benzylidene-2-(3-nitrophenyl)hydrazine derivatives.This method is simple and effective in terms of its short reaction time, excellent yields, and the formation of a single product.The various spectral data of these (E)-1-benzylidene-2-(3nitrophenyl)hydrazines have been utilized for studying the substituent effects through Hammett correlations.The antimicrobial activity of all the (E)-1-benzylidene-2-(3-nitrophenyl)hydrazine compounds tested for Bauer-Kirby method.

Physical Measurement
All the chemicals used in the present investigation were purchased from Sigma-Aldrich and E-Merck chemical companies.Melting points were determined with open glass capillaries on SUNTEX melting point apparatus and are uncorrected.The UV absorption spectrum of all the synthesized (E)-1-benzylidene-2-(3nitrophenyl)hydrazines were recorded in SHIMADZU-1650 spectrophotometer (max, nm) in spectral grade ethanol.Fourier-transform infrared spectra (KBr, 4000-400 cm -1 ) were recorded as KBr pellets on SHIMADZU-2010 Fourier transform spectrometer.Bruker AV400 NMR spectrometer operating at 400 MHz has been used for recording 1 H spectra and that operating at 100 MHz for recording 13 C spectra in CDCl3 solvent using TMS as internal standard.

Preparation of SiO2-H3PO4 catalyst
In a 50 mL Borosil beaker, 3 g of silica and 3 mL of orthophosphoric acid have been taken and mixed thoroughly with glass rod at room temperature.This mixture has been heated on a hot air oven at 100 C for 1 h, cooled to room temperature, stored in a borosil bottle and tightly capped [31].

General procedure for synthesis of substituted (E)-1-benzylidene-2-(3-nitrophenyl)hydrazines
A mixture of appropriate 3nitrophenylhydrazine (2 mmole), substituted benzaldehyde (2 mmole) and SiO2-H3PO4 (0.5 g) was thoroughly ground with a pestle in an open mortar at room temperature for 5-10 minutes.The initial syrupy reaction mixtures get solidified within 3-5 minutes.The completion of the reaction was monitored by use of TLC.After the completion of the reaction 10 mL of dichloromethane was added and the organic layer separated by filtration, yielding solid product on evaporation.The solid, on recrystallization with ethanol, afforded glittering orange solid.The insoluble catalyst has been recycled by washing with ethyl acetate (10 mL) followed by drying in an air oven at 100°C for 1h and reused for further reactions.The general reaction of substituted (E)-1-benzylidene-2-(3-nitrophenyl)hydrazines is as shown in Scheme 1. Scheme 1. Synthesis of substituted (E)-1benzylidene-2-(3-nitrophenyl)hydrazines.

RESULTS AND DISCUSSION
In the present research work, we adopt a mild, efficient, high yielding process for the synthesis of substituted (E)-1-benzylidene-2-(3nitrophenyl)hydrazines from different aromatic aldehydes with 3-nitrophenylhydrazine in SiO2-H3PO4 catalyst under solvent-free condition.This method involves shorter reaction time with excellent yield (87-95%).This catalyst was reused for further reaction runs in the synthesis of the parent hydrazine with equal molar quantities of 3-nitrophenylhydrazine (2 mmole) benzaldehyde (2 mmole) and SiO2-H3PO4 (0.5 g) catalyst (entry 3a).In this reaction, the first run gave 90 % product.The second, third and fourth runs gave 89.5 % product.The fifth run gave 89% product.There is no appreciable change of percentage of product observed in these reaction runs.It is noteworthy to mention that the green route method requires simple work-up procedure involving simple filtration to isolate the products as they are insoluble in water.The physical constants of synthesized (E)-1benzylidene-2-(3-nitrophenyl)hydrazines shown in Table -1

Uv-Vis spectral study
The recorded ultraviolet absorption maximum λmax C=N (nm) values of all the substituted (E)-1-benzylidene-2-(3-nitrophenyl)hydrazine compounds are presented in Table-2.This data has been correlated with different Hammett substituent constants and Swain-Lupton's F and R parameters using spectral correlation analyses according to the approach of John Shorter [33].The results of single regression analyses [33] are presented in Table -3  where λ0 is the frequency for the parent member of the series.
All the correlations regarding UV spectral data shows positive ρ values.This positive ρ value indicates the operation of normal substituent effects.In view of the inability of the Hammett constants to produce individually satisfactory correlations with the UV absorption maximum values, the authors think that, it is worthwhile to seek multi-correlation analysis of Hammett σI and σR constants and Swain-Lupton's [40] F and R parameters.The multicorrelation analyses have shown much better correlations for all the substituents and are shown in the following equations ( 2) and (3).

IR Spectral study
The assigned infrared νC=N (cm -1 ) stretching frequency values of all the substituted (E)-1benzylidene-2-(3-nitrophenyl)hydrazine compounds are presented in Table-2.These IR frequency νC=N (cm -1 ) values are correlated with different Hammett substituent constants and F and R parameters using single-linear and multi-linear regression analyses [34][35][36][37][38][39].The results of the statistical analysis [40] are presented in Table-3.In this correlation the structure parameter Hammett equation employed is as shown in given equation (4).
where νo is the frequency for the parent member of the series.
The results of the spectral correlation analyses are presented in Table-3.From this Table, it is evident that the infrared νC=N (cm -1 ) stretching frequency values of all the (E)-1-benzylidene-2-(3nitrophenyl)hydrazine compounds, except those with 4-NO2 substituent, has shown good linear correlations with Hammett substituent constant σ (r = 0.904) and σ + (r = 0.905) parameters.The substituents that have been given exception were reducing the correlations considerably when they are included in regression.However the IR stretching frequency νC=N (cm -1 ) values of all the (E)-1-benzylidene-2-(3nitrophenyl)hydrazine compounds, have shown unsatisfactory correlations (r < 0.900) with the remaining Hammett substituent constants viz., σI, σR and F and R parameters.The unsatisfactory correlation is due to the weak polar, resonance, inductive and field effects of the substituents to predict their reactivity through the conjugative structure shown in Figure 1.All the correlations have shown positive ρ value except resonance parameters.It indicates the operation of normal substituent effect with respect to infrared stretching frequency νC=N (cm -1 ) values in all the (E)-1-benzylidene-2-(3nitrophenyl)hydrazine compounds.

NMR Spectral study 1 H NMR spectral study
The 1 H NMR spectra of all the synthesized (E)-1-benzylidene -2-(3-nitrophenyl)hydrazine compounds in the present investigation are recorded in CDCl3 using TMS as internal standard.The signals of the imine protons have been assigned and their chemical shift values are presented in Table-2.The chemical shift δCH=N (ppm) values of all the (E)-1benzylidene-2-(3-nitrophenyl)hydrazine compounds have been correlated with Hammett substituents constants and F and R parameters using single-linear and multi-linear regression analysis [34][35][36][37][38][39].In this correlation the structure parameter Hammett equation employed is as shown in equation (7).δ = ρσ +δ0 (7) where δ0 is the chemical shift of the corresponding parent compound.
The results of the statistical analysis are presented in Table-3.From this Table, it is evident that the 1 H NMR chemical shift δCH=N (ppm) values of all the substituted (E)-1-benzylidene-2-(3nitrophenyl)hydrazine compounds reveal that all the substituents have shown poor (r < 0.900) correlations [33][34][35][36][37][38] with Hammett substituent constants namely σ, σ + , σI, σR and Swain-Lupton's F and R parameters [39].The poor correlation is attributed to the conjugative structure shown in Figure 1.All the correlations have shown positive ρ values.This positive ρ value indicates the operation of normal substituent effect.

Antibacterial activity
The antibacterial sensitivity assay has been performed by using Kirby-Bauer [41]

Antifungal activity
The antifungal activities of all the synthesized substituted (E)-1-benzylidene-2-(3nitrophenyl)hydrazine compounds have been studied against three fungal species namely, Aspergillus niger, Mucor species and Trigoderma viride.The antifungal activities of all the substituted (E)-1benzylidene-2-(3-nitrophenyl)hydrazine compounds have been studied and are shown in Figure 4 for Plates (1-6) and the corresponding clustered column chart is shown in Figure 5.The antifungal sensitivity assay has been performed using Kirby-Bauer [41]
The synthesized substituted (E)-1benzylidene-2-(3-nitrophenyl)hydrazine compounds are characterized by their physical constants, UV, FT-IR, and NMR spectral data.The UV, FT-IR and NMR spectral data of the substituted (E)-1-benzylidene-2-(3-nitrophenyl)hydrazine compounds have been correlated with Hammett constants σ, σ + , σI, σR and swain-Lupton's F and R parameters using single and multi-regression analyses.From the results of correlation analysis Hammett substituent constants σ and σ + produce satisfactory with FT-IR spectral data.The UV and 1 H NMR spectral data have shown poor correlations.The 13 C NMR spectral data produce satisfactory correlations with σ, σ + , σR and Swain-Lupton's R parameters.Multi-regression analyses produce satisfactory correlations for all the Hammett substituent constants and F and R parameters.The antimicrobial activities of all the compounds have shown good to moderate activities against all the microorganisms.

Table - 4
compounds have been studied and the zone of inhibition values of the effect is given in Table-5.From Table-5 reveals that all the compounds have moderate to good antifungal activity .Antibacterial activity of substituted (E)-1-benzylidene-2-(3-nitrophenyl)hydrazines.