Available online at www.worldscientificnews.com
World Scientific News
8 (2015) 211-226 EISSN 2392-2192
Synthesis, Assessment of substituent effect and Antimicrobial activities of some substituted (E)-N-
benzylidene-4H-1,2,4-triazol-4-amines
R. Senbagama, M. Rajarajana, R. Vijayakumara, V. Manikandana, S. Balajia, G. Vanangamudia, G. Thirunarayananb,*
aPG & Research Department of Chemistry, Government Arts College, C-Mutlur,
Chidambaram - 608 102, India
bDepartment of Chemistry, Annamalai University, Annamalainagar - 608 002, India
*E-mail address: [email protected]
ABSTRACT
A series of heterocyclic Schiff base compounds have been synthesized from 4H-1,2,4-triazol-4-
amine with various substituted benzaldehydes and were refluxed for 3h with 20 mL of absolute
ethanol. The purity of all the Schiff base compounds has been checked using their physical constants
and spectral data. The UV λmax(nm), IR νC=N(cm-1
), NMR δ(ppm) of CH=N and C=N spectral data
have been correlated with Hammett substituent constants and Swain-Lupton’s parameters using single
and multi-linear regression analysis. From the results of statistical analysis, the effect of substituents
on the above spectral data has been studied. The single parameter correlation with few Hammett
constants and Swain-Lupton’s parameters gave satisfactory correlation coefficients whereas all
multiple correlations gave satisfactory correlation coefficients with Inductive, Resonance, Field and
Swain-Lupton’s parameters. The antimicrobial activities of all the Schiff bases have been studied
using standard methods.
Keywords: synthesis; heterocyclic Schiff bases; UV; IR and NMR spectra; QSAR study and
antimicrobial study
1. INTRODUCTION
Schiff bases are prepared by condensation of primary amine with a compound
containing an active carbonyl group [1]. They are also known as ‘azomethines’, ‘anils’ or
‘imines’. Among the large number of synthetic and naturally occurring nitrogen donor
molecules, schiff bases are of the greatest interest. In general, Schiff bases are represented by
the general formula RCH=NR’ where >C=N is the azomethine group. The colour of the schiff
bases is due to the presence of this azomethine (>C=N) linkage and can vary by introducing
World Scientific News 8 (2015) 211-226
-212-
other auxochromic groups. Schiff basess are characterized by the –N=CH– group and this
finds importance in elucidating the mechanism of transamination and racemization reactions
in biological systems [2,3]. They have been used in the study of asymmetric catalysis [4],
magnetic properties [5], dyes and photographic chemicals[6], corrosion inhibitors [7,8] and in
the preparation of polymers[9].
Schiff’s bases have attracted considerable attention of organic chemists due to their
significant biological activities like anticancer [10], antitumor [11], anti-inflammatory agents
[12], insecticidal [13], antituberculosis [14], antimicrobial [15], anticonvulsant [16] activity.
In the present day the correlation analysis of Schiff base compounds has become one of
the important studies for researchers to study their substituent effect. Recently, the correlation
analysis of Schiff base compounds have been reported [17-19]. The author also synthesized
ten number of heterocyclic Schiff base compounds from 4H-1,2,4-triazol-4-amine and
substituted benzaldehydes and studied their substituent effects using single and multi-linear
regression analysis. The biological activities of these Schiff base derivatives have been
studied.
2. EXPERIMENTAL
2. 1. General
All the chemicals used have been purchased from Sigma–Aldrich and E-Merck
chemical companies. Melting points of all of substituted (E)-N-benzylidene-4H-1,2,4-triazol-
4-amine compounds have been determined in open glass capillaries on a Mettler FP51
melting point apparatus and are uncorrected. The UV spectra of all synthesized (E)-N-
benzylidene-4H-1,2,4-triazol-4-amine compounds have been recorded using SHIMADZU-
1650 SPECTROMETER in spectral grade methanol. IR spectra (KBr, 4000-400 cm-1
) have
been recorded on AVATAR-300 Fourier transform spectrophotometer. The NMR spectra of
all the substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds have been
recorded in BRUKER 400 spectrometer operating at 400 MHz for 1H NMR spectra and 100
MHz for 13
C NMR spectra in CDCl3 solvent using TMS as internal standard.
2. 2. Synthesis of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds
An appropriate mixture of equimolar quantities of 4H-1,2,4-triazol-4-amine (0.01mol),
substituted benzaldehyde (0.01mol) and 0.5ml acetic acid were refluxed for 3h with 20 mL of
absolute ethanol [17].
Scheme 1. X = H, 3-Br, 4-Br, 3-Cl, 4-Cl, 4-F, 4-CH3, 4-OCH3, 3-NO2, 4-NO2
World Scientific News 8 (2015) 211-226
-213-
The completion of the reaction was monitored by TLC continuously. The resultant
mixture was cooled at room temperature. Then the precipitate obtained, was filtered at the
filter pump and washed several times with cold water. A pale yellow solid was obtained as the
final product. This crude product was recrystallized from ethanol to get glittering colorless
solid, and their melting points have been noted. The general reaction is shown in Scheme 1.
Table 1. The UV, IR and NMR spectroscopic data of substituted
(E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds.
Entry X M.F. M. W. M. p.
(°C)
λmax
(nm)
ν C=N
(cm-1
)
δ1H
CH=N
(ppm)
δ13
C
C=N
(ppm)
1 H C9H10N4 174 134-135 275.20 1602.99 8.644 156.82
2 3-Br C9H7BrN4 251 113-114 273.00 1619.03 8.620 167.71
3 4-Br C9H7BrN4 251 174-175 284.00 1588.11 8.573 158.62
4 3-Cl C9H7ClN4 206 107-108 272.80 1620.19 8.301 155.44
5 4-Cl C9H7ClN4 206 150-151 282.00 1593.23 8.654 155.39
6 4-F C9H7FN4 190 120-121 277.20 1602.40 8.649 153.77
7 4-CH3 C10H10N4 18 128-129 283.00 1607.74 8.647 157.17
8 4-OCH3 C10H10N4O 202 87-88 305.40 1604.76 8.598 163.43
9 3-NO2 C9H7N5O2 217 108-109 259.60 1578.31 8.730 155.34
10 4-NO2 C9H7N5O2 217 228-229 295.40 1592.32 8.684 154.65
3. RESULTS AND DISCUSSION
All the compounds synthesized in the present investigation have been confirmed by
their physical constants and UV, IR and NMR spectral data as shown in Table 1. The spectral
data of all the synthesized substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine
compounds have been correlated with Hammett substituent constants and F and R parameter
and are shown in Table-2.
3. 1. UV-visble spectral correlations of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-
amine compounds
The assigned UV absorption maximum λmax (nm) values of all the substituted (E)-N-
benzylidene-4H-1,2,4-triazol-4-amine compounds are presented in Table 1. These UV
absorption maximum values are correlated with different Hammett substituent constants and
F and R parameters using single and multi-linear regression analyses [17-21]. Hammett
equation employed, for the correlation analysis, involving the UV absorption maximum is
shown in equation (1).
λ = ρ σ + λ0 … (1)
where λ0 is the absorption maximum of the parent member of this series.
The results of statistical analysis [17-21] are shown in Table 2. The assigned UV
absorption maximum λmax (nm) values of all the substituted (E)-N-benzylidene-4H-1,2,4-
triazol-4-amine compounds, except that with 4-NO2 substituent have shown satisfactory
World Scientific News 8 (2015) 211-226
-214-
1 2
3
4
5
6
7
8
9
10
250,00
260,00
270,00
280,00
290,00
300,00
310,00
-1 -0,5 0 0,5 1
λmax
(n
m)
σ+
correlation with Hammett substituent constants σ+ (r = 0.902) only. All the compounds
except those with H, 4-CH3 and 4-NO2 substituents have shown satisfactory correlation with
Hammett constant σI (r = 0.952) only.
In addition, all the substituents except those with 4-F and 4-NO2 substituents have
shown satisfactory correlation with R parameter (r = 0.905). When those substituents that
have been exception are included in regression they reduce the correlations considerably. The
single linear plot of UV absorption maximum λmax (nm) values against Hammett constant σ+
is shown in the following Fig. 1.
Fig. 1. Single linear plot of λmax (nm) values of substituted
(E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds Vs σ+
Table 2. Results of statistical analysis of UV λmax (nm), IR νC=N (cm-1
), NMR δ1H(ppm) CH=N and
δ13
C (ppm) C= N of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds with
Hammett substituent constants σ, σ+, σI, σR and F and R parameters.
Frequency Constants r I ρ s n Correlated derivatives
λmax(nm) σ 0.846 284.20 -14.704 12.31 10
H, 3-Br, 4-Br, 3-Cl,
4-Cl, 4-F, 4-CH3, 4-OCH3,
3-NO2, 4-NO2
σ+ 0.915 283.05 -13.637 11.54 9
H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-F, 4-CH3, 4-OCH3, 3-NO2
σI 0.952 284.31 -9.053 13.27 7 3-Br, 4-Br, 3-Cl, 4-Cl, 4-F,
4-OCH3, 3-NO2,
σR 0.833 277.92 -20.664 12.69 10
H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-F, 4-CH3, 4-OCH3, 3-NO2,
4-NO2
11. H
12. 3-Br
13. 4-Br
14. 3-Cl
15. 4-Cl
16. 4-F
17. 4-CH3
18. 4-OCH3
19. 3-NO2
20. 4-NO2
1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
World Scientific News 8 (2015) 211-226
-215-
r = correlation coefficient, I = intercept, ρ = slope, s = standard deviation, n = number of correlated derivatives
F 0.875 284.02 -8.002 13.31 10
H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-F, 4-CH3, 4-OCH3, 3-NO2,
4-NO2
R 0.905 277.06 -19.426 12.48 8 H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-CH3, 4-OCH3, 3-NO2
νC=N(cm-1
) σ 0.937 1604.24 -14.261 12.99 8 H, 4-Br, 4-Cl, 4-F, 4-CH3,
4-OCH3, 3-NO2, 4-NO2
σ+ 0.934 1602.47 -9.347 13.20 8
H, 4-Br, 4-Cl, 4-F, 4-CH3,
4-OCH3, 3-NO2, 4-NO2
σI 0.903 1608.93 -20.425 13.01 8 H, 4-Br, 4-Cl, 4-F, 4-CH3,
4-OCH3, 3-NO2, 4-NO2
σR 0.843 1597.12 -27.591 12.66 10
H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-F, 4-CH3, 4-OCH3, 3-NO2,
4-NO2
F 0.835 1608.57 -18.794 13.11 10
H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-F, 4-CH3, 4-OCH3, 3-NO2,
4-NO2
R 0.839 1596.84 -21.380 12.87 10
H, 3-Br, 4-Br, 3-Cl, 4-Cl,
4-F, 4-CH3, 4-OCH3, 3-NO2,
4-NO2
δCH=N(ppm) σ 0.909 8.60 0.032 0.123 9 H,3-Br, 4-Br, 4-Cl, 4-F,
4-CH3, 4-OCH3, 3-NO2, 4-NO2
σ+ 0.902 8.61 0.005 0.123 9
H,3-Br, 4-Br, 4-Cl, 4-F,4-CH3,
4-OCH3, 3-NO2, 4-NO2
σI 0.893 8.60 0.014 0.124 10 H, 3-Br, 4-Br, 3-Cl, 4-Cl, 4-F,
4-CH3, 4-OCH3, 3-NO2, 4-NO2
σR 0.928 8.63 0.163 0.118 9 H,3-Br, 4-Br, 4-Cl, 4-F,4-CH3,
4-OCH3, 3-NO2, 4-NO2
F 0.913 8.59 0.049 0.123 9 H,3-Br, 4-Br, 4-Cl, 4-F,4-CH3,
4-OCH3, 3-NO2, 4-NO2
R 0.922 8.63 0.106 0.121 9 H,3-Br, 4-Br, 4-Cl, 4-F,4-CH3,
4-OCH3, 3-NO2, 4-NO2
δC=N(ppm) σ 0.901 158.55 -3.073 4.54 8 H, 4-Br, 3-Cl, 4-Cl, 4-F, 4-CH3,
3-NO2, 4-NO2
σ+ 0.902 158.24 -2.455 4.51 8
H, 4-Br, 3-Cl, 4-Cl, 4-F, 4-CH3,
3-NO2, 4-NO2
σI 0.816 158.99 -2.938 4.62 10 H, 3-Br, 4-Br, 3-Cl, 4-Cl, 4-F,
4-CH3, 4-OCH3, 3-NO2, 4-NO2
σR 0.823 157.15 -4.939 4.56 10 H, 3-Br, 4-Br, 3-Cl, 4-Cl, 4-F,
4-CH3, 4-OCH3, 3-NO2, 4-NO2
F 0.826 159.49 -4.063 4.56 10 H, 3-Br, 4-Br, 3-Cl, 4-Cl, 4-F,
4-CH3, 4-OCH3, 3-NO2, 4-NO2
R 0.927 156.88 -5.002 4.50 8 H, 4-Br, 3-Cl, 4-Cl, 4-CH3,
4-OCH3, 3-NO2, 4-NO2
World Scientific News 8 (2015) 211-226
-216-
However, UV absorption maximum λmax (nm) values of all the substituted (E)-N-
benzylidene-4H-1,2,4-triazol-4-amine compounds have shown poor correlations (r < 0.900)
with the remaining Hammett constant σR and F parameter.
The poor in correlation is attributed to weak resonance and field effect of the
substituents for predicting the reactivity through resonance as per the resonance conjugative
structure shown in Fig. 2.
Fig. 2. The resonance-conjugative structure.
All the correlations with Hammett constants and F and R parameters have shown
negative ρ values. This indicates the operation of reverse substituent effect with respect to UV
absorption maximum λmax (nm) values of all the substituted (E)-N-benzylidene-4H-1,2,4-
triazol-4-amine compounds.
Since, some of the single regression analyses have shown poor correlation with few
Hammett constants and F and R parameters. Hence, the authors think that worthwhile to seek
the multi regression analysis of the UV absorption maximum λmax (nm) values of all the
substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds with inductive, resonance
and swain-Lupton’s[15] parameters produces satisfactory correlations as shown in equations
(2) and (3).
λmax (nm) = 280.780 (±9.272) - 6.89 (± 2.584) σI - 19.618 (±3.809) σ R … (2)
(R = 0.900, n = 10, P > 90%)
λmax (nm) = 280.574 (± 8.772) - 8.751 (±2.378) F - 19.778 (±2.726) R … (3)
(R = 0.900, n = 10, P > 90%)
3. 2. IR Spectral correlations of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine
compounds
The assigned infrared stretching frequency νC=N (cm-1
) values of all the substituted
(E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds are presented in Table 1.
These infrared stretching frequency values are correlated [17-21] using single and
multi-linear regression analyses. The structure parameter correlation involving group
frequencies, the employed Hammett equation is shown in equation (4).
ν = ρ σ + ν0 … (4)
where ν0 is the frequency of the parent member of this series.
World Scientific News 8 (2015) 211-226
-217-
1
2
3
4
5
6
7 8
9
10
1575,00
1580,00
1585,00
1590,00
1595,00
1600,00
1605,00
1610,00
1615,00
1620,00
1625,00
-0,4 -0,2 0 0,2 0,4 0,6 0,8 1
νC=N
(cm
-1)
σ
The results of the statistical analysis [17-21] are presented in Table-2, it is evident that
the infrared that the infrared stretching frequency νC=N (cm-1
) values of all the substituted
(E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds, except those with 3-Br and 3-Cl
substituents have shown satisfactory correlations with Hammett constants σ (r = 0.937), σ+ (r
= 0.934) and σI (r = 0.903). When these substituents that have been given exception are
included in regression they reduce the correlations considerably. The single linear plot of IR frequency νC=N (cm
-1) values against Hammett constant σ is shown in the following Fig. 3.
However the infrared stretching frequency νC=N (cm-1
) values of all the substituted
(E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds, have shown poor correlations (r <
0.900) with the Hammett constant σR and F & R parameters. This is attributed to weak
resonance and field effect of the substituents to predict the reactivity on frequency through
resonance as per conjugative structure shown in Fig. 2.
Fig. 3. Single linear plot of IR frequency νC=N (cm-1
)values of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds Vs σ
All the correlations have shown negative ρ values. This indicates the operation of
reverse substituent effect with respect to infrared stretching frequency νC=N (cm-1
) values of
all the substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amines.
Since some of the single regression analyses, have shown poor correlations with
Hammett constant and F and R parameters. It is decided to go for multi regression analysis.
The multi regression analysis of the stretching frequency νC=N(cm-1
) values of all Schiff base
compounds with inductive, resonance and Swain-Lupton’s [22] parameters produce
satisfactory correlations as shown in equations (5) and (6).
νC=N (cm-1
) = 1604.44 (±8.721) - 17.679 (±5.82) σI - 24.908 (±4.151) σR… (5)
(R = 0.999, n = 10, P > 95%)
νC= N (cm-1
) = 8.608 (± 0.085) + 0.084 (± 0.021) F + 0.108 (± 0.051) R … .(6)
(R = 0.999, n = 10, P > 95%)
31. H
32. 3-Br
33. 4-Br
34. 3-Cl
35. 4-Cl
36. 4-F
37. 4-CH3
38. 4-OCH3
39. 3-NO2
40. 4-NO2
21. H
22. 3-Br
23. 4-Br
24. 3-Cl
25. 4-Cl
26. 4-F
27. 4-CH3
28. 4-OCH3
29. 3-NO2
30. 4-NO2 1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
World Scientific News 8 (2015) 211-226
-218-
1 2
3
4
5 6 7 8
9
10
8,250
8,300
8,350
8,400
8,450
8,500
8,550
8,600
8,650
8,700
8,750
8,800
-0,4 -0,2 0 0,2 0,4 0,6 0,8 1
δC
H=
N(p
pm
)
σ
3. 3. NMR spectral correlations of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-
amine compounds
The observed chemical shift values (ppm) of all the substituted (E)-N-benzylidene-4H-
1,2,4-triazol-4-amine compounds are presented in Table 1. These chemical shift values
(ppm) are correlated with different Hammett substituent constants and F and R parameters
using single and multi-linear regression analyses [17-21]. In this correlation, the structure-
parameter equation employed is shown in equation (7).
δ = ρσ + δ0 … (7)
where δ0 is the chemical shift of the corresponding parent compound.
3. 3. 1. 1
H NMR spectral correlation:
From Table 2, it is evident that the 1H NMR chemical shift δCH=N (ppm) values of all
the substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds except that with 3-Cl
substituent have shown satisfactory correlations with σ(r = 0.909), σ+(r = 0.902), σR (r =
0.928), F (r = 0.913) and R (r = 0.922) parameters.
The single linear plot of 1H NMR chemical shift δCH=N (ppm) values against Hammett
constant σ is shown in the following Fig. 4.
Fig. 4. Single linear plot of 1H NMR chemical shift δCH=N (ppm) values of
substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds Vs σ
The remaining Hammett constant σI has shown poor correlation (r < 0.900) with all the
substituents. The failure in correlation is due to the reason that has been stated earlier with
resonance conjugative structure as shown in Fig. 2.
All the correlations with Hammett substituent constants F and R parameters have
shown positive ρ values. It indicates the operation of normal substituent effect with respect to 1H NMR spectral data of all the compounds. While seeking the multi–correlation collectively
the inductive, resonance and field effects [22] have shown satisfactory correlation as shown in
the equations (8) and (9).
1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
World Scientific News 8 (2015) 211-226
-219-
1
2
3
4 5
6
7
8
9
10
152,00
154,00
156,00
158,00
160,00
162,00
164,00
166,00
168,00
170,00
-0,4 -0,2 0 0,2 0,4 0,6 0,8 1
δC
=N
(pp
m)
σ
δC= N (ppm) = 8.633 (±0.087) - 0.0933 (±0.022) σI + 0.1639 (±0.05)σR … (8)
(R = 0.999, n = 10, P > 95%)
δC= N (ppm) = 8.608 (±0.085) + 0.054 (± 0.002) F + 0.108 (± 0.051) R … (9)
(R = 0.999, n = 10, P > 95%)
3. 3. 2. 13
C NMR spectral correlation
The results of the statistical analysis [17-21] are presented in Table-2. It is evident that
the 13
CNMR chemical shift δ C=N (ppm) values of all the substituted (E)-N-benzylidene-4H-
1,2,4-triazol-4-amine compounds except 3-Br and 4-OCH3 substituents have shown
satisfactory correlations with Hammett constants σ (r = 0.901) and σ+(r = 0.902). All the
compounds except 3-Br and 4-F have also shown satisfactory correlation with R (r = 0.927)
parameter.
The single linear plot of 13
C NMR chemical shift δC=N (ppm) values against Hammett constant σ is shown in the following Fig. 5.
Fig. 5. Single linear plot of 13
C NMR chemical shift δC=N (ppm) values of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds Vs σ
The remaining Hammett constants σI, σR and F parameter have shown poor correlations
(r < 0.900). The failure in correlation is due to the reason that has been stated earlier with
resonance conjugative structure as shown in Fig. 2. All the Hammett constants F and R
parameters have shown positive ρ values. This shows that the normal substituent effect
operates in all systems. While seeking the multi-correlation, collectively with inductive,
resonance and field effects [22] satisfactory correlation as shown in equations (10) and (11).
δC = N(ppm) = 158.16(±3.329) - 2.434(±0.41)σI - 4.578(±1.52) σR …(10)
(R = 0.904, n = 10, P > 95%)
δC= N(ppm) = 158.59(±3.11) – 4.259(±1.52)F - 5.17(±1.72) R …(11)
(R = 0.904, n = 10, P > 95%)
1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
World Scientific News 8 (2015) 211-226
-220-
3. 4. Antimicrobial activities of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine
compounds
3. 4. 1. Antibacterial activity
Antibacterial sensitivity assay has been performed by using [23] disc diffusion
technique. In each Petri plate about 0.5 ml of the test bacterial sample has been spread
uniformly over the solidified Mueller Hinton agar using sterile glass spreader. Then the discs
with 5mm diameter made up of Whatmann No.1 filter paper, impregnated with the solution of
the compound have been placed on the medium using sterile forceps. The plates have been
incubated for 24 hours at 37 ºC by keeping the plates upside down to prevent the collection of
water droplets over the medium. After 24 hours, the plates have been visually examined and
the diameter values of the zone of inhibition were measured. Triplicate results have been
recorded by repeating the same procedure.
The antibacterial screening effect of all the synthesized substituted (E)-N-benzylidene-
4H-1,2,4-triazol-4-amine compounds is shown in (Fig. 6) (Plates 1-10). The antibacterial
activities have been studied against three gram positive pathogenic strains Bacillus substilis,
Micrococcus luteus, Staphylococcus aureus and two gram negative strains Escherichia coli
and Pseudomonas aurogenosa. The disc diffusion technique was followed, at a concentration
of 250μg/mL with ciprofloxacin taken as the standard drug. The zone of inhibition is
compared using Table 3 and the corresponding clustered column chart is shown in (Fig. 7). A
good antibacterial activity has been possessed by all substituents on the microorganisms in
general. The substituents 3-Cl, 4-F, 3-NO2 and 4-NO2 have shown very good antibacterial
activity against Bacillus subtilis. The substituents 4-Br and 4-NO2 have shown very good
activity against Micrococcus luteus. The substituent 4-CH3 has shown very good activity
against Staphylococcus aureus. The substituents 3-Cl, 4-F, 4-CH3, and 4-NO2 have shown
very good antibacterial activity against E.coli. The substituents 3-Cl, 4-Cl, 4-F, 4-CH3, 3-NO2
and 4-NO2 have shown very good antibacterial activity against Pseudomonas.
Table 3. Antibacterial activity of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds.
S.No. Substituents
Zone of inhibition (mm)
Gram positive Bacteria Gram negative Bacteria
B.subtilis M.luteus S.aureus E.coli P.aeruginosa
1 H 7 7 6 7 6
2 3-Br 6 9 6 10 7
3 4-Br 7 11 6 8 7
4 3-Cl 10 7 9 10 9
5 4-Cl 6 8 6 7 8
6 4-F 10 9 6 11 9
7 4-CH3 8 7 12 10 8
8 4-OCH3 7 8 7 9 7
9 3-NO2 13 9 9 9 9
10 4-NO2 12 11 8 11 8
Standard Ciprofloxacin 9 9 10 8 7
Control DMSO 0 0 0 0 0
World Scientific News 8 (2015) 211-226
-221-
Plate-1 Plate-2
Plate-3 Plate-4
Plate-5 Plate-6
Plate-7 Plate-8
Plate-9 Plate-10
Fig. 6. Antibacterial activity of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine
compounds -petri dishes.
World Scientific News 8 (2015) 211-226
-222-
0
2
4
6
8
10
12
14
zon
e of
inh
ibit
ion
(m
m)
substituents
Antibacterial Activity
Bacillus
M. luteus
S. aureus
E. coli
P. aurogenosa
Fig. 7. The antibacterial activities of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine
compounds -clustered column chart.
3. 4. 2. Antifungal activities
The antifungal activities of all the synthesized heterocyclic Schiff base compounds have
been studied against Aspergillus niger, Mucour species and Tricoderma viride using [23] disc
diffusion technique. PDA medium was prepared and sterilized as above. It has been poured
(ear bearing heating condition) in the Petri-plate which has been already filled with 1 ml of
the fungal species. The plates have been rotated clockwise and counter clock-wise for uniform
spreading of the species. The discs have been impregnated with the test solution. The test
solution has been prepared by dissolving 15mg of the Schiff base compounds in 1ml of
DMSO solvent. The medium have been allowed to solidify and kept for 24 hours. Then the
plates have been visually examined and the diameter values of zone of inhibition have been
measured. Triplicate results have been recorded by repeating the same procedure.
The antifungal activities of substituted Schiff base have been studied and are shown in
(Fig. 8 for Plates (1-4) and the zone of inhibition values of the effect is given in Table 4. The
clustered column chart, shown in (Fig. 9). A good antifungal activity has been possessed by
all substituents on the microorganisms in general. The substituents 4-F, 3-NO2 and 4-NO2
have shown very good fungal activity against A. niger. All the compounds have shown
moderate antifungal activity against the fungal species Peniciliumscup.
1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
World Scientific News 8 (2015) 211-226
-223-
sTable 4. Antifungal activity of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds.
S.No. Substituents Zone of inhibition(mm)
A. niger M. species T. species
1 H 5 8 6
2 3-Br 6 8 7
3 4-Br 7 5 8
4 3-Cl 6 6 7
5 4-Cl 6 5 6
6 4-F 9 8 7
7 4-CH3 6 9 7
8 4-OCH3 8 8 6
9 3-NO2 12 9 9
10 4-NO2 10 7 5
Standard Ciprofloxacin 8 10 6
Control DMSO 0 0 0
Fig. 8. Antifungal activity of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine
compounds -petri dishes.
Plate-1 Plate-2
Plate-3 Plate-4
Plate-5 Plate-6
World Scientific News 8 (2015) 211-226
-224-
0
2
4
6
8
10
12
14
Zon
e o
f in
hib
itio
n (
mm
)
substituents
Antifungal Activity
A. niger
M. species
T. viride
Fig. 9. Antifungal activity of substituted (E)-N-benzylidene-4H-1,2,4-triazol-4-amine compounds -
clustered column chart.
4. CONCLUSIONS
A series of ten number of heterocyclic Schiff base compounds viz., substituted (E)-N-
benzylidene-4H-1,2,4-triazol-4-amine compounds have been synthesized by condensation
method. These compounds were confirmed by their physical constants and spectral data. The
spectral data of these compounds have been correlated with Hammett sigma constants and F
and R parameters using single and multi-linear regression analysis. From the results of
statistical analysis, most of the correlations were found to be satisfactory. The antimicrobial
activities of these heterocyclic Schiff base compounds were found to be moderate to good
activity.
ACKNOWLEDGEMENT
The authors thank DST NMR Facility, Department of Chemistry, Annamalai University, Annamalainagar - 608
002, for recording NMR spectra of all compounds.
References
[1] Schiff H., Justus Liebigs Annalen der Chemie. 131(1) (1864) 118-9.
[2] Lau K. Y., Mayr A., Cheung K. K., Inorgnic Chimica Acta. 285 (1999) 223-232.
[3] Shawali A. S., Harb N. M. S., Badahdah K. O., Journal of Heterocyclic Chemistry. 22
(1985) 1397-1403.
[4] Gupta K. C., Sutar A. K., Coordination Chemistry Review. 252 (2008) 1420-1450.
1. H
2. 3-Br
3. 4-Br
4. 3-Cl
5. 4-Cl
6. 4-F
7. 4-CH3
8. 4-OCH3
9. 3-NO2
10. 4-NO2
World Scientific News 8 (2015) 211-226
-225-
[5] Yuan M., Zhao F., Zhang W., Wang Z. M., Gao S., Inorgnic Chemistry. 46 (2007)
11235-42.
[6] Nakaic T., Meddu S., Kurahashi T., Japan Patent. 7389932 (1973); Chemical
Abstracts. 81(1974) 65182.
[7] Quraishi Harion M. A., Sharma K., Journal of Materials Chemistry and Physics.
78 (2002) 18-21.
[8] Ramesh S., Rajeshwari S., Elctrochimica Acta. 49 (2004) 811-820.
[9] Colter R. J., Matzner M., Ring terming poly, Part B-1, ‘Heterocyclic Ring’ Academic,
New Youk, (1972).
[10] Popp. F. D., Journal of Organic Chemistry. 26 (1961) 1566-1568.
[11] Rao X., Huang X., He L., Song J., Song Z., Shang S., Combinatorial Chemistry &
High Throughput Screening. 15 (2012) 840-844.
[12] Hadjipavlou-litina D. J., Geronikaki A. A., Letters in Drug Design & Discovery. 15
(1996) 199-206.
[13] Tiwary M., Naik S. N., Tiwari D. K., Mittal P. K., Yadav S., Journal of vector Brone
Diseases. 44 (2007) 198-204.
[14] Solak N., Rollas S., Arkivoc. (2006) 173-181.
[15] Wadher S. J., Puranik M. P., Karande N. A., Yeole P. G., International Journal of
Pharm Tech Research 1 (2009) 22-33.
[16] Cates A. L., Rasheed S. M., Pharmaceutical Research 6 (1984) 271-273.
[17] Sakthinathan S. P., Suresh R., Mala V., Sathiyamoorthi K.,
Kamalakkannan D., Ranganathan K., John Joseph S., Vanangamudi G.,
Thirunarayanan G., International Journal of Scientific Research and Knowledge,
1(11) (2013) 472.
[18] Sakthinathan S. P., Suresh R., Mala V., Sathiyamoorthi K.,
Kamalakkannan D., Ranganathan K., Arulkumaran R., Vijayakumar S., Sundararajan
R., Vanangamudi G., Thirunarayanan G., International Letters of Chemistry, Physics
and Astronomy. 6 (2013) 77.
[19] Suresh R., Kamalakkannan D., Ranganathan K., Arulkumaran R.,
Sundararajan R., Sakthinathan S. P., Vijayakumar S., Sathiyamoorthi K.,
Mala V., Vanangamudi G., Thirumurthy K., Mayavel P., Thirunarayanan G.,
Spectrochim. Acta, 101A (2013) 239.
[20] Arulkumaran R., Vijayakumar S., Sakthinathan S.P., Kamalakkannan D., Ranganathan
K., Suresh R., Sundararajan R., Vanangamudi G., Thirunarayanan G., Journal of
Chilean Chemical Society. 2 (2013) 58.
[21] Thirunarayanan G., Gopalakrishnan M., Vanangamudi G., Spectrochemica Acta 67A
(2007)1106-1112
[22] Swain C. G., Lupton E. C., Journal of American Chemical Society.
90 (1968) 4328-4337.