Synthesis, structure investigation, spectral characteristics and biological activities of some novel azodyes

Article abstract of DOI:10.1016/j.saa.2010.12.037

Four novel azo compounds were synthesized; o-phenylazo- (C ₁₄H₁₃N₃O₂) (I), p-bromo-o-phenylazo- (C₁₄H₁₃BrN₃O₂) (II), p-methoxy-o-phenaylazo- (C₁₅H₁₆

Full text of DOI:10.1016/j.saa.2010.12.037

Spectrochimica Acta Part A 78 (2011) 1027–1036
Contents lists available at ScienceDirect
Spectrochimica Acta Part A: Molecular and
Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa
Synthesis, structure investigation, spectral characteristics and biological
activities of some novel azodyes
M.A. Zayed , Gehad G. Mohamed , S.A.M. Abdullahb
a,∗
a
a
Chemistry Department, Faculty of Science, Cairo University, 12613 Giza, Egypt
University of Workers,10 Aswan, Egypt
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 17 September 2010
Received in revised form
Four novel azo compounds were synthesized; o-phenylazo- (C14H13N O ) (I), p-bromo-o-phenylazo-
3
2
(C14H13BrN3O2) (II), p-methoxy-o-phenaylazo- (C15H16N3O3) (III) and p-nitro-o-phenylazo-p-
acetamidophenol (C14H13N4O4) (IV). These compounds were carefully investigated using elemental
2
5 November 2010
1
analyses, UV–vis, FT-IR, H NMR and mass spectra. Also, the effects of p-substituents such as bromo,
Accepted 10 December 2010
methoxy and nitro groups on the mass fragmentation pathways of these dyes were studied using
Hammet’s effects. This research aimed chiefly to threw lights on the structures–stability relationship
of four novel newly prepared azo derivatives of p-acetoamidophenol. The data obtained referred to
the variation of mass fragmentation pathways with the variation of p-substituent of these dyes which
can be used in industry for various dyeing purposes. This variation is also correlated and verified by
molecular orbital calculations which were done on ionic forms of these dyes using semi empirical PM3
program. The biological activities of these dyes were also investigated and its structure relationship was
correlated.
Keywords:
Acetoamidophenolazo compounds
UV–vis
FT-IR
1
H NMR
Mass spectra
MO calculations
Biological activity
©
2010 Elsevier B.V. All rights reserved.
1
. Introduction
yields of diarylethenes [12,13]. The potential of organic dyes and
their intermediates to adversely impact human health and the envi-
ronment has moved toxicological considerations to the forefront of
their molecular design [14].
Mass spectrometry plays pivotal role in the structural char-
acterization of organic molecules [15]. In conjunction with mass
spectrometric analysis [16,17], computational quantum chemistry
can provide additional information about the atoms and bonds,
which can be used successfully in an interpretation of experimental
results [18].
Recently, several studies have been published on the syn-
thesis and spectral properties of azo dyes [1–3]. This reflects
their widely important applications in different fields, such as
polyester fiber [4] and disperses dyes [5], as well as their involve-
ment in many biological reactions and in analytical chemistry
[
6].
Photochromic compounds are receiving more interest due to
their potential applications in photonic devices such as optical
memories, photo-switches, and full color displays [7–9]. The pho-
tochromic properties of the some dyes depend on several factors,
such as the nature of the heteroaryl moieties, the conformation
of the open-ring isomer, electron donor/acceptor substituents, and
the p-conjugation length of the hetero-aryl groups, among others
The aim of the present work is to carry out experimental and
theoretical investigation of the four azo compounds of the biologi-
1
cal activity of p-acetamidophenol using UV–vis, FT-IR, H NMR and
EI mass spectral (MS) fragmentation at 70 eV. Also, MO calculations
are performed using PM3 procedure, on the charged molecular
ion to investigate bond length, bond order, heats of formation,
ionization energy and charge distribution. These calculations are
correlated with the EI-MS experimental results to obtain informa-
tion about the stability of the studied compounds and prediction
of the site of primary fragmentation step and subsequent ones.
Also, it is used to discuss the effect of Hammet values of sub-
stituents on experimental and computational results. The biological
activities of the azodyes under investigation are reported using Tri-
bolium confusum. The structures of the azodyes (I–IV) are given in
Fig. 1.
[
10]. Electron-donating substituents can increase the absorption
coefficients and decrease the cycloreversion quantum yields of
some dyes, while electron-withdrawing groups do not affect the
cycloreversion quantum yield but can shift the absorption maxima
to longer wavelengths [11]. The longer the p-conjugation length
of the hetero-aryl groups, the lower the cycloreversion quantum
∗ Corresponding author. Tel.: +20 105776675; fax: +2 002 35728843/7556.
E-mail address: mazayed429@yahoo.com (M.A. Zayed).
1
386-1425/$ – see front matter © 2010 Elsevier B.V. All rights reserved.
doi:10.1016/j.saa.2010.12.037

1
028
M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
was used in this study. The EI-MS spectra were obtained at ionizing
energy value of 70 eV, ionization current of 60 ␮A and vacuum is
−
6
better that 10 Torr.
The molecular orbital calculations (MOCs) were performed
using semi-empirical molecular orbital calculation. The method
used in these computations is the parametric method (PM3)
described by Stewart [20]. The default criteria for terminating all
optimizations were increased by a factor of 100 (keyword PRECISE).
Vibrational frequencies were computed for the studied structures
(keyword FORCE) so as to check whether the newly designed
geometries are local minima. All the molecular orbital calcula-
tions were carried out at the unrestricted Hartree–Fock level (UHF)
for there positively charged ions using PM-3 method followed by
full optimization of all geometrical variables (bond lengths, bond
angles, and dihedral angles), without any symmetry constraint. All
structures were optimized to a gradient normalization of 0.01–0.05,
using the eigenvector following (EF) routine [21]. All the semi
empirical MO calculations were performed with the MOPAC2000
software package [22] implemented on an Intel Pentium IV 3.0 G
Hz computer.
Fig. 1. Structure of azodyes (I–IV).
2
. Experimental
2.5. Biological activity
2.1. Reagents
Adult of T. confusum were laboratory reared on wheat flour at
All the chemicals used in this work were of analytical grade.
◦
2
7.5 ± 1.5 C and 70% ± 5% (R.H.) according to the method of Fred-
They included p-acetamidophenol, aniline, p-bromoaniline, p-
methoxyaniline, and p-nitroaniline (Sigma) sodium nitrite and
sodium hydroxide (Adwik). Organic solvents (spectroscopic pure
from BDH) used included absolute ethyl alcohol, diethylether and
dichloromethane.
eric et al. [23] with some modifications.
T. confusum adult was topically treated with 10 ␮m of each
compound according to the protocol described by Delobel et al.
24] as follows: thirty insects divided on three replicates (10
adult/replicate) were topically and mortality was then monitored
after 24 h. Thirty adults of control experiment were used in three
replicates without treatment. The adult mortality was estimated
according to Abbot Method [25]. Estimation of LD50 values was
made using probity analysis as given by Finney [26].
[
2
.2. Preparation of dyes (I–IV)
p-Acetamidophenol-azo-derivatives (I–IV) were prepared by
coupling p-acetamidophenol with aryl, p-bromo-, p-methoxy- and
p-nitro-diazonium chloride, in an ice bath, in the presence of
sodium hydroxide [19]. The precipitates were left in refrigerator
over night, filtered and crystallized from acetic acid (yield 78–86%).
3
. Results and discussion
The structure elucidation of the prepared azodyes (I–IV) is car-
2
.3. Solutions
ried out by using different techniques namely elemental analyses,
UV–vis, FT-IR and 1H NMR and mass spectroscopy, in addition to
molecular orbital calculations.
The solutions used in UV–vis. measurements are obtained
by dissolving the accurate weight of the corresponding dye in
ethanol or in methylene chloride. The concentrations of the stud-
The elemental analyses of the prepared dyes referred to the
general formulae of o-phenylazo- (C14H13N O ) (I), p-bromo-
3
2
−3
−3
−3
ied dyes are 0.1574 × 10 M, 0.126 × 10 M, 0.143 × 10 M and
o-phenylazo- (C14H13BrN O ) (II), p-methoxy-o-phenaylazo-
3 2
−2
0
.137 × 10 M for dyes I, II, III and IV, respectively.
(C15H16N O ) (III) and p-nitro-o-phenylazo-p-acetamidophenol
3 3
(
C14H13N O ) (IV) compounds, respectively.
4 4
2.4. Instruments
3.1. UV–vis absorption spectra
Elemental microanalyses of the separated solid dyes for C,
H and N were performed at the Microanalytical Center, Cairo
University, using CHNS-932 (LECO) Vario Elemental Analyzers.
The UV–vis electronic spectra of dyes (I–IV) were measured in
methanol and in methylene chloride (MC) solvents using Shimadzu
recording spectrophotometer UV/VIS/NIR 3101 PC model in the
region of ꢀ = 200–800 nm. The infrared spectra were recorded on
a PerkinElmer FT-IR type 1650 spectrophotometer in wave num-
These structures are confirmed by the UV–visible spectra in
ethanol (Fig. 2a) and in methylene chloride (Fig. 2b). The correlation
between the structural formulae of these dyes and the absorption
spectral data is given in Table 1. The electronic transition n–␲*
3
−1
−1
(
ε ≈ 10 L mol cm , ꢀmax ≈ 330–439 nm) generally occurs in
amide carbonyl in dyes I–III together with that occurs in NO group
2
4
−1
−1
,
−1
in dye IV and ␲–␲* electronic transition (ε ≈ 10 L mol cm
ber region 4000–400 cm . The spectra were recorded as KBr
pellets. 1H NMR spectra were measured using an instrument of
ꢀ
≈ 230–260 nm) occurs in azo group/and or other unsaturated
max
bonds in all dyes (I–IV). These transitions in ethanol are of less molar
absorptivity than in methylene chloride for all dyes. This is may be
attributed to the effect of polarity difference of the two solvents.
The position of their bands (Fig. 2) is varied from one dye to the
other and from one solvent to another which may be attributed to
the p-phenylazo substituent variable donating power.
Model Gemini 2000 Switzerland using duterated dimethylsulphox-
ide (DMSO-d ) and measured in Micro Analytical Center, Cairo
6
University. Electron ionization mass spectra (EI-MS) of the studied
dyes were obtained using Shimadzu GC-MS-Qp 1000 PX quadru-
ple mass spectrometer with electron multiplier detector equipped
with GC–MS data system. The direct probe (DP) for solid material

M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
1029
Fig. 2. (a) The absorption spectra of dyes I–IV in ethonal. (b) The absorption spectra of dyes I–IV in methylene chloride.
3.3. 1H NMR spectra
3.2. Infrared spectral studies
The 1H MNR spectra of these dyes (I–IV) are given in Table 3.
These 1H NMR data are correlated with the proposed structures
of these dyes. In dye I (Fig. 4), the chemical shifts appeared at
ı = 1.700–2.502, 3.416–4.000, and 7.056–7.707 are attributed to
FT-IR spectra (Fig. 3) of these dyes are correlated with their pro-
posed structural formulae (Table 2). These data refer to assignment
of main bands of the active groups in the moiety of these dyes,
such as 510–618, 1173–1310, 1419–1595, 1657, 3028–3056 and
−1
3
284–3560 cm for ␯(C–N) amide, ␯(C–O) amide, ␯(CH ) amide,
protons of CH , NH and OH of acetamidophenol, aromatic, pro-
3
3
␯
(N N azo), ␯(CO) amide, ␯(NH amide) and ␯(OH phenolic) in dyes
tons of phenylazo group, and aromatic protons of phenyl group
of acetamidophenol, respectively. The main proton chemical shifts
of other dyes (II–IV) appeared at different values depending upon
the donating power effect of p-substituent of p-phenylazogroups.
These ı values are appeared in case of dye II, at 7.606–7.767 of aro-
matic protons of p-Br-phenylazo group; and in case of dye III, at
(
I–IV), respectively. The positions of these bands are varied from
one dye to another due to the variation of donating power effect
of p-substituent of phenyl azo group. Extra bands are appeared at
5
−
1
−
1
15–585 cm for ␯C–Br in case of dye II and 832 cm of ␯NO2 in
case of dye IV.
Table 1
The UV–vis spectral data description of a – 2 ml of 0.01 g/ml ethanol, b – 2 ml of 0.01 g/ml methylene chloride of some acetoamidophenolazo derivatives (I–IV).
−
1
−1
Compound
ꢀmax (nm)
Absorbance
ε (L mol cm )
Electronic transition
I, 0.1574 × 10−
3
M
a – 333.5
a-0.9214
2.4440
5.853 × 10
3
n–␲* in CO group
␲–␲* in azo group
4
260.5
1.552 × 10
3
b – 333.4
b – 0.5460
1.1659
3.46 × 10
n–␲* in CO group
␲–␲* in azo group
4
231.0
0.74 × 10
II, 0.126 × 10−
3
M
a – 338.5
51.0
a – 1.1135
2.0793
8.83 × 10
3
n–␲* in CO group
␲–␲* in azo group
4
2
1.65 × 10
3
b – 340.5
38.5
b – 0.8574
1.3026
6.8 × 10
n–␲* in CO group
␲–␲* in azo group
4
2
1.033 × 10
III, 0.143 × 10−
3
M
M
a – 363.0
56.5
a – 0.8537
3.4323
5.97 × 10
3
n–␲* in CO group
␲–␲* in azo group
4
2
2.40 × 10
3
n- ␲* in CO group
␲–␲* in azo group
b – 355.0
37.0
b – 1.0859
2.5099
7.59 × 10
4
2
1.75 × 10
IV, 0.137 × 10−
2
a – 420.5
a – 0.5730
1.6976
1.8818
4.18 × 10
3
n–␲* in CO group
␲–␲* in azo group
␲–␲* in NO2 group
4
3
2
34.5
50.5
1.23 × 10
4
1.37 × 10
3
b – 439.5
b – 0.2860
0.9830
1.1261
2.08 × 10
n–␲* in CO group
␲–␲* in azo group
n–␲* in NO2 group
4
337.0
30.5
0.717 × 10
4
2
0.821 × 10
I—o-phenylazo-pacetamedophenol,
acetamedophenol.
II—p-bromo-o-phenylazo-p-acetamedophenol,
III—p-methoxy-o-phenylazo-p-acetamedophenol,
IV—p-nitro-o-phenylazo-p-

1
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M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
Fig. 3. The FT-IR spectra of dye I.
Fig. 4. The 1H NMR spectra of dye I.
1
.674–2.502 of CH3 protons of aliphatic CH O; in case of dye IV at
3. Selection of the most probable decomposition pathways in vitro
3
8
.182–8.43 of aromatic protons of p-NO -phenylazo group.
system using MS technique.
2
4
. Substituent effects on experimental and computational results.
3.4. Mass spectral fragmentation
Electron ionization (EI) mass spectra of the four dyes (I–IV)
at 70 eV were recorded and investigated (e.g. of dye I, Fig. 5).
The proposed of principal and important fragmentation pathways
following electron impact is shown in Schemes 1–4. The signal
Knowledge of the decomposition mechanisms of these dyes
is very important to understand its chemical structure–reactivity
relationship. It is difficult to establish the exact major fragmenta-
tion pathway in EI using conventional MS. However, combination
of the experimental techniques (UV–vis, FT-IR, 1H NMR and MS)
and MO calculations is very important to understand the following
topics:
1
2
. Primary site fragmentation process and its major fragmentation
pathway.
. Stability of the studied dyes as positively charged molecules in
solid state phase and molecular ions in gas phase.
Fig. 5. EI-MS of dye I.

M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
1031
Scheme 1. Mass spectral fragmentation pathways of dye I.
appeared in the mass spectra of dyes I, II, III and IV at m/z = 255,
33, 286 and 300 are due to the formation of molecular ions,
general formula and agrees well with their elemental analyses. In
case of dye I (Scheme 1) mass spectra (Fig. 5) refer to the appear-
ance of mole fragments in three pathways. The signals appeared at
3
+
+
+
(
C14H13N O , RI = 25%), (C14H13N O Br , RI = 24%), (C15H16N O
,
3
2
3
2
3
3
+
RI = 20%) and (C14H13N O , RI = 15%) for these dyes, respectively.
m/z = 58 (RI = 50%) due to fragment ion HCOCH , 77 (RI = 100%, base
4
4
3
The appearance of these molecular ions confirms their proposed
peak) due to fragment ion C H5 and the fragment ions containing
6
+
+
OH
OH
N
N
N
N
-
COCH 2
m/z = 253, R.I.= 1 %
NHCOCH 3
m/z = 211
R.I. = 16 %
Path 1
-
HBr
NH2
+
+
OH
OH
Br
COCH 2
N
N
Br
N
N
Path 2
-
m/z = 291, R.I. = 23.8 %
NHCOCH 3
NH2
OH
C -N
9
OH
m/z = 333, R.I. = 23.8%
3
Cleavage
+
C14H13N O Br
+
2
2
+
Path 3
-
COCH 2
NH2
m/z =108, R.I. = 9.6 %
NHCOCH 3
m/z = 150, R.I. = 100 %
Scheme 2. The mass spectral fragmentation pathways of the dye II.

1
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M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
Scheme 3. Mass fragmentation scheme of dye III.
azo group appeared at m/z = 105 (RI = 60%) due to C H5–N NH, and
one fragment ion is the only one containing p-nitro-azo group. The
lower RI% values may refer to the instability of p-nitro-azodye IV
in a similar way like p-bromoazodye II. This means that the elec-
tron withdrawing substituents are weakening the azodye stability.
This also confirmed by its absorption spectra at longer wavelengths
than the main azo dye I. Therefore, we may order the stability of
these dye during ionization with 70 eV electron beam as dye I > dye
III > dye II > dye IV. This is agrees well with the Hammet values
in relation to some MOCs parameters such as heat of formation,
dipole-moment, electron affinity and ionization potential (Table 4).
6
at 165 (RI = 60%) due to CH CO–C H (OH)–N NH. The appearance
3
6
4
of the fragment ions containing azo group of high RI of 60% refer
to the stability of azodye I even at high energy ionization source
7
0 eV. In case of dye II (Scheme 2) its mass spectra show fragmen-
tation of this dye in three parallel pathways. It shows in path I,
fragment ions at m/z = 253 (RI = 1%) after the loss of HBr from the
molecular ion; at m/z = 211 (RI = 16%) due fragment ion contain-
ing azo group; at m/z = 291 (RI = 30%) in path containing also azo
group; and at m/z = 150 (RI = 100%; base peak) containing no azo
group. The low RI values of fragment ions containing azo group may
refer to the low stability of p-bromoazodye II on its ionization with
the high energy electron beam of 70 eV. In case of p-methoxy azo
dye III mass, its fragmentation occurs in three pathways as given
by Scheme 3. It shows three fragment ions containing azo group of
m/z = 245 (RI = 30) and m/z = 105 (RI = 23%) in path 1; m/z = 226 (25%)
in both paths 2 and 3. The middle values of RI % of these fragment
ions containing azo group may refer to the middle stability of p-
methoxyazo dye III. It is finally decomposed to give the fragment ion
3.5. MO calculations
Molecular orbital calculations (Tables 4–8) give valuable infor-
mation about the structure and the reactivity of a molecule and
its molecular ions. Computational data can be used to support the
experimental data. The much import parameters calculated using
MO calculation are geometries, bond order, bond strain, charge
distribution, heat of formation and ionization energy. Investigation
of the molecular structure of four dyes I–IV was interest in the
present work aiming to help in illumination of experimental data
of m/z = 151 (RI = 60%) of p-acetamidophenol parent compound and
+
m/z = 77 (RI = 100%) as a base peak of C H5 stable fragment ion. The
6
EI-MS of p-nitroazodye IV, are explained in Scheme 4. It shows three
fragmentation pathways. This scheme shows three fragment ions
containing azo groups. These are at m/z = 254 (RI = 1%), at m/z = 212
(i.e. prediction of the weakest bond cleavage and the stability
of the cationic forms of these dyes). The weakness of the bond
is indicated by its longer bond length, by the less value of the
bond order and the high value of bond strain and vise versa.
(
RI = 28%) in path 1 and at m/z = 258 (RI = 12.5%) in path 2. The last

M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
1033
+
+
OH
OH
N
N
N
N
-
COCH2
m/z = 254, R.I .< 1 %
m/z = 212
R.I. = 2.8 %
Path 1
-
NO2
NH2
NHCOCH3
+
+
OH
OH
NO2
N
N
N
N
NO2
Path 2
-COCH2
m/z = 258, R.I. = 12.5 %
NHCOCH3
NH2
OH
C -N
9
OH
m/z = 300, R.I. = 15.3 %
3
Cleavage
+
C14H13N O
+
4
4
+
Path 3
-
COCH2
NH2
m/z =108, R.I. = 81.3 %
NHCOCH3
m/z = 150, R.I. = 21%
Scheme 4. The mass spectral fragmentation pathways of the dye IV.
Table 3
Fig. 6 shows the numbering system of the dyes I–IV skeleton that
helps in ordering the calculated bond order and charge distribu-
tion (of different atoms) of the charged species. The calculated
geometrical parameters are shown in Tables 4–8. From the calcu-
lated geometries (dipole moment, heat of formation, ionization
potential, Table 4) and (bond length, and bond order) for dye I
The 1H NMR spectral data description of some p-acetoamidophenolazo derivatives
(
I–IV).
Compound
I
Chemical shift values, ꢁ
Description
1.700–2.502
Protons of CH3 of
acetamidophenol
3.416–4.000
7.056–7.366
7.357–7.707
Protons of NH and OH of
acetamidophenol
Aromatic protons of phenylazo
group
Aromatic protons of phenyl
group of acetamidophenol
(
Table 5) it is clear that the order of increasing stability of bonds
is:
C16–C17 < N11–C16 < C3–N9 < N13–C19 < C5–N11 < N9–N13.
Table 2
The FT-IR spectral data description of acetoamidophenolazo derivatives (I–IV).
−
Compound
I
Wave number (cm 1
)
Description
II
2.024–2.497
.00–4.00
CH3 protons of
acetamidophenol group
Protons of NH and OH groups
of acetamidophenol
510–618
␯C–N amid, C–O amid
␯CH3 amid
␯N N azo
␯CO amid
␯OH phenolic
␯NH amid
3
1
1
1
3
3
173–1310
419–1595
657
028–3056
284–3560
7.606–7.767
Aromatic protons of
p-Br-phenylazo group
Aromatic protons of phenyl
group of acetamidophenol
9.900–10.300
II
515–585
007–1068
420–1664
663
087
284–3684
␯C–Br, ␯C–O amid
␯CH3 amid
III
1.674–2.502
CH3 protons of aliphatic CH3O
CH3 protons of acetamido
group
Protons of phenolic and of NH
amid group
Protons of substituted
phenylazo group
Aromatic protons of
acetamidophenol
1
1
1
3
3
3.372–3.904
6.672–7.175
7.197–8.038
␯N
N
␯CO amid
␯OH phenolic
␯NH amid
III
IV
511–585
105–1203
426–1666
666
108
321–3648
␯C–N amid, ␯C–O
␯CH3 amid
1
1
1
3
3
9.903–10.303
2.035–2.500
␯N
N
␯CO amid
␯OH phenolic
␯NH amid
IV
CH3 protons of acetamido
group
Protons of NH and OH of
acetamidophenol
Aromatic protons of
p-NO2-phenylazo group
Aromatic protons of
acetamidophenol
7.048–8.058
522–640
␯C–O amid, ␯N–O
␯NO2
␯CH3 amid
␯CO amid
␯N N
␯OH phenolic
␯NH amid
832
8.182–8.436
1
1
1
2
3
176–1293
660
439–1660
835–3069
291–3600
9.955–10.639
I—o-phenylazo-pacetamedophenol,
acetamedophenol,
IV—p-nitro-o-phenylazo-p-acetamedophenol.
II—p-bromo-o-phenylazo-p-
III—p-methoxy-o-phenylazo-p-acetamedophenol,
I—o-phenylazo-pacetamedophenol,
acetamedophenol,
IV—p-nitro-o-phenylazo-p-acetamedophenol.
II—p-bromo-o-phenylazo-p-
III—p-methoxy-o-phenylazo-p-acetamedophenol,

1
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M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
Table 4
The MOCs parameters and Hammet’s subtituents values of dyes I–IV.
The dye no. and state
Heat of formation (kcal/mol)
Dipole moment (Debye)
Electron affinity (eV)
Ionization potential (eV)
Hammet constant
I, p-H Cationic state
II, p-Br Cationic state
III, p-OCH3Cationic state
IV, p-NO2 Cationic state
184.37137
193.633
144.964
181.990
6.160
15.942
8.707
4.421
4.482
4.333
4.705
12.71
12.78
12.62
13.03
0.01
0.23
−0.27
17.920
0.78
This means that the first cleavage bond is C16–C17 fol-
lowed by N11–C16, C3–N9 and N13–C19. The most stable
bonds are azo group (N9–N13) and its neighbor bond C5–N11.
The calculated geometries of the cationic form of dye II
m/z = 150 (RI = 24%) of p-acetamidophenol parent, followed by the
rupture of C16–N11, in path 1. In path 2 it starts its fragmentation
with C19–N13 and C3–N9 bonds cleavage leading to the forma-
tion of fragment ion of m/z = 165 (RI = 60%) containing azo group.
Also path 3 refer to the cleavage of bond C19–N13 to give the
most abundant fragment of m/z = 77 (RI = 100%). The geometries
also refer to the highest stable bond is the azogoup. This conclu-
sion agree well with mass, UV–vis, and IR data, that dye I is the
most stable dye (heat of formation = 184.37 kcal/mol). Dye II in path
1 also starts its mass fragmentation by the weakest bond C22–Br25
leading to phenylazoacetamidophenol of m/z = 253 (RI = 1%), fol-
lowed by the rupture of the bond C16–N11 to form a fragment
(
Table 6) refer to the order of increasing stability of bonds is:
C22–Br25 < C16–C17 < N11–C16 < C3–N9 < N13–C19 < C5–N11 <
N9–N13. This means that the first cleavage bond is C22–Br25,
C16–C17 followed by N11–C16, C3–N9 and N13–C19.
The most stable bonds are azo group (N9–N13) and its
neighbor bond C5–N11. Also there is highly stable bond
C16–O18. In case of dye III, the geometries (Table 7) indi-
cate that, the order of increasing stability of bonds is:
N11–C16 < C16–C17 < O25–C26 < C3–N9 < N13–C19 < C5–N11 <
N9–N13. This means that the first cleavage bond is N11–C16
and C16–C17 followed by O25–C26, C3–N9 and N13–C19.
The most stable bonds are azo group (N9–N13) and its
neighbor bond C5–N11. Also there is another stable bond
C16–O18. The molecular orbital geometries of dye IV (Table 8)
indicate that, the order of increasing stability of bonds is:
C22–N25 < N11–C16 < C16–C17 < N13–C19 < C3–N9 < N9–N13.
This means that the first cleavage bond is C22–N25 followed by
N11–C16, C16–C17, C3–N9 and N13–C19. The most stable bonds
are azo group (N9–N13) and its neighbor bond C5–N11. Also there
are other stable bonds C16–O18, N25–O38 and N25–O37 of nitro
group.
Table 6
The calculated geometries of dye II in cationic state.
Dye-II-cation, p-Br
Atom Atom ID Partial charge Bond
Bond length (Å) Bond order
O8
N9
8
9
−0.128
−0.014
0.196
0.075
0.252
−0.143
−0.276
−0.138
0.060
C2–O8
C3–N9
1.326
1.443
1.374
1.229
1.471
1.498
1.211
1.437
1.405
1.863
1.229
1.031
1.307
1.903
0.944
0.959
1.901
1.034
1.367
0.984
N11
N13
C16
C17
O18
C19
Br25
H30
11
13
16
17
18
19
25
30
C5–N11
N9–N13
N11–C16
C16–C17
C16–O18
N13–C19
C19–C24
C22–Br25
0.086
3.6. Comparison between EI-MS and MO calculations
The scope of the present investigation is restricted to a search
Table 7
The calculated geometries of dye III in cationic state.
or prediction and discerns features of initial bond ruptures dur-
ing the course of fragmentation of dyes (I–IV) molecules. Empirical
observations indicate that the course of subsequent fragmentation
is determined to large extent by the initial bond ruptures of the
molecular ion in MS [26]. It is quite acceptable to say that the com-
putational quantum chemistry can provide additional data which
can be used successfully to interpret MS experimental results.
These theoretical data can, particularly, be valuable for mass spec-
trometry scientists; in which the gas-phase species can be handled
much more easily by quantum chemistry than those surrounded
by solvent [27].
Dye-III-cation (p-methoxy)
Atom Atom ID Partial charge Bond
Bond length (Å) Bond order
O8
N9
8
9
−0.128
−0.032
0.188
0.097
0.252
−0.143
−0.280
−0.196
0.047
C2–O8
C3–N9
1.326
1.439
1.226
1.040
1.295
1.875
0.950
0.959
1.895
1.054
1.081
0.979
N11
N13
C16
C17
O18
C19
C26
H27
11
13
16
17
18
19
26
27
C5–N11 1.376
N9–N13 1.232
N11–C16 1.469
C16–C17 1.499
C16–O18 1.211
N13–C19 1.429
C22–O25 1.369
O25–C26 1.410
Comparing computational data and the previously EI-MS of dyes
I–IV indicate that, dye I fragmentation starts with the cleavage of
C3–N9 to give m/z = 105 (RI = 60%) containing phenylazo group and
0.036
Table 8
The calculated geometries of dye IV in cationic state.
Table 5
Dye-IV cat, p-nitro
The calculated geometries of dye I in cationic state.
Atom Atom ID Partial charge Bond
Bond length (Å) Bond order
Dye I, cation, p-H
N9
9
11
13
16
17
18
19
30
31
32
0.003
0.205
0.057
C3–N9
1.446
1.023
1.318
1.923
0.938
0.959
1.905
1.019
0.879
1.518
1.517
Atom Atom ID Partial charge Bond
Bond length (Å) Bond order
N11
N13
C16
C17
O18
C19
H30
H31
H32
C5–N11 1.373
N9–N13 1.228
N11–C16 1.474
C16–C17 1.498
C16–O18 1.210
N13–C19 1.443
C22–N25 1.512
N25–O37 1.212
N25–O38 1.212
O8
N9
8
9
−0.128
−0.016
0.193
C2–O8
C3–N9
1.326
1.442
1.229
1.035
1.303
1.898
0.947
0.959
1.899
1.037
0.252
N11
N13
C16
C17
O18
C19
11
13
16
17
18
19
C5–N11 1.375
N9–N13 1.230
N11–C16 1.471
C16–C17 1.499
C16–O18 1.211
N13–C19 1.435
−0.143
−0.271
−0.082
0.086
0.087
0.107
0.081
0.252
−0.143
−0.278
−0.145

M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
1035
Fig. 6. Numbering systems of dyes I–IV.
of m/z = 211 (RI = 16%) containing azo group. In path 2, dye II starts
its decomposition by the rupture of C16–N11 to form a fragment
ion containing azo group of m/z = 291 (RI = 21.8%). In path 3, dye
II gives the most abundant p-acetamidophenol parent fragment of
m/z = 150 (RI = 100%) via the rupture of the C3–N9 bond and this is
followed by the rupture of the bond C16–N11 to form a fragment ion
of m/z = 108 (RI = 9.6%) of p-aminophenol. The computational data
also refer to the high stability of the azo bond N9–N13 (heat of for-
mation of dye II = 193.63 kcal/mol), but unfortunately the presence
of Br substituent facilitates the azo group decomposition in path
ment ion of dye I of m/z = 254 (RI = 1%), it follows with the rupture
of the bond C16–N11 leading to the formation of fragment ion still
containing azo group of m/z = 212 (RI = 28%). In path 2, the fragment
ion containing azo group of m/z = 258 (RI = 12.5%) is formed via the
rupture of C16–N11, keeping the NO2 in its entity. Therefore, the
presence of NO2 decreases abundance of this azo fragment ion if
compared with that of m/z = 212 fragment ion free from this group.
In path 3, dye IV gives the parent p-acetoamidophenol fragment ion
of m/z = 150 (RI = 21%) by the loss of p-nitro-phenylazo group and
cleavage of the weak bond C3–N9. Therefore, we can conclude that
3
. It also makes the dye II less stable than dye I. Dye III in path 1
the presence of NO decreases abundance of azo fragment ions and
2
starts its decomposition by the rupture of the weak bond C16–N11
to give p-methoxy-phenylazo fragment ion of m/z = 245 (RI = 30%)
followed by the loss of p-methoxy group via the rupture of the
second weak bond C22–O25 to give a fragment ion of m/z = 105
consequently dye IV stability (heat of formation = 181.99 kcal/mol).
Consequently comparison of practical mass results of dyes with
theoretical MOCs facilitates to understand stability of these com-
pounds, fragmentation pathways, and the bond cleavage order of
each dye.
(
RI = 23%) and finally it gives the most abundant fragment ion of
m/z = 77 (RI = 100%) in the same way like dye I. In path 2, dye III still
keeping the p-methoxy group after rupture of the bond C5–N11
leading to the formation of the fragment ion of m/z = 226 (RI = 25%).
In path 3, the dye III loosed the p-methoxy-phenylazo group via
the rupture of the weak bond C3–N9 leaving the parent fragment
ion of p-acetoamidophenol of m/z = 151 (RI = 60%). This comparison
conclude that dye III (heat of formation = 144.96 kcal/mol) is more
stable than dye II and less stable than dye I. Dye IV mass shows three
pathways, it starts in path 1 with cleavage of the bond C22–N25 by
loss of NO2 group leading to the formation of low abundant frag-
3.7. The biological activities of dyes
The biological activity of four dyes was studied and data
obtained are represented in Table 9. From these data it concluded
that [28–33] the dye II (P–Br) is the most biologically active dye
where it produced 38 and 100% mortalities of the T. confusum at
concentration of 10 and 30%, respectively. The same previous two
concentrations of the dye III (p-OCH3) caused 15 and 40 mortal-
ities of adult T. confusum; the dye IV (p-NO ) caused 12 and 30
2
mortalitis and finally the dye I (p-H) caused 9 and 20 of T. confusum
mortality, respectively. On the other hand the most effective con-
centration was 30 in case of dye II while the concentration of 50%
was the highest bioactive in case of dye III, dye IV and dye I, respec-
tively. In order of toxicity, LC50 of the phenylazo compounds was
Table 9
Effect of the phenylazo compounds on the Tribolium confusum.
Phenyazo compounds
%
Conc.
Dye III (p-OCH3)
Dye IV (p-NO2)
Dye II (p-Br)
38
Dye I (p-H)
3
6, 48, 15 and 64 of the dye III, dye IV, dye II and dye I, respec-
1
3
5
0
0
0
15
40
68
36
00
12
30
55
48
00
9
20
36
64
00
tively. It is finally concluded that presence of p-substituent near
to the azo group make the dye biologically active. The order of
reactivity of these dyes is more or less the reverse of their stabil-
ity.
100
00
15
00
LC50
Control

1
036
M.A. Zayed et al. / Spectrochimica Acta Part A 78 (2011) 1027–1036
4
. Conclusion
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2333–2338.
techniques for structure elucidation of four novel acetoamidophe-
nol azodyes I–IV. Also, computational investigation is applied
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species of these compounds I–IV. From the mass results and MOCs
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during ionization with 70 eV electron beam is: dye I > dye III > dye
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support this order. Consequently comparison of practical mass
results of dyes with theoretical MOCs facilitates to understand sta-
bility of these compounds, fragmentation pathways, and the bond
cleavage order of each dye. These data can be used in industrial
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