Triazene 1-oxide compounds: Synthesis, characterization and evaluation as fluorescence sensor for biological applications

Article abstract of DOI:10.1016/j.molstruc.2013.12.055

Triazene compounds have been known for over 100 years. This class of compounds is versatile because they have different applications as chemical and biological reagents. Currently, the tendency of applications of triazenes is as fluorescence sensors with biological and environmental functions. This work discusses the synthesis and structural characterization through IR, MS (EI), ¹³C NMR, ¹H NMR, DSC and TGA analyzes of two triazene compounds, namely, 1-methyl-3-(p-carboxyphenyl)triazene 1-oxide (1) and 1-methyl-3-(phenyl)triazene 1-oxide (2). A comparative fluorescence study between these triazenes as potential compounds for application in the chemical sensors has also been carried out. The molecular and supramolecular structure of compound (1) is also determined by X-ray diffraction on single crystal, where the classic hydrogen bonds give the tridimensional arrangement. The presence of para-carboxylic group in compound (1) as well as the polarity and viscosity of the tested organic solvents resulted in a great influence on the evaluation of the fluorescence effect. These experimental findings show the potential use of triazenes as fluorescent biosensors, since the compounds (1) and (2) present antimicrobial activity.2014 Elsevier B.V. All rights reserved.

Full text of DOI:10.1016/j.molstruc.2013.12.055

Journal of Molecular Structure 1060 (2014) 264–271
Contents lists available at ScienceDirect
Journal of Molecular Structure
journal homepage: www.elsevier.com/locate/molstruc
Triazene 1-oxide compounds: Synthesis, characterization and evaluation
as fluorescence sensor for biological applications
a
a
Aline Joana Rolina Wohlmuth Alves dos Santos a, , Patrícia Bersch , Huéder Paulo Moisés de Oliveira ,
⇑
Manfredo Hörner b, Gustavo Luiz Paraginski b
a Universidade Federal de Pelotas – UFPel, Campus Universitário do Capão do Leão, Centro de Ciências Químicas, Farmacêuticas e de Alimentos, Caixa Postal 354, CEP 96.010-900
Pelotas, RS, Brazil
b Universidade Federal de Santa Maria – UFSM, Departamento de Química, Caixa Postal 5031, 97.105-970 Santa Maria, RS, Brazil
g r a p h i c a l a b s t r a c t
h i g h l i g h t s
The compound 1-methyl-3-p-(carboxyphenyl)triazene 1-oxide is unidimensionally polymerized by clas-
sical hydrogen bonds in the direction of the crystallographic axis a. The synthesis and structural charac-
terization by spectroscopic and non-spectroscopic methods as well as the influence of solvents on the
fluorescent properties of triazenes 1-oxide are discussed in this work.
ꢀ
Spectroscopic and structural
characterization of two triazene
1-oxide compounds.
X-Ray determinations on single
crystal.
ꢀ
ꢀ
ꢀ
Comparative fluorescence study.
Evaluation as potential fluorescence
sensor.
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 3 August 2013
Received in revised form 20 December 2013
Accepted 20 December 2013
Available online 3 January 2014
Triazene compounds have been known for over 100 years. This class of compounds is versatile because
they have different applications as chemical and biological reagents. Currently, the tendency of applica-
tions of triazenes is as fluorescence sensors with biological and environmental functions. This work dis-
cusses the synthesis and structural characterization through IR, MS (EI), 13C NMR, 1H NMR, DSC and TGA
analyzes of two triazene compounds, namely, 1-methyl-3-(p-carboxyphenyl)triazene 1-oxide (1) and
1-methyl-3-(phenyl)triazene 1-oxide (2). A comparative fluorescence study between these triazenes as
potential compounds for application in the chemical sensors has also been carried out. The molecular
and supramolecular structure of compound (1) is also determined by X-ray diffraction on single crystal,
where the classic hydrogen bonds give the tridimensional arrangement. The presence of para-carboxylic
group in compound (1) as well as the polarity and viscosity of the tested organic solvents resulted in a great
influence on the evaluation of the fluorescence effect. These experimental findings show the potential use
of triazenes as fluorescent biosensors, since the compounds (1) and (2) present antimicrobial activity.
Ó 2014 Elsevier B.V. All rights reserved.
Keywords:
Fluorescence
Biological sensor
Triazene 1-oxide
1. Introduction
(N@NANH), which act as Lewis bases when deprotonated and,
then, coordinate metal ions [2]. They are capable of forming supra-
The chemistry of triazenes has been researched since 1859
when Griess synthesized the first triazene, 1,3-bis(phenyl) triazene
[1]. Triazenes are azo compounds with diazoaminic aliphatic chain
molecular aggregates through intermolecular bonds, metal–ligand
interactions, and secondary ligand–ligand interactions [3].
Triazenes 1-oxide are basic compounds O(N) with negative
charge in the deprotonated form, where the proximity of the two
coordination sites (O and N) confers to these compounds good
chelating ability and a tendency to stabilize in the formation of five
⇑
Corresponding author. Fax: +55 3275 7533.
E-mail address: alinejoana@gmail.com (A.J.R.W.A. dos Santos).
0022-2860/$ - see front matter Ó 2014 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.molstruc.2013.12.055

Aline Joana Rolina Wohlmuth Alves dos Santos et al. / Journal of Molecular Structure 1060 (2014) 264–271
265
equipment; the differential thermogravimetry analysis (TGA) in a
Shimadzu-60 equipment; and the fluorescence analysis in a spec-
trofluorimeter Perkin Elmer model LS55.
R
N
R'
R
N
R'
N
N
O
N
H
N
O
H
I
II
2.2. Synthesis and structural characterization
Fig. 1. Tautomerism between 3-hydroxitriazene (I) and triazene 1-oxide (II). R,
The compound 1-methyl-3-(p-carboxyphenyl)triazene 1-oxide
(1), Fig. 2, was synthesized from the diazotization of p-aminoben-
zoic acid (0.5 g, 0.0036 mol) with sodium nitrite (0.3 g, 0.0043 mol)
in acid solution containing 5 mL of hydrochloric acid and 5 mL of
distilled water and by coupling of the diazonium salt formed with
N-methylhydroxylamine hydrochloridrate (0.169 g, 0.0020 mol).
The solution was maintained at 0–5 °C and neutralized with aque-
ous solution of sodium acetate. The yellow powder formed was fil-
tered. Yield 29.2% from N-methylhydroxylamine hydrocloridrate,
m.p.: 215–217 °C. The adopted conditions for the thermogravimet-
ric analysis (TGA) and calorimetry analysis (DSC) were: 1.218 mg
of the compound (1), heating rate of 5 °C minꢁ1, range 40–300 °C
and flow of nitrogen gas (50 mL minꢁ1). Single crystals suitable
for X-ray diffraction analysis were obtained by slow evaporation
of a solution of compound (1) (0.05 g) in 10 mL of dimethylsulfox-
ide and 10 mL of toluene.
R0 = aryl, alkyl.
atom rings with transition metal ions [4]. In the solid form, these
compounds have rather a deprotonated hydroxyl group (AO)
bonded to the N(1) atom of the triazene chain. The result is a
tautomeric equilibrium characteristic of this class of compounds
between solid form (triazene 1-oxide) and solution form
(3-hydroxitriazene) (Fig. 1) [3].
This class of compounds is versatile because they have different
applications as chemical and biological reagents for the metal
determination in analytical chemistry [5] as well as intermediate
compounds in organic synthesis [6]. They are not only important
in the study of supramolecular arrangements in metallic
complexes [3,4] but also because they present antibacterial
[7–9], antileukemic [10], anticancer [11] and antitripanosomal
[12] activities. Currently, the tendency of application of triazenes
is as fluorescence sensors with biological and environmental
functions [3–5,13].
Azo compounds evidence fluorescent capacity and due to this
reason triazenes and triazene 1-oxide are studied as chemical sen-
sors. [14,15] This work shows and discusses the synthesis and the
structural characterization of two triazene 1-oxide representatives
of the azo compounds class, 1-methyl-3-(p-carboxyphenyl)tria-
zene 1-oxide (1) and 1-methyl-3-(phenyl)triazene 1-oxide (2) as
well as the comparative fluorescence study between them as
potential compounds for the application in chemical sensors.
Structural characterizations of (1): IR (KBr/cmꢁ1
) m: 3297,
1683, 1607, 1520, 1417, 1246. 1H NMR (300 MHz, DMSO-d6,
273 K, ppm): d = 11.50 (s, COOH), 7.84 (2d, 2H, H3 and H5,
J3-2 = J5-6 = 8,79 Hz), 7.29 (2d, 2H, H2 and H6, J2-3 = J6-5 = 8,79 Hz),
3.95 (s, 3H, CH3), 1.90 (s, OH). 13C NMR (75 MHz, DMSO-d6,
273 K, ppm): d = 167.61 (ACOOH), 144.08 (C1, Carom) 125.17 (C4,
Carom), 130.81 (2C, C3, C5, Carom), 113.25 (2C, C2, C6, Carom), 50.63
(CH3). MS (EI, 70 eV, 280 °C, inlet) (M/z): 195 (molecular ion),
149, 121, 77 and 65 (base peak) [15]. DSC:
201.37 °C [16]. TGA: 74.5% weight loss at 180–280 °C [17].
D
H = ꢁ963.38 J gꢁ1 at
The compound 1-methyl-3-(phenyl)triazene 1-oxide (2), Fig. 2,
was synthesized from the diazotization of aniline (1.2 mL,
12.1 mmol) with sodium nitrite (1 g, 14.5 mmol) in acidic solution
containing 30 mL of hydrochloric acid and 60 ml of distilled water
and by coupling with N-methylhydroxylamine hydrocloridrate
(1 g, 12.1 mmol). The solution was maintained at 0–5 °C and
neutralized with aqueous solution of sodium acetate. The yellow
powder formed was filtered. [9] Yield 45% from N-methylhydroxyl-
amine hydrocloridrate, m.p.: 67–68 °C. The adopted conditions for
the thermogravimetric analysis (TGA) and calorimetric analysis
(DSC) were: 1.730 mg of the compound (2), heating rate of
2. Experimental
2.1. Materials and methods
Reagents were of PA grade and were used without purification,
except the tetrahydrofuran and acetonitrile, which were of spec-
troscopic grade.
The melting point was determined in the equipment PFM II; the
infrared spectroscopy analysis in a spectrophotometer Shimadzu IR
Prestige-21 using potassium bromide tablet (KBr); the 1H NMR
spectroscopy (300 MHz) and the 13C NMR spectroscopy (75 MHz)
in a multinuclear BRUKER equipment; the mass spectrometry (EI)
in a Triple Quadrupole detector – LC–MS–MS; the ultraviolet–
visible spectroscopy analysis in a spectrophotometer Perkin Elmer
Model Lambda 25; the structural analysis by X-ray diffraction on
single crystal in the diffractometer BRUKER APEX II-CCD; the differ-
ential scanning calorimetry analysis (DSC) in a SHIMADZU-60
5 °C minꢁ1
,
range 40–300 °C and flow of nitrogen gas
(50 mL minꢁ1).
Structural characterizations of (2): IR (cmꢁ1): 3219, 3048, 2947,
1598, 1517, 1351, 1205, 757, 686. 1H NMR (300 MHz, CDCl3, 273 K
ppm): 10.14 (s, NH)., 7.32 (t, 2H, H3 and H5, J3-2 = J3-4 = J5-4 = J5-6
=
7,7 Hz), 7.13 (d, 2H, H2 and H6, J2-3 = J6-5 = 7,7 Hz), 7.32 (t, 1H, H4,
J4-3 = J4-5 = 7,6 Hz), 3.97 (s, 3d, ACH3), 1.76 (s, OH). 13C NMR
(75 MHz, CDCl3, 273 K, ppm): 139.99 (C1, Carom), 129.63 (2C, C3,
C5, Carom), 122.83 (C4, Carom), 114.48 (2C, C2, C6, Carom), 50.14
5
5
HOOC
4
6
6
4
N
H
C
3
N
H
C
3
3
3
1
1
N
N
O
N
H
N
O
2
2
H
(2)
(1)
Fig. 2. Molecular structure of 1-methyl-3-p(carboxyphenyl) triazene 1-oxide (1) and 1-methyl-3-phenyl(triazene) 1-oxide (2).

266
Aline Joana Rolina Wohlmuth Alves dos Santos et al. / Journal of Molecular Structure 1060 (2014) 264–271
(CH3). MS (EI) (M/z): 100 (base peak), 87, 73, 60, 45 [15]. DSC:
H = 119.62 J gꢁ1 at 65.62 °C and H = 78.74 J gꢁ1 at 154.45 °C
[18,19]. TGA: 95.7% weight loss at 100–150 °C [20].
employing direct methods using the program SHELXS-97 [22].
The refinements were performed with programs of SHELXS-97
[22] using the method full matrix/least square structural factors
F2 with anisotropic thermal displacement parameters for all
non-hydrogen atoms and for H1 and H2 hydrogens. The other
hydrogens in the structure were geometrically located. The projec-
tions of the crystal structures were performed using the program
DIAMOND [23] and ORTEP [24].
D
D
2.3. Spectroscopic studies
Solutions for the fluorescence study were prepared from a stan-
dard solution of 1.0 ꢂ 10ꢁ4 mol Lꢁ1 of the compounds (1) and (2) in
dimethylsulfoxide. The solvents employed were Milli-Q water, eth-
anol (MeOH), dimethylsulfoxide (DMSO), N,N-dimethylformamide
(DMF), acetonitrile (MeCN), methanol (MeOH), and tetrahydrofu-
ran (THF). The absorption and emission spectra of compounds (1)
The compound (1) is found as monoclinic system with space
group C2/c and final disagreement indices [I > 2
r (I)] as
R1 = 0.0325, wR2 = 0.0977 and disagreement indices (all data) as
R1 = 0.0374, wR2 = 0.1016. Data and intensity collection by X-ray
diffraction on single crystal and refinement of the crystal structure
of (1) are in Table 1. The molecular structure of compound (1) is
shown in Fig. 3 and has the molecular formula C8H9N3O3.
and (2) were obtained in the final concentration of 1.0 ꢂ 10ꢁ5
-
mol Lꢁ1 in all solvents used. The excitation wavelength was
selected based on the evaluation of the maximum absorption in
the UV–vis spectrum. The analyses were performed in quartz
cuvettes. The quantum yield values were determined by compari-
son to a solution of naphthalene in cyclohexane used as a standard
for fluorescence, at the same concentrations as (1) and (2)
solutions [21].
The bond lengths observed for (1) in the NNN chain of the tria-
zene are 1.2718(13) Å for N11AN12 and 1.3389(14) Å for
N12AN13. The bond lengths are 1.2769(12), 0.880(17) and
0
0.98(3) AÅ, for N11AO1, N13AH1 and O22AH2, respectively. The
bond angle between N12AN11AO1 corresponds to 124.13(10)°
and the angle between N11AN12AN13 corresponds to
112.12(10)°. These values are not significantly different from
the values reported for the compound (2) [9] and other
compounds structurally related to (1) [25]. The atoms of the
3. Results and discussion
3.1. Structure of compound (1) [C8H9N3O3]
p-carboxyl group of the compound (1) have bond lengths of
0
1.2380(15) and 1.2949(14) ÅA for C2AO21 and O22AC2, respec-
tively. The angle between O21AC2AO22 is 122.93(11)°. These
values are very similar to the values found in the literature for
triazene compounds with substituent o-carboxyl in the phenyl
[25,26] (see Table 1S).
The molecule of compound (1) is almost planar (r.m.s.
0.0192 Å), and this planarity is confirmed when the interplanar an-
gles between the main fragments of the molecular structure are
analyzed in Table 2. The compound (2) exhibits a deviation from
planarity slightly larger (r.m.s. 0.0505 Å) than a deviation observed
for (1) [9].
The compound (1) is unidimensionally polymerized by classical
hydrogen bonds in the direction of the crystallographic axis a. This
polymerization gives independent chains (Fig. 4). These intermo-
lecular bonds in the compound (1) are of the type N13AH1. . .O10
[code of symmetry (0) ꢁx, 2 ꢁ y, ꢁz], O22AH2. . .O2100 [code of
symmetry (00)½ ꢁ x, ꢁ½ ꢁ y, ꢁz]. There are inversion centers in
the center of the hydrogen bonds and in the center of the unit cell.
The spectroscopic analysis in the infrared region shows the
band
m
(NAH) of 3297 cmꢁ1, suggesting that this compound in
the solid state presents preferably the triazene 1-oxide tautomeric
form (form II). Other bands are: 1683 cmꢁ1
[
m
m
(C@O)]; 1607 cmꢁ1
(N@N)], 1246 cmꢁ1
[
[
m
m
(C@C)]; 1520 cmꢁ1 [d(NAH)]; 1417 cm-1 [
(NAN)] [15] (see Fig. 1S).
Mass spectrometry (EI) of the compound (1) shows the fragments
(m/z): 195 [M+, 10%], 149 [M-46, 28%, fragment ANNC6H4COOH],
121 [M-74, 90%, fragment AC6H4COOH], 77 [M-118, 30%, fragment
AC6H5] and 65 [base peak, M-130, fragment AC5H5) [15] (see
Fig. 2S).
Through the 1H nuclear magnetic resonance (NMR) analysis it
can be observed at 11.50 ppm the hydrogen of the ACOOH group.
Each doublet at 7.84 and 7.29 ppm refers of two aromatic hydro-
gens. At 3.95 ppm it can be observed a singlet of three hydrogens
of the ACH3 group, which is directly bonded to the nitrogen chain
NNN of the triazene. At 1.90 ppm, the singlet of the hydrogen of the
group AOH appears, indicating that the compound in the solution
has preferably the 3-hidroxytriazene tautomeric form (form I). At
2.50 ppm, hydrogens of the solvent DMSO-d6 appears [15] (see
Fig. 3S).
Table 1
Data and intensity collection by X-ray diffraction on single crystal and refinement of
the crystal structure.
The analysis of the 13C NMR shows at 167.61 ppm the carbon of
the ACOOH group. Between 144.08 and 125.17 ppm we can ob-
serve two substituted carbons of the phenyl ring. The other signals
at 130.81 and 113.25 ppm refer to the other carbons of the phenyl
ring. The signal of the carbon of the ACH3 group appears at
50.63 ppm, which is directly bonded to the nitrogen chain NNN
of the triazene. The heptet at 39.50 ppm corresponds to the solvent
DMSO-d6 [15] (see Fig. 4S).
Compound (1)
Cell parameters (Å)
a = 27.2046(8)
b = 5.02180(10) b = 113.13(1)°
c = 14.4503(5)
1815.47(9)
Volume (Å3)
Number of elementary formulas (Z)
8
Calculated density (M g/m3)
1.428
0.112
816
Linear absorption coefficient (mmꢁ1
F (000)
)
For compound (1) the melting point is 215–217 °C and the
calorimetry analysis (DSC) shows an exothermic peak at
201.37 °C of the decomposition of azo group with release of N2
Crystal size (mm3)
Scan region 2h (°)
Region indices
0.45 ꢂ 0.21 ꢂ 0.08
1.63–25.49
ꢁ32 6 h 6 30, ꢁ6 6 k 6 6,
ꢁ17 6 l 6 17
8515
(
D
H = ꢁ963.38 J gꢁ1) [16]. (see Fig. 5S) On the other hand, the ther-
Number of reflections collected
Number of independent reflections 1688 [R(int) = 0.0172]
Scan region 2h (°)
mogravimetric analysis (TGA) shows a single loss between 180 and
280 °C corresponding to a loss of 74.5% by mass (0.908 mg) [17]
(see Fig. 6S).
25.49%, 99.9%
0.9911 and 0.9510
1688/0/136
Max. and min. transmission
Data/restrictions/refined
parameters
The data collection of X-ray diffraction of the compound (1) was
performed on a diffractometer Bruker APEX II Kappa CCD (Charge
Goodness-of-fit on F2
Residual electron density (e Åꢁ3
1.126
0.162 and ꢁ0.193
0
Coupled Device Detector), using Mo K
a radiation (0.71073 ÅA) with
)
a graphite monochromator, at 293 K. The structure was solved

Aline Joana Rolina Wohlmuth Alves dos Santos et al. / Journal of Molecular Structure 1060 (2014) 264–271
267
3.2. Structure of compound (2) [C7H9N3O1]
The spectroscopy analysis in the infrared region shows the band
m
(NAH) (very strong) at 3219 cmꢁ1, suggesting that both com-
pounds (1) and (2) in the solid state present preferably the triazene
1-oxide tautomeric form (form II). Other bands are: 3048 cmꢁ1
[
m
(CAH) ꢁo1f the phenyl], 2947 cmꢁ1
[
m
(CAH) of the methyl],
1598 cm
1205 cmꢁ1
Fig. 7S).
[m [m(C@N)],
(C@C)], 1517 cmꢁ1 [d(NAH)], 1351 cmꢁ1
[m
(N@N ? O)], 757 and 686 cmꢁ1 [d(CAH)] [15] (see
Through the mass spectrometry (EI) of compound (2) we
can observe the fragments (m/z): 100 [base peak, M-51, fragment
AC2H2NNNOHCH3), 87 [M-64, 40%, fragments ANNOHCH3 and
AC2H3], 73 [M-78, 30%, fragment ANNNOCH3], 69 [M-82, 45%,
fragments ANC3H2 and AOH], 45 [M-106, 75%, fragment ANOCH3]
[15] (see Fig. 8S).
Fig. 3. Projection of the molecular structure of compound (1) (ORTEP [24]). Thermal
ellipsoids represented in probability level of 50%.
1H NMR and 13C NMR of compound (2) were performed in deu-
terated chloroform (CDCl3). 1H NMR (300 MHz) shows a hydrogen
of the NH group at 10.14 ppm, indicating the presence of com-
pound (2) as triazene 1-oxide (tautomeric form II). The singlet at
3.97 ppm refers to the hydrogens of the CH3 group, which is di-
rectly bonded to the nitrogen chain NNN of the triazene (see
Fig. 9S).
The signal at 7.26 ppm represents the CHCl3. The signals for the
monosubstituted aromatic ring are observed as triplet at 7.32 ppm
(2H), as doublet at 7.13 ppm (2H), and as triplet at 7.32 ppm (1H)
[15].
Table 2
Interplanar angles (Å) between the main fragments of the molecule 1-methyl-3-(p-
carboxyphenyl) triazene 1-oxide (1).
Fragments
22C2O21/C11AC16
11AC16/N13N12N11O1C1
Planarity (r.m.s. Å) Interplanar angle (°)
O
C
0.0000/0.0064
0.0064/0.0093
1.6(2)
1.0(1)
0.8(1)
N13N12N11O1C1/whole molecule 0.0093/0.0192
Through the 13C NMR of compound (2), it can be observed at
77.23 ppm the signal from solvent CDCl3. The signal at
139.99 ppm refers to the quaternary carbon of the phenyl ring.
At 129.63 ppm (2C), 122.83 ppm (C) and 114.48 ppm (2C) we can
observe the other aromatic carbons. At 50.14 ppm, there is a signal
referent carbon of the methyl group [15] (see Fig. 10S).
The compound (2) has a melting point at 67–68 °C [9], and the
calorimetric analysis (DSC) shows an endothermic peak at 65.62 °C
On the other hand, the compound (2) has a centrosymmetric
dimmer form with bifurcated hydrogen bonds and classical and
non-classical intermolecular interactions. The intramolecular
interactions are of type NAH. . .O and intermolecular interactions
of the type NAH. . .O0 and CAH. . .O0 with symmetry code (0) ꢁ x,
ꢁy, ꢁz [9].
All complementary analysis methods of structural characteriza-
tion of the compound (1) are in agreement with the data obtained
by X-ray diffraction on single crystal.
(D
H = 119.61 J gꢁ1
154.45 °C (
H = 78.74 J gꢁ1), indicating a fusion process possibly
) [22] and another endothermic peak at
D
Besides the proven structure, the compound (1) also has bacte-
riostatic and bactericidal activities well defined. Evaluation of min-
imal inhibitory concentration (MIC) and minimum bactericidal
concentration (MBC) for a range of gram positive and gram nega-
tive bacteria showed the most significant result against the bacte-
from the rearrangement of the structure after the first melting pro-
cess [19] (see Fig. 11S). On the other hand, the thermogravimetric
analysis (TGA) shows only a loss between 100 and 150 °C corre-
sponding to a loss of 95.7% by mass (1.657 mg) [20] (see Fig. 12S).
rium Streptococcus agalactiae with MIC of 16
32 g/mL. The highest dilution used for compound (1) was
128 g/mL [8]. The same evaluation was reported in the literature
lg/mL and MBC of
l
l
3.3. UV–visible and fluorescence spectroscopic studies
for the compound (2). It showed the activity against the bacterium
Micrococcus spp. with MIC of 128 mg/mL and appeared inactive
against gram-negative bacteria [8,9].
The influence of solvents on the spectroscopic properties of
triazenes 1-oxide (1) and (2) was evaluated. The polar solvents
used for the analysis were dimethylsulfoxide (non-protic),
Fig. 4. Projection of one-dimensional polymerization of molecules of compound (1) in the crystallographic direction [100]. The classical hydrogen bonds are represented by
dotted lines. (DIAMOND [23]).

268
Aline Joana Rolina Wohlmuth Alves dos Santos et al. / Journal of Molecular Structure 1060 (2014) 264–271
Fig. 5. UV–visible spectrum of triazenes 1-oxide (1) and (2) in organic solvents.
Table 3
Table 4
Parameters of organic solvents.
Photophysical properties of compounds (1) and (2) in organic solvents.
Solvent ETa(kcal/mol) [30]
l
b(Debye)
e
c(°C)
g
(cP)d[30]
D
fe
Solvent kabs a(max/
kem b(max/
nm)
D
m
e
Uf
e
25 °C
[30]
(25 °C)
nm)
c(cmꢁ1
)
d(Mꢁ1 cmꢁ1
)
MeOH
EtOH
MeCN
DMSO
DMF
55.4
51.9
45.6
45.1
43.2
37.4
1.7
1.7
3.2
3.96
3.8
32.6
22.4
37.5
46.6
36.7
7.6
0.6
0.849
0.417
1.143
0.151
0.618
0.841
Compound (1)
1.08
0.38
2.0
0.82
0.55
MeOH
EtOH
MeCN
DMSO
DMF
318
321
320
327
325
325
389
399
389
404.5
429.5
390.5
6089.9
24765.0
27497.7
29339.9
27410.4
27986.2
27925.6
0.0006
0.0156
0.0017
0.01
0.0140
0.0073
5739.6
5543.0
5232.5
7486.3
5161.0
THF
1.75
TFH
a
b
c
Solvent polarity.
Dipole moment.
Compound (2)
MeOH
EtOH
MeCN
DMSO
DMF
307
308
306
312
310
310
380.5
382.5
381.5
387.5
355
6292.0
6323.7
6467.4
6244.8
4089.0
6250.2
19506.8
19649.6
19405.9
19640.6
19839.1
20882.7
0.0092
0.0139
0.0019
0.0040
0.0096
0.0016
Dielectric constant.
Absolute viscosity.
Parameter of solvent polarity.
d
e
THF
384.5
a
b
c
Maximum absorption.
Maximum emission.
Stokes shift.
Molar absorptivity.
Quantum yield.
N,N-dimethylformamide (non-protic), methanol (protic), ethanol
(protic), tetrahydrofuran (non-protic), and acetonitrile (non-
protic). The absorption spectra are shown in Fig. 5. In the spectra
of compounds (1) and (2) there is a large band between 300 and
350 nm because of the union of bands of the chromophore groups
AN@NA for compounds (1) and (2) and AC@O for compound (1)
and auxochrome group AOH for compounds (1) and (2), character-
istic of compounds 3-hydroxytriazenes. Another characteristic
band of this class of compounds can be observed for the compound
(2) and is located in the region between 280 and 300 nm [13,27].
The intramolecular transfer of electrons from the nonbonding
orbitals of the occupied molecular orbital of the highest energy
d
e
(HOMO) of the donor NAOH to the unoccupied molecular orbital
of the lowest energy (LUMO) of the receptor N-Phenyl increases
the electron density on this nitrogen atom, which is directly
bonded to the aromatic ring. Furthermore, the excitation of
nonbonding electrons from the oxygen of NAOH decreases the
electron density on the oxygen, weakening the OAH bond. Thus,
Fig. 6. Emission spectra of fluorescence in organic solvents for the compounds 1-methyl-3(p-carboxyphenyl)triazene 1-oxide (1) and 1-methyl-3-(phenyl)triazene
1-oxide (2).

Aline Joana Rolina Wohlmuth Alves dos Santos et al. / Journal of Molecular Structure 1060 (2014) 264–271
269
identify a band of less absorption intensity in the region between
280 and 300 nm.
When the solvent polarity decreases (Table 3) there is a bacto-
cromic effect (positive) in the studied solvents when compared to
the solvent of higher polarity (methanol). This effect is observed
for both compounds. The bathochromic effect of the compounds
(1) and (2) is higher in non-protic solvents, which indicates
rearrangement of the load of solute molecules under excitation
electronics [28].
For diazoaminebenzene derivatives, whose benzene ring
presents withdrawing groups or donor groups of electron, as in
compounds (1) and (2), a solvatochromic effect in the region
between 300 and 350 nm is observed due to the increase of the
donor–acceptor character of the substitutents in solution [13]. This
band can be referred to the low-energy transition
p ? p
* or transi-
tion n ?
p
*, involving electrons of the azo group AN@NA and azo-
methine group AN@NACH3 [28,29]. For compound (2), the band
located at 280–300 nm can be attributed to a higher electron
density on the nitrogen atom of the triazene chain bonded to the
aromatic ring, as well as to the moderate energy to transitions
Fig. 7. Relation between maximum absorption and polarity parameter for com-
pounds (1) (N—) e (2) (j—).
p
?
p* of the aromatic rings [28].
The change of polarity of the organic solvents for compounds
the hydrogen of OAH migrates to the nitrogen bonded to the
aromatic ring [13]. This migration is responsible for tautomerism
in this class of compounds.
(1) and (2) analyzes have significant influence on the maximum
emission (Fig. 6). Thus, a batocromic effect is observed in the emis-
sion spectra when the polarity of the solvent decreases, except for
THF in the spectrum for compound (1) and for DMF in the
spectrum for compound (2) [31].
A
significant interference from electronic absorption of
compounds (1) and (2) with the polarity of the solvent is observed
through a bathochromic effect of the bands in the region of
300–350 nm, more generally, for all the studied solvents for
compounds (1) and (2). For compound (2) it is also possible to
Compound (1) has more resonance capacity, which could
increase the fluorescence effect of this compound; however, this
Fig. 8. Relation between the Stokes shift and the solvent polarity parameter for compounds (1) and (2).
Fig. 9. Relation between quantum yield and the solvent polarity parameter for compounds (1) and (2).

270
Aline Joana Rolina Wohlmuth Alves dos Santos et al. / Journal of Molecular Structure 1060 (2014) 264–271
Fig. 10. Relation between the quantum yield and the solvent viscosity for compounds (1) and (2).
was not observed in the experimental tests (Fig. 6). Compounds (1)
and (2) show the N@N group that decreases the fluorescence effect.
The only difference between the structures of compounds (1) and
(2) is the carboxylic group that decreases the fluorescence effect
in the emission spectra of compound (1) [21]. Table 4 shows the
spectral properties of compounds (1) and (2) in organic solvents
[21].
spectroscopic and fluorescence analyzes of compounds (1) and
(2) demonstrated the fluorescent effect of the triazene 1-oxide
class. The presence of para-carboxylic group in compound (1)
and the polarity and viscosity of the organic solvents resulted in
great influence on the evaluation of the fluorescence effect. These
observations show triazenes as possible application as fluores-
cence sensors. Thus, further researches have to be made with other
substituent groups in the NNN triazene chain that can increase the
fluorescent effect of this compound class for the study of fluores-
cent biosensors, since the compounds (1) and (2) show antimicro-
bial activity.
Fig. 7 shows that there is an increase of maximum absorption
with the increase of the solvent polarity when the solvent interacts
with the compound. This increased polarity effect is greater for com-
pound (1) compared to compound (2) because there is a negative
tendency when the polarity parameter decreases thus there is an
influence of the solvents on the structure of compound (1).
This may explain why compound (1) has a carboxylic group at para
position in the aromatic ring. Furthermore, the increase of the
maximum absorption with the increase of the solvent polarity indi-
cates greater stability of the compound in the excited state [32].
In Fig. 8, the positive tendency when the polarity parameter
decreases shows that the dipole moment for compound (1) is
greater in the ground state than in the excited state. For compound
(2), the negative tendency indicates that the dipole moment is
lower in the ground state than in the excited state. The Stokes shift
increases with the increase of the solvent polarity for
compound (2), which indicates a strong solvation of compound
(2) [31].
Acknowledgements
CAPES/CNPq/Brazil (research Grant, proc. 552197/2011-4).
Appendix A. Supplementary material
Supplementary data associated with this article can be found,
in the online version, at http://dx.doi.org/10.1016/j.molstruc.2013.
12.055.
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