New long alkyl side-chain benzo[a]phenoxazines as micellisation probes

Article abstract of DOI:10.1016/j.tetlet.2009.05.055

Several novel fluorescent benzo[a]phenoxazinium chlorides possessing a long aliphatic chain substituent at the 5-amino function of the heterocycle were efficiently synthesised. All compounds obtained absorbed and emitted at longer wavelengths with moderat

Full text of DOI:10.1016/j.tetlet.2009.05.055

Tetrahedron Letters 50 (2009) 4470–4474
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
New long alkyl side-chain benzo[a]phenoxazines as micellisation probes
Carla M. A. Alves a, Sarala Naik a, Paulo J. G. Coutinho b, M. Sameiro T. Gonçalves a,
*
a Centro de Química, Universidade do Minho, Campus de Gualtar, 4710-057 Braga, Portugal
b Centro de Física, Universidade do Minho, Campus de Gualtar, 4710-057 Braga, Portugal
a r t i c l e i n f o
a b s t r a c t
Article history:
Several novel fluorescent benzo[a]phenoxazinium chlorides possessing a long aliphatic chain substituent
at the 5-amino function of the heterocycle were efficiently synthesised. All compounds obtained
absorbed and emitted at longer wavelengths with moderate to good fluorescent quantum yields. The
photophysics of N-[10-methyl-5-(octylamino)-9H-benzo[a]phenoxazin-9-ylidene]ethanaminium chlo-
ride and N-[10-methyl-5-(dodecylamino)-9H-benzo[a]phenoxazin-9-ylidene]ethanaminium chloride
was studied in TritonÒ X-100 and in cetyltrimethylammonium bromide micellar media, demonstrating
the capability of these fluorophores in detecting the micellisation process.
Received 31 March 2009
Revised 12 May 2009
Accepted 18 May 2009
Available online 22 May 2009
Keywords:
Benzo[a]phenoxazines
Nile Blue
Ó 2009 Elsevier Ltd. All rights reserved.
Long-wavelength dyes
Lipid bilayer
Micelles
1. Introduction
new benzo[a]phenoxazines with aliphatic side chains that would
function as anchors in biological structures (membranes, proteins,
Contemporary analytical methodologies based on fluorescent
spectroscopy techniques have been well used for biological pur-
poses.1–3 In this field, suitable probes include fluorophores with
absorption and emission at longer wavelengths (600–1000 nm),
which showed minimal background interference from biological
material as well as high sensitivity.4,5 Benzo[a]phenoxazines, such
as Nile Blue and its derivatives possessing a positively charged, oxi-
dised, phenoxazine system, are among the fluorochromophores
used in bioapplications as non-covalent and covalent labels.
As is well known, a number of biological macromolecules and
structures have hydrophobic and hydrophilic zones. The presence
of a long hydrocarbon chain in the fluorescence probe allows it
to easily bind to the hydrophobic parts of biomolecules or bio-
membranes, enabling the fluorophore moiety to probe its
environment.6
Micelles are a simple model of biomolecules as the hydrophobic
effect7 determines the micellisation process and is also the main
driving force for self-association of phospholipids in biological
membranes, as well as one of the factors that contributes to pro-
tein conformation. In addition, like biological membranes, micelles
have a polar surface and a hydrophobic interior. Thus, interaction
studies of fluorophores with micelles ascertain their ability as bio-
logical fluorescent probes.
etc.), as a preliminary study, we decided to synthesise new amphi-
phatic fluorochromophores possessing octyl or dodecyl substitu-
ents at the required amine function of the heterocycle, and used
three different combinations of substituents (including H, Me and
Et groups) in their 9-amine positions. All benzo[a]phenoxazines
obtained absorb and fluoresce, in ethanol, from 627 nm to
638 nm and from 654 to 678 nm, respectively.
The interaction of the synthesised N-[10-methyl-5-(octylami-
no)-9H-benzo[a]phenoxazin-9-ylidene]ethanaminium
chloride
(1b) and N-[10-methyl-5-(dodecylamino)-9H-benzo[a]phenoxa-
zin-9-ylidene]ethanaminium chloride (1e) with surfactant mi-
celles was studied by means of fluorescence spectroscopic
methods. The results showed that cationic phenoxazine com-
pounds with long alkyl side chains are capable of detecting the
critical micellar concentration, even in the case of cationic surfac-
tants, in which there is a repulsive electrostatic interaction be-
tween the micelles and the fluorescent probe.
Benzo[a]phenoxazinium chlorides 1a–g were synthesised by
condensation of 5-alkylamino-2-nitrosophenol hydrochlorides
2a–c with N-alkyl-naphthylamines 3a–c in an acidic medium
(Scheme 1).11 The required 5-alkylamino-2-nitrosophenol hydro-
chlorides 2a–c were synthesised using a standard procedure12,8
involving treatment of the corresponding 3-alkylaminophenol
with sodium nitrite in an acid solution. Intermediates 3a–c were
prepared by alkylation, in methanol (3a) or ethanol (3b and 3c),
of 1-naphthylamine with the appropriate bromo-derivative, 1-bro-
mopropane, 1-bromooctane and 1-bromododecane, respectively.13
After dry chromatography purification, these compounds were
Bearing this in mind, in connection with our current research
interests,8–10 and considering the fact that we intended to obtain
* Corresponding author. Tel.: +351 253 604372; fax: +351 253 604382.
E-mail address: msameiro@quimica.uminho.pt (M. S. T. Gonçalves).
0040-4039/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2009.05.055

C. M. A. Alves et al. / Tetrahedron Letters 50 (2009) 4470–4474
4471
NO
OH
R
N
N
R
HCl/EtOH
R1
R2
+
H
H
reflux
R1
N+
Cl-
R2
O
N
N
(CH2)nCH3
(CH2)nCH3
2
3
1
a n = 2, R = Me, R1 = H, R2 = Et
b n = 7, R = Me, R1 = H, R2 = Et
c n = 7, R = H, R1 = R2 = Et
d n = 7, R = H, R1 = R2 = Me
a R = Me, R1 = H, R2 = Et
b R = H, R1 = R2 = Et
c R = H, R1 = R2 = Me
a n = 2
b n = 7
c n = 11
e n = 11, R = Me, R1 = H, R2 = Et
f n = 11, R = H, R1 = R2 = Et
g n = 11, R = H, R1 = R2 = Me
Scheme 1.
Table 1
Yields, UV/visible and fluorescence data in ethanol for compounds 1a–g
Compound
Yield (%)
kabs (nm) (
e
[Mꢀ1 cmꢀ1])
kem (nm)
UF
Stokes’ shift (nm)
1a
1b
1c
1d
1e
1f
92
98
88
45
83
69
43
628 (46,000)
627 (67,000)
636 (68,000)
628 (45,000)
627 (63,000)
638 (46,000)
629 (56,000)
654
655
678
676
655
677
676
0.46
0.49
0.21
0.21
0.48
0.20
0.21
26
28
42
48
28
39
47
1g
obtained as oils (3a, 60%; 3b, 51%; 3c, 44%) and were characterised
by high resolution mass spectrometry, IR and NMR (1H and 13C)
spectroscopies.
ues of kabs, which are dependent on the amine substituents present
in the opposite position. Figure 1 shows the normalised absorbance
spectra of compounds 1b–d, in ethanol.
Condensation
of
5-ethylamino-4-methyl-2-nitrosophenol
The evaluation of the fluorescent properties of compounds 1a–g
in ethanol using Oxazine 1 as a standard (fluorescent quantum
yield, UF = 0.11 in ethanol14) was performed (Table 1). All com-
pounds were excited at 590 nm and the results showed that these
hydrochloride 2a with N-propylnaphthalen-1-amine 3a, in the
presence of hydrochloric acid, produced the benzo[a]pheno-
xazinium chloride 1a. Based on the nitroso compounds 2a,
5-diethylamino-2-nitrosophenol hydrochloride 2b and 5-dimeth-
ylamino-2-nitrosophenol hydrochloride 2c, and using N-octyl-
naphthalen-1-amine 3b, compounds 1b, 1c and 1d, respectively,
were obtained. N-Dodecylnaphthalen-1-amine 3c also reacted with
nitrosophenol hydrochlorides 2a, 2b and 2c, giving the correspond-
ing benzo[a]phenoxazine derivatives 1e–g.
fluorophores have wavelengths of maximum emission (kem
)
superior to 654 nm (1a), the highest value being 678 nm (1c).
Considering compounds 1a–g, it appears that the values of kem
are the same for all fluorophores bearing the same combination
of R, R1 and R2 substituents and regardless of the size of the alkyl
chain at the amine of the 5-position of the heterocycle. Thus, this
occurs when R = Me, R1 = H, R2 = Et, kem is 655 nm (1b, 1e) or
654 nm (1a). The higher values of kem arise with the combination
R = H, R1 = R2 = Me, where kem is 676 nm (1d, 1g) and R = H,
R1 = R2 = Et, where kem is 677 nm (1f) or 678 nm (1c). Figure 2
shows the normalised fluorescence spectra of compounds 1b, 1c
and 1d, in ethanol.
After purification by dry chromatography on silica gel, cationic
dyes 1a–g were isolated as solid materials in good to excellent
yields (Table 1) and were fully characterised by the usual analytical
techniques.
Electronic absorption spectra of 10ꢀ6 or 10ꢀ5 M solutions of
fluorophores 1a–g in degassed absolute ethanol were measured.
Summarised data of this study are presented in Table 1. These
compounds showed maximum absorption (kabs) at long wave-
Regarding the Stokes’ shift, benzo[a]phenoxazines with the
diethyl or dimethyl amine at the 9-position (R = H, R1 = R2 = Et or
lengths with molar absorptivities (e) between 45,000 (1d) and
68,000 (1c) Mꢀ1 cmꢀ1. Through the comparison of kabs of com-
pounds 1b–d possessing the same number of methylenic groups
(n = 7) and different substituents R, R1 and R2, it was possible to
see that compound 1c (R = H, R1 = R2 = Et) showed maximum
absorption at the longest wavelength.
1
1b
0.8
1c
By comparison of compounds 1e–g, with
a similar alkyl
0.6
0.4
0.2
0
1d
chain (n = 11) at the amine function in the 5-position of benzo[a]-
phenoxazine ring, there is the same trend, that is, a bathochromic
shift from the compound with the combination of substituents,
R = Me, R1 = H and R2 = Et to compounds with dialkylated amine
at the 9-position of the heterocycle, R = H, R1 = R2 = Me (2 nm)
and R = H, R1 = R2 = Et (11 nm).
400
450
500
550
600
650
700
Wavelength (nm)
Considering this behaviour, increasing the size of the alkyl chain
(change in the values of n) of the amine function of 5-position of
the benzo[a]phenoxazine ring does not seem to influence the val-
Figure 1. Normalised absorbance spectra of compounds 1b, 1c and 1d measured in
ethanol (C = 1 ꢁ 10ꢀ6 M).

4472
C. M. A. Alves et al. / Tetrahedron Letters 50 (2009) 4470–4474
tional methylenic groups in the same position, is only water-solu-
ble in the sub M range. This behaviour indicates that there are no
micellar aggregates of this type of molecules as they would result
in increased solubility. Furthermore, the fluorescence intensity of
compound 1b was found to be linear with concentration in the
1.00
0.80
0.60
0.40
0.20
0.00
1b
1c
1d
l
0.1–10-lM range (data not shown), which would not be the case
if fluorophores were positioned side by side in
a micellar
arrangement.
As a preliminary evaluation of the suitability of these long-alkyl
side-chain benzo[a]phenoxazine derivatives as biological probes,
we report a fluorescence study of N-[10-methyl-5-(octylamino)-
9H-benzo[a]phenoxazin-9-ylidene]ethanaminium chloride (1b)
and N-[10-methyl-5-(dodecylamino)-9H-benzo[a]phenoxazin-9-
ylidene]ethanaminium chloride (1e) in aqueous solutions of a neu-
tral (TritonÒ X-100, TX100) or positive (cetyltrimethylammonium
bromide, CTAB) surfactant shown in Figures 3 and 5. TX100 and
CTAB form micelles above a critical micellar concentration of
0.25 mM15 and 0.89 mM,16 respectively. The photophysical behav-
iour of compound 1b depended on the type of surfactant used.
Upon micellisation of TX100, 8 nm and 16 nm red shifts were ob-
served in emission and excitation, respectively (Fig. 3). Most fluo-
rescence probes have pp* transitions, which show a blue shift10 as
a result of localisation in the more hydrophobic core of the micelle.
From results of our previous work,8 we can conclude that the
nature of the transition was indeed pp*. The observed red shift
600
650
700
750
800
Wavelength (nm)
Figure 2. Normalised fluorescence spectra of compounds 1b, 1c and 1d measured
in ethanol (C = 1 ꢁ 10ꢀ6 M).
R = H, R1 = R2 = Me) showed higher values (39–48 nm) than fluoro-
phores possessing the monoalkylated amine (R1 = H, R2 = Et) as
well as the methyl group at 10-position of the heterocycle (26 or
28 nm).
Considering the fluorescent quantum yields of all compounds
(1a–g), it appears that values are located in two intervals, related
to the benzene ring substituents (R, R1 and R2) and regardless of
the size of the alkyl side chain of the amine in the naphthalene
ring. The values of UF with the combination R = Me, R1 = H,
R2 = Et (1a, 1b and 1e) were between 0.46 (1a) and 0.49 (1b) and
the other fluorophores (1c, 1d, 1f and 1g) occurred at 0.20 and
0.21. This behaviour is consistent with our previous results8 for
other benzo[a]phenoxazines with different types of substituents
at the 5-amine position of the heterocycle, namely, it was found
that electron-donating groups in the R position increased the fluo-
rescence quantum yields (up to 4 times upon changing H to Me).
The photophysical properties in organic solvents was the result
of the coexistence of acid and basic forms depending on the pro-
ton-acceptance nature of the solvent. The basic form occurred in
the 500 nm spectral region and the fluorescence quantum yield
was almost one magnitude order lower. In aqueous solutions it
was influenced by non-fluorescent H-aggregates (ꢂ50 nm blue
shift), which was dependent on the substituent groups (R, R1 and
R2).
1.2
below CMC
above CMC
0.8
increasing
[TX100]
0.4
0
400
450
500
550
600
650
700
750
The water solubility of benzo[a]phenoxazines 1a–g markedly
depends on the alkyl side chain length. Compound 1b, possessing
the alkyl substituent with 8 carbons, is easily soluble at least up
Wavelength (nm)
Figure 4. Normalised absorption spectra of compound 1e with TX100. The
concentration of compound 1e was 2 ꢁ 10ꢀ6 M.
to 10
lM concentration range, while compound 1e, with four addi-
Excitation Emission
1
665
7
6
660
5
4
655
3
2
650
CMC
1
below CMC
0
645
below
CMC
0.5
0
0.5
1
[TX-100] (mM)
above
above
CMC
CMC
0
500
550
600
650
700
Wavelength (nm)
750
800
850
900
Figure 3. Normalised fluorescence emission and excitation spectra of compound 1b with TX100. The inset shows the plot of the maximum fluorescence intensity, IF, ( , })
and the corresponding fluorescence wavelengths (N,
D) as a function of TX100 concentration. The filled and unfilled symbols are used, respectively, for compounds 1b and 1e.
The concentration of compounds 1b and 1e was 2 ꢁ 10ꢀ6 M.

C. M. A. Alves et al. / Tetrahedron Letters 50 (2009) 4470–4474
4473
3.5
3.0
2.5
2.0
1.5
1.0
670
665
660
655
650
1.0
0.5
0.0
CMC
2.0
0.0
0.5
1.0
1.5
2.5
[CTAB] (mM)
in water
Increasing [CTAB]
600
650
700
Wavelength (nm)
750
800
Figure 5. Normalised fluorescence spectra of compound 1b with CTAB. The inset shows the plot of the fluorescence intensity, IF, ( ) and the corresponding fluorescence
wavelength (N) as a function of CTAB concentration. The concentration of compound 1b was 2 ꢁ 10ꢀ6 M.
was probably due to the formation of ground state complexes
between the aromatic moieties of both compound 1b and TX100,
which disappeared upon micellisation. The possibility of variation
of the fraction of H-aggregates8 upon addition of the surfactant
cannot explain this result, since no variation would occur in the
shape of the excitation spectra. This is due to the fact that H-aggre-
gates do not fluoresce. Concerning the use of this type of probes in
proteins, the interaction of compound 1b with TX100 is interesting,
since all fluorescent amino acids (tyrosine, tryptophan and phenyl-
alanine) have aromatic fluorochromes.
In the case of compound 1b, there was obviously no ground
state association with CTAB molecules and the photophysical
behaviour can be interpreted by a rapid decrease in the amount
of H-aggregate fraction upon association of compound 1b with
CTAB micelles, which appear above the critical micellar concentra-
tion. This association occurred even though there was a charge
repulsion as a consequence of the effect of the octylamino substi-
tuent as an anchor in the hydrophobic core of the micelle. Due to
the hydrophobic effect7 the energetic expenditure would be far
higher if one octylamino substituent of the side chain was solvated
by water molecules.
In order to further confirm the ground state complexation, we
obtained the absorption spectra of compound 1b in TX100 water
solutions and found a similar red shift to that observed in excita-
tion spectra upon micellisation (data not shown). Compound 1e
showed a similar photophysical behaviour, but the variations in
absorption spectra were much more complex (Fig. 4) indicating
that various types of surfactant-dye aggregates existed and possi-
bly with the presence of the neutral basic form of 1e, which
showed an absorption maximum near 500 nm.8 In addition, a
much higher increase in fluorescence intensity upon micellisation
was observed for compound 1e than for compound 1b (60 times
instead of three times). The more pronounced increase could be
due to a sudden change in the amount of the basic form of 1e
which should have a ꢂ10 times lower fluorescence quantum yield
than the acid form. In the case of compound 1b, no basic form was
detected in the absorption spectra (data not shown) and we con-
clude that the quantum yields of 1b in water and in TX100 micelles
are similar with the increase in fluorescence intensity originating
mainly from a ꢂ4 times increase in absorbance.
2. Conclusion
In this work, 5,9-diaminobenzo[a]phenoxazinium dyes 1a–g,
containing long-alkylamino side chains at 5-position of the fluoro-
chromophores were efficiently synthesised.
These cationic dyes, with absorption in ethanol in the 627–
638 nm range, were highly fluorescent and revealed a maximum
wavelength between 654 and 678 nm. Preliminary results of the
study of N-[10-methyl-5-(octylamino)-9H-benzo[a]phenoxazin-9-
ylidene]ethanaminium chloride (1b), with TritonÒ X-100 and
cetyltrimethylammonium bromide showed the potential of this
fluorophore in sensing the micellisation process of neutral and cat-
ionic surfactants. The longer side chain derivative, N-[10-methyl-
5-(dodecylamino)-9H-benzo[a]phenoxazin-9-ylidene]ethanamini-
um chloride (1e) showed a distinct photophysical behaviour and is
better in the detection of the micellisation of TritonÒ X-100, but is
unable to detect the self-organisation of the CTAB surfactant. The
results obtained in this investigation suggested that this type of
5,9-diaminobenzo[a]phenoxazines presents great potential as bio-
logical fluorescent probes in the near-infrared spectral region.
Regarding the positive surfactant, CTAB, no spectral shifts were
observed from the fluorescence measurements obtained but the
micellisation process was still detected (Fig. 5) through a sudden
increase in fluorescence intensity. From the absorption measure-
ments (data not shown), it was observed that compound 1e again
had a very distinct behaviour when compared to compound 1b. In
the latter, there were no significant spectral variations while, for
the former, there were complicated spectral features involving
mainly the neutral basic form (bands from 450 to 550 nm in the
spectral region) with very little contribution from the usual acid
form near 600 nm. As a result, there was no trend in fluorescence
measurements, which allowed for the detection of the micellisa-
tion process of CTAB using compound 1e. Both the dependence
of fluorescence intensity and maximum emission wavelength
showed an oscillating behaviour (data not shown). Further studies
will be undertaken to better understand the behaviour of
benzo[a]phenoxazine derivatives with long alkyl (>8 carbons) side
chains.
Acknowledgements
Thanks are due to the ‘Fundação para a Ciência e Tecnologia’
(Portugal) for its financial support to the ‘Centro de Química’ and
‘Centro de Física’ (Universidade do Minho), as well as a Post-PhD
grant to Sarala Naik (SFRH/BPD/37840/2007).
References and notes
1. Neuhaus, W.; Trzeciak, J.; Lauer, R.; Lachmann, B.; Noe, C. R. J. Pharm. Biomed.
Anal. 2006, 40, 1035–1039.
2. Sevestre, A.; Charmantray, F.; Hélaine, V.; Lásiková, A.; Hecquet, L. Tetrahedron
2006, 62, 3969–3976.
3. Gonçalves, M. S. T. Chem. Rev. 2009, 109, 190–212.
4. Frangioni, J. V.; Onishi, S. U.S. Patent 082423A2, 2005.

4474
C. M. A. Alves et al. / Tetrahedron Letters 50 (2009) 4470–4474
5. Jose, J.; Burgess, K. Tetrahedron 2006, 62, 11021–11037.
44.40 (NHCH2CH3), 92.44 (C-6), 93.22 (C-8), 123.46 (Ar-C), 123.99 (C-1), 125.54
(C-4), 127.06 (C-10), 129.66 (Ar-C), 129.94 (C-3), 130.57 (Ar-C), 130.98 (C-11),
131.61 (C-2), 133.48 (Ar-C), 146.72 (Ar-C), 150.57 (Ar-C), 154.03 (C-9), 156.71
(C-5) ppm. The assignments were supported by HMBC and HMQC techniques.
HRMS: m/z (FAB): calcd for C27H34N3O [M+] 416.2702; found 416.2708.
12. Crossley, M. L.; Turner, R. J.; Hofmann, C. M.; Dreisbach, P. F.; Parker, R. P. J. Am.
Chem. Soc. 1952, 74, 578–584.
13. Typical procedure for the synthesis of compounds 3a–c (described for 3b): To a
solution of 1-naphthylamine (1.0 g; 6.98 ꢁ 10ꢀ3 mol) in ethanol (2 mL), 1-
bromooctane (1.97 mL; 9.81 ꢁ 10ꢀ3 mol) was added and the resulting mixture
was refluxed for 33 h 30 min., and monitored by TLC (silica: chloroform). The
solvent was removed under reduced pressure and the crude mixture was
purified by dry chromatography on silica gel using chloroform/n-hexane, 1:1).
N-Octylnaphthalen-1-amine 3b was obtained as a brown oil (0.455 g, 51%).
6. Pozharski, E. V.; Macdonald, R. C. Anal. Biochem. 2005, 341, 230–400.
7. Tanford, C. In The Hydrophobic Effect: Formation of Micelles and Biological
Membranes; John Wiley & Sons: New York, 1980.
8. Frade, V. H. J.; Gonçalves, M. S. T.; Coutinho, P. J. G.; Moura, J. C. V. P. J.
Photochem. Photobiol. A. 2007, 185, 220–230.
9. Frade, V. H. J.; Barros, S. A.; Moura, J. C. V. P.; Coutinho, P. J. G.; Gonçalves, M. S.
T. Tetrahedron 2007, 63, 12405–12418.
10. Coutinho, P. J. G.; Castanheira, E. M. S.; Rei, M. C.; Oliveira, M. E. C. D. R. J. Phys.
Chem. B 2002, 106, 12841–12846.
11. Typical procedure for the synthesis of compounds 1a–g (described for 1b): To a
cold solution (ice bath) of 5-ethylamino-4-methyl-2-nitrosophenol
hydrochloride 2a (0.087 g; 4.82 ꢁ 10ꢀ4 mol) in ethanol (2 mL), N-
octylnaphthalen-1-amine (3b) (0.307 g; 1.20 ꢁ 10ꢀ3 mol) and concentrated
hydrochloride acid (5.0 ꢁ 10ꢀ2 mL) were added. The mixture was refluxed for
4 h and monitored by TLC (silica: dichloromethane/methanol 59:1). The
solvent was removed under reduced pressure and the crude mixture was
purified by dry chromatography on silica gel using dichloromethane/ n-hexane
and dichloromethane/methanol, mixtures of increasing polarity as the eluent.
N-[10-methyl-5-(octylamino)-9H-benzo[a]phenoxazin-9-ylidene]ethanaminium
chloride (1b) was obtained as a blue solid (0.49 g, 98%). Mp 147.5–149.0 °C.
Rf = 0.61 (silica: chloroform/n-hexane, 1:2). FTIR (neat):
2925, 2855, 2469, 1625, 1605, 1582, 1531, 1465, 1410, 1377, 1344, 1282, 1254,
1216, 1173, 1142, 1081, 1016, 953, 861, 800, 770, 724, 666 cmꢀ1 1Y NMR
mmax 3389, 3058, 2956,
.
(CDCl3, 300 MHz): d 0.97 (3H, t J 7.2 Hz, CH3), 1.30–1.60 (10 H, m, 5 ꢁ CH2),
1.78–1.90 (2H, m, NHCH2CH2), 3.33 (2H, t J 7.5 Hz, NHCH2CH2), 6.85 (1H, d J
7.2 Hz, 4-H), 7.35 (1H, d J 8.1 Hz, 2-H), 7.45 (1H, t J 8.1 Hz, 3-H), 7.38–7.56 (2H,
m, 6-H and 7-H), 7.80–7.88 (1H, m, 8-H), 7.90–7.98 (1H, m, 5-H) ppm. 13C NMR
(CDCl3, 75.4 MHz): d 13.99 (CH3), 22.50 (2 ꢁ CH2), 26.86 (CH2), 27.13 (CH2),
29.03 (CH2), 31.62 (CH2), 49.08 (NHCH2CH2), 114.57 (C-4), 120.81 (C-5), 124.75
(C-7), 125.65 (C-2), 126.17 (C-4a), 126.47 (C-3), 126.57 (C-6), 128.73 (C-8),
134.33 (C-8a), 136.21 (C-1) ppm. The assignments were supported by HMBC
and HMQC techniques. HRMS: m/z (EI): calcd. for C18H25N [M+] 255.1987;
found 255.1985.
Rf = 0.45 (silica: dichloromethane/methanol, 5:1). FTIR (KBr):
2853, 1642, 1592, 1561, 1544, 1519, 1498, 1451, 1435, 1315, 1295, 1259, 1232,
1185, 1163, 1129, 1085, 1054, 1010, 881, 817, 773, 733, 708, 666 cmꢀ1 1Y NMR
(CDCl3, 300 MHz): d 0.86 (3H, t J 6.6 Hz, NH(CH2)7CH3), 1.20–1.50 (13 H, 2 ꢁ m,
5 ꢁ CH2 and NHCH2CH3), 1.85 (2H, br s, NHCH2CH2), 2.45 (3H, s, CH3), 3.20–
3.30 (2H, m, NHCH2CH2), 3.57 (2H, br s, NHCH2CH3), 6.30 (2H, br s, 8-H and 6-
H), 7.49 (1H, s, 11-H), 7.87 (2H, br s, 2-H and 3-H), 8.80–8.90 (1H, m, 1-H), 9.18
(1H, br s, 4-H), 10.81 (1H, br s, NH) ppm. 13C NMR (CDCl3, 75.4 MHz): d 13.95
(NHCH2CH3), 14.03 (NH(CH2)7CH3), 18.60 (CH3), 22.55 (CH2), 27.12 (CH2),
28.65 (NHCH2CH2), 29.14 (CH2), 29.24 (CH2), 31.71 (CH2), 38.72 (NHCH2CH2),
mmax 3195, 2925,
.
14. Sens, R.; Drexhage, K. H. J. Lumin. 1981, 24, 709–712.
15. Gharibi, H.; Razavizadeh, B. M.; Hashemianzaheh, M. Colloid Surf. A-
Physicochem. Eng. Asp. 2000, 174, 375–386.
16. Paredes, S.; Sepulveda, L.; Tribout, M. J. Phys. Chem. 1984, 88, 1871–1875.