Synthesis and fluorescence properties of benzoxazole-1,4-dihydropyridine dyads achieved by a multicomponent reaction

Article abstract of DOI:10.1039/c4nj00777h

Photoactive 2-(2′-hydroxyphenyl)benzoxazole-1,4-dihydropyridine (HBO-DHP) dyads were obtained by a multicomponent one-pot Hantzsch synthesis using a fluorescent aldehyde, a 1,3-dicarbonylic compound and ammonium acetate. The key step in this synthetic methodology was the synthesis of the formyl benzoxazole derivative through a Duff-modified functionalization protocol. UV-Vis absorption and fluorescence emission spectroscopies were also applied to better understand the photophysics of these compounds. The three novel fluorescent compounds were obtained in moderate yields as stable solids with absorption in the UV region and emission in the blue-green region. Preliminary results indicate that after excitation both HBO and DHP fluorophores behave independently in the HBO-DHP structure. the Partner Organisations 2014.

Full text of DOI:10.1039/c4nj00777h

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Synthesis and fluorescence properties of
benzoxazole-1,4-dihydropyridine dyads achieved
Cite this:DOI: 10.1039/c4nj00777h
by a multicomponent reaction†
Ricardo Ferreira Affeldt,a Antonio Cesar de Amorim Borges,a Dennis Russowsky*b
ˆ
´
and Fabiano Severo Rodembusch*a
Photoactive 2-(20-hydroxyphenyl)benzoxazole-1,4-dihydropyridine (HBO–DHP) dyads were obtained by a
multicomponent one-pot Hantzsch synthesis using a fluorescent aldehyde, a 1,3-dicarbonylic compound
and ammonium acetate. The key step in this synthetic methodology was the synthesis of the formyl
Received (in Montpellier, France)
13th May 2014,
benzoxazole derivative through
a Duff-modified functionalization protocol. UV-Vis absorption and
Accepted 2nd July 2014
fluorescence emission spectroscopies were also applied to better understand the photophysics of these
compounds. The three novel fluorescent compounds were obtained in moderate yields as stable solids
with absorption in the UV region and emission in the blue-green region. Preliminary results indicate that
after excitation both HBO and DHP fluorophores behave independently in the HBO–DHP structure.
DOI: 10.1039/c4nj00777h
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Introduction
The multicomponent synthesis is an important tool for the
construction of compound libraries with high atom economy,
allowing a large number of derivatives from combinations of
different reagents, also being explored by the combinatorial
chemistry field.1 In contrast to the multistep linear strategy,
these one-pot reactions are versatile and efficient at achieving
small molecules for biological screening and drug design.2
1,4-Dihydropyridines (1,4-DHP) 1 are small molecules that
exhibit pronounced biological activity and are synthesized by the
Hantzsch multicomponent reaction.3 Their activity as calcium
channel modulators has important consequences on the treatment
of heart diseases such as hypertension and angina.4 Other
promising activities include their role as antioxidants, broncho-
dilators, antiartherosclerotic and AD therapeutic agents.5
The scope of the Hantzsch reaction is not restricted to
1,4-dihydropyridine synthesis only, but polycyclic derivatives of
Scheme 1 Chemical structure of the dihydropyridine core 1, polyhydro-
quinoline 2, polyhydroacridinedione 3 and NADH 4.
the aromatic analogues can also be obtained, such as polyhydro- The NADH coenzyme system 4 is structurally related to 1,4-dihydro-
quinolines 2 and polyhydroacridinedione 3 (Scheme 1).6 pyridine compounds found in living cells and is responsible for
electron transport and energy production in metabolic reactions. It
possesses two oxidation states – i.e. NAD+ and NADH – that differ in
a
´
ˆ
´
Grupo de Pesquisa em Fotoquımica Organica Aplicada, Instituto de Quımica/
UFRGS, Av. Bento Gonçalves, 9500. CEP 91501-970, Porto Alegre, RS, Brazil.
E-mail: fabiano.rodembusch@ufrgs.br
one proton and two electrons. NADH is also a powerful natural
antioxidant. The different photophysical behaviours of the two
species make them useful for protein identification in enzymatic
colorimetric essays.7 Additionally, it can also be applied as a
fluorescence sensor for identification of explosives8 and monitoring
of biological reactions and microbial fermentations.9
b
´
´
ˆ
´
Laboratorio de Sıntese Organica, Instituto de Quımica/UFRGS, Av. Bento
Gonçalves, 9500. CEP 91501-970, Porto Alegre, RS, Brazil.
E-mail: dennis@iq.ufrgs.br
† Electronic supplementary information (ESI) available: Spectral and HRMS data
for the new compounds, as well as additional data for fluorophore 9, and details
of supplementary photophysical experiments and attempts to achieve the quino-
line synthesis and salicylic acid formylation. See DOI: 10.1039/c4nj00777h
We have recently described a photophysical study of 4-aryl-
substituted dihydropyridines allied with theoretical calculations,
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which showed that these systems are close to the well-known paper also deals with the photophysical characterization of
NADH. Their fluorescence emission can be ascribed to a normal the new compounds by means of UV-Visible absorption and
relaxation process or an intramolecular charge transfer in fluorescence emission spectroscopies in solution.
dimethylamino-substituted compounds.10 Albini et al. have also
studied the mechanistic pathways for the excited state of these
dihydropyridines by ultraviolet absorption, fluorescence and
Experimental
General procedures
phosphorescence spectroscopies, leading to different roles of
substituted 1,4-dihydropyridines.11–14 It is desirable that a
All reagents and solvents were purchased from commercial sup-
fluorophore shows fluorescence in a well-defined region of
pliers and used without further purification. Reactions were
monitored using thin-layer chromatography (TLC) carried out on
silica gel 60 F254 pre-coated aluminium plates. The visualization
the electromagnetic spectrum and a high value of Stokes’ shift,
i.e. a large separation between emission and absorption maxima
wavelengths, since the interaction with biological systems can
was achieved under UV light (254 nm) or by staining with I2.
result in a hypsochromic shift of fluorescence favoring self-
absorption and fluorescence quenching.15
Chromatographic separations were achieved on silica gel columns
(70–230 mesh). An ultrasound bath of 40 kHz (50 W) was used for
Hydroxyphenylbenzazole heterocycles assume an important
mixture homogenization. The 1H and 13C NMR spectra were
role as wide-application dyes, hence they show a high Stokes’
recorded in CDCl3 at 300 MHz and 75.4 MHz, respectively. The
shift due to an excited state intramolecular proton transfer
chemical shifts (d) are reported in parts per million (ppm) relative
(ESIPT) mechanism (Fig. 1), a phototautomerization strongly
1
to TMS (0.00 ppm) for H NMR and to the central line of CDCl3
influenced by the solvent, in which usually a dual fluorescence
(77.0 ppm) for 13C NMR. Coupling constants J are reported in hertz
emission takes place in solution or in the solid state.16 This
(Hz). The following abbreviations are used for the multiplicities: s,
mechanism can be related to a normal species (or enol band)
singlet; d, doublet; dd, double-doublet; dq, double-quartet; t,
and a keto tautomer (or ESIPT band). This behavior confers to
triplet; q, quartet; m, multiplet; br s, broad signal. FTIR spectra
these compounds physical and chemical properties that are
were obtained using KBR pellets with a resolution of 4 cmꢀ1
attractive from both synthetic and technological points of
between 400 and 4000 cmꢀ1. Fragmentation patterns were
view.17 Fluorescence properties of 1,4-dihydropyridines could be
obtained using a quadrupole analyzer equipped with a 70 eV
explored for determination and characterization of pharmacological
electron impact ionization source by direct insertion of samples
mechanisms through interactions between molecules and biological
diluted in chloroform (1 mL).
systems, such as membrane calcium channels in cardiac cells with
fluorimetric essays or other specific targets.18
Spectroscopic grade solvents dichloromethane, acetonitrile,
1,4-dioxane and ethanol were used in fluorescence emission
Considering the high Stokes’ shift originating from 2-(20-
and UV-Vis absorption spectroscopy measurements. UV-Vis
hydroxyphenyl)benzoxazole (HBO), we decided to establish a
absorption spectra were recorded using a Shimadzu UV-2450
synthetic methodology for its functionalization to make it a
spectrophotometer, and steady state fluorescence spectra were mea-
suitable component for one-pot synthesis of new 1,4-dihydro-
sured using a Shimadzu spectrofluorometer model RF-5301PC.
pyridine derivatives starting from an ESIPT derivative. This
Spectral correction was performed to enable measurements of true
spectra by eliminating instrumental response such as wavelength
characteristics of the monochromator or the detector using Rhoda-
mine B as an internal standard (quantum counter). The fluorescence
quantum yield (ffl) measurement was made in spectroscopic grade
solvents using the dilute optical methodology. Quinine sulphate
(Riedel) in 1 M H2SO4 (ffl = 0.55) was used as a quantum yield
standard.19 All experiments were performed at room temperature.
High resolution mass spectrometry electrospray ionization (HRMS-
ESI) data, in a positive mode, were collected using a Micromass
Q-Tof instrument. Samples were infused by a 100 mL syringe at a flow
rate 30 mL minꢀ1 for all samples. The following typical operating
conditions were used: a capillary voltage of 2980 V, a sample cone
voltage of 30 V, an extraction cone voltage of 3.0 V, and a desolvation
gas temperature of 60 1C. N2 was used as the desolvation gas and
HPLC grade acetonitrile was the solvent used with the samples.
Syntheses
5-Formylsalicylic acid (6). 22 mL of glacial acetic acid was
added to an equimolar mixture of salicylic acid (5) (14.5 mmol,
Fig. 1 Photophysical pathways for ESIPT-exhibiting dyes: normal (or enol)
2.00 g) and hexamethylenetetramine (14.5 mmol, 2.03 g) in a
100 mL round-bottom vessel and the solution was heated to
emission (green) and ESIPT (or tautomer) emission (red). The asterisk
indicates the excited state.
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reflux for 8 hours, after which 16 mL of hot distilled water and (10 mol%), followed by 2.5 mmol of ethyl acetoacetate (13)
10 mL of concentrated hydrochloric acid were added with (0.317 mL), 1.25 mmol of previously synthesized 9 (0.299 g) and
vigorous stirring. The solution was then cooled to ambient 2.5 mmol of ammonium acetate (12) (0.193 g) were added to a
temperature and the formed precipitate was filtered using a round-bottom vessel. The mixture was dissolved in isopropanol
sintered funnel followed by washings with 50 mL of distilled (10 mL) and refluxed for 3 hours. The solution was cooled and
water (40 1C and ambient temperature). The pale yellow solid the precipitate filtered and washed with cold isopropanol (4 ꢁ
was dried leading to 0.24 g of 5-formylsalicylic acid in 16% yield 5 mL). The catalyst was separated from the product by redissolution
and its characterization is in agreement with the literature.20 of the solid in dichloromethane and micropore (0.45 mm) filtration.
1
M.p. 250 1C (decomp.). H NMR (DMSO-d6, 300 MHz): d = 7.12 The solution obtained was concentrated and dried leading to 0.26 g
3
4
(d, J = 8.5 Hz, 1H); 7.99 (dd, J = 8.5 and J = 2.0 Hz, 1H); 8.34 of the pure product (46%), a white solid with blue fluorescence
(d, J = 2.0 Hz, 1H); 9.84 (s, 1H). 13C NMR (DMSO-d6, 75.4 MHz): when exposed to UV radiation. M.p. = 233–236 1C. 1H NMR (CDCl3,
d = 114.3; 118.9; 128.9; 134.6; 135.8; 166.4; 171.8; 191.9.
300 MHz): d = 1.27 (t, 6H, J = 7.1 Hz); 2.39 (s, 6H); 4.07–4.22 (m, 4H);
2-(20-Hydroxyphenyl)benzoxazole (8). An equimolar mixture 5.05 (s, 1H); 5.75 (s, 1H); 7.00 (d, J = 8.7 Hz, 1H); 7.38–7.4 (m, 3H);
of salicylic acid (5) (18.3 mmol, 2.53 g) and 2-aminophenol (7) 7.60–7.63 (m, 1H); 7.72–7.73 (m, 1H); 7.83 (d, J = 2.1 Hz, 1H). 13C
(18.3 mmol, 2.00 g) in polyphosphoric acid (10 mL) was stirred NMR (CDCl3, 75.4 MHz): d = 14.5; 19.9; 39.3; 60.0; 104.2; 109.8;
at 180 1C for 5 h. After cooling, the mixture was poured onto ice- 110.8; 117.1; 119.3; 125.1; 125.3; 126.6; 133.9; 139.6; 144.1; 149.3;
cold water and the precipitate was filtered, washed with water 157.4; 163.5; 164.9; 167.8. FTIR (KBr, n = cmꢀ1): 3323 (NH), 3096,
and dried in a stove. The solid was purified by column 2980, 1681, 1649, 1548, 1491, 1302, 1212. MS: m/z (%) = 462.2 (10)
chromatography using dichloromethane as an eluant leading [M+], 433.1 (10), 389.3 (19), 252.1 (100), 211.0 (5), 196.0 (25). HRMS
to 2.09 g of the product (54%). M.p. = 125–126 1C. 1H NMR (ESI-qTOF) m/z: [M + H]+ calcd for C26H26N2O6 463.1869; found
4
(CDCl3, 300 MHz): d = 6.89 (t, J = 8.1 Hz, 1H); 7.01 (dd, J = 8.2 463.1865 (Dppm = 0.9).
and 3J = 0.9 Hz, 1H); 7.24–7.35 (m, 3H); 7.46–7.49 (m, 1H); 7.58–
Ethyl 4-[2-(20-hydroxyphenyl)benzoxazolyl]-1,4,5,6,7,8,-hexahydro-
7.62 (m, 1H); 7.90 (dd, 4J = 7.9 and 3J = 1.8 Hz, 1H); 11.37 (br s, 2,7,7-trimethylquinolin-5(1H,4H,6H)-one 3-carboxylate (16). 0.106 g
1H). 13C NMR (CDCl3, 75.4 MHz): d = 110.8; 117.6; 119.4; 119.7; of In/SiO2 (10 mol%), followed by 1.25 mmol of 5,5-dimethyl-
125.1; 125.5; 127.3; 133.7; 140.1; 149.2; 158.9; 163.0; 164.9.
cycloexane-1,3-dione (14) (0.175 g), 1.25 mmol of ethyl aceto-
2-(50-Formyl-20-hydroxyphenyl)benzoxazole (9). Method A acetate (13) (0.158 mL), 1.25 mmol of previously synthesized 9
(cyclization): polyphosphoric acid (10 mL) followed by 6.4 mmol of (0.299 g) and 2.5 mmol of ammonium acetate (12) (0.193 g)
5-formylsalicylic acid (6) (1.06 g) and 6.4 mmol of 2-aminophenol (7) were added to a round-bottom vessel. The mixture was dis-
(0.70 g) were added to a round-bottom flask. The mixture was heated solved in isopropanol (10 mL) and refluxed for 3 hours. The
at 170 1C for 5 hours. The crude mixture was poured onto ice-cold solution was cooled and the precipitate filtered and washed
water with vigorous stirring and the precipitate was filtered with cold isopropanol (4 ꢁ 5 mL). The catalyst was separated
yielding a dark brown solid. The solid was extracted by using a from the product by redissolution of the solid in dichloro-
Soxhlet extractor with dichloromethane and then purified by methane and micropore (0.45 mm) filtration. The solution
column chromatography using dichloromethane as the eluent, obtained was concentrated and dried leading to 0.39 g of the
yielding, after solvent evaporation, 0.46 g of a white solid (30%) pure product (66%), a white solid with green fluorescence when
exhibiting intense green fluorescence when exposed to UV exposed to UV radiation. M.p. 4250 1C. 1H NMR (CDCl3,
radiation. Method B (formylation): 10 mL of polyphosphoric 300 MHz): d = 0.96 (s, 3H, CH3); 1.09 (s, 3H, CH3); 1.23 (t, J =
acid (10 mL) followed by 4.7 mmol of previously synthesized 7.1 Hz, 3H, CH3); 2.14–2.40 (m, 4H, 2CH2); 2.4 (s, 3H, CH3); 4.08
HBO (8) (0.99 g) and 5 mmol of hexamethylenetetramine (0.70 g) (dq, J = 1.1 Hz and J = 7.1 Hz, 2H, CH2); 5.08 (s, 1H); 5.97 (s, 1H);
were added to a round-bottom flask. The temperature was raised 6.97 (d, J = 8.7 Hz, 1H); 7.34–7.39 (m, 3H); 7.58–7.61 (m, 1H);
to 100 1C and the mixture stirred for 4 hours. After the heating, 7.69–7.72 (m, 1H); 7.99 (d, J = 2.4 Hz, 1H); 11.34 (br s,1H). 13C
30 mL of ice-cold water was added leading to a white precipitate, NMR (CDCl3, 75.4 MHz): d = 14.2; 19.5; 27.1; 29.4; 32.7; 35.9;
which was then filtered and washed with water before drying. 41.1; 50.6; 59.9; 106.0; 109.6; 110.6; 112.0; 117.0; 119.1; 124.8;
The solid was purified by column chromatography using dichloro- 125.1; 126.5; 133.5; 138.6; 140.1; 143.4; 147.9; 149.1; 157.1;
methane as an eluant leading to 0.38 g of the product (34%). White 163.2; 167.3; 196.0. FTIR (KBr, n = cmꢀ1): 3289, 3222, 2963,
needles crystals. M.p. = 165–170 1C. 1H NMR (CDCl3, 300 MHz): d = 1706, 1608, 1487, 1379, 1262. MS: m/z (%) = 472 (39) [M+], 399
7.22 ppm (d, 2H, J = 8.6 Hz); 7.39–7.45 (m, 2H); 7.62–7.66 (m, 1H); (20), 262 (100), 234 (28). HRMS (ESI-qTOF) m/z: [M + H]+ calcd
7.73–7.76 (m, 1H); 7.96 (dd, 3J = 8.6 Hz and 4J = 2.0 Hz); 8.55 (d, J = for C26H28N2O5 473.2076; found 473.2058 (Dppm = 3.8).
2.0 Hz); 9.95 (s, 1H); 12.17 (br s, 1H). 13C NMR (CDCl3, 75.4 MHz):
3,3,6,6-Tetramethyl-9-[2-(20-hydroxyphenyl) benzoxazolyl]-3,4,6,7-
d = 111.2; 118.6; 119.7; 125.7; 126.3; 129.2; 130.5; 134.3; 139.7; 149.4; tetrahydro acridine-1,8-(2H,5H,9H,10H)-dione (17). 0.106 g of In/
161.9; 163.7; 165.0; 190.2. FTIR (KBr, n = cmꢀ1): 3061, 2848, 2735, SiO2 (10 mol%), followed by 2.5 mmol of 5,5-dimethylcyclohexane-
1699, 1630, 1490. MS: m/z (%) = 240.1 (16), 239.1 (100) [M+], 238.1 1,3-dione (14) (0.350 g), 1.25 mmol of previously synthesized 9
(90), 210.2 (10), 182.0 (29), 63.1 (32). HRMS (ESI-qTOF) m/z: [M + H]+ (0.299 g) and 2.5 mmol of ammonium acetate (12) (0.193 g) were
calcd for C14H9NO3 240.0660; found 240.0620 (Dppm = 4.2).
added to a round-bottom vessel. The mixture was dissolved in
Diethyl 2,6-dimethyl-4-[2-(20-hydroxyphenyl) benzoxazolyl]- isopropanol (10 mL) and refluxed for 3 hours. The solution was
1,4-dihydropyridine 3,5-dicarboxylate (15). 0.106 g of In/SiO2 cooled and the precipitate filtered and washed with cold isopropanol
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(4 ꢁ 5 mL). The catalyst was separated from the product by
redissolution of the solid in dichloromethane and micropore
(0.45 mm) filtration. The solution obtained was concentrated and
dried leading to 0.30 g of the pure product (50%), a white solid with
1
blue fluorescence when UV exposed. M.p. = 212–215 1C. H NMR
(CDCl3, 300 MHz): d = 1.15 (s, 6H, 2CH3); 1.35 (s, 6H, 2CH3); 2.33–
2.55 (m, 8H, 4CH2); 5.53 (s, 1H); 7, 03 (d, J = 8.7 Hz, 1H); 7. 16 (d, J =
8.7 Hz); 7.36–7.38 (m, 2H); 7.48–7.51 (m, 1H); 7.71–7.74 (m, 1H);
7.78–7.81 (m, 1H); 11.23 (br s, 1H, NH); 12.01 (s, 1H, OH). 13C NMR
(CDCl3, 75.4 MHz): d = 27.4; 30.2; 31.6; 32.3; 48.7; 47.3; 110.6; 115.7;
117.5; 119.5; 125.2; 125.5; 126.8; 129.2; 132.4; 140.4; 149.2; 157.0;
163.1; 165.0; 169.7; 180.9. FTIR (KBr, n = cmꢀ1): 2958, 2925, 1593,
1372. MS: m/z (%) = 361.1 (100) [M+], 318.0 (8), 277.0 (16), 263.0 (9),
249.0 (30), 140.0 (13), 83.0 (49). HRMS (ESI-qTOF) m/z: [M + H]+ calcd
for C30H30N2O4 483.2284; found 483.2293 (Dppm = 1.9).
Fig. 2 Proposed HBO–DHP structure.
Results and discussion
Synthesis
We first attempted to apply aminobenzazoles on benzoxazolyl-
quinoline synthesis without success using multicomponent
and even classical cyclization protocols involving anilines, such
as Skaup glycerol acidic reactions and Combes, Doebner-Miller
and Conrad-Limpach approaches (see ESI†).
Therefore it was decided to bypass this drawback by preparing a
formyl derivative which has, to the best of our knowledge, not been
described in the literature, although 3,5-formyl-hydroxyphenyl-
benzoxazole has been recently described as an intermediate for a
fluorescent pyrophosphate sensor.21 In Hantzsch multicomponent
condensation giving 1,4-dihydropyridines, it has been observed that
aromatic and heteroaromatic aldehydes bearing both electron-
withdrawing as well as electron-releasing substituted groups do
not significantly affect the reaction yields.22 On the other hand,
the synthesis of 2-(20-hydroxyphenyl)benzoxazole (HBO) is well
established in the literature.23 In this way, a synthetic method
Scheme 2 Reaction pathways for obtaining formyl functionalized HBO
(9). Conditions: (i) Hexamethylenetetramine (HMTA), acetic acid (AcOH),
reflux, 8 h; (ii) and (iii) 7, polyphosphoric acid (PPA), 170 1C, 5 h, (iv) HMTA,
PPA, 100 1C, 4 h.
was proposed to afford functionalization of HBO by introducing mixture of the 3-formyl and 5-formyl derivatives (11 and 6, respec-
a formyl group, which was predicted to react with two equiva- tively) could be obtained, whose separation was quite difficult.
lents of a 1,3-dicarbonyl compound and ammonia, aiming for Therefore, a simple procedure was chosen: filtering the solid pre-
the synthesis of fluorescent dyad HBO–DHP, the structure of cipitate after hydrolysis and washing with hot water portions. This
which is depicted in Fig. 2.
method has proved to be useful since 5-formylsalicylic acid (6) is less
In this way, as depicted in Scheme 2, two different methodologies soluble in water than 3-formylsalicylic acid (11); however, product 6
(routes A and B) were used to synthesize the formyl functionalized was obtained in low yield (16%).
fluorophore 9. Duff’s formylation for salicylic acid (5) using hexam-
1H-NMR analysis of the pale yellow precipitate produced a
ethylenetetramine (HMTA) involves stable and low-cost reagents, spectrum in agreement with that of pure 5-formylsalicylic acid
although low yields are usually obtained.20 The procedure was (6). Pure 3-formylsalicylic acid (11) could also be obtained by
employed for salicylic acid (5) and later for condensation of recrystallization (m.p. = 178–179 1C). It is worth noting that no
5-formylsalicylic acid (6) with 2-aminophenol (7). At the same formylated product was observed when the reaction was carried
time, formylation of HBO 8 obtained by condensation of in pure water up to 16 hours.
salicylic acid (5) and 2-aminophenol (7) was tested.
The condensation procedure to yield the novel fluorescent
It was expected that the rate of Duff’s formylation reaction formylated compound 9 was adapted from the literature,23 i.e.
was highly dependent on the successful hydrolysis of iminium reacting the previously synthesized 5-formylsalicylic acid (6)
cation 10 formed by protonated HMTA and the activated and 2-aminophenol (7) in polyphosphoric acid (PPA) at 170 1C
phenol derivative in acidic media (Scheme 3). We performed for 5 hours. The product was purified by column chromatography
this reaction by refluxing salicylic acid (5) and an excess of yielding (9) (yield 30%), a white solid with green fluorescence, quite
HMTA in acetic acid for 8 hours. After several attempts, a similar to the unfunctionalized HBO fluorophore 8. It was expected
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Scheme 3 Iminium hydrolysis in Duff’s formylation of salicylic acid (5).
that an inverse procedure starting with the HBO synthesis and its
further formylation could improve the yields in both reaction steps
(Scheme 2, route B).
Precursor 8 was prepared as reported in the literature24 (yield
57%) and the formylation methodology was investigated in three
acidic solvents. The results are summarized in Table 1. PPA was
found to be suitable for both steps of cyclization and formylation.
The structure of 9 was confirmed by 1H and 13C NMR, FTIR and MS
analyses (see ESI†). The signal pattern in the region between 7 and
9 ppm in the 1H NMR spectrum (Fig. 3) was crucial for the
identification of the formyl group in the 50 position of the phenolic
ring. The novel fluorophore precursor shows good thermal stability
and no decomposition was observed up to 250 1C.
The Hantzsch three-component synthesis of 4-(2-(20-hydroxy-
phenyl)benzoxazolyl)-1,4-dihydropyridine 15 was performed by
mixing one equivalent of 9 with ammonium acetate (12) and two
equivalents of ethyl acetoacetate (13), employing a In/SiO2
heterogeneous Lewis-acid catalyst in refluxing isopropanol
(Scheme 4).22 The product was isolated after 3 hours as a blue
fluorescent solid in moderate yield (46%) by simply filtering the
heterogeneous isopropanol mixture. It is worth mentioning that
when acetylacetone was used, a complex mixture was obtained.
Considering the antimicrobial activity of decahydroacridine-
diones and hexahydroquinolines with a fused dimedone
Scheme 4 Multicomponent synthesis of HBO–DHPs 15–17.
moiety,25 for further investigations, two N-heterocycles, 16
and 17, bearing the HBO moiety and the DHP core were
synthesized using the same methodology. The compounds were
obtained with significant yields (66% and 50% for 16 and 17,
respectively) without any additional chromatographic purification.
All synthesized novel heterocycles were white solids and fluorescent
in the blue-green region when exposed to UV 365 nm radiation
(solid or in solution).
Photophysics
The photophysical study of the synthesized compounds was
performed in solution (10ꢀ5 M) using four different organic
solvents with a wide range of dielectric constants. This investigation
was focused on the influence of the structural changes, as well as the
oxidation state of the dihydropyridine core in the ESIPT emission.
Fig. 4 presents the UV-Vis absorption and fluorescence emission
spectra of 9. The fluorescence emission spectra were obtained using
the absorption maxima as the excitation wavelengths. The relevant
photophysical data are summarized in Table 2.
Table 1 Experimental conditions for the HBO’s formylation
Entry Solvent
Time (h) Temp. (1C) Yield (%)
1
2
3
Acetic acid (AcOH)
Trifluoroacetic acid (TFAA)
Polyphosphoric acid (PPA)
6
6
4
Reflux
Reflux
100
8
15
34
The UV-Vis absorption spectra of 9 in all solvents exhibit an
intense band located at 266 nm and a red-shifted one at 320–335 nm
with molar absorptivity coefficient, e, values in agreement with p–p*
transitions. No significant solvatochromism in the ground state was
observed for this dye.
The fluorescence emission spectra show a major emission
band located at 474–483 nm. As already presented in the
literature, the measured Stokes’ shift (B9150 cmꢀ1) can be
ascribed to the ESIPT mechanism.17 Due to this dye the ground-
state enol-cis conformer (N) absorbs UV radiation leading to the
singlet excited state enol-cis (N*), which tautomerizes to the
excited-state keto (T*). This conversion is responsible for energy
loss with a large Stokes’ shift, and the excited keto decays to the
ground state (T) emiting fluorescence. The single red-shifted
emission band ascribed to (T)S0 ’ (T*)S1 and consequently the
absence of a blue-shifted band from the locally excited enol
species (N)S0 ’ (N*)S1 is quite similar to that of the HBO
Fig. 3 1H NMR spectrum of 9.
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Fig. 4 Normalized UV-Vis absorption and fluorescence emission spectra
of 9 in different solvents.
Table 2 Relevant UV-Vis absorption and fluorescence emission data of 9,
where labs and lem are the absorption and emission maxima, respectively
in nanometers, and e is the molar extinction coefficient in 103 L molꢀ1 cmꢀ1
DlST (cmꢀ1
)
a
Solvent
labs (e)
lem
CHCl3
MeCN
EtOH
1,4-Dioxane
334(9.68), 272(19.1)
330(13.3), 266(28.9)
332(10.9), 269(18.9)
333(13.6), 265(28.5)
474
478
475
483
8843
9382
9068
9326
a
Stokes’ shift was obtained from the difference of the emission and
absorption maxima (DlST = lem ꢀ labs).
spectra.24 These results strongly suggest that the formyl group
in the 50-position of the phenolic ring does not favour an
equilibrium between the conformers in solution in the ground
state, as observed for amino derivatives in the same position.16
Excitation spectra show similar profiles to the absorption
curves (not shown, see ESI†), where the structured bands
ascribed to the HBO (322 and 334 nm) and benzoxazolyl
moieties (B270 nm) can be observed.
Fig. 5 Normalized UV-Vis absorption and fluorescence emission spectra
of 15–17 in different solvents.
Fig. 5 presents the absorption and emission spectra of HBO–
DHPs 15–17 in solution. The relevant data from the photophysical
study of these compounds are summarized in Table 3.
the so-called short wavelength (SW) and long wavelength (LW),
The compounds exhibit absorption maxima in the UV region respectively. In these compounds, the fluorescence emission
(334–340 nm) with electronic transitions ascribed to p–p*. can be influenced by internal electron transfer and/or the
Changes in the solvent polarity, where only a slight solvato- energy transfer character of the corresponding DHP core
chromism was observed, as well as modifications in the DHP (blue-shifted band)11–14 or by deactivation of the keto tautomer,
core, seem not to affect the photophysics of these compounds which arises from the ESIPT mechanism (red-shifted band). It
in the ground state. The fluorescence emission was also is worth mentioning that the normal emission from enol
obtained using the absorption maxima as the excitation wave- conformers was discarded since the measured Stokes’ shifts
lengths. The compounds in the solid state exhibit emission in was much higher (B5200–7000 cmꢀ1) than those attributed to
the blue-green region (Fig. 6).
this emission in similar compounds (B3500 cmꢀ1).16,17 An
Although ESIPT emission could be observed in the studied exception can be observed in the case of compound 17 in
HBO–DHPs, as already observed for compound 9, the fluores- acetonitrile, where the blue-shifted band could be related to
cence spectra of compounds 15–17 seem to be more complex the normal emission. The nature of the emission spectra of
being dependent on both the solvent polarity and the DHP these compounds indicates that both HBO and DHP behave,
moiety. Generally, an emission spectrum presents a dual after excitation, as independent fluorophores in the HBO–DHP
fluorescence emission located at around 425 nm and 488 nm, structure. Concerning the intricate photophysics of the DHPs,11–14
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Table 3 Relevant UV-Vis absorption and fluorescence emission data of the
HBO–DHPs 15–17, where the concentration is B10ꢀ6 mol Lꢀ1, labs and lem
are the absorption and emission maxima, respectively in nanometers, DlST is
the Stokes’ shift, e is the molar extinction coefficient in 103 L molꢀ1 cmꢀ1 and
acetate. The first attempt at using different ESIPT fluorescent amino
derivatives as nitrogen sources did not provide the desired
cyclization compounds. A new photoactive aldehyde derivative
of 2-(20-hydroxyphenyl)benzoxazole was successfully obtained
through a Duff-modified functionalization protocol.
The new HBO–DHPs were obtained in moderate yields as
thermal and photostable solids. The photophysical study in
solution shows an absorption maximum in the UV region and
fluorescence emission in the blue-green region depending on
the compound. Preliminary results indicate that after excitation
both HBO and DHP behave as independent fluorophores in the
HBO–DHP structure. Since a solid synthetic methodology to
incorporate ESIPT compounds into the DHP structures could
be established, additional experiments are in progress to better
understand the photophysics of these dyes.
ffl is the fluorescence quantum yield
lem
DlST (cmꢀ1
)
Solvent
labs (e)
SW
LW
SW
LW
ffl
15
CHCl3
MeCN
EtOH
339(24.1)
336(10.2)
336(15.5)
339(24.5)
424
433
441
426
483
—
—
5914
—
—
8795
6667
7086
8880
0.19
0.07
0.11
0.11
1,4-Dioxane
485
6024
16
CHCl3
MeCN
EtOH
339(24.1)
336(10.2)
336(15.5)
339(24.5)
424
433
441
426
483
—
—
5914
—
—
8795
6667
7086
8880
0.31
0.08
0.08
0.11
1,4-Dioxane
485
6024
17
Acknowledgements
CHCl3
MeCN
EtOH
340(8.17)
337(13.7)
334(21.3)
339(23.9)
—
382
—
499
492
484
506
—
3456
—
7140
9348
9279
9736
0.35
0.23
0.33
0.17
The authors thank the Conselho Nacional de Desenvolvimento
1,4-Dioxane
—
—
´
´
Cientıfico e Tecnologico (CNPq) and the Instituto Nacional de
˜
´
Inovaçao em Diagnosticos para Sau´de Pu´blica (INDI-Sau´de) for
financial support.
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