Spontaneous Formation of Fluorescent Nanofibers from Self-Assembly

Feb 1, 2008 - Marion Mille,†,‡ Jean-François Lame`re,†,‡ Fernanda Rodrigues,†,‡ and. Suzanne Fery-Forgues*,†,‡. UniVersite´ Paul Sab...
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Langmuir 2008, 24, 2671-2679

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Spontaneous Formation of Fluorescent Nanofibers from Self-Assembly of Low-Molecular-Weight Coumarin Derivatives in Water Marion Mille,†,‡ Jean-Franc¸ ois Lame`re,†,‡ Fernanda Rodrigues,†,‡ and Suzanne Fery-Forgues*,†,‡ UniVersite´ Paul Sabatier, Laboratoire des Interactions Mole´ culaires Re´ actiVite´ Chimique et Photochimique, and CNRS UMR 5623, 31062 Toulouse cedex 9, France ReceiVed July 20, 2007. In Final Form: NoVember 28, 2007 A solvent-exchange process was applied to three fluorescent dyes belonging to the 2-benzimidazolyl-7diethylaminocoumarin series (namely Coumarin 7 (1), Coumarin 30 (2), and one of their derivatives bearing a butyl chain (3)). The three compounds only differ by the substitution of the nitrogen atom of the benzimidazolyl group. They were first dissolved in acetone and then suddenly placed in an aqueous environment where they generated molecular assemblies. The size and shape of the latter were studied by fluorescence microscopy and transmission and scanning electron microscopy. It appeared that 1 gave aggregates and flat microcrystals that evolved toward elongated structures. 2 formed straight nanofibers that are 10-20 µm long and exhibit a crystal structure. 3 gave short fibers (1 µm × 25 nm), which finally arrange into entangled solid nanofibers. The formation of fibers arising from lowmolecular-weight molecules was particularly interesting. The optical properties of the free-standing particles in suspension were analyzed and compared to those of highly dilute dyes, with the aim to get additional information about the dye arrangement in the nanostructures.

Introduction Owing to their unique spectroscopic and photophysical properties, highly organized assemblies of organic dyes have received increasing attention. For instance, fluorescent nanocrystals, nanofibers, and π-conjugated polymers are now considered to be potential functional materials for photovoltaics, optoelectronics,1-3 and optics.4 They could also find applications such as photosensitizers5 and fluorescent sensors for the analysis of biological molecules and pollutants by fluorimetry.6 A pragmatic way to obtain well-defined nano-objects is to exploit the self-organization properties of the dye molecules. However, predicting the type of molecular assembly that will be obtained * To whom correspondence should be addressed. Phone: +33 5 61 55 68 05. Fax: +33 5 61 55 81 55. E-mail: [email protected]. † Universite Paul Sabatier. ‡ CNRS UMR 5623. (1) (a) Yanagi, H.; Ohara, T.; Morikawa, T. AdV. Mater. 2001, 13, 1452. (b) Yanagi, H.; Morikawa, T. Appl. Phys. Lett. 1999, 75, 187. (2) (a) Laquai, F.; Wegner, G.; Im, C.; Bu¨sing, A.; Heun, S. J. Chem. Phys. 2005, 123, 074902. (b) Chen, C. T. Chem. Mater. 2004, 16, 4389-4400. (3) Swanson, S. A.; Wallraff, G. M.; Chen, J. P.; Zhang, W.; Bozano, L. D.; Carter, K. R.; Salem, J. R.; Villa, R.; Scott, J. C. Chem. Mater. 2003, 15, 23052312. (4) (a) Lim, S. J.; An, B. K.; Jung, S. D.; Chung, M. A.; Park, S. Y. Angew. Chem., Int. Ed. 2004, 43, 6346-6350. (b) Spagnoli, S.; Block, D.; BotzungAppert, E.; Colombier, I.; Baldeck, P. L.; Ibanez, A.; Corval, A. J. Phys. Chem. B 2005, 109, 8587-8591. (c) Kaneko, Y.; Shimada, S.; Fukuda, T.; Kimura, T.; Yokoi, H.; Matsuda, H.; Onodera, T.; Kasai, H.; Okada, S.; Oikawa, H.; Nakanishi, H. AdV. Mater. 2005, 17, 160-163. (d) Kaneko, Y.; Onodera, T.; Kasai, H.; Okada, S.; Oikawa, H.; Nakanishi, H.; Fukuda, T.; Matsuda, H. J. Mater. Chem. 2005, 15, 253-255. (e) Onodera, T.; Yoshida, M.; Okazoe, S.; Fujita, S.; Kasai, H.; Okada, S.; Oikawa, H.; Nakanishi, H. Int. J. Nanosci. 2002, 1, 737-741. (f) Oikawa, H.; Kasai, H.; Nakanishi, H. In Anisotropic Organic Materials; ACS Symposium Series 798; Glaser, R., Kasizynski, P., Ed.; American Chemical Society: Washington, DC, 2002; Chapter 12, pp 169-178. (5) Kim, H. Y.; Bjorklund, T. G.; Lim, S.-H.; Bardeen, C. J. Langmuir 2003, 19, 3941-3946. (6) (a) Zhou, Y.; Bian, G.; Wang, L.; Dong, L.; Wang, L.; Kan, J. Spectrochim. Acta, Part A 2005, 61, 1841-1845. (b) Wang, L.; Wang, L.; Dong, L.; Bian, G.; Xia, T.; Chen, H. Spectrochim. Acta, Part A 2005, 61, 129-133. (c) Wang, L.; Xia, T.; Wang, L.; Chen, H.; Dong, L.; Bian, G. Microchim. Acta 2005, 149, 267-272. (d) Wang, L.; Wang, L.; Xia, T.; Dong, L.; Bian, G.; Chen, H. Anal. Sci. 2004, 20, 1013-1017. (e) Jinshui, L.; Lun, W.; Feng, G.; Yongxing, L.; Yun, W. Anal. Bioanal. Chem. 2003, 377, 346-349. (f) Botzung-Appert, E.; Monnier, V.; Ha Duong, T.; Pansu, R.; Ibanez, A. Chem. Mater. 2004, 16, 1609-1611.

is still a difficult task, and knowing how the molecular arrangement will influence the spectroscopic and photophysical properties is even more complicated.7 A thorough understanding of the optical behavior of self-assembled systems is therefore of obvious interest. In our team, we have been studying for the past few years the formation of organic microcrystals prepared by a solventexchange process, called the “reprecipitation method”.8 A particular attention was brought to fluorescent dyes of the nitrobenzoxadiazole (NBD) series, which have the distinct advantage to give microcrystals whose size and shape vary strongly according to the experimental conditions.9 In the present work, we would like to extend the reprecipitation method to another series of dyes, to see which type of particles can be formed and which fluorescence behavior can be encountered. We chose to work in the coumarin series, and more particularly with the commercially available Coumarin 7 (1) and Coumarin 30 (2), and with one of their derivatives that was synthesized purposely (3). In these three dyes, the basic chromophore is a coumarin heterocycle substituted by a diethylamino group in the 7-position, and by a benzimidazole group in the 3-position. The nitrogen atom on the benzimidazole ring bears a hydrogen atom in 1, a methyl group in 2, and a butyl chain in 3 (Figure 1). The three dyes are thus made of the same chromophore with very small modifications of the chemical structure. Different reasons motivated the choice of these compounds. (i) First of all, the spectroscopic and photophysical behavior of 1 and 2 in solution (7) (a) Birks, J. B. Photophysics of Aromatic Molecules; Wiley: London, 1970. (b) Silinsh, E. A. Organic Molecular Crystals; Springer-Verlag: Berlin, 1980. (8) Nakanishi, H.; Oikawa, H. In Single Organic Nanoparticles; Masuhara, H., Nakanishi, H., Sasaki, K., Ed.; Springer-Verlag: Berlin, 2003; Chapter 2, pp 17-31. (9) (a) Bertorelle, F.; Lavabre, D.; Fery-Forgues, S. J. Am. Chem. Soc. 2003, 125, 6244-6253. (b) Abyan, M.; Bertorelle, F.; Fery-Forgues, S. Langmuir 2005, 21, 6030-6037. (c) Birla, L.; Bertorelle, F.; Rodrigues, F.; Badre´, S.; Pansu, R.; Fery-Forgues, S. Langmuir 2006, 22, 6256-6265. (d) Bertorelle, F.; Rodrigues, F.; Fery-Forgues, S. Langmuir 2006, 22, 8523-8531. (e) Abyan, M.; Bıˆrlaˇ, L.; Bertorelle, F.; Fery-Forgues, S. C. R. Chim. 2005, 8, 1276-1281. (f) Bertorelle, F.; Al-Ali, F.; Fery-Forgues, S. Int. J. Photoenerg. 2004, 6, 221-225.

10.1021/la702197h CCC: $40.75 © 2008 American Chemical Society Published on Web 02/01/2008

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Figure 1. Chemical structure of compounds 1-3.

are already very well known.10-14 The presence of the benzimidazolyl group increases the molar absorption coefficient and leads to a red-shift of the absorption and emission spectra.10 It also bestows excellent photostability on the dyes.15 All this constitutes a distinct advantage for spectroscopic applications. (ii) Dyes 1 and 2 are hydrophobic, their solubility in water being estimated below 10-5 M.16 Dye 3, with its butyl chain, should be the most hydrophobic of the three. Consequently, these compounds should form aggregates in water. (iii) It is established that their fluorescence efficiency is almost not quenched by water molecules in organic solvents containing up to 50% water.12,14 Moreover, looking at the dyes in their packaging flask reveals that they are highly fluorescent in the solid state. (iv) 1 and 2 have proved to be very useful for applications in fields as varied as the coloring of textile fibers and other materials,17,18 medical imaging,19 lasers,15,20 and photonic devices based on nonlinear optics.21 They are also used as photosensitizers,22 fluorescent probes,16,23 and doping agent for organic blue light-emitting diodes.3,24 For all these reasons, the compounds appeared as excellent candidates for the reprecipitation method, which allows free-standing nano- and microparticles of dyes to be obtained in (10) (a) Christie, R. M.; Lui, C. H. Dyes Pig. 2000, 47, 79-89. (b) Christie, R. M.; Lui, C. H. Dyes Pig. 1999, 42, 85-93. (11) Senthilkumar, S.; Nath, S.; Pal, H. Photochem. Photobiol. 2004, 80, 104111. (12) Jones, G., II.; Jackson, W. R.; Choi, C. Y.; Bergmark, W. R. J. Phys. Chem. 1985, 89, 294-300. (13) Nemkovich, N. A.; Baumann, W.; Reis, H.; Zvinevich, Yu. V. J. Photochem. Photobiol. A 1997, 109, 287-292 and references therein. (14) Raju, B. B.; Costa, S. M. B. Phys. Chem. Chem. Phys. 1999, 1, 35393547. (15) Schinitschek, E. J.; Trais, J. A.; Hammond, P. R.; Henry, R. A.; Atkins, R. L. Opt. Commun. 1976, 16, 313-316. (16) Jones, G., II.; Jimenez, J. A. C. J. Photochem. Photobiol. B 2001, 65, 5-12. (17) Christie, R. M. ReV. Prog. Color. 1993, 23, 1-18. (18) (a) Szuster, L.; Kazmierska, M.; Krol, I. Fiber. Text. East. Eur. 2004, 12, 70-75. (b) Czajkowski, W.; Kazmierska, M. AdV. Colour Sci. Technol. 2003, 6, 91-94. (19) Kudo, K.; Suzuki, M.; Suemoto, T.; Okamura, N.; Shiomitsu, T.; Shimazu, H. Patent no. JP 2004250411. Chem. Abst. 2004, 741782. (20) (a) Tuccio, S. A.; Drexhage, K. H.; Reynolds, G. A. Opt. Commun. 1973, 7, 248-252. (b) Drexhage, K. H. In Dye Lasers. Topics in Applied Physics; Scha¨fer, F. P., Ed.; Springer: Berlin, 1973; Vol. 1. (c) Yenagi, J. V.; Gorbal, M. R.; Savadatti, M. I.; Palit, D. K. Spectrosc. Lett. 1992, 25, 63-72. (d) Basov, N. G.; Logunov, O. A.; Startsev, A. V.; Stoilov, Yu. Yu.; Zuev, V. S. J. Mol. Struct. 1982, 79, 119-123. (e) Richardson, J. H.; Steinmetz, L. L.; Deutscher, S. B.; Bookless, W. A.; Schmelzinger, W. L. Z. Naturforsch. A: Phys. Sci. 1978, 33A, 1592-1593. (f) Kopylova, T. N.; Mayer, G. V.; Reznichenko, A. V.; Samsonova, L. G.; Svetlichnyi, V. A.; Dolotov, S. M.; Ponomarenko, E. P.; Tavrizova, M. A. Quantum Electron. 2003, 33, 498-502. (g) Deshpande, A. V.; Namdas, E. B. J. Lumin. 2000, 91, 25-31. (h) Kozlov, V. G.; Parthasarathy, G.; Burrows, P. E.; Forrest, S. R.; You, Y.; Thompson, M. E. Appl. Phys. Lett. 1998, 72, 144-146. (21) Acebal, P.; Blaya, S.; Carretero, L. Bol. Soc. Esp. Ceram. Vidrio 2004, 43, 467-469. (22) Fang, G.; Xu, J.; Yang, Y. J. Photopolym. Sci. Technol. 1999, 12, 339342. (23) Clancy, S. O.; Padmaperuma, A. B.; Harper, A. W. Proc. SPIE 2003, 5224, 113-120. (24) (a) Gautier-Thianche, E.; Sentein, C.; Nunzi, J. M.; Lorin, A.; Denis, C.; Raimond, P. Synth. Met. 1997, 91, 323-324. (b) Gautier-Thianche, E.; Sentein, C.; Lorin, A.; Denis, C.; Raimond, P.; Nunzi, J. M. J. Appl. Phys. 1998, 83, 4236-4241. (c) Mori, Y. In Organic Electroluminescent Materials and DeVices; Mitaya, S., Nalwa, H. S., Eds.; Gordon and Breach: Amsterdam, 1997; pp 391414.

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aqueous solution. Moreover, it can be expected that the particles formed will display properties different from those of the dissolved dyes, possibly useful for practical applications. With compound 3, we added a third member to this series to see whether a minor modification of the chemical structure, not involving the conjugated electron system, was enough to modify the selfassembling properties and, hence, the optical properties. In the following work, we proceeded with reprecipitation of the three dyes in water. The shape and size of the particles formed was studied by fluorescence microscopy, as well as by transmission and scanning electron microscopy. Information concerning their crystal or amorphous nature was looked for. Finally, the optical properties of the resulting free-standing suspensions were investigated by absorption and fluorescence spectroscopy. Experimental Section Materials. Acetone (Prolabo) and high-pressure demineralized water (resistivity 16 MΩ cm) prepared with a Milli-Q apparatus (Millipore) were used as solvents. Laser grade 3-(2′-benzimidazolyl)7-N,N-diethylaminocoumarin (Coumarin 7, 1) and 3-(2′-N-methylbenzimidazolyl)-7-N,N-diethylaminocoumarin (Coumarin 30, 2) were purchased from Acros. Dye 3 (3-(2′-N-butylbenzimidazolyl)7-N,N-diethylaminocoumarin) was prepared as follows. 2-Benzimidazolylacetonitrile (Aldrich) was reacted with 1-iodobutane (Aldrich) according to a method described in ref 25 to give in low yield 2-(N-butyl)benzimidazolylacetonitrile, which was purified by HPLC (eluent: CH3CN/H2O 70:30, column prepared with 60RP-18 silica gel). This compound was then condensed with 4-N,N(diethylamino)salicylaldehyde (Aldrich) in ethanol with piperidine as catalyst, according to the method of Christie.10a The product was purified by TLC on silica plate using CH3CN as eluent and checked for structure and purity by conventional methods. Apparatus. Spectroscopic measurements were conducted at 25 °C in a temperature-controlled cell. UV/vis absorption spectra were recorded on a Hewlett-Packard 8452A diode-array spectrophotometer. Corrected steady-state fluorescence spectra were recorded with a Photon Technology International (PTI) Quanta Master 1 spectrofluorometer. The fluorescence quantum yields of the suspensions (Φ) were determined using the classical formula: Φx ) (As × Fx × nx2 × Φs)/(Ax × Fs × ns2) where A is the absorbance at the excitation wavelength, F the area under the fluorescence curve, and n the refraction index. Subscripts s and x refer to the standard and to the sample of unknown quantum yield, respectively. Coumarin 6 in ethanol (Φ ) 0.78) was taken as the standard.26 The fluorescence quantum yields were measured by exciting the samples near their absorption maximum. Fluorescence decay was measured with the stroboscopic technique utilizing a Strobe Master fluorescence lifetime spectrophotometer from PTI. The excitation source was a flash lamp filled with a mixture of nitrogen and helium (30:70). Data were collected over 200 channels with a time-base of 0.1 ns per channel. Excitation was performed at 337 nm, and emission was at 490-494 nm, except for suspensions of 3 (510 nm). Analysis of fluorescence decay was performed using the multiexponential method software from PTI. The size and shape of the micro/nanoparticles were observed with a Zeiss Axioskop fluorescence microscope equipped with a standard camera. The objective was made up of lenses of either ×40 or ×100 magnification, the eyepiece had a lens of ×2.5 magnification. The excitation wavelength was 430-450 nm, and the emission wavelength was set at around 500-530 nm, using suitable filters. Electron microscopy was performed at the Service Commun de Microscopie Electronique de l’Universite´ Paul Sabatier. For transmission electron microscopy, a TEMSCAN 200CX microscope, and a JEOL JEM 1011 microscope equipped with a SIS Megaview III camera were used. To prepare the samples, a droplet of coumarin aqueous solution was taken and put on a carbon grid (25) Anisimova, V. A.; Askalepova, O. I.; Bagdasarov, K. N.; Chernov’yants, M. S. Khim. Geterotsiklicheskikh Soedin. 1988, 3, 345-349. (26) Reynolds, G. A.; Drexhage, K. H. Opt. Commun. 1975, 13, 222-225.

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after reprecipitation was complete. The excess liquid was drawn off with paper and the sample was revealed with ammonium molybdate (2%, pH ) 5) as a contrasting agent and allowed to dry for 12 h under vacuum at room temperature. Scanning electron microscopy was performed with a JEOL JSM6700F microscope. In this case, the dye particles were prepared by the reprecipitation method, filtered on a microfilter, and dried under vacuum before observation. The same type of sample was used for the powder X-ray diffraction analysis. The diffraction patterns were collected in transmission mode, on capillaries samples, on a θ-θ XPert Pro Panalytical diffractometer, with λ (Cu KR1, KR2) 1.54059,1.54439 Å. The extraction of peak positions for indexing was performed with the fitting program, available in the PC software package Highscore+ supplied by Panalytical. Preparation of the Suspensions. A stock solution of coumarin derivative (1 × 10-3 M for 1 and 2, and 3.9 × 10-4 M for 3) in acetone was prepared. Then, a small volume of the dye solution (40 µL for 1 and 2, and 80 µL for 3) was transferred into a cell containing 1.96 or 1.92 mL of water, respectively. The dye concentration in the mixture was 2.0 × 10-5 M for 1 and 2 and 1.6 × 10-5 for 3. The proportion of acetone in water was 2% v/v for 1 and 2 and 4% v/v for 3. Determination of the Solubility Threshold. An aliquot of the dye stock solution in acetone (1 mL, 1 × 10-3 M for 1 and 2, and 2 mL, 5 × 10-4 M for 3) was placed in a 50 mL flask, which was filled with water. The suspension was stirred at room temperature for 24-48 h and then filtered with a Bu¨chner funnel with a paper filter. The resulting clear solution was evaporated off and the deposit was dissolved again in 5 mL of acetone. The amount of dye in acetone was determined by UV/vis absorption spectroscopy, using the molar absorption coefficients previously determined, 51 200, 42 900, and 23 500 M-1 cm-1 for 1, 2, and 3, respectively.

Results Dye Reprecipitation Monitored by UV/Vis Absorption Spectroscopy. First of all, we thought that it was necessary to determine the exact solubility of the three dyes in the aqueous medium that will be used throughout this work. The solubility limit was then measured in water containing 2% acetone for 1 and 2, and it was found to be 1.8 × 10-6 and 3.5 × 10-7 M, respectively. For compound 3, it was measured in water containing 4% acetone and was about 6.0 × 10-7 M. It must be noted that these values must be considered as an upper limit of solubility because small aggregates may pass through the filter (see Experimental Section for the method used) and lead to overestimated results. The principle of the reprecipitation method used to prepare organic micro/nanoparticles is very simple. A concentrated solution of a hydrophobic compound in an organic solvent is poured into a large amount of water, in which the solvent is fully miscible. The abrupt modification of the medium induces the precipitation of the compound, which generally forms micro/ nanoparticles or micro/nanocrystals. This method was applied here to the three coumarin derivatives (see Experimental Section). Acetone was taken as the organic solvent because the coumarin derivatives were more soluble in this medium than in alcohols and higher concentrations were attained. Stock solutions of 1 and 2 were made at a concentration of 1.0 × 10-3 M. For dye 3, such a solution tended to become cloudy quite quickly. So, to get a steady solution, the concentration was reduced to 3.9 × 10-4 M. An aliquot of the organic stock solution was then mixed with water. The dye concentration was 2.0 × 10-5 M for 1 and 2 and 1.6 × 10-5 M for 3, and the proportion of acetone in water was 2% v/v for 1 and 2, and 4% v/v for 3. This corresponds to high dye supersaturation in the medium. The mixture was kept at 26 °C under constant stirring. To the naked eye, just after mixing, the coumarin solutions were yellow. Then, the solutions faded and became turbid.

Figure 2. Evolution of the UV/vis absorption spectrum of dyes1-3 (2.0 × 10-5 M for 1 and 2, and 1.6 × 10-5 M for 3) during the reprecipitation process at 26 °C: (a) Dye 1 in water containing 2% acetone, one measurement every 30 s; (b) Dye 2 in water containing 2% acetone, one measurement every 10 s; (c) Dye 3 in water containing 4% acetone, one measurement every 2 min. Inset: First stage of reprecipitation of 3, one measurement every 5 s.

The reprecipitation process was monitored by UV/vis absorption spectroscopy. Just after mixing, the spectrum of the three dyes was quite close to that of the dilute dyes in the water/ acetone mixture (vide infra). Then, as time elapsed, absorbance was strongly decreased. Such an evolution of the UV/vis absorption spectrum is characteristic of the formation of aggregates in the reprecipitation medium. Compounds 1 and 2 showed a constant decrease of absorbance with time (Figure 2a and b), with a marked increase of the baseline for 2 indicating

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Figure 3. Fluorescence microscopy image of aqueous suspensions of compounds 1 (a and b), 2 (c), and 3 (d). Dye concentration: 2 × 10-5 M for 1 and 2 and 1.6 × 10-5 M for 3.

the presence of particles in suspension. The duration of the process was evaluated precisely. For instance, the whole process was completed within 12 min for compound 1 and within less than 4 min for compound 2. The behavior of compound 3 was more complicated. The spectrum showed a very fast decrease during the first 30 s, then absorbance remained steady for 6-8 min before it was decreased again (Figure 2c). The whole process took more than 40 min. This behavior was surprising because reprecipitation was expected to be fast for compound 3, which is highly hydrophobic, even in a medium containing 4% acetone. It must be noted that the baseline was rather high at the beginning of the experiment, which reveals the presence of very small particles from the very early stage of reprecipitation. The fact that the baseline remains low during the second phase of the process can correspond to sticking of micro/nanoparticles on the vessel walls and on the stirrer. Observations by Fluorescence Microscopy. At the end of the reprecipitation process, a drop of the suspensions was placed between two slides of glass and observed under a fluorescence microscope. For compound 1, shapeless agglomerates with a green fluorescence were observed (Figure 3a). Some of them measured several tens of micrometers. When the suspension was left to stand under stirring for a couple of hours, small regular sticks appeared, emitting strong yellow light (Figure 3b). Compound 2 gave fluorescent ultrafine fibers that remained rather well distinct from each others. These fibers had homogeneous size, measuring about 20 µm long (Figure 3c). No evolution was detected with aging of the suspension. Compound 3 led to fibers that seemed to be about a hundred micrometers long and assembled as tight bundles (Figure 3d). Observations by Electron Microscopy. The samples were first observed by TEM. To do so, the dye suspensions were prepared in the same conditions as before. When reprecipitation was complete, a drop of suspension was deposited on a grid, dried, and stained by a contrasting agent. For compound 1, a thin film was formed on the grids quite homogeneously. Many aggregates were visible (Figure 4a), but they did not show clear electron diffraction spectra. Only some points of diffraction

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appeared, which can be attributed to a beginning of crystallization. When the suspension was left to stand under stirring for 1 h, little objects with well-defined shape were observed (Figure 4b). Their size was variable, some of them measuring more than 10 µm. They generated a clear diffraction spectrum and were assigned to crystallized 1. For 2, straight fibers with homogeneous size were observed (Figure 4c). They were between 5 and 10 µm long and below 0.5 µm wide. During the first investigation by TEM microscopy, the fibers appeared to be terminated by bevel-edged extremities, so that we thought that they were hollow. However, further investigation by SEM on samples prepared by filtration and drying of the suspension clearly indicated that the fibers were solid (Figure 4d and e). For 3, solid fibers were also observed (Figure 4f-h). Most of them were about 1 µm long and 25 nm wide. They were so closely entangled that they seemed to be very long. No electron diffraction spectra could be obtained for the fibers formed by compounds 2 and 3. This does not necessarily mean that we are in the presence of amorphous structures because micro/nanocrystals can easily decompose under the electron beam. Nature of the Molecular Arrangement in the Micro/ Nanoparticles. An evolution with time was noted when observing the suspensions of 1 under the electron microscope. The initial amorphous state (aggregates) led to geometrical structures that exhibit all the characteristics of microcrystals. This propensity to crystallize is in line with the fact that, according to the bibliography, large rod-shaped yellowish crystals have been obtained from an organic solvent mixture.27 Their shape is reminiscent of that of our microcrystals. This indicates that the crystals of 1 show a preferred growth axis. The crystal structure of 1 has been determined.27 In contrast, no XRD data were reported for 2 and our attempts to grow monocrystals were unsuccessful. Thus, we wondered whether the fibers formed in suspension are in amorphous or crystal form. Consequently, fibers prepared by the reprecipitation method were filtered and dried, and an X-ray powder diffraction (XRPD) analysis was performed. This sample gave a diffraction pattern very close to that of the commercial dye used as received (Figure 5). This shows that 2 is a microcrystalline powder in both cases and that the molecular arrangement was not altered by the reprecipitation process. No XRPD analysis was performed on the particles obtained from compound 3 because the amount of fibers obtained after filtration of the suspension was not enough. Spectroscopic Characteristics of the Particle Suspensions and Comparison with the Dissolved Dyes. The benzimidazolelinked coumarin dyes 1-3 are of a typical donor-acceptor chromogenic type.17,28 The diethylamino group is a strong electron donor, and the carbonyl group plays the role of the electron acceptor. The molecule is polar in the ground state, and its polar character is reinforced in the excited state. The absorption spectrum of this type of dye displays an intense band at long wavelengths, resulting from an intramolecular charge-transfer transition from a π orbital, which is significantly weighted at the amine moiety, to the π* orbital centered on the benzo ring associated to the carbonyl group. Emission involves the same orbitals than absorption, in a reverse process.10,13,15 The absorption and fluorescence spectra were recorded here for the dye suspensions in the native aqueous mixture at the end of the reprecipitation process. The spectra were also recorded in the same medium for dye concentrations lower than or close to (27) Chinnakali, K.; Sivakumar, K.; Natarajan, S. Acta Crystallogr. 1990, C46, 405-407. (28) Griffiths, J.; Millar, V.; Bahra, G. S. Dyes Pigm. 1995, 28, 327-339.

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Figure 4. Transmission electron microscopy (TEM) image of the structures formed in the aqueous suspensions of compounds 1 (a, b), 2 (c), and 3 (f-h). Scanning electron microscopy (SEM) image of the nanofibers formed by 2 (d, e).

the solubility limit. The data are gathered in Table 1. For the sake of comparison, the spectroscopic data recorded in acetone, a typical organic solvent, are reported as Supporting Information. It can be noted that for compounds 2 and 3 the absorption spectra of the suspensions were situated at the same wavelengths as those of the corresponding highly dilute dyes, although they were somewhat flattened and widened. Only the absorption

spectrum of the suspensions of 1 was slightly blue-shifted with respect to that of the dilute dye. From a general viewpoint, compound 1 absorbs at longer wavelengths than the two other dyes, whether it is dilute or in suspension. Regarding the fluorescence properties, the excitation spectra of the dilute dyes and those of the suspensions were very close to the absorption spectra, except a slight shift observed in the

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Figure 5. XRPD patterns for nanocrystals of 2. Blue line: nanofibers obtained by the reprecipitation method. Red line: commercial compound used as received. Table 1. Spectroscopic Characteristics of the Dye Suspensions after Completion of the Reprecipitation Process and Comparison with Highly Diluted Solutionsa λabs λex λem dye (nm) (nm) (nm)

Φf

τ (ns)

particle suspensionsb

1 2 3

442 424 418

451 418 420

505 496 490

0.080 ( 0.020 1.2 ( 0. 1 0.046 ( 0.020 2.8 ( 0. 1 0.094 ( 0.020 1.5 ( 0. 4

highly dilute dyec

1 2 3

456 424 422

451 425 424

504 494 491

0.27 ( 0.05 0.11 ( 0.05 0.08 ( 0.05

1.3 ( 0. 2 0.9 ( 0. 1 1.1 ( 0. 2

a λ , maximum absorption wavelength; λ , maximum excitation abs ex wavelength; λem, maximum emission wavelength; Φf, fluorescence quantum yield with excitation at the maximum absorption wavelength; τ, fluorescence lifetime; sh: shoulder. Proportion of acetone in water: 2% for 1 and 2 and 4% for 3. b Total dye concentration 2.0 × 10-5 M for 1 and 2 and 1.6 × 10-5 M for 3. c Dye concentration: 2.0 × 10-7, 3.7 × 10-7, and 5.8 × 10-7 M for Compounds 1-3, respectively. A cell of 10 cm optical pathway was used for absorption measurements of the dilute solutions.

case of 1 (Figure 6). The emitting species is therefore the one that is responsible for the absorption spectrum. Besides, the shape and position of the excitation spectra were almost independent of the emission wavelength, indicating that only one fluorescent species is present in the medium. The emission spectrum of the three dyes displayed only one intense, unresolved band. The position of this band was very close for the dilute dyes and for the particle suspensions. No significant variations of the emission spectrum with the excitation wavelength were detected. The fluorescence quantum yield of the dilute dyes was between 0.27 and 0.08. It was thus rather low if we compare it to the quantum yield of these dyes in organic solvent (see Supporting Information). This was unexpected for the following reasons. It is generally accepted that the presence of the benzimidazolyl group complicates the normal donoracceptor behavior of the coumarin heterocycle10 and that the additional electron release experienced by this type of dye upon excitation is not very intense. Consequently, the fluorescence deactivation mechanisms that involve charged excited states, such as those formed by twisted intramolecular chargetransfer and umbrellalike motions, should not take place in benzimidazolyl-7-aminocoumarins,12 contrary to other 7-amino-

Figure 6. UV/vis absorption spectrum (squares), excitation and emission spectra (plain line) of suspensions of dyes 1-3 after the reprecipitation process at 26 °C: (a) Dye 1 in water containing 2% acetone, λex ) 450 nm, λem ) 510 nm; (b) Dye 2 in water containing 2% acetone, λex ) 390 nm, λem ) 500 nm; (c) Dye 3 in water containing 4% acetone, λex ) 422 nm, λem ) 490 nm. Total dye concentration: 2.0 × 10-5 M for 1 and 2 and 1.6 × 10-5 M for 3.

Nanoparticles of Coumarin DeriVatiVes

coumarins.12,29 This reason has been invoked to explain why the fluorescence quantum yield and lifetime of dyes 1 and 2 remain high in polar and protic solvents (containing between 20% and 50% water).12,14 In this work, it appears that the fluorescence quantum yield and the lifetime of the dyes are noticeably decreased when the proportion of water becomes very high. This effect could be due to the fact that the dyes are not totally dissolved and that aggregates are formed in our water/acetone mixture, despite the caution that has been brought to the experiments. When passing from dilute solution to particles, the quantum yield was significantly decreased for dyes 1 and 2. This can be attributed to intermolecular interactions that induce nonradiative deactivation of the excited states. For compound 3, the quantum yield did not change much. The fluorescence lifetime of all samples was in the nanosecond range. It hardly varied for 1 and 3 when passing from dilute dyes to suspensions. In contrast, the lifetime of the suspension of 2 was surprisingly high, comparable to that found in organic solvent. It can be noted that the fluorescence decay was found to be monoexponential in every case, which is in line with the presence of only one type of fluorophore in the medium.

Discussion In the reprecipitation method, organic compounds are suddenly placed in an aqueous environment. Thus, the first question that arises is about the protonation state of these compounds in water. In principle, all 7-aminocoumarins can be protonated on the diethylamino group, but such modifications have not been observed for pH ranging from 4.0 to 10.0.12 However, compared with other 7-aminocoumarins, the compounds under study have an additional site of protonation with the benzimidazolyl group. The acid-base properties of 1 and 2 have been reported. In a 30:70 ethanol/water mixture, their pKa values fall in the range of 4.5-5.0.30 In a 50:50 ethanol/water mixture, their pKa values are around 4.20.31 Consequently, a significant proportion of the molecules should be protonated in water at pH ) 5.5. It has been shown that protonation on the benzimidazolyl group leads to a strong shift of the absorption and emission maxima.30,31 For instance, the absorption and emission maxima for 1 have been reported to be 454 and 498 nm, respectively, in water containing 30% ethanol, and 479 and 513 nm after addition of 1.7 M trifluoroacetic acid. For 2, the values are 425 and 489 nm, respectively, in water with 30% ethanol, and 450 and 504 nm in the presence of acid.30 These values can be compared with our data, taking into account the fact that solvatochromism can lead to a slight red-shift in our medium that contains 98% water. It appears that in our working conditions the absorption spectra are rather close to those of unprotonated dyes. Additionally, we observed that addition of NaOH traces in a solution of 1 (2.0 × 10-7 M) until pH ) 8.7 lead to a blue-shift of the absorption maximum by 6 nm, but the position of the emission and excitation spectra remained unchanged. Therefore, the presence of protonated molecules in our solutions has a weak effect upon the fluorescence properties. The problem of molecular conformation is also important because it influences directly the self-association properties and the spectroscopic properties. Since rotation is permitted between the coumarin moiety and the benzimidazole ring, different (29) Lo´pez Arbeloa, Y.; Lo´pez Arbeloa, F.; Tapia, M. J.; Lo´pez Arbeloa, I. J. Phys. Chem. 1993, 97, 4704-4707. (30) Jones, G., II.; Jimenez, J. A. C. Tetrahedron Lett. 1999, 40, 8551-8555. Jones, G., II.; Jimenez, J. A. C. J. Photochem. Photobiol. B. 2001, 65, 5-12. (31) (a) Sizova, Z. A.; Karasev, A. A.; Lukatskaya, L. L.; Rubtsov, M. I.; Doroshenko, A. O. Theor. Exp. Chem. 2002, 38, 168-172 and reference therein.

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conformations are possible. The existence of different conformers for dyes 1 and 2 has been investigated in the literature. An X-ray analysis of crystals of 1 showed that the dye is almost planar in the solid state, the benzimidazolyl group displaying an angle of 9.5° with respect to the coumarin moiety.27 This close conformation is probably favored by the strong intramolecular hydrogen bond that takes place between the carbonyl group of the coumarin ring and the NH group of the benzimidazole ring, although another planar conformation is also possible.13 In compound 2, the NH group has been replaced by a NCH3 group, and so no hydrogen bond can be formed with the carbonyl group. Calculation reveals that compound 2 has two planar conformers, as well as a gauche conformer in which the planes of the coumarin and Nmethylbenzimidazole rings display an angle of 150°. Evidence for the existence of the gauche conformer has been given in paraffin oil, a viscous medium in which fluorescence spectroscopy reveals clearly the presence of two species.13 The bibliography gives no information about the conformation of 3. Knowing the conformation of the molecules allows their spectroscopic characteristics to be better understood. Actually, the benzimidazolyl group is involved in the conjugated electron system and optimal electron delocalization over the whole molecule is attained for planar structures. This is one of the reasons that explains that the absorption and fluorescence spectra of 1 are red-shifted with respect to those of the two other dyes. Conversely, some information about conformation can be deduced from our spectroscopic study. Since the optical properties of 2 are very close to those of 3, at least in solution, it can be assumed that the conformation of the two molecules is rather similar. Let us now turn our attention toward the association properties of our compounds. They are particularly interesting. All three molecules end up forming rodlike structures if left to stir for a period of time. Among the three dyes, compound 1 is the only one for which flat microcrystals were first observed, and then elongated structures were finally formed (Figure 3b). Compound 2 quickly leads to regular, distinct straight nanofibers, while compound 3 gives small fragments that finally arrange into intertwined long fibers. It is very interesting to see that lengthening the side chain by two carbon atoms favors the formation of fiber bundles. This suggests that the butyl chain is situated at the outer side of the fiber and can promote interactions between neighboring fibers. The structures formed by 1 and 2 were shown to be microcrystals. We do not know the exact molecular arrangement in the fibers of 2 and 3. However, it can be thought that this arrangement is not very different from that encountered for 1 because the optical properties show strong similarities. In any case, the anisotropic growth of the fibers suggests a preferred direction of assembly and thus some degree of order. It must be underlined that increasing the size of the alkyl substituent brings steric hindrance and conformational flexibility that does not favor crystallization. This could explain why the whole reprecipitation process of compound 3 took so much time. We showed that reprecipitation of 3 occurs from a two-step process. A first aggregation step would take place very rapidly in the very first minutes of reprecipitation due to the hydrophobicity of the compound. Then, molecular ordering would take place more slowly, giving rise to the nanofibers that are thermodynamically more stable than a disordered aggregate. The X-ray structure of macroscopic monocrystals of 1 is known.27 The space group is P1h, with two centrosymmetrically related molecules. The planar conformation of 1 favors the stacking of the molecules in the crystal. It can be assumed that the microcrystals formed by reprecipitation of 1 are similar to the crystallized dye that was studied by X-ray analysis. The

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molecular packing of the dye molecules in the crystal can thus explain the emission of fluorescence in the solid state. In fact, the molecules are displayed in a head-to-tail manner, an arrangement that usually stabilizes dipolar dyes in the excited state. Regarding the spectroscopic properties of these micro/ nanostructures, the most striking fact was to see that the characteristics of the suspensions were rather close to those of the dilute dyes in an aqueous environment. This can be due to the surrounding presence of water, and possibly to the presence of water into the fibers. This shows that the good emission properties exhibited by the dye powder in the package flask are not a sufficient condition for the particle suspensions obtained by the reprecipitation method to be strongly emissive. To end, we must underline how intriguing is the formation of fibers from our coumarin derivatives. From a general viewpoint, several organic systems have been reported to self-organize into fluorescent solid micro- or nanofibers (the particular case of hollow cylindrical structures, namely nanotubes, will not be considered here32-34). The molecules generally contain a large aromatic moiety that is prone to stacking and a lipophilic moiety that provides supplementary van der Waals attractions between adjacent molecules within the stacks and between neighboring stacks to give nanostructures.35 Some organic gelators spontaneously give fluorescent fibers in organic solvents. Among them are rod-shaped alkoxy-tetracene and alkoxy-anthracene,36 and cholesterol-appended phenanthroline.37 This is also the case for cyanostilbene derivatives where a strong π-π stacking interaction arises from rigid rodlike aromatic segments and additional intermolecular interactions are induced by CF3 groups.38 Highly dipolar merocyanine dyes,39 p-phenylenevinylene40 and perylene diimide trimers,41 all associated to fatty chains, were also reported to give helical stacks, although the molecular arrangement is different for each of these compounds. The presence of an ionized group in the compound gives amphiphilic properties, and so water is allowed to be used as a solvent for thin film deposition.42 It must also be noted that other compounds, such as crowded aromatics and hexa-peri-hexabenzocoronene linked to fatty chains, which are known to give columnar liquid crystals, can afford micrometer-long fibers.43 If considering now small molecules without alkyl chains, it is striking to see that only a small number of them have been (32) Shimizu, T.; Masuda, M.; Minamikawa, H. Chem. ReV. 2005, 105, 14011443 and references therein. (33) Yui, H.; Guo, Y.; Koyama, K.; Sawada, T.; John, G.; Yang, B.; Masuda, M.; Shimizu, T. Langmuir 2005, 21, 721-727. (34) Balakrishnan, K.; Datar, A.; Zhang, W.; Yang, X.; Naddo, T.; Huang, J.; Zuo, J.; Yen, M.; Moore, J. S.; Zang, L. J. Am. Chem. Soc. 2006, 128, 6576-6577. (35) Hoeben, F. J. M.; Jonkheijm, P.; Meijer, E. W.; Schenning, A. P. H. J. Chem. ReV. 2005, 105, 1491-1546. (36) (a) Reichwagen, J.; Hopf, H.; Del Guerzo, A.; Belin, C.; Bouas-Laurent, H.; Desvergne, J. P. Org. Lett. 2005, 7, 971-974. (b) Shklyarevskiy, I. O.; Jonkheijm, P.; Christianen, P. C. M.; Schenning, A. P. H. J.; Del Guerzo, A.; Desvergne, J. P.; Meijer, E. W.; Maan, J. C. Langmuir 2005, 21, 2108-2112. (c) Del Guerzo, A.; Olive, A. G. L.; Reichwagen, J.; Hopf, H.; Desvergne, J. P. J. Am. Chem. Soc. 2005, 127, 17984-17985. (37) Sugiyasu, K.; Fujita, N.; Shinkai, S. J. Mater. Chem. 2005, 15, 27472754. (38) An, B. K.; Lee, D. S.; Lee, J. S.; Park, Y. S.; Song, H. S.; Park, S. Y. J. Am. Chem. Soc. 2004, 126, 10232-10233. (39) (a) Yao, S.; Beginn, U.; Gress, T.; Lysetska, M.; Wu¨rthner, F. J. Am. Chem. Soc. 2004, 126, 8336-8348. (b) Wu¨rthner, F.; Yao, S.; Beginn, U. Angew. Chem., Int. Ed. 2003, 42, 3247-3250. (40) Jeukens, C. R. L. P. N.; Jonkheijm, P.; Wijnen, F. J. P.; Gielen, J. C.; Christianen, P. C. M.; Schenning, A. P. H. J.; Meijer, E. W.; Maan, J. C. J. Am. Chem. Soc. 2005, 127, 8280-8281. (41) Yan, P.; Chowdhury, A.; Holman, M. W.; Adams, D. M. J. Phys. Chem. B 2005, 109, 724-730. (42) Everett, T. A.; Twite, A. A.; Xie, A.; Battina, S. K.; Hua, D. H.; Higgins, D. A. Chem. Mater. 2006, 18, 5937-5943. (43) (a) Bushey, M. L.; Nguyen, T. Q.; Nuckolls, C. J. Am. Chem. Soc. 2003, 125, 8264-8269. (b) Pisula, W.; Kastler, M.; Wasserfallen, D.; Pakula, T.; Mu¨llen, K. J. Am. Chem. Soc. 2004, 126, 8074-8075.

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reported to give nanofibers. Luminescent nanowires can be obtained by introducing the organic fluorophore into a nanoporous inorganic medium and then dissolving the matrix.44 Some aromatic compounds may also give needlelike crystals that can be seen like nanofibers. This is the case in particular for p-hexaphenyl studied independently by the team of Yanagi and by that of Balzer and Rubahn, but these structures were only obtained by high-vacuum sublimation and epitaxial growth on a mineral substrate.1,45 Concerning the reprecipitation method, the formation of nanowire crystals has already been reported for 1,3-diphenyl-2-pyrazoline46 and a phenylenediamine derivative,47 as well as for the polydiacetylene studied by the team of Nakanishi.48 It can be noted that particular experimental conditions, such as keeping the reprecipitation mixture at high temperature48 or adding surfactant micelles to the medium,46 can be required to obtain such structures. The spontaneous formation of fluorescent nanofibers from dyes of low molecular weight, deprived of any fatty chain, is therefore rather uncommon.

Conclusion In this work, we started with a compound (1) that tends to crystallize anisotropically, with a preferred growth axis, giving macroscopic rod-shaped crystals. The reprecipitation method allows the size of the crystals to be limited to some micrometers. We have seen that when tiny modifications were introduced in the chemical structure of 1 by increasing the length of the alkyl chain, resulting in compounds 2 and 3, the major crystallization trend was retained. Actually, the ultimate fate of all three molecules studied under reprecipitation conditions appears to be mostly solid nanofibers. However, the structural modifications brought to the basic structure lead to marked differences in the size and appearance of the fibers. It must be underlined that the spontaneous formation of nanofibers was rather unexpected considering the chemical structure of the dye molecules. Nanofibers can be used as templates for metal nanowire formation.31,49 According to their high surface area, they can promote cell attachment for tissue engineering.50 If made of fluorescent molecules, they can also be used to study the propagation of light in self-assembled systems1,38,44,45 and could find applications as new materials for sub-micrometer-sized optoelectronics.33,51 It is nice to see that these structures can be obtained in a reproducible way from simple molecules, with easy handling and safe preparation from aqueous solution. This is a distinct advantage for practical applications of nanoassemblies in the near future. Acknowledgment. We thank Mr. Lucien Datas and Mr. Laurent Weingarten at the Service Commun de Microscopie (44) Lee, J. K.; Koh, W. K.; Chae, W. S.; Kim, Y. R. Chem. Commun. 2002, 138-139. (45) (a) Balzer, F.; Bordo, V. G.; Simonsen, A. C.; Rubahn, H.-G. Phys. ReV. B 2003, 67, 115408. (b) Balzer, F.; Rubahn, H.-G. AdV. Funct. Mater. 2005, 1, 17-24. (46) Fu, H.; Xiao, D.; Yao, J.; Yang G. Angew. Chem., Int. Ed. 2003, 42, 2883-2886. (47) Li, S.; He, L.; Xiong, F.; Li, Y.; Yang, G. J. Phys. Chem. B 2004, 108, 10887-10892. (48) (a) Nakanishi, H.; Katagi, H. Supramol. Sci. 1998, 5, 289-295. (b) Oshikiri, T.; Kasai, H.; Katagi, H.; Okada, S.; Oikawa, H.; Nakanishi, H. Mol. Cryst. Liq. Cryst. 1999, 337, 25-30. (c) Onodera, T.; Oshikiri, T.; Katagi, H.; Kasai, H.; Okada, S.; Oikawa, H.; Terauchi, M.; Tanaka, M.; Nakanishi, H. J. Cryst. Growth 2001, 229, 586-590. (49) Reches, M.; Gazit, E. Science 2003, 300, 625-627. (50) Schindler, M.; Ahmed, I.; Kamal J., Nur-E-Kamal, A.; Grafe, T. H.; Young, C. H.; Meiners, S. Biomaterials 2005, 26, 5624-5631. (51) Henrichsen, H. H.; Kjelstrup-Hansen, J.; Engstrom, D.; Clausen, C. H.; Boggild, P.; Rubahn, H. G. Org. Electron. 2007, 8, 540-544.

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Electronique de l’Universite´ Paul Sabatier for their kind help in TEM and SEM measurements and Dr. Laure Vendier at the Service Diffraction X du Laboratoire de Chimie de Coordination de Toulouse for the powder X-ray diffraction patterns. We are also indebted to the referees for their valuable comments.

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Supporting Information Available: Spectroscopic characteristics of 1-3 in acetone. This material is available free of charge via the Internet at http://pubs.acs.org. LA702197H