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Vesicle and Stable Monolayer Formation from Simple “Click” Chemistry Adducts in Water Santanu Bhattacharya*,†,‡,§ and Joydeep Biswas† †
Department of Organic Chemistry, Indian Institute of Science, Bangalore 560 012, India, and ‡Chemical Biology Unit of JNCASR, Bangalore 560 064, India § J.C. Bose Fellow, DST, New Delhi, India Received May 28, 2010
Click chemistry has been successfully extended into the field of molecular design of novel amphiphatic adducts. After their syntheses and characterizations, we have studied their aggregation properties in aqueous medium. Each of these adducts forms stable suspensions in water. These suspensions have been characterized by dynamic light scattering (DLS) studies and transmission electron microscopy (TEM). The presence of inner aqueous compartments in such aggregates has been demonstrated using dye (methylene blue) entrapment studies. These aggregates have been further characterized using X-ray diffraction (XRD), which indicates the existence of bilayer structures in them. Therefore, the resulting aggregates could be described as vesicles. The temperature-induced order-to-disorder transitions of the vesicular aggregates and the accompanying changes in their packing and hydration have been examined using high-sensitivity differential scanning calorimetry, fluorescence anisotropy, and generalized polarization measurements using appropriate membrane-soluble probe, 1,6-diphenylhexatriene, and Paldan, respectively. The findings of these studies are consistent with each other in terms of the apparent phase transition temperatures. Langmuir monolayer studies confirmed that these click adducts also form stable monolayers on buffered aqueous subphase at the air-water interface.
1. Introduction Copper(I)-catalyzed cycloaddition between an alkyne and an azide is popularly known as “click chemistry”.1-3 Click chemistry has been introduced by Sharpless, and it describes a step that is tailored to generate adducts quickly and reliably by joining suitable molecular units together. This is inspired by the fact that nature also generates larger molecules by joining small molecular units. Click chemistry has already found widespread use in diverse fields such as bioconjugation of oligonucleotides,4 glycoclusters syntheses,5 design of enzyme inhibitors,6 and even in polymer and materials science.7 Further this reaction has been adapted to join lipids with proteins8 and used to generate synthetic analogues of a type of lipids that occur in archaebacteria.9 However, so far there has been no attempt to exploit this useful chemistry for the design of amphiphilic systems that would aggregate in aqueous media to produce nanoscopic ensembles. Here we present four novel adducts (Figure 1) synthesized using click chemistry: an appropriately chosen monomer (M1) along with a dimer (D1) and a tetramer (T1). A long-chained variant, M2 of the monomer, M1, was also prepared for comparison. The syntheses of M1, M2, D1, and T1 were accomplished by click *Corresponding author: e-mail
[email protected]; Ph (91)-80-22932664; Fax (91)-80-2360-0529. (1) Kolb, H. C.; Finn, M. G.; Sharpless, K. B. Angew. Chem., Int. Ed. 2001, 40, 2004. (2) Tornoe, C. W.; Christensen, C.; Meldal, M. J. Org. Chem. 2002, 67, 3057. (3) Hassane, F. S.; Frisch, B.; Schuber, F. Bioconjugate Chem. 2006, 17, 849. (4) Morvan, F.; Meyer, A.; Pourceau, G.; Vidal, S.; Chevolot, Y.; Souteyrand, E.; Vasseur, J.-J. Nucleic Acids Symp. Ser. 2008, 52, 47. (5) Chevolot, Y.; Bouillon, C.; Vidal, S.; Morvan, F.; Meyer, A.; Cloarec, J.-P.; Jochum, A.; Praly, J.-P.; Vasseur, J.-J.; Souteyrand, E. Angew. Chem., Int. Ed. 2007, 46, 2398. (6) Srinivasan, R.; Li, J.; Ng, S. L.; Kalesh, K. A.; Yao, S. Q. Nature Protoc. 2007, 2, 2655. (7) Binder, W. H.; Sachsenhofer, R. Macromol. Rapid Commun. 2007, 28, 15. (8) Musiol, H.-J.; Dong, S.; Kaiser, M.; Bausinger, R.; Zumbusch, A.; Bertsch, U.; Moroder, L. ChemBioChem 2005, 6, 62. (9) O’Neil, E. J.; DiVittorio, K. M.; Smith, B. D. Org. Lett. 2007, 9, 199.
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coupling between each of the corresponding propargylated ester precursors and appropriate either mono-, or di-, or tetra-azides, respectively (Schemes 1 and 2). Since these compounds were synthesized by click chemistry, they are exclusively made up of 1,4substituted 1,2,3-triazoles. In the present set of adducts, appropriate alkyl groups replace the tautomeric H-atom of the triazole backbone. It occurred to us that since the triazoles are significantly polar,10 the same part of the adduct molecules would get hydrated preferentially than their hydrophobic segments if brought into contact with water. On the basis of the published reports11 on the basicity constant for N-methyl-1,2,3-triazole, each adduct described herein should remain mainly protonated in water at pH e 3.0, as shown in Chart 1. Upon brief sonication each of these adducts formed stable suspensions in aqueous medium, which were characterized by dynamic light scattering (DLS) studies and transmission electron microscopy (TEM) to determine the average sizes of the aggregates. Such aggregates were found to possess inner aqueous compartments as evidenced from dye entrapment studies, indicating that these adducts on dispersal in aqueous media afforded vesicles. Cast films formed from each of these aggregates were also characterized by X-ray diffraction (XRD) studies, which indicate the formation of a tilted bilayer type arrangements from the aggregates of M1 whereas regular bilayer structures were predominant from the aggregates derived from each of M2, D1, and T1. The temperature-induced order-to-disorder transitions of the vesicles and the accompanying changes in packing and hydration were examined using high-sensitivity differential scanning calorimetry (DSC), fluorescence anisotropy, and generalized polarization measurements using a membrane-soluble probe, DPH, and a polarity (10) Nulwala, H.; Takizawa, K.; Odukale, A.; Khan, A.; Thibault, R. J.; Taft, B. R.; Lipshutz, B. H.; Hawker, C. J. Macromolecules 2009, 42, 6068. (11) (a) Elguero, J.; Katritzky, A. R.; Denisko, O. V. Adv. Heterocycl. Chem. 2000, 76, 1. (b) Abboud, J.-L. M.; F-Foces, C.; Notario, R.; Trifonov, R. E.; Volovodenko, A. P.; Ostrovskii, V. A.; Alkorta, I.; Elguero, J. Eur. J. Org. Chem. 2001, 3013.
Published on Web 02/03/2011
DOI: 10.1021/la104851g
1581
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Bhattacharya and Biswas
Figure 1. Molecular structures of the triazole-based adducts that have been synthesized and used in the present investigation. Scheme 1 a
a Reagents and conditions: (i) DCC, DMAP, THF, rt, 6 h; (ii), (iii), (iv), and (v) NaN3, DMF, 80 °C, 10 h.
sensor, Paldan, respectively. Interestingly, each of these adducts also formed a stable film at the air-water interface as characterized by the Langmuir isotherm studies. Click chemistry thus allows a simple preparation of novel amphiphiles with assorted variation in their head/tail groups and adds to the growing list of new amphiphiles.12a-f
2. Experimental Section 2.1. Materials and Methods. All reagents, solvents, and chemicals used in these studies were obtained from the best known commercial sources. The solvents were dried prior to use. Column chromatography was performed using a 60-120 mesh silica gel. NMR spectra were recorded using a Jeol JNM λ-300 (300 MHz (12) (a) Bhattacharya, S.; Biswas, J. Langmuir 2010, 26, 4642. (b) Menger, F. M.; Shi, L.; Rizvi, S. A. A. J. Colloid Interface Sci. 2010, 344, 241. (c) Ahmad, R. K.; Faure, D.; Goddard, P.; Oda, R.; Bassani, D. M. Org. Biomol. Chem. 2009, 7, 3173. (d) Aime, C.; Manet, S.; Satoh, S.; Ihara, H.; Park, K.-Y.; Godde, F.; Oda, R. Langmuir 2007, 23, 12875. (e) Jiao, T.; Liu, M. Langmuir 2006, 22, 5005. (f) Grinberg, S.; Linder, C.; Kolot, V.; Waner, T.; Wiesman, Z.; Shaubi, E.; Heldman, E. Langmuir 2005, 21, 7638. (g) Haldar, J.; Aswal, V. K.; Goyal, P. S.; Bhattacharya, S. Angew. Chem., Int. Ed. 2001, 40, 1228. (h) Bhattacharya, S.; De, S.; Subramanian, M. J. Org. Chem. 1998, 63, 7640. (i) Bhattacharya, S.; Kumar, V. P. Langmuir 2005, 21, 71. (j) Bhattacharya, S.; Snehalatha, K.; Kumar, V. P. J. Org. Chem. 2003, 68, 2741. (k) Bhattacharya, S.; Haldar, J. Langmuir 2004, 20, 7940. (l) Haldar, J.; Kondaiah, P.; Bhattacharya, S. J. Med. Chem. 2005, 48, 3823.
1582 DOI: 10.1021/la104851g
for 1H) spectrometer. The chemical shifts (δ) are reported in ppm downfield from the internal standard, TMS, for 1H NMR. Mass spectra were recorded on a MicroMass ESI-TOF spectrometer. Infrared (IR) spectra were recorded using neat samples on a Jasco FT-IR 410 spectrometer. Caution! Reactions of azide salts with organic compounds and transformations involving organic azides are potentially explosive. Only small amounts of materials should be used and cautiously handled. Although we did not experience any problem, proper caution must be exercised while handling with all new azide compounds. 2.2. Synthesis. Prop-2-ynyl Dodecanoate (1). Propargyl alcohol (616 mg, 11.00 mmol) was added to a stirred solution of lauric acid (2.0 g, 10.00 mmol), DMAP (1.59 g, 13.00 mmol), and DCC (2.68 g, 13.00 mmol) in dry THF (12 mL), and the resultant mixture was stirred at room temperature for 6 h. The reaction mixture was then diluted with acetone and filtered. The filtrate was concentrated, diluted with ethyl acetate (50 mL), and washed with saturated aqueous KHSO4 solution (20 mL 3) followed by water (20 mL) and finally using saturated brine (20 mL). The organic phase was separated, dried over anhydrous Na2SO4, and evaporated, leaving a residue. The product residue, prop-2-ynyl dodecanate (1), was found to be of sufficient purity for direct use in the next step without any need for further purification. Yield: light yellow oil, 2.0 g (8.4 mmol, 84.0%). IR (neat) (cm-1): 3313, 2927, 2856, 2131, 1747, and 1158. 1H NMR (300 MHz, CDCl3): δ 0.86 (3H, m), 1.34-1.18 (16H, m), 1.62 (2H, m), 2.36 (2H, t), 2.44 (1H, t), 4.65 (2H, d). ESI-MS calcd. for [C15H26O2 þ Naþ]: 261.3555; found: 261.183. Azidoethane (2). NaN3 (598 mg, 9.20 mmol) was added to a solution of bromoethane (200 mg, 1.84 mmol) in DMF (5 mL) kept in a screw-top pressure tube, and the resulting mixture was heated to 80 °C for 10 h. Then the solvent was evaporated under vacuum to leave a residue, to which EtOAc was added. This afforded an organic layer, which was washed with water (20 mL 2) and followed by saturated brine (20 mL). Because of the potential explosiveness of azides,13 the organic layer was dried over anhydrous Na2SO4 and concentrated gently below 40 °C under vacuum. The crude residue thus obtained was purified by column chromatography using EtOAc/hexane (2:98), which afforded azidoethane (2) as a brown oil, 120 mg (1.69 mmol, 92.0%). IR (neat) (cm-1): 2930, 2867, 1598, 1495, 1467, 1366, 1188, and 1174. 1H NMR (CDCl3, 300 MHz): δ 1.74 (3H, t), 3.39 (2H, q). ESI-MS calcd for [C2H5N3 þ Naþ]: 94.0711; found: 94.0381. (13) Klapotke, T. M.; Rienacker, C. M. Propellants, Explos., Pyrotech. 2001, 26, 43.
Langmuir 2011, 27(5), 1581–1591
Bhattacharya and Biswas
Article Scheme 2 a
a
Reagents and conditions: (i), (ii), (iii), and (iv) CuI, DIEA, MeOH, rt, 6-8 h.
Chart 1
1-Azidotetradecane (3). NaN3 (234 mg, 3.60 mmol) was added to a solution of 1-bromotetradecane (200 mg, 0.72 mmol) in DMF (20 mL) in a round-bottom flask, and the resulting mixture was warmed to 80 °C for 10 h. Then the solvent was evaporated under vacuum to leave a residue, to which EtOAc was added. This afforded an organic layer, which was washed with water (20 mL 2) and followed by saturated brine (20 mL). Because of the potential explosiveness of azides,13 the organic layer was dried over anhydrous Na2SO4 and concentrated gently below 40 °C under vacuum. The crude product thus obtained was purified by column chromatography using EtOAc/hexane (2:98) which afforded 1-azidotetradecane (3) as a brown oil, 164 mg (0.69 mmol, 95.0%). IR (neat) (cm-1): 2930, 2867, and 2104. 1H NMR (CDCl3, 300 MHz): δ 0.78-0.84 (3H, t), 1.18-1.32 (22H, m), 1.641.82 (2H, m), 3.30-3.36 (2H, t). ESI-MS calcd for [C14H29N3 þ Naþ]: 262.3901; found: 262.2259. 1,3-Diazidopropane (4). NaN3 (322 mg, 4.95 mmol) was added to a solution of 1,3-dibromopropane (200 mg, 0.99 mmol) in DMF (5 mL) in a round-bottom flask, and the resulting mixture was warmed gently to 80 °C. After 10 h, the solvent was evaporated under vacuum. The reaction mixture was diluted with EtOAc. The organic layer was washed with water (20 mL 2) and followed by saturated brine (20 mL). Because of the potential explosiveness of azides,13 the organic layer was dried over anhydrous Na2SO4 and concentrated under vacuum gently
