pubs.acs.org/Langmuir © 2010 American Chemical Society
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1,2,3-Triazole-Containing Molecular Pockets Derived from Cholic Acid: The Influence of Structure on Host-Guest Coordination Properties Jiawei Zhang,†,‡ Matthias J. N. Junk,§ Juntao Luo,†, Dariush Hinderberger,§ and X. X. Zhu*,† †
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D epartement de Chimie, Universit e de Montr eal, C. P. 6128, Succursale Centre-ville, Montr eal, Quebec H3C 3J7 Canada, ‡Institute of Polymer Chemistry, Nankai University, Tianjin, 300071 China, and § Max Planck Institute for Polymer Research, Ackermannweg 10, 55128 Mainz, Germany. Present address: Division of Hematology & Oncology, Department of Internal Medicine, University of California, Davis, Cancer Center, CA 95817. Received May 27, 2010. Revised Manuscript Received July 3, 2010 Two cholic acid-containing trimers with 1,2,3-triazole groups close to the connecting point (“top”) and at the end of the cholic acid arms (“bottom”) were synthesized. These molecules are able to form hydrophobic pockets and solubilize pyrene and other hydrophobic molecules in polar media due to the facial amphiphilicity of cholic acid. Heavy metal ions such as Cu(II) can also be coordinated by the 1,2,3-triazole groups, as shown by EPR spectroscopy. Due to the vicinity of metal ions and pyrene in the pockets, the fluorescence of pyrene is quenched. However, the position of the 1,2,3-triazole groups has a crucial influence on the metal ion complexation. The trimer with 1,2,3-triazole groups at the top is found to bind metal ions more effectively and gives rise to a significantly enhanced fluorescence quenching efficiency. Here, the metal ions act as one tridentate chelating agent, while the triazoles on the bottom rather behave as single entities without any cooperativity in binding to the metal. In the latter case, the quenching effect is reduced considerably despite the fact that both trimers are able to bind Cu(II). This indicates that the specific and strong binding of Cu(II) at the top leads to closer spatial proximity between metal ion and pyrene when compared to the Cu(II) bound on the bottom.
Introduction Bile acids are biological compounds with interesting properties. They exhibit facial amphiphilic properties due to a hydrophilic concave side with hydroxyl groups and a carboxylic acid group and a hydrophobic convex side with three methyl groups. Due to this facial amphiphilicity and the rigidity of the steroidal polycyclic backbone, they are attractive building blocks for biomimetic materials.1 These materials based on bile acids still preserve the capabilities to self-assemble and respond to the chemical environment by exposing either their hydrophilic or hydrophobic faces. Various polymers and oligomers have been prepared from bile acids for potential biological and pharmaceutical applications.2-9 More recently, bile acids became attractive candidates in the construction of star-shaped derivatives called “molecular pockets” that are sensitive to environmental stimuli such as changes in solvent polarity. Various molecular pockets
have been prepared as drug delivery vehicles,10-15 molecular containers,16-19 nonpolymeric hydrogelators,20,21 and chemosensors for metal ions.22 We have previously reported the synthesis and properties of di-, tri-, and tetra-armed molecular pockets.23,24 These molecules can reversibly form hydrophobic cavities in polar solvents or hydrophilic cavities in nonpolar solvents due to the facial amphiphilicity of cholic acid.23,25 Cu(I)-catalyzed 1,3-dipolar cycloaddition, also known as “click chemistry”,26,27 can be performed under various conditions with high yields and constitutes a versatile and efficient method to synthesize bile acid-based molecular pockets. Moreover, the 1,2,3-triazole moieties generated through the cycloaddition are able to complex heavy metal ions.28,29 Recently, we introduced cholic acid oligomers with 1,2,3-triazole groups close to the connecting point of the arms as promising chemosensors for the detection of trace amounts of metal ions.22 Concerning the metal
*To whom correspondence should be addressed. (1) Mukhopadhyay, S.; Maitra, U. Curr. Sci. 2004, 87, 1666–1683. (2) Zhang, J. W.; Zhu, X. X. Sci. China, Ser. B Chem. 2009, 52, 849–861. (3) Zhu, X. X.; Nichifor, M. Acc. Chem. Res. 2002, 35, 539–546. (4) Zhong, Z. Q.; Zhao, Y. Org. Lett. 2007, 9, 2891–2894. (5) Gautrot, J. E.; Zhu, X. X. Angew. Chem., Int. Ed. 2006, 45, 6872–6874. (6) Gauthier, M. A.; Zhang, Z.; Zhu, X. X. ACS Appl. Mater. Interfaces 2009, 1, 824–832. (7) Chen, W.; Wei, H.; Li, S.; Feng, J.; Nie, J.; Zhang, X.; Zhuo, R. Polymer 2008, 49, 3965–3972. (8) Zhao, Y. J. Org. Chem. 2009, 74, 7470–7480. (9) Zhu, X. X.; Avoce, D.; Liu, H. Y.; Benrebouh, A. Macromol. Symp. 2004, 207, 187–191. (10) Janout, V.; Jing, B. W.; Regen, S. L. Bioconjugate Chem. 2002, 13, 351–356. (11) Janout, V.; Jing, B. W.; Regen, S. L. J. Am. Chem. Soc. 2005, 127, 15862– 15870. (12) Janout, V.; Lanier, M.; Regen, S. L. J. Am. Chem. Soc. 1996, 118, 1573– 1574. (13) Janout, V.; Regen, S. L. J. Am. Chem. Soc. 2005, 127, 22–23. (14) Janout, V.; Regen, S. L. Bioconjugate Chem. 2009, 20, 183–192. (15) Janout, V.; Zhang, L. H.; Staina, I. V.; Di Giorgio, C.; Regen, S. L. J. Am. Chem. Soc. 2001, 123, 5401–5406.
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(16) Ryu, E. H.; Yan, J.; Zhong, Z. Q.; Zhao, Y. J. Org. Chem. 2006, 71, 7205– 7213. (17) Ryu, E. H.; Zhao, Y. Org. Lett. 2004, 6, 3187–3189. (18) Ryu, E. H.; Zhao, Y. J. Org. Chem. 2006, 71, 9491–9494. (19) Zhao, Y.; Ryu, E. H. J. Org. Chem. 2005, 70, 7585–7591. (20) Maitra, U.; Mukhopadhyay, S.; Sarkar, A.; Rao, P.; Indi, S. S. Angew. Chem., Int. Ed. 2001, 40, 2281–2283. (21) Mukhopadhyay, S.; Maitra, U.; Ira; Krishnamoorthy, G.; Schmidt, J.; Talmon, Y. J. Am. Chem. Soc. 2004, 126, 15905–15914. (22) Zhang, J. W; Luo, J.; Zhu, X. X.; Junk, M. J. N.; Hinderberger, D. Langmuir 2010, 26, 2958–2962. (23) Chen, Y.; Luo, J.; Zhu, X. X. J. Phys. Chem. B 2008, 112, 3402–3409. (24) Luo, J.; Chen, Y.; Zhu, X. X. Synlett 2007, 2201–2204. (25) Luo, J.; Chen, Y.; Zhu, X. X. Langmuir 2009, 25, 10913–10917. (26) Rostovtsev, V. V.; Green, L. G.; Fokin, V. V.; Sharpless, K. B. Angew. Chem., Int. Ed. 2002, 41, 2596–2599. (27) Tornøe, C. W.; Christensen, C.; Meldal, M. J. Org. Chem. 2002, 67, 3057– 3064. (28) Chang, K. C.; Su, I. H.; Senthilvelan, A.; Chung, W. S. Org. Lett. 2007, 9, 3363–3366. (29) Huang, S.; Clark, R. J.; Zhu, L. Org. Lett. 2007, 9, 4999–5002.
Published on Web 07/20/2010
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binding properties, many questions can be posed on whether the position of 1,2,3-triazole moieties can influence the complexing effect. Taking into account the structural flexibility of the individual cholic acid arms, one could imagine that the metal ions may “close” the pocket and hence enhance the encapsulation properties if the 1,2,3-triazole moieties were moved to the end of the arms. In this article, we discuss the relationship between the molecular structure and host-guest coordination properties. Two star-shaped cholic acid derivatives with 1,2,3-triazole moieties close to the connecting point of the three arms (“top”) and at the end of the cholic acid arms (”bottom”) were prepared by click chemistry (Scheme 1). The position of the 1,2,3-triazole groups in the molecular pockets strongly affects the complexation of the metal ions by the trimers, as demonstrated by electron paramagnetic resonance (EPR) spectroscopy. The strength of this coordination manifests itself in the fluorescence quenching efficiency when pyrene is also present in the hydrophobic pocket in polar solvents. Thus, the coordination strength has direct consequences for the pockets’ capability to detect heavy metal ions, which will be the main subject of this paper.
Scheme 1. Molecular Structures of the Studied Star-Shaped Cholic Acid Derivatives
Experimental Section Pyrene (Sigma-Aldrich, 99%), ethanol (Sigma-Aldrich, HPLC grade), and glycerol (Fluka, 87%) were used without further purification. Milli-Q water was used throughout the experiments. Scheme 1 shows the chemical structures of trimers 1 and 2. Trimer 1 was prepared following a reported procedure.24 The synthetic details of trimer 2 are provided in the Supporting Information. Fluorescence Experiments. Stock solutions of trimers 1 and 2 were prepared in methanol and diluted with Milli-Q water to obtain the desired concentrations. A stock solution of pyrene (20 μM) was prepared in methanol and added to the solution of the trimer to obtain the desired concentration of pyrene. For the study of host-guest complexes, the final concentration of pyrene was 0.2 μM. For the fluorescence quenching experiments stock solutions of Cu2þ (5 mM in water) were added to obtain the desired concentrations of Cu2þ and to set the final concentration of pyrene to 0.1 μM. The order of addition of the compounds can help to determine whether the pockets may remain accessible to pyrene after the addition of Cu2þ (1 and 2 equiv.). After degassing and flushing the samples with nitrogen, steady-state fluorescence spectra were recorded at room temperature on a Varian fluorescence spectrophotometer equipped with a Xe-900 lamp with an excitation wavelength set at 335 nm. The slit widths of excitation and emission were 10 and 2.5 nm, respectively. CW EPR Spectroscopy. Continuous wave (CW) EPR spectra of the Cu-triazole complexes were recorded on a Miniscope MS200 (Magnettech, Berlin, Germany) benchtop spectrometer working at X-band (νmw ∼ 9.4 GHz) with a modulation amplitude of 0.2 mT, a sweep width of 140 mT, a sweep time of 120 s, and a microwave power of 6.2 mW. A manganese standard reference (Mn2þ in ZnS, Magnettech) was used to calibrate the magnetic field of the spectrometer. All the EPR spectra were recorded in a mixture of ethanol, water, and glycerol (9:1:2, v/v) at 78 K. The spectra were background-corrected and simulated with a custom-built program in Matlab (The Mathworks, Inc.) using the EasySpin program package for EPR.30 Pulse EPR Spectroscopy. All pulse EPR experiments were performed at X-band frequencies (9.2-9.4 GHz) with a Bruker Elexsys 580 spectrometer equipped with a Bruker Flexline split-ring resonator ER4118X_MS3. The temperature was set to 10 K by cooling with a closed cycle cryostat (ARS AF204, customized for pulse EPR by ARS, Macungie, PA). First, an echo-detected spectrum was recorded. Hyperfine sublevel correlation (HYSCORE) spectra were then measured with the pulse (30) Stoll, S.; Schweiger, A. J. Magn. Reson. 2006, 178, 42–55.
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sequence π/2-τ-π/2-t1-π-t2-π/2-τ- echo at the maximum spectral position Bmax and at Bmax - 25 mT.31 The length of all pulses was set to tP = 16 ns. The time intervals of t1 and t2 were varied from 300 to 4396 ns in steps of 16 ns. An eight-step phase cycle was used to eliminate unwanted echoes. The time traces of the HYSCORE spectra were baseline-corrected with a thirdorder polynomial, apodized with a Gauss window, and zero-filled. After two-dimensional Fourier transformation, the absolute value spectra were calculated. To account for blindspot artifacts, HYSCORE spectra were recorded with two τ values (132 ns and 164 or 200 ns). The spectra were simulated with a Matlabinterfaced program written by Madi et al.32
Results and Discussion Hydrophobic Pockets Formed by Trimers 1 and 2 in Water. It was previously reported that star-shaped cholic acid oligomers can form hydrophobic pockets (with the hydrophilic faces of the cholic acid arms pointing outward) in polar solvents or hydrophilic pockets (with the hydrophobic faces pointing outward) in nonpolar solvents due to the facial amphiphilicity (31) H€ofer, P.; Grupp, A.; Nebenf€uhr, H.; Mehring, M. Chem. Phys. Lett. 1986, 132, 279–282. (32) Madi, Z. L.; Van Doorslaer, S.; Schweiger., A. J. Magn. Reson. 2002, 154, 181–191.
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Figure 1. Ratios I3 to I1 of pyrene (0.2 μM in H2O, λex = 335 nm) as a function of the concentration of trimer 1 (9) and trimer 2 (2). The red lines are drawn as visual guides.
of cholic acid.19,25 In this study, pyrene was used as a hydrophobic fluorophore to sample the hydrophilicity of its immediate surroundings.22-25,33,34 The ratio of the peak intensities I3 to I1 of the fluorescence spectra indicates the polarity of the microenvironment.35,36 In Figure 1, the I3/I1 ratios of pyrene in water in the presence of trimers 1 and 2 are plotted against the concentration of the respective hosts (trimer 1 or 2). Their increase along with the concentration of the host molecules indicates that an increasing number of pyrene molecules enter a nonpolar environment. It can be concluded that both trimers 1 and 2 form hydrophobic cavities in aqueous media, in which pyrene is included. Upon a further increase of the host concentration, both curves attain a plateau value due to the limited solubility of the hosts, which then shelter the maximum amount of pyrene molecules. While the shapes of the two curves are similar, the host concentrations at which the I3/I1 plateau are reached and the slopes of the curves are different. Nonetheless, when using the Benesi-Hildebrand type double reciprocal equations (Supporting Information, Figures S1 and S2),37 we found that both trimers 1 and 2 complexed pyrene in a 1:1 ratio. Hence, the formation of the pocket and the encapsulation of pyrene by the two trimers are similar. Apparently, the position of the 1,2,3-triazole groups in the pockets does not strongly affect the ability to host pyrene. Since trimer 1 contains three carboxyl groups and trimer 2 has a tertiary amine, one may expect that the solution pH will have large effects on the formation and the ionic nature of the pockets. The pKa of cholic acid is about 5-6.5,38-41 and the alkalinity of acylamide is very weak. Therefore, both trimers 1 and 2 should remain in a normal state in pure water. We performed the hostguest binding experiment of trimer 1 and pyrene at pH 10 (to increase the solubility of trimer 1), and found trimer 1 could form hydrophobic pocket and form a 1:1 complex with pyrene under these conditions.22 On the basis of these results, we think the ionic state would not significantly influence the formation of the pocket and the guest-binding. (33) Dyck, A. S. M.; Kisiel, U.; Bohne, C. J. Phys. Chem. B 2003, 107, 11652– 11659. (34) Hashimoto, S.; Thomas, J. K. J. Am. Chem. Soc. 1985, 107, 4655–4662. (35) Dong, D.; Winnik, M. A. Can. J. Chem. 1984, 62, 2560–2565. (36) Kalyanasundaram, K.; Thomas, J. K. J. Am. Chem. Soc. 1977, 99, 2039– 2044. (37) Benesi, H. A.; Hildebrand, J. H. J. Am. Chem. Soc. 1949, 71, 2703–2707. (38) Roda, A.; Minutello, A.; Angellotti, M. A.; Finit, A. J. Lipid. Res. 1990, 31, 1433–1443. (39) Kurdi, P.; van Veen, H. W.; Tanaka, H.; Mierau, I.; Konings, W. N.; Tannock, G. W.; Tomita, F.; Yokota, A. J. Bacteriol. 2000, 182, 6525–6528. (40) Cabral, D. J.; Hamilton, J. A.; Small, D. M. J. Lipid. Res. 1986, 27, 334–343. (41) Kurdi, P.; Tanaka, H.; van Veen, H. W.; Asano, K.; Tomita, F.; Yokota, A. Microbiology 2003, 149, 2031–2037.
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Figure 2. Ratio I3 to I1 of the pyrene fluorescence spectra (2 μM, λex = 335 nm) in the absence (O) and presence of trimer 1 (20 μM) (9) or trimer 2 (20 μM) (2) as a function of the water content in a THF-water mixture. The red lines are drawn as visual guides.
Hydrophobic Pockets Formed by Trimers 1 and 2 in THF/ H2O. As reported earlier, a conformational change of the starshaped cholic acid derivatives can be induced by changing the polarity of the solvent. This conformational adaption can be detected by the ratio I3/I1 of the sheltered pyrene molecules.23,25 Thus, fluorescence spectra of pyrene in the absence and presence of trimer 1 or 2 were recorded as a function of the composition of the binary solvent system THF/water. As shown in Figure 2, the ratio I3/I1 decreases with increasing water content when no host molecules are added. This trend is expected since the polarity of the solvent increases steadily. In the presence of trimer 1 or 2, the ratio I3/I1 follows the same trend as in the absence of trimer up to a water content of approximately 90% (95% for trimer 1 and 85% for trimer 2) before increasing again. The latter clearly indicates that the water-rich solvent mixture triggers the formation of hydrophobic pockets of both trimers 1 and 2, in which pyrene is sheltered. That was also a clear evidence that both trimers 1 and 2 remain facially amphiphilic even after the chemical modifications. The turning point of I3/I1 in the presence of trimer 2 is at a water content of 85 vol%, which is similar to that found in our previous study of hydrophobic pockets formed by star-shaped cholic acid derivatives.23,25 In the presence of trimer 1, the I3/I1 ratio only starts to rise at a higher water contents (95 vol %), which indicates that trimer 1 requires a higher solvent polarity to form a hydrophobic cavity. Although the binding between trimer 1 and pyrene is somewhat stronger (Table S1) due to the tetrahedral structure of the central carbon of trimer 1, the formation of the pocket is more difficult for trimer 1 than for trimer 2. This could be due to the bulkiness of the triazole groups located “on top” that may restrict the rotation of the cholate arms. In trimer 2 with the 1,2,3-triazole groups at the bottom of the cholate arms, the rotation of the cholate arms should not be significantly affected by the triazole moieties. In conclusion, the amphiphilic molecular pockets made of cholic acid can form hydrophobic cavities triggered by solvophobic effects as the driving force when the overall solvent polarity reaches a threshold value. Fluorescence Quenching Studies of Trimers 1 and 2 by Cu2þ. We have previously reported that trimer 1 can be used as a potential chemosensor to reveal the presence of heavy metal ions in aqueous media at the high ppb level.22 One molecule of trimer 1 is capable of hosting one pyrene molecule due to solvophobic interactions. In addition, one metal cation can be hosted through coordination to the 1,2,3-triazole moieties. Therefore, metal ions and pyrene can be brought together in close proximity to cause fluorescence quenching of pyrene via a photoinduced electron transfer (PET) process. Note that even a metal ion concentration DOI: 10.1021/la102158a
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Figure 3. Fluorescence quenching as shown by the changes in the ratio I/I0 of pyrene (0.1 μM, λex = 335 nm) by Cu2þ in the absence (O) and presence of 25 μM trimer 1 (9), trimer 2 (2) in H2O (5 vol % methanol). The red lines are drawn as visual guides.
of only 1 μM is sufficient to induce a significant fluorescence quenching of pyrene.22 In contrast, the fluorescence quenching was largely prohibited, when a trimer without triazole groups was used as it could shield pyrene from the metal ions in the polar medium.22 To examine the role of the position of the triazole groups within the oligomers for their fluorescence quenching enhancing abilities, one can compare the fluorescence quenching curves of pyrene by Cu2þ in the presence of trimers 1 and 2. We used the ratio of the peak intensities, I/I0, to estimate the quenching effect, where I0 and I denote the intensities of the first peak in the fluorescence spectra in the absence and presence of Cu2þ, respectively (Figure 3). With 1,2,3-triazole groups at the end of the arms, trimer 2 should also be able to coordinate metal ions. However, the Cu(II)induced fluorescence quenching of the trimer 2-pyrene complex is much less efficient compared to that of the trimer 1-pyrene complex. Surprisingly, the ratio I/I0 of the trimer 2-pyrene complex is similar to that of free pyrene. While I/I0 of the trimer 1-pyrene complex reaches a constant level when the amount of Cu2þ equals the amount of host, much more than one equivalent of Cu2þ is needed to reach a constant level in case of trimer 2 (Figure 3). To unravel the origin of this fundamental difference, EPR spectroscopy was applied to study the complexation of Cu2þ by trimer 2 in detail (see below). Influence of Cu2þ on the Inclusion Properties of Trimer 2. One may speculate that coordination of a metal ion by several 1,2,3-triazole groups “on the bottom” may bring the three cholate arms so close to together that the encapsulation of pyrene is hampered. In the extreme case one may even think of a “closure” of the pocket preventing guest molecules from entering. One can examine whether pyrene still enter the pocket formed by trimer 2 if Cu2þ is added before pyrene, in other words, whether the complexation with Cu2þ can close the pocket and prevent the entrance of pyrene into the cavity. We have performed the following experiments to test for such effects. Various concentrations of trimer 2 in aqueous solution were mixed with 1 equiv of Cu2þ. The mixtures were then equilibrated overnight to allow enough time for the complexation between the 1,2,3-triazole groups and Cu2þ ions. Then pyrene was added and the fluorescence spectra were recorded. A series of comparative experiments without Cu2þ were also performed. The results are shown in Figure 4. For both series of solutions (with and without the addition of Cu2þ), the ratio I3/I1 of pyrene increases with increasing concentration of trimer 2, indicating that pyrene molecules have entered the hydrophobic cavities of trimer 2. The ratio I3/I1 is only slightly decreased when 1 equiv of Cu2þ was 13418 DOI: 10.1021/la102158a
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Figure 4. Ratio I3 to I1 of pyrene in H2O (0.2 μM, λex = 335 nm) as a function of the concentration of trimer 2 in the absence (0) and presence (9) of 1 equiv Cu2þ. The red lines are drawn as visual guides.
Figure 5. CW EPR spectra of 1:1 and 1:2 mixtures of CuCl2 and trimer 2 at 78 K. The spectra consist of three spectral contributions, “free” (green), “loosely bound 1” (blue), and “loosely bound 2” (ochre), which are shown below in the low-field region. The spectral positions of the low-field peaks are highlighted by vertical lines. The isolated contributions of both “loosely bound” species were obtained by linear combinations of the spectra; the contribution of the “free” species was obtained by a CuCl2 reference without any cholic acid derivative. All spectra were normalized to their double integral. The low field regions of all spectral contributions are magnified by a factor of 10.
added, which means that pyrene molecules can still enter the hydrophobic cavities with relative ease. This could either be indicative of the molecular pockets not yet being “closed” by 1 equiv of Cu2þ or of the gap between two cholic acid moieties being large enough for pyrene to enter the pocket. The EPR spectroscopic results presented in the next section give a better understanding (from the Cu2þ-ions’ point of view) on which of these pictures may be more accurate. Further increasing the Cu2þ concentration did not induce any changes (Supporting Information, Figure S3). We should point out that similar experiments of trimer 1 cannot be performed, since trimer 1 induces a significant fluorescence quenching of the entrapped pyrene. The fluorescence signal intensity is so much reduced that the error of the I3/I1 ratio would be too large to be meaningful. Characterization of the Cu2þ-Triazole Coordination by EPR Spectroscopy. CW EPR spectra of Cu2þ-trimer 2 Langmuir 2010, 26(16), 13415–13421
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Article Table 1. EPR Spectroscopic Parameters of the Observed Copper Species in Trimers 1-22 and 2-CuCl2 Mixtures
trimer 1/2 trimer 2 trimer 1
g
A /MHz
aiso(N)/MHz
triazoles “coupled”
free loosely bound 1 loosely bound 2 intermediate bound
2.084 2.077 2.072 2.063 2.054
2.419 2.370 2.330 2.323 2.241
403.6 453.6 498.4 514.0 623.0
∼6 ∼6 45.8 48.1
1 2 2 3
)
)
) )
mixtures are displayed in Figure 5. Obviously, the spectra do not contain any 14N superhyperfine couplings in the high field region. This is in strong contrast to trimer 1, where these couplings constitute the most prominent feature of the spectra (Figure S4).22 As explained later in the text, the coordination of Cu2þ by the triazole units of trimer 2 is too weak and too undefined to be resolved in a CW EPR spectrum. A more detailed analysis reveals that the spectra contain contributions from three different Cu2þ species, which is in analogy to trimer 1. While free, uncoordinated copper is present in all mixtures of Cu2þ and the cholic acid derivatives irrespective of the structure of the oligomer, the two additional spectral contributions deviate significantly from those observed in trimer 1. The low-field contributions of each species to the Cu(II) spectra in trimer 2 are illustrated in Figure 5. The corresponding spectral simulations are displayed in Figure S5, and the extracted EPR parameters are listed in Table 1. The two newly observed Cu(II) species in trimer 2-CuCl2 mixture, denoted “loosely bound 1” and “loosely bound 2”, originate from Cu2þ weakly coordinated by triazole units. The weak coordination manifests itself in a slight decrease of the g tensor components g^ and g and in a slight increase of the hyperfine coupling constant A as compared to free Cu2þ. The values of the “loosely bound 2” species may be regarded as similar to those of the “intermediate” species (observed in trimer 1), while the values of the “loosely bound 1” species are in between those of “intermediate” and “free” Cu2þ. For both species, the ligand field experienced by the metal ion is significantly decreased compared to the “bound” Cu2þ species in trimer 1, which is coordinated by three triazole moieties. The number of coupled triazoles can be inferred from the magnitudes of g and A . Since the parameters for species “loosely bound 2” coincide with those of the species “intermediate”, a coordination to two triazole units is likely. Species “loosely bound 1” experiences a weaker ligand field, which most likely originates from a coupling to one triazole group. This conclusion is corroborated by the fact that Cu2þ adducts with triazole-modified single cholic acid arms exhibit the same species (data not shown). Note that no species is observed to be bound to all three triazole groups available in one trimer 2 molecule. This is in strong contrast to the Cu2þ coordination in trimer 1. Apparently, the strong chelating effect that was observed for trimer 1 is absent in case of trimer 2. The absence of this chelating effect results in a substantial decrease in the strength of the triazole-metal coordination as revealed by HYSCORE spectroscopy. Hence, the term “loosely bound” was chosen. HYSCORE is a two-dimensional pulse EPR method to resolve weak electron-nuclear hyperfine couplings.31 Representative spectra for mixtures of CuCl2 and trimers 1 and 2 are shown in Figure 6. The spectra exhibit marked differences, which are explained in detail in the next paragraphs. The spectrum of the CuCl2-trimer 2 mixture contains a multitude of distinct resonances in the (-,þ) quadrant of the spectrum, while these resonances are almost completely absent for
)
g^
)
Cu2þ species
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Figure 6. HYSCORE spectra of 1:2 mixtures of (a) CuCl2-trimer
1 and (b) CuCl2-trimer 2. The spectra were recorded at the EPR spectral maximum (corresponding to g^, a) and at a 25 mT lower magnetic field position (b). Note that trimer 1 did not exhibit resonances from strongly coupled 14N atoms at any spectral position and τ value (see Supporting Information, Figure S6). The spectra were recorded at 10 K and τ was set to 132 ns.
the CuCl2-trimer 1 complex. Resonances in this quadrant originate from coupled nuclei with hyperfine couplings greater than twice the nuclear Larmor frequency. Spectral simulations suggest that these resonances are due to a 14N atom with an isotropic hyperfine coupling aiso ∼ 6 MHz (cf. Supporting Information, Figure S7). This 14N atom is assigned as the part of the triazole group, which directly coordinates to the Cu2þ ion. For trimer 1, this directly coupled nitrogen nuclear spin exhibits an isotropic hyperfine coupling constant of ∼48 MHz, which is strong enough to result in superhyperfine features in the CW EPR spectrum (Table 1). Such large couplings cannot be observed in HYSCORE spectra. Hence, in the absence of a chelating effect, the Cu-N coupling strength is decreased by a factor of ∼8. Both spectra contain features that originate from at least one weakly coupled 14N nucleus near the exact cancellation limit |A/2| = |ωI|. These remote nitrogen nuclei give rise to the two prominent matrix peaks in the (þ,þ) quadrant at 0.92 and 1.83 MHz and to a broader feature at ∼4 MHz. The observed frequencies are similar to those found for the remote 14N atom of an imidazole group. Here, marked resonances at ∼0.8, 1.6, and 4.2 MHz were observed and assigned to the nuclear quadrupole DOI: 10.1021/la102158a
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Table 2. EPR-Spectral Contribution of Different Cu2þ Species in the CuCl2-Trimer 2 Systems
Scheme 2. Depiction of Possible Binding Situation of Trimers 1 (Left) and 2 (Right) with Pyrene and a Copper Ion
relative spectral contribution ratio CuCl2:trimer 2
free
loosely bound 1
loosely bound 2
1:2 1:1
