Photoionic Supermolecules - American Chemical Society

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Photoionic Supermolecules: Mobilizing the Charge and Light Brigades A. Prasanna de Silva, Thorfinnur Gunnlaugsson, and Colin P. McCoy School of Chemistry, Queen’s University, Belfast BT9 5AG, Northern Ireland Much of the information about our environment reaches us in the form of photons via our eyes. Our eyes contain molecular systems capable of receiving these environmental signals and eventually converting them into currents of sodium and potassium ions within the optic nerves. These connect with our brains, where the messages are processed further, stored, and acted upon. The rhodopsin systems within the retinal cells in our eyes (1) are perhaps the most important examples of photoionic supermolecules. However, chemists around the world have been designing and constructing simpler examples for a variety of uses that are the subject of this article. Many of these uses stem from the fact that various ions, both metallic and nonmetallic, are responsible for maintaining our health or for destroying it. In general, a photoionic supermolecule possesses at least two molecular components with sites that interact with photons and ions, respectively (2–4). Such a supermolecule becomes particularly interesting when these two components also interact with each other in some way. It is easy to find molecular sites that interact with photons or ions when we recall that ions contain electronic charges, and that photons and electromagnetic waves are the two faces of light. The electrons within molecules provide a natural medium for enmeshing with either of these electric species. Molecules that interact with photons are most obviously available among the chromophores of dyes—the most visible products of chemistry. These absorb light of particular colors, including wavelengths that are invisible to us. Some dyes also act as fluorophores or phosphors by emitting some of the light energy that was absorbed in the first place. The molecular components that interact with ions can be found among the ligands and receptors of coordination chemistry and its younger cousin, supramolecular chemistry. By drawing upon these sources of molecular components, the designers of photoionic supermolecules can work with light in a myriad of colors and ions in a variety of charges, shapes, and sizes.

wine” color transformation. The colorless 1 changes into the red 2 when confronted with aqueous sodium hydroxide. A quick examination of structure 2 reveals an extensively delocalized π-electron system stretching all the way across the molecule, which is responsible for its bright red color. The photon interaction site therefore encompasses the entire molecule. Further inspection exposes the phenolate units whose oxygen centers act as proton receptor sites, which remain proton-free in alkaline solution. Upon joining with protons, 2 is converted to 1 with several smaller π-electron systems. In this instance the supermolecule 2 contains the proton receptor site overlapped with the proton interaction site. This simple design feature has been employed in many modern optical sensors for ions, following the success of the classical case of 1 2. Molecule 3 was used as a color sensor for Ba 2+ ions at the University of Bonn by Fritz Vögtle (6). This structure contains an azo dye unit stretching from the amino nitrogen atom to the nitro group. Of course, azo dyes are well-known chromophores in industry. For his ion receptor, Vögtle chose a nitrogen-substituted crown ether unit. Crown ethers burst upon the scene in 1967 from Charles Pedersen’s laboratories at DuPont in Wilmington, Delaware (7). A molecule with a hole, as represented by crown ethers, remains an icon of supramolecular chemistry to this day (8). In the case of sensor 3, its absorption spectrum shifts from a peak at 477 nm to 357 nm on encountering Ba2+ ions (Fig. 1). Optical sensors for ions are of growing importance because of the increasing need to monitor the concentrations of various ions in biological fluids by means of a simple test.

Ion-Triggered Color Since the time of the earliest alchemists, a change in color has been one of the most direct means of signaling molecular interactions. This is no small feat when we remember that the molecules concerned are a billionfold smaller than their observers. Some of the bestknown examples of photoionic supermolecules are based on light absorption with transmitted or reflected light serving as the output signal (5). For instance, the pH indicator or sensor phenolphthalein (1) is probably the first molecule to have made a visible impression on several generations of chemistry students in schools across the world as the central character behind the “water into

Figure 1. Absorption spectra of sensor 3 with and without Ba2+ ions. Reprinted with permission from Acc. Chem. Res. 1985, 18, 65. Copyright (1985) American Chemical Society.

Vol. 74 No. 1 January 1997 • Journal of Chemical Education

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HO

O–

OH

CO2– CO2–

–O C 2 –O C 2

N

N O

O

O O

O CO2–

NH

1

O

2

CO2–

O

O

N

N N

O

NO2

O

3 Harnessing the Brighter Light Fluorescent sensors for ions can be even more sensitive than cases based on light absorption such as 3 because of the easy detection of fluorescence against a dark background. This sensitivity means that very small quantities of the sensor are needed for a given use. One of the most interesting and useful applications of fluorescent sensor supermolecules is as spies for ions within living cells. There is little danger of poisoning the cells because the amount of the sensor needed is well below the lethal dose. From a more general viewpoint, spies are less likely to be detected and neutralized or be fed false information by the host system if the number of spies is a minimum. In addition, fluorescence emission provides the most visual way of conveying the information gathered by spies. A fluorescent microscope can create an image of ion concentration by using the light emitted by the spies distributed within the cell. Of course, the fluorescence image follows the ion movements in real time. Therefore this category of photoionic supermolecules provides a method for biologists literally to watch, as at a cinema, the ionic happenings within cells that are living under an assortment of conditions. Roger Tsien, now at the University of California–San Diego, is one of the pioneers behind this vital chemical service to the life science community and 4 is one of his sensors that enjoys considerable popularity for monitoring intracellular Ca2+ (9). The upper part of 4 contains a cleftlike receptor for Ca2+, which is lined with carboxylate groups, amine nitrogens, and ether oxygens. The fluorophore includes the indole carboxylate and the aminophenyl group connected to it. So the components for interaction with photons and Ca2+ overlap considerably. Ion binding to 4 and relatives leads to shifts in the fluorescence emission or excitation spectra, which generally correspond to the absorption spectral shifts seen with 3. Thus dual wavelength monitoring is feasible where the fluorescence intensity change at one wavelength is the key observation, while the corresponding change at a second wavelength allows internal referencing to compensate for several problems that can arise in intracellular monitoring situations. Such problems include variabilities in the amounts of sensor incorporation, optical thicknesses, and degrees of fluorescence quenching at different points across the cell.

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4 A powerful class of photoionic supermolecules useful as fluorescent sensors employs spatially separated sites to receive photons and ions, respectively (10). Their mechanisms of action draw inspiration from the lifesustaining process of photosynthesis. Absorption of sunlight within the chloroplasts of plants causes the transfer of an electron between chlorophyll and quinone units embedded separately within a protein matrix. We arrange for a similar photoinduced electron transfer (PET) between a fluorophore and an ion receptor across a deliberately placed spacer. Of course, the presence of a PET channel serves as a sink for the energy deposited in the fluorophore by light absorption. Thus the supermolecule is, at best, weakly fluorescent. However, this is true only in the absence of any ions that interact with the receptor. When the correct ions arrive at a high enough concentration, the receptor becomes ion-bound and unable to participate in a PET process. The energy of absorbed light is therefore returned as fluorescence—a vivid signal of ion entry into the receptor. An example is 5, which signals the presence of Na+ with blue fluorescence. Note the crown ether receptor for Na + and the cyano anthracene fluorophore clearly separated by a methylene spacer. These sensor systems give “off–on” switching of fluorescence rather than the wavelength shift seen within 4, for example. O O O O O

CN

5 Another type of sensor supermolecule uses two fluorophores in order to give rise to a special fluorescence signature when a normal singlet-excited state of a fluorophore sticks to an unexcited fluorophore of its own kind. Such excited dimers or excimers usually are molecular sandwiches. Their emission wavelengths are longer than those of their parent monomer because of the more delocalized π-electron system. Their lifetimes also tend to be longer. The balance between excimer and monomer can be tipped by ion binding if receptors are built into the supermolecule at suitable locations. Once again, the presence of

Journal of Chemical Education • Vol. 74 No. 1 January 1997


the ion can be signaled by changes in the fluorescence spectra. Let us consider an example 6, arising from a collaboration between Jean-Marie Lehn at Strasbourg and JeanPierre Desvergne and Henri Bouas-Laurent at Bordeaux (11). Sensor 6 traps H3+N(CH2)6N+H3 in between its anthracene units due to the complexation of each ammonium terminal by a nitrogen-substituted crown ether group. The “hot-dog” structure obtained by inserting the rodlike alkane diammonium ion between the planar anthracene prevents an excimer sandwich from developing. The rather rigid construction of 6 allows selective fluorescence signaling of the C6 alkane diammonium ion with respect to its shorter or longer colleagues.

O

O N

O

O

O

O

O

O O

O N

N

O

O

N

6 Using Delayed Emission The general area of medical diagnostics has recently begun to benefit from photoionic supermolecules with a special feature of delayed light emission. Organic compounds composed of carbon, hydrogen, and a few other light atoms usually emit light within a short period of several nanoseconds after being excited. On the other hand, when an organic ligand binds a heavy metal ion, large nuclear electric charge and, subsequently, heavy magnetic storms are introduced. Among other effects, these cause long-lived excited states to emerge in the microsecond or millisecond time regime. The lanthanide metal ions are the outstanding examples, possessing the nearly unique ability to absorb and emit light by themselves. However, their innate ability can be greatly amplified by complexation with ligands. The ligands act as d ætennas by funneling the incoming light energy from the π-system to the lanthanide ion. Another role of the ligands is to shield the lanthanide ion from water molecules, which can drain the energy from the metal excited state via O–H vibrations. The Eu3+ complex (7) is a recent example of this type arising from Juan Rodriguez-Ubis’s laboratory at University Autonoma de Madrid (12). If we employ a short light pulse for excitation, any light absorbed by the everpresent matrix molecules would be promptly returned as fluorescence. If we use the strategy of delayed observation, the matrix fluorescence would be ignored, and the delayed light emission from the metal complex would then stand out as a beacon.

N N

N

N EuIII

N N

N

CO2– – CO2– O2C

–O C 2

7 Practical observation of real-life environments can be made much simpler by using such metal complexes, and a quiet revolution has been occurring in hospital laboratories. The elegant analytical technique of radioimmunoassay has permitted the assay of drugs and other species of clinical interest. Conceptually, this technique is brilliant because it combines the extreme sensitivity of detection of a radioactive label with the power of the immune system to learn and then selectively bind a biological target. Practically, however, there have been ever-tightening controls on the handling of radioactive material. Detection of delayed light emission from antibodies tagged with lanthanide complexes avoids any radioactivity hazard but still offers good sensitivity for assays (13). Delayed emission from organic excited states can also be incorporated into photoionic systems. As in the case of metal ion complexation, substitution with heavy nonmetallic atoms such as bromine in 8 can allow the emergence of triplet excited states that can potentially emit light over a period of milliseconds. However, unlike their singlet relatives, the inherent magnetism of triplet excited states makes them prone to energy sapping interactions with other magnetic species. Molecular oxygen is a principal culprit even at very low concentrations. Self-annihilation by collision of two triplet excited states is also a common occurrence, helped no doubt by their inherently long lifetimes. Thus it comes as no surprise to learn that light emission from molecular triplet states or phosphorescence in room-temperature fluid solutions is far less common than that seen from excited singlet states. However, if the process of phosphorescence can be arranged to occur, the advantage of delayed emission can be put to good use as with the lanthanide complexes. Our pH sensor 9 is such a case (14). We need to prevent collisional interactions with the potentially phosphorescent triplet excited state of 8, while allowing exciting photons in and emission photons out. NEt2 Br NEt2

Br

β-cyclodextrin

8

9


55


up from several glucose units and related molecules as transparent containers for organic phosphors such as 1-bromonaphthalene. However, if 8 is to function as a pH sensor, it is important that the amino group remain accessible to serve as a receptor for protons in the environment. In other words, the bromonaphthalene unit must be encapsulated within the cyclodextrin, while the amine group must protrude into the water phase. Part of the solution is that since supermolecule 8 has spatially separate sites for interaction with photons and protons, respectively, the mutually exclusive requirements of the two sites can be accommodated by correctly orienting 8 within the cyclodextrin. Interestingly, 8 performs this act of regioselective self-assembly because the more hydrophobic bromonaphthalene unit hides away from water by burrowing into the cyclodextrin, while the less hydrophobic amine group points outwards. The proton-sensing action of host–guest complex 9 is based on the “switching on” of phosphorescence when the amino group is accosted by protons. A PET process suppresses phosphorescence when the amino group is proton-free, rather similar to the fluorescent sensor 5 for Na +. So 9 serves as a phosphorescent pH sensor capable of functioning in inherently fluorescent neighborhoods. Ionically Releasing Pent-up Light So far we have concentrated on the exploitation of photoionic supermolecules as optical sensors. Since sensing is a continuous process, a power supply needs to be available to sustain the sensor capability. Optical molecular sensors, whether based on absorption or emission, obtain their power from the exciting light. Excitation light sources introduce a complication of their own when extremely sensitive determination is required by the observer. Especially in nonhomogeneous solutions commonly encountered by biologists, some of the exciting radiation is scattered into the light detection system. This can interfere with the observation of fluorescence if the excitation and emission wavelengths are close together. However, there are many applications in biological and medical diagnostics where only one measurement is needed instead of continuous monitoring. In such situations it is possible to avoid the use of excitation light sources altogether and yet arrange for light emission. This results in one of the most sensitive molecular detection methods known. Electronic energy caged inside some special molecules may be thermally unleashed as light. Some special organisms can enzymatically arrange such light release. Low-level light arising from chemi/ bioluminescent events has fascinated people since antiquity and initiated a ghost story or two. If such emissions can be ionically triggered, photoionic devices with rather unique features would appear. For example, the marine protein aequorin will “switch on” its luminescence when presented with Ca2+ (15). In fact, aequorin played an important part in the initial measurements of intracellular Ca2+ levels in living systems. Smaller molecules with similar features are available. Dioxetane (10) is almost nonchemiluminescent until it is deprotonated at high pH, when the dioxetane ring collapses and releases light (16). This destabilizing effect of electron-rich phenolate groups on dioxetanes has been exploited for the design of very sensitive assays for enzymes such as alkaline phosphatase (17). A whole range of biologically important analyses can then be assayed by conjugating the enzyme to nucleic acid probes or antibodies (13, 18).

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HO O O O

O

10 Molecular Logic with Ions and Light Besides the sensing and diagnostics applications, photoionic supermolecules are also beginning to find use in the design of molecular information processors. The prospect of synthetic molecular computers has made some inroads into the popular imagination. The fact that we all have excellent examples within our heads has contributed to this appeal. Nevertheless, we can now begin to take a few steps towards this Holy Grail. The digital electronics revolution has left an indelible mark on late 20th-century life. Among the principal revolutionaries are the logic gates whose truth tables are introduced to us as teenagers. For example, a two-input AND gate will pass an output of 1 only if both input data bits are 1. If either or both input bits are 0, the output will be 0. Molecular mimicry of the behavior of these logic elements would therefore be an important milestone. What was aimed for was not a slavish reproduction of these devices at the molecular level but rather the emulation of their input/output characteristics. Unlike their larger solidstate counterparts, molecular electronic logic devices present serious difficulties even at the design stage. Since all the input and output signals are qualitatively the same (i.e., electronic), “cross talk” between the different channels can be expected to be severe across these molecular distances. This situation can be rescued by employing a set of signals and a power supply where all of them are qualitatively different and therefore distinguishable. For the different inputs, we can employ several ions because ion recognition is a well-trodden path in chemistry. The output signal can be fluorescence photons with the power supply being the incident light of shorter wavelength. The distinguishability of different ions and of light of different colors one from the other makes the photoionic approach unique for the present purpose. Furthermore, a supermolecule can self-select the correct ions into the matching receptor modules and the incident photons into the fluorophore compartment. The mechanism of fluorescence switching based on PET, outlined previously for designing sensors, can now be used to design the logic device 11 (19). System 11 incorporates a cyano anthracene fluorophore, which receives the power supply and produces the output fluorescence. The ion inputs Na+ and H+ are collected by benzo crown ether and amine receptors, respectively, and 10{3 M H+ and 10 {2 M Na + are used to provide the “high” levels of the two inputs. Either receptor can engage in PET with the fluorophore and therefore kill off the enTable 1. Truth Table for the Photoionic AND Logic Gate 11 First input (H+) level – (low) – (low) 10–3 M (high) 10–3


M (high)

Second input (Na+) level

Fluorescence Output Level

– (low)

low

10–2

M (high)

– (low) 10–2

M (high)

low low high


ergy deposited by the incoming photon, i.e., the fluorescence output is “switched off”. The binding of the appropriate cation (Na+ or H +) to its receptor will close off that particular PET pathway, but fluorescence will be “switched on” only as long as both receptors are occupied by their respective guest cations (Fig. 2). The corresponding truth table is shown in Table 1. Such an output behavior represents the action of a photoionic AND logic gate.

would soak up salts during the day, leaving fresh water to be pumped off before dusk. During the night the polymer lining would disgorge the salts and would be ready to recommence the good work at dawn. Time will tell whether this scenario will be realized.

O O

O

O

O

NC

O

O

O O

O

O O

O N

O

O

N

O

O O

N

O N

11

Light-Driven Ion Pumps The importance of rhodopsin among photoionic supermolecules was pointed out in the introduction. Bacteriorhodopsin, which is found within bacterial dwellers in salt flats, is an example of a photocontrolled ion transporter for energy production rather than for signal processing (20). Chemists have applied their ingenuity to build synthetic models of these beautiful systems. One approach exploits the light-induced geometry changes of the azobenzene molecule. The cis and trans isomers of azobenzene can be reversibly interconverted with different wavelengths of light. Also, the cis form can be thermally driven to its trans counterpart. An important connection is the control of ion binding by the geometry of the ligand or receptor: one of the first gener-

N

trans-12 cis-12 The phenolate receptor for protons has been combined with the azobenzene chromophore by Paul Haberfield at the City University of New York in some remarkably simple experiments to demonstrate the pumping of protons under the influence of light (22). Figure 3 gives a schematic representation. The hybrid structure in this case is a 2-hydroxy-azobenzene, 13, where the trans form (dominant in the dark) has a low acidity due to an internal hydrogen bond. On the other hand, illumination produces cis-13 whose increased acidity allows the proton donation along with a counter ion into the receiving phase of aqueous NaOH. The cis phenolate 13 migrates into the dark region of the toluene liquid membrane. It is accompanied and stabilized by a tetrabutylammonium ion, which is a popular component of phase transfer systems. Thermal isomerization of cis phenolate 13 to trans phenolate 13 produces the stronger base, which extracts a proton along with a counter ion from the donating phase of aqueous NaOH in the dark. The net effect is the pumping of H+ from the illuminated aqueous donating phase to the dark receiving phase.

Cl

HO

Cl

Cl

Cl

N

Cl N

N N H O

trans-13 Figure 2. Fluorescence emission spectra of the photoionic AND logic gate 11 for different ionic inputs. Reprinted with permission from Nature 1993, 364, 42. Copyright (1993) Macmillan Magazines, Limited.

alizations of supramolecular chemistry. These two ideas have been fused together by Seiji Shinkai, currently at Kyushu University, by incorporating the azobenzene moiety into a crown ether (21). The hybrid structure 12 binds Na+ in the cis form but not in the trans form. Illumination of trans-12 causes the formation of cis12 and subsequent uptake of Na+ , whereas the thermal return of cis-12 to trans-12 would squeeze out the Na +. The availability of 12 and relatives allows, at least in principle, the solar-powered desalinization of sea water. Here, coastal tanks lined with polymer-appended 12

Cl

cis-13

The azobenzene unit features heavily, though not exclusively, in the design of “caged ions”, where light serves as the key that unlocks the door of the cage and sets the ion free (23). Such photorelease of ions need not be reversible for several applications in physiology, and some designed systems take advantage of one-way photochemical reactions to destroy ion receptors. Some illuminated systems can even eject ions within the lifetime of the excited state. The reader is referred to a recent review for discussion of these enlightened molecules (24). To conclude, designed supermolecules allow transactions with light and ions that result in optical sensors, tags, switches, and pumps. In general, photoionic supermolecules can provide tools that can assist scientists across a wide range of disciplines to address problems of intense human interest.


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13

13

Figure 3. Operation of proton pump 13.

Acknowledgments What has been presented here is just an aperitif for the large amount of very interesting material served up by many laboratories. Therefore only leading references are given. We appreciate the support of Queen’s University, University of Colombo, EPSRC/SERC, DENI, Nuffield Foundation, IAESTE, and NATO (grant 921408 shared with J.-Ph. Soumillion at Université Catholique de Louvain). The help of Nimal Gunaratne and Ian Gibson is specially acknowledged. Literature Cited 1. Principles of Neural Science, 3rd ed.; Kandel, E. R.; Schwartz, J. H.; Jessell, T. M., Eds.; Elsevier: New York, 1991. 2. Lehn, J.-M. Supramolecular Chemistry; VCH: Weinheim, 1995. 3. Balzani, V.; Scandola, F. Supramolecular Photochemistry; EllisHorwood: Chichester, 1991. 4. Bissell, R. A.; de Silva, A. P; Gunaratne, H. Q. N.; Lynch, P. L. M.; Maguire, G. E. M.; Sandanayake, K. R. A. S. Chem. Soc. Rev. 1992, 21, 187. 5. Indicators; Bishop, E., Ed.; Pergamon: Oxford, 1972. 6. Löhr, H.-G.; Vögtle, F. Acc. Chem. Res. 1985, 18, 65. 7. Pedersen, C. J. J. Am. Chem. Soc. 1967, 89, 7017. 8. Chenevert, R.; Rodrique, A. J. Chem. Educ. 1984, 61, 465. 9. Tsien, R. Y. Am. J. Physiol. 1992, 263, C723. 10. Bissell, R. A.; de Silva, A. P; Gunaratne, H. Q. N.; Lynch, P. L. M.; Maguire, G. E. M.; McCoy, C. P.; Sandanayake, K. R. A. S. Top. Curr. Chem. 1993, 168, 223. 11. Fages, F.; Desvergne, J.-P.; Kampke, K.; Bouas-Laurent, H.; Lehn, J.-M; Meyer. M.; Albrecht-Gary, A.-M. J. Am. Chem. Soc. 1993, 115, 3658. 12. Remuinan, M. J.; Roman, H.; Alonso, M. T.; Rodriguez-Ubiz, J. J. Chem. Soc. Perkin Trans. 2 1993, 1099. 13. Mayer, A.; Neuenhofer, S. Angew. Chem. Int. Ed. Engl. 1994, 33, 1044. 14. Bissell, R. A.; de Silva, A. P. J. Chem. Soc. Chem. Commun. 1991, 1148. 15. Cobbold, P. A.; Lee, J. A. C. In Cellular Calcium: A Practical Approach; McCormack, J. G.; Cobbold, P. A., Eds.; IRL: Oxford, 1991; p 55. 16. Schaap, A. P.; Gagnon, S. D. J. Am. Chem. Soc. 1982, 104, 3504. 17. Schaap, A. P.; Sandison, M. D.; Handley, R. S. Tetrahedron Lett. 1987, 28, 1159. 18. Clyne, J. M.; Running, J. A.; Stempien, M.; Stephens, R. S.; Akhavan-Tafti, H.; Schaap, A. P.; Urdea, M. S. J. Biolumin. Chemilumin. 1989, 4, 357. 19. de Silva, A. P.; Gunaratne, H. Q. N.; McCoy, C. P. Nature 1993, 364, 42. 20. El-Sayed, M. A. Acc. Chem. Res. 1992, 25, 279. 21. Shinkai, S.; Manabe, O. Top. Curr. Chem. 1984, 121, 67. 22. Haberfield, P. J. Am. Chem. Soc. 1987, 109, 6177, 6178. 23. Biological Applications of Photochemical Switches; Morrison, H., Ed.; Wiley: New York, 1993. 24. Valeur, B.; Bardez, E. Chem. Br. 1995, 31, 216.

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Photoionic Supermolecules - American Chemical Society

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