In the beginning, beauty is enough. But later, one wonders: What's it good for?
And so it was with dendritic polymers, those highly branched, tree-like molecules that chemist Donald A. Tomalia and coworkers at Dow Chemical grew for the first time between 1979 and 1980. At first, few labs took an interest in developing synthetic routes to these aesthetically pleasing macromolecules. But in the past six years, as it became more and more apparent that dendritic polymers have something very special to offer, research efforts in this area have mushroomed.
"It's amazing," says chemistry professor Jeffrey S. Moore of the University ofIllinois, Urbana-Champaign, referring to the fact that whenever he visits other labs, he's always encountering researchers who, unbeknown to him, have jumped into the field. Many of these research efforts are so new that they haven't yet registered as blips on the dendritic radarscreen. Tomalia estimates that at least 120 groups worldwide are working with dendritic molecules.
This rapidly growing international community of researchers is exploring or developing a variety of uses for dendritic macromolecules. These include nanoscale catalysts and reaction vessels, micelle mimics, magnetic resonance imaging agents, immuno-diagnostics, agents for delivering drugs orgenes into cells, chemical sensors, information-processing materials, high-performance polymers, adhesives and coatings, separation media, and molecular antennae for absorbing light energy and funneling it to a central core (asoccurs in photosynthetic systems). Researchers also are enthusiastic about using dendritic molecules as building blocks for synthesizing even more complex supermolecules and supramolecular structures.
These applications spring from the unusual architecture and properties of dendritic macromolecules, which come in two basic structural types. The first type that was made - by Tomalia and coworkers - is the dendrimer. It has a globular structure in which well-defined branches radiate from a central core, becoming more branched and crowded as they extend out to the periphery. Some dendrimers have a diameter of more than 10 nm and a molecular weight exceeding 1 million daltons. These molecules, Tomalia points out, are "some of the most precise synthetic polymers produced by nonbiological methods."
The second type of dendritic structureis the hyperbranched polymer, development of which was spearheaded by Young H. Kim and Owen W. Webster at DuPont, and by other groups. This type of polymer also has a fractal pattern of chemical bonds, but its branches don't emanate from a central core. Hyperbranched polymers can have either random or fairly regular architectures. Tomalia, now at the Michigan Molecular Institute in Midland, Mich., and his coworkers have prepared hyperbranched polymers with comb like tructures and molecular weights greater than 10 million daltons.
The dendritic structure of these polymers is responsible for their low intrinsic viscosity, high solubility and miscibility, and high reactivity (from the presence of many chain-ends).
"The beauty of dendrimers is that their size and architecture can be specifically controlled in their synthesis, "writes chemistry professor Thomas W. Bell of the University of Nevada, Reno. Using multistep repetitive syntheses, chemists can construct the dendrimer layer by layer (generation by generation). This makes for a difficult and tedious synthesis, but it does allow exquisite tailoring of architectural features.
Dendritic polymers differ from linear polymers in that they don't have entangled chains and they do have numerous chain-ends that can be functionalized. Because of this, dendritic molecules can be constructed with discrete domains having different properties. For example, a dendrimer can be designed to have large hydrophobic cavities in its interior and a hydrophilic surface, vice versa.
The potential of dendrimers as vessels or hosts for other molecules was strikingly demonstrated in 1994 by E. W.(Bert) Meijer, chemistry professor at Eindhoven University of Technology in the Netherlands, and his coworkers Johan F. G. A. Jansen (then a postdoctoral associate at Eindhoven) and Ellen M. M.de Brabander-van den Berg, a chemist at DSM Research in Geleen, the Netherlands. They described a "dendritic box" about 5 nm in diameter that can trap smaller molecules in the box's internal cavities [Science, 266, 1226 (1994)].
Meijer stumbled on the dendritic box in his quest for a chiral dendrimer. His starting point was a globular poly (propylene imine) dendrimer with 64 branches on the periphery. DSM produces this dendrimer on a commercial scale (C&EN, Aug. 16, 1993, page 20). The synthesis involves reacting 1,4-diaminobutane with four equivalents of acrylonitrile in a Michael addition reaction. The product, which becomes the core of the dendrimer, has four branches, each tipped with a cyano group. This tetracyano intermediate is hydrogenated to a tetraamine, which is then treated with eight equivalents of acrylonitrile to give a more highly branched octacyano molecule. Thus, each generation results in a doubling of the number of terminal groups. By the fifth generation, the dendrimer has 64 functional groups at the end of as many arms.
The dendritic box came about when Meijer and coworkers attached a protected amino acid to the end of the 64 branches, giving the box a chiral shell. Their best results have been obtained using l-phenylalanine, where the amino group is protected with a tert-butyloxycarbonyl (t-BOC) group. Because the amino acid derivatives are bulky, sterically crowded, and hydrogen-bonded,the shell is "rigid" compared to the spacious and flexible interior of the box. (The rigidity also leads to zero optical activity, they found out later.)
When the last step in building the box - the addition of the dense shell - is o carried out in the presence of guest molecules, those molecules are stably encapsulated inside the box. Some of the imprisoned molecules are fairly large - for example, Rose Bengal (a polyhalogenated tetracyclic carboxylic acid dye also known as Bengal Rose). The Dutch researchers find that there is no measurable diffusion of the guest molecules out of the box, even after prolonged heating or sonication.
It is, however, possible to release the trapped guests in a controlled manner, based on the shape of the guest molecules. In a more recent report [J. Am.Chem. Soc., 117, 4417 (1995)], Meijer, Jansen, and de Brabander describe experiments in which they trapped four molecules of Rose Bengal and eight to 10 molecules of p-nitrobenzoic acid in each box. On addition of formic acid, the bulky t-BOC protecting groups arehydrolyzed away, giving the outershell a more open structure. Dialysis (membrane filtration) of the reaction mixture using aqueous acetone extracts all the p-nitrobenzoic acid molecules out of the perforated box. The punctures aren't big enough, though, to let the much larger Rose Bengal molecules through, so they remain trapped.
The Rose Bengal guests can be liberated, however, by hydrolyzing the outer shell using 12 N hydrochloric acid under reflux for two hours, followed by dialysis with water. This procedure removes the amino acid residues, yielding the original, flexible dendrimer having 64 amino-tipped branches. This dendrimer is recovered in 50 to 70% yield, the researchers note.
This two-step "shape-selective liberation" can be "tuned considerably" because changing the amino acid and its protecting group can change the volume of the cavities and the porosity of the shell, Meijer points out. His group also has observed that the number of guests that can be entrapped in the dendritic box is determined by the shape of the guest and the architecture of the box and its cavities.
Source: Meijer and coworkers, Polym. Mater. Sci. Eng., 73, 123 (1995)
This particular dendritic box isn't a suitable vehicle for drug delivery, Meijer says. But his group is trying to design boxes that could be opened enzymatically. He's also looking at boxes that could be opened photochemically.
The dendritic box has a number of other interesting and potentially useful properties. For instance, trapping a Rose Bengal molecule inside the box allows it to fluoresce strongly as it is isolated from other molecules that could quench its emission. The dye's fluorescence is relatively insensitive to solvent effects, the researchers find. Thus, such dye-in-the-box systems might be useful as fluorescent markers for pores in the nanometer range, Meijer says. And they might allow the photochemistry and photophysics of isolated molecules to be studied in a well-defined microenvironment.
There are a lot of applications one could envision for these materials, Meijer tells C&EN, but each will require hard work to realize. "We have ideas, [and so do] many of the companies approaching us. But every idea needs a lot of experimentation" and development work. "It is all very, very difficult chemistry," he stresses. The stepwise synthesis of large dendrimers is tedious, and it often involves more than 200 reactions on the same molecule. Statistically speaking, then, it should not be surprising that the product, rather than being pure, is a mixture or ensemble of similar structures. When the Dutch chemists make a dendrimer, only 20% of the product molecules are defect-free.
That's certainly not the way nature goes about building large structures. As Steven C. Zimmerman, professor of chemistry at the University of Illinois, Urbana-Champaign, notes, when nature "wants to make a viral capsid - a covering for a virus particle - it doesn'tmake a polypeptide that is hundreds of thousands of amino acids long because [at some point] it would make an error. So what it does instead is make small subunits. But it designs them in such away that they'll noncovalently self-assemble into the larger structure that it's interested in. As chemists, we're interested in mimicking that because it'spotentially a very powerful way to make big structures without having to go through the labor of synthesizing them bond by bond."
With nature as both his inspiration and challenger, Zimmerman asked himself, "Can we self-assemble dendrimers to make a larger dendrimer?" As he and his coworkers demonstrated, publishing the work earlier this year in Science [271,1095 (1996)], the answer is a resounding yes.
In simple terms, Zimmerman's group prepared a wedgelike molecule with adendritic tail and allowed six of these wedges to assemble themselves into a pielike hydrogen-bonded aggregate. This "pie" is about 9 nm in diameter and 2 nm thick, according to a molecular modeling study. At the center of the hexameric aggregate is a large cavity, which is encircled by dendritic "spinach." The hexamer's size and molecular weight (34,000 daltons) are comparable to those of small proteins, according to Zimmerman and his coworkers, FanwenZeng, David E. C. Reichert, and Sergei V. Kolotuchin.
The design of the molecular wedge was crucial. The wedge has a polycyclic backbone to which are attached two isophthalic acid units, one 7 Å above the other in a double-decker arrangement. Thus, each wedge contains four carboxylic acid groups that can hydrogen-bond to two other carboxylic acid groups on each side.
Hydrogen-bonding is the key to the hexamer's stability, Zimmerman tells C&EN, but it may not be the whole story. He suspects that van der Waals interactions between adjacent dendrimer segments also help to stabilize the aggregate, although he doesn't have any evidence for that as yet.
The stability of the hexamer is affected by other factors as well. For example, says Zimmerman, "If you dilute the solution, the aggregate will start to break apart into monomer. And if we move from a relatively nonpolar solvent like methylene chloride to tetrahydrofuran, it goes from hexamer to monomer. And I'm sure that if we raised the temperature, the aggregate would not be phenomenally stable - it would start to break up into monomer at some temperature."
The hexamer's relative fragility is due to its noncovalent nature. "One of the next steps would be to see if we can take this noncovalent aggregate and maybe start to form covalent bonds between subunits," Zimmerman says. Such an approach might allow the synthesis of large covalent structures without having to resort to iterative synthesis. "We're starting to do some rather simple model studies with single dendrimers to see if we can start to polymerize the arms." Eventually, this work might lead to a method for synthesizing ultrapure dendrimers having uniform molecular weights.
Zimmerman also envisions making even larger dendritic aggregates. The largest one he's made so far - the 9-nm-wide disk - is a fourth-generation hexamer. He hopes that higher generation dendrimers will self-assemble into a more spherical structure.
The fourth-generation aggregate has a central cavity 14 Å across - large enough to bind a couple of porphyrins, Zimmerman says. But so far he hasn't been able to bind anything in the cavity except solvent molecules. "We're not entirely sure why that is," he adds, but he hasn't given up. Lining such a cavity with appropriate functional groups could be used for molecular complexation or reaction catalysis, he notes.
The University of Nevada's Bell sees Zimmerman's self-assembled dendrimers as potential building blocks for the self-assembly of nanostructured materials. Although Zimmerman sayshis research efforts are not aimed in that direction, he does allow that materials scientists would be interested to see whether these types of aggregates might organize themselves into a liquid-crystal structure.
The dendritic structures created by Zimmerman and Meijer, because of their novelty and potential utility, have sparked a lot of excitement in the field. Their work is "stimulating," says chemistry professor Jean M. J. Fréchet of Cornell University. "Clearly, it's a new ballgame."
Fréchet: it's a new ball
gameFréchet should know. For years, he's been a key player in the field of dendritic polymers. Much of his research hasbeen geared toward developing better ways to synthesize dendritic materials, such as the self-condensing vinylpolymerization method his group announced last year (C&EN, Aug. 28,1995, page 7).
But Fréchet also is interested in the applications end - specifically, creating molecular-scale devices, such as sensors, using dendritic building blocks. A functionalized dendritic molecule that is adsorbed to a surface could be the basis of a sensor, Fréchet explains. The molecule's interactions with a chemical species in solution could be designed to trigger a change in a measurable property, such as surface conductivity or refractive index.
"We are looking at molecules that respond to changes in the environment, "Fréchet says. In April, for example, he and senior research associate Ivan Gitsov published some preliminary resultsn on macromolecules that change shape when the polarity of the solvent changes[J. Am. Chem. Soc., 118, 3785 (1996)]. These macromolecules consist of four long poly(ethylene glycol) (PEG) chains extending out from a central carbonatom. Each of these hydrophilic PEG "arms" is terminated with a hydrophobic wedgelike dendritic group based on3,5-dihydroxybenzyl alcohol. The arms are long enough and flexible enough to allow this "hybrid star" to assumeseveral very different conformations insolution.
In tetrahydrofuran, the hybrid star forms a unimolecular micelle that has a hydrophilic core of tightly packed PEG arms, surrounded by a loose hydrophobic shell of dendritic wedges. In chlorinated solvents such as chloroform, in which both building blocks are readily soluble, the PEG arms (and their dendritic extremities) extend outward, making the core more accessible. In polar or aqueous media such as methanol, the hydrophilic PEG arms loop around the hydrophobic dendritic wedges, pushing the wedges into the star's core and leaving an outer PEG layer exposed to "the outside world,"Fréchet says. The hybrid stars rearrange their two types of components in this way to minimize the overall free energy of the system.
Experiments involving this particular star copolymer were aimed at proving the concept that such a hybrid material can drastically change its micellar structure depending on solvent polarity. But these molecules"could easily be wrapped around something, and they could easily release that something," Fréchet points out, implying that this system could be applied to drug delivery.
In future reports, Fréchet promises, his group will use such "stimuli-responsive" macromolecules to do some useful things. "I have a terrific example" of an application, he tells C&EN, but he's not at liberty to discuss it yet. In any case, he and his coworkers are working to commercialize some of their more advanced materials.
One molecular device that the Cornell chemists will soon disclose is based on the assembly of several polyether dendrimers around a central rare-earth cation. Because of its dendritic "overcoat," each cation is isolated from all the others, so they cannot quencheach other when excited by light of the appropriate wavelength. The target application is signal amplification for fiber-optic communication systems (telephone), but that's about all Fréchet is willing to say for now.
A couple of years ago, Fréchet gave a seminar on his dendrimer work at TexasA&M University, College Station. One of his listeners - associate professor of chemistry Richard M. Crooks - came away impressed with the potential of these materials for chemical sensors, one of his areas of interest. Crooks, a surface and analytical chemist, wasn't working with dendritic materials at the time, but he is now. His lab is exploring the possibilities of using both hyperbranched polymers and dendrimers as chemically sensitive interfaces for sensing applications.
Crooks sees these highly branched molecules as a way to combine the best qualities of two other sensor materials - self-assembled monolayers (SAMs) and polymer thin films. SAMs can be made simply by dipping a solid substrate into the liquid that one wants to coat the surface. The resulting monolayer films make "dandy sensor materials" for a number of reasons, Crooks says, one of them being that the analyte molecules don't need to spend time diffusing through a membrane - they can attach themselves to the exposed monolayer surface right away. By contrast, polymerthin films do impede analyte diffusion, but because of their thickness, they can take up a lot of analyte, and so their sensitivity is usually very high.
"What we set out to do was to make something sort of halfway between a self-assembled monolayer and a polymer - to combine the best of both worlds," Crooks says. "And that's what these hyperbranched polymers are."
Crooks and his coworkers - David E.Bergbreiter, a chemistry professor at Texas A&M University, and postdoctoral associates Yuefen Zhou and Merlin L. Bruening - start out with a mercaptoalkanoic acid monolayer on a gold substrate. Using a series of repeated steps, the researchers chemically graft onto this monolayer a hyperbranched poly(acrylicacid) polymer. The end result is a layered, treelike polymer film in which each new layer contains more polymerbranches than the previous one, and thus is more tightly packed [J. Am. Chem.Soc., 118, 3773 (1996)]. Films thicker than 1,000 Å can be prepared using this method.
Because these hyperbranched films contain a high density of carboxylic acid groups, they can selectively bind metalions with greater sensitivity than SAMs. At the same time, they can be designed to have a fairly open structure, allowing the metal ions to permeate the film more easily than in conventional polymer films. The detection of the metal ions is fairly straight forward: For example, the mass they add to the film on binding can be measured using an extremely sensitive quartz microbalance.
The Crooks and Bergbreiter groups are working to transform the carboxylic acid groups in their hyperbranched films to different kinds of receptors, such as pyrenes and calixarenes. That will allow the sensor system to be tailored for different kinds of analytes.
The films also can be tailored by growing layers with different chemical or physical properties. Crooks and Bergbreiter have developed chemistry where the carboxylic acid sites are reacted with a perfluoroalkylamine to yield a perfluorinated film hundreds of angstroms thick. This extremely hydrophobic material can be converted into a hydrophilic material by covalently capping it with one or more layers of poly(acrylic acid). In this way, the Texas A&M chemists have been able to create a fluorinated, layered nanocomposite on gold that is fully water wettable.
This covalent layering approach could be useful for making biocompatible sensors. In such a sensor, the sensing layers could be protected by a protein-resistant coating of poly(ethylene oxide). "The outermost layer would prevent fouling of the sensor surface by proteins, while the inner layer would [remain accessible] to an ion or molecule that could penetrate the outer coating," Crookssays. He calls this a "restricted access material - it's like a molecular filter." And it's one of his "big hopes" that his lab will realize this objective. Says Crooks, "It doesn't seem like there's any reason why we can't."
The University of Illinois 'Moore says he's "pretty impressed" with Crooks's work and is optimistic about its commercial potential. "I think Dick has shown that these are the polymeric materials of choice for sensor applications."
Because such hyperbranched films can be grown in layers, they also could be useful as corrosion-inhibiting coatings, Crooks points out. Each layer could be tailored to block passage of a different corrosive agent. And it isn't even necessary to put a self-assembled monolayer on the surface to begin with, he says, because the polymer film can be grown directly on many metal surfaces. According to Crooks, a venture capital firm is seeking to license the process.
In addition to these hyperbranched polymer films, Crooks also is making a different kind of thin film for sensor applications using globular dendrimersas the building blocks. He and graduate student Mona Wells carried out the preliminary work using poly(amidoamine) (PAMAM) dendrimers, which were the ones originally developed by Tomalia and coworkers at Dow Chemical. These dendrimers are available in several generations (sizes) from Aldrich Chemical Co. and Dendritech, a Midland, Mich., firm cofounded by Tomalia.
Source: J. Am. Chem. Soc., 118, 3988 (1996).
As in the preparation of the hyperbranched films, Crooks and Wells first coat a gold substrate with a self-assembled monolayer having carboxylic acid groups on the top surface. Then they attach PAMAM dendrimers, the branches of which are tipped with amino groups, to the monolayer by forming amide bonds. According to Crooks, these are the first dendrimer monolayer films that have been covalently attached to a surface.
The best results have been obtained with the fourth-generation PAMAMo dendrimer, which Crooks characterizes as "a soft spheroid" with an accessible interior. In solution, it is globular. But when stuck to a surface, it appears to be slightly squashed, like a drop of water on a windowpane. "If the dendrimers weren't somewhat flattened, then we would see unreacted carboxylic acid groups on the surface" between the dendrimers, Crooks points out. Spectroscopically, they don't see such unreacted groups, suggesting that "almost the entire surface is covered with dendrimer." Experiments to check that are in the works, he says.
The most important finding about these dendrimer films, Crooks says, is that "you can use them as sensors." He and Wells exposed the films to volatile organic compounds and measured the amount of absorption using a highly sensitive device called a surface acoustic wave mass balance. Hydrocarbons like benzene absorb very little. Alcohols, which can hydrogen-bond to the receptors (amines on the outside of the dendrimer, amides on the inside), absorb more. And propionic acid, which can transfer a proton to the amine group,"absorbs like crazy," Crooks says.
In their first report on this work [J. Am.Chem. Soc., 118, 3988 (1996)], the TexasA&M chemists point out that this system has some of the essential attributes of an ideal chemical sensor: The response to test chemicals is very rapid and typically is completely reversible, and the device has an excellent signal-to-noise ratio.
Furthermore, the free amine groups on the outside of the dendrimer can be readily converted into other chemically sensitive groups for a variety of sensor applications. Crooks also is planning to use other types of dendrimers as building blocks for chemically sensitive films.
"The real power of dendrimers as chemically sensitive interfaces lies in their use as elements of sensor arrays,"Crooks points out. Such arrays offer enhanced selectivity because they group dendrimers that are responsive to different classes of analytes. Crooks is pursuing this strategy in collaboration with Antonio J. Ricco of the microsensor R&D group at Sandia National Laboratories in Albuquerque, N.M.
"This paper is just the starting point," Crooks says. "By tinkering with the outside - and inside - of the dendrimer, we hope to make the films very highly selective" - though their selectivity isn't that bad now.
Crooks's work with dendritic materials is a good example of applications-driven research. Other investigators are more interested in finding answers to fundamental questions about these materials. There's plenty of room for both approaches. As Eindhoven's Meijer says about his own work: "We're really trying to push the frontiers of science. And from that, new applications will emerge."
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