Anion Recognition in Water: Recent Advances from a Supramolecular and Macromolecular Perspective

Abstract The recognition of anions in water remains a key challenge in modern supramolecular chemistry, and is essential if proposed applications in biological, medical, and environmental arenas that typically require aqueous conditions are to be achieved. However, synthetic anion receptors that operate in water have, in general, been the exception rather than the norm to date. Nevertheless, a significant step change towards routinely conducting anion recognition in water has been achieved in the past few years, and this Review highlights these approaches, with particular focus on controlling and using the hydrophobic effect, as well as more exotic interactions such as C−H hydrogen bonding and halogen bonding. We also look beyond the field of small‐molecule recognition into the macromolecular domain, covering recent advances in anion recognition based on biomolecules, polymers, and nanoparticles.


Introduction
Thec hemical recognition of anions has progressed in leaps and bounds in the last twenty years or so,during which time ahuge array of selective receptors and sensors have been developed. However,t he proposed applications are overwhelmingly in the biological, medical, environmental, and industrial arenas and, therefore,t ypically require aqueous conditions, [1,2] while the majority of research to date has taken place in organic media (in contrast to the earliest hosts [3] ). The design of selective anion receptors that function in aqueous solution is,t herefore,akey challenge in supramolecular chemistry:i np articular, examples of receptors that are neutral or of low charge and operate in organic-aqueous mixtures are uncommon, and those that function in 100 % water are rarer still. Water itself is ahighly competitive polar solvent, able to hydrate the host and guest through hydrogenbond donation and acceptance,w hilst anions themselves possess intrinsic properties that make them difficult to bind effectively in aqueous solution. Anions are heavily solvated in water, typically more so than analogous cations of the same charge and similar size, [4] they also display am uch greater range of hydrophilicity/hydrophobicity, [5] and their recognition is further complicated by the multiple protonation equilibria that many exhibit. Furthermore,a nions are in general relatively large,t hereby leading to weaker electrostatic interactions with ap ositively charged host. From as ynthetic chemistry perspective,t he design and synthesis of water-soluble receptors is also demanding,a nd requires careful consideration of potential competing self-assembly processes arising from amphiphilicity.M any such artificial anion receptors suffer from poor aqueous solubility,t hus necessitating the use of organic-aqueous solvent mixtures for anion-binding investigations.N evertheless,o vercoming this range of challenges is essential if applications of synthetic anion receptors in competitive as well as biologically and environmentally relevant aqueous media are to be exploited.
Anion recognition in water has been central to one of the most interesting science stories to hit the mainstream news in recent years.In2011, it was reported that astrain of bacteria taken from Mono Lake in California (USA) could grow in media rich in arsenate but free of phosphate. [6] Such ar esult implied that arsenic could replace phosphorus in the bacterial genome,a nd would be the first instance of life comprised of alternative elements-thus meaning that the search for life outside Earth should be significantly broadened in terms of possible environments. However,the report was immediately questioned by many in the scientific community on anumber of different fronts,and it was discovered that the bacteria possessed such highly specific phosphate-binding proteins that they could survive on the merest of trace levels of phosphate. [7] Themicroorganisms have shown that such precise discrimination is possible in water-now the scientific community is aiming to achieve this level of selectivity with tunable synthetic recognition systems.
Stefan Kubik, ap ioneer in the use of neutral host molecules for anion binding in aqueous solutions,h as reviewed the state of the art in the field of anion recognition in water using supramolecular host systems,t he most recent of which was published in 2010. [8,9] However,i nt he past five years numerous exciting and significant advances in this rapidly expanding field have emerged, in particular exotic interactions such as CÀHh ydrogen bonding and halogen bonding have been exploited in anion host systems capable of operating in water. Theh ydrophobic effect in particular has attracted much interest in the context of anion recognition in water by electroneutral hosts and, yet, of all of the factors driving host-guest complexation, it is arguably the least well The recognition of anions in water remains akey challenge in modern supramolecular chemistry,and is essential if proposed applications in biological, medical, and environmental arenas that typically require aqueous conditions are to be achieved. However,s ynthetic anion receptors that operate in water have,i ng eneral, been the exception rather than the norm to date.Nevertheless,asignificant step change towards routinely conducting anion recognition in water has been achieved in the past few years,and this Review highlights these approaches,with particular focus on controlling and using the hydrophobic effect, as well as more exotic interactions such as C À H hydrogen bonding and halogen bonding.W ealso look beyond the field of small-molecule recognition into the macromolecular domain, covering recent advances in anion recognition based on biomolecules, polymers,a nd nanoparticles.
understood. Recent studies have highlighted and investigated the nuanced relationship between the hydrophobic effect, the Hofmeister series,i on-solute interactions,a nd the thermodynamic contributions that dominate these phenomena, which has been aided by the recent design and synthesis of hydrophobic host systems capable of strongly binding anions in water.
This Review commences with the recent advances in the field of supramolecular anion coordination chemistry in water, from 2010 to the present day.W e have chosen to only include examples of synthetic receptors which operate in solvent mixtures containing 50 %ormore water, with particular emphasis laid on those capable of functioning in 100 %a queous solutions.W ewill also look beyond the field of smallmolecule recognition into the macromolecular domain, covering recent advances in anion recognition based on biomolecules,p olymers,a nd nanoparticles in ! 50 %w ater. Since this wider perspective has not yet been reviewed, the time constraint is somewhat more relaxed.

Charged Hydrogen-Bonding Anion Receptors
Anion recognition in water by small-molecular supramolecular receptors has been predominantly the domain of positively charged hosts.E xamples include the protonated macrobicyclic anion receptors,d escribed by Park and Simmons in seminal studies, [3] theq uaternary ammonium cages from Schmidtchen, [10] polyammonium macrocycles,c ryptands, [11] and macrocyclic guanidinium-based receptors from Lehn and co-workers, [12] as well as many more recent examples.
Arguably the most frequently employed motif to date is the ammonium group,u sed to bind anions through electrostatic and hydrogen-bonding interactions in aqueous solution at low pH values (to ensure protonation of the amine groups), [13] and an umber of notable examples have very recently been reported. Forexample,Bencini and co-workers described as eries of 1,4,7-triazacyclononane ( [9]aneN 3 ) macrocycles linked by rigid aromatic motifs such as com-pound 1 (Figure 1) that bind phosphate anions in acidic media with size selectivity arising from the distance between the polyammonium macrocyclic units. [14] Similarly,t he same research group reported as tructurally related Binol-based receptor 2 that is capable of the impressive enantiomeric recognition of the tartrate anion in aqueous buffer. [15] The [9]aneN 3 -derived host exhibited high selectivity in the binding and fluorescence sensing of (S,S)-tartrate (log K = 6.1 in 0.1m tris-buffer at pH 7) over (R,R)/meso-tartrate,w hich bound approximately three orders of magnitude more weakly. Furthermore,s electivity of up to three orders of magnitude was observed over (S)/(R)-malate,s uccinate,m aleate,f umarate,and (S)/(R)-lactate.
Theg uanidinium motif remains protonated over am uch wider pH range than ammonium groups and, importantly, remains protonated at physiological pH. Furthermore,i ts ability to form two strong,p arallel hydrogen bonds is particularly advantageous for the binding of oxoanions such as carboxylates or phosphates.F or these reasons,such motifs continue to find use in receptors designed for oxoanion recognition in pH-neutral solutions.K uchelmeister and Schmuck recently reported ag uanidinium-based tweezerslike receptor (3), which binds nucleotide anions in buffered water solution at ap hysiological pH value. [16] Notably,t he receptor binds monoanionic adenosine monophosphate (AMP; K % 10 4 m À1 )w ith an unprecedented selectivity over trianionic adenosine triphosphate (ATP), which associates an order of magnitude more weakly. [16] Similarly,K ataev et al.

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Reviews have recently reported ab isguanidinium receptor capable of the selective binding of orthophosphate in 1:1m ethanol/ aqueous buffer solution. [17] Although less frequently utilized for anion recognition than the ubiquitous NH hydrogen-bond donor,C ÀHh ydrogen bonds can be usefully employed in receptor design, and such motifs have lately been incorporated within anion receptors capable of operating in aqueous solvents.F or example,Y ou and co-workers have described at etrakis(imidazolium) macrocycle 4 ( Figure 2) for the recognition of sulfate in water. [18] As trong 2:1s toichiometric complex is formed between 4 and sulfate in which the anion is bound by eight CHÀOhydrogen bonds,with an association constant of 8.6 10 9 m À2 .N otably,s ulfate binding is concomitant with an increase in the fluorescence response of the macrocycle, thereby enabling the receptor to act as an optical sulfate sensor in aqueous solution. Kim and co-workers have also used the tetrakis(imidazolium) cyclophane 5 for the selective detection of guanosine-5'-triphosphate and iodide. [19] Binding of these species by charge-assisted C À Hh ydrogen-bonding interactions in aqueous solution at pH 7.4 results in quenching of the fluorescence from the anthracene motif.F urthermore,H ay,S essler, and co-workers have investigated the anion recognition properties of ap yrrole-based triazoliumphane 6,which contains NH and cationic CH hydrogen-bond donor groups. [20] Them acrocycle recognizes tetrahedral oxoanions with high affinity and selectivity over ar ange of monoanions and trigonal planar anions in 2:3a cetone/water solution at pH 7.2.

Metal Complexes
Arguably the strongest interaction that can be used for anion binding in water is direct coordination to ametal cation. Metal-containing receptors are typically designed around an organic scaffold that binds metal cations with high kinetic and thermodynamic stability such that at least one coordination site on the metal center remains vacant or coordinated to aw eakly bound substrate and thus available for anion binding.I nt he past five years,n umerous metal-based anion receptors have been reported and, as with examples from the more distant past, they are dominated by Zn II ,C u II ,a nd, to alesser extent, trivalent lanthanide cations.
Zn II -dipicolylamine-based receptors,s uch as peptide derivative 7 [21] (Figure 3), remain ap opular class of compounds for the recognition of biologically relevant polyoxoanions in water. They exhibit high affinity for pyrophosphate anions in particular, and have been coupled to ar ange of fluorophores to facilitate applications in sensing and detection. Such receptors and their uses have been reviewed exte nsively by Jolliffe and co-workers,a nd so will not be further discussed in this Review. [22] Although direct anion to metal coordination can result in receptors exhibiting very high anion-binding affinities,t he   selectivities of simple metal coordination compounds can often be poor.H owever, by integrating metal cations into organic scaffolds which impose structural geometric constraints and/or provide supplementary supramolecular interactions,ahigher degree of guest selectivity can be realized. Forexample,T ripier and co-workers have demonstrated that ad ual metal-coordination/hydrogen-bonding approach can be effective for anion recognition in water, whereby the pendant polyammonium arm in Zn II complex 8 enhances the recognition capability towards ATPand glyphosate anions. [23] Theh ydrophobic effect has also be exploited to further enhance the anion-recognition capability of metal complexes. Reinaud et al. have used Cu II coordination to bind fluoride anions with high affinity (log K = 4.9) in water at pH 5.9, by embedding the metal center within ah ydrophobic cavity of acalix [6]arene 9 ( Figure 3). [24] Crucially,control experiments revealed that no binding of fluoride occurs in the absence of either the metal or the hydrophobic cavity,w hilst computational studies indicate that the fluoride guest interacts with two water molecules within the cavity,which suggests that the receptor binds ap artially hydrated anion. This lowers the dehydration cost of binding,w hilst further stabilization may occur through OH À p interactions with the aromatic walls of the calixarene.
Theu se of luminescent lanthanide complexes is established as an effective approach to anion binding and sensing in water, and has recently been extensively reviewed by Butler and Parker, [25] and so only selected recent examples are highlighted here.T he binding of anions to the vacant coordination site of aL n III complex is dominated by electrostatic interactions;h owever,s electivity can be achieved by tuning the nature of the ligand and the lanthanide cation itself.
Parker and co-workers have recently reported atriazacyclononane-Eu III complex 10 ( Figure 4) which binds oxoanions at the metal center by displacement of coordinated water molecules.T he complex was used to quantify bicarbonate anions in human serum by means of aratiometric analysis of the bright europium-based luminescence. [26] Vilar and coworkers have coupled aG d III complex with Zn II -dipicolylamine moieties to devise ap yrophosphate-responsive magnetic resonance imaging (MRI) contrast reagent 11. [27] The binding of pyrophosphate anions in water solution at aphysiological pH value modulates the relaxivity of the Gd III complex sufficiently to be observed in imaging experiments. In contrast, asupramolecular dimerization approach has been exploited by Tr ipier and co-workers to bind fluoride anions with high affinity in water. [28] Theaddition of fluoride to Eu III complex 12 results in the formation of astrong 2:1host-guest complex (log b = 13): structural data from single-crystal X-ray analysis reveals that, in addition to the bridging fluoride anion between the two Eu centers,the complex is further stabilized by p-stacking interactions and hydrogen bonding.T he strong fluoride association in water leads to very sensitive detection of the anion, to alimit of 0.46 ppb.
Whilst lanthanide complexes comprise the vast majority of examples of f-metal coordination compounds for anion binding,S chiaffino and co-workers have shown that actinide complexes can also be exploited for this purpose. [29] They have devised an eutral, water-soluble uranyl-salophen complex 13 ( Figure 4) that is able to bind the nucleotide polyanions ADP and ATPw ith high affinity (log K > 4) in aqueous buffer solution by direct coordination to the bound UO 2 2+ motif.

Neutral and Negatively Charged Anion Receptors
Theu se of charged receptors is undoubtedly the primary approach used for anion recognition in water. However, the disadvantage of such charged hosts is that they rely on nondirectional electrostatic interactions which can result in poor anion selectivity,w hilst accompanying counterions compete with the guest for the binding site.N eutral hosts are not affected by these disadvantages,b ut in general interact more weakly than their charged analogues with anions in solution, and as ar esult are usually confined to operating in low-polarity organic solvents.However, in recent years several research groups have reported elegant examples of electroneutral anion receptors that are able to operate in water, by exploiting both complementary hydrogen-bonding interactions and the hydrophobic effect to enhance the overall complex stability.F urthermore,e xamples of negatively charged hosts,w here anion binding occurs in ac entral hydrophobic cavity of ar eceptor solubilized by peripheral appended carboxylate groups,h ave also been developed. Examples of neutral or negatively charged hydrophobic hosts remain uncommon;however,the area is the subject of active current interest, and in the past few years ahandful of notable examples have been reported, and are discussed here.  Figure 5). [30] They subsequently showed that 1:1host-guest stoichiometric complexes could be prepared by joining two cyclopeptide macrocycles together with af lexible linker. [31] Thes ame research group has now reported ar elated host system 15 ( Figure 5) which exhibits enhanced aqueous solubility,thus enabling the recognition of iodide and sulfate in 95:5 D 2 O/CD 3 OD (log K = 3.70 and 3.96 respectively). [32] In both cases, ah ighly favorable entropic contribution is the dominant driving force for complex formation. Although not strictly belonging to this section, enhanced sulfate recognition was achieved in related cyclopeptide-derived hosts decorated with pendent ammonium groups to enable high-affinity binding of the anion (log K = 5.1) in 1:1m ethanol/water solution at pH 4.8, driven by both favorable entropy and enthalpy. [33] Very recently,Sindelar and co-workers have raised the bar on what is achievable in terms of anion-recognition strength in water by hydrophobic host molecules,w ith their report of aw ater-soluble variant of the bambusuril macrocycle 16. [34] Theh ost binds various inorganic anions in water within the hydrophobic cavity in an enthalpy-driven process,w hilst additional stabilization of the complex arises from multiple CHÀanion hydrogen-bonding interactions.T he receptors bind the anion guests with high affinity in phosphate buffer solution, with the affinity ranging from ca. 10 3 m À1 for Cl À ,to ca. 10 5 m À1 for Br À and NO 3 À ,with the strongest binding being observed for I À and ClO 4 À (K % 10 7 m À1 ). Thea uthors ascribe these remarkable affinities to the total isolation of the bound anion from water molecules by inclusion within the hydrophobic bambusuril cavity.
Ar elated cyclic hexamer has also been shown to bind anions in water by Pittelkow and co-workers. [35,36] They reported the halide-anion-templated synthesis of am acrocyclic 6 + 6h examer 17 of d-biotin and formaldehyde,a nd demonstrated that the macrocycle binds ar ange of monovalent anions in water at pH 7.5 in phosphate buffer.T he binding of softer anions such as À SCN,I À ,a nd SeCN À is favored, with association constants ranging from log K = 4.5 for SCN À down to log K = 1.8 for Cl À .Thermodynamic anionbinding titrations using isothermal titration calorimetry (ITC) revealed that anion association in all cases is enthalpically driven, characteristic of anon-classical hydrophobic effect in which the anion liberates high-energy water molecules from the hydrophobic cavity of the macrocycle.F urthermore,t he single-crystal X-ray structure of 17 with iodide provided evidence for C À Hh ydrogen-bonding interactions between the host and anion.
Thec ombination of C À Hh ydrogen bonding and the hydrophobic effect has also been elegantly used by Flood and co-workers for the recognition of chloride anions by the aryltriazole foldamer 18 in organic-aqueous solution (Figure 6a). [104] Hydrophobic interactions are responsible for organizing and stabilizing the foldamer,w hich forms a2 :1 stoichiometric complex with chloride in 1:1C H 3 CN/H 2 O solution, with about 80 %o ft he hydrophobic p surfaces buried within the duplex in the process.T he chloride anion is bound by eleven triazole C-H-Cl À hydrogen-bonding interactions within ahydrophobic pocket, which shields the anion from the surrounding solvent. Thef ormation of the chloride duplex is driven almost entirely by favorable enthalpy,with an overall stability constant of b 2 = 910 12 m À2 .T he addition of al arge excess of chloride results in the equilibrium being pushed in favor of a1 :1 stoichiometric single helix complex, the crystal structure of which is shown in Figure 6b.I nt his case,o nly about 50 %o ft he hydrophobic p surfaces are buried in the foldamer,thus accounting for its lower stability than the duplex structure (K 1 = 210 5 m À1 ). It is noteworthy that the formation of the 2:1d uplex is ah ighly cooperative process,w ith a K 2 /K 1 ratio of 165 (where K 2 is the stepwise duplex association constant, and K 1 is the association constant for the 1:1c omplex).
Thea forementioned examples exploit the hydrophobic effect to achieve the strong binding of anions in aqueous solution. However,aspreviously mentioned, the hydrophobic effect is arguably the least well understood factor that drives host-guest complexation in solution. Indeed, the traditional view of describing the (classical) hydrophobic effect in terms of entropic arguments does not hold true for many examples of host-guest complexation, in which ad ominant favorable enthalpy drives the association, and may be described instead by the non-classical hydrophobic effect. Them icroscopic origin of the favorable negative enthalpy thermodynamic signature is ascribed to the release of high-energy water

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Reviews molecules that occupy the hydrophobic binding domain of the host (and thus form fewer hydrogen bonds than in bulk water), thereby leading to the formation of more cohesive water-water interactions and ag ain in enthalpy.T his phenomenon was first described by Diederich and co-workers in 1991 in the context of the recognition of arene guests by cyclophanes, [37] but recent studies have revealed high-affinity binding of anions in water in which the non-classical hydrophobic effect is proposed to be the dominant thermodynamic driving force.I nb oth the biotin hexamer reported by Pittelkow and co-workers and the foldamer reported by Flood and co-workers,s trong anion binding is driven exclusively by favorable enthalpy:the thermodynamic signature of the non-classical hydrophobic effect.
Ar elated phenomenon often cited in relation to anion recognition is the Hofmeister effect. [38] TheHofmeister effect has historically been viewed in terms of makers and breakers of water structure,o riginally in the context of protein solubility (kosmotropic anions increase water structure,t hus enhancing the hydrophobic effect and decreasing protein solubility,w hilst chaotropic anions decrease water structure, thus weakening the hydrophobic effect and increasing protein solubility). However,the role of ion-solute interactions is also important, and the nuanced relationship between these effects and anion-recognition phenomena is only now beginning to be understood and investigated in detail.
In an elegant recent study,Gibb and Gibb investigated the Hofmeister effect and its relationship with the hydrophobic effect in the context of anion binding within acavitand host 19 (Figure 7). [39] Thea uthors demonstrated that kosmotropes, such as fluoride,acetate,and sulfate,enhanced the binding of ah ydrophobic guest (adamantane carboxylate) within the cavitand, whilst chaotropes,s uch as perchlorate,d ecreased the association. This latter effect was ascribed to competitive binding of these anions within the cavitands hydrophobic pocket. Perchlorate,inparticular,was shown to bind surprisingly strongly (K a = 95 m À1 ), and am ore recent investigation has shown that the binding of the anion is dependent on both the nature and concentration of co-salts. [40] Theoverall trend (in which chaotropic anions decreased perchlorate anion association whilst kosmotropic anions enhanced it) directly reflects the Hofmeister series.T he authors proposed that the former may be explained in terms of competitive binding of relatively poorly hydrated and polarizable anions leading to an overall decrease in perchlorate association, whilst, in contrast, strongly hydrated salts enhance anion association as ar esult of cation binding to the cavitands peripheral carboxylate groups and thus reduce the net negative charge of the host. These results demonstrate an unprecedented molecular insight into the role of ion-solute interactions in the Hofmeister effect, and provide al ink between this hitherto ambiguous concept and those of hydrophobicity and anion binding.
Theb inding of chaotropic anions within hydrophobic cavities has also recently been explored by Nau and coworkers,w ho investigated the binding of negatively charged dodecaborate clusters of the form B 12 X 12 2À and B 12 X 11 Y 2À (where X = H, Cl, Br,a nd I; and Y = OH, SH, NH 3 + ,a nd NR 3 + )w ithin g-cyclodextrin. [41] Such clusters were found to bind with remarkably high affinity (log K = 3-6), but with low selectivity between anions of differing size,a nd driven exclusively by as trong enthalpic contribution to the binding whilst disfavored by entropy.T he authors ascribe these data to two effects:1 )increased dispersion interactions between the anion and the cyclodextrin compared to those between the anion its solvation shell, and 2) the so called "chaotropic effect", in which the borate clusters act as chaotropic anions and disrupt the local water structure.F ollowing the latter argument, the negative complexation enthalpy is explained by the re-formation of the broken hydrogen bonds in the bulk aqueous solution upon binding of the anion in the hydrophobic cavity.T he negative entropy change arises from the loss of structural entropy in the bulk water upon binding of the cluster and the subsequent re-formation of hydrogen bonds.T his recovery of hydrogen bonds as as upramolecular driving force is analogous to the common description of the non-classical hydrophobic effect, such as in the case of anion binding by the aforementioned biotin cyclic hexamer 17, where the release of high-energy water molecules from within

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Reviews the macrocyclic cavity,a so pposed to those surrounding the chaotropic anion, is driven by the enthalpically favorable reformation of hydrogen bonds with the bulk solvent. This study highlights the need for considering in detail the solvation of both the guest and host, and the influence of high-energy water molecules within both the host cavity and surrounding ac haotropic anion on the thermodynamics of hydrophobicdriven anion binding in water.

Halogen-Bonding Anion Receptors
Halogen bonding, [42] the attractive interaction between an electron-deficient halogen atom and aL ewis base,h as in recent years attracted much interest as an intermolecular interaction analogous to the ubiquitous hydrogen bond. [43] Whilst extensively utilized in the solid state,its application in the solution phase is less developed, but has recently been shown to be an effective interaction for anion recognition in organic solvents. [44] However,itwas only in the last year that seminal examples of halogen-bonding receptors capable of anion recognition in water were developed. In 2014, we reported ar otaxane host 20 capable of exclusively binding halide anions over ar ange of oxoanions in aqueous organic solvent mixtures through four convergent halogen bonds in the cavity formed between the interlocked components ( Figure 8). [45] An incorporated luminescent rhenium(I) bipyridyl motif in the macrocycle facilitated the study of the anionbinding properties of the rotaxane by allowing the change in the Re I metal-ligand-charge-transfer (MLCT) emission to be monitored. This revealed aselectivity trend of I À > Br À > Cl À in 1:1CH 3 CN/H 2 Osolution, with remarkably high affinity for ad oubly charged receptor (log K a = 4.4, 3.8, and 2.7, respectively).
We later published as eminal study to quantify halogenbonding interactions in 100 %w ater, and demonstrated that the intermolecular interaction is superior to hydrogen bonding for anion recognition in aqueous solution. [46] Anionbinding titrations in D 2 Owith aseries of acyclic and rotaxanebased receptors containing the bidentate 3,5-bisiodotriazole pyridinium halogen bond donor motif,a nd solubilized with permethylated b-cyclodextrin derivatives,revealed aremarkable enhancement in the binding affinity for halide anions when compared to analogous C À Ha nd amide N À Hh ydrogen-bond donors.F or example,X Br otaxane 22 (Figure 9) binds iodide with high affinity in water (K a = 2200 m À1 ), two orders of magnitude more strongly than the analogous 3,5bisprototriazole pyridinium CÀHhydrogen-bonding rotaxane system (K a = 40 m À1 ). Remarkably,t he halogen-bonding acyclic receptor 21 is also capable of anion recognition in water despite its simplicity and low charge.V ant Hoff analysis of the iodide recognition process by XB-rotaxane 22 revealed that the binding is enthalpically driven. This is in contrast to the analogous rotaxane hydrogen-bonding systems,i nw hich the association is driven by entropy,t hus reflecting both the charge-transfer contribution to XB-anion interactions [47] and differing desolvation thermodynamics of XB donors that contribute to the remarkable anion-binding properties.A later study,i nw hich the solubilizing b-cyclodextrin derivatives of ah ydrogen-bonding rotaxane were modified, revealed that these stoppers play am inimal contribution to the overall anion-binding affinity of the host in water, thus suggesting that aw ider range of solubilizing stopper groups may be employed without adversely affecting the anionrecognition capability of the host system. [48] Indeed, we have since employed polyethylene glycol derivatives to solubilize ab isiodotriazolium-based acyclich alogen-bonding receptor 23,w hich is capable of enhanced perrhenate recognition in

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Reviews D 2 Oc ompared to the hydrogen-bonding analogue,w hich is again enthalpically driven. [49]

Macromolecular Recognition of Anions in Water
Small-molecule receptors have dominated anion-binding and anion-sensing technologies to date,largely because of the potential for the direct rational design and simple delineation of structure-function relationships.However,there is much to be said for macromolecular and nanotechnological approaches to anion recognition, particularly when faced with the challenges of aqueous media. Larger systems allow the construction of solvent-excluded microdomains to maximize binding strength, the exploitation of multivalencya nd cooperative recognition, signal amplification, and fine control of material properties such as solubility and rheology.
With the exception of ah andful of small-molecule anion transporters such as prodigiosin, it is macromolecules which have been selected in nature to perform highly selective anion-binding tasks.Inrecent years there have been anumber of important advances in the study of anion binding by biomacromolecules,w hich we will summarize in Section 3.1; the discussion will not be exhaustive (the number of proteins which bind ATPm akes that task somewhat herculean), but we will consider the most significant advances from the viewpoint of supramolecular design.

Biomolecules
Living systems must manipulate anions in water in ahuge number of sophisticated ways and, therefore,b iology is an important point of reference for anion recognition in water.
Thescientific response to the apparent finding of bacteria (GFAJ-1) living in an arsenic-rich, phosphate-free environment, [6] and suggesting the existence of arseno-DNA, was comprehensive:chemists noted that the rates of hydrolysis [50] and reduction [51] of arsenate esters would predict ap rohibitively low stability for arseno-DNA; furthermore,b iologists showed that trace levels of phosphate were present and sufficient for bacteria, [7] and found no detectable arsenic in their DNA. [52] It transpired that the bacteria are able to tolerate high arsenate/low phosphate environments because they can be extremely selective in their uptake of the two: ap hosphate-binding protein (PBP) up-regulated by GFAJ-1inphosphate-poor conditions has a4500-fold preference for phosphate over arsenate.T his is especially striking considering that the sizes of the anions differ by only 4%,t heir pK a values are nearly identical, and their oxygen atoms are very similarly charged. Them olecular basis for this discrimination was elucidated the following year [53] by Elias,W ellner et al.,w ho discovered that an etwork of dipole-anion interactions and repulsive interactions were responsible. Both phosphate and arsenate anions are bound in their monoprotonated forms,a nd the locations of the three nonprotonated oxygen atoms are firmly anchored, thus maximizing the directionality of the size difference ( Figure 10). In the case of arsenate,this results in asteric clash between the protonated oxygen atom with two Cb atoms on nearby residues and distorts al ow-barrier hydrogen bond between the OH group of the anion and ap roximal aspartate side chain. Conversely,all these parameters are optimal when the cleft is occupied by phosphate.T his astounding example of natural selectivity serves as ag old standard for synthetic anion hosts:i ti sp ossible not just to bind anions strongly in water, but to have total control over selectivity between very similar targets.
Another important finding for anion-binding proteins was the identification by Watson and Milner-White of aparticular fold, named the "nest", which can occur with am inimum of just two residues (Figure 11 a). [54] Then est is described by residues with alternating main-chain dihedral angles of f,y % À90,08 8 and f,y % 70,408 8 (or their negative). When two adjacent residues display this conformation, an atom-sized concave space is formed into which the two main-chain NH groups plus that of the following amino acid point, thus creating as pace suitable for coordination of an anionic or d À atom (Figure 11 a). Thea rrangement and spacing of NH groups is analogous to that of oxygen atoms in crown ethers. This observation generalized an umber of previously known specific anion-binding motifs such as the alpha turn and the paperclip/Schellman loop. [55] Compound nests consist of longer tracts of residues displaying these dihedral angles and results in larger cavities such as those known as P-loop,w hich is the most commonly found ATPorguanosine triphosphate (GTP) binding motif in proteins and contains af our-residue nest (Figure 11 b). In serine protease,a nother four-residue nest is used to stabilize atetrahedral oxyanion intermediate,while nests of up to eight residues are often found supporting iron-sulfur clusters. [58] These observations have led to new anion hosts,afirst example of which is ahelical hairpin eicosapeptide which was Figure 10. Binding environment of hydrogen phosphate (orange) and hydrogen arsenate (blue) by GFAJ-1 phosphate-binding protein (PDB 4F19 and 4F18, respectively). Note the clamping of unprotonated oxo groups, which causes the protonated oxygen atom to be maximally displaced, and subsequent distortion of hydrogen-bonding geometry at the protic site of the anion.

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Reviews found to crystallize (albeit from organic solvents) with an included acetate anion in an est bridging the two helices, [59] and in 2012 am inimal hexapeptide was designed that contained aP -loop-type nest which was shown to bind HPO 4 2À strongly in water at neutral pH. [60] Interestingly,i t appeared that adeprotonated amine from alysine side chain accepts ah ydrogen bond from the phosphate,a ss een in phosphate-binding proteins.
As imilar anion-recognition moiety to the nest is the C a NN motif,w hich consists of two main-chain amide hydrogen bonds and aC ÀHhydrogen bond from the next a-carbon atom (Figure 11 c). [61] Thesystem was initially put forward as ap hosphate-recognition unit (Figure 11 d), but by attaching Leu-Gly-Lys-Gln, aC a NN fragment, to the Nterminus of an anchor helix, Sheet and Banerjee observed sulfate recognition in water manifested as achange in the conformation that was typical of an "induced fit" system. [62] Computational analysis confirmed earlier crystallographic studies showing that both sulfate and phosphate binding triggers aconformational switch from anonhelical to ahelical morphology. [63] In addition to these findings,there have been anumber of incremental discoveries related to the binding of oxoanions [64] which reveal interesting aspects of natural anion-coordination strategies.T he binding of nitrate by cyanobacteria provides nitrogen fixation for entire aqueous ecosytems.I nt he open ocean, such ions are scarce,a nd so the cyanobacteria need highly evolved capture systems.I nN rtA, an itrate binding protein which is tethered to the extracellular side of the cyanobacterial membrane as part of at ransporter system, nitrate is bound in ad istinctly asymmetric fashion. [65] While all three of the nitrate oxygen atoms are indistinguishable in water, when bound, the charge is localized on just one oxygen atom, with complementary positively charged groups focused about just that atom. Theo ther two oxygen atoms are more weakly hydrogen bonded, with one in am arkedly hydrophobic cleft, thereby enabling the protein to also host the nitrite anion. Thesystem is highly homologous with ac yanobacterial bicarbonate binding protein, CmpA, which may also relate to its asymmetric coordination mode. [66] Another nitrate receptor was discovered as part of the nitrate regulatory element (Nre) of staphylococci, which enables the bacteria to reduce the oxoanion instead of oxygen during anaerobic respiration. Thebinding pocket is primarily hydrophobic,p rovides only four hydrogen bonds,a nd could also host iodide anions;unusually,the proteins biological function could also be activated in the presence of iodide. [67] It was discovered in 2009 that bacterial uptake of molybdate and tungstate was enabled through modification of the anion itself:u pt ot hat time all known metalloprotein structures containing Mo or Wdisplayed tetracoordinate metal centers, but octahedral coordination was encountered in transporter proteins,w ith two carboxylate groups providing extra coordination directly to the metal centers while hydrogen-bond donors complemented the oxygen atoms. [68] Such af inding could provide inspiration for new strategies to bind multiatomic anions.M otivated by the possibility of noncovalent encapsulation of radionuclides for nuclear medicine,t he molybdate-binding protein has been re-engineered to show selectivity for perrhenate as am odel for pertechnetate (for 99m Tc) and for b-emitting 188 Re and 186 Re in its own right. [69] Nucleic acids,t he other major class of biopolymer,m ay seem unlikely candidates for anion coordination because of the multiply negatively charged phosphate backbone,b ut there are some interesting possibilities raised by the prospect of aptamers.N ucleic acid aptamers are single-stranded oligonucleotides which fold in ap articular way to bind as pecific substrate [70] in aqueous media (nucleic acids being poorly soluble in anything else). Charge screening required for folding and recognition is achieved through the addition of inorganic cations.A ptamers are alternatives to antibodies, and can be "raised" through am ore simple in vitro selection process against any potential target, [71,72] ranging from monoatomic ions through small molecules and proteins,u pt o whole cells.T odate they have been only sparsely explored in anion recognition, despite examples of metal-ion binding being quite numerous. [73,74] Aptamers for nucleoside phosphates are well known. [75] In particular, an ATPa ptamer has been discovered which is highly selective for the longer oligophosphates over the shorter analogues,which shows that nucleic acids can even bind polyanions strongly and selectively (in the presence of Mg 2+ ions). [76] An aptamer for AMP has been coupled to fluorescent nanoparticles,t hereby providing ah ighly selective sensor for that target. [77] Besides this,a ptamers have been reported for biologically relevant anionic porphyrins, [78] again highlighting the possibilities for further exploitation in aqueous anion-recognition chemistry. Very recently,D NA-like oligomers with synthetic units replacing nucleobases were reported for pattern-based detection of anion pollutants in water. [79] Al ibrary of 1296 tetrameric strands was created on beads from six monomers, including two metal-binding ligands and three organic fluorophores.M etals were added to provide binding sites, Figure 11. a) Simplified illustration of the most basic nest motif. b) GTP analogue GppNp bound by hydrogen bonds from five consecutive main-chain amides in an est formation (PDB 5P21 [56] ). c) Simplified illustration of the C a NN motif. d) Binding of aphosphate group by aC a NN motif located at the end of ahelix (PDB 1AJS [57] ). and the fluorescent response of the beads examined using 17 anions (including underexamined examples such as selenite, arsenate,b romate,a nd permanganate) in aqueous buffer solution. Them ost responsive systems were selected and narrowed down to aset of eight, which in combination could discriminate between all the anions and measure their concentrations at micromolar concentrations.

Liposomes and Membrane Transport
Biological anion binding frequently relates to crossmembrane transport, and synthetic mimicry of anion binding is,t herefore,o ften motivated by related biological and medicinal possibilities.T ransmembrane anion channels [80] have been made,a sw ell as transporter systems,l argely led by Gale and co-workers. [81][82][83] In these studies,a na nion host (typically an uncharged small molecule) is added to abuffered aqueous solution containing liposomes with an anion concentration gradient imposed across the membrane.T he resultant transport across the membrane is then measured. It is currently an open question as to whether anion coordination occurs in water with the anionophores freely entering the aqueous phase (where some can certainly be present [83] )o ra tt he membrane interface with transporters confined within the leaflets.Some studies report the strength of the anion-host interaction, but only in low-water media. [84] Them obility of the anionophore likely relates to lipophilic balance [83] -the transporter must be able to get through the hydrophobic membrane,w hile still bind the anion-and also potential for self-assembly within the bilayer to give channels. [85,86] Whatever the case,the entire supramolecular system can be viewed as an anion host. One outstanding example of this area is use of small molecular isophthalamides for DNA transfection by Gokel and co-workers ( Figure 12). [87] By employing av ery simple anion host system, it was possible to transfect plasmids of > 20 kbase with ampicillin resistance across the membrane into bacterial cells,thereby resulting in robust colony formation. Importantly,t he plasmids trans-fected were larger than those previously thought possible by chemical means.

Polymers
Polymers are particularly attractive for recognition applications as aconsequence of their multivalent effects,creation of microdomains,tunability of solubility and physical properties,a nd processability for materials production. Despite these advantages,t he field is largely undeveloped. Foundational studies on the interaction of anions with polyolefins were carried out aq uarter of ac entury ago,w hen Jhon and co-workers established that NMR line broadening of the 127 I signal in water in the presence of polymers such as PHEMA, PA AM, and PVP was due to binding (Figure 13 a). [88] In the case of PVP,the binding strength was found to correlate with the Hofmeister series,w ith the more chaotropic anions interacting most strongly. [89] These studies were picked up only in the last ten years with PNIPAM, which is al ower critical solution temperature (LCST) polymer,m eaning that it is insoluble in water above ac ertain temperature.C remer and co-workers found that that the transition temperature was lowered by more hydrated (i.e.kosmotropic) anions and correlated with the hydration entropy. [90] Less hydrated anions (ClO 4 À > SCN À > Br À > NO 3 À ), on the other hand, actually salted in the polymer at low concentration, thereby raising the transition temperature,w hile at high concentrations the increased surface tension resulted in salting out. Analysis of the concentration/LCST curves revealed ac omponent of direct anion binding was responsible for the improved solubility,w hich was presumed to be occurring through the amide group as ah ydrogen-bond donor. The structural basis for salting-in was analyzed in more detail using PDEA, an LCST polymer with at ertiary rather than secondary amide,a nd hence no apparent anion-binding site. [91] Combined NMR and IR spectroscopic studies showed that the same chaotropic anions were interacting with the a-CH proton on the polymer backbone.W hile this may seem unlikely at first glance,i ti sc onsistent with the involvement of a-CH peptide protons in anion binding,asin the C a NN motif (see Section 3.1).
Theutilization of polymers for anion recognition in water has centered on exploitation of the photophysical properties of conjugated systems for sensing, [92] for the most part by inducing the kind of desolvation discussed above.F or example,b yd ecorating poly(phenylene-ethynylene) with cationic primary ammonium side chains (Figure 13 b), it is possible to sense pyrophosphate through ar ecognitioninduced aggregation mechanism that gives ar atiometric fluorescent response. [93] Conversely,p oly(thiophene) decorated with pendant quaternary ammonium groups (Figure 13 c) disaggregates in water in the presence of iodide and sulfonate-based surfactants to give ar ed-to-yellow color change with as ensitivity down to the nanomolar regime. [94] Poly(fluorene-b-thiophene) has also been functionalized with tertiary ammonium groups on the thiophene block (Figure 13 d), and the fluorescence of that section was found to quench with halides in water (Cl À > Br À > I À ), although no Figure 12. Transfection of DNA macromolecules across cell membranes for genetic modification using small anion hosts. Adapted from Ref. [87] with permission.C opyright2 012 American ChemicalS ociety.

Angewandte Chemie
Reviews change of aggregation state was observed in this case. [95] By using the constant fluorescence of the poly(fluorene) block, it was possible to measure DNAc oncentrations down to the sub-micromolar level.

Nanomaterials
Aggregation-induced spectral changes also dominate examples of synthetic nanomaterial systems for anion recognition in water. Thep lasmon resonance band of gold nanoparticles (AuNPs) is highly sensitive to the aggregation state, and by using anions to bring nanoparticles together,i ti s possible to construct sensors.A mide ligands grafted onto poly(vinyl alcohol)-stabilized AuNPs (Figure 14 a) result in discrimination between the isomeric dicarboxylates fumarate and maleate in water:t he former give ap urple solution caused by bunching of the particles,whereas other dicarboxylates give red solutions. [96] Aqueous solutions of thiouronium-decorated AuNPs (Figure 14 b) also give selective aggregation-induced color changes in the presence of hydrophobic anions.Inthis case,ani nteresting approach was used to generate afluoride sensor by adding aphenylboronic acid to the solution and sensing the resultant fluoroborate. [97] Apparently,d irect fluoride-induced aggregation was seen with thioglucose-capped AuNPs in buffer (Figure 14 c). [98] Cationic imidazolium ligands on AuNPs (Figure 14 d) gave strong aggregation in the presence of hydrophobic anions such as tetrafluoroborate and hexafluoroacetate,t hereby making the constructs insoluble in water and facilitating phase transfer into ionic liquid media. [99] Theuse of the same ligands on magnetic iron oxide NPs enabled DNAcatch-andrelease through stepwise anion exchange:s tarting chloride anions were displaced by DNA, which could then be released through displacement by hydrophobic anions. [100] Similarly, imdazolium-functionalized multiwalled carbon nanotubes are soluble in water only in the presence of chloride, thus providing an anioncontrolled aggregation switch. [101] Although there are few examples of synthetic macromolecules for anion recognition in water, there are further types of nanosystems which have not yet been explored at all. Dendritic systems,f or example,h ave yet to be explored. This is despite the systems potentially displaying the "dendritic effect" that gives rise to signal amplification, as observed by Astruc and co-workers,i nl esswater-soluble ferrocene-based systems. [102,103] Thew hole range of nanocarbons has barely been touched, while nanoparticles of non-gold elements are also virgin territory.There is vast scope in these fields for researchers aiming to create potent and functional materials for the challenge of anion recognition in water.

Summary and Outlook
Thef indings surveyed above clearly illustrate that an umber of approaches can now be exploited to achieve the recognition of anions in water. Effective strategies both for the binding of guests and solubilization of small-molecule hosts have been outlined. Some are obvious,s uch as charge, but others such as the hydrophobic effect require much more in-depth research to fully understand. New anion-binding methods such as utilizing CH hydrogen bonds and halogen bonding feed into this theme and allow the creation of hydrophobic,s olvent-separated binding domains for finetuning the recognition through control of the microenvironment. Thew orld of biomolecules continues,a se ver,t o provide new insights and inspiration for tuning selectivity, while polymers and nanomaterials are just starting to emerge as promising candidates for anion extraction and sensing materials of tomorrow.M ost importantly,a sa queous anion coordination chemistry becomes more mainstream, many of the long-term proposed applications in the industrial, environmental, biological, and medical arenas will develop into real-life technologies.