Journal of the Mexican Chemical Society ISSN: X Sociedad Química de México México

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1 Journl of the Mexicn Chemicl Society ISS: X Sociedd Químic de México México Rosete-Lun, Shron; Medrno, Felipe; Bernl-Uruchurtu, Mrgrit I.; Godoy-Alcántr, Crolin Crown Ether Ditopic Receptors for Ammonium Slts with High Affinity for Amino Acid Ester Slts Journl of the Mexicn Chemicl Society, vol. 53, núm. 4, octubre-diciembre, 2009, pp Sociedd Químic de México Distrito Federl, México Avilble in: How to cite Complete issue More informtion bout this rticle Journl's homepge in redlyc.org Scientific Informtion System etwork of Scientific Journls from Ltin Americ, the Cribben, Spin nd Portugl on-profit cdemic project, developed under the open ccess inititive

2 Article Crown Ether Ditopic Receptors for Ammonium Slts with High Affinity for Amino Acid Ester Slts Shron Rosete-Lun, Felipe Medrno, Mrgrit I. Bernl-Uruchurtu, nd Crolin Godoy-Alcántr* Centro de Investigciones Químics, Universidd Autónom del Estdo de Morelos, Cuernvc, Morelos, México. Tel/Fx: , Received August 13, 2009; ccepted ovember 19, 2009 J. Mex. Chem. Soc. 2009, 53(4), , Sociedd Químic de México ISS X Abstrct. Two bis crown ether receptors were synthesized nd tested s host molecules for protonted forms of lkyl mines nd mino cid esters. Moleculr recognition studies were conducted in CH 2 Cl 2 :MeH (92:8) by spectrophotometric UV/Vis titrtions nd by spectrometric 1 H MR titrtions in CDCl 3. The clculted binding constnts re in the rnge M -1. A high ffinity for L-mino cid methyl ester derivtives ws found. A theoreticl study t the DFT level of the synthesized receptor nd some nlog lignds with three different mmonium ions helps to rtionlize the experimentlly found trends. Key words: Ditopic Receptors, Binding Constnt; Crown Ethers, Alkyl Ammonium, Amino Acid. Introduction Resumen. En este trbjo se sinterizron dos receptores del tipo bis eter coron y fueron estudidos como moléculs nfitrión de lquilmins y ésteres de minoácidos protondos. Se relizron estudios de reconocimiento moleculr por titulciones espectrofotométrics UV/Vis en CH 2 Cl 2 :MeH (92:8) y por titulciones espectrométrics de RM 1 H en CDCl 3. Ls constntes de enlce clculds están en el intervlo M -1. Se encontró un lt finidd por los ésteres metílicos de los L-minoácidos estudidos. Un estudio teórico nivel DFT de los receptores sinterizdos y lgunos ligndos nálogos con tres diferentes iones monio permite explicr ls tendencis experimentles encontrds. Plbrs clve: Receptores ditópicos, constnte de enlce, éteres coron, lquilmonio, minoácido. Moleculr recognition of mines is n interesting field due to their widespred presence in nture. Mny biogenic mines such s histmine, cdverine, nd putrescine re used s probes to mesure the qulity of mrine food products [1]. Tyrmine is potent bcteril mutgen tht induces tumors t multiple sites in rodents [2]. Dopmine is well-known neurotrnsmitter, nd epinephrine (drenline) is n drenl hormone. The entire clss of lkloids comprises toxic mines while liphtic mines hve mny pplictions in the chemicl industry some of them re considered environmentl hzrds. The development of effective receptors for mines hs been n re of considerble interest for mny yers. Crown ethers nd heterocrown ethers constituted the erliest hosts or chemosensors for mines, which were recognized primrily in their mmonium forms [3]. Metlloporphyrin hosts were developed to bind neutrl mines by coordintion to metl ctions [4]. These hosts were ble to recognize mines in both orgnic solvents nd wter [5]. Dynmic covlent chemistry dded new dimension to mine recognition, for exmple, the rection of neutrl, primry mines with crbonyl groups in dyes to form fluorescent imines [6]. Zimmermn nd co-workers provided n element of selectivity by prepring moleculrly imprinted polymer dye for the detection of mines [7]. However some of these hosts hve solubility problems, i.e., they re soluble in nonpolr orgnic solvents while the mmonium ions require polr solvents which limited the potentil nlyticl procedures of study to liquid-liquid extrctions [8-13] nd trnsport experiments [14-17] nd only some host-guest complex hve been studied in solution by 1 H MR [8, 18, 19] or by clorimetric titrtions [20, ]. In Tble 1 we summrize the nlyticl methods used for their study nd their corresponding stbility constnts. Functionlized benzocrown ethers re desirble cndidtes for mine receptor molecules becuse of their well known bility to form strong complexes with orgnic ctions [3d]. In this work we report the use of two ditopic bis(crown) ethers to detect protonted mine nd mino cid derivtives in solution. In Scheme 1 the generl preprtion method used is presented nd in Chrt 1 the guest molecules used re indicted. The receptor molecules hve in their structure two crown ether moieties linked by conjugted romtic bis zomethine unit. These structurl units cn bind the mmonium group of the guest molecule through H-bonds. The romtic nd conjugted systems in the receptor molecules open the possibility for dditionl ϖ-ϖ interctions or vn der Wls contcts with the lkyl or romtic groups of mmonium guest. The ionophores re electroniclly connected which might introduce llosteric control over the recognition properties. This prticulr type of ditopic receptors hs been used s hosts for lkli nd lkline metls [22] but, to the best of our knowledge, mine nd mino cid complexes hve not been studied yet. In this work we report the ffinity of the bis crown ether receptors towrds protonted mines nd mino cid ester derivtives in solution. Using theoreticl methods t the DFT level we propose the structure of these complexes nd some insight on the origin of the trends of the selectivity experimentlly observed.

3 0 J. Mex. Chem. Soc. 2009, 53(4) Shron Rosete-Lun et l. Tble 1. Binding constnts for selected host-guest complexes of mmonium orgnic ctions. Host Guest K (M -1 ) Method Solvent Reference chirl 18-crown-6 derivtives with lipophilic chins clix(4)-crown ethers bis-crown ether peptidic receptors 18-crown-6 nd mcrocyclic xnthone-bsed receptor bis(crown-ether) nlogue of Tröger s bse tetrzmcrocyclic (cyclidene) nickel(ii) complex bering two crown-ether residues meso-ternphthlene bering two crown ethers 18-crown-6 18-crown-6 benzo-18-crown-6 chirl mmonium picrtes lkyl mmonium picrtes bismmonium lkyl picrtes H MR Extrction experiments CD 3 C CHCl 3 :H 2 ( ) 10 4 Extrction experiments CH 2 Cl 2 :H Extrction experiments CHCl 3 :H mino cids 1-9 Extrction experiments CHCl 3 :H 2 11 monommonium nd bismmonium chlorides bismmonium thiocyntes or perchlortes (6-7.4) 10 3 (5-78) H MR CDCl 3 :MeH-d H MR CD 3 C 19 bismmonium picrtes Extrction experiments CHCl 3 :H 2 12 orgnic mmonium hlides protonted mino cid methyl esters protonted mino cid methyl esters Clorimetric titrtion MeH Clorimetric titrtion Clorimetric titrtion MeH MeH 8 Rtio between the two ssocition constnts of both distereomeric complexes. H H + Reflux 6h Ethnol H H H + H 2 Reflux 6h Ethnol Scheme 1. Synthesis of the host molecules studied in this work.

4 Crown Ether Ditopic Receptors for Ammonium Slts with High Affinity for Amino Acid Ester Slts 1 Cl H 3 CH C CH Cl - H 3 3 CH C CH - H Cl - Cl - H 3 CH C CH 3 CH 2 D CH 2 CH 3 A F H 3 Cl- B H C E H 3 Cl - H 3 Cl - H 3 Cl- G H Chrt 1. Chemicl structures of the guest molecules studied in this work. Results nd Discussion Electronic spectr of receptors ssocited with ion binding Using mixture of methylene chloride nd methnol (92:8) the bsorption mxim of receptors 1 nd 2 ppers t 387 nd 352 nm, respectively. The observed trnsitions re of ϖ-ϖ* nture nd re ttributed to the extended ϖ system formed by the three romtic rings connected by two imine groups present in the receptor structure. The bnd is highly sensitive to the polrity of the solvent, methnol ddition to CH 2 Cl 2 solution of receptor 1, leds to hypochromic effect of 380 nm bnd (Supplementry mteril) This solvtochromic behviour hs been explined s due to hydrogen bond formtion between imine groups of the receptor molecule nd the hydroxyl group of the methnol [23]. Addition of the mmonium slts result in hypsochromic nd hypochromic shifts of the receptors bnds s it is shown in Fig. 1. These shifts might be explined in terms of the conformtionl chnges occurring upon complextion on the, -(p-phenylenedimethylidyne)diniline structurl unit tht connects both crown ethers (vide infr). Receptor molecules re fluorescent in cetonitrile solution with two emission bnds locted t 307 nm nd 355 nm when 250 nm excittion rdition is used. Titrtion of the receptors with lkyl mmonium slts induces modest enhncement of fluorescence intensity which cn be rtionlized in terms of photoinduced electron trnsfer effect due to the coordintion of non-bonded oxygen electron pirs of the crown ethers. However, clcultion of ny ssocition constnt from these dt ws not possible. Binding constnts The binding constnts between receptors 1 nd 2 nd the mmonium slt guests were determined by UV/Vis nd 1 H Absorbnce λ (nm) Fig. 1. Chnges of the bsorption spectr during the titrtion of 1 ( M) with F ( M) in CH 2 Cl 2 :MeH (92:8). The rrow indictes the direction of spectrl chnges occurring with the incresing concentrtion of F. MR mesurements. Complex stoichiometry for both receptor molecules ws determined by mens of continuous vrition (Job) method [24], using UV/Vis spectroscopy, which gve minimum t mole rtio of 0.5 for 1:1 stoichiometry which corresponds to the stronger complex formed, however Job grphs were no symmetricl suggesting the presence of complexes of higher stoichiometry (Fig. 2). Although the Job s plots indicte the formtion of 1:1 host-guest complex the experimentl dt suggested more complex ssocition scheme, i.e., 1:2 host:guest stoichiometry. The chnges in the observed bsorbnce s function of the guest concentrtion

5 2 J. Mex. Chem. Soc. 2009, 53(4) Shron Rosete-Lun et l A λ = 420 nm Absorbnce λ = 420 nm x x x x x10-4 X 1 [mmonium], M A λ = 380 nm Absorbnce λ = 380 nm Fig. 2. Continuous vrition curve (Job s method) for the () 1-G nd (b) 2-G complexes observing the vrition of the bsorbnce in host s function of mole rtio of host. X 2 b x x x x x x x10-4 [mmonium slt], M b Fig. 3. Typicl titrtion plots for interctions of receptors 1 [ M] () nd 2 [ M] (b) with mmonium chlorides in CH 2 Cl 2 : MeH (92:8). pen circles: A, down tringles: E nd squres: G. The solid lines re the fitting curves in ccordnce with eqution 1. were fitted to eqution (1) tht considers formtion of both 1:1 nd 1:2 host:guest complexes. A obs 2 ( A0 + A11 K11[ G] + A K K G H T [ ] T )[ ] T = 2 (1) 1 K G K K G + [ ] + [ ] 11 T where A obs is the observed bsorbnce, A 0, A 11 nd A 12 re the bsorbnces of the free host nd its 1:1 nd 1:2 respective complexes, K 11 nd K 12 re the corresponding stepwise formtion constnts, nd [G T ] nd [H] T re the totl concentrtions T of the guest nd host, respectively. The eqution is vlid for conditions with high excess of the guest over the host, which were fulfilled in the titrtion experiments. In Fig. 3 some exmples of the typicl titrtion plots re shown. The verge binding constnts clculted for the different guests re listed in Tble 2. The binding constnts were lso clculted by fitting the 1 H MR chemicl shifts of the host protons in the receptors s function of the guest (A nd G) concentrtion using eqution 2 tht considers 1:1 stoichiometry nd ssumes tht the totl concentrtions of host nd guest re similr to ech other [25].

6 Crown Ether Ditopic Receptors for Ammonium Slts with High Affinity for Amino Acid Ester Slts [ H ] + [ G] + [ H ] + [ G] + K K δ = δ δ obs H 2 T T T T [ H ] [ G ] 4 T T (2) [ H ] T where δ H is the chemicl shift of given proton of the host, Δd is the difference between the chemicl shift of the proton in the complex nd in the free host (complextion-induced shift t sturtion, CIS), [H] T is the totl concentrtion of the host, [G] T is the totl concentrtion of the guest nd K is the binding constnt. CIS vlues for the protons of the receptors used in this work re given in Tble 3. Typicl titrtion plots re Tble 2. Binding constnts for the host-guest complexes in CH 2 Cl 2 : MeH: (92:8) determined by UV/Vis spectroscopy (25 C). shown in Fig. 4 nd verged binding constnts re collected in Tble 4. Tble 2 contins the clculted vlues of K 11 nd K 12 for both host molecules. From these dt it results evident tht, for ll the studied cses, the 1:1 complex is more stble thn the corresponding 1:2 complex in greement with the shrpness of the Job s plot. The K 11 dt obtined for receptor 1 shows tht the binding constnts vlues for the protonted mines increse in the order D < E < G < H < A F. At glnce, the order found suggests tht stbility might be relted with the mount of contct surfce between host nd guest s well s the type of the interctions estblished between them. Ammonium ion (D) binds to the crown ether only with hydrogen bonds wheres, lkyl mmonium ions (E, G) cn estblish vn der Wls Host 8.58 Guest 1 λ = 420 nm 2 λ = 380 nm 8.57 K 11 (M -1 ) K 12 (M -1 ) K 11 (M -1 ) K 12 (M -1 ) A ± ± ± ± 19 B ± ± 5 C ± ± 8 D ± ± ± ± 1 E 542 ± ± ± ± 6 F ± ± ± 34 G ± ± ± ± 14 H ± ± ± 295 o interction detected. δ H (7) (ppm) [mmonium slt], M Tble 3. Complextion-induced 1 H MR chemicl shifts (CIS) of the host protons in host-guest complexes with the mmonium chlorides A nd G in CDCl 3 (in ppm). A Guest Host proton CH 2 (13,14) b - b CH 2 (15,16) b - b - b CH 2 (,22) - b - b - b H (1) - b - b H (2) - b - b H (3) - b H (4) - b - b H (6) b H (11) - b - b - b umbers in prentheses correspond to lbels of the protons s shown in Scheme 1. b ot determined becuse of signl overlpping. G δ H (3) (ppm) [mmonium slt], M b Fig. 4. Typicl titrtion plots for interctions of receptors 1 [ M] () nd 2 [ M] (b) with mmonium chlorides in CDCl 3. Squres: A nd circles: G. The solid lines re the fitting curves in ccordnce with eqution 2.

7 4 J. Mex. Chem. Soc. 2009, 53(4) Shron Rosete-Lun et l. Tble 4. Binding constnts for the host-guest complexes determined by 1 H MR in CDCl 3 t 20 C. Host, K (M -1 ) Guest 18C6 B18C6 B18C6 1 2 A 3114 ± ± ± ± ± 375 G 2551 ± ± ± ± ± 51 interctions tht increse with the length of the chin, nd if the guest molecule hs n romtic ring (H, F) locted t the pproprite distnce it cn lso estblish ϖ-ϖ interctions, ll of them resulting in n enhncement of the overll binding. For tht reson, it is quite surprising tht smll mino cid derivtive s L-lnine methyl ester (A) hs binding comprble to tht of F while for lrger mino cids, like B or C, no interction ws detected. In the cse of receptor 2 regulr trend of binding ffinity ws not found. However, from the clculted binding constnts presented in Tble 2 it is evident tht the complexes with L-mino cid methyl esters hve the lrgest stbility constnts of the studied guests but ll these vlues re lower thn those reported from extrction [10, 12] or from MR [18,19] experiments with similr receptors. A similr trend ws observed by 1 H MR titrtions (Tble 4) in which it is possible to compre the ssocition constnts for heptyl mmonium nd L-lnine methyl ester. For ll the studied cses the K 12 is t lest one order of mgnitude smller thn K 11 thus, the K 12 /K 11 rtio explins the strong negtive llosteric effect observed. This coopertive effect is well known nd hs been mply discussed in the literture [26]. It is now ccepted tht the two crown units ct independently with respect to complextion when the rtio of ssocition constnts K 12 /K 11 is lrger or equl to A negtive coopertive effect occurs when this rtio is smller thn 0.25, positive effect is sid to hppen in the opposite cse [27]. To compre the reltive contributions to the stbilities coming from the different structurl units in the host molecules the complextion of A nd G, s typicl guests of mine nd mino cid slts ws studied with the 18-crown-6 (18C6), 18-benzocrown-6 (B18C6) nd the monoimine (B18C6). The results obtined by 1 H MR titrtions show tht receptor 2 hs the highest selectivity with K A /K G rtio of c. 5 (Tble 4). The presence of benzo zo methine substituent on the B18C6 increses the ffinity of the receptors. In the cse of receptor 2 the presence of bis zo methine increses the selectivity. The interction between the receptors 1 nd 2 with bismmonium guests (s 1,6-hexnedimmonium nd m-xililendimmonium) ws explored by 1 H MR experiments. However, when one equivlent of bismmonium slt is dded to solution of the host the hydrolysis of imine bond ws observed s ws deduced by the rise of ldehyde signl in the MR spectrum. Computtionl studies In order to gin insight into the chrcteristics of the interctions between the guest ions nd receptor 1, the stble gsphse structures of three single receptor molecules, 18C6, B18C6, nd B18C6 s well s their complexes with D, E nd A were studied using DFT methods (B3LYP/6-31G(p)). The optimized stble structures of B18C6 nd its complex with A re shown in Fig. 5. The properties of these complexes re compred with the properties of the optimized stble structures of receptor 1 complexes. A stble structure of receptor 1 is shown in Fig. 6 nd two of the studied complexes re presented in Fig. 7. The optimized geometry of 18C6 corresponds to the S 6 conformer tht hs been described s the most stble conformer predicted with DFT nd MP2 b initio methods [28]. Fig. 5. B3LYP/6-31G* optimized structures of B18C6, () nd its complex with L-lnine methyl ester (b) The C= H 2 C distnces re in Å nd the highlighted toms (yellow) form the dihedrl ngle C benzo --C met -C met mentioned in the text. Fig. 6. B3LYP/6-31G* optimized structure of receptor 1.

8 Crown Ether Ditopic Receptors for Ammonium Slts with High Affinity for Amino Acid Ester Slts 5 Fig. 7. Frgment of the B3LYP/6-31G* optimized structures of the complex formed between receptor 1 nd propil mmonium () nd L-lnine methyl ester (b). H-bonds nd close contct distnces re in Å. The white dot over the romtic ring corresponds to the clculted centroid. For B18C6 ws found tht romtic ring lies in the sme plne s the two the donor oxygen toms directly ttched to it. As some uthors hve pointed out [29], the size of the cvity is not ffected by the presence of the benzo group however, the bsicity of the donor groups nd the shpe of the cvities re different. The structure of the B18C6 frgment is preserved in B18C6 nd in receptor 1. In these free lignds, the phenyl group of the benzo crown ether nd the romtic bis-imine moiety form n extended conjugted system nd the ngle formed between the plnes contining them is ~29. This vlue is in good greement with wht is found in the crystl structure of, -(m-phenylenedimethylidyne)diniline [30]. It hs been discussed tht the size of the H 4 + ion (1.4 Å) nd the H 3 + group mke of this species idel guests on the 18C6 cvity (~1.42 Å) where they re stbilized through three hydrogen bonds disposed in three-fold symmetry pttern [29, 31]. In the cse of mmoni (D), the verge -H distnce ws found to be 1.83 Å. For the other guest molecules, the presence of the R- substituent on the H 3 + group ffects the fit leding to less symmetricl coordintion pttern with longer H-bonds, 1.87±0.01 Å for E nd 1.92±0.05 Å for A. In the cse of the B18C6 complexes it ws found tht the three H-bonds hve different lengths for ll the studied cses. The lck of symmetry in this ltter cse is not only relted with the differences on the ether conformtion but lso with conformtionl chnge occuring s consequence of the complex formtion; the dihedrl ngle C benzo --C met -C met chnges from ~80 to ~160. It is expected tht electronic chnges go long with this structurl modifiction. Besides the H-bonds, the intermoleculr distnces between the host nd the guest molecule strongly suggest the presence of dditionl interctions: the methylene groups of E re close to one of the ether oxygens ( Å) nd in the cse of A the oxygen toms of the crbonyl ester group re close to the methylene hydrogens of the crown ether (2.4 nd 2.8 Å). Upon complex formtion, receptor 1 undergoes further conformtionl chnges, the twist between the romtic rings connecting the crown ether rings decreses by c.. 10 on the side where the complex is formed but this is counterblnced by n equivlent increse in the twist of the free side. However, on the double complex (1:2) the decrese in the twist is symmetric. The interction energies for ll the studied complexes, were obtined s: E int = E complex (E host + ne guest ), where E complex, E host, nd E guest, re the totl energies of the corresponding optimized structures nd n is the number of guest molecules per complex. The interction energies of mmonium ion complexes re lrger thn tht of E nd A with ll the lignds considered in this study. For tht reson the interction energy clculted for the 18C6-D complex will be considered s n internl reference for the following discussion. The reltive interction int X + H int energies, E = E E 4, re presented in Tble 5. The differences on the interction energies of 18C6 nd B18C6 complexes re not very lrge nd cn be rtionlized in terms of two situtions: first, the incresed bsicity of the two oxygen toms ttched to the benzo group with respect to the others does not result in stronger H-bonds. And second, the energetic cost ssocited with the conformtionl chnge occurring on the ether ring, estimted from scn of the pertining torsion brrier, for both guest molecules is less thn 1.5 kcl/mol. Both situtions led to very smll difference between 18C6 nd B18C6 for E but more fvourble bonding sitution for A with B18C6. The interction energies of the B18C6 complexes reflect not only the steric effect of the ether ring substituent but lso the electronic effects of the deloclized system in it. For this reson, it is better lignd thn B18C6 for E nd A, but no for the mmonium ion. Furthermore, it is lso possible tht the dditionl non-covlent interctions, C-H type, might be smll contribution to the H G 18C6 D Tble 5. Reltive interction energies, E = Eint Eint, for the 1:1 complexes with the studied lignds. All vlues re in kcl/mol. Lignds Guest 18C6 B18C6 B18C6 1 D A E

9 6 J. Mex. Chem. Soc. 2009, 53(4) Shron Rosete-Lun et l. stbility. In this cse, their presence, suggested by the distnce vlues between the relevnt groups, does not hve noticeble effect in the totl interction energies. Further investigtion of the role these interctions hve in the stbility of these complexes requires the use of b initio methods [32]. From the interction energies of the complexes formed with receptor 1 it is evident tht the electronic effects relted with the presence of the lrge conjugted system ttched to the ether ring mke of this lignd much better host for both E nd A thn ny other of the considered lignds. The interction energies for the double complex of receptor 1 (1:2) were clculted in the sme wy tht for the 1:1 complexes. The comprison of these interction energies with the corresponding vlues for the 1:1 complexes of receptor 1 is mesure of the coopertive effects in the system. For the three studied ctions the interction energy of the double complex is smller thn the double of the corresponding interction energy of the 1:1 complex. However, there re significnt differences in the non-dditivity for ech system. For D, the interction energy is 16.2% less ttrctive thn the expected vlue for fully dditive system; this difference ws considerbly more importnt for E, 31.3% nd rther smll for A, only 6.6%. Discussion In this study, the theoreticl results re useful to better understnd the experimentlly observed behviour of these receptors. The single most importnt chnge tking plce upon complextion is the conformtionl chnge occurring in the lignds, in the crown ether ring nd the romtic link between the two host groups. This structurl modifiction is lrgely responsible of the chnges observed in the UV/Vis spectr nd the shifting of some protons in the crown ether to low field s observed on the CIS. The negtive llosteric effect found is lso consequence of this conformtionl chnge. The binding of ny guest ion to one of the benzocrown units ffects the electronic density of the oxygen toms on the unoccupied crown ether lowering its bsicity. In conjugted bis-crown receptors, the first ction, which is electron withdrwing, cn modify the electron environment of the second crown. Electrosttic repulsion cn lso hppen when the complexed crowns lie in close proximity. In previous studies, it ws found tht in benzo nd dibenzo crown ethers [33], chnges in the chrge on the romtic oxygen in the mcrocycle, brought bout by chnge in the nture of the substituent, produced profound effects on the binding constnts. Detiled nlysis of the complextion dt indictes tht in generl two fctors influence the binding bility of the ether ring; the polrizbility of the electrons on the ring oxygen nd the constrints on the conformtionl chnges tht re required to occur for the ring to chieve minimum conformtion round the solvted ction. The lrge selectivity observed for the L-lnine ction cn be understood s result of severl combined fctors; this guest cn engge in wek interctions dditionl to the H-bonds tht might confer more dynmicl stbility to the complex; the romtic substitution on the crown ethers fvour the complex formtion with it nd, to smller extent but lso importnt, the lrge difference on coopertive effects found from the interction energies of the 1:1 nd 1:2 complexes reflects tht the modifictions induced by its presence in one of the crown ether rings of receptor 1 does not severely impir the other to ct s resonbly good host for nother ction. Conclusions The binding constnts obtined in this work re comprble with those reported from MR experiments [18, 19] or clorimetric titrtions [20] for mmonium ctions nd one order of mgnitude lrger thn those reported for benzo-18-crown-6 for protonted mino cid methyl esters obtined from clorimetric titrtion in methnol []. It ws found tht the receptors reported in this work hve high ffinity for ctionic mino cid slts. This study confirms the interction energies of the complex formed re resonbly good pproch to understnd the stbility constnts but it is only the simultneous considertion of the energetic nd structurl chnges occurring during complex formtion tht the selectivity of these receptors cn be explined. Experimentl Mterils nd spectroscopic mesurements All regents nd solvents were purchsed from Aldrich nd Fluk nd were used s received. Amine slts were prepred following literture procedures [34] nd dried under vcuum. 1 H MR spectr were recorded in CDCl 3 t 400 MHz t probe temperture of 20 C on Vrin UITY IVA with TMS s internl reference. The electronic spectr were recorded t 25 C on Hewlett Pckrd 8453A diode rry spectrophotometer. FAB mss spectr were obtined on JEL JMS- SMX102A instrument. Elementl nlysis ws obtined on n ELEMETAR Vrio ELIII (C, H,, S) instrument. UV/Vis titrtions The spectroscopic titrtions were performed t 25 C, in CH 2 Cl 2 :MeH: (92:8). Aliquots of concentrted solution of guest ( M for mmonium slts D-H nd M for A-C) were dded to solution of 1 ( M) or 2 ( M). 1 H MR titrtions Guests nd hosts molecules were dissolved in CDCl 3. Titrtions were performed t 18 ºC by dding seven liquots of

10 Crown Ether Ditopic Receptors for Ammonium Slts with High Affinity for Amino Acid Ester Slts 7 G ( M) or A ( M) guest stock solutions to M solutions of the hosts. The experimentl dt were fitted using non-liner lestsqures regression with Microcl rigin 7 progrm. Computtionl method DFT methodologies were chosen to optimize the structure of the bis crown ether receptors, the mine slts nd the complexes becuse they provide good qulity description of the conformtionl fetures of this type of systems [28, 35] nd qulittive correct picture of the reltive stbilities of hydrogen bonded complexes [32, 33]. In this cse, the B3LYP functionl ws used with double-ζ split vlence bsis set with one set of polriztion functions on ll toms, 6-31G(p). The reltive stbilities of the studied complexes t this level of clcultion llow for qulittive picture of the interctions involved in the binding process. The study of some structurl modifictions of the receptor molecule will help to understnd the structurl nd electronic effects resulting of connecting two crown ether units. All clcultions were done using the Gussin 03 suite of progrms [36]. 23), 3.70 (m, 2H, H-22), 3.74 (m, 2H, H-18), 3.74 (m, 2H, H-), 3.78 (m, 2H, H-19), 3.78 (m, 2H, H-20), 3.95 (dd, 2H, H-16), 3.95 (dd, 2H, H-17), 4.20 (dd, 2H, H-14), 4.20 (dd, 2H, H-15), 6.83 (d, 1H, H-8), 6.85 (d, 1H, H- 10) 6.90 (dd, 1H, H-11), 7.56 (dd,1h, H-3), 8.00 (d, 1H, H-2, H-4), 8.36 (s, 1H, H-1), 8.55 (s, 1H, H-7). 13 C MR (CDCl 3, 100 MHz, reference TMS): δ 69.18(C-15), 69.58(C- 14), 69.80(C-16), 69.89(C-17), 70.97(C-23), 70.97(C- 22), 70.97(C-), 70.97(C-18), 71.10(C-20), 71.10(C-19), (C-10), (C-8), (C-11), (C-1), (C-3), (C-2, C-4), (C-5, C-6), (C- 9), (C-13), (C-12), (C-7). MS(FAB + ) m/z = 754 [M+H] +.mp C. MeH/CH 2 Cl 2 (8:92) ε 352 nm = ± 529 cm -1 M -1. Monoimine (B18C6) ws prepred from benzldehyde ccording to procedure described to receptors 1 nd 2. Acknowledgements We thnk Professor A. K. Ytsimirsky for the dvice nd fruitful discussion of the results. SRL thnks CACyT for the Ph. D. Fellowship. This work ws supported by CACyT, projects Q nd Synthesis of receptors The synthetic route [22] for the receptors is shown in Scheme 1. Molecules 1 nd 2 were synthesized by the procedure below described using molecule 1 s n exmple g (0.292 mmol) of terephthldehyde nd g (0.592 mmol) of 4-minobezo-18-crown-6 were dissolved in ethnol (10 ml) in 25 ml round bottom flsk nd stirred for 6 hours under reflux. When the rection ws completed yellowish fine solid ws formed. The product ws recovered by filtrtion, wshed with cetone (2 5 ml) nd dried under vcuum. Yield 80%. Anl. Clc. for C 40 H : C, 63.81; H, 6.95;, 3.72% found C, 63.20; H, 7.01;, 3.89%. 1 H MR (CDCl 3, 400 MHz, reference TMS, see Scheme 1 for lbels): δ 3.65 (m, 2H, H-23), 3.65 (m, 2H, H-22), 3.68 (m, 2H, H-18), 3.68 (m, 2H, H-), 3.74 (m, 2H, H-19), 3.74 (m, 2H, H-20), 3.90 (dd, 2H, H-16), 3.90 (dd, 2H, H-17), 4.15 (dd, 2H, H-14), 4.15 (dd, 2H, H-15), 6.80 (d, 1H, H-8), 6.85 (d, 1H, H-10), 6.87 (dd, 1H, H-11), 7.92 (s,1h, H-1), 8.47 (s, 1H, H-7). 13 C MR (CDCl 3, 100 MHz, reference TMS): δ 69.25(C-15), 69.63(C-14), 69.82(C-16), 69.91(C- 17), 71.00(C-23), 71.00(C-22), 71.00(C-), 71.00(C-19), 71.13(C-20), 71.13(C-18), (C-10), (C-8), (C-11), (C-1), (C-5, C-6), (C-9), (C-12), (C-13), (C-7). MS (FAB+) m/z = 754 [M+H] +. mp C. MeH/CH 2 Cl 2 (8:92) ε 387nm = ± 472 cm -1 M Yield 60%. Anl. Clc. for C 40 H : C, 63.81; H, 6.95;, 3.72% found C, 62.99; H, 6.91;, 3.79%. 1 H MR (CDCl 3, 400 MHz, reference TMS): δ 3.70 (m, 2H, H PPM (F2) PPM (F1) PPM (F2) PPM (F1) Fig. S1. HSQC nd HMBC spectr for 1 nd 2 in CDCl 3. Effect of the methnol concentrtion over 380 nm bsorbnce bnd of receptor 1 in CH 2 Cl 2. b

11 8 J. Mex. Chem. Soc. 2009, 53(4) Shron Rosete-Lun et l. PPM (F2) PPM (F1) Fig. S3. Effect of the methnol concentrtion over 380 nm bsorbnce bnd of receptor 1 ( M) in CH 2 Cl 2. References PPM (F2) PPM (F1) b Guy, P. A.; Gremud, E.; Richoz, J.; Turesky, R. J. J. Chromtogr., A 2000, 883, Fig. S2. ) HSQC nd b) HMBC for 2 in CDCl () hgki, H.; Tkym, S.; Sugimur, T. Mutt. Res. 1991, 259, (b) Wkbyshi, K.; go, M.; Esumi, H.; Sugimur, T. Cncer Res.1992, 52, 2092S-2098S. 3. () Sutherlnd, I.. Pure Appl. Chem. 1989, 61, (b) Voyer,.; Lmothe, J. Tetrhedron 1995, 51, (c) Gokel, G. W.; Abel, E. Comprehensive Suprmoleculr Chemistry; Vol. 1, Elsevier: xford, Englnd, 1996;.(d) Gokel, G. W.; Leevy, W. M.; Weber, M. E. Chem. Rev. 2004, 104, () Hyshi, T.; onogushi, M.; Ay, T.; goshi, H. Tetrhedron Lett.1997, 38, (b) Em, T.; Misw, S.; emugki, S.; Ski, T.; Utk, M. Chem. Lett. 1997, ()Tsubcki, K.; Kusumoto, T.; Hyshi,.; uruzzmn, M.; Fuji, K. rg. Lett. 2002, 4, (b) de Silv, A. P.; Sndnyke, K. R. A. S. Angew. Chem. Int. Ed. Engl. 1990, 29, () Feuster, E. K.; Glss, T. E. J. Am. Chem. Soc. 2003, 125, (b) Mohr, G. J. Chem. Eur. J. 2004, 10, () Zimmermn, S. C.; Wendlnd, M. S.; Rkow,. A.; Zhrov, I.; Suslick, K. S. ture 2002, 418, (b) Mertz, E.; Zimmermn, S. C. J. Am. Chem. Soc. 2003, 125, Coler, M.; Costero, A. M.; Gviñ, P.; Gil, S. Tetrhedron: Asymmetry 2005, 16, Jung, Y. E., Song, B. M., Chng, S.-K. J. Chem. Soc. Perkin Trns , Voyer,.; Deschênes, D.; Bernier, J.; Roby, J. J. Chem. Soc., Chem. Commun. 1992, Hernández, J. V.; liv, A. I.; Simón, L.; Muñiz, F. M.; Grnde, M.; Morán, J. R. Tetrhedron Lett. 2004, 45, Tsubki, K., Tnk, H., Furut, T., Tnk, K., Kinoshit, T., Fuji, K. Tetrhedron 2002, 58, Glán, A.; Andreu, D.; Echvrren, A. M.; Prdos, P.; De Mendoz, J. J. Am. Chem. Soc. 1992, 114, Reetz, M. T.; Huff, J.; Rudolph, J.; Töllner, K.; Deege, A.; Goddrd, R. J. Am. Chem. Soc. 1994, 116, Brboiu, M. D.; Hovnnin,. D.; Luc, C.; Cot, L. Tetrhedron 1999, 55, Demirel,.; Bulut, Y.; Hosgören, H. Tetrhedron: Asymmetry 2004, 15, Brecci, P.; Vn Gool, M.; Pérez-Fernández, R.; Mrtín- Sntmrí, S.; Ggo, F.; Pdros, P.; De Mendoz, J. J. Am. Chem. Soc. 2003, 125, Hnsson, A. P.; orrby, P.-.; Wärnmrk, K. Tetrhedron Lett. 1988, 39, Krytov,. P., Kolchinski, A. G., Rybk-Akimov, E. V. Tetrhedron 2003, Iztt, R. M.; Lmb, J. D.; Iztt,. E.; Rossiter, B. E. Jr., Christensen, J. J., Hymore. B. L. J. Am. Chem. Soc. 1979, 101, Buschmnn, H.-J.; Schollmeyer, E.; Mutihc, L. J. Inclusion Phenom. Mcrocyclic Chem. 2001, 40, () Defen, W.; Xioqing, S.; Dengjin, W.; Hongwen, H. Youji Huxue 1987, 3, (b) Xiochun, Ch.; Defen, W.; Denjing, W.; Hongwen, H. Chem. Res. Chin. Univ. 1991, 7, (c) Dengjin, W.; Defen, W.; Xioqing, S.; Hongwen, H. Godeng Xuexio Huxue Xuebo 1990, 11(7), (d) Defen, W.; Denjing, W.; Hongwen, H. Godeng Xuexio Huxue Xuebo 1990, 11(3), Jffe, H.H.; rchin, M. Theory nd Applictions of Ultrviolet Spectroscopy. John Wiley & Sons, USA, 1962, p Connors, K. A. Binding Constnts. The Mesurement of Moleculr Complex Stbility. John Wiley & Sons, USA, 1987, p Schneider, H.-J.; Ytsimirsky, A. K.: Principles nd Methods in Suprmoleculr Chemistry. John Wiley & Sons, Englnd, 2000, p Mrquis, D.; Desvergne, J.-P.; Bous-Lurent, H. J. rg. Chem. 1995, 60,

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