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Diaza-18-crown-6-Based Saccharide Receptor Bearing Two Boronic Acids. Possible Communication between Bound Saccharides and Metal Cations Kazuaki Nakashima, Ritsuko Iguchi, and Seiji Shinkai* Chemotransfiguration Project, Japan Science and Technology Corporation, 2432 Aikawa, Kurume, Fukuoka 839-0861, Japan
A novel diaza-18-crown-6-based saccharide receptor (4) bearing two boronic acid groups has been developed. In 4 boronic acid groups in the side arms interact as Lewis acids with amines and boron atoms are changed to sp3 hybridization through the B‚‚‚N interaction. Because of this hybridization, the boronic acid groups can bind saccharides even in a neutral pH region. Judging from the distance between two boronic acid groups in 4 assuming a syn conformation, one can expect that 4 forms 1:1 cyclic complexes with monosaccharides. These are confirmed by circular dichroism spectroscopy of 4 in the presence of glucose and allose. UV photometric titration studies showed that the boronic acid groups and metal cation bound to the crown cavity competitively interact as Lewis acids with the crown amines. As a result, saccharides and metal cations “communicate” with each other in an allosteric manner. It was shown that added metal cations can affect the saccharide binding ability of 4 through the competitive interaction with the crown amines and the conformational change of the crown ether ring. Introduction Multipoint recognition based on hydrogen-bonding interactions is the primary tool for recognition of guest molecules.1,2 However, because saccharides show practical solubility only in water, hydrogen-bonding interactions can be applied for saccharide recognition only in limited cases.3-6 To solve this problem, boronic acids have been employed to “touch” and “recognize” saccharides in water.7-20 However, boronic acids have a merit and a demerit: they can rapidly and reversibly form complexes with diol groups in saccharides in an aqueous system, but such complexation occurs significantly only in a basic pH region because cyclic boronate esters are stabilized only in a sp3-hybridized boron atom and OHis indispensable for this. It was shown later that boronic acids bearing an appropriate intramolecular amine can bind saccharides even in a neutral pH region because the boron atom has been changed to sp3 hybridization through the B‚‚‚N interaction.17,18 By using this concept, several novel reading-out-type saccharide interfaces have successfully been developed.17,19,20 Meanwhile, diboronic acid derivatives bearing crown ether(s) 1-3 were synthesized to mimic allosteric interaction occurring in nature.13,21,22 In these compounds metal and saccharide binding sites “communicate” with each other through a conformational change caused by metal binding to the crown cavity (Figure 1). In the present study a diaza-18-crown-6-based saccharide receptor 4 is designed as a new class of allosteric metal-saccharide receptor. One can expect for 4 that boronic acid groups in the side arms and a metal cation bound to the crown cavity competitively interact as Lewis acids with two nitrogens in diaza-18-crown-6. * To whom correspondence should be addressed. Tel.: +81-942-39-9011. Fax: +81-942-39-9012. E-mail: seijitcm@ mbox.nc.kyushu-u.ac.jp.
Therefore, metal cation bound to the crown cavity can “communicate” with the boronic acid groups not only through the conformational change but also through this competitive interaction. This system should be useful as a metal-controllable saccharide receptor or a saccharide-controllable metal receptor and act as a novel allosteric saccharide recognition system.23
10.1021/ie000225i CCC: $19.00 © 2000 American Chemical Society Published on Web 09/08/2000
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Figure 1. Schematic representation of a negative allosteric system between metal ion and saccharide.
N,N′-Bis(2-borophenylmethyl)-4,13-diaza-18crown-6 (4). To a suspension of 2,4,6-tris[2-(bromomethyl)phenyl]boroxin (0.85 g, 1.32 mmol), potassium carbonate (0.55 g, 3.98 mmol), and potassium iodide (50 mg, catalyst) in refluxing acetonitrile (20 mL) was added dropwise over 30 min 4,13-diaza-18-crown-6 (0.52 g, 1.98 mmol) in acetonitrile (18 mL). The mixture was stirred and refluxed for 8 h under a nitrogen stream. After the mixture cooled to room temperature, inorganic salt was removed by filtration. The solvent was removed, and the residue was dissolved in 0.01 N NaOH. The pH of the mixture was adjusted to 7 with a 10% aqueous HCl solution. The colorless solid was collected by filtration and washed with water. Finally, it was recrystallized from methanol-water to afford 4 as a colorless powder (0.85 g, 71%): mp 291.6-293.2 °C; IR (KBr) 3304 (O-H), 1376 (B-O); 1H NMR (CD3OD) δ 3.23 (8H, t, J ) 5 Hz, NCH2CH2O), 3.63 (8H, s, OCH2CH2O), 3.80 (8H, t, J ) 5 Hz, NCH2CH2O), 4.22(4H, s, CH2Ar), 7.15-7.65 (8H, m, aromatic); 13C NMR (CD3OD) δ 53.21, 62.17, 67.90, 71.73, 128.37, 128.53, 130.47, 135.29, 139.55. Results and Discussion
Experimental Section General Procedures. Melting points were measured on a Yanaco MP-500D micro melting point apparatus and are uncorrected. 1H and 13C NMR spectra were recorded on a Bruker ARX-300 spectrometer. IR spectra were recorded on a Shimadzu FT-IR 8100 spectrometer using KBr disks. UV spectra were measured on a Jasco V-570 spectrometer. Circular dichroism (CD) spectra were measured on a Jasco J-720WI spectrometer. Curve fitting based on a nonliniar least-squares method was carried out in a data analysis software package KaleidaGraph (Synergy Software, Reading, PA). Materials. Solvents (methanol and water) used for UV and CD measurements were of spectroscopic grade and used as received. All other chemicals were of reagent grade and were used without further purification. Acetonitrile was dried over molecular sieve 4A. Potassium carbonate was finely powdered and dried in a vacuum oven overnight. (2-Methylphenyl)boronic acid was purchased from Aldrich Co. 4,13-Diaza-18crown-6, choline hydroxide (50% solution in water), and choline chloride were purchased from Tokyo Kasei Kogyo Co. Calcium thiocyanate, strontium thiocyanate, barium thiocyanate, D-fructose, and D-glucose were purchased from Wako Pure Chemicals Industries (Osaka). L-Glucose and D-allose were purchased from Sigma Chemical Co. 2,4,6-Tris[2-(bromomethyl)phenyl]boroxin. This compound was prepared from (2-methylphenyl)boronic acid according to the literature procedure.24
Complexation with Monosaccharides. Boronic acid appended diaza-18-crown-6 4 was prepared by the reaction of 4,13-diaza-18-crown-6 and 2,4,6-tris[2-(bromomethyl)phenyl]boroxin in the presence of potassium carbonate as the base. Previous studies demonstrate that compounds bearing two boronic acid groups arranged in an appropriate spatial orientation show high selectivity for their complementary saccharides.11,13,19 In these compounds, each boronic acid group binds two OH groups in saccharides to form chiral macrocyclic 1:1 complexes. Because two benzene rings are immobilized in a chiral twist in these compounds, the saccharide binding can be easily “read out” by CD spectroscopy. In glucose, boronic acids are considered to bind a 1-OH/2-OH diol group and a 4-OH/6-OH (in a pyranose form) or 5-OH/6-OH (in a furanose form) diol group. In compound 4, upon adoption of a syn conformation, the distance between two boronic acid groups is comparable to that of these two diol groups in monosaccharides such as glucose. These facts prompted us to measure CD spectra of 4 in the presence of monosaccharides (D- and L-glucose, D-allose, and D-fructose). As shown in Figure 2, exciton-coupled CD spectra were observed in the presence of D- and L-glucose and D-allose. Clearly, these CD bands are indicative of the immobilization of two benzene rings by ring closure with a monosaccharide, and they reflect the chirality of added saccharides. The plots for [θ]max ([θ]223.2 for D-glucose and [θ]225.4 for D-allose) vs [monosaccharide] are shown in Figure 3. These titration curves are fitted with a model assuming the formation of cyclic 1:1 complexes: thus, the association constants are estimated according to the Benesi-Hildebrand equation (Kass ) 2.7 × 104 mol-1 dm3 for D-glucose and 1.9 × 102 mol-1 dm3 for D-allose). In contrast, 4 gives no CD signal in the presence of D-fructose, which tends to form noncyclic D-fructose/ boronic acid 2:1 complexes rather than cyclic 1:1 complexes. This result also supports the view that 4 forms CD-active macrocyclic 1:1 complexes with glucose and allose.
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Figure 2. CD spectra of 4-D-glucose, 4-L-glucose, and 4-D-allose complexes (1.0 × 10-3 mol dm-3) in methanol-water (9:1, v/v) containing 0.05 mol dm-3 choline (pH 8.5, 25 °C).
Figure 4. pH-dependent spectral change of 4 (3.4 × 10-4 mol dm-3 in methanol-water (9:1, v/v) containing 0.05 mol dm-3 choline): (A) pH 3.01-7.91; (B) pH 7.91-12.69.
Figure 5. Plot of A272 vs pH.
Figure 3. Plot of [θ]max vs [monosaccharide] [at 223.2 nm for D-glucose (A) and 225.4 nm for D-allose (B)].
Estimation of pKa Values for Amines and Boronic Acid Groups. pKa values for amines and boronic acid groups were estimated by using both photometric and potentiometric titrations. To avoid the possible metal-crown interaction arising from buffer species, we used choline [(2-hydroxyethyl)trimethylammonium] hydroxide as the base. The pH-dependent change in the absorption spectra is shown in Figure 4.
Although the absorption band at around 270 nm decreases monotonically, the change apparently includes three pKa points. A plot of pH vs A272 is shown in Figure 5. A total of 2 equiv of choline OH- was consumed at pH 3-8 and 8-11, respectively. Judging from our previous studies,17 they are assigned to first and second deprotonation of amines and dissociation of two boronic acid groups. The total equilibria of amines and boronic acid groups in 4 are illustrated in Scheme 1. On the basis of this equilibrium, a curve fitting of the plot shown in Figure 4 was carried out and three pKa values were obtained: pKaN1 ) 4.87, pKaN2 ) 6.77, and pKaB ) 12.2. The photometric titrations of 4 were also carried out in the presence of metal cation (Ca2+) and/or saccharide (D-glucose or D-fructose), and the plot was analyzed in a manner similar to estimation of the pKa values. The results are summarized in Table 1. In the presence of saccharides (3.4 × 10-2 mol dm-3), pKaN1 and pKaN2 shifted to lower pH, whereas pKaB is affected only to a
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Scheme 1. Equilibria of Amines and Boronic Acid Groups in 4
Table 2. Saccharide Association Constants in the Absence and Presence of Metal Cationsa metal cation
Kass(D-glucose) (104 mol-1 dm3)
Kass(D-allose) (102 mol-1 dm3)
none Ca2+ Sr2+ Ba2+
2.65 (1.00)b 1.65 (0.62)b 1.43 (0.54)b 2.25 (0.85)b
1.95 (1.00)b 1.48 (0.76)b 1.07 (0.55)b 1.13 (0.58)b
a [4] ) 1.0 × 10-3 mol dm-3, [metal cation] ) 2.0 × 10-2 mol dm-3 in methanol-water (9:1) containing 0.05 mol dm-3 choline, pH ) 8.5, 25 °C. b Values in parentheses are relative values.
Table 1. pKa Values of Amines and Boronic Acid Groups in 4 in the Absence and Presence of Guestsa guest
pKaN1
pKaN2
pKaB
none Ca2+ b D-fructosec D-glucosec Ca2+ and D-fructoseb,c Ca2+ and D-glucoseb,c
4.87 6.33 4.47 3.97 4.70 3.82
6.77
12.2 10.3 11.6 12.1 10.3 11.3
5.56 6.01 6.23 6.14
a [4] ) 3.4 × 10-4 mol dm-3 in methanol-water (9:1) containing choline (5.0 × 10-2 mol dm-3), 25 °C. b [Ca2+] ) 3.4 × 10-2 mol dm-3. c [saccharide] ) 3.4 × 10-3 mol dm-3.
smaller extent, as shown in Table 1. The finding indicates that saccharide-boronic acid complexation, which makes the acidity of the boron atom stronger,8,17 facilitates dissociation of the nitrogen protons through the B‚‚‚N interaction. Because of this interaction, two boronic acid groups can bind saccharides even in a neutral pH region. When Ca2+ was added, pKa values of two amines became equal because the stepwise deprotonation from the crown nitrogens no longer takes place. Thus, we obtained two pKa values for amine deprotonation and OH- adduct formation with boronic acid groups (pKaN ) 6.33 and pKaB ) 10.3). Ca2+ competes with the boronic acid groups for the nitrogens; i.e., the B‚‚‚N interaction is weakened by the Ca2+‚‚‚N interaction, so that pKaN is higher than pKaN1 in the absence of Ca2+. Similarly, in dissociation of the boronic acid groups (i.e., OHadduct formation with boronic acid groups), the B‚‚‚N interaction is weakened because of the Ca2+‚‚‚N interaction, so that pKaB is lowered compared with pKaB in
the absence of Ca2+. Thus, the foregoing complexation processes shown in Scheme 2 are proposed. When saccharide and Ca2+ coexist, as seen from Table 1, the pKa values of two amines split into two and pKaN1 shifts to a lower pH region than that in the presence of only Ca2+. On the other hand, pKaB is scarcely changed. Because the acidity of boronic acid is strengthened by saccharide complexation,8,17 the shift of pKaN1 and pKaN2 is attributable to the strengthened B‚‚‚N interaction. On the other hand, the minor change in pKaB implies that Ca2+ scarcely affects the boronic acid dissociation step: that is, the saccharide-enhanced acidity has made the boronic acid groups more advantageous in the competitive interaction with the nitrogens. It is not clear yet, however, why the pKa values of two amines become different in the presence of both Ca2+ and saccharide. Negative Allosterism between Boronic Acid Groups and Metal Cation Bound to a Crown Cavity. The result of photometric titration suggested the competitive interaction of boronic acid groups and metal cation bound to a crown cavity in 4. To estimate the effect of this competitive interaction to the saccharide binding quantitatively, the association constants for D-glucose and D-allose in the presence of metal cations (Ca2+, Sr2+, and Ba2+) were determined by CD titration. The results are summarized in Table 2. In all cases the association constants decreased in the presence of metal cations. This implies that a metal cation bound to the crown cavity interacts with nitrogen, which hampers the B‚‚‚N interaction and suppresses the boronic acid-saccharide interaction in a neutral pH region. The order of the association constants is Ca2+ > Ba2+ > Sr2+ for D-allose and Ba2+ > Ca2+ > Sr2+ for D-glucose. Judging from the saccharide structures, the distance between two diol groups in D-allose is shorter than that in D-glucose. This difference suggests that the
Scheme 2. Effect of Saccharide and Metal Cation on the Dissociation of Amines and Boronic Acid Groups
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binding of a “bigger” metal cation changes the crown ring conformation into one that is unfavorable for the binding of a “smaller” saccharide guest. Conclusions In conclusion, the present study showed a novel example for the design of artificial saccharide receptors in which saccharide and metal cation interact with the binding sites competitively. In this system metal and saccharide binding sites interact not only through a conformational change but also through a competitive interaction with nitrogens in diaza-18-crown-6 ring as Lewis acids. Consequently, metal and saccharide binding sites couple in a negative allosteric manner, and metal cations make the saccharide association constants smaller. One can expect that the metalinduced saccharide release or vice versa is achieved using this system. This negative allosteric concept is very important in practical, dynamic molecular recognition systems, for example, to release saccharides only in the presence of some threshold concentration of metal ions, to regenerate metal (or saccharide)-selective electrodes by saccharides (or metal ions), to control the membrane transport rate of metal ions (or saccharides) by saccharides (or metal ions), etc. Literature Cited (1) Rebek, J., Jr. Molecular Recognition with Model Systems. Angew. Chem., Int. Ed. Engl. 1990, 29, 245-255 and references therein. (2) Hamilton, A. D. Synthetic Studies on Molecular Recognition. Bioorg. Chem. Front. 1991, 2, 115-174 and references therein. (3) Kano, K.; Yoshiyasu, K.; Hashimoto, S. Enantioselective Complexation of Bilirubin with Cyclodextrin and Noncyclic Oligosaccharides. J. Chem. Soc., Chem. Commun. 1988, 801-802. (4) Aoyama, Y.; Tanaka, Y.; Toi, H.; Ogoshi, H. Polar HostGuest Interaction. Binding of Nonionic Polar Compounds with a Resorcinol-Aldehyde cyclooligomer as a Lipophilic Polar Host. J. Am. Chem. Soc. 1988, 110, 634-635. (5) Kikuchi, Y.; Kobayashi, K.; Aoyama, Y. Complexation of Chiral Glycols, Steroidal Polyols, and Sugars with a Multibenzenoid, Achiral Host As Studied by Induced Circular Dichroism Spectroscopy: Exciton Chirality Induction in Resorcinol-Aldehyde Cyclotetramer and Its Use as a Supramolecular Probe for the Assignment of Stereochemistry of Chiral Guests. J. Am. Chem. Soc. 1992, 114, 1351-1358. (6) Inouye, M.; Miyake, T.; Furusyo, M.; Nakazumi, H. Molecular Recognition of β-Ribofuranosides by Synthetic Polypyridines Macrocyclic Receptors. J. Am. Chem. Soc. 1995, 117, 1241612425. (7) Wulff, G.; Krieger, S.; Ku¨hneweg, B.; Steigel, A. Occurrence of Strong Circular Dichroism during Measurement of CD spectra Due to Intramolecular Cyclization. J. Am. Chem. Soc. 1994, 116, 409-410. (8) Yoon, J.; Czarnik, A. W. Fluorescent Chemosensors of Carbohydrates. A means of Chemically Communicating the Binding of Polyols in Water Based on Chelation-Enhanced Quenching. J. Am. Chem. Soc. 1992, 114, 5874-5875.
(9) Paugan, M.-F.; Smith, B. D. Active Transport of Uridine Through a Liquid Organic Membrane Mediated by Phenylboronic Acid and Driven by a Fluoride Ion Gradient. Tetrahedron Lett. 1993, 34, 3723-3726. (10) Nagai, Y.; Kobayashi, K.; Toi, H.; Aoyama, Y. Stabilization of Sugar-Boronic Esters of Indolylboronic Acid in Water via Sugar-Indole Interaction: A Notable Selectivity in Oligosaccharides. Bull. Chem. Soc. Jpn. 1993, 66, 2965-2971. (11) Shiomi, Y.; Kondo, K.; Saisho, M.; Harada, T.; Tsukagoshi, K.; Shinkai, S. Specific Complexation of Saccharides with Dimeric Phenylboronic Acid That Can Be Detected by Circular Dichroism. Supramol. Chem. 1993, 2, 11-17. (12) James, T. D.; Murata, K.; Harada, T.; Ueda, K.; Shinkai, S. Chiral Discrimination of Monosaccharides through Gel Formation. Chem. Lett. 1994, 273-276. (13) Deng, G.; James, T. D.; Shinkai, S. The Allosteric Interaction of the Metal Ions with Saccharides in a Crowned Diboronic Acid. J. Am. Chem. Soc. 1994, 116, 4567-4572. (14) Nakashima, K.; Shinkai, S. Sugar-Assisted Chirality Control of Tris(2,2′-bipyridine)-Metal Complexes. Chem. Lett. 1994, 1267-1270. (15) Murakami, H.; Nagasaki, T.; Hamachi, I.; Shinkai, S. Sugar Sensing Utilizing Aggregation Properties of a Boronicacid-appended Porphyrin. Tetrahedron Lett. 1993, 34, 6273-6276. (16) James, T. D.; Sandanayake, K. R. A. S.; Shinkai, S. Novel Photoinduced Electron-transfer Sensor for Saccharides Based on the Interaction of Boronic Acid and Amine. J. Chem. Soc., Chem. Commun. 1994, 477-478. (17) Sandanayake, K. R. A. S.; Shinkai, S. Novel Molecular Sensors for Saccharides Based on the Interaction of Boronic Acid and Amines: Saccharide Sensing in Neutral Water. J. Chem. Soc., Chem. Commun. 1994, 1083-1084. (18) Wulff, G. Selective Binding to Polymers via Covalent Bonds. The Construction of Chiral Cavities as Specific Receptor Sites. Pure. Appl. Chem. 1982, 54, 2093-2102. (19) James, T. D.; Sandanayake, K. R. A. S.; Shinkai, S. A Glucose-Selective Molecular Fluorescent Sensor. Angew. Chem., Int. Ed. Engl. 1994, 33, 2207-2209. (20) James, T. D.; Sandanayake, K. R. A. S.; Shinkai, S. Chiral Discrimination of Monosaccharides Using a Fluorescent Molecular Sensor. Nature 1995, 374, 345-347. (21) Ohseto, F.; Yamamoto, H.; Matsumoto, H.; Shinkai, S. Allosteric Communication between the Metal-binding Lower Rim and the Sugar-binding Upper Rim on a Calix[4]crown Platform. Tetrahedron Lett. 1995, 38, 6911-6914. (22) James, T. D.; Shinkai, S. A Diboronic Acid ‘Glucose Cleft’ and a Biscrown Ether ‘Metal Sandwich’ are Allosterically Coupled. J. Chem. Soc., Chem. Commun. 1995, 1483-1484. (23) Preliminary communication: Nakashima, K.; Shinkai, S. Diaza-18-crown-6-based Saccharide Receptor Bearing Two Boronic Acids. Possible Communication between Bound Saccharides and Metal Cations. Chem. Lett. 1995, 443-444. (24) Snyder, H. R.; Reedy, A. J.; Lennarz, W. J. Synthesis of Boronic acid groups. Aldehydo Boronic acid groups and a Boronic Acid Analogue of Tyrosine. J. Am. Chem. Soc. 1958, 80, 835-838.
Received for review February 14, 2000 Revised manuscript received June 13, 2000 Accepted June 13, 2000 IE000225I
