Fluorescent Chemo-Sensor for Organic Guests Based on Regioselectively Modified 6 A,6 B -, 6 A,6 C -, and 6 A,6 D -Bisdansylglycine-Modified

Transcription

1 ANALYTICAL SCIENCES DECEMBER 999, VOL The Japan Society for Analytical Chemistry 99 Fluorescent Chemo-Sensor for Organic Guests Based on Regioselectively Modified 6 A,6 B -, 6 A,6 C -, and 6 A,6 D -Bisdansylglycine-Modified -Cyclodextrins Mayumi SATO, Miyuki NARITA, Nobuaki OGAWA and Fumio HAMADA Department of Materials-process Engineering and Applied Chemistry for Environments, Faculty of Engineering and Resource Science, Akita University, Tegata, Akita , Japan Flexible hosts, 6 A,6 B -, 6 A,6 C -, and 6 A,6 D -bis-dansylglycine-modified β-cyclodextrins (β-, β-2, and β-3, respectively) have been synthesized as a fluorescent chemo-sensor for organic guests including terpenoids and bile acids. Molecular sensing abilities obtained as fluorescence spectral changes of these hosts on accommodation of guests were almost twice those of γ -analogues. The sequence of binding ability of β-analogues is β-2>β->β-3. The molecular recognition behaviors of dansyl moieties of the title compounds are studied by induced circular dichroism (ICD), fluorescence, and absorption spectra without and with the guest. These spectral variations suggest that the appended moieties move out far from the cavity of β-cyclodextrin when host-guest complexation occurs. Keywords Modified β-cyclodextrin, dansylglycine, fluorescent chemo-sensor, pattern recognition The molecular recognition by fluorescent sensing system of modified cyclodextrins is a current topic in hostguest chemistry. Cyclodextrins, torus-shaped cyclic oligomers of D-glucopyranose named α-, β-, and γ - for hexa-, hepta-, and octamers, respectively, can make host-guest complexation with a variety of organic compounds in their cavity in aqueous solution. 2 So far, binding abilities of mono-dansylmodified cyclodextrins have been studied. 3 6 β-analogues show higher sensitivity for organic guest such as bile acids than γ -analogues. Ueno and co-workers have discussed the effect on the position of modification on the cyclodextrin cavity for fluorescent chemo-sensing. 7,8 In these papers, they described how modification of the primary hydroxy side on the cyclodextrin is quite useful for the molecular sensing. Recently, we described the fluorescent chemo-sensors of four bis-dansylglycine-appended γ -cyclodextrins, which are 6 A,6 B -, 6 A,6 C -, 6 A,6 D -, and 6 A,6 E -modified γ -cyclodextrins. 9 These hosts indicate much higher sensitivity and selectivity for the guests such as bile acids than do mono-dansyl modified γ -analogues; however, it is hard to detect small guests such as terpenes. For further extension of our work, we synthesized regioselectively modified bis(dansylglycyl)-βcyclodextrins, which are 6 A,6 B -, 6 A,6 C -, and 6 A,6 D - bis(dansylglycyl)-β-cyclodextrins, and examined the sensing ability of these compounds. In this paper, we would like to describe the host-guest binding systems based on these hosts. Compound β-2 exhibits high sensitivity and selectivity particularly for ursodeoxycholic To whom correspondence should be addressed. acid and ( )-borneol. Experimental Preparations of 6 A,6 B -, 6 A,6 C -, and 6 A,6 D -diiodo-βcyclodextrins (I, II, and III, respectively) A mixture of 6 A,6 B -ditosyl-β-cyclodextrin 0 (300 mg, 0.2 mm) and potassium iodide (345 mg, 2.08 mm) in 4 ml of DMF was heated at 80 C for 3 h. After cooling, the reaction mixture was poured into 250 ml of acetone. The resulting precipitates were filtered and dried. II and III were prepared by the same procedure as I. The crude product of I was used as the starting material for β-. I: R f 0.55 (butanol:ethanol:water=5:4:3 by volume; TLC; silicagel 60F254). II: R f 0.53 (butanol:ethanol:water= 5:4:3 by volume; TLC; silicagel 60F254). III: R f 0.55 (butanol:ethanol:water=5:4:3 by volume; TLC; silicagel 60F254). Preparations of 6 A,6 B -, 6 A,6 C -, and 6 A,6 D -bis- (dansylglycyl)-β-cyclodextrins (β-, β-2, and β-3, respectively) To a solution of 6 A,6 B -diiodo-β-cyclodextrin (297 mg, 0.22 mm) in 4 ml of DMF was added sodium N-dansylglycine (204 mg, 0.62 mm). The reaction mixture was heated at 80 C for 24 h. After cooling, the reaction mixture was concentrated in vacuo and poured into 300 ml of acetone. The resulting precipitates were filtered and dissolved in 4 ml of DMF. The DMF soluble fraction was applied to reversed-phase column chromatography (Lober column LiChroprep RP8). After step-

2 200 ANALYTICAL SCIENCES DECEMBER 999, VOL. 5 Fig. Preparations of β-, β-2, and β-3. wise elution with 500 ml of 0 vol.%, 400 ml of 30 vol.%, 400 ml of 40 vol.%, 400 ml of 50 vol.%, 400 ml of 55 vol.%, and 500 ml of 60 vol.% aqueous MeOH were applied, β- was obtained in a pure form. Compounds β-2, and β-3 were prepared and purified by the same procedure as for β-. β-: Yield 0.%. R f 0.58 (butanol:ethanol:water= 5:4:3 by volume, TLC; silicagel 60F254) and 0.59 (methanol:water=2: by volume, TLC; RP-8F254S; Merck Ltd.). H-NMR (DMSO-d 6 )=2.82 (2H, s, N CH 3 ), (48H, m, CH 2 and C2 C6H of cyclodextrin), (5H, br, O6H of cyclodextrin), (7H, m, CH of cyclodextrin), (4H, m, O2H and O3H of cyclodextrin), 7.23 (2H, d, J=8. Hz, aromatic-h), 7.57 (4H, t, J=8.0 Hz, aromatic-h), 8.09 (2H, t, J=6.6 Hz, aromatic-h), 8.27 (2H, d, J=8. Hz, aromatic-h), 8.43 (2H, d, J=8. Hz, aromatic-h). Elemental analyses. Calcd. for C 70 H 98 O 4 N 4 S 2 6H 2 O: C, 46.09; H, 6.08; N, 3.07%. Found: C, 45.95; H, 6.4; N, 3.30%. MS (FAB): 75 ([M+H] + ). β-2: Yield 7.0%. R f 0.58 (butanol:ethanol:water=5:4:3 by volume, TLC; silicagel 60F254) and 0.6 (methanol:water=2: by volume, TLC; RP-8F254S; Merck Ltd.). H-NMR (DMSO-d 6 )=2.82 (2H, s, N CH 3 ), (48H, m, CH 2 and C2 C6H of cyclodextrin), (5H, br, O6H of cyclodextrin), (7H, m, CH of cyclodextrin), (4H, m, O2H and O3H of cyclodextrin), 7.24 (2H, d, J=6.9 Hz, aromatic-h), 7.57 (4H, m, J=7.8 Hz, aromatic-h), 8.09 (2H, d, J=6.9 Hz, aromatic-h), 8.26 (2H, d, m, aromatic-h), (2H, m, aromatic-h). Elemental analyses. Calcd. for C 70 H 98 O 4 N 4 S 2 6H 2 O: C, 45.65; H, 6.3; N, 3.04%. Found: C, 45.94; H, 6.6; N, 3.40%. MS (FAB): 75 ([M+H] + ). β-3: Yield 29.7%. R f 0.56 (butanol:ethanol:water=5:4:3 by volume, TLC; silicagel 60F254) and 0.70 (methanol:water=2: by volume, TLC; RP-8F254S; Merck Ltd.). H-NMR (DMSO-d 6 )=2.84 (2H, s, N CH 3 ), (48H, m, CH 2 and C2 C6H of cyclodextrin), (5H, br, O6H of cyclodextrin), (7H, m, CH of cyclodextrin), 7.28 (2H, d, J=7.8 Hz, aromatic-h), 7.60 (4H, t, J=8.0 Hz, aromatic-h), 8. (2H, d, J=7.2 Hz, aromatic-h), 8.28 (2H, d, J=8.7 Hz, aromatic-h), 8.47 (2H, t, J=7.6 Hz, aromatic-h). Elemental analyses. Calcd. for C 70 H 98 O 4 N 4 S 2 3H 2 O: C, 47.50; H, 5.92; N, 3.%. Found: C, 47.55; H, 6.20; N, 3.92%. MS (FAB): 75 ([M+H] + ). Measurements Ultraviolet, fluorescence, and circular dichroism spectra were measured at 25 C with a Perkin Elmer Lambda 40 UV/Vis spectrophotometer, a Perkin Elmer LS 40B fluorescence spectrometer, and a JASCO J-700 spectropolarimeter, respectively. For the fluorescence measurements, the excitation wavelength of the fluorescence spectra was 340 nm and excitation and emission slits were 0 nm wide. An aqueous solution containing ethylene glycol (0 vol.%) was used as solvent for hosts for the spectroscopic measurements because their solubility in pure water is poor. Five microliter aliquots of guest solution (0.5, 0.05 and M) in dimethyl sulfoxide (DMSO) or MeOH were injected into a 0 vol.% ethylene glycol aqueous solution of host (2.5 ml) to make sample solutions with a host concentration of M and guest concentrations of 0.0, 0. and.0 mm, respectively.

3 ANALYTICAL SCIENCES DECEMBER 999, VOL Fig. 2 Induced circular dichoism spectra of β-, β-2, and β-3 (0 4 M) in a 0vol.% ethylene glycol aqueous solution (25 C) in the absence ( ) and presence ( ) of ursodeoxycholic acid (0 4 M). Results and Discussion Preparations of β-, β-2, and β-3 Bis-dansylglycine-modified γ -cyclodextrins were prepared directly from ditosylated γ -cyclodextrins. 9 However, β-analogues were not obtained in the same way. These compounds were synthesized from diiodoβ-cyclodextrins (I, II, and III) and sodium N-dansylglycine in DMF at 80 C as shown in Fig.. Induced circular dichroism (ICD), UV-Vis, and fluorescence spectra Figure 2 shows ICD spectra of three hosts, β-, β-2, and β-3, alone or in the presence of ursodeoxycholic acid in a 0vol.% ethylene glycol aqueous solution. The ICD spectra of the host compounds, alone, show a positive band at around 265 nm and negative bands at around 230 and 355 nm, whose intensities are decreased upon addition of a guest. The ICD patterns of the hosts are basically similar. However, the ICD intensities of β-2 at the positive and the negative bands with the guest are decreased much more than those of β- and β-3. The intensities of β- at the negative bands are decreased more than those of β-3 on accommodation of the guest, whereas the intensity of β- at the positive band in the presence of the guest is decreased less than that of β-3. These results suggest that the binding ability of β-2 for the guest is larger in comparison with those of β- and β-3, and dansyl moieties move from the interior of the hydrophobic cyclodextrin cavity toward the outside bulk water environment while simultaneously a guest is included in the cyclodextrin cavity. Figure 3 shows fluorescence spectra of β-2 in a 0vol.% ethylene glycol aqueous solution in the presence and absence of ursodeoxycholic acid. The fluorescence spectra of these hosts are composed of almost pure monomer emission with a peak around 526 nm, and the fluorescence intensity decreases with increasing ursodeoxycholic acid concentration. It is reported that the guest-induced fluorescence decrease means that the appended moiety is moving out of the cyclodextrin cavity 4, whereas an enhancement means the appended moiety is moving more deeply into the cavity. 9 Figure 4 shows absorption spectra of β-2 in a 0vol.% ethylene glycol aqueous solution in the presence and absence of ursodeoxycholic acid. The absorption spectra of β-, β-2, and β-3 in the presence of this guest are changed with isosbestic point at 280 and 338, 79 and 344, and 277 and 345 nm, respectively. The results obtained as ICD, fluorescence, and absorption spectral changes of these hosts suggest that

4 202 ANALYTICAL SCIENCES DECEMBER 999, VOL. 5 Fig. 3 Fluorescence spectra of β-2 in a 0vol.% ethylene glycol aqueous solution (0 6 M, 25 C) at various concentrations of ursodeoxycholic acid (: 0, 2: , 3: , 4: , 5: , 6: M). Fig. 4 Absorption spectra of β-2 in a 0vol.% ethylene glycol aqueous solution (0 4 M, 25 C) at various concentrations of ursodeoxycholic acid (: 0, 2: , 3: , 4: , 5: M). Scheme Host-guest complexation mechanism for bis-dansylglycine-modified β-cyclodextrins. the dansylglycine moieties move out from the cyclodextrin cavity upon guest binding and act, as a hydrophobic cap, as illustrated in Scheme. As reported previously, the sensing ability of host compounds was displayed using I/I 0 value as a sensitivity parameter. 9,0,2 Figure 5 shows the parameter values of β-, β-2, and β-3 with steroids at 0. mm except for terpenoids (0. mm) and lithocholic acid (7), which was examined at 0.0 mm because 0. mm of lithocholic acid is not soluble in a 0 vol.% ethylene glycol aqueous solution. It is recognized that the data obtained are reproducible. It is evident that ursodeoxycholic acid (9) is detected with remarkably high sensitivity, exhibiting values of 0.527, 0.469, and for β-2, β-, and β-3, respectively. These values of the hosts for ursodeoxycholic acid are almost twice as large as those of γ -analogues reported previously 9, and chenodeoxycholic acid (8), which is the diastereoisomer of ursodeoxycholic acid, was detected with high sensitivity. Lithocholic acid (7), which bears one hydroxy group, was detected with lower sensitivity. Deoxycholic acid (6), which is different from the other steroids only in the position of one hydroxy group, and cholic acid (0), which has one more hydroxy group than 8 and 9, were hardly detected at all. It is probable that there is some strong interaction such as hydrogen bonding between the bile acids such as ursodeoxycholic acid (9) and chenodeoxycholic acid (8) and β- analogues. The sensing factors of bile acids by the hosts decrease in the sequence 9>8>7>0>6. The sensing ability of the three hosts is in the order β-2>β->β- 3. It indicates that regioselectively modified cyclodextrin cavity affects the sensing ability, as shown in the case of γ -analogues, in which β-2 show highest sensing ability. All hosts show only little sensitivity for ketosteroids which have two or three hydroxy groups. Progesterone (), which bears no hydroxy group and is more hydrophobic than the other ketosteroids, was detected with values of 0.050, 0.040, and for β-2, β-, and β-3, respectively, which is higher than those of other ketosteroids. ( )-Borneol (), (+)-fenchone (2), and ( )-fenchone (3), which are bicyclic derivatives, were detected with high sensitivity. In particular, ( )-borneol () shows sensing values of 0.559, 0.405, and for β-2, β-, and β-3, respectively, indicating that the sensing parameters are almost six times as large as those of γ -analogues. Cyclohexanol (4), cyclooctanol (5), and ( )-menthol (6), which are monocyclic derivatives, benzhydrol (7), which bears two aromatic rings, and nerol (8), which is a noncyclic compound, were detected with high sensitivity. These values are positive, whereas γ -analogues showed negative sensing parameter for these guests. The guest-induced fluorescence variation at 526 nm was employed to figure out the binding constants of these hosts by using Eq. () as reported previously. 4

5 ANALYTICAL SCIENCES DECEMBER 999, VOL Fig. 5 Sensitivity factors of β- ( ), β-2 ( ), and β-3 ( ) in a 0 vol.% ethylene glycol aqueous solution (0 6 M, 25 C) for all guests examined. Scheme 2 Guest molecules. = = I f I f0 a[cd] 0 b[cd] 0 K [G] 0 () Here, I is the fluorescence intensity at 526 nm (I f : for complex, I f0 : for the host alone), [CD] 0 is the total host concentration, [G] 0 is the total guest concentration, and a and b are constants. The binding constants of the : complex of the three hosts with several guests were obtained to examine the correlation between the fluorescence variations and the binding ability of the hosts. The results are listed in Table. The binding constants are in the orders 7>6>>9>8> for β -, and 7>6>9>8>> for β-2 and β-3. The order of the binding constants of each host for these guests is not parallel with the order of the sensitivity factors. This means that the sensitivity value gives a relative but not absolute sensing ability. Figure 6 shows response curves of β-, β-2, and β-3 for a couple of guests such as lithocholic acid, ursodeoxycholic acid, and cholic acid. Since these guests were detected with remarkably different lower detection limits by the hosts with the order of lithocholic acid<ursodeoxycholic acid<cholic acid, they are expected to have different response ranges when the

6 204 ANALYTICAL SCIENCES DECEMBER 999, VOL. 5 Table Binding constants (K/mol dm 3 ) of β-, β-2, and β-3 in a 0 vol.% ethylene glycol agueous solution (0 6 M, 25 C) a Guest β- β-2 β-3 Progesterone () 2300± 40 b 2600± ± 400 Deoxycholic acid (6) 6600± ± ±3000 Lithocholic acid (7) 65000± ± ±6600 Chenodeoxycholic acid (8) 4500± ± ± 600 Ursodeoxycholic acid (9) 7300± ± ± 630 Borneol () 870± ± ± 20 a. The K values were obtained from guest-induced fluorescence variations. b. The errors were assessed by statistical tests. Fig. 6 Fluorescence variations of β-, β-2, and β-3 in a 0 vol.% ethylene glycol aqueous solution (0 6 M, 25 C) for lithocholic acid ( ), ursodeoxycholic acid ( ), and cholic acid ( ) as a function of guest concentration. concentrations are varied. All hosts give clear concentration dependency for the guests, reflecting the sensitivities of the system for the guests with response ranges of , , and above 0 4 M for lithocholic acid, ursodeoxycholic acid, and cholic acid, respectively. Figure 7 shows the sensing parameter ( I/I) of seven hosts for each guest at the guest concentration of.0 mm, 0. mm or 0.0 mm. In this presentation, progesterone and lithocholic acid are detected by γ -analogues with higher sensitivity than β-analogues. On the other hand, chenodeoxycholic acid and ursodeoxycholic acid give expanded lines with distorted shapes of heptagon. Host β-2 detects ( )-borneol, (+)-fenchone, ( )-fenchone, cyclohexanol, and cyclooctanol with remarkably high sensitivities and host β- and β-3 recognize these small guests with high sensing parameters. These results demonstrate that each guest has its own shape and the shape representation is an indication of the molecular recognition of the hosts. In conclusion, three analogues of bis-dansylglycinemodified β-cyclodextrins have been prepared to investigate their sensing ability for organic guests, including steroids and terpenoids. The variation of these fluorescence intensities was used as a parameter to describe sensing ability. These host compounds recognize larger guests such as bile acids, because chenodeoxycholic acid and ursodeoxycholic acid were detected with remarkably higher sensitivity than those of γ -cyclodextrins, and also detect smaller guests. The position of modification affects the binding ability of these hosts, which may cause the highest sensitivity by 6 A,6 C -modification, probably due to ease of movement of the appended moieties. It is suggested that the appended moieties of these hosts work as a hydrophobic cap to increase the binding ability. The fluorescence chemosensor system using such modified cyclodextrins is a very convenient and useful method, because the chemical modification of a guest, even if it is spectroscopically inert, is not necessary; a guest can be examined directly in this system.

7 ANALYTICAL SCIENCES DECEMBER 999, VOL Fig. 7 Variations of sensitivity factors of seven hosts (0 6 M, 25 C) induced by various organic guests. This study was supported by a Grant-in-Aid for Specially Promoted Research (No. 404: Molecular Synchronization for Design of New Materials System) from the Ministry of Education, Science, Sports, and Culture of Japan. References. J.-M. Lehn, Supramolecular Chemistry, VCH, Verlagsgesellshaft, J. Szejtli, Cyclodextrin Technology, Kluwer, Dordrecht, A. Ueno, S. Minato, I. Suzuki, M. Fukushima, M. Ohkubo, T. Osa, F. Hamada and K. Murai, Chem. Lett., 605 (990). 4. F. Hamada, Y. Kondo and R. Ito, J. Inclusion Phenom. Mol. Recognit. Chem., 5, 273 (993). 5. H. F. M. Nelissen, F. Venema, R. M. Uittenbogaard, M. C. Feiters and R. J. M. Nolte, J. Chem. Soc., Perkin Trans. 2, 2045 (997). 6. K. Hamasaki, S. Usui, H. Ikeda, T. Ikeda and A. Ueno, Supramol. Chem., 8, 25 (997). 7. Y. Wang, T. Ikeda, A. Ueno and F. Toda, Chem. Lett., 863 (992). 8. Y. Wang, T. Ikeda, H. Ikeda, A. Ueno and F. Toda, Bull. Chem. Soc. Jpn., 67, 598 (994). 9. M. Narita, F. Hamada and M. Sato, J. Inclusion Phenom. Mol. Recognit. Chem., 34, 42 (999). 0. M. Narita, F. Hamada, I. Suzuki and T. Osa, J. Chem. Soc., Perkin Trans. 2, 275 (998).. G. Nerso, G. Patonay and I. M. Warner, J. Include. Phenom., 6, 277 (988). 2. S. Ito, M. Narita and F. Hamada, Int. J. of Mat. Eng. for Resources, 7, 56 (999). (Received June 30, 999) (Accepted September 9, 999)