J. CHEM. SOC. FARADAY TRANS.. 1994. 90(61. 845-84" Cyclodextrin-Monosaccharide Interactions in Water Angela F. Danil de Namor," Peter M. Blackett, Mercedes C. Cabaleirot and Jasim M. A. Al RawiS Laboratory of Thermochemistry, Department 0'' Chemistry, University of Surrey, Guildford, Surrey, UK GU25XH Stability constants and derived Gibbs energies, enthalpies and entropies of complexation of D-monosaccharides with 2-and /I-cyclodextrinll in water at 298.15 < obtained by titration microcalorimetry are reported. The results show that x-cyclodextrin interacts with D-glucose. o-fructose, D-xylose, D-mannose and o-galactose. No heat was evolved with D-arabinose. However, /I-cyclodextrin is able to recognise aldopentoses (o-xylose and D-arabinose) but not aldohexoses.13CNMR studies on these systems are discussed. Cyclodextrins are cyclic oligosaccharides constituted by Y-J-glucopyranose units and are known to interact with a ~itie variety of substrates. An important feature of cyclodextrins is that these compounds are characterised by their low degree of toxicity and therefore. these ligands have found numerotis applications in the pharmaceutical and food industries.'.' Despite the large number of contributions in this area. thermodynamic studies on the binding (A, G .A, H-,A, S I *)f these ligands with different substrates are relatively scarce It seems appropriate to emphasise that the Gibbs energy )f complexation is a most relevant parameter sincc it reflects the selectivity of a host for a given guest.However, interpretation based solely on A, C;' values c:!n be misleading and therefore it is always desirable to evalu,i!e the enthalpy and entropy associated with these processes. Stability constant data so far reported in cyclodevtrin chemistry3 are within the range required to be measured accurately by titration calorimetry and therefore this tec:i-nique offers the advantage that not only the stability but also the enthalpy can be accurately derived. particularly when highly sensitive microcalorimetric systems are used for t htse purposes. This paper reports: (i) Thermodynamic data for the con -plexation of D-monosaccharides and x-and p-cyclodext rir:s in water at 298.15 K derived from titration microcalorimetr! : (ii) 13C NMR studies on various monosaccharides and 5-cyclodextrin in D,O at the same temperature. Experimental x-(from Aldrich) and /j-(from Sigma) cyclodextrins wore dried in a vacuum oven for 3 days at 343 K prior to ujc.11-Glucose. D-, L-, and m-xylose, D-fructose, methyl-fl-u-g! L-copyranoside, methyi-/I-galactopyranoside and methyl 2-14-glucopyranoside (all from Fluka), D-mannose. 11-arabinoe and 3-O-methyl glucose (Aldrich) and 1,-galactose (BDt-i) were dried under vacuum for a few hours at 313 K before IM. Heats of complexation of cyclodextrins and monosaccha -rides were measured in a Thermal Activity Monitor micit- calorimetric system purchased from Thermonierric. Aqueou solutions of cyclodextrins (0.08--0.10rnol dm- '1 in the calcl-metric vessel were titrated with the monosaccharide n ate-t Present address: Instituto de Quimica Organica.Departamrnt!, de Quimica & Ingenieria Quimica. Universidad Nacional del Stir. Bahia Blanca 8000. R. Argentina. Present address: Department of Chemistry. University of Sanxi. Yemen. r-Cyclodextrin = cyclomaltohexaose: /I-cyclodaxtrin = cycia-maltoheptaose. solution', (1-3 rnol dm-3) contained in a 500 p1 gas-tight motor driven Hamilton syringe. All calorimetric measure- ments ivere performed at 298.15 K and the solvent was doubly distilled deionised water. The reliability of the calo- rimeter was checked using the test calibration of Ba" and 18 crwn 6 ether in water at 298.15 K suggested by Briggner and Wadso.' A value of -31.86 & 0.32 kJ mol-' was obtained.in excellent agreement with the literature value of -3 1.42 & 0.20 kJ mol- '. All calorimetric data experiments performed in triplicate. 'c' NMR experiments were carried out on a Bruker AC-300 MHz spectrometer with a wide bore magnet oper- ating in the Fourier-transform mode using proton decoupling for I 'C. Spectra were internally referenced with 1P-dioxane relative to tetramethylsilane (TMS). For the 13C NMR experiments. the spectra of r-cyclodextrin (ca. 1 x rnol dm 3, and then, with an excess (1 x 10.' rnol dm-3) of the appropmte monosaccharide were recorded. These studies were also carried out in the reverse order by recording the spectra of the monosaccharide (0.01 rnol dm-3) and then adding an excess of x-cyclodextnn (0.1 rnol dm -').Results and Discussion Thermodynamic Data of Complexation From calorimeter data, values for the stability constant (expressed as log K,) and the enthalpy of complexation A, H were calculated using a minimisation program developed by Karlson and Kullberg.' The thermodynamic data fitted a model which corresponds to a 1 : 1 (monosaccharide: cyolodext rin) stoichiometry complex. Therefore, thermodyna- mic data for the interaction of cyclodextrins (CD)with mono- saccharides (M) in water at 298.15 K are referred to the process. M(H,O) + CD(H20j-+ M *CD(H20) (1) Table 1 lists log K, and derived Gibbs energies, A,G'. enth-alpies. Ac Fi'. and entropies, A, S values for the complexation process [qn.(I)]. The individual errors in K, and Ac H' are expressed as the standard deviation (IT)of the mean using the expression (T = (Xui -.f):(n -1). where n is the number of steps (at least six) considered in each calorimetric run. The CJ values given in Table 1 are the weighted average of three calorimetric runs for each monosaccharide considered. Note that it was the availability of a highly sensitive micro- calorimetric system which made possible the quantitative evaluation of the enthalpic contribution. Indeed. enthalpy data could not be obtained by classical titration calorimetry. Thermodynamic data show the following. J. CHEM. SOC. FARADAY TRANS., 1994, VOL. 90 Table 1 Thermodynamic parameters of complexation of (a-and B-)cyclodextrins with D-monosaccharides in water at 298.15 K monosaccharide 1% K, Ac G"/kJ mol - Ac H"/kJ mol - A,So/J K-' mol-' D-glucose D-fructose 1.56 k0.23 1.72 f0.24 a-cyclodext rin -8.90 f1.31 -9.82 f1.30 -0.14 -0.05 f0.03 k0.01 29.4 f4.4 32.8 f4.6 D-xylose D-mannose 1.57 & 0.11 1.77 f0.17 -8.96 f1.37 -10.10 f0.97 -0.09 f0.01 -0.11 f0.01 29.8 f2.1 33.5 f3.2 D-galactose D-arabinose 1.19 f0.31 -6.79 f1.77 -0.32 _+ 0.12 21.7 _+ 5.9 D-arabinose D-xylose 1.21 & 0.32 1.22 * 0.10 B-cyclodextrin -6.91 f1.82 -6.96 f0.57 -0.42 -0.62 0.22 k0.06 21.8 & 6.1 21.3 f1.1 (a) In water, low-stability complexes are formed. This is conformers) in water or indeed to a very small enthalpy mainly attributed to strong host-water and guest-water associated with the binding process. This is now being inves- interactions.As far as the monosaccharides are concerned, tigated further. these molecules are able to interact with water through The most striking feature of these results is the ability of hydrogen-bond formation. Attachment of water to cyclo- /3-cyclodextrin to interact with aldopentoses but not with dextrins occurs through the hydroxy groups occupying both aldohexoses in water. This was previously reported by As a Aoyama et aL7 on the basis of 13C NMR studies ofrims of the cone as well as the inside of the ~avity.~ result of the individual attraction shown by the host and the aligosugar-sugar interactions in water. In fact, these authors guest for the solvent, the binding between the cyclodextrin attributed the lack of interaction of this macrocycle with and the monosaccharide is relatively weak. Stability con-aldohexoses to steric factors, suggesting that the presence stants for monosaccharides and /3-cyclodextrins in water have (aldohexoses) or the absence (aldopentoses) of a substituent been reported by Aoyama et aL7 However, the data reported in the C-5 of the pyranoside ring could be a key factor as far by these authors can be only regarded as tentative values as /3-cyclodextrin -monosaccharide interactions are con-since the overall error reported (520%) is relatively large cerned.In summary, the broad conclusions drawn from and the temperature at which these measurements were Table 1 are that (i) a-cyclodextrin is unable to recognise these carried out was not reported. monosaccharides selectively as assessed from stability con- (b)The complexation of monosaccharides to cyclodextrins stant data (D-galactose lies just outside of the range) and (ii) (exothermic reaction) must be accompanied by strong desol- /?-cyclodextrin distinguishes between aldopentoses and aldo- vation of both the host and the guest molecules (endothermic hexoses.reaction) upon complexation, leading to a small enthalpy Encouraged by the realisation that cyclodextrins could dis- change for this process. Positive entropies are often found for tinguish between aldohexoses and aldopentoses, enantio- reactions in which the host, guest or both undergo strong meters of xylose were investigated in order to assess if desolvation upon complexation, and this is also observed in cyclodextrins could selectively recognise one isomer from these systems (see Table 1).No heat was detected when a- another. The stability constant should determine this, and cyclodextrin was titrated with D-arabinose. This result on its these data are shown in Table 2. Also reported in this table own does not provide evidence that this monosaccharide are the contributions of the enthalpy and entropy to the does not interact with a-cyclodextrin. It could be that the Gibbs energy of these processes. In order to test whether a-heat associated with this process is too small to be measured, cyclodextrin is able to discriminate between the isomers, the in which case calorimetry is not a suitable reporter for this 'student's' t-distribution test was applied to these data and it system.However, it should be stressed that, among the was concluded that at the 95% confidence interval, a claim monosaccharides considered, the fC conformer predomi-that a-cyclodextrin is able to discriminate between these nates, except for arabinose.* For this monosaccharide, the isomers is not justified. iC conformer is the predominant species in solution. There- fore, it is most interesting to investigate whether the absence Blocked Monosaccharid4 yclodextrin Interactions of heat observed is attributed to the lack of interaction A few blocked monosaccharides (methyl-a-D-glucopyrano-between a-cyclodextrin and arabinose (in which case this side, methyl-/3-glucopyranoside, 3-0-methyl glucose and ligand could have a potential role in the recognition of methyl-/3-D-galactopyranoside)were chosen to try to locate Table 2 Thermodynamic parameters of complexation of xyloses with (a-and b-)cyclodextrins in water at 298.15 K monosaccharide 1% K, Ac Go/kJ mol -' Ac H"/kJ mol-' AcSo/J K-' mol-' a-c yclodextrin D-xylose 1.57 k0.1 1" -8.9 f0.63 -0.09 &-0.01" 29.8 k2.1 L-xyiose 2.07 5 0.37 -11.81 5 2.1 1 -0.12 f0.01 39.2 & 7.1 DL-xylose 1.42 f0.13 -8.10 f0.74 -0.11 * 0.01 26.8 & 2.5 B-cyclodextrin D-X ylose 1.22 f0.10" -6.96" f0.57 -0.62 f0.06" 21.3 f1.9 L-xylose 1.32 -t 0.13 -7.53 f0.74 -0.87 rf 0.12 22.3 f2.5 DL-xylose 1.06 -t 0.07 -6.05 f0.40 -1.12 f0.18 16.5 f 1.5 a From data listed in Table 1.J. CHEM.SOC. FARADAY TRANS., 1994, VOL 90 Table 3 A6 values in the 13CNMR of monosaccharides in the pres- ence of r-cyclodextrin at 298.15 K A6 D-mannose' D-~~UCOS~' D-fructose" D-xylose" c-1 0.10 0.11 0.03 0.13 c-2 0.11 0.12 0.04 0.11 c-3 0.11 0.14 0.06 0.13 c-4 0.07 0.02 0.02 0.11 c-5 0.14 0.12 0.07 0.14 C-6 0.07 0.07 0.06 -The chemical shifts (6) in ppm of the carbons of the monosaccha- rides are as follows: D-mannose; = 94.48; 6,, = 72.03; 6c-3~1 73.86; 6,, = 67.66; dCm5= 76.98 ; dC-6 = 6 1.79 ; D-glucose; dc-96.72; 6,, = 74.95; 6,, = 76.76; 6,, = 70.46; dc-5 = 76.57; dCa6== 61.57; D-fructose; dC-, = 98.88; aC-, = 75.27; 6c.3 = 76.20; 8c-4 = 64.70; 6c-5 = 76.20; = 63.48 and D-xylose; = 97.43; 6,, = 73.63; 6c-3 = 76.63;aC-, = 70.22 and 6c-5 = 66.00.the site of complexation on the pyranose ring for glucose. No heat was observed for the blocked monosaccharides with 3-cyclodextrin except for methyl z-D-ghcopyranoside (log K,= 1.32 k0.23; AcGo= -7.53 & 1.31 kJ mol-'; Ac H"= -0.27 f0.04 kJ mol-' and AcSo= 24.4 & 4.4 J K-'mol-'). It appears that as far as the receptor binding to the monosaccharide is concerned, the r-conformer is the important site. Indeed no heat was detected between the p-conformer or with the monosaccharide substituted at C-3. Table 4 A6 values in the I3C NMR of a-cyclodextrin in the pres- ence of monosaccharides at 298..15 K AJO D-mannose mglucose D-fructose D-xylose c-1 0.10 0.12 0.00 0.13 c-2 0.08 0.09 0.01 0.19 c-3 0.11 0.11 0.02 0.14 c-4 0.11 0.11 0.01 0.14 c-5 0.09 0.09 0.00 0.12 C-6 0.05 0.05 -0.03 0.08 'The chemical shifts (6) in ppm of the carbons of r-cyclodextrin in D,O at 298.15 K are: 6,, = 102.27; a,--, = 74.11; 6,, = 72.85; dc-4 = 82.09;6c-5= 72.49 and 6c-6= 61.16.Further studies are being carried out to investigate whether or not this is applicable to other monosaccharides. I3C NMR Studies A8 values for the I3C NMR of monosaccharides in the pres- ence of x-cyclodextrin and vice uersa are shown in Tables 3 and 4. The 13C NMR signals of the carbons of mannose, glucose and xylose (Table 4) particularly those arising from carbons 1-6 show, in the presence of z-cyclodextrin, shifts towards lower fields.The much smaller shifts observed for fructose are also reflected in the AcH" value for this mono- saccharide and x-cyclodextrin shown in Table 1. Since all these monosaccharides can exist as ;C,conformers corre- sponding to a configuration in which the 3-OH is axial, it should be reasonable to assume that the ring oxygen and the 3-OH play an important role in the monosaccharide-r- cyclodextrin interaction that affects the resonance frequencies of the carbon atoms. This suggestion seems to be supported by the lack of heat observed (see text) when the 3-OH of the monosaccharide is in equatorial position or methylated. Again, the fact that methyl-b-D-glucopyranose does not give detectable heat with a-cyclodextrin could be explained on steric grounds, since the methoxy group could prevent the interaction of the neighbouring ring oxygen. P.M.B. thanks SERC (UK) for a scholarship. References D. Duchgne, in Cyclodextrins and the Industrial b'ses, ed. D. DuchCne, Editions de Sante, Paris, 1987. ch. 6. H. Hashimoto, in Proc. 4th Int. Symp. on Cyclodextrins, ed. 0. Huber and J. Szejtli, Kluwer, Dordrecht, 1988, p. 533. R.Traboulssi, Ph.D. Thesis, University of Surrey, 1990. L. E. Brignner and I. Wadso, J. Biochem. Biophys., 1991,22, 101. R.Karlsson and L. Kullberg, Chem. Scr., 1976,9, 54. A. Cesaro, in Thermodynamic Data for Biochemistry and Biotech- nology, ed. H-J. Hinz, Springer-Verlag, 1986, ch. 6, p. 178. Y. Aoyama, Y. Nagai, J. Otsuki, K. Kobayashi and H. Toi. Anyew. Chem., Int. Ed. Engl., 1992, 31, 745. J. F. Stoddard, Stereochemistry of Carbohydrates, Wiley Inter- science, New York, 1971. Paper 3/04644C; Received 3rd August, 1993