Macromol. Rapid Commun. 2002, 23, 179–182 179

Synthesis of a-Cyclodextrin-Conjugated Poly(e-lysine)sand Their Inclusion Complexation Behavior

Kang Moo Huh, Hajime Tomita, Won Kyu Lee, Tooru Ooya, Nobuhiko Yui*

School of Materials Science, Japan Advanced Institute of Science and Technology, 1-1 Asahidai, Tatsunokuchi,Ishikawa 923-1292, JapanFax: +81-761-51-1645; E-mail: yui@jaist.ac.jp

Keywords: cyclodextrins; host-guest systems; inclusion chemistry; phase separation;

IntroductionCyclodextrins (CDs) have been extensively investigatednot only for their widespread applications in pharmaceu-tical chemistry, food chemistry and analytical chemistry,but also as building blocks for constructing supramolecu-lar structures due to their unique noncovalent bindingproperties with various guest molecules.[1–4] a-, b- and c-CDs, the most widely used CDs as hosts, consist of six,seven, and eight d-glucopyranose residues, respectively,and can be chosen considering the dimensions and struc-ture of corresponding guest molecules.[5] Other advanta-geous aspects of CDs that make them useful include awell-defined chemical structure, providing many poten-tial sites for chemical modification, water-solubility, lowtoxicity, and positive contributions to the solubility, thestability and the bioavailability of the guest molecules.[6]

CDs have found some potential applications in polymersynthesis. For example, Ritter et al. have established thefree-radical polymerization in aqueous solution, wherehydrophobic monomers, such as styrene or methacry-lates, are solubilized by complexation with methylated b-CD, finally giving water-insoluble polymers in highyield.[7]

Recently, various types of CD-containing polymershave been synthesized to enhance or modulate the physi-

cochemical properties such as inclusion complexation ofCDs for specific applications.[8, 9] In such polymeric sys-tems, CD molecules come to gather closer to one anotherso that they can participate in the complexation processsimultaneously and cooperatively. For example, using ana-CD-based molecular tube, we demonstrated more stableinclusion complexation with sodium alkyl sulfonate[10]

and supramolecular network formation with [poly(ethyl-ene oxide) monocetyl ether]-graft-dextran.[11]

In this study, to obtain a novel functional polymericsystem utilizing noncovalent binding properties of CDs,a-CDs are conjugated to poly(e-lysine) (PL), which is abiodegradable and cationic polymer,[12] as side groups.The unique interaction between a-CD-conjugated PL and3-(trimethylsilyl)propionic acid (TPA) as a model guestis investigated in aqueous media. An interesting phase-separation phenomenon is observed as a result of theinteraction between polymeric host and model guest,which has not been demonstrated and reported for normalCD molecules and other CD-containing polymers. Thiswater-soluble polymeric host would be very useful forvarious applications, especially in the biomedical field,due to the biodegradability and low toxicity of their con-stituents, and an unique functionality, represented as acomplexation-induced phase separation.

Communication: Novel functional polymers utilizingspecific host/guest interactions were designed byintroducing a-CD host molecules into poly(e-lysine)chains as side groups. An interesting phase separation wasobserved as a result of the inclusion complexationbetween the polymeric host and 3-(trimethylsilyl)propio-nic acid as a model guest in aqueous media. This water-soluble polymeric host would be useful for various appli-cations, particularly drug delivery, due to its biodegrad-ability, low toxicity, and unique functionality representedas a complexation-induced phase separation.

Macromol. Rapid Commun. 2002, 23, No. 3 i WILEY-VCH Verlag GmbH, 69469 Weinheim 2002 1022-1336/2002/0302–0179$17.50+.50/0

180 K. M. Huh, H. Tomita, W. K. Lee, T. Ooya, N. Yui

Experimental Part

Materials

a-CD was purchased from Bio-Research Co., Ltd, Yoko-hama, Japan, and purified by recrystallization from distilledwater, followed by drying in vacuo at 60 8C. PL (10% aque-ous solution, M

—w = 4700, M

—w/M

—n = 1.14) was kindly supplied

by Chisso Co., Ltd, Tokyo, Japan, and freeze-dried beforeuse. Dimethyl sulfoxide (DMSO, Wako) was dried overCaH2 and distilled. The other synthetic reagents were used asreceived without further purification.

Synthesis of Monoaldehyde a-CD (Ald-CD, 2)[13]

Ten grams of a-CD (10.3 mmol) were dissolved in 250 ml ofDMSO. Two equivalents of Dess-Martin periodinane pre-pared according to the method reported in the literature[14]

was added and the reaction mixture was stirred for 1 h atroom temperature. The solution was poured into an excessiveamount of acetone and kept at –208C for 2 h. The productwas collected by filtration and dissolved again in distilledwater to remove insoluble impurities by filtering. The aque-ous solution was concentrated and recrystallized from coldacetone, followed by filtration and drying in vacuo at 608C;yield: 88%.

1H NMR (DMSO-d6): d = 9.68 (s, formyl proton), 5.44 (m,12H, sec. OH), 4.75 (s, 5H, H-1), 4.42 (m, 5H, prim. OH),4.00–3.00 (4 m, CD protons).

IR (KBr): 3384 (s, OH), 2930 (s, C1H), 1731 (m, alde-hyde C2O), 1500–1200 (m, C1H and OH), 1154 (m,C1O), 1032 cm–1 (m, C1O).

a-CD-Conjugated PLs (4)[15]

a-CD-conjugated PLs were prepared by coupling reactionbetween Ald-CD (2) and PL, as shown in Scheme 1. Forinstance, PL (5.0 g, 1.02 mmol) was allowed to react withAld-CD (75.85 g, 78.1 mmol) in 0.5 m acetate buffer(pH4.4) at room temperature for 1 h. Sodium cyanoborohy-dride (12.28 g, 195.4 mmol) was added to the resulting solu-tion. The mixture was stirred at room temperature for 96 h,and then neutralized with 2 m sodium hydrate, followed bydialysis against water (MWCO = 10000) and lyophilization.The a-CD content in the polymers was calculated by meansof 1H NMR spectroscopy by comparing the integrationvalues originating from the peaks of the anomeric protons ofglucose units and a and e protons of lysine units); yield:64%.

1H NMR (D2O): d = 4.96 (s, 6H, H-1, CD), 4.30–3.35 (2m, CD protons), 3.35–2.6 (3 m, a and e protons, PL), 2.00–1.00 (3 m, b, c, and d protons, PL).

IR (KBr): 3413 (s, OH), 2929 (s, C1H), 1637 (s, C2O),1559 (m, N1H), 1458 (m, C1H), 1154 (m), 1077 (m),1031 cm–1 (m, C1O).

Results and DiscussionThree kinds of a-CD-conjugated PLs that contain differ-ent amounts of a-CD were synthesized according to thesynthetic route shown in Scheme 1. The results are sum-

marized in Table 1. The a-CD content (CD/Lys) in thepolymers prepared in this experiment was 0.28, 0.41, and0.64 mol-%, respectively, and could be controlled byvarying the feed ratio between PL and Ald-CD (2). Allthe polymers obtained showed high water solubility overthe whole range of pHs, irrespective of the a-CD content.The chemical composition of the polymers was con-firmed by means of 1H NMR and FT-IR spectroscopy.Figure 1 shows a representative 1H NMR spectrum of thea-CD-conjugated PL, demonstrating the presence ofcovalently bound a-CD moieties in the polymers.

The inclusion properties of a-CD and 4 with TPA as amodel guest were studied by means of 1H NMR spectro-scopy in D2O and compared with each other. The 1HNMR spectrum of a solution of TPA was measured, andthen a-CD was added to this solution in small aliquots.Figure 2 shows the changes in the 1H NMR spectra ofTPA as a function of the amount of a-CD added to TPA

Scheme 1. Synthesis of a-CD-conjugated PLs.

Table 1. Synthetic results derived from a-CD-conjugated PLs.

CodeNo.

Molar feedratio Ald-a-

CD/Lys

MnaÞ

g=molMn

bÞ

g=molCD

contentcÞ

mol-%

Yieldwt:-%

4a 0.5 :1 12100 14550 28 404b 0.8 :1 15200 15000 41 324c 2.0 :1 22600 22800 64 43

a) Calculated from the peak integration of 1H NMR spectra.b) Measured by means of MALDI-TOF mass spectrometry.c) Calculated from the peak integration of 1H NMR spectra.

Figure 1. 1H NMR spectrum of a-CD-conjugated PL in D2O.

Synthesis of a-Cyclodextrin-Conjugated Poly(e-lysine)s and Their Inclusion Complexation Behavior 181

solution. In the absence of a-CD, the methyl protons ofTPA gave a sharp and single signal (A) around d = 0.0(Figure 2a), while with the introduction of a-CD theresonance for TPA gradually broadened and was dimin-ished in intensity. At the same time, a new resonanceresulting from the methyl groups of TPA complexed witha-CD grew in proportion to the amounts of a-CD added.Subsequently, further addition of a-CD resulted in a shar-pening of new signal A9 and the disappearance of originalsignal A. The methylene protons of TPA (B) showedsimilar patterns.

The corresponding 1H NMR spectra of TPA for a-CD-conjugated PL are represented in Figure 3 as a functionof the amount of added 4a. While the methyl protons ofTPA gave a single signal, the introduction of 4a to the

TPA solution induced the marked broadening of the sig-nal together with the appearance of a new peak corre-sponding to complexation. These results are due to theinclusion complexation of a-CD molecules, which areconjugated to PL backbones. Consequently, the a-CDmoieties covalently bound to PL chains were found toretain their inclusion ability. There have been severalsimilar reports on more stable inclusion complexes ofCD-containing polymers with guest molecules due to acooperation effect by multiple binding between the adja-cent CD molecules along the polymers. The cooperativitydepended on the distance between neighboring CD mole-cules in the polymer structure.[16]

In addition to the inclusion ability, these CD-contain-ing polymers were found to exhibit an interesting phe-nomenon resulting from inclusion complexation withTPA, represented as a complexation-induced phaseseparation. Except of 4a, which contained the lowest CDcontent, the a-CD-conjugated PLs showed phase-separa-tion phenomena after adding the polymers to TPA solu-tion. There was no phase separation when TPA wasadded to an a-CD or PL solution, indicating that theinclusion complexation of a-CD with TPA molecules andthe possible ionic interaction between the amino-groupsof PL and TPA cannot directly induce such phase separa-tion. In addition to intra- and intermolecular multivalentbinding between the polymers and a-CD molecules, theionic interactions between the amino groups of PL andTPA molecules included into the CD cavities of the otherpolymers can also contribute to the phase separation ofthe solution. As a result, TPA acts as a noncovalent cross-linker via inclusion complexation and ionic interaction.The contribution of the ionic interaction to phase separa-

Figure 2. Changes in the 1H NMR spectra of TPA as a functionof the amount of added a-CD; [a-CD]/[TPA] = 0 :1 (a), 0.5 :1(b), 1 :1 (c), 1.5 :1 (d), and 4:1 (e).

Figure 3. Changes in the 1H NMR spectra of TPA as a functionof the amount of added a-CD-conjugated PL; [a-CD]/[TPA] =0 :1 (a), 1 :1 (b), 2 :1 (c), and 3 :1 (d).

Figure 4. Schematic representation of the interaction betweena-CD and TPA (a) and the interaction between a-CD-conjugatedPL and TPA (b).

182 K. M. Huh, H. Tomita, W. K. Lee, T. Ooya, N. Yui

tion could be confirmed by the fact that the ionic strengthin aqueous media significantly affected the solution prop-erties and resulting phase-separation behavior (data notshown). Therefore, such a phenomenon should be aunique property due to the interactions between the poly-meric host and TPA molecules exclusively. Figure 4shows a schematic representation of the interactionbetween a-CD and TPA (a) and the interaction betweena-CD-conjugated PL and TPA (b).

More detailed studies for inclusion complexation andsolution properties are in progress. Such complexation-induced phase separation properties could be useful forpotential applications resulting from the specificity andselectivity of the CD molecules toward a great number ofguests including drugs and proteins.

Acknowledgement: A part of this study was financially sup-ported by Grants-in-Aid from the Mitsubishi Foundation, Japanand Scientific Research for Priority Area “Molecular Synchroni-zation for Construction of New Materials System” (No. 404/11167238) from MESSC, Japan.

Received: November 12, 2001Revised: January 3, 2002

Accepted: January 7, 2002

[1] G. Wenz, Angew. Chem. Int. Ed. Engl. 1994, 33, 803.[2] J. Szejtli, Chem. Rev. 1998, 98, 1743.[3] M. O. Ahmed, I. El-Gibaly, S. M. Ahmed, Int. J. Pharm.

1998, 171, 111.[4] C. C. Rusa, A. E. Tonelli, Macromolecules 2000, 33, 1813.[5] L. Huang, A. E. Tonelli, J. Macromol. Sci. – Macromol.

Chem. Phys. 1998, C38(4), 781.[6] K. Uekama, Adv. Drug Deliv. Rev. 1999, 36, 1.[7] [7a] S. Bernhardt, P. Glöckner, A. Thesis, H. Ritter,

Macromolecules 2001, 34, 1647; [7b] P. Casper, P. Glöck-ner, H. Ritter, Macromolecules 2000, 33, 4361; [7c] J.Jeromin, H. Ritter, Macromol. Rapid Commun. 1998, 19,377.

[8] A. Ruebner, G. L. Statton, M. R. James, Macromol. Chem.Phys. 2000, 201, 1185.

[9] Ger. 19612768 (1997), BASF AG, invs.: J. Huff, A. Kisten-macher, G. Schornick, M. Weickenmeier, G. Wenz; Chem.Abstr. 1997, 127, 294896z.

[10] [10a] T. Ikeda, T. Ooya, N. Yui, Polym. Adv. Technol.2000, 11, 830; [10b] T. Ikeda, E. Hirota, T. Ooya, N. Yui,Langmuir 2001, 17, 234.

[11] T. Ikeda, T. Ooya, N. Yui, Macromol. Rapid Commun.2000, 21, 1257.

[12] M. Kunioka, H. J. Choi, J. Appl. Polym. Sci. 1995, 58, 801.[13] M. J. Cornwell, Tetrahedron Lett. 1995, 36, 8371.[14] R. E. Ireland, L. Liu, J. Org. Chem. 1993, 58, 2899.[15] T. Tojima, J. Polym. Sci. Part A: Polym. Chem. 1998, 36,

1965.[16] A. Deratani, B. Pöpping, Macromol. Chem. Phys. 1995,

196, 343.