Anal. Chem. 1998, 70, 580-589
A Family of Single-Isomer Chiral Resolving Agents for Capillary Electrophoresis. 3. Heptakis(2,3-dimethyl-6-sulfato)-β-cyclodextrin Hong Cai, Thanh V. Nguyen, and Gyula Vigh*
Department of Chemistry, Texas A&M University, College Station, Texas 77845-3255
The third member of a new family of single-isomer charged cyclodextrins, the sodium salt of heptakis(2,3-dimethyl6-sulfato)-β-cycldextrin, has been synthesized, characterized, and used for the capillary electrophoretic separation of the enantiomers of neutral, acidic, basic, and zwitterionic analytes. Though heptakis(2,3-dimethyl-6-sulfato)β-cyclodextrin complexes much less strongly with any of the analytes tested here than the previously synthesized heptakis(2,3-diacetyl-6-sulfato)-β-cyclodextrin and heptakis-6-sulfato-β-cyclodextrin, it offers excellent enantioselectivities, complementary to those of the other two single-isomer, differently functionalized charged cyclodextrins. Confirming the predictions of the charged resolving agent migration model, heptakis(2,3-dimethyl6-sulfato)-β-cyclodextrin allowed for the reversal of the migration order of the enantiomers of neutral analytes as the cyclodextrin concentration was increased. Just as with the previous two single-isomer charged resolving agents, separation selectivity for the acidic, basic, and zwitterionic analytes could increase, decrease, or pass a maximum as the cyclodextrin concentration was increased, depending on the respective binding strength of the enantiomers and the ionic mobilities of both the complexed and noncomplexed forms of the enantiomers. In view of the increasing role of capillary electrophoresis (CE) in the efficient separation of enantiomers,1,2 our laboratory initiated a long-term project with multiple objectives: (i) to provide a comprehensive theoretical framework for the description of the electrophoretic migration of the enantiomers in order to identify the operating variables and their ranges that are most likely to offer successful separations (implemented in the charged resolving agent migration model, or the CHARM model3), (ii) to synthesize charged cyclodextrins that offer unique intermolecular interactions by having different functional groups connected to the 2- and 3-carbon atoms of the glucopyranose subunits of cyclodextrins,4,5 and (iii) to eliminate the most common drawback of the commercially available charged cyclodextrins, namely the presence (1) St. Claire, R. L. Anal. Chem. 1996, 68, 569R. (2) Fanali, S. J. Chromatogr. A, 1996, 735, 77. (3) Williams, B. A.; Vigh, Gy. J. Chromatogr. 1997, 776, 295. (4) Vincent, J. B.; Sokolowski, A. D.; Nguyen, T. V.; Vigh, Gy. Anal. Chem. 1997, 69, 4226. (5) Vincent, J. B.; Kirby, D. M.; Nguyen, T. V.; Vigh, Gy. Anal. Chem. 1997, 69, 4419.
580 Analytical Chemistry, Vol. 70, No. 3, February 1, 1998
of numerous isomers which makes these preparations ill-defined complex mixtures with ill-defined mixed properties.6-14 In parts 1 and 2 of this series,4,5 we described the synthesis and use of single-isomer β-cyclodextrin derivatives which are completely sulfated in the 6-positions and completely substituted on their larger rims with hydrophilic (hydroxy)5 and moderately hydrophobic (acetyl) functional groups.4 This paper describes the synthesis, characterization, and use of the third member of the new single-isomer, fully charged cyclodextrin family, a derivative which carries the hydrophobic methyl functional group on its larger rim, heptakis(2,3-dimethyl-6-sulfato)-β-cyclodextrin. EXPERIMENTAL SECTION Synthesis of Heptakis(2,3-dimethyl-6-sulfato)-β-cyclodextrin. Except for β-cyclodextrin (CD), which was a gift from Cerastar (Hammond, IN), all chemicals were purchased from Aldrich Chemical Co. (Milwaukee, WI). The sodium salt of heptakis(2,3-dimethyl-6-sulfato)-β-cyclodextrin was synthesized according to Figure 1. First, heptakis-6-dimethyl-tert-butylsilyl-βcyclodextrin was obtained by reacting β-CD with dimethyl-tertbutylchlorosilane according to ref 15 and purifying the raw reaction mixture by preparative gradient elution column chromatography16 using a silica gel column and the n-hexane/ethyl acetate/ethanol eluent system.15 (The 50-mm-i.d., 300-mm-long preparative HPLC column packed with 30-nm pore size, 10-µm irregular silica (Merck, Darmstadt, Germany) was generously loaned to us by Dr. Y. Y. Rawjee of Smith-Kline Beecham, King of Prussia, PA.) The pure intermediate was then methylated with iodomethane in the presence of NaH,15 and the resulting reaction mixture was (6) Tait, R. J.; Skanchy, D. J.; Thompson, D. O.; Chetwyn, N. C.; Dunshee, D. A.; Rajewsky, R. A.; Stella, V. J.; Stobaugh, J. F. J. Pharm. Biomed. Anal. 1992, 10, 615. (7) Tait, R. J.; Thompson, D. O.; Stobaugh, J. F. Anal. Chem. 1994, 66, 4013. (8) Endresz, G.; Chankvetadze, B.; Bergenthal, D.; Blaschke, G. J. Chromatogr. A 1996, 732, 133. (9) Chankvetadze, B.; Endresz, G.; Schulte, G.; Bergenthal, D.; Blaschke, G. J. Chromatogr. A 1996, 732, 143. (10) Szeman, J.; Ganzler, K.; Salgo, A.; Szejtli, J. J. Chromatogr. A 1996, 728, 423. (11) Chankvetadze, B.; Schulte, G.; Blaschke, G. J. Chromatogr. A 1996, 732, 183. (12) Stalcup, A. M.; Gahm, K. H. Anal. Chem. 1996, 68, 1360. (13) Gahm, K. H.; Stalcup, A. M. Chirality 1996, 8, 316. (14) Jung, M.; Francotte, E. J. Chromatogr. A 1996, 755, 81. (15) Takeo, K.; Hisayoshi, M.; Uemura, K. Carbohydr. Res. 1989, 187, 203. (16) Vigh, Gy.; Quintero, G.; Farkas, Gy. J. Chromatogr. 1989, 484, 237. S0003-2700(97)00822-6 CCC: $15.00
© 1998 American Chemical Society Published on Web 01/06/1998
Figure 1. Synthesis scheme for heptakis(2,3-dimethyl-6-sulfato)β-cyclodextrin.
extensively purified by preparative gradient elution column chromatography using the n-hexane/ethyl acetate eluent system.15 This second intermediate was then reacted with HF in ethanol17 to remove the tert-butyldimethyl silyl protecting group. The excess HF was carefully neutralized with NaHCO3, and the inorganic components were removed from the reaction mixture by filtration. After removal of the alcohol-water solvent mixture by rotary evaporation, the crude reaction mixture containing mostly heptakis(2,3-dimethyl)-β-CD was once again repurified by preparative gradient elution column chromatography using the n-hexane/ethyl acetate eluent system.15 The pure third intermediate was then sulfated with pyridinesulfonate18 in N,N′-dimethylformamide as solvent. Completeness of the sulfation reaction was monitored by indirect UV detection CE19 using a 20 mM p-toluenesulfonic acid (PTSA) background electrolyte,4 whose pH was adjusted to 8 with tris(hydroxymethyl)aminomethane (TRIS). Once the sulfation reaction was complete, the reaction mixture was poured into a 10-fold excess of acetone, and the gummy product was separated, redissolved in water, and titrated with NaOH to liberate pyridine. The sodium sulfate byproduct was removed by repeated partial precipitation using ethanol. Finally, heptakis(2,3-dimethyl-6-sulfato)-β-cyclodextrin was obtained by pouring the sodium sulfate-free aqueous solution into excess ethanol, collecting the precipitate, and carefully drying it in a vacuum oven overnight. The indirect UV detection electropherogram of a typical heptakis(2,3-dimethyl-6-sulfato)-β-CD sample, obtained with a typical
