Enantiomeric separation of sulfonium ions by capillary electrophoresis using neutral and charged cyclodextrins

Article abstract of DOI:10.1021/ac971379j

Capillary zone electrophoresis was successfully applied, for the first time, to the chiral separation of structurally related sulfonium ions, using sodium phosphate buffer pH 2.5 withy β-cyclodextrin (β-CD) or sutfated-β-cyclodextrin (S-β-CD) as the chira

Full text of DOI:10.1021/ac971379j

Anal. Chem. 1998, 70, 3612-3618
En a n t io m e ric S e p a ra t io n o f S u lfo n iu m Io n s b y
Ca p illa ry Ele c t ro p h o re s is Us in g Ne u t ra l a n d
Ch a rg e d Cyc lo d e x t rin s
Fra nc is c o A. Va le nzue la , Thom a s K. Gre e n, a nd Da rw in B. Da hl*
Department of Chemistry, Western Kentucky University, Bowling Green, Kentucky 42101
Capillary zone electrophoresis was successfully applied,
for the first time, to the chiral separation of structurally
related sulfonium ions, using sodium phosphate buffer
pH 2 .5 with â-cyclodextrin (â-CD) or sulfated-â-cyclodex-
trin (S-â-CD) as the chiral selector with tetrabutylammo-
nium bromide (TBA). For this study, a series of struc-
turally related sulfonium ions in which one of the alkyl
chains varied in length were synthesized from a common
sulfide. The resolution of the ions was found to be
dependent on the type of cyclodextrin used, the presence
or absence of TBA, and the structure of the sulfonium ion.
â-CD was found to be effective only for ions containing
two aromatic groups, while the S-â-CD was effective only
for ions containing one aromatic group. The chiral
separation of thiophenium ions was also studied under
the conditions established. Chiral separation of sulfo-
nium ions in a binary buffer system containing methanol
was explored as well. Separations were achieved for all
but one sulfonium ion and one thiophenium ion. The data
presented show the effectiveness of the enantiomeric
separation using CZE with cyclodextrin-modified buffer
for these types of ions.
sulfonium salts have found certain applications, i.e., cationic
initiators of polymerization,7 the importance of chiral sulfonium
salts has yet to be fully appreciated. Chirality around a sulfonium
center is often found to play an integral role in biological systems.
It has been determined that methyltransferases use substrates
containing chiral sulfonium centers as their methyl donors.10
-9
S-Adenosyl-L-methionine is an important methyl donor that has
such a center, and it has been additionally established that
preference is given to one enantiomeric conformation over the
other.1
1-13
One of the most promising applications investigated
so far is the use of optically active sulfonium ions as asymmetric
alkylating agents.14 The synthetic value of such an agent is
evident. The synthesis of enriched or optically pure enantiomers
requires the use of an analytical method to assess the enantiomeric
excess. Conventional methods, i.e., polarimetry, for the analysis
may not prove to be satisfactory. Although an NMR procedure
for determining enantiomeric excess using chiral shift reagents
has been developed,15 it requires extended sample preparation
time and the use of expensive shift reagents.
CZE has been shown to be a very effective analytical technique
for a wide range of compounds and, therefore, has received
considerable attention in efforts to separate difficult mixtures and
racemic compounds. Separation of analytes in CZE is achieved
by the difference in the net electrophoretic mobility of charged
compounds under the influence of an electric field. The migration
time will vary according to the mass-to-charge ratio of the species.
Neutral compounds can also be analyzed, but typically this is
achieved via different electrophoretic methods, i.e., micellar
electrokenetic chromatography (MEKC).16 A very important area
of application is the enantiomeric separation of optically active
The analysis of tertiary sulfonium compounds using currently
available methods often encounters drawbacks in terms of ef-
ficiency and sensitivity. Previously reported procedures for
1
2
analysis include ion chromatography, liquid chromatography,
paper electrophoresis, and thin-layer electrophoresis.4 The use
of capillary zone electrophoresis (CZE) offers an unprecedented
analytical advantage over these published methods. Valenzuela
et al. have reported preliminary data concerning the analysis of
3
(
7) Hamazu, F.; Akashi, S.; Koizumi, T.; Takata, T.; Endo, T. J. Polym. Sci. Polym.
Chem. 1 9 9 1 , 29, 1675-1680.
sulfonium compounds using CZE and determined the technique
was both rapid and highly effective.5
(8) Hamazu, F.; Akashi, S.; Koizumi, T.; Takata, T.; Endo, T. J. Polym. Sci. Polym.
Chem. 1 9 9 1 , 29, 1845-1851.
Little has been reported on the study of chiral sulfonium salts,
and only a handful of these compounds have actually been
assigned absolute configurations.6 Although racemic mixtures of
(
9) Sundell, P. E.; Jonsson, S.; Hult, A. J. Polym. Sci. Polym. Chem. 1 9 9 1 , 29,
1535-1543.
(10) Eichler, D. C. Biochimie 1 9 9 4 , 76, 1115-1122.
11) Segal, D. M.; Eichler D. C. Arch. Biochem. Biophys. 1 9 8 9 , 275, 334-343.
(
*
Corresponding author: E-mail: darwin.dahl@wku.edu. Fax: 502-745-5361.
(12) Wagner, J.; Hirth, Y.; Piriou, F.; Zakett, D.; Claverie, N.; Danzin, C. Biochem.
(
(
(
(
1) Hoffman, J. L. J. Chromatogr. 1 9 9 1 , 588, 211-216.
2) Massardier, V.; Vialle, J. Anal. Chim. Acta. 1 9 9 3 , 281, 391-396.
3) Mazurkiewicz, B. J. Chromatogr. 1 9 8 5 , 328, 439-442.
Biophys. Res. Commun. 1 9 8 5 , 133, 546-553.
(13) Zappia, V.; Zydek-Cwick, C. R.; Schlenk, F. Biochim. Biophys. Acta 1 9 6 9 ,
178, 185-187.
4) Gorham, J.; Coughlan S. J.; Storey, R.; Jones, W. R. G. J. Chromatogr. 1 9 8 1 ,
(14) Umemura, K.; Matsuyama, H.; Watanabe, N.; Kobayashi, M.; Kamigata, N.
2
10, 550-554.
5) Valenzuela, F. A.; Dahl, D.; Green, T. J. Chromatogr. A 1 9 9 8 , 802, 395-
98.
6) Andersen, K. K.; Caret, R. L.; Ladd, D. L. J. Org. Chem. 1 9 7 6 , 41, 3096-
100.
J. Org. Chem. 1 9 8 9 , 54, 2374-2383.
(15) Green, T. K.; Whetstine, J. R.; Son, E. J. R. Tetrahedron: Asymmetry 1 9 9 7 ,
8, 3175-3181.
(
(
3
(16) Marina, M. L.; Benito, I.; D ´ı ez-Masa, J. C.; Gonz a´ les, M. J. Chromatographia
3
1 9 9 6 , 42, 269-272.
3612 Analytical Chemistry, Vol. 70, No. 17, September 1, 1998
S0003-2700(97)01379-6 CCC: $15.00 © 1998 American Chemical Society
Published on Web 08/05/1998

Fig u re 1 . Molecular structures and names for the ions used in the study. All compounds are BF4- salts.
compounds. The significance of the different activity of enanti-
omers in biological systems is well understood. Stereospecific
reactions are carried out preferring one enantiomeric form of a
substrate or drug over the other. Thus, enantiomeric separations
are of principal importance in the pharmaceutical analysis of chiral
drugs.17,18 Enantiomeric separation is typically achieved by the
addition of a chiral resolving agent such as chiral surfactants,
crown ethers, bile salts, or cyclodextrins to the separation
buffer.19,20
example, typical commercial CDs development kits include
â-cyclodextrin, 2-hydroxypropyl-â-cyclodextrin, heptakis(2,6-di-O-
methyl)-â-cyclodextrin, heptakis(2,3,6-tri-O-methyl)-â-cyclodextrin,
and γ-cyclodextrin which can be used to optimize specific
analyses.26
In this report, we describe, for the first time, the effectiveness
of â-CD-modified enantiomeric separation of chiral sulfonium ions
using CZE. In this study, we investigated the use of â-CD and
sulfated-â-cyclodextrin (S-â-CD) with buffer additives of tetrabu-
tylammonium bromide (TBA) and methanol in an effort to
separate chiral sulfonium ions. Four series of chiral sulfonium
ions with varying alkyl chain lengths (Figure 1) were synthesized
from a common parent sulfide by alkylation with the appropriate
iodoalkane. The extent of enantiomeric resolution for each
racemic mixture was found to be dependent on the type of
cyclodextrin used, buffer additive, and the general structure of
the sulfonium ion. Additionally, the enantiomeric separation of
chiral thiophenium ions was also briefly explored using the
conditions established for the chiral sulfonium ions.
The most versatile chiral resolving agents used thus far in CZE
are the cyclodextrins (CDs). The most common CDs, R-, â-, or
γ-forms, are able to resolve chiral compounds by forming dif-
ferentiated, transient inclusion complexes with the two enanti-
omers dissolved in the separation buffer. The degree of the
interaction changes the migration time of the enantiomers enough
to achieve separation. Cyclodextrins have been used extensively
for the enantioseparation of drugs of pharmaceutical importance.2
Although â-CD appears to be the most commonly used chiral
selector, it is somewhat limited in the types of structural
compounds it can resolve. Further derivatization of the native
CDs can greatly enhance their selectivity and specificity. For
1-25
EXPERIMENTAL SECTION
Materials. â-Cyclodextrin, sulfated-â-cyclodextrin (degree of
substitution 7-11), acetonitrile, HPLC grade methanol, tetrabu-
tylammonium bromide, dichloroethane (DCE), all sulfides,
thiophenes, and iodoalkanes were purchased from Aldrich Chemi-
cal Co. (Milwaukee WI) and were used without further purifica-
tion. Silver tetrafluoroborate was purchased from Alfa Aesar
(Ward Hill, MA). Sodium phosphate buffer (0.1 M, pH 2.5), was
obtained from Bio-Rad (Hercules, CA). Dichloroethane was kept
(
(
17) Ariens, E. J. Drug Design I; Academic press, Inc: London, 1971: pp 77-81,
49-157.
18) Kuhn, R.; Hoffstetter, S. Capillary Electrophoresis: Principles and Practice;
Springer-Verlag: Berlin, Heidelberg, 1993; pp 261-273.
19) Novotny, M.; Soini, H.; Stefansson M. Anal. Chem. 1 9 9 4 , 66, 646A-655A.
20) Ward, T. J. Anal. Chem. 1 9 9 4 , 66, 633A-639A.
21) Quang, C.; Khaledi, M. G. Anal. Chem. 1 9 9 3 , 65, 3354-3358.
22) Nielen M. W. F. Anal. Chem. 1 9 9 3 , 65, 887-893.
23) Lurie, I. S.; Klein, R. F. X.; Cason, T. A. D.; LeBelle, M. J. Anal. Chem. 1 9 9 4 ,
1
(
(
(
(
(
6
6, 4019-4026.
(
(
24) Wang, Z.; Sun, Y.; Sun, Z. J. Chromatogr. A 1 9 9 6 , 735, 295-301.
25) Bjørnsdottir, I.; Hansen, S. H. Chirality 1 9 9 5 , 7, 219-225.
(26) Majors, R. E. LC-GC 1 9 9 7 , 15, 412-422.
Analytical Chemistry, Vol. 70, No. 17, September 1, 1998 3613

dry over 4-Å molecular sieves. HPLC grade water was used for
injection sample preparation.
chromophore, this group not only facilitates detection in the UV
range but also plays a central role in the currently accepted model
of inclusion interaction within the cyclodextrin cavity.2
8-31
All
Instrumentation. The CZE separations were accomplished
using a BioFocus 3000 capillary electrophoresis system from Bio-
Rad. An uncoated fused silica capillary column of 50 µm i.d., with
a total length of 50 cm (45.4 cm to detector window), mounted
on a user-assembled cartridge was used. Sample injection was
achieved using the pressure mode set at 2 psi‚s. The working
potential was operated at 17 kV (+ to -, constant voltage), and
the current limit was set at 100 µA. The cartridge and carousel
temperature was preset to 20 °C. The detection wavelength was
fixed at 220 nm. The capillary column was cleaned after each
run by flushing for 2 min with the running buffer, followed by
flushing for 5 min with deionized water. Prior to storage, the
column was purged with nitrogen for 2 min. It was necessary to
clean the electrodes every five consecutive runs to remove
adsorbed salts by dipping them in methanol for 2 min.
sulfonium salts are stable for analysis in an acidic buffer.
However, the thiophenium ions, particularly S-methylthiophenium,
are subject to nucleophilic attack by water molecules27 and thus
slowly revert back to the corresponding thiophene after addition
to the buffer. The instability did not present a problem in the
time frame of the analysis. It has been shown that a strong
interaction occurs between â-CD and dibenzylmethylsulfonium
tetrafluoroborate. Studies revealed that the transformations sul-
fonium ions undergo when subjected to alkaline conditions in the
absence or presence of â-CD are different. Dibenzylmethylsul-
fonium, for example, typically undergoes Stevens rearrangement
to give a 1,2-diphenylethane derivative, but in the presence of â-CD
it undergoes a Sommelet rearrangement to give a diphenyl-
methane derivative.32 As a result of this strong interaction, â-CD
was chosen as the resolving agent for chiral separation. The first
set of chiral separations attempted were carried out using standard
parameters: acidic pH (2.50) and an uncoated capillary with
varying concentrations of â-CD in 0.1 M phosphate buffer.
Sulfonium ions containing both a benzyl and a phenyl group were
initially investigated because these groups are believed to have
the strongest interaction within the â-CD cavity.32
Synthesis of Sulfonium Salts. A modification of the proce-
dure developed by Acheson and Harrison27 was used for the
preparation of the sulfonium and thiophenium salts. In this
procedure, 1 mmol of the appropriate sulfide or thiophene and
1
.2 mmol of the corresponding iodoalkane are dissolved in
approximately 2 mL of dry DCE in a 5-mL conical vial. A DCE
solution of silver tetrafluoroborate (1.2 mmol of AgBF dissolved
4
in 1 mL of DCE) is then added dropwise over a 5-min period.
The mixture is then stirred overnight. The yellow AgI precipitate
is centrifuged and washed twice with 1 mL of acetonitrile. The
solvents are then removed by rotary vaporization at 40 °C. The
resulting oil or crystal is dried under a vacuum overnight. After
the product is thoroughly dry, the structure is confirmed by NMR
Under acidic conditions, the electroosmotic flow (EOF) is
directed toward the cathode. EOF velocities in phosphate buffers,
from pH 2.00 to 8.00, have been well established by St. Pierre
and Sentell.33 They showed that, at low pH, the EOF velocity is
relatively low. The uncharged â-CD molecules move with the
velocity of the EOF and thus can be considered to form a
pseudostationary phase.21 The net mobility of positively charged
species, which also migrate toward the cathode, is thus the
summation of the electrophoretic mobility of the ions and the EOF.
It has been shown that a number of buffer additives, particularly
tetraalkylammonium (TAA) salts and organic solvents, alter the
polarity and the viscosity of the buffer system, resulting in a
lowering of the EOF.21,34, The alteration of the EOF thus affects
the net electrophoretic mobility of the analytes, often improving
resolution but at the expense of analysis time. Reports show that
some chiral separations are achieved only in the presence of these
modifiers. Short-chain tetraalkylammonium salts effectively sup-
press or even reverse the EOF in the low pH range.21,34 Quang
and Khaledi established EOF velocities in the presence of different
TAA salts. They found that a phosphate buffer at pH 2.5
containing 50 mM TBA showed a low magnitude reversal in
EOF.34 Under these conditions, the EOF counteracts the migra-
tion of the enantiomers, thus improving resolution. Figure 2A
shows the electropherogram of a mixture of ions 1 and 2 , from
series I, with 5 mM â-CD added to the phosphate buffer. In the
absence of TBA, incomplete resolution of these ions results.
Figure 2b shows the effect of adding 50 mM TBA for the
6
analysis using acetone-d as the solvent.
Sample and Buffer P reparation. Stock solutions of the
compounds were prepared by dissolving 10-20 mg of the
sulfonium salts in 1 mL of acetonitrile. The solutions were stored
at 4 °C. The buffers used were prepared by dissolving TBA, â-CD,
and S-â-CD, in the required amounts, directly into the standard
0
.10 M phosphate buffer (pH 2.5) or the 30% methanol (v/ v) 0.10
M phosphate buffer (pH 2.5). The 30% methanol/ phosphate
buffer required the addition of NaH PO to compensate for the
2
4
dilution of the 0.10 M phosphate buffer by the addition of methanol
and the final salt concentration was brought back up to 0.10 M.
All buffers made were filtered through a 0.45-µm membrane filter
(
Gelman Sciences) prior to use. The injection samples were
prepared by diluting the necessary aliquot of the stock solution
with a 50/ 50 (v/ v) water/ appropriate running buffer for each run
to yield a final concentration of 2.5 mM. Resolution of selected
ions was calculated by using the equation
R ) 2[(t - t )/ (w + w )]
(1)
s
2
1
2
1
where t is the migration time of the enantiomers and w is the
peak width.22
(
28) Szejtli, J. Cyclodextrins and their Inclusion Complexes; Akad e´ miai Kiad o´ :
Budapest, 1982; pp 95-100, 271.
RESULTS AND DISCUSSION
(29) Rogan M. M.; Altria, K. D.; Goodall, D. M. Chirality 1 9 9 4 , 6, 25-40.
(
(
(
30) Leli e` vre F.; Gareil, P. Anal. Chem. 1 9 9 7 , 69, 385-392.
31) Guttman, A.; Brunet, S.; Jurado, C.; Cooke, N. Chirality 1 9 9 5 , 7, 409-414.
32) Szejtli, J. Cyclodextrins and Their Inclusion Complexes; Akad e´ miai Kiad o´ :
Budapest, 1982; p 174.
The general structures for the synthesized series of sulfonium
ions and the two thiophenium ions are shown in Figure 1. For
convenience, all compounds synthesized included at least one
(
33) St. Pierre, L. A.; Sentell, K. B. J. Chromatogr. B 1 9 9 4 , 657, 291-300.
(
27) Acheson, R. M.; Harrison, D. R. J. Chem. Soc. 1 9 7 0 , 13, 1764-1776.
614 Analytical Chemistry, Vol. 70, No. 17, September 1, 1998
(34) Quang, C.; Khaledi G. J. Chromatogr. A 1 9 9 5 , 692, 253-265.
3

Fig u re 2 . Electropherograms showing the chiral separation of ions
1
and 2. Conditions: capillary, uncoated fused silica 50 µm i.d. × 50
cm (45.4 cm effective length); voltage, 17 kV; injection, 2 psi‚s; current
limit, 100 µA; temperature, 20 °C; detection, 220 nm; concentration
of sulfonium ions, 2.5 mM; buffer, 5 mM â-CD in 0.1 M sodium
phosphate buffer pH 2.5. Separation (A) without TBA and (B) with
50 mM TBA.
Fig u re 3 . Impact of â-CD concentration on the resolution of ion 2
holding TBA concentration constant at 50 mM, (A) Resolution vs
migration time, (B) migration time of the first-eluted enantiomer vs
â-CD concentration, and (C) effect of TBA concentration on resolution
on ion 2, plot of the resolution vs migration time, holding â-CD
concentration constant at 5 mM. Other conditions are as described
for Figure 2.
separation of the same sulfonium ions, 1 and 2 . The two sets of
enantiomers are clearly separated, and peak symmetry is dramati-
cally improved, as is the resolution of each pair of enantiomers.
The sulfonium ion containing the p-tolyl group yielded better
resolution, presumably due to a stronger interaction with the â-CD
cavity. As expected, an increase in migration time due to the
reversed EOF is also observed.
Effects of TBA and â-CD concentration. Several factors
make TBA a desirable additive. The low conductivity of the salt
allows the use of a wide range of concentrations, it increases the
solubility of â-CD and, the low hydrophobicity prevents micelle
formation as well as minimizes competitive effects with the
positively charged analytes for the â-CD cavity.21 The combined
effects of varying concentrations of the buffer additives were
explored. Figure 3A shows the effects of varying â-CD concentra-
tions, while holding TBA constant at 50 mM, on the resolution of
a single sulfonium ion, benzylmethyl-p-tolylsulfonium ion (2 ). No
separation was observed using a 1 mM â-CD concentration;
however, baseline resolution can be achieved easily with a â-CD
concentration as low as 5 mM. It is evident that the resolution
increases substantially as the â-CD concentration is increased.
The trade-off between higher resolution and analysis time is clearly
seen in Figure 3B, which shows the migration time of the first-
eluted enantiomer of ion 2 versus â-CD concentration. The
migration time of the enantiomer increases dramatically at higher
â-CD concentrations. Figure 3C shows the effects of varying TBA
concentrations, while holding â-CD constant at 5 mM, on the
resolution of the same ion. A gradual increase in resolution is
observed as the concentration of TBA increases as well as a small
corresponding increase in migration time.
Substituent Effects on Resolution. The first series of ions
studied, 1 -3 , differ only in the size of one of the aromatic groups.
Figure 4 shows the separation of all three ions from this series
using 5 mM â-CD and 50 mM TBA. These concentrations were
chosen to minimize analysis time while still affording good
resolution. Resolution increases in the order phenyl < p-tolyl <
naphthyl substituent. The enhanced resolution of ion 3 may be
attributed to a greater degree of interaction of the hydrophobic
naphthyl substituent with the â-CD cavity.
To further study substituent effects, series II, ions 4 -8 , were
synthesized. In this particular series, the aromatic substituents
are kept constant while the length of the alkyl chain is increased
from methyl to pentyl. Separations of these five ions are shown
in Figure 5. Figure 5A shows the separation carried out in the
absence of TBA with 5 mM â-CD. Although chiral resolution is
Analytical Chemistry, Vol. 70, No. 17, September 1, 1998 3615

Fig u re 4 . Electropherogram showing the chiral separation of series
I, ions 1-3, using 5 mM â-CD and 50 mM TBA. Other conditions are
as described for Figure 2.
achieved, peak tailing and only partial resolution for ion 5 is
observed. The addition of TBA, Figure 5B, results in improved
resolution of the same five ions as expected, as well as increasing
the migration time of each set of enantiomers by about 2 min.
A general trend in resolution can be observed under these
conditions: that is, as the alkyl chain lengthens, the resolution
decreases. The diminishing resolution can be attributed, in part,
to decreased interactions within the â-CD as the alkyl chain
lengthens. It is possible that the longer alkyl chain sterically
hinders the interaction between the aromatic groups with the
hydrophobic cavity of â-CD. In the absence of this steric
interaction, the resolution should increase as the length of the
alkyl group increases due to column residence time alone.
However, since resolution clearly decreases, it is concluded that
the alkyl chain is actively involved in the interaction.
Effect of Methanol. The general solubility of these particular
types of sulfonium ions is not very large in an aqueous buffer
system. The addition of organic solvents (10-80%) has been used
to enhance the solubility and also to improve CZE separations of
highly hydrophobic substances. Among the solvents used are
methanol, acetonitrile, 1-propanol, THF, ethanol, and 2-pro-
panol.35,36 The improved solubility of analytes in a binary buffer
system of this type significantly improves peak shape, thus
potentially enhancing resolution. An increase of the EOF at low
pH has been noted on previous studies when organic modifiers
of this type have been employed.37 The effects of methanol on
the chiral separation of the sulfonium ions was briefly explored
to determine whether an enhancement on the overall chiral
separation was, in fact, observed. Figure 5c shows the separation
of ions 4 -8 using a buffer containing 50 mM TBA, 5 mM â-CD,
Fig u re 5 . Electropherograms showing the chiral separation of series
II, ions 4-8, (A) with 5 mM â-CD, (B) 5 mM â-CD and 50 mM TBA,
and (C) with 5 mM â-CD, 50 mM TBA, and 30% methanol. Other
conditions are as described for Figure 2.
Use of Sulfated-â-Cyclodextrin. Series III, ions 9 -1 2 , and
series IV, ions 1 3 -1 7 , were synthesized next to study the extent
of interaction of a lone aromatic group and two alkyl substituents
with the â-CD cavity. Again, two of the groups were held constant,
aromatic and methyl substituents, while the second alkyl chain
increased from ethyl to pentyl. Surprisingly, no chiral separation
was achieved when using â-CD, even at high concentrations, >20
mM, in the absence or presence of TBA (data not shown). A
negatively charged cyclodextrin was postulated to have a greater
affinity for sulfonium ions, in general, due to the additional
electrostatic attraction of opposite charges. Sulfated-â-cyclodextrin
was obtained for evaluation as a potential chiral selector for these
particular ions. Figure 6A shows the separation of series III, ions
3
0% (v/ v) methanol, and 0.1 M phosphate (pH 2.5). It is obvious
from Figure 5C that chiral selectivity was completely lost by the
addition of methanol. Sz o¨ k _o et al. reported a similar effect: that
is, a lowering of binding constants, and thus resolution, was
observed for drug enantiomers as the result of increasing the
concentration of methanol.36 The enhanced solubility of the ions
9
-1 2 , with S-â-CD. An approximate concentration of 4 mM was
in the binary buffer system essentially eliminated the hydrophobic
used since the molecular mass is not accurately known due to
the nature of the product, a mixture of randomly substituted
molecules. It was necessary to keep the concentration relatively
low to avoid exceeding the current limit of 100 µA and to keep
Joule heating at a minimum. Although it is possible to work at
interaction within the â-CD cavity necessary for chiral separation.
(
(
35) Zhang, C.; Heeren, F. V.; Thorman, W. Anal. Chem. 1 9 9 5 , 67, 2070-2077.
36) Sz o¨ k _o , E.; Gyimesi, J.; Barcza, L.; Magyar, K. J. Chromatogr. A. 1 9 9 6 , 745,
1
81-187.
(
37) Fujiwara, S.; Honda, S. Anal. Chem. 1 9 8 7 , 59, 487-490.
3616 Analytical Chemistry, Vol. 70, No. 17, September 1, 1998

Fig u re 6 . Electropherograms showing the chiral separation of series
III, ions 9-12, using (A) 4 mM S-â-CD and (B) 4 mM S-â-CD and 50
mM TBA. Other conditions are as described for Figure 2.
Fig u re 7 . Electropherograms showing the chiral separation of series
IV, ions 13-16, using (A) 4 mM S-â-CD and (B) 4 mM S-â-CD and
50 mM TBA. Other conditions are as described for Figure 2.
higher concentrations of S-â-CD by switching to reversed polarity
3
8,39
or reversed mode,
it was deemed unnecessary for this study
since, even at this low concentration of 4 mM, S-â-CD was able
to adequately resolve each pair of enantiomers. Although some
peak tailing is observed, the overall resolution is acceptable. No
obvious trend is observed for this series. The resolution decreases
in going from ion 9 to 1 0 , and then it increases, with ion 1 2
having the greatest resolution, as shown in Figures 6A and 8. Ion
1
0 , with the propyl substituent, showed the lowest resolution. It
Fig u re 8 . Plot of the resolution vs alkyl chain length for series III
(b) and series VI (2) using the conditions described for Figures 7
and 8.
possible that the overall diameter of the structure imparts a similar
degree of interaction for the two enantiomers with S-â-CD or that
there is a reversal in binding constants when going from propyl
to butyl substituent. The lack of nonracemic standards prohibited
experimental verification of this hypothesis.
The overall separation looks similar to that of series III, in that
there is a minimum in resolution when the alkyl group is propyl
TBA was tested to determine whether any improvement is
observed upon its addition. Figure 6B shows the separation of
the same ions in the presence of 50 mM TBA. The effect was
detrimental to the chiral analysis for ions containing the smaller
ethyl and propyl substituents. As shown, complete loss of
resolution was observed for ions 9 and 1 0 . The observed results
can, perhaps, be caused by competitive effects between the
positively charged TBA and the sulfonium ions, having small alkyl
substituents, for the negatively charged S-â-CD cavity due to the
strong electrostatic attraction. Ions 1 1 and 1 2 were relatively
unaffected in terms of resolution.
(
ion 1 4 ), as shown in Figures 7A and 8. In fact, there is no chiral
resolution for ion 1 4 at all. This result may be explained by the
enantiomers of ions 1 4 having virtually the same degree of
interaction with the S-â-CD. Ion 1 4 was not resolved under any
of the conditions studied thus far. Figure 7B shows the effect of
adding 50 mM TBA to the same separation buffer. The resolution
remained virtually the same for the analytes aside from ion 1 3 ,
having the small ethyl substituent, whereby resolution decreased.
All of the sulfonium ions 1 -1 6 possess a chiral sulfur center.
The successful resolution of all but one of these sulfonium ion
suggested that a sulfonium ions possessing two chiral centers
might be resolved into four stereoisomers. To test this hypothesis,
ion 1 7 was synthesized to contain two adjacent chiral centers, an
additional chiral carbon present on the sec-butyl group. If there
is a strong interaction at a chiral carbon, the S-â-CD should be
Figure 7A shows the separation of series IV with 4 mM S-â-
CD, with the exception of ion 17, which was analyzed individually.
(
(
38) Gahm, K. H.; Stalcup, A. M. Chirality 1 9 9 6 , 8, 316-324.
39) Stalcup, A. M.; Gahm, K. H. Anal. Chem. 1 9 9 6 , 68, 1360-1368.
Analytical Chemistry, Vol. 70, No. 17, September 1, 1998 3617

Fig u re 1 0 . Electropherogram showing the chiral separation for the
two thiophenium ions, 18 and 19, using 4 mM S-â-CD. Other
conditions are as described for Figure 2.
established for the chiral separation of sulfonium ions are also
applicable. No chiral separation was observed for these ions when
using â-CD-modified buffers. However, ion 1 8 was baseline
resolved when using 4 mM S-â-CD-modified buffer, as shown of
Figure 10. Ion 1 9 was not resolved under any of the conditions
studied. We are currently investigating the use of different
cyclodextrin derivatives which may prove to be better candidates
for chiral separation of thiophenium ions.
Fig u re 9 . Electropherograms of the chiral separation of ion 17
showing all four enantiomer peaks, (R,R), (S,S), (R,S), and (S,R),
not necessarily in that order, using (A) 4 mM S-â-CD and (B) 4 mM
S-â-CD and 50 mM TBA. Other conditions are as described for Figure
CONCLUSION
The chiral analysis of a variety sulfonium and thiophenium ions
was successful using cyclodextrin/ TBA-modified capillary zone
electrophoresis employing uncoated fused silica capillaries. Suc-
cessful chiral resolution of the series of ions analyzed was found
to be dependent on the type of substituents on the sulfonium ions,
the type of cyclodextrin used, and the presence of TBA. The
established parameters can be easily optimized further to obtain
baseline resolution of particular ions of interest. The reproduc-
ibility of migration time was found to yield a %RSD of 0.37, based
on six replicate injections. In conclusion, the use of cyclodextrin-
modified CZE for the analysis of sulfonium ions offers an effective,
inexpensive, and rapid method of analysis.
2
.
able to recognize and resolve all four stereoisomers: (R,R), (S,S),
R,S), and (S,R). Figure 9A shows the separation of ion 1 7 using
mM S-â-CD. Although the four stereoisomers are separated,
(
4
they are not well resolved. The addition of 50 mM TBA improved
resolution considerably, as shown Figure 9B.
It is important to note that, even though S-â-CD was able to
effectively resolve sulfonium ions containing only one aromatic
substituent, it failed to resolve the first two series of ions
containing two aromatic substituents. Trial analysis showed that,
when a 4 mM S-â-CD modified buffer was used to analyze series
I and II, the ions failed to elute from the capillary within 1 h. The
peaks were so broad that no useful data could be obtained. When
a very low concentration of the S-â-CD was employed, <1 mM,
peaks were seen at a reasonable time, but they were again broad
and showed no chiral resolution (data not shown). This result
suggests that the interaction between these ions and the S-â-CD
is sufficiently strong that they are detained significantly by the
oppositely migrating S-â-CD, thereby limiting chiral separation.
Thiophenium Ion Analysis. Two chiral thiophenium ions,
ACKNOWLEDGMENT
The authors acknowledge the NSF-ILI award DUB-9650340 for
the acquisition of the CE system used in this work. T.K.G.
acknowledges the support of the American Chemical Society PRF
Grant 29413-B1,2.
Received for review December 29, 1997. Accepted July 1,
1998.
1
8 and 1 9 , were synthesized to determine if the conditions
AC971379J
3618 Analytical Chemistry, Vol. 70, No. 17, September 1, 1998