A tricatecholic receptor for carbohydrate recognition: Synthesis and binding studies

Article abstract of DOI:10.1021/jo0702286

(Chemical Equation Presented) A new tripodal receptor bearing three catechol subunits on a benzene platform has been synthesized in four steps from 1,3,5-triethylbenzene and pyrogallol. The binding ability of the tricatecholic receptor was investigated to

Full text of DOI:10.1021/jo0702286

studied.3 In this context, we recently reported a new tripodal
receptor 1 featuring ureidic H-bonding groups for the recognition
of saccharides, which showed affinities in the millimolar range
and moderate selectivities toward the octyl glycosides of a set
of monosaccharides in CDCl3.4
A Tricatecholic Receptor for Carbohydrate
Recognition: Synthesis and Binding Studies
Martina Cacciarini,†,§ Elisa Cordiano,† Cristina Nativi,†,§ and
Stefano Roelens*,‡,§
Dipartimento di Chimica Organica and Laboratorio di
Progettazione Sintesi e Studio di Eterocicli Biologicamente
AttiVi (HeteroBioLab), UniVersita` di Firenze, and Istituto di
Metodologie Chimiche (IMC), Consiglio Nazionale delle
Ricerche (CNR), Polo Scientifico e Tecnologico, Via della
Lastruccia 13, I-50019 Sesto Fiorentino, Firenze, Italy
stefano.roelens@unifi.it
ReceiVed February 2, 2007
In search for new receptors of improved recognition ability,
we thought that the ureidic groups on the side arms of 1 could
be conveniently replaced by catecholic ligands, in the belief
that acidic hydroxyls in a chelating arrangement would advan-
tageously substitute the ureidic NH functions as H-bonding
partners for glycosidic hydroxyls, still retaining the flexibility
of the tripodal architecture and the binding geometry of the
bidentate ligands, which was shown to provide the correct
geometry for binding monosaccharides.4 From a literature
search, it appeared that catechol ligands, widely found in cation
complexes and in natural and synthetic ionophores,5 are yet
unexplored as neutral H-bonding ligands for carbohydrates; thus,
tripodal catecholic receptors may be interesting new candidates.6
We wish to report the synthesis and carbohydrate binding
properties of a new tripodal benzene-based catecholic receptor,
showing preference for R-mannoside among octyl glycosides
of biologically relevant monosaccharides.
A new tripodal receptor bearing three catechol subunits on
a benzene platform has been synthesized in four steps from
1,3,5-triethylbenzene and pyrogallol. The binding ability of
the tricatecholic receptor was investigated toward several
monosaccharides in CDCl3, where multiple equilibria were
detected, and compared to that of a previously reported
trisureidic receptor of analogous structure. Association
1
constants were measured by H NMR titrations, and the
corresponding affinities were assessed through the BC50
parameter, a binding descriptor univocally defining the
affinity of a host for a guest in multi-equilibrium systems.
Results show that the tripodal catecholic receptor binds the
octyl glycosides with affinities ranging from 0.87 to 5.2 mM
and with a 6-fold selectivity factor for the R-mannoside over
the â-glucoside. Although the affinity for glycosides was not
appreciably improved with respect to the ureidic receptor, a
significant change in selectivity was obtained by the H-
bonding group replacement.
(1) (a) Lutzen, A. In Highlights in Bioorganic Chemistry; Schmuck, C.,
Wennemers, H., Eds.; Wiley-VCH: Weinheim, Germany, 2004; pp 109-
119. (b) Striegler, S. Curr. Org. Chem. 2003, 7, 81-102. (c) Dam, T. K.;
Brewer, C. F. Chem. ReV. 2002, 102, 387-429. (d) Mellet, C. O.; Defaye,
J.; Garc´ıa Ferna´ndez, J. M. Chem.sEur. J. 2002, 8, 1982-1990. (e) Host-
Guest Chemistry. Mimetic Approaches to Study Carbohydrate Recognition.
In Topics in Current Chemistry; Penade´s, S., Ed.; Springer-Verlag:
Heidelberg, 2002; Vol. 218. (f) Davis, A. P.; Wareham, R. S. Angew. Chem.,
Int. Ed. 1999, 38, 2978-2996. (g) James, T. D.; Sandanayake, K. R. A. S.;
Shinkai, S. Angew. Chem., Int. Ed. Engl. 1996, 35, 1910-1922. (h) Lee,
Y. C.; Lee, R. T. Acc. Chem. Res. 1995, 28, 321-327.
(2) (a) Zachara, N. E.; Hart, G. W. Chem. ReV. 2002, 102, 431-438.
(b) Lundquist, J. J.; Toone, E. J. Chem. ReV. 2002, 102, 555-578. (c) Ernst,
B.; Hart, W.; Sinay¨, P. Carbohydrates in Chemistry and Biology; Wiley-
VCH: Weinheim, Germany, 2000; Part I, Vol. 2 and Part II, Vol. 4. (d)
Lindhorst, T. K. Essentials of Carbohydrate Chemistry and Biochemistry;
Wiley-VCH: Weinheim, Germany, 2000. (e) Mammen, M.; Choi, S.-K.;
Whitesides, G. M. Angew. Chem., Int. Ed. 1998, 37, 2754-2794.
(3) For a recent comprehensive review on the subject, see: Davis, A.
P.; James, T. D. In Functional Synthetic Receptors; Schrader, T., Hamilton,
A. D., Eds.; Wiley-VCH: Weinheim, Germany, 2005; pp 45-109.
(4) Vacca, A.; Nativi, C.; Cacciarini, M.; Pergoli, R.; Roelens, S. J. Am.
Chem. Soc. 2004, 126, 16456-16465.
Molecular recognition of carbohydrates has been an actively
investigated area of research in the past few years1 because
selective recognition of saccharides plays a crucial role in
various biological processes, such as cell adhesion, regulation,
and growth.2 Structurally diverse synthetic receptors have been
employed to tackle the challenging task of understanding the
molecular basis of the recognition processes occurring in Nature,
among which neutral hosts designed to interact with mono- or
oligosaccharides through H-bonding have been by far the most
(5) See, for example: (a) Seeber, G.; Tiedemann, B. E. F.; Raymond,
K. N. Top. Curr. Chem. 2006, 265, 147-183. (b) Budzikiewicz, H. Mini-
ReV Org. Chem. 2004, 1, 163-168. (c) Albrecht, M. Chem. Soc. ReV 1998,
27, 281-288. (d) Pierpont, C. G.; Lange, C. W. Prog. Inorg. Chem. 1994,
41, 331-442. (e) Raymond, K. N. Pure Appl. Chem. 1994, 66, 773-781.
(f) Raymond, K. N.; McMurry, T. J.; Garrett, T. M. Pure Appl. Chem 1988,
60, 545-548.
* To whom correspondence should be addressed. Phone: +39-055-457-3546.
Fax: +39-055-457-3570.
† Dipartimento di Chimica Organica.
§ HeteroBioLab.
(6) A phenolic tripodal receptor is the closest example reported: Abe,
H.; Aoyagi, Y.; Inouye, M. Org. Lett. 2005, 7, 59-61.
‡ CNR-IMC.
10.1021/jo0702286 CCC: $37.00 © 2007 American Chemical Society
Published on Web 04/20/2007
J. Org. Chem. 2007, 72, 3933-3936
3933

SCHEME 1. Synthesis of the Tripodal Catecholic Receptor 7
CHART 1
were measured by 1H NMR titrations in CDCl3 at T ) 298 K.
Glucose (Glc), galactose (Gal), and mannose (Man) were
selected among the most frequently encountered monosaccha-
ridic epitopes, present as terminal R- or â-glycosides in more
complex oligo- or polysaccharides on cell surfaces and in
glycoconjugates.
Addition of increasing amounts of 7 to the glycosidic guests
induced a significant shift of the signals of both reagents, clearly
indicating binding phenomena under fast exchange, which were
particularly evident for catecholic and glycosidic hydroxyl
protons and for some of the glycosidic CH protons. The upfield
shift of the latter protons and the downfield shift of the former
protons (except for OH(A), see later), which could be monitored
if acidic impurities were carefully removed from the medium,
were consistent with the shielding effect of the aromatic ring
current and the deshielding effect from H-bonding, respectively,
in agreement with the formation of a host-guest complex in
which the glycoside is located in the cleft of the receptor. The
titration of âGal with 7 is reported in Figure 1 as a typical
example.
An in depth investigation of all the aspects involved in the
correct determination of binding affinities toward glycosides
by NMR titrations (reactant self-association, chemical model,
choice of signals, etc.) has been described in a previous paper.4
In the present study, we closely followed the established titration
protocol and the optimized analysis of data, which consisted of
diluting the receptor with a stock solution of the glycoside and
of the simultaneous fit of all the available signals of both
reagents to the appropriate model of chemical equilibria,
respectively. As with the ureidic receptor 1, experimental data
were not adequately fitted by a simple 1:1 association. Binding
of the receptor to the investigated glycosides required a model
including 1:1 and 2:1 association equilibria in all cases, while
no evidence of reagent self-association was found from dilution
experiments. The excellent agreement between experimental and
calculated shifts for all the detected signals can be easily
appreciated from the plots in Figure 1. The results obtained with
the glycosides of Chart 1 are reported in Table 1 as cumulative
log â values for the formation of the 1:1 and 2:1 receptor to
glycoside adducts. Because more than one complex species is
present in all cases, binding constants cannot be directly used
to assess the overall affinity of the receptor for each glycoside.
This can be conveniently determined using the BC50 parameter,4
a binding descriptor univocally defining the affinity of a host
for a guest in multi-equilibrium systems, which can be calculated
from the measured binding constants.8 The BC50 parameter is
defined as the total concentration of receptor necessary to bind
Synthesis. Catecholic substituents were conveniently intro-
duced into the tripodal scaffold by ether linkages through the
synthetic steps reported in Scheme 1.
1,3,5-Tris(bromomethyl)-2,4,6-triethylbenzene (4), prepared
from the commercially available 1,3,5-triethylbenzene 2,7 was
reacted with the protected pyrogallol derivative 5 to afford the
protected tricatecholic derivative 6. Protection as the orthoester
resulted in being the method of choice; the alternative protection
of pyrogallol as the acetonide proved not to be a viable
alternative, as subsequent deprotection failed under several
conditions. Attempts to protect pyrogallol by reacting 3 with
di-tert-butyldichlorosilane or ethyl propiolate failed, as well.
On the other hand, protection of the hydroxyl groups was
mandatory, as reacting 4 with unprotected pyrogallol consistently
gave untractable mixtures. Deprotection of 6 to the catecholic
receptor 7 was eventually accomplished by acidic cleavage.
Binding Studies. The binding affinities of receptor 7 toward
the octyl glycosides of monosaccharides depicted in Chart 1
(8) To expedite the calculation of BC50 and BC050, a utility program
based on the general treatment described in ref 4 has been developed and
made available for free upon request at the corresponding author’s e-mail
address.
(7) Walsdorff, C.; Saak, W.; Pohe, S. J. Chem. Res. (M) 1996, 1601-
1618.
3934 J. Org. Chem., Vol. 72, No. 10, 2007

TABLE 1. Cumulative Association Constants (log ân) with
Standard Deviations (σ) for 1:1 and 2:1 Adducts, Intrinsic Median
Binding Concentration (B C050), and Relative Affinities (RA) for
Complexes of Receptor 7 with Octyl Glycosidesa
glycoside
log â11
log â21
BC050 (mM)
RA
RMan
âMan
âGal
RGlc
RGal
âGlc
3.03 ( 0.04
2.59 ( 0.07
2.46 ( 0.03
2.39 ( 0.06
2.33 ( 0.04
2.25 ( 0.05
5.04 ( 0.04
4.42 ( 0.13
3.85 ( 0.21
3.94 ( 0.16
3.98 ( 0.08
3.48 ( 0.26
0.87 ( 0.07
2.2 ( 0.3
3.2 ( 0.2
3.6 ( 0.4
4.0 ( 0.3
5.2 ( 0.6
6.0
2.4
1.6
1.4
1.3
1.0
1
a Measured by H NMR (400 MHz) from titration experiments at T )
298 K in CDCl3 on 0.8-1.2 mM stock solutions of glycoside using receptor
concentrations up to 15 mM. Binding constants â11 (M-1) and â21 (M-2
)
were calculated by simultaneous nonlinear least-square fit of all the available
signals shifts. BC0 values were calculated from log âi values using the
50
BC50 Calculator program; see ref 8.
receptor is substantially insensitive to the described binding
group substitution, although their H-bonding properties were
expected to be quite different. In contrast, the selectivity trend
exhibited by 7 toward the investigated glycosides is significantly
different from that observed for the ureidic progenitor: the most
remarkable feature is that, while RMan is the least effectively
bound glycoside in the case of 1, it is the most effectively bound
to 7, showing a 6-fold preference factor over âGlc. Such an
inversion of selectivity, although not impressive in size, is
certainly noteworthy and is most likely related to a change in
binding geometry achieved by the catecholic receptor with
respect to the ureidic host. Unfortunately, a convincing inter-
pretation of the complex structure and geometry could not be
inferred from the chemical shift variations of the signals of both
reagents upon binding. The only clear indication seems to be
the systematic upfield shift contribution to the catecholic OH
protons, which for the hydroxyl closest to the aromatic platform
largely overrides the downfield shift due to H-bonding, indicat-
ing that a conformational rearrangement occurs upon binding
that orientates the catechol hydroxyl groups toward the interior
of the cleft, where they experience the ring current shielding
effect. Besides this evidence, it does not appear fortuitous that
the axial mannose hydroxyl is the only glycosidic OH signal
of the entire investigation shifting upfield upon complexation,
and that concomitantly, apart from the H-2 signal, the largest
upfield shifts observed are those of the axial H-4 proton of the
glycoside and the OH(A) proton of the catechol moiety,
suggesting that all four signals reside in the shielding cone of
the arene’s ring current. This is compatible with a complex’s
geometry in which the â face of the mannoside is facing the
scaffold’s arene moiety and in which the inner catecholic
hydroxyl lies in close proximity to the axial mannoside hydroxyl
and in a correct orientation for H-bonding. Whatever the
structural situation, experimental evidence demonstrates that the
selectivity of the tripodal receptor can be modulated by
appropriately changing the bidentate H-bonding groups, switch-
ing preference from âGal to RMan by replacing catecholic for
ureidic ligands.
FIGURE 1. Plot of complexation induced shifts versus receptor’s total
concentration in the titration of âGal 1.1 mM with 7 in CDCl3 at 400
MHz and T ) 298 K. Bottom: galactose OH signals (b OH-6; 9 OH-
2; 2 OH-3; ( OH-4). Middle: galactose CH signals (r CH-2; 1 CH-
7; f CH-7′; + CH-4; × CH-1). Top: receptor’s catechol OH signals
(b OH-B; 9 OH-A). Symbols are experimental data points; solid lines
are best fit curves obtained through nonlinear regression by simulta-
neous fit of all data.
50% of the ligand and is most conveniently employed as its
limiting value for dilute solutions, the intrinsic median binding
concentration BC050, which is independent of the concentration
of the ligand. BC050 values calculated from the binding con-
stants are reported in Table 1 for each glycoside, together with
the corresponding relative affinity derived from normalized B
C050 values.
The overall impression from results in Table 1 is that
replacement of the catecholic for the ureidic binding groups in
the tripodal scaffold did not bring a significant improvement in
the binding ability of receptor 1. Indeed, binding affinities
ranging from nearly 1 to 5 mM were observed for 7, very similar
to those obtained for 1 (1-6 mM),4 indicating that the tripodal
In conclusion, the development of structures bearing different
subunits on a benzene-based platform reveals the possibility of
modulating the binding selectivity toward different series of
monosaccharides depending on the nature of the binding groups.
Although a significant improvement in binding ability was not
achieved by replacing catecholic for ureidic binding groups in
the receptor’s side arms, this work demonstrated that the tripodal
catecholic receptor 7 is able to bind the set of octyl glycosides
J. Org. Chem, Vol. 72, No. 10, 2007 3935

7 (0.230 g, 0.399 mmol, 78%) as a white solid: Mp 178-180 °C;
1H NMR (CDCl3, 400 MHz) δ 6.86-6.82 (m, 3H), 6.72-6.66 (m,
6H), 5.37 (br s, 3H, OH(A)), 5.30 (br s, 3H, OH(B)), 5.15 (s, 6H),
2.86 (q, J ) 7.6 Hz, 6H), 1.25 (t, J ) 7.6 Hz, 9H); 13C NMR
(CDCl3, 50 MHz) δ 147.0, 146.3, 144.4, 132.7, 130.9, 119.9, 109.4,
103.9, 64.9, 22.9, 16.7; MS-EI m/z (%) 577(M+, 17), 448 (23),
326 (23), 199 (26). Anal. Calcd for C33H36O9: C, 68.74; H, 6.29.
Found: C, 68.96, H, 6.30.
in Chart 1 with millimolar affinities in CDCl3 and with a
significant selectivity for RMan over âGlc.
Experimental Section
1,3,5-Tris(2′-methoxy-1′,3′-benzodioxol-4′-oxymethyl)-2,4,6-
triethylbenzene (6): To a solution of 4 (1.81 g, 4.10 mmol) and 5
(2.09 g, 12.4 mmol) in acetone (120 mL) was added K2CO3 (5.30
g, 38.4 mmol), and the mixture was heated under reflux for 4 h.
The reaction mixture was cooled to room temperature, filtered, and
concentrated under vacuum to give 6 (1.74 g, 2.48 mmol, 61%) as
a white solid, which was recrystallized from methanol: Mp 92-
Acknowledgment. The authors wish to thank Prof. A.
Vacca, Universita` di Firenze, for helpful discussions. The
contribution of Dr. P. Gans, University of Leeds, and of Prof.
A. Sabatini, Universita` di Firenze, to the upgrade of the
HypNMR program is gratefully acknowledged. Ente Cassa di
Risparmio di Firenze (Italy) is acknowledged for granting a 400
MHz NMR spectrometer.
1
94 °C; H NMR (CDCl3, 200 MHz) δ 6.96-6.61 (m, 12H), 5.25
(s, 6H), 3.42 (s, 9H), 2.56 (q, J ) 7.4 Hz, 6H), 1.28 (t, J ) 7.4 Hz,
9H); 13C NMR (CDCl3, 50 MHz) δ 147.5, 146.6, 142.5, 134.5,
130.8, 122.0, 119.2, 109.9, 102.2, 66.1, 49.9, 23.1, 16.6. Anal. Calcd
for C39H42O12: C, 66.66; H, 6.02. Found: C, 66.52, H, 5.91.
1,3,5-Tris(2′,3′-dihydroxyphenoxymethyl)-2,4,6-triethylben-
zene (7): To a solution of 6 (0.360 g, 0.510 mmol) in CHCl3 (5
mL) was added p-toluenesulfonic acid (0.15%) as catalyst. The
reaction mixture was stirred at room temperature for 30 min, then
washed with saturated solution of NaHCO3, and dried over Na2-
SO4. Concentration of the solvent and purification of the crude by
flash column chromatography (silica gel, CHCl3/CH3OH 9:1) gave
Supporting Information Available: General methods and
materials. Synthetic procedure for compound 5. Titration procedure,
data analysis, and plots of experimental and calculated shifts for
the titration of RMan with 7. NMR spectral data. This material is
available free of charge via the Internet at http://pubs.acs.org.
JO0702286
3936 J. Org. Chem., Vol. 72, No. 10, 2007