Water-soluble cavitands: synthesis of methylene-bridged resorcin[4]arenes containing hydroxyls and phosphates at their feet and bromomethyls and thiomethyls at their rims

Article abstract of DOI:10.1021/jo980305k

The synthesis of rim-functionalized methylene-bridged resorcin[4]arenes ("cavitands") containing hydrophilic propanol or water-solublilizing propylphosphate feet is described. The cavitands possess the synthetically useful benzylthiol (cavitands 6 and 16)

Full text of DOI:10.1021/jo980305k

6824
J . Org. Chem. 1998, 63, 6824-6829
Wa ter -Solu ble Ca vita n d s: Syn th esis of Meth ylen e-Br id ged
Resor cin [4]a r en es Con ta in in g Hyd r oxyls a n d P h osp h a tes a t Th eir
F eet a n d Br om om eth yls a n d Th iom eth yls a t Th eir Rim s
Adam R. Mezo and J ohn C. Sherman*
Department of Chemistry, University of British Columbia, 2036 Main Mall,
Vancouver, British Columbia, Canada, V6T 1Z1
Received February 18, 1998
The synthesis of rim-functionalized methylene-bridged resorcin[4]arenes (“cavitands”) containing
hydrophilic propanol or water-solublilizing propylphosphate feet is described. The cavitands possess
the synthetically useful benzylthiol (cavitands 6 and 16) or benzylbromide (cavitands 9 and 11)
functionalities at their rims, which are suitable for further derivatization near the hydrophobic
cavity of the cavitand. These water-soluble cavitands represent new building blocks that are ideal
for use in aqueous supramolecular chemistry. As an example of their synthetic utility in
supramolecular studies, we have reacted phosphate-footed cavitands 11 and 16 with cysteine-
containing peptide 17 and chloroacetylated peptide 19, respectively, to afford the corresponding de
novo proteins 18 and 20.
In tr od u ction
so-called “feet” in structure 1) which limits their solubility
to organic solvents and hence, their versatility as models
for supramolecular study. We recently reported the first
The study of supramolecular assemblies in water is an
important stepping stone toward understanding and
emulating the noncovalent interactions found in nature.1
The development of new supramolecular building blocks
that are soluble in water would be a great asset to this
area. Methylene-bridged resorcin[4]arenes,2 or cav-
itands, of the general structure 1 have been used
extensively in supramolecular assemblies because of their
rigidity, enforced cavities and synthetic availability.2e,3
They have been used to (1) synthesize carceplexes, where
chemical and physical properites of molecules can be
altered by encapsulation,3a,4 (2) bind neutral guest mol-
ecules,5 useful, for example, in the removal of toxins from
wastewater,5f and (3) form monolayers which have po-
tential as molecular sensors.6 However, these cavitands
have all incorporated a hydrophobic pendent group (R,
water-soluble cavitand,7 but only by incorporating water-
solubilizing groups at the rim positions (R′); this position
is vital for the incorporation of functional groups near
the cavity. To effectively break into aqueous cavitand
chemistry, it is essential to incorporate water-solubilizing
functionalities into the pendent group (R, cavitand 1)
while retaining useful functionalities at the rim position
(R′) suitable for further derivatization.
In 1989 Cram and co-workers introduced a method to
incorporate hydroxyl groups into the pendent groups
during the initial condensation step to form butanol-
footed, unbridged resorcin[4]arenes.8 More recently, our
group developed a method to selectively bridge the
phenolic hydroxyls to afford the first hydroxyl-footed
methylene-bridged cavitands.9 Here, we use this meth-
odology in conjunction with a series of orthogonal protec-
tion and functionalization steps to yield rim-functional-
(1) (a) For a review of cyclodextrins, see: Wenz, G. Angew. Chem.,
Int. Ed. Engl. 1994, 33, 803-822. (b) Diederich, F.; Dick, K.; Griebel,
D. Chem. Ber. 1985, 118, 3588-3619. (c) Kobayashi, K.; Asakawa, Y.;
Kato, Y.; Aoyama, Y. J . Am. Chem. Soc. 1992, 114, 10307-10313.
(2) For recent examples of unbridged resorcin[4]arenes, see: (a)
Lewis, P. T.; Davis, C. J .; Saraiva, M. C.; Treleaven, W. D.; McCarley,
T. D.; Strongin, R. M. J . Org. Chem. 1997, 62, 6110-6111. (b) Botta,
B.; Monache, G. D.; Salvatore, P.; Gasparrini, F.; Villani, C.; Botta,
M.; Corelli, F.; Tafi, A.; Gacs-Baitz, E.; Santini, A.; Carvalho, C. F.;
Misti, D. J . Org. Chem. 1997, 62, 932-938. (c) Fujimoto, T.; Shimizu,
C.; Hayashida, O.; Aoyama, Y. J . Am. Chem. Soc. 1997, 119, 6676-
6677. For recent reviews, see: (d) Schneider, H. J .; Schneider, U. J .
Inclusion Phenom. 1994, 19, 67-83. (e) Timmerman, P.; Verboom, W.;
Reinhoudt, D. N. Tetrahedron 1996, 52, 2663-2704.
(3) (a) Cram, D. J .; Cram, J . M. Container Molecules and Their
Guests, Monographs in Supramolecular Chemistry; Stoddart, J . F., Ed.;
Royal Society of Chemistry: Oxford, UK, 1994; Vol. 4.; (b) Moran, J .
R.; Karbach, S.; Cram, D. J . J . Am. Chem. Soc. 1982, 104, 5826-5828.
(4) (a) Sherman, J . C. Tetrahedron 1995, 51, 3395-3422. (b)
Sherman, J . C. In Large Ring Molecules; Semlyen, J . A., Ed.; J . Wiley
& Sons: West Sussex, England, 1996; pp 507-524. (c) Yoon, J .; Cram,
D. J . J . Chem. Soc. Chem. Commun. 1997, 497-498.
(6) (a) Schierbaum, K. D.; Weiss, T.; Thoden van Velzen, E. U.;
Engbersen, J . F. J .; Reinhoudt, D. N.; Go¨pel, W. Science 1994, 265,
1413-1415. (b) Thoden van Velzen, E. U.; Engbersen, J . F. J .; de
Lange, P. J .; Mahy, J . W. G.; Reinhoudt, D. N. J . Am. Chem. Soc. 1995,
117, 6853-6862.
(5) (a) Tucker, J . A.; Knobler, C. B.; Trueblood, K. N.; Cram, D. J .
J . Am. Chem. Soc. 1989, 111, 3688-3699. (b) Rudkevich, D. M.;
Hilmersson, G.; Rebek, J . J . Am. Chem. Soc. 1997, 119, 9911-9912.
(c) Careri, M.; Dalcanale, E.; Mangia, A.; Ruffini, M. Anal. Comm. 1997,
34, 13-15. (d) Vincetti, M.; Dalcanale, E. J . Chem. Soc., Perkin Trans.
2 1995, 1069-1076. (e) Boerrigter, H.; Verboom, W.; Reinhoudt, D. N.
J . Org. Chem. 1997, 62, 7148-7155. (f) Dalcanale, E.; Costantini, G.;
Soncini, P. J . Inclusion Phenom. 1992, 13, 87-92.
(7) Fraser, J . R.; Borecka, B.; Trotter, J .; Sherman, J . C. J . Org.
Chem. 1995, 60, 1207-1213.
(8) Tunstad, L. M.; Tucker, J . A.; Dalcanale, E.; Weiser, J .; Bryant,
J . A.; Sherman, J . C.; Helgeson, R. C.; Knobler, C. B.; Cram, D. J . J .
Org. Chem. 1989, 54, 1305-1312.
(9) Gibb, B.C.; Chapman, R. G.; Sherman, J . C. J . Org. Chem. 1996,
61, 1505-1509.
S0022-3263(98)00305-3 CCC: $15.00 © 1998 American Chemical Society
Published on Web 09/09/1998

Water-Soluble Cavitands
J . Org. Chem., Vol. 63, No. 20, 1998 6825
Sch em e 1
ized water-soluble cavitands. More specifically, we present
a method to introduce benzylbromides or benzylthiols at
the rim position (R′), while incorporating either hydroxyl
or phosphate groups into the feet of the cavitands.
Among the many potential water-solubilizing groups, we
chose phosphates because of their synthetic viability and
to incorporate as many charges as possible into the feet.
We are also interested in developing templates for the
organization of peptide structure. We recently linked
four 14-residue peptides to a cavitand containing methyl
feet to create a de novo protein based upon a hydrophobic
template.10 In this paper, we illustrate the synthetic
utility of phosphate-footed cavitands by reacting suitably
activated peptides with both phosphate-footed bromom-
ethyl and phosphate-footed thiomethyl cavitands, thus
forming a de novo protein linked to a hydrophilic tem-
plate.
thetically versatile aryl methyl group, which can lead to
various functional groups on the rim of the cavitand.11
In accord with the second criteria, 2,3-dihydrofuran was
used as the latent aldehyde to introduce short propanol
feet and thus aid in reducing the cavitand’s overall
hydrophobicity. The resulting acid-catalyzed condensa-
tion reaction of 2-methylresorcinol and 2,3-dihydrofuran
proceeded in 78% yield to form C4v-symmetric dodecol 2
(Scheme 1). Selective bridging9 of the phenolic hydroxyl
groups with CH2BrCl afforded the rigid methylene-
bridged 2-methyl cavitand 3 (42%).
Our first target was hydroxyl-footed benzylthiol 6
accessible via Sorrell’s bromination method.11 Initial
attempts to directly brominate hydroxyl-footed cavitand
3 failed due to poor solubility in the desired solvent CCl4.
Acetylation of the pendent hydroxyl groups with acetic
anhydride in pyridine afforded cavitand 4 (87%), which
was soluble in CCl4. Radical bromination with NBS and
benzoyl peroxide as an initiator afforded acetyl-protected
benzyl bromide 5 (68%). Treatment of cavitand 5 with
thiourea followed by base hydrolysis produced benzylthi-
ols and regenerated the hydroxyls in one step to form
hydroxyl-footed benzylthiol 6 (48%). This route to a
benzylthiol cavitand was found to be more efficient than
one previously developed by Cram.12
Resu lts a n d Discu ssion
Syn th esis. Two criteria were born in mind in design-
ing the water-soluble cavitands: (1) the rims of the
cavitands must be easily derivatized and (2) their feet
must be short and water-soluble. With respect to the first
criteria, we chose 2-methylresorcinol to exploit the syn-
(10) Gibb, B. C.; Mezo, A. R.; Sherman, J . C. Tetrahedron Lett. 1995,
36, 7587-7590.
(11) Sorrell, T. N.; Pigge, F. C. J . Org. Chem. 1993, 58, 784-785.

6826 J . Org. Chem., Vol. 63, No. 20, 1998
Mezo and Sherman
Sch em e 2
While planning the syntheses for both phosphate-
footed benzyl bromide 11 and phosphate-footed ben-
zylthiol 16, it was evident that positions R and R′
(structure 1) on the cavitand should possess orthogonal
reactivities. As base-sensitive benzyl bromides were one
target for position R′, we chose to use base-sensitive
protecting groups in position R′ and acid-sensitive groups
in position R. The chosen acid-sensitive protecting group
for the hydroxyl feet must be stable to NBS yet be
removable without affecting the acetal bridges of the
cavitand. Thus, cavitand 3 was treated with tert-butyl-
diphenylsilyl chloride (TBDPS-Cl) and imidazole to form
TBDPS-protected cavitand 7 (87%). Subsequent selective
bromination with NBS and benzoyl peroxide afforded
benzyl bromide 8 (54%). Removal of the TBDPS protect-
ing groups with HBr afforded hydroxyl-footed benzyl
bromide 9 (85%).
For the synthesis of cavitand 11, we chose to phospho-
rylate the hydroxyl feet with di-tert-butyl N,N-dieth-
ylphosphoramidite13 (DDP) followed by oxidation with
H2O2. The use of DDP has four main advantages: DDP
(1) eliminates the need to generate alkoxides in base-
sensitive compounds such as cavitand 9, (2) provides acid-
labile phosphate protecting groups consistent with our
protection strategy, (3) generates phosphite esters in high
yields, and (4) is quite stable, thus allowing convenient
storage. Phosphorylation of cavitand 9 with DDP fol-
lowed by oxidation with hydrogen peroxide at -78 °C
afforded tert-butyl-protected phosphate 10. Subsequent
removal of the tert-butyl groups with TFA afforded
phosphate-footed benzyl bromide 11. Unfortunately, the
phosphorylation did not proceed to completion (most
likely a function of the purity of the starting material
available only in technical grade),13 and resulted in
residual amounts of the corresponding tris-phosphate
impurity, which were evident in the 1H NMR and LSIMS
mass spectra. Attempts to further purify the compound
before and after removal of the tert-butyl groups were
unsuccessful. We hoped that subsequent derivatization
would facilitate purification. Indeed, in the pursuit of
de novo proteins, reaction of impure benzyl bromide 11
with cysteine-containing peptide 17 in pH 8.5 phosphate
buffer gave a mixture of products that was separable by
reversed phase HPLC to afford de novo protein 18
(Scheme 2).14
by treatment of benzyl bromide 8 with thioacetic acid and
diisopropylethylamine to afford S-acetyl cavitand 12
(64%). Removal of the TBDPS groups with HCl afforded
hydroxyl-footed S-acetyl cavitand 13 (93%). Phosphory-
lation of the hydroxyl groups with DDP followed by
oxidation with H2O2 yielded S-acetyl tert-butyl-protected
phosphate 14 (79%). Removal of the tert-butyl groups
with TFA followed by removal of the acetyl groups with
NaOH afforded phosphate-footed tetrathiol 16 (93% for
2 steps). Both phosphate-footed cavitands 11 and 16
have eight potential negative charges and are thus
soluble in water (pH 8). As an example of the synthetic
utility of cavitand 16, we reacted tetrathiol 16 with
chloroacetylated peptide 19 in phosphate buffer (pH 8.5)
to afford de novo protein 20 (Scheme 2).14
Con clu sion
We have developed synthetic pathways for both hy-
droxyl-footed and phosphate-footed cavitands that are
functionalized at their rims. These cavitands possess
either electrophilic benzyl bromides or nucleophilic ben-
zylthiols at their rims which are suitable for further
manipulation. These rigid, water-soluble cavitands open
the door to a variety of new areas of cavitand chemistry
ideal for supramolecular study. A number of studies are
currently underway in our labs, including the investiga-
tion of water-soluble carceplexes and hemicarceplexes.15
Another program of research in our lab entails the use
of cavitands as templates for the organization of peptide
structure.10,16 We have shown in this paper that both
benzyl bromide 11 and benzylthiol 16, when reacted with
suitably activated peptides, provide four peptides that
are linked by highly charged templates. Studies on the
structure and stability of these de novo proteins are
currently underway. We are also extending this meth-
En route to phosphate-footed benzylthiol 16, it was
necessary to introduce a protected thiol from TBDPS-
protected benzyl bromide 8; the S-protecting group must
be stable to the acidic conditions required to remove the
TBDPS group (HCl), the mildly basic conditions required
for phosphorylation (1H-tetrazole), the oxidatitive condi-
tions to generate the phosphate (H2O2, -78 °C), and must
be removable in the presence of a phosphate. To this end,
the S-acetyl protecting group was chosen and introduced
(12) Cram, D. J .; Karbach, S.; Kim, Y. H.; Baczynshkyj, L.; Marti,
K.; Sampson, R. M.; Kalleymeyn, G. W. J . Am. Chem. Soc. 1988, 110,
2554-2560.
(13) Perich, J . W.; J ohns, R. B. Synth. Commun. 1988, 142-144.
Attempts to purify DDP were unsuccessful.
(14) The amino acid sequences of peptides 17 and 19 were designed
such that each peptide would have a high propensity to form an
amphiphilic R-helix and bundle with the other helices attached to the
cavitand (ref 10). In addition, peptide 17 included a N-terminal cysteine
for reaction with benzyl bromide 11 while peptide 19 included a
N-terminal chloroacetyl group for reaction with benzylthiol 16. The
structures and stabilities of de novo proteins 18 and 20 will be reported
elsewhere. Standard one letter code abbreviations have been used for
the amino acids: C ) cysteine; E ) glutamic acid; G ) glycine; K )
lysine; L ) leucine.
(15) The first water-soluble hemicarceplex was recently reported;4c
however, its aqueous solubility was derived from its inter-bowl
linkages. Such a method to attain water solubility greatly limits the
variety of potential inter-bowl bridges and thus limits the cavity size.
By attaining water solubility via the cavitand’s pendent groups,
carceplexes and hemicarceplexes are far more versatile candidates for
supramolecular study and potential drug delivery systems.
(16) Gibb, B. C.; Mezo, A. R.; Causton, A. S.; Fraser, J . R.; Tsai, F.
C. S.; Sherman, J . C. Tetrahedron 51, 8719-8732.

Water-Soluble Cavitands
J . Org. Chem., Vol. 63, No. 20, 1998 6827
column chromatography (CHCl3:MeOH, 96:4) to afford acety-
lated 2-methyl cavitand 4 as a white solid (0.76 g, 87%): mp
131 °C (dec.); 1H NMR (CDCl3) δ 6.93 (s, 4H), 5.86 (d, J ) 6.9
Hz, 4H), 4.82 (t, J ) 9.1 Hz, 4H), 4.23 (d, J ) 6.9 Hz, 4H),
4.15 (t, J ) 6.5 Hz, 8H), 2.26 (m, 8H), 2.03 (s, 12H), 1.95 (s,
12H), 1.69 (m, 8H); MS (TG) m/z 993 ((M + H)+, 100). Anal.
Calcd for C56H64O16: C, 67.73; H, 6.50. Found: C, 67.65; H,
6.30.
odology to study the aqueous binding of neutral organic
molecules by phosphate-footed cavitands. Cram and co-
workers have shown that methyl-footed cavitands bind
molecules such as CH2Cl2 in a 1:1 stoichiometry in
organic solvents.5a Moreover, we have demonstrated
binding of neutral guests in a 2:1 cavitand:guest stoichi-
ometry.17 Such complexation may indeed be far stronger
in aqueous solution.
Acetyla ted Ben zyl Br om id e 5. A solution of 2-methyl
acetylated cavitand 4 (0.74 g, 0.75 mmol), NBS (0.58 g, 3.3
mmol), and benzoyl peroxide (10 mg) in CCl4 (50 mL) was
refluxed for 16 h. The reaction mixture was then cooled to
room temperature and filtered, and the solvent was evaporated
in vacuo. The crude product was purified by column chroma-
tography (EtOAc:hexanes, 1:1) to afford benzyl bromide 5 as
a white solid (0.66 g, 68%): mp 136 °C (dec.); 1H NMR (CDCl3)
δ 7.09 (s, 4H), 6.01 (d, J ) 6.7 Hz, 4H), 4.82 (t, J ) 8.1 Hz),
4.55 (d, J ) 6.7, 4H), 4.39 (s, 8H), 4.15 (t, J ) 6.5 Hz, 8H),
2.28 (m, 8H), 2.05 (s, 12H), 1.67 (m, 8H); MS (TG) m/z 1229
((M - Br + H)+, 100), 1309 ((M + H)+, 80). Anal. Calcd for
Exp er im en ta l Section
Gen er a l. Chemicals were reagent grade (Aldrich) except
for di-tert-butyl N,N-diethylphosphoramidite (Toronto Re-
search Chemicals) which was technical grade and used without
further purification. THF was distilled under N2 from sodium
benzophenone ketyl. DMF and N,N-dimethylacetamide (DMA)
were dried over 4 Å molecular sieves. LSIMS were run in
positive mode (unless otherwise noted) and recorded using
thioglycerol (TG), m-nitrobenzyl alcohol (NBA), chloroform
(CHCl3), dimethyl sulfoxide (DMSO), or methanol (MeOH) as
the matrixes. MALDI-TOF mass spectra were run using 2,5-
dihydroxybenzoic acid (DHB) as the matrix. Melting points
are uncorrected. Silica gel (230-400 mesh, BDH) was used
for column chromatography, and silica gel glass-backed ana-
lytical plates (0.2 mm, Aldrich) were used for TLC with UV
detection. Size exclusion chromatography was performed
using Sephadex LH-20. All products were dried overnight at
room temperature and 0.1 Torr.
2-Meth yld od ecol (2). 2,3-Dihydrofuran (3.04 mL, 40.3
mmol) was added over 4 h with a syringe pump to a solution
of 2-methylresorcinol (5.00 g, 40.3 mmol) and concentrated HCl
(7.6 mL) in MeOH (30 mL) and stirred for 16 h under N2. The
reaction mixture was then heated to 50 °C for 5 days. The
precipitate was filtered, washed rigorously with H2O, and dried
in vacuo for 24 h. The solid was then suspended in THF,
sonicated, and filtered to afford dodecol 2 as an off-white solid
(6.1 g, 78%): mp > 250 °C; 1H NMR (DMSO-d6) δ 8.62 (s, 8H),
7.27 (s, 4H), 4.19 (t, J ) 7.8 Hz, 4H), 4.10 (br s, 4H), 3.44 (t,
J ) 6.7 Hz, 8H), 2.24 (m, 8H), 1.94 (s, 12H), 1.35 (m, 8H); MS
(TG + DMSO) m/z 776 ((M + H)+, 90), 717 ((M - (CH2)3OH +
H)+, 100). Anal. Calcd for C44H56O12‚H2O: C, 66.48; H, 7.35.
Found: C, 66.71; H, 7.28.
C
56H60Br4O16: C, 51.40; H, 4.62. Found: C, 51.60; H, 4.42.
Hyd r oxyl-F ooted Ben zylth iol 6. Thiourea (5.1 mg, 0.072
mmol) was added to a solution of bromocavitand 5 (20 mg,
0.015 mmol) in degassed DMF (2 mL), and the mixture was
stirred for 2 h. The reaction mixture was poured onto degassed
2 M NaOH (2 mL) and stirred for 1 h. The reaction mixture
was evaporated in vacuo, dissolved in water, and acidified with
5% acetic acid. The precipitate was filtered, thoroughly
washed with water, dissolved in CHCl3:MeOH (9:1), dried over
MgSO4, and evaporated in vacuo. The crude product was
purified by column chromatography (9:1, CHCl3:MeOH) to
afford tetrathiol 6 as a white solid (7.0 mg, 48%): mp > 250
°C; 1H NMR (DMSO-d6) δ 7.56 (s, 4H), 5.95 (d, J ) 7.5 Hz,
4H), 4.59 (t, J ) 7.9 Hz, 4H), 4.43 (m, 8H), 3.47 (m, 16H), 2.71
(t, J ) 7.8 Hz, 4H), 2.38 (m, 8H), 1.42 (m, 8H); MS (LSIMS-,
NBA) m/z 917 ((M - SH - H)-, 100), 951 ((M - H)-, 50). Anal.
Calcd for C48H56O12S4‚H2O: C, 59.36; H, 6.02. Found: C,
59.43; H, 5.73.
TBDP S-P r otected 2-Meth yl Ca vita n d 7. TBDPS-Cl
(6.06 mL, 23.3 mmol) was added to a solution of cavitand 3
(2.40 g, 2.91 mmol) and imidazole (3.16 g, 46.6 mmol) in DMF
(20 mL) and stirred overnight under N2 at room temperature.
The DMF was then evaporated in vacuo, CHCl3 (20 mL) was
added, and the organic layer was washed with water (3 × 10
mL), dried over MgSO4, and evaporated in vacuo. The crude
product was purified by column chromatography (9:1, hexanes:
EtOAc) to afford cavitand 7 as a white foam (4.62 g, 87%): 1H
NMR (CDCl3) δ 7.60 (m, 16H), 7.28 (m, 24H), 6.91 (s, 4H), 5.85
(d, J ) 6.9 Hz, 4H), 4.77 (t, J ) 8.2 Hz, 4H), 4.23 (d, J ) 6.9
Hz, 4H), 3.66 (t, J ) 6.4 Hz, 8H), 2.22 (m, 8H), 1.95 (s, 12H),
1.59 (m, 8H), 0.99 (s, 36H); MS (LSIMS-, NBA + TG + CHCl3)
m/z 1824 ((M - H)-, 40), 1931 ((M‚thioglycerol - H)-, 100).
Anal. Calcd for C112H128O12Si4: C, 75.64; H, 7.25. Found: C,
75.49; H, 7.12.
2-Meth yl Ca vita n d 3. Dodecol 2 (20.0 g, 25.8 mmol) was
dissolved in degassed DMA (100 mL) and added via a syringe
pump over 48 h under N2 at room temperature to a solution
of DMA (700 mL), bromochloromethane (7.50 mL, 113 mmol),
and potassium carbonate (46.3 g, 335 mmol). After an ad-
ditional 24 h at room temperature, CH2BrCl (7.50 mL, 113
mmol) was added, and the reaction mixture was heated to 45
°C. An additional aliquot of CH2BrCl (7.50 mL, 113 mmol)
was added the next day, and the reaction mixture was heated
to 65 °C. After another 24 h at 65 °C, the reaction mixture
was evaporated in vacuo followed by careful neutralization of
the carbonate salts with 2 M HCl. The crude precipitate was
filtered, washed with water until the filtrate was neutral,
dissolved in THF, and dried over MgSO4, and the solvent was
evaporated in vacuo. The solid was purified by column
chromatography (CHCl3:MeOH, 9:1) to afford 2-methyl cav-
TBDP S-P r otected Ben zyl Br om id e 8. NBS (2.93 g, 16.5
mmol) and benzoyl peroxide (0.10 g, 0.41 mmol) were added
to a solution of cavitand 7 (6.83 g, 3.74 mmol) in CCl4. The
solution was refluxed for 18 h under N2 at which point the
reaction mixture was cooled to room temperature and filtered.
The filtrate was evaporated in vacuo, and the crude residue
was purified by column chromatography (hexanes:EtOAc, 95:
5; followed by 92:8) to afford benzyl bromide 8 as a white foam
(2.41 g, 54%): 1H NMR (CDCl3) δ 7.59 (m, 16H), 7.30 (m, 24H),
7.06 (s, 4H), 6.00 (d, J ) 6.2 Hz, 4H), 4.78 (t, J ) 7.9 Hz, 4H),
4.54 (d, J ) 6.2 Hz, 4H), 4.39 (s, 8H), 3.65 (t, J ) 6.3 Hz, 8H),
2.22 (m, 8H), 1.56 (m, 8H), 0.99 (s, 36H); MS (LSIMS-, NBA
1
itand 3 as an off-white solid (8.84 g, 42%): mp > 250 °C; H
NMR (DMSO-d6) δ 7.43 (s, 4H), 5.86 (d, J ) 7.4 Hz, 4H), 4.59
(t, J ) 6.1 Hz, 4H), 4.43 (t, J ) 5.1 Hz, 4H), 4.19 (d, J ) 7.4
Hz, 4H), 3.48 (m, 8H), 2.35 (m, 8H), 1.88 (s, 12H), 1.42 (m,
8H); MS (TG) m/z 825 ((M + H)+, 100). Anal. Calcd for
C
48H56O12‚H2O: C, 68.39; H, 6.93. Found: C, 68.07; H, 6.82.
Acetyla ted 2-Meth yl Ca vita n d 4. 2-Methyl cavitand 3
+ CHCl3) m/z 2093 ((M - H)-, 100). Anal. Calcd for C112H124
Br4O12Si4: C, 64.24; H, 5.97. Found: C, 64.64; H, 5.96.
-
(0.72 g, 0.87 mmol) was dissolved in pyridine (8 mL) and acetic
anhydride (8 mL) and stirred overnight under N2. The solvent
was removed in vacuo, and the crude product purified by
Hyd r oxyl-F ooted Ben zyl Br om id e 9. HBr (48%, 420 µL,
2.49 mmol) was added to a solution of tetrabromocavitand 8
(80 mg, 0.038 mmol) in THF:MeOH (4:1, 4 mL), and the
reaction was stirred for 4 h. The reaction mixture was then
evaporated in vacuo and purified by column chromatography
(9:1, CHCl3:MeOH) to afford hydroxyl-footed cavitand 9 as a
white solid (37 mg, 85%): mp > 250 °C; 1H NMR (acetone-d6)
(17) Chapman, B. C.; Sherman, J . C. J . Am. Chem. Soc. 1995, 117,
9081-9082.
(18) The 1H NMR signals from the tris-phosphate appeared as broad
signals under those of the tetraphosphate.
(19) Rink, H. Tetrahedron Lett. 1987, 28, 3787-3788.

6828 J . Org. Chem., Vol. 63, No. 20, 1998
Mezo and Sherman
δ 8.02 (s, 4H), 6.03 (d, J ) 7.5 Hz, 4H), 4.73 (t, J ) 8.2 Hz,
4H), 4.55 (s, 8H), 4.49 (d, J ) 7.5 Hz, 4H), 3.96 (t, J ) 5.9 Hz,
4H), 3.64 (dt, J ) 5.9 Hz, J ) 5.9 Hz, 8H), 2.70 (m, 8H), 1.51
(m, 8H); MS (LSIMS-, TG + CHCl3) m/z 1140 ((M)-, 60), 1219
((M‚Br)-, 100). Anal. Calcd for C48H52Br4O12‚H2O: C, 49.91;
H, 4.72. Found: C, 50.17; H, 4.62.
tert-butyl diethylphosphoramidite (0.25 mL, 0.89 mmol), and
the resulting mixture stirred for 10 min under N2. The
reaction mixture was then cooled to -78 °C, at which point
H2O2 (0.12 mL, 1.1 mmol) was added and the reaction mixture
was allowed to warm to room temperature over 30 min. The
reaction mixture was then poured onto H2O and extracted
three times with CHCl3. The combined CHCl3 extracts were
then washed with 10% NaHSO3 and H2O, dried over MgSO4,
and evaporated. The residue was purified by size exclusion
chromatography (EtOAc:MeOH:H2O; 40:10:4) followed by col-
umn chromatography (CHCl3:MeOH; 96:4) to afford tert-butyl-
protected S-Ac phosphate 14 as a white solid (0.13 g, 79%):
ter t-Bu tyl P h osp h or yla ted Ben zyl Br om id e 10. 1H-
Tetrazole (46 mg, 0.66 mmol) was added to a THF (10 mL)
solution of hydroxyl-footed benzyl bromide cavitand 9 (25 mg,
0.022 mmol) and di-tert-butyl diethylphosphoramidite (61 µL,
0.22 mmol), and the mixture was stirred for 15 min at room
temperature. The reaction mixture was then cooled to -78
°C, H2O2 (30 µL, 0.26 mmol) was added, and the reaction
mixture was allowed to warm to room temperature over 30
min. The reaction mixture was poured onto H2O, extracted
three times with CHCl3, dried over MgSO4, and evaporated
in vacuo. The crude product was purified by size exclusion
chromatography (EtOAc:MeOH:H2O, 40:10:4) followed by col-
umn chromatography (CHCl3:MeOH; 96:4) to afford a white
solid, which was a mixture of tetraphosphate 10 and ca. 15%
of the corresponding tris-phosphate derivative; tetraphosphate
10 was not purified further.18 For 10: 1H NMR (CDCl3) δ 7.15
(bs, 4H), 5.99 (d, J ) 6.5 Hz, 4H), 4.82 (t, J ) 7.5 Hz, 4H),
4.54 (d, J ) 6.5 Hz, 4H), 4.38 (s, 8H), 4.00 (m, 8H), 2.35 (bm,
8H), 1.67 (bm, 8H), 1.45 (s, 72H); 31P NMR (CDCl3) δ -9.97
(s, 4P); MS (NBA) m/z 1461 ((M - 8(C(CH3)3) + 9H)+, 100),
1798 (M - 2(C(CH3)3) + 9H)+, 20), 1855 (M - C(CH3)3 + 9H)+;
10); HRMS (NBA) calcd for (M - 8(C(CH3)3) + 9H)+ 1460.8910,
found 1460.8924.
1
mp 100 °C (dec); H NMR (CDCl3) δ 7.04 (s, 4H), 5.82 (d, J )
7.3 Hz, 4H), 4.76 (t, J ) 8.1 Hz, 4H), 4.25 (d, J ) 7.3 Hz, 4H),
4.00 (m, 16H), 2.29 (br m, 20H), 1.65 (m, 8H), 1.44 (s, 72H);
31P NMR (CDCl3) δ -9.96 (s, 4P); MS (NBA + CHCl3) m/z 1441
(M - (C(CH3)3)8 + 9H)+, 100). Anal. Calcd for C88H132O28
P4S4: C, 55.92; H, 7.04. Found: C, 56.10; H, 7.00.
-
S-Acetyl P h osp h a te-F ooted Ca vita n d 15. TFA (0.40
mL, 5.2 mmol) was added to a solution of cavitand 14 (46 mg,
0.024 mmol) in DCM, and the reaction mixture was stirred
for 10 min. The reaction mixture was evaporated in vacuo to
afford cavitand 15 as a white solid (34 mg, 98%): mp 100 °C
1
(dec); H NMR (methanol-d4) δ 7.35 (s, 4H), 5.82 (d, J ) 7.3
Hz, 4H), 4.75 (t, J ) 8.0 Hz, 4H), 4.32 (d, J ) 7.3 Hz, 4H),
4.10 (m, 16H), 2.44 (m, 8H), 2.05 (br s, 12H), 1.60 (m, 8H); 31
P
NMR (methanol-d4) δ 0.00 (s, 4P); MS (LSIMS-, NBA +
MeOH) m/z 1439 ((M - H)-, 100). Anal. Calcd for C56H68O28
P4S4‚3H2O: C, 44.98; H, 4.99. Found: C, 45.06; H, 4.72.
-
P h osp h a te-F ooted Ben zyl Br om id e 11. TFA (0.32 mL,
4.2 mmol) was added to a CH2Cl2 (4 mL) solution of impure
tetraphosphate 10 (25 mg of the mixture described above, ca.
0.01 mmol), and the mixture was stirred for 10 min. The
reaction mixture was evaporated in vacuo to afford a white
solid, which was a mixture of tetraphosphate-footed benzyl
bromide 11 and ca. 15% of the corresponding tris-phosphate
derivative (19 mg); tetraphosphate 11 was not purified fur-
ther.18 For 11: 1H NMR (CDCl3 + 20% acetone-d6) δ 7.36 (bs,
4H), 5.95 (d, J ) 7.3 Hz, 4H), 4.69 (bt, J ) 7.6 Hz, 4H), 4.43
(m, 12H), 4.15 (bm, 8H), 2.42 (bm, 8H), 1.59 (bm, 8H); 31P NMR
(CDCl3 + 20% acetone-d6) δ -0.53 (s, 4P); MS (LSIMS-, NBA
+ CHCl3 + MeOH) m/z 1379 ((M - PO3H2 - H)-, 60), 1459
((M - H)-, 100); HRMS (LSIMS-, NBA + CHCl3 + MeOH)
calcd for (M - H)- 1458.8754, found 1458.8765.
S-Acetyl TBDP S Ca vita n d 12. Thioacetic acid (219 µL,
3.08 mmol) was added to a solution of benzyl bromide 8 (1.50
g, 0.701 mmol) and diisopropylethylamine (535 µL, 3.08 mmol)
in DMF (20 mL). The reaction mixture was stirred for 16 h,
evaporated in vacuo, and purified by column chromatography
(hexanes:EtOAc, 3:1) to afford cavitand 12 as a white foam
(930 mg, 64%): 1H NMR (CDCl3) δ 7.59 (m, 16H), 7.30 (m,
24H), 6.96 (s, 4H), 5.88 (d, J ) 7.2 Hz, 4H), 4.73 (t, J ) 8.1
Hz, 4H), 4.26 (d, J ) 7.2 Hz, 4H), 4.01 (s, 8H), 3.64 (t, J ) 6.3
Hz, 8H), 2.31 (s, 12H), 2.19 (m, 8H), 1.55 (m, 8H), 0.99 (s, 36H);
MS (MALDI+, DHB) m/z 2097 (M‚Na+)+, 100), 2113 ((M‚K+)+,
90). Anal. Calcd for C120H136O16S4Si4: C, 69.46; H, 6.61.
Found: C, 69.60; H, 6.51.
P h osp h a te-F ooted Ben zylth iol 16. A 0.190 M degassed
solution of NaOH (1.39 mL, 0.264 mmol, 20 equiv) was added
to a degassed suspension of S-acetylated cavitand 15 (19 mg,
0.0132 mmol) in MeOH and H2O (1:1, 7 mL). The reaction
mixture was stirred for 30 min under N2 at which point
prewashed Amberlite ion-exchange resin (H+) was added via
a sidearm until the solution was no longer basic. The mixture
was stirred for 2 min and then cannulated to another flask so
as to leave the resin behind. The solution was evaporated in
vacuo to afford tetrathiol 16 as a white solid (16 mg, 95%):
1
mp 100 °C (dec.); H NMR (methanol-d4) δ 7.32 (s, 4H), 5.92
(d, J ) 7.3 Hz, 4H), 4.77 (t, J ) 8.3 Hz, 4H), 4.60 (d, J ) 7.3
Hz, 4H), 4.03 (dt, J ) 5.8 Hz, J ) 5.8 Hz, 8H), 3.61 (s, 8H),
2.45 (m, 8H), 1.64 (m, 8H); 31P NMR (methanol-d4) δ 3.24 (s,
4P); MS (LSIMS-, TG + MeOH) m/z 1271 (M - H)-, 100).
Anal. Calcd for C48H60O24P4S4‚4H2O: C, 42.86; H, 5.10.
Found: C, 42.53; H, 4.73.
P ep tid e Syn th esis. Peptides were synthesized on an
Applied Biosystems 431A peptide synthesizer using standard
Fmoc/t-Bu chemistry, HOBt (1-hydroxybenzotriazole)/HBTU
(2-(1H-benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluo-
rophosphate) activation of the amino acids, and Rink’s amide
resin19 to afford C-terminal amides. The last cycle in the
synthesis of peptide 17 entailed acetylation of the N-terminus
with acetic anhydride in NMP. Peptide 17 was then cleaved
from the resin by a 2 h treatment of a mixture of TFA (95%),
H2O (2.5%), and 1,2-ethanedithiol (2.5%). For peptide 19, the
last cycle included chloroacetylation of the N-terminus by
treatment with chloroacetyl chloride and DIEA as reported
previously.10 Peptide 19 was cleaved from the resin by a 2 h
treatment with 95% TFA. Each crude peptide was then
concentrated, precipitated with ice-cold ether, and filtered off.
Both peptides 17 and 19 were purified by reversed phase
HPLC using linear gradients of aqueous acetonitrile containing
0.1% TFA on a Phenomenex Selectosil C18 column (250 × 22.5
mm) at a flow rate of 10 mL/min. The peptides were lyophi-
lized and characterized by analytical HPLC and MS (LSIMS+,
TG). For peptide 17: m/z 1745 ((M + H)+, 100). For peptide
19: m/z 1814 ((M + H)+, 100).
S-Acetyl Hyd r oxyl-F ooted Ca vita n d 13. Concentrated
HCl (150 µL, 1.82 mmol) was added to a solution of S-Ac-
TBDPS cavitand 12 (100 mg, 0.0472 mmol) in THF:MeOH (4:
1, 5 mL). The reaction mixture was stirred for 4 h, neutralized
with saturated NaHCO3, and evaporated in vacuo. The crude
product was dissolved in CHCl3:MeOH (9:1, 10 mL), dried over
MgSO4, evaporated in vacuo, and purified by column chroma-
tography (9:1, CHCl3:MeOH) to afford cavitand 13 as a white
1
solid (49 mg, 93%): mp >250 °C; H NMR (DMSO-d6) δ 7.58
(s, 4H), 5.79 (d, J ) 7.7 Hz, 4H), 4.55 (t, J ) 8.1 Hz, 4H), 4.44
(br s, 4H), 4.27 (d, J ) 7.7 Hz, 4H), 3.94 (s, 8H), 3.47 (t, J )
6.5 Hz, 8H), 2.37 (m, 8H), 2.29 (s, 12H), 1.40 (m, 8H); MS
(MALDI+, DHB) m/z 1144 ((M + Na+)+, 90), 1160 ((M + K)+,
100). Anal. Calcd for C120H136O16S4‚H2O: C, 59.03; H, 5.84.
Found: C, 59.42; H, 5.72.
De Novo P r otein 18. Peptide 17 (20.4 mg, 11.2 µmol) and
impure cavitand 11 (3.7 mg, ca. 2.3 µmol) were stirred in
degassed 150 mM sodium phosphate pH 8.5 buffer (4 mL) for
1 h at room temperature under N2. The crude product was
purified twice by reversed phase HPLC (conditions as above).
The major peak eluted at ca. 45% acetonitrile, separable from
the corresponding tris-phosphate derivative, and was lyoph-
ter t-Bu tyl P h osp h or yla ted S-Acetyl Ca vita n d 14. 1H-
Tetrazole (0.19 g, 2.7 mmol) was added to a THF solution of
hydroxyl-footed S-Ac-cavitand 13 (0.10 g, 0.089 mmol) and di-

Water-Soluble Cavitands
J . Org. Chem., Vol. 63, No. 20, 1998 6829
ilized to afford a white solid (1.9 mg, 9%). Purity was assessed
by the observation of one peak by analytical HPLC, and its
identity was confirmed by electrospray MS to give a mass of
8394.6 ( 1.2 Da [calcd 8393.7 Da (average isotope composi-
tion)].
De Novo P r otein 20. Peptide 19 (20.0 mg, 11.5 µmol) and
cavitand 16 (3.6 mg, 2.9 µmol) were stirred in a degassed 150
mM sodium phosphate pH 8.5 buffer (4 mL) for 4 h at room
temperature under N2. The crude product was purified twice
by reversed phase HPLC (conditions as above). The major
peak eluted at ca. 45% acetonitrile and was lyophilized to
afford a white solid (7.8 mg, 33%). Purity was assessed by
the observation of one peak by analytical HPLC, and its
identity was confirmed by electrospray MS to give a mass of
8110.6 ( 1.2 Da [calcd 8109.4 Da (average isotope composi-
tion)].
Ack n ow led gm en t. We thank NSERC (Canada) for
financial support in the form of a grant to J .C.S. and a
post-graduate fellowship to A.R.M. Acknowledgment is
made to the donors of the Petroleum Research Fund,
administered by the ACS, for partial support of this
research. Warm thanks to Professor Bruce C. Gibb for
critical reading of this manuscript.
J O980305K