A New Generation of "Cholaphanes": Steroid-Derived Macrocyclic Hosts with Enhanced Solubility and Controlled Flexibility

Article abstract of DOI:10.1021/jo971272w

The macrocyclic "cholaphanes" 3a - c were synthesized from the inexpensive steroid cholic acid. Like earlier relatives they feature substantial cavities with inward-directed hydroxyl groups, suitable for binding polar molecules such as carbohydrates in nonpolar media. New features are the externally directed alkyl chains, promoting solubility in organic solvents, and (in the case of 3b/c) reduced conformational freedom resulting from truncation of the steroidal side-chain. In particular, modeling shows that the smallest macrocycle 3c possesses very little flexibility, preferring an open conformation which is also revealed in the X-ray crystal structure of its pentahydrate. NMR studies indicated that all three cholaphanes form 1:1 complexes with octyl β-D-glucoside in CDCl₃, with K_a = 600-1560 M^-1. Cholaphanes 3b/c proved able to extract methyl β-D-glucoside from aqueous solutions into CHCl₃. The transport of methyl β-D-glucoside across a chloroform barrier was also demonstrated for 3c.

Full text of DOI:10.1021/jo971272w

J . Org. Chem. 1997, 62, 8463-8473
8463
A New Gen er a tion of “Ch ola p h a n es”: Ster oid -Der ived Ma cr ocyclic
Hosts w ith En h a n ced Solu bility a n d Con tr olled F lexibility
Khadga M. Bhattarai, Anthony P. Davis,* J ustin J . Perry, and Christopher J . Walter
Department of Chemistry, Trinity College, Dublin 2, Ireland
Stephan Menzer and David J . Williams
Department of Chemistry, Imperial College of Science and Technology,
South Kensington, London SW7 2AY, U.K.
Received J uly 14, 1997X
The macrocyclic “cholaphanes” 3a -c were synthesized from the inexpensive steroid cholic acid.
Like earlier relatives they feature substantial cavities with inward-directed hydroxyl groups, suitable
for binding polar molecules such as carbohydrates in nonpolar media. New features are the
externally directed alkyl chains, promoting solubility in organic solvents, and (in the case of 3b/c)
reduced conformational freedom resulting from truncation of the steroidal side-chain. In particular,
modeling shows that the smallest macrocycle 3c possesses very little flexibility, preferring an open
conformation which is also revealed in the X-ray crystal structure of its pentahydrate. NMR studies
indicated that all three cholaphanes form 1:1 complexes with octyl â-D-glucoside in CDCl3, with Ka
) 600-1560 M-1. Cholaphanes 3b/c proved able to extract methyl â-D-glucoside from aqueous
solutions into CHCl3. The transport of methyl â-D-glucoside across a chloroform barrier was also
demonstrated for 3c.
In tr od u ction
Conformational control is a central challenge in the
practice of supramolecular chemistry. The programmed
association of molecules must be based on specific three-
dimensional complementarities, which are hard to achieve
in the absence of well-defined conformational preferences.
Control in most (synthetic) supramolecular systems
derives largely from the use of rigid “construction mod-
ules” which restrict the freedom of the overall structure.
Thus benzene rings, aromatic heterocycles, alkyne units,
and aliphatic six-membered rings have all featured
strongly in supramolecular architecture.1
The steroid nucleus can make unique contributions to
this area owing to its size, chirality, and rigid polycyclic
framework.2 The bile acids, such as cholic acid (1), have
proved especially useful, because of their availability and
useful levels of functionalization.2b,c On one hand, the
presence of functionality at both ends of these units
suggests the construction of macrocyclic structures which
may possess a high level of inward-directed polar func-
tionality.3 On the other, the codirected hydroxyl groups
present in most bile acids may be exploited (directly or
indirectly) in podand-type receptors,4 linear dimeric
hosts,5 or “facial amphiphiles”.6
The typical bile acid structure, as in 1, may be viewed
as two segments, the large rigid tetracyclic nucleus and
the smaller, flexible side-chain. Derived architectures
often inherit this combination of rigidity and flexibility,
which is useful for many applications. However, in the
case of synthetic receptors, even moderate levels of
flexibility may be undesirable. The restricted preorga-
nization may limit the selectivity attainable7 and reduce
the strength of binding to well-matched substrates.
Almost certainly, it will hamper attempts to model the
system and to predict and interpret its behavior.
This problem arose in our early studies on the
“cholaphanes” 2,2a,8 the first of the above-mentioned
macrocyclic systems, and among the first synthetic
receptors for carbohydrate derivatives in nonpolar sol-
(3) Recent examples: (a) Davis, A. P.; Walsh, J . J . J . Chem. Soc.,
Chem. Commun. 1996, 449. (b) Davis, A. P.; Menzer, S.; Walsh, J . J .;
Williams, D. J . J . Chem. Soc., Chem. Commun. 1996, 453. (c) Davis,
A. P.; Gilmer, J . F.; Perry, J . J . Angew. Chem., Int. Ed. Engl. 1996,
35, 1312. (d) Kolehmainen, E.; Tamminen, J .; Lappalainen, K.; Torkkel,
T.; Seppala, R. Synthesis 1996, 1082. (e) Irie, S.; Yamamoto, M.;
Kishikawa, K.; Kohmoto, S.; Yamada, K. Synthesis 1996, 1135. (f)
Kohmoto, S.; Fukui, D.; Nagashima, T.; Kishikawa, K.; Yamamoto, M.;
Yamada, K. J . Chem. Soc., Chem. Commun. 1996, 1869. (g) Bonar-
Law, R. P. J . Am. Chem. Soc. 1995, 117, 12397. (h) Bonar-Law, R. P.;
Sanders, J . K. M. J . Am. Chem. Soc. 1995, 117, 259. (i) Albert, D.;
Feigl, M. Tetrahedron Lett. 1994, 35, 565.
(4) Boyce, R.; Li, G.; Nestler, H. P.; Suenaga, T.; Still, W. C. J . Am.
Chem. Soc. 1994, 116, 7955. Cheng, Y. A.; Suenaga, T.; Still, W. C. J .
Am. Chem. Soc. 1996, 118, 1813. Hsieh, H.-P.; Muller, J . G.; Burrows,
C. J . J . Am. Chem. Soc. 1994, 116, 12077. Maitra, U.; D′Souza, L. J .
J . Chem. Soc., Chem. Commun. 1994, 2793. Davis, A. P.; Perry, J . J .;
Williams, R. P. J . Am. Chem. Soc. 1997, 119, 1793.
(5) McKenna, J .; McKenna, J . M.; Thornthwaite, D. W. J . Chem.
Soc., Chem. Commun. 1977, 809. Burrows, C. J .; Sauter, R. A. J .
Inclusion Phenom. 1987, 5, 117. J anout, V.; Lanier, M.; Regen, S. L.
J . Am. Chem. Soc. 1996, 118, 1573.
(6) Cheng, Y.; Ho, D. M.; Gottlieb, C. R.; Kahne, D.; Bruck, M. A. J .
Am. Chem. Soc. 1992, 114, 7319. Venkatasan, P.; Cheng, Y.; Kahne,
D. J . Am. Chem. Soc. 1994, 116, 6955.
(7) For recent illustrations of this principle, featuring inverse
correlations between flexibility and enantioselectivity, see: (a) Yoon,
S. S.; Georgiadis, T. M.; Still, W. C. Tetrahedron Lett. 1993, 34, 6697.
(b) Cuntze, J .; Owens, L.; Alcazar, V.; Seiler, P.; Diederich, F. Helv.
Chim. Acta 1995, 78, 367.
X Abstract published in Advance ACS Abstracts, November 1, 1997.
(1) (a) Lehn, J .-M., Ed. Comprehensive Supramolecular Chemistry;
Pergamon: Oxford, 1996. Other leading refs: (b) Diederich, F. N.
Cyclophanes; Royal Society of Chemistry: Cambridge, 1991. (c) Rebek,
J ., J r. Angew. Chem., Int. Ed. Engl. 1990, 29, 245. (d) Vo¨gtle, F.
Cyclophane Chemistry; Synthesis, Structures, Reactions; Wiley: Chich-
ester, 1993. (e) Philp, D.; Stoddart, J . F. Angew. Chem., Int. Ed. Engl.
1996, 35, 1155. (f) Kilburn, J . D.; Patel, H. K. Contemp. Org. Synth.
1994, 1, 259. (g) Dowden, J .; Kilburn, J . D.; Wright, P. Contemp. Org.
Synth. 1995, 2, 289.
(2) (a) Davis, A. P. Chem. Soc. Rev. 1993, 22, 243. (b) Davis, A. P.;
Bonar-Law, R. P.; Sanders, J . K. M. in ref 1a, Vol. 4, p 257. (c) Miyata,
M.; Sada, K. in ref 1a, Vol. 6. (d) Li, Y. X.; Dias, J . R. Chem. Rev.
(Washington, D.C.) 1997, 97, 283.
S0022-3263(97)01272-3 CCC: $14.00 © 1997 American Chemical Society

8464 J . Org. Chem., Vol. 62, No. 24, 1997
Bhattarai et al.
vents.9 While these hosts showed pleasing affinities and
Sch em e 1
selectivities for alkyl glucosides in chloroform,8b,c
a
detailed interpretation of the binding results proved
unrealistic. Molecular modeling showed that the frame-
work of 2 could adopt various conformations, arising from
rotations about the series of single bonds connecting the
inflexible arylsteroid units.8b,e Consideration of all low
energy conformers, combined with an adequate range of
substrate orientations, was beyond our computational
resources.
flexibility is controlled through steroidal side-chain ad-
justment, and (b) solubility in nonpolar solvents is
maintained by four externally directed butyl esters.
Preliminary binding studies, computer-based molecular
modeling and X-ray crystallography suggest that 3c, in
particular, has unusual potential for the selective (and
interpretable) recognition of carbohydrates in nonpolar
media
A second problem encountered in our early work was
the limited solubility of 2 (R ) H), the more effective of
the two receptors. Maximum concentrations in CDCl3
(ca. 2 mM) were barely sufficient for the binding mea-
surements. An increase in rigidity appeared likely to
exacerbate this difficulty, and we therefore sought a way
to separate the two problems, i.e., to tune solubility
independently of the central framework.
The answer seemed to lie in Scheme 1, a sequence for
introducing both an aryl spacer group and an externally
directed 3â solubilizing group, employing a chemoselec-
tive organometallic reagent. Preliminary work confirmed
that this sequence was practical for E ) CN,10 and that
the product could be converted to a cholaphane.11 We
now report the synthetic culmination of this program, the
preparation of a range of cholaphanes 3 in which (a) core
(8) (a) Bonar-Law, R. P.; Davis, A. P. J . Chem. Soc., Chem. Commun.
1989, 1050. (b) Bonar-Law, R. P.; Davis, A. P.; Murray, B. A. Angew.
Chem., Int. Ed. Engl. 1990, 29, 1407. (c) Bhattarai, K. M.; Bonar-Law,
R. P.; Davis, A. P.; Murray, B. A. J . Chem. Soc., Chem. Commun. 1992,
752. (d) Bonar-Law, R. P.; Davis, A. P. Tetrahedron 1993, 49, 9829.
(e) Bonar-Law, R. P.; Davis, A. P. Tetrahedron 1993, 49, 9845. (f)
Bonar-Law, R. P.; Davis, A. P.; Dorgan, B. J . Tetrahedron 1993, 49,
9855.
Resu lts a n d Discu ssion
Syn th esis of 3a a n d 3b. The syntheses of 3a , from
cholic acid (1), and 3b, from norcholic acid (5), followed
parallel paths as outlined in Schemes 2 and 3. Although
our previous cholaphane syntheses had relied mainly on
acetyl protection for the 7,12R-OH groups,8,11 removal had
proved troublesome. Considering the presence of the
dibutylmalonyl groups in 3, we decided to use the more
labile formyl groups in this case.12 The preparation of
norcholic acid (5), and the formylation of 1 and 5 to give
6a and 6b, respectively, were accomplished using high-
yielding literature procedures.13,14 Esterification with
diazomethane, selective deformylation of the equatorial
C3-oxygens, and oxidation to ketones 8 proceeded smoothly
as expected.
(9) For other studies on carbohydrate recognition, see e.g. refs 3b,
3f, 3h, 7b, and: (a) Aoyama, Y.; Tanaka, Y.; Sugahara, S. J . Am. Chem.
Soc. 1989, 111, 5397. (b) Kikuchi, Y.; Tanaka, Y.; Sutaro, S.; Kobayashi,
K.; Toi, H.; Aoyama, Y. J . Am. Chem. Soc. 1992, 114, 10302. (c) Liu,
R.; Still, W. C. Tetrahedron Lett. 1993, 34, 2573. (d) Savage, P. B.;
Gellman, S. H. J . Am. Chem. Soc. 1993, 115, 10448. (e) Das, G.;
Hamilton, A. D. J . Am. Chem. Soc. 1994, 116, 11139. (f) Huang, C.-Y.;
Cabell, L. A.; Anslyn, E. V. J . Am. Chem. Soc. 1994, 116, 2778. (g)
Cotero´n, J . M.; Vicent, C.; Bosso, C.; Penade´s, S. J . Am. Chem. Soc.
1993, 115, 10066. (h) Eliseev, A. V.; Schneider, H.-J . J . Am. Chem.
Soc. 1994, 116, 6081. (i) Mohler, L. K.; Czarnik, A. W. J . Am. Chem.
Soc. 1993, 115, 2998. (j) Morin, G. T.; Hughes, M. P.; Paugam, M.-F.;
Smith, B. D. J . Am. Chem. Soc. 1994, 116, 8895. (k) Deng, G.; J ames,
T. D.; Shinkai, S. J . Am. Chem. Soc. 1994, 116, 4567. (l) J ames, T. D.;
Sandanayake, K. R. A. S.; Shinkai, S. Angew. Chem., Int. Ed. Engl.
1996, 35, 1911. (m) Mizutani, T.; Murakami, T.; Matsumi, N.; Kura-
hashi, T.; Ogoshi, H. J . Chem. Soc., Chem. Commun. 1995, 1257. (n)
Anderson, S.; Neidlein, U.; Gramlich, V.; Diederich, F. Angew. Chem.,
Int. Ed. Engl. 1995, 34, 1596. (o) Neidlein, U.; Diederich, F. J . Chem.
Soc., Chem. Commun. 1996, 1493. (p) Inouye, M.; Miyake, T.; Furusyo,
M.; Nakazumi, H. J . Am. Chem. Soc. 1995, 117, 12416.
The next step, malonylidenation of the 3-keto group,
had appeared the major obstacle to success at the start
of our program. While the Knoevenagel condensation
between malonates and aldehydes is a well-established
(10) Davis, A. P.; Egan, T. J .; Orchard, M. G.; Cunningham, D.;
McArdle, P. Tetrahedron 1992, 48, 8725.
(11) Davis, A. P.; Orchard, M. G. J . Chem. Soc., Perkin Trans 1 1993,
919.
(12) cf. refs 3a-c.
(13) Schteingart, C. D.; Hofmann, A. F. J . Lipid Res. 1988, 29, 1387.
(14) Tserng, K.-Y.; Klein, P. D. Steroids 1977, 29, 635.

A New Generation of “Cholaphanes”
J . Org. Chem., Vol. 62, No. 24, 1997 8465
Sch em e 2
Sch em e 3
a
Reaction conditions. (a) HCO2H, HClO4 (cat.), Ac2O; (b) CH2N2,
CH2Cl2, MeOH; (c) NaHCO3, MeOH, 0 °C; (d) pyridinium chloro-
chromate, CH2Cl2, SiO2.
process,15 the reaction between malonates and ketones
does not proceed well using traditional methodology.
However, as described elsewhere,16 the problem was
overcome through a “forcing” equivalent of the Knoev-
enagel reaction, employing 2,2-dibromomalonates and
tributylstibine, and driven by oxidation of Sb(III) to Sb-
(V). Application of this method to ketones 8 gave dibutyl
malonylidene derivatives 9 in excellent yields.
Introduction of the spacer group required an organo-
metallic reagent which could add to C3 chemoselectively
(in the presence of 5 ester groups) and stereoselectively
(equatorial attack) and which bore a substituent capable
of conversion to an aminomethyl group. Previous work
on related systems10,11 implied that a “higher-order
cuprate” derived from a Grignard reagent would have the
necessary chemo- and stereoselectivity. Regarding the
substituent, our choice was limited by the dearth of
N-protecting groups which are compatible with Grignard
and organocuprate reagents.17 Although an (N-pyrrolo)-
methyl group had proved viable in similar circum-
stances,18 the deprotection method required a hydrolysis
step which would probably have been incompatible with
the formate esters. We therefore reverted to our earlier
procedure,11 involving a tert-butoxymethyl substituent
which is subsequently converted to hydroxymethyl, me-
sylated, treated with azide, reduced to aminomethyl, and
Boc-protected. The sequence from 9 through to 13
proceeded well in both series, with overall yields of 61%
(n ) 2) and 68% (n ) 1).
a
Reaction conditions. (a) SbBu3 (3.5 equiv), dibutyl 2,2-dibro-
momalonate (1.4 equiv), THF, 60-65 °C; (b) Ar2Mg2Br2CuCN,
THF, -60 °C [Ar ) p-(C6H4)CH2Ot-Bu]; (c) TFA, CH2Cl2, 50 °C,
then NH3 (aq), Et2O; (d) MsCl, i-Pr2NEt, CH2Cl2, -14-0 °C, then
(Me2N)2CNH2+N3-, 0-30 °C; (e) Ph3P, THF, H2O, 60 °C, then
(Boc)2O; (f) NaOH, MeOH, THF, then HCO2H, HClO4 (cat.), Ac2O,
then (Boc)2O, Et2O, THF, NaHCO3 (aq); (g) C6F5OH, DCC, CH2Cl2;
(h) TFA, CH2Cl2, then DMAP, CH2Cl2; (i) K2CO3, THF, MeOH,
H2O, 60 °C.
groups, cyclodimerization, and deformylation. The first
of these steps proved somewhat troublesome, as hydroly-
sis of the methyl esters could not be accomplished without
removal of the formyl groups in positions 7 and 12. The
latter could be reinstated, but the acidic conditions
necessary removed the N-Boc protection. However, after
reprotection of the amino groups and treatment with
DCC/pentafluorophenol, the activated esters 14 were
obtained in acceptable yields. N-Deprotection and cy-
clodimerization at high dilution (ca. 1 mM) gave macro-
cycles 4a and 4b in yields of 77% and 44%, respectively,
the difference perhaps reflecting an element of strain in
4b . Finally, deformylation gave the tetrahydroxy-
cholaphanes 3a /b.
Conversion of 13 to macrocycles 3 required activation
of the side-chain carboxyls, deprotection of the amino
(15) J ones, G. Org. React. 1967, 15, 204.
(16) (a) Davis, A. P.; Bhattarai, K. M. Tetrahedron 1995, 51, 8033.
See also: (b) Liao, Y.; Huang, Y.-Z. Tetrahedron Lett. 1990, 31, 5897.
(c) Chen, C.; Huang, Y.-Z.; Shen, Y.; Liao, Y. Heteroatom Chem. 1990,
1, 49.
Syn th esis of 3c. The sequence leading to the smallest
cholaphane, 3c, is shown in Scheme 4. The synthesis
differs from those in Schemes 2 and 3 in that side-chain
degradation was accomplished in two widely separated
stages. Removal of C(24) through oxidative decarboxy-
(17) Most common N-protecting groups contain acidic protons or
polarized multiple bonds which will quench
a Grignard reagent.
Obvious exceptions, such as silyl groups, do not eliminate N-basicity,
which can interfere with the reactivity of a derived cuprate. For further
discussion see ref 11.
(18) Davis, A. P.; Egan, T. J . Tetrahedron Lett. 1992, 33, 8125.

8466 J . Org. Chem., Vol. 62, No. 24, 1997
Bhattarai et al.
Sch em e 4
F igu r e 1. The structure of 3c in the crystal showing also the
entrapped hydrogen bonded solvent water molecules. Nonter-
minated bonds represent hydrogen bonds to symmetry-related
molecules.
hydroxylation,20 treatment with sodium metaperiodate,
and oxidation of the resulting aldehyde with sodium
chlorite21 ). In this case the carboxyl group was activated
as the (more reactive) pentafluorothiophenyl ester22 to
compensate for the sterically hindered environment of
the carbonyl carbon. Cyclodimerization to 4c (in 67%
yield) was followed by deformylation, to give the target
cholaphane 3c.
Str u ctu r a l a n d Recogn ition P r op er ties of 3a -c.
As discussed earlier, the design of 3a -c was influenced
by two major considerations. First, we wished to control
and, in particular, lower the flexibility of the cholaphane
framework, to increase selectivity and binding power (to
well-matched substrates). Second, we wished to main-
tain solubility in nonpolar organic media, irrespective of
the core structure. Initial observations implied that we
had succeeded in both respects. On one hand, all the
tetrahydroxycholaphanes were freely soluble in CDCl3,
to the level of at least 25 mg/mL. On the other, computer-
based molecular modeling23 showed that, while 3a and
3b could undertake a variety of conformations (including
collapsed structures with no substantial cavity), the
macrocyclic framework of 3c had very little freedom.
Open toroidal or bowl-shaped structures dominated the
lower reaches of the energy spectrum, the only important
mode of flexibility being a folding motion with the Ar-
CH2-N bonds acting as hinges.
The structure of 3c, and its potential as a host for polar
molecules, was confirmed by X-ray crystallography (Fig-
ure 1). Crystals were grown from a solution in 1:1 CH2-
Cl2-MeOH, which was allowed to evaporate in an
atmosphere of methanol. Although there was no deliber-
ate introduction of water, the crystals were found to be
heavily hydrated with each molecule of 3c accompanied
by five water molecules. The X-ray analysis reveals the
molecule in an open, approximately C2-symmetric con-
formation which might be described as a “skewed bucket”
by analogy to e.g. cyclodextrins (Figure 2). The transan-
nular separations of the pairs of C(12) and C(7) hydroxyl
oxygen atoms are 7.4 and 5.5 Å respectively. These
effectively determine the breadth of the cavity while the
a
Reaction conditions. (a) Pb(OAc)4, Cu(OAc)2 cat., py, C6H6; (b)
NaOAc‚3H2O, MeOH; (c) pyridinium chlorochromate, CH2Cl2,
SiO2; (d) SbBu3 (3 equiv), dibutyl 2,2-dibromomalonate (1.2 equiv),
THF, 60-65 °C; (e) Ar2Mg2Br2CuCN, THF, -60 °C [Ar
)
p-(C6H4)CH2Ot-Bu]; (f) TFA, CH2Cl2, 50 °C, then NH3 (aq), Et2O;
(g) MsCl, i-Pr2NEt, CH2Cl2, -14-0 °C, then (Me2N)2CNH2+N3
,
-
0-30 °C; (h) Ph3P, THF, H2O, 60 °C, then (Boc)2O; (i) N-
methylmorpholine N-oxide, OsO4 (cat.), py, Bu4NOAc; (j) NaIO4,
CH3CN, H2O, then NaClO2, H2NSO3H, CH3CN, H2O; (k) C6F5SH,
DCC, CH2Cl2; (l) TFA, CH2Cl2, then DMAP, CH2Cl2; (m) K2CO3,
THF, MeOH, H2O, 60 °C.
(19) Carlson, G. L.; Belobaba, D. T. E.; Hofmann, A. F.; Wedmid, Y.
Steroids 1977, 30, 787.
(20) Carlsen, P. H. J .; Katsuki, T.; Martin, V. S.; Sharpless, K. B.
J . Org. Chem. 1981, 46, 3936.
(21) Lindgren, B. O.; Nilsson, T. Acta Chem. Scand. 1973, 27, 888.
(22) Davis, A. P.; Walsh, J . J . Tetrahedron Lett. 1994, 35, 4865.
(23) Details to be published separately.
lation of triformate 6a yielded the 22,23-alkene in 15.3a,19
As in previous work involving “bis-nor” monomer units,3a,b
this group was employed as a “masked carboxyl”, being
carried through the introduction of the spacer group and
then revealed via oxidative cleavage (OsO4-catalyzed bis-

A New Generation of “Cholaphanes”
J . Org. Chem., Vol. 62, No. 24, 1997 8467
of 3a the motions were consistent with a binding constant
(Ka) of 1305 M-1, and a limiting chemical shift change
(∆δ) of 1.35 ppm. This binding constant is significantly
lower than that of 3100 M-1 observed previously for 2 (R
) H),8c perhaps implying that the rigidly constrained
aromatic spacer group interferes with the preferred
binding geometry of the carbohydrate. In the case of 3b,
a still lower Ka of 600 M-1 was measured (∆δ ) 0.84
ppm). For the smallest and least flexible macrocycle 3c,
we naturally hoped for a substantial increase in binding
constant, reflecting the improved preorganization of the
cavity. In fact, the measured value of 1560 M-1 (∆δ )
1.07 ppm) was somewhat disappointing. However, we
had no reason to assume, a prioiri, that 25 would be
especially complementary to 3c, and this rather modest
figure may be eclipsed when we survey a wider range of
substrates.
F igu r e 2. Side view of the macrocyclic portion of 3c showing
the skewed conformation.
The tetrahydroxycholaphanes were also tested for their
abilities to extract the hydrophilic methyl â-D-glucoside
(26) from aqueous solution into an organic medium.25
Solutions of the macrocycles in chloroform (1 mM) were
stirred with aqueous 26 (concentrations from 2.5 M down
to 1 M) and the separated organic phases analyzed for
carbohydrate using TLC. No extraction was detected in
the absence of receptor. The technique could be operated
in a semiquantitative fashion by applying the samples
in a standard manner using a micropipet and making
comparisons with solutions of 26 at known concentra-
tions. When the method was applied to 3c, extraction
was clearly detectable over the full range of glucoside
concentrations. With [26] (aq) ) 1.75 M, the analysis
indicated that roughly 0.2 equiv of the glucoside were
carried into the organic phase. When the full range of
cholaphanes [including 2 (R ) H)] were tested under the
same conditions, the results were somewhat surprising.
The order of effectiveness appeared to be 3c > 3b > 2 (R
) H) > 3a . In the case of 3a , the degree of extraction
was below the limit of detection. Although the poor
performance of 2 (R ) H) and 3a was unexpected, in view
of their effectiveness in the homogeneous binding studies,
it may be noted that different substrates (octyl vs methyl
glucosides) and conditions (dry vs water-saturated CHCl3)
are involved in the two types of experiment.
F igu r e 3. Side view of the packing of molecules of 3c in the
crystal. The water molecules that occupy the helical channel
passing through the macrocycles are depicted as large spheres.
surfaces of the aromatic spacer units (ring-centroid‚‚‚-
ring-centroid distance 11.5 Å) define its length. The
inward-directed hydroxyls and amide N-H groups create
a hydrophilic interior within which the five included
water molecules are bound via O-H‚‚‚O and N-H‚‚‚O
hydrogen bonds.24 It is interesting to note that the
included water molecules are concentrated toward one
end of the macrocyclic cavity, and also that only one of
the amide N-H groups is involved in hydrogen bonding.
Inspection of the packing of the molecules in the crystal
shows one of the crystallographic screw axes to pass
through this region of the macrocycle. The effect of this
is to produce partial overlap of the cavities of symmetry
related macrocycles creating helical water-containing
channels that extend in the b direction (Figure 3).
As demonstrated in previous work, the three-dimen-
sional arrangement of polar functionality in cholaphanes
is particularly suitable for the recognition of carbohydrate
nuclei in nonpolar media.8b,c In a preliminary test of their
carbohydrate-binding properties, macrocycles 3 were
subjected to NMR titration experiments in CDCl3 with
octyl â-D-glucoside (25) as guest. Addition of 25 to 3
resulted in downfield shifts of the receptor NH NMR
signals, analyzable in terms of 1:1 binding. In the case
The extraction ability of 3c was confirmed by a
transport experiment conducted in a U-tube apparatus,
employing a gently stirred solution of the cholaphane in
chloroform (2 mM), overlaid by aqueous 26 (2 M) on one
side and an aqueous receiving phase on the other. The
carbohydrate could be detected in the receiving phase
after 24 h and increased steadily in concentration over a
number of days. Transport was not observed in the
absence of the receptor.
In conclusion, we have developed a new method for
constructing cholaphanes with heightened solubility in
organic solvents and applied it to the synthesis of a series
of receptors which vary in size and flexibility. The
smallest and least flexible example (3c) possesses a well-
(24) There are 15 O-H‚‚‚O hydrogen bonds which range in distance
from 2.73 to 3.05 Å, and one N-H‚‚‚O hydrogen bond (3.07 Å) from
one of the amide nitrogen atoms to one of the included water molecules.
(25) For the extraction of 26 into CCl4, as a 2:1 host:guest complex,
see ref 9b.

8468 J . Org. Chem., Vol. 62, No. 24, 1997
Bhattarai et al.
22.24, 18.6, 12.00. Anal. Calcd for C26H40O7: C, 67.22; H, 8.68.
Found: C, 66.69; H, 8.66.
defined central famework and a highly preorganized,
polar interior. The structural and preliminary recogni-
tion studies on this new cholaphane suggest that it will
prove useful in investigations of carbohydrate binding
and transport in nonpolar organic media.
Meth yl 3-Oxo-7r,12r-bis(for m yloxy)-5â-ch ola n -24-oa te
(8a ). Silica gel (7 g) was dried overnight at 110 °C and then
stirred under nitrogen with pyridinium chlorochromate (5.6
g, 26 mmol) and CH2Cl2 (50 mL). A solution of 7a (6.81 g,
14.2 mmol) in CH2Cl2 (50 mL) was added dropwise. Analysis
by TLC after 2 h showed the reaction to be complete. The
mixture was filtered and the insoluble material washed with
ether. Concentration of the filtrate and washings gave a
greenish yellow residue which was dissolved in ether, washed
with water and brine, dried (MgSO4), and filtered through a
short pad of silica. Evaporation of the eluent and crystalliza-
tion from CH2Cl2-hexane to yielded 8a as white needles (5.93
g, 87%): TLC Rf 0.6 (CH2Cl2-ether, 4:1); IR (film from CDCl3)
2965, 2880, 1716, 1180 cm-1; 1H NMR (300 MHz) δ 8.16 (1H,
s), 8.10 (1H, s), 5.31 (1H, br s), 5.16 (1H, br s), 3.66 (3H, s),
3.01 (1H, t, J ) 15 Hz), 1.04 (3H, s), 0.85 (3H, d, J ) 6.4 Hz),
0.79 (3H, s); 13C NMR δ 211.52, 174.33, 160.34, 160.25, 75.0,
70.47, 51.44, 47.10, 44.94, 44.44, 42.76, 42.06, 37.61, 36.43,
36.05, 34.62, 33.33, 30.93, 30.75, 30.51, 29.39, 27.03, 25.77,
22.66, 21.45, 17.36, 12.07. Anal. Calcd for C27H40O7: C, 68.04;
H, 8.46. Found: C, 67.70; H, 8.47.
Exp er im en ta l Section
Gen er a l Meth od s. 1H and 13C NMR spectra were recorded
in CDCl3 on Bruker MSL300 (1H at 300 MHz, 13C at 75 MHz)
or WP80 (1H at 80 MHz) spectrometers, with TMS and CDCl3,
respectively, as internal standards. IR spectra were recorded
on a Perkin-Elmer 298 spectrometer. Flash chromatography26
was performed using Kieselgel 60 (400-230 mesh) purchased
from Merck. Reactions were monitored by TLC on DC Alu-
folien Kieselgel 60 F254 (0.2 mm) and visualized by charring
or developing with iodine as appropriate. CHCl3 and CDCl3
used for extraction and binding experiments were deacidified
and partially dried by storage over potassium carbonate.
Anhydrous solvents were prepared by following standard
procedures.27 Melting points were uncorrected.
Sta r tin g Ma ter ia ls. Cholic acid (1) was supplied by
Freedom Chemical Diamalt GmbH of Raubling, Germany.
Triformate 6a was prepared from 1, formic acid, and acetic
anhydride, employing a modification of the procedure of Tserng
and Klein.14 24-Norcholic acid (5) was prepared from trifor-
mate 6a via R-nitrosation-fragmentation to the nornitrile,
using a method based on that of Schteingart and Hofmann.13
Formylation of 5, as above, gave triformate 6b. Details of
these preparations are given as Supporting Information.
Meth yl 3r-Hyd r oxy-7r,12r-bis(for m yloxy)-5â-ch ola n -
24-oa te (7a ). Diazomethane was bubbled into a solution of
6a (20 g, 41 mmol) dissolved in CH2Cl2 (400 mL) and methanol
(10 mL) maintained at -5 °C to 0 °C. After the completion of
the reaction as judged by TLC (hexane-ethyl acetate, 7:3),
evaporation of solvent yielded the corresponding methyl ester.
This material (19.5 g, ca. 39 mmol) was dissolved in methanol
(300 mL) and cooled to 0 °C. Solid sodium bicarbonate (5 g,
60 mmol) was added in portions with stirring. Deformylation
was followed by TLC (CH2Cl2-ether, 1:1). After completion
of the reaction (1.5 to 2 h) unreacted bicarbonate was neutral-
ized with dilute acetic acid and the solvent evaporated to give
a residue. The residue was dissolved in ether, washed with
water, dried over Na2SO4, and filtered. Evaporation of the
filtrate gave crude 7a as a gum (18 g, ca. 92% overall from
6a ), homogeneous by TLC. A portion of this material was
further purified by flash chromatography (ether as eluent) to
give 7a as a white solid: TLC Rf 0.3 (CH2Cl2-ether, 1:1); IR
(film from CDCl3), 3422, 1718, 1180 cm-1; 1H NMR (300 MHz)
δ 8.14 (1H, s), 8.10 (1H, s), 5.26 (1H, br s), 5.05 (1H, s), 3.65
(3H, s), 3.49 (1H, m), 0.92 (3H, s), 0.83 (3H, d, J ) 6.2 Hz),
0.75 (3H, s); 13C NMR δ 174.4, 160.67, 160.5, 75.19, 71.31,
70.75, 51.42, 47.05, 44.85, 42.83, 40.85, 38.38, 37.61, 34.76,
34.62, 34.14, 31.34, 30.76, 30.53, 30.20, 28.43, 27.04, 25.38,
22.66, 22.25, 17.34, 11.99. Anal. Calcd for C27H42O7‚H2O: C,
65.32; H, 8.46. Found: C, 65.34; H, 8.65.
Meth yl 3-Oxo-7r,12r-bis(for m yloxy)-24-n or -5â-ch ola n -
23-oa te (8b). 7b (4.95 g, 10.65 mmol) was treated as above
with silica gel (5 g) and pyridinium chlorochromate (3.8 g, 18.6
mmol) in CH2Cl2 (80 mL), to give 8b as white needles (4.25 g,
86%): TLC Rf 0.6 (CH2Cl2-ether, 4:1); IR (film from CDCl3)
1
2963, 2878, 1715, 1178, 730 cm-1; H NMR (300 MHz) δ 8.16
(1H, s), 8.09 (1H, s), 5.30 (1H, br s), 5.15 (1H, br d), 3.65 (3H,
s), 3.01 (1H, t, J ) 14 Hz), 1.04 (3H, s), 0.91 (3H, d, J ) 6.2
Hz), 0.83 (3H, s); 13C NMR δ 211.48, 173.46, 160.23, 74.87,
70.40, 51.34, 47.09, 45.02, 44.43, 42.79, 42.02, 40.92, 37.60,
36.41, 36.02, 34.31, 33.02, 30.92, 29.37, 27.15, 25.74, 22.65,
21.44, 18.63, 12.08. Anal. Calcd for C26H38O7: C, 67.51; H,
8.28. Found: C, 67.63; H, 8.33.
Met h yl 3-[Bis(b u t oxyca r b on yl)m et h ylid en e]-7r,12r-
bis(for m yloxy)-24-n or -5â-ch ola n -23-oa te (9b). According
to the previously reported method,16a tributylstibine (8.82 g,
8 mL, 30.1 mmol) was injected slowly into a stirred solution
of ketone 8b (4.0 g, 8.65 mmol) and dibutyl dibromomalonate
(4.5 g, 12.04 mmol) in dry THF (30 mL) under nitrogen at room
temperature. The reaction vessel was lowered into an oil bath
at 60-65 °C and stirring was maintained for 72 h. After this
time interval, TLC analysis (eluent CH2Cl2) of the reaction
mixture indicated complete disappearance of starting ketone
(visualization by 2,4-DNP) and appearance of the faster-
moving product. The reaction mixture was cooled and stirred
open to the atmosphere for 2 h. The solvent was evaporated
and the residue dissolved in a minimum volume of hexane-
ether (4:1). This solution was applied to to a column of silica
gel column (2.5 × 20 cm) packed using hexane and eluted with
hexane (200 mL), hexane-ether (200 mL, 9:1), hexane-ether
(200 mL, 4:1) and finally with hexane-ether (7:3) until no
further product emerged. Fractions containing the product
were evaporated to yield 9b (5.50 g, 96%): TLC Rf 0.33
(hexane-ether, 1:1); IR (film from CHCl3) 2960, 2878, 1719,
1634, 1204, 1174, 1064 cm-1; 1H NMR (300 MHz) δ 8.14 (1H,
s), 8.09 (1H, s), 5.28 (1H, br s), 5.11 (1H, br s), 4.15 (4H, t, J
) 5.5 Hz), 3.65 (3H, s), 0.81 (3H, s); 13C NMR δ 173.40, 165.65,
165.42, 160.81, 160.30, 121.49, 74.90, 70.36, 64.67, 64.56,
51.26, 47.04, 44.89, 43.05, 42.77, 40.87, 37.53, 37.16, 34.66,
34.55, 32.97, 31.23, 30.34, 29.06, 27.13, 26.97, 25.56, 22.60,
Met h yl 3r-Hyd r oxy-7r,12r-b is(for m yloxy)-24-n or -5â-
ch ola n -23-oa te (7b). A solution of 6b (15 g, 31 mmol)
dissolved in CH2Cl2 (200 mL) and CH3OH (20 mL) was treated
with diazomethane as described above to give the correspond-
ing methyl ester. Deformylation as above with sodium bicar-
bonate (4.1 g, 49 mmol) in methanol (300 mL), followed by flash
chromatography with ether as eluent, gave 7b (13.84 g, 95%):
TLC Rf 0.3 (CH2Cl2-ether, 1:1); IR (film from CDCl3) 3422,
21.65, 18.92. 18.56, 13.49, 11.99. Anal. Calcd for C37H56O10
C, 67.24; H, 8.54. Found: C, 67.29; H, 8.20.
:
Meth yl 3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[(p-ter t-bu -
toxym eth yl)p h en yl]-7r,12r-bis(for m yloxy)-5â-ch ola n -24-
oa te (10a ). A solution of tert-butyl p-bromobenzyl ether (5 g,
20.56 mmol) in dry THF (30 mL) was added in one portion to
magnesium turnings (0.7 g, 28.5 mmol) in flame-dried ap-
paratus under argon. The Grignard reaction was initiated by
warming and sonication. Sonication continued for 1 h after
which the solution was transferred via cannula to a second
flame dried reaction vessel. The magnesium-free Grignard
reagent was cooled to ca. -10 °C (ice-salt bath), and CuCN
(0.895 g, 10 mmol) was added with stirring against a strong
1
2958, 2878, 1714, 1186, 908, 734 cm-1; H NMR (300 MHz) δ
8.14 (1H, s), 8.09 (1H, s), 5.25 (1H, br s), 5.05 (1H, br s), 3.65
(3H, s), 3.49 (1H, tt), 0.92 (3H, d, J ) 6.0 Hz), 0.88 (3H, s),
0.79 (3H, s); 13C NMR δ 173.55, 160.68, 160.38, 75.06, 71.25,
70.70, 51.29, 47.10, 44.93, 42.86, 40.95, 40.84, 38.38, 37.62,
34.75, 34.12, 33.02, 31.35, 30.19, 28.40, 27.17, 25.35, 22.63,
(26) Still, W. C.; Kahn, M.; Mitra, A. J . Org. Chem. 1978, 43, 2923.
(27) Perrin, D. D.; Armanego, W. F.; Perrin, D. R. Purification of
Laboratory Chemicals; 2nd ed.; Pergamon: Oxford, 1980.

A New Generation of “Cholaphanes”
J . Org. Chem., Vol. 62, No. 24, 1997 8469
flow of argon. Stirring was continued as the temperature was
allowed to rise to ca. 0 °C, and the resulting homogeneous
solution was then cooled to -60 °C. A solution of 9a 16a (5.00
g, 7.4 mmol) in THF (20 mL) was added dropwise, and then
the mixture was allowed to attain room temperature. The
reaction was quenched with saturated NH4Cl (aq) and the
mixture extracted with ether. The extract was washed with
water and brine, dried over MgSO4, and evaporated to give a
gummy residue. Flash chromatography eluting with hexane-
ether (4:1, followed by 7:3), gave 10a (5.39 g, 87%): TLC Rf
0.5 (hexane-ether, 1:1); IR (film from CHCl3) 2966, 2876, 1722,
(film from CDCl3) 3548, 1721, 1177 cm-1; 1H NMR (300 MHz)
δ 8.05 (1H, s), 8.03 (1H, s), 7.46, 7.31 (4H, ABq, J ) 8.6 Hz),
5.23 (1H, br s), 5.06 (1H, br s), 4.67 (2H, s), 4.08 (1H, s), 4.04-
3.87 (4H, m), 3.63 (3H, s), 1.04 (3H, s), 0.89-0.82 (9H, m), 0.78
(3H, s); 13C NMR δ 173.60, 168.12, 168.01, 160.52, 145.05,
138.98, 127.40, 126.04, 74.93, 70.60, 64.93, 64.83, 54.38, 51.36,
47.18, 44.96, 42.87, 42.63, 41.02, 38.08, 37.87, 37.65, 34.28,
33.07, 32.03, 31.54, 30.28, 29.33, 28.58, 27.21, 25.52, 22.92,
22.69, 18.97, 18.93, 18.63, 13.57, 12.06. Anal. Calcd for
C44H64O11: C, 68.72; H, 8.39. Found: C, 69.06; H, 8.41.
Meth yl 3â-[Bis(bu toxycar bon yl)m eth yl]-3r-[(p-azidom -
et h yl)p h en yl]-7r,12r-b is(for m yloxy)-5â-ch ola n -24-oa t e
(12a ). A solution of 11a (3.28 g, 4.19 mmol) in CH2Cl2 (20
mL) was cooled to -14 °C (ice-salt bath) under nitrogen.
Methanesulfonyl chloride (0.42 mL, 5.32 mmol) and then N,N-
diisopropylethylamine (0.95 mL, 5.47 mmol) were added
dropwise with stirring. The mixture was allowed to warm to
0 °C. TLC analysis of the reaction mixture after 1 h indicated
conversion of the starting material into two faster moving
products.29 N,N,N′,N′-Tetramethylguanidinium azide30 (1.29
g, 8.15 mmol) was added with stirring. The mixture was
allowed to reach room temperature then warmed to 30 °C for
30 min. The mixture was diluted with ether (40 mL) and
filtered through a short pad of silica, which was then washed
with ether. The filtrate and washings were combined, washed
with water and saturated brine, dried (MgSO4), and evapo-
rated to yield 12a (3.8 g) of sufficient purity for further
elaboration. A portion of the crude product (0.05 g) was
purified by flash chromatography using hexane-ether (4:1)
as eluent to yield pure 12a (0.033 g): TLC Rf 0.57 (hexane-
ethyl acetate, 3:2); IR (film from CDCl3) 2963, 2879, 2110,
1722, 1468, 1243, 1176 cm-1; 1H NMR (300 MHz) δ 8.06 (1H,
s), 8.03 (1H, s), 7.42, 7.25 (4H, ABq, J ) 8.5 Hz), 5.24 (1H, br
s), 5.06 (1H, br s), 4.33 (2H, s), 4.07 (1H, s), 4.03-3.84 (4H,
m), 3.64 (3H, s), 1.04 (3H, s), 0.89-0.79 (9H, m), 0.74 (3H, s);
13C NMR δ 174.35, 168.05, 167.95, 160.53, 145.75, 133.56,
127.65, 127.08, 75.03, 70.59, 64.82, 54.40, 51.41, 47.10, 44.86,
42.85, 42.65, 38.07, 37.76, 37.62, 34.65, 34.22, 31.99, 31.50,
30.79, 30.56, 30.21, 29.25, 28.62, 27.06, 25.56, 22.91, 22.67,
18.89, 17.34, 13.49, 12.02. Anal. Calcd for C45H65N3O10: C,
66.89; H, 8.11; N, 5.2. Found: C, 66.83; H, 8.16, N, 4.96.
Meth yl 3â-[Bis(bu toxycar bon yl)m eth yl]-3r-[p-(azidom -
et h yl)p h en yl]-7r,12r-b is(for m yloxy)-24-n or -5â-ch ola n -
23-oa te (12b). 11b (3.57 g, 4.65 mmol) was treated as above
with methanesulfonyl chloride (0.47 mL, 6.04 mmol) and
N,N-diisopropylethylamine (1.1 mL, 6.3 mmol), followed by
N,N,N′,N′-tetramethylguanidinium azide (1.43, g, 9.06 mmol),
to yield crude 12b (3.96 g). Flash chromatography of a portion
of this material (0.06 g) yielded pure 12b: mp 130-133 °C;
TLC Rf 0.5 (hexane-ethyl acetate, 3:2); IR (film from CDCl3)
2964, 2880, 2101, 1723, 1178 cm-1; 1H NMR (300 MHz) δ 8.06
(1H, s), 8.03 (1H, s), 7.43, 7.25 (4H, ABq, J ) 8.5 Hz), 5.24
(1H, br t), 5.07 (1H, br d), 4.33 (2H, s), 4.08 (1H, s), 4.07-3.84
(4H, m), 3.63 (3H, s), 1.04 (3H, s), 0.89-0.82 (9H, m), 0.78 (3H,
s); 13C NMR δ 173.53, 168.05, 167.94, 160.48, 145.73, 133.56,
127.65, 127.08, 74.90, 70.53, 64.82, 54.40, 51.33, 47.15, 44.94,
42.88, 42.62, 40.98, 38.05, 37.75, 37.61, 34.22, 33.05, 31.96,
31.50, 30.21, 29.25, 28.58, 27.19, 25.52, 22.91, 22.66, 18.92,
18.61, 13.52, 12.03. Anal. Calcd for C44H63N3O10: C, 66.56;
H, 7.99; N, 5.29. Found: C, 66.48; H, 8.05; N, 4.83.
1618, 1467, 1439, 1389, 1364, 1243, 1178, 1063, 1020 cm-1
;
1H NMR (300 MHz) δ 8.05 (1H, s), 8.01 (1H, s), 7.34, 7.26 (4H,
ABq, J ) 8.6 Hz), 5.23 (1H, br s), 5.05 (1H, br s), 4.38 (2H, s),
4.06 (1H, s), 4.03-3.86 (4H, m), 3.65 (3H, s), 1.28, (9H, s), 1.03
(3H, s), 0.89-0.80 (9H, m), 0.73 (3H, s); 13C NMR δ 174.41,
168.14, 160.63 and 160.55, 144.50, 137.58, 127.13, 126.67,
74.92, 73.16, 70.57, 64.73, 63.76, 54.63, 51.41, 47.02, 44.80,
42.64, 42.5, 38.01, 37.89, 37.60, 34.62, 34.22, 32.09, 31.49,
30.76, 30.53, 30.23, 29.10, 28.54, 27.58, 27.05, 25.42, 22.78,
22.69, 18.93, 17.32, 13.54, 11.98. Anal. Calcd for C49H74O11
C, 70.13; H, 8.89. Found: C, 70.42; H, 8.83.
:
Meth yl 3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[(p-ter t-bu -
t oxym e t h yl)p h e n yl]-7r,12r-b is(for m yloxy)-24-n or -5â-
ch ola n -23-oa te (10b). Organocuprate addition to 9b (5.00
g, 7.6 mmol) as above yielded 10b (5.7 g, 91%): TLC Rf 0.38
(hexane-ether, 1:1); IR (film from CHCl3) 2966, 2876, 1722,
1174 cm-1 1H NMR (300 MHz) δ 8.05 (1H, s), 8.00 (1H, s),
;
7.35, 7.26 (4H, ABq, J ) 8.5 Hz), 5.23 (1H, br s), 5.05 (1H, br
d), 4.38 (2H, s), 4.05 (1H, s), 4.03-3.86 (4H, m), 3.64 (3H, s),
1.28 (9H, s), 1.02 (3H, s), 0.89-0.82 (9H, m), 0.77 (3H, s); 13
C
NMR δ 173.65, 168.17, 168.06, 160.55, 144.53, 137.62, 127.16,
126.71, 74.82, 73.20, 70.55, 66.77, 63.78, 54.6, 51.37, 47.16,
44.92, 42.72, 42.52, 41.03, 38.03, 37.92, 37.64, 34.25, 33.10,
32.11, 31.52, 30.26, 27.61, 27.23, 25.42, 22.79, 22.70, 18.96,
18.91, 18.63, 13.57, 12.04. Anal. Calcd for C48H72O11
69.87; H, 8.80. Found: C, 69.76; H, 8.81.
: C,
Met h yl 3â-[Bis(b u t oxyca r b on yl)m et h yl]-3r-[(p -h y-
d r oxym et h yl)p h en yl]-7r,12r-b is(for m yloxy)-5â-ch ola n -
24-oa te (11a ). To a solution of 10a (5.08 g, 6.04 mmol) in
dry CH2Cl2 (50 mL) under inert atmosphere was added
trifluoroacetic acid (5 mL, 65 mmol). The mixture was heated
at 50 °C. After 5 h a small aliquot was withdrawn and treated
with aqueous alkali.28 Analysis by TLC confirmed the disap-
pearance of the starting material. The reaction mixture was
poured into a biphasic mixture of ether (400 mL) and aqueous
ammonia (100 mL, 4 M) under vigorous stirring. After 5 h
the hydrolysis was complete as evidenced by TLC. The phases
were separated, and the aqueous phase was extracted with
ether (2 × 20 mL). The organic phases were combined, washed
with brine until neutrality, and dried (MgSO4). Evaporation
and flash chromatography with hexane-ether (2:3) as eluent
gave 11a (3.87 g, 82%) as a solid: TLC Rf 0.3 (hexane-ethyl
acetate, 3:2); IR (film from CDCl3) 3528, 2962, 2879, 1721,
1174, 1062, 1018; 1H NMR (300 MHz) δ 8.04 (1H, s), 8.02 (1H,
s), 7.40, 7.31 (4H, ABq, J ) 8.4 Hz), 5.24 (1H, br s), 5.06 (1H,
br s), 4.67 (2H, s), 4.08 (1H, s), 4.04-3.87 (4H, m), 3.64 (3H,
s), 1.04 (3H, s), 0.89-0.80 (9H), 0.73 (3H, s); 13C NMR δ 174.43,
168.13, 168.04, 160.61, 145.08, 138.97, 127.41, 126.05, 75.06,
70.65, 64.95, 64.84, 53.77, 51.47, 47.12, 44.88, 42.83, 42.64,
38.09, 37.88, 37.65, 34.67, 34.29, 31.99, 31.55, 30.83, 30.58,
30.26, 29.33, 28.58, 27.09, 25.56, 22.94, 22.72, 18.98, 17.37,
13.58, 12.06. Anal. Calcd for C45H66O11: C, 69.02; H, 8.49.
Found: C, 68.82; H 8.50.
Meth yl 3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[[[p-(ter t-
bu toxycar bon yl)am in o]m eth yl]ph en yl]-7r,12r-bis(for m y-
loxy)-5â-ch ola n -24-oa te (13a ). To a solution of azide 12a
(3.75 g) in THF (20 mL) were added triphenylphosphine (2.0
g, 7.6 mmol) and water (3 mL). The resulting mixture was
heated to 60 °C for 1 h. Di-tert-butyl dicarbonate (1.47 g, 6.5
mmol) in THF (5 mL) was added, and the reaction mixture
Met h yl 3â-[Bis(b u t oxyca r b on yl)m et h yl]-3r-[(p -h y-
d r oxym et h yl)p h en yl]-7r-12r-b is(for m yloxy)-24-n or -5â-
ch ola n -23-oa te (11b). 10b (5.76 g, 6.98 mmol) was treated
as above with trifluoroacetic acid (5 mL, 65 mmol), followed
by aqueous ammonia, to yield 11b as a white solid (4.8 g,
89%): mp 120-123 °C; TLC Rf 0.25 (hexane-ether, 2:3); IR
(29) The benzyl mesylate (major component) is accompanied by the
corresponding benzyl chloride. Both react in the second step to yield
azide 12a . The second step is difficult to monitor by TLC as the benzyl
chloride and 12a have very similar Rf values.
(28) Cleavage of the tert-butyl ether unit by TFA yields a trifluo-
roacetate which must then be hydrolyzed under basic conditions. The
trifluoroacetate is not readily distinguished by TLC from the starting
ether, necessitating hydrolysis of reaction samples as well as the bulk
product.
(30) CAUTION: when used in chlorinated solvents, this reagent can
lead to explosive side-products. Workers intending to repeat this
preparation should seek an alternative to dichloromethane as solvent.
See ref 11 and: Li, C.; Shih, T.-L.; J eong, J . U.; Arasappan, A.; Fuchs,
P. L. Tetrahedron Lett. 1994, 35, 2645.

8470 J . Org. Chem., Vol. 62, No. 24, 1997
Bhattarai et al.
was cooled to room temperature, sonicated for 30 min, and
then left to stir overnight. The solvent was evaporated and
the residue dissolved in ether (150 mL), washed with water
and then brine, and dried (MgSO4). Evaporation gave a
residue which was dissolved in minimum volume of ether and
cooled (ice-salt bath). Insoluble material (Ph3PO) was re-
moved by filtration. Evaporation of the filtrate gave a residue
which was purified by flash chromatography eluting with
hexane-ethyl acetate (9:1 and then 2:5) to yield 13a (3.21 g,
86% from 11a ): TLC Rf 0.32 (hexane-ethyl acetate, 2:1); IR
(film from CDCl3) 3411, 2963, 2878, 1722, 1515, 1367, 1246,
which on flash chromatography eluting with hexane-ether (1:
1) gave 14a (0.327 g, 56%): TLC Rf 0.3 (hexane-ether, 1:1);
IR (film from CDCl3) 3414, 2964, 2879, 1791, 1755, 1722, 1519,
1469, 1174; δH (300 MHz) δ 8.07 (1H, s), 8.03 (1H, s), 7.37,
7.22 (4H, ABq, J ) 8.4 Hz), 5.26 (1H, br s), 5.07 (1H, br s),
4.87 (1H, br t), 4.30 (2H, d, J ) 5.9 Hz), 4.08 (1H, s), 4.04-
3.86 (4H, m), 1.46 (9H, s), 1.05 (3H, s), 0.89-0.83 (9H, m), 0.77
(3H, s). Anal. Calcd for C55H72NO12F5: C, 63.87; H, 7.02; N,
1.35. Found: C, 63.25; H, 6.96; N, 1.15.
P en t a flu or op h en yl 3â-[Bis(bu t oxyca r bon yl)m et h yl]-
3r-[[[p-(ter t-bu toxyca r bon yl)a m in o]m eth yl]p h en yl]-7r,-
12r-bis(for m yloxy)-24-n or -5â-ch ola n -23-oa te (14b). Meth-
yl ester 13b (0.4 g, 0.46 mmol) was converted as above to the
corresponding carboxylic acid (0.26 g). This was treated as
above with pentafluorophenol (0.062 g, 0.33 mmol) and DCC
(0.07 g, 0.33 mmol) to give 14b (0.262 g, 57%): TLC Rf 0.25
(hexane-ether, 1:1); IR (film from CDCl3) 3414, 2964, 2880,
2455, 1789, 1757, 1722, 1520, 1176 cm-1; 1H NMR (300 MHz)
δ 8.07 (1H, s), 8.03 (1H, s), 7.37, 7.22 (4H, ABq, J ) 8.3 Hz),
5.26 (1H, br s), 5.08 (1H, br s), 4.83 (1H, br t), 4.30 (2H, d),
4.07 (1H, s), 4.04-3.86 (4H, m), 1.05 (3H, s), 1.00 (3H, d, J )
6.4 Hz), 0.90-0.83 (6H, m), 0.81 (3H, s); 13C NMR δ 168.69,
168.07, 167.96, 160.47, 155.85, 144.66, 136.90, 127.40, 126.35,
79.40, 74.73, 70.50, 64.78, 54.33, 46.78, 44.98, 44.07, 42.90,
42.55, 40.10, 38.03, 37.80, 37.62, 34.23, 33.29, 31.97, 31.48,
30.23, 29.27, 28.57, 28.29, 27.27, 25.52, 22.86, 22.64, 18.92,
18.32, 13.50, 11.97. Anal. Calcd for C54H70NO12F5: C, 63.57;
H, 6.92; N, 1.35. Found: C, 62.97; H, 6.84; N, 1.30.
Tetr a k is(for m yloxy) Ch ola p h a n e 4a . Trifluoroacetic
acid (1 mL) was added to a well-stirred solution of 14a (0.327
g, 0.31 mmol) in dry CH2Cl2 (2 mL) cooled to 0 °C under argon.
The mixture was allowed to warm gradually to ambient
temperature. After 4 h, removal of organic volatiles under
high vacuum gave a solid residue. CCl4 (5 mL) was added,
the mixture stirred, and solvent removed under high vacuum.
The above procedure was repeated twice to afford the corre-
sponding ammonium salt as a white solid. This material was
dissolved in dry CH2Cl2 (50 mL) and added dropwise to a well-
stirred solution of DMAP (0.58 g) in dry CH2Cl2 (300 mL) at
ambient temperature under an atmosphere of argon. The
reaction mixture was stirred for 3 days, after which evapora-
tion and flash chromatography, eluting with CHCl3-ethyl
acetate (7:3), gave macrocycle 4a (0.194 g, 77%): TLC Rf 0.36
(CHCl3-ethyl acetate, 7:3); IR (film from CDCl3) 3405, 2961,
2877, 1724, 1662, 1519, 1467, 1178 cm-1; 1H NMR (300 MHz)
δ 8.01 (2H, s), 7.99 (2H, s), 7.37, 7.23 (8H, ABq, J ) 8.5 Hz),
5.60 (2H, t, J ) 5.3 Hz), 5.18 (2H, br s), 5.04 (2H, br s), 4.40,
4.26 (4H, dq, ABX, J AB ) 13.7 Hz, J A/BX ) 5.5 and 5.0 Hz),
4.05 (2H, s), 4.01-3.83 (8H, m, 4 × OCH2), 1.03 (6H, s), 0.87-
0.80 (18H, m), 0.74 (6H, s); 13C NMR δ 172.82, 168.01, 167.93,
160.39, 145.17, 136.27, 127.77, 127.64, 75.45, 70.79, 64.85,
54.31, 45.94, 44.77, 43.55, 42.97, 42.56, 38.04, 37.67, 37.59,
34.29, 34.26, 31.97, 31.93, 31.57, 31.38, 30.25, 29.25, 28.49,
27.26, 25.67, 22.93, 22.67, 18.94, 17.46, 13.56, 11.95. Anal.
Calcd for C88H126N2O18‚4H2O:31 C, 67.24; H, 8.59; N, 1.78.
Found: C, 67.41; H, 8.16; N, 1.59; FABMS m/z 1521, 100%
[M + Na]+.
Tetr a k is(for m yloxy) Ch ola p h a n e 4b. 14b was depro-
tected as above with trifluoroacetic acid to yield the corre-
sponding ammonium salt. This material was treated as above
with DMAP (0.472 g) in dry CH2Cl2 (250 mL) to yield 4b (0.082
g, 44%): TLC Rf 0.4 (CHCl3-ethyl acetate, 7:3); IR (film from
CDCl3) 3410, 2961, 2877, 1754, 1748, 1727, 1718, 1680, 1667,
1514, 1467 cm-1; 1H NMR (300 MHz) δ 8.12 (2H, s), 8.04 (2H,
s), 7.33, 7.13 (8H, ABq, J ) 8.3 Hz), 5.36 (2H, d, J ) 7.4 Hz),
5.11 (2H, br s), 5.01 (2H, br s), 4.80 (2H, ABX, J ) 8.0, 12.9
Hz), 4.02-3.82 (12H, br m), 1.00 (6H, s), 0.88-0.78 (24H, m);
13C NMR δ 171.18, 167.95, 167.84, 161.43, 160.45, 145.98,
135.84, 128.04, 76.50, 71.02, 64.93, 64.83, 55.70, 49.17, 45.25,
44.21, 43.86, 43.74, 42.62, 42.56, 38.13, 38.01, 37.59, 33.84,
32.76, 31.53, 30.37, 29.66, 26.72, 26.19, 22.38, 22.32, 18.98,
1174 cm-1 1H NMR (300 MHz) δ 8.05 (1H, s), 8.02 (1H, s),
;
7.36, 7.22 (4H, ABq, J ) 8.5 Hz), 5.23 (1H, br s), 5.06 (1H, br
s), 4.89 (1H, br t), 4.29 (2H, d, J ) 5.8 Hz), 4.07 (1H, s), 4.04-
3.86 (4H, m), 3.64 (3H, s), 1.46 (9H, s), 1.04 (3H, s), 0.89-0.79
(9H, m), 0.74 (3H, s); 13C NMR δ 174.29, 168.02, 167.92, 160.45,
155.74, 144.63, 136.90, 127.37, 126.32, 79.23, 74.97, 70.55,
64.72, 54.30, 51.35, 47.03, 44.80, 44.05, 42.76, 42.52, 38.01,
37.76, 37.57, 34.59, 34.20, 31.95, 31.46, 30.73, 30.50, 30.19,
29.25, 28.53, 28.28, 27.01, 25.49, 22.86, 22.63, 18.85, 17.20,
13.48, 11.96. Anal. Calcd for C50H75NO12: C, 68.10; H, 8.57,
N, 1.58. Found: C, 67.89; H, 8.61; N, 1.32.
Meth yl 3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[[[p-(ter t-
bu toxycar bon yl)am in o]m eth yl]ph en yl]-7r,12r-bis(for m y-
loxy)-24-n or -5â-ch ola n -23-oa te (13b). 12b (3.9 g) was
treated as above with triphenylphosphine (1.9 g, 7.2 mmol)
and water (3 mL) in THF (20 mL), followed by di-tert-butyl
dicarbonate (1.65 g, 7.3 mmol), to yield 13b (3.41 g, 84% from
11b): TLC Rf 0.44 (hexane-ethyl acetate, 3:2); IR (film from
CDCl3) 3410, 2965, 2879, 1723, 1173, 733 cm-1; 1H NMR (300
MHz) δ 8.05 (1H, s), 8.02 (1H, s), 7.36, 7.22 (4H, ABq, J ) 8.5
Hz), 5.23 (1H, br s), 5.06 (1H, br s), 4.94 (1H, br t), 4.29 (2H,
d, J ) 5.8 Hz), 4.07 (1H, s), 4.04-3.86 (4H, cm), 1.04 (3H, s),
0.89-0.82 (9H, m), 0.78 (3H, s); 13C NMR δ 173.36, 167.94,
167.84, 160.34, 155.68, 144.54, 136.88, 127.29, 126.23, 79.13,
74.76, 70.43, 64.62, 54.25, 51.16, 47.02, 44.82, 43.98, 42.73,
42.45, 40.85, 37.93, 37.70, 37.50, 34.13, 32.93, 31.88, 31.39,
30.10, 29.17, 28.45, 28.20, 27.06, 25.39, 22.76, 22.54, 18.77,
18.48, 13.39, 11.89. Anal. Calcd for C49H73NO12: C, 67.80;
H, 8.47; N, 1.61. Found: C, 67.70; H, 8.53; N, 1.55.
P en t a flu or op h en yl 3â-[Bis(b u t oxyca r bon yl)m et h yl]-
3r-[[[p-(ter t-bu toxyca r bon yl)a m in o]m eth yl]p h en yl]-7r,-
12r-bis(for m yloxy)-5â-ch ola n -24-oa te (14a ). Aqueous so-
dium hydroxide (3 mL, 1.5 M) was added to a stirred solution
of 13a (0.5 g, 0.57 mmol) in THF-methanol (12 mL, 2:1). The
mixture was stirred at room temperature for 24 h and then
neutralized with NaHSO4 (0.400 g, 3.3 mmol). Evaporation
gave a residue which was dissolved in ether, washed thor-
oughly with water then brine, and dried (Na2SO4). Evapora-
tion of the solvent and drying under high vacuum gave crude
dihydroxy acid as a white solid. This was dissolved in formic
acid (5 mL), a catalytic amount of 70% perchloric acid was
added, and the solution was heated at 50 °C (bath tempera-
ture) for 24 h. The mixture was cooled to 40 °C. Acetic
anhydride (2 drops) was added, and the mixture was stirred
for 1 h and then evaporated under high vacuum at ambient
temperature. The residue was dissolved in THF-ether (10
mL, 1:1) and neutralized with a slight excess of 1% aqueous
sodium bicarbonate. A solution of di-tert-butyl dicarbonate
(0.140 g, 0.64 mmol) in ether (2 mL) was added with stirring.
The mixture was left overnight and then diluted with ether
and dried (MgSO4). Evaporation gave a residue which was
purified by flash chromatography eluting with CHCl3-
methanol (99:1 then 98:2) to give 3â-[bis(butoxycarbonyl)-
methyl]-3R-[[[p-(tert-butoxycarbonyl)amino]methyl]phenyl]-
7R,12R-bis(formyloxy)-5â-cholan-24-oic acid (0.361 g): TLC Rf
0.3 (ether). This material was dissolved in CH2Cl2 (3 mL) with
pentafluorophenol (0.084 g, 0.456 mmol) and cooled to -10 °C
under an inert atmosphere. Dicyclohexyl carbodiimide (DCC)
(0.094 g, 0.50 mmol) in CH2Cl2 (2 mL) was added dropwise.
The mixture was left stirring overnight, after which a drop of
distilled water was added and stirring continued under an
open atmosphere for 1 h. The mixture was cooled to -10 °C,
precipitated dicyclohexyl urea was separated by filtration, and
the insoluble material washed with ether-hexane (1:1) pre-
cooled to -10 °C. Evaporation of the filtrate gave a residue
(31) In line with previous experience (refs 8d, 11) analytical samples
of cholaphanes 3 and 4 appeared to be solvated. In all cases the data
was consistent with integral numbers of water molecules, possibly
acquired from atmospheric exposure after drying.

A New Generation of “Cholaphanes”
J . Org. Chem., Vol. 62, No. 24, 1997 8471
17.57, 13.52, 12.19, 12.24. Anal. Calcd for C86H122N2O18‚2H2O:
31 C, 68.50; H, 8.42; N, 1.86. Found: C, 68.67; H, 8.29; N, 1.71.
FABMS m/z 1494, 100% [M + Na]+.
alcohol 16 as a white solid (0.089 g, 97.8%): 1H NMR (80 MHz)
δ 8.13 (1H, s), 8.08 (1H, s), 5.8-5.4 (1H, m), 5.19 (1H, br t),
5.16-4.71 (3H, m, 1H), 3.4 (1H, m), 0.88 (3H, s), 0.74 (3H, s).
A portion of this material (0.040 g, 0.096 mmol) was oxidized
with pyridinium chlorochromate (0.034 g, 0.16 mmol), as
described above for 7a , to yield 17 (0.035 g, 85% overall): IR
(film from CDCl3) 3025, 2963, 2878, 1716, 1642, 1220, 1179
Tetr a h yd r oxy Ch ola p h a n e 3a . To a stirred solution of
4a (0.045 g, 0.028 mmol) in THF (4 mL) was added a 3%
solution of K2CO3 in methanol-water (3 mL, 4:1). The
temperature was raised to 60 °C for 24 h, after which analysis
by TLC showed total consumption of the starting material.
Evaporation gave a residue to which CH2Cl2 and a small
volume of 1% aqueous sodium hydrogen sulfate were added.
The organic phase was washed with water and saturated brine
till neutrality and then dried (Na2SO4). Evaporation gave a
residue which was subjected to flash chromatography eluting
with CHCl3-ethyl acetate-methanol (44:5:1 and then 40:5:5)
to give 3a (0.034 g, 86%): TLC Rf 0.25 (CHCl3-methanol, 9:1);
IR 3340, 3332, 3325, 2963, 2875, 2336, 1755, 1722, 1649, 1535,
cm-1 1H NMR (300 MHz) δ 8.17 (1H, s), 8.09 (1H, s), 5.62
;
(1H, m), 5.30 (1H, br t), 5.15 (1H, br d), 4.88 (2H, m), 3.01
(1H, t, J ) 14 Hz), 1.04 (3H, s), 0.96 (3H, d, J ) 6.6 Hz), 0.81
(3H, s); 13C NMR δ 211.56, 160.29, 160.41, 144.06, 112.4, 74.99,
70.58, 46.87, 45.01, 44.54, 42.89, 42.16, 40.45, 37.73, 36.50,
36.15, 34.44, 31.04, 29.62, 27.22, 25.91, 22.76, 21.54, 19.57,
12.41. Anal. Calcd for C25H36O5: C, 72.08; H, 8.71. Found:
C, 71.80; H, 8.79.
3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[(p-(ter t-bu toxym -
et h yl)p h en yl]-7r,12r-b is(for m yloxy)-24-n or -5â-ch ola n -
22(23)-en e (19). Prepared by treating 1816a (3.98 g, 6.47
mmol) with the organocuprate derived from tert-butyl p-
bromobenzyl ether (4.23 g, 16.5 mmol), magnesium (0.56 g,
23 mmol), and CuCN (0.77 g, 8.59 mmol), as described above
for 10a . Flash chromatography of the crude product eluting
with hexane-ether (9:1, 200 mL; 17:3, 200 mL; 4:1, remaining
fractions) yielded 19 (3.51 g, 70%):32 TLC Rf 0.3 (hexane-ether,
7:3); IR (film from CDCl3) 2967, 2877, 1758, 1724, 1639, 1467,
1
1463, 1415, 754 cm-1; H NMR (300 MHz) δ 7.38, 7.13 (8H,
ABq, J ) 8.3 Hz), 5.73 (2H, br t), 4.39, 4.21 (4H, ABX, J AB
)
13.9, J A/BX ) 5.4 and 4.6 Hz), 4.08 (2H, s), 4.02-3.80 (8H, m,
4 x OCH2), 1.04 (6H, s), 0.93 (6H, d, J ) 6.0 Hz), 0.88-0.81
(12H, m), 0.67 (6H, s); 13C NMR δ 173.19, 168.35, 168.24,
145.48, 135.84, 127.76, 127.45, 72.76, 68.08, 64.76, 54.92,
46.43, 46.36, 43.65, 42.84, 41.80, 39.60, 38.67, 38.33, 35.13,
34.68, 34.54, 32.87, 32.49, 31.45, 30.30, 29.30, 28.35, 27.45,
26.89, 23.03, 22.94, 18.99, 17.40, 13.59, 12.47. Anal. Calcd
for C84H126N2O14‚2H2O:31 C, 70.85; H, 9.20; N, 1.96. Found:
C, 70.96; H, 9.27; N, 1.58. FABMS m/z 1409, 100% [M + Na]+.
Tetr a h yd r oxy Ch ola p h a n e 3b. 4b was treated as above
with K2CO3 in methanol-water. Flash chromatography of the
crude product, eluting with CHCl3-methanol (98:2, then 95:5)
gave 3b (0.03 g, 81%): IR 3415, 2966, 2877, 1755, 1724, 1662
cm-1; 1H NMR (300 MHz) δ 7.35, 7.12 (8H, ABq, J ) 8.4 Hz),
5.37 (2H, br s), 4.5 (2H, brq), 4.14-3.81 (16H, m), 0.97-0.94
(12H, d), 0.84 (12H, q, J ) 7.1 Hz), 0.72 (6H, s); 13C NMR δ
171.95, 168.12, 146.12, 135.56, 128.05, 72.77, 67.95, 65.82,
64.76, 54.94, 48.99, 46.54, 44.96, 43.88, 42.39, 39.34, 38.92,
38.37, 34.59, 34.34, 33.35, 32.66, 30.33, 28.47, 27.14, 22.72,
18.99, 18.05, 15.25, 13.6, 12.52. Anal. Calcd for C82H122N2O14‚-
2H2O:31 C, 70.56; H, 9.10; N, 2.01. Found: C, 70.68; H, 9.05;
N, 1.60. FABMS m/z 1381, 100% [M + Na]+.
1
1382, 1364, 1239, 1188, 1066, 1021, 959, 733 cm-1; H NMR
(300 MHz) δ 8.07 (1H, s), 8.05 (1H, s), 7.35, 7.26 (4H, ABq, J
) 8.5 Hz), 5.59 (1H, m), 5.23 (1H, br s), 5.05 (1H, br s), 4.85
(2H, m), 4.39 (2H, s), 4.05 (1H, s), 3.95 (4H, m), 1.03 (3H, s),
0.86 (3H, d, J ) 6.9 Hz), 0.75 (3H, s); 13C NMR δ 168.16,
168.07, 160.65, 160.55, 144.53, 144.20, 137.65, 127.16, 126.68,
112.17, 74.82, 73.19, 70.62, 64.76, 63.79, 54.70, 46.73, 45.82,
42.70, 42.50, 40.40, 38.11, 37.96, 37.68, 34.29, 32.17, 31.34,
30.28, 29.17, 28.72, 27.62, 27.23, 25.51, 22.82, 22.74, 19.46,
18.97, 13.56, 12.26. Anal. Calcd for C47H70O9: C, 72.46; H,
9.06. Found: C, 72.72; H, 8.88.
3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[p-(h yd r oxym eth -
yl)p h en yl]-7r,12r-bis(for m yloxy)-24-n or -5â-ch ol-22(23)-
en e (20). Prepared from 19 (8.27 g, 10.60 mmol), as described
above for 11a . Flash chromatography of the crude product
eluting with hexane-ethyl acetate (3:2) yielded 20 (7.11 g,
92.7%): TLC Rf 0.3 (hexane-ether, 3:7); IR (film from CHCl3)
3546 (br), 2961, 2876, 1753, 1720, 1176; δH (300 MHz) δ 8.06
(1H, s), 8.01 (1H, s), 7.40, 7.31 (4H, ABq, J ) 8.6 Hz), 5.59
(1H, m), 5.24 (1H, br t), 5.06 (1H, br s), 4.85 (2H, m), 4.67
(2H, s), 4.09 (1H, s), 3.96 (4H, m), 1.05 (3H, s), 0.86 (3H, d, J
) 6.4 Hz), 0.87 (6H, 2 t, J ) 6.7 Hz), 0.76 (3H, s); 13C NMR δ,
168.14, 168.03, 160.63, 160.55, 145.08, 144.16, 138.97, 127.42,
126.05, 112.23, 74.93, 70.66, 64.93, 64.83, 54.38, 46.75, 44.83,
42.86, 42.64, 40.46, 38.10, 37.87, 37.64, 34.31, 32.04, 31.54,
30.28, 29.33, 28.69, 27.24, 25.59, 22.94, 22.71, 19.46, 18.98,
13.58, 12.29. Anal. Calcd for C43H62O9: C, 71.43; H, 8.64.
Found: C, 71.52, H, 8.71.
3â-[Bis(bu toxyca r bon yl)m eth yl]-3r-[p-(a zid om eth yl)-
p h e n yl]-7r,12r-b is(for m yloxy)-24-n or -5â-ch ol-22(23)-
en e (21). Prepared from 20 (0.51 g, 0.705 mmol), methane-
sulfonyl chloride (0.09 g, 61 µL, 0.77 mmol), and N,N-
diisopropylethylamine (0.14 mL, 0.80 mmol), followed by
N,N,N′,N′-tetramethylguanidinium azide (0.22 g, 1.39 mmol),
as described above for 12a . The crude azide obtained (0.535
g) was of sufficient purity for further elaboration. A portion
(0.05 g) was purified by flash chromatography, eluting with
hexane-ether (4:1) to yield 21 (0.04 g): TLC Rf 0.2 (hexane-
ether, 4:1); IR (film from CDCl3) 2963, 2879, 2100, 1756, 1722,
1639, 1516, 1468, 1176 cm-1; 1H NMR (300 MHz) δ 8.07 (1H,
s), 8.03 (1H, s), 7.43, 7.23 (4H, ABq, J ) 8.2 Hz), 5.64-5.52
(1H, m), 5.24 (1H, br s), 5.06 (1H, br s), 4.91-4.79 (2H, m),
4.33 (2H, s), 4.08 (1H, s), 3.99-3.86 (4H, m), 1.05 (3H, s), 0.92-
0.82 (9H, m), 0.76 (3H, s); 13C NMR δ 168.04, 167.94, 160.52,
145.7, 144.11, 133.54, 127.64, 127.07, 112.18, 74.87, 70.58,
3r,7r,12r-Tr is(for m yloxy)-24-n or -5â-ch ol-22(23)-en e (15).
The procedure was closely based on that of Carlson et al.19
A
mixture of cholic acid triformate (6a ) (10.96 g, 22.2 mmol),
cupric acetate monohydrate (0.5 g, 2.5 mmol), and pyridine
(1.2 mL, 14.8 mmol) in benzene (600 mL) under nitrogen was
subjected to azeotropic removal of water for 3 h. During this
period three 20 mL portions of benzene and water were
collected in a Dean-Stark apparatus. The mixture was cooled
to about 40 °C, and lead tetraacetate (40 g, 86 mmol) was
added in one portion. The reaction vessel was then lowered
into an oil bath maintained at 90 °C, and the mixture was
vigorously stirred. Progress of the reaction was monitored by
TLC eluting with hexane-ethyl acetate (7:3). After 4 h, the
mixture was cooled and filtered through a pad of Celite. The
insoluble lead salts remaining on the filter were thoroughly
washed with benzene. The combined filtrates were washed
successively with HCl (aq) and water, dried, and evaporated.
The residue was dissolved in ether and filtered through a pad
of silica, which was then washed with further ether. Filtrate
and washings were combined, concentrated, and recrystallized
from hexane to afford 15 (7.32 g, 76%): mp 185-188 °C (lit.19
mp 186-188 °C); IR 1720, 1710, 1605 cm-1 1H NMR (300
;
MHz) δ 8.17 (1H, s), 8.09 (1H, s), 8.02 (1H, s), 5.61 (1H, m),
5.26 (1H, br t), 5.06 (1H, br d), 4.87 (2H, m), 4.73 (1H, m),
0.95 (3H, s), 0.94 (3H, d, J ) 6.5 Hz), 0.78 (3H, s); 13C NMR δ,
160.42, 144.07, 112.2, 75.01, 73.61, 70.57, 46.76, 44.86, 42.89,
40.71, 40.37, 37.61, 34.43, 34.35, 34.20, 31.23, 28.55, 27.17,
26.47, 25.48, 22.67, 22.23, 19.45, 12.26.
3-Oxo-7r,12r-b is(for m yloxy)-24-n or -5â-ch ol-22(23)-
en e (17). Sodium acetate trihydrate (0.050 g, 0.37 mmol) was
added to a stirred solution of 15 (0.100 g, 0.22 mmol) in
methanol (24 mL). Analysis by TLC, using ethyl acetate as
eluent, showed complete disappearance of starting material
after 96 h. The solvent was evaporated, and the residue was
dissolved in ether, washed with water and then brine, and
dried. Evaporation and drying under high vacuum afforded
(32) A minor slower-running impurity proved to be 3â-[bis(butoxy-
carbonyl)methyl]-3R-[p-(tert-butoxymethyl)phenyl]-7R-formyloxy-12R-
hydroxy-24-nor-5â-cholan-22(23)-ene, presumably arising from de-
formylation of 19. In large-scale work this was reformylated and carried
through to 20.

8472 J . Org. Chem., Vol. 62, No. 24, 1997
Bhattarai et al.
64.82, 54.34, 46.69, 44.78, 42.60, 40.42, 38.04, 37.72, 37.58,
34.23, 31.95, 31.48, 30.19, 29.23, 28.67, 27.20, 25.57, 22.92,
22.66, 19.41, 18.91, 13.60, 12.25. Anal. Calcd for C43H61NO8:
C, 68.92; H, 8.25; N, 5.16. Found: C, 69.05; H, 8.22; N, 5.62.
3â-[Bis(b u t oxyca r b on yl)m et h yl]-3r-[[[p -(ter t-b u t oxy-
ca r bon yl)a m in o]m eth yl]p h en yl]-7r,12r-bis(for m yloxy)-
24-n or -5â-ch ola n -22(23)-en e (22). Triphenylphosphine (2.75
g, 10.5 mmol) was added to a solution of 21 (6.48 g, ca. 8.66
mmol) in THF-water mixture (6:1, 70 mL). The mixture was
heated with stirring at 60 °C. After the completion of the
reduction, as indicated by TLC, the mixture was cooled to
ambient temperature. Di-tert-butyl dicarbonate (2.46 g, 13.06
mmol) and triethylamine (2.4 mL, 17.3 mmol) were added, and
the mixture stirred overnight. The solvent was evaporated
and the residue dissolved in ether (150 mL), washed with
water and then brine, and dried (MgSO4). Evaporation gave
a residue which was dissolved in minimum volume of ether
and cooled (ice-salt bath). Insoluble material (Ph3PO) was
removed by filtration. Evaporation of the filtrate gave a
residue which was purified by flash chromatography, eluting
with hexane-ethyl acetate (9:1 and then 4:1), to yield 22 (6.10
g, 86% from 20): TLC Rf 0.35 (hexane-ethyl acetate, 4:1); IR
(film from CDCl3) 3410, 2965, 2879, 1721, 1517, 1468, 1417,
1245, 1176 cm-1; 1H NMR (300 MHz) δ 8.07 (1H, s), 8.02 (1H,
s), 7.37, 7.22 (4H, ABq, J ) 8.4 Hz), 5.65-5.28 (1H, m), 5.23
(1H, br s), 5.05 (1H, br s), 4.91-4.79 (3H, m), 4.30 (2H, d, J )
5.8 Hz), 4.08 (1H, s), 4.04-3.86 (4H, m), 1.46 (9H, s), 1.04 (3H,
s), 0.97-0.83 (9H, m), 0.76 (3H, s); 13C NMR δ 168.12, 168.02,
160.64, 155.81, 144.71, 144.15, 136.95, 127.45, 126.41, 112.22,
79.37, 74.92, 70.65, 64.82, 54.31, 46.73, 44.81, 44.12, 42.84,
42.57, 40.46, 38.09, 37.81, 37.62, 34.30, 32.01, 31.52, 30.25,
29.31, 28.66, 28.35, 27.24, 26.84, 25.60, 22.95, 22.70, 19.45,
18.96, 13.57, 12.27. Anal. Calcd for C48H71O10N: C, 70.13;
H, 8.70; N, 1.70. Found: C, 69.96; H, 8.78; N, 1.38.
dried (Na2SO4). Evaporation yielded 3â-[bis(butoxycarbonyl)-
methyl]-3R-[[[p-(tert-butoxycarbonyl)amino]methyl]phenyl]-
7R,12R-bis(formyloxy)-23,24-bisnor-5â-cholan-22-al: (TLC Rf
0.80, ether). The crude aldehyde was dissolved in acetoni-
trile-water (3:1, 40 mL) and cooled to -20 °C. Sulfamic acid
(0.250 g, 2.57 mmol) was added with stirring, followed, after
a delay of 5 min, by dropwise addition of a solution of sodium
chlorite (1.33 mmol) in water (10 mL). Stirring was continued
at 0 °C until analysis by TLC showed consumption of the
aldehyde. Sodium metabisulfite (2 g, 10.5 mmol) was added
in one portion and the mixture stirred for 30 min, diluted with
ether, and filtered. Evaporation of filtrate and washings gave
a residue which was dissolved in ether (100 mL), washed with
water and brine, and dried (Na2SO4). Evaporation yielded 3â-
[bis-(butoxycarbonyl)methyl]-3R-[[[p-(tert-butoxycarbonyl)ami-
no]methyl]phenyl]-7R,12R-bis(formyloxy)-23,24-bisnor-5â-cholan-
22-oic acid: (TLC Rf 0.36, ether). A solution of this material
and pentafluorothiophenol (0.36 g, 0.24 mL, 1.74 mmol) in CH2-
Cl2 (10 mL) was cooled to -10 °C under an inert atmosphere
and stirred for 10 min. DCC (0.326 g, 1.6 mmol) in CH2Cl2 (3
mL) was added dropwise. The mixture was left stirring
overnight, after which a drop of water was added and the
mixture stirred for 1 h and then cooled to -10 °C. Precipitated
dicyclohexylurea was separated by filtration and washed with
ether-hexane mixture (1:1) cooled to -10 °C. Evaporation of
the filtrate and washings gave a residue which on flash
chromatography, eluting with hexane-ether (1:1), gave 24
(1.123 g, 85%): TLC Rf 0.4 (hexane-ether, 1:1); IR (film from
CDCl3) 3419, 2968, 2939, 2880, 1757, 1724, 1689, 1516, 1495,
1472, 1176, 983, 962 cm-1; 1H NMR (300 MHz) δ 8.09 (1H, s),
8.02 (1H, s), 7.37, 7.22 (4H, ABq, J ) 8.3 Hz), 5.20 (1H, br s),
5.07 (1H, br s), 4.87 (1H, br t), 4.29 (2H, d, J ) 5.8 Hz), 4.07
(1H, s), 4.04-3.88 (4H, m), 2.68 (1H, m), 1.19 (3H, d, J ) 7.1
Hz), 1.06 (3H, s), 0.88-0.81 (9H, m); 13C NMR δ 194.86, 168.09,
168.01, 160.44, 155.82, 144.65, 137.03, 134.65, 127.44, 126.43,
79.42, 74.25, 70.41, 64.84, 54.42, 51.33, 45.35, 44.62, 44.16,
42.59, 42.38, 38.05, 37.84, 37.69, 34.32, 32.14, 32.07, 31.52,
30.29, 29.31, 28.77, 28.73, 28.36, 26.10, 25.63, 24.65, 22.90,
18.97, 16.92, 13.60, 12.35, 12.30.
3â-[Bis(b u t oxyca r b on yl)m et h yl]-3r-[[[p -(ter t-b u t oxy-
ca r bon yl)a m in o]m eth yl]p h en yl]-7r,12r-bis(for m yloxy)-
24-n or -5â-ch ola n e-22(R,S)-23-d iol (23). To a well-stirred
heterogeneous mixture of pyridine (0.1 mL, 1.2 mmol), N-
methylmorpholine N-oxide (1.51 g, 12.5 mmol), aqueous Bu4-
NOAc (1.4 M, 2.5 mL, 3.5 mmol), and potassium acetate (0.1
g, 1 mmol) in acetone-distilled water mixture (20 mL, 10:1,
v/v) was added a 2% solution of regenerated OsO4 (NaIO4) in
dioxane water mixture (1:1, 7 mL, ca. 0.5 mmol). After 5 min,
a solution of 22 (3.6 g, 4.37 mmol) in acetone-water (20 mL,
10:1) was added. Stirring was maintained for 5 h at room
temperature and then overnight at 40 °C. Analysis by TLC
showed complete consumption of 22. Sodium metabisulfite
(2.7 g, 14.2 mmol) was added after cooling, and the mixture
was stirred for 1 h. The mixture was diluted with ether (50
mL) and dried (Na2SO4) and filtered. Concentration of the
filtrate and washings gave crude 23 (4.1 g, ca. 90%) of
sufficient purity for further elaboration. A portion of the crude
diol was purified by flash chromatography eluting with hex-
ane-ethyl acetate (2:3): TLC Rf 0.17 (ether); IR (film from
CDCl3) 3410, 2963, 2877, 1754, 1721, 1513, 1466, 1414, 1384,
Tetr a k is(for m yloxy) Ch ola p h a n e (4c). Prepared by
treating 24 (1.123 g, 1.10 mmol) with TFA (2 mL) and then
DMAP (3.07 g) in dry CH2Cl2 (1.2 l), as described above for
4a . The cyclodimerization was carried out over a period of 6
days. Flash chromatography of the crude product, eluting with
CHCl3-ethyl acetate (3:2) and drying under high vacuum at
50-55 °C for 4 h, gave 4c (0.550 g, 67%): TLC Rf 0.3 (CHCl3-
ethyl acetate, 3:2); IR (film from CDCl3) 3404, 3300, 2962,
2876, 1753, 1722, 1659, 1517, 1466, 1414, 1384, 1179, 755
cm-1 1H NMR (300 MHz) δ 8.06 (2H, s), 7.88 (2H, s), 7.42,
;
7.23 (8H, ABq, J ) 8.3 Hz), 5.62 (2H, d, J ) 5.6 Hz), 5.15 (2H,
br s), 5.02 (2H, br d), 4.68 (2H, dq, J ) 7.1, 13.8 Hz), 4.07 (2H,
s), 4.03-3.82 (10H, m), 2.45 (2H, br d), 1.04 (6H, d, J ) 6.1
Hz), 1.03 (6H, s), 0.88-0.80 (12H, m), 0.78 (6H, s); 13C NMR
δ 175.29, 167.84, 160.28, 159.96, 145.20, 136.08, 127.86,
127.76, 75.01, 71.02, 64.79, 54.03, 45.17, 44.98, 44.82, 43.76,
43.39, 42.41, 38.09, 37.73, 37.52, 34.11, 31.89, 31.34, 30.23,
29.47, 29.21, 26.42, 25.99, 23.08, 22.67, 18.84, 17.16, 13.48,
12.18. Anal. Calcd for C84H118N2O18‚3H2O:31 C, 67.36; H, 8.34;
N, 1.87. Found: C, 67.13; H, 8.03; N, 1.76. FABMS m/z 1465,
100% [M + Na]+.
1
1367, 1245, 1173, 1063, 755 cm-1; H NMR (300 MHz) δ 8.04
(1H, s), 8.02 (1H, s), 7.35, 7.20 (4H, ABq, J ) 8.3 Hz), 5.19
(1H, br s), 5.06 (1H, br s), 4.93 (1H, br t), 4.28 (2H, d, J ) 5.6
Hz), 4.06 (1H, s), 4.03-3.86 (4H, m), 3.69 (1H, br d), 3.56-
3.36 (2H, br m), 1.04 (3H, s), 0.89-0.80 (9H, m), 0.75 (3H, s);
13C NMR δ 168.04, 167.94, 160.44, 155.81, 144.61, 136.84,
127.33, 126.33, 79.32, 75.02, 73.48, 70.56, 64.74, 62.19, 54.34,
45.13, 44.19, 44.09, 42.56, 42.42, 39.31, 37.98, 37.80, 37.64,
34.19, 31.95, 31.46, 30.19, 29.26, 28.57, 28.28, 26.55, 25.46,
Tetr a h yd r oxy Ch ola p h a n e (3c). Prepared by treatment
of 4c (0.087 g, 0.056 mmol) with K2CO3 in methanol-water,
as described above for 3a . Flash chromatography of the crude
product, eluting with CH2Cl2-methanol (99:1, 98:2, and then
95:5) gave 3b (0.072 g, 90%): TLC Rf 0.25 (CHCl3-methanol,
9:1); IR (film from CDCl3) 3615, 3400, 2966, 2880, 1756, 1724,
1652, 1517, 1468, 1438, 1383, 1144, 1096 cm-1; 1H NMR (300
MHz) δ 7.42, 7.16 (8H, ABq, J ) 8.3 Hz), 5.71 (2H, br d), 4.75
(2H, dq, J ) 7.1, 13.8 Hz), 4.11 (2H, s), 4.01-3.84 (14H, m),
1.17 (6H, d, J ) 6.9 Hz), 1.00 (6H, s), 0.84 (12H, m), 0.69 (6H,
s); 13C NMR δ 176.14, 168.39, 168.29, 145.64, 135.67, 127.65,
127.54, 72.62, 67.98, 64.75, 54.60, 46.61, 45.29, 44.79, 43.57,
42.92, 41.73, 39.47, 38.49, 38.35, 34.67, 34.34, 32.35, 30.23,
29.59, 28.63, 26.81, 26.68, 23.04, 18.94, 18.91, 16.98, 13.57,
12.56. Anal. Calcd for C80H118N2O14‚2H2O:31 C, 70.25; H, 8.99;
22.82, 18.87, 13.46, 12.01, 11.72. Anal. Calcd for C48H73
-
NO12: C, 67.34; H, 8.59; N, 1.63. Found: C, 67.05; H, 8.62;
N, 1.38.
P en t a flu or op h en yl 3â-[Bis(b u t oxyca r bon yl)m et h yl]-
3r-[[[p-(ter t-bu toxyca r bon yl)a m in o]m eth yl]p h en yl]-7r,-
12r-b is(for m yloxy)-23,24-b isn or -5â-ch ola n e-22-t h iola t e
(24). Sodium metaperiodate (0.3 g, ca. 1.4 mmol) was added
to a stirred solution of 23 (1.01 g, 1.18 mmol) in acetonitrile-
water (20 mL, 3:1). After complete oxidation of the diol
(analysis by TLC), the mixture was filtered. Evaporation of
filtrate and washings gave a residue which was dissolved in
ether, washed with sodium bisulfite, water, and brine, and

A New Generation of “Cholaphanes”
J . Org. Chem., Vol. 62, No. 24, 1997 8473
N, 2.05. Found: C, 70.31; H, 8.99; N, 1.93. FABMS m/z 1354,
100% [M + Na]+.
added gently to the “departing arm”. The organic phase was
stirred gently (ca. 60-80 rpm). After 24 h, all three phases
were analyzed by TLC, eluting with ethyl acetate-methanol
(4:1) and visualizing with 2,4-DNP. The analyses were
repeated after 48, 72, and 168 h. 26 was detected at ap-
proximately static levels in the organic phase, and in small
but increasing concentrations in the receiving phase. A similar
experiment in the absence of 3c showed no detectable amount
of glucoside in the “receiving arm” or organic layer.
NMR Titr a tion Stu d ies. Solutions of cholaphanes 3a -c
(1.2-1.8 mM) in CDCl3 were placed in NMR tubes. Solutions
in CDCl3 of octyl â-D-glucopyranoside (25) (70-90 mM) were
added in portions. The resulting downfield shifts of the NH
NMR signals were analyzed using an iterative nonlinear least-
squares curve-fitting program, with weighting of data points
according to the error analysis of Deranleau,33 to give the
binding constants and limiting chemical shifts quoted in the
text. Binding curves are provided as Supporting Information.
Extr a ction Exp er im en ts. Aqueous solutions of methyl-
â-D-glucoside (26) (3 mL, 1-2.5 M) and CHCl3 solutions of
cholaphanes 3a -c and 2 (R ) H) (3 mL, 1 mM) were placed
in sealed flasks and stirred for 24 h in the dark. The two
phases were separated by removing the bulk of the aqueous
layer with a pipet and then filtering the organic layer through
hydrophobic filter paper. The resulting clear organic phase
was analyzed by TLC, spotting with a 20 µL pipet onto glass-
backed plates, and eluting with ethyl acetate-methanol-
water (8:1:1). Visualization with resorcinol/H2SO4 (2-propanol
as solvent) and heating with a heat gun gave dark spots for
the cholaphanes (Rf 0.8-0.95) and for 26 (Rf 0.05-0.1). For
quantitation, extracts were evaporated, redissolved in metha-
nol-CHCl3 (4:1), and compared against standard solutions of
26 spotted from the same solvent mixture.
Ack n ow led gm en t. This work was supported by
Forbairt, the Irish Science and Technology Agency, with
additional funding from the EU Human Capital and
Mobility Program and BioResearch Ireland. We thank
N. Hanly for molecular modeling, Dr. B. Dorgan for
supporting synthetic work, Dr. S. Draper for advice on
crystal growth for X-ray crystallography, Mr. P. Ashton
(University of Birmingham) for mass spectra, and
Freedom Chemical Diamalt GmbH for generous gifts of
cholic acid. K.M.B. thanks the Tribhuvan University,
Amrit Campus, Nepal, for sabbatical leave, and D.J .W.
thanks the Wolfson Foundation for equipment and
computing grants.
Tr a n sp or t Exp er im en t. A CHCl3 solution of cholaphane
6c (6 mL, 2.5 mM) was placed in a “U-tube” transport
apparatus with an extra port allowing access to the organic
phase. The “receiving arm” was charged with distilled water
(2 mL) and a solution of 26 in distilled water (2 mL, 2M) was
Su p p or tin g In for m a tion Ava ila ble: Synthetic proce-
dures for 5, 6a , and 6b; binding curves from NMR titrations
(6 pages). This material is contained in libraries on microfiche,
immediately follows this article in the microfilm version of the
journal, and can be ordered from the ACS; see any current
masthead page for ordering information.
(33) Deranleau, D. A. J . Am. Chem. Soc. 1969, 91, 4044. Connors,
K. A. Binding Constants; Wiley: New York, 1987; p 66.
J O971272W