Total synthesis and antifungal activity of a carbohydrate ring-expanded pyranosyl nucleoside analogue of nikkomycin B

Article abstract of DOI:10.1021/jo701814b

(Chemical Equation Presented) In a study aimed at investigating an as yet unknown structure-activity relationship of the nikkomycin family of antifungal peptidyl nucleoside antibiotics, the present research reports the synthesis and antifungal evaluation

Full text of DOI:10.1021/jo701814b

Total Synthesis and Antifungal Activity of a Carbohydrate
Ring-Expanded Pyranosyl Nucleoside Analogue of Nikkomycin B†
Christina S. Stauffer,‡ Pushpal Bhaket,‡ Annette W. Fothergill,§ Michael G. Rinaldi,§ and
Apurba Datta*,‡
Department of Medicinal Chemistry, The UniVersity of Kansas, 1251 Wescoe Hall DriVe, Lawrence,
Kansas 66045-2500, and Department of Pathology, Fungus Testing Laboratory, UniVersity of Texas
Health Science Center at San Antonio, San Antonio, Texas 78229-3900
adutta@ku.edu
ReceiVed August 17, 2007
In a study aimed at investigating an as yet unknown structure-activity relationship of the nikkomycin
family of antifungal peptidyl nucleoside antibiotics, the present research reports the synthesis and antifungal
evaluation of a carbohydrate ring-expanded pyranosyl nucleoside analogue of nikkomycin B. Employing
a convergent synthetic route, independent synthesis of the N-terminal amino acid side chain and a
stereoselective de novo construction of the desired pyranosyl nucleoside amino acid fragment was followed
by peptidic coupling of the two components, leading to the first synthesis of a carbohydrate ring-enlarged
pyranosyl nikkomycin B analogue. In vitro biological evaluation of the above analogue against a variety
of human pathogenic fungi demonstrated significant antifungal activity against several fungal strains of
clinical significance.
Introduction
(neopolyoxins) (Figure 1), displayed selective and potent
inhibitory activity against fungi such as Pyricularia oryzae and
Rhizoctonia solani.4
The complex peptidyl nucleoside antibiotics are a unique class
of secondary microbial metabolites, with potent antifungal
activity against a wide spectrum of pathogenic fungi.1 Among
the various peptidyl nucleoside antibiotics, polyoxins (Figure
1) were the first family of compounds to be isolated in the 1960s
from Streptomyces cacaoi var. asoensis.2 The polyoxins were
found to exhibit potent antimicrobial activity against various
fungi and find use as efficient agricultural fungicides with no
adverse side effects.3 Subsequently, another class of Strepto-
myces-derived peptidyl nucleoside antibiotics, the nikkomycins
The original interest in the polyoxin and nikkomycin families
of compounds stemmed from their highly selective activity
against various fungi, while being nontoxic to bacteria, plants,
and animals. Acting via a novel mode of action, these natural
products were found to be strong competitive inhibitors of chitin
synthases (Ki range: 0.1-1 µM),2,4 the fungal glycosyl process-
ing enzymes responsible for the formation of chitin, an essential
fungal cell wall component. Chitin, a â-(1f4)-linked polymer
of N-acetylglucosamine, is responsible for imparting shape,
strength, and rigidity to the fungal cell wall.5 Inhibition of chitin
* To whom correspondence should be addressed. Phone: +1 785 864 4117.
Fax: +1 785 864 5326.
† Dedicated to the memory of Dr. C. V. Asokan, a fellow chemist, educator,
and a good friend.
(4) (a) Kobinata, K.; Uramoto, M.; Nishi, M.; Kusakabe, H.; Nakamura,
G.; Isono, K. Agric. Biol. Chem. 1980, 44, 1709-1711. (b) Dahn, U.;
Hagenmeier, H.; Hohne, H.; Konig, W. A.; Wolf, G.; Zahner, H. Arch.
Microbiol. 1976, 107, 143-160.
(5) For reviews, see: (a) Ruiz-Herrera, J.; Ruiz-Medrano, R. Chitin
biosynthesis in fungi. Mycology Ser. 2004, 20, 315-330. (b) Gooday, G.
W. In Biochemistry of Cell Walls and Membranes in Fungi; Kuhn, P. J.,
Trinci, A. P., Jung, M. J., Goosey, M. W., Copping, L. G., Eds.; Springer-
Verlag: Berlin, Germany, 1990; p 61. (c) Cabib, E. AdV. Enzymol. 1987,
59, 59-101.
‡ The University of Kansas.
§ University of Texas Health Science Center at San Antonio.
(1) (a) Isono, K. Current progress on nucleoside antibiotics. Pharmacol.
Ther. 1991, 52, 269-286. (b) Isono, K. J. Antibiot. 1988, 41, 1711-1739.
(2) (a) Isono, K.; Suzuki, S. Heterocycles 1979, 13, 333-351. (b) Isono,
K.; Ashai, K.; Suzuki, S. J. Am. Chem. Soc. 1969, 91, 7490-7505.
(3) Ko, K. Pesticide Chemistry. In Human Welfare and the EnVironment;
Miyato, J., Ed.; Pergamon Press: Elmsford, NY, 1983; p 247.
10.1021/jo701814b CCC: $37.00 © 2007 American Chemical Society
Published on Web 11/15/2007
J. Org. Chem. 2007, 72, 9991-9997
9991

Stauffer et al.
nucleobase component, and replacement of the furanose ring
oxygen with carbon, sulfur, or nitrogen functionalities.9a Sur-
prisingly, thus far there have been no studies investigating the
effect of the carbohydrate ring size of the above nucleosides
on their biological activity. Interestingly, there are several
examples of bioactive complex nucleoside antibiotic natural
products containing a four-membered (“ring contracted”) car-
bohydrate core (e.g., oxetanocins),11 or a six-membered (“ring
expanded”) carbohydrate nucleoside core (e.g., amipurimycin,
miharamycin, blasticidin, gougerotin, etc.).12 The occurrence and
impressive spectrum of biological activities of the above natural
products suggest that size and conformation of the carbohydrate
ring can be potentially important handles toward improving and
“fine-tuning” the activity profile of the peptidyl nucleosides.
Accordingly, as part of a research program directed at the
synthesis, structural modification studies, and biological evalu-
ation of the peptidyl nucleoside antibiotics,13 we report herein
the synthesis and preliminary antifungal evaluation of a hitherto
unreported pyranosyl nucleoside analogue of nikkomycin B (1
in Figure 2).
Results and Discussion
FIGURE 1. Structures of representative polyoxins and nikkomycins.
Our strategy for the synthesis of the desired pyranosyl
nikkomycin B (1) is shown in Figure 2. The plan envisaged a
convergent synthetic route based upon initial stereocontrolled
synthesis of the left-hand side amino acid side chain fragment
2, and the right-hand side R-amino acid nucleoside subunit 3,
followed by peptidic coupling of the above fragments to
construct the complete structural framework of 1 (Figure 2).
A key reaction in the formation of the side chain amino acid
2 envisaged a stereoselective crotylboration reaction between
4-pivaloyloxybenzaldehyde (4) and an appropriate chiral borane
reagent. On the other hand, following our earlier reported
protocol,13b the pivotal pyranosyl nucleoside amino acid 3 could
be synthesized from the D-serine-derived chiral pyranone
intermediate 5.
biosynthesis has been found to cause osmotic sensitivity,
abnormal morphology, and fungal growth arrest, ultimately
leading to cell death. As the chitin biosynthetic pathway is absent
in humans and other mammals,5c,6 chitin synthase and the
cellular mechanisms that regulate the activity of this enzyme
are considered to be excellent targets for pharmaceutical and
agricultural pathogen management.7
With demonstrated in vitro and in vivo activity against various
human pathogenic mycotic infections, the nikkomycin and
polyoxin family of complex peptidyl nucleosides represent
exciting leads in the continuing search for new, effective, and
nontoxic antifungal agents.8 However, direct clinical application
of the peptidyl nucleosides has been compromised by their
attenuated in vivo activity, largely due to their inefficient uptake
into the fungal cell.7 A detailed understanding of the pharma-
cophore of these compounds and the ability to fine-tune
structural features toward optimization of antifungal potency
and pharmacodynamics are therefore critical needs in accelerat-
ing the transfer of these promising antifungal peptidyl nucleoside
lead compounds into clinically useful therapeutic agents.
In view of their exciting antifungal potential, nikkomycins
and polyoxins continue to be the focus of total syntheses and
structure-activity relationship investigations.9,10 Most of the
reported SAR studies on these compounds have, however, been
directed at modification of the amino acid side chain, the
Accordingly, following Barrett’s procedure,14 a highly enan-
tio- and diastereoselective addition of (-)-E-crotyldiisopi-
(9) For reviews, see: (a) Zhang, D.; Miller, M. J. Curr. Pharm. Des.
1999, 5, 73-99 and references cited therein. (b) Garner, P. Synthetic
approaches to complex nucleoside antibiotics. In Studies in Natural Products
Chemistr; Atta-ur-Rahman, Ed.; Elsevier: Amsterdam, The Netherlands,
1988; Stereoselective synthesis (Part A), Vol. 1, pp 397-435. (c) Knapp,
S. Chem. ReV. 1995, 95, 1859-1876 and references cited therein.
(10) For some recent reports, see: (a) Li, F.; Brogan, J. B.; Gage, J. L.;
Zhang, D.; Miller, M. J. J. Org. Chem. 2004, 69, 4538-4540. (b) Suda,
A.; Ohta, A.; Sudoh, M.; Tsukuda, T.; Shimma, N. Heterocycles 2001, 55,
1023-1028. (c) Obi, K.; Uda, J.-I.; Iwase, K.; Sugimoto, O.; Ebisu, H.;
Matsuda, A. Bioorg. Med. Chem. Lett. 2000, 10, 1451-1454. (d) Uda, J.-
I.; Obi, K.; Iwase, K.; Sugimoto, O.; Ebisu, H.; Matsuda, A. Nucleic Acids
Symp. Ser. 1999, 42, 13-14.
(6) For reviews, see: (a) Bulawa, C. E. Annu. ReV. Microbiol. 1993, 47,
505-534. (b) Cabib, E.; Shematek, E. M. In Biology of Carbohydrates;
Ginsburg, V., Robius, P. W., Eds.; John Wiley and Sons: New York, 1981;
Vol. I, p 51.
(11) (a) Hoshino, H.; Shimizu, N.; Shimada, N.; Takita, T.; Takeuchi,
T. J. Antibiot. 1987, 40, 1077-1078 and references cited therein. (b)
Shimada, N.; Hasegawa, S.; Harada, T.; Tomisawa, T.; Fujii, A.; Takita,
T. J. Antibiot. 1986, 39, 1623-1625.
(12) (a) Seto, H.; Koyama, M.; Ogino, H.; Tsuruoka, T.; Inouye, S.;
Otake, N. Tetrahedron Lett. 1983, 24, 1805-1808. (b) Iwasa, T.; Kishi,
T.; Matsuura, K.; Wakae, O. J. Antibiot. 1977, 30, 1-10. (c) Kanzaki, T.;
Higashide, E.; Yamamoto, H.; Shibata, M.; Nakazawa, K.; Iwasaki, H.;
Takewaka, T.; Miyake, A. J. Antibiot., Ser. A 1962, 15, 93-97. (d) Takeuchi,
S.; Hirayama, K.; Ueda, K.; Sakai, H.; Yonehara, H. J. Antibiot., Ser. A
1958, 11, 1-5.
(13) (a) Khalaf, J. K.; Datta, A. J. Org. Chem. 2005, 70, 6937-6940.
(b) Bhaket, P.; Stauffer, C. S.; Datta, A. J. Org. Chem. 2004, 69, 8594-
8601.
(7) For reviews, see: (a) Behr, J. B. Curr. Med. Chem.: Anti-Infect.
Agents 2003, 2, 173-189 and references cited therein. (b) Ruiz-Herrera,
J.; San-Blas, G. Chitin synthesis as a target for antifungal drugs. Curr. Drug
Targets: Infect. Disord. 2003, 3, 77-91 and references cited therein.
(8) (a) Anderson, Mark, B.; Roemer, T.; Fabrey, R. Progress in antifungal
drug discovery. Ann. Rep. Med. Chem. Doherty, A. M., Ed.; ElseVier-
Academic Press: New York, 2003, 38, 163-172 and references cited therein.
(b) Watkins, W. J.; Renau, T. E. Progress with antifungal agents and
approaches to combat fungal resistance. Annu. Rep. Med. Chem. Doherty,
A. M. Ed.; Academic Press: New York, 2000, 35, 157-166 and references
cited therein. (c) Andriole, V. T. Int. J. Antimicrob. Agents 2000, 16, 317-
321.
(14) Barrett, A. G. M.; Lebold, S. A. J. Org. Chem. 1991, 56, 4875-
4884.
9992 J. Org. Chem., Vol. 72, No. 26, 2007

Carbohydrate Ring-Enlarged Pyranosyl Nikkomycin B Analogue
interactions between the bulky dihydroxylating reagent and the
â-configured substituent at the benzylic stereocenter, situated
two carbons away from the olefinic reactive site. Interestingly,
dihydroxylation of 8 with osmium tetroxide (OsO4) under
standard conditions afforded the required 2,4-syn-diol 9 with
somewhat improved selectivity (4:1, by 1H NMR). It is assumed
that, compared to AD-mix-â, the relatively smaller size of OsO4
probably reduces the steric interaction with the distal benzylic
stereocenter, resulting in the observed improvement in selectiv-
ity.
Employing Rychnovsky’s 13C NMR method for stereochem-
ical determination of 1,3-diol systems,16 the assigned stereo-
chemistry of the 1,3-syn- and 1,3-anti-diols 9 and 10 was con-
firmed via analysis of the 13C NMR spectra of the corresponding
acetonide derivatives 9A and 10A, respectively (Figure 3).
FIGURE 2. Pyranosyl nikkomycin B (1): retrosynthetic strategy and
approach.
FIGURE 3. Diagnostic 13C NMR shifts of the acetonides derived from
the diols 9 and 10.
nocampheylborane to 4-pivaloyloxybenzaldehyde 7 (Scheme 1)
and subsequent protection of the resulting secondary hydroxy
group as its TBS-ether resulted in the expected homoallylic
alcohol derivative 8 (Scheme 1) as the only product. NMR (1H
and 13C) spectral studies and HPLC analysis of 8 confirmed its
structural and stereochemical integrity.
Selective TBS-protection of the primary hydroxy group
allowed easy chromatographic separation of the product mixture,
affording the required triol derivative 11 in good overall yield
(Scheme 2). Following a standard Mitsunobu procedure,17 the
free hydroxy group of 11 was converted to the corresponding
SCHEME 1
SCHEME 2
Toward introduction of the required R-amino acid functional-
ity as present in the nikkomycin B side chain, stereoselective
dihydroxylation of the terminal olefinic moiety of 8 was next
investigated. Disappointingly, attempted Sharpless asymmetric
dihydroxylation of 8 with AD-mix-â (a matched pair)15 resulted
in a mixture of the expected 2,4-syn-diol 9 along with the
unwanted 2,4-anti-diol 10 with only a moderate selectivity (3:
1
1, by H NMR) in favor of 9.
In retrospect, the unsatisfactory selectivity in the above
reaction can probably be attributed to possible unfavorable steric
(16) Rychnovsky, S. D.; Rogers, B.; Yang, G. J. Org. Chem. 1993, 58,
3511-3515.
(17) (a) Pearson, W. H.; Hines, J. V. J. Org. Chem. 1989, 54, 4235-
4237. (b) Lal, B.; Pramanik, B. N.; Manhas, M. S.; Bose, A. K. Tetrahedron
Lett. 1977, 1977-1980.
(15) Johnson, R. A.; Sharpless, K. B. Catalytic asymmetric dihydroxy-
lation-discovery and development. In Catalytic Asymmetric Synthesis, 2nd
ed.; Ojima, I., Ed.; Wiley-VCH: New York, 2000; pp 357-398.
J. Org. Chem, Vol. 72, No. 26, 2007 9993

Stauffer et al.
azide 12, with concomitant inversion of the stereocenter
involved. Reduction of the azide to amine and protection of
the resulting amine uneventfully afforded the corresponding
carbamate derivative 13 in high overall yields. Camphorsulfonic
acid assisted selective deprotection at the primary silylether
linkage18 to generate 14, followed by oxidation of the resulting
free hydroxy group to carboxylic acid culminated in an efficient
route to the orthogonally protected nikkomycin B side chain
N-terminal amino acid derivative 2. It is worth mentioning that,
in terms of brevity and efficiency, the above synthesis of the
strategically protected nikkomycin B side chain (2) compares
well with the other literature reported methods,9a and is expected
to be an attractive alternative for the synthesis of this unusual
amino acid and related analogues.
Biological Evaluation
To evaluate the effect of the above carbohydrate ring
enlargement on the antifungal activity of nikkomycins, in vitro
susceptibility testing of the pyranosyl analogue 19 against
clinical isolates of five different fungi, Candida albicans,
Candida glabrata, Aspergillus fumigatus, Cryptococcus neo-
formans, Coccidioides immitis, and Blastomyces dermatitidis,
were performed. The antifungal drug amphotericin B (Ampho-
B) and commercially available nikkomycin Z (Nik-Z) (Sigma)
were used as reference standards in these antifungal assays. The
results (MIC) of the biological evaluation are given in Table 1.
The MIC was designated as the lowest concentration where
visible growth was absent.
TABLE 1. Antifungal Activity (MIC) of Pyranosyl Nikkomycin B
(19)
Following our earlier reported method, a de novo synthesis
of the fully protected right-hand side pyranosyl nucleoside amino
acid fragment 15 (Scheme 3) was achieved utilizing D-serine
as a chiral starting material.13b Subsequent hydrogenolytic
removal of the Cbz-functionality generated the free amine 3. A
BOP-Cl mediated peptidic coupling between 3 and the side chain
amino acid 2 provided the fully protected pyranosyl nikkomycin
B analogue 16.
pyranosyl
Nik-B (19)
[MIC ) µg/mL]
Nik-Z
[MIC )
µg/mL]
Ampho-B
[MIC )
µg/mL]
fungal species
(no. of isolates)
C. albicans (3)
C. glabrata (2)
A. fumigatus (3)
C. neoformans (1)
C. immitis (1)
>16
>16
4, 8, and >16
>16
0.125
0.125 and 0.25
0.25-0.5
0.25
>16
2-4
e0.03
0.25
e0.03
e0.03
0.5
0.25
0.25
B. dermatitidis (1)
0.03
SCHEME 3
As evident, the pyranosyl nikkomycin B analogue 19 was
found to be inactive against Candida albicans, Candida gla-
brata, and Aspergillus fumigatus. This is not entirely unexpected,
as nikkomycin Z (the most potent among the natural nikkomy-
cins) itself is not a very efficient antifungal agent against most
strains of Candida and Aspergillus. Gratifyingly, against human
pathogenic fungal strains of Cryptococcus neoformans and
Coccidioides immitis, the analogue 19 exhibited strong inhibitory
activity. Thus, while the antifungal activity of 19 against
Cryptococcus neoformans was equipotent to that of nikkomycin
Z (and significantly better than that of amphotericin B), the MIC
of 19 against Coccidioides immitis was much lower than both
the references amphotericin B and nikkomycin Z. Although not
as active as amphotericin B or nikkomycin Z, the pyranosyl
analogue 19 also displayed some inhibitory activity against
Blastomyces dermatitidis.
It is worth mentioning that, although relatively less common,
fungal infections caused by C. neoformans (cryptococcosis), C.
immitis (coccidiodomycosis), and B. dermatitidis (blastomyco-
sis) are nonetheless pathogenic systemic (lungs, brain, bone,
GI tract, etc) fungal infections of increasingly serious concern.
Interestingly, while Amphotericin B is a commonly used agent
for the treatment of these infections, in our antifungal assay,
involving clinical isolates of the above human pathogenic strains,
the newly synthesized pyranosyl nik-B analogue 19 was found
to be about 10 times more active than the Amphotericin B
standard. We believe that this is a significant observation and
can provide important design parameters toward future optimi-
zation of the nikkomycin structural lead.
In conclusion, the present total synthesis and structure-
activity relationship study aimed at a previously unexplored
structural component of the natural nikkomycins indicate that,
carbohydrate core ring-enlargement of the nikkomycin family
of compounds could be a potentially beneficial strategy toward
optimizing the antifungal potency of the above peptidyl nucleo-
sides. Studies are currently underway toward the synthesis of
further modified analogues based on the above pyranosyl
nucleoside scaffold and eventual antifungal evaluation of these
Standard removal of the silyl protecting group to form 18,
followed by hydrolytic cleavage of the four ester functionalities
yielded the polyhydroxylated carboxylic acid derivative 18.
Finally, hydrogenolysis of the amine protecting group followed
by crystallization induced purification of the resulting product
via oxalate salt formation culminated in the desired carbohydrate
ring expanded nikkomycin B pyranosyl analogue 19.
(18) Nelson, T. D.; Crouch, R. D. Synthesis 1996, 1031-1069.
9994 J. Org. Chem., Vol. 72, No. 26, 2007

Carbohydrate Ring-Enlarged Pyranosyl Nikkomycin B Analogue
141.4, 150.3, 177.5; HRMS calcd for C22H40NO3Si m/z (M + NH4)+
394.2777, found 394.2797. HPLC: CHIRALPAK AD-RH, 95%
MeOH/H2O, 0.8 mL/min, 254 nm, retention time ) 4.55 min
(>95%).
analogues against various pathogenic fungi. Results from the
above studies are expected to provide a better understanding of
the role of the carbohydrate core ring size in the biological
activity of the nikkomycin family of antifungal antibiotics. We
hope that continued investigation of the pyranosyl nikkomycin
analogues will help us fully explore and optimize the structural
features of the nikkomycins family of novel lead compounds
toward the potential development of a new class of therapeuti-
cally useful antifungal agents.
(1S,2S,3R)-1-[(tert-Butyldimethylsilyl)oxy]-2-methyl-1-[4-(piv-
aloyloxy)phenyl]butane-3,4-diol (9) and (1S,2S,3S)-1-[(tert-Bu-
tyldimethylsilyl)oxy]-2-methyl-1-[4-(pivaloyloxy)phenyl]butane-
3,4-diol (10). To an ice-cooled, well-stirred solution of the olefin
8 (4 g, 10.6 mmol) and 4-methylmorpholine N-oxide (3.7 g, 31.6
mmol) in acetone (80 mL) and H2O (20 mL) was added a solution
of OsO4 (4 mL, 5% solution in toluene) dropwise. The resulting
solution was stirred at 0 °C for 12 h. The reaction was quenched
by the addition of saturated aqueous Na2SO3 solution (15 mL),
diluted by EtOAc (100 mL), and stirred at room temperature for
30 min. The two layers were separated and the aqueous layer was
extracted with EtOAc (3 × 20 mL). The combined organic extracts
were dried over anhydrous Na2SO4 and the solvent removed in
vacuo. The residue was purified by flash column chromatography
(hexanes/EtOAc ) 4:1) to yield a mixture of the diols 9 and 10
Experimental Section
4-Pivaloyloxybenzaldehyde (7). 4-Hydroxybenzaldehyde 6 (12
g, 98.8 mmol) was dissolved in anhydrous CH2Cl2 (120 mL),
followed by addition of anhydrous pyridine (13.5 mL, 167 mmol)
and a catalytic amount of DMAP. The reaction mixture was cooled
to 0 °C, and freshly distilled pivaloyl chloride (18.5 mL, 150 mmol)
was added dropwise with stirring. After completion of the addition,
the reaction mixture was brought to room temperature and stirred
for another hour. The reaction was quenched by the addition of
ice-cold water (100 mL) and stirred for 5 min. The two layers were
separated and the aqueous layer was extracted with CH2Cl2 (3 ×
20 mL). The combined organic extracts were dried over anhydrous
Na2SO4 and the solvent was removed in vacuo. The residual oil
was purified by flash chromatography (hexane/EtOAc ) 19:1) to
yield 4-pivaloyloxybenzaldehyde (7) as a low melting solid
1
(4:1 by H NMR) as viscous oil (4 g, 91%).
Diols 9 and 10: IR (neat) 3430, 1752, cm-1; 1H NMR (400 MHz,
CDCl3, diastereomeric mixture) δ -0.26 and -0.19 (2s, 3H), 0.06
and 0.09 (2s, 3H), 0.65 and 0.99 (2d, J ) 8, 3H), 0.89 and 0.93
(2s, 9H), 1.37 (s, 9H), 1.82 and 2.0 (2 m, 1H), 2.2 (br t, 1H), 3.56-
3.93 (m, 3H), 4.31 and 4.42 (2s, 1H), 4.60 and 4.78 (2d, J ) 8 Hz,
1H), 7.03 (d, J ) 6.7 Hz, 2H), 7.33 (d, J ) 6.6 Hz, 2H); 13C NMR
(100.6 MHz, CDCl3, diastereoisomeric mixture) δ -4.8, -4.7,
-4.1, -3.9, 12.3, 12.4, 18.4, 26.1, 26.2, 27.5, 39.5, 42.7, 44.0,
64.9, 65.6, 71.6, 75.4, 80.1, 80.6, 121.3, 121.5, 121.6, 127.6, 128.6,
140.6, 140.9, 150.9, 177.5; HRMS calcd for C22H42NO5Si m/z (M
+ NH4)+ 428.2832, found 428.2859.
1
(21.0 g, 92%): IR (neat) 1752, 1680 cm-1; H NMR (400 MHz,
CDCl3) δ 1.38 (s, 9H), 7.27 (d, J ) 6.0 Hz, 2H), 7.92 (d, J ) 6.0
Hz, 2H), 10.0 (s, 1H); 13C NMR (100.6 MHz, CDCl3) δ 26.9, 39.1,
122.2, 131.0, 133.7, 155.8, 176.2, 190.8. HRMS calcd for C12H15O3
m/z (M + H)+ 207.1021, found 207.1012.
(1S,2S,3R)-1,4-Bis[(tert-butyldimethylsilyl)oxy]-2-methyl-1-[4-
(pivaloyloxy)phenyl]butane-3-ol (11) and (1S,2S,3S)-1,4-Bis[(tert-
butyldimethylsilyl)oxy]-2-methyl-1-[4-(pivaloyloxy)phenyl]butane-
3-ol (minor isomer). A solution of the mixture of the diols 9 and
10 (4.0 g, 9.8 mmol), imidazole (1.0 g, 14.7 mmol), a catalytic
amount of DMAP, and TBDMSCl (1.8 g, 11.9 mmol) in anhydrous
CH2Cl2 (40 mL) was stirred at room temperature for 12 h. Upon
completion of the reaction (tlc monitoring), ice-cold water (100
mL) was added to the reaction mixture. The two layers were
separated and the aqueous layer was extracted with CH2Cl2 (3 ×
25 mL). The combined organic extracts were dried (Na2SO4) and
concentrated. Purification of the residue by flash column chroma-
tography (hexanes/EtOAc ) 97:3) yielded the required alcohol
isomer 11 as a colorless oil (3.94 g, 77%): [R]D -12.6 (c 1.02,
CHCl3); IR (neat) 3462, 1745 cm-1; 1H NMR (400 MHz, CDCl3)
δ -0.15 (s, 3H), 0.05, 0.08 and 0.11 (3s, 9H), 0.70 (d, J ) 10.4
Hz, 3H), 0.91 (s, 18 H), 1.37 (s, 9H), 2.02 (m, 1H), 2.97 (d, J )
3.0 Hz, 1H), 3.33 (m, 1H), 3.50 (dd, J ) 7, 10 Hz, 1H), 3.69 (dd,
J ) 3.2, 10 Hz, 1H), 5.01 (d, J ) 5.4 Hz, 1H), 7.02 (d, J ) 9.0
Hz, 2H), 7.35 (d, J ) 8.5 Hz, 2H); 13C NMR (100.6 MHz, CDCl3)
δ -5.0, -4.9, -4.7, -4.2, 10.4, 18.5, 18.7, 26.2, 26.3, 27.5, 39.5,
44.2, 65.5, 73.1, 75.5, 121.0, 128.4, 140.0, 150.4, 177.6; HRMS
calcd for C28H52O5Si2Li m/z (M + Li)+ 531.3513, found 531.3521.
Minor alcohol isomer. Colorless oil (0.99 g, 19%): [R]D -30.6
(3S,4S)-4-[(tert-Butyldimethylsilyl)oxy]-3-methyl-4-[4-(piv-
aloyloxy) phenyl]butene (8). Step 1: To a well-stirred solution of
t-BuOK (1 M solution in THF, 38 mL, 38 mmol) at - 78 °C was
added trans-2-butene (7 mL, 74.7 mmol), followed by dropwise
addition of n-BuLi (1.6 M in hexane, 24 mL, 38 mmol). After
completion of addition, the reaction mixture was warmed to
-40 °C, stirred for 30 min, then recooled to -78 °C, then (-)-B-
methoxydiisopinocampheyl-borane (1 M in ether, 44.5 mL, 44.5
mmol) was added to it dropwise. After stirring at -78 °C for 30
min, BF3‚OEt2 (6.4 mL, 51 mmol) was added to the reaction
mixture, followed by the slow addition of a solution of the aldehyde
7 (5 g, 21.2 mmol) in anhydrous ether (30 mL). After stirring at
the same temperature for another 4.5 h, the reaction was quenched
by careful addition of MeOH (3 mL) and the mixture was allowed
to attain room temperature. After removal of the volatile compo-
nents in vacuo, the residue was dissolved in MeOH (50 mL), and
8-hydroxyquinoline (7 g) was added to the solution. After stirring
at room temperature for 12 h, the precipitated 8-HQ-B(Ipc)2 crystals
were filtered off and the filtrate was concentrated to afford the crude
adduct as a thick syrup along with some pinene-derived impurities.
Step 2: The crude product obtained from step 1 above was
dissolved in anhydrous DMF (20 mL), followed by the addition of
imidazole (3.5 g, 51.4 mmol), TBDMSCl (5 g, 31.1 mmol), and a
catalytic amount of DMAP. The mixture was stirred at 60 °C for
12 h, cooled to room temperature, and diluted by the addition of
ice-cold water (30 mL) and ether (100 mL). The two layers were
separated and the aqueous layer was extracted with ether (3 × 25
mL). The combined organic extracts were dried over anhydrous
Na2SO4, the solvent was removed under vacuum, and the residue
was purified by flash column chromatography (hexanes/EtOAc )
97:3) to provide the TBS-protected olefin 8 as a viscous oil (6.8 g,
84% over two steps): [R]D -36.7 (c 0.96, CHCl3); IR (neat) 1752
cm-1; 1H NMR (400 MHz, CDCl3) δ -0.18 (s, 3H), 0.03 (s, 3H),
0.90 (m, 12H), 1.33 (s, 9H), 2.42 (m, 1H), 4.48 (d, J ) 5.8 Hz,
1H), 4.97 (m, 2H), 5.82 (m, 1H), 7.01 (d, J ) 8.5 Hz, 2H), 7.27
(d, J ) 8.5 Hz, 2H); 13C NMR (100.6 MHz, CDCl3) δ -4.6, -4.2,
16.4, 18.6, 26.2, 27.6, 39.5, 46.8, 78.9, 115.1, 121.0, 128.0, 141.1,
1
(c 1.00, CHCl3); H NMR (400 MHz, CDCl3) δ -0.19 (s, 3H),
0.02 (s, 3H), 0.04 (s, 3H), 0.08 (s, 3H), 0.86 and 0.91 (2 br s, 21H),
1.38 (s, 9H), 1.59-1.83 (m, 1H), 3.00 (d, J ) 2.0 Hz, 1H), 3.51-
3.57 (m, 2H), 4.01 (t, J ) 6.5 Hz, 1H), 4.74 (d, J ) 6.0 Hz, 1H),
7.03 (d, J ) 8.5 Hz, 2H), 7.31 (d, J ) 8.5 Hz, 2H); 13C NMR
(100.6 MHz, CHCl3) δ -5.0, -4.9, -4.8, -4.2, 11.0, 18.5, 18.6,
26.2, 26.3, 27.5, 39.5, 42.4, 65.6, 70.5, 79.0, 121.4, 127.8, 141.5,
150.5, 177.4.
(1S,2S,3S)-3-Azido-1,4-bis[(tert-butyldimethylsilyl)oxy]-2-meth-
yl-1-[4-(pivaloyloxy)phenyl]butane (12). To a well-stirred solution
of alcohol 11 (0.10 g, 0.2 mmol) in dry THF (1 mL) at 0 °C was
added diisopropyl azodicarboxylate (0.05 mL, 0.25 mmol) and
triphenylphosphine (0.063 g, 0.24 mmol), followed by dropwise
addition of diphenylphosphoryl azide (0.05 mL, 0.23 mmol). After
J. Org. Chem, Vol. 72, No. 26, 2007 9995

Stauffer et al.
completion of addition, the reaction mixture was brought to room
temperature and stirred for 1 h. Upon completion of the reaction
(monitored by tlc), excess solvent was removed under vacuum and
the residue was purified by flash chromatography (hexanes/EtOAc
) 98:2) to obtain the azide 12 as an oil (0.096 g, 92%): [R]D -22.5
(c 1.04, CHCl3); IR (neat) 2094, 1752 cm-1; 1H NMR (400 MHz,
CDCl3) δ -0.29 (s, 3H), 0.10, 0.11, and 0.12 (3s, 9H), 0.49 (d, J
) 6.8 Hz, 3H), 0.89 (s, 9H), 0.94 (s, 9H), 1.37 (s, 9H), 1.78 (m,
1H), 3.71 (dd, J ) 4.4, 10.4 Hz, 1H), 3.82 (dd, J ) 8.9, 10.2 Hz,
1H), 4.21 (m, 1H), 4.41 (d, J ) 9.2 Hz, 1H), 7.05 (d, J ) 8.3 Hz,
2H), 7.29 (d, J ) 8.4 Hz, 2H); 13C NMR (100.6 MHz, CDCl3) δ
-5.1, -5.0, -4.9, -4.0, 10.6, 18.5, 18.6, 26.2, 26.3, 27.5, 39.5,
42.7, 64.0, 66.5, 76.8, 121.6, 128.5, 141.5, 150.8, 177.5; HRMS
calcd for C28H55N4O4Si2 m/z (M + NH4)+ 567.3762, found
567.3780. HPLC: IProSIL 120-5 Si 5.0 µm, 98% hexanes/2%
EtOAc, 1.0 mL/min, 254 nm, retention time ) 6.23 min.
(1S,2S,3S)-3-[(Benzyloxycarbonyl)amino]-1,4-bis[(tert-butyldi-
methylsilyl)oxy]-2-methyl-1-[4-(pivaloyloxy)phenyl]butane (13).
Step 1: To a well-stirred room temperature solution of the azide
12 (5.3 g, 9.65 mmol) in anhydrous EtOAc (50 mL) was added
10% palladium on activated carbon (1 g) and the mixture was stirred
under H2 atmosphere for 4 h. After completion of the reaction (tlc
monitoring), the catalyst was filtered off and washed thoroughly
with EtOAc. The combined filtrate was concentrated under vacuum
and then dried under high vacuum for 1 h to yield the crude amine
as a light yellow oil.
66.0, 67.3, 78.6, 121.4, 121.7 127.6, 127.9, 128.5, 128.6, 128.7,
128.8, 128.9, 136.8, 140.5, 150.8, 177.4; HRMS calcd for C30H45-
NO6SiNa m/z (M + Na)+ 566.2914, found 566.2772.
(2S,3S,4S)-2-[(Benzyloxycarbonyl)amino]-4-[(tert-butyldi-
methylsilyl)oxy]-3-methyl-4-[4-(pivaloyloxy)phenyl]butyric Acid
(2). To a stirring, room temperature solution of CH3CN/CCl4/H2O
(1.5:0.5:1, 3 mL) was added NaIO4 (0.08 g, 0.36 mmol) and RuCl3‚
3H2O (0.005 g, 0.018 mmol) sequentially. The resulting mixture
was stirred for 30 min, after which it was added into a solution of
the alcohol 14 (0.1 g, 0.18 mmol) dissolved in CH3CN (3 mL). To
the resulting dark solution an additional lot of NaIO4 (0.04 g, 0.18
mmol) was added and the mixture was stirred at room temperature
for 30 min. The inorganic solids were removed by filtration through
a celite pad, washing thoroughly with EtOAc. The combined filtrate
was concentrated and the residue purified by flash chromatography
(hexanes/EtOAc ) 6:4) to obtain the carboxylic acid 2 as a foamy
white solid (0.079 g, 77%): mp 71-74 °C; [R]D -20.0 (c 0.50,
1
CHCl3); IR (neat) 3308, 1746, 1726 cm-1; H NMR (500 MHz,
CDCl3) δ -0.33 (s, 3H), 0.03 (s, 3H), 0.75 (d, J ) 6.8 Hz, 3H),
0.88 (s, 9H), 1.38 (s, 9H), 2.41 (m, 1H), 4.5 (d, J ) 8.2 Hz, 1H),
4.71 (br s, 1H), 5.12 (d, J ) 12.2 Hz, 1H), 5.23 (d, J ) 12.2 Hz,
1H), 5.67 (br d, J ) 8.1 Hz, 1H), 7.03 (d, J ) 8.2 Hz, 2H), 7.28
(d, J ) 7.8 Hz, 2H), 7.49 (m, 5H), 9.27 (br s, 1H); 13C NMR (125.7
MHz, CDCl3) δ -5.5, -4.6, 12.3, 17.9, 25.7, 27.0, 38.9, 42.9, 55.0,
67.0, 76.7, 121.3, 127.6, 128.0, 128.1, 128.4, 136.2, 140.0, 150.5,
156.5, 176.9, 177.2; HRMS calcd for C30H47N2O7Si m/z (M +
NH4)+ 575.3153, found 575.3127.
Synthesis of the Fully Protected Pyranosyl Nikkomycin B
(16). Step 1: To a solution of the nucleoside amino acid derivative
15 (0.62 g, 1.16 mmol)13b in anhydrous EtOAc (10 mL) was added
10% palladium on activated carbon (0.7 g), and the mixture was
stirred under a H2 atmosphere for 3 h. The reaction mixture was
filtered through celite, and the residue was washed with EtOAc
and MeOH. The filtrate was concentrated to yield the amine as a
foamy solid (0.45 g, 97% crude), which was taken on to the next
step without further purification.
Step 2: The crude amine (5.04 g, 9.63 mmol) obtained from
step 1 above was dissolved in a 4:1 mixture of EtOAc (40 mL)
and saturated aqueous NaHCO3 solution (10 mL), and cooled in
an ice-bath, followed by dropwise addition of benzylchloroformate
(1.64 mL, 11.55 mmol) with vigorous stirring. After completion
of addition, the reaction mixture was allowed to come to room
temperature and stirred for 3 h. The aqueous and organic layers
were separated and the aqueous layer was extracted with EtOAc
(2 × 10 mL). The combined organic layers were washed with brine,
dried (Na2SO4), and concentrated. The residue was purified by flash
chromatography (hexanes/EtOAc ) 97:3) to yield the desired
carbamate 13 as a clear viscous oil (5.38 g, 85% over two steps):
Step 2: The carboxylic acid 2 (0.75 g, 1.35 mmol) was dissolved
in anhydrous CH
2Cl2 (8 mL) and cooled to 0 °C, followed by the
[R]D -34.3 (c 1.00, CHCl3); IR (neat) 3416, 1757, 1721, 1705 cm-1
;
addition of N,N-diisopropylethylamine (0.23 mL, 1.35 mmol) and
BOP-Cl (0.34 g, 1.35 mmol). The reaction was stirred at 0 °C for
45 min. The crude amine (0.45 g, 1.12 mmol) as obtained from
step 1 above was dissolved in anhydrous CH2Cl2 (7 mL) and was
added to the activated acid, followed by N,N-diisopropylethylamine
(0.23 mL, 1.35 mmol). The reaction was allowed to come to rt and
stirred for 12 h. After the reaction was quenched with H2O (10
mL) the two layers were separated, and the aqueous layer was
extracted with EtOAc (3 × 10 mL). The combined organic layers
were washed with brine (1 × 15 mL), dried (Na2SO4), and
concentrated under vacuum, and the residue was purified by flash
chromatography (EtOAc/hexanes ) 6:4 to 8:2) to yield the fully
protected pyranosyl nikkomycin analogue 16 as a white solid (0.83
g, 77%): mp 134-136 °C; [R]D -18.8 (c 1.05, CHCl3); IR (neat)
3293, 1752, 1696 cm-1; 1H NMR (400 MHz, CDCl3) δ -0.28 (s,
3H), 0.03 (s, 3H), 0.80 (d, J ) 7.1 Hz, 3H), 0.85 (s, 9H), 1.36 (s,
9H), 1.97-2.07 (m, 4H), 2.18-2.25 (m, 4H), 2.33 (br s, 1H), 3.78
(s, 3H), 4.28-4.35 (m, 2H), 4.66-4.73 (m, 2H), 4.90 (d, J ) 7.2
Hz, 1H), 4.90-5.20 (m, 2H), 5.57 (s, 1H), 5.73 (d, J ) 6.5 Hz,
1H), 6.03 (d, J ) 9.6 Hz, 1H), 6.10 (br s, 1H), 6.96 (d, J ) 8.5 Hz,
2H), 7.18-7.37 (m, 9H), 9.10 (br s, 1H); 13C NMR (100.6 MHz,
CHCl3) δ -4.8, -4.2, 12.8, 18.4, 20.9, 21.5, 26.2, 27.5, 32.0, 39.5,
43.2, 53.2, 55.2, 56.4, 67.3, 67.6, 68.9, 74.9, 78.9, 103.9, 121.8,
127.7, 128.7, 128.9, 136.5, 139.5, 140.2, 150.5, 150.9, 157.1, 163.0,
169.1, 170.1, 170.3, 172.1, 177.5; HRMS calcd for C46H66N5O15Si
m/z (M + NH4)+ 956.4325, found, 956.4293. HPLC: Agilent
ZORBAX SB-C18, 3.0 × 150 mm, 3.5 µm, 2% acetonitrile/water
containing 0.1% TFA and TEA, 0.4 mL/min, 225 nm, retention
time ) 7.57 min.
1H NMR (400 MHz, CDCl3) -0.32 (s, 3H), 0.27 (s, 9H), 0.72 (d,
J ) 6.6 Hz, 3H), 0.73 and 0.88 (2s, 18H), 1.38 (s, 9H), 2.17 (m,
1H), 3.59 (t, J ) 8.4 Hz, 1H), 3.76 (br s, 1H), 4.01 (br s, 1H), 4.53
(d, J ) 7.4 Hz, 1H), 5.07-5.17 (m, 3H), 7.0 (d, J ) 8.1 Hz, 2H),
7.24 (m, 2H), 7.33-7.40 (m, 5H); 13C NMR (100.6 MHz, CDCl3)
δ -5.1, -4.9, -4.8, -4.1, 12.2, 18.4, 18.5, 26.2, 27.5, 39.5, 41.8,
52.7, 64.1, 66.9, 77.9, 121.5, 128.1, 128.4, 128.6, 128.9, 137.1,
141.4, 150.7, 156.5, 177.4; HRMS calcd for C36H59NO6Si2Na m/z
(M + Na)+ 680.3779, found 680.3787.
(1S,2S,3S)-3-[(Benzyloxycarbonyl)amino]-1-[(tert-butyldi-
methylsilyl)oxy]-2-methyl-1-[4-(pivaloyloxy)phenyl]butan-4-ol
(14). To an ice-cooled solution of the carbamate 13 (2.46 g, 3.7
mmol) in MeOH/CH2Cl2 (1:1, 30 mL) was added with stirring small
portions of camphorsulfonic acid (0.007 g, 0.032 mmol) until pH
3. Stirring was continued at 0 °C for 1 h. Upon completion of the
reaction (tlc monitoring), saturated NaHCO3 solution (10 mL) was
added to the mixture, the two layers were separated, and the aqueous
layer was extracted with CH2Cl2 (3 × 10 mL). The combined
extracts were dried over anhydrous Na2SO4 and the solvent was
removed in vacuo. The residue was purified by flash column
chromatography (hexanes/EtOAc ) 7:3) to yield the free primary
alcohol derivative 14 as a viscous low melting solid (1.75 g,
91%): [R]D -52.0 (c 1.00, CHCl3); IR (neat) 3421, 3385, 1746,
1705 cm-1 1H NMR (400 MHz, CDCl3, rotameric mixture) δ
;
-0.26 and 0.30 (2s, 3H), 0.02 and 0.06 (2s, 3H), 0.90 (br s, 12H),
1.37 (s, 9H), 2.07 (m, 1H), 3.45 (br s, exchangeable with D2O,
1H), 3.61 (t, J ) 7.0 Hz, 1H), 3.77 (br s, 2H), 4.60-4.62 (2d, J )
6.0 Hz, 1H), 5.11 (m, 2H), 6.02 (br s, 1H), 7.01 (d, J ) 8.5 Hz,
2H), 7.24 (d, J ) 8.5, 2H), 7.35 (m, 5H); 13C NMR (100.6 MHz,
CDCl3) δ -4.9, -4.1, -3.9, 13.6, 18.4, 26.2, 27.5, 39.5, 42.6, 55.6,
Silyl Deprotection of the Coupled Product 16 to the Alcohol
17. To a stirring solution of the pyranosyl analogue 16 (0.745 g,
9996 J. Org. Chem., Vol. 72, No. 26, 2007

Carbohydrate Ring-Enlarged Pyranosyl Nikkomycin B Analogue
0.79 mmol) in anhydrous THF (12 mL) at 0 °C was added TBAF
(1 M in THF, 1.6 mL, 1.6 mmol). The reaction was stirred at 0 °C
for 2 h, and then quenched with saturated aqueous ammonium
chloride (5 mL), followed by addition of EtOAc (20 mL). The two
layers were separated, and the aqueous layer was extracted with
EtOAc (3 × 5 mL). The combined organic layers were dried (Na2-
SO4) and concentrated under vacuum, and the residue was purified
by flash chromatography (EtOAc/hexanes ) 7:3 to 8:2) to afford
the deprotected alcohol 17 as a white solid (0.535 g, 87%): mp
154-156 °C; [R]D 22.7 (c 1.00, CHCl3); IR (neat) 3298, 1747,
7.31-7.43 (m, 5H), 7.61 (d, J ) 7.9 Hz, 1H); 13C NMR (100.6
MHz, CD3OD) δ 11.0, 33.9, 43.0, 48.9, 56.2, 57.6, 67.1, 68.0, 69.9,
74.2, 75.4, 81.0, 102.4, 115.1, 128.0, 128.1, 128.3, 128.6, 134.2,
137.0, 142.0, 151.9, 157.0, 158.1, 165.1, 172.5, 174.2; HRMS calcd
for C30H38N5O12 m/z (M + NH4)+ 660.2517, found 660.2523.
Pyranosyl Nikkomycin B Oxalate Salt (19). To a room
temperature solution of the hydroxy acid 18 (0.052 g, 0.08 mmol)
in anhydrous MeOH (3 mL) was added 10% palladium on activated
carbon (0.06 g), and the mixture was stirred under a H2 atmosphere
for 2 h. The reaction mixture was filtered through celite, and the
residue was washed with MeOH. The filtrate was concentrated to
yield the free amine as a white solid (0.04 g, 95%, crude). Toward
further purification via crystallization, a saturated solution of oxalic
acid (0.1 g) in ether (5 mL) was added to the above amine dissolved
in a minimum amount of MeOH. The white precipitate was
collected by filtration to yield the fully deprotected pyranosyl
nikkomycin B (19) as its oxalic acid salt (0.036 g, 76%): mp
>200 °C dec; [R]D -13.8 (c 0.13, H2O); IR (KBr) 3447, 1670
cm-1; 1H NMR (400 MHz, D2O) δ 0.79 (d, J ) 7.1 Hz, 3H), 1.93
(d, J ) 15 Hz, 1H), 2.10 (t, J ) 12.9 Hz, 1H), 2.48 (t, J ) 5.2 Hz,
1H), 3.81 (dd, J ) 2.3 and 9.6 Hz, 1H), 4.31-4.42 (m, 4H), 4.55
(d, J ) 8.1 Hz, 1H), 5.81-5.86 (m, 2H), 6.86 (d, J ) 8.3 Hz, 2H),
7.23 (d, J ) 8.3 Hz, 2H), 7.68 (d, J ) 8.1 Hz, 1H); 13C NMR (125
MHz, D2O) δ 11.2, 33.0, 40.7, 54.3, 57.3, 67.3, 68.6, 72.4, 75.1,
80.8, 102.9, 115.5, 128.1, 133.7, 142.2, 152.1, 155.4, 165.9, 168.7,
172.9; HRMS calcd for C22H29N4O10 (free amine) m/z (M + H)+
509.1884, found 509.1880. HPLC: ZORBAX SB-C18 column (3.5
µm; 3.0 × 150 mm), 2% MeCN/H2O (water contained 0.1% TFA
and 0.1% Et3N), 0.4 mL/min, 225 nm, retention time ) 7.57 min
(100%).
1
1699 cm-1; H NMR (500 MHz, CDCl3) δ 0.53 (d, J ) 6.7 Hz,
3H), 1.35 (s, 9H), 1.87 (s, 3H), 2.04-2.10 (m, 4H), 2.17-2.28
(m, 3H, 1H exchangeable with D2O), 3.84 (s, 3H), 4.21 (dd, J )
3.8 and 9.7 Hz, 1H), 4.40 (d, J ) 12.1 Hz, 1H), 4.71 (d, J ) 3.8
Hz, 1H), 4.89 (d, J ) 9.0 Hz, 1H), 4.98 (br s, 1H), 5.11 and 5.20
(2d, J ) 12.3 Hz, 2H), 5.58 (d, J ) 2.8 Hz, 1H), 5.71 (d, J ) 6.5
Hz, 1H), 6.14 (d, J ) 7.9 Hz, 1H), 6.20 (d, J ) 9.8 Hz, 1H), 6.98
(d, J ) 8.4 Hz, 2H), 7.17 (d, J ) 8.2 Hz, 1H), 7.23 (d, J ) 8.4 Hz,
2H), 7.35-7.39 (m, 5H), 8.20 (d, J ) 8.7 Hz, 1H), 9.52 (s,
exchangeable with D2O, 1H); 13C NMR (125 MHz, CHCl3) δ 11.0,
20.3, 20.7, 27.0, 31.8, 39.0, 45.0, 53.1, 54.4, 54.8, 66.7, 67.4, 67.6,
74.8, 75.3, 78.6, 103.8, 121.3, 127.9, 128.0, 128.2, 128.5, 136.0,
138.7, 139.7, 150.4, 150.7, 157.3, 162.2, 169.5, 169.9, 170.1, 170.9,
176.9; HRMS calcd for C40H49N4O15 m/z (M + H)+ 825.3194,
found 825.3195.
Alkaline Hydrolysis of the Ester Linkages of 17 to the
Hydroxy Acid 18. The pyranosyl nucleoside derivative 17
(0.214 g, 0.25 mmol) was dissolved in THF (12 mL) and cooled
to 0 °C, followed by addition of a solution of LiOH‚H2O (0.1 g,
2.6 mmol) in H2O (3 mL). The reaction was stirred at 0 °C for 2.5
h, and then diluted with H2O (2 mL) and EtOAc (15 mL). The
aqueous layer was acidified to pH 2-3 with 10% KHSO4. The
two layers were separated, and the aqueous layer was extracted
with EtOAc (5 × 10 mL). The combined organic layers were dried
(Na2SO4) and concentrated. The residue was purified by flash
chromatography (CHCl3/MeOH/H2O ) 7:2.8:0.2 to 5:4.5:0.5) to
yield the polyhydroxy acid 18 as a white solid (0.088 g, 54%):
mp >185 °C dec; [R]D 13.8 (c 0.55, MeOH); IR (Teflon film) 3348,
Acknowledgment. We thank the Herman Frasch Foundation
(American Chemical Society) for financial support. Pre-doctoral
research fellowships to C.S.S, from the American Chemical
Society Medicinal Chemistry Division (2004-2005) (sponsored
by Aventis), and the American Foundation for Pharmaceutical
Education (2005-2006) are also gratefully acknowledged.
Supporting Information Available: General experimental
details, copies of NMR spectra (1H and 13C) of all the new
compounds, and a copy of the HPLC chromatogram of compound
19. This material is available free of charge via the Internet at
http://pubs.acs.org.
1
1693, 1614 cm-1; H NMR (400 MHz, CD3OD) δ 0.58 (d, J )
6.8 Hz, 3H), 1.94 (d, J ) 12.5 Hz, 1H), 2.18 (t, J ) 12.2 Hz, 1H),
2.40 (br s, 1H), 3.61 (d, J ) 8.7 Hz, 1H), 4.23 (d, J ) 9.2 Hz, 2H),
4.44 (br d, J ) 10.6 Hz, 1H), 4.51 (s, 1H), 4.82 (s, 1H), 5.09 and
5.19 (2d, J ) 12.4 Hz, 2H), 5.68 (d, J ) 7.9 Hz, 1H), 5.92 (d, J )
7.3 Hz, 1H), 6.74 (d, J ) 8.3 Hz, 2H), 7.12 (d, J ) 8.2 Hz, 2H),
JO701814B
J. Org. Chem, Vol. 72, No. 26, 2007 9997