Deacylation studies on furanose triesters using an immobilized lipase: Synthesis of a key precursor for bicyclonucleosides

Article abstract of DOI:10.1039/b618426j

Lipozyme TL IM immobilized on silica catalyses the deacylation of 4-C-acyloxymethyl-3,5-di-O-acyl-1,2-O-(1-methylethylidene)-β-l-threo- pentofuranose to form 3,5-di-O-acyl-4-C-hydroxymethyl-1,2-O-(1-methylethylidene) -α-d-xylo-pentofuranose in a highly se

Full text of DOI:10.1039/b618426j

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Deacylation studies on furanose triesters using an immobilized lipase:
Synthesis of a key precursor for bicyclonucleosides{
a
ab
a
a
a
Ashok K. Prasad,* Neerja Kalra, Yogesh Yadav, Rajesh Kumar, Sunil K. Sharma, Shamkant Patkar,
c
c
Lene Lange, Jesper Wengel* and Virinder S. Parmar*
b
a
Received (in Cambridge, UK) 18th December 2006, Accepted 20th February 2007
First published as an Advance Article on the web 27th March 2007
DOI: 10.1039/b618426j
Lipozyme1 TL IM immobilized on silica catalyses
the deacylation of 4-C-acyloxymethyl-3,5-di-O-acyl-1,2-O-(1-
methylethylidene)-b-L-threo-pentofuranose to form 3,5-di-O-
acyl-4-C-hydroxymethyl-1,2-O-(1-methylethylidene)-a-D-xylo-
pentofuranose in a highly selective and efficient manner.
of pentofuranose 1 in very poor yield (Scheme 2). In our modified
procedure, the two primary hydroxyl groups of 4 were first
11
acetylated to give its diacetyl derivative 5, which upon
debenzylation furnished the novel diacetate 4-C-acetyloxymethyl-
5-O-acetyl-1,2-O-(1-methylethylidene)-b-L-threo-pentofuranose (6).
The diacetate 6, on treatment with K CO –methanol, led to the
2
3
The synthesis of novel nucleoside analogues is gaining importance
because of their applications as key intermediates in the
development of antisense and/or antigene oligonucleotides to
formation of target compound 1 in an overall yield of 9% from
D-glucose (Scheme 2). The trihydroxy sugar derivative 1 was
converted to its triacylated derivatives 2a, 2b and 2c using acetic
anhydride, propanoic anhydride and butanoic anhydride, respec-
tively, in the presence of a catalytic amount of DMAP in yields of
80–85% (Scheme 1).
1–4
regulate targeted gene expression, and for their direct utilization
5
,6
as anti-tumor or anti-viral compounds. One of the major
problems emphasized in the synthesis of modified nucleosides is
the presence of multiple functionalities of nearly identical
Based on our experience of biocatalytic transacylation reactions
on polyhydroxy compounds and their peracylated derivatives, we
chose to use the enzymes Candida antarctica lipase-B immobilized
on polyacrylate (Lewatit), commonly known as Novozyme-435,
porcine pancreatic lipase (PPL), Candida rugosa lipase (CRL),
Theremomyces lanuginosus lipase immobilized on silica
(Lipozyme1 TL IM) and the Candida antarctica lipase-B
immobilized on accurel [CAL B-L(A)] for selective deacylation
of the triacylated pentofuranose derivatives 2a–2c in different
organic solvents in the presence of n-butanol as the acyl acceptor.
Lipozyme1 TL IM in toluene was found to be the most efficient
biocatalyst for the deacylation of compounds 2a–2c (Scheme 1,
Table 1). The other lipases, i.e. Novozyme-435, [CAL B-L(A)],
PPL and CRL, did not accept any of the acylated sugar derivatives
as substrates.
7,8
reactivity, which are difficult to protect and deprotect selectively.
In the present study, we have developed a highly efficient
chemo-enzymatic route for the diastereo- and regioselective
deacylation of one out of the three acyloxy groups of 2a–2c
(Scheme 1), derived from the two primary and one secondary
hydroxyl groups of the potentially useful triol 1, which is an
important precursor for the synthesis of different types of
bicyclonucleosides.
The trihydroxy sugar derivative 1 was synthesized, starting from
9
D-glucose, by following a modified procedure of Youssefyeh et al.;
debenzylation of 3-O-benzyl-4-C-hydroxymethyl-1,2-O-(1-methy-
1
0
lethylidene)-b-L-threo-pentofuranose (4) resulted in the formation
The structure of the diacetylated compound 3a was established
as 3,5-di-O-acetyl-4-C-hydroxymethyl-1,2-O-(1-methylethylidene)-
1
a-D-xylo-pentofuranose on the basis of its IR, H NMR,
13
C
1
NMR, HRMS and H NOE experiments, and comparison of its
1
H NMR spectrum with that of the starting triacetate 2a. Thus,
Scheme 1 Lipase-catalyzed selective deacylation studies on triacylated
pentofuranoses 2a–2c.
a
Bioorganic Laboratory, Department of Chemistry, University of Delhi,
Delhi 110 007, India. E-mail: virparmar@gmail.com;
Fax: +91 11 2766 7206; Tel: +91 11 2766 6555
Nucleic Acid Center, Department of Chemistry, University of Southern
b
Denmark, Campusvej 55, DK-5230 Odense M, Denmark
Novozymes A/S, Denmark, Smormosevej 25, DK-2880 Bagsvaerd,
Denmark
c
{ Electronic supplementary information (ESI) available: Experimental
procedures and NMR characterisation data. See DOI: 10.1039/b618426j
Scheme 2 Synthesis of compound 1 from glucose.
2
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Table 1 Diastereoselective deacylation of 4-C-acyloxymethyl-3,5-di-O-acyl-1,2-O-(1-methylethylidene)-b-L-threo-pentofuranose 2a–2c catalyzed
a
by Lipozyme1 TL IM in toluene in the presence of n-butanol
Reaction
time/h
Percentage
yield
Substrate
Product
4
3
4
a
-C-Acetoxymethyl-3,5-di-O-acetyl-1,2-O-
1-methylethylidene)-b-L-threo-pentofuranose (2a)
,5-Di-O-propanoyl-1,2-O-(1-methylethylidene)-4-C-
propanoyloxymethyl-b-L-threo-pentofuranose (2b)
-C-Butanoyloxymethyl-3,5-di-O-butanoyl-1,2-O-
3,5-Di-O-acetyl-4-C-hydroxymethyl-1,2-O-
9
7
4
98
88
93
(
(1-methylethylidene)-a-D-xylo-pentofuranose (3a)
3,5-Di-O-propanoyl-4-C-hydroxymethyl-1,2-O-
(1-methylethylidene)-a-D-xylo-pentofuranose (3b)
3,5-Di-O-butanoyl-4-C-hydroxymethyl-1,2-O-
(1-methylethylidene)-a-D-xylo-pentofuranose (3c)
(1-methylethylidene)-b-L-threo-pentofuranose (2c)
All of these reactions, when performed under identical conditions but without adding the lipase Lipozyme1 TL IM, did not yield any
product.
13,14
temperature
11
and acylating agents on the selectivity of lipases.
the C-3 proton in product 3a and in the starting triacetate 2a
resonated at practically the same d value, i.e. at 5.29 and 5.35,
respectively, which indicated that the acetoxy function at the C-3
position, present in the starting triester 2a, is intact in the product
The present study demonstrates the selectivity of lipases due to
their immobilization on different solid supports. This may be
because of the fact that the protein adopts different conformations
and geometries on different support surfaces. Also, there are ways
to differentiate between primary and secondary hydroxyl groups,
but discrimination between two primary hydroxyl groups or their
derivatives is, in general, not possible by classical chemical
methods alone. The very efficient and convenient enzymatic
method discovered for the discrimination between two primary
hydroxyl groups of the furanose derivatives herein may find
applications in the ‘green’ synthesis of bicyclonucleosides,
important precursors for the preparation of antisense or antigene
3a. Furthermore, two pairs of doublets, resonating at d 4.36 and
4
.29, and at 4.27 and 4.10, were observed due to C–19H and C–5H
1
in the H NMR spectrum of the starting triacetate 2a; one pair of
these doublets, resonating at d 4.36 and 4.29, shifted upfield to d
3
1
.82 and 3.73 in the H NMR spectrum of the product diacetate
3a.
The formation of a single monodeacetylated product indicates
that the lipase selectively catalysed the deacetylation of the acetoxy
function, either at the C-5 or C-19 position, in compound 2a. The
NOE study carried out on product diacetate 3a proved that
the acetoxy function at the C-19 position in 2a is deacetylated in
the presence of the lipase. Thus, the irradiation of the upfield-
shifted signals of C–19H, i.e. those resonating at d 3.82 and 3.73 in
compound 3a, exhibited appreciable NOE (4%) on the C-3 proton
resonating at d 5.29. If the upfield-shifted signals at d 3.82 and 3.73
had been due to the C-5 protons, their irradiation would have not
affected the signal of the C-3 proton because they are not in close
proximity.
15
oligonucleotides.
We thank the Danish Natural Science Research Foundation
and the Department of Biotechnology, Government of India (New
Delhi) for their financial support of this work.
Notes and references
1
2
3
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It has been observed that increasing the chain length of the acyl
moiety increases the rate of the lipase-catalyzed deacylation
reaction (Table 1). Thus, the rate of deacylation of propanoylated
sugar derivative 2b is about 1.3 times faster than the rate of
deacylation of acetylated sugar derivative 2a. Accordingly, the rate
of deacylation of butanoylated sugar derivative 2c is about 2.3 and
4
5
B. R. Babu, P. J. Hrdlicka, C. J. McKenzie and J. Wengel, Chem.
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1.8 times faster than the rate of deacylation of acetylated and
1199–1210.
propanoylated sugar derivatives 2a and 2b, respectively. The
structures of all the novel compounds obtained in this study (2a–
7
8
9
P. Collins and R. Ferrier, Monosaccharides: Their Chemistry and Their
Roles in Natural Products, Wiley, Chichester, UK, 1995, pp. 431.
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c, 3a–3c and 6) were unambiguously established on the basis of
1347–1360.
1
13
their spectral (IR, H NMR, C NMR, HRMS and NOE)
analyses. The structures of the known compounds mentioned in
Scheme 2 were further confirmed by comparison of their physical
and/or spectral data with those reported in the literature. All
deacylation reactions, when performed under identical conditions
but without adding any lipase, did not proceed to any extent.
Lipozyme1 TL IM discriminated between the three ester
functions derived from two primary hydroxyl groups and a
secondary hydroxyl group in novel sugar derivatives. Ourselves
R. D. Youssefyeh, J. P. H. Verheyden and J. G. Moffatt, J. Org. Chem.,
1979, 44, 1301–1309.
10 T. F. Tam and B. Fraser-Reid, Can. J. Chem., 1979, 57, 2818–2822.
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5, 3380–3386 and references cited therein.
4 V. S. Parmar, A. K. Prasad, P. K. Singh and S. Gupta, Tetrahedron:
1
1
1
1
3
1
Asymmetry, 1992, 3, 1395–1398.
15 J. Wengel, Acc. Chem. Res., 1999, 32, 301–310.
12
and others have previously demonstrated the effect of solvent,
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