Enzymatic synthesis of a branched-chain hexulose: 5-Deoxy-5-C- hydroxymethyl-β-L-xylo-hex-2-ulopyranose

Article abstract of DOI:10.1002/(SICI)1099-0690(199906)1999:6<1415::AID-EJOC1415>3.0.CO;2-2

The coupling of dihydroxyacetone phosphate with 3-hydroxy-2- (hydroxymethyl)propanal (1) in the presence of rabbit muscle aldolase afforded a branched-chain hexulose phosphate isolated in 64% yield as the barium salt 3. The parent, dephosphorylated sugar 4 was converted to its anhydro derivative 8 and glycoside 9.

Full text of DOI:10.1002/(SICI)1099-0690(199906)1999:6<1415::AID-EJOC1415>3.0.CO;2-2

FULL PAPER
Enzymatic Synthesis of a Branched-Chain Hexulose:
-Deoxy-5-C-hydroxymethyl-β--xylo-hex-2-ulopyranose
5
Serge David[a]
Keywords: Enzymatic synthesis / Rabbit muscle aldolase / Branched-chain sugars / Anhydro sugars / Glycosidation
The coupling of dihydroxyacetone phosphate with 3-
hydroxy-2-(hydroxymethyl)propanal (1) in the presence of
rabbit muscle aldolase afforded a branched-chain hexulose
phosphate isolated in 64% yield as the barium salt 3.The
parent, dephosphorylated sugar 4 was converted to its
anhydro derivative 8 and glycoside 9.
Introduction
Discussion
Presumably, the authors[2] had examined an aqueous
solution of 1 directly obtained by acidic hydrolysis of its
diethyl acetal, 1,1-diethoxy-3-hydroxy-2-(hydroxymeth-
yl)propane, since the isolation of 1 appears problematic.
The preparation of such a solution with the help of an ion-
The difficulties linked with the introduction of a branch
of the >CHR type with a determined configuration in a
sugar are well known.[1] On the other hand this mode of
branching is the more useful when sugars are used as chiral
units in total synthesis. As a new approach to the problem,
we report the preparation (Scheme 1) of the branched-chain
ketose 4 by the enzymatic condensation of aldehyde 1 with
dihydroxyacetone phosphate (DHAP, 2) catalyzed by rabbit
muscle aldolase (from now on simply called aldolase in this
paper). Aldehyde 1 is one of a collection of 64 aldehydes
which were tested by the Whitesides group as potential sub-
ϩ
exchange resin in the H form (26 mL per mmol of acetal)
has been already reported.[ Removal of the water gave an
oil with the expected composition; however the authors
considered it difficult to separate this concentration step
from a subsequent chemical transformation, the elimination
of water from this β-hydroxylated aldehyde, yielding the
substituted acrylic acid.[ For this reason some authors
adopted a 1-h standing at room temperature in aqueous
sulfuric acid, pH ϭ 2.5, as conditions for the hydrolysis of
the acetal. Catalytic hydrogenation of the crude product
gave 2,2-dihydroxy-1,3-propanediol in quantitative yield.[
In our system, these conditions did not result in a quantitat-
ive yield, as 65% of unreacted acetal could be recovered by
ether extraction. The acidic solvent was a 0.1  aqueous
solution of DHAP, partially neutralized to the required pH
with 1  KOH. Acceptable conditions are reported below.
The precursor acetal has been prepared by reduction of
diethyl 2,2-diethoxymethylmalonate, a compound easily
available on the mole scale.[ For this reduction, the au-
3]
4]
strates for aldolase by Vmax estimation.[2] The Vmax was 0.34
times that of -glyceraldehyde phosphate, the natural sub-
strate, and thus the reaction was considered potentially use-
ful in synthesis. However, although the handling of an elu-
sive aldehyde such as 1 cannot be straightforward, no de-
tails were given nor was the isolation of the product ketose
phosphate reported. In this paper, we describe a prepara-
tive-scale synthesis of this compound, and some chemical
transformations.
5]
3]
thors utilized 3 mol of LiAlH per mol of diester. Such
4
excess is not necessary: We obtained this acetal in 82% yield
with a 1.5:1 ratio and conventional distillation. We found
that the hydrolysis of this acetal was practically complete
after 2 h at pH ϭ 2.00 at room temperature by enzymatic
estimation of released ethanol with alcohol dehydrase and
oxidized diphosphopyridine nucleotide.
Aqueous solutions of DHAP were prepared by the
method of Effenberger.[ Their molarities were about 0.1 
but this figure coud be somewhat raised by more sparingly
using water in the last steps. The pH was about 1.0. Three
equivalents of acetal were added to the solution of DHAP,
and the pH was quickly raised to 2.0 with 1  KOH and
kept for 2 h at this value. There was a slight turbidity, but
this had no adverse effect on the enzyme. Then the pH was
6]
Scheme 1. The aldolase reaction with aldehyde 1
[
a]
Institut de Chimie Mol e´ culaire de lЈUniversit e´ de Paris-Sud,
B aˆ t. 420, F-91405 Orsay cedex, France
Eur. J. Org. Chem. 1999, 1415Ϫ1420

WILEY-VCH Verlag GmbH, D-69451 Weinheim, 1999
1434Ϫ193X/99/0606Ϫ1415 $ 17.50ϩ.50/0
1415

S. David
FULL PAPER
1
raised to 7.20 with 1  KOH and aldolase was added. After Table 1. H-NMR data for backbone and methyl protons of pyra-
noses and pyranosides[
a]
2
d at room temperature, the product ketose phosphate 3
was isolated as the barium salt in about 70% yield, on a 20-
mmol scale. Treatment with phosphatase gave the free sugar
Compound3
Chemical shifts (δ)
4
17
18
16
4
in nearly quantitative yield as a hygroscopic syrup which
crystallized on standing for several months at room tem- 1-H
3.44
3.70
3.44
3.69
3.48
3.68
1.86
3.72
3.77
3.59
4.07
4.26
5.17
5.35
2.33
3.61
3.85
4.1
3.62
3.72
3.87
3.93
2.16
3.62
3.86
3.53
3.56
3.29
3.60
3.77
5.43
5.43
2.22
3.70
3.94
3.57
3.57
3.16
1
3
4
Ј-H
-H
-H
perature. This solid melted at 106°C. The solubility is of
the order of 0.1  in dioxane at room temperature, and this
allows to remove coloured impurities originating from the 5-H
[
c]
[c]
3.62
1.84
3.62
3.95
3.52
[
b]
6
6
a-H
e-H
7-H
phosphatase. Thus, a colourless compound was obtained
with the expected composition, C H O . Both the sugar
7
14
6
and its phosphate appeared homogeneous according to 7Ј-H
4.1
1
Me
Coupling constants (J, in Hz)
3.27
their H-NMR spectra (Table 1). The configurations and
conformations given in Scheme 1 are indicated by the fol-
lowing considerations: Formula 5 (Scheme 2) is a Fischer
representation of the ketonic tautomer of this sugar, as ex-
pected from the known course of the aldolase reaction.
There are two identical branches (pro-R) and (pro-S) on C-
J
J
J
J
J
J
J
J
1,1Ј
3,4
10
12
9
10
10
3.5
11.5
10
10
12
5
10
10
10
10
5
10
10
11
11
5
4,5
10
9.5
6
7
5
11.5
5,6a
5,6e
5,7
5
. The correct name is 5-deoxy-5,5-di-C-hydroxymethyl--
threo-pentulose, as there are only two permanent chiral
centres. However, for the sake of clarity, we prefer to con-
sider the cyclic tautomers as derivatives of 5-deoxyhex-
uloses (-xylo, 6) and (-arabino, 7). In these, the carbon
atom of one of the branches, either the (pro-R) in 6, or the
4
5,7Ј
7,7Ј
12
[a]
2 2 3 3
Solvents: D O for 4, D O with a drop of HNO for 3, CDCl
[b]
for the other compounds. Ϫ Proton 5-H has 5 neighbours, which
(pro-S) in 7, is considered as C-6 of a hexulose, the other
[c]
could lead in principle to a signal of 32 lines. Ϫ J/∆ν > 2.
hydroxymethyl group being a branch on C-5 of this hexu-
lose.
β--xylo configuration appears much favoured, while we
may expect to observe the presence of small quantities of
the β--arabino sugar.
Scheme 3. Possible cyclic tautomers of hexulose 5
Scheme 2. Some drawings of the acyclic tautomer of hexulose 4
1
In the H-NMR spectrum of the pyranoses in Scheme 3
Four possible configurations can be drawn on paper for (see 4 in Scheme 3 for numbering), we expect the signal of
the pyranose hemiacetals derived from 5 (Scheme 3). Cycli- 5-H to appear at high fields, well separated from the signals
zation involving the (pro-R)-hydroxymethyl group would of the other skeleton protons. Then the β--xylo configura-
give the two anomeric -xylo-hexopyranoses (α- and β-). tion should be easily recognized, as this is the only one in
Cyclization involving the (pro-S)-hydroxymethyl group which this proton is axial and coupled to two axial protons.
would give the two -arabino derivatives. These have been One further large coupling with one of the hydroxymethyl
drawn in chair form, in the conformation presumed to be protons, 7-H or 7Ј-H, and two smaller couplings can be
the most stable. Configurations at C-2 are imposed by two expected. In fact, the signal of 5-H in phosphate 3 (or the
overwhelming factors: the anomeric effect and the en- corresponding acid) was a broad multiplet at δ ϭ 1.94,
hanced conformational energy of a hydroxymethyl substitu- spanning 38 Hz. The signal of 5-H was very similar in the
ent when next to the ring oxygen atom in a pyranose.[ The spectrum of the free sugar, identical in most respects with
7]
1416
Eur. J. Org. Chem. 1999, 1415Ϫ1420

Enzymatic Synthesis of a Branched-Chain Hexulose
FULL PAPER
that of 3, in agreement with configuration 4. The signal of
5
-H, and the observation of coupling constants in the vicin-
ity of 10 Hz between 3-H and 4-H, 4-H and 5-H, 5-H and
a-H in the NMR spectrum of other derivatives described
6
below, confirmed the permanence of this configuration and
conformation in the transformation products (Table1). Fur-
thermore the sharpness of the peaks of 3 and 4 indicated
the homogeneity of the compound. Thus, of the four pos-
sible configurations for this product, only one was observed
for conformational reasons. The cyclization reaction in-
volves the (pro-R)-hydroxymethyl group.
Sugar 4 was converted to an anhydro derivative 8 by
treatment with acid in a non-hydroxylated solvent (Scheme
4
). The reaction in a 0.1  solution of camphorsulfonic acid
in dioxane proceeded very slowly at room temperature after
ca. 80% conversion. This reaction may be equilibrated. In
the spectrum of the product (see Scheme 4 for the number-
ing) the signal of 5-H appears as an unresolved multiplet at
δ ϭ 2.10, spanning only 12 Hz (Table 2). Only weak coup-
lings were observed for protons 3-H, 4-H and 5-H. The
same observations could be made in the spectra of deriva-
tives 10 and 11 (Scheme 5) and confirm structure 8 for the
Scheme 5. Derivatives of 8 and 9
1
Table 2. H-NMR data for backbone protons of anhydro deriva-
anhydro sugar. Both prochiral branches of 4 are involved in tives[
a]
this reaction. Observation of a model shows that this can
Compound2
Chemical shifts (δ)
10
11
14
12
13
be done in only one way and the given numbering for the
carbon atoms of 8 has been chosen consistent with this ori-
gin: Thus, carbon atom 7 in 8 and carbon atom 7 in 4 both
originate from the (pro-S)-carbon atom in 5.
1
-H
3.50
3.50
3.70
3.93
2.06
4.07
4.14
3.96
3.96
3.86
4.20
5.12
5.01
2.22
4.06
4.09
4.20
4.25
3.38
3.62
3.90
3.83
2.16
3.93
3.98
4.13
4.27
3.67
3.67
3.79
ca 4.1
2.04
3.91
3.64
3.64
4.12
3.92
1.72
3.79
3.89
3.96
4.39
3.46
3.46
5.50
3.92
1.64
3.87
4.46
3.90
3.62
1Ј-H
3
4
5
6
6
7
7
-H
-H
-H
[b]
a-H
e-H
-H
4.10
4.22
Ј-H
Coupling constants (J, in Hz)
J
J
J
J
J
J
J
1,1Ј
3,4
17
2.5
5
12.5
2.5
10.5
10.5
4
13
0
4,5
5
5,6a
5,6e
6,6
2.5
1
9
7
10
9
8.5
9
10
10
9
9
9
7,7Ј
[
a]
3
Solvent: CDCl . Long-range couplings may be expected between
3
-H and 5-H, 4-H and 6-H, 6Ј-H and 7Ј-H. Couplings of the order
Scheme 4. Reactions of hexulose 4 in acidic media
of 0.5Ϫ2 Hz are visible on the spectra, but have not been recorded
here. Number 7 has been given to the only proton which cannot
[b]
exhibit W-type long-range coupling. Ϫ
half heigth, 9Ϫ12 Hz.
Broad signals: width at
The rate of ring closure is higher than the rate of glyco-
side formation in methanol in the presence of an acidic re-
sin at room temperature. Under some conditions (see Ex-
perimental Section) complete disappearance of 4 was ob- trum of the acetyl derivative 13, on which the signal of the
served after 2 h. The estimation of the composition of the only displaced proton appears as a sharp peak. Compound
medium at this stage from the relative intensities of the 12 was isolated in 20% yield.
NMR signals of 5-H of 8 and 9 was misleading, as it greatly
The next eluted product was the methyl glycoside 15, sily-
underrated the concentration of 9. On the other hand, deri- lated on positions 1 and 7, obtained in 22% yield. This was
vatization with tert-butylchlorodiphenylsilane[ gave a mix- characterized as the acetate 16. In the H-NMR spectrum
ture of three silyl ethers which were cleanly separated on a of 16 the distance between the signals of 3-H and 4-H is
silica-gel column in the expected order of polarities. The smaller than their coupling and accordingly, a first-order
first eluted product was the monoalcohol 12, silylated on analysis is not possible. The observed pattern corresponds
8]
1
1
positions 1 and 4. This was indicated by the H-NMR spec- to ν Ϫ ν ϭ 2 Hz, J ϭ 10 Hz, J4,5 ϭ 8 Hz, that is, a
3
4
3,4
Eur. J. Org. Chem. 1999, 1415Ϫ1420
1417

S. David
FULL PAPER
trans-diaxial disposition of 3-H, 4-H and 5-H. The most conditions being achieved by the operation of the classical
polar derivative was the anhydro sugar 14, silylated at its ring-chain tautomerism of sugars. The maximum theoreti-
primary position (19%). Thus, the total yield of derivatives cal yield is 100%. Furthermore, to reach the aldolase-me-
of 8 is 39%, while the derivative of the methyl glycoside is diated equilibrium in reasonable time, the above-mentioned
obtained in 22% yield. Under the conditions of the deriva- authors have added much more enzyme, in fact 22 times as
tization (excess of reagent) we may assume that these figures much as we have used per mol of final product.
truly reflect the proportions of 8 and 9 in the glycosidation
medium after 2 h and that there is roughly twice as much neck in the scaling up of this preparation. All the starting
anhydro sugar as methyl glycoside. materials are easily available, and aldolase could be immo-
From the preparative point of view, we foresee no bottle-
On the other hand, acetylation of the mixture obtained bilized if necessary. Finally, we note that treatment of com-
after several days of reaction at room temperature gave the pound 15 or some analogue with NaIO and NaBH (the
4
4
tri-O-acetyl derivative 17 of the methyl glycoside in 66% so-called Smith degradation of polysaccharides) would give
yield from the crude sugar 4 (about 50% yield from a chiral tris(hydroxymethyl)methane. There is current inter-
1
[10]
DHAP). Configuration 17 is proved by H-NMR spec- est in these compounds.
troscopy. This result indicates that the methyl glycoside 9 is
thermodynamically more stable that the anhydro sugar 8.
Zempl e´ n deacetylation of 17 followed by benzylation in the Experimental Section
usual way gave the tri-O-benzyl ether 18, again with the
same configuration.
General: Melting points are uncorrected. Ϫ Optical rotations:
JASCO DIP 370 polarimeter; 1-cm cells; all values measured at
Thus, the aldolase-mediated synthesis gave only one room temperature. Ϫ Thin-layer chromatography (TLC): Silica gel
SO
pyranose configuration, 4. This configuration could be sta- coated glass plates (Merck); detection by spraying with 5% H
2
4
in ethanol and charring. Ϫ Column chromatography: Open col-
umns filled with 20 g of silical gel (particle size 60 µm) per g of
bilized by conversion to the glycoside 9. Under equilibrium
conditions the anhydro derivative 8 was also obtained as a
by-product. Compound 8 can be considered as a derivative
of the same configuration as the free sugar, originating from
the attack of the hemiacetal carbon atom by the branch on
C-5. In fact, in the acidic medium of glycosidation, the ace-
tal function in 8 might open in two ways, and lead, in prin-
1
product. Ϫ UV/Vis: Cary Varian 1E. Ϫ H NMR: Bruker AC 250
(
250 MHz); CDCl
3 4 2
as solvent, Me Si as internal standard; D O as
solvent, δ ϭ 4.80. The chemical shifts of the protons bound to
the carbon skeletons of the sugars are given in Table 1 and 2; the
signals of substituents in derivatives, that is, acetyl, phenyl, benzyl,
and tert-butyl are as predicted in locations and intensities and have
Η
ciple, to an equilibrium mixture of the four possible methyl not been reported. Ϫ Rabbit muscle aldolase, phosphatase, -gly-
glycosides, but free enthalpy differences between these in- cerophosphate dehydrase and ethanol dehydrase were Sigma prod-
ucts. DHAP was prepared by the method of Effenberger[ and esti-
6]
sure that 9 is the only one that can be isolated.
mated with -glycerophosphate dehydrase and NADH according
It is remarkable that both the starting materials are achi-
ral and that aldolase is the only source of chirality. It should
be noted that the discrimination between the prochiral
branches of 5 is not a consequence of the use of an enzyme.
It is well known, that the aldolase-mediated condensation
of DHAP with racemic glyceraldehyde gives the -fructo
and -sorbo configuration in roughly equal amounts. In our
work, aldolase only insured an easy preparation of the
branched-chain ketose, and its locking in the  series (the
acyclic configuration being used to define the series, as it
should).
[
11]
to ref.
Ethanol in water was estimated by the reduction of
NAD in the presence of ethanol dehydrase, at pH ϭ 10.0 in 0.1
glycine buffer with the help of the equilibrium constant measured
ϩ

by Racker:[ k ϭ [CH
12]
CHO][NADH]/[CH CH OH][NAD ] ϭ
ϩ
3 3 2
0
.115.
1
5
,1-Diethoxy-3-hydroxy-2-(hydroxymethyl)propane: A solution of
0.4 g (0.192 mol) of diethyl diethoxymethylmalonate in 100 mL
of ether was added dropwise in the course of 2 h to a solution of
1 g (0.29 mol) of LiAlH in 200 mL of ether, and the mixture was
refluxed for 16 h. Then 11 mL of water, 11 mL of 15% aqueous
NaOH, and 11 mL of water were added in succession. The suspen-
1
4
The prevalence of a β--xylo configuration in an aldolase sion was filtered, and the precipitate was extracted twice with boil-
condensation has already been reported by Durrwachter et ing ether (4 h). Distillation of the combined ether solutions gave
[
9]
al. Nevertheless, there is only a superficial similarity with 28.3 g of the diethyl acetal (82%), liquid, b.p. 93°C (0.05 Torr). Ϫ
1
H NMR (D
J ϭ 6.5 and 7.5 Hz, CH(CH
H, J ϭ 6.5 Hz, CH(OEt) ].
2
O): δ ϭ 1.18 (t, 6 H, J ϭ 7 Hz, 2 Me), 1.94 [dq 1 H,
the reaction reported in our paper. The substrate utilized
by these authors is a mixture of enantiomers, (R,S)-2-
2
OH) ], 3.55Ϫ3.62 (m, 8 H), 4.63 [d,
2
1
2
(hydroxymethyl)-4-pentenal, added in excess. The product
obtained under equilibrium conditions corresponds to the 5-Deoxy-5-C-hydroxymethyl-β-
L
-xylo-hex-2-ulopyranose 1-(Barium
Phosphate) (3): The above acetal (17.6 g; 96.8 mmol) was added to
51 mL of a 0.123  solution of DHAP in water, and the pH was
raised to 2.00 with aqueous KOH. After 2 h at room temperature,
the pH was raised to 7.19 with 1  KOH, 5 mg of NaN , 29 mg
practically exclusive utilization of the (S) enantiomer. This
is a consequence of the reversibility of the catalysis by aldo-
lase, which regenerates DHAP from the less stable hexulose.
Thus, in these experiments, the substrate is chiral, there is
no de novo creation of a chiral centre at C-5, the presence
of aldolase is necessary, and the maximum theoretical yield
is 50% of the racemic substrate. On the other hand, in our
2
3
of tricyclohexylammonium phosphoglycollate and 1 mL (275 units)
of aldolase suspension were added. After 64 h at room temperature,
the amount of DHAP had dropped to 5.3 mmol. Then the pH was
raised to 8.4 with 1  KOH. Barium acetate (15.24 g; 0.6 mol) was
experiment the substrate is achiral, the enzyme does not par- added, the pH which had dropped to 7.0 was again adjusted to 8.4
ticipate in the creation of chirality at C-5, the equilibrium with 1  KOH. Barium phosphate was removed by filtration, and
1418
Eur. J. Org. Chem. 1999, 1415Ϫ1420

Enzymatic Synthesis of a Branched-Chain Hexulose
00 mL of 95% ethanol was added to the filtered solution. The
FULL PAPER
butylchlorodiphenylsilane, and 0.4 g of imidazole were added.[8]
8
precipitate was collected, washed with 150 mL of ethanol/water After 6 h standing at room temperature, the mixture was par-
(
4:1), ethanol and ether, and air-dried to give 8.03 g of crude prod-
titioned between water and diethyl ether . The silyl ethers in the
ethereal extract (1.118 g) were separated by chromatography. Elu-
tion with light petroleum ether/ethyl acetate (8:1) gave 204 mg of
uct, which, from its NMR spectrum still contained 9.4% of barium
acetate (δ ϭ 1.86 in D
Ϫ
2 3 2
O, CH CO , shifting to δ ϭ 2.00 by acidifi-
cation with HNO
3
). The corrected yield of 3 was 58% (70% ac- 2,7-anhydro-1,4-di-O-tert-butyldiphenylsilyl-5-deoxy-5-C-
counting for recovered DHAP). This product was used as such for
the next step.
hydroxymethyl-β--xylo-hex-2-ulopyranose (12; 0.31 mmol; 20%).
Ϫ TLC ixght petroleum ether/ethyl acetate (4:1)]: R ϭ 0.63. Ϫ
α] ϭ Ϫ5.2 (c ϭ 0.89, dichloromethane). Ϫ C39 Si (652):
calcd. C 71.78, H 7.36; found C 71.79, H 7.61. Ϫ Elution with light
petroleum ether/ethyl acetate (4:1) gave 240 mg (0.35 mmol; 22%)
of the bis(ether) 15 which was characterized as the diacetate (see
below). Finally, elution with ethyl acetate gave 355 mg of a mixture
which was further purified by chromatography [light petroleum
ether/ethyl acetate (1:1)] to give 123 mg (0.30 mmol; 19%) of 2,7-
anhydro-1-O-tert-butyldiphenylsilyl-5-deoxy-5-C-hydroxymethyl-β-
f
[
D
H
48
O
5
2
5
-Deoxy-5-C-hydroxymethyl-β-
L
-xylo-hex-2-ulopyranose (4):
A
solution of 7.6 g (18.6 mmol) of 3, dissolved in a little water, was
ϩ
passed through a 90 meq column of Dowex-50 (H ) ion-exchange
resin. The effluent was concentrated to 40 mL at 45Ϫ50°C, the
volume was adjusted to 100 mL, and the pH was brought to 4.25
with 1  KOH. Wheat germ phosphatase (120 units) was added
and the solution was kept for one week at room temperature under
a layer of toluene. Then it was heated to 90°C for a few minutes,
cooled and separated from the coagulated proteins by filtration,
and deionized by successive passages through a 25 meq Dowex-50

-xylo-hex-2-ulopyranose (14) as a syrup which crystallized on
standing, m.p. 120Ϫ123°C. Ϫ TLC [light petroleum ether/ethyl
acetate (1:1)]: R ϭ 0.23. Ϫ [α] ϭ Ϫ20.8 (c ϭ 1.25, dichlorometh-
ane). Ϫ C23 Si (414): calcd. C 66.66, H 7.25; found C 66.66,
f
D
ϩ
Ϫ
(
H ) and a 30 meq Dowex-1 (AcO ) ion-exchange resin column.
30 5
H O
Concentration gave 3.0 g (83.6%) of 4, as a hygroscopic syrup
slowly crystallizing in a few months at room temperature. Dissol-
ution in dioxane at 80°C (10 mL per mmol), filtration and concen-
tration removed some coloured impurities; m.p. 106°C. Ϫ TLC [2-
H 7.39. Ϫ Acetylation of 12 (acetic anhydride/pyridine) gave the 3-
O-acetyl derivative 13. Acetylation of 15 gave methyl 3,4-di-O-ace-
tyl-1-O-tert-butyldiphenylsilyl-5-tert-butyldiphenylsilyloxymethyl-
5
-deoxy-β--xylo-hex-2-ulopyranoside (16; 100%), which was crys-
tallized from pentane, m.p. 90Ϫ98°C. Ϫ [α] ϭ 24.8 (c ϭ 1.7, di-
Si (768): calcd. C 68.75, H 7.29; found
propanol/water (3:1)]: R
ϭ 0.51; [methanol/dichloromethane (1:4)]: R
Ϫ24.3 (c ϭ 2.0, H O). Ϫ C (194): calcd. C 43.30, H 7.22,
O 49.48; found C 43.11, H 7.25, O 49.22.
f
ϭ 0.71; [methanol/dichloromethane (1:1)]:
D
R
f
f
ϭ 0.11.Ϫ [α]
D
ϭ
56 8
chloromethane). Ϫ C44H O
2
7 14 6
H O
C 68.83, H 7.39.
2
,7-Anhydro-5-deoxy-5-C-hydroxymethyl-β-
L
-xylo-hex-2-
Methyl 5-C-Acetoxymethyl-1,3,4-tetra-O-acetyl-5-deoxy-β-L-xylo-
ϩ
ulopyranose (8): The addition of 194 mg of 4 to 11 mL of a 0.1 
camphorsulfonic acid solution in dioxane gave a clear solution
which was kept for 2 d at room temperature. Then K CO (415 mg;
2 3
hex-2-ulopyranoside (17): Dowex-50 ion-exchange resin in the H
form was stirred for 2 d in anhydrous methanol, with frequent re-
moval of the liquid. A sample of this resin (1.8 mL) was added to
3
mmol) was added and the suspension was stirred for 2 d at room a solution of 842 mg (4.42 mmol) of 4 in dry methanol (20 mL),
temperature, then filtered, and dioxane was removed from the
liquid phase by evaporation. TLC examination indicated that the
residue was a mixture of 4 and 8 in a ratio of ca 3:1. Chromatogra-
and the mixture was stirred for 3 d at room temperature. The resin
was filtered off, 0.2 mL of pyridine was added to the solution which
was concentrated to dryness. To the resin were added 3.4 mL of
phy dichloromethane/methanol (6:1)] separated 8 (95 mg; 54%) acetic anhydride and 4.3 mL of pyridine, and the mixture was kept
m.p. 135Ϫ137°C. Ϫ TLC dichloromethane/methanol (4:1)]: R
.33. Ϫ [α] ϭ Ϫ3.4 (c ϭ 1, methanol). Ϫ C (176): calcd.
C 47.72, H 6.82, O 45.45; found C 47.61, H 6.71, O 45.29.
f
ϭ
for 2 d at room temperature, then concentrated to dryness to give
a crude product (1.372 mg) which was purified by chromatography
[methanol/dichloromethane (1:100)]. Elution first gave 376 mg of a
fraction which was a 1.4:1 mixture of 17 and 10, as estimated from
the intensities of the signals of 3-H and 4-H in the NMR spectrum.
Continued elution gave 848 mg (52%) of 17 as a syrup. Ϫ TLC
0
D
7 12 5
H O
1
,3,4-Tri-O-acetyl-2,7-anhydro-5-deoxy-5-C-hydroxymethyl-β-
xylo-hex-2-ulopyranose (10): Prepared from 8 in the usual way
pyridine/acetic anhydride) and purified by column chromatogra-
phy [light petroleum ether/ethyl acetate (8:1)]. Syrup; TLC light
petroleum ether/ethyl acetate (1:1)]: R ϭ 0.46. Ϫ [α] ϭ Ϫ9.2 (c ϭ
.98, dichloromethane). Ϫ C13 (302): calcd. C 51.68, H 6.12;
found C 51.96, H 6.12.
L-
(
[methanol/dichloromethane (2.5:100)]: R ϭ 0.37. Ϫ [α]D ϭ Ϫ13.2
f
(c ϭ 2.1, dichloromethane). Ϫ C H O (376): calcd. C 51.07, H
1
6
24 10
f
D
6.45, O 42.31; found C 51.25, H 6.56, O 42.63.
0
18 8
H O
Methyl 1,3,4-Tri-O-benzyl-5-C-5-benzyloxymethyl-β-L-xylo-hex-2-
ulopyranoside (18): To a solution of sodium methoxide, prepared
from 56 mg of Na in 5.6 mL of methanol, was added 400 mg (1.06
mmol) of 17. The solution was kept for 4 h at room temperature,
refluxed for 90 min and concentrated to dryness. Benzylation in 11
mL of DMF, with 1.0 mL (8.5 mmol) of benzyl bromide and 250
mg (6.3 mmol) of 60% NaH suspension in oil, by heating at 90°C
for 4 h, was incomplete. The product was recovered as described
for 18 at the end of this section, dissolved in 12 mL of DMF, and
2
,7-Anhydro-1,3,4-tri-O-benzyl-5-deoxy-5-C-hydroxymethyl-β-
xylo-hex-2-ulopyranose (11): Prepared from 8 in the usual way
NaH and benzyl bromide in DMF) and purified by column chro-
matography [light petroleum ether/ethyl acetate (4:1)]. Syrup; TLC
light petroleum ether/ethyl acetate (1:1)]: R ϭ 0.74. Ϫ [α] ϭ Ϫ0.6
(446): calcd. 75.33, H
.72, O 17.93; found C 75.19, H 7.01, O 17.8.
L-
(
[
f
D
30 5
(c ϭ 1.8, dichloromethane). Ϫ C28H O
6
Study of the First Steps of the Glycosidation Reaction: Sugar 4 (304 heated for 6 h at 95°C in the presence of 300 mg of NaH suspen-
mg; 1.57 mmol) was dissolved in a suspension of 1 mL of Dowex-
sion. Methanol (1 mL) was added, the volatiles were evaporated,
and the residue was dissolved with ether which was washed with
ϩ
50 (H ) ion-exchange resin in 10 mL of methanol. The mixture
was stirred at room temperature for 2 h, when TLC [methanol/ water to neutrality. Purification by chromatography [light petro-
dichloromethane (2:3)] indicated the disappearance of the starting
material. The resin was filtered off and the methanol evaporated.
The residue was dissolved in 1.5 mL of DMF and 0.81 mL of tert-
leum ether/ethyl acetate (7:1)] separated 18 (428 mg; 71%). Ϫ
[α] ϭ Ϫ8.6 (c ϭ 1.2, dichloromethane). Ϫ C36 (568): calcd.
C 76.05, H 7.04, O 16.90; found C 75.97, H 7.11, O 17.08.
D
40 6
H O
Eur. J. Org. Chem. 1999, 1415Ϫ1420
1419

S. David
FULL PAPER
[
[
5]
6]
A. T. Nielsen, D. W. Moore, B. D. Schuetze, Jr., Pol. J. Chem.
Acknowledgments
1
981, 55, 1393Ϫ1403.
F. Effenberger, A. Straub, Tetrahedron Lett. 1987, 28,
The author is indebted to Professor A. Lubineau for the hospitality
of his laboratory and to Mme A. Malleron for her valuable assist-
ance in some physical determinations.
1
641Ϫ1644.
[
[
7]
8]
J. Aug e´ , S. David, Tetrahedron 1984, 40, 2101Ϫ2106.
S. Hanessian, P. L. Lavall e´ e, Can. J. Chem. 1975, 53,
2
975Ϫ2977.
[
9]
J. R. Durrwachter, C.-H. Wong, J. Org. Chem. 1988, 53,
4175Ϫ4181.
[
[
1]
2]
J. Yoshimura, Adv. Carbohydr. Chem. Biochem. 1984, 42,
9Ϫ134.
[10]
6
L. Banfi, G. Guanti, Eur. J. Org. Chem. 1998, 745Ϫ757.
R. L. Colbran, J. K. N. Jones, N. K. Matheson, I. Rozema,
Carbohydr. Res. 1967, 4, 355Ϫ358.
[11]
M. D. Bednarski, E. S. Simon, N. Bischofberger, W. D. Fessner,
M.-J. Kim, W. Lees, T. Saito, H. Waldmann, G. M. Whitesides,
J. Am. Chem. Soc. 1989, 111, 627Ϫ635.
[12]
E. Racker, J. Biol. Chem. 1950, 184, 313Ϫ319.
Received November 2, 1998
[
[
3]
4]
J. E. Vik, Acta Chem. Scand. 1973, 27, 239Ϫ250.
J. E. Vik, Acta Chem. Scand.1973, 27, 251Ϫ257.
[O98498]
1420
Eur. J. Org. Chem. 1999, 1415Ϫ1420