Photoinduced electron transfer and chemical α-deoxygenation of D-galactono-1,4-lactone. Synthesis of 2-deoxy-D-lyxo-hexofuranosides

Article abstract of DOI:10.1016/S0008-6215(02)00118-0

Two simple procedures for the synthesis of 2-deoxy-D-lyxo-hexono-1,4-lactone are described. Reductive cleavage of a 2-O-tosyl derivative of D-galactono-1,4-lactone in the presence of sodium iodide afforded the 2-deoxy derivative. On the other hand, α-deoxygenation of D-galactono-1,4-lactone was easily achieved by photochemical electron transfer deoxygenation of HO-2 as the 3-(trifluoromethyl)benzoate. Methyl 2-deoxy-β-D-lyxo-hexafuranoside ('methyl 2-deoxy-β-D-galactofuranoside') was synthesized and tested as substrate for exo β-D-galactofuranosidase from Penicillium fellutanum. The reaction was followed by HPAEC, showing that methyl 2-deoxy-β-D-galactofuranoside was not hydrolyzed by incubation with the enzyme. Neither the 2-deoxy lactone, nor the 2-deoxy-β-D-galactofuranoside acted as inhibitors of the reaction with the 4-nitrophenyl β-D-galactofuranoside. The present and our previous results show that the hydroxyl groups at C-2, C-3 and C-6 of the galactofuranoside are essential for interaction with the exo β-D-galactofuranosidase.

Full text of DOI:10.1016/S0008-6215(02)00118-0

Carbohydrate Research 337 (2002) 2119–2126
www.elsevier.com/locate/carres
Photoinduced electron transfer and chemical a-deoxygenation of
-galactono-1,4-lactone. Synthesis of
2-deoxy- -lyxo-hexofuranosides
D
D
Alejandro Chiocconi, Carla Marino, Eugenio Otal, Rosa M. de Lederkremer*
CIHIDECAR, Departamento de Qu´ımica Orga´nica, Facultad de Ciencias Exactas y Naturales, Uni6ersidad de Buenos Aires. Pabello´n II,
Ciudad Uni6ersitaria, 1428 Buenos Aires, Argentina
Received 8 March 2002; accepted 8 May 2002
Dedicated to Professor Derek Horton on the occasion of his 70th birthday
Abstract
Two simple procedures for the synthesis of 2-deoxy-
derivative of -galactono-1,4-lactone in the presence of sodium iodide afforded the 2-deoxy derivative. On the other hand,
a-deoxygenation of -galactono-1,4-lactone was easily achieved by photochemical electron transfer deoxygenation of HO-2 as the
3-(trifluoromethyl)benzoate. Methyl 2-deoxy-b- -lyxo-hexafuranoside (‘methyl 2-deoxy-b- -galactofuranoside’) was synthesized
and tested as substrate for exo b- -galactofuranosidase from Penicillium fellutanum. The reaction was followed by HPAEC,
showing that methyl 2-deoxy-b- -galactofuranoside was not hydrolyzed by incubation with the enzyme. Neither the 2-deoxy
lactone, nor the 2-deoxy-b- -galactofuranoside acted as inhibitors of the reaction with the 4-nitrophenyl b- -galactofuranoside.
The present and our previous results show that the hydroxyl groups at C-2, C-3 and C-6 of the galactofuranoside are essential
for interaction with the exo b- -galactofuranosidase. © 2002 Elsevier Science Ltd. All rights reserved.
D-lyxo-hexono-1,4-lactone are described. Reductive cleavage of a 2-O-tosyl
D
D
D
D
D
D
D
D
D
Keywords: Deoxy sugars; exo b-D-Galactofuranosidase; 2-Deoxy-D-lyxo-hexofuranosides; 2-Deoxy-D-lyxo-hexono-1,4-lactone
1. Introduction
co-workers2,3 We have synthesized substrates4,5 and in-
hibitors,6,7 and we recently studied the influence of the
inhibitors on the culture of the microorganism.8 An
affinity chromatography system for purification of the
enzyme was reported.9
The deoxy analogues of glycosides are useful for
studies of the specificity of glycosidases. Galactofura-
nosidases and galactofuranosyltransferases are of great
interest, because they are involved in the construction
and metabolism of important glycoconjugates of patho-
genic bacteria, protozoa and fungi.1 As galactose in the
furanoic configuration is not present in mammalian
glycoconjugates, these enzymes are good targets for the
development of antimicrobial agents. In this context,
Concerning the specificity of the exo b-D-galactofura-
nosidase, we have previously described simple ap-
proaches for the synthesis of 3-deoxy10 and 6-deoxy
analogues of the substrate,11 showing that the hydroxyl
groups at C-6 and C-3 of the galactofuranoside are
necessary for recognition by the enzyme. The synthesis
of glycosides deoxygenated at the 2-position is de-
scribed in the present work in order to gain further
knowledge about the glyconic specificity of this enzyme.
we have studied different aspects of the exo b-D-galac-
tofuranosidase from Penicillium fellutanum, with the
purpose of understanding the enzyme–substrate inter-
action. This enzyme was first studied by Gander and
D-Galactono-1,4-lactone (1) was chosen as precursor
of the furanoic sugar. The lactone was a-deoxygenated
by two alternative methods. The chemical synthesis
involved reductive cleavage of a 2-O-tosyl derivative in
the presence of sodium iodide to afford the 2-deoxy
* Corresponding author. Fax: +54-114-5763352
E-mail address: lederk@qo.fcen.uba.ar (R.M. de Lederkre-
mer).
0008-6215/02/$ - see front matter © 2002 Elsevier Science Ltd. All rights reserved.
PII: S0008-6215(02)00118-0

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A. Chiocconi et al. / Carbohydrate Research 337 (2002) 2119–2126
derivative. The other method is based on deoxygena-
tion via a photoinduced electron-transfer mechanism,
of HO-2 as the 3-(trifluoromethyl)benzoyl derivative.
The 2-deoxylactones were selectively reduced to the
corresponding sugars with diisoamyl borane12 and
glycosylated.
after interchange with D2O. In the 13C NMR spectrum
of 3 (Table 2), the signal corresponding to C-2 was
shifted downfield (79.1 ppm) with respect to the C-2
signal (74.6 ppm) of 2.
Successive treatment of 3 with acetic acid and either
benzoylation or acetylation led to compounds 5a,b.
Paulsen and Eberstein17 transformed a,b-epoxy-
uloses into a-deoxy-uloses by the action of sodium
iodide in acetone. In this reaction, the epoxide is
opened by the iodide, leading to an iodohydrin interme-
diate. The iodide of this intermediate is reduced to the
deoxy derivative by the action of another iodide. We
extended this reaction to the a-tosylated galactonolac-
tone. Thus, we treated derivative 5a,b with sodium
iodide in the presence of TFA to produce the 2-deoxy-
lactone 8a,b in 70% yield (Scheme 2). The presence of
two double doublets at high fields, with a large Jgem
On the other hand, it was interesting to test the
behavior of the 2-deoxy-
oxygalactofuranosides’) as substrates or inhibitors of
-galactofuranosidase. In this respect, methyl 2%-
D-lyxo-hexofuranosides (‘2-de-
the b-
D
deoxy-b-lactoside is
a
potent inhibitor of b-D-
galactopyranosidase.13
2. Results and discussion
1
We have previously obtained crystalline per-O-ben-
zoyl-a,b- -galactofuranose by direct benzoylation of
-galactose at high temperature.14 The mixture of the
(ꢀ18 Hz) in the H NMR spectrum (Table 1), and
signals at 34.9, 34.6 ppm in the 13C NMR spectrum
(Table 2) confirmed the identity of the deoxy lactones
8a,b. However, the acetylated derivative 5b afforded a
considerable proportion of the product of b-elimination
9, identified on the basis of the 13C NMR spectra of
analogous enonolactones18 (Table 2).
D
D
furanosic benzoates was used for the preparation of
galactofuranosyl glycosides and disaccharides.15 The
simplicity of the procedure led us to try direct benzoy-
lation of commercially available 2-deoxy-
D-lyxo-hexose
(‘2-deoxy- -galactose’) for the synthesis of the fura-
D
The other route relied on deoxygenation of the 3-
(trifluoromethyl)benzoyl derivatives 6 or 7 (Schemes 1
and 2), accomplished via a photoinduced electron-
transfer (PET) reaction using 9-methylcarbazole as
photosensitizer. This reaction was first used for the
synthesis of 2-deoxyribonucleosides19,20 and later ap-
plied to the deoxygenation of a disaccharide analogue
of moenomycin A.21 Now, we extended the PET reac-
tion to the synthesis of 2-deoxylactones. In our case,
photochemical deoxygenations were carried out with a
Heraeus TQ, medium-pressure Hg lamp in a Pyrex
reaction vessel. We first tried deoxygenation of the
perbenzoylated derivative 7 that was obtained similarly
to 5a. Monitoring the reaction by 13C NMR spec-
troscopy (Table 2), we observed in the methylene re-
gion, not only the signal at 34.9 ppm corresponding to
C-2 of compound 8a, but also signals at l 32.6 and 27.9
(see Section 3), indicating that the deoxygenation was
not regioselective and that benzoyl groups had been
also reduced. On the other hand, the 3-(trifl-
uoromethyl)benzoyl derivative 6, with only the 2-O-es-
ter group, was deoxygenated in 1 h. The NMR spectra
nosic benzoates. In this case a mixture with a low
proportion of a,b-furanosic forms (ꢀ30%), was ob-
tained as indicated by a 13C NMR spectrum.
As for the preparation of the 3-deoxy10 and 6-deoxy
analogues,11 we used
D-galactono-1,4-lactone (1) as the
precursor of the 5-membered ring, with the advantage
of the differential reactivity of the HO-2 due to the
inductive b-carbonyl effect. Selectivity between HO-2
and -6 is not possible with the more common protective
groups like benzoyl. With the aim of obtaining a
derivative of 1 with HO-2 selectively substituted, we
treated 5,6-O-isopropylidene-D-galactono-1,4-lactone
(2)16 with one equivalent of 4-toluenesulfonyl chloride
in acetone–pyridine at 0 °C. This reaction gave re-
giospecifically compound 3 (Scheme 1), as evidenced
from its NMR spectra (Tables 1 and 2). The signal
corresponding to H-3 (4.69 ppm) was coupled (J3,OH 2.9
Hz) with that observed at 3.5 ppm, which disappeared
confirmed that 2-deoxy- -lyxo-hexono-1,4-lactone (10)
D
was obtained as the only product (Tables 1 and 2).
Lactone 10 was previously prepared by oxidation of
2-deoxy-D
-lyxo-hexose with bromine.22,23
The lactonic group of derivatives 8a,b was reduced to
the corresponding lactols 11a,b (Scheme 3) with di-
isoamylborane.12 After workup and several evapora-
tions with methanol to eliminate boric acid, TLC
analysis showed the anomeric mixture of the free lactol
as the main product along with a less polar product
Scheme 1. (i) 2,2-Dimethoxypropane–acetone; (ii) R1Cl, pyri-
dine, −10 °C; (iii) 4:1 AcOH–H2O, 50–60 °C; (iv) R2Cl,
pyridine.

A. Chiocconi et al. / Carbohydrate Research 337 (2002) 2119–2126
2121
Table 1
1H NMR (200.1 MHz, CDCl3) chemical shifts and JH,H coupling constants values for compounds 3–8, 10, 13, 14
Compd
H-1
H-2
H-2%
H-3
H-4
H-5
H-6
H-6%
(J1,2) (J1,2%
)
(J2,3
)
(J2%,3) (J2,2%
)
(J3,4
)
(J4,5
)
(J5,6
)
(J5,6%) (J6,6%)
3
4.99
(7.9)
5.56
(7.6)
5.63*
4.69
(8.2)
4.67
(7.3)
5.63*
(5.12)
5.42
(5.48)
4.65
(8.4)
5.93
(5.8)
5.55
4.17
(3.3)
4.34
(3.3)
4.91
(2.19)
4.54
(2.2)
4.40
(1.8)
5.06
(2.7)
5.04
(2.2)
4.67
4.36
(6.8)
4.43
(7.0)
5.91
4.13
4.15
4.00
(7.0) (8.8)
4.11
(6.6) (8.8)
4.66*
4
5a
5b
5.23
(5.84)
5.94
(9.8)
6.05
(6.2)
3.01
(7.7)
2.91
(7.3)
3.01
(7.2)
2.37
(7.2)
2.37
(8.4)
2.19
(6.8)
2.29
(7.5)
5.32
(5.20)
3.76
4.28
4.20
(6.2) (11.7)
4.47*
6a
7
6.00
(6.3)
5.98
(5.1)
5.36
(5.5)
3.89
(5.5)
5.25
(4.1)
5.29
(4.4)
4.79
4.72
4.33
3.64
4.34
4.32
4.70
(6.2) (12.0)
4.64
(7.0) (11.7)
4.21
(6.6) (11.7)
3.60
(7.2) (11.5)
4.17*
(6.4) (11.8)
4.21
(7.1) (11.6)
3.56
8a
2.75
(1.8) (17.8)
2.54
(2.2) (18.6)
2.51
(2.8) (18.7)
2.07
(–) (13.5)
1.95
(2.5) (14.5)
2.10
(6.5) (13.8)
1.85
(2.5) (14.4)
8b
5.19
10b
13bac
13bbc
14ac
14bc
4.57
(5.5)
5.21
4.52
(2.6)
4.17*
(3.2)
4.23
(3.9)
3.84
(5.0)
3.97
(3.7)
5.09
(2.2) (5.4)
5.09
(1.1) (5.5)
5.15
(2.4) (5.4)
5.12
4.97
(4.5)
4.40
3.66*
4.30
3.75
(4.6)
3.65
(7.8)
3.57
(7.8) (11.7)
(5.5) (1.2)
a DMSO-d6.
b D2O.
c 500 MHz.
* Center of a complex multiplet.
that was isolated by column chromatography and char-
acterized as the anomeric methyl glycosides 13a,b
(15%). In the case of the acetylated lactone 8b, rear-
rangement to the pyranosic lactol was also observed.
This result, and the fact that b-elimination of 5b could
not be avoided during the reduction with NaI, led us to
choose the benzoylated derivatives for the synthetic
sequence.
Lactol 11a was acetylated to give 12 (Scheme 3) with
the purpose to activate the anomeric HO for glycosyla-
tion reactions. However, many efforts for glycosylation
of 12 were unsuccessful. Treatment of 12 with SnCl4,
and addition of either methanol24 or 4-nitrophenol11 (or
the tetrabutylammonium salt of 4-nitrophenol5), did
not afford the corresponding glycosides, indicating that
in comparison with the galactofuranose derivative, the
absence of HO-2 dramatically changes the reactivity of
the anomeric center.
promoter of the glycosylation reaction. Thus, com-
pounds 11a,b or 12 were treated with BF3·MeOH, and
methyl glycosides 13a,b were obtained in both cases in
ꢀ90% yield as a 1:1 anomeric mixture. The lack of
stereoselectivity in this reaction was predictable, due to
the absence of a participating group at C-2. The
anomeric mixture 13a was chromatographically homo-
geneous, but 13b could be resolved by column chro-
1
matography. However, the H NMR spectra of both
anomers of 13b were almost identical because of the
absence of a substituent at C-2. Nevertheless, the
anomeric configuration could be assigned on the basis
of the coupling constants observed in the 1H NMR
spectrum (Table 1).
The anomeric mixture 13a was debenzoylated, and
the free glycosides 14a and 14b were separated by fast
silicagel column chromatography. The hydrolytic insta-
bility of the 2-deoxyglycofuranosides was shown by the
The fact that lactols 11a,b had been easily glycosy-
lated during the workup of the diisoamylborane reduc-
tion suggested that boron trifluoride could be a good
formation of 2-deoxy-D-galactose during the purifica-
tion. The more polar product (48%) gave [h]D −65°,
1
suggesting a b configuration, in agreement with the H

2122
A. Chiocconi et al. / Carbohydrate Research 337 (2002) 2119–2126
Table 2
D
-lyxo-hexofuranoside (14a) was isolated with 48%
13C NMR chemical shifts (50.3 MHz, CDCl3) for compounds
yield and gave [h]D 40°. The methyl a,b glycosides 14,
were prepared by Fisher glycosylation, but they were
not fully characterized.23
We also considered the advantage of performing the
deoxygenation step after the reduction of the lactone
group and with further glycosylation so the anchimeric
effect of the substituent at HO-2 would be effective for
stereoselective b-glycosylation. However, neither chemi-
cal deoxygenating of the 2-O-tosyl glycosides with NaI,
2–14
Compd C-1
C-2
74.6
C-3
74.4*
C-4
79.9
C-5
C-6
2
3
4
174.5
74.1* 65.0
166.7
169.4
166.7
166.5
170.1
171.6
173.7
173.7
79.1
77.6
76.0
75.8
76.0
73.6
34.9
34.6
73.6
73.8
73.6
72.6
69.9
73.0
72.1
71.0
79.6
79.7
79.5
78.7
79.9
78.8
83.2
82.4
73.0
73.3
69.5
68.3
67.9
69.8
70.8
70.5
64.9
65.0
62.1
61.4
61.6
62.3
62.3
61.5
58.6
58.3
62.9
63.9
63.4
63.1
63.1
62.8
62.8
63.4
5a
5b
6a
7
8a
8b
9Z
9E
10b
nor PET deoxygenation of b-D-galactofuranosides
derivatized with 3-(trifluoromethyl)benzoyl at HO-2,
were successful.
This is the first report on the PET reaction applied to
an aldonolactone, and the effectiveness of this strategy
could be attributed to the combination of two factors:
the selectivity of the substitution because of the lactonic
carbonyl group, and the activation of the photochemi-
cal reaction, due to the formation of a radical interme-
diate stabilized by conjugation with the carbonyl group.
However, the stability of the resulting radical would
not account for deoxygenation at other ring carbons
when benzoyl groups are present. Further work is in
progress to identify the products and to study the
kinetics of this reaction.
138.6 124.5
140.3 121.0
146.5 110.7
146.9 109.8
180.4
38.4
39.4
39.3
38.4
38.5
39.0
39.1
41.0
41.2
71.7
75.1
74.6
74.1
74.6
73.9
73.7
73.0
72.2*
88.9
82.3
82.3
83.2
84.3
81.6
80.6
86.3
86.1
69.8
71.8
71.1
70.8
71.0
70.9
70.2
71.7
11aa,b 104.4
103.7
98.5
97.9
12a,b
13ba,b 105.2
104.9
105.9
106.3
14ab
14bb
72.4* 63.6
a DMSO-d6.
The biological interactions of the exo b-D-galacto-
b D2O.
furanosidase from P. fellutanum with the 2-deoxylac-
tone 10 and the 2-deoxyglycosides 14a and 14b were
* Interchangeable.
investigated. We have previously reported6 that
D-
NMR spectrum (Table 1). The H-2%, in a trans relation-
ship with both H-3 and H-1, indicated that these hydro-
gens were oriented to the same face, so the anomeric
configuration was b. The less polar methyl 2-deoxy-a-
galactono-1,4-lactone (1) is a good inhibitor of b-
D
-
galactofuranosidase. Thus, we studied the inhibitory
properties of the 2-deoxylactone 10 by incubating the
enzyme in the presence of both the substrate 4-nitro-
Scheme 2. (i) NaI, TFA, acetone; (ii) 9-methylcarbazole, 9:1 n-PrOH–H2O, hw; (iii) HCl, aq MeOH, reflux.

A. Chiocconi et al. / Carbohydrate Research 337 (2002) 2119–2126
2123
Scheme 3. (i) DSB, THF; (ii) Ac2O, pyridine; (iii) BF3, MeOH; (iv) MeONa, MeOH.
phenyl b-
D
-galactofuranoside4,6 and compound 10. We
200 spectrometer at 200 MHz (1H) and 50 MHz (13C)
or with a Bruker AM 500 spectrometer at 500 MHz
(1H) and 125 MHz (13C).
also examined the hydrolytic activities on the 2-deoxy-
galactofuranosides 14a and 14b and their inhibitory
properties.
Benzoylation of 2-deoxy-
perature.—The procedure previously described for
the preparation of penta-O-benzoyl-a,b- -galacto-
furanose14 was adapted for 2-deoxy-
-lyxo-hexose. A
solution of 2-deoxy- -lyxo-hexose (0.5 g, 2.77 mmol) in
D-lyxo-hexose at high tem-
After incubation, the enzymatic mixtures were ana-
lyzed by high-pH anion-exchange chromatography with
pulse amperometric detection (HPAEC–PAD), which
proved to be an excellent method for the resolution of
2-deoxygalactose and the methyl glycosides 14a,b (Fig.
1). The absence of 2-deoxygalactose indicated that no
hydrolysis of 14b occurred (not shown). On the other
hand, the activity of the enzyme towards 4-nitrophenyl
D
D
D
anhyd Py (7.0 mL) was stirred for 2 h in a water bath
at 100 °C. The temperature was lowered to 60 °C, and
BzCl was added dropwise. After 1.5 h at 60 °C the
mixture was poured into ice-water, and the syrup that
separated was dissolved in CH2Cl2 and sequentially
washed with 5% HCl, satd aq NaHCO3, and then dried
(Na2SO4). The syrup obtained after evaporation of the
solvent was a mixture of a-pyranosic and a,b-furanosic
forms. 13C NMR (CDCl3) inter alia, l: 99.3, 99.2 (C-1
a,b-f ), 92.4 (C-1 a-p), 84.9, 83.6 (C-4 a,b-f ), 63.4 (C-6
a,b-f ), 62.4 (C-6 a-p), 38.9 (C-2 a,b-f ), 31.5 (C-2 a-p).
From the C-2 signals a 30% content of furanosic forms
was estimated.
b-D-galactofuranoside as substrate, was not affected by
the presence of 10, 14a or 14b, indicating that these
compounds do not act as inhibitors, and thus no inter-
action with the enzyme takes place in the absence of
HO-2. The methyl b-
D-galactofuranoside had been used
as substrate for the determination of the exo b-
D-galac-
tofuranosidase of P. fellutanum.2 Many enzymes show
broad substrate specificity, and deoxygenated analogues
of the substrates can be hydrolyzed with a characteristic
kinetics,25 or act as inhibitors of the glycosidase.12
However, the present and our previous results9,10 show
that the exo b-D-galactofuranosidase from P. fellu-
tanum has a strict selectivity against the glycon struc-
ture, and that the hydroxyl groups at C-2,3 and 6 of the
substrate are essential for interaction with the enzyme.
3. Experimental
General
methods.—Thin-layer
chromatography
(TLC) was carried out on 0.2 mm Silica Gel 60 F254 (E.
Merck) aluminum supported plates, using the following
solvents: (a) 9:1 toluene–EtOAc, (b) 20:1 toluene–
EtOAc, (c) 8:1 EtOAc–MeOH, (d) 19:1 EtOAc–
MeOH. Detection was effected by exposure to UV light
and by spraying with 10% (v/v) H2SO4 in EtOH and
charring. Column chromatography was performed on
Silica Gel 60 (200–400 mesh, E. Merck). Optical rota-
tions were measured with a Perkin–Elmer 343 polar-
imeter. NMR spectra were recorded with a Bruker AC
Fig. 1. HPAEC–PAD separation of 2-deoxy-
(I), methyl 2-deoxy-b- -lyxo-hexofuranoside (II) and methyl
2-deoxy-a- -lyxo-hexofuranoside (III). A CarboPac MA-10
D-lyxo-hexose
D
D
anion-exchange column under conditions described in Section
3 was used.

2124
A. Chiocconi et al. / Carbohydrate Research 337 (2002) 2119–2126
5,6-Di-O-isopropylidene-2-O-tosyl-
lactone (3).—To a solution of 5,6-di-O-isopropylidene-
-galactono-1,4-lactone16 (2, 5.8 g, 26 mmol) in dry Py
D
-galactono-1,4-
procedure described for 5a, but using Ac2O instead of
BzCl; mp 155–156 °C, [h]D +15° (c 1, CHCl3).
3,5,6-Tri-O-benzoyl-2-O-[3 -(trifluromethyl)benzoyl]-
D
D-galactono-1,4-lactone (7).—Compound 6 (1.50 g,
(25 mL), cooled at 0 °C, 4-toluenesulfonyl chloride (5.6
g, 29 mmol) diluted in acetone (12 mL) was added
during 1 h. After stirring for 4 h at 0 °C, the mixture
was kept at 4 °C overnight. The mixture was then
poured into ice-water and the syrup that separated was
dissolved in CH2Cl2, and washed with HCl (5%), water,
satd aq NaHCO3, water, and then dried (MgSO4).
After evaporation of the solvent, the syrup that was
obtained was recrystallized from toluene to afford com-
pound 3 (6.8 g, 70%), mp 140–141 °C, [h]D −94° (c 1,
CHCl3). Anal. Calcd for C16H20O8S: C, 51.61; H, 5.41.
Found: C, 51.85; H, 5.23.
4.10 mmol) was treated with 1:1 BzCl–Py (10.0 mL) at
0 °C for 3 h. After workup, compound 7 was obtained
as a syrup (2.65 g, 93%), Rf 0.63 (solvent a), [h]D+2° (c
1, CHCl3). Anal. Calcd for C37H32F3O10: C, 64.07; H,
4.65. Found: C, 64.32; H, 4.50.
Deoxygenation step—(a) Chemical h-deoxygenation.
3,5,6-Tri-O-benzoyl-2-deoxy-D-lyxo-hexono-1,4-lactone
(8a). To a solution of compound 5a (1.0 g, 1.6 mmol) in
acetone (18 mL), trifluoroacetic acid (2 mL) and NaI
(5.0 g, 33.0 mmol) were added. After 16 h of stirring the
dark, mixture was diluted with CH2Cl2, washed sequen-
tially with satd aq solutions of NaHSO3, NaHCO3,
NaCl, and water and dried (MgSO4). The syrup ob-
tained after evaporation of the solvent was purified by
column chromatography (20:1 toluene–EtOAc), recrys-
tallized from EtOH (0.53 g, 70%), to give a solid with
mp 137–138 °C, [h]D −35° (c 1, CHCl3). Anal. Calcd
for C27H22O8: C, 68.35; H, 4.67. Found: C, 68.50; H,
4.50.
5,6-Di-O-isopropylidene-2-O-[3-(trifluromethyl)benz-
oyl]-
D
-galactono-1,4-lactone (4).—To a solution of 5,6-
di-O-isopropylidene-
D
-galactono-1,4-lactone16 (2, 1.0 g,
4.5 mmol) in CH2Cl2 (15 mL), cooled at −15 °C, dry
Py (1.5 mL) was added. 3-(Trifluromethyl)benzoyl chlo-
ride (0.81 mL, 5.4 mmol) diluted in CH2Cl2 (5.0 mL)
was added during 2 h. After stirring for 4 h at −15 °C,
the mixture was poured over ice-water and the resulting
syrup was dissolved in CH2Cl2, and washed with HCl
(5%), water, satd aq NaHCO3, water, and then dried
(MgSO4). Evaporation of the solvent and recrystalliza-
tion from toluene afforded compound 4 (2.23 g, 68%),
mp 166–168 °C, [h]D 21° (c 1.3, CHCl3). Anal. Calcd
for C17H17F3O7: C, 52.31; H, 4.39. Found: C, 52.26; H,
4.62.
3,5,6-Tri-O-acetyl-2-deoxy-D-lyxo-hexono-1,4-lactone
(8b).—Compound 8b was prepared from 5b (1.0 g, 2.2
mmol) by the procedure described for 8a. After workup
TLC examination of the syrup obtained showed a main
product (Rf 0.25, solvent b) and minor amount of a
faster moving product (Rf 0.40), which were separated
by column chromatography (20:1 toluene–EtOAc). The
product of Rf 0.40 was spectroscopically identified as
6-O-acetyl-2-O-tosyl-hexa-2,4-dien-4-olide (9) by com-
parison with the spectra for 2,6-dibenzoyloxy-2,4-hexa-
dien-4-olide.18 Fractions containing the product of Rf
0.25 were evaporated, and the syrup obtained (8b, 0.38
g, 60%) gave the same spectroscopic properties as those
reported.22
2-O-[3-(Trifluromethyl]benzoyl-D-galactono-1,4-lac-
tone (6).—Compound 4 (2.0 g, 5.73 mmol) was treated
with 4:1 AcOH–water (40 mL) for 1.5 h at 60 °C. The
solvent was evaporated under reduced pressure, and the
remaining acid was removed by several evaporations
with water. A white solid was obtained (1.68 g, 90%)
that upon recrystallization from water gave Rf 0.65
(solvent c), mp 176–177 °C, [h]D −7° (c 1, CHCl3).
Anal. Calcd for C15H16F3O7: C, 49.32; H, 4.41. Found:
C, 49.27; H, 4.18.
(b) Photochemical deoxygenation. General proce-
dure.—In
a
custom-made Pyrex reaction vessel
equipped with a cold finger, a solution containing 0.15
mmol of the substrate, Mg(ClO4)2 (66 mg, 0.3 mM),
and methylcarbazole (3 mg, ꢀ0.015 mmol) in 100 mL
of 10% deionized water–2-propanol was degassed by
bubbling UHP Ar through the solution for 30 min. The
reaction was photolyzed with a Heraeus TQ medium-
pressure Hg lamp, while the temperature was main-
tained at 25 °C with a circulating water bath. The
following compounds were photolyzed:
3,5,6-Tri-O-benzoyl-2-O-tosyl-D-galactono-1,4-lac-
tone (5a).—Compound 2 (1.5 g, 4.5 mmol) was treated
with 4:1 AcOH–water (30 mL) at 60 °C during 1.5 h.
After evaporation of the solvent and several co-evapo-
rations with water, the syrup obtained was dried and
treated with a solution of Py–BzCl (1:1, 10 mL) at
0 °C. After 2 h of stirring the mixture was poured over
ice-water. The resulting syrup was extracted with
CH2Cl2, and the extract was washed as usual and dried.
Benzoic acid was eliminated by dissolution of the syrup
in Et2O and precipitation with hexane. Compound 5a
(2.30 g, 90%) was characterized as a syrup that gave
[h]D+38° (c 1, CHCl3). Anal. Calcd for C34H28O11S: C,
63.35; H, 4.38. Found: C, 63.10; H, 4.50.
(i) 3,5,6-Tri-O-benzoyl-2-O-[3-(trifluromethyl)benzo-
yl]- -galactono-1,4-lactone (7, 0.10 g, 0.15 mmol). TLC
D
analysis after 3 h of reaction showed a product with the
same Rf as 8a (Rf 0.25, solvent a). The reaction mixture
was evaporated to remove the 2-PrOH, and the aq
residue was extracted with CH2Cl2. The organic phase
was washed with water, dried (MgSO4) and evaporated.
13C NMR (CDCl3) methylene region, l 34.9, 32.6 and
27.9.
3,5,6-Tri-O-acetyl-2-O-tosyl-D-galactono-1,4-lactone
(5b).—Compound 5b was obtained in 96% yield by the

A. Chiocconi et al. / Carbohydrate Research 337 (2002) 2119–2126
2125
(ii) 2-O-[3-(Trifluromethyl]benzoyl-
D
-galactono-1,4-
incubated with 4-nitrophenyl b-
D
-galactofuranoside4 as
lactone, (6, 0.06 g, 0.16 mmol). After 1 h TLC analysis
showed a single spot (Rf 0.64, solvent c). The solvent
was evaporated, and the residue was dissolved in
MeOH and filtered, and the filtrate was evaporated to
a control reaction, or with the deoxylactone 10 or the
methyl glycosides 14a,b, for studying the specificity of
the enzyme, or with each of these compounds together
with 4-nitrophenyl b-D-galactofuranoside, for the inhi-
give 2-deoxy-
D-lyxo-hexono-1,4-lactone (10, 0.02 g,
bition tests. The assays were carried out by incubating
100 mL of the culture medium containing the enzyme
(20 mg of protein) with 60 mL of a 5 mM solution of the
substrate, 100 mL of 66 mM AcONa buffer (pH 4.0),
and the inhibitors (1 mM) in a final volume of 500 mL.
The enzymatic reaction was stopped after 1.5 h incuba-
tion at 37 °C by adding 1 mL of 0.1 M Na2CO3 buffer
(pH 9.0). The released 4-nitrophenol was measured
spectrophotometrically at 410 nm.
88%), [h]D −10° (c 1, water), in agreement with litera-
ture data.23
3,5,6-Tri-O-benzoyl-2-deoxy- -lyxo-hexose (11a).—
D
To a solution of freshly prepared bis(3-methyl-2-bu-
tyl)borane (5.6 mmol) in anhyd THF (5.0 mL),12
compound 8a (0.66 g, 1.4 mmol) was added. The
solution was stirred for 16 h at rt and then processed as
already described.11 After column chromatography
purification (10:1 toluene–EtOAc), compound 11a
(0.63 g, 95%) was obtained as an amorphous solid, [h]D
−19° (c 1, CHCl3). Anal. Calcd for C27H24O8: C,
68.06; H, 5.08. Found: C, 68.30; H, 4.90.
For the specificity studies, incubation at 37 °C was
performed overnight. The sample was centrifuged for
25 min at 10000×g through an Ultrafree-MC centrifu-
gal filter (MW 5000). The filtrate was analyzed by TLC
and HPAEC–PAD.
1-O-Acetyl-3,5,6-tri-O-benzoyl-2-deoxy-h,i-D-lyxo-
hexofuranose (12).—Compound 11a (0.67 g, 1.4 mmol)
was conventionally acetylated (1:1, Ac2O–Py, 10.0 mL)
to afford compound 12 (0.70 g, 96%), as a 1:1 anomeric
mixture that gave [h]D −26° (c 1, CHCl3). Anal. Calcd
for C29H26O9: C, 67.18; H, 5.05. Found: C, 67.04; H,
5.16.
HPAEC–PAD analysis.—Analysis by HPAEC–
PAD was performed using a Dionex DX 300 high
performance liquid chromatography (HPLC) system
with pulse amperometric detection (PAD), set at 30 nA
and E1= +0.05 V, E2= +0.60 V, and E3= −0.60 V.
The column used was a CarboPac MA-10 anion-ex-
change analytical column (4×250 mm) equipped with
a guard column MA-10 (4×50 mm). The separations
were performed isocratically in 70 mM NaOH at a flow
Methyl 3,5,6-tri-O-benzoyl-2-deoxy-D-lyxo-hexofura-
noside (13a).—A solution of either 11a (0.27 g, 0.57
mmol) or 12 (0.29 g, 0.57 mmol) in 14% BF3–MeOH
(10.0 mL) was stirred for 3 h at rt. The solvent was
evaporated, and the syrup obtained was dissolved in
CH2Cl2, washed with NaHCO3, water and dried
(MgSO4). After evaporation of the solvent, the syrup
was purified by column chromatography (20:1 toluene–
EtOAc). Compound 13a (0.25 g, 90%) was obtained as
an anomeric mixture. Anal. Calcd for C28O26O8. C,
68.56; H, 5.34. Found: C, 68.44; H, 5.16.
rate of 0.4 mL min−1
.
Acknowledgements
We are indebted to Agencia Nacional de Promocio´n
Cient´ıfica (ANPCYT, BID 802/OC-AR PICT 06-3036
and BID 1201/OC-AR-PICT 06-05133), University of
Buenos Aires and CONICET (Consejo Nacional de
Investigaciones Cient´ıficas y Tecnolo´gicas) for financial
support. R.M. de L. is a Research member of
CONICET.
Methyl 2-deoxy-h-D-lyxo-hexofuranoside (14a) and
methyl 2-deoxy-i- -lyxo-hexofuranoside (14b).—The
D
anomeric mixture of 13a (0.40 g, 0.80 mmol) was
suspended in a 0.5 M solution of NaOMe in MeOH
(5.0 mL) and stirred until complete dissolution occurred
(2 h). The solution was neutralized with Dowex 50W
(H+) and concentrated. Methyl benzoate was elimi-
nated by repeated evaporation with water. TLC exami-
nation showed the presence of two products of Rf 0.40
and 0.30 (solvent d). After column chromatography
purification (19:1 EtOAc–MeOH) compound 14a (0.08
g, 55%) was obtained as a syrup that gave [h]D 40° (c
0.3, water). Fractions containing the product of Rf 0.3
(14b), were evaporated (0.06 g, 41%) and gave [h]D
−65° (c 0.3, water). Anal. Calcd for C7H14O5: C, 47.19;
H, 7.92. Found: C, 46.41; H, 8.09.
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