Synthesis of potassium (2R)-2-O-α-D-mannopyranosyl-(1→2)- α-D-glucopyranosyl-2,3-dihydroxypropanoate: A naturally compatible solute

Article abstract of DOI:10.1002/ejoc.201100934

An expedient synthesis of the potassium salt of (2R)-2-O-α-D- mannopyranosyl-(1→2)-α-D-glucopyranosyl-2,3-dihydroxypropanoic acid (MGG)a-a recently isolated, rare, compatible solutea-was accomplished. A bis-acetal-protected thioglucoside, 6-OTBDPS, with a

Full text of DOI:10.1002/ejoc.201100934

FULL PAPER
DOI: 10.1002/ejoc.201100934
Synthesis of Potassium (2R)-2-O-α-
D
-Mannopyranosyl-(1Ǟ2)-α- -
D
glucopyranosyl-2,3-dihydroxypropanoate: A Naturally Compatible Solute
Eva C. Lourenço[a] and M. Rita Ventura*[a]
Keywords: Glycosylation / Hypersolutes / Natural products / Thioglycosides / Solid-phase synthesis
An expedient synthesis of the potassium salt of (2R)-2-O-α-
-mannopyranosyl-(1Ǟ2)-α- -glucopyranosyl-2,3-dihydroxy-
propanoic acid (MGG)Ϫa recently isolated, rare, compatible
soluteϪwas accomplished. A bis-acetal-protected thiogluco-
side, 6-OTBDPS, with a 2-OH group was used as the ac-
ceptor in the first glycosylation reaction with tetraacetylman-
nosyl trichloroacetimidate, and as the donor in the glycosyl-
ation reaction with the glycerate derivative. The α anomer
was the only product of both glycosylation reactions, as ex-
pected for the formation of the α-mannoside. The formation
of the 1,2-cis glucoside was more challenging.
D
D
Introduction
Compatible solutes are low-molecular-mass organic
compounds that accumulate in the cytoplasm of many halo-
philic or halotolerant organisms to counterbalance the os-
motic pressure of the external medium.[1] Hyperthermo-
philes accumulate compatible solutes that exhibit thermo-
protection against heat denaturation of a variety of pro-
teins.[2,3] The thermostabilization properties of hypersolutes
have enormous potential for industrial and health-related
applications.
In contrast with the neutral or zwitterionic nature of the
solutes commonly found in mesophiles, organisms with op-
timal growth temperatures above 60 °C accumulate mainly
negatively charged compounds, such as α-d-mannosyl-
(1Ǟ2)-d-glycerate (MG), α-d-glucosyl-(1Ǟ2)-d-glycerate
(GG), α-d-glucosyl-(1Ǟ6)-α-d-glucosyl-(1Ǟ2)-d-glycerate
(GGG) (Figure 1) and di-myo-inositol-1,3Ј-phosphate
(DIP). The trisaccharide GGG, which was isolated from hy-
perthermophile Persephonella marina, has recently been
synthesized, along with related GG (Figure 1).[4]
A new solute, the structure of which was established to
be (2R)-2-O-α-d-mannopyranosyl-(1Ǟ2)-α-d-glucopyrano-
syl-2,3-dihydroxypropanoate [1, α-d-mannopyranosyl-
(1Ǟ2)-α-d-glucopyranosyl-(1Ǟ2)-d-glycerate (MGG)], was
isolated from the thermophilic bacteria Petrotoga mi-
otherma[5] and Petrotoga mobilis[6] (Figure 1). MGG is a
rare solute and it serves as an osmolyte during low-level
osmotic adaptation in Ptg. miotherma, but it is not used
against oxidative or heat stress by the cell.[5] In contrast, in
Ptg. mobilis, MGG is the major compatible solute under
osmotic and thermal stresses.[5] It was shown that MGG
Figure 1. Natural glycerate solutes.
was an efficient protector of pig-heart malate dehydro-
genase against heat inactivation and freeze-drying and has
been patented for the stabilization and preservation of bio-
materials.[5]
This solute is available in very small quantities from its
natural sources and chemical synthesis was considered the
best route to obtain 1 in larger amounts.
Results and Discussion
During the analysis of the structure of 1, two immediate
critical issues emerged: the need for the C-2 hydroxy group
of glucose to be free[7] to form a glycosidic bond with man-
nose and the stereoselective formation of the 1,2-cis glucos-
idic bond with glycerate. Additionally, we hoped to perform
the synthesis on a solid support to avoid the need for diffi-
cult and time-consuming separations, such as silica and sol-
vent-consuming chromatographies. A two-way strategy was
[a] Instituto de Tecnologia Química e Biológica,
Universidade Nova de Lisboa,
Apartado 127, 2780-901 Oeiras, Portugal
E-mail: rventura@itqb.unl.pt
6698
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Synthesis of a Naturally Compatible Solute
considered to be suitable; therefore, we needed to prepare a
donor/acceptor with the 6-OH of the glucose unit differen-
tially exposed to attach this donor/acceptor to a suitable
resin. However, a solution-phase synthesis was carried out
to test the strategy and to later compare this strategy with
the solid-phase one.
Butane-2,3-bis-acetals have been used in chiral memory
studies with tartaric acid[8] and extensively used to protect
vicinal diequatorial hydroxy groups in sugars.[9,10] An easily
separable 1:1 mixture of bisacetals 2 and 3 was formed
(Scheme 1)[11,12] by treating phenylthioglucopyranoside[13]
with 2,2,3,3-tetramethoxybutane (TMB). It was possible to
recycle isomer 3 by heating it in methanol at reflux with
catalytic CSA overnight; this again afforded a 1:1 mixture
of isomers, increasing the overall yield of 2 to 70%
(Scheme 1).
Scheme 2. Reagents and conditions: (a) TBDPSCl, imidazole,
DMF, r.t., 89%; (b) trimethylsilyl trifluoromethanesulfonate
(TMSOTf), CH2Cl2, –20 °C, 58%; (c) N-iodosuccinimide (NIS),
trifluoromethanesulfonic acid (TfOH), 4 Å molecular sieves,
CH2Cl2, 0 °C, 60%.
Scheme 1. Reagents and conditions: (a) Camphorsulfonic acid
(CSA), MeOH, Δ, 91%.
Selective silylation of the primary hydroxy group of 2 by
using tert-butyldiphenylsilyl chloride (TBDPSCl) and imid-
azole afforded 4 in 89% yield (Scheme 2). A glycosylation
reaction between thioglucoside 4 as the acceptor and man-
nose trichloroacetimidate 5, using TMSOTf as the pro-
moter,[14] afforded exclusively, as expected, the correspond-
ing α-mannoside 6 in 58% yield. The second glycosylation
reaction, using disaccharide 6 as the donor and glycerate 7
by employing NIS/TfOH as the activating system,[12] was
also highly selective and afforded α-glucoside 8 in 60% yield
(Scheme 2). It has been reported that the α selectivity of
glucosylations was increased when bulky substituents, such
as TBDPS, were introduced at the 6-position of the glucosyl
donor.[15] The main reason for the moderate yields was the
partial hydrolysis of the acetal under the glycosylation con-
ditions, which we were unable to avoid.
Scheme 3. Reagents and conditions: (a) TBAF, THF, r.t., 71%;
(b) TFA/CH2Cl2/H2O, r.t., 60%; (c) MeONa, MeOH, r.t., quant.
(d) KOH, H2O, r.t., quant.
Selective fluorolysis of the silyl ether of 8 with tetra- ment of 12 to Tentagel MB-NH2 resin was successfully ac-
butylammonium fluoride (TBAF; 71%), followed by complished by using standard carbodiimide coupling.
cleavage of the acetal with trifluoroacetic acid (TFA) in Resin-bound 13 was characterized by high-resolution magic
dichloromethane/water (60%), methanolysis of the acetates angle spinning (HR-MAS) NMR spectroscopy experi-
of the mannosyl unit, and hydrolysis of the methyl ester of ments. This technique was an excellent way of monitoring
the glycerate afforded 1 (Scheme 3).
the solid-supported synthesis by the analysis of high-quality
The NMR spectroscopic data for the synthesized product 1H, 13C, and HSQC NMR spectra. From the dry-weight
were identical to those described in the literature[5] for natu- difference, an initial loading of 0.32 mmolgϪ1 of resin was
ral MGG (Table 1).
estimated.
The glycosylation reactions were performed with poly-
For the solid-supported synthesis of 1, our strategy con-
sisted of attaching the thioglucoside to a Tentagel MB NH2 mer-bound glucoside 13, as indicated in Scheme 4, and we
resin through the formation of an amide bond between the obtained immobilized fully protected 15. Both glycosylation
carboxylic acid of the hemisuccinic acid moiety at C-6 and reactions afforded the corresponding α products, which was
the amino group of the Tentagel resin.[16–18] The hemisuc- expected in the first reaction. For subsequent formation of
cinate was selectively formed at the C-6 hydroxy of diol 2 the 1,2-cis glucoside 15, the succinate ester exerted the same
by using succinic anhydride to afford 12 in good yield (90%; orienting effect as the acetate ester in the 6-position of the
Scheme 4) and left the hydroxy group at C-2 free. Attach- thioglucoside donor.[19] In these two sequential glycosyl-
Eur. J. Org. Chem. 2011, 6698–6703
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E. C. Lourenço, M. R. Ventura
FULL PAPER
Table 1. Comparison of 13C NMR chemical shifts for the synthetic potassium salt of 1 with data from the natural product.[4]
13C, δ [ppm]
α(1Ǟ2)mannosyl moiety
C-2 C-3 C-4 C-5
α(1Ǟ2)glucosyl moiety
C-2 C-3 C-4 C-5
Glyceryl moiety
C-1 C-2 C-3
C-1
C-6
C-1
C-6
Natural
Synthetic
99.90 72.53 72.88 69.33 75.47 63.50 96.60 76.76 73.85 72.24 74.62 63.30 179.2 81.0
100.1 72.7 73.1 70.6 75.7 63.9 96.8 76.9 74.1 72.5 74.8 63.5 179.5 81.3
65.7
65.9
Scheme 4. Reagents and conditions: (a) Succinic anhydride, iPr2NEt, 4-dimethylaminopyridine (DMAP), CH2Cl2, r.t., 90%; (b) Tentagel
MB-NH2, diisopropylcarbodiimide (DIC), 1-hydroxybenzotriazole (HOBt), CH2Cl2, r.t.; (c) 5, TMSOTf, CH2Cl2, –20 °C; (d) 7, NIS,
TfOH, 4 Å molecular sieves, CH2Cl2, 0 °C; (e) MeONa, MeOH, r.t.; (f) Ac2O, pyridine, DMAP, r.t., quant.
ations no additional purification was needed and capping to compare with the product obtained through the solution-
with acetate made no difference to the overall efficiency, the phase method by using a bidirectional (non-linear) ap-
excess of reagents were simply filtered and washed off the proach, whereby the central glucose moiety was attached to
Tentagel-bound products.
the resin such that the donor and acceptor atoms were free
Cleavage from the resin was accomplished by using so- to react with their counterparts under adequate conditions.
dium methoxide in methanol, with simultaneous hydrolysis A partially deprotected product was obtained in 18% over-
of the acetate groups of the mannose moiety (Scheme 4). all yield by this method.
To better purify and analyze the product obtained from the
resin, the free hydroxy groups were acetylated to give per-
Experimental Section
acetate 17. After chromatographic purification, the spectro-
General: 1H NMR spectra were obtained with a Bruker Avance II+
scopic data obtained confirmed the success of the polymer-
400 MHz spectrometer at 400 MHz in CDCl3 or D2O with chemi-
cal shift values (δ) in ppm downfield from tetramethylsilane in the
bound glycosylations but an overall yield of 18%, based on
calculated immobilized 12, was disappointing.
case of CDCl3, and 13C NMR spectra were obtained at
100.61 MHz in CDCl3 or D2O. Assignments are supported by 2D
correlation NMR spectroscopy studies. Preparative chromato-
graphic separations were carried out by using medium-pressure col-
umn chromatography with Merck 60 H silica gel. Analytical TLC
was performed on aluminum-backed Merck 60 F254 silica gel plates.
Specific rotations ([α]2D0) were measured by using a Perkin–Elmer
241 automatic polarimeter. Reagents and solvents were purified
and dried according to ref.[20] All of the reactions were carried out
under an inert atmosphere (argon), except when the solvents were
not dried.
Conclusions
A solution-phase synthesis of the natural solute 1 has
been achieved with an overall yield of 15%. Differentiation
of the 2-OH (acceptor) group of the glucose moiety was
accomplished in just two steps through the dioxane bis-acet-
al, followed by selective protection of the more reactive 6-
OH group. Different activating groups were used to carry
out glycosylation reactions in the presence of the other
Phenyl 6-O-tert-Butyldiphenylsilyl-3,4-di-O-(2,3-dimethoxybutane-
groups. A similar solid-phase synthesis of 1 was carried out 2,3-diyl)-1-thio-β-D-glucopyranoside (4): TBDPSCl (1.28 mL,
6700
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Eur. J. Org. Chem. 2011, 6698–6703

Synthesis of a Naturally Compatible Solute
4.92 mmol) was added to a solution of 2[10] (0.634 g, 1.64 mmol) in
dry DMF (5 mL) at room temperature, followed by imidazole
(0.391 g, 5.74 mmol). After 12 h at room temperature, the reaction
was quenched with H2O (5 mL), extracted with CH2Cl2 (3ϫ5 mL),
and the combined organic phases were dried (MgSO4) and concen-
trated. Purification by flash column chromatography on silica gel
(30:70 EtOAc/hexane) afforded the product 4 as a white solid
4.29–4.18 (m, 3 H), 3.97 (dd, J = 5.4, 10.5 Hz, 1 H), 3.91–3.83 (m,
2 H), 3.80–3.71 (m, 4 H), 3.72 (s, 3 H), 3.28 (s, 3 H), 3.17 (s, 3 H),
2.17 (s, 3 H), 2.13 (s, 3 H), 2.04 (s, 3 H), 1.97 (s, 3 H), 1.27 (s, 6
H), 1.01 (s, 9 H), 1.00 (s, 9 H) ppm. 13C NMR (CDCl3): δ = 170.7,
170.3, 169.8, 169.6, 169.5, 135.8, 135.6, 135.5, 135.4, 133.7, 133.2,
132.7, 129.8, 129.7, 129.6, 129.5, 127.8, 127.7, 127.5, 127.4, 99.6,
99.5, 94.6 [C-1(α) mannose], 93.9 [C-1(α) glucose], 74.6, 70.9, 70.4,
69.6, 69.1, 68.5, 67.8, 66.3, 65.8, 64.4, 62.3, 61.7, 51.9, 47.9, 47.8,
26.7, 26.5, 20.9, 20.7, 20.69, 20.67 19.3, 19.1, 17.6, 17.6 ppm.
C62H82O20Si2 (1203.49): calcd. C 61.88, H 6.87; found C 61.90, H
(0.908 g, 89%). [α]2D0 = +34.1 (c = 0.17, CH2Cl2). M.p. 69.2–71.4 °C.
1
FTIR (film): ν = 3496 (O–H) cm–1. H NMR (CDCl ): δ = 7.67–
˜
3
7.72 (m, 4 H), 7.57–7.55 (m, 2 H), 7.42–7.31 (m, 6 H), 7.23–7.19
(m, 3 H), 4.57 [d, J = 9.3 Hz, 1 H, 1-H(β)], 3.99–3.90 (m, 2 H), 6.75.
3.82–3.73 (m, 2 H), 3.59–3.52 (m, 2 H), 3.31 (s, 3 H), 3.19 (s, 3 H),
Methyl (2R)-2-O-[2,3,4,6-Tetra-O-acetyl-α-
(1Ǟ2)-3,4-di-O-(2,3-dimethoxybutane-2,3-diyl)-1-O-α-
D
-mannopyranosyl-
-glucopyr-
1.33 (s, 3 H), 1.28 (s, 3 H), 1.06 (s, 9 H) ppm. 13C NMR (CDCl3):
δ = 135.8, 135.5, 133.7, 133.1, 132.8, 131.9, 129.6, 129.5, 129.0,
127.9, 127.66, 127.63, 99.7, 99.4, 88.5 [C-1(β)], 78.9, 73.7, 69.1,
64.9, 61.9, 48.2, 48.0, 26.9, 19.3, 17.7, 17.6 ppm. C34H44O7SSi
(624.86): calcd. C 65.35, H 7.10, S 5.13; found C 65.00, H 6.86, S
5.01.
D
anosyl]-2,3-dihydroxypropanoate (9): TBAF (1 m in THF; 0.22 mL,
0.22 mmol) was added to a solution of 8 (0.246 g, 0.20 mmol) in
THF (3 mL) at room temperature. The reaction mixture was stirred
for 3 h and then water was added. The mixture was extracted with
EtOAc, dried (MgSO4) and concentrated to give a yellow viscous
residue. Purification by column chromatography (EtOAc) afforded
product 9 as a viscous colorless foam (0.103 g, 71%). [α]2D0 = +155.2
Phenyl 2,3,4,6-Tetra-O-acetyl-α-
butyldiphenylsilyl-3,4-di-O-(2,3-dimethoxybutane-2,3-diyl)-1-thio-β-
-glucopyranoside (6): Acceptor 4 (0.254 g, 0.41 mmol) was added
D-mannopyranosyl-(1Ǟ2)-6-O-tert-
D
(c = 1.82, CH Cl ). FTIR (film): ν = 3463 (O–H), 1747, 1636
˜
2
2
(C=O) cm–1. 1H NMR (CDCl3): δ = 5.39–5.37 (m, 1 H), 5.35–5.28
(m, 2 H), 5.19 [d, J = 3.8 Hz, 1 H, C-1(α) glucose], 5.13 [s, 1 H, C-
1(α) mannose], 4.50–4.48 (m, 1 H), 4.39–4.36 (m, 1 H), 4.26–4.17
(m, 2 H), 4.10 (t, J = 10.0 Hz, 1 H), 3.95–3.81 (m, 5 H), 3.75–3.67
(m, 2 H), 3.73 (s, 3 H), 3.34 (s, 3 H), 3.24 (s, 3 H), 2.17 (s, 3 H),
2.13 (s, 3 H), 2.04 (s, 3 H), 1.98 (s, 3 H), 1.30 (s, 3 H), 1.26 (s, 3
H) ppm. 13C NMR (CDCl3): δ = 170.7, 170.1, 169.9, 169.74,
169.70, 99.8, 99.7, 94.9 [C-1(α) mannose], 94.6 [C-1(α) glucose],
75.4, 71.4, 70.0, 69.3, 69.2, 68.7, 67.4, 66.0, 65.8, 63.3, 62.1, 60.9,
52.2, 47.9, 47.8, 20.8, 20.7, 17.6, 17.5 ppm. HRMS: calcd. for
C30H46O20Na+ [M+ + Na] 749.2475; found 749.2451.
to a solution of trichloroacetamidate 5 (0.250 g, 0.51 mmol) in dry
CH2Cl2 (4 mL). The solution was cooled to –20 °C and TMSOTf
(92.30 μL, 0.51 mmol) was slowly added. When the reaction was
completed, a saturated aqueous solution of NaHCO3 (2 mL) was
added, followed by extractions with CH2Cl2. The combined organic
phases were dried (MgSO4) and concentrated. The residue was
purified by flash column chromatography on silica gel (20:80
EtOAc/hexane) to afford 6 as a white foam (0.237 g, 58%). [α]2D0
=
+54.15 (c = 1.30, CH Cl ). FTIR (film): ν = 1752 (C=O) cm–1. 1H
˜
2
2
NMR (CDCl3): δ = 7.73–7.70 (m, 4 H), 7.55–7.53 (m, 2 H), 7.43–
7.19 (m, 9 H), 5.51 [s, 1 H, 1-H(α) mannose], 5.35–5.33 (m, 3 H),
4.73–4.72 (m, 1 H), 4.65 [d, J = 8.7 Hz, 1 H, 1-H(β) glucose], 4.29
(dd, J = 3.6, 12.0 Hz, 1 H), 4.18–4.15 (m, 1 H), 3.95–3.88 (m, 2
H), 3.79–3.76 (m, 3 H), 3.32 (s, 3 H), 3.17 (s, 3 H), 2.20 (s, 3 H),
2.12 (s, 3 H), 2.01 (s, 3 H), 1.99 (s, 3 H), 1.26 (s, 3 H), 1.25 (s, 3
H), 1.06 (s, 9 H) ppm. 13C NMR (CDCl3): δ = 170.8, 169.9, 169.8,
169.6, 135.8, 135.5, 133.9, 133.6, 133.0, 131.9, 131.5, 129.6, 129.5,
129.0, 127.6, 127.5, 99.9, 99.3, 97.1 [C-1(α) mannose], 88.7 [C-1(β)
glucose], 78.5, 73.4, 72.8, 69.2, 69.1, 68.6, 66.0, 65.1, 62.0, 48.0,
47.9, 26.8, 20.9, 20.71, 20.70, 20.6, 19.3, 17.6, 17.3 ppm.
C48H62O16SSi (955.15): calcd. C 60.36, H 6.54; found C 60.50, H
6.40.
Methyl (2R)-2-O-[2,3,4,6-Tetra-O-acetyl-α-
D-mannopyranosyl-
(1Ǟ2)-1-O-α- -glucopyranosyl]-2,3-dihydroxypropanoate (10): A
D
mixture of TFA/H2O (5:1) (0.8 mL) was added to a solution of 9
(0.093 g, 0.13 mmol) in CH2Cl2 (1 mL) at room temperature. The
reaction mixture was stirred until all of the starting material had
been consumed and then it was concentrated to give a viscous resi-
due. Purification by column chromatography (1:4, MeOH/CH2Cl2)
afforded product 10 as a viscous colorless foam (0.055 g, 70%).
[α]2D0 = +72.17 (c = 0.46, EtOH). FTIR (film): ν = 3460 (O–H),
˜
1748 (C=O) cm–1. 1H NMR (CDCl3): δ = 5.43–5.42 (m, 1 H), 5.38
(dd, J = 3.2, 10.0 Hz, 1 H), 5.26 (t, J = 10.0 Hz, 1 H), 5.20 [d, J =
3.6 Hz, 1 H, 1-H(α) glucose], 5.17 [d, J = 1.4 Hz, 1 H, 1-H(α) man-
nose], 4.48 (t, J = 4.2 Hz, 1 H), 4.46–4.39 (m, 2 H), 4.15–4.12 (m,
1 H), 3.88–3.68 (m, 7 H), 3.69 (s, 3 H), 3.43 (t, J = 9.4 Hz, 1 H),
2.18 (s, 3 H), 2.09 (s, 3 H), 2.06 (s, 3 H), 1.98 (s, 3 H) ppm. 13C
NMR (CDCl3):δ = 173.7, 173.1, 172.8, 172.7, 172.0, 95.4 [C-1(α)
mannose], 94.8 [C-1(α) glucose], 75.9, 75.4, 72.2, 70.9, 69.7, 69.3,
68.4, 65.5, 62.3, 61.8, 60.3, 52.7, 20.1, 20.0 ppm. HRMS: calcd. for
C24H36O18Na+ [M+ + Na] 635.1794; found 635.2451.
Methyl 3-O-tert-Butyldiphenylsilyl-(2R)-2-O-[2,3,4,6-tetra-O-
acetyl-α-
D
-mannopyranosyl-(1Ǟ2)-6-O-tert-butyldiphenylsilyl-3,4-
-glucopyranosyl]-2,3-di-
di-O-(2,3-dimethoxybutane-2,3-diyl)-1-O-α-
D
hydroxypropanoate (8): A suspension of thioglycoside 6 (0.387 g,
0.40 mmol), glycerate 7 (0.143 g, 0.40 mmol) and 4 Å molecular si-
eves in CH2Cl2 (3 mL) was stirred at room temperature for 1 h.
The solution was cooled to 0 °C and N-iodosuccinimide (0.099 g,
0.44 mmol) and TMSOTf (36 μL, 0.20 mmol) were added. After all
of the starting material had been consumed, a 10% aqueous solu-
tion of Na2S2O3 (2 mL) and a saturated aqueous solution of
NaHCO3 (1 mL) were added and the mixture was extracted with
CH2Cl2; the combined organic phases were dried (MgSO4), filtered
and the solvent was removed in vacuo. The crude product was puri-
fied by column chromatography (30:70 EtOAc/hexane) to afford
Methyl (2R)-2-O-[α-D-Mannopyranosyl-(1Ǟ2)-1-O-α-D-glucopyr-
anosyl]-2,3-dihydroxypropanoate (11): A 1 n solution of NaOMe
(34 μL, 0.03 mmol) in MeOH was added to a stirred solution of 10
(0.036 g, 0.06 mmol) in MeOH (1 mL) at 0 °C. After 1 h, previously
activated Dowex-H+ resin was added until the pH of the reaction
mixture reached 7. After filtration with MeOH and water, the sol-
glycoside 8 as a colorless viscous foam (0.286 g, 60%). [α]2D0 = +83.3
1
(c = 1.00, CH Cl ). FTIR (film): ν = 1641, 1753 (C=O) cm–1. H
vent was removed in vacuo to yield 11 as a viscous colorless foam
˜
2
2
1
NMR (CDCl3): δ = 7.68–7.62 (m, 8 H), 7.42–7.30 (m, 12 H), 5.46 (0.026 g, quantitative). [α]2D0 = +135.61 (c = 2.03, H2O). H NMR
(dd, J = 3.4, 9.9 Hz, 1 H), 5.40–5.39 (m), 5.29 (t, J = 10.0 Hz, 1
H), 5.20 [d, J = 1.4 Hz, 1 H, C-1(α) mannose], 5.13 [d, J = 3.8 Hz,
1 H, C-1(α) glucose], 4.52–4.48 (m, 1 H), 4.37 (dd, J = 5.2, 3.8 Hz),
(D2O): δ = 5.26 [d, J = 3.4 Hz, 1 H, 1-H(α) glucose], 5.09 [d, J =
1.3 Hz, 1 H, 1-H(α) mannose], 4.52 (t, J = 3.8 Hz, 1 H), 4.03 (q, J
= 1.6 Hz, 1 H), 3.91–3.90 (m, 2 H), 3.87–3.63 (m, 10 H), 3.77 (s, 3
Eur. J. Org. Chem. 2011, 6698–6703
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E. C. Lourenço, M. R. Ventura
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H), 3.45 (t, J = 9.0 Hz, 1 H) ppm. 13C NMR (D2O): δ = 174.6,
Synthesis of Resin-Bound 15: Resin-bound donor 14 (0.350 g,
99.6 [C-1(α) mannose], 96.8 [C-1(α) glucose], 77.9, 76.3, 75.2, 74.7,
0.10 mmol) was swelled in CH2Cl2 (2 mL) with 4 Å molecular si-
73.5, 72.8, 72.4, 71.8, 69.1, 64.9, 63.4, 62.9, 55.3 ppm. HRMS: eves. Acceptor 7 (0.280 g, 0.78 mmol) was dissolved in CH2Cl2,
calcd. for C16H28O14Na+ [M+ + Na] 467.1371; found 467.1372.
transferred by cannula to the reaction flask and the reaction mix-
ture was shaken at room temperature for 1 h. The reaction was
cooled to –20 °C, NIS (0.037 g, 0.12 mmol) and TMSOTf
(0.013 mL, 0.07 mmol) was added, and the reaction shaken at
–20 °C for min. The solution phase was filtered out and the resin
was washed three times with DMF, MeOH, Et2O and CH2Cl2. The
resin was dried in vacuo over phosphorus pentoxide. HR-MAS 1H
NMR spectroscopy indicated successful coupling with a theoretical
resin loading of 0.186 mmolgϪ1, as calculated from resin weight
gain.
Potassium (2R)-2-O-α- -Mannopyranosyl-(1Ǟ2)-α- -glucopyran-
D
D
osyl-2,3-dihydroxypropanoate (1): A solution of 2 m KOH (320 μL)
was added to a stirred solution of ester 11 (0.026 g, 0.06 mmol) in
H2O (1 mL). After all of the starting material had been consumed,
the pH was adjusted to 7 with 10% HCl and the solvent was evapo-
rated to afford 1 as a viscous colorless foam (0.027 g, quantitative).
The NMR spectroscopic data were identical to those of a natural
sample. [α]2D0 = +67.76 (c = 0.85, H O). FTIR (film): ν = 3315,
˜
2
1655 cm–1. H NMR (D2O): δ = 5.12 [d, J = 3.4 Hz, 1 H, 1-H(α)
1
glucose], 5.06 [d, J = 1.12 Hz, 1 H, 1-H(α) mannose], 4.13 (dd, J
= 3.2, 7.0 Hz, 1 H), 4.04–4.02 (m, 1 H), 3.85–3.58 (m, 12 H), 3.39
(t, J = 9.3 Hz, 1 H) ppm. 13C NMR (D2O): δ = 176.5, 97.1 [C-1(α)
mannose], 93.8 [C-1(α) glucose], 78.3, 73.9, 72.7, 71.8, 71.1, 70.1,
69.7, 69.5, 66.6, 62.9, 60.9, 60.5 ppm. HRMS: calcd. for
C15H26O14K+ [M+ + H] 469.09541; found 469.0954.
Cleavage from the ResinϪMethyl 3-O-tert-Butyldiphenylsilyl-(2R)-
2-O-[α-
2,3-diyl)-1-O-α-
D
-mannopyranosyl-(1Ǟ2)-3,4-di-O-(2,3-dimethoxybutane-
-glucopyranosyl]-2,3-dihydroxypropanoate (16):
D
Resin 15 (0.430 g) was swelled in CH2Cl2 for 30 min and then sus-
pended in MeOH. A 1 n solution of NaOMe (0.6 equiv.) was added
and the reaction was stirred for 5 h at room temperature. The resin
was filtered and washed six times with CH2Cl2 and MeOH. The
solution phase was treated with previously activated Dowex-H+,
filtered with MeOH and concentrated under reduced pressure. Pu-
rification was accomplished by preparative TLC (1:9, MeOH/
CH2Cl2) to give the desired product 16 (2 mg, 18%). This product
was very polar and for further characterization it was acetylated to
6-O-[Phenyl-3,4-di-O-(2,3-dimethoxybutane-2,3-diyl)-1-thio-β-D-glu-
copyranosyl] Succinate (12): Succinic anhydride (0.041 g,
0.41 mmol), followed by diisopropylethylamine (171 μL,
0.98 mmol) and a catalytic amount of DMAP were added to a
solution of 2 (0.160 g, 0.41 mmol) in CH2Cl2 (2 mL) at 0 °C. The
reaction mixture was stirred as the temperature was allowed to rise
to room temperature. After 3 h, the reaction was quenched and
washed with H2O. The aqueous phase was extracted with AcOEt
(3ϫ5 mL) and the combined organic phases were dried (MgSO4)
and concentrated. Purification by column chromatography (80:20
EtOAc/hexane) afforded product 12 as a viscous colorless oil
1
afford 17. Compound 16: H NMR (CDCl3): δ = 7.68–7.64 (m, 4
H), 7.42–7.39 (m, 6 H), 5.16–5.12 [m, 2 H, 1-H(α) mannose and
glucose], 4.41–4.39 (m, 1 H), 4.15–3.58 (m, 14 H), 3.75 (s, 3 H),
3.21 (s, 3 H), 3.17 (s, 3 H), 1.26 (s, 3 H), 1.22 (s, 3 H), 1.02 (s, 9
H) ppm.
(0.180 g, 90%). [α]2D0 = +110.5 (c = 0.19, CH Cl ). FTIR (film): ν
˜
Methyl 3-O-tert-Butyldiphenylsilyl-(2R)-2-O-[2,3,4,6-tetra-O-acet-
2
2
= 3488 (O–H), 1736 (C=O) cm–1. 1H NMR (CDCl3): δ = 7.56–7.53
(m, 2 H), 7.33–7.29 (m, 3 H), 4.52 [d, J = 9.3 Hz, 1 H, 1-H(β)],
4.48 (dd, J = 1.9, 11.8 Hz, 1 H), 4.21 (dd, J = 5.4, 11.9 Hz, 1 H),
3.77–3.71 (m, 2 H), 3.59 (t, J = 9.8 Hz, 1 H), 3.48 (t, J = 9.32 Hz,
1 H), 3.30 (s, 3 H), 3.21 (s, 3 H), 2.72–2.63 (m, 4 H), 1.32 (s, 3 H),
1.28 (s, 3 H) ppm. 13C NMR (CDCl3): δ = 176.6, 171.8, 133.5,
130.9, 128.9, 128.5, 99.8, 99.7, 88.0 [C-1(β)], 75.7, 73.3, 69.0, 65.6,
62.8, 48.05, 48.02, 28.7, 28.6, 17.6, 17.5 ppm. HRMS: calcd. for
C22H30O10SNa+ [M+ + Na] 509.1452; found 509.1454.
yl-α-
D
-mannopyranosyl-(1Ǟ2)-6-O-acetyl-3,4-di-O-(2,3-dimeth-
-glucopyranosyl]-2,3-dihydroxypropano-
oxybutane-2,3-diyl)-1-O-α-
D
ate (17): Acetic anhydride (5 μL, 5.3ϫ10–2 mmol) and a catalytic
quantity of DMAP were added to a solution of 16 (2 mg,
2.5ϫ10–3 mmol) in pyridine (0.5 mL) at room temperature. After
12 h at room temperature, the reaction was quenched with H2O
(1 mL), extracted with EtOAc (3ϫ2 mL) and the combined or-
ganic phases were dried (MgSO4) and concentrated. Purification by
flash column chromatography on silica gel (30:70 EtOAc/hexane)
afforded product 17 as viscous colorless oil (3 mg, 100%). [α]2D0
=
Synthesis of Resin-Bound 13: Thioglucoside 12 (0.130 g, 0.27 mmol)
was dissolved in CH2Cl2 (2 mL) and cooled to 0 °C. HOBt (0.036 g,
0.27 mmol) was added, followed by DIC (0.041 mL, 0.37 mmol),
and the solution was stirred at 0 °C for 10 min. The reaction mix-
ture was transferred by cannula to Tentagel MB-NH2 resin
(0.601 g, 0.24 mmol) swelled in CH2Cl2 (2 mL), and the reaction
was shaken at room temperature for 6 h. The resin was filtered, and
rinsed three times with DMF, MeOH and CH2Cl2, and dried in
vacuo over phosphorus pentoxide. HR-MAS 1H NMR spec-
troscopy indicated successful coupling, with a theoretical resin
loading of 0.32 mmolgϪ1, as calculated from the resin weight gain.
+101.5 (c = 1.99, CH Cl ). FTIR (film): ν = 1747 (C=O) cm–1. 1H
˜
2
2
NMR (CDCl3): δ = 7.70–7.66 (m, 4 H), 7.43–7.42 (m, 6 H), 5.45
(dd, J = 3.4, 9.9 Hz, 1 H), 5.39–5.37 (m, 1 H), 5.30 (t, J = 10.0 Hz,
1 H), 5.18 [d, J = 3.7 Hz, 1 H, C-1(α) glucose], 5.16 [d, J = 0.96 Hz,
1 H, C-1(α) mannose], 4.50–4.46 (m, 1 H), 4.44 (dd, J = 6.0, 3.6 Hz,
1 H), 4.23–4.10 (m, 5 H), 4.07–4.00 (m, 2 H), 3.93–3.88 (m, 2 H),
3.71–3.64 (m, 1 H), 3.68 (s, 3 H), 3.29 (s, 3 H), 3.19 (s, 3 H), 2.17
(s, 3 H), 2.13 (s, 3 H), 2.04 (s, 3 H), 2.03 (s, 3 H), 1.97 (s, 3 H),
1.27 (s, 3 H), 1.26 (s, 3 H), 1.04 (s, 9 H) ppm. 13C NMR (CDCl3):
δ = 170.7, 170.6, 170.0, 169.8, 169.7, 169.6, 135.6, 135.5, 133.1,
132.8, 129.8, 129.7, 127.9, 127.86, 127.84, 99.8, 99.7, 94.7, 94.2,
75.0, 70.8, 69.4, 69.1, 68.5, 67.5, 67.4, 66.2, 66.0, 64.3, 62.1, 62.0,
60.3, 52.0, 47.8, 47.7, 26.6, 20.9, 20.76, 20.71, 20.70, 20.6, 19.1,
17.6, 17.5, 14.1 ppm.
Synthesis of Resin-Bound 14: Resin-bound thioglucoside acceptor
13 (0.711 g, 0.23 mmol) was swelled in CH2Cl2 (2 mL) with 4 Å
molecular sieves. Donor 5 (0.340 g, 0.69 mmol) was dissolved in
CH2Cl2, transferred by cannula to the reaction flask and shaken for
1 h. The reaction was cooled to –20 °C and TMSOTf (0.049 mL,
0.27 mmol) was added. The solution phase was filtered after 15 min
and the resin washed three times with DMF, MeOH, Et2O and
CH2Cl2. The resin was dried in vacuo over phosphorus pentoxide.
Acknowledgments
We wish to acknowledge the analysis department at the ITQB for
providing elemental analyses. We acknowledge the generous finan-
cial support provided by Fundação para a Ciência e Tecnologia
(FCT) PTDC/BIO/70806/2006. E. L. acknowledges the FCT for a
1
HR-MAS H NMR spectroscopy indicated successful coupling,
with a theoretical resin loading of 0.28 mmolgϪ1, as calculated
from the resin weight gain.
6702
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© 2011 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2011, 6698–6703

Synthesis of a Naturally Compatible Solute
[9]
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work of the National Program for Scientific Re-equipment, con-
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(FEDER) and FCT.
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Received: June 28, 2011
Published Online: September 19, 2011
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www.eurjoc.org
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