Synthesis, surface tension properties and antibacterial activities of amphiphilic d-galactopyranose derivatives

Article abstract of DOI:10.1016/j.ejmech.2012.12.032

Several amphiphilic d-galactopyranose derivatives were synthesized in which the glycosidic moiety was separated from the hydrophobic alkyl chain (along 8 or 12 carbon atoms) by a spacer arm (butyl, butynyl or benzyl) in order to increase their surfactant properties and to obtain new antibacterial compounds. The surface tensions of the products were analyzed by Critical Micelle Concentration (CMC) and γ_CMC measurements and the antimicrobial activities were assayed against 10 bacterial species by Minimum Inhibitory Concentration (MIC) determination in liquid broth. The introduction of an aliphatic spacer arm increased the amphiphilic properties of the compounds and the CMC values were 40-500 times lower than their analogs without spacer arm. In the same manner, the spacer arms significantly increased the antibacterial power of the compounds. The products 4d and 4e exhibiting a C12 alkyl chain and an aliphatic spacer arm (butyl and butynyl) were the best surfactants (CMC = 0.023 and 0.032 mmol/L, respectively) and presented also the best antibacterial activities (MIC = 15.62 and 3.91 μg/mL for Micrococcus luteus, respectively). But the antibacterial activity of the newly synthesized products seemed to depend more on the cell wall composition of the bacteria than only on the amphiphilic character of the compounds.

Full text of DOI:10.1016/j.ejmech.2012.12.032

European Journal of Medicinal Chemistry 62 (2013) 177e186
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis, surface tension properties and antibacterial activities
of amphiphilic
D-galactopyranose derivatives
,
b
,
c
a
Ludovic Chaveriat a , Isabelle Gosselin
, Cécile Machut , Patrick Martin
*
**
a Centre Technologique des Agroressources (CTA), Université d’Artois, IUT de Béthune, 1230 rue de l’Université, BP 819, 62408 Béthune Cedex,
France
b Unité de Génie Enzymatique et Cellulaire (GEC), UMR CNRS 6022, Université de Picardie Jules Verne, UFR des Sciences, 33 rue Saint Leu,
80039 Amiens cedex, France
c Unité de Catalyse et de Chimie du Solide (UCCS), UMR CNRS 8181, Université d’Artois, IUT de Béthune, 1230 rue de l’Université, BP 819,
62408 Béthune Cedex, France
a r t i c l e i n f o
a b s t r a c t
Article history:
Several amphiphilic D-galactopyranose derivatives were synthesized in which the glycosidic moiety was
Received 12 July 2012
Received in revised form
11 December 2012
Accepted 16 December 2012
Available online 25 December 2012
separated from the hydrophobic alkyl chain (along 8 or 12 carbon atoms) by a spacer arm (butyl, butynyl or
benzyl) in order to increase their surfactant properties and to obtain new antibacterial compounds. The
surface tensions of the products were analyzed by Critical Micelle Concentration (CMC) and gCMC measure-
ments and the antimicrobial activities were assayed against 10 bacterial species by Minimum Inhibitory
Concentration (MIC) determination in liquid broth. The introduction of an aliphatic spacer arm increased the
amphiphilic properties of the compounds and the CMC values were 40e500 times lower than their analogs
without spacer arm. In the same manner, the spacer arms significantly increased the antibacterial power of
the compounds. The products 4d and 4e exhibiting a C12 alkyl chain and an aliphatic spacer arm (butyl
and butynyl) were the best surfactants (CMC ¼ 0.023 and 0.032 mmol/L, respectively) and presented also
Keywords:
D
-galactopyranoside
Alkyl chain
Spacer arm
Surfactant
the best antibacterial activities (MIC ¼ 15.62 and 3.91
mg/mL for Micrococcus luteus, respectively). But the
Antimicrobial
antibacterial activity of the newly synthesized products seemed to depend more on the cell wall
composition of the bacteria than only on the amphiphilic character of the compounds.
Ó 2012 Elsevier Masson SAS. All rights reserved.
1. Introduction
membrane integrity and cause the cellular lysis [13,14]. But
generally, the carbohydrate-derived surfactants synthesized for
Carbohydrate-derived surfactants are non-ionic amphiphilic
compounds in which the carbohydrate moiety is linked to a long
alkyl chain. In recent years, this class of detergents has been
extensively studied [1e7] due to several interesting properties in
addition to their emulsifying properties such as non-toxicity or
biodegradability, and because carbohydrates are attractive starting
materials for chemical elaboration due to their status as renewable
resources [8,9]. An overall review on the synthesis and the prop-
erties of carbohydrate-based amphiphilic molecules has been
published recently [10]. Carbohydrate-derived surfactants are also
known for their antimicrobial activity due to their capacity to
interact with biological membranes [11,12]. Indeed surfactants can
solubilize the lipids constituting the membrane bilayer, impair the
antimicrobial purposes were derivatives of amino ether [6,15e19],
thio ether [20,21], aminoglycosides [22e25], or fatty acid ester
[26e29]. Only few works deal with the antimicrobial activity of
carbohydrate ether derivatives [30e34].
The aim of this study was to deepen the knowledge on the
antimicrobial properties of carbohydrate ether derivatives in the
actual context of bacterial resistance to antibiotics and the
increasing need to discover new antibacterial molecules [35,36]. In
that purpose, we synthesized carbohydrate ethers deriving from
caprylic and lauric acids, C8 and C12 fatty acid respectively, well
known for their strong antibacterial properties [33,37e43]. Con-
cerning the glycosidic part, previous studies on carbohydrate esters
showed that the configuration of hydroxyl groups in the carbohy-
drate moiety influenced the antibacterial activities, and the galac-
tose derivatives were more bactericide than the glucose, fructose or
sucrose ones [44]. Then, we synthesized 6-O-alkyl-galactose deriv-
atives with a C8 or a C12 alkyl chain and tested the tension surface
characteristics as well as the antimicrobial activities of these
compounds. In previous works, the amphiphilic and liquid crystal
properties of these molecules have been shown to strongly decrease
Abbreviations: CMC, critical micelle concentration; MIC, minimum inhibitory
concentration.
* Corresponding author. Tel.: þ33 3 21 63 23 25; fax: þ33 3 21 63 02 94.
** Corresponding author. Tel.: þ33 3 22 82 74 73; fax: þ33 3 22 82 75 95.
E-mail
addresses:
Ludovic.Chaveriat@univ-artois.fr
(L.
Chaveriat),
Isabelle.Gosselin@u-picardie.fr (I. Gosselin).
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
http://dx.doi.org/10.1016/j.ejmech.2012.12.032

178
L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
when the alkyl chain contained more than eight carbon atoms
[45,46]. This phenomenon, which decreased the hydrophilic char-
acter, resulted from the intramolecular hydrogen bonding between
the cis-hydroxyl groups located on C-3 and C-4 [47]. It was proposed
that the C-6 hydrophobic alkyl chain with a certain length, sterically
crowded the C-3 and C-4 hydroxyl groups, thus preventing them to
be engaged in intermolecular hydrogen bonding with water mole-
cules [48]. In order to circumvent this problem, we prepared gluci-
configuration [45,46,49] (see representative NMR spectra in
Supplementary Data).
After synthesis, the deprotected compounds 4aef were evalu-
ated for their surfactant properties. The Critical Micelle Concen-
trations (CMC) was determined by surface tension measurement
(g) using the Wilhelmy method and the measured values were
plotted against the molar concentrations. The surface tension
characteristics of the galactose derivatives were reported in Table 1
and the corresponding graphs in Figs. 1 and 2. The results were
compared to the corresponding analogs without spacer arm 6a and
doamphiphile derivatives from D-galactose in which the glucidic
moiety and the hydrophobic alkyl chain were separated by a spacer
arm with various length and rigidity (butyl, butynyl, benzyl). After-
ward these molecules and their derivatives without spacer armwere
all tested for their surface-active properties and their antimicrobial
activities on a large range of bacterial strains.
6b (6-O-alkyl-D-galactopyranoses), which were already described
in the literature [48] but were synthesized again here (Scheme 3) to
compare the influence of the spacer arm on the tension surface
properties and hereafter on the antimicrobial efficiency.
The compounds 4a, 4b, 4d and 4e (derivatives with non
aromatic spacer arms) exhibited a large decrease in surface tension
from 72 to 30e42 mN/m approximatively when the concentration
raised (Figs. 1 and 2) which is a classical surfactant behavior. In
addition, all of them presented a characteristic phenomenon of
micellization. For the compounds 4c and 4f with an aromatic spacer
arm, the profiles were different. The surface tension for the
compound 4c decreased from 72 to 54 mN/m (Fig.1), but no CMC or
gcmc could be determined because the solution became turbid over
0.6 mmol/L. This compound seemed to have a hydrotropic behavior
which prevented the molecule organization in micelle and induced
2. Results and discussion
Firstly, the synthesis of the hydrophobic alkyl chain including
spacer arm began with the monoalkylation of commercial diol with
n-dodecyl bromide or n-octyl bromide in DMF (Scheme 1). For this
step, sodium hydride was used when the starting material was
butan-1,4-diol or butyn-1,4-diol and sodium hydroxide to prevent
degradation of hydroquinone. The corresponding alkylated alcohol
1aef was obtained with a yield of 60e90%. Then, the activation of
hydroxyl group was performed using p-toluenesulfonyl chloride
and dimethylaminopyridine in dichloromethane. Tosylated deriv-
atives 2aef were isolated with a 73e92% yield.
no break in the
g slope (Fig. 1). For the compound 4f, the same
decrease in surface tension as 4c was observed; however, a CMC
could be measured but with a 100 times worse value than the
compounds with aliphatic spacer arm, i.e. 3.1 mmol/L instead of
0.023 and 0.032 mmol/L for 4d and 4e, respectively.
The 6-O-alkyl-
1,2:3,4-di-O-isopropylidene-
D
-galactopyranoses wereprepared byalkylation of
-galactopyranose using appropriate
D
tosylated alkyl chain and potassium hydroxide for derivatives with
spacer arm (Scheme 2), or suitable bromoalkanes and sodium
hydride for derivatives without spacer arm (Scheme 3).
The comparison between the CMC values obtained for the
galactose derivatives with and without non aromatic spacer arm
(Table 1) showed that the distance between the glucidic moiety and
the hydrophobic alkyl chain greatly improved the surfactant prop-
erties of the molecules. For the C8 derivatives, the CMC decreased
from 12 mmol/L for 6a to 0.3 and 0.06 mmol/L for butyl 4a and
butynyl 4b spacer arm, respectively. And for the C12 derivatives, no
CMC could be measured for the product 6b without spacer arm, in
accordance to the literature [48], whereas the values were 0.023 and
0.032 mmol/L for butyl 4d and butynyl 4e spacer arm, respectively.
The final condensation of the tosylated derivatives with 1,2:3,4-
di-O-isopropylidene-
presence of potassium hydroxide produced the 3aef intermediate
6-O-alkyl-1,2:3,4-di-O-isopropylidene- -galactopyranoses. These
D-galactopyranose in tolueneeMe2SO in the
D
compounds were subsequently deprotected with CF3COOH to
obtain 4aef with a 75e90% yield. The experimental conditions for
the deacetalization reaction allowed the maintenance of the
anomeric integrity to provide deprotected compounds in the
a-
a
b
HO E OH
HO E OCnH2n+1
TsO E OCnH2n+1
60-90%
73-92%
1a-f
2a-f
n
E
Product
1a, 2a
8
butyl
(CH2)4
8
butynyl 1b, 2b
benzyl 1c, 2c
H2C
C
C
CH2
spacer arm E =
8
12
12
12
butyl
1d, 2d
butynyl 1e, 2e
benzyl 1f, 2f
Scheme 1. Synthesis of alkyl chains including spacer arm. Reagents and conditions: (a) BrCnH2nþ1, NaH or NaOH, DMF, RT; (b) TsCl, DMAP, CH2Cl2, 0 ꢀC.

L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
179
OH
OH
(CH2)4
O
O
OCnH2n+1
e
O
(CH2)4
O
O
OCnH2n+1
OH
O
HO
HO
O
O
b
Me2C
a
Me2C
O
HO
O
OH
O
OH
O
C
OH
O
O
OH
C
D-galactose
Me2
Me2
3a, 3d
4a, 4d
c
n= 8 or 12
d
OCnH2n+1
O
O
O
CH2
C
C CH2
O
OCnH2n+1
O
O
Me2C
Me2C
O
O
O
O
O
O
C
C
Me2
Me2
3c, 3f
3b, 3e
e
e
OCnH2n+1
CH2
C
C CH2
O
OCnH2n+1
HO
HO
O
HO
HO
O
O
OH
OH
OH
OH
4b, 4e
4c, 4f
Scheme 2. Synthesis of 6-O-alkyl-D-galactopyranose derivatives with spacer arm: Reagents and conditions: (a) acetone, H2SO4, 54%; (b) TsO(CH2)4OCnH2nþ1, KOH, DMF, RT, 85e91%;
(c) TsOCH2C^CCH2OCnH2nþ1, KOH, DMF, 74e81%; (d) TsOPhOCnH2nþ1, KOH, DMF, 62e66%; (e) TFAeH20, 0 ꢀC, 75e90%.
The nature of the spacer arm affected the surface tension prop-
erties in relation to the length of the alkyl chain of the galactose
derivatives (Table 1). Therefore, for the galactose derivatives with
butyl spacer arm, the C8 derivative showed a CMC value 10 times
higher than the C12 derivatives (CMC ¼ 0.3 and 0.023 mmol/L for 4a
and 4d, respectively) and 2 times higher with the butynyl spacer
arm (CMC ¼ 0.06 and 0.032 mmol/L for 4b and 4e, respectively).
However when the alkyl chain was longer (4d, 4e), the nature of the
spacer arm (alkyne or alkane) had less influence and resulted in
equivalent CMC values (0.023 and 0.032 mmol/L, respectively). The
lengthening of the hydrocarbon chain adjusted the hydrophilice
lipophilic balance in favor of a more important emulsifying power.
For molecules with a C8 alkyl chain, the introduction of a rigid
spacer arm was effective to prevent the folding of the alkyl chain
only when the spacer was aliphatic. The compounds 4a and 4b
showed very interesting surface tension properties with
a CMC ¼ 0.3 and 0.06 mmol/L, respectively, instead of 12 mmol/L
for C8 derivative without spacer arm 6a.
chain increased and no significant difference was observed
between the CMC of the compounds 4d and 4e.
The aromatic spacer arm didn’t seem appropriate since the
micellization didn’t appear clearly in the surface tension graphs (4c
in Fig. 1 and 4f in Fig. 2). The benzene ring made the final sugar
completely hydrophobic and was probably too rigid to form a stable
supramolecular structure.
The surface tension properties of all the synthesized derivatives,
except those with an aromatic spacer arm, were comparable to
the most effective neutral surfactant, Tween 20, exhibiting
a CMC ¼ 0.05 mmol/L [50].
To complete this study, the antimicrobial activity of all the 6-O-
alkyl-galactose derivatives (C8 or C12 alkyl chain, with or without
spacer arm, protected or deprotected) was determined by liquid
broth serial dilutions. The Minimum Inhibitory Concentration
(MIC) was assayed on 10 bacterial species potentially pathogens for
humans, animals or plants: 6 Gram negative bacteria (Citrobacter
freundii ATCC 6750, Enterobacter cloacae ATCC 13047, Erwinia
chrysanthemi EC3937, Escherichia coli ATCC 10536, Pseudomonas
aeruginosa CIP A22 and Sinorhizobium meliloti Rm1021) and 4 Gram
positive bacteria (Bacillus stearothermophilus ATTC 12980, Micro-
coccus luteus CIP 53.45, Mycobacterium smegmatis CIP 73.26 and
Staphylococcus aureus subsp aureus CIP 53.154). Chloramphenicol
When the alkyl chain was relatively short (8 carbons), the
insertion of a spacer arm a little more rigid supported the
deployment of the alkyl chain and the supramolecular assembly of
the compounds in aqueous solution. However the nature of the
spacer arm became less important when the length of the alkyl
OH
O
OCnH2n+1
OCnH2n+1
HO
O
O
a
O
O
Me2C
b
Me2C
O
O
HO
O
O
OH
O
O
OH
C
C
Me2
Me2
5a n=8
5b n=12
6a n=8
6b n=12
Scheme 3. Synthesis of 6-O-alkyl-D-galactopyranose derivatives without spacer arm: Reagents and conditions: (a) BrCnH2nþ1, NaH, DMF, 69e81%, (b) TFAeH20, 0 ꢀC, 95%.

180
L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
Table 1
positive strains, whereas the Gram negative bacteria seemed to be
more resistant to these products, with the only exception of
S. meliloti with 4e (MIC ¼ 31.25 g/mL). Among Gram positive
strains, the lowest MIC values were obtained on M. luteus and
g/mL).
On another hand, the C8 derivatives (compounds 3aec, 4aec)
showed poor antibacterial activities and no MIC values under
62.5 mg/mL could be measured whatever was the strain tested.
Surface tension characteristics of the 6-O-alkyl-D-galactopyranose derivatives.
m
Compound
CMC (mmol/L)
gCMC (mN/m)
OC8H17
HO
HO
B. stearothermophilus (MIC ¼ 3.91e31.25
m
O
6a
6b
12
28
OH
OH
The products without spacer arm 5ae6b gave no MIC, showing
the importance to move the alkyl chain away from the glycosidic
part to present antibacterial properties.
OC12H25
HO
HO
O
ea
ea
The newly synthesized 6-O-alkyl-galactose derivatives exhibi-
ted tension surface properties related to the antibacterial activities
and the best surfactants 4d and 4e were also the best antimicro-
bials. Nevertheless, the amphiphilic character is not enough to
explain the bactericidal power and the compounds exhibited
a strain-dependant action in which the Gram negative species were
more resistant than the Gram positive bacteria. This was already
observed with fatty acid derivatives [29,37,51] and this resistance
was attributed to the presence of the cell wall lipopolysaccharide
(LPS) and of the outer membrane, which are two structures both
absent in Gram positive bacteria and could prevent the active
compounds to reach the cytoplasmic membrane of the Gram
negative species [37,52].
The only exception in the resistance to the products by the Gram
negative species is the case of S. meliloti with 4e, which was the best
antibacterial compound. This special feature could be explained by
the unusual LPS structure of S. meliloti which is composed of an
atypical very long-chain fatty acid with up to 30 carbon atoms [53]
and could interact differently with the alkyl chain of the
compounds.
Among the Gram positive bacteria, some species were more
sensitive to the 6-O-alkyl-galactose derivatives like M. luteus or
B. stearothermophilus, whereas others were more resistant like
M. smegmatis or S. aureus. This implies that the specific strain
composition of the cell wall or the cytoplasmic membrane is
important to explain the bactericidal effects of the products [54].
For example, M. luteus is known to possess mannosyl residus in its
membrane phospholipids [55] and M. smegmatis a unique arabi-
nogalactan feature [53]. Each strain presents a specific cell wall
composition and a deeper study on the molecular interactions
between the products and the bacterial components is necessary to
understand the bactericidal mechanisms.
OH
OH
O(CH2)4OC8H17
O
HO
HO
4a
4b
4c
0.3
0.06
ea
30
30
ea
OH
OH
OCH2C CCH2OC8H17
O
HO
HO
OH
OH
OC8H17
O
HO
HO
O
OH
OH
O(CH2)4OC12H25
O
HO
HO
4d
4e
4f
0.023
0.032
3.1
48
35
54
OH
OH
OCH2C CCH2OC12H25
O
HO
HO
OH
OH
OC12H25
O
HO
O
HO
OH
OH
a
No CMC or gCMC were recorded during surface tension experiment.
3. Conclusion
The introduction of a spacer arm between the galactose moiety
and the alkyl chain prevented the invasion of the hydrophobic part
in the region of the OH groups. Hence the intermolecular
hydrogen bonds with water molecules were allowed and the
newly synthesized compounds showed good amphiphilic proper-
ties. In order to check the correlation between the glucidoam-
phiphile structure and the surface tension characteristics, as well
as the antimicrobial activities of the surfactant molecules, we
currently extend the synthesis pathway to different molecules
containing other types of rigid spacer arm and one or more alkyl
groups.
was used as a standard to compare the synthesized products to
a large spectrum antibiotic. The results are presented in Table 2.
The alkyl-galactose derivatives exhibiting the best antimicrobial
activities were the C12 derivatives with non aromatic spacer arms,
whether they were protected or not, i.e. the products 3d, 3e, 4d and
4e. These compounds gave MIC values as low as 3.91
mg/mL
depending on the strain, which was similar to the chloramphenicol
action. The aromatic spacer arm seemed to cancel the antimicrobial
activity of the C12-derived products and no significant MIC could
be detected for compounds 3f or 4f, with only one exception of the
3f tested on M. luteus (MIC ¼ 31.25
mg/mL).
The MIC results were independant of the nature of the aliphatic
spacer arm and were equivalent with a butyl or a butynyl group.
Moreover, the protected galactose derivatives had similar antimi-
crobial activities as the deprotected ones (3d and 4d, or 3e and 4e)
exhibiting analogous MIC values.
4. Experimental section
All chemicals were purchased from Acros and Aldrich Chemicals
in their highest purity degree. All solvents were used as-supplied
without further purification. Distilled water was used in all
experiments. Analytical thin-layer chromatography (TLC) was per-
formed on E. Merck aluminum-backed silica gel (Silica Gel F254).
The antibacterial effects were clearly strain-dependant. The
compounds 3d, 3e, 4d and 4e had a better activity against Gram

L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
181
90
80
70
60
50
40
30
γ
20
1,00E-01
1,00E-07
1,00E-06
1,00E-05
1,00E-04
1,00E-03
1,00E-02
Fig. 1. Surface tension of 6-O-octyl-D-galactopyranose derivatives 4a, 4b and 4c with butyl-, butynyl- and benzyl-spacer arm respectively, and 6a without spacer arm, as a function
of concentration.
Compounds were identified using UV fluorescence and/or staining
with a solution of phosphomolybdic acid in aqueous sulfuric acid
and ethanol.
Melting points were determined on an electrothermal appa-
ratus and are uncorrected. Optical rotations, for solutions in CHCl3
or methanol, were measured at room temperature with a digital
polarimeter ATAGO model Polax-2L.
NMR spectra were recorded on a Bruker DRX300 spectrometer
operating at 300 MHz for 1H nuclei and 75 MHz for 13C nuclei. CDCl3
(99.50% isotopic purity) and DMSO d6 (99.80% isotopic purity) were
purchased from Euriso-Top. 1H NMR data are reported as chemical
shift, multiplicity (s, singlet; d, doublet; m, multiplet), relative
integral, coupling constant (J in hertz).
tension measurements at 293 K. A concentrated solution was
installed in a syringe and the addition of small volumes to ultrapure
water enhanced the solution concentration. After each addition, the
solution was gently stirred for 30 s. Equilibrium surface tension was
measured for each concentration. All surface tension values were
mean quantities of at least three measurements. The standard
deviation of the mean never deviated more than ꢁ1.5% of the mean.
The precision of the force transducer of the surface tension appa-
ratus was 0.1 mN/m and before each experiment, the platinum
plate was cleaned in red/orange color flame.
4.1. General procedure of monoalkylation using sodium hydroxide
The processor tensiometer Sigma 70 (KSV) and the Wilhelmy
plate method for airewater interface have been used for the surface
Hydroquinone (9.08 mmol, 1 equiv) and potassium hydroxide
(9.08 mmol, 1 equiv) were stirred at room temperature for 1 h in
90
γ
80
70
60
50
40
30
20
1,00E-07
1,00E-06
1,00E-05
1,00E-04
1,00E-03
1,00E-02
Fig. 2. Surface tension of 6-O-dodecyl-D-galactopyranose derivatives 4d, 4e and 4f with butyl-, butynyl- and benzyl-spacer arm respectively, and 6b without spacer arm, as
a function of concentration.

182
L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
DMF (40 mL), then n-alkyl bromide (10.9 mmol, 1.2 equiv) was
added slowly. After 24 h, the reaction was quenched with acid resin
DOWEX 50WX4 and the organic layer was extracted with ethyl
acetate, dried with Na2SO4 and concentrated under reduced pres-
sure. The monoalkylated derivative was isolated as a white solid
after purification by column chromatography with 7:3 hexanee
ethyl acetate.
4.2. General procedure of monoalkylation using sodium hydride
Sodium hydride (3.3 mmol, 1.1 equiv) was added at room
temperature to a solution of alcohol (3 mmol, 1 equiv) in DMF
(20 mL). The medium was stirred at room temperature during 1 h
and n-alkyl bromide (3.6 mmol, 1.2 equiv) was added slowly. After
1 h at room temperature, water was added to quench the reaction
and the organic layer was extracted with ethyl acetate, dried with
Na2SO4 and concentrated under reduced pressure. The mono-
alkylated derivative was isolated as a colorless oil after purification
by column chromatography with 7:3 hexaneeethyl acetate.
4.3. General procedure of tosylation
p-Toluenesulfonyl chloride (5.91 mmol, 1.5 equiv) in dichloro-
methane (10 mL) was added dropwise to a solution of mono-
alkylated diol (3.94 mmol, 1 equiv) and dimethylaminopyridine
(5.91 mmol, 1.5 equiv) in dichloromethane (20 mL) at ꢂ10 ꢀC. After
7 h at room temperature, the solution was filtered and concentrated
under reduced pressure. The tosylated derivative was isolated as
a viscous liquid after purification by column chromatography with
9:1 hexaneeethyl acetate.
4.4. General procedure of condensation between tosylated
derivatives and protected sugar
Finely powdered potassium hydroxide (10.6 mmol, 2 equiv) was
added to a solution of 1,2:3,4-di-O-isopropylidene-D-galactopyr-
anose (5.3 mmol, 1 equiv) in tolueneeMe2SO 3:2 (10 mL) at room
temperature. The reaction was stirred during 1 h at room temper-
ature and a solution of the tosylated derivative (5.3 mmol, 1 equiv)
in tolueneeMe2SO 3:2 (5 mL) was added to the medium. After 15 h,
the mixture was filtered and the filtrate neutralized with saturated
aq. NH4Cl. The organic phase was separated, washed with water
(twice), dried (Na2SO4) and concentrated under reduced pressure.
The 6-O-alkyl-1,2:3,4-di-O-isopropylidene-D-galactopyranose was
isolated as a white solid after purification by column chromatog-
raphy with 9:1 hexaneeethyl acetate.
4.5. General procedure for deprotection of diacetal derivatives
Diacetal derivative (3.2 mmol, 1 equiv) was added to a stirred
solution of 9:1 CF3COOHeH2O (5 mL) at room temperature. Cold
diethyl ether was added after 1 h and the solution was cooled at
ꢂ20 ꢀC. The desired products were filtered off, sucked dry, washed
with diethyl ether (twice) and recrystallized from THF to give 6-O-
n-alkyl-a-D-galactopyranose.
4.6. 4-O-n-Octyl-butan-1-ol (1a)
Colorless oil (8.70 g, 43.06 mmol, 78%); 1H NMR (CDCl3): 3.51
(2H, t, J 6.8 Hz, CH2OH), 3.39 (4H, m, CH2OR), 1.53 (m, 6H,
CH2CH2O), 1.29 (m, H alkyl chain), 0.82 (t, 3H, J 6.4 Hz, CH3); 13C
NMR (CDCl3): 71.4, 71.1 (CH2OR), 62.5 (CH2OH), 32.4e22.9 (CH2
alkyl chain), 14.4 (CH3).

L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
183
4.7. 4-O-n-Dodecyl-butan-1-ol (1d)
3H, J 6.3 Hz, CH3); 13C NMR (CDCl3): 145.4, 133.6, 130.2, 128.3 (Caro),
86.3 (C^), 70.8, 67.4 (CH2OR), 58.3 (CH2OTs), 32.2e23.0 (CH2 alkyl
chain), 22.1 (CH3 tosyl),14.5 (CH3).
Colorless oil (3.46 g, 13.43 mmol, 90%); 1H NMR (CDCl3): 3.53
(2H, t, J 6.8 Hz, CH2OH), 3.35 (4H, m, CH2OR), 1.54 (m, 6H,
CH2CH2O), 1.19 (m, H alkyl chain), 0.81 (t, 3H, J 6.4 Hz, CH3); 13C
NMR (CDCl3): 71.4, 71.1 (CH2OR), 62.6 (CH2OH), 32.3e23.0 (CH2
alkyl chain), 14.4 (CH3).
4.15. 4-O-n-Dodecyl-but-2-yn-1-yl tosylate (2e)
White solid (2.41 g, 5.91 mmol, 75%); 1H NMR (CDCl3): 7.80, 7.34
(4H, d, J 8.1 Hz, Haro), 4.74 (2H, m, ^CCH2OTs), 4.02 (2H, m,
ROCH2C^), 3.38 (t, 2H, CH2OCH2C^, J 6.7 Hz), 2.45 (s, 3H, CH3
tosyl),1.52 (m, 2H, CH2CH2OCH2C^), 1.29 (m, H alkyl chain), 0.88 (t,
3H, J 7.1 Hz, CH3); 13C NMR (CDCl3): 145.4, 133.5, 130.2, 128.5 (Caro),
86.3 (C^), 70.8, 67.2 (CH2OR), 58.3 (CH2OTs), 32.2e23.0 (CH2 alkyl
chain), 22.0 (CH3 tosyl),14.2 (CH3).
4.8. 4-O-n-Octyl-but-2-yn-1-ol (1b)
Colorless oil (8.99 g, 45.40 mmol, 78%); 1H NMR (CDCl3): 4.21 (2H,
m, ^CCH2OH), 4.12 (2H, m, ROCH2C^), 3.43 (t, 2H, CH2OCH2C^, J
6.8 Hz),1.48 (m, 2H, CH2CH2OCH2C^),1.22 (m, H alkyl chain), 0.82 (t,
3H, J 7.0 Hz, CH3); 13C NMR (CDCl3): 85.9 (C^), 69.7, 58.2 (CH2OR),
51.3 (CH2OH), 32.2e23.0 (CH2 alkyl chain), 14.2 (CH3).
4.16. p O-n-Octyl-phenyl tosylate (2c)
4.9. 4-O-n-Dodecyl-but-2-yn-1-ol (1e)
White solid (2.81 g, 7.47 mmol, 90%); 1H NMR (CDCl3): 7.66, 7.30
(4H, d, J 8.1 Hz, Haro), 6.85, 6.74 (4H, d, J 8.0 Hz, Haro), 3.87 (2H, t,
CH2OR, J 6.3 Hz), 2.45 (s, 3H, CH3 tosyl), 1.73 (m, 2H, CH2CH2OR, J
6.8 Hz), 1.27 (m, H alkyl chain), 0.88 (t, 3H, J 7.2 Hz, CH3); 13C NMR
(CDCl3): 158.2, 145.7, 115.3 (4 ꢄ Caro), 145.4, 133.5, 130.2, 128.5 (Caro
tosyl)68.7 (CH2OR), 32.3e22.1 (CH2 alkyl chain), 22.0 (CH3 tosyl),14.5
(CH3).
Colorless oil (5.87 g, 23.03 mmol, 79%); 1H NMR (CDCl3): 4.29
(2H, m, ^CCH2OH), 4.16 (2H, m, ROCH2C^), 3.49 (t, 2H,
CH2OCH2C^, J 6.7 Hz), 1.84 (m, 2H, CH3CH2(CH2)10), 1.58 (m, 2H,
CH2CH2OCH2C^), 1.26 (m, H alkyl chain), 0.87 (t, 3H, J 7.0 Hz, CH3);
13C NMR (CDCl3): 85.9 (C^), 69.7, 58.2 (CH2OR), 51.3 (CH2OH),
32.2e23.0 (CH2 alkyl chain), 14.2 (CH3).
4.17. p O-n-Dodecyl-phenyl tosylate (2f)
4.10. p-O-n-Octyl-phenol (1c)
White solid (3.12 g, 7.22 mmol, 92%); 1H NMR (CDCl3): 7.66, 7.30
(4H, d, J 8.1 Hz, Haro), 6.85, 6.74 (4H, d, J 8.0 Hz, Haro), 3.85 (2H, t,
CH2OR, J 6.5 Hz), 2.47 (s, 3H, CH3 tosyl), 1.71 (m, 2H, CH2CH2OR, J
6.9 Hz), 1.29 (m, H alkyl chain), 0.85 (t, 3H, J 7.2 Hz, CH3); 13C NMR
(CDCl3): 158.2, 145.7, 115.3 (4 ꢄ Caro), 145.4, 133.5, 130.2, 128.5 (Caro
tosyl) 68.5 (CH2OR), 32.1e22.2 (CH2 alkyl chain), 21.8 (CH3 tosyl),
14.4 (CH3).
White solid (6.05 g, 27.24 mmol, 60%); 1H NMR (CDCl3): 6.79
(4H, m, Haro), 3.88 (2H, t, CH2OR, J 8.9 Hz), 1.69 (m, 2H, CH2CH2OR),
1.24 (m, H alkyl chain), 0.94 (t, 3H, J 6.5 Hz, CH3); 13C NMR (CDCl3):
153.4, 150.1, 116.2, 115.9 (4 ꢄ Caro), 69.3 (CH2OR), 32.3e23.1 (CH2
alkyl chain), 14.6 (CH3).
4.11. p-O-n-Dodecyl-phenol (1f)
4.18. 6-O-(4-O-n-Octyl-but-1-yl)-1,2:3,4-di-O-isopropylidene-a-D-
galactopyranose (3a)
White solid (8.08 g, 29.06 mmol, 64%); 1H NMR (CDCl3): 6.79
(4H, m, Haro), 3.91 (2H, t, CH2OR, J 9.0 Hz), 1.77 (m, 2H, CH2CH2OR, J
6.8 Hz), 1.29 (m, H alkyl chain), 0.91 (t, 3H, J 6.3 Hz, CH3); 13C NMR
(CDCl3): 153.6,149.8,116.4,116.1 (4 ꢄ Caro), 69.3 (CH2OR), 32.3e23.1
(CH2 alkyl chain), 14.6 (CH3).
White solid (1.21 g, 2.72 mmol, 85%); (Found: C, 64.7; H, 10.0.
C24H44O7 requires C, 64.8; H, 10.0%); m.p. 83 ꢀC; ½a D26
ꢅ
ꢂ18.2ꢀ (c 0.7;
CHCl3); 1H NMR (CDCl3):
d 5.51 (1H, d, J1e2 5.0 Hz, H-1), 4.59 (1H,dd,
J3,4 7.9 Hz, H-3), 4.31 (1H, dd, J2,3 2.3 Hz, H-2), 4.26 (1H, dd, J4,5
1.8 Hz, H-4), 3.97 (1H, m, H-5), 3.48 (6H, m, H-6, CH2O), 1.65e1.40
(12H, s, CH3 iso), 1.27 (m, H alkyl chain), 0.88 (t, 3H, J 6.4 Hz, CH3);
4.12. 4-O-n-Octyl-but-1-yl tosylate (2a)
White solid (2.61 g, 7.33 mmol, 74%); 1H NMR (CDCl3): 7.79, 7.32
(4H, d, J 8.4, Haro), 4.05 (2H, t, J 6.3 Hz, CH2OTs), 3.34 (4H, m,
CH2OR), 2.44 (s, 3H, CH3 tosyl), 1.72e1.26 (m, H alkyl chain), 0.87 (t,
3H, J 6.4 Hz, CH3); 13C NMR (CDCl3): 145.0, 133.6, 130.2, 128.3 (Caro),
71.4, 70.9, 70.1(CH2OR), 32.2e23.0 (CH2 alkyl chain), 22.0 (CH3
tosyl), 14.4 (CH3).
13C NMR (CDCl3):
d 109.6, 108.9 (Ciso), 96.8 (C-1), 71.7 (C-4), 71.6
(CH2O), 71.4 (C-3), 71.0 (2 ꢄ CH2O, C-2), 69.7 (C-5), 67.1 (C-6), 32.3e
23.0 (CH2 alkyl chain), 26.7, 25.1, 23.2, 22.9 (4 ꢄ CH3 iso), 14.5 (CH3).
4.19. 6-O-(4-O-n-Dodecyl-but-1-yl)-1,2:3,4-di-O-isopropylidene-
a-D
-galactopyranose (3d)
4.13. 4-O-n-Dodecyl-but-1-yl tosylate (2d)
White solid (1.83 g, 3.66 mmol, 91%); (Found: C, 67.1; H, 10.6.
C28H52O7 requires C, 67.2; H,10.5%); m.p. 95 ꢀC; ½a D26
ꢅ
¼ ꢂ21.3ꢀ (c 0.8;
White solid (2.64 g, 6.40 mmol, 82%); 1H NMR (CDCl3): 7.80, 7.34
(4H, d, J 8.1, Haro), 4.07 (2H, t, J 6.1 Hz, CH2OTs), 3.32 (4H, m, CH2OR),
2.46 (s, 3H, CH3 tosyl), 1.80e1.29 (m, H alkyl chain), 0.85 (t, 3H, J
6.6 Hz, CH3); 13C NMR (CDCl3): 144.9, 133.4, 130.2, 128.4 (Caro), 71.2,
70.9, 69.9 (CH2OR), 32.1e23.2 (CH2 alkyl chain), 21.9 (CH3 tosyl),
14.3 (CH3).
CHCl3); 1H NMR (CDCl3):
d 5.53 (1H, d, J1e2 5.1 Hz, H-1), 4.59 (1H,dd,
J3,4 7.8 Hz, H-3), 4.31 (1H, dd, J2,3 2.4 Hz, H-2), 4.23 (1H, dd, J4,5 1.8 Hz,
H-4), 3.97 (1H, m, H-5), 3.48 (6H, m, H-6, CH2O), 1.65e1.34 (12H, s,
CH3 iso), 1.25 (m, H alkyl chain), 0.89 (t, 3H, J 6.5 Hz, CH3); 13C NMR
(CDCl3): d 109.6, 108.9 (Ciso), 96.8 (C-1), 71.7 (C-4), 71.6 (CH2O), 71.4
(C-3), 71.0 (2 ꢄ CH2O, C-2), 69.7 (C-5), 67.1 (C-6), 32.4e22.7 (CH2
alkyl chain), 26.7, 25.1, 23.4, 23.2 (4 ꢄ CH3 iso), 14.4 (CH3).
4.14. 4-O-n-Octyl-but-2-yn-1-yl tosylate (2b)
4.20. 6-O-(4-O-n-Octyl-but-2-yn-1-yl)-1,2:3,4-di-O-
isopropylidene-a-D-galactopyranose (3b)
White solid (2.59 g, 7.37 mmol, 73%); 1H NMR (CDCl3): 7.80, 7.34
(4H, d, J 8.1 Hz, Haro), 4.75 (2H, m, ^CCH2OTs), 4.03 (2H, m,
ROCH2C^), 3.39 (t, 2H, CH2OCH2C^, J 6.6 Hz), 2.46 (s, 3H, CH3
tosyl), 1.54 (m, 2H, CH2CH2OCH2C^), 1.30 (m, H alkyl chain), 0.87 (t,
White solid (1.07 g, 2.43 mmol, 74%); (Found: C, 65.4; H, 9.1.
C24H40O7 requires C, 65.4; H, 9.15%); m.p. 78 ꢀC; ½a D26
ꢅ
ꢂ20.1ꢀ (c 0.6;

184
L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
CHCl3); 1H NMR (CDCl3):
d
5.54 (1H, d, J1e2 4.8 Hz, H-1), 4.60 (1H,
4.25. 6-O-(4-O-n-Dodecyl-but-1-yl)-a-D-galactopyranose (4d)
dd, J3,4 8.0 Hz, H-3), 4.31 (1H, dd, J2,3 2.5 Hz, H-2), 4.26 (3H, m, H-4,
^CCH2), 4.16 (2H, m, ROCH2C^), 3.90 (1H, m, H-5), 3.75 (1H, dd,
J6ae6b 11.2 Hz, J6ae5 5.1 Hz, H-6a), 3.65 (1H, dd, J6be5 6.9 Hz, H-6b),
3.48 (2H, m, CH2O), 1.54e1.33 (12H, s, CH3 iso), 1.27 (m, H alkyl
White solid (1.38 g, 3.29 mmol, 90%); (Found: C, 62.8; H, 10.6.
C22H44O7 requires C, 62.8; H, 10.5%); m.p. 121 ꢀC; ½a D26
ꢅ
24.1ꢀ (c 0.5;
MeOH); 1H NMR (DMSO d6):
d 4.92 (1H, d, J1e2 3.0 Hz, H-1), 3.93
chain), 0.88 (t, 3H, J 6.4 Hz, CH3); 13C NMR (CDCl3):
d
109.7, 109.0
(1H, m, H-5), 3.65 (1H, dd, J4,3 3.1 Hz, H-4), 3.50 (1H, dd, J3,2 7.2 Hz,
H-3), 3.48 (1H, dd, H-2), 3.43 (6H, m, CH2O), 3.38 (1H, dd, J6ae6b 9.1,
H-6a), 3.35 (1H, dd, J6be5 5.5 Hz, H-6b), 1.53e1.21 (m, H alkyl chain),
(Ciso), 96.7 (C-1), 83.2, 82.3 (C^), 71.6 (C-4), 71.1 (C-3), 70.9 (C-2)
70.7 (CH2O), 69.0 (C-5), 61.8 (C-6), 59.2, 58.6 (OCH2C^), 32.3e23.0
(CH2 alkyl chain), 26.5, 25.3, 23.0 (4 ꢄ CH3 iso), 14.5 (CH3).
0.83 (t, 3H, J 6.9 Hz, CH3); 13C NMR (DMSO d6):
d 92.5 (C-1), 70.1 (C-
6), 69.3 (C-4), 69.1 (C-3), 68.4 (CH2O), 68.3 (C-2), 67.9 (C-5), 31.5e
21.6 (CH2 alkyl chain), 13.3 (CH3).
4.21. 6-O-(4-O-n-Dodecyl-but-2-yn-1-yl)-1,2:3,4-di-O-
isopropylidene-a-D-galactopyranose (3e)
4.26. 6-O-(4-O-n-Octyl-but-2-yn-1-yl)-a-D-galactopyranose (4b)
White solid (1.26 g, 3.03 mmol, 81%); (Found: C, 67.7; H, 9.7.
White solid (0.77 g, 2.14 mmol, 88%); (Found: C, 59.9; H, 8.9.
C28H48O7 requires C, 67.7; H, 9.7%); m.p. 91 ꢀC; ½a D26
ꢅ
ꢂ23.4ꢀ (c 0.7;
C18H32O7 requires C, 60.0; H, 8.95%); m.p. 108 ꢀC; ½a D26
ꢅ
20.3ꢀ (c 0.5;
CHCl3); 1H NMR (CDCl3):
d 5.56 (1H, d, J1e2 5.0 Hz, H-1), 4.57 (1H,
MeOH); 1H NMR (DMSO d6):
d 4.91 (1H, d, J1e2 3.3 Hz, H-1), 4.21
dd, J3,4 7.9 Hz, H-3), 4.31 (1H, dd, J2,3 2.6 Hz, H-2), 4.27 (3H, m, H-4,
^CCH2), 4.16 (2H, m, ROCH2C^), 3.92 (1H, m, H-5), 3.77 (1H, dd,
J6ae6b 10.9 Hz, J6ae5 5.0 Hz, H-6a), 3.66 (1H, dd, J6be5 7.0 Hz, H-6b),
3.48 (2H, m, CH2O), 1.52e1.28 (12H, s, CH3 iso), 1.25 (m, H alkyl
(2H, m, H-4, ^CCH2), 4.12 (2H, m, ROCH2C^), 4.01 (1H, m, H-5),
3.68 (1H, dd, J4,3 3.1 Hz, H-4), 3.53 (1H, dd, J3,2 7.8 Hz, H-3), 3.48 (1H,
dd, H-2), 3.45 (2H, m, CH2O), 3.39 (1H, dd, J6ae6b 9.1, H-6a), 3.32
(1H, dd, J6be5 5.7 Hz, H-6b), 1.50e1.25 (m, H alkyl chain), 0.88 (t, 3H,
chain), 0.85 (t, 3H, J 6.4 Hz, CH3); 13C NMR (CDCl3):
d 109.7, 109.0
J 6.5 Hz, CH3); 13C NMR (DMSO d6):
d 92.5 (C-1), 70.1 (C-6), 69.3 (C-
(Ciso), 96.7 (C-1), 83.2, 82.3 (C^), 71.6 (C-4), 71.1 (C-3), 70.9 (C-2)
70.7 (CH2O), 69.0 (C-5), 61.8 (C-6), 59.2, 58.6 (OCH2C^), 32.6e22.8
(CH2 alkyl chain), 26.5, 25.3, 23.0 (4 ꢄ CH3 iso), 14.5 (CH3).
4), 69.1 (C-3), 68.6 (CH2O), 68.5 (C-2), 68.3 (C-5), 31.2e22.0 (CH2
alkyl chain), 13.6 (CH3).
4.27. 6-O-(4-O-n-Dodecyl-but-2-yn-1-yl)-a-D-galactopyranose
4.22. 6-O-(p O-n-Octyl-phen-1-yl)-1,2:3,4-di-O-isopropylidene-
-galactopyranose (3c)
a-
(4e)
D
White solid (0.99 g, 2.39 mmol, 79%); (Found: C, 63.3; H, 9.7.
White solid (1.46 g, 3.15 mmol, 62%); (Found: C, 67.1; H, 8.7.
C22H40O7 requires C, 63.4; H, 9.7%); m.p. 113 ꢀC; ½a D26
ꢅ
21.4ꢀ (c 0.5;
C26H40O7 requires C, 67.2; H, 8.7%); m.p. 95 ꢀC; ½a D26
ꢅ
ꢂ23.7ꢀ (c 0.9;
MeOH); 1H NMR (DMSO d6):
d 4.95 (1H, d, J1e2 3.7 Hz, H-1), 4.19
CHCl3); 1H NMR (CDCl3):
d
7.77, 7.29 (4H, d, J 8.1 Hz, Haro), 5.40 (1H,
(2H, m, H-4, ^CCH2), 4.11 (2H, m, ROCH2C^), 3.98 (1H, m, H-5),
3.70 (1H, dd, J4,3 3.4 Hz, H-4), 3.53 (1H, dd, J3,2 7.9 Hz, H-3), 3.45 (1H,
dd, H-2), 3.42 (2H, m, CH2O), 3.39 (1H, dd, J6ae6b 9.5, H-6a), 3.35
(1H, dd, J6be5 5.9 Hz, H-6b), 1.54e1.19 (m, H alkyl chain), 0.91 (t, 3H,
d, J1e2 5.1 Hz, H-1), 4.53 (1H, dd, J3,4 7.8 Hz, H-3), 4.25 (1H, dd, J2,3
5.4 Hz, H-2), 4.16 (2H, m, H-4 H-6a), 4.05 (2H, m, H-5 H-6b),1.27 (m,
H alkyl chain), 1.17 (t, 3H, J 6.4 Hz, CH3); 13C NMR (CDCl3):
d 145.2,
133.1, 130.1, 128.4 (Caro), 109.9, 109.2 (Ciso), 96.5 (C-1), 70.8 (C-4),
70.7 (C-3), 70.6 (C-2) 68.6 (CH2O), 66.2 (C-5), 60.7 (C-6), 32.3e23.0
(CH2 alkyl chain), 14.5 (CH3).
J 6.5 Hz, CH3); 13C NMR (DMSO d6):
d 92.5 (C-1), 70.3 (C-6), 69.1 (C-
4), 68.9 (C-3), 68.6 (CH2O), 68.5 (C-2), 68.1 (C-5), 31.6e21.5 (CH2
alkyl chain), 13.6 (CH3).
4.28. 6-O-(p O-n-Octyl-phen-1-yl)-a-D-galactopyranose (4c)
4.23. 6-O-(p O-n-Dodecyl-phen-1-yl)-1,2:3,4-di-O-isopropylidene-
a
-
D
-galactopyranose (3f)
White solid (0.91 g, 2.36 mmol, 75%); (Found: C, 62.5; H, 8.4.
C20H32O7 requires C, 62.5; H, 8.4%); m.p. 139 ꢀC; ½a D26
ꢅ
28.2 (c 0.5;
White solid (1.02 g, 1.96 mmol, 66%); (Found: C, 69.1; H, 9.2.
MeOH); 1H NMR (DMSO d6):
d 7.80, 7.32 (4H, d, J 8.3 Hz, Haro), 4.91
C30H48O7 requires C, 69.2; H, 9.3%); m.p. 98 ꢀC; ½a D26
ꢅ
ꢂ25.2ꢀ (c 0.7;
(1H, d, J1e2 3.3 Hz, H-1), 3.96 (1H, m, H-5), 3.69 (1H, dd, J4,3 2.7 Hz,
H-4), 3.59 (1H, dd, J3,2 7.7 Hz, H-3), 3.55 (1H, dd, H-2), 3.45 (2H, m,
CH2O), 3.38 (1H, dd, J6ae6b 9.7 Hz, H-6a), 3.32 (1H, dd, J6be5 6.1 Hz,
H-6b), 1.49e1.27 (m, H alkyl chain), 0.85 (t, 3H, J 6.4 Hz, CH3); 13C
NMR (DMSO d6): d 151.2, 143.1, 138.1, 131.4 (Caro), 92.8 (C-1), 71.1 (C-
6), 69.1 (C-4), 68.9 (C-3), 68.4 (CH2O), 68.1 (C-2), 59.9 (C-5), 31.2e
22.0 (CH2 alkyl chain), 13.4 (CH3).
CHCl3); 1H NMR (CDCl3):
d 7.79, 7.30 (4H, d, J 8.0 Hz, Haro), 5.45 (1H,
d, J1e2 5.3 Hz, H-1), 4.53 (1H, dd, J3,4 7.7 Hz, H-3), 4.27 (1H, dd, J2,3
5.3 Hz, H-2), 4.16 (2H, m, H-4 H-6a), 4.07 (2H, m, H-5 H-6b),1.26 (m,
H alkyl chain), 1.15 (t, 3H, J 6.4 Hz, CH3); 13C NMR (CDCl3):
d 145.2,
133.1, 130.1, 128.4 (Caro), 109.8, 109.4 (Ciso), 96.4 (C-1), 71.1 (C-4),
70.8 (C-3), 70.7 (C-2) 68.4 (CH2O), 66.3 (C-5), 60.7 (C-6), 32.7e22.8
(CH2 alkyl chain), 14.3 (CH3).
4.29. 6-O-(p O-n-Dodecyl-phen-1-yl)-a-D-galactopyranose (4f)
4.24. 6-O-(4-O-n-Octyl-but-1-yl)-a-D-galactopyranose (4a)
White solid (0.65 g, 1.49 mmol, 76%); (Found: C, 65.4; H, 9.2.
White solid (0.84 g, 2.31 mmol, 85%); (Found: C, 59.2; H, 10.0.
C24H40O7 requires C, 65.4; H, 9.15%); m.p. 128 ꢀC; ½a D26
ꢅ
26.7ꢀ (c 0.5;
C18H36O7 requires C, 59.3; H, 10.0%); m.p. 119 ꢀC; ½a D26
ꢅ
22.4ꢀ (c 0.8;
MeOH); 1H NMR (DMSO d6):
d 7.79, 7.35 (4H, d, J 8.5 Hz, Haro), 4.93
MeOH); 1H NMR (DMSO d6):
d
4.93 (1H, d, J1e2 3.1 Hz, H-1), 3.91
(1H, d, J1e2 3.6 Hz, H-1), 3.95 (1H, m, H-5), 3.71 (1H, dd, J4,3 3.0 Hz,
H-4), 3.57 (1H, dd, J3,2 7.9 Hz, H-3), 3.56 (1H, dd, H-2), 3.42 (2H, m,
CH2O), 3.34 (1H, dd, J6ae6b 9.9 Hz, H-6a), 3.29 (1H, dd, J6be5 6.3 Hz,
H-6b), 1.53e1.23 (m, H alkyl chain), 0.84 (t, 3H, J 6.4 Hz, CH3); 13C
(1H, m, H-5), 3.62 (1H, dd, J4,3 2.9 Hz, H-4), 3.53 (1H, dd, J3,2 7.5 Hz,
H-3), 3.50 (1H, dd, H-2), 3.45 (6H, m, CH2O), 3.40 (1H, dd, J6ae6b 9.4,
H-6a), 3.36 (1H, dd, J6b-5 5.8 Hz, H-6b), 1.49e1.27 (m, H alkyl chain),
0.85 (t, 3H, J 6.4 Hz, CH3); 13C NMR (DMSO d6):
d
92.5 (C-1), 70.1 (C-
NMR (DMSO d6): d 151.2, 143.1, 138.1, 131.4 (Caro), 92.7 (C-1), 71.1 (C-
6), 69.3 (C-4), 69.1 (C-3), 68.6 (CH2O), 68.5 (C-2), 68.3 (C-5), 31.2e
22.0 (CH2 alkyl chain), 13.6 (CH3).
6), 69.3 (C-4), 68.7 (C-3), 68.5 (CH2O), 68.1 (C-2), 59.7 (C-5), 31.6e
21.5 (CH2 alkyl chain), 13.5 (CH3).

L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
185
mannitols displaying amphiphilic and mesophasic properties, Carbohydr. Res.
339 (2004) 1817e1821.
4.30. Analysis of antimicrobial properties
[6] M.V. de Almeida, M. Le Hyaric, G.W. Amarante, M.C.S. Lourenço, M.L.L. Brandão,
Synthesis of amphiphilic galactopyranosyl diamines and amino alcohols as
antitubercular agents, Eur. J. Med. Chem. 42 (2007) 1076e1083.
[7] L. Chaveriat, C. Meyer, D. Beaupère, G. Demailly, I. Stasik, 6-S-alkyl-6-
thiohexonolactones and 6-S-alkyl-6-thiohexitols: efficient synthesis and
new smectic liquid crystal phases, J. Mol. Liq. 142 (2008) 17e21.
[8] K. Holmberg, Novel surfactants. Preparation, applications and biodegrad-
ability, second ed., in: M. Dekker (Ed.), Surfactant Science Series, vol. 114,
New York, 2003, pp. 1e192.
[9] S. Warwel, F. Brüse, C. Demes, M. Kunz, M. Rüsch gen Klaas, Polymers and
surfactants on the basis of renewable resources, Chemosphere 43 (2001) 39e48.
[10] Y. Queneau, S. Chambert, C. Besset, R. Cheaib, Recent progress in the synthesis
of carbohydrate-based amphiphilic materials: the examples of sucrose and
isomaltulose, Carbohydr. Res. 343 (2008) 1999e2009.
[11] M. le Maire, P. Champeil, J.V. Møller, Interaction of membrane proteins and
lipids with solubilizing detergents, Biochim. Biophys. Acta 1508 (2000) 86e111.
[12] J. Piao, S. Kishi, S. Adachi, Surface tensions of aqueous solutions of 1-O-
monoacyl sugar alcohols, Colloids Surf. A Physicochem. Eng. Aspects 277
(2006) 15e19.
4.30.1. Organisms and culture conditions
The antimicrobial activity of the 6-O-alkyl-galactose derivatives
was tested on ten bacterial strains: B. stearothermophilus ATTC
12980, C. freundii ATCC 6750, E. cloacae ATCC 13047, E. chrysanthemi
EC3937, E. coli ATCC 10536, M. luteus CIP 53.45, M. smegmatis CIP
73.26, P. aeruginosa CIP A22, S. meliloti Rm1021 and S. aureus subsp
aureus CIP 53.154.
Stock cultures were maintained in Luria Bertani (LB) broth
supplemented with 20% glycerol at ꢂ80 ꢀC. Cultures were realized
from 1% (v/v) stock cultures in a nutritive broth identical for all the
strains (Yeast extract 1 g/L, Meat extract 2 g/L, Peptone 5 g/L and
NaCl 5 g/L) at 30 ꢀC, except for M. smegmatis which was incubated
in a LB/Tween 80 medium (Tryptone 10 g/L, Yeast extract 5 g/L, NaCl
5 g/L and Tween 80 at 0.05%) at 37 ꢀC. Working suspensions were
obtained from an overnight culture, eventually diluted in nutritive
broth (or LB/Tween 80 medium for M. smegmatis), to reach
a bacterial density equivalent to a 0.5 McFarland standard, corre-
sponding approximatively to 1.5 ꢄ 108 cfu/mL.
[13] M.N. Jones, Surfactants in membrane solubilisation, Int. J. Pharm. 177 (1999)
137e159.
[14] H. Ahyayauch, M. Bennouna, A. Alonso, F.M. Goñi, Detergent effects on
membranes at subsolubilizing concentrations: transmembrane lipid motion,
bilayer permeabilization, and vesicle lysis/reassembly are independent
phenomena, Langmuir 26 (2010) 7307e7313.
[15] J.D. Rose, J.A. Maddry, R.N. Comber, W.J. Suling, L.N. Wilson, R.C. Reynolds,
Synthesis and biological evaluation of trehalose analogs as potential inhibitors
of mycobacterial cell wall biosynthesis, Carbohydr. Res. 337 (2002) 105e120.
[16] N. Tewari, V.K. Tiwari, R.P. Tripathi, V. Chaturvedi, A. Srivastava, R. Srivastava,
P.K. Shukla, A.K. Chaturvedi, A. Gaikwad, S. Sinha, B.S. Srivastava, Synthesis of
galactopyranosyl amino alcohols as a new class of antitubercular and anti-
fungal agents, Bioorg. Med. Chem. Lett. 14 (2004) 329e332.
[17] D. Katiyar, V.K. Tiwari, N. Tewari, S.S. Verma, S. Sinha, A. Gaikwad,
A. Srivastava, V. Chaturvedi, R. Srivastava, B.S. Srivastava, R.P. Tripathi,
Synthesis and antimycobacterial activities of glycosylated amino alcohols and
amines, Eur. J. Med. Chem. 40 (2005) 351e360.
[18] R.P. Tripathi, V.K. Tiwari, N. Tewari, D. Katiyar, N. Saxena, S. Sinha, A. Gaikwad,
A. Srivastava, V. Chaturvedi, Y.K. Manju, R. Srivastava, B.S. Srivastava,
Synthesis and antitubercular activities of bis-glycosylated diamino alcohols,
Bioorg. Med. Chem. 13 (2005) 5668e5679.
[19] A.F. Taveira, M. Le Hyaric, E.F.C. Reis, D.P. Araújo, A.P. Ferreira, M. Aparecida de
Souza, L.L. Alves, M.C.S. Lourenço, F.R.C. Vicente, M.V. de Almeida, Preparation
and antitubercular activities of alkylated amino alcohols and their glycosy-
lated derivatives, Bioorg. Med. Chem. 15 (2007) 7789e7794.
4.30.2. Assay of antimicrobial activity in liquid medium
Stock solutions of the 6-O-alkyl-galactose derivatives were
prepared at 1 g/L in DMSO and were then serially diluted in sterile
nutritive broth (or LB/Tween 80 medium for M. smegmatis) to a final
volume of 2 mL. Two milliliters of bacterial working suspension
were added to each tube and the final concentration of 6-O-alkyl-
galactose derivatives ranged from 1:2 to 1:512. Several controls
were realized for each microorganism and each synthesized
compound, as previously described [29]. A supplementary control
consisted in inoculated nutritive broth without compounds but
with chloramphenicol, which is a broad-spectrum antibiotic, in
order to compare its antimicrobial activity to those of the 6-O-alkyl-
galactose derivatives.
Then each tube was incubated for 24 h in a rotary shaker
(150 rpm) at 30 ꢀC, except M. smegmatis at 37 ꢀC. Bacterial density
was determined spectrophotometrically at 600 nm. The MIC was
defined as the lowest concentration of compound that showed no
increase in Optical Density (OD) values for all the replicates
compared to the negative control after incubation. Each experiment
was replicated three times.
[20] C.B. Davis, R.D. Hartnell, P.D. Madge, D.J. Owen, R.J. Thomson, A.K. Chong,
R.L. Coppel, M. von Itzstein, Synthesis and biological evaluation of gal-
actofuranosyl alkyl thioglycosides as inhibitors of mycobacteria, Carbohydr.
Res. 342 (2007) 1773e1780.
[21] A.K. Sanki, J. Boucau, P. Srivastava, S.S. Adams, D.R. Ronning, S.J. Sucheck,
Synthesis of methyl 5-S-alkyl-5-thio-D-arabinofuranosides and evaluation of
their antimycobacterial activity, Bioorg. Med. Chem. 16 (2008) 5672e5682.
[22] S. Quader, S.E. Boyd, I.D. Jenkins, T.A. Houston, Multisite modification of
neomycin B: combined Mitsunobu and click chemistry approach, J. Org. Chem.
72 (2007) 1962e1979.
[23] J. Zhang, F.-I. Chiang, L. Wu, P.G. Czyryca, D. Li, C.-W. Tom Chang, Surprising
alteration of antibacterial activity of 500-modified neomycin against resistant
bacteria, J. Med. Chem. 51 (2008) 7563e7573.
[24] S. Pal, K. Mitra, S. Azmi, J.K. Ghosh, T.K. Chakraborty, Towards the synthesis of
sugar amino acid containing antimicrobial noncytotoxic CAP conjugates with
gold nanoparticles and a mechanistic study of cell disruption, Org. Biomol.
Chem. 9 (2011) 4806e4810.
Acknowledgments
This work was supported by the Ministère de l’Enseignement
Supérieur et de la Recherche. The authors thank Adrien Valingot
and Cécile Poret for their excellent microbiological support.
[25] N. Gernigon, V. Bordeau, F. Berrée, B. Felden, B. Carboni, Synthesis and anti-
bacterial activity of novel neamine derivatives: preponderant role of the
substituent position on the neamine core, Org. Biomol. Chem. 10 (2012)
4720e4730.
[26] R.P. Tripathi, R. Tripathi, V.K. Tiwari, L. Bala, S. Sinha, A. Srivastava,
R. Srivastava, B.S. Srivastava, Synthesis of glycosylated beta-amino acids as
new class of antitubercular agents, Eur. J. Med. Chem. 37 (2002) 773e781.
[27] M. Ferrer, J. Soliveri, F.J. Plou, N. López-Cortés, D. Reyes-Duarte,
M. Christensen, J.L. Copa-Patiño, A. Ballesteros, Synthesis of sugar esters in
solvent mixtures by lipases from Thermomyces lanuginosus and Candida
antarctica B, and their antimicrobial properties, Enzyme Microb. Technol. 36
(2005) 391e398.
Appendix A. Supplementary data
Supplementary data related to this article can be found at http://
dx.doi.org/10.1016/j.ejmech.2012.12.032.
References
[1] T. Plat, R.J. Linhardt, Syntheses and applications of sucrose-based esters,
J. Surfactants Deterg. 4 (2001) 415e421.
[2] M. Abert, N. Mora, J.-M. Lacombe, Synthesis and surface-active properties of
[28] A. Smith, P. Nobmann, G. Henehan, P. Bourke, J. Dunne, Synthesis and anti-
microbial evaluation of carbohydrate and polyhydroxylated non-
carbohydrate fatty acid ester and ether derivatives, Carbohydr. Res. 343
(2008) 2557e2566.
[29] P. Nobmann, A. Smith, J. Dunne, G. Henehan, P. Bourke, The antimicrobial
efficacy and structure activity relationship of novel carbohydrate fatty acid
derivatives against Listeria spp. and food spoilage microorganisms, Int. J. Food
Microbiol. 128 (2009) 440e445.
a new class of surfactants derived from D-gluconic acid, Carbohydr. Res. 337
(2002) 997e1006.
[3] D. Pilakowska-Pietras, K. Lunkenheimer, A. Piasecki, Synthesis of novel N,N-di-
n-alkylaldonamides and properties of their surface chemically pure adsorption
layers at the air/water interface, J. Colloid Interface Sci. 271 (2004) 192e200.
[4] C. Satgé, R. Granet, B. Verneuil, Y. Champavier, P. Krausz, Synthesis and
properties of new bolaform and macrocyclic galactose-based surfactants ob-
tained by olefin metathesis, Carbohydr. Res. 339 (2004) 1243e1254.
[30] A.K. Pathak, V. Pathak, L. Seitz, J.A. Maddry, S.S. Gurcha, G.S. Besra, W.J. Suling,
[5] L. Chaveriat, I. Stasik, G. Demailly, D. Beaupère, Direct syntheses of S-alkylthio-
D-galactono-, D-mannono-1,4-lactones, S-alkylthio-L-galactitols and D-
R.C. Reynolds, Studies on (
b,1/5) and (b,1/6) linked octyl Galf disaccharides

186
L. Chaveriat et al. / European Journal of Medicinal Chemistry 62 (2013) 177e186
as substrates for mycobacterial galactosyltransferase activity, Bioorg. Med.
Chem. 9 (2001) 3129e3143.
[31] A.K. Pathak, V. Pathak, W.J. Suling, S.S. Gurcha, C.B. Morehouse, G.S. Besra,
J.A. Maddry, R.C. Reynolds, Studies on n-octyl-5-( -arabinofuranosyl)-b-D-
Propionibacterium acnes: its therapeutic potential for inflammatory acne
vulgaris, J. Invest. Dermatol. 129 (2009) 2480e2488.
[44] T. Watanabe, S. Katayama, M. Matsubara, Y. Honda, M. Kuwahara, Anti-
bacterial carbohydrate monoesters suppressing cell growth of Strepto-
coccus mutansin the presence of sucrose, Curr. Microbiol. 41 (2000) 210e
213.
a
-D
galactofuranosides for mycobacterial glycosyltransferase activity, Bioorg.
Med. Chem. 10 (2002) 923e928.
[32] F.V.M. Silva, M. Goulart, J. Justino, A. Neves, F. Santos, J. Caio, S. Lucas,
A. Newton, D. Sacoto, E. Barbosa, M.-S. Santos, A.P. Rauter, Alkyl deoxy-ara-
bino-hexopyranosides: synthesis, surface properties, and biological activities,
Bioorg. Med. Chem. 16 (2008) 4083e4092.
[45] P. Bault, P. Godé, G. Goethals, J.W. Goodby, J.A. Haley, S.M. Kelly, G.H. Mehl,
G. Ronco, P. Villa, Liquid crystalline derivatives of galactose and galactitol:
dependence of thermotropic mesomorphism on carbohydrate form, Liq. Cryst.
25 (1998) 31e45.
[33] P. Nobmann, P. Bourke, J. Dunne, G. Henehan, In vitro antimicrobial activity
and mechanism of action of novel carbohydrate fatty acid derivatives against
Staphylococcus aureus and MRSA, J. Appl. Microbiol. 108 (2010) 2152e2161.
[34] L. Legentil, J.L. Audic, R. Daniellou, C. Nugier-Chauvin, V. Ferrières, Studies of
a furanoside as antimycobacterial agent loaded into a biodegradable PBAT/
sodium caseinate support, Carbohydr. Res. 346 (2011) 1541e1545.
[35] H.C. Neu, The crisis in antibiotic resistance, Science 257 (1992) 1064e1073.
[36] J.L. Martinez, A. Fajardo, L. Garmendia, A. Hernandez, J.F. Linares, L. Martínez-
Solano, M.B. Sánchez, A global view of antibiotic resistance, FEMS Microbiol.
Rev. 33 (2009) 44e65.
[37] B. Ouattara, R.E. Simard, R.A. Holley, G.J.-P. Piette, A. Bégin, Antibacterial
activity of selected fatty acids and essential oils against six meat spoilage
organisms, Int. J. Food Microbiol. 37 (1997) 155e162.
[38] G. Bergsson, J. Arnfinnsson, O. Steingriacute;msson, H. Thormar, Killing of
gram-positive cocci by fatty acids and monoglycerides, APMIS 109 (2001)
670e678.
[46] P. Bault, P. Godé, G. Goethals, J.W. Goodby, J.A. Haley, S.M. Kelly, G.H. Mehl,
G. Ronco, P. Villa, An homologous series of 6-O-n-alkyl-alpha-D-galactopyra-
noses: synthesis and thermotropic mesomorphic properties, Liq. Cryst. 24
(1998) 283e293.
[47] M.P. Savelli, P. Van Roeckeghem, O. Douillet, G. Cavé, P. Godé, G. Ronco,
P. Villa, Effect of tail alkyl chain length (n), head group structure and junction
(Z) on amphiphilic properties of 1-Z-R-D, L-xylitol compounds, Int. J. Pharm.
182 (1999) 221e236.
[48] G. Goethals, A. Fernàndez, P. Martin, M. Minana-Pérez, C. Scorza, P. Villa,
P. Godé, Spacer arm influence on glucidoamphiphile compound properties,
Carbohydr. Polym. 45 (2001) 147e154.
[49] P. Bault, P. Godé, G. Goethals, J.W. Goodby, J.A. Haley, S.M. Kelly, G.H. Mehl,
P. Villa, Synthesis and mesomorphism of 6-Z-n-alkyl-
Liq. Cryst. 26 (1999) 985e997.
a-D-galactopyranoses,
[50] L. Di Marzio, C. Marianecci, M. Petrone, F. Rinaldi, M. Carafa, Novel pH-
sensitive non-ionic surfactant vesicles: comparison between Tween 21 and
Tween 20, Colloids Surf. B Biointerfaces 82 (2011) 18e24.
[51] A.D. Russell, Mechanisms of bacterial resistance to non-antibiotics: food
additives and food and pharmaceutical preservatives, J. Appl. Bacteriol. 71
(1991) 191e201.
[39] M.K. Nair, H. Abouelezz, T. Hoagland, K. Venkitanarayanan, Antibacterial effect
of monocaprylin on Escherichia coli O157:H7 in apple juice, J. Food Prot. 68
(2005) 1895e1899.
[40] T. Kitahara, Y. Aoyama, Y. Hirakata, S. Kamihira, S. Kohno, N. Ichikawa,
M. Nakashima, H. Sasaki, S. Higuchi, In vitro activity of lauric acid or myristyl-
amine incombination withsixantimicrobial agentsagainstmethicillin-resistant
Staphylococcus aureus (MRSA), Int. J. Antimicrob. Agents 27 (2006) 51e57.
[41] A. Kollanoor, P. Vasudevan, M.K.M. Nair, T. Hoagland, K. Venkitanarayanan,
Inactivation of bacterial fish pathogens by medium-chain lipid molecules
(caprylic acid, monocaprylin and sodium caprylate), Aquacult. Res. 38 (2007)
1293e1300.
[52] T.J. Silhavy, D. Kahne, S. Walker, The bacterial cell envelope, Cold Spring Harb.
Perspect. Biol. 2 (2010) a000414.
[53] A.F. Haag, S. Wehmeier, A. Muszynski, B. Kerscher, V. Fletcher, S.H. Berry,
G.L. Hold, R.W. Carlson, G.P. Ferguson, Biochemical characterization of Sino-
rhizobium meliloti mutants reveals gene products involved in the biosynthesis
of the unusual lipid A very long-chain fatty acid, J. Biol. Chem. 286 (2011)
17455e17466.
[42] H.I. Jang, M.S. Rhee, Inhibitory effect of caprylic acid and mild heat on Cro-
nobacter spp. (Enterobacter sakazakii) in reconstituted infant formula and
determination of injury by flow cytometry, Int. J. Food Microbiol. 133 (2009)
113e120.
[43] T. Nakatsuji, M.C. Kao, J.-Y. Fang, C.C. Zouboulis, L. Zhang, R.L. Gallo,
C.-M. Huang, Antimicrobial property of lauric acid against
[54] R.M. Epand, R.F. Epand, Domains in bacterial membranes and the action of
antimicrobial agents, Mol. Biosyst. 5 (2009) 580e587.
[55] J. de Bony, A. Lopez, M. Gilleron, M. Welby, G. Lanéelle, B. Rousseau,
J.P. Beaucourt, J.F. Tocanne, Transverse and lateral distribution of phospho-
lipids and glycolipids in the membrane of the bacterium Micrococcus luteus,
Biochemistry 28 (1989) 3728e3737.