Preparation of regioselectively feruloylated p-nitrophenyl α-l-arabinofuranosides and β-d-xylopyranosides-convenient substrates for study of feruloyl esterase specificity

Article abstract of DOI:10.1016/j.carres.2010.03.034

p-Nitrophenyl α-l-arabinofuranoside and β-d-xylopyranoside mono-O-ferulates were prepared by 4-O-acetylferuloylation of corresponding enzymatically prepared di-O-acetates followed by deacetylation. An alternative mild acylation catalysed by zinc oxide was tested on xylopyranoside derivatives. The chemoselective methanolysis of the acetyl groups using neutral catalyst dibutyltin oxide at reflux was used as deacetylation method. Under these conditions a significant feruloyl migration was observed mainly on p-nitrophenyl 3-O-feruloyl-β-d-xylopyranoside resulting in low yields of the positional isomers. Investigation of substrate and positional specificity of different types of feruloyl esterases on the presented compounds in enzyme-coupled assays was reported previously.

Full text of DOI:10.1016/j.carres.2010.03.034

Carbohydrate Research 345 (2010) 1094–1098
Contents lists available at ScienceDirect
Carbohydrate Research
journal homepage: www.elsevier.com/locate/carres
Preparation of regioselectively feruloylated p-nitrophenyl
a-L-arabinofuranosides and b-D-xylopyranosides—convenient substrates
for study of feruloyl esterase specificity
*
Mária Mastihubová , Peter Biely
Institute of Chemistry, Center for Glycomics, Slovak Academy of Sciences, Dúbravská cesta 9, 845 38 Bratislava, Slovakia
a r t i c l e i n f o
a b s t r a c t
Article history:
p-Nitrophenyl
a-L-arabinofuranoside and b-D-xylopyranoside mono-O-ferulates were prepared by
Received 11 January 2010
Received in revised form 19 March 2010
Accepted 25 March 2010
Available online 8 April 2010
4-O-acetylferuloylation of corresponding enzymatically prepared di-O-acetates followed by deacetyla-
tion. An alternative mild acylation catalysed by zinc oxide was tested on xylopyranoside derivatives.
The chemoselective methanolysis of the acetyl groups using neutral catalyst dibutyltin oxide at reflux
was used as deacetylation method. Under these conditions a significant feruloyl migration was observed
mainly on p-nitrophenyl 3-O-feruloyl-b-D-xylopyranoside resulting in low yields of the positional iso-
Keywords:
mers. Investigation of substrate and positional specificity of different types of feruloyl esterases on the
presented compounds in enzyme-coupled assays was reported previously.
Ó 2010 Elsevier Ltd. All rights reserved.
p-Nitrophenyl glycoside ferulates
Feruloyl esterase substrates
ZnO acylation
Bu2SnO deacetylation
1. Introduction
investigated, the specificity of FAEs regarding the saccharide moiety
and the position of acylation is not well understood. A few reports
Ferulic acid esterases (FAEs, E.C. 3.1.2.73) are well-defined com-
ponents of microbial plant cell wall-degrading enzyme systems.1–5
As a subclass of the carboxylic acid esterases, they are able to
hydrolyse the ester bond between saccharides and hydroxycin-
namic acids or their dehydrodimers. The classification of FAEs into
four types (A–D) was proposed based on protein sequence identi-
ties and similarities in hydrolytic activity profiles against methyl
esters of partly hydroxylated or methoxylated hydroxycinnamic
acids or their dimers.2 Despite this effort the structure–function
relationship of FAEs is far from being understood. The reason is
the great diversity of esterification of ferulic acid in plants.
indicate that microorganisms produce at least two types of FAEs,
which differ in the affinity for 2-O- and 5-O-feruloylated a-L-arabi-
nofuranoside residues.1,24
Understanding of the role of FAEs in biodegradation of plant cell
walls requires detailed knowledge of their catalytic properties.
Consequently, there is a growing demand for a variety of regiose-
lectively feruloylated p-nitrophenyl glycosides that could serve as
chromogenic substrates for investigation of catalytic properties of
these enzymes. A number of substrates in which ferulic acid was
esterified with chromogenic non-saccharidic alcohol25,26 or with
hydroxyl groups of
L
-arabinofuranoside27–30 have previously been
Ferulic acid [(2E)-3-(4-hydroxy-3-methoxyphenyl)-2-prope-
noic acid] has been found regioselectively attached to polysaccha-
ride backbone of hemicelluloses. In dicotyledonous plants, such as
spinach and sugar beet, the acid is attached to primary C-6 hydrox-
synthesised and used in FAE assays.24,26,27,30,31 Our synthesis of
p-nitrophenyl 2-O- and 5-O-feruloyl
a-L-arabinofuranoside was
published earlier as a preliminary communication.29 These com-
pounds were found to be convenient chromogenic substrates for
determination of activity and differentiation of FAEs according to
yls of 1,4-galactans and to C-2 hydroxyls of 1,5-
monocotyledonous plants and grasses, such as bamboo, sugar cane,
wheat and maize, ferulic acid is esterified by C-4 hydroxyls of -xy-
-arabinose in
L
-arabinans.6,7 In
substrate specificity in an
a-L-arabinofuranosidase-coupled UV-
D
spectrophotometric assay.24
lose in xyloglucans and by primary C-5 hydroxyls of
L
In this full paper we present improved and integral synthesis of
all positional isomers of p-nitrophenyl -arabinofuranoside and
b- -xylopyranoside mono-O-ferulates using enzymatic synthesis
arabinoxylans.8–12
a-L
Although a relationship between the specificity of FAEs in hydro-
lysis of methyl phenylalkanoates13,14 and release of ferulic acid or
their dehydrodimers from plant cell walls saccharides15–23 has been
D
of suitably acetylated starting compounds in combination with
convenient acylations and chemoselective deacetylations under
mild conditions. These products, mimicking the naturally occur-
ring feruloylated pentoses, served as substrates in enzyme-coupled
assays to study the substrate and positional specificity of types A, B
* Corresponding author. Tel.: +421 2 59410246; fax: +421 2 59410222.
E-mail address: chemjama@savba.sk (M. Mastihubová).
0008-6215/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2010.03.034

M. Mastihubová, P. Biely / Carbohydrate Research 345 (2010) 1094–1098
1095
and C FAEs.32 The coupling of the action of FAEs with that of Asper-
gillus niger -arabinofuranosidase or A. niger b- -xylosidase,
respectively, renders feruloylated p-nitrophenyl pentosides to
Table 1
Dibutyltin oxide-catalysed deacetylation
a-L
D
Starting compound
Producta
Migration productsa
NPhGlyca,b
chromogenic substrates of FAEs.
7
8
9
10
11
12
13 (80%)
14 (79%)
15 (85%)
16 (78%)
17 (63%)
18 (84%)
—
—
—
—
—
—
—
19 (12%)
19 (11%)
19 (9%)
20 (10%)
20 (13%)
20 (8%)
2. Results and discussion
17 (6%)
16 (9%)
—
18 (5%)
—
Several different strategies have been published for regioselec-
tive preparation of feruloyl glycosides. The straightforward methods
were described like was the acylation exploiting a slightly higher
reactivity of sterically less hindered primary hydroxyl of methyl
a
Isolated yields, the residues were partially deacetylated starting compounds.
NPhGlyc-4-nitrophenyl glycoside.
b
a-L
-arabinofuranoside27,29 or dibutyltin oxide-mediated one-step
acylation of methyl b-D
-xylopyranoside.33 More complicated pro-
4–6 are solid compounds, we have tested the reaction in dioxane,
acetonitrile and dichloromethane. Using dry dichloromethane
and 0.5 equiv of ZnO, the desired products 10 (79%), 11 (77%)
and 12 (80%) were obtained within 1 h. Formation of positional
isomers due to possible acyl migration was not observed. To our
knowledge, ZnO has not been previously employed in acylations
of carbohydrates.
The next step of our study was the investigation of conditions for
chemoselective deacetylations on 7–12 which would cleave only
acetyl esters and preserve the ferulate ester bond. In addition, the
conditions should prevent acyl migration. The deacetylations with
K2CO3 in CH2Cl2/MeOH (2:1) successfully applied to per-O-acety-
lated 2-O- and 5-O-feruloylated
or pyrrolidine in ethanol previously used by Helm et al.28 failed in
the case of -xylopyranoside derivatives 10–12 due to acyl migra-
tion along the xylopyranoside ring. The migration intermediates
eventually decomposed to 4-nitrophenyl b- -xylopyranoside and
tection–deprotection strategies limited to suitable deprotection of
primary position of per-O-silylated28 or per-O-acetylated29,30
a-
L-arabinofuranosides were developed as well. Diacetates of
p-nitrophenyl -arabinofuranoside 1–3 and p-nitrophenyl b-D-
a
-L
xylopyranoside 4–6 serving as starting compounds in our syntheses
were prepared either by regioselective acetylation of corresponding
glycosides or by deacetylation of their per-O-acetylated derivatives
by commercial immobilised lipases34,35 (Chart 1).
The free hydroxyl groups in di-O-acetates 1–6 were acylated
with 4-O-acetylferuloyl chloride25,27 using DMAP and Et3N to give
4-O-acetylferuloyl glycosides 7–12 in high yields within 3 h. An
L
-arabinofuranosides (7 and 9)29
exception was the acetylferuloylation of 2,4-di-O-acetyl-b-
xylopyranoside 5 which needed more than 6 h of reaction time.
D-
D
Our previous study of stability of acetylated xylopyranosides in
aqueous media revealed that the acetyl moiety easily migrates
along the flexible b-D
-xylopyranoside ring.35 Hence, acylations of
D
ferulic acid. Similarly, attempts to use several mild basic or acidic
xylopyranosides should be executed with carefully selected
reagents and reaction conditions. Zinc oxide (ZnO) as a recently
introduced efficient and eco-friendly acylation catalyst36,37 was
therefore applied in acylations of partially acetylated xylopyrano-
sides 4–6. The previously described heterogeneous reactions using
liquid acylation reagents have been accomplished in solvent-free
conditions.36,37 Since 4-O-acetylferuloyl chloride and glycosides
ways of deacetylation of 2-O-(40-O-acetylferuloyl)-3,4-di-O-acetyl-
b-D-xylopyranoside 10 failed due to unsatisfactory reactivity or che-
moselectivity. In 2002, Liu et al. published a neutral, chemoselective
and efficient methanolysis of O-acetyl groups with dibutyltin oxide
(Bu2SnO) in dry methanol.38 Using this method, deacetylations of
compounds 7–12 with Bu2SnO (2 equiv to glycoside) in MeOH/
CH2Cl2 (9:1) under mild reflux (60 °C in bath, 6 h) afforded the de-
sired ferulates 13–18 in yields 63–85% (Table 1). Minor de-feruloy-
lated products 19 and 20 were also isolated in 8–13%.
Feruloyl migration has been anyway observed in a limited extent
due to the presence of protic solvent and higher reaction tempera-
ture. After deacetylation of 2,4-di-O-acetyl-3-O-(40-O-acetylferu-
ONPh
O
ONPh
O
OR2
OR2
R3O
R3O
OR1
OR1
loyl)-b-D-xylopyranoside 11, products 16 and 18 have been
isolated in 9% and 5%, respectively (Table 1, Scheme 1). Similarly,
3,4-di-O-acetyl-2-O-(40-O-acetylferuloyl) derivative 10 afforded a
small amount (6%) of 3-O-feruloyl product 17. The mono-O-feruloy-
R1
1: H
R2
R3
Ac
Ac
H
R1
4: H
R2
R3
Ac
Ac
H
Ac
H
Ac
H
lated
a-L-arabinofuranosides 13–15 as well as 4-O-feruloyl-b-D-
2: Ac
3: Ac
5: Ac
6: Ac
xylopyranoside 12 showed remarkable stability to feruloyl
migration. This phenomenon is in accordance with our previous
results with partially acetylated 4-nitrophenyl glycosides.34,35 There
Ac
Ac
7: AcFe Ac
Ac
10: AcFe Ac
Ac
is no acetyl group migration along the L-arabinofuranosyl ring,
8: Ac
9: Ac
13: Fe
14: H
15: H
19: H
AcFe Ac
11: Ac
12: Ac
16: Fe
17: H
18: H
20: H
AcFe Ac
neither between 5-O position and other free hydroxyl groups. In
all cases, the deprotection of glycosides with Bu2SnO left the 4-nitro-
phenyl aglycon fully intact.
Ac
H
AcFe
Ac
H
AcFe
H
H
H
Fe
H
Fe
H
Fe
H
H
3. Conclusion
Fe
H
H
H
Synthesis of targeted substrates p-nitrophenyl 2-O-, 3-O- and
5-O-feruloyl-
a-
L-arabinofuranosides 13–15 as well as p-nitro-
phenyl 2-O-, 3-O- and 4-O-feruloyl-b-
D-xylopyranosides 16–18
O
O
was accomplished. The effective mild acylation method of starting
compounds by zinc oxide was compared with conventional ways
using partially acetylated xylopyranosides 4–6 as feruloyl accep-
tors. The neutral chemoselective de-O-acetylation procedure by
dibutyltin oxide has been successfully applied for preparation of
H3CO
H3CO
AcO
HO
AcFe =
Fe =
4’-O-acetylferuloyl
feruloyl
Chart 1.
mono-O-feruloyl-D-glycosides 13–18. The compounds 13–18 have

1096
M. Mastihubová, P. Biely / Carbohydrate Research 345 (2010) 1094–1098
O
HO
ONPh
ONPh
HO
OFe
16
17
O
O
O
a
AcO
AcFeO
HO
FeO
HO
HO
ONPh
ONPh
OAc
OH
OH
20
11
O
FeO
HO
ONPh
OH
18
Scheme 1. Reagents and conditions: (a) Bu2SnO (2 equiv), CH2Cl2/MeOH (1:9), 60 °C, 6 h.
already been used as the substrates for determination of activity
and differentiation of FAEs according to substrate specificity in a
UV-spectrophotometric assay.24,32
(m, 3H, H-4, H-5a, H-5b), 3.88 (s, 3H, OCH3), 2.33 (s, 3H, COCH3),
2.18 (s, 3H, COCH3), 2.12 (s, 3H, COCH3). 13C NMR (CDCl3): d
170.5 (COCH3), 170.0 (COCH3), 168.7 (COCH3), 165.3 (O–C@O),
160.8 (C-10), 151.5 (C-300), 146.4 (C-A), 142.8 (C-40), 142.0 (C-400),
132.7 (C-10), 2 ꢀ 125.8 (C-30, C-50), 123.4, 121.5 (C-600, C-500),
2 ꢀ 116.6 (C-20, C-60), 116.3 (C-B), 111.4 (C-200), 103.9 (C-1), 82.2
(C-3), 81.2 (C-2), 76.8 (C-4), 63.0 (C-5), 55.9 (OCH3), 20.7
(2 ꢀ COCH3), 20.6 (COCH3). Anal. Calcd for C27H27NO13: C, 56.55;
H, 4.75; N, 2.44. Found: C, 56.28; H, 5.02; N, 2.17.
4. Experimental
4.1. General methods
Solvents were dried and distilled before use. The reagent
Bu2SnO (BDH Chemicals) was used as purchased without further
purification. The ZnO powder (Aldrich) was activated for one hour
at 400 °C before use. All reactions were monitored by TLC on Silica
Gel 60 F254 plates (0.25 mm, E. Merck, Darmstadt, Germany). Spots
were detected under UV lamp (kmax = 254 nm) followed by dipping
the plate in 5% ethanolic H2SO4 and heating at ca. 200 °C. Column
chromatography was performed on silica gel (0.035–0.070 mm,
4.2.1.2. p-Nitrophenyl 3-O-(40-O-acetylferuloyl)-2,5-di-O-acetyl-
a-L-arabinofuranoside (8). The title compound 8 (0.538 g, 94%)
was obtained from diacetate 2 as a white foam; ½a D20
ꢂ102 (c 1.0,
ꢁ
1
CHCl3); d H NMR (CDCl3): d 8.22 (d, 2H, J 9.3 Hz, H-30, H-50),
7.71 (d, 1H, J 16.0 Hz, H-A), 7.17 (d, 2H, J 9.3 Hz, H-20, H-60),
7.18–7.07 (m, 3H, H-200, H-500, H-600), 6.43 (d, 1H, H-B), 5.82 (s,
1H, H-1), 5.49 (d, 1H, J2,3 1.4 Hz, H-2), 5.27 (dd, 1H, J3,4 4.4 Hz, H-
3), 4.51–4.46 (m, 2H, H-4, H-5a), 4.34 (dd, 1H, J5a,5b 12.7, J4,5b
6.4 Hz, H-5b), 3.88 (s, 3H, OCH3), 2.33 (s, 3H, COCH3), 2.19 (s, 3H,
COCH3), 2.13 (s, 3H, COCH3). 13C NMR (CDCl3): d 170.5 (COCH3),
169.6 (COCH3), 168.7 (COCH3), 165.8 (O–C@O), 160.8 (C-10),
151.5 (C-300), 146.1 (C-A), 142.8 (C-40), 141.9 (C-400), 132.8 (C-10),
2 ꢀ 125.8 (C-30, C-50), 123.4, 121.5 (C-600, C-500), 3 ꢀ 116.6 (C-20, C-
60, C-B), 111.4 (C-200), 103.9 (C-1), 82.2 (C-4), 81.1 (C-2), 77.2 (C-
3), 63.0 (C-5), 56.0 (OCH3), 20.7 (COCH3), 20.6 (2 ꢀ COCH3). Anal.
Calcd for C27H27NO13: C, 56.55; H, 4.75, N 2.44. Found: C, 56.68;
H, 5.09; N, 2.28.
pore diameter ca. 6 nm, Acros Organics). Melting points were
¯
determined on a Kofler hot-stage and are uncorrected. Optical rota-
tions were measured with a Perkin–Elmer 241 polarimeter at
20 °C. 1H NMR spectra (300 MHz, Me4Si as internal standard) and
13C NMR spectra (75 MHz, shifts referenced to internal solvent)
were recorded with Bruker AM 300. The assignment of resonances
in the 1H and 13C NMR spectra of products was made by two-
dimensional homonuclear and heteronuclear shift correlation
experiments. Microanalyses were performed with a Fisons EA
1108 analyser.
4.2. General procedure for acetylferuloylation of 1–6
4.2.1.3. p-Nitrophenyl 5-O-(40-O-acetylferuloyl)-2,3-di-O-acetyl-
a-L-arabinofuranoside (9). The starting compound 9 (0.498 g,
4.2.1. Procedure A using triethylamine/dimethylaminopyridine
Di-O-acetates of 4-nitrophenyl glycosides 1–634,35 (0.355 g,
1 mmol) and 4-O-acetylferuloyl chloride25,27 (0.306 g, 1.2 mmol)
were dissolved in dry CH2Cl2 (10 ml). Triethylamine (0.139 ml,
1 mmol) and 4-dimethylaminopyridine (0.031 g, 0.25 mmol) were
added at 0 °C. The reaction mixture was then stirred for 3 h at lab-
oratory temperature. Then the mixture was diluted with CH2Cl2
(40 ml), washed with 1% HCl (10 ml), water (2 ꢀ 20 ml), dried over
Na2SO4 and concentrated under reduced pressure. Products 7–12
were isolated by column chromatography of the residues on silica
gel (toluene/EtOAc, 2:1).
87%) was obtained from 3 as a white solid; mp 78–80 °C (EtOH);
½
a 2D0
ꢁ
ꢂ56 (c 1.0, CHCl3); 1H NMR (CDCl3): d 8.21 (d, 2H, J 9.2 Hz,
H-30, H-50), 7.69 (d, 1H, J 16.0 Hz, H-A), 7.16 (d, 2H, J 9.3 Hz, H-20,
H-60), 7.14–7.05 (m, 3H, H-200, H-500, H-600), 6.41 (d, 1H, H-B), 5.81
(s, 1H, H-1), 5.41 (d, 1H, J2,3 1.1 Hz, H-2), 5.21 (br d, 1H, J3,4
3.3 Hz, H-3), 4.59–4.41 (m, 3H, H-4, H-5a, H-5b), 3.86 (s, 3H,
OCH3), 2.32 (s, 3H, COCH3), 2.17 (s, 3H, COCH3), 2.15 (s, 3H, COCH3).
13C NMR (CDCl3): d 170.0 (COCH3), 169.5 (COCH3), 168.7 (COCH3),
166.2 (O–C@O), 160.8 (C-10), 151.4 (C-300), 145.1 (C-A), 142.7 (C-40),
141.7 (C-400), 133.0 (C-10), 2 ꢀ 125.7 (C-30, C-50), 123.3, 121.3 (C-600,
C-500), 117.3 (C-B), 2 ꢀ 116.5 (C-20, C-60), 111.3 (C-200), 103.8 (C-1),
82.2 (C-4), 81.1 (C-2), 76.7 (C-3), 62.9 (C-5), 55.9 (OCH3), 20.7
(2 ꢀ COCH3), 20.6 (COCH3). Anal. Calcd for C27H27NO13: C, 56.55;
H, 4.75; N, 2.44. Found: C, 56.54; H, 4.94; N, 2.22.
4.2.1.1. p-Nitrophenyl 2-O-(40-O-acetylferuloyl)-3,5-di-O-acetyl-
a-L-arabinofuranoside (7). The starting compound 1 gave 7 (0.
516 g, 90%) as white solid crystals; mp 126–128 °C (EtOH); ½a D20
ꢁ
1
ꢂ90 (c 1.0, CHCl3); H NMR (CDCl3): d 8.22 (d, 2H, J 9.2 Hz, H-30,
4.2.1.4. p-Nitrophenyl 2-O-(40-O-acetylferuloyl)-3,4-di-O-acetyl-
H-50), 7.73 (d, 1H, J 16.0 Hz, H-A), 7.17 (d, 2H, J 9.2 Hz, H-20, H-
60), 7.16–7.06 (m, 3H, H-200, H-500, H-600), 6.41 (d, 1H, H-B), 5.88 (s,
1H, H-1), 5.51 (s, 1H, H-2), 5.23 (bd, 1H, J3,4 2.9, H-3), 4.49–4.32
b-D-xylopyranoside (10). The compound 10 (0.517 g, 90%) was
obtained from diacetate 4 as a white solid; mp 114–116 °C (EtOH);

M. Mastihubová, P. Biely / Carbohydrate Research 345 (2010) 1094–1098
1097
½
a 2D0
ꢁ
ꢂ23.0 (c 1.0, CHCl3); 1H NMR (CDCl3): d 8.21 (d, 2H, J 9.2 Hz,
4.3. General procedure for deacetylation of 7–12
H-30, H-50), 7.71 (d, 1H, J 15.9 Hz, H-A), 7.11–7.07 (m, 5H, H-20,
H-60, H-200, H-500, H-600), 6.36 (d, 1H, J 15.9 Hz, H-B), 5.39–5.34 (m,
3H, H-2, H-1, H-3), 5.06 (m, 1H, H-4), 4.27 (dd, 1H, J5a,5b 12.3,
J4,5b 4.3 Hz, H-5a), 3.86 (s, 3H, OCH3), 3.66 (dd, 1H, J4,5b 6.7 Hz, H-
5b), 2.32 (s, 3H, COCH3), 2.11 (s, 3H, COCH3), 2.10 (s, 3H, COCH3).
13C NMR (CDCl3): d 2 ꢀ 169.8 (COCH3), 168.7 (COCH3), 165.0 (O–
C@O), 161.1 (C-10), 151.5 (C-300), 146.1 (C-A), 143.1 (C-40), 141.9
(C-400), 132.7 (C-10), 2 ꢀ 125.8 (C-30, C-50), 123.4, 121.6 (C-600, C-
500), 2 ꢀ 116.6 (C-20, C-60), 116.5 (C-B), 111.3 (C-200), 97.8 (C-1),
69.8, 69.4 (C-2, C-3), 68.2 (C-4), 62.0 (C-5), 55.9 (OCH3), 20.8
The respective di-O-acetyl-mono-O-(40-O-acetylferuloyl)-glyco-
sides 7–12 (0.287 g, 0.5 mmol) were suspended in a dry solution of
MeOH/CH2Cl2 (9:1, 10 ml), followed by addition of Bu2SnO
(0.250 g, 1 mmol) in one portion. The reaction mixture was then
kept under slight reflux (60 °C) for 6 h. The resulting homogeneous
mixture was concentrated under reduced pressure and the product
was isolated by column chromatography using gradient toluene/
EtOAc (2:1?0:1). Completely deacylated 4-nitrophenyl glycosides
were also isolated as minor products. Their physicochemical data
were in accordance with literature.39,40 Moreover, minor migration
products were isolated after deacetylation of 10 and 11.
(COCH3), 20.7 (COCH3), 20.6 (COCH3). Anal. Calcd for C27H27NO13
C, 56.55; H, 4.75; N, 2.44. Found: C, 56.49; H, 5.05; N, 2.07.
:
4.2.1.5. p-Nitrophenyl 3-O-(40-O-acetylferuloyl)-2,4-di-O-acetyl-
b- -xylopyranoside (11). The compound 11 was isolated from
4.3.1. p-Nitrophenyl 2-O-feruloyl-a-L-arabinofuranoside (13)
The compound 13 (0.172 g, 77%) was obtained as a pale yellow
D
solid; mp 81–83 °C (CH2Cl2); ½a D20
ꢁ
ꢂ152 (c 1.0, CH3OH); 1H NMR
acetylferuloylation of diacetate 5 according to the general method
described above. Due to slower process the reaction mixture was
stirred for 6 h at laboratory temperature. The isolation of product
was accomplished by the manner described above. The compound
11 (0. 510 g, 89%) was obtained from diacetate 5 as a white solid;
(CDCl3): d 8.23 (d, 2H, J 9.3 Hz, H-30, H-50), 7.69 (d, 1H, JA,B
15.9 Hz, H-A), 7.16 (d, 2H, J 9.3 Hz, H-20, H-60), 7.10 (dd, 1H, J5 ,6
00 00
8.2, J2 ,6 1.8 Hz, H-600), 7.03 (d, 1H, H-200), 6.94 (d, 1H, H-500), 6.32
00 00
(d, 1H, H-B), 5.93 (d, 1H, J1,2 1.3 Hz, H-1), 5.31 (d, 1H, J2,3 2.1 Hz,
H-2), 4.30 (m, 2H, H-3, H-4), 3.99 (dd, 1H, J5a,5b 12.6, J4,5a 2.4 Hz,
H-5a), 3.95 (s, 3H, OCH3), 3.84 (dd, 1H, J4,5b 3.0 Hz, H-5b). 13C
NMR (CDCl3): d 167.7 (O–C@O), 161.1 (C-10), 148.7, 146.9, 142.6
(C-40, C-400, C-300), 147.3 (C-A), 126.3 (C-100), 2 ꢀ 125.8 (C-30, C-50),
123.6 (C-600), 2 ꢀ 116.5 (C-20, C-60), 114.9 (C-500), 113.3 (C-B),
109.5 (C-200), 103.9 (C-1), 87.2 (C-2), 84.3, 76.2 (C-3, C-4), 61.2
(C-5), 56.0 (OCH3). Anal. Calcd for C21H21NO10: C, 56.38; H, 4.73;
N, 3.13. Found: C, 56.09; H, 5.04; N, 2.82. The minor deacylated
mp 110–112 °C (from EtOH); ½a D20
ꢁ
+14 (c 1.0, CHCl3); 1H NMR
(CDCl3): d 8.23 (d, 2H, J 9.2 Hz, H-30, H-50), 7.70 (d, 1H, J 16.0 Hz,
H-A), 7.16–7.07 (m, 5H, H-20, H-60, H-200, H-500, H-600), 6.38 (d, 1H,
J 16.0 Hz, H-B), 5.43–5.27 (m, 3H, H-2, H-1, H-3), 5.11 (m, 1H, H-
4), 4.28 (dd, 1H, J5a,5b 12.2, J4,5b 4.5 Hz, H-5a), 3.89 (s, 3H, OCH3),
3.66 (dd, 1H, J4,5b 7.0 Hz, H-5b), 2.33 (s, 3H, COCH3), 2.10 (s, 3H,
COCH3), 2.08 (s, 3H, COCH3). 13C NMR (CDCl3): d 169.8 (COCH3),
169.3 (COCH3), 168.7 (COCH3), 165.4 (O–C@O), 161.1 (C-10),
151.5 (C-300), 146.0 (C-A), 143.1 (C-40), 141.9 (C-400), 132.8 (C-10),
2 ꢀ 125.8 (C-30, C-50), 123.4, 121.7 (C-600, C-500), 3 ꢀ 116.6 (C-20, C-
60, C-B), 111.3 (C-200), 97.8 (C-1), 70.1, 69.6 (C-2, C-3), 68.2 (C-4),
62.1 (C-5), 56.0 (OCH3), 20.8 (COCH3), 20.6 (2 ꢀ COCH3). Anal.
Calcd for C27H27NO13: C, 56.55; H, 4.75; N, 2.44. Found: C, 56.81;
H, 5.93 N, 2.60.
product p-nitrophenyl
a-L-arabinofuranoside was also isolated
(0.016 g, 12%).
4.3.2. p-Nitrophenyl 3-O-feruloyl-a-L-arabinofuranoside (14)
The compound 8 afforded product 14 (0.176 g, 79%) as a pale
yellow foam; ½a 2D0
ꢁ
ꢂ174 (c 1.0, CH3OH); 1H NMR (CDCl3): d 8.21
(d, 2H, J 9.2 Hz, H-30, H-50), 7.68 (d, 1H, JA,B 15.9 Hz, H-A), 7.14 (d,
2H, J 9.2 Hz, H-20, H-60), 7.11 (dd, 1H, J5 ,6 8.2, J2 ,6 1.8 Hz, H-600),
7.04 (d, 1H, H-200), 6.95 (d, 1H, H-500), 6.33 (d, 1H, H-B), 5.82 (s,
1H, H-1), 5.12 (dd, 1H, J2,3 1.7, J3,4 4.4 Hz, H-3), 4.53 (d, 1H, H-2),
4.44–4.41 (m, 1H, H-4), 3.99–3.95 (m, 2H, H-5a, H-5b), 3.95 (s,
00 00
00 00
4.2.1.6. p-Nitrophenyl 4-O-(40-O-acetylferuloyl)-2,3-di-O-acetyl-
b-D-xylopyranoside (12). The title compound 12 (0.521 g, 91%),
was obtained from 6 as a white solid; mp 181–183 °C (EtOH);
½
a 2D0
ꢁ
ꢂ52.0 (c 1.0, CHCl3); 1H NMR (CDCl3): d 8.22 (d, 2H, J 9.2 Hz,
13
3H, OCH3). C NMR (CDCl3): d 167.5 (O–C@O), 161.3 (C-10),
H-30, H-50), 7.69 (d, 1H, JA,B 15.9 Hz, H-A), 7.18–7.06 (m, 5H, H-20,
H-60, H-200, H-500, H-600), 6.38 (d, 1H, JA,B 15.9 Hz, H-B), 5.38–5.33
(m, 2H, H-1, H-3), 5.24 (dd, 1H, J2,3 7.3, J1,2 5.5 Hz, H-2), 5.13 (m,
1H, H-4), 4.32 (dd, 1H, J5a,5b 12.2, J4,5a 4.3 Hz, H-5a), 3.88 (s, 3H,
OCH3), 3.68 (dd, 1H, J4,5b 6.9 Hz, H-5b), 2.33 (s, 3H, COCH3), 2.11
(s, 3H, COCH3), 2.10 (s, 3H, COCH3). 13C NMR (CDCl3): d 169.8
(COCH3), 169.3 (COCH3), 168.7 (COCH3), 165.5 (O–C@O), 161.0
(C-10), 151.5 (C-300), 145.9 (C-A), 143.1 (C-40), 141.9 (C-400), 132.8
(C-10), 2 ꢀ 125.8 (C-30, C-50), 123.4, 121.6 (C-600, C-500), 116.7 (C-
B), 2 ꢀ 116.5 (C-20, C-60), 111.3 (C-200), 97.6 (C-1), 69.8, 69.5 (C-2,
C-3), 68.2 (C-4), 61.9 (C-5), 56.0 (OCH3), 20.7 (3 ꢀ COCH3). Anal.
Calcd for C27H27NO13: C, 56.55; H, 4.75; N, 2.44. Found: C, 56.44;
H, 5.09; N, 2.26.
148.6, 146.8, 142.5 (C-40, C-400, C-300), 146.8 (C-A), 126.5 (C-100),
2 ꢀ 125.8 (C-30, C-50), 123.5 (C-600), 2 ꢀ 116.4 (C-20, C-60), 114.9
(C-500), 113.8 (C-B), 109.5 (C-200), 106.4 (C-1), 84.6 (C-4), 79.9 (C-
3), 79.8 (C-2), 61.6 (C-5), 56.0 (OCH3). Anal. Calcd for
C21H21NO10: C, 56.38; H, 4.73; N, 3.13. Found: C, 56.74; H, 4.93;
N, 2.84. The minor deacylated product p-nitrophenyl a-L-arabino-
furanoside was isolated (0.015 g, 11%).
4.3.3. p-Nitrophenyl 5-O-feruloyl-a-L-arabinofuranoside (15)
The compound 15 (0.191 g, 85%) was obtained as a pale yellow
solid; mp 140–142 °C (EtOAc/toluene); ½a D20
ꢁ
ꢂ87 (c 1.0, CH3OH); 1H
NMR (CD3OD): d 8.20 (d, 2H, J 9.2 Hz, H-30, H-50), 7.63 (d, 1H, JA,B
15.9 Hz, H-A), 7.20 (d, 2H, J 9.3 Hz, H-20, H-60), 7.17 (d, 1H, J2 ,6
00 00
1.6 Hz, H-200), 7.05 (dd, 1H, J5 ,6 8.2, J2 ,6 1.7 Hz, H-600), 6.79 (d,
1H, H-500), 6.38 (d, 1H, H-B), 5.70 (d, 1H, J1,2 1.5 Hz, H-1), 4.44
(dd, 1H, J5a,5b 11.7, J4,5a 3.2 Hz, H-5a), 4.35–4.23 (m, 3H, H-2, H-4,
H-5b), 4.06 (dd, 1H, J2,3 3.9, J3,4 6.2 Hz, H-3), 3.87 (s, 3H, OCH3).
13C NMR (CD3OD): d 168.9 (O–C@O), 163.3 (C-10), 150.8, 149.4,
143.7 (C-40, C-400, C-300), 147.4 (C-A), 127.7 (C-100), 2 ꢀ 126.7 (C-30,
C-50), 124.3 (C-600), 2 ꢀ 117.7 (C-20, C-60), 116.5 (C-500), 115.1 (C-
B), 111.7 (C-200), 107.8 (C-1), 84.2 (C-4), 83.7 (C-3), 78.9 (C-2),
64.8 (C-5), 56.5 (OCH3). Anal. Calcd for C21H21NO10: C, 56.38; H,
4.73; N, 3.13. Found: C, 56.24; H, 5.02; N, 2.93. Minor p-nitro-
00 00
00 00
4.2.2. Procedure B using zinc oxide
Di-O-acetates 4–6 (0.1725 g, 0.5 mmol) and 4-O-acetylferuloyl
chloride (0.153 g, 0.6 mmol) were dissolved in dry CH2Cl2 (2 ml).
Zinc oxide (0.02 g) was added in one portion. The heterogeneous
mixture was then stirred for 1 h at laboratory temperature. Then
the reaction mixture was diluted with CH2Cl2 (10 ml) and filtered.
After concentration under reduced pressure, the residues were
purified by column chromatography on silica gel (toluene/EtOAc,
3.5:1) to afford 10 (0.226 g, 79%), 11 (0. 221 g, 77%) and 12
(0.229 g, 80%) as white solids. The physicochemical data were
identical to those of compounds 10–12 described above.
phenyl
a-L-arabinofuranoside (0.012 g, 9%) as a product of total
deacylation was also isolated.

1098
M. Mastihubová, P. Biely / Carbohydrate Research 345 (2010) 1094–1098
4.3.4. p-Nitrophenyl 2-O-feruloyl-b-D-xylopyranoside (16)
Acknowledgements
The compound 10 afforded product 16 (0.174 g, 78%) as a pale
yellow solid; mp 170–171 °C (EtOAc/cyclohexane); ½a D20
ꢂ31.0 (c
ꢁ
This work was supported by the Slovak Grant Agency for
Science VEGA No. 2/0145/08 and the Slovak State Programme
Project No. 2003 SP 200280203.
1
1.0, CH3OH); H NMR (CD3OD): d 8.18 (d, 2H, J 9.3 Hz, H-30, H-50),
7.66 (d, 1H, JA,B 15.9 Hz, H-A), 7.17 (d, 1H, J2 ,6 1.8 Hz, H-200), 7.14
00 00
(d, 2H, J 9.3 Hz, H-20, H-60), 7.06 (dd, 1H, J5 ,6 8.2, J2 ,6 1.9 Hz, H-
600), 6.79 (d, 1H, H-500), 6.39 (d, 1H, H-B), 5.31 (d, 1H, J1,2 7.5 Hz,
H-1), 5.11 (dd, 1H, J2,3 9.0, J1,2 7.6 Hz, H-2), 4.03 (dd, 1H, J5a,5b
11.3, J4,5a 4.3 Hz, H-5a), 3.72–3.68 (m, 2H, H-3, H-4), 3.86 (s, 3H,
OCH3), 3.52 (dd, 1H, J4,5b 9.8 Hz, H-5b). 13C NMR (CD3OD): d
168.2 (O–C@O), 163.3 (C-10), 150.8, 149.4, 144.2 (C-40, C-400, C-300),
147.6 (C-A), 127.7 (C-100), 2 ꢀ 126.7 (C-30, C-50), 124.3 (C-600),
2 ꢀ 117.7 (C-20, C-60), 116.5 (C-500), 115.1 (C-B), 111.8 (C-200),
100.3 (C-1), 74.5 (C-2), 75.7, 70.9 (C-3, C-4), 67.1 (C-5), 56.5
(OCH3). Anal. Calcd for C21H21NO10: C, 56.38; H, 4.73; N, 3.13.
Found: C, 56.33; H, 4.74; N, 3.13. The migration product 17
00 00
00 00
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4.3.5. p-Nitrophenyl 3-O-feruloyl-b-D-xylopyranoside (17)
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ꢁ
+67.0 (c 1.0, CH3OH);
1H NMR (CD3OD): d 8.22 (d, 2H, J 9.2 Hz, H-30, H-50), 7.69 (d, 1H, JA,B
15.9 Hz, H-A), 7.22 (d, 2H, J 9.2 Hz, H-20, H-60), 7.21 (d, 1H, J2 ,6
00 00
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Williamson, G. J. Sci. Food Agric. 1999, 79, 428–434.
1.8 Hz, H-200), 7.10 (dd, 1H, J5 ,6 8.2, J2 ,6 1.8 Hz, H-600), 6.82 (d,
1H, H-500), 6.45 (d, 1H, H-B), 5.19 (d, 1H, J1,2 7.3 Hz, H-1), 5.12
(dd, 1H, J2,3 9.0, J3,4 9.0 Hz, H-3), 4.03 (dd, 1H, J5a,5b 11.4, J4,5a
5.2 Hz, H-5a), 3.90 (s, 3H, OCH3), 3.83 (m, 1H, H-4), 3.72 (dd, 1H,
H-2), 3.58 (dd, 1H, J4,5b 10.0 Hz, H-5b). 13C NMR (CD3OD): d 168.9
(O–C@O), 163.6 (C-10), 150.7, 149.5, 144.1 (C-40, C-400, C-300), 147.1
(C-A), 127.9 (C-100), 2 ꢀ 126.7 (C-30, C-50), 124.1 (C-600), 2 ꢀ 117.7
(C-20, C-60), 116.6 (C-500), 115.7 (C-B), 111.9 (C-200), 102.0 (C-1),
78.1 (C-3), 72.7 (C-2), 69.3 (C-4), 66.9 (C-5), 56.5 (OCH3). Anal.
Calcd for C21H21NO10: C, 56.38; H, 4.73; N, 3.13. Found: C, 56.09;
H, 5.00; N, 2.87. The migration products 16 (0.020 g, 9%) and 18
00 00
00 00
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´
25. Mastihubová, M.; Mastihuba, V.; Kremnicky, L.; Willet, J. L.; Côté, G. L. Synlett
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4.3.6. p-Nitrophenyl 4-O-feruloyl-b-
D-xylopyranoside (18)
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pale yellow solid; mp 184–186 °C (EtOAc/cyclohexane);
½ ꢁ
a 2D0
ꢂ7.0 (c 1.0, CH3OH); 1H NMR (CD3OD): d 8.22 (d, 2H, J 9.2 Hz, H-
30, H-50), 7.68 (d, 1H, JA,B 15.9 Hz, H-A), 7.23 (d, 2H, J 9.3 Hz, H-20,
H-60), 7.20 (d, 1H, J2 ,6 1.6 Hz, H-200), 7.09 (dd, 1H, J5 ,6 8.2, J2 ,6
00 00
00 00
00 00
1.8 Hz, H-600), 6.82 (d, 1H, H-500), 6.40 (d, 1H, H-B), 5.13 (d, 1H,
´
J1,2 7.3 Hz, H-1), 4.93–4.84 (m, 1H, H-4), 4.13 (dd, 1H, J
11.5,
5a,5b
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7844.
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231–244.
J4,5a 5.3 Hz, H-5a), 3.90 (s, 3H, OCH3), 3.79 (dd, 1H, J2,3 9.0, J3,4
9.0 Hz, H-3), 3.62 (dd, 1H, H-2), 3.57 (dd, 1H, J4,5b 9.7 Hz, H-5b).
13C NMR (CD3OD): d 168.5 (O–C@O), 163.7 (C-10), 150.9, 149.5,
144.1 (C-40, C-400, C-300), 147.6 (C-A), 127.7 (C-100), 2 ꢀ 126.7 (C-
30, C-50), 124.3 (C-600), 2 ꢀ 117.7 (C-20, C-60), 116.6 (C-500), 115.0
(C-B), 111.9 (C-200), 102.1 (C-1), 74.7, 74.6 (C-2, C-3), 72.8 (C-4),
64.0 (C-5), 56.5 (OCH3). Anal. Calcd for C21H21NO10: C, 56.38; H,
4.73; N, 3.13. Found: C, 55.99; H, 5.11; N, 2.93. The deacylated
product p-nitrophenyl b-D-xylopyranoside (0.011 g, 8%) was also
isolated.