Cyclomaltodextrin glucanotransferase-catalyzed transglycosylation from dextrin to alkanol maltosides

Article abstract of DOI:10.1271/bbb.80295

Maltosides of butanol, octanol, and lauryl alcohol were found for the first time to serve as substrates for cyclomaltodextrin glucanotransferase (CGTase), and glycosyl residue was transfered from dextrin to the substrate affording novel maltosides with 3-

Full text of DOI:10.1271/bbb.80295

Biosci. Biotechnol. Biochem., 72 (11), 3006–3010, 2008
Note
Cyclomaltodextrin Glucanotransferase-Catalyzed Transglycosylation
from Dextrin to Alkanol Maltosides
Haisuo ZHAO,1 Hiroyuki NAITO,1 Yoshimi UEDA,1 Katsuhide OKADA,2
Kenji SADAGANE,1 Minoru IZUMI,1 Shuhei NAKAJIMA,1 and Naomichi BABA
y
1;
1Division of Bioscience, Graduate School of Natural Science and Technology, Okayama University,
Tsushimanaka, Okayama 700-8530, Japan
2Research and Development Center, Hayashibara Biochemical Laboratories, Inc.,
2-3 Shimoishii 1-chome, Okayama 700-0970, Japan
Received May 1, 2008; Accepted July 15, 2008; Online Publication, November 7, 2008
[doi:10.1271/bbb.80295]
Maltosides of butanol, octanol, and lauryl alcohol
were found for the first time to serve as substrates for
cyclomaltodextrin glucanotransferase (CGTase), and
glycosyl residue was transfered from dextrin to the
substrate affording novel maltosides with 3–4 glucose
units.
glycerol, sugars, and flavonoids.11) It has been said,
however, that when the substrate polyols are somewhat
hydrophobic, they are not good substrates as glycosyl
acceptors for the enzyme. In fact, there appears to be no
example in which saccharides bearing highly lipophilic
substituents serve as good substrates for CGTase.
Hayashibara Biochemical Laboratories, Inc., developed
a new CGTase produced by Bacillus stearothermophilus
having improved catalytic activity with broader sub-
strate specificity. This enzyme has been used there
practically to increase the water solubility of hesper-
idine, making it usable as a food additive. Our recent
study demonstrated that sucrose esters of lauric acid (12
carbons) can serve as substrates of CGTase, producing
new glycosyl derivatives having 2–4 glucose units.12)
This study suggests that glycosides of hydrophobic
alcohols may also be able to serve as glycosyl acceptors
for the enzymatic reactions. No example of enzymatic
reactions, however, appears to be reported so far on this
type of substrate. Based on structural similarities to
sucrose esters that have two glycosyl units (glucose and
fructose), alkanol maltosides 3a–3c that also have two
glycosyl units (glucose and glucose) are thought to be
potential glycosyl acceptors of CGTase-catalyzed trans-
glycosidation. Also, they are thought to have aqueous
solubility similar to sucrose esters. Results from the
glycosylation of the alkanol maltosides, as to whether it
occurs or not, should provide useful information on the
reactivity of other related substrates with different sugar
parts and hydrophobic substituents. Thus in our present
study, maltoside of alcohols 3a–3c that were synthesized
according to Scheme 1 were subjected to CGTase-
catalyzed transglycosylation. Anhydrous maltose was
acetylated with acetic anhydride to afford peracetylated
maltose, which was converted to the bromide 2 with
titanium tetrabromide in dry dichloromethane in a
nitrogen atmosphere at room temperature for 5 h. In
Key words: glucanotransferase; CGTase; transglycosi-
dation; surfactant; amphiphile
As non-ionic amphiphiles, glycosides of fatty alcohols
are in wide use in a variety of fields, including the food
technology, cosmetics, and pharmaceutical industries.1)
In the production of these materials, chemical and
enzymatic reactions have been extensively studied to
improve synthetic conditions and yields. Especially in
recent years, the syntheses through biotransformation
have been developed, as exemplified by the employment
of supersaturated substrate solutions,2,3) aqueous-organic
biphasic system,4) enzyme immobilization,5) and by
searches for new enzyme sources.6,7) Physicochemical
properties of non-ionic surfactants, such as water
solubility and critical micelle concentration are largely
dependent on the structure of the sugar part and the
hydrophobic lipid group. The desired structure and
properties of such compounds are of course different in
different practical uses. Most of the approaches to
producing non-ionic surfactants with different properties
have focused on alkylation as well as esterification of
sugars, including mono- and disaccharides with different
fatty acids or different alcohols.8–10) Another approach to
tuning the hydrophilic/lipophilic balance in non-ionic
surfactans is to alter the size-length of the saccharide
chain by adding or reducing the number of sugar units in
the glycosides or glycosyl esters.
CGTase catalyzes the transfer of dextrin units from
cyclodextrin or longer dextrins to polyols, such as
y
To whom correspondence should be addressed. Tel: +81-86-251-8292; Fax: +81-86-251-8388; E-mail: babanaom@cc.okayama-u.ac.jp

CGTase-Catalyzed Glycosylation of Alcohol Maltosides
3007
OH
OAc
O
O
AcO
HO
HO
OAc
OH
NaOCOCH3
Ac2O
TiBr4
AcO
O
OAc
O
OH
CH2Cl2, Ethyl acetate
OH
AcO
O
HO
O
OAc
Br
OH
2 (89%)
1
OH
O
3a: R= -(CH2)3CH3
3b: R= -(CH2)7CH3
HO
HO
1N NaOH-MeOH
CHCl3
AgCO3, ROH
CH2Cl2
OH
O
OH
O
OR
3c: R= -(CH2)11CH3
HO
OH
OR
O
4 (Glc3-Bu, Butyl)
OH
O
5 (Glc4-Bu, Butyl)
6 (Glc3-Oc, Octyl)
7 (Glc4-Oc, Octyl)
8 (Glc3-La, Laulyl)
3a
HO
Dextrin, CGTase
HO
OH
3b
3c
OH
Acetate buffer
(pH 5.7, 0.1 M),
50 °C, 9 h
O
O
n
HO
OH
9 (Glc4-La, Lauryl)
OH
OH
OH
OH
6'''
6''
6'
4'
6
3
4
4'''
4''
O
O
O
2''
HO
HO
2'''
2
5
5'''
2'
5''
3''
OR
5'
1'''
1'
HO
1''
HO
HO
3'''
1
OH
3'
OH
OH
OH
O
O
O
Scheme 1. Synthesis of the Alkyl Maltosides 3a–3c and the CGTase-Catalyzed Gucanoyl Transfer Reaction Affording the Products 4–9.
Numbering of the product glycosides of alcohols is also given here.
this experiment, titanium tetrabromide has been found to
be a good reagent as to product yield in the bromination
of heptaacetyl maltose.13) After purification by silica gel
column, the bromide was submitted to reaction with
alcohols at room temperature for 5 h in the presence of
silver carbonate.13) The products were purified by silica
gel column chromatography, and each acetyl maltoside
was submitted to deacetylation using aqueous sodium
hydroxide. The maltosides 3a–3c thus obtained were
submitted to reaction with CGTase-catalyzed transgly-
cosylation using dextrin (10–15 glucose units) as a
glycosyl donor. The reaction conditions were basically
the same as those applied in the CGTase-catalyzed
glycosylation of mono-lauroyl sucrose.12) The spectral
data for the isolated products are given in Table 1 and
ref. 14. Typically, a solution of octyl maltoside (3b,
240 mg, 0.47 mmol), dextrin (0.74 g, 10–15 units of
glucose), and CGTase solution (0.74 ml, 1500 units/ml)
in a mixture of acetate buffer (6.5 ml, pH 5.6, 0.1 M) and
distilled water (1 ml) was stirred at 55 ꢀC for 24 h. Then,
the enzyme was inactivated with boiling water for 5 min,
followed by filtration and concentration under reduced
pressure. The reaction mixture was analyzed by silica
gel TLC (Fig. 1).
found to be contaminated with unidentified products.
This difficulty in isolation has been due to the overlap of
the reaction products with butyl glycosides and oligo-
glycoside fragments formed by the enzymatic hydrolysis
of the dextrin. CGTase is an enzyme that can catalyse
1
both transglycosidation and hydrolysis. H, 13C NMR
(CD3OD), and ESI-MS spectral data for the fractions at
Rf ¼ 0:29, 0.44 and 0.61 are given in Table 1. Since all
the yields were evaluated based on their isolated products
and complete isolation was not possible due to the partial
overlap with others, spot intensity on TLC plate was not
pallalel with the % yield described in Table 1. For the
fraction at Rf ¼ 0:44 in Fig. 1A, as entry 1 in Table 1
shows, the ratio of the sum of the signal intensity for all
the protons on the sugar rings without the hydroxy
protons and that of the butyl group was 2.33, approx-
imately equal to the value (2.31) calculated from
structure 4. This indicates that a glucose unit was added
to the maltosyl moiety in 3a. Moreover, three 1H signals
were observed at ꢀ 5.13, 5.16 (ꢁ-H, doublet, J ¼ 3:8 Hz,
ꢂ-1,4-linkages between two glucosyl parts) and 4.28 (ꢂ-
H, doublet, J ¼ 7:8 Hz, ꢂ-proton at the anomeric carbon
adjacent to the butoxy group), indicating that there were
three anomeric protons in the product obtained from the
fraction at Rf ¼ 0:44, with an ꢂ-glucosidyl linkage. This
is not incompatible with the general feature that CGTase
catalyzes cleavage or formation of a glucosidic bond in
an ꢂ-1, 4 manner. The 13C NMR (CD3OD) spectrum also
indicated three anomeric signals, at ꢀ 100.4, 101.7, and
Figure 1A shows the TLC for the reaction mixture of
butyl maltoside and dextrin. Several spots appeared, and
the fractions of Rf ¼ 0:29, 0.44 and 0.61 alone were
isolated pure by silica gel chromatography eluted with
CHCl3/CH3OH (3:1). The other eluted fractions were

3008
H. ZHAO et al.
Table 1. 1H, 13C NMR, ESI MS Data and Chemical Yields in the CGTase-Catalyzed Glycosyl Transfer Reaction from Dextrin to Alkanol
Maltosides
Lipooligo-
saccharide
Sug./Lip.ꢁ
found
Sug./Lip.ꢁ
calculated
A.P.Sꢁꢁ
(ꢀ, ppm)
A.C.Sꢁꢁꢁ
(ꢀ, ppm)
m=z
found
m=z (M + H)þ
calculated
Yield
(%)
Entry
1
4
2.3
2.3
5.16
5.13
4.28
103.2
101.7
100.4
578.2
578.3ꢁꢁꢁꢁ
14.1
2
5
3.1
3.1
5.19
5.17
5.14
4.30
103.8
101.5
100.7
100.3
740.4
740.3
22.3
3
4
6
7
1.2
1.7
1.2
1.7
5.12
5.10
4.18
104.4
102.9
101.6
634.3
796.4
634.2
796.4
10.8
17.5
5.10
5.07
5.05
4.18
104.4
102.8
101.3
100.9
5
6
8
9
0.9
1.2
0.8
1.1
5.05
5.04
4.17
104.4
102.9
101.6
690.4
852.6
690.4
852.4
7.1
3.8
5.14
5.11
5.05
4.22
104.4
102.9
101.3
100.0
ꢁNumber of protons on the sugar carbons/Number of protons on lauryl carbon chain
ꢁꢁChemical shift of anomeric proton signals (600 MHz, Varian INOVA UNITY 600)
ꢁꢁꢁChemical shift of anomeric carbon signals (150 MHz, Varian INOVA UNITY 600)
ꢁꢁꢁꢁm=z (M + NH4)þ
0.78
0.72
0.77
0.70
0.77
Glc -La
2
Glc -Oc
2
(substrate)
(substrate)
0.61
0.44
Glc -Bu
2
(substrate)
0.52
Glc -La
3
0.46
0.38
Glc -Oc
Glc -Bu
3
3
0.36
0.29
0.30
0.18
Glc -La
4
Glc -Bu
4
0.25
0.12
Glc -Oc
4
0.13
1C
1B
1A
Fig. 1. TLC of the Reaction Mixture Obtained by CGTase-Catalyzed Glucanoyl Transfer Reaction from Dextrin to Alkyl Maltosides 3a–3c.
TLC conditions: silica gel TLC plate F254 developed withchloroform/methanol/water (65:25:4) detected with 10% sulfuric acid in ethanol.
103.2. ESI-MS (positive mode) of the fraction at Rf ¼
0:44 gave a molecular ion peak at m=z 578.2 as an
ammonium ion, which coincided with m=z 578.3
[M + NH4]þ, calculated from glucosylated maltosyl
butylate 4. Combining all of these spectral data, the
compound at the fraction of Rf ¼ 0:44 should have tri-
glucosylated structure 4. Likewise, the fraction at Rf ¼
0:29 (entry 2) was found to be tetraglucosyl butylate 5,
which was formed by the addition of two glucose units.
The fraction at Rf ¼ 0:61 was found to be the substrate
butyl maltoside 3a itself. Although not all the other
fractions could be isolated pure, the fraction of Rf ¼ 0:77
was considered to be a product that formed by the
CGTase-catalyzed disproportionation of products Glc3-

CGTase-Catalyzed Glycosylation of Alcohol Maltosides
3009
2) Millqvist-Fureby, A., Gill, I. S., and Vulfson, E. N.,
Enzymatic transformations in supersaturated substrate
solutions: I. A general study with glycosidases. Bio-
technol. Bioeng., 60, 190–196 (1998).
3) Millqvist-Fureby, A., MacManus, D. A., Davies, S., and
Vulfson, E. N., Enzymatic transformations in super-
saturated substrate solutions: II. Synthesis of disacchar-
ides via transglycosylation. Biotechnol. Bioeng., 60,
197–203 (1998).
4) Kobayashi, T., Adachi, S., Nakanishi, K., and Matsuno,
R., Synthesis of alkyl glycosides through ꢁ-glucosidase-
catalyzed condensation in an aqueous-organic biphasic
system and estimation of the equilibrium constants for
their formation. J. Mol. Cat. B: Enzymatic, 11, 13–21
(2000).
5) Gargouri, M., Smaali, I., Maugard, T., Legoy, M. D., and
Marzouki, N., Fungus ꢁ-glycosidases: immobilization
and use in alkyl-ꢁ-glycoside synthesis. J. Mol. Cat. B:
Enzymatic, 29, 89–94 (2004).
Bu 4 and Glc4-Bu 5 as well as substrate 3a. The fractions
from the starting point (Rf ¼ 0:0) to that of Rf ¼ 0:13
might have been a mixture of polar butyl glycosides with
more glucose units and/or dextrins, including cyclo-
dextrins that formed from the substrate dextrin also
catalyzed by CGTase.
The reaction mixture obtained from the glycosylation
of octyl maltoside 3b under the same conditions also
gave several spots on TLC, as shown in Fig. 1B. In this
case, too, the fractions at Rf ¼ 0:25, 0.46 and 0.70 were
isolated pure, and the compound at Rf ¼ 0:25 was found
to be octyl tetraglucoside 7, that of Rf ¼ 0:46 to be octyl
triglucoside 6, and that at Rf ¼ 0:70 to be octyl maltoside
(substrate 3b). The other fractions at Rf ¼ 0:12, 0.38 and
0.77 that could not be isolated pure might have been
mixtures of dextrins, possibly including cyclodextrins,
some small dextrins, and a product that formed by
the CGTase-catalyzed disproportionation of products
Glc3-Oc 6 and Glc4-Oc 7 as well as the substrate 3b,
respectively, as described above. A similar result was
observed, as seen in Fig. 1C, for the reaction of lauryl
maltoside 3c. Again, three products, lauryl tetraglucosde
7, triglucoside 6, and substrate 3c, were isolated and
identified. Although the other fractions at Rf ¼ 0:18 and
0.78 could not be identified due to nonhomogeneity, the
first one might have been a mixture similar to those of
Rf ¼ 0:13 in Fig. 1A and 0.12 in Fig. 1B, and that at
Rf ¼ 0:78 might also have been similar to ones in
Fig. 1A and B, as described above.
In the case of stearyl maltoside, with a longer carbon
chain and more hydrophobic, new spots on TLC were
found close to the starting point. The proton NMR
spectrum of the product isolated showed, however, that
the glycosyl moiety was much larger than those of the
stearyl carbon chain. Therefore, it was not possible to
determine correctly the number of glucose units in the
product. Although we do not know what happens in this
reaction, this result does not indicate that stearyl
maltoside can not be a substrate of CGTase.
6) Lirdprapamongkol, K., and Svasti, J., Alkyl glucoside
synthesis using Thai rosewood ꢁ-glucosidase. Biotech-
nol. Lett., 22, 1889–1894 (2000).
7) Park, T.-H., Choi, K.-W., Park, C.-S., Lee, S.-B., Kang,
H.-Y., and Shon, K.-J., Substrate specificity and trans-
glycosylation catalyzed by a thermostable ꢁ-glucosidase
from marine hyperthermophile Thermotoga neapolitana.
Appl. Microbiol. Biotechnol., 69, 411–422 (2005).
8) Gao, C., Mayon, P., MacManus, D. A., and Vulfson,
E. N., Novel enzymatic approach to the synthesis of
flavonoid glycosides and their esters. Biotechnol. Bio-
eng., 71, 235–243 (2001).
9) Plou, F. J., Cruces, M. A., Ferrer, M., Fuentes, G.,
Pastor, E., Bernabe, M., Christensen, M., Comelles, F.,
Parra, J. L., and Ballesteros, A., Enzymatic acylation of
di- and trisaccharides with fatty acids: choosing the
appropriate enzyme, support and solvent. J. Biotechnol.,
96, 55–66 (2002).
10) van Doren, H. A., Smits, E., Pestman, J. M., Engbert,
J. B. F. N., and Kellogg, R. M., Mesogenic sugars. From
aldose to liquid crystals and surfactants. Chem. Soc.
Rev., 29, 183–199 (2000).
11) Kometani, T., Terada, Y., Nishimura, T., Takii, H., and
Okada, S., Synthesis of hesperidin glycosides by cyclo-
dextrin glucanotransferase from an alkalophilic Bacillus
species in alkaline pH and properties of hesperidin
glucosides. Biosci. Biotechnol. Biochem., 58, 1990–1994
(1994).
In summary, the present study indicates for the first
time that CGTase can catalyse glycosyl transfer from
dextrin to maltosides of primary alcohols with 4–12
carbon atoms. This ability of CGTase might potentially
contribute to tuning of hydrophilic/lipophilic balance of
alkyl glycosides affording neutral non-ionic surfactants
with desired properties.
12) Okada, K., Zhao, H., Izumi, M., Nakajima, S., and Baba,
N., Glucosylation of sucrose laurate with cyclodextrin
glucanotransferase. Biosci. Biotechnol. Biochem., 71,
826–829 (2007).
13) Paulsen, H., and Paal, M., Blocksynthesis von O-
Glycopeptiden und anderen T-Antigen Strukturen. Car-
bohydr. Res., 135, 71–84 (1984).
Acknowledgments
We are acknowledged to the laboratory of SC-NMR
and the laboratory of API III mass spectrometry in
Okayama University.
14) The numbering of carbons in glycoside moiety is
described in Scheme 1. Glc2-Bu (3a): 1H NMR ꢀH
(CD3OD): 0.83 (3H, t, J ¼ 7:4 Hz, CH3), 1.31 (2H,
m, CH2CH3), 1.50 (2H, m, CH2CH2CH3), 3.16–3.81
(14H, OCH2, and protons on the carbons of the
maltose), 4.16 (1H, d, J ¼ 8:1 Hz, 1-H), 5.08 (1H, d,
J ¼ 3:9 Hz, 10-H). ESI-MS [M + NH4]þ m=z 416.3.
Calcd. for [M + NH4]þ, 416.2. Glc3-Bu (4): 1H NMR
ꢀH (CD3OD): 0.93 (3H, t, J ¼ 7:6 Hz, CH3), 1.40 (2H,
References and Notes
1) Hill, K., and Rhode, O., Sugar-based surfactants for
consumer products and technical applications. Lipid-
Fett., 101, 25–33 (1999).

3010
H. ZHAO et al.
m, CH2CH3), 1.59 (2H, m, CH2CH2CH3), 3.22–3.39
2 ꢂ CH2), 3.17–4.00 (26H, OCH2, and protons on the
sugar carbons), 4.32 (1H, d, J ¼ 7:8 Hz, 1-H), 5.05 (1H,
d, J ¼ 3:9 Hz, 10-H), 5.07 (1H, d, J ¼ 3:9 Hz, 100-H)
5.10 (1H, d, J ¼ 3:9 Hz, 1000-H). ESI-MS (M + NH4)þ
m=z 796.4. Calcd. for [M + NH4]þ, 796.4. Glc2-La
(3c): 1H NMR ꢀH (CD3OD): 0.89 (3H, t, J ¼ 6:6 Hz,
CH3), 1.29 (14H, m, 7 ꢂ CH2), 1.35 (2H, m, CH2CH3),
1.61 [4H, m, OCH2(CH2)2], 3.19–3.90 (14H, OCH2,
and protons on the sugar carbons), 4.27 (1H, d,
J ¼ 8:2 Hz, 1-H), 5.16 (1H, d, J ¼ 3:7 Hz, 10-H). ESI-
MS [M + NH4]þ m=z 528.5. Calcd. for [M + NH4]þ,
(20H, OCH2, and protons on the sugar carbons), 4.28
(1H, d, J ¼ 7:8, 1-H), 5.13 (1H, d, J ¼ 3:8 Hz, 10-H),
5.16 (1H, d, J ¼ 3:8 Hz, 100-H). ESI-MS [M + NH4]þ
m=z 578.2. Calcd. for [M + NH4]þ, 578.3. Glc4-Bu (5):
1H NMR ꢀH (CD3OD): 0.92 (3H, t, J ¼ 7:6 Hz, CH3),
1.40 (2H, m, CH2CH3), 1.60 (2H, m, CH2CH2CH3),
3.25–3.89 (26H, OCH2, and protons on the sugar
carbons), 4.30 (1H, d, J ¼ 7:8 Hz, 1-H), 5.14 (1H, d,
J ¼ 3:8 Hz, 10-H), 5.17 (1H, d, J ¼ 3:8 Hz, 100-H), 5.19
(1H, d, J ¼ 3:8 Hz, 1000-H). ESI-MS [M + NH4]þ m=z
740.4. Calcd. for [M + NH4]þ, 740.3. Glc2-Oc (3b):
1H NMR ꢀH (CD3OD): 0.81 (3H, t, J ¼ 6:6 Hz, CH3),
1.21 (8H, m, 4 ꢂ CH2), 1.53 (4H, m, 2 ꢂ CH2), 3.05–
3.86 (15H, OCH2, and protons on the sugar carbons),
3.94 (1H, d, J ¼ 7:8 Hz, 1-H), 5.04 (1H, d, J ¼ 3:7 Hz,
10-H). ESI-MS [M + NH4]þ m=z 472.2. Calcd. for
[M + NH4]þ, 472.2. Glc3-Oc (6): 1H NMR ꢀH
(CD3OD): 0.81 (3H, t, J ¼ 6:9 Hz, CH3), 1.20 (8H,
m, 4 ꢂ CH2), 1.52 (4H, m, 2 ꢂ CH2), 3.10–3.64 (20H,
OCH2, and protons on the sugar carbons), 4.18 (1H, d,
1-H), 5.10 (1H, d, J ¼ 3:7 Hz, 10-H), 5.12 (1H, d,
J ¼ 3:7 Hz, 100-H). ESI-MS [M + NH4]þ m=z 634.2.
Calcd. for [M + NH4]þ, 634.3. Glc4-Oc (7): 1H NMR
ꢀH (CD3OD): 0.91 (3H, t, J ¼ 6:9 Hz, CH3), 1.29 (6H,
m, 3 ꢂ CH2), 1.38 (2H, m, CH2CH3), 1.61 (4H, m,
1
528.3. Glc3-La (8): H NMR ꢀH (CD3OD): 0.78 (3H, t,
J ¼ 6:6 Hz, CH3), 1.17 (14H, m, 7 ꢂ CH2), 1.26 (2H,
m, CH2CH3), 1.50 [4H, m, OCH2(CH2)2], 3.11–3.81
(20H, OCH2, and protons on the sugar carbons), 4.17
(1H, d, J ¼ 8:2 Hz, 1-H), 5.04 (1H, d, J ¼ 3:8 Hz, 10-
H), 5.05 (1H, d, J ¼ 3:7 Hz, 100-H). ESI-MS [M +
NH4]þ m=z 690.4. Calcd. for [M + NH4]þ, 690.4.
Glc4-La (9): 1H NMR ꢀH (CD3OD): 0.84 (3H, t,
J ¼ 6:6 Hz, CH3), 1.27 (14H, m, 7 ꢂ CH2), 1.32 (2H,
m, CH2CH3), 1.56 [4H, m, OCH2(CH2)2], 3.19–3.88
(26H, OCH2, and protons on the sugar carbons), 4.22
(1H, d, J ¼ 7:8 Hz, 1-H), 5.05 (1H, d, J ¼ 3:7 Hz, 10-
H), 5.11 (1H, d, J ¼ 3:9 Hz, 100-H), 5.14 (1H, d, J ¼
3:9 Hz, 1000-H). ESI-MS [M + NH4]þ m=z 852.6. Calcd.
for [M + NH4]þ, 852.4.