Structural characterization of a series of 10-carbon sugar derivatives by electrospray-ionization MSn mass spectrometry

Article abstract of DOI:10.1016/j.carres.2005.06.032

Electrospray-ionization MSⁿ mass spectrometry (ESI-MS ⁿ) with low-energy, collision-induced dissociation (CID) was used to establish the fragmentation behavior of sodium ion adducts of higher-carbon amino spiro-sugar derivatives. Their fragmentation pathways are proposed on the basis of the MSⁿ studies and deuteration experiments. Some of the rings of these derivatives opened under the conditions of electrospray ionization. Novel fragmentations were observed and their mechanisms are proposed. This study demonstrates the power of modern mass spectrometry for rapid elucidation of the structure of higher-carbon sugar derivatives.

Full text of DOI:10.1016/j.carres.2005.06.032

Carbohydrate
RESEARCH
Carbohydrate Research 340 (2005) 2411–2421
Structural characterization of a series of 10-carbon sugar
derivatives by electrospray-ionization MSn mass spectrometry
Da-Peng Zou, Shu-Xia Cao, Wei-Chao Xu and Hong-Min Liu*
Department of Chemistry, Zhengzhou University, Zhengzhou 450052, PR China
Received 2 February 2005; accepted 20 June 2005
Available online 26 August 2005
Abstract—Electrospray-ionization MSn mass spectrometry (ESI-MSn) with low-energy, collision-induced dissociation (CID) was
used to establish the fragmentation behavior of sodium ion adducts of higher-carbon amino spiro-sugar derivatives. Their fragmen-
tation pathways are proposed on the basis of the MSn studies and deuteration experiments. Some of the rings of these derivatives
opened under the conditions of electrospray ionization. Novel fragmentations were observed and their mechanisms are proposed.
This study demonstrates the power of modern mass spectrometry for rapid elucidation of the structure of higher-carbon sugar
derivatives.
ꢀ 2005 Elsevier Ltd. All rights reserved.
Keywords: Higher-carbon sugars; Amino spiro-sugars; Structural characterization; Electrospray ionization; MSn mass spectrometry
1. Introduction
important. Electrospray ionization (ESI), a soft ioniza-
tion technique, is a powerful tool for structure elucida-
tion of nonvolatile and thermally labile compounds,
especially biological materials9–12 and carbohydrate
derivatives.13–17 Many investigations concerning metal-
ion adducts of carbohydrates in ESI-MS experiments
have been reported recently,18–20 but to the best of our
knowledge, the ESI-MS fragmentation characteristics
of higher-carbon amino spiro-sugar derivatives have
not been studied. Here, we report the ESI-MSn spectra
of a series of such compounds (4a–h, 5a–h, 6a–f, and
7a–f in Scheme 1) to study their fragmentation behavior
and establish characteristic fragments suitable for the
identification of similar carbohydrates.
Higher-carbon sugar derivatives having backbones long-
er than the usual five or six carbon atoms are an impor-
tant class of carbohydrates. The presence of such units
in natural products confers unique biological properties.
Several of these 7–11 carbon carbohydrates play prom-
inent roles in biological processes.1,2 Thus, the higher-
carbon sugars are interesting synthetic targets, and
several approaches towards their synthesis have been
reported.3–7 In our previous work, an efficient and
economical synthetic method for C10 higher-carbon
sugars from D-xylose was reported.8 Hence, we employ
substituted anilines in C10 higher-carbon sugars to
alter their stereochemistry and search structure–
activity relationships, and some higher-carbon amino
spiro-sugars useful as chiral synthons were therefore
synthesized. Some of these have glycosidase-inhibitor
activities.
2. Results and discussion
Under the experimental conditions, it was found that the
sodium ion adducts were the base peaks in the ESI-MS
spectra of all compounds in Scheme 1. The sodium ion
was probably derived from glass containers and as an
impurity picked up during the preparation of the sample.
Figure 1a shows the ESI mass spectrum of a methanol
For structural elucidation of these compounds, stud-
ies on their ESI-MS fragmentation characteristics are
*
Corresponding author. Tel./fax: +86 371 7767200; e-mail:
liuhm@zzu.edu.cn
0008-6215/$ - see front matter ꢀ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2005.06.032

2412
D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
12
11
O
O
X
O
O
13
O
O
O
O
Y
R
O
O
NH2
O 10
NaBH4
O
O
O
9
O
7
4
O
8
5
6
1
O
O
3
N
2
HN
O
O
O
15
O
14
16
R
R
1
4a-4h
5a-5h
O
O
O
O
O
O
O
O
O
O
O
O
R
O
O
O
O
O
NH2
O
NaBH4
PDC, Ac2O
O
O
O
O
O
O
O
O
N
HN
O
O
HO
O
R
R
2
3
7a-7f
6a-6f
a R = p-CH3; b R = o-CH3; c R = p-F; d R = p-Cl; e R = p-Br; f R = p-OCH3; g R = H; h R = m-CH3
Scheme 1. Structures of compounds 1–7.
2.1. MSn spectra analysis of compounds 4a–h
Intens.
x106
a
b
c
d
+MS
452.1
1.0
The compounds 4a–h obviously followed a common
dissociation pathway. Studies by MSn mass spectrometry
showed losses of 58 and 46 Da from the precursor
[M+Na]+ ion. For example, the mass spectrum of com-
pound 4a showed the base peak [M+Na]+ at m/z 452.
In the ESI-MS2 spectrum of compound 4a (Fig. 1b), the
[M+Na]+ ion of the most abundant fragment-ion ap-
peared at m/z 394, obviously formed by the loss of
58 Da, the molecular mass of acetone. According to the
structure of compound 4a (Scheme 1), there were two pos-
sibilities to yield the fragment ion at m/z 394, namely by
the loss of a molecule of acetone, either from the 1,2-O-
or the 9,10-O-isopropylidene group of the molecule. In
addition to the ion at m/z 394, a weak ion peak at m/z
406 in the ESI-MS2 spectrum was also observed. This sig-
nal can be rationalized in terms of an [M+NaÀ
H2OÀCO]+ ion. We presumed that the presence of the
double bond in the C-7–C-8 position may cause the X
and Y rings (Scheme 1) to be unstable under the experi-
mental conditions and they would cleave to an open-
chain structure. Therefore, we proposed that the
[M+NaÀH2OÀCO]+ ion was probably formed by elimi-
nation of carbon monoxide and water simultaneously
after the X and Y rings were opened, as shown in Scheme
2. In addition, the presence of the C@N double bond
would stabilize the [M+NaÀH2OÀCO]+ fragment ion.
The ESI-MS3 spectrum of the [M+NaÀH2OÀCO]+ ion
shows the elimination of 58 Da to produce the single
ion corresponding to loss of a molecule of acetone. Fur-
thermore, the ESI-MS3 spectrum of the [M+NaÀCH3-
COCH3]+ ion at m/z 394 shows the formation of an
394.1
394.1
372.1
0.5
314.0
244.0
336.1
336.1
484.1
+MS2(452.0)
0.0
x104
4
2
452.1
406.1
376.1
282.0
0
2000
+MS3(452.0->406.0)
+MS3(452.0->394.0)
348.0
1500
1000
500
0
406.1
366.1
318.0
376.1
1000
500
0
336.0
348.0
394.1
278.0
250
300
350
400
450
500
m/z
Figure 1. (a) ESI-MS spectrum of a methanol solution of compound
4a; (b) MS2 spectrum of the [M+Na]+ (m/z 452) of compound 4a; (c)
MS3 spectrum of the m/z 406 of compound 4a; (d) MS3 spectrum of
the m/z 394 of compound 4a.
solution of (1aS,2aR,4S,7E,9aR,10aR)-1,2:9,10-di-O-
isopropylidene-3-(p-toluidino)-3,5,6-tri-deoxy-dec-7-enos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (4a) without
any other additive. Like all other compounds studied
here, no other significant molecular ions beside the base
peak, which was the sodium ion adduct, appeared in the
spectrum except for the solvated molecular ion
[M+CH3OH+Na]+. Therefore, the [M+Na]+ ions were
studied in detail to investigate some new general rules
for the interpretation of the fragment ion spectra of high-
er-carbon amino spiro-sugar derivatives.

D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
2413
O
O
O
O
O
O
O
N
O
+Na+
O
+Na+
O
+Na+
O
O
+Na+
+Na+
O
O
OH
O
O
O
-CH3COCH3
-CH3COCH3
O
O
O
O
O
O
N
N
N
-
N
O
O
4a
H3C
[M+Na-58-58-58]+
[M+Na-46]+
m/z 406
[M+Na-46-58]+
m/z 348
[M+Na-58-58]+
m/z 336
[M+Na]+
H3C
H3C
H3C
H3C
m/z 278
m/z 452
O
O
O
O
+Na+
+Na+
O
O
O
O
O
+Na+
O
O
O
+Na+
O
+Na+
O
O
O
OH
OH
OH
O
-H2O
O
-CH3COCH3
O
O
N
N
O
O
N
O
N
N
O
4a
[M+Na]+
m/z 452
[M+Na-58]+
m/z 394
H3C
[M+Na-58-76]+
H3C
H3C
[M+Na-58-18]+
[M+Na-58-28]+
H3C
H3C
m/z 376
m/z 366
m/z 318
-CO
-CH3COCH3-H2O
Scheme 2. Proposed fragmentation pathway of compound 4 during electrospray MSn mass spectrometry with CID.
[M+NaÀCH3-COCH3ÀH2OÀCO]+ ion at m/z 348 as
shown in Figure 1d. Simultaneously, the loss of 18, 28,
58, or 76 Da is also observed in the ESI-MS3 spectrum
of m/z 394. The major fragmentation pathways for com-
pound 4 can be summarized and depicted as shown in
Scheme 2, using compound 4a as a representative
example.
ion [M+NaÀCH3COCH3]+ at m/z 396 was derived
from the [M+Na]+ ion of compound 5a by the elimina-
tion of a molecule of acetone. In the ESI-MS3 spectra,
the [M+NaÀCH3COCH3]+ ion then lost 18, 58, and
76 Da, successively. Furthermore, the ESI-MS4 spec-
trum of the [M+NaÀ2CH3COCH3]+ ion at m/z 338
showed the fragment ions at m/z 320 and 310.
In order to confirm the fragmentation pathways and
to investigate the mechanism of formation of the
[M+NaÀ58À76]+ ion in the spectra of compound 5,
the deuterated analogues of compounds 4 and 5 under
the same experimental conditions were studied in detail.
As shown in Figure 2a, the MS2 spectrum of the deute-
rium-exchanged [M+Na]+ (m/z 452) ion of compound
4a was the same as that in Figure 1b, because there
was no active hydrogen in the structure of compound
4. In the CID spectra of deuterated analogues of com-
pound 5a (Fig. 2b), the [M+Na]+ ion produced the most
abundant fragment ion at m/z 397 by the elimination of
a molecule of acetone. Figure 2c and d shows the ESI-
MS3 and ESI-MS4 spectra of deuterated analogues of
compound 5a, respectively. The ESI-MS3 spectrum of
the [M+NaÀCH3COCH3]+ ion at m/z 397 showed elim-
ination of 58 and 19 Da fragments, corresponding to the
loss of a molecule of acetone and of water (DHO).
Simultaneously, the elimination of 18 Da was also
2.2. MSn spectra analysis of compounds 5a–h
Compound 5 has active hydrogen at the nitrogen atom
in its structure, thus differing from compound 4. In the
MS2 spectra of the [M+Na]+ ion of compound 5, the
base peak was the peak corresponding to the ion
[M+NaÀCH3COCH3]+, as with compound 4, whereas,
the fragment ion [M+NaÀH2OÀCO]+ was not found in
the MS2 spectra. After elimination of a molecule of ace-
tone from the precursor ion, the ESI-MS3 spectra of
[M+NaÀCH3COCH3]+ ion showed the sequential elim-
ination of 58 and 18 Da, corresponding to loss of a mole-
cule of acetone and water, respectively. For example, the
first stage MS of (1aS,2aR,3aS,4S,7E,9aR,10aR)-
1,2:9,10-di-O-isopropylidene-3-(p-toluidino)-3,5,6-tri-
deoxy-dec-7-enos-4-ulose 1,4:7,10-difuranose-4,8-pyra-
nose (5a) showed the base peak [M+Na]+ at m/z 454.
In the ESI-MS2 spectrum, the most abundant fragment

2414
D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
Intens
x105
pathways for the dehydration process, and the
+MS2(452.0)
a
b
c
d
exchangeable hydrogen at the nitrogen atom partici-
pated in one of the dehydration processes of compound
5a. Therefore, as shown in Scheme 3, the deuterated
[M+NaÀCH3COCH3]+ ion at m/z 397 lost a molecule
of deuterium-exchanged water (DHO) and of water
(H2O) to form the [M+NaÀCH3COCH3ÀDHO]+ ion
at m/z 378 and the [M+NaÀCH3COCH3ÀH2O]+ ion
at m/z 379, respectively. In Scheme 3, we propose a frag-
mentation mechanism for compound 5 using compound
5a as a representative example. The values in parenthe-
ses are the observed m/z values of the corresponding
peaks in the CID spectra of deuterium-exchanged mole-
cules. This is also applicable in all the other schemes
depicting fragmentation of the sodium adduct ions.
The corresponding product ions observed for deute-
rium-exchanged compounds also match the suggested
formation pathways (Schemes 2 and 3).
1.5
1.0
0.5
393.7
451.7
405.7
347.6 365.6
0.0
x105
+MS2(455.0)
1.0
396.7
396.6
0.5
454.7
377.7
377.6
338.6
0.0
8000
+MS3(455.0->397.0)
6000
4000
2000
0
338.5
319.5
320.5
302.3
+MS4(455.0->397.0->339.0
310.6
200
100
0
338.6
292.6
302.3
250
275
300
325
350
375
400
425
450
475 m/z
Figure 2. (a) MS2 spectrum of the [M+Na]+ (m/z 452) ion of
compound 4a after treatment with deuterated methanol; (b) MS2
spectrum of the [M+Na]+ (m/z 455) ion of compound 5a after
treatment with deuterated methanol; (c) MS3 spectrum of m/z 397 of
the deuterated analogue of 5a; (d) MS4 spectrum of the m/z 339 ion of
the deuterated analogue of 5a.
2.3. MSn spectra analysis of compounds 6a–f
The structure of compound 6 differs from compound 4
by the absence of a double bond in the X ring. As
expected, there was almost no [M+NaÀH2OÀCO]+
ion in the CID spectrum of compound 6, because cleav-
age of the X and Y rings to an open-chain structure is
not as favorable as that in the compound 4. The MS3
observed in the ESI-MS3 spectrum of the [M+NaÀCH3-
COCH3]+ ion. This indicated that there might be two
+Na+
O
O
+Na+
O
+Na+
O
O
+Na+
O
O
O
O
+Na+
O
O
O
O
O
O
N
O
N
O
O
or
or
O
O
HN
O
O
O
O
HN
HN
O
CH3
CH3
H3C
5a
H3C
H3C
[M+Na]+
[M+Na-58-58-18]+
m/z 320(321)
[M+Na-58-58-18]+
m/z 320(320)
[M+Na-58-18]+
m/z 378(378)
[M+Na-58-18]+
m/z 378(379)
m/z 454(455)
-H2O
O
O
O
+Na+
O
+Na+
O
+Na+
O
O
+Na+
+Na+
O
O
O
O
OH
O
OH
OH
O
OH
-CO
O
O
-CH3COCH3
OH
-CO
O
O
O
O
O
HN
O
HN
HN
O
HN
HN
5a
[M+Na]+
m/z 454(455)
H3C
[M+Na-58-58-28]+
m/z 310(311)
[M+Na-58-28]+
m/z 368(369)
[M+Na-58-58]+
m/z 338(339)
H3C
H3C
[M+Na-58]+
m/z 396(397)
H3C
H3C
-CH3COCH3
Scheme 3. Proposed fragmentation pathway of compound 5 during electrospray MSn mass spectrometry with CID.

D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
2415
Intens.
x104
so simple as the spectra of compound 6. The main frag-
mentation pathway of the [M+Na]+ ion is still the
sequential elimination of 58 Da to form the [M+NaÀ
CH3COCH3]+ and [M+NaÀ2CH3COCH3]+ ions. In
addition to the loss of 58 Da, the losses of 76, 100, and
116 Da were also observed in the ESI-MS2 spectrum of
the [M+Na]+ ion. From the ESI-MS3 spectrum of the
[M+NaÀCH3COCH3]+ ion, it can be deduced that a
[M+NaÀCH3COCH3ÀH2O]+ ion and a [M+NaÀ
2CH3COCH3]+ ion were formed by elimination of 18
and 58 Da from the [M+NaÀCH3COCH3]+ ion. For-
mation of the [M+NaÀ100]+ ion may be rationalized
by the presence of active hydrogen at the nitrogen atom.
The deuterated analogues of compound 7 under the same
experimental conditions were also investigated in detail.
For example, Figure 4c showed the MS2 spectrum of
deuterium-exchanged [M+Na]+ (m/z 473) of (1aS,2aR,
3aS,4S,7aS,8bR,9aR,10aR)-1,2:9,10-di-O-isopropylid-
ene-3-(p-methoxyanilino)-3,5,6-trideoxydecos-4-ulose
+MS2(454.0)
a
b
6
395.8
4
2
453.9
337.7
337.6
0
4000
+MS3(454.0->396.0)
3000
2000
1000
0
395.7
200
250
300
350
400
450
m /z
Figure 3. (a) MS2 spectrum of the [M+Na]+ ion (m/z 454) of
compound 6a; (b) MS3 spectrum of the m/z 396 ion of compound 6a.
spectra of [M+NaÀCH3COCH3]+ of compound 6 was
considerably simpler than that of compounds 4 and 5.
This could be explained by the absence of a double bond
in the X ring and the absence of active hydrogen at the
nitrogen atom. The presence of a double bond in the X
ring would result in an intense peak corresponding to
the open-chain structure, and the active hydrogen at
the nitrogen atom would result in the formation of
the [M+NaÀCH3COCH3ÀH2O]+ and [M+NaÀCH3-
COCH3ÀCO]+ ions. For example, the [M+Na]+ ion
of (1aS,2aR,4S,7aS,8bR,9aR,10aR)-1,2:9,10-di-O-iso-
propylidene-3-(p-toluidino)-3,5,6-trideoxy-dec-4-uloal-
dose 1,4:7,10-difuranose-4,8-pyranose (6a) yielded the
most abundant fragment ion at m/z 396 by the loss of
a molecule of acetone, as shown in Figure 3a. Further-
more, Figure 3b shows the ESI-MS3 spectrum of m/z
396, and the single product ion is [M+NaÀ2CH3-
COCH3]+ ion at m/z 338. The fragmentation mechanism
for compound 6 is shown in Scheme 4 with compound
6a as a representative example.
IIntens.
x104
+MS2(472.0)
a
413.8
2
1
0
372.3
355.7
471.9
395.7
395.8
+MS3(472.0->414.0)
b
355.6
2000
413.8
309.5
395.7
232.3
0
x104
6
+MS2(473.0)
c
d
395.7
371.7
356.7
4
2
414.8
396.7
372.7
472.9
0
6000
395.7
+MS3(473.0->415.0)
396.7
4000
2000
356.6
350
414.7
310.5
300
221.4
0
2.4. MSn spectra analysis of compounds 7a–f
200
250
400
450
m/z
Figure 4. (a) MS2 spectrum of the [M+Na]+ (m/z 472) ion of
compound 7f; (b) MS3 spectrum of the m/z 414 ion of compound 7f;
(c) MS2 spectrum of the deuterium-exchanged [M+Na]+ (m/z 473) ion
of compound 7f; (d) MS3 spectrum of the m/z 415 ion of deuterium-
exchanged compound 7f.
Compound 7 does not contain a double bond in the X
ring, but there is an active hydrogen at the nitrogen
atom, as in the structure of compound 5. Hence, the
MSn spectra of the [M+Na]+ ion of compound 7 is not
O
O
O
O
O
+Na+
O
O
+Na+
O
+Na+
O
O
O
O
O
O
-CH3COCH3
-CH3COCH3
O
O
N
N
N
O
O
6a
[M+Na]+
m/z 454
[M+Na-58]+
m/z 396
[M+Na-58-58]+
m/z 338
H3C
H3C
H3C
Scheme 4. Proposed fragmentation pathway of compound 6 during electrospray MSn mass spectrometry with CID.

2416
D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
1,4:7,10-difuranose-4,8-pyranose (7f). The most abun-
dant fragment ion at m/z 415 was formed by the elimina-
tion of acetone, as in Figure 4a. Because of the presence
of both the product ion [M+NaÀ100]+ at m/z 373 and
the [M+NaÀ101]+ at m/z 372 in the MS2 spectrum of
the deuterium-exchanged [M+Na]+ (m/z 473) ion of
compound 7f, we could presume that there are two path-
ways to yield the [M+NaÀ100]+ ion by the loss of a mole-
cule of 1,2-O-isopropylideneethenediol. One pathway in-
volves the active hydrogen and the other does not, as
shown in Scheme 5. After loss of acetone, the
[M+NaÀCH3COCH3]+ ion at m/z 414 also had two
pathways for elimination of a water molecule. Therefore,
in the MS3 spectrum of the m/z 415 ion of deuterium-
exchanged compound 7f, there were two ions at m/z
396 and at m/z 397 as shown in Figure 4d. The major
fragmentation pathways for compound 7 are summa-
rized and depicted as shown in Scheme 5. Compound
7f is an example.
tra were recorded on a Bio-Rad FTS-40 instrument using
KBr disks in the 400–4000 cmÀ1 region. The 1H and 13
C
NMR spectra were acquired using a Bruker AVANCE
DPX-400 spectrometer with chemical shifts (d) given in
parts per million relative to Me4Si as the internal stan-
dard. Liquid secondary-ion mass spectrometry spectra
were taken with a ZAB-2SE double-focusing mass spec-
trometer (VG Analytical, Manchester, UK).
3.1.1. General procedure for the preparation of com-
pounds 4a–h.21 A solution of (1aS,2aS,4S,7E,9aR,10-
aR)-5,6-dideoxy-1,2:9,10-di-O-isopropylidene-dec-7-enos-
3,4-diulose 1,4:7,10-difuranose-4,8-pyranose (1) (340 mg,
1.0 mmol) and a substituted aniline (1.1 mmol) in benz-
ene (10 mL) was stirred at rt for 48 h in the presence of
p-toluenesulfonic acid (3 mg). The mixture was filtered
through a short silica-gel column and the solvents evap-
orated. The residue was crystallized from MeOH to af-
ford 4a–h as white needles.
3.1.1.1. (1aS,2aR,4S,7E,9aR,10aR)-1,2:9,10-Di-O-
isopropylidene-3-(p-toluidino)-3,5,6-trideoxy-dec-7-enos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (4a). Obtained
from 1 (340 mg, 1.0 mmol); 236 mg (55%); mp 154–
3. Experimental
3.1. Chemicals
1
156 ꢁC; IR (KBr); m 1504 and 1380 cmÀ1 (Ph); H and
Melting points were determined on a WC-1 melting-
point apparatus and are uncorrected. Elemental analyses
were carried out on a MOD 1106 analyzer. Infrared spec-
13C NMR data were in agreement with those
published;22 Anal. Calcd for C23H27NO7: C, 64.32; H,
6.34; N, 3.26. Found: C, 64.23; H, 6.19; N, 3.25.
O
O
O
HO
O
+Na+
+Na+
+Na+
+Na+
O
O
HO
O
O
HO
OH
O
OH
O
O
O
O
O
HN
or
O
O
HN
N
O
HN
H3CO
[M+Na-100]+
m/z 372(373)
7f
H3CO
H3CO
H3CO
[M+Na]+
[M+Na-100]+
[M+Na-58-58]+
m/z 472(473)
m/z 372(372)
m/z 356(357)
O
O
O
O
O
O
+Na+
OH
O
O
+Na+
+Na+
+Na+
O
O
HO
HO
O
OH
OH
N
O
O
O
-CH3COCH3
-H2O
or
O
O
O
O
HN
HN
O
HN
O
OCH3
[M+Na-58-18]+
m/z 396(396)
7f
[M+Na]+
H3CO
[M+Na-58]+
H3CO
[M+Na-58-18]+
H3CO
m/z 472(473)
m/z 414(415)
m/z 396(397)
Scheme 5. Proposed fragmentation pathway of compound 7 during electrospray MSn mass spectrometry with CID.

D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
2417
3.1.1.2. (1aS,2aR,4S,7E,9aR,10aR)-1,2:9,10-Di-O-iso-
propylidene-3-(p-toluidino)-3,5,6-trideoxy-dec-7-enos-4-
ulose 1,4:7,10-difuranose-4,8-pyranose (4b). Obtained
from 1 (340 mg, 1.0 mmol); 219 mg (51%); mp 154–
lished;22 Anal. Calcd for C22H25NO7: C, 63.60; H,
6.07; N, 3.37. Found: C, 63.77; H, 6.20; N, 3.28.
3.1.1.8. (1aS,2aR,4S,7E,9aR,10aR)-1,2:9,10-Di-O-
isopropylidene-3-(m-toluidino)-3,5,6-trideoxy-dec-7-enos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (4h). Obtained
from 1 (340 mg, 1.0 mmol); 287 mg (67%); mp 125–
1
155 ꢁC; IR (KBr); m 1505 and 1379 cmÀ1(Ph); H and
13C NMR data were in agreement with those pub-
lished;22 Anal. Calcd for C23H27NO7: C, 64.32; H,
6.34; N, 3.26. Found: C, 64.13; H, 6.14; N, 3.15.
1
126 ꢁC; IR (KBr); m 1592 and 1381 cmÀ1 (Ph); H and
13C NMR data were in agreement with those pub-
lished;22 Anal. Calcd for C23H27NO7: C, 64.32; H,
6.34; N, 3.26. Found: C, 64.21; H, 6.19; N, 3.22.
3.1.1.3. (1aS,2aR,4S,7E,9aR,10aR)-3-(4-Fluoroani-
lino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-7-
enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (4c). Ob-
tained from 1 (340 mg, 1.0 mmol); 303 mg (70%); mp
3.1.2. General procedure for the preparation of compound
5a–h. A solution of 4a–h (0.1 mmol) and NaBH4 (11.0
mg, 0.29 mmol) in EtOH (5 mL) was stirred at rt for 3 h.
After evaporation of the solvent under reduced pressure,
the residue was dissolved in water (5 mL), extracted with
EtOAc (3 · 10 mL), dried (Na2SO4), and the solvents
evaporated. After recrystallization from MeOH, the
products 5a–h were obtained as colorless needles.
1
138–140 ꢁC; IR (KBr); m 1503 and 1380 cmÀ1 (Ph); H
and 13C NMR data were in agreement with those pub-
lished;22 Anal. Calcd for C22H24FNO7: C, 60.96; H,
5.58; N, 3.23. Found: C, 60.85; H, 5.45; N, 3.14.
3.1.1.4. (1aS,2aR,4S,7E,9aR,10aR)-3-(4-Chloroani-
lino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-7-
enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (4d). Ob-
tained from 1 (340 mg, 1.0 mmol); 301 mg (67%); mp
3.1.2.1. (1aS,2aR,3aS,4S,7E,9aR,10aR)-1,2:9,10-Di-
O-isopropylidene-3-(p-toluidino)-3,5,6-trideoxy-dec-7-enos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (5a). Obtained
from 4a (43 mg, 0.1 mmol); 40 mg (93%); mp 182–
184 ꢁC; IR (KBr); m 3369 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.90 (d, 2H, J 8.4 Hz, H-
arom), 6.73 (d, 2H, H-arom), 5.92 (d, 1H, J10,9 5.2 Hz,
H-10), 5.83 (d, 1H, J1,2 3.2 Hz, H-1), 5.21 (m, 2H, H-
9), 5.18 (d, 1H, NH), 4.83 (dd, 1H, J2,3 5.2 Hz, H-2),
4.25 (dd, 1H, J3,NH 9.6 Hz, H-3), 2.15 (s, 3H, CH3),
2.20–2.05 (m, 3H, H-5b, 6a, 6b), 1.70–1.60 (m, 1H, H-
5a), 1.50 (s, 3H, CH3), 1.40 (s, 3H, CH3), 1.37 (s, 3H,
CH3), 1.31 (s, 3H, CH3); Anal. Calcd for C23H29NO7:
C, 64.02; H, 6.77; N, 3.25. Found: C, 64.23; H, 6.79;
N, 3.25.
1
152–153 ꢁC; IR (KBr); m 1481 and 1382 cmÀ1 (Ph); H
and 13C NMR data were in agreement with those pub-
lished;22 Anal. Calcd for C22H24ClNO7: C, 58.73; H,
5.38; N, 3.11. Found: C, 58.65; H, 5.41; N, 3.22.
3.1.1.5. (1aS,2aR,4S,7E,9aR,10aR)-3-(4-Bromoani-
lino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-7-
enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (4e). Ob-
tained from 1 (340 mg, 1.0 mmol); 296 mg (60%); mp
1
159–160 ꢁC; IR (KBr); m 1478 and 1382 cmÀ1 (Ph); H
and 13C NMR data were in agreement with those pub-
lished;22 Anal. Calcd for C22H24BrNO7: C, 53.45; H,
4.89; N, 2.83. Found: C, 53.36; H, 4.91; N, 2.94.
3.1.1.6. (1aS,2aR,4S,7E,9aR,10aR)-1,2:9,10-Di-O-iso-
propylidene-3-(4-methoxyanilino)-3,5,6-trideoxy-dec-7-
enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (4f). Ob-
tained from 1 (340 mg, 1.0 mmol); 320 mg (72%); mp 58–
3.1.2.2. (1aS,2aR,3aS,4S,7E,9aR,10aR)-1,2:9,10-Di-
O-isopropylidene-3-(o-toluidino)-3,5,6-trideoxy-dec-7-enos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (5b). Obtained
from 4b (43 mg, 0.1 mmol); 41 mg (95%); mp 86–88 ꢁC;
IR (KBr); m 3443 cmÀ1 (NH); 1H NMR (400 MHz,
Me2SO-d6): d 7.04 (m, 2H, H-arom), 6.70 (d, 1H, J
8.0 Hz, H-arom), 6.60 (t, 1H, H-arom), 5.94 (d, 1H,
J10,9 5.6 Hz, H-10), 5.87 (d, 1H, J1,2 3.6 Hz, H-1), 5.25
(m, 1H, H-9), 4.96 (m, 1H, H-2), 4.31 (m, 2H, H-3,
NH), 2.20–2.05 (m, 3H, H-5b, 6a, 6b), 2.12 (s, 3H,
CH3), 1.70–1.60 (m, 1H, H-5a), 1.53 (s, 3H, CH3),
1.41 (s, 3H, CH3), 1.38 (s, 3H, CH3), 1.36 (s, 3H,
CH3); Anal. Calcd for C23H29NO7: C, 64.02; H, 6.77;
N, 3.25. Found: C, 64.15; H, 6.82; N, 3.28.
1
60 ꢁC; IR (KBr); m 1507 and 1380 cmÀ1 (Ph); H NMR
(400 MHz, CDCl3): d 7.21 (d, 2H, J 8.8 Hz, H-arom),
6.92 (d, 2H, H-arom), 6.02 (d, 1H, J1,2 3.6 Hz, H-1),
5.94 (d, 1H, J10,9 5.2 Hz, H-10), 5.30 (m, 1H, H-9),
4.80 (d, 1H, H-2), 3.83 (s, 3H, OCH3), 2.51–2.50 (m,
1H, H-6b), 2.45–2.30 (m, 2H, H-5b, 6a), 2.15–2.05 (m,
1H, H-5a), 1.56 (s, 6H, CH3), 1.48 (s, 3H, CH3), 1.41
(s, 3H, CH3); Anal. Calcd for C23H27NO8: C, 62.01; H,
6.11; N, 3.14. Found: C, 62.17; H, 6.21; N, 3.38.
3.1.1.7. (1aS,2aR,4S,7E,9aR,10aR)-3-Anilino-1,2:9,10-
di-O-isopropylidene-3,5,6-trideoxy-dec-7-enos-4-ulose 1,4:
7,10-difuranose-4,8-pyranose (4g). Obtained from 1
(340 mg, 1.0 mmol); 224 mg (54%); mp 158–160 ꢁC
3.1.2.3. (1aS,2aR,3aS,4S,7E,9aR,10aR)-3-(4-Fluoro-
anilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-7-
enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (5c). Ob-
tained from 4c (43 mg, 0.1 mmol); 41 mg (95%); mp
1
(dec); IR (KBr); m 1589 and 1380 cmÀ1 (Ph); H and
13C NMR data were in agreement with those pub-

2418
D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
186–188 ꢁC; IR (KBr); m 3384 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.93 (m, 2H, H-arom), 6.82
(m, 2H, H-arom), 5.92 (d, 1H, J10,9 5.2 Hz, H-10),
5.84 (d, 1H, J1,2 3.2 Hz, H-1), 5.45 (d, 1H, NH), 5.21
(m, 1H, H-9), 4.82 (dd, 1H, J2,3 5.2 Hz, H-2), 4.25 (dd,
1H, J3,NH 9.6 Hz, H-3), 2.20–2.05 (m, 3H, H-5b, 6a,
6b), 1.70–1.60 (m, 1H, H-5a), 1.51 (s, 3H, CH3), 1.40
(s, 3H, CH3), 1.37 (s, 3H, CH3), 1.31 (s, 3H, CH3); Anal.
Calcd for C22H26FNO7: C, 60.68; H, 6.02; N, 3.22.
Found: C, 60.78; H, 6.11; N, 3.28.
3.1.2.7. (1aS,2aR,3aS,4S,7E,9aR,10aR)-3-Anilino-
1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-7-enos-4-
ulose 1,4:7,10-difuranose-4,8-pyranose (5g). Obtained
from 4g (42 mg, 0.1 mmol); 40 (94%); mp 174–176 ꢁC;
IR (KBr); m 3384 cmÀ1 (NH); 1H NMR (400 MHz,
Me2SO-d6): d 7.09 (t, 2H, J 7.6 Hz, H-arom), 6.82 (t,
2H, H-arom), 6.59 (t, 1H, H-arom), 5.92 (d, 1H, J10,9
5.2 Hz, H-10), 5.84 (d, 1H, J1,2 3.2 Hz, H-1), 5.40
(d, 1H, NH), 5.22 (m, 1H, H-9), 4.84 (dd, 1H, J2,3
5.2 Hz, H-2), 4.29 (dd, 1H, J3,NH 9.2 Hz, H-3), 2.20–
2.05 (m, 3H, H-5a, 6a, 6b), 1.70–1.60 (m, 1H, H-5a),
1.51 (s, 3H, CH3), 1.40 (s, 3H, CH3), 1.38 (s, 3H,
CH3), 1.32 (s, 3H, CH3); Anal. Calcd for C22H27NO7:
C, 63.30; H, 6.52; N, 3.36. Found: C, 63.51; H, 6.49;
N, 3.25.
3.1.2.4. (1aS,2aR,3aS,4S,7E,9aR,10aR)-3-(4-Chloro-
anilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-
7-enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (5d). Ob-
tained from 4d (45 mg, 0.1 mmol); 41 mg (91%); mp
183–185 ꢁC (dec); IR (KBr); m 3390 cmÀ1 (NH); 1H
NMR (400 MHz, Me2SO-d6): d 7.11 (d, 2H, J 8.8 Hz,
H-arom), 6.85 (d, 2H, H-arom), 5.92 (d, 1H, J10,9
5.2 Hz, H-10), 5.85 (d, 1H, J1,2 3.2 Hz, H-1), 5.72 (d,
1H, NH), 5.21 (m, 1H, H-9), 4.83 (dd, 1H, J2,3 5.2 Hz,
H-2), 4.28 (dd, 1H, J3,NH 9.2 Hz, H-3), 2.20–2.05 (m,
3H, H-5b, 6a, 6b), 1.65–1.55 (m, 1H, H-5a), 1.51 (s,
3H, CH3), 1.40 (s, 3H, CH3), 1.37 (s, 3H, CH3), 1.31
(s, 3H, CH3); Anal. Calcd for C22H26ClNO7: C, 58.47;
H, 5.80; N, 3.10. Found: C, 58.52; H, 5.91; N, 3.20.
3.1.2.8. (1aS,2aR,3aS,4S,7E,9aR,10aR)-1,2:9,10-Di-
O-isopropylidene-3-(m-toluidino)-3,5,6-trideoxy-dec-7-
enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (5h). Ob-
tained from 4h (43 mg, 0.1 mmol); 39 mg (90%); mp
146–148 ꢁC; IR (KBr); m 3381 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.97 (t, 1H, J 8.0 Hz, H-arom),
6.66 (s, 1H, H-arom), 6.62 (d, 1H, H-arom), 6.42 (d, 1H,
H-arom), 5.92 (d, 1H, J10,9 5.2 Hz, H-10), 5.84 (d, 1H,
J1,2 3.2 Hz, H-1), 5.28 (d, 1H, NH), 5.21 (m, 1H, H-
9), 4.82 (dd, 1H, J2,3 5.6 Hz, H-2), 4.29 (dd, 1H, J3,NH
9.6 Hz, H-3), 2.18 (s, 3H, CH3), 2.20–2.05 (m, 3H,
H-5b, 6a, 6b), 1.60–1.50 (m, 1H, H-5a), 1.51 (s,
3H, CH3), 1.40 (s, 3H, CH3), 1.37 (s, 3H, CH3),
1.31 (s, 3H, CH3); Anal. Calcd for C23H29NO7: C,
64.02; H, 6.77; N, 3.25. Found: C, 64.21; H, 6.79; N,
3.27.
3.1.2.5. (1aS,2aR,3aS,4S,7E,9aR,10aR)-3-(4-Bromo-
anilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-dec-
7-enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (5e). Ob-
tained from 4e (50 mg, 0.1 mmol); 48 mg (96%); mp
175–176 ꢁC; IR (KBr); m 3404 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 7.22 (d, 2H, J 8.8 Hz, H-
arom), 6.81 (d, 2H, H-arom), 5.92 (d, 1H, J10,9 4.4 Hz,
H-10), 5.84 (d, 1H, J1,2 3.2 Hz, H-1), 5.73 (d, 1H,
NH), 5.21 (m, 1H, H-9), 4.82 (dd, 1H, J2,3 5.2 Hz, H-
2), 4.28 (dd, 1H, J3,NH 9.2 Hz, H-3), 2.20–2.05 (m, 3H,
H-5b, 6a, 6b), 1.65–1.55 (m, 1H, H-5a), 1.51 (s, 3H,
CH3), 1.50 (s, 3H, CH3), 1.37 (s, 3H, CH3), 1.31 (s,
3H, CH3); Anal. Calcd for C22H26BrNO7: C, 53.24; H,
5.28; N, 2.82. Found: C, 53.41; H, 5.22; N, 2.85.
3.1.3.
(1aS,2aS,4S,7aS,8bR,9aR,10aR)-5,6-Dideoxy-
1,2:9,10-di-O-isopropylidene-decosdialdos-3,4-diulose 1,4:
7,10-difuranose-4,8-pyranose (3). A solution of (1aS,2aR,
3aS,4S,7aS,8bR,9aR,10aR)-5,6-dideoxy-1,2:9,10-di-O-iso-
propylidene-decosdialdos-4-ulose 1,4:7,10-difuranose-4,8-
pyranose (2)8 (1.0 g, 2.9 mmol) and PDC (pyridinium
dichromate) (0.8 g, 2.0 mmol) in CH3CN (8 mL) was
stirred at 80 ꢁC for 6 h in the presence of Ac2O
(0.9 mL). After evaporation of the solvent under reduced
pressure, the residue was dissolved in EtOAc (30 mL),
filtered through a short silica-gel column, washed with
satd aq NaHCO3 (3 · 20 mL) and the solvents evapo-
rated. Recrystallization from Et2O yielded the product
3.1.2.6. (1aS,2aR,3aS,4S,7E,9aR,10aR)-1,2:9,10-Di-
O-isopropylidene-3-(4-methoxyanilino)-3,5,6-trideoxy-dec-
7-enos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (5f). Ob-
tained from 4f (45 mg, 0.1 mmol); 43 mg (95%); mp 155–
156 ꢁC; IR (KBr); m 3405 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.77 (d, 2H, J 9.2 Hz, H-
arom), 6.71 (d, 2H, H-arom), 5.92 (d, 1 H J10,9 5.2 Hz,
H-10), 5.83 (d, 1H, J1,2 3.2 Hz, H-1), 5.21 (m, 1H, H-
9), 5.07 (d, 1H, NH), 4.82 (dd, 1H, J2,3 4.8 Hz, H-2),
4.20 (dd, 1H, J3,NH 9.2 Hz, H-3), 3.64 (s, 3H, OCH3),
2.20–2.10 (m, 3H, H-5b, 6a, 6b), 1.70–1.60 (m, 1H, H-
5a), 1.50 (s, 3H, CH3), 1.40 (s, 3H, CH3), 1.37 (s, 3H,
CH3), 1.31 (s, 3H, CH3); Anal. Calcd for C22H29NO8:
C, 61.73; H, 6.53; N, 3.13. Found: C, 61.95; H, 6.59;
N, 3.21.
3
as white needles (0.9 g, 90%); mp 128–130 ꢁC;
IR (KBr); m 1778 cmÀ1 (C@O); 1H NMR (400 MHz,
CDCl3): d 6.19 (d, 1H, J1,2 4.0 Hz, H-1), 5.71 (d, 1H,
H-10), 4.74 (dd, 1H, J9,10 4.8 Hz, H-9), 4.65 (d, 1H, H-
2), 3.96 (dd, J8,7 4.8, J8,9 5.2 Hz, 1H, H-8), 3.88 (m, 1H,
H-7), 2.37 (m, 1H, H-6b), 2.13 (m, 2H, H-5b, 6a),
1.85 (m, 1H, H-5a), 1.65 (s, 3H, CH3), 1.47 (s, 3H,
CH3), 1.45 (s, 3H, CH3), 1.42 (s, 3H, CH3); Anal.
Calcd for C16H22O8: C, 56.13; H, 6.48. Found: C, 56.35;
H, 6.62.

D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
2419
3.1.4. General procedure for the preparation of compound
6a–f. A solution of (1aS,2aS,4S,7aS,8bR,9aR,10aR)-
5,6-dideoxy-1,2:9,10-di-O-isopropylidene-decosdialdos-
3,4-diulose 1,4:7,10-difuranose-4,8-pyranose (3) (342
mg, 1.0 mmol) and substituted aniline (1.1 mmol) in
EtOH (10 mL) was stirred at 70 ꢁC for 48 h. After evap-
oration of the solvent under reduced pressure, the resi-
due was recrystallized from EtOH and afforded 6a–f as
white needles.
3.1.4.4. (1aS,2aR,4S,7aS,8bR,9aR,10aR)-3-(4-Chloro-
anilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxydecos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (6d). Obtained
from 3 (342 mg, 1.0 mmol); 284 mg (63%); mp 192–
194 ꢁC; IR (KBr); m 1503 and 1379 cmÀ1 (Ph); 1H
NMR (400 MHz, Me2SO-d6): d 7.43 (d, 2H, J 8.8 Hz,
H-arom), 7.09 (d, 2H, H-arom), 5.96 (d, 1H, J1,2
4.0 Hz, H-1), 5.64 (d, 1H, H-10), 5.23 (d, 1H, H-2),
4.75 (t, 1H, J9,10 5.6 Hz, H-9), 4.20 (dd, J8,7 4.4, J8,9
5.6 Hz, H-8), 3.89 (m, 1H, H-7), 2.40–2.30 (m, 1H, H-
6b), 2.00–1.80 (m, 3H, H-5a, 5b, 6a), 1.53 (s, 3H,
CH3), 1.36 (s, 3H, CH3), 1.31 (s, 3H, CH3), 1.25 (s,
3H, CH3); Anal. Calcd for C22H26ClNO7: C, 58.47; H,
5.80; N, 3.10. Found: C, 58.35; H, 6.01; N, 3.15.
3.1.4.1. (1aS,2aR,4S,7aS,8bR,9aR,10aR)-1,2:9,10-
Di-O-isopropylidene-3-(p-toluidino)-3,5,6-trideoxydecos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (6a). Obtained
from 3 (342 mg, 1.0 mmol); 181 mg (42%); mp 164–
165 ꢁC; IR (KBr); m 1504 and 1375 cmÀ1 (Ph); 1H
NMR (400 MHz, Me2SO-d6): m 7.18 (d, 2H, J 8.4 Hz,
H-arom), 6.97 (d, 2H, H-arom), 5.95 (d, 1H, J1,2
3.6 Hz, H-1), 5.63 (d, 1H, H-10), 5.14 (d, 1H, H-2),
4.75 (dd, 1H, J9,10 4.8 Hz, H-9), 4.22 (dd, J8,9 5.2, J8,7
9.2 Hz, H-8), 3.89 (m, 1H, H-7), 2.38–2.32 (m, 1H, H-
6b), 2.30 (s, 1H, CH3), 2.00–1.85 (m, 3H, H-5a, 5b,
6a), 1.52 (s, 3H, CH3), 1.38 (s, 3H, CH3), 1.31 (s, 3H,
CH3), 1.26 (s, 3H, CH3); Anal. Calcd for C23H29NO7:
C, 64.02; H, 6.77; N, 3.25. Found: C, 62.24; H, 6.61;
N, 3.40.
3.1.4.5.
(1aS,2aR,4S,7aS,8bR,9aR,10aR)-3-(4-
Bromoanilino)-1,2:9,10-di-O-isopropylidene-3,5,6-tride-
oxydecos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (6e).
Obtained from 3 (342 mg, 1.0 mmol); 253 mg (51%);
mp 220–222 ꢁC; IR (KBr); m 1480 and 1379 cmÀ1 (Ph);
1H NMR (400 MHz, Me2SO-d6): d 7.55 (d, 2H, J
8.4 Hz, H-arom), 7.02 (d, 2H, H-arom), 5.96 (d, 1H,
J1,2 4.0 Hz, H-1), 5.64 (d, 1H, H-10), 5.22 (d, 1H,
H-2), 4.75 (dd, 1H, J9,10 4.8 Hz, H-9), 4.19 (dd, 1H,
J8,7 4.0, J8,9 5.2 Hz, H-8), 3.89 (m, 1H, H-7), 2.40–2.30
(m, 1H, H-6b), 2.00–1.80 (m, 3H, H-5a, 5b, 6a), 1.53
(s, 3H, CH3), 1.36 (s, 3H, CH3), 1.31 (s, 3H, CH3),
1.25 (s, 3H, CH3); Anal. Calcd for C22H26BrNO7: C,
53.24; H, 5.28; N, 2.82. Found: C, 53.28; H, 5.32; N,
2.90.
3.1.4.2. (1aS,2aR,4S,7aS,8bR,9aR,10aR)-1,2:9,10-
Di-O-isopropylidene-3-(o-toluidino)-3,5,6-trideoxydecos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (6b). Obtained
from 3 (342 mg, 1.0 mmol); 194 mg (45%); mp 122–
1
124 ꢁC (dec); IR (KBr); m 1505 and 1378 cmÀ1 (Ph); H
NMR (400 MHz, Me2SO-d6): d 7.22 (t, 1H, J 7.2 Hz,
H-arom), 7.17 (d, 1H, H-arom), 7.06 (t, 1H, H-arom),
6.89 (d, 1H, H-arom), 5.99 (d, 1H, J1,2 4.0 Hz, H-1),
5.64 (d, 1H, H-10), 5.14 (d, 1H, H-2), 4.77 (dd, 1H,
J9,10 4.8 Hz, H-9), 4.20 (dd, 1H, J8,7 4.0, J8,9 5.6 Hz, H-
8), 3.91 (m, 1H, H-7), 2.45–2.35 (m, 1H, H-6b), 2.12 (s,
3H, CH3), 2.00–1.85 (m, 3H, H-6a, 5a, 5b), 1.54 (s, 3H,
CH3), 1.37 (s, 3H, CH3), 1.32 (s, 3H, CH3), 1.23 (s, 3H,
CH3); Anal. Calcd for C23H29NO7: C, 64.02; H, 6.77;
N, 3.25. Found: C, 62.23; H, 6.56; N, 3.39.
3.1.4.6. (1aS,2aR,4S,7aS,8bR,9aR,10aR)-1,2:9,10-
Di-O-isopropylidene-3-(4-methoxyanilino)-3,5,6-trideoxy-
decos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (6f). Ob-
tained from 3 (342 mg, 1.0 mmol); 188 mg (42%); mp
1
134–136 ꢁC; IR (KBr); m 1513 and 1376 cmÀ1 (Ph); H
NMR (400 MHz, Me2SO-d6): d 7.12 (d, 2H, J 8.8 Hz,
H-arom), 6.94 (d, 2H, H-arom), 5.96 (d, 1H, J1,2
4.0 Hz, H-1), 5.63 (d, 1H, H-10), 5.18 (d, 1H, H-2),
4.74 (dd, J9,10 4.4 Hz, H-9), 4.22 (dd, J8,9 5.2, J8,7
9.6 Hz, H-8), 3.90 (m, 1H, H-7), 3.76 (s, 3H, OCH3),
2.40–2.30 (m, 1H, H-6b), 2.00–1.80 (m, 3H, H-5a,
5b, 6a), 1.52 (s, 3H, CH3), 1.38 (s, 3H, CH3), 1.31 (s,
3H, CH3), 1.29 (s, 3H, CH3); Anal. Calcd for
C23H29NO8: C, 61.73; H, 6.53; N, Found: C, 61.62; H,
6.35; N, 3.33.
3.1.4.3. (1aS,2aR,4S,7aS,8bR,9aR,10aR)-3-(4-Fluoro-
anilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxydecos-
4-ulose 1,4:7,10-difuranose-4,8-pyranose (6c). Obtained
from 3 (342 mg, 1.0 mmol); 196 mg (45%); mp 162–
164 ꢁC; IR (KBr); m 1504 and 1378 cmÀ1 (Ph); 1H
NMR (400 MHz, Me2SO-d6): d 7.21 (m, 2H, H-arom),
7.12 (m, 2H, H-arom), 5.96 (d, 1H, J1,2 4.0 Hz, H-1),
5.64 (d, 1H, H-10), 5.22 (d, 1H, H-2), 4.74 (dd, 1H,
J9,10 4.8 Hz, H-9), 4.21 (dd, 1H, J8,7 4.4, J8,9 5.6 Hz,
H-8), 3.89 (m, 1H, H-7), 2.40–2.30 (m, 1H, H-6b),
2.00–1.85 (m, 3H, H-5a, 5b, 6a), 1.53 (s, 3H, CH3),
1.36 (s, 3H, CH3), 1.31 (s, 3H, CH3), 1.25 (s, 3H,
CH3); Anal. Calcd for C22H26FNO7: C, 60.68; H, 6.02;
N, 3.22. Found: C, 60.84; H, 6.23; N, 3.42.
3.1.5. General procedure for the preparation of compound
7a–f. A solution of compound 6a–f (0.1 mmol) and
NaBH4 (11.0 mg, 0.29 mmol) in EtOH (5 mL) was stir-
red at rt for 3 h. After evaporation of the solvent under
reduced pressure, the residue was dissolved in water
(10 mL), extracted with EtOAc (3 · 10 mL), dried
(Na2SO4), and the solvents evaporated. After recrystal-
lization from EtOH, the product 7a–f was obtained as
colorless needles.

2420
D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
3.1.5.1.
(1aS,2aR,3aS,4S,7aS,8bR,9aR,10aR)-1,2:
H-1), 5.58 (d, 1H, H-10), 5.55 (d, 1H, NH), 4.81 (t,
1H, J2,3 3.6 Hz, H-2), 4.65 (t, 1H, J9,10 4.4 Hz, H-9),
4.10 (dd, 1H, J3,NH 10.0 Hz, H-3), 3.99 (t, 1H, J8,9
4.4 Hz, H-8), 3.76 (m, 1H, H-7), 1.90–1.70 (m, 4H, H-
5, 6), 1.50 (s, 3H, CH3), 1.46 (s, 3H, CH3), 1.30 (s,
3H, CH3), 1.26 (s, 3H, CH3); Anal. Calcd for
C22H28ClNO7: C, 58.21; H, 6.22; N, 3.09. Found: C,
63.86; H, 7.33; N, 3.20.
9,10-Di-O-isopropylidene-3-(p-toluidino)-3,5,6-trideoxy-
decos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (7a). Ob-
tained from 6a (43 mg, 0.1 mmol); 40 mg (92%); mp
132–134 ꢁC; IR (KBr); m 3415 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.90 (d, 2H, J 8.4 Hz,
H-arom), 6.73 (d, 2H, H-arom), 5.79 (d, 1H, J1,2
4.0 Hz, H-1), 5.57 (d, 1H, H-10), 5.04 (d, 1H, NH),
4.78 (dd, 1H, J2,3 5.6 Hz, H-2), 4.64 (dd, 1H, J9,10
4.8 Hz, H-9), 4.07 (dd, 1H, J3,NH 10.4 Hz, H-3), 3.98
(dd, 1H, J8,7 5.2, J8,9 5.6 Hz, H-8), 3.76 (m, 1H, H-7),
2.15 (s, 3H, CH3), 1.90–1.70 (m, 4H, H-5, 6), 1.50
(s, 3H, CH3), 1.47 (s, 3H, CH3), 1.30 (s, 3H, CH3),
1.26 (s, 3H, CH3); Anal. Calcd for C23H31NO7: C,
63.73; H, 7.21; N, 3.23. Found: C, 63.86; H, 7.33; N,
3.20.
3.1.5.5. (1aS,2aR,3aS,4S,7aS,8bR,9aR,10aR)-3-(4-
Bromoanilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-
decos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (7e). Ob-
tained from 6e (50 mg, 0.1 mmol); 48 (96%); mp 122–
124 ꢁC; IR (KBr); m 3412 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 7.21 (d, 2H, J 8.8 Hz, H-
arom), 6.81 (d, 2H, H-arom), 5.80 (d, 1H, J1,2 4.0 Hz,
H-1), 5.59 (d, 1H, H-10), 5.58 (d, 1H, NH), 4.65 (t,
J9,10 4.8 Hz, H-9), 4.81 (dd, 1H, J2,3 5.6 Hz, H-2),
4.10 (dd, 1H, J3,NH 10.4 Hz, H-3), 3.98 (t, J8,9 4.8 Hz,
H-8), 3.76 (m, 1H, H-7), 1.90–1.70 (m, 4H, H-5, 6),
1.50 (s, 3H, CH3), 1.46 (s, 3H, CH3), 1.30 (s, 3H,
CH3), 1.26 (s, 3H, CH3); Anal. Calcd for C22H28BrNO7:
C, 53.02; H, 5.66; N, 2.81. Found: C, 53.22; H, 5.58; N,
2.92.
3.1.5.2.
(1aS,2aR,3aS,4S,7aS,8bR,9aR,10aR)-1,2:
9,10-Di-O-isopropylidene-3-(o-toluidino)-3,5,6-trideoxy-
decos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (7b). Ob-
tained from 6b (43 mg, 0.1 mmol); 41 mg (95%); mp 80–
81 ꢁC; IR (KBr);
m
3439 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 7.04 (m, 2H, H-arom), 6.79
(d, 1H, J 8.0 Hz, H-arom), 6.60 (t, 1H, J 7.2 Hz, H-
arom), 5.85 (d, 1H, J1,2 4.0 Hz, H-1), 5.61 (d, 1H, H-
10), 4.89 (t, 1H, J2,3 4.0 Hz, H-2), 4.67 (t, 1H, J9,10
4.8 Hz, H-9), 4.16 (m, 2H, H-3, NH), 4.11 (t, 1H, J8,9
4.8 Hz, H-8), 3.84 (m, 1H, H-7), 2.12 (s, 3H, CH3),
1.90–1.60 (m, 4H, H-5, 6), 1.51 (s, 3H, CH3), 1.50 (s,
3H, CH3), 1.32 (s, 3H, CH3), 1.30 (s, 3H, CH3); Anal.
Calcd for C23H31NO7: C, 63.73; H, 7.21; N, 3.23.
Found: C, 63.80; H, 7.42; N, 3.32.
3.1.5.6.
(1aS,2aR,3aS,4S,7aS,8bR,9aR,10aR)-1,2:
9,10-Di-O-isopropylidene-3-(4-methoxyanilino)-3,5,6-tri-
deoxydecos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (7f).
Obtained from 6f (45 mg, 0.1 mmol); 43 mg (95%); mp
102–104 ꢁC; IR (KBr); m 3429 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.77 (d, 2H, J 9.2 Hz, H-
arom), 6.72 (d, 2H, H-arom), 5.79 (d, 1H, J1,2 4.0 Hz,
H-1), 5.58 (d, 1H, H-10), 4.82 (d, 1H, NH), 4.77 (dd,
1H, J2,3 5.6 Hz, H-2), 4.64 (dd, 1H, J9.10 4.8, J9,8
5.6 Hz, H-9), 4.10–3.90 (m, 2H, H-3, 8), 3.77 (m, 1H,
H-7), 3.65 (s, 3H, OCH3), 1.70–1.90 (m, 4H, H-5,6),
1.50 (s, 3H, CH3), 1.48 (s, 3H, CH3), 1.30 (s, 3H,
CH3), 1.26 (s, 3H, CH3); Anal. Calcd for C23H31NO8:
C, 61.46; H, 6.95; N, 3.12. Found: C, 61.65; H, 6.80;
N, 3.23.
3.1.5.3. (1aS,2aR,3aS,4S,7aS,8bR,9aR,10aR)-3-(4-
Fluoroanilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-
decos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (7c). Ob-
tained from 6c (43 mg, 0.1 mmol); 41 mg (95%); mp 92–
94 ꢁC; IR (KBr);
m
3428 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 6.92 (m, 2H, H-arom), 6.83
(m, 2H, H-arom), 5.80 (d, 1H, J1,2 4.0 Hz, H-1), 5.58
(d, 1H, H-10), 5.27 (d, 1H, NH), 4.79 (dd, 1H, J2,3
4.8 Hz, H-2), 4.64 (t, 1H, J9,10 4.8 Hz, H-9), 4.06 (dd,
1H, J3,NH 9.6 Hz, H-3), 3.98 (dd, 1H, J8,9 4.8, J8,7
5.2 Hz, H-8), 3.77 (m, 1H, H-7), 1.90–1.70 (m, 4H, H-
5, 6), 1.50 (s, 3H, CH3), 1.47 (s, 3H, CH3), 1.29 (s,
3H, CH3), 1.26 (s, 3H, CH3); Anal. Calcd for
C22H28FNO7: C, 60.40; H, 6.45; N, 3.20. Found: C,
60.54; H, 6.49; N, 3.34.
3.2. Mass spectrometry
The chemicals used were commercially available and of
analytical grade. HPLC grade MeOH was used to dis-
solve the samples. MeOH-d was purchased from the Sig-
ma Company. Mass spectra were recorded on a Bruker
Esquire 3000 ion trap mass spectrometer (Bruker Dalto-
nik, Bremen, Germany) equipped with an electrospray
ion source. Samples were typically dissolved in MeOH
at a concentration of about 10À5 mol/L. The deuterated
analogues were prepared by dissolving the sample in
MeOH-d. They were introduced into the electrospray
needle by a Cole–Parmer 74900 syringe pump (Cole–
Parmer Instrument Company) at a flow rate of 4 lL/
min. The ESI source potentials were capillary 4.0 kV,
3.1.5.4. (1aS,2aR,3aS,4S,7aS,8bR,9aR,10aR)-3-(4-
Chloroanilino)-1,2:9,10-di-O-isopropylidene-3,5,6-trideoxy-
decos-4-ulose 1,4:7,10-difuranose-4,8-pyranose (7d). Ob-
tained from 6d (45 mg, 0.1 mmol); 40 mg (89%); mp
130–132 ꢁC; IR (KBr); m 3412 cmÀ1 (NH); 1H NMR
(400 MHz, Me2SO-d6): d 7.09 (d, 2H, J 8.4 Hz, H-
arom), 6.85 (d, 2H, H-arom), 5.80 (d, 1H, J1,2 3.6 Hz,

D.-P. Zou et al. / Carbohydrate Research 340 (2005) 2411–2421
2421
lens 1 5.0 V, lens 2 60.0 V, and capillary exit offset is
References
75.9 V. The mass spectrometer was scanned at the rate
300 mass units per second. At least 10 scans were aver-
aged to obtain each spectrum. Nitrogen as nebulizer gas
with a flow of 4 L/min (nebulizer pressure of 7 psi) at
300 ꢁC was used. The MSn spectra were obtained by
low-energy CID with helium after isolation of the
appropriate precursor ions. An isolation width of 2.0
m/z was used to isolate the selected peaks. The fragmen-
tation low mass cut-off was set to 27% of the precursor
ion mass. The collision conditions were ramped from
0.60% to 1.20% of the fragmentation amplitude and
set over the 40 ms fragmentation time. Data acquisition
and processing were carried out using Data analysis 5.0
software supplied with the instrument.
1. (a) Iwasa, T.; Kusuka, T.; Suetomi, K. J. Antibiot. 1978,
31, 511–518; (b) Harada, S.; Kishi, T. J. Antibiot. 1978, 31,
519–524; (c) Harada, S.; Mizuta, E.; Kishi, T. Tetrahedron
1981, 37, 1317–1327.
2. (a) Takatsuki, A.; Arima, K.; Tamura, G. J. Antibiot.
1971, 24, 215–223; (b) Takatsuki, A.; Tamura, G. J.
Antibiot. 1971, 24, 224–231; (c) Takatsuki, A.; Tamura, G.
J. Antibiot. 1971, 24, 232–238; (d) Takatsuki, A.; Kawa-
mura, K.; Okina, M.; Kodama, Y.; Ito, T.; Tamura, G.
Agric. Biol. Chem. 1977, 41, 2307–2309.
3. (a) Mootoo, D. R.; Fraser-Reid, B. J. Org. Chem. 1987,
52, 4511–4517; (b) Mootoo, D. R.; Fraser-Reid, B. J. Org.
Chem. 1989, 54, 5548–5550; (c) Jørgensen, M.; Iversen, E.
H.; Madsen, R. J. Org. Chem. 2001, 66, 4625–4629; (d)
´
Jarosz, S.; Skora, S.; Szewczyk, K.; Ciunik, Z. Tetrahe-
dron: Asymmetry 2001, 12, 1895–1905; (e) Jarosz, S. J.
Carbohydr. Chem. 2001, 20, 93–107; (f) Marco-Cotelles, J.;
Opazo, E. D.; Arroyo, N. Tetrahedron 2001, 57, 4729–
4739; (g) Junker, H.-D.; Phung, N.; Fessner, W.-D.
Tetrahedron Lett. 1999, 40, 7063–7066; (h) Jarosz, S.;
4. Conclusions
´
Salanski, P.; Mach, M. Tetrahedron 1998, 54, 2583–2594.
Electrospray-ionization MSn mass spectrometry (ESI-
MSn) with CID was used to establish the fragmentation
behavior of four series of sodium ion adducts of higher-
carbon amino spiro-sugar derivatives. In the ESI-MS
spectra of compounds 4–7, the base peaks are the so-
dium ion adducts and a dominant fragmentation path-
way is the loss of one or two molecule of acetone.
Their fragmentation pathways have been proposed on
the basis of the MSn studies. Some interesting ions at
[M+NaÀ46]+, [M+NaÀ58À76]+, and [M+NaÀ100]+
have been found and the mechanism of formation is
proposed. The [M+NaÀ46]+ ion in the spectra of com-
pounds 4a–h was probably formed by the opening of
two rings followed by simultaneous elimination of car-
bon monoxide and water. The MSn spectra of the
[M+NaÀ58]+ ion of compound 6 was considerably sim-
pler than that of compounds 4 and 5. Formation of the
[M+NaÀ100]+ ion of compounds 7a–f may be rational-
ized by the presence of active hydrogen at the nitrogen
atom. Deuterated analogues were prepared for confir-
mation of the proposed mechanism. The observations
may assist in the interpretation of fragment ion spectra
of related derivatives.
4. Araki, Y.; Endo, T.; Arai, Y.; Tanji, M.; Ishido, Y.
Tetrahedron Lett. 1989, 30, 2829–2832.
5. Kim, K. S.; Cho, I. H.; Joo, Y. H.; Yoo, I. Y.; Song, J. H.;
Ko, J. H. Tetrahedron Lett. 1992, 33, 4029–4032.
6. Eyrisch, O.; Keller, M.; Fessner, W.-D. Tetrahedron Lett.
1994, 35, 9013–9016.
7. Haines, A. H.; Lamb, A. J. Carbohydr. Res. 2000, 325,
323–339.
8. Liu, H. M.; Zou, D. P.; Zhang, F.; Zhu, W. G.; Peng, T.
Eur. J. Org. Chem. 2004, 10, 2103–2106.
9. Przybylski, M.; Glocker, M. O. Angew. Chem., Int. Ed.
1996, 35, 806–813.
10. Loo, J. A. Int. J. Mass Spectrom. 2000, 200, 175–186.
11. Hua, Y.; Wainhaus, S. B.; Yang, Y.; Shen, L.; Xiong, Y.;
Xu, X.; Zhang, F.; Bolton, J. L.; Breemen, R. B. J. Am.
Soc. Mass Spectrom. 2001, 12, 80–87.
12. Jeric, I.; Versluis, C.; Horvat, S.; Heck, A. J. R. J. Mass
Spectrom. 2002, 37, 803–811.
13. Desaire, B.; Leary, J. A. Anal. Chem. 1999, 71, 1997–2002.
14. Penn, S. G.; Cancilla, M. T.; Lebrilla, C. B. Anal. Chem.
1996, 68, 2331–2339.
ˇ
´
´
15. Banoub, J.; Thibault, P.; Goueth, P. Y.; Ronco, G.; Villa,
P. J. Mass Spectrom. 1997, 32, 109–121.
16. Young, M. K.; Dinh, N.; Williams, D. Rapid Commun.
Mass Spectrom. 2000, 14, 1462–1467.
´
´
´
17. Sanchez-Rabaneda, F.; Jauregui, O.; Casals, I.; Andres-
´
Lacueva, C.; Izquierdo-Pulido, M.; Lamuela-Raventos,
M. L. J. Mass Spectrom. 2003, 38, 35–42.
18. Levery, S. B.; Toledo, M. S.; Straus, A. H.; Takahashi, H.
K. Rapid Commun. Mass Spectrom. 2001, 15, 2240–2258.
19. Madhusudanan, K. P.; Raj, K.; Bhaduri, A. P. J. Mass
Spectrom. 2000, 35, 901–911.
Acknowledgments
20. Cui, M.; Song, F.; Liu, Z.; Liu, S. Rapid Commun. Mass
Spectrom. 2001, 15, 586–595.
21. Liu, H. M.; Zhang, F.; Wang, S. Org. Biomol. Chem. 2003,
1, 1641–1642.
22. Zou, D. P.; Zhu, W. G.; Xu, W. C.; Liu, H. M.; Zhang,
H. J.; Li, X. Bopuxue Zazhi 2004, 3, 305–310.
We are grateful to the NNSF of PR China (No.
20272054) for financial support of this research. We also
express our deep appreciation to Dr. Marc A. Grundl
of Stuttgart University for the revision of the
manuscript.