An efficient synthesis of 2′-O-(β-d-glucopyranosyl)- and 2′-O-(2-acetamido-2-deoxy-β-d-glucopyranosyl)-l-biopterins

Article abstract of DOI:10.3987/COM-05-S(T)28

N²-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1′,2′-di-O-trimethylsilyl-L-biopterin (6) was prepared from L-biopterin in 5 steps. Glycosylation of 6 with 2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl trichloroacetimidate (9) and 1,3,4

Full text of DOI:10.3987/COM-05-S(T)28

HETEROCYCLES, Vol. 67, No. 1, 2006
299
HETEROCYCLES, Vol. 67, No. 1, 2006, pp. 299 - 310. © The Japan Institute of Heterocyclic Chemistry
Received, 6th July, 2005, Accepted, 24th August, 2005, Published online, 26th August, 2005. COM-05-S(T)28
AN EFFICIENT SYNTHESIS OF 2’-O-(β-D-GLUCOPYRANOSYL)- AND 2’-
O-(2-ACETAMIDO-2-DEOXY-β-D-GLUCOPYRANOSYL)-L-BIOPTERINS
Tadashi Hanaya,* Kazuyuki Soranaka, Koichiro Harada, Hiroshi Yamaguchi,
Ryo Suzuki, Yusumi Endo, Hiroshi Yamamoto, and Wolfgang Pfleiderer†
Department of Chemistry, Faculty of Science, Okayama University, Tsushima-
naka, Okayama 700-8530, Japan
† Fakultät für Chemie, Universität Konstantz, Postfach 5560, D-78434 Konstanz,
Germany
Abstract – N2-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1’,2’-
di-O-trimethylsilyl-L-biopterin (6) was prepared from L-biopterin in 5 steps.
Glycosylation of 6 with 2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl trichloro-
acetimidate (9) and 1,3,4,6-tetra-O-acetyl-2-phtalimido-β-D-glucopyranose (10)
respectively afforded the corresponding 2’-O-(β-D-glucopyranosyl) derivatives
(12b, 12c) as major products. Removal of protecting groups of 12c provided
naturally occurring limipterin (2).
INTRODUCTION
Some pteridine glycosides having a glycosidated side-chain at C–6 of the pterin ring are known to occur
in nature. For example, 2’-O-(α-D-glucopyranosyl)-L-biopterin (1a) was isolated from cyanobacterium,
Anacystis nidulans,1 Synechococcus sp. PCC 7942,2 and Spirulina (Arthrospira) platensis,3 whereas
limipterin [2’-O-(2-acetamido-2-deoxy-β-D-glucopyranosyl)-L-biopterin] (2) was isolated from a green
sulfur photosynthetic bacterium Chlorobium limicola f. thiosulfatophilum NCIB 8327.4 Although
various types of glycosides of biopterin and related pteridins are considered to be of interest from the
viewpoint of their biological activities and functions as well as the structural proof of hitherto reported
O
N
OH
O
N
OH
N
N
N
HN
H N
HN
H N
O
O
N
2
2
OH
HO
HO
OH
O
OH
O
HO
OH
NHAc
1a
2

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HETEROCYCLES, Vol. 67, No. 1, 2006
natural products, attempts for preparation of these compounds by glycosylation have not been reported so
far. We describe herein the first efficient synthesis of 2’-O-(D-glucopyranosyl)-L-biopterins.5
RESULTS AND DISCUSSIN
Because of the effectively stabilized intramolecular hydrogen bondings in the solid state,6 many pterin
derivatives including biopterin are little soluble in nonpolar aprotic solvents in which glycosidation
reactions smoothly proceed. To overcome this probelem, L-biopterin (3) can be converted into a
suitably protected and sufficiently solubilized derivative, N2-(N,N-dimethylaminomethylene)-3-[2-(4-
nitrophenyl)ethyl]-L-biopterin (5)7 (Scheme 1). Namely, treatment of 3 with N,N-dimethylformamide
dimethyl acetal in DMF and the following acetylation of hydroxy groups afforded the 1’,2’-di-O-acetyl-
N2-(N,N-dimethylaminomethylene) derivative (4). The Mitsunobu reaction of 4 with p-nitrophenylethyl
(NPE) alcohol in the presence of triphenylphosphine and diethyl azodicarboxylate (DEAD),8 followed by
methanolysis of acetyl groups provided a versatile intermediate (5). The reported procedures7 for these
steps were slightly modified to give 4 and 5 in higher yields.
For the purpose of raising the solubility of the N2, N(3)-protected biopterin (5) in dichloromethane, 5 was
temporarily silylated with 1,1,1,3,3,3-hexamethyldisilazane (HMDS) in the presence of ammonium
1) NPEOH, PPh
3
O
N
OH
O
N
OAc
1) Me NCH(OMe)
2
2
DEAD, THF, rt
rt, 12 h, 91%
3
2'
6
N
N
N
N
DMF, rt, 3 h
3'
HN
HN
1'
OH
OAc
2) Ac O, Py
2
2) TEA, MeOH
30 °C, 4 h, 99%
7
H N
2
N
N
2
rt, 12 h
3
96% (2 steps)
4
O
OH
O
OTMS
NPE
NPE
N
N
N
N
HMDS, (NH ) SO
4 2
7–10
4
N
N
OH
OR
OTMS
CH Cl , reflux, 48 h
2
activators
2
N
N
N
N
N
N
CH Cl , rt, 18-24 h
2
2
99%
5
6
O
O
N
OH
O
N
OR
NPE
NPE
NPE
N
N
N
N
N
N
N
+
+
OH
OR
OR
N
N
N
N
N
N
N
N
N
11a–c
12a–c
BzO
13a–c
AcO
AcO
AcO
O
OAc
O
OBz
O
OAc
a R =
b R =
c R =
AcO
BzO
OAc
OBz
NPhth
Scheme 1

HETEROCYCLES, Vol. 67, No. 1, 2006
301
sulfate in dichloromethane under reflux for 12 h to give 1’,2’-di-O-trimethylsilyl derivative (6)
quantitatively. Glycosylation of 6 with various glucosyl donors (7–10) was extensively investigated
under various conditions in the presence of activators. The yields of 1’-O- (11), 2’-O-glycosyl (12), and
1’,2’-di-O-glycosyl compounds (13) depended upon the molar ratio of 6 to glycosyl donors, as
summarized in Table 1.
AcO
AcO
BzO
BzO
BzO
BzO
AcO
AcO
OAc
O
OAc
O
OAc
O
OBz
O
OBz
Br
OAc
Br
OBz
O
NH
CCl
BzO
NPhth
3
8
7
9
10
Table 1. Glycosilation of L-biopterin derivative (6) a
——————————————————————————————————————————————
b
Entry
Glycosyl donor (mol equiv.) Activator (mol equiv.)
Products (yield)
——————————————————————————————————————————————
1
7 (1.5)
7 (2.0)
7 (2.5)
7 (3.0)
8 (2.0)
8 (3.0)
9 (2.0)
9 (3.0)
10 (2.5)
10 (2.5)
SnCl4 (4.0)
SnCl4 (4.0)
SnCl4 (4.0)
SnCl4 (4.0)
SnCl4 (4.0)
SnCl4 (4.0)
TMSOTf (0.4)
TMSOTf (0.6)
SnCl4 (4.0)
SnCl4 (8.0)
11a (27%)
11a (31%)
11a (22%)
11a (16%)
11b (11%)
11b (15%)
11b (7%)
11b (5%)
11c (<1%)
11c (<1%)
12a (32%)
12a (38%)
12a (26%)
12a (18%)
12b (28%)
12b (41%)
12b (47%)
12b (36%)
12c (40%)
12c (68%)
13a (5%)
13a (8%)
13a (16%)
13a (27%)
13b (5%)
13b (14%)
13b (0%)
13b (9%)
13c (5%)
13c (8%)
2
3
4
5
6
c
c
7
8
9
10
——————————————————————————————————————————————
a
b
All reactions were carried out in CH2Cl2 at rt (except for Entries 7, 8) for 18–24 h.
Yields of inseparable
c
monoglycosides (11) and (12) were estimated by 1H NMR spectra.
At –30 °C.
Glycosylation of 6 with 1.5–3.0 mol equiv. of tetera-O-acetyl-α-D-glucopyranosyl bromide (7)9 in the
presence of tin(IV) chloride (4.0 mol equiv.) in dichloromethane at room temperature for 24 h afforded
1’-O-(β-D-glucopyranosyl)-L-biopterin (11a), 2’-O-(β-D-glucopyranosyl) isomer (12a), and 1’,2’-di-O-
(β-D-glycopyranosyl) derivative (13a) (Entries 1–4).10 Although the mono- and di-glycosyl compounds
were chromatographically separable with a silica gel column, separation of the monoglucosides (11a) and
(12a) has not been achieved. The 1’-O-substituted structure of 11a was derived from a doublet of H-1’
signal, whereas a triplet of H-1’ for 12a indicated the presence of 1’-OH group (Table 2). As for
glucopyranosyl moieties of these compounds, all of their anomeric configurations were found to be β-
form on the evidence of their J1,2 values (7.7–7.9 Hz). Use of 2.0 mol equiv. of 7 (Entry 2) gave
monoglycosides (11a, 12a) in the highest yield among Entries 1–4, but 2’-O-preference of glycosylation

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HETEROCYCLES, Vol. 67, No. 1, 2006
was little observed: 1’-O-glycoside: 2’-O-glycoside = 45:55. Meanwhile, in the case of same treatment
of 6 with tetra-O-benzoyl-α-D-glucopyranosyl bromide (8),11 3 mol equiv. of 8 rather than 2 mol afforded
monoglycosides (11b, 12b) in a higher yield and better 2’-O-selectivity (27:73) (Entries 5,6). An
improvement in the 2’-O-selectivity is presumably ascribed to use of a larger protecting group of the
glycosyl donor (8) than that of 7. The 1’-O-monoglycoside (11b) and the 2’-O-glycoside (12b) were
also obtained as an inseparable mixture and separation of them was achieved after having converted them
into the corresponding acetates (see below).
Treatment of 6 with 2 mol equiv. of 2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl trichloroacetimidate
(9)12 in the presence of trimethylsilyl triflate (TMSOTf)13 at –30 °C revealed an improvement of 2’-O-
selectivity (13:87) and absence of diglycoside (13b) (Entry 7). Use of 3 mol equiv. of 9 resulted in a
lower yield of 12b and formation of 13b (Entry 8).
Glycosylation of 6 with 1,3,4,6-tetra-O-acetyl-2-deoxy-2-phthalimido-β-D-glucopyranose (10)14 in the
presence of tin chloride (4.0 equiv.) mainly afforded 2’-O-monoglycoside (12c) together with 1’,2’-di-O-
glycoside (13c) (Entry 9). Although a trace amount of byproduct speculated to be the 1’-O-glycoside
(11c) in the fraction of 12c was detectable from 1H NMR spectrum, 11c was removed by recrystallization
of the fraction to give pure 12c. Use of a larger amount (8.0 equiv.) of the activator brought about the
highest yield (68%) of 12c among this work. The stereoselective β-glycoside formation of all these
products (11–13a,b,c) was mainly attained by participation of neighboring groups (acyloxy of 7–9 and
phthalimido of 10).
O
N
OAc
O
O
N
OR
NPE
N
N
N
N
N
N
HN
RNH
1) MeONa, MeOH
rt, 1 h
O
N
1) Ac O, Py
2) NH OH, MeOH
2
4
BzO
RO
rt, 12 h
rt, 12 h
O
O
12b
OBz
OR
2) separation
3) DBU, DMF
rt, 12 h
(+ 11b)
BzO
RO
OBz
OR
89% (3 steps)
14
1b R = H
15 R = Ac
Ac O, Py
2
O
OAc
O
N
OH
N
NPE
N
N
N
HN
H N
O
O
AcNH
N
N
HO
2
1) NH OH, MeOH
1) MeNH , MeOH
4
2
AcO
rt, 24 h
rt, 12 h
O
O
12c
OAc
OH
2) Ac O, Py
2) DBU, DMF
rt, 12 h
2
AcO
HO
rt, 12 h
NHAc
NHAc
91% (2 steps)
95% (2 steps)
16
2
Scheme 2

HETEROCYCLES, Vol. 67, No. 1, 2006
303
Removal of the protecting groups of the 2’-O-(tetra-O-benzoyl-β-D-glucopyranosyl)-L-biopterin
derivative (14) isolated as the 1’-O-acetate from 12b was carried out according to the following steps
(Scheme 2). Namely, 14 was treated with sodium methoxide in methanol at room temperature for 2 h to
cleave acetyl and benzoyl groups, and then treated with aqueous ammonia-methanol at room temperature
for 12 h, to cleave the N,N-dimethylaminomethylene group. The NPE group was then removed by the
treatment with DBU in DMF at room temperature for 12 h,7,8 thus providing 2’-O-(β-D-glucopyranosyl)-
L-biopterin (1b) in 89% (overall yield from 14). Structure of 1b was established as the corresponding
per-O- and N2-acetylated derivative (15) obtained by treatment of acetic anhydride in pyridine (Table 2).
For deprotection of 2’-O-(tetra-O-acetyl-2-deoxy-2-phthalimido-β-D-glucopyranosyl)-L-biopterin
derivative (12c), conversion of phthalimido group into acetamido group was carried out as an initial step.
Namely, removal of the phthaloyl group of 12c with methylamine in methanol, followed by acetylation
with acetic anhydride in pyridine, afforded di-N2:1’-O-acetyl-2’-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-
β-D-glucopyranosyl)-3-[2-(4-nitrophenyl)ethyl]-L-biopterin (16).
Treatment of 16 with aqueous
ammonium hydroxide, followed by the action of DBU, afforded limipterin (2). The spectral data of the
synthetic compound (2) were in all respects with those of the natural product.4
By employing similar procedures described for those of 2 from 16, deprotection of a mixture of 1’-O-
glycoside (11a) and 2’-O-glycoside (12a) afforded a mixture of 1’-O-(β-D-glucopyranosyl)-L-biopterin
(17) and 1b. Although these compounds were converted into the corresponding acetylated derivatives
for separation and structural assignment, isoration of di-N2:2’-O-acetyl-1’-O-(2,3,4,6-tetra-O-β-D-
glucopyranosyl)-L-biopterin (18) from 15 was not achieved. The similar treatment of 1’,2’-di-O-(β-D-
glucopyranosyl) derivatives (13a, 13b) both afforded the same L-biopterin di-glycoside (19), which was
converted into the acetylated compound (20).
RO
RO
OR
OR
RO
RO
O
O
OR
OR
O
N
O
O
N
O
N
N
N
N
HN
RNH
HN
RNH
OR
O
RO
O
OR
17 R = H
18 R = Ac
19 R = H
RO
OR
20 R = Ac
The present work thus demonstrates an efficient way for preparation of 2’-O-(β-D-glucopyranosyl)-L-
biopterin (1b) and natural product limipterin (2). Improvement of selectivity for 2’-O-
monoglycosylation as well as preparation of naturally occurring 2’-O-(α-D-glucopyranosyl) derivative
(1a), is in progress.

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HETEROCYCLES, Vol. 67, No. 1, 2006
1
Table 2. 500 MHz H-NMR Spectral Parameters for 1’- and 2’-O-(β-D-glucopyranosyl)-L-biopterin
derivatives (11–16, 18, 20) in CDCl
——————————————————————————————————————————————
3
Chemical shifts / δ (coupling constants / Hz)
Biopterin moiety
Me2NCH=N-2
———————— ——————————— ——————————
pounds H-7 H-1’ H-2’ H3-3’ Me2N CH=N Ar(m) Ar(o) CH2CH2N Other signals
(J1’,2’) (J2’,3’ (Jo,m
(3JH,H
——————————————————————————————————————————————
ArCH2CH2-N(3)
Com-
)
)
)
11a
12a
13a
11b
12b
13b
12c
13c
14
8.80 5.07 4.30 1.07 3.24, 3.19 8.86
(3.7) (6.7)
8.15 7.43
(8.6)
3.18, 4.61
(7.5)
2.68 (HO-2’, J2’,OH = 6.1)
8.89 4.95 4.25 1.24 3.24, 3.19 8.86
(5.0) (6.4)
8.14 7.42
(8.6)
3.18, 4.61
(7.5)
3.27 (HO-1’, J1’,OH = 5.2)
8.80 5.12 4.57 1.12 3.24, 3.19 8.81
(2.8) (6.7)
8.15 7.42
(8.6)
3.19, 4.62
(7.5)
8.72 5.14 4.32 1.03 3.24, 3.18 8.81
(3.4) (6.7)
8.14 7.45
(8.5)
3.14, 4.58
(7.6)
2.82 (HO-2’, J2’,OH = 5.9)
3.15 (HO-1’, J1’,OH = 4.3)
8.85 4.93 4.35 1.22 3.24, 3.19 8.84
(5.2) (6.4)
8.14 7.41
(8.5)
3.14, 4.55
(7.6)
8.75 4.93 4.50 1.23 3.24, 3.19 8.79
(3.4) (6.4)
8.14 7.41
(8.6)
3.12, 4.50
(7.6)
8.67 4.31 4.26 1.22 3.25, 3.20 8.83
(5.2) (6.4)
8.31 7.41
(8.7)
3.13, 4.55
(7.6)
4.79 (HO-1’, J1’,OH = 5.3)
8.21 4.65 4.38 1.29 3.26, 3.21 8.89
(6.0) (6.4)
8.14 7.42
(8.6)
3.14, 4.53
(7.8)
8.65 5.90 4.58 1.34 3.25, 3.21 8.82
(5.5) (6.4)
8.15 7.45
(8.6)
3.19, 4.59
(7.9)
1.74 (AcO-1’)
a
15
8.86 5.94 4.47 1.29
(5.2) (6.4)
10.08 [H-N(3)]
2.15 (AcO-1’)
2.44 (AcN-2), 12.57 (NH-2)
a
16
8.70 5.91 4.44 1.26
(5.2) (6.4)
8.19 7.50
(8.8)
3.17, 4.55
(7.6)
2.16 (AcO-1’)
2.34 (AcN-2), 13.9 (NH-2)
a
18
8.94 5.13 5.32 1.24
(5.2) (6.4)
9.35 [H-N(3)]
2.12 (AcO-2’)
2.47 (AcN-2 ), 12.39 (NH-2)
2.36 (AcN-2 ), 12.40 (NH-2)
20
8.90 5.15 4.57 1.10
(2.7) (6.4)
9.35 [H-N(3)]
——————————————————————————————————————————————
a
b
The assignments of acetyl groups may have to be interchanged.
The assignments of H-1,2,3 signals may have
c
to be interchanged.
Upper two lines for 1’-O-glucopyranosyl moiety, lower two lines for 2’-O-glucopyranosyl
d
moiety. Correlations of protons were confirmed by 2D-COSY measurement.
The assignments of Bz(o)
e
signals may have to be interchanged.
The assignments of phthaloyl groups may have to be interchanged.

HETEROCYCLES, Vol. 67, No. 1, 2006
305
——————————————————————————————————————————————
Glucopyranosyl moiety
H-1
H-2
H-3
H-4
H-5
Ha-6
Hb-6
Other signals
(J1,2) (J2,3) (J3,4) (J4,5) (J5,6a
)
(J6a,6b
)
(J5,6b)
——————————————————————————————————————————————
b
b
b
11a
12a
13a
4.52
(7.7) (9.5) (9.5) (9.8) (5.5)
4.64 4.96 5.14 5.04 3.69
(7.9) (9.5) (9.5) (9.8) (5.5)
5.11
5.10
5.09
3.68
4.25
4.17
(2.4)
4.13
(2.1)
4.12
(2.1)
4.17
(2.4)
4.47
(5.6)
4.49
(5.8)
4.51
(5.8)
4.37
(5.8)
4.17
(2.4)
4.17
(2.4)
4.11
(2.4)
4.48
(5.5)
4.11
(2.4)
4.13
(2.1)
4.12
(2.4)
4.12
(2.1)
4.16
(2.4)
2.13, 2.03, 2.01, 1.99 (AcO-2,3,4,6)
2.08, 2.04, 2.02, 1.98 (AcO-2,3,4,6)
2.19, 2.09, 1.99, 1.98 (AcO-2,3,4,6)
2.16, 2.14, 2.02, 1.98 (AcO-2,3,4,6)
(12.2)
4.23
(12.2)
4.34
c
b
b
b
4.33
(7.9) (9.5) (9.5) (9.8) (4.6)
5.11 4.90 5.32 5.08 3.80
(7.9) (9.5) (9.5) (9.8) (4.7)
4.96 5.65 5.85 5.72 4.17
(7.9) (9.4) (9.7) (9.9) (2.8)
5.08 5.50 5.89 5.65 4.19
(7.9) (9.8) (9.6) (9.8) (3.1)
4.85 5.19 5.72 5.46 4.17
(7.9) (9.5) (9.9) (9.8) (3.0)
5.07 5.47 5.88 5.57 4.03
(7.9) (9.6) (9.8) (9.8) (3.0)
5.53 4.29 5.70 5.13 3.91
(8.6) (10.7) (9.2) (10.1) (5.2)
5.13 3.83 5.52 5.00 3.73
(8.5) (10.7) (9.2) (10.0) (5.5)
5.24 4.16 5.65 5.03 3.81
(8.2) (10.6) (9.2) (10.0) (5.2)
5.16 5.44 5.90 5.61 4.22
(7.9) (9.8) (9.5) (10.1) (3.1)
4.62 4.85 5.10 5.00 3.68
(7.9) (9.5) (9.5) (9.8) (5.2)
4.82 4.20 5.27 5.00 3.77
(8.5) (10.5) (9.5) (9.9) (5.5)
5.12
5.10
5.08
3.57
(12.2)
4.27
(12.2)
4.73
11b
12b
13b
8.09, 7.98, 7.89, 7.80 [Bz(o)]
7.54–7.26 [Bz(m,p)]
(12.1)
4.64
8.03, 7.91, 7.84, 7.80 [Bz(o)]
7.54–7.26 [Bz(m,p)]
(12.2)
4.56
c
d
8.10, 7.98, 7.92, 7.79 [Bz(o)]
(12.2)
4.57
7.55–7.15 [Bz(m,p)]
d
7.98, 7.92, 7.80, 7.57 [Bz(o)]
(12.2)
4.28
7.55–7.15 [Bz(m,p)]
2.11, 2.02,1.82 (AcO-3,4,6)
7.76, 7.65 (Phth)
12c
(12.0)
4.29
c
a
13c
2.08, 1.98, 1.77 (AcO-3,4,6)
e
(12.2)
4.25
7.69, 7.56 (Phth)
a
2.15, 2.00, 1.79 (AcO-3,4,6)
e
(12.2)
4.64
7.78, 7.69 (Phth)
14
15
16
18
8.02, 7.91, 7.77, 7.76 [Bz(o)]
7.54–7.24 [Bz(m,p)]
(11.6)
4.21
a
2.08, 2.02, 1.96, 1.94 (AcO-2,3,4,6)
(12.2)
4.22
a
2.02, 2.02, 1.86 (AcO-3,4,6)
a
(12.2)
4.20
2.08 (AcN-2), 5.83 (NH-2)
b
b
b
a
a
a
4.46
5.06
5.07
5.08
3.61
2.00, 1.99, 1.97, 1.94 (AcO-2,3,4,6)
(7.9) (9.5) (9.5) (9.8) (5.2)
(12.2)
4.33
.
c
b
b
b
20
4.31
(7.9) (9.5) (9.5) (9.8) (4.6)
5.11 4.91 5.33 5.09 3.80
(7.9) (9.5) (9.5) (9.8) (4.6)
5.12
5.10
5.08
3.58
2.14, 2.06, 2.03, 1.99 (AcO-2,3,4,6)
(12.2)
4.26
2.21, 2.09, 2.04, 2.00 (AcO-2,3,4,6)
(12.2)
——————————————————————————————————————————————

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HETEROCYCLES, Vol. 67, No. 1, 2006
EXPERIMENTAL
All reactions were monitored by TLC (Merck silica gel 60F, 0.25 mm) with an appropriate solvent system
[(A) AcOEt, (B) 1:9 MeOH–CHCl3, and (C) 5:3:1 2-PrOH–AcOEt–H2O]. Column chromatography was
performed by Daiso Silica Gel IR-60/210w. Components were detected by exposing the plates to UV
light and/or spraying them with 20% sulfuric acid–ethanol (with subsequent heating). The NMR spectra
were measured in CDCl3 with Hitachi R-1900 (90 MHz) and Varian VXR-500 (500 MHz) spectrometers
at 23 °C. Chemical shifts are reported as δ values relative to CHCl3 (7.26 ppm) as an internal standard.
The MS spectra were taken on a VG-70SE instrument and are given in terms of m/z (relative intensity)
compared with the base peak.
N2-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-L-biopterin (5).7
The following modification of the literature procedures7 was made. To a suspension of 3 (100 mg,
0.421 mmol) in dry DMF (6.0 mL) was added N,N-dimethylformamide dimethyl acetal (0.150 mL, 1.13
mmol). The mixture was stirred at rt for 3 h and evaporated in vacuo. The residue was dissolved in
5% MeOH-CHCl3 and stirred at rt for 2 h. The mixture was evaporated in vacuo and the residue was
dissolved in pyridine (5.0 mL) and acetic anhydride (2.5 mL) at 5 °C. The mixture was stirred at rt for
12 h and evaporated in vacuo. The residue was purified by column chromatography with 2% MeOH-
CHCl3 as an eluant to give 1’,2’-di-O-acetyl-N2-(N,N-dimethylaminomethylene)-L-biopterin (4) (152 mg,
96% from 3) as pale yellow prisms: mp 192–193 °C (from AcOEt-hexane, lit.,7 mp 194–195 °C, 77%
yield); Rf = 0.43 (B).
To a solution of 4 (152 mg, 0.404 mmol), NPE alcohol (81. 0 mg, 0.485 mmol), and triphenylphosphine
(159 mg, 0.608 mmol) in dry THF (4.0 mL) was added DEAD (0.100 mL, 0.654 mmol). The mixture
was stirred at rt for 12 h and evaporated in vacuo. The residue was purified by column chromatography
with 1:2 AcOEt-hexane and then 2% MeOH-CHCl3 to give the 3-NPE derivative (193 mg, 91%) as pale
yellow crystals: mp 204–205 °C (from AcOEt-hexane, lit.,7 mp 205–206 °C, 80% yield); Rf = 0.60 (B).
To a solution of the above product (193 mg, 0.367 mmol) in MeOH (3.0 mL) was added TEA (1.0 mL,
7.2 mmol). The solution was stirred at 30 °C for 4 h and evaporated in vacuo. The residue was
recrystallized from EtOH to give 5 (160 mg, 99%) as pale yellow crystals: mp 136–138 °C (lit.,7 mp
138–140 °C, 97% yield); Rf = 0.42 (B).
N2-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1’,2’-di-O-trimethylsilyl-L-biopterin
(6).
A suspension of 5 (96.0 mg, 0.217 mmol), ammonium sulfate (66.2 mg, 0.502 mmol), and HMDS (0.680
mL, 3.28 mmol) in dry CH2Cl2 (5.0 mL) was refluxed for 48 h. The precipitate was filtered off and the
filtrate was evaporated in vacuo. The residue was purified by short-path column chromatography with
1% MeOH-CHCl3 to give 6 (126 mg, 99%) as a pale yellow foam: Rf = 0.65 (B); 1H NMR (90 MHz) δ =
0.06, 0.09 (3H each, 2s, SiMe3), 1.07 (3H, d, J2’,3’ = 6.4 Hz, H3-3’), 3.09 [2H, m, CH2–C–N(3)], 3.17,
3.22 (3H each, 2s, NMe2), 4.15 (1H, dq, J1’,2’ = 4.0 Hz, H-2’), 4.60 [2H, m, CH2–N(3)], 4.89 (1H, d, H-

HETEROCYCLES, Vol. 67, No. 1, 2006
307
1’), 7.41, 8.12 (2H each, 2d, Jo,m = 8.7 Hz, C6H4NO2), 8,85 (1H, s, CH=N2), 8.93 (1H, s, H-7).
N2-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1’-O-(2,3,4,6-tetra-O-acetyl-β-D-
glucopyranosyl)-L-biopterin (11a), its 2’-O-(2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl) isomer (12a),
and
N2-(N,N-dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1’,2’-bis-O-(2,3,4,6-tetra-O-
acetyl-β-D-glucopyranosyl)-L-biopterin (13a).
To a solution of 79 (180 mg, 0.44 mmol) in CH2Cl2 (2.0 mL) was added SnCl4 (0.10 mL, 0.88 mmol) at
0 °C. After stirring at 0 ° C for 0.5 h, a solution of 6 (130 mg, 0.222 mmol) in CH2Cl2 (1.5 mL) was
added and then the mixture was stirred at rt for 24 h. After addition of saturated aqueous NaHCO3, the
mixture was extracted with CHCl3. The organic layer was dried (Na2SO4) and evaporated in vacuo.
The residue was separated by column chromatography with 1:2 AcOEt-hexane and then 2:3 AcOEt-
CHCl3 into two fractions.
The faster-eluting fraction [Rf = 0.63 (B)] gave 13a (18 mg, 7.3%) as a pale yellow powder: mp
1
184–186 °C (from AcOEt-hexane): H NMR, see Table 2; FAB MS m/z 1102 (M+1; 72), 772 (14), 727
(12), 424 (26), 247 (16), 154 (100). Found: m/z 1102.3760. Calcd for C48H60N7O23: M+1, 1102.3742.
The slower-eluting fraction [Rf = 0.47 (B)] gave a pale yellow powder (117 mg), which consisted of 11a
1
1
(53 mg, 31%) and 12a (64 mg, 38%), the ratio being estimated by H NMR spectrum: H NMR for 11a
and 12a, see Table 2; FAB MS m/z 772 (M+1, 15), 424 (12), 307 (20), 289 (14), 154 (100). Found: m/z
772.2780. Calcd for C34H42N7O14: M+1, 772.2792.
N2-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1’-O-(2,3,4,6-tetra-O-benzoyl-β-D-
glucopyranosyl)-L-biopterin (11b), its 2’-O-(2,3,4,6-tetra-O-benzoyl-β-D-glucopyranosyl) isomer
(12b), and N2-(N,N-dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-1’,2’-bis-O-(2,3,4,6-tetra-
O-benzoyl-β-D-glucopyranosyl)-L-biopterin (13b).
(A) From 8. By use of the same procedures described above, compound (6) (54.0 mg, 0.0922 mmol)
was treated with 811 (183 mg, 0.278 mmol) and SnCl4 (0.044 ml, 0.38 mmol) at rt for 20 h. The
products were separated by column chromatography with 1:2 AcOEt-hexane and then 2% MeOH-CHCl3
into two fractions.
1
The faster-eluting fraction [Rf = 0.75 (B)] gave 13b (20.8 mg, 14%) as a pale yellow powder: H NMR,
see Table 2. Anal. Calcd for C88H75N7O23: C, 66.12; H, 4.73. Found: C, 66.38; H, 4.82.
The slower-eluting fraction [Rf = 0.53 (B)] gave a pale yellow powder (52.8 mg), which consisted of 11b
1
1
(14.0 mg, 15%) and 12b (38.8 mg, 41%), the ratio being estimated by H NMR: H NMR for 11b and
12b, see Table 2.
(B) From 9. To a solution of 912 (121 mg, 0.164 mmol) in CH2Cl2 (2.0 mL) was added TMSOTf (0.006
mL, 0.033 mmol) at –30 °C. After stirring at same temperature for 0.5 h, a solution of 6 (47.8 mg,
0.0816 mmol) in CH2Cl2 (1.0 mL) was added and then the mixture was stirred at –30 °C for 18 h. After
addition of saturated NaHCO3, the mixture was extracted with CHCl3. The organic layer was dried
(Na2SO4) and evaporated in vacuo. The residue was separated by column chromatography to give an
inseparable mixture (45.2 mg) of 11b (6.0 mg, 7%) and 12b (39.2 mg, 47%).

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HETEROCYCLES, Vol. 67, No. 1, 2006
N2-(N,N-Dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-2’-O-(3,4,6-tri-O-acetyl-2-deoxy-2-
phthalimido-β-D-glucopyranosyl)-L-biopterin (12c) and N2-(N,N-dimethylaminomethylene)-3-[2-(4-
nitrophenyl)ethyl]-1’,2’-di-O-(3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-β-D-glucopyranosyl)-L-
biopterin (13c).
By use of the same procedures as those for preparation of 11–13a, compound (6) (315 mg, 0.538 mmol)
was treated with 1014 (644 mg, 1.35 mmol) and SnCl4 (0.51 mL, 4.3 mmol) at rt for 24 h. The products
were separated by column chromatography with 1:2 AcOEt-hexane and then 2% MeOH-CHCl3 into two
fractions.
The faster-eluting fraction [Rf = 0.58 (B)] gave 13c (56 mg, 8.2%) as pale yellow crystals: mp
1
134–135 °C (from AcOEt-hexane); H NMR, see Table 2. Anal. Calcd for C60H61N9O23: C, 56.469; H,
4.82. Found: C, 56.528; H, 4.90.
The slower-eluting fraction [Rf = 0.48 (B)] gave pale yellow solid (324 mg) of 12c contaminated with a
trace amount of 11c, which was recrystallized from AcOEt-hexane to give 12c (315 mg, 68%) as pale
yellow crystals: mp 166–167 °C; 1H NMR, see Table 2. Anal. Calcd for C40H42N8O14: C, 55.94; H, 4.93.
Found: C, 56.02; H, 5.01.
1’-O-Acetyl-N2-(N,N-dimethylaminomethylene)-3-[2-(4-nitrophenyl)ethyl]-2’-O-(2,3,4,6-tetra-O-
benzoyl-β-D-glucopyranosyl)-L-biopterin (14).
A 13:87 mixture (34.1 mg) of 11b and 12b was dissolved in pyridine (1.0 mL) and then acetic anhydride
(0.40 mL, 4.2 mmol) was added at 0 °C. The mixture was stirred at rt for 12 h and evaporated in vacuo.
The residue was chromatographed on a silica gel with AcOEt as an eluant to give 14 (25.1 mg, 81% from
12b) as a yellow syrup: Rf = 0.24 (A); 1H NMR, see Table 2. Anal. Calcd for C56H51N7O15: C, 63.33; H,
4.84. Found: C, 63.24; H, 4.99.
Besides pure 14, a mixture (8.2 mg) which consisted of 14 and the 2’-O-acetyl derivative of 11b was
obtained: Rf = 0.24–0.20 (A).
Di-N2:1’-O-acetyl-2’-O-(2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl)-L-biopterin (15).
Compound (14) (56.3 mg, 0.0530 mmol) was dissolved in MeOH (2.0 mL) and a 28% methanolic
NaOMe (0.03 mL, 0.15 mmol) was added at 0 °C. The mixture was stirred at rt for 1 h and neutralized
with Amberlite IR-120(H+). The resin was filtered off and the filtrate was evaporated in vacuo. The
residue was dissolved in MeOH (4.0 mL) and 28% aqueous ammonia solution (4.0 mL) was added. The
mixture was stirred at rt for 12 h and evaporated in vacuo. The residue was dissolved in DMF (1.0 mL)
and DBU (0.050 mL, 0.32 mmol) was added. The mixture was stirred at rt for 12 h and evaporated in
vacuo. The residue was washed with CHCl3 and dried under reduced pressure to give 2’-O-(β-D-
glucopyranosyl)-L-biopterin (1b) (18.8 mg) as a yellow powder: Rf = 0.23 (C).
Compound (1b) was dissolved in pyridine (2.0 mL) and then acetic anhydride (1.0 mL) was added at 0 °C.
The mixture was stirred at rt for 12 h and evaporated in vacuo. The residue was purified by column
chromatography with AcOEt to give 15 (29.7 mg, 86% from 14) as a pale yellow syrup: Rf = 0.52 (B); 1H
NMR, see Table 2. Anal. Calcd for C27H33N5O14: C, 49.77; H, 5.10. Found: C, 49.92; H, 5.02.

HETEROCYCLES, Vol. 67, No. 1, 2006
309
Di-N2:1’-O-acetyl-3-[2-(4-nitrophenyl)ethyl]-2’-O-(2-acetamido-3,4,6-tri-O-acetyl-2-deoxy-β-D-
glucopyranosyl)-L-biopterin (16).
Compound (12c) (100 mg, 0.116 mmol) was dissolved in MeOH (5.0 mL) and 40% methanolic
methylamine (2.0 mL) was added. The mixture was stirred at rt for 12 h and evaporated in vacuo. The
residue was dissolved in pyridine (2.0 mL) and then acetic anhydride (1.0 mL, 11 mmol) was added at
0 °C. The mixture was stirred at rt for 12 h and evaporated in vacuo. The residue was purified by
column chromatography with 1:2 AcOEt-hexane (to remove impurities) and then 2% MeOH-CHCl3 as an
eluant to give 16 (84.7 mg, 91%) as pale yellow crystals: mp 142–144 °C (from AcOEt-hexane): Rf =
0.42 (B); 1H NMR, see Table 2. Anal. Calcd for C35H41N7O15: C, 52.56; H, 5.17. Found: C, 52.39; H,
5.29.
2’-O-(2-Acetamido-2-deoxy-β-D-glucopyranosyl)-L-biopterin (2).
Compound (16) (84.0 mg, 0.105 mmol) was dissolved in MeOH (8.0 mL) and 28% aqueous ammonia
solution (3.0 mL) was added. The mixture was stirred at at for 24 h and evaporated in vacuo. The
residue was dissolved in DMF (2.0 mL) and DBU (0.10 mL, 0.64 mmol) was added. The mixture was
stirred at rt for 12 h and evaporated in vacuo. The residue was washed with CHCl3 and dried under
reduced pressure to give 2 (43.8 mg, 95%) as pale yellow crystals: mp 192–195 °C (from H2O–2-PrOH):
Rf = 0.38 (C). Anal. Calcd for C17H24N6O8: C, 46.36; H, 5.49. Found: C, 46.19; H, 5.58.
Di-N2:2’-O-acetyl-1’-O-(2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl)-L-biopterin (18).
By use of the same procedures for 2 from 16, a 45:55 mixture (130 mg) of 11a and 11b was converted
into a mixture of 1’-O-(β-D-glucopyranosyl)-L-biopterin (17) and 1b. The mixture was acetylated with
acetic anhydride (1.0 mL, 11 mmol) in pyridine (2.0 mL) to give an inseparable mixture (53 mg) of 18
and 15 as a pale yellow syrup: overall yield 81 % from 11a and 11b; Rf = 0.50 (B); 1H NMR for 18, see
Table 2.
N2-Acetyl-1’,2’-bis-O-(2,3,4,6-tetra-O-acetyl-β-D-glucopyranosyl)-L-biopterin (20).
By use of the same procedures for 2 from 16, compound (13a) (43.2 mg) was converted into 1’,2’-bis-O-
(β-D-glucopyranosyl)-L-biopterin (19) (20.3 mg, 92%) as a yellow powder: Rf = 0.19 (C). By use of the
same procedures for 14 from 1b, compound (13b) (20.8 mg) was also converted into 19 (6.4 mg, 88%).
Compound (19) (25.5 mg) was acetylated with acetic anhydride (0.40 mL, 4.2 mmol) in pyridine (1.0
1
mL) to give 20 (38.7 mg, 91%) as a pale yellow syrup: Rf = 0.47 (B); H NMR, see Table 2. Anal.
Calcd for C39H49N5O22: C, 49.84; H, 5.26. Found: C, 50.02; H, 5.29.
ACKNOWLEDGEMENTS
We are grateful to the SC-NMR Laboratory of Okayama University for the NMR measurements.

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HETEROCYCLES, Vol. 67, No. 1, 2006
REFERENCES AND NOTES
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