JOURNAL OF CHEMICAL RESEARCH 2016 359
Table 1 Optimisation of the reaction conditions for the base-catalysed synthesis of cytidine-5’-phosphomorpholide 2 from cytidine 6 and
a,c
morpholinophosphordichlorodate 7 (Scheme 1, route b)
b
Entry
n6:n7
Solvent
Temp/°C
Time/h
Base
Yield/%
1
2
3
4
5
6
7
8
9
1:1
1:2
1:3
1:2
1:2
1:2
1:2
1:2
1:2
1:2
1:2
1:2
MeCN
MeCN
MeCN
MeCN
MeCN
MeCN
MeCN
MeCN
dioxane
pyridine
DMF
r.t.
r.t.
r.t.
50
0
0
0
0
0
1
1
1
1
1
2
2
2
2
2
2
3
TEA
TEA
TEA
TEA
TEA
32
46
41
39
63
72
81
68
64
76
56
73
TEA
DMAP
pyridine
DMAP
DMAP
DMAP
DMAP
1
0
0
0
0
1
1
12
MeCN
a
Reaction conditions: morpholinophosphorodichloridate 7 (1, 2 or 3 equiv.) was added slowly to a stirred mixture of cytidine 6 (1 equiv.) and an organic base in various solvents for various
times.
b
Isolated yield.
c
TEA = triethylamine, DMAP = 4-dimethylaminopyridine.
When the reaction of 6 and 7 (1:1) was conducted in MeCN
10 mL) at room temperature for 1 h using triethylamine TEA
as the base, 2 was obtained in 32% yield (Table 1, entry 1). The
yield was increased to 46% when 2 equiv. of 7 was employed
multiplet (m). High resolution mass spectra were obtained with a 3000
mass spectrometer, using a Waters Q-TofMS/MS system. All reagents
and solvents were purchased from commercial sources and purified
before use.
(
(
4
(
entry 2). Increasing the amount of 7 led to a lower yield of
1% due to the side reactions of the 2’ or 3’ hydroxyl groups
entry 3). A lower yield was also observed at 50 °C and this led
Cytidine-5’-phosphomorpholide (2): Cytidine (0.243 g, 1.0 mmol)
and DMAP (0.183 g, 1.5 mmol) in MeCN (10 mL) were stirred
slowly and cooled to 0 °C, and 7 (2.0 mmol) was added slowly. The
mixture was heated to 50 °C and kept at this temperature for 2 h.
The solvent was removed in vacuo and the residue was purified by
recrystallisation from EtOH to give 2 as a white semi-solid (0.318 g);
us to conduct the reaction at a lower temperature to improve
the yield (entry 4). Gratifyingly, the reaction at 0 °C gave a
yield of 63% (entry 5). Doubling the reaction time improved
the yield to 72% (entry 6). Other bases such as DMAP or
pyridine were assessed and DMAP gave the best yield of 81%
1
yield 81%; m.p. 62–64 °C; H NMR (400 MHz, DMSO-d ) δ 8.43 (d,
6
J = 7.6 Hz, 1H), 7.39 (s, 2H), 7.19 (d, J = 7.6 Hz, 1H), 5.77 (d, J = 2.8
Hz, 1H), 5.51 (d, J = 4.8 Hz, 1H), 5.18 (t, J = 5.2 Hz, 1H), 5.08 (d, J =
5.6 Hz, 1H), 3.76–3.71 (m, 1H), 3.61–3.56 (m, 1H), 3.45–3.42 (m, 4H),
(
entry 7). Other solvents such as dioxane, pyridine and DMF
were also examined but failed to give better results (entries
9
7
13
−11). Extending the reaction time to 3 h reduced the yield to
3% (entry 12). Thus, the optimised reaction conditions were
3.03–2.99 (m, 4H); C NMR (100 MHz, DMSO-d ) δ 166.5, 157.8,
6
145.9, 141.6, 88.1, 86.2, 74.3, 70.7, 65.1, 65.0, 60.3, 60.2; HRMS calcd
+
determined as described in entry 7.
for C H N O P [M + H] 393.1170, found: 393.1172.
13
22
4
8
The traditional method for the synthesis of 2 with POCl gave
CDP-choline (1): 2 (0.392 g, 1.0 mmol) was added to MeOH (10 mL)
followed by the addition of 3 (0.310 g, 1.2 mmol) and was stirred at
room temperature for 10 min. Then 98% H SO (0.005 mL, 10 mol%)
3
only a 40% yield and needed three steps. We surmised that the
better yield of our method was due to the good selectivity for 5’
hydroxylation rather than 2’ or 3’ hydroxylation because of the
steric hindrance of morpholino group.
2
4
was added. The mixture was kept at 50 °C for 3 h. The solvent was
removed in vacuo and the residue was purified by recrystallisation
1
With 2 in hand, next, we conducted the reaction of 2 and 3 to
give the final product CDP-choline. After intensive experiment
and optimisation, CDP-choline was synthesised in 85% yield
catalysed by 10 mol% H SO at 50 °C for 3 h.
from EtOH to give 1 as a white solid (0.410 g); yield 85%. H NMR
(400 MHz, D O) δ 7.86 (s, 2H), 6.04 (d, J = 5.2 Hz, 1H), 5.91 (d,
2
J = 5.2 Hz, 1H), 4.32 (brs, 2H), 4.26–4.22 (m, 2H), 4.18 (brs, 2H), 4.11
13
(t, J = 3.2 Hz, 1H), 3.60 (t, J = 2.4 Hz, 2H), 3.14 (s, 9H); C NMR (100
2
4
To evaluate the reproducibility and the stability of the
reaction, the synthesis of 2 and CDP-choline on a larger
scale was performed. The synthesis of 2 by the selective
phosphorylation on a 200 g scale was carried out and the
reaction still worked well in 80% yield. The synthesis of CDP-
choline on a 200 g scale also gave a yield of 80%. An added
virtue is that CDP-choline could be purified by recrystallisation,
thus chromatography or a lengthy purification process was not
required, which made this route more attractive for industrial
applications.
MHz, D O) δ 166.1, 157.7, 141.5, 96.6, 89.3, 82.6, 74.1, 69.3, 66.0, 65.9,
2
+
64.8, 59.9, 54.0; HRMS calcd for C H N O P [M + H] 489.1146,
14
27
4
11
2
found: 489.1140.
For the synthesis of 1 from 6 on a 200 g scale, see the Electronic
Supplementary Information (ESI).
We are grateful to the Key Scientific Research Projects in the
Universities of Henan Province (16A150042) and Science and
Technology Innovation Fund of Xinxiang University (15ZP06)
for funding.
Experimental
Melting points were recorded with a micro melting point apparatus
and are uncorrected. NMR spectra were recorded on a Bruker AV/400
Electronic Supplementary Information
The ESI is available through:
stl.publisher.ingentaconnect.com/content/stl/jcr/supp-data
1
13
spectrometer (400 MHz for H NMR and 100 MHz for C NMR).
Chemical shifts δ are given in ppm relative to tetramethylsilane as
1
13
Paper 1603957 doi: 10.3184/174751916X14628025243831
Published online: 17 May 2016
internal standard, or residual DMSO-d or D O for H or C NMR
6
2
spectroscopy. Multiplicities are reported as follows: singlet (s),
doublet (d), doublet of doublets (dd), triplet (t), quartet (q) and
Xia, Ran
