Synthesis and Structure Elucidation of (R)-3-(2′-hydroxyprop-1-yl) adenine

Article abstract of DOI:10.1002/jhet.2139

(R)-3-(2′-hydroxyprop-1-yl) adenine 4 was obtained by alkylation of adenine 1 with R-propylene carbonate 2 in the presence of pulverized sodium hydroxide in the synthetic process of tenofovir. The elucidation of the structure of 4 was confirmed by single crystal X-ray diffraction and NMR experiments such as 1D ¹H, ¹³C, and DEPT, as well as 2D COSY, HSQC, and HMBC spectra.

Full text of DOI:10.1002/jhet.2139

March 2016
Synthesis and Structure Elucidation of
579
(R)-3-(2′-hydroxyprop-1-yl) adenine
Yang Gao,a,b Jia-Liang Zhong,a,b Tai-Zhi Wu,a,b Lin-Yong Jin,a,b and Fu-Li Zhanga,b
*
aDepartment of Chemical Pharma, Shanghai Institute of Pharmaceutical Industry, Shanghai 200437, China
bState Key Laboratory of New Drug and Pharmaceutical Process, Shanghai Institute of Pharmaceutical Industry, Shanghai
200437, China
*
E-mail: zhang.fuli@sipi.com.cn
Additional Supporting Information may be found in the online version of this article.
Received March 20, 2013
DOI 10.1002/jhet.2139
Published online 8 May 2015 in Wiley Online Library (wileyonlinelibrary.com).
(R)-3-(2′-hydroxyprop-1-yl) adenine 4 was obtained by alkylation of adenine 1 with R-propylene
carbonate 2 in the presence of pulverized sodium hydroxide in the synthetic process of tenofovir. The
elucidation of the structure of 4 was confirmed by single crystal X-ray diffraction and NMR experiments
1
such as 1D H, 13C, and DEPT, as well as 2D COSY, HSQC, and HMBC spectra.
J. Heterocyclic Chem., 53, 579 (2016).
INTRODUCTION
(Scheme 1). This compound had a different HPLC retention
time with that of the unknown impurity which showed that
they were not the same compound.
(R)-9-(2′-hydroxyprop-1-yl) adenine (3) is an important
intermediate in the synthetic process of tenofovir disoproxil
fumarate, a nucleoside analog used in the treatment of
HIV/AIDS and hepatitis B. An unknown impurity was found
in an approximately 10:1 product: impurity ratio in the
synthesis of compound 3 by alkylation of adenine 1 with
R-propylene carbonate 2 in the presence of pulverized
sodium hydroxide.
The compound 3 with the impurity were synthesized and
separated according to the given procedure reported by
1
Raić [1]. The melting point and the data of H NMR and
13C NMR of the impurity were in accord with these
reported by Raić. According to the reported assignment
1
by Raić, the assignment of C-2 (152.70 ppm, JCH = 197.5
Hz) relatively downfield to C-8 (144.42 ppm, 1JCH = 207.1
Hz) was in accordance with the literature data for adenine,
purine, and other related molecules [4]. What is more,
doublet for C-2 and doublet of triplets for C-8 were
observed in the gated decoupled spectra. The aforementioned
data were in agreement with the N-7 isomer. However, the
HMBC spectrum was given displaying cross peak between
Raić [1] had stated that the impurity was N-7 isomer (5) by
1
means of one-dimensional and two-dimensional H-NMR
and 13C-NMR spectroscopy. The melting point of the
impurity reported by Raić was 275–277°C.
Ripin [2] has reported the regioisomer was N-(Purine-6-yl)-
1-amino-2-propanol (6). The compound 6 was synthesized by
reaction of 6-chloropurine with 2-amino-1-propanol [3]
© 2015 HeteroCorporation

580
Y. Gao, J-L. Zhong, T-Z. Wu, L-Y. Jin, and F-L. Zhang
Vol 53
Table 1
Crystal data and refinement parameters for compound 4.
Scheme 1. Synthetic route of 6-NH2 substituted impurity 6.
Molecular formula
C8H11N5O
Molecular weight
193.22
Temperature
Wavelength
296(2) K
1.54178 Å
Crystal system
Space group
Orthorhombic
C 2 2 21
C-4 and H-1′ rather than between C-5 and H-1′, and one can
also see the strong cross peaks between C-5 and H-2 and be-
tween C-6 and H-8. These results were unreasonable with the
N-7 isomer. In order to solve the aforementioned problem, the
single crystals were grown, and we have found that N-9 and
N-3 isomers 3 and 4 were formed, respectively, but not the
N-7 one. We report here the single crystal X-ray diffraction data
and the assignment of 1H-NMR and 13C-NMR spectra. (Fig. 1)
a
b
c
7.636(1) Å
8.310(1) Å
28.937(6) Å
1836.1(6) Å3
8
Volume
Z
Calculated density
Absorption coefficient
F(000)
1.398 g/cm3
0.826 mmÀ1
816
Crystal size
0.15 × 0.12 × 0.10 mm
Theta range for data collection 3.05–67.34 deg.
Limiting indices
À8 < =h < =8, À9 < =k < =9,
À34 < =l < =34
6279/1617 [R(int) = 0.0162]
99.0%
RESULTS AND DISCUSSION
Reflections collected / unique
Completeness to theta = 67.34
Absorption correction
The grown crystal was analyzed by single crystal X-ray
diffraction method. The X-ray data were collected at room
temperature by Bruker SMART APEX-II (Bruker Com-
pany, Karlsruhe, Switzerland) using CuKa radiation
(λ = 1.54178 Å). Refinement was conducted by full-matrix
least-squares calculation. Direct method (Shelxs97) and Fou-
rier technique were used to solve the crystal structure. The
crystallographic data and experimental information
are summarized in Table 1. CCDC927180 contains the
supplementary crystallographic data for this article. The
data can be obtained free of charge from the Cambridge
Crystallographic Data Centre via www.ccdc.cam.ac.uk/
community/requestastructure. Figure 2 gives a perspective
view of the molecular structure of compound 4. The
2′-Hydroxypropyl group is located at N-3 atom of adenine ring.
The assignments of the carbon and hydrogen atoms were
finally confirmed by the single crystal structure and one-
Semi-empirical from
equivalents
Max. and min. transmission
Refinement method
0.9220/0.8861
Full-matrix least-squares on F2
1617/0/127
Data/restraints/parameters
Goodness-of-fit on F2
1.101
Final R indices [I > 2σ (I)]
R indices (all data)
Absolute structure parameter
Largest diff. peak and hole
R1 = 0.0308 wR2 = 0.0813
R1 = 0.0310 wR2 = 0.0816
0.1(3)
0.243/À0.189 e.AÀ3
1
dimensional and two-dimensional H-NMR and 13C-NMR
spectra such as DEPT, HSQC, and HMBC (Tables 2 and 3).
The part of HMBC spectrum (Fig. 3) of 4 is given
displaying the coupling between C-4 and H-1′. In a hypo-
thetical N-7 regioisomer, the coupling between the C-5 and
H-1′ rather than between C-4 and H-1′ would be expected.
One can see the strong coupling between C-5 and δ7.74
and between C-6 and δ8.20, which shows that H-2
(8.20 ppm) is relatively downfield to H-8 (7.74 ppm). The
corresponding C-2 (144.42 ppm) and C-8 (152.70 ppm)
Figure 2. Molecule three-dimensional structure of compound 4. [Color
figure can be viewed in the online issue, which is available at
wileyonlinelibrary.com.]
Figure 1. Structures of compounds 4, 5 and 6.
Journal of Heterocyclic Chemistry
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March 2016
Synthesis and Structure Elucidation of (R)-3-(2′-hydroxyprop-1-yl) adenine
581
Table 2
1H-NMR chemical shifts (δ/ppm) of compounds 3 and 4.
Assignment
3
4
2-H
8-H
8.13(s)
8.03(s)
7.06(s)
4.97(s)
4.10 a)
3.99
8.20(s)
7.74(s)
7.86(br)
5.15(d, J = 4.4 Hz)
4.35(dd, J = 13.2 Hz, J = 3.2 Hz)4.05(dd, J = 13.2 Hz, J = 8.4 Hz)
4.13(m)
1.14(d, J = 6.0 Hz)
NH2
OH
1′-H
2′-H
–CH3
1.05(d, J = 5.6 Hz)
Table 3
13C-NMR chemical shifts(δ/ppm)a , DEPT, HSQC, and HMBC of compound 4.
Assignment
δC
DEPT
HSQC
HMBC
6-C
8-C
4-C
2-C
5-C
2′-C
1′-C
3′-C
155.51
152.70
150.39
144.42
120.89
64.12
q
t
q
t
q
t
s
p
H-2(strong), H-8
7.74
H-2, H-8, H-1′
H-1′
H-2, H-8(strong)
H-1′, H-3′,OH
H-2, H-2′, H-3′, OH
H-1′, OH
8.20
4.13
4.34, 4.05
1.12– 1.14
56.74
21.33
was considered essential, as natural purine nucleosides
are substituted at N-9 [5]. Yet, the alkylation pattern
(N-3: N-7: N-9 site selectivity) for the ambident adenine
nucleophile in DMF is dependent on the nature of SN2
transition state involved with different alkylation agents
[6]. Imoto [7] has reported on simultaneous formation
of the N-9 and N-3 regioisomers in the alkylation
reaction of adenine and hypoxanthine. Rasmussen [6]
had investigated the alkylation of adenine with a variety
of alkylating agents under the normal alkaline conditions
most frequently used for the synthesis of 9-alkylated
adenine systems. The N-9 alkylated products were
clearly dominant, but substantial amounts of N-3 isomers
and small amounts of the corresponding N-7 alkylated
adenines were also detected. No N-1 alkylation was
found. On the other hand, Raić [8] has reported on
concurrent formation of N-9 and N-7 regioisomers in
the alkylation reactions of 6-(N-pyrrolyl) purine, as
Figure 3. A part of the HMBC spectrum of 4.
1
confirmed by H-NMR and 13C-NMR spectroscopy.
On the basis of single crystal X-ray diffraction analysis,
it shows that (R)-3-(2′-hydroxyprop-1-yl) adenine 4 with 3
were obtained by alkylation of adenine 1 with R-propylene
carbonate 2 in the presence of pulverized sodium hydrox-
ide. With knowing the structure of compound 4, we have
confirmed the assignments of all carbon and hydrogen
atoms by the one-dimensional and two-dimensional
1H-NMR and 13C-NMR spectra such as DEPT, HSQC, and
HMBC. In N-3 isomer H-2 (8.20 ppm) is more deshielded
than H-8 (7.74 ppm). The corresponding C-8 (152.70 ppm)
can be determined by HSQC spectra. The cross peak
between C-2 (144.42 ppm) and H-1′ can be seen in the
HMBC spectra. One can see the strong cross peaks
between C-5 and δ7.74 and between C-6 and δ8.20, which
is also in agreement with the aforementioned speculation.
In the synthesis of purine nucleosides, four regioisomers
with acyclic substituents at N-9, N-7, N-3, or N-1 may be
theoretically obtained. However, the N-9 regioselectivity
of the coupling of the heterocycle with the acyclic residue
Journal of Heterocyclic Chemistry
DOI 10.1002/jhet

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Y. Gao, J-L. Zhong, T-Z. Wu, L-Y. Jin, and F-L. Zhang
Vol 53
containing the solution was placed in a clean and quiet
environment before the formation of crystal.
is relatively downfield to C-2 (144.42 ppm), contrary to the
assignments of the spectra reported by Raić. It is also con-
trary to the literature data for adenine, purine, and other re-
lated molecules [4] where C-2 is always relatively
downfield to C-8. It is difficult to explain this abnormal
phenomenon and more researches are needed to be performed.
N-(Purine-6-yl)-1-amino-2-propanol (6).
To a 250 mL,
three necked, round bottom flask equipped with condenser was
added 6-chloropurine (10.3 g, 0.066 mol), 1-amino-2-propanol
(6.4 g, 0.085 mol), sodium carbonate (3.5 g, 0.033 mol), sodium
hydroxide (1 pellet), and water (120 mL). The mixture was
heated at reflux for 6 h and then cooled overnight. The
precipitated solids were isolated, washed with water, and dried
to yield 6 (9.7 g, 76.2%). The crude product was crystalized
from water (100 mL), and the pure 6 was obtained (7.8 g,
61.2%), mp 232.6–232.7°C. 6 elutes at a retention time of
9.1 min, while 4 elutes at a retention time of 3.5 min.
EXPERIMENTAL
Melting points were determined on a WRS-1B Digital
Melting-point Apparatus (Shanghai YiCe Instrument Co., LTD
1
China). The H-NMR and 13C-NMR spectra were recorded on a
Bruker 400 instrument (Bruker Company, Karlsruhe, Switzerland).
Chemical shifts were reported in parts per million from internal
TMS standard. The solvent for NMR spectra was DMSO. The
single crystal data were recorded on a Bruker SMART APEX-II
instrument (Bruker Company, Karlsruhe, Switzerland), and Shelxs97
was used to analyze the single crystal structure.
Analytical HPLC test method. C18 column (4.6 × 250 mm,
5 μm particles), isocratic at 1.0 mL/min, mobile phase 900 mL of
10mM phosphate buffer adjusted to pH5.5 mixed with 100mL
1:1 acetonitrile/methanol.
(R)-3-(2′-hydroxyprop-1-yl) adenine (4).
A mixture of
REFERENCES AND NOTES
adenine (1) (5.50 g, 40 mmol), pulverized sodium hydroxide
(0.12 g, 3 mmol), and R-propylene carbonate (2) (5.70 g,
56 mmol) in dry DMF (25 mL) was heated for 8 h under reflux.
The reaction mixture was cooled to room temperature and toluene
(30 mL) was added. The solid was filtered by suction and washed
with toluene (5mL). The yield was 7.07 g (90%), mp 193°C. The
crude product was separated by double column chromatography
with CH2Cl2–MeOH in volume ratio (v/v) 4:1 as the solvent.
The (R)-9-(2′-hydroxyprop-1-yl) adenine (3) was obtained as a
major product 5.65g (80%), white powder, mp 196.9–197.1°C,
RF ~ 0.5 (CH2Cl2–MeOH = 4:1). The pure N-3 isomer 4 was
obtained as a minor product 0.56g (8%), white powder, mp
272.9–273.3°C, UV (CH2CH3OH): λ max 278 mμ. RF ~ 0.2
(CH2Cl2–MeOH = 4:1), 1H-NMR (Table 2), 13C-NMR (Table 3).
Single crystal growth of (R)-3-(2′-hydroxyprop-1-yl)
adenine (4). The single crystal of (R)-3-(2′-hydroxyprop-1-yl)
adenine (4) was grown by solvent evaporation growth technique.
Methanol has been chosen as the solvent for the growth. The tube
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Journal of Heterocyclic Chemistry
DOI 10.1002/jhet