SYNTHEIS OF N-(3-AZIDO-2-HYDROXYPROPYL), N-(3-PHTHALIMIDO-2-HYDROXYPROPYL) AND N-(3-AMINO-2-HYDROXYPROPYL)DERIVATIVES OF HETEROCYCLIC BASES

Article abstract of DOI:10.1135/cccc19941153

Alkylation of heterocyclic bases with azidomethyloxirane (I) under basic catalysis with potassium or cesium carbonate afforded N-(3-azido-2-hydroxypropyl) derivatives II.Hydrogenation of these compounds over palladium on carbon gave the corresponding 3-am

Full text of DOI:10.1135/cccc19941153

N-(3-Amino-2-hydroxypropyl) Azaheterocycles
1153
SYNTHESIS OF N-(3-AZIDO-2-HYDROXYPROPYL), N-(3-PHTHALIMIDO-
2-HYDROXYPROPYL) AND N-(3-AMINO-2-HYDROXYPROPYL)
DERIVATIVES OF HETEROCYCLIC BASES*
Maria SPASSOVA**, Hana DVORAKOVA, Antonin HOLY, Milos BUDESINSKY
and Milena MASOJIDKOVA
Institute of Organic Chemistry and Biochemistry,
Academy of Sciences of the Czech Republic, 166 10 Prague 6, The Czech Republic
Received November 1, 1993
Accepted December 8, 1993
Alkylation of heterocyclic bases with azidomethyloxirane (I) under basic catalysis with potassium or
cesium carbonate afforded N-(3-azido-2-hydroxypropyl) derivatives II. Hydrogenation of these com-
pounds over palladium on carbon gave the corresponding 3-amino-2-hydroxypropyl derivatives III.
The same compounds III were prepared by alkylation of heterocyclic bases with phthalimido-
methyloxirane (VII) in the presence of cesium carbonate and subsequent reaction of the formed
N-(3-phthalimido-2-hydroxypropyl) derivatives VIII with hydrazine. The phthalimido derivatives VIII
are easily hydrolyzed already in weakly alkaline aqueous medium to give 9-[3-(o-carboxybenzoyl-
amino)-2-hydroxypropyl] derivatives IX and X.
Within the framework of investigation on preparation and biological properties of nu-
cleoside analogs we studied the preparation and properties of compounds bearing in the
side-chain amino groups in addition to the hydroxyl functionalities. Some time ago we
prepared the isomeric 9-azidohydroxypropyl derivatives of adenine and converted them
into the corresponding aminohydroxypropyl derivatives by catalytic hydrogenation1,2
(Scheme 1). It appeared that already 9-(3-azido-2-hydroxypropyl)adenine (IIa) ex-
hibited marked mutagenic effect on Salmonella typhimurium3.
In the light of this finding and of the already known mutagenicity of related 1-azido-
propane-2,3-diol4 we decided to study how the heterocyclic base affects the mutagenic
activity of the 3-azido-2-hydroxypropyl derivatives II. However, the main stimulus for
seeking a general approach to these compounds was the prospect of their further syn-
* This paper was in part published in a preliminary form: Spassova M., Holy A.: Collect. Czech. Chem.
Commun. 55, Special Issue No. 1, 65 (1990).
**On leave from the Institute of Molecular Biology, Bulgarian Academy of Sciences, 1113 Sofia,
Bulgaria.
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SCHEME 1
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N-(3-Amino-2-hydroxypropyl) Azaheterocycles
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thetic utilization, e.g. in the synthesis of 3-amino-2-hydroxypropyl derivatives III or
3-amino-2-phosphonomethoxypropyl derivatives2 IV, analogs of antivirals of the HPMP
series (ref.5) or NAD analogs6.
Until now, synthetic methods leading to azido derivatives of acyclic nucleoside
analogs of the type II consisted either in nucleophilic substitution of the tosyl group on
the alkyl chain1, 7 or in alkylation of the heterocyclic base with a protected derivative of
1-azido-2-hydroxy-3-propyl tosylate2. The former method requires conversion of hy-
droxyl into a sulfonate group; in spite of this it appears to be more advantageous than
the direct conversion of the hydroxyl into the azide functionality by reaction with tri-
phenylphosphine, lithium azide and carbon tetrabromide8,9 or tetraiodide10, as described
for syntheses of acyclic azidothymidine analogs, or by the Mitsunobu reaction of the
hydroxyl with triphenyl phosphine, DEAD and azoimide11. The use of protected
3-azido-2-hydroxypropyl tosylate4 for the alkylation of bases finds its application
mainly in the preparation of optically active forms of 3-amino-2-hydroxypropyl deriva-
tives6 III. In the present study we make use of alkylation of the heterocyclic base with
azidomethyloxirane (I) which is readily accessible from epichlorohydrin6. The reaction
takes place under catalysis with potassium or cesium carbonate in dimethylformamide
as solvent.
As expected, the reaction proceeded smoothly with adenine, its 3-deaza analog,
6-chloropurine, 6-methylthiopurine and 2,6-diaminopurine, affording predominantly
the 9-isomers IIa, IIc, IIe, IIj and IIb, respectively. High regiospecificity in the alkyla-
tion of the base with oxirane I was also found for 4-aminopyrazolo[5,6-d]pyrimidine: in
this case the alkylation takes place predominantly on N7, leading to compound IIk. Also
4-methoxy-2-pyrimidone and 4-methoxy-5-methyl-2-pyrimidone afforded the expected
N1-isomers IIl and IIm; the former was ammonolyzed to give the cytosine derivative IIp
which was also prepared by direct alkylation of cytosine with compound I.
A less unequivocal reaction course under the conditions described was observed with
2-amino-6-chloropurine which afforded, in addition to the N9-isomer IIf, some N7-alky-
lation product. As could be expected, alkylation of 2-mercaptoadenine with oxirane I
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Spassova, Dvorakova, Holy, Budesinsky, Masojidkova:
afforded compound V as the primary product of alkylation on the sulfur atom, together
with the N9,S-disubstituted derivative VI.
Acid hydrolysis of 6-chloro derivatives IIe and IIf afforded 9-(3-azido-2-hydroxy-
propyl) derivatives of hypoxanthine and guanine IIh and IIi, respectively. They were
reacted with thiourea to give the respective 6-mercaptopurine and 6-thioguanine deriva-
tives IIr and IIs. In the same manner, the corresponding derivatives of 4-methoxy-2-
pyrimidone and 4-methoxy-5-methyl-2-pyrimidone IIl and IIm were converted into the
uracil and thymine derivatives IIn and IIo.
The azido compounds II were easily and in good yields converted into 3-amino-2-
hydroxypropyl derivatives III by hydrogenation, e.g. over a palladium catalyst in ethanol.
The only exception was the preparation of 2-amino-6-chloropurine derivative IIIf in
which the reduction was accompanied by reductive dehalogenation and the resulting
product was 9-(3-amino-2-hydroxypropyl)-2-aminopurine (IIIg).
The structure of azido derivatives II and amino derivatives III was proved by the
1
usual techniques, including UV and H NMR spectroscopy. In the determination of
structures of the bases in the derivatives II, III, V and VI the limited information from
1
the H NMR spectra (sharp signals of heteroaromatic protons and broad signals of NH
or OH protons, see Table I) was extended by that from the 13C NMR data, particularly
as concerns quaternary carbon atoms (Table II). The structural assignment of carbon
atom signals was based on the experimental distinction of tertiary and quaternary atoms
and their comparison with the spectra of the free bases. In contrast to the very broad
carbon signals (halfwidths 10 – 100 Hz) in the spectra of free purine bases (particularly
of C-4 and C-5), the corresponding signals in our azido and amino derivatives II and III
were narrow (1 – 2 Hz), reflecting evidently the absence of the N-7 ↔ N-9 tautomer-
ism in the substituted compounds. The only exception was compound V which, how-
ever, is the product of S-alkylation in position 2 (vide infra) and in which the
mentioned tautomerism can exist. The presence of 3-azido- or 3-amino-2-hydroxy-
propyl groups was clearly demonstrated by signals in the region of aliphatic carbon
atoms in 13C NMR spectra (Table II). Whereas the signals of C-2′ and C-3′ reflect first
of all character of substituents in these positions and differ only little within the series
of azido or amino derivatives, signals due to C-1′ are markedly influenced by the
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N-(3-Amino-2-hydroxypropyl) Azaheterocycles
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N-(3-Amino-2-hydroxypropyl) Azaheterocycles
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character of the base: for purine derivatives they appear in the region δ 46.2 – 48.3, for
pyrimidine derivatives they range from δ 51.1 to 53.1. The considerable upfield shifts
of C-1′ signals in the spectra of V and VI (δ 34.9) indicate bonding to the sulfur atom.
The position of the alkyl substituent – N-9 in the purine and N-1 in the pyrimidine
derivatives – was determined by comparison with 13C NMR data of the corresponding
free bases obtained under identical conditions. For most of the studied compounds II,
III and VI we observed a marked alkylation shift of the C-8 signal (about 2 ppm down-
field) for purine derivatives (and their deaza analogs) and of the C-6 signal (about 4.5
ppm downfield) for pyrimidine derivatives as compared with the free bases, in accord
with the N-9 alkylation of purines12 and N-1 alkylation of of pyrimidines13. Opposite
alkylation effects – downfield shifts of the C-5 and C-6 signals (about 7.5 and 5.5 ppm)
and upfield shifts of C-4 and C-8 signals (about −7 and −1.5 ppm) – were found for
hypoxanthine and 6-mercaptopurine derivatives. Similar effects connected with N-9
alkylation may be also expected with guanine and 6-thioguanine derivatives for which,
however, reference data for the free bases are lacking (they were not found in the
literature and also our attempted measurements failed). The reason is apparently the
very low solubility and extremely broad signals due to lactam–lactim or thione–thiol
tautomerism11. The five-spin system of hydrogen atoms in the 3-azido- or 3-amino-2-
1
hydroxypropyl substituent leads to more or less complex H NMR spectra (2nd order
effects). In some cases the chemical shifts and coupling constants had to be determined
by simulation of the spectra and in the case of compounds IIIg and VI a complete
analysis was not possible. The different vicinal coupling constants for both methylene
groups in the whole series of the compounds studied (Table I) indicate a limited flexi-
bility of the three-carbon alkyl which is more expressive around the C-1′−C-2′ bond
(average J(H-1′,H-2′) = 3.7 and 8.0 Hz and J(H-2′,H-3′) = 3.3 and 6.6 Hz). Under the
usual simplifying assumption of staggered rotamers, the J values show a markedly dif-
ferent population of the two possible rotamers with an anti-relation of hydrogen atoms.
Since the stereospecific assignment of protons of the methylene groups is not possible,
we cannot determine the predominant anti-rotamer from the J-values. Nevertheless, we
may assume that, for steric reasons, the most populated rotamer will be that with anti-
arrangement of the bulkiest substituents, i.e. the base moiety and the C(3′)H2NR group
attached to the C-1′−C-2′ bond and the groups C(1′)H2−B and NR on the C-2′−C-3′
bond.
As an alternative synthetic approach to amino derivatives III we also studied the
possible utilization of the easily accessible N-(3-phthalimido-2-hydroxypropyl) deriva-
tives VIII which could be converted into amino derivatives III, e.g. by hydrazinolysis.
They were synthesized in quantitative yields during several hours by reaction of the
corresponding bases with the commercially accessible N-(2,3-epoxypropyl)phthalimide
(VII) in the presence of catalytic amount of cesium carbonate in dimethylformamide; in
the case of purine derivatives (adenine, 2,6-diaminopurine and 2-amino-6-chloropurine)
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Spassova, Dvorakova, Holy, Budesinsky, Masojidkova:
principal amounts of the pure N9-isomers (VIIIa, VIIIb, VIIIf) were obtained by boiling
the crude reaction mixture with methanol and filtration while hot (Scheme 2). Analo-
gously, alkylation of cytosine with oxirane VII afforded predominantly the N1-isomer
VIIIp without any noticeable O2-alkylation.
SCHEME 2
Phthalimido compounds VIII were smoothly converted to amino derivatives III by
reaction with 98% hydrazine hydrate in boiling ethanol. (The recommended use of
N-methylhydrazine14 was not successful.) In this manner we prepared derivatives of
adenine IIIa and 2,6-diaminopurine IIIb which were identical with samples synthesized
from the azides II. Since the reaction of phthalimido derivatives with hydrazine is suf-
ficiently rapid, even in the case of the cytosine derivative VIIIp there was no undesired
transformation of the cytosine ring15 and the synthesized compound IIIp was identical
(HPLC) with that obtained by the former synthetic way.
In this context, synthesis of the guanine derivative IIIi represented a difficult metho-
dological problem. For known reasons, direct alkylation of guanine with oxirane VII
was of no advantage; on the other hand, use of the azide alternative for 2-amino-6-
chloropurine with subsequent acid hydrolysis of the C−Cl bond and reduction also did
not represent the best solution.
Therefore, our synthetic goal was also to utilize the 2-amino-6-chloropurine phthali-
mido derivative VIIIf for synthesis of the guanine analog IIIi. Attempted hydrolysis of
the C−Cl bond by heating compound VIIIf with 0.4 M triethylammonium hydrogen
carbonate in dimethylformamide resulted, however, in opening the phthalimide group-
ing under formation of phthalamic acid derivative IX whereas the base remained un-
changed. The same result was obtained with the adenine derivative VIIIa. Structure of
the compounds IX and X was unequivocally proved by 13C NMR spectroscopy (see
Table IV).
The failure of an easy conversion of chloro derivative VIIIf into the guanine deriva-
tive led us to try the phthalimide method with another guanine precursor, 2-amino-6-
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N-(3-Amino-2-hydroxypropyl) Azaheterocycles
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benzyloxypurine (XI). Although this known base16 was used in studies of regiospeci-
ficity of alkylation with other reagents17, no particular attention has been paid to its
preparation. As the method of choice we have found the reaction of 2-amino-6-chloro-
purine with sodium benzylate in an aprotic solvent, toluene. As a side-product, the
reaction afforded 2-benzylamino-6-benzyloxypurine (XII); this compound was, how-
ever, practically the sole product when the reaction was conducted in dimethylforma-
mide. The alkylation of 2-amino-6-benzyloxypurine (XI) with oxirane VII proceeded
similarly as in the case of compounds VIIIa, VIIIb and VIIIf giving the product VIIIq in
a relatively high yield during 4 h. Compound VIIIq may serve as a suitable precursor
for the preparation of other compounds of the guanine series.
Both the described methods leading to 3-amino-2-hydroxypropyl derivatives of nu-
cleic bases are sufficiently general and for the given purpose they appear to be more
suitable than the originally used syntheses based on transformation of azides obtained
from the corresponding tosyl or mesyl derivatives. The phthalimide alternative in com-
bination with hydrazinolysis is thus comparable and has its substantiation, particularly
in cases where the amino group in the side-chain has to be protected in further synthetic
steps, e.g. introduction of the phosphonomethylether functionality5. The high sensitiv-
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Spassova, Dvorakova, Holy, Budesinsky, Masojidkova:
ity of N-(phthalimido-2-hydroxypropyl) derivatives VIII to hydrolysis should, of
course, be kept in mind.
The structure of compounds VIII – XII was determined by 1H and 13C NMR spectros-
copy (Tables III and IV). The proton and carbon signals, assigned similarly as in the
above-discussed compounds II and III, enabled us to prove the structure of the base.
The N-phthalimido-2-hydroxypropyl substituent was confirmed by signals of the three-
membered aliphatic fragment, a multiplet of four aromatic protons and particularly four
signals of eight sp2-carbon atoms of the symmetrical phthalimide moiety (Table IV).
Whereas compounds VIIIa, VIIIb, VIIIf, VIIIp and VIIIq exhibited alkylation effects
characteristic for N-9 substitution on the purine, or N-1 substitution on the pyrimidine
nuclei (see the discussion above), for the minor product XIII we observed markedly
different alkylation effects (for C-4 and C-8 downfield shifts of about 9 ppm and for
C-5 and C-6 upfield shifts of about −8.5 and −7 ppm) indicating N-7 alkylation18. The
number of signals in 13C NMR spectra of compounds IX and X proves a loss of sym-
metry of the phthalimide part resulting from opening the phthalimide to the phthalamic
acid. The vicinal coupling constants of the three-carbon alkyl in compounds VIII – X
indicate similar conformational properties as in the compounds II and III discussed
above.
Biological Activity
The azidohydroxypropyl and aminohydroxypropyl derivatives II and III were tested for
activity against a standard choice of DNA and RNA viruses and retroviruses in the
Laboratory of Professor E. De Clercq (Rega Institute, Catholic University, Leuven,
Belgium). Neither of them showed any marked antiviral effect. The mutagenic effect of
azido derivatives II was already reported elsewhere3.
EXPERIMENTAL
Methods. The melting points were determined on a Kofler block and are uncorrected. Solvents
were evaporated on a rotatory evaporator at 40 °C/2 kPa. Analytical samples were dried at 25 °C and
6.5 Pa for 8 h. NMR spectra were measured on Varian UNITY-200 (1H at 200 MHz and 13C at 50.3
MHz) or UNITY-500 (1H at 500 MHz and 13C at 125.7 MHz) instruments in CD3SOCD3. Chemical
shifts of protons were referenced to TMS as internal standard, chemical shifts of carbon atoms
against the solvent signal using the relationship δ(CD3SOCD3) = 39.7 ppm. In addition to the com-
mon proton-decoupled 13C NMR spectra were measured the J-modulated spectra (“attached proton
test pulse sequence”, ref.19) or proton-coupled spectra, enabling distinction of carbon signals of the
type C, CH, CH2 and CH3. UV spectra were measured on a Pye Unicam 8800 UV/VIS spectrometer;
the wavelengths of the extrema are given in nm. Thin-layer chromatography (TLC) was performed
on Silufol UV254, column chromatography on Silpearl (both Kavalier, Votice, The Czech Republic).
Solvent systems for TLC: S1 chloroform–methanol (3 : 1), S2 chloroform–methanol (4 : 1), S3
chloroform–methanol (9 : 1), S4 chloroform–methanol (95 : 5), S5 2-propanol–concentrated aqueous
ammonia–water (7 : 1 : 2). High performance liquid chromatography was performed on columns
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(250 × 4 mm or 250 × 17 mm) packed with Separon SGX C18 (5 µm, Laboratorni Pristroje, Prague,
The Czech Republic); isocratic elution (1 ml/min) with 0.05 M triethylammonium hydrogen carbo-
nate, pH 7.5 (S6) containing 5% (v/v) (S7) or 7% (S8) of acetonitrile, detection at 254 nm. Prep-
arative chromatography on C18 silica gel was performed on a column of octadecyl silica gel (30 µm,
260 ml) in water and water–methanol mixtures. Paper electrophoresis was carried out on a Whatman
3 MM paper at 40V/cm for 1 h in 0.05 M triethylammonium hydrogen carbonate, pH 7.5 (S9).
Materials and reagents. Adenine, dimethylformamide, sodium azide, cesium carbonate, epichloro-
hydrin and bromotrimethylsilane were Janssen (Belgium) products, 2-amino-6-chloropurine was ob-
tained from Mack (Germany), cytosine and phthalimidomethyloxirane from Fluka (Switzerland),
6-chloropurine from Sigma, 2, 6-diaminopurine from Kasyo (Japan), 6-methylthiopurine from Loba-
Chemie (Austria) and 4-aminopyrazolo[5,6-d]pyrimidine from Lachema (The Czech Republic).
4-Methoxy-2-pyrimidone was prepared according to a described procedure20. Dimethylformamide,
dichloromethane and acetonitrile were dried by distillation from phosphorus pentoxide and kept over
molecular sieves.
Azidomethyloxirane (I)
A solution of sodium azide (19.5 g, 0.3 mol) in water (70 ml) was added dropwise during 90 min to
a stirred solution of epichlorohydrine (25 g, 0.27 mol) in water (50 ml). After stirring for further
20 min, the bottom layer was separated and the upper one extracted with dichloromethane (3 × 40 ml).
The combined extracts and the bottom layer were dried over sodium sulfate, the solvent was evap-
orated under diminished pressure and the residue was distilled in vacuo (the receiver cooled to −60 °C);
b.p. 22 – 24 °C/13 Pa. This product was used in the alkylation reactions without further purification.
Preparation of N-(3-Azido-2-hydroxypropyl) Derivatives II. General Procedure
A mixture of the heterocyclic base (10 mmol), potassium carbonate (0.1 g), azidomethyloxirane (I,
1.44 g, 15 mmol) and dimethylformamide (25 ml) was heated under stirring at 80 – 140 °C for 5 – 8 h
(calcium chloride protecting tube), filtered while hot and the precipitate was washed with dimethyl-
formamide. The filtrate was concentrated in vacuo, the residue codistilled with toluene and extracted
with boiling chloroform. The extract was filtered, the filtrate concentrated in vacuo and the residue
chromatographed on a column of silica gel in chloroform with increasing concentration of methanol.
The product-containing fractions were combined, the solvent was evaporated in vacuo and the residue
was crystallized. Table V gives the reaction conditions and some characteristics of the prepared com-
pounds II.
9-(3-Azido-2-hydroxypropyl)hypoxanthine (IIh) and 9-(3-Azido-2-hydroxypropyl)guanine (IIi)
A solution of compound IIe (0.47 g, 2 mmol) in 80% acetic acid (50 ml) was heated at 85 °C for 6 h.
After evaporation in vacuo, the residue was codistilled with water (4 × 20 ml) and column chroma-
tographed on C18 silica gel; elution with a water–methanol gradient. The product fraction was evap-
orated and the product crystallized from ethanol. Yield 70% of compound IIh (for analysis see Table V).
Similarly, compound IIf (0.42 g, 1.5 mmol) vas converted into derivative IIi; yield 0.36 g (87%).
For analysis see Table V.
9-(3-Azido-2-hydroxypropyl)uracil (IIn) and 9-(3-Azido-2-hydroxypropyl)thymine (IIo)
A solution of compound IIl (0.255 g, 1 mmol) in 80% acetic acid (50 ml) was heated at 80 °C for 3 h.
After evaporation of the solvent in vacuo, the residue was codistilled with water (2 × 20 ml) and
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crystallized from tetrahydrofuran–light petroleum to give compound IIn in the yield of 70%. For ana-
lysis see Table V.
Similarly, compound IIm was converted into derivative IIo in 80% yield. For analysis see Table V.
9-(3-Azido-2-hydroxypropyl)cytosine (IIp)
A solution of compound IIl (1.0 g, 4.4 mmol) in 30% methanolic ammonia (35 ml) was heated in a
pressure vessel at 110 °C for 4 h. The mixture was concentrated to dryness in vacuo and the residue
was chromatographed on a column of C18 silica gel; elution with a gradient of water–methanol. The
product-containing fraction gave, after evaporation in vacuo and crystallization from methanol–ether,
compound IIp (0.74 g, 80%). For analysis see Table V.
9-(3-Azido-2-hydroxypropyl)-6-mercaptopurine (IIr) and 9-(3-Azido-2-hydroxypropyl)-6-thio-
guanine (IIs)
A solution of compound IIe (0.47 g, 2 mmol) and thiourea (0.30 g, 4 mmol) in ethanol (25 ml) was
refluxed for 5 h. After evaporation of the solvent in vacuo, the residue was chromatographed on a
preparative loose layer of silica gel (50 × 18 × 0.4 cm) in chloroform–methanol (9 : 1). The product
band was eluted with methanol and the residue crystallized from ethanol to give 0.36 g (72%) of
compound IIr. For m.p. and analysis see Table V.
Similarly, reaction of compound IIf (0.84 g, 3 mmol) with thiourea (0.45 g, 6 mmol) in ethanol
(50 ml) afforded compound IIs; yield 0.63 g (79%). For m.p. and analysis see Table V.
2-(3-Azido-2-hydroxypropyl)thioadenine (V) and 2,9-Bis(3-azido-2-hydroxypropyl)thioadenine (VI)
Azidomethyloxirane (I; 1.98 g, 22.5 mmol) was added at 100 °C in portions to a stirred suspension
of 2-mercaptoadenine (2.51 g, 15 mmol) and potassium carbonate (0.15 g) in dimethylformamide
(60 ml). The mixture was refluxed for 10 h (calcium chloride protecting tube) and filtered while hot.
The filtrate was concentrated in vacuo and the residue extracted with chloroform (200 ml). The chlo-
roform extract was filtered through Celite and the solvent was evaporated in vacuo. The residue was
chromatographed on silica gel in chloroform (elution with chloroform–methanol 9 : 1) to give a mix-
ture of both products which were further separated on a column of octadecyl silica gel (200 ml) by
elution with a gradient of water–methanol (0 – 100%). Crystallization from tetrahydrofuran–methanol
(1 : 5) afforded 1.43 g (36%) of compound V, m.p. 218 °C, RF 0.51 (S3). For C8H10N8OS (266.2)
calculated: 36.09% C, 3.79% H, 42.09% N; found: 36.05% C, 3.69% H, 41.42% N. Further elution
afforded 1.73 g (32%) of compound VI, m.p. 145 °C (tetrahydrofuran–methanol 1 : 5). For
C11H15N11O2S2 (365.3) calculated: 36.16% C, 4.13% H, 42.18% N; found: 36.04% C, 4.01% H,
42.04% N.
Preparation of N-(3-Amino-2-hydroxypropyl) Derivatives III by Hydrogenation of Azido
Compounds II. General Procedure
Compound II (5 mmol) was hydrogenated in 50% aqueous ethanol (120 ml) over 10% palladium on
carbon (0.67 g) at room temperature overnight. After filtration through Celite and washing the solid
with 50% aqueous ethanol (25 ml), the filtrate was made alkaline with ammonia, concentrated in
vacuo and the residue was crystallized from ethanol–ether. Compound IIIg was isolated by chroma-
tography on C18 silica gel in 50% aqueous methanol. The yields and properties of the thus-obtained
compounds III are given in Table VI.
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N-(3-Amino-2-hydroxypropyl) Azaheterocycles
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General Procedure for Preparation of Phthalimido Derivatives VIII
N-(2,3-Epoxypropyl)phthalimide (VII; 4.8 g, 24 mmol) was added to a mixture of the corresponding
base (20 mmol), cesium carbonate (0.4 g, 1.2 mmol) and dimethylformamide (100 ml). The mixture
was heated at 120 °C for 6 h under stirring and protection from moisture (calcium chloride tube)
until the starting base disappeared (TLC, S2). After evaporation of the dimethylformamide and
codistillation with toluene (3 × 50 ml), the residue was boiled with methanol (200 ml) and undis-
solved compound was collected by filtration of the hot mixture. This procedure afforded predominant
amount of the pure N9- (or N1-) isomers whereas the mother liquors contained a mixture of both
isomers. Further crops of products were obtained by chromatography of the mother liquors on silica
gel.
9-(3-Phthalimido-2-hydroxypropyl)adenine (VIIIa); yield 3.97 g (60%), m.p. 250 °C, RF 0.59 (S2).
For C16H14N6O3 (338.3) calculated: 56.80% C, 4.17% H, 24.83% N; found: 56.85% C, 4.37% H,
24.51% N.
9-(3-Phthalimido-2-hydroxypropyl)-2,6-diaminopurine(VIIIb); yield 5.49 g (78%), m.p. 237 –239 °C,
RF 0.41 (S2). For C16H15N7O3 (353.3) calculated: 54.39% C, 4.28% H, 27.74% N; found: 54.44% C,
4.36% H, 27.68% N.
TABLE VI
N-(3-Amino-2-hydroxypropyl) derivatives III
Calculated/Found
M.p., °C
Yield, %
Formula
M.w.
a
Compound
RF
% C
% H
% N
IIIa
IIIb
IIIc
IIId
IIIg
IIIi
176 – 178
0.53
0.50
C8H12N6O . 2H2O
244.3
39.33
39.40
6.60
6.12
34.44
34.06
42
205 – 206
C8H13N7O . H2O
241.3
39.82
39.49
5.43
5.99
40.61
40.50
63
192
48
C9H13N5O . H2O
225.2
47.98
48.12
6.72
6.45
31.10
30.96
162 – 163
C9H13N5O . H2O
225.2
47.98
47.77
6.72
6.82
31.10
31.34
76
249 – 251
0.47
0.26
0.49
0.27
C8H12N6O . H2O
226.2
42.47
42.81
6.24
5.89
37.15
37.47
55
223 – 225
C8H12N6O2 . H2O
242.2
39.66
39.40
5.82
5.62
34.47
34.06
66
IIIo
IIIp
131 – 133
C8H13N3O3 . H2O
217.2
44.23
44.25
6.96
6.81
19.34
19.25
54
203 – 205
C7H12N4O2 . H2O
202.2
41.57
41.75
6.98
6.42
27.71
27.86
55
a
Solvent system S1.
Collect. Czech. Chem. Commun. (Vol. 59) (1994)

1172
Spassova, Dvorakova, Holy, Budesinsky, Masojidkova:
9-(3-Phthalimido-2-hydroxypropyl)-2-amino-6-chloropurine (VIIIf) and 7-(3-Phthalimido-2-hydroxy-
propyl)-2-amino-6-chloropurine (XIII)
The residue obtained by concentration of the reaction mixture was boiled with a 1 : 1 mixture of
methanol and chloroform and the crystalline material was collected by filtration of the hot mixture;
yield 1.6 g of pure N9-isomer VIIIf. The mother liquor was chromatographed on silica gel (50 g)
in chloroform. The product VIIIf was eluted with chloroform–methanol (9 : 1). Total yield of the
N9-isomer was 4.46 g (60%), m.p. 248 – 250 °C, RF 0.68 (S2). For C16H13ClN6O3 (372.7) calculated:
51.56% C, 3.52% H, 22.54% N, 9.51% Cl; found: 51.64% C, 3.69% H, 22.24% N, 9.05% Cl.
Further elution afforded 0.60 g (8%) of the N7-isomer XIII, m.p. >250 °C, RF 0.58 (S2). For
C16H13ClN6O3 (372.7) calculated: 51.56% C, 3.52% H, 22.54% N, 9.51% Cl; found: 51.98% C,
3.57% H, 22.36% N, 10.28% Cl.
1-(3-Phthalimido-2-hydroxypropyl)cytosine (VIIIp)
Chromatography of the mother liquor on silica gel (125 g) in chloroform afforded 1.14 g of product
VIIIp (elution with chloroform–methanol 4 : 1). Total yield 4.44 g (71%), m.p. >250 °C, RF 0.25
(S2). For C15H14N4O4 (314.3) calculated: 57.33% C, 4.49% H, 17.82% N; found: 57.52% C, 4.46% H,
17.97% N.
9-(3-Phthalimido-2-hydroxypropyl)-2-amino-6-benzyloxypurine (VIIIq)
The reaction mixture after codistillation with toluene was chromatographed on silica gel (600 g) in
chloroform. The product was eluted with chloroform–methanol (9 : 1). Crystallization from ethanol
afforded 3.8 g (43%) of compound VIIIq, RF 0.52 (S3). For C23H20N6O4 (444.4) calculated: 62.16% C,
4.54% H, 18.90% N; found: 61.80% C, 4.59% H, 18.98% N.
9-[3-(o-Carboxybenzoyl)amino-2-hydroxypropyl]-2-amino-6-chloropurine (IX)
A mixture of phthalimido derivative VIIIf (2.0 g, 6 mmol), 0.4 M triethylammonium hydrogen carbo-
nate solution (50 ml) and dimethylformamide (50 ml) was kept at 50 °C for 30 h. After evaporation
and codistillation with toluene (3 × 100 ml), the white crystalline residue was triturated with water,
collected on filter and washed with acetone and ether. Yield 1.68 g (72%) of compound IX, m.p.
195 – 196 °C, RF 0.36 (S2), EUp 0.4. For C16H15ClN6O4 (390.7) calculated: 49.17% C, 4.39% H,
21.50% N, 9.07% Cl; found: 48.74% C, 4.01% H, 21.41% N, 8.81% Cl. Mass spectrum (m/z): 391
(M + H).
9-[3-(o-Carboxybenzoyl)amino-2-hydroxypropyl]adenine (X)
A mixture of compound VIIIa (1.0 g, 3 mmol), ammonia (10 ml) and water (20 ml) was stirred at
room temperature for 24 h. After evaporation, the reaction mixture was separated by preparative
HPLC (elution with water for 30 min, then with a gradient 0.5% methanol/min to 40% methanol).
Yield 0.5 g (50%) of compound X, m.p. 200 – 202 °C, k 26.5 (0.05 M TEAB 30 min, gradient to 10%
acetonitrile in 0.05 M TEAB/10 min), EUp 0.4. For C16H16N6O4 . H2O (374.3) calculated: 51.34% C,
4.85% H, 22.44% N; found: 51.29% C, 4.63% H, 22.05% N. Mass spectrum (m/z): 357.2 (M + H).
Collect. Czech. Chem. Commun. (Vol. 59) (1994)

N-(3-Amino-2-hydroxypropyl) Azaheterocycles
1173
Preparation of N-(3-Amino-2-hydroxypropyl) Derivatives III by Hydrazinolysis
of N-(Phthalimido-2-hydroxypropyl) Derivatives VIII. General Procedure
A mixture of phthalimido derivative VIIIa, VIIIb or VIIIp (1 mmol), ethanol and 98% hydrazine hy-
drate (0.055 ml) was refluxed for 8 h (calcium chloride protecting tube). The reaction mixture was
concentrated and the amino derivative was separated from phthalazine on Dowex 1 (AcO−, 50 ml) by
elution with water. After deionization on Dowex 50 (H+ form, 50 ml), evaporation and codistilla-
tion with ethanol (3 × 20 ml), the amino derivatives III were crystallized from ethanol. The obtained
compounds III were identical with authentic samples (vide supra) according to HPLC (S6 for IIIp, S7
for IIIb and S8 for IIIa). Yields: compound IIIa 93%, compound IIIb 82% and compound IIIp 87%.
2-Amino-6-benzyloxypurine (XI)
Benzyl alcohol (50 ml, 460 mmol) was added dropwise at room temperature to a stirred mixture of
sodium hydride (60% dispersion, 2.5 g, 62 mmol) and toluene (250 ml) and stirring was continued
for 1 h. 2-Amino-6-chloropurine (5 g, 30 mmol) was added, the mixture was refluxed for 6 h and
filtered while hot. The crystalline material collected on the filter represented the main portion of
compound XI and was purified by crystallization from ethanol; yield 2.4 g. The mother liquor after
filtration of the reaction mixture was chromatographed on silica gel (300 g) in chloroform. Elution
with chloroform–methanol (98 : 2) afforded dibenzyl derivative XII (0.8 g), RF 0.39 (S3), elution
with chloroform–methanol (96 : 4) gave further amount (1.6 g) of the monobenzyl derivative XI.
Total yield 4.0 g (56%) of compound XI, m.p. 190 – 192 °C, RF 0.24 (S3). For C12H11N5O (241.2)
calculated: 59.75% C, 4.60% H, 29.20% N; found: 59.21% C, 4.63% H, 28.83% N. Mass spectrum
(m/z): 242.1 (M + H). UV spectrum (methanol): λmax 284.0 nm (εmax 7 420), λmax 241.0 nm (εmax 7 420).
2-Benzylamino-6-benzyloxypurine (XII)
Benzyl alcohol (13.5 g, 125 mmol) was added dropwise under stirring to an ice-cooled mixture of
sodium hydride (60% dispersion, 5.0 g, 125 mmol) and dimethylformamide (250 ml) and the stirring
was continued for 1 h. 2-Amino-6-chloropurine (4.25 g, 25 mmol) was added to the obtained solution
and the mixture was stirred at 60 °C for 8 h (calcium chloride tube). After cooling, the reaction mix-
ture was neutralized with acetic acid, the solvent was evaporated and the residue was codistilled with
toluene. Chromatography on silica gel (500 g) in chloroform–methanol (97 : 3) afforded as the prin-
cipal product the dibenzyl derivative which was then crystallized from ethyl acetate. Yield 3.1 g
(37%), m.p. 165 – 167 °C, RF 0.39 (S3). For C19H17N5O (331.3) calculated: 68.87% C, 5.17% H,
21.13% N; found: 68.69% C, 5.16% H, 20.44% N. Mass spectrum (m/z): 332.2 (M + H). UV spec-
trum (methanol): λmax 284.0 nm (εmax 7 420), λmax 241.0 nm (εmax 7 420).
REFERENCES
1. Holy A.: Collect. Czech. Chem. Commun. 43, 3444 (1978).
2. Holy A.: Collect. Czech. Chem. Commun. 54, 446 (1989).
3. Gichner T., Holy A., Spassova M., Veleminsky J., Gruz P.: Mutagenesis 6, 55 (1991).
4. Juricek M., Gichner T., Kocisova J., Yefremova G. I., Veleminsky J., Stanek J., Moravcova J.,
Jary J.: Mutat. Res. 179, 175 (1987).
5. De Clercq E., Sakuma T., Baba M., Pauwels R., Balzarini J., Rosenberg I., Holy A.: Antiviral
Res. 8, 261 (1987).
6. Juricova K., Holy A., Smrckova S., Spassova M., Dvorakova H.: Collect. Czech. Chem.
Commun. 58, Special Issue, 244 (1993).
Collect. Czech. Chem. Commun. (Vol. 59) (1994)

1174
Spassova, Dvorakova, Holy, Budesinsky, Masojidkova:
7. Holy A.: Collect. Czech. Chem. Commun. 47, 173 (1981).
8. Ogilvie K. K., Nguyen-Ba N., Gillen M. F., Radatus B. K., Cheriyan U. O., Hanna H. R., Smith
K. O., Galloway K. S.: Can. J. Chem. 62, 241 (1984).
9. Koomen G. J., Provoost L. M., van Maarschalkerwaart D. A., Willard N. P.: Nucleosides
Nucleotides 11, 1297 (1992).
10. Ogawa T., Takaku H., Yamamoto N.: Nucleosides Nucleotides 8, 499 (1989).
11. Vemishetti P., El Subbagh H. I., Abushanab E., Panzica R. P.: Nucleosides Nucleotides 11, 739
(1992).
12. Chenon M.-T., Pugmire R. J., Grant D. M., Panzica R. P., Townsend L. B.: J. Am. Chem. Soc.
97, 4636 (1975).
13. Still I. W. J., Plavac N., McKinnon D. M., Chauhan M. S.: Can J. Chem. 56, 725 (1978).
14. Harnden M. R., Parkin A., Parratt J., Perkins R. M.: J. Med. Chem. 36, 1343 (1993).
15. Baron F., Brown D. M.: J. Chem. Soc. 1955, 2855.
16. Bowles W. A., Schneider F. H., Lewis L. R., Robins R. K.: J. Med. Chem. 6, 471 (1963).
17. Yu K. L., Bronson J. J., Yang H., Patick A., Alam M., Brankovan V., Datema R., Hitchcock M.
J. M., Martin J. C.: J. Med. Chem. 35, 2958 (1992).
18. Chenon M.-T., Pugmire R. J., Grant D. M., Panzica R. P., Townsend L. B.: J. Am. Chem. Soc.
97, 4627 (1975).
19. LeCocq C., Lallemand J.-Y.: J. Chem. Soc., Chem. Commun. 1981, 150.
20. Holy A., Ivanova G. S.: Nucleic Acids Res. 1, 19 (1974).
Translated by M. Tichy.
Collect. Czech. Chem. Commun. (Vol. 59) (1994)