Synthesis and biological activity of the 2-amino-4-(4-iodophenyl)amino-6- methylpyrimidine isosteric analogs

Article abstract of DOI:10.1134/S1070363213030213

Transformation of the structure of 6-methylisocytosine results in the isosteric analogs of 2-amino-4-(4-iodophenyl)amino-6-methylpyrimidine exhibiting tuberculocidal properties. The level of biological activity of the synthesized compounds depends on the

Full text of DOI:10.1134/S1070363213030213

ISSN 1070-3632, Russian Journal of General Chemistry, 2013, Vol. 83, No. 3, pp. 538–541. © Pleiades Publishing, Ltd., 2013.
Original Russian Text © A.V. Erkin, V.I. Krutikov, 2013, published in Zhurnal Obshchei Khimii, 2013, Vol. 83, No. 3, pp. 485–488.
Synthesis and Biological Activity
of the 2-Amino-4-(4-iodophenyl)amino-6-methylpyrimidine
Isosteric Analogs
A. V. Erkin and V. I. Krutikov
St. Petersburg State Institute of Technology (Technical University),
Moskovskii pr. 26, St. Petersburg, 190013 Russia
e-mail: kruerk@yandex.ru
Received October 4, 2012
Abstract―Transformation of the structure of 6-methylisocytosine results in the isosteric analogs of 2-amino-4-
(4-iodophenyl)amino-6-methylpyrimidine exhibiting tuberculocidal properties. The level of biological activity
of the synthesized compounds depends on the site of location of the halogen atom.
DOI: 10.1134/S1070363213030213
A factor determining the ability of 2-amino-4-
arylamino-6-methylpyrimidines (I) to inhibit the
growth of Mycobacterium smegmatis cell culture at
low concentrations is the presence of halogen atoms in
meta- or para-position of the aromatic ring [1, 2 ]. The
migration of the halogen to the hetero-ring causes the
loss of antimycobacterial activity due to the formation
of respective inactive 2-amino-5-halo-6-methyl-4-
phenylaminopyrimidine (II) [3]. In order to find out
whether this trend occurs in the series of 2-amino-4-(4-
iodophenyl)amino-6-methyl-pyrimidine hydrochloride
(III) and its isosteric analogs inhibiting the cell growth
of the related culture, Mycobacterium tuberculosis [4],
we synthesized 2-amino-4-(3-iodophenyl)amino-6-methyl-
pyrimidine (IV) and 2-amino-5-iodo-6-methyl-4-
phenylaminopyrimidine (V) hydrochlorides and invest-
tigated the tuberculocidal properties of these compounds.
Iodophenylaminopirimidine IV formed at the acidic
hydrolysis of the intermediate 4-(3-iodophenyl)amino-
6-methyl-2-formylaminopyrimidine VI, which, in turn,
resulted from the condensation of 6-methyl-2-formyl-
amino-4-chloropyrimidine VII with 3-iodophenyl-
amine at elevated temperature in the absence of a
solvent and was used without further purification in the
deformylation step.
I
HN
I
Cl
H2N
N
HN
I
N
HCl
N
HCl
N
N
Me
N
N
Me
O
H2N
N
Me
O
H
H
VI
VII
IV
In the 1H NMR spectrum of compound IV there are
the characteristic proton signals of primary and
secondary amino groups near 8.0 and 10.9 ppm
respectively, and the signal of the proton H5 is shifted
upfield compared with similar parameter of the
substrate VII.
To obtain iodopyrimidine V we performed se-
quential modification of the 2-amino-5-iodo-6-methyl-
pyrimidin-4(3H)-one VIII.
The acetylation of iodoisocytosine VIII with excess
of acetic anhydride led to 2-acetylamino-5-iodo-6-
538

SYNTHESIS AND BIOLOGICAL ACTIVITY
539
O
O
Cl
I
I
Me
I
Ac2O
POCl3
HN
H2N
HN
N
Ac
Ac
N
Me
N
N
N
N
Me
H
H
VIII
IХ
Х
PhNH2
NHPh
NHPh
NHPh
I
I
I
HCl
KOH
N
N
N
HCl
Ac
H2N
N
Me
H2N
N
Me
N
H
N
Me
V
ХII
ХI
methyl-pyrimidin-4(3H)-one IX, which at treating with
phosphorus oxychloride was converted into 2-acetyl-
amino-5-iodo-6-methyl-4-chloropyrimidine X. The protec-
tion of amino group of compound VIII was carried out
in order to avoid its possible [4] interaction with the
chlorinating agent. The condensation of dihalo-
pyrimidine X with phenylamine in ethanol gave 2-
acetylamino-5-iodo-6-methyl-4-phenyl-aminopyrimidine
XI. The latter underwent deacetylation under alkaline
hydrolysis to 2-amino-5-iodo-6-methyl-4-phenylamino-
pyrimidine XII. The quaternization of the free base
XII with hydrogen chloride in an inert solvent caused
the formation of the target iodopyrimidine V.
where R = 2-amino-6-methyl-4-oxo-3,4-dihydropyr-
imidin-5-yl.
1H NMR spectrum of formylaminopyrimidine XIV
contains the characteristic signals of the protons of the
methyl group around 2.3 ppm and formamide group
around 6.9 ppm, with integral intensities ratio 3:2.
We succeeded to perform exchange chlorination of
compounds IX limiting the contact of the reactant by
the period of homogenization of the initially formed
suspension. At more prolonged heating a significant
tarring of the reaction mixture occurred, and it was
difficult to purify the isolated dihalopyrimidine X from
the low-melting impurities of unknown origin.
In contrast to the proposed [5] method of iodination
of 2-amino-6-methyl-pyrimidin-4(3H)-one XIII with
iodine in the presence of hydrogen peroxide under the
influence of infrared irradiation, we used a less
laborious method, based on the processing compound
XIII with iodine in aqueous alkaline solution that
ensured a 60% yield of reaction product VIII.
O
The amination of compound X proceeded rather
difficultly. In the course of heating the equimolar
mixture with phenylamine in ethanol in the absence of
hydrogen chloride acceptor in the amount needed for
the obtaining N2-acetyl-5-iododiaminopyrimidine XI
directly as the hydrochloride, we observed the forma-
tion of a new compound, a measurable amount of
which was extracted from the reaction mixture at the
complete removal of the solvent. The double crystal-
lization of the combined product from 30% aqueous
ethanol resulted in a chromatographically pure com-
I2
HN
VIII
H2N
N
Me
1
pound XI as a free base. H NMR spectrum of N2-
ХIII
acetyl-5-iododiaminopyrimidine XI contained the
signals of the protons of methyl groups around 2.1 and
2.5 ppm, aromatic ring at 7.0–7.9 ppm, and secondary
amino groups at about 8.0 and 10.1 ppm After con-
centration of the mother liquor in vacuo, the residue
showed a positive test for the presence of chloride ion
with the aqueous solution of silver nitrate. The
crystallization from 30% ethanol did not increase the
chromatographic purity of the substance, although the
Attempted formylation of iodoisocytosine VIII by
heating the substrate with 85% formic acid resulted in
formation of 5-iodo-6-methyl-2-formyl-aminopyrimidin-
4(3H)-one XIV in a yield not exceeding 10%: the com-
pound VIII mostly underwent deiodination according
to the redox reactions:
2RI + 2H3O+ + 2e–→ 2RH + I2 + H2O,
HCOO– – 2e–→ CO2 + H+,
RUSSIAN JOURNAL OF GENERAL CHEMISTRY Vol. 83 No. 3 2013

540
ERKIN, KRUTIKOV
character of its melting corresponded to a purified
product. H NMR spectrum showed that the substance
EXPERIMENTAL
1
1H NMR spectra were recorded on a spectrometer
Bruker WM-400 (operating frequency 400.13 MHz) in
DMSO-d6, internal reference the residual proton
signals of the solvent. Individuality of compounds was
monitored by TLC on Silufol UV-254 plates in the
systems: 1-butanol–acetic acid–water, 1:1:1 (eluent A)
and acetone–heptane, 2:1 (eluent B). The development
of spots was performed under UV irradiation. Elemen-
tal analyzes were carried out on a Hewlett Packard B-
185 analyzer. Water or water-ethanol solutions of
compounds IV and V gave a positive test for the
presence of chloride ion with an aqueous solution of
silver nitrate. 6-Methyl-2-formylamino-4-chloropyr-
imidine VII was prepared by the method [4], 2-amino-
6-methyl-pyrimidin-4(3H)-one XIII, by the method
[6].
was a mixture of N2-acetyl-5-iododiaminopyrimidine
XI and 2-acetylamino-6-methyl-4-phenylaminopyr-
imidine XV, identified by the proton signals of the
methyl groups about 2.3 and 2.5 ppm, methine group
at 6.2 ppm, the secondary amino groups near 8.2 and
6.10 ppm, and the quaternized nitrogen atom in the
region of 12–13 ppm.
NHPh
PhNH2
N
Х
ХI
+
HCl
Ac
N
N
Me
H
ХV
We succeeded to avoid deiodination of compound
XI in situ by performing reaction of dihalopyrimidine
X and phenylamine in the presence of triethylamine. In
this case we met the difficulty at the purification the
resulting N2-acetyl-5-iododiaminopyrimidine XI from
the impurities of unknown nature, and therefore we
carried out alkaline hydrolysis of the crude substrate
and obtained 5-iododiaminopyrimidine XII in a yield
of about 40%. Despite the simplification of the
synthesis procedure, compound XII isolated from the
reaction mixture required repeated crystallization to
reach the chromatographic purity and narrow tem-
perature range of melting. In the 1H NMR spectrum of
the purified 5-iododiaminopyrimidine XII there were
the signals of the protons of methyl group around
2.4 ppm, of the primary and secondary amino groups
at about 6.2 and 7.6 ppm, respectively, and of the
aromatic fragment at 7.0–7.7 ppm.
2-Amino-4-(3-iodophenyl)amino-6-methylpyrimidine
(IV). A mixture of 0.41 g of formylaminochloropyr-
imidine VII and 0.53 g of 3-iodophenylamine was kept
at 100°C until solidification. The melt was mecha-
nically ground and dispersed in 10 ml of concentrated
hydrochloric acid. The mixture was heated to homo-
genization, kept in a boiling water bath for 30 min,
then cooled, the precipitate formed was filtered off,
and the filtrate was evaporated to dryness in a vacuum.
The combined products were recrystallized from
ethanol and dried at 60°C for 6 h. 0.33 g (42%) of
compound IV was obtained as the hydrochloride, mp
1
260°C (decomp.), Rf 0.78 (A). H NMR spectrum, δ,
ppm: 2.31 s (3H, Me), 6.29 s (1H, CH), 7.11–8.09 m
(6H, Ar, NH2), 10.91 s (1H, NH), 13.15 s ( 1H, N+H).
Found, %: C 36.45, H 3.17, N 15.06. C11H11IN4·HCl.
Calculated, %: C 36.44, H 3.34, N 15.45.
The results of the biological screening of the iso-
steric analogs of diaminopyrimidine III indicate that
its tuberculocidal effect is due to, among other po-
tential factors, the strict localization of the halogen
atom in the para-position of the aromatic ring. When
the iodine is in the meta-position of the ring the
respective compound IV does not exhibit the ability to
inhibit Mycobacterium tuberculosis, but inhibits to
100% the cell growth of the related culture Myco-
bacterium smegmatis, at a concentration of 25 mg ml–1.
The migration of the halogen from the para-positions
of the aromatic ring of diaminopyrimidine III in the
hetero-ring results in the loss of inhibiting properties of
the arising iodopyrimidine V with respect to the cells
of both cultures even at increasing its concentrations to
100 μg ml–1.
2-Amino-5-iodo-6-methyl-4-phenylaminopyrimidine
(V) hydrochloride. Through the solution of 117 mg of
compound XII in 15 ml of anhydrous benzene was
passed a flow of dry hydrogen chloride until saturation.
The resulting suspension was filtered off, the solid was
washed with water-free benzene and dried at 60°C for
6 h to afford 112 mg (86%) of compound V, mp 181°C
(decomp.), Rf 0.74 (A). Found, %: C 36.18%, H 3.21,
N 15.23. C11H11IN4·HCl. Calculated, %: C 36.44%, H
3.34; N 15.45.
2-Amino-5-iodo-6-methyl-pyrimidin-4(3H)-one
(VIII). To a solution of 4 g of isocytosine XIII in 10%
aqueous potassium hydroxide prepared from 6 g of
alkali and 54 ml of water was added dropwise at
vigorous stirring a solution of 8.13 g of iodine in
RUSSIAN JOURNAL OF GENERAL CHEMISTRY Vol. 83 No. 3 2013

SYNTHESIS AND BIOLOGICAL ACTIVITY
541
aqueous potassium hydroxide of the same concentra-
tion. The mixture was stirred for 1 h and after 24 h
neutralized with concentrated hydrochloric acid. The
precipitate formed was filtered off, washed with water,
and recrystallized from 60% DMF. After thorough
washing with water and drying at 80°C for 8 h 5.17 g
(64%) of compound VIII was obtained, mp. 203°C
(decomp.) [published data [5]: mp 225–226°C (decomp.)],
ethylamine in 20 ml of ethanol was refluxed for 1 h.
After removal of ethanol in vacuo, the residue was
triturated with 15 ml of ethyl acetate to form a slurry,
and left to stand for 2 days. The precipitate formed was
filtered off, washed with ethyl acetate, and dried at 60°C
for 2 h. 0.93 g (53%) of crude 2-acetylamino-5-iodo-6-
methyl-4-phenylamino-pyrimidine XI was obtained. It
was dissolved in 20 ml of 60% ethanol containing 0.51 g
of potassium hydroxide. The mixture was kept in a
boiling water bath for 1 h, then evaporated in a vacuum
to dryness, the residue was mixed with 10 ml of water,
insoluble part was filtered off, washed with water, and
dried at 60°C for 6 h. The dry product was recrystal-
lized three times from cyclohexane to achieve chro-
matographic purity and narrow melting temperature
range. After drying in a high vacuum 117 mg (7.5%
based on the original dihalopyrimidine X) of
compound XII was obtained, mp 143°C, Rf 0.31 (B).
1H NMR spectrum, δ, ppm: 2.39 s (3H, Me), 6.16 s
(2H, NH2), 7.01 m (1H, Ph), 7.28 m (2H, Ph), 7.57 s
(1H, NH) , 7.65 d (2H, Ph). Found, %: C 40.29%, H
3.35, N 16.89. C11H11IN4. Calculated, %: C 40.51%, H
3.40, N 17.18.
1
Rf 0.76 (A). H NMR spectrum, δ, ppm: 2.19 s (3H,
Me), 6.60 br.s (1H, NH2), 11.12 br.s (1H, NH). Found,
%: C 23.63%, H 2.57, N 16.56. C5H6IN3O. Calculated,
%: C 23.92%, H 2.41; N 16.74.
2-Acetylamino-5-iodo-6-methylpyrimidin-4(3H)-
one (IX). A mixture of 2.43 g of iodoisocytosine VIII
and 36 ml of freshly distilled acetic anhydride was
boiled for 1 h, the formed precipitate was filtered off
and recrystallized from 30% acetic acid. After
thorough washing with water and drying at 60°C for
6 h, 1.2 g (42%) of compound IX was isolated, mp
1
215°C (decomp.), Rf 0.88 (A). H NMR spectrum, δ,
ppm: 2.16 s (3H, Me), 2.44 s (3H, Ac), 11.72 s (1H,
NH), 11.93 s (1H, NH). Found, %: C 28.76%, H 2.43,
N 13.96. C7H8IN3O2. Calculated, %: C 28.69%, H
2.75, N 14.34.
ACKNOWLEDGMENTS
The authors are grateful to Professor V.V. Tets and
coworkers (Pavlov State Medical University at St.
Petersburg) for microbiological studies.
2-Acetylamino-5-iodo-6-methyl-4-chloropyrimidine
(X). A mixture of 1.2 g of acetylaminopyrimidinone IX
and 15 ml of freshly distilled phosphorus oxychloride
was heated at a temperature not exceeding 115°C until
dissolution. Excess of phosphorus oxychloride was
com-pletely removed in vacuo, the residue was mixed
with finely crushed ice and ground to form a slurry.
The precipitate was filtered off, washed with water,
and recrystallized from 40% ethanol. After washing
with water and drying at 60°C for 6 h 0.76 g (59%) of
compound X was isolated, mp 167°C, Rf 0.59 (B).
Found, %: C 26.87%, H 2.37, N 13.61. C7H7ClIN3O.
Calculated, %: C 26.99%, H 2.27, N 13.49.
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2-Amino-5-iodo-6-methyl-4-phenylaminopyrimidine
(XII), free base. A mixture of 1.49 g of dihalopyr-
imidine X, 0.44 g of phenylamine, and 0.48 g of tri-
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2011, vol. 81, no. 8, p. 1354.
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