New amino derivatives of 1,2,3-triazolo[4,5-d]pyrimidines and their affinity towards A1 and A(2A) adenosine receptors

Article abstract of DOI:10.1016/S0223-5234(99)00205-6

Starting from the appropriate azides (4-chlorobenzyl-, 2- thiophenemethyl-, 2-fluorobenzyl-, and 4-fluorobenzylazides) in which the variation of the substituent is at the basis of the four series of derivatives (a-d), the 7-aminosubstituted 1,2,3-triazolo[4,5-d]pyrimidines 4 were prepared by a well known synthetic route. The biological activity of compounds 4 was expected on the basis of the presence of particular substituents on N(7), and these substituents were introduced by the reaction of the 7 lactamic carbonyl function, present on precursors 3, with cycloalkyl-, aralkyl- and arylamines. Radioligand binding assays at bovine brain adenosine A₁ and A(2A) receptors showed that some compounds possessed a high affinity and selectivity for the A₁ receptor subtype. Furthermore, biological results indicated that the p-chlorobenzyl substituent lowered receptor binding, compared with the previously prepared benzyl and 2-chlorobenzyl derivatives, suggesting certain particular steric requirements of the lipophilic region which interacts with the benzyl substituent. The thiophenemethyl substituent conferred more activity than the benzyl one. The presence of a fluorine atom on the benzyl group determined a high affinity, especially when it was in the ortho position. Compounds 4c.1 (R = 2-fluorobenzyl, R' = cyclopentyl, Ki = 10.5 nM), 4c.2 (R = 2-fluorobenzyl, R' = cyclohexyl, Ki = 19.5 nM) and 4d.1 (R = 4- fluorobenzyl, R' = cyclopentyl, Ki = 26 nM) were the most active for A₁ receptors.

Full text of DOI:10.1016/S0223-5234(99)00205-6

Eur. J. Med. Chem. 34 (1999) 867−875
© 1999 Éditions scientifiques et médicales Elsevier SAS. All rights reserved
867
Short communication
New amino derivatives of 1,2,3-triazolo[4,5-d]pyrimidines and
their affinity towards A1 and A2A adenosine receptors
Laura Bettib, Giuliana Biagia, Gino Giannaccinib, Irene Giorgia, Oreste Livia*, Antonio Lucacchinib,
Clementina Maneraa, Valerio Scartonia
aDipartimento di Scienze Farmaceutiche, Facoltà di Farmacia, Università di Pisa, via Bonanno 6, 56126 Pisa, Italy
bDipartimento di Psichiatria, Neurobiologia, Farmacologia e Biotecnologie, Università di Pisa, via Bonanno 6, 56126 Pisa, Italy
(Received 19 December 1998; revised 7 April 1999; accepted 21 April 1999)
Abstract – Starting from the appropriate azides (4-chlorobenzyl-, 2-thiophenemethyl-, 2-fluorobenzyl-, and 4-fluorobenzylazides) in which
the variation of the substituent is at the basis of the four series of derivatives (a–d), the 7-aminosubstituted 1,2,3-triazolo[4,5-d]pyrimidines
4 were prepared by a well known synthetic route. The biological activity of compounds 4 was expected on the basis of the presence of
particular substituents on N(7), and these substituents were introduced by the reaction of the 7 lactamic carbonyl function, present on
precursors 3, with cycloalkyl-, aralkyl- and arylamines. Radioligand binding assays at bovine brain adenosine A1 and A2A receptors showed
that some compounds possessed a high affinity and selectivity for the A1 receptor subtype. Furthermore, biological results indicated that the
p-chlorobenzyl substituent lowered receptor binding, compared with the previously prepared benzyl and 2-chlorobenzyl derivatives,
suggesting certain particular steric requirements of the lipophilic region which interacts with the benzyl substituent. The thiophenemethyl
substituent conferred more activity than the benzyl one. The presence of a fluorine atom on the benzyl group determined a high affinity,
especially when it was in the ortho position. Compounds 4c.1 (R = 2-fluorobenzyl, R≠ = cyclopentyl, Ki = 10.5 nM), 4c.2 (R = 2-fluorobenzyl,
R≠ = cyclohexyl, Ki = 19.5 nM) and 4d.1 (R = 4-fluorobenzyl, R≠ = cyclopentyl, Ki = 26 nM) were the most active for A1 receptors. © 1999
Éditions scientifiques et médicales Elsevier SAS
1,2,3-triazoles / 1,2,3-triazolo[4,5-d]pyrimidines / A1-adenosine and A2A-adenosine receptor antagonists
1. Introduction
a fan-shape with respect to the NH function, which was
engaged as a hydrogen bond donor.
Our previous studies on 7-aminosubstituted 1,2,3-
triazolopyrimidines [1] indicated that certain compounds,
bearing a benzyl or 2-chlorobenzyl as the lipophilic
substituent in the 3 position, showed a high affinity and
selectivity towards the A1 receptor subtype.
On the basis of our previous results, the investigations
into these structures were continued in order to study the
mode of binding of these compounds with A1-receptors
by comparative SAR analysis. We report here the synthe-
sis and biological evaluation of a series of 1,2,3-
triazolo[4,5-d]pyrimidines (4) characterized by new
benzyl-type substituents (R = 4-chlorobenzyl, 2-thenyl,
2-fluorobenzyl and 4-fluorobenzyl). In the 7 position,
amino substituents which conferred a high biological
activity to previously prepared molecules (cyclopentyl-
and cyclohexylamino, meta- and para-toluidino,
α-methylbenzylamino, amphetamino) as well as other
amino substituents (isopropylamino, 2-butylamino,
2-pentylamino, 3-pentylamino, p-bromoanilino, furfury-
lamino, 2-pyridylamino and 2-hydroxy-cyclohexyl-
amino) were introduced.
These and other studies of ours on 1,2,3-triazolo-
pyrimidines [2–6], in accordance with the receptor bind-
ing site models proposed for adenosine agonists and
antagonists [7], revealed three lipophilic binding regions
(corresponding to the N-6, C-2 and N-9 positions of the
azapurine ring) in the A1 adenosine receptors, arranged as
*Correspondence and reprints

868
lipophilic substituent in the
3
position:
a
(4-
chlorobenzyl), b (2-thiophenemethyl), c (2-fluorobenzyl)
and d (4-fluorobenzyl) (table I).
The structures of all the new compounds were assigned
on the basis of the well-known reaction mechanisms
already regularly carried out in our laboratory: 1,3-
dipolar cycloaddition of azides to activated methylenic
compounds, formation of the pyrimidine ring, chlorina-
tion and nucleophilic displacement of the halogen by
amines. The structures were also confirmed by analytical
and spectroscopic data.
3. Biochemistry
The 7-(aminosubstituted)-1,2,3-triazolo[4,5-d]pyrimi-
dines were tested in radioligand binding assays for
affinity at A1 and A2A adenosine receptors in bovine brain
cortical membranes and in bovine brain striatal mem-
branes respectively. [3H]R-(-)-N6-cyclohexyl-adenosine
(CHA) was used as the A1 radio-ligand and [3H]-2-{[[p-
(2-carboxyethyl)phenyl]ethyl]amino}-5≠-(N-ethylcarba-
moyl)adenosine (CGS 21680) as the A2A radioligand.
The experimental details of the receptor binding assays
were reported in a previous paper [1].
Figure 1. Synthetic route for compounds 4.
4. Results and discussion
2. Chemistry
The results of the A1 and A2A adenosine receptor
binding assays, expressed as inhibition constants (Ki,
nM) in table II show that introduction of the
p-chlorobenzyl substituent on the triazole ring in the 3
position (compounds 4a.1–7) reduces A1 and A2A ad-
enosine receptor binding compared with the
2-chlorobenzyl substituent [1]. The azapurine compound
4a.1, a cyclopentylamino derivative, is an exception, with
an A1 affinity constant of Ki = 128 nM, followed by the
cyclohexylamino derivative 4a.2 with a Ki = 1 360 nM.
Both compounds showed a high receptor selectivity
(table II); the inhibition percentages of the A2A adenosine
receptor binding assays were so low that the correspond-
ing Ki values were not calculated. The shift of the
chlorine from the ortho to the para position suggested a
moderate and well-defined extent of the lipophilic region
which receives the benzyl substituent. Whereas the ortho-
chlorine atom on the benzyl group increased the affinity
towards the A1 receptor [1], the results of the 4a series
indicated that the chlorine atom in the para position
decreased the capacity to bind with the receptor site. This
fact could mean that the para-chloro substitution gener-
ally caused a steric repulsion within the receptor, owing
to the limited depth of the lipophilic site.
The scheme for the preparation of compounds 4 started
from the appropriate azide, which determined the R
substituent (series a–d) and followed
experimented synthetic route (figure 1). All the azides
have been reported in the literature (4-
chlorobenzylazide, [8]; 2-thiophenemethylazide, [9];
2-fluorobenzylazide [10]; and 4-fluorobenzylazide [11])
but preparation of 2-thiophenemethylazide has not been
described.
The appropriate azide was reacted with cyanoaceta-
mide in the presence of sodium ethoxide in refluxing
ethanol to give the 1-substituted-4-carboxamido-5-
amino-1H-1,2,3-triazoles 1a–d in good yields. These
compounds, by heating with formamide, easily cyclized
to the corresponding 7-hydroxy-triazolopyrimidines 2a–d
which, by reaction with thionyl chloride in chloroform,
provided the expected chloroderivatives 3a–d. Com-
pounds 1a, 2a and 3a have been described in the
literature [8]. The chlorine atom in the 7 position was
reactive enough to undergo a nucleophilic displacement
by primary aliphatic or aromatic amines (consequently,
several triazolopyrimidine derivatives 4 were prepared,
corresponding to the four series characterized by the
a
well-

869
Table I. Physicochemical data of triazolopyrimidine derivatives and intermediates.
Compound Yield%
crystall.
solvent
M.p. °C Molecular formula
(molecular weight)
MS
M+
m/z
base peak
1b
1c
1d
2b
2c
2d
3b
3c
56
73
87
59
56
52
27
63
61
18
36
65
50a
83
84
16
49
42
10a
18a
56
73
46
71
27a
42a
54
71
44
84
52
45a
68
41a
23a
42
59
31
67a
78
54
45
88
84
71
66
73
65
EtOH
EtOH
EtOH
EtOH
EtOH
EtOH
60–80 °C petr. ether
60–80 °C petr. ether
60–80 °C petr. ether
MeOH
60–80 °C petr. ether
EtOH
EtOH
EtOH
EtOH
MeOH
MeOH
MeOH
EtOH
MeOH
EtOH
EtOH
MeOH
MeOH
EtOH
EtOH
EtOH
MeOH
EtOH
EtOH
EtOH
EtOH
MeOH
EtOH
EtOH
MeOH
MeOH
60–80 °C petr. ether
EtOH
EtOH
EtOH
100–140 °C petr. ether
EtOH
EtOH
Toluene
EtOH/H2O
60–80 °C petr. ether
60–80 °C petr. ether
197–198 C8H9N5OS (223.25)
186–188 C10H10N5OF (235.22)
221–223 C10H10N5OF (235.22)
225–230 C9H7N5OS (233.25)
221–224 C11H8N5OF (245.22)
225–228 C11H8N5OF (245.22)
223
235
235
233
216 (M+-29)
245
251
263
263
328
342
364
379
350
350
316 (M+-65)
300
314
259 (M+-91)
336
322
322
312
326
362
271 (M+-91)
334
334
342
324
399
321
286
300
314
97
109
109
204
109
109
97
109
109
125
125
125
125
125
125
125
203
97
79–80
85–88
74–76
C9H6N5SCl (251.69)
C11H7N5FCl (263.66)
C11H7N5FCl (263.66)
3d
4a.1
4a.2
4a.3
4a.4
4a.5
4a.6
4a.7
4b.1
4b.2
4b.3
4b.4
4b.5
4b.6
4c.1
4c.2
4c.3
4c.4
4c.5
4c.6
4c.7
4c.8
4c.9
4c.10
4c.11
4c.12
4c.13
4c.14
4d.1
4d.2
4d.3
4d.4
4d.5
115–116 C16H17N6Cl (328.80)
109–111 C17H19N6Cl (342.83)
110–112 C19H17N6Cl (364.84)
158–161 C20H19N6Cl-HCl (415.33)
175–176 C18H15N6Cl (350.81)
163–164 C18H15N6Cl (350.81)
115–117 C17H12N7O2Cl (381.78)
120–122 C14H16N6S (300.38)
132–135 C15H18N6S (314.41)
148–152 C18H18N6S-HCl (386.90)
139–142 C17H16N6S-HCl (372.88)
149–151 C16H14N6S (322.39)
170–172 C16H14N6S (322.39)
109–112 C16H17N6F (312.35)
107–109 C17H19N6F (326.38)
159–162 C20H19N6F-HCl (398.87)
181–185 C20H19N6F-HCl (398.87)
139–142 C18H15N6F (334.36)
139–141 C18H15N6F (334.36)
153–156 C17H19N6OF (342.38)
170–173 C16H13N6OF (324.32)
196–198 C17H12N6BrF (399.22)
182–185 C16H12N7F-HCl (357.78)
127–129 C14H15N6F (286.31)
160–165 C15H17N6F-HCl (336.80)
167–170 C16H19N6F-HCl (350.83)
107–109 C16H19N6F (314.37)
121–124 C16H17N6F (312.35)
115–118 C17H19N6F (326.38)
154–157 C20H19N6F-HCl (398.87)
184–186 C18H15N6F (334.36)
139–141 C18H15N6F (334.36)
142–144 C17H19N6OF (342.38)
195–198 C16H13N6OF (324.32)
205–208 C17H12N6BrF (399.22)
214–217 C16H12N7F (321.32)
97
97
97
97
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
109
b
314
312
326
362
334
334
342
324
399
321
300
314
4d.6
4d.7
4d.8
4d.9
4d.10
4d.11
4d.12
99–101
82–84
90–92
C15H17N6F (300.34)
C16H19N6F (314.37)
C16H19N6F (314.37)
314
b
28
aIsolated as a hydrochloride; Specific rotation of the free base [α]D = – 42.3° (c = 1.30, CHCl3).

870
Table II. A1 and A2A adenosine receptor binding of triazolopyrimidines 4a–d.
Compound
R
R1
A1
A2A
Ki A2A/Ki A1
Ki nM
Ki nM
4a.1
128 ± 12
> 10 000
> 78
4a.2
4a.3
″
″
1 360 ± 82
> 10 000
> 10 000
> 10 000
> 7.3
–
4a.4
″
> 10 000
> 10 000
–
4a.5
4a.6
″
″
> 10 000
> 10 000
> 10 000
> 10 000
–
–
4a.7
″
> 10 000
> 10 000
–
-----------------------------------------------------------------------------------------------------------------------------------------
4b.1
37 ± 3
1 460 ± 131
39.4
4b.2
4b.3
″
″
75 ± 5
39 ± 3
1 740 ± 156
> 10 000
23.2
> 256
4b.4
4b.5
″
″
68 ± 4
42 ± 3
798 ± 56
616 ± 43
11.7
14.6
4b.6
″
172 ± 7
1 305 ± 104
7.5

871
Table II. continued.
Compound
R
R1
A1
A2A
Ki A2A/Ki A1
325.9
Ki nM
Ki nM
4c.1
10.5 ± 0.7
3 422 ± 205
4c.2
4c.3
″
″
19.5 ± 1.2
73.4 ± 6.6
3 973 ± 198
2 181 ± 153
203.7
29.7
4c.4
″
91 ± 6
932 ± 83
10.2
4c.5
4c.6
″
″
56 ± 4
1 906 ± 172
> 10 000
34.0
402 ± 32
> 24.8
4c.7
4c.8
″
″
1 188 ± 95
> 10 000
> 8.4
1 650 ± 149
9 800 ± 686
5.9
4c.9
″
″
112 ± 7
45 ± 3
> 10 000
> 89
4c.10
2 613 ± 183
58.0
4c.11
4c.12
4c.13
″
″
″
204 ± 18
81 ± 6
> 10 000
> 10 000
> 10 000
> 49
> 123
> 38
262 ± 23
4c.14
″
55 ± 4
4 000 ± 240
72.7

872
Table II. continued.
Compound
R
R1
A1
A2A
Ki A2A/Ki A1
Ki nM
Ki nM
4d.1
26 ± 2
> 10 000
> 384
4d.2
4d.3
″
″
72 ± 6
> 10 000
> 10 000
> 138
> 34
287 ± 26
4d.4
4d.5
″
″
254 ± 18
> 10 000
> 10 000
> 39
> 8
1 190 ± 83
4d.6
4d.7
″
″
1 137 ± 102
> 10 000
> 10 000
> 10 000
> 8
–
4d.8
4d.9
″
″
377 ± 26
429 ± 30
> 10 000
> 10 000
> 26
> 23
4d.10
4d.11
4d.12
″
″
″
119.5 ± 9.5
268 ± 24
84 ± 6
> 10 000
> 83
> 37
90.3
> 10 000
7 586 ± 683
The 2-thiophenemethyl (2-thenyl) substituent (series
b) was chosen because it could partially imitate the
2-chlorobenzyl substituent in view of its steric and
electronic characteristics. This substituent appeared to be
actively involved in A1 adenosine receptor binding: the
triazolopyrimidines 4b.1–5 (table II) showed a high A1
adenosine affinity (Ki < 100 nM) except for the
m-toluidino derivative 4b.6 (Ki = 172 nM). The most
active compound of this series was again the cyclopen-
tylamino derivative 4b.1 (Ki = 37 nM) but its affinity was
lower than those of the 2-chlorobenzyl (Ki = 21 nM) [1]
and 2-fluorobenzyl 4c.1 (Ki = 10.5 nM) derivatives.
These results further confirmed the ability of the A1
receptor to interact with lipophilic substituents bearing a

873
sulphur or a chlorine atom in the 2 position. The
amphetamino 4b.3 and the p-toluidino 4b.5 derivatives
possessed equivalent affinities (Ki = 39 and 42 nM
respectively), confirming the effectiveness of the amphet-
amino substituent and of the para-methyl position, com-
pared with the meta-methyl one (4b.6, Ki = 173 nM).
This effect was also recorded in the para-fluorobenzyl
series (4d.4, Ki = 254 nM; 4d.5, Ki = 1 190 nM). Among
compounds of the b series, 4b.3 showed the best
selectivity (Ki A2A/Ki A1 > 256) and the most active
compound 4b–1 followed with a ratio Ki A2A / Ki A1 =
39.5.
(cyclopentyl derivative, Ki = 26 nM), 4d.2 (cyclohexyl
derivative, Ki = 72 nM) and 4d.12 (3-pentyl derivative,
Ki = 84 nM). Thus in this series, a high affinity towards
A1 adenosine receptors was conferred by aliphatic amino-
substituents, which also showed a high receptor selectiv-
ity. The other aminosubstituents induced a noticeable
decrease in binding affinity compared with the corre-
sponding derivatives of the 2-chlorobenzyl [1] or
2-fluorobenzyl series.
5. Experimental protocols
The selection of the fluorobenzyl substituents, to com-
pare with the corresponding chlorobenzyl substituents,
was based upon the consideration that the strong elec-
tronegativity of the fluorine, which is capable of accept-
ing a hydrogen bond, allowed us to evaluate the effect of
possible electronic, as well as steric factors on receptor
binding. As regards the 2-fluorobenzyl substituent, used
for the preparation of the triazolopyrimidines 4c.1–14
(table II), a generalized increase in affinity was observed
towards the A1 adenosine receptors, compared with the
2-chlorobenzyl derivatives [1], excluding the amphet-
amino derivatives 4c.3 and 4c.4. The most active com-
pounds were once again the cyclopentylamino 4c.1 (Ki =
10.5 nM) and the cyclohexylamino 4c.2 (Ki = 19.5 nM)
derivatives, which also showed the best selectivity to-
wards the A1 adenosine receptors (Ki A2A/Ki A1 = 326
and 203, respectively); the introduction of an alcoholic
OH function on the cyclohexyl ring (compound 4c.7) (see
GR-79326, GlaxoWellcome A1 adenosine agonist) mark-
edly decreased binding affinity. The amphetamino sub-
stituent maintained a good affinity (Ki < 100 nM), but it
is worth noting that the racemic derivative 4c.3 was found
to be slightly more effective than the levorotatory isomer
4c.4. The new furfurylamino substituent (4c.8) markedly
lowered receptor binding. As regards the known
7-arylamino substituents, the binding affinity was con-
firmed to be high with the para-toluidino derivative 4c.5
(Ki = 56 nM), while it decreased with the meta-toluidino
derivative 4c.6. Also the new pyridylamino substituent of
4c.10 induced a high receptor affinity (Ki = 45 nM) and
a good selectivity. As for the four derivatives bearing
branched aliphatic amines not previously tested by us,
coming from the simplification or opening of the cyclo-
pentyl ring, it is interesting to point out the high affinity
of 4c.14 (3-pentylamino derivative, Ki = 55 nM) and
4c.12 (2-butylamino derivative, Ki = 81 nM).
5.1. Chemistry
Melting points were determined on a Kofler hot-stage
and are uncorrected. IR spectra in nujol mulls were
recorded on a Perkin-Elmer Mod. 1310 spectrometer.
Mass spectra were performed with a Hewlett Packard
MS/System 5988. Elemental analyses (C, H, N) were
within ± 0.4% of theoretical values and were performed
on a Carlo Erba Elemental Analyzer Mod. 1106 appara-
tus. Optical rotations were measured with a Violet AA-5
polarimeter.
5.2. 2-Azidomethyl-thiophene
NaN3 (7.68 g, 118 mmol) was added to a solution of
2-chloromethyl-thiophene (5.24 g, 39.5 mmol) in 25 mL
of MeOH and 4 mL of H2O, and the suspension was
stirred at room temperature for 24 h. The inorganic
precipitate was removed by filtration and the filtrate was
concentrated, treated with H2O and extracted with
CHCl3. The chloroform extract was dried and evaporated
in vacuo to give the title compound as a yellow oil which
was used without purification: 5.19 g, 94.5% yield. The
azide was short distilled in a tubular oven at 40–45 °C,
1.8 mm Hg; IR (cm–1): 2 105 (N3).
5.2.1. 1-Substituted-4-carboxamido-5-amino-1H-1,2,3-
triazoles 1b–d
0.934 g (11 mmol) of cyanacetamide was added to a
stirred solution of sodium ethoxide (0.253 g, 0.011 g
atom of Na) in 10 mL of absolute EtOH, and stirring was
continued for 30 min. A solution of 10.0 mmol of the
suitable
azide
(2-thenyl-,
2-fluorobenzyl-
or
4-fluorobenzylazide) in 10 mL of absolute EtOH was
slowly added to the suspension, and then the mixture was
heated under reflux for 1.5–2 h. The reaction mixture was
concentrated in vacuo and treated with H2O, and the
insoluble material, consisting of the title compounds, was
collected and purified by crystallization (table I).
Finally, as regards the 4-fluorobenzyl substituent used
for the preparation of the triazolopyrimidine derivatives
4d.1–12, the results reported in table II show that inhibi-
tion constants Ki < 100 nM were obtained for 4d.1

874
5.2.2. 3-Substituted-7-hydroxy-1,2,3-triazolo[4,5-d]pyri-
midines 2b–d
A solution of 10.0 mmol of the suitable triazole deriva-
tive (1b, 1c or 1d) in 8 mL of formamide was refluxed for
2 h. After cooling the reaction mixture was diluted with
H2O and the solid precipitate was collected by filtration
and recrystallized (table I).
cyclopentyl-, cyclohexyl-, (±)-α-methylphenethyl-, (±)-
α-methylbenzylamine, para- or meta-toluidine) in 10 mL
of absolute EtOH was heated under reflux for 2.5 h. The
reaction mixture was evaporated in vacuo and the residue
was treated with H2O and purified by crystallization to
give compounds 4b.1, 4b.2, 4b.5, and 4b.6. For the
isolation of 4b.3 and 4b.4, the residue was extracted with
Et2O, and then Et2O-HCl was added to the ether solution
to precipitate the derivatives as hydrochlorides which
were collected and crystallized (table I).
5.2.3. 3-Substituted-7-chloro-1,2,3-triazolo[4,5-d]pyri-
midines 3b–d
0.8 ml of DMF and 4.5 mL of SOCl2 were added to a
suspension of 5.0 mmol of the suitable triazolopyrimidine
(2b, 2c or 2d) in 22 mL of boiling anhydrous CHCl3. The
reaction mixture was refluxed for 2 h, the solvent was
evaporated in vacuo (temperature ≤ 35 °C), and the
residue, after cooling at 0 °C, was triturated with crushed
ice. The solid formed was collected by filtration, dried
and extracted repeatedly with boiling 60–80 °C petro-
leum ether. The combined extracts were evaporated in
vacuo to give the title compounds as white solids (ta-
ble I).
5.2.6. 3-(2-Fluorobenzyl)-7-(substituted amino)-1,2,3-
triazolo[4,5-d]pyrimidines 4c.1–14
A mixture of 3c (0.30 g, 1.14 mmol), triethylamine
(0.16 mL, 1.15 mmol) and the suitable amine (1.40 mmol
of cyclopentyl-, cyclohexyl-, (±)-α-methylphenethyl- or
(–)-α-methylphenethylamine; 4.0 mmol of para or meta-
toluidine; 2.2 mmol of trans-2-cyclohexanol-, furfuryl-,
4-bromophenyl-, 2-pyridyl-, 2-butyl-, 2-pentyl- or
3-pentylamine) in 10 mL of absolute EtOH (10 mL of
toluene for the pyridylamino derivative 4c.10) was heated
under reflux for 2.5 h. For the isolation of 4c.5, 4c.6, 4c.8,
4c.9 and 4c.14, the reaction mixture was allowed to cool
and the crystallized precipitate was collected by filtration
(table I). For the isolation of 4c.1, 4c.2, 4c.7 and 4c.10,
the reaction mixture was evaporated in vacuo, the residue
was treated with H2O and 10% HCl and the insoluble
material was collected by filtration and crystallized (ta-
ble I). For the isolation of 4c.3, 4c.4, 4c.12 and 4c.13, the
reaction mixture was evaporated in vacuo, the residue
was washed with H2O and 5% HCl and dissolved in Et2O
or absolute EtOH or MeOH. Addition of Et2O-HCl to the
solution precipitated the compounds as hydrochlorides
which were purified by crystallization (table I). The
isopropylamino derivative 4c.11 was obtained by heating
the 7-chloro-triazolopyrimidine 3c (0.39 g, 1.48 mmol) in
2 mL of isopropylamine at 100–110 °C in a closed tube
for 2.5 h and treating the reaction mixture with H2O to
precipitate the expected derivative (table I).
5.2.4. 3-(4-Chlorobenzyl)-7-(substituted amino)-1,2,3-
triazolo[4,5-d]pyrimidines 4a.1–7
A
mixture of 3-(4-chlorobenzyl)-7-chloro-1,2,3-
triazolo[4,5-d]pyrimidine 3a (0.40 g, 1.43 mmol), tri-
ethylamine (0.24 mL, 1.70 mmol) and the suitable amine
(1.70 mmol of cyclopentyl-, cyclohexyl-, (±)-α-methyl-
phenethylamine, 3.70 mmol of (±)-α-methylbenzyl-
amine, 5.0 mmol of para- and meta-toluidine or
1.0 mmol of para-nitroaniline) in 10 mL of absolute
EtOH was refluxed for 2.5 h. For the isolation of com-
pounds 4a.1 and 4a.2, the reaction mixture was evapo-
rated in vacuo, the residue was treated with H2O and 5%
HCl (pH 3) and the insoluble material was collected
and purified by crystallization (4a.1) or by extraction
with 60–80 °C petroleum ether (4a.2) respectively. For
the isolation of compounds 4a.3, 4a.5, 4a.6, and 4a.7, the
reaction mixture was allowed to cool and the crystallized
precipitate was collected by filtration. Isolation of 4a.7
required further treatment of the precipitate with 10%
HCl to remove unreacted p-nitroaniline. For the isolation
of 4a.4, the reaction mixture was evaporated in vacuo, the
residue was dissolved in MeOH and the derivative was
precipitated as a hydrochloride by addition of Et2O-HCl
(table I).
5.2.7. 3-(4-Fluorobenzyl)-7-(substituted amino)-1,2,3-
triazolo[4,5-d]pyrimidines 4d.1–12
A mixture of 3d (0.30 g, 1.14 mmol), triethylamine
(0.16 mL, 1.15 mmol) and the suitable amine (1.40 mmol
of cyclopentyl-, cyclohexyl-, (±)-α-methylphenethyl- or
furfurylamine; 4.0 mmol of para or meta-toluidine;
2.3 mmol of trans-2-cyclohexanol-, 4-bromophenyl-,
2-pyridyl-, 2-butyl-, 2-pentyl- or 3-pentylamine) in
10 mL of absolute EtOH (10 mL of toluene for the
pyridylamino derivative 4d.9) was heated under reflux
for 2.5 h. The reaction mixture was evaporated in vacuo,
5.2.5.
3-(2-Thenyl)-7-(substituted
amino)-1,2,3-tri-
azolo[4,5-d]pyrimidines 4b.1–6
A mixture of 3b (0.25 g, 1.0 mmol), triethylamine
(0.17 mL, 1.2 mmol) and the suitable amine (1.2 mmol of

875
the residue was washed with H2O and 5% HCl (pH 3)
and the insoluble material was collected and purified by
crystallization (4d.1, 4d.6, 4d.10, 4d.11 and 4d.12) or by
extraction with 60–80 °C petroleum ether (4d.2) (table I).
For the isolation of 4d.3, the liquid residue obtained after
the rinses was dissolved in Et2O or MeOH, and Et2O-HCl
was added to the solution to precipitate the derivative as
a hydrochloride (table I). For the isolation of 4d.4, 4d.5,
4d.7, 4d.8 and 4d.9 the reaction mixture was allowed to
cool and the crystallized precipitate was collected by
filtration (table I).
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Acknowledgements
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We wish to thank the Consiglio Nazionale delle
Ricerche (C.N.R.) for financial support.
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