Synthesis, reactions, and antimicrobial activity of some fused thieno[2,3-d]pyrimidine derivatives

Article abstract of DOI:10.1080/10426500701247151

Some new indeno[1',2' :4,5]thieno[2,3-d]pyrimidines were prepared from the reaction of 2-aminoindeno[2,1-b]thiophene-3-carbonitrile (1) with different reagents. Also, some indeno[1',2' :4,5]thieno[3,2-e]tetrazolo[1,5-c]pyrimidines and indeno[1',2' :4,5]th

Full text of DOI:10.1080/10426500701247151

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Synthesis, Reactions, and
Antimicrobial Activity of Some
Fused Thieno[2,3-d]pyrimidine
Derivatives
Mohamed I. Hegab a , Nasser A. Hassan a , Aymn E.
Rashad a , Afaf A. Fahmy a & Farouk M. E. Abdel-
Megeid a
a National Research Centre, Dokki, Cairo, Egypt
Version of record first published: 07 Jun 2007.
To cite this article: Mohamed I. Hegab , Nasser A. Hassan , Aymn E. Rashad , Afaf A.
Fahmy & Farouk M. E. Abdel-Megeid (2007): Synthesis, Reactions, and Antimicrobial
Activity of Some Fused Thieno[2,3-d]pyrimidine Derivatives, Phosphorus, Sulfur, and
Silicon and the Related Elements, 182:7, 1535-1556
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Phosphorus, Sulfur, and Silicon, 182:1535–1556, 2007
Copyright © Taylor & Francis Group, LLC
ISSN: 1042-6507 print / 1563-5325 online
DOI: 10.1080/10426500701247151
Synthesis, Reactions, and Antimicrobial Activity of Some
Fused Thieno[2,3-d]pyrimidine Derivatives
Mohamed I. Hegab
Nasser. A. Hassan
Aymn E. Rashad
Afaf A. Fahmy
Farouk M. E. Abdel-Megeid
National Research Centre, Dokki, Cairo, Egypt
Some new indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidines were prepared from the
reaction of 2-aminoindeno[2,1-b]thiophene-3-carbonitrile (1) with different
reagents. Also, some indeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e]tetrazolo[1,5-c]pyrimidines and
indeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidines and their isomeric
[1,2,4]triazolo[1,5-c]pyrimidines were prepared. The antimicrobial evaluation of
some prepared products showed that many of them revealed promising antimicro-
bial activity.
Keywords Antimicrobial evaluation; indenothienopyrimidines; rearrangement;
thienotetrazolopyrimidines; thienotriazolopyrimidines
INTRODUCTION
Thieno[2,3-d]pyrimidine derivatives represent one of the most ac-
tive classes of compounds possessing a wide spectrum biological
activity. They bear structure analogy and isoelectronic relation to
purine.1 Besides, some active compounds have been described when
the thiophene moiety is fused to a carbocyclic ring,2 also prominent
biological activities have been reported for fused thienopyrimidine
derivivatives.3,4 So, this article is aimed at the synthesis of new
compounds related to indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidine deriva-
tives, their fused [1ꢀ,2ꢀ:4,5]thieno[3,2-e]tetrazolo[1,5-c]pyrimidines,
Received July 27, 2006; accepted December 4, 2006.
We wish to thank the National Research Centre, Dokki,Cairo, Egypt, for financing and
for all the facilities provided to us, and we wish also to thank Dr. I. Shaban, Department
of Chemistry of Natural Products, NRC for biological evaluation.
Address correspondence to Farouk Abdel-Megeid, National Research Centre, EL-
Bhoos Street, 12622, Dokki, Cairo, Egypt. E-mail: faroukezat@yahoo.com
1535

1536
M. I. Hegab et al.
indeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine
deriva-
tives, and their isomeric [1,2,4]triazolo[1,5-c]pyrimidines, and the
study of some of their reactions, as well as the evaluation of some of
their antimicrobial activities.
RESULTS AND DISCUSSION
When a mixture of 2-aminoindeno[2,1-b]thiophene-3-carbonitrile (1)6
and conc. sulfuric acid was stirred at room temperature or when heated
in a mixture of polyphosphoric acid /orthophosphoric acid (1:1), hydroly-
sis of the cyano group took place to give 2-aminoindeno[2,1-b]thiophene-
3-carboxylic acid amide (2) (Scheme 1).
SCHEME 1
Refluxing of compound 1 with benzoyl isothiocyanate, led to the for-
mation of 2-(3-benzoylthioureido)indeno[2,1-b]thiophene-3-carbonitrile
(3), which was annulated to a pyrimidine derivative by refluxing
its ethanolic sodium hydroxide solution to give the tetracyclic prod-
uct, 4-aminopyrimidine-2(1H)-thione derivative 4 (Scheme 1). The 4-
pyrimidinone derivatives were prepared by reacting compound 1 with
formic acid or with acetic acid/conc. hydrochloric acid (1:1) under re-
flux temperature to give derivatives 5a or 5b, respectively (Scheme 1).

Fused Thieno[2,3-d]pyrimidine Derivatives
1537
Products 5a and 5b were formed presumably via the intermediacy of the
corresponding oxazinimine derivative,2,7 which then rearranged under
the conditions of the reaction. Besides, products 5a and 5b were also
obtained by treating compound 2 with triethyl orthoformate or triethyl
orthoacetate, respectively under reflux temperature.
Products 5a and 5b were converted into their corresponding 4-chloro
derivatives 6a and 6b, respectively by treating with phosphorus oxy-
chloride under reflux temperature (Scheme 1).
Compounds 6a and 6b were converted into their corresponding 4-
thiono derivatives by refluxing their ethanolic solution with thiourea
8
following the procedure of Shiba et al. to give products 7a and 7b,
respectively (Scheme 1).
When compounds 1a or 1b were refluxed with triethyl orthoformate
or triethyl orthoacetate, they afforded derivatives 8a or 8b, respectively
(Scheme 1). On the other hand, when the ethanolic solutions of 8a or
8b were stirred at room temperature with hydrazine hydrate, they af-
forded the 4-iminopyrimidin-3-ylamine derivatives 9a or 9b, respec-
tively. However, the pyrimidin-4-ylhydrazine derivatives 10a and 10b
were obtained by treatment of compounds 6a or 6b and 8a or 8b with
hydrazine hydrate under reflux temperature (Scheme 2).
SCHEME 2

1538
M. I. Hegab et al.
Also, compounds 9a and 9b were isomerized to their corresponding
more stable 4-hydrazino derivatives 10a and 10b, respectively upon
refluxing in ethanol in the presence of hydrazine hydrate. Actually,
hydrazine hydrate acts as a base in this Dimroth type rearrangement,
which involves a sequence of ring opening and ring closure reactions.9,10
The synthesis of 4-amino pyrimidine derivatives was achieved
by heating compound 8b with methylamine to give the N-
methylpyrimidin-4-ylamine derivative 11, or by heating compounds
8a or 8b with ammonium hydroxide solution to give the pyrimidin-
4-ylamine derivatives 12a and 12b, respectively.
The displacement of the 4-chlorine atom of both compounds 6a
or 6b in glacial acetic acid with sodium azide, afforded the poly-
cyclic tetrazolo[1,5-c]pyrimidine derivatives 13a and 13b, respectively
as judged by the IR spectra, since both compounds did not show
the characteristic azide absorption bands near the region at ν 2100–
2200 cm−1 11−13
.
The reduction of compounds 13a or 13b was accomplished by treat-
ment of both compounds in glacial acetic acid with zinc dust, to afford
the pyrimidin-4-ylamine derivatives 12a and 12b, respectively; pre-
sumably via the formation of the azido intermediates 14a and 14b
(Scheme 2).
Many reports14,15 have described the synthesis of [1,2,4]triazolo[4,3-
c]pyrimidine derivatives and [1,2,4]triazolo[1,5-c]pyrimidine fused
to different nitrogen containing heterocyclic moieties. In the mean-
time, the synthesis of [1,2,4]triazolo[4,3-c]pyrimidine derivatives and
[1,2,4]triazolo[1,5-c]pyrimidine derivatives fused to a thiophene moiety
have attracted the attention of many investigators.3,10,16−18 Actually,
previous observations revealed that thieno[3,2-e][1,2,4]triazolo[4,3-
c]pyrimidines can isomerizes under different suitable reaction
conditions to the more stable thieno[3,2-e][1,2,4]triazolo[1,5-
c]pyrimidines.10,19 Nevertheless, this pattern of isomerization appears
to have been overlooked by some workers.14,16,20
In this investigation, when compounds 10a or 10b were treated with
triethyl orthoacetate (TEOA)] under reflux temperature, they afforded
the polycyclic [1,2,4]triazolo[4,3-c]pyrimidine derivatives 15a and 15b,
respectively. On the other hand, when the 4-imino-3-amino derivatives
9a or 9b were treated with a mixture of acetic acid/acetic anhydride
(1:1) they gave the [1,2,4]triazolo[1,5-c]pyrimidine derivatives 16a and
16b, respectively. In the meantime, compounds 16a and 16b could be
obtained by isomerization of compounds 15a and 15b, respectively upon
treatment with a base under reflux temperature (Scheme 3). It is note-
worthy to mention that the1H NMR spectra, in general, revealed that
the signals of the C3-H, or C3-CH3 and C5-H or C5-CH3 protons of the

Fused Thieno[2,3-d]pyrimidine Derivatives
1539
SCHEME 3
thieno[3,2-e][1,2,4]triazolo[4,3-c]pyrimidine derivatives whenever they
exist in this text are more deshielded and appeared at a more down-
field when compared with the corresponding protons of the thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidines derivatives (c.f. Exp.). These obser-
vations are in accordance with the results of Shishoo et al.16,17 Actually,
this type of isomerization to a more stable products in the presence of
a catalytic amount of sodium ethoxide, was also described by some au-
thors as a Dimroth type rearrangement,10,21 which presumably involves
a sequence of ring opening and ring closure reactions.
When compound 10b was treated with triethyl orthoformate
(TEOF) under reflux temperature it afforded the [1,2,4]triazolo[4,3-
c]pyrimidine derivative 17b.

1540
M. I. Hegab et al.
Moreover, compounds 9a or 9b and 10a or 10b were treated with
formic acid under reflux temperature, in both cases they gave one and
the same identical product in all respects assigned the structure of
the [1,2,4]triazolo[1,5-c]pyrimidine derivative 18a or 18b, respectively.
In fact, the [4,3-c]triazolo products could not be isolated even when
the reaction mixture was heated at low temperature (40–50◦C) and at
short time, since tlc monitoring revealed the formation of more than one
spot during the reaction course until the reaction ended by formation
of one spot giving the final product. It is noteworthy to mention that
the [1,2,4]triazolo[4,3-c]pyrimidine 17b, which was obtained before,
rearranged into its isomeric[1,2,4]triazolo[1,5-c]pyrimidine deriva-
tive 18b upon heating with formic acid under reflux temperature
(Scheme 3).
Also, the [1,2,4]triazolo[1,5-c]pyrimidine-2(1H)-thione derivative 20
was prepared by reacting compound 9b or 10b in dry pyridine with car-
bon disulfide under reflux temperature (presumably via the intermedi-
acy of the triazolo[4,3-c] isomer 19 in case of the reaction of compound
10b, which then rearranged under the conditions of the reaction to the
triazolo[1,5-c] product 20.
Moreover, a series of triazolo[1,5-c]pyrimidine derivatives was pre-
pared by reacting compounds 9b or 10b with some bifunctional
one-carbon donor cyclizing agents. So, when compounds 9b or 10b,
in ethanol, were treated with benzaldehyde in the presence of few
drops of acetic acid under reflux temperature or when compound
9b, in dioxane, was treated with benzoyl chloride in the presence
of a few drops of triethylamine under reflux temperature, they
afforded in both cases one and the same product: the 2-phenyl-
[1,2,4]triazolo[1,5-c]pyrimidine derivative 21. Also, when compound 9b
was treated with chloroacetyl chloride, α-bromopropionyl bromide in
the presence of few drops of triethyl amine (TEA), or dichloroacetyl
chloride under reflux temperature, it gave the corresponding 2-
substituted 5-methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e][1,2,4]triazolo[1,5-c]
pyrimidine derivatives 22a,b, and 23, respectively (Scheme 4).
Compounds 2, 4, 6a,7a,7b,10a,21, 22b, and 23 were selected to rep-
resent the newly synthesized compounds and tested for their antimi-
crobial activity. Streptomycin (S), Amoxicillin (Am), and Fusidic acid
(FA) were used as reference drugs and the biological results can be
summarized in Table I.
CONCLUSION
Starting from 2-aminoindeno[2,1-b]thiophene-3-carbonitrile (1), we
have demonstrated a convenient procedure for the preparation of some

Fused Thieno[2,3-d]pyrimidine Derivatives
1541
SCHEME 4
new indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidines, indeno [1ꢀ,2ꢀ:4,5]thieno-
[3,2-e]tetrazolo[1,5-c]pyrimidines and indeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e]-
[1,2,4]triazolo[4,3-c]pyrimidines and their isomeric [1,2,4]triazolo[1,5-
c]pyrimidines were prepared. As shown in Table I, the 2-substituted-
5-methyindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e][1,2,4]triazolo[1,5-c]pyrimidine

1542
M. I. Hegab et al.
TABLE I Test of the Synthesized Compounds
The tested compoundsa
C. albicans
S.aures
M.phlei
B.subtilis
E.coli
(S)b
(Am)c
(FA)d
2
4
6a
7a
7b
10a
21
22b
23
++++
−
−
−
−
+
++++
−
++++
+++
−
−
+++
−
+
++++
+
−
−
+++
++
−
++
+
++
−
+
+
−
−
−
−
−
−
+
++
+++
−
−
+
−
+++
+
+
+
−
−
−
++
++
+++
++
++
+++
−
−
−
++
+
++
–1
aγ = 2 µg mL–1 in DMSO; bγ = 25 µg/mL−1; cγ = 4 µg mL
;
dγ = 4 µg/mL−1
;
+++—Highly sensitive (inhibition zone = 21–25 mm); ++—Fairly sensitive (inhibition
zone = 16–20 mm); +—Slightly sensitive (inhibition zone =10–15 mm); and- —Not
sensitive (inhibition zone = 0–9 mm).
derivative 23 revealed the highest antimicrobial activity than the
other tested compounds and some of the reference drugs.
EXPERIMENTAL
All melting points were uncorrected and measured using Electro-
thermal IA 9100 apparatus (Shimadzu, Japan). IR spectra were
recorded as potassium bromide pellets on a Perkin-Elmer 1650 spec-
trophotometer (Shimadzu, Japan).1H NMR and 13C NMR spectra were
recorded on Varian Gemini 180 spectrometer (Varian, UK), and chemi-
cal shifts were expressed as (ppm) values against TMS as internal ref-
erence. Mass spectra were recorded on Gc Ms-QP 1000 EX (Shimadzu,
Japan). Column Chromatography was performed on (Merck) Silica gel
60 (particle size 0.06–0.20 mm).
Synthesis of 2-Aminoindeno[2,1-b]thiophene-3-carbonitrile (1)
Method A
A mixture of 1-indanone (1.32 g, 0.01 mol), malononitrile (0.66 g, 0.01
mol) and elemental sulfur (0.32 g, 0.01 mol) in pyridine (20 mL) was
heated under reflux temperature for 3 h, cooled, poured into ice-water
with stirring for 1 h, neutralized with conc. hydrochloric acid, filtered,

Fused Thieno[2,3-d]pyrimidine Derivatives
1543
dried off, and purified on silica gel using petroleum ether 60–80◦C :
ethyl acetate (4:1) to give compound 1.
Method B
Step 1. A mixture of 1-indanone (1.32 g, 0.01 mol), malononitrile
(0.66 g, 0.01 mol), ammonium acetate (5 g), glacial acetic acid (20 mL)
and dry benzene (50 mL) was heated under reflux temperature for 6 h
using a Dean-Stark water separator, cooled, filtered, dried, and recrys-
tallized from ethanol to give 1-indanylidenmalononitrile (1.50 g, 83%).
1
IR; v 2280, 2200 (NH2), 2210 (CN). H NMR: δ 3.75 (s, 2H, CH2), 4.80
(s, 2H, NH2, D2O exchangeable), 7.20–7.60 (m, 3H, Ar H), 7.75 (d, J =
6.00 Hz, 1H, Ar H). MS: m/z 212 (M+, 100), 196 (1.71), 184 (4.20).
Step 2. A mixture of 1-indanylidenmalononitrile (1.80 g, 0.01 mol)
and elemental sulfur (0.32 g, 0.01 mol) was heated under reflux temper-
ature in 20 mL dry pyridine for 3 h, cooled, poured into ice-water with
stirring for 1 h, neutralized with conc. hydrochloric acid, filtered, dried,
and purified as in method A to give compound 1. Compound 1 prepared
by this method is identical in all respects (physical and spectral data)
to those prepared by method A.
Synthesis of 2-Aminoindeno[2,1-b]thiophene-3-carboxylic
acid amide (2)
Method A
Compound 1 (0.212 g, 1 mmol) was added portion wise to 20 mL
conc. sulfuric acid, with stirring for 1 h. The reaction mixture was
poured into ice-water, neutralized, filtered, dried, and recrystallized
from DMF to give compound 2. Yield 87%, M. p. 196–198◦C. IR; v 3470,
3396 (CONH2), 3290, 3196 (NH2), 1642 (C O).1H NMR: δ 3.60 (s, 2H,
CH2), 3.80–4.20 (br s, 2H, NH2, D2O exchangeable), 6.60 (s, 2H, CONH2,
D2O exchangeable), 7.00–7.50 (m, 3H, Ar H), 7.80 (d, J = 10.80 Hz, 1H,
Ar H). MS: m/z 230 (M+, 81.80), 213(100), 185 (38.00).
Method B
Compound 1 (0.212 g, 1 mmol) was added to a mixture of 20 mL or-
thophosphoric acid:polyphosphoric acid (1:1) with stirring for 1 h at
room temperature, then the reaction mixture was refluxed for 6 h,
cooled, poured into water, filtered, dried, and recrystallized from DMF
to give compound 2. Compound 2 prepared by this method is identi-
cal in all respects (physical and spectral data) to those prepared by
method A.

1544
M. I. Hegab et al.
Synthesis of 2-(3-Benzoylthioureido)indeno[2,1-b]-
thiophene-3-carbonitrile (3)
Compound 1 (0.212 g, 1 mmol) was added to 50 mL dry acetone contain-
ing 1 mmol of benzoyl isothiocyanate (prepared in situ from potassium
isothiocyanate and benzoyl chloride) then the reaction mixture was re-
fluxed for 2 h, cooled, filtered, dried, and recrystallized from DMF to
give compound 3. Yield 75%, M. p. 248–250◦C. IR: v 3292 (NH), 3245
(NH), 2211 (CN), 1664 (C O). MS: m/z 375 (M+, .87), 316 (43.79), 251
(10.50), 211(4.87), 195 (2.47). Anal. Calcd. For C20H13N3OS2 (375); C,
64.30; H, 3.60; N, 11.03; S, 17.30%. Found: C, 64.00; H, 3.46; N, 11.20;
S, 17.06.
Synthesis of 4-Aminoindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidine-
2(1H)-thione (4)
Compound 3 (0.75 g, 2 mmol) was refluxed in 10 mL ethanol contain-
ing 5.5 ml in sodium hydroxide for 1 h, cooled, poured into water, and
neutralized with drops of conc. hydrochloric acid, filtered, dried, and re-
crystallized from DMF to give compound 4. yield 91%, M. p. 318–20◦C.
IR: v 3501, 3402 (NH2), 3187 (NH), 1677 (C N).1H NMR: δ 3.90 (s, 2H,
CH2), 6.00–6.30 (s, 2H, NH2, D2O exchangeable); 7.10–7.60 (m, 4H, 3
Ar H, NH, D2O exchangeable), 7.90 (d, J = 10.80 Hz, 1H, Ar H); 13C
NMR (DMSO-d6, ppm): 35.11 (C-9), 120.66–126.65 (Ar C), 136.69 (C-
4b), 137.10 (C-9a), 137.40 (C-10a), 145.90 (C-4a), 153.87 (C-4), 174.00
(C S). MS: m/z 271(M+, 35.46), 212 (23.5). Anal. Calcd. for C13H9N3S2
(271.36); C, 57.44; H, 3.16; N, 15.22%. Found: C, 57.53; H, 3.34; N, 15.48.
Synthesis of Indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidin-4(3H)-one
(5a)
Method A
Compound 1 (0.212 g, 1 mmol) was heated under reflux temperature
in 20 mL formic acid for 6 h. The reaction mixture was cooled, poured
into water, filtered, dried, and the residue was recrystallized from diox-
ane/methanol (1:1) to give compound 5a. Yield 92%, M. p. 310–312◦C.
IR: v 3200 (NH), 1700 (C O), 1590 (C N). 1H NMR: δ 4.00 (s, 2H, CH2),
7.20–7.50 (m, 3H, Ar H), 8.10 (s, 1H, C2-H), 8.40 (d, J = 9.45 Hz, 1H,
Ar H), 12.60 (s, 1H, NH, D2O exchangeable); 13C NMR (DMSO-d6): δ
35.30 (C-9), 118.90–126.74 (Ar-C), 138.27 (C-4a), 140.00 (C-4b), 140.23
(C-9a), 145.50 (C-10a), 168.50 (C O). MS: m/z 240 (M+, 100), 212 (5.67),
197(1.48), 185 (15.53). Anal. Calcd. for C13H8N2OS (240); C, 64.94; H,
3.40; N, 11.42; S, 13.63%. Found: C, 65.00; H, 3.30; N, 11.66; S, 13.33.

Fused Thieno[2,3-d]pyrimidine Derivatives
1545
Method B
Compound 5a was also obtained by treating compound 2 (0.23 g, 1
mmol) with 20 mL triethyl orthoformate in the presence of few drops
of piperidine under reflux temperature for 4 h, then the reaction mix-
ture was cooled, filtered, dried, and the residue was recrystallized from
dioxane/methanol (1:1) to give compound 5a. Compound 5a prepared
by this method is identical in all respects (physical and spectral data)
to those prepared by method A.
Synthesis of 2-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidin-
4(3H)-one (5b)
Method A
A mixture of 20 mL glacial acetic acid/conc. hydrochloric acid (1:1)
and compound 1 (0.212 g, 1 mmol) was heated under reflux temperature
for 3 h, filtered while hot, washed with water, then methanol, dried, and
recrystallized from DMF to give compound 5b. Yield 79%, M. p. 290–
292◦C. IR: v 3250 (NH), 1665 (C O), 1590 (C N). 1H NMR (DMSO-d6):
δ 2.66 (s, 3H, C2-CH3), 3.90 (s, 2H, CH2), 7.20–7.50 (m, 3H, Ar H), 8.50
(d, J = 12.15 Hz, 1H, Ar H), 11.60 (s, 1H, NH, D2O exchangeable). MS:
m/z 254 (M+, 100), 213 (13.15), 195 (0.88), 183 (9.20). Anal. Calcd. for
C14H10N2OS (254.31); C, 66.44; H, 4.10; N, 10.96; S, 12.61%. Found:
C,66.14; H, 3.93; N, 11.02; S, 12.59.
Method B
Compound 5b was also obtained by treating compound 2 (0.23 g, 1
mmol) with 20 mL triethyl orthoacetate in the presence of few drops
of piperidine under reflux temperature for 3 h, then the reaction mix-
ture was cooled, filtered, dried, and the residue was recrystallized from
DMF to give compound 5b. Compounds 5b prepared by this method are
identical in all respects (physical and spectral data) to those prepared
by method A.
Synthesis of 4-Chloroindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidine
(6a)
Compound 5a (0.24 g, 1 mmol) was heated under reflux temperature in
20 mL phosphorus oxychloride for 5 h, then the reaction mixture was
poured into ice-water, stirred, filtered, and purified on silica gel using
petroleum ether (40–60◦C) : ethyl acetate (4 : 1) to give compound 6a.
1
Yield 89%, M. p. 152–154◦C. H NMR: δ 4.20 (s, 2H, CH2), 7.20–7.60
(m, 3H, Ar H), 8.45 (d, J = 10.80 Hz, 1H, Ar H), 8.90 (s, 1H, C2-H). MS:

1546
M. I. Hegab et al.
m/z 260 (M+, Cl37, 47.50), 258 (M+, Cl35, 100), 223 (13.76), 195 (32.85).
Anal. Calcd. for C13H7ClN2S (258.00); C, 60.54; H, 2.90; N, 10.59; S,
12.11%. Found: C, 60.46; H, 2.71; N, 10.85; S, 12.40.
Synthesis of 4-Chloro-2-methylindeno[1ꢀ,2ꢀ:4,5]-
thieno[2,3-d]pyrimidine (6b)
Compound 5b (0.254 g, 1 mmol) was heated under reflux temperature
in 20 mL phosphorus oxychloride for 6 h, then the reaction mixture
was cooled, poured into ice-water. The solid that formed was filtered
off, dried, and purified on silica gel using petroleum ether (40–60◦C) :
ethyl acetate (4:1) as an eluent to give compound 6b. Yield 55%, M. p.
1
168–170◦C. H NMR: δ 2.85 (s, 3H, C2-CH3), 4.00 (s, 2H, CH2), 7.30–
7.60 (m, 3H, Ar H), 8.50 (d, J = 10.80 Hz, 1H, Ar H). MS: m/z 274
(M+, Cl37, 39.91), 272 (M+, Cl35, 100), 237 (65.12), 30 (52.04), 10 (4.49),
195 (77.69). Anal. Calcd. for C14H9ClN2S (272.50); C, 61.76; H, 3.30; N,
10.29; S, 11.76%. Found: C, 61.76; H, 3.30; N, 10.29; S, 11.76.
Synthesis of Indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidine-4(3H)-
thione (7a)
A mixture of compound 6a (0.258 g, 1 mmol) dissolved in 20 mL absolute
ethanol, thiourea (2 mmol) was heated for 6 h, cooled, filtered, washed
with a little amount of methanol, dried, and recrystallized from DMF
1
to give compound 7a. Yield 78%, M. p. 320–322◦C. H NMR: δ 4.00
(s, 2H, CH2), 7.20–7.50 (m, 3H, Ar H), 8.10 (d, J = 10.80 Hz, 1H, Ar
H), 9.40 (s, 1H, C2-H), 13.90 (s, 1H, NH, D2O exchangeable). 13C NMR
(DMSO-d6): δ 35.20 (C-9), 124.40–129.60 (Ar C), 138.40 (C-4a), 140.50
(C-4b), 143.60 (C-9a), 144.18 (C-10a), 145.70 (C2), 177.50 (C S). MS:
m/z 256 (M+, 100), 240 (3.36), 228 (14.45), 196 (21.45). Anal. Calcd. for
C13H8N2S2 (256.00); C, 61.30; H, 3.14; N, 11.00; S, 25.07%. Found: C,
60.93; H, 3.12; N, 10.93; S, 25.00.
Synthesis of 2-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidine-
4(3H)-thione (7b)
Compound 6b (0.272 g, 1 mmol) was dissolved in 30 mL absolute
ethanol, then thiourea (2 mmol) was added and the reaction mixture
was refluxed for 4 h, cooled, filtered, washed with a little amount of wa-
ter, then ethanol, dried, and recrystallized from DMF to give compound
7b. Yield 93%, M. p. 324–326◦C. 1H NMR: δ 3.40 (s, 3H, C2-CH3), 4.00
(s, 2H, CH2), 7.20–7.50 (m, 3H, Ar H), 9.40 (d, J = 10.80 Hz, 1H, Ar

Fused Thieno[2,3-d]pyrimidine Derivatives
1547
H), 13.80 (s, 1H, NH, exchangeable with D2O). 13C NMR (DMSO-d6):
δ 20.50 (CH3), 35.20 (C-9), 124.30–126.30 (Ar C), 133.00 (C-4a), 138.00
(C-4b), 140.00 (C-9a), 146.00 (C-10a), 154.00 (C-2), 178.00 (C S). MS:
m/z 270 (M+, 71.70), 253 (1.45), 228 (100), 211 (22.15), 196 (21.19), 184
(4.50). Anal. Calcd. for C14H10N2S2 (270.00); C, 62.60; H, 3.45; N, 11.00;
S, 10.23%. Found: C, 62.22; H, 3.70; N, 10.37.
Synthesis of Ethyl N-{3-cyanoindeno[2,1-b]thien-2-yl}-
methanimidate (8a)
A mixture of compound 1 (0.212 g, 1 mmol) and 20 mL triethyl ortho-
formate was heated under reflux temperature for 4 h, then evaporated
under reduced to give compound 8a. Yield 93%, M. p. 134–136◦C. IR
(KBr): v 3265 (NH).1H NMR: δ1.40 (t, J = 8.45 Hz, 3H, CH3), 3.80 (s,
2H, CH2), 4.40 (q, J = 1.80 Hz, 2H, OCH2), 7.20–7.50 (m, 3H, Ar H),
7.80 (d, J = 8.10 Hz, 1H, Ar H), 8.05 (s, 1H, N=CHOEt). MS: m/z 268
(M+, 1.75), 252 (36.20), 254 (13.17).
Synthesis of 2-Ethyl N-{3-cyanoindeno[2,1-b]thien-2-yl}-
ethanimidate (8b)
A mixture of compound 1 (0.212 g, 1 mmol) and 20 mL triethyl orthoac-
etate was heated under reflux temperature for 5 h, then evaporated
under reduced pressure, and the residue was recrystallized from diox-
ane to give compound 8b. Yield 89%, M. p. 140–142◦C.1H NMR: δ 1.40
(t, J = 8.50 Hz, 3H, CH3), 3.20 (s, 3H, CH3), 3.80 (s, 2H, CH2), 4.35 (q,
J = 8.20 Hz, 2H, OCH2), 7.20-7.50 (m, 3H, Ar H), 7.80 (d, J = 8.10 Hz,
1H, Ar H). MS: m/z 282 (M+, 1.16), 268 (100), 252 (33.80).
Synthesis of
4-Iminoindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidin-3-ylamine (9a)
A mixture of compound 8a (0.268 g, 1 mmol) dissolved in 20 mL of
absolute ethanol and 3 mL of hydrazine hydrate (99%), was stirred for
1 h at room temperature. The solid that formed was filtered, washed
with a little amount of methanol, dried, and recrystallized from ethanol
to give compound 9a. Yield 91%, M. p. 223–225◦C. IR (KBr): v 3250,
3200 (NH2), 3100 (NH), 1645 (C N). 1H NMR: δ 3.95 (s, 2H, CH2), 5.70
(s, 2H, NH2, D2O exchangeable); 7.20–7.50 (m, 4H, 3 Ar H, NH, D2O
exchangeable), 8.40 (d, J = 10.80 Hz, 1H, Ar H), 8.95 (s, 1H, C2-H). MS:
m/z 254 (M+, 100), 238.10(13.70), 211 (42.70), 195 (2.34).

1548
M. I. Hegab et al.
Synthesis of 2-Methyl-4-iminoindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-
d]pyrimidin-3-ylamin (9b)
A mixture of compound 8b (0.282 g, 1 mmol) dissolved in 20 mL absolute
ethanol containing 3 mL of hydrazine hydrate (99 %), was stirred for 1
h at room temperature, the solid that formed was filtered, washed with
a little amount of cold methanol, dried, and recrystallized from ethanol
to give compound 9b. Yield 93%, M. p. 324–326◦C. IR (KBr): v 3295,
1
3200 (NH2), 3150 (NH), 1640 (C N). H NMR: δ 2.60 (s, 3H, C2-CH3),
3.80 (s, 2H, CH2), 4.80 (s, 2H, NH2, D2O exchangeable), 7.20 (s, 1H,
NH, D2O exchangeable), 7.30–7.50 (m, 3H, Ar H), 7.95 (d, J = 10.85,
1H, Hz, Ar H). 268 (M+, 100), 252 (36), 210 (38), 195 (10), 183 (8). Anal.
Calcd. for C14H12N4S (268.00); C, 62.40; H, 4.35; N, 20.60%. Found: C,
62.68; H, 4.47; N, 20.89.
Synthesis of Indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidin-4-
ylhydrazine (10a)
Compound 8a (0.258, g 1 mmol) was dissolved in 20 mL absolute
ethanol, then 2 mL of hydrazine hydrate (99%) were added, and the re-
action mixture was heated under reflux temperature for 3 h; it was then
evaporated under reduced pressure, and the residue was recrystallized
from dioxane to give compound 10a. Yield 79%, M. p. 200–202◦C.1H
NMR: δ 4.00 (s, 2H, CH2), 5.00–5.50 (br. s, 2H, NH2, D2O exchange-
able), 7.10–7.50 (m, 4H, 3 Ar H + NH, D2O exchangeable), 8.05 (d, J =
10.80 Hz, 1H, Ar H), 8.40 (s, 1H, C2-H); 13C NMR (DMSO-d6, δ ppm):
35.00 (C-9), 122.00–127.00 (Ar C), 130.00 (C-4a), 136.00 (C-4b), 138.00
(C-9a), 142.00 (C-10a), 145.00 (C-2), 160.00 (C-4). MS: m/z 254 (M+, 60),
239 (60), 224 (10.70), 210 (50.35), 196 (14.15), 183 (11.89). Anal. Calcd.
for C13H10N4S (254.00); C, 61.20; H, 4.14; N, 22.39; S, 12.40%. Found:
C, 61.41; H, 3.93; N, 22.04; S, 12.59.
Synthesis of 2-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]-
pyrimidin-4-ylhydrazine (10b)
Compound 8b (0.272 g, 1 mmol) was dissolved in 20 mL absolute
ethanol, then 3 mL hydrazine hydrate (99%) were added then and re-
action mixture was heated under reflux temperature for 2 h, evapo-
rated under reduced pressure, and the residue was recrystallized from
ethanol to give compound 10b. Yield 75%, M. p. 213–215◦C. 1H NMR: δ
2.65 (s, 3H, C2-CH3), 3.90 (s, 2H, CH2), 4.30 (s, 2H, NH2, D2O exchange-
able), 6.60 (s, 1H, NH, D2O exchangeable), 7.20–7.60 (m, 3H, Ar H), 7.90
(d, J = 10.80 Hz, 1H, Ar H). MS: m/z 268 (M+, 62.10), 252 (14.73), 238

Fused Thieno[2,3-d]pyrimidine Derivatives
1549
(6.79), 226 (4.13), 210 (100), 196 (34.61), 183 (23.67) Anal. Calcd. for
C14H12N4S (268); C, 62.35; H, 4.50; N, 20.46%. Found: C, 62.68; H, 4.47;
N, 20.89.
Isomerization of 9a and 9b to 10a and 10b
Compounds 9a (0.254 g, 1 mmol) and 9b, (0.268 g, 1 mmol) were dis-
solved in 20 mL ethanol and then drops of hydrazine hydrate were
added, then the reaction mixture was heated under reflux temperature
for 2 h, and evaporated under reduced pressure to give compounds. 10a
(0.24 g, 94%) and compound 10b (0.25 g, 93 %), respectively. Products
10a and 10b obtained from these isomerizations are identical in all
respects with compounds 10a and 10b obtained before.
Synthesis of N-Methyl-2-methylindeno[1ꢀ,2ꢀ:4,5]
thieno[2,3-d]pyrimidin-4-ylamine (11)
A mixture of compound 8b (0.28 g, 1mmol), 20 mL absolute ethanol,
and 10 mL alcoholic methylamine was stirred on a water-bath for 5
h, the reaction mixture was cooled, filtered off, and the residue was
recrystallized from dioxane to give compound 11. Yield 94%, M.p. 168–
1
170◦C. IR (KBr): v 3273 (NH), 1611 (C N). H NMR: δ 2.30 (d, 3H,
NHCH3), 2.40 (s, 3H, C2-CH3), 3.00 (s, 2H, CH2), 3.40 (s, 1H, NH, D2O
exchangeable), 7.20–7.70 (m, 4H, Ar H).
Synthesis of Indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidin-4-ylamine
(12a)
A mixture of compound 8a (0.268, g 1 mmol), in 20 mL absolute ethanol,
and 10 mL ammonium hydroxide solution was stirred at room tem-
perature for 10 min then, for 2 h on a water-bath, evaporated un-
der reduced pressure, and the residue was recrystallized from ethanol
to give indeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]pyrimidin-4-ylamine (12a). Yield
1
80%, M.p. 202–204◦C. H NMR: δ 4.10 (s, 2H, CH2), 7.10 (s, 2H, NH2,
D2O exchangeable); 7.30–7.70 (m, 3H, Ar H), 8.40 (d, J = 11.60 Hz,
1H, Ar H), 8.90 (s, 1H, C2-H). MS: m/z 239 (M+, 100), 211 (21.40), 195
(10.34). Anal. Calcd. for C13H9N3S (239); C, 65.03; H, 4.00; N, 17.30%.
Found: C, 65.20; H, 3.79; N, 17.50.
Synthesis of 2-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[2,3-d]-
pyrimidin-4-ylamine (12b)
A mixture of Compound 8b (0.282 g, 1 mmol), dissolved in 20 mL ab-
solute ethanol, and 10 mL ammonium hydroxide solution was stirred

1550
M. I. Hegab et al.
at room temperature for 10 min, then for 2 h on a water-bath, evapo-
rated under reduced pressure, and the residue was recrystallized from
ethanol to give compound 12b. Yield 87%, M.p. 260–262◦C. IR: v 3453,
418 (NH2), 1661 (C N).1H NMR: δ 2.45 (s, 3H, C2-CH3), 4.30 (s, 2H,
CH2), 4.50-5.00 (br. s, 2H, NH2, D2O exchangeable), 7.20–7.70 (m, 4H,
Ar H).
Synthesis of Indeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e]tetrazolo[1,5-c]-
pyrimidine (13a)
Compound 6a (0.258 g, 1 mmol) was added to 20 mL glacial acetic acid
containing sodium azide (0.13 g, 2 mmol) with stirring at 50–70◦C for 2
h, then, the reaction mixture was cooled, filtered, dried, and the residue
was recrystallized from dioxane to give compound 13a. Yield 79%, M.p.
234–236◦C. 1H NMR: δ 4.25 (s, 2H, CH2), 7.20–7.50 (m, 3H, Ar H), 8.80
(d, J = 10.80 Hz, 1H, Ar H), 9.50 (s, 1H, C5-H). MS: m/z 265 (M+, 35.00),
237(65.63), 236 (100), 210 (49.69).
Synthesis of 5-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e]tetrazolo[1,5-c]pyrimidine (13b)
Compound 6b (0.272 g, 1 mmol) was added to 20 mL glacial acetic acid
containing sodium azide (0.13 g, 2 mmol) with stirring at 50–70◦C for
2 h, the reaction mixture was cooled filtered, dried, and the residue
was recrystallized from dioxane to give compound 13b. Yield 79%, M.p.
1
238–240◦C. H NMR: δ 3.20 (s, 3H, C5-CH3), 4.10 (s, 2H, CH2), 7.20–
7.60 (m, 3H, Ar H), 8.70 (d, J = 10.90 Hz, 1H, Ar H). MS: m/z 279 (M+,
37.75), 250 (100), 224 (8.89), 210 (27.12). Anal. Calcd. for C14H9N5S
(279); C60.50; H, 3.14; N, 25.42%. Found: C, 60.21; H, 3.22; N, 25.08.
Reduction of Tetrazopyrimidines 13a, and 13b to 12a and 12b
To a solution of both compounds 13a (0.265 g, 1 mmol) and 13b (0.279
g, 1 mmol) in 20 mL glacial acetic acid, zinc dust (0.24 g, 3 mmol) was
1
added with stirring for / h at room temperature then refluxed for
2
10 h; the reaction mixture was cooled, poured into water, extracted with
chloroform, the organic layer was evaporated under reduced pressure,
and the residue was recrystallized from ethanol to give indeno[1ꢀ,2ꢀ:
4,5]thieno[2,3-d]pyrimidin-4-ylamine (12a) and 2-methylindeno[1ꢀ,2ꢀ:
4,5]thieno[2,3-d]pyrimidin-4-ylamine (12b), respectively. Products 12a
and 12b are identical in all respects with compound 12a and 12b ob-
tained before.

Fused Thieno[2,3-d]pyrimidine Derivatives
1551
Synthesis of 3-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[4,3-c]pyrimidine (15a)
A mixture of compound 10a (0.254 g, 1 mmol) and 20 mL triethyl or-
thoacetate was heated under reflux temperature for 10 h. The reaction
mixture was cooled, filtered, dried, and the residue was recrystallized
1
from dioxane to give compound 15a. Yield 43%, M.p. 298–300◦C. H
NMR: δ 2.80 (s, 3H, C3-CH3), 4.10 (s, 2H, CH2), 7.20–7.60 (m, 3H, Ar
H), 8.70 (d, J = 11.6 Hz, 1H, Ar H), 9.70 (s, 1H, C5-H). MS: m/z 278
(M+, 100), 263 (0.54), 236 (9.91).
Synthesis of 3,5-Dimethylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[4,3-c]pyrimidine(15b)
Compound 10b (0.268 g, 1 mmol) was heated under reflux in 20 mL
triethyl orthoacetate for 10 h, then evaporated under reduced pressure
and the residue was recrystallized from DMF to give compound 15b.
1
Yield 82%, M.p. 328–330◦C. H NMR: δ 2.95 (s, 3H, C3-CH3), 3.10 (s,
3H, C5-CH3), 4.20 (s, 2H, CH2), 7.20–7.60 (m, 3H, Ar H), 8.80 (d, J =
12.15 Hz, 1H, Ar H). MS: m/z 292 (M+, 100), 277 (1.00), 250 (5.25),
210 (20.70), 195 (4.62), 183 (8.20). Anal. Calcd. for C16H12N4S (292); C,
65.61; H, 4.42; N, 19.25%. Found: C, 65.75; H, 4.10; N, 19.17.
Synthesis of 2-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidine (16a)
Compound 9a (0.254 g, 1 mmol) was heated under reflux temperature in
20 mL glacial acetic acid/acetic anhydride (1:1) for 6 h; then, the reaction
mixture was cooled, poured into water, filtered, dried, and recrystallized
1
from ethanol to give compound 16a. Yield 72%, M.p. 222–224◦C. H
NMR: δ 2.60 (s, 3H, C2-CH3) 4.10 (s, 2H, CH2), 7.20–7.60 (m, 3H, Ar H),
8.55 (d, J = 10.80 Hz, 1H, Ar H), 9.50 (s, 1H, C5-H). MS: m/z 278 (M+,
100), 263 (3), 236(4.9), 210 (33.82), 195 (1.69), 183 (8.20). Anal. Calcd.
for C15H10N4S (278); C, 64.56; H, 3.44; N, 20.32%. Found: C, 64.74; H,
3.59; N, 20.14.
Synthesis of 2,5-Dimethylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidine (16b)
Compound 9b (0.268 g, 1 mmol) was heated under reflux in 20 mL
glacial acetic acid/acetic anhydride (1:1) for 6 h; the reaction mixture
was cooled, filtered, dried, and the residue was recrystallized from
ethanol to give compound 16b. Yield 69%, M.p. 268–270◦C. 1H NMR: δ

1552
M. I. Hegab et al.
2.75 (s, 3H, C2-CH3), 3.00 (s, 3H, C5-CH3), 4.00 (s, 2H, CH2), 7.20–7.60
(m, 3H, Ar H), 8.70 (d, J = 11.00 Hz, 1H, Ar H). MS: m/z 292 (M+, 100),
277 (2.69), 250 (2.28), 210 (28.66), 195 (2.39), 183 (5.58). Anal. Calcd.
for C16H12N4S (292); C, 65.63; H, 4.32; N, 19.00%. Found: C, 65.75; H,
4.10; N, 19.17.
Isomerization of 15a and 15b to 16a and 16b
Compound 15a (0.278 g, 1 mmol) and 15b (0.292 g, 1 mmol) were heated
under reflux temperature in 20 mL absolute ethanol containing few
drops of base [piperidine or sodium ethoxide] for 6 h (compound 15a)
and 16 h (compound 15b); then, the reaction mixture was evaporated
under reduced pressure and the residue was recrystallized from ethanol
to give products 16a and 16b, respectively, which are identical in all
respects with compounds 16a and 16b obtained in the previous exper-
iments.
Synthesis of 5-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[4,3-c]pyrimidine (17b)
Compound 10b (0.268 g, 1 mmol) was heated under reflux temperature
in 20 mL triethyl orthoformate for 2 h; then, the reaction mixture was
cooled, filtered, dried, and the residue was recrystallized from DMF to
give compound 17b. Yield 90%, M.p. 270–272◦C. 1H NMR: δ 2.90 (s, 3H,
C5-CH3), 4.15 (s, 2H, CH2), 7.30–7.60 (m, 3H, Ar H), 8.75 (d, J = 11.20
Hz, 1H, Ar H), 9.60 (s, 1H, C3-H).
Synthesis of Indeno[1ꢀ,2ꢀ:4,5]thieno[3,2-e][1,2,4]triazolo-
[1,5-c]pyrimidine (18a)
Method A
Compound 9a (0.254 g, 1 mmol) was heated under reflux in 20 mL
formic acid for 5 h; then, the reaction mixture was cooled, poured into
water, filtered off, dried, and purified on silica gel using petroleum ether
(40–60◦C) : ethyl acetate (3:2) to give compound 18a (0.19 g, 72 %), m.p.
256–258◦C. 1H NMR: δ 4.20 (s, 2H, CH2), 7.20–7.60 (m, 3H, Ar H), 8.60
(d, J = 11.07 Hz, 1H, Ar H), 8.80 (s, 1H, C2-H), 9.70 (s, 1H, C5-H).
MS: m/z 264 (M+, 100), 236 (5), 210 (28.5), 195 (2.66). Anal. Calcd. for
C14H8N4S (264); C, 63.80; H, 3.30; N, 21.10; S, 12.00%. Found: C, 63.63;
H, 3.03; N, 21.21; S, 12.12.
Method B
The same as in method A but compound 10a (0.254 g, 1 mmol) was
refluxed for 8 h, to give compound 18a.

Fused Thieno[2,3-d]pyrimidine Derivatives
1553
Synthesis of 5-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidine (18b)
Method A
A mixture of Compound 9b (0.268 g, 1 mmol) and formic acid (20 mL)
was heated under reflux temperature for 5 h, cooled, filtered, dried, and
recrystallized from ethanol to give compound 18b. Yield 76%, M.p. 255–
257◦C. 1H NMR: δ 2.95 (s, 3H, C5-CH3), 4.25 (s, 2H, CH2), 7.30–7.60 (m,
3H, Ar H), 8.70 (d, J = 11.00 Hz, 1H, Ar H), 8.80 (s, 1H, C2-H). MS: m/z
278 (M+, 100), 264 (4.0), 210 (72.0), 195 (6.0), 183 (16.0). Anal. Calcd.
for C15H10N4S (278.00); C, 65.11; H, 3.35; N, 20.46%. Found: C, 64.74;
H, 3.59; N, 20.14.
Method B
Compound 10b (0.268 g, 1 mmol) was heated under reflux in 20 mL
formic acid for 10 h; then, the reaction mixture was cooled, filtered,
dried, and recrystallized from ethanol to give compound 18b. Both prod-
ucts obtained in experiments A and B are identical with each other in
all respects.
Isomerization of 17b to 18b
Compound 17b (0.278 g, 1 mmol) was heated under reflux temperature
in 20 mL formic acid for 4 h; then, the reaction mixture was cooled,
poured into water, filtered, dried, and recrystallized from ethanol to
give product 18b. The product of this experiment is identical in all
respects with compound 18b obtained in the previous experiment.
Synthesis of 2-Methylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidine-2(1H)-thione (20)
To a solution of compound 9b or 10b (0.268 g, 1 mmol) in 20 mL dry
pyridine, carbon disulfide (10 mL) was added then the reaction mixture
was refluxed for 5 h (compound 9b) or 3 h (compound 10b); the reaction
mixture was cooled, poured into cold water, then 1 mL of concentrated
hydrochloric acid was added, filtered, dried, and recrystallized from
DMF to give in both cases one and the same identical product 20 (0.21
g, 68% from compound 9b and 0.25 g, 81% from compound 10b). M.p.
1
290–292◦C. H NMR: δ 3.20 (s, 3H, C5-CH3) 4.10 (s, 2H, CH2), 7.30–
7.60 (m, 3H, Ar H), 8.30 (d, J = 11.34 Hz, 1H, Ar H), 14.60 (s, 1H, NH,
D2O exchangeable). MS: m/z 310 (M+, 100), 295 (46.17), 236(5.38), 2 10
(31.69).

1554
M. I. Hegab et al.
Synthesis of 5-methyl-2-phenylindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidine (21)
Method A
To a solution of compound 9b (0.268 g, 1 mmol) in 20 mL dry diox-
ane containing 5 drops of triethyl amine, benzoyl chloride (0.50 mL)
was added, then the reaction mixture was heated under reflux tem-
perature for 10 h, evaporated under reduced pressure, and the residue
was recrystallized from dioxane to give compound 21 (0.31 g, 89%). M.p.
1
250–252◦C. H NMR: δ 3.00 (s, 3H, C5-CH3), 4.00 (s, 2H, CH2), 7.20–
8.80 (m, 10H, Ar H). MS: m/z 354 (M+, 100), 277 (1.84), 210 (28.40), 195
(4.17), 183 (9.2). Anal. Calcd. for C21H14N4S (354); C, 71.13; H, 4.17; N,
15.69; S, 9.01%. Found: C, 71.18; H, 3.95; N, 15.81; S, 9.03.
Method B
To a solution of compound 9b or 10b (0.268 g, 1 mmol) in ethanol, ben-
zaldehyde (1 mmol) was added followed by addition of 2–3 drops glacial
acetic acid; then, the reaction mixture was heated under reflux tem-
perature for 10 h (compound 9b) and 4 h (compound 10b), evaporated
under reduced pressure to give in both cases one and the same product
21 (0.25 g, 71% from compound 9b and 0.30 g, 85% from compound 10b).
Products 21 obtained from both method A and B are identical with each
other in all respects.
Reaction of 9b with: ClCH2COCl, CH3CHBrCOBr, and
Cl2CHCOCl
To a solution of compound 9b (0.268 g, 1 mmol) in 20 mL dry dioxane
containing 5 drops of triethylamine, chloroacetyl chloride (3 mL), α-
bromopropionyl bromide (1 mL), or dichloroacetyl chloride (1 mL) were
added; then, the reaction mixture was heated under reflux temperature
for 5, 5, and 9 h, respectively. The reaction mixture was then evapo-
rated under reduced pressure, and the residue was recrystallized from
dioxane to give the 2-substituted-5-methyindeno[1ꢀ,2ꢀ:4,5]thieno[3,2-
e][1,2,4]triazolo[1,5-c]pyrimidine derivatives 22a, 22b, and 23, respec-
tively.
1
22a: Yield 80%, M.p. 234–236◦C. H NMR: δ 3.00 (s, 3H, C5-CH3),
4.00 (s, 2H, CH2), 4.90 (s, 2H, CH2Cl), 7.20–7.60 (m, 3H, Ar H), 8.70 (d,
J = 10.80 Hz, 1H, Ar H). MS: m/z 328 (M+, Cl37, 39.90), 326 (M+, Cl35,
100), 291.1 (14.1), 277 (7.5), 250 (8.2), 210 (32.6), 195 (13.5). Anal. Calcd.
for C16H11ClN4S (326); C, 58.63; H, 3.45; N, 17.04; S, 9.60%. found: C,
58.80; H, 3.37; N, 17.17; S, 9.82.

Fused Thieno[2,3-d]pyrimidine Derivatives
1555
22b: Yield 78%, M.p. 200–202◦C. 1H NMR: δ 2.30 (d, 3H, CH3), 3.00
(s, 3H, C5-CH3), 3.95 (s, 2H, CH2), 5.50 (q, J = 8.20 Hz, 1H, CH), 7.30–
7.55 (m, 3H, Ar H), 8.70 (d, J = 10.80 Hz, 1H, Ar H). MS: m/z 386
(M+, Br81, 39), 384 (M+, Br79, 36), 305 (67.15), 264 (2.85), 210 (7.76),
195 (5.00), 183 (4.15). Anal. Calcd. for C17H13BrN4S (384); C, 52.90; H,
3.59; N, 14.41; S, 8.05%. Found: C, 53.12; H, 3.38; N, 14.58; S, 8.33.
1
23: Yield 83%, M.p. 262–264◦C. H NMR: δ 3.00 (s, 3H, C5-CH3),
4.00 (s, 2H, CH2), 7.00–7.50 (m, 4H, Ar H), 11.60 (s, 1H, NH, D2O
exchangeable). MS: m/z 364 (M+, 2Cl37, 14), 362 (M+, Cl37, Cl35 69.29),
360 (M+, 2Cl35, 100), 324 (18), 277 (9.84), 236 (2.38), 210 (9.30), 195
(4.78), 183 (3.08).
Antimicrobial Activity
The in vitro antimicrobial activity of the synthesized compounds was
investigated against several pathogenic representative Gram-positive
bacteria (Staphylococcus aureus ATCC 29231, Bacillus subtilis, ATCC
10783 and Mycobacterium phlei, ATCC 1014); Gram-negative bacte-
ria (Escherichia coli ATCC 11105) and yeast (Candida albicans ATCC
10231). All microorganisms used were obtained from the culture collec-
tion of the Department of Natural and Microbial Products, National Re-
search Centre, Dokki, Cairo, Egypt. Compounds were tested against Es-
cherichia coli andStaphylococcus aureus in a nutrient broth, pH = 7.0,
against Bacillus subtilis, Mycobacterium phlei in the Bacto brain heart
infusion broth, pH = 7.0, and against Candida albicans in a broth con-
taining 1% peptone, 2% dextrose, pH = 5.7. Escherichia coli of known
antibiotic sensitivity served for control purposes. Media for disc sen-
sitivity tests were the nutrient agar and Muller-Hinton agar (MHA)
purchased from Difco (USA). Nonsterile powder of the tested compound
was dissolved in sterile DMSO to yield 2.0 µg mL−1, and passed through
0.2 µm membrane filters (Millipore Corp., USA). The filtrates were dis-
pensed as 2 mL samples into sterile, small screw-capped vials, frozen
and kept stored at −15◦C. The vials were refrozen after thawing. Disc
diffusion sensitivity test was done in the manner identical to that of
Bauer et al.5 DMSO showed no inhibition zones. Streptomycine, Amox-
icilin (Bioanalyse, Turkey) and Fusidic acid (Sigma, Aldrich, USA) were
used as reference substances.
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