Design, synthesis, antibacterial and antifungal activity of novel spiro-isoxazolyl bis-[5,5']thiazolidin-4-ones and spiro-isoxazolyl thiazolidin-4-one-[5,5']-1,2-4 oxdiazolines

Article abstract of DOI:10.1080/17415993.2010.492474

The synthesis of novel isoxazolyl 1,6-dithia-4,9-diazaspiro[4.4]nonane-3,8- diones (4a-h) and isoxazolyl 1-oxa-6-thia-2,4,9-triazaspiro[4.4]non-2-ene-8-ones (5a-h) analogs is described. Reaction of N-1-{3-methyl-5-[(E)-2-aryl-1- ethenyl]-4-isoxazolyl}-2-c

Full text of DOI:10.1080/17415993.2010.492474

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Design, synthesis, antibacterial and
antifungal activity of novel spiro-
isoxazolyl bis-[5,5′]thiazolidin-4-ones
and spiro-isoxazolyl thiazolidin-4-one-
[5,5′]-1,2-4 oxdiazolines
Eligeti Rajanarendar a , Enugala Kalyan Rao a , Firoz Pasha Shaik a
, Modugu Nagi Reddy a & Manda Srinivas a
a Department of Chemistry , Kakatiya University , Warangal, AP,
506 009, India
Published online: 12 Aug 2010.
To cite this article: Eligeti Rajanarendar , Enugala Kalyan Rao , Firoz Pasha Shaik , Modugu Nagi
Reddy & Manda Srinivas (2010) Design, synthesis, antibacterial and antifungal activity of novel
spiro-isoxazolyl bis-[5,5′]thiazolidin-4-ones and spiro-isoxazolyl thiazolidin-4-one-[5,5′]-1,2-4
oxdiazolines, Journal of Sulfur Chemistry, 31:4, 263-274, DOI: 10.1080/17415993.2010.492474
To link to this article: http://dx.doi.org/10.1080/17415993.2010.492474
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Journal of Sulfur Chemistry
Vol. 31, No. 4, August 2010, 263–274
Design, synthesis, antibacterial and antifungal activity of novel
spiro-isoxazolyl bis-[5,5ꢀ]thiazolidin-4-ones and spiro-isoxazolyl
thiazolidin-4-one-[5,5ꢀ]-1,2-4 oxdiazolines
Eligeti Rajanarendar*, Enugala Kalyan Rao, Firoz Pasha Shaik, Modugu Nagi Reddy
and Manda Srinivas
Department of Chemistry, Kakatiya University, Warangal, AP 506 009, India
(Received 3 February 2010; final version received 8 May 2010)
The synthesis of novel isoxazolyl 1,6-dithia-4,9-diazaspiro[4.4]nonane-3,8-diones (4a–h) and
isoxazolyl 1-oxa-6-thia-2,4,9-triazaspiro[4.4]non-2-ene-8-ones (5a–h) analogs is described. Reaction
of N-1-{3-methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl}-2-chloroacetamide (2) with aryl isothio-
cyanates yielded 3,3-methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl-2-(arylimino)-1,3-thiazolan-4-ones
(3). Cyclocondensation of 3 with mercaptoacetic acid furnished novel isoxazolyl-1,6-dithia-4,9-
diazaspiro[4.4]nonane-3,8-diones (4a–h). Cycloaddition of 3 with benzonitrile oxides afforded novel isox-
azolyl 1-oxa-6-thia-2,4,9-triazaspiro[4.4]non-2-ene-8-ones (5a–h). Compounds 4a–h and 5a–h showed
significant biological activity against all the standard strains.
Keywords: cyclocondensation reaction; spirothiazolidinones; cycloaddition reaction; antibacterial activ-
ity; antifungal activity
1. Introduction
The importance of heterocycles in biological systems has stimulated interest in designing and
constructing new heterocyclic systems using a molecular modification approach. Among various
heterocyclic systems, isoxazole holds a prominent role, because it is present as a core unit in a
number of compounds possessing a broad spectrum of biological activities (1). The chemistry
of isoxazole unit continues to draw the attention of synthetic organic chemists, as it consti-
tutes an easily accessible nucleus that is present in a number of natural and pharmacological
compounds (2).
Isoxazole derivatives are used as potent antitumor (3), as CNS-active (4), as analgesic (5), as
antimicrobial (6), as muscle relaxant (7), for the treatment of hypercholesterolemia and hyperlipi-
demia (8), as organic electrolytes for non-aqueous batteries (9), in photographic emulsions (10),
as synthetic intermediates (11) and as chemotherapeutic agents (12). Spirothiazolidinones have
attracted our attention because of their enhanced activities in medicinal and biological chemistry
*Corresponding author. Email: eligeti_rajan@yahoo.co.in
ISSN 1741-5993 print/ISSN 1741-6000 online
© 2010 Taylor & Francis
DOI: 10.1080/17415993.2010.492474
http://www.informaworld.com

264 E. Rajanarendar et al.
(13, 14). Several of these derivatives are known to possess antitubercular (15), anticonvulsant
(16), anti-inflammatory (17), fungistatic (18), bacteriostatic (19) and antimicrobial activity (20).
A literature survey revealed that when two heterocyclic moieties are joined together in a sin-
gle molecular framework, the resulting compound is expected to possess enhanced bioactivity
(21). Realizing the importance of the above biodynamic heteryl nuclei, and in continuation of
our interest in designing the synthesis of biologically active nitrogen and sulfur heterocycles
linked to isoxazole ring (22), it was thought worthwhile to undertake the synthesis of novel
spirothiazolidinones containing isoxazole moiety and to evaluate their antimicrobial activity.
2. Chemistry
The synthesis of compounds 2, 3, 4 and 5 was accomplished by the synthetic sequence shown in
Scheme 1. The reaction of 4-amino-3-methyl-5-styrylisoxazole (1) (23) with chloroacetyl chloride
in the presence of triethyl amine in benzene furnished N-1-{3-methyl-5-[(E)-2-aryl-1-ethenyl]-
4-isoxazolyl}-2-chloroacetamide (2) (24). Compound 2 on treatment with aryl isothiocyanates
in the presence of K2CO3 in CH3CN afforded the key intermediates, viz., 3-3-methyl-5-[(E)-
2-aryl-1-ethenyl]-4-isoxazolyl-2-(arylimino)-1,3-thiazolan-4-ones (3). The mechanism involves
the addition of amide derivative to isothiocyanate in the presence of a base and subsequent
cyclization takes place by nucleophilic substitution of chlorine by the sulfur atom of isothio-
cyanate. This mechanism was in agreement with an earlier observation (25). Cyclocondensation
of 3 with mercaptoacetic acid in dry benzene led to the formation of novel 4-3-{methyl-5[(E)-
2-aryl-1-ethenyl]-4-isoxazolyl}-9-aryl-1,6-dithia-4,9-diazaspiro[4.4]nonane-3,8-diones (4). The
other possible regioisomer was not formed in the reaction. This selectivity in the cycloconden-
sation giving only one of the two possible regioisomers may be ascribed to the polarization
of the imine double bond in 3 toward the nitrogen rendering the imine carbon positively
charged. So, the mercapto functional group of mercaptoacetic acid makes a nucleophilic
attack on the imine carbon giving only 4. The styryl double bond, which is not polarized, is
inert toward the nucleophilic attack of mercaptoacetic acid, hence the chemoselectivity was
observed only in the imine group. Cycloaddition of 3 with benzonitrile oxides generated in
situ from benzhydroxamoyl chloride in the presence of triethyl amine at ice-cold temperature
furnished the novel 9-{3-methyl-5[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-3,4-diphenyl-1-oxa-
6-thia-2,4,9-triazaspiro[4.4]non-2-ene-8-ones (5) by the 1,3-dipolar cycloaddition (Scheme 1).
The other regioisomer could not be obtained in the reaction. This selectivity in cycloaddition
leading to the formation of only one of the possible region isomers may be due to the polarization
of the imine double bond of 3 toward nitrogen, making imine carbon positively charged. So, the
oxygen of benzonitrile oxide attacks on this carbon giving only 5. When the reaction was per-
formed using excess of benzonitrile oxide, only one regioisomer 5 was obtained, indicating that
the olefinic double bond of the styryl moiety is inert, hence the chemoselectivity was achieved
with the participation of imine double bond. The structures of products 3, 4 and 5 have been
elucidated on the basis of spectral (IR, 1H NMR, and MS) and microanalytical data.
Compound 3 displayed characteristic absorption bands in the IR spectra around 1630 and
1685 cm−1 due to C N and C O functional groups, respectively, confirming cyclocondensation.
1H NMR spectra of 3 exhibited a sharp singlet around δ 4.2 due to newly formed thiazolan-
4-one ring methylene protons. Mass spectrum of the product 3 agrees well with the cyclized
structure, which showed a molecular ion peak at [M + H]+ at m/z 376. The 1H NMR spectrum of
4 displayed two distinct singlets around δ 4.2 and 4.4 due to methylene protons of thiazolidinone
rings, confirmingtheformationofspirothiazolidinonerings.Themassspectrumof 4 confirmedthe
structure by exhibiting the molecular ion peak [M + H]+ at m/z 450. The 1H NMR spectrum of 5
showed a prominent singlet around 4.0 due to thiazolidinone ring CH2 protons. The mass spectrum
=
=

Journal of Sulfur Chemistry 265
Scheme 1. Synthesis of spiro-isoxazolyl bis-[5,5ꢀ]thiazolidin-4-ones and spiro-isoxazolyl
thiazolidin-4-one-[5,5ꢀ]-1,2,4-oxazdiazolines.
of 5 also agrees well with the structure, which showed the molecular ion peak [M + H]+ at m/z
495 confirming the cycloaddition reaction. Elemental analyses are satisfactory and confirmed
elemental composition and purity of the newly synthesized compounds 3-5.
3. Antibacterial activity
Antibacterial activity of 4a–h and 5a–h in acetone was performed by the broth dilution
method using nutrient agar against Gram-negative bacteria Pseudomonas aeruginosa, Klebsiella

266 E. Rajanarendar et al.
Table 1. Antibacterial activity of 4a–h.
Minimum inhibitory concentration
Gram-negative
Gram-positive
Compound
B. subtilis
B. sphaericus
S. aureus
P. aeruginosa
K. aerogenes
C. violaceum
4a
4b
4c
4d
4e
4f
4g
4h
16
13
21
20
19
18
14
15
23
21
12
18
23
19
13
17
22
25
25
14
16
28
20
17
11
24
30
20
12
10
21
18
20
15
11
22
18
9
17
15
19
17
16
12
11
20
22
16
18
15
12
14
11
20
Ciprofloxacin
Notes: Negative control (acetone) – no activity. Values are indicated in μg/ml.
aerogenes, Chormobacterium violaceum and Gram-positive bacteria Bacillus subtilis, Bacillus
sphaericus and Staphylococcus aureus at 100 μg/ml concentration. The minimum inhibitory con-
centration was done by the broth dilution method (26). Ciprofloxacin was used as standard for
comparison. The ready-made nutrient broth medium (Himedia, 24 g) was suspended in distilled
water (100 ml) and heated until it dissolved completely. The medium and test tubes were auto-
claved at a pressure of 15 lb/inc2 for 20 min. A set of sterilized test tubes with nutrient broth
medium was capped with cotton plugs. The test compound was dissolved in acetone and a con-
centration of 100 μg/ml of the test compound was added in the first test tube, which was serially
diluted. A fixed volume of 0.5 ml of overnight culture was added in all the test tubes which were
incubated at 37◦C for 24 h. After 24 h, these tubes were measured for turbidity. Results are given
in Tables 1 and 2.
The results of antibacterial screening (Tables 1 and 2) reveal that compounds 4a–h and 5a–h
displayed a better activity and are more active than the standard drug ciprofloxacin. In series
4, compounds 4c, 4f and 4g possessing chloro, bromo and methyl groups as substituents on the
benzene ring showed a better activity, whereas in series 5, compounds 5b and 5f carrying methoxy
and chloro groups as substituents on the benzene ring imparted a remarkable activity. However,
the degree of inhibition varied both with the test compound and with the bacteria used in the
present investigation. In conclusion, almost all the compounds, 4a–h and 5a–h, exhibited the
maximum activity by inhibiting the growth of all the six bacteria to a greater extent in comparison
Table 2. Antibacterial activity of 5a–h.
Minimum inhibitory concentration
Gram-positive
Gram-negative
Compound
B. subtilis
B. sphaericus
S. aureus
P. aeruginosa
K. aerogenes
C. violaceum
5a
5b
5c
5d
5e
5f
5g
5h
16
8
15
11
15
17
16
8
15
17
20
16
12
18
15
20
12
20
16
25
20
8
18
20
17
8
15
20
30
16
13
15
18
19
13
15
17
25
13
15
15
15
16
10
17
16
25
16
18
20
10
15
19
20
Ciprofloxacin
Notes: Negative control (acetone) – no activity. Values are indicated in μg/ml.

Journal of Sulfur Chemistry 267
with the standard drug streptomycin. The antibacterial activity of some of these compounds is
promising compared with standard ciprofloxacin, and they can be exploited for the formation of
bactericide after further study.
4. Antifungal activity
The antifungal activity of 4a–h and 5a–h was performed by the agar cup bioassay method
(27) using clotrimazole as the standard. The compounds were tested for their antifungal activ-
ity against five test organisms, Aspergillus niger, Chrysosporium tropicum, Rhizopus oryzae,
Fusarium moniliforme and Curvularia lunata using the agar cup bioassay method at 100 μg/ml
concentration. The ready-made potato dextrose agar medium (Himedia, 39 g) was suspended in
distilled water (1000 ml) and heated until it dissolved completely. The medium and Petri dishes
were autoclaved at a pressure of 15 lb/inc2 for 20 min. The medium was poured into sterile Petri
dishes under aseptic conditions in a laminar flow chamber. When the medium in the plates solid-
ified, 0.5 ml of the (week-old) culture of test organism was inoculated and uniformly spread over
the agar surface with a sterile L-shaped rod. Solutions were prepared by dissolving plant extract
in acetone (100 μg/ml). Agar inoculation cups were scooped out with 6 mm sterile cork borer
and the lids of the dishes were replaced. To each cup, 100 μg/ml of test solution was added. Con-
trols were maintained with acetone and clotrimazole (100 μg/ml). The treated and the controls
were kept at room temperature for 72–96 h. Inhibition zones were measured and the diameter was
calculated in millimeters. Three to four replicates were maintained for each treatment. The results
are given in Tables 3 and 4.
Table 3. Antifungal activity of 4a–h.
Zone of inhibition in mm
Compound
A. niger
C. tropicum
R. oryzae
F. moniliforme
C. lunata
4a
4b
4c
4d
4e
4f
4g
4h
26
50
50
25
26
25
20
30
29
20
45
55
20
20
24
23
25
30
15
55
50
25
19
26
23
30
28
19
50
45
20
18
30
15
20
23
17
45
40
20
18
35
15
20
20
Clotrimazole
Note: Negative control (acetone) – no activity.
Table 4. Antifungal activity of 5a–h.
Zone of inhibition in mm
Compound
A. niger
C. tropicum
R. oryzae
F. moniliforme
C. lunata
5a
5b
5c
5d
5e
5f
5g
5h
45
50
30
50
40
45
35
55
26
45
55
40
55
50
40
35
55
29
35
55
30
40
55
35
40
55
23
35
55
35
50
50
40
45
55
27
40
55
40
45
50
45
45
50
28
Clotrimazole
Note: Negative control (acetone) – no activity.

268 E. Rajanarendar et al.
The antifungal activity results (Tables 3 and 4) indicated that compounds 4a–h and 5a–h are
significantly toxic toward all the five fungi and they are lethal at 100 μg/ml concentration. In
series 4, compounds 4b and 4c exhibited a high antifungal activity which may be due to the
presence of methoxy and chloro substituents on the benzene ring. In series 5, compounds 5b
and 5h possessing methoxy and nitro groups are highly toxic toward some fungi. However, the
degree of spore germination inhibited varied with the test compound as well as with the fungi.
The antifungal activity of these compounds was compared with the standard drug clotrimazole,
and they have a promising activity, when compared with the standard drug. In conclusion, almost
all the series of compounds, 4a–h and 5a–h, are highly toxic toward the fungi under investigation
and they are lethal even at 100 μg/ml concentration in comparison with standard clotrimazole at
the same concentration. It is noteworthy that some of the compounds may be exploited for control
of wilt diseases of different crops as fungicides after detailed study.
5. Experimental
Melting points were recorded on an Electrothermal type 9100 melting point apparatus and are
uncorrected. The IR spectra were recorded on Nicolet Impact 410 FTIR spectrophotometer using
KBr pellets. 1H NMR spectra were recorded on Bruker Ac 300 spectrometer in CDCl3 and with
TMS as the internal standard. The mass spectra were obtained on a Varian MATCH-7 instrument
at 70 eV. Elemental analyses were carried out using Perkin-Elmer 240C CHN-analyzer.
5.1. Synthesis of 3-3-methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl-2-(arylimino)-1,3-
thiazolan-4-one derivatives (3a–h) – general procedure
A mixture of N-1-{3-methyl-5[(E)-2-aryl-1-ethenyl]-4-isoxazolyl}-2-chloroacetamide (2)
(0.01 mol) and aryl isothiocyanate (0.01 mol) was taken in acetonitrile (15 ml) in the presence
of K2CO3 (0.5 g). The reaction mixture was refluxed while stirring for about 6 h. The solvent was
removed under reduced pressure and the residue was purified by recrystallization from methanol
to produce 3a–h in high yields.
5.2. 3-{3-Methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl}-2-(arylimino)-1,3-thiazolan-4-
one (3a)
Pale yellow powder; yield 85%, m.p. 180–181◦C; IR (KBr) cm−1: 1630 (C N), 1685 (C O);
=
=
1H NMR (300 MHz, CDCl3) δ (ppm): 2.32 (s, 3H, CH3), 4.30 (s, 2H, CH2), 6.82, 7.20 (2d,
+
J = 15.2 Hz, 2H, CH CH), and 7.35–8.03 (m, 10H, ArH). EI-MS[M + H] m/z 376. 13C NMR
=
(75 MHz, CDCl3) δ (ppm): 11.41 (C-6ꢀꢀ), 28.35 (C-5), 109.87 (C-7ꢀꢀ), 115.20 (C-8ꢀꢀ), 115.27
(C-4ꢀꢀ), 120.10 (Ar–C), 126.87 (Ar–C), 127.35 (Ar–C), 127.81 (Ar–C), 129.01 (Ar–C), 130.25
(Ar–C), 130.88 (Ar–C), 132.29 (Ar–C), 134.65 (Ar–C), 137.69 (Ar–C), 138.39 (Ar–C), 141.09
(Ar–C), 156.28 (C-3ꢀꢀ), 158.85 (C-5ꢀꢀ), 160.70 (C-2), 174.60 (C-4).Anal. Calcd. for C21H17N3O2S
(375.45): C, 67.20; H, 4.53; N, 11.20. Found: C, 67.17; H, 4.49; N, 11.16%.
5.3. 3-{5-[(E)-2-(4-Methoxyphenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-2-(phenylimino)-1,3-
thiazolan-4-one (3b)
1
Pale yellow powder; yield 85%, m.p. 130–131◦C; IR (KBr) cm−1: 1620 (C N), 1685 (C O); H
=
=
NMR (300 MHz, CDCl3) δ: 2.40 (s, 3H, CH3), 3.82 (s, 3H, OCH3), 4.10 (s, 2H, CH2), 6.76, 7.35
(2d, J = 15.2 Hz, 2H, CH CH), and 7.30–8.22 (m, 9H, ArH). EI-MS [M + H] m/z 406. 13C
+
=

Journal of Sulfur Chemistry 269
NMR (75 MHz, CDCl3) δ (ppm): 11.39 (C-6ꢀꢀ), 31.50 (C-5), 53.05 (Ar–OCH3), 109.87 (C-7ꢀꢀ),
114.85 (C-8ꢀꢀ), 115.05 (C-4ꢀꢀ), 119.25 (Ar–C), 125.56 (Ar–C), 126.65 (Ar–C), 127.85 (Ar–C),
129.50 (Ar–C), 130.55 (Ar–C), 131.00 (Ar–C), 132.85 (Ar–C), 133.95 (Ar–C), 137.10 (Ar–C),
138.05 (Ar–C), 142.05 (Ar–C), 155.25 (C-5ꢀꢀ), 159.56 (C-3ꢀꢀ), 165.33 (C-2), 185.50 (C-4). Anal.
Calcd. for C22H19N3O3S (405.47): C, 65.18; H, 4.69; N, 10.37. Found: C, 65.17; H, 4.67; N,
10.32%.
5.4. 3-{5-[(E)-2-(4-Chlorophenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-2-(phenylimino)-1,3-
thiazolan-4-one (3c)
Pale yellow powder; yield 85%, m.p. 145–146◦C; IR (KBr) cm−1: 1615 (C N), 1690 (C O);
=
=
1H NMR (300 MHz, CDCl3) δ: 2.33 (s, 3H, CH3), 4.31 (s, 2H, CH2), 6.90, 7.25 (2d, J =
+
=
15.2 Hz, 2H, CH CH), and 7.40–8.12 (m, 9H, ArH); EI-MS [M + H] m/z 410. Anal. Calcd.
for C21H16N3O2SCl (409.89): C, 61.61; H, 3.91; N, 10.26. Found: C, 61.57; H, 3.89; N, 10.21%.
5.5. 3-{5-[(E)-2-(2-Chlorophenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-2-(phenylimino)-1,3-
thiazolan-4-one (3d)
Pale yellow powder; yield 90%, m.p. 185–186◦C; IR (KBr) cm−1: 1625 (C N), 1695 (C O); H
1
=
=
NMR (300 MHz, CDCl3) δ: 2.22 (s, 3H, CH3), 4.23 (s, 2H, CH2), 7.05, 7.36 (2d, J = 15.2 Hz,
+
=
2H, CH CH), and 7.48–8.26 (m, 9H, ArH); EI-MS [M + H] m/z 410. Anal. Calcd. for
C21H16N3O2SCl (409.89): C, 61.61; H, 3.91; N, 10.26. Found: C, 61.59; H, 3.87; N, 10.22%.
5.6. 2-[(4-Methoxyphenyl)imino]-3-{3-methyl-5-[(E)-2-(4-methylphenyl)-1-ethenyl]-4-
isoxazolyl}-1,3-thiazolan-4-one (3e)
Pale yellow powder; yield 80%, m.p. 175–176◦C; IR (KBr) cm−1: 1625 (C N), 1680 (C O);
=
=
1H NMR (300 MHz, CDCl3) δ: 2.30 (s, 3H, CH3), 2.52 (s, 3H, CH3), 3.71 (s, 3H, OCH3), 4.33
=
(s, 2H, CH2), 6.62, 7.25 (2d, J = 15.2 Hz, 2H, CH CH), and 7.33–8.05 (m, 8H, ArH). EI-MS
[M + H]+ m/z 420. Anal. Calcd. for C23H21N3O3S (419.50): C, 65.87; H, 5.01; N, 10.02. Found:
C, 65.82; H, 5.00; N, 10.01%.
5.7. 2-[(4-Bromophenyl)imino]-3-{3-methyl-5-[(E)-2-phenyl)-1-ethenyl]-4-isoxazolyl}-1,3-
thiazolan-4-one (3f)
1
Pale yellow powder; yield 85%, m.p. 170–171◦C; IR (KBr) cm−1: 1615 (C N), 1680 (C O); H
=
=
NMR (300 MHz, CDCl3) δ: 2.35 (s, 3H, CH3), 4.35 (s, 2H, CH2), 6.82, 7.25 (2d, J = 15.2 Hz,
+
=
2H, CH CH), and 7.22–8.14 (m, 9H, ArH). EI-MS [M + H] m/z 454. Anal. Calcd. for
C21H16N3O2SBr (454.34): C, 55.62; H, 3.53; N, 9.27. Found: C, 55.60; H, 3.50; N, 9.23%.
5.8. 3-{3-Methyl-5-[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-2-[(4-methylphenyl)imino]-1,3-
thiazolan-4-one (3g)
1
Pale yellow powder; yield 80%, m.p. 165–166◦C; IR (KBr) cm−1: 1640 (C N), 1700 (C O); H
=
=
NMR (300 MHz, CDCl3) δ: 2.35 (s, 3H, CH3), 2.52 (s, 3H, CH3), 4.25 (s, 2H+, CH2), 6.77, 7.32
=
(2d, J = 15.2 Hz, 2H, CH CH), 7.48–8.18 (m, 9H, ArH); EI-MS [M + H] m/z 390. Anal.
Calcd. for C22H19N3O2S (389.47): C, 67.86; H, 4.88; N, 10.79. Found: C, 67.84; H, 4.84; N,
10.75%.

270 E. Rajanarendar et al.
5.9. 2-[(4-Chlorophenyl)imino]-3-{-5-[(E)-2-(4-methoxyphenyl)-1-ethenyl]-3-methyl-4-
isoxazolyl}-1,3-thiazolan-4-one (3h)
Pale yellow powder; yield 85%, m.p. 195–196◦C; IR (KBr) cm−1: 1638 (C N), 1680 (C O);
=
=
1H NMR (300 MHz, CDCl3) δ: 2.40 (s, 3H, CH3), 3.82 (s, 3H, OCH3), 4.12 (s, 2H, CH2), 6.75,
+
=
7.12 (2d, J = 15.2 Hz, 2H, CH CH), and 7.13–8.26 (m, 8H, ArH). EI-MS [M + H] m/z 440.
Anal. Calcd. for C22H18N3O3SCl (439.92): C, 60.13; H, 4.10; N, 9.56. Found: C, 60.10; H, 4.08;
N, 9.52%.
5.10. 4-{3-Methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl}-9-aryl-1,6-dithia-4,9-diazaspiro
[4.4]nonane-3,8-dione derivatives (4a–h) – general procedure
To a solution of 3-3-methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl-2-(arylimino)-1,3-thiazolan-
4-ones (3) (0.01 mol) in dry benzene (25 ml) mercaptoacetic acid (0.01 mol) was added and the
contents were refluxed in a Dean-stark apparatus for 6 h. The solvent was distilled off and the
gummy material obtained was triturated with light petrol repeatedly. The product was purified by
passing through column. Elution with petroleum ether–benzene (1:1) yielded compounds 4a–h.
5.11. 4-{3-Methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl}-9-aryl-1,6-dithia-4,9-
diazaspiro[4.4]nonane-3,8-dione (4a)
White powder; yield 70%, m.p. 210–211◦C; IR (KBr) cm−1: 1710 (C O), 1275 (C–S); H NMR
1
=
(300 MHz, CDCl3) δ: 2.25 (s, 3H, CH3), 4.20 (s, 2H, CH2), 4.42 (s, 2H, CH2), 6.85 (d, J= 15.2 Hz,
+
1H, CH CH), 7.22–7.86 (m, 10H,ArH and 1H, CH CH). EI-MS [M + H] m/z 450. 13C NMR
(75 MHz, CDCl3) δ (ppm): 11.05 (C-6ꢀꢀ), 27.80 (C-2), 28.33 (C-7), 80.05 (C-5), 109.20 (C-4ꢀꢀ),
109.90 (C-7ꢀꢀ), 115.55 (C-8ꢀꢀ), 120.08 (Ar–C), 125.11 (Ar–C), 125.85 (Ar–C), 127.30 (Ar–C),
127.99 (Ar–C), 128.05 (Ar–C), 128.52 (Ar–C), 129.15 (Ar–C), 130.00 (Ar–C), 130.10 (Ar–C),
134.60 (Ar–C), 139.02 (Ar–C), 156.65 (C-5?), 158.90 (C-3?), 176.35 (C-3), 177.05 (C-8). Anal.
Calcd. for C23H19N3O3S2 (449.55): C, 61.46; H, 4.23; N, 9.35. Found: C, 61.44; H, 4.20; N,
9.32%.
=
=
5.12. 4-{5-[(E)-2-(4-Methoxyphenyl)-1-ethenyl]-3-methyl4-isoxazolyl}-9-phenyl-1,6-dithia-
4,9-diazaspiro[4.4]nonane-3,8-dione (4b)
White powder; yield 65%, m.p. 225–226◦C; IR (KBr) cm−1: 1720 (C O), 1280 (C–S); H NMR
1
=
(300 MHz, CDCl3) δ: 2.22 (s, 3H, CH3), 3.78 (s, 3H, OCH3), 4.20 (s, 2H, CH2), 4.41 (s, 2H,
=
=
CH2), 6.73 (d, J = 15.2 Hz, 1H, CH CH), 7.35-7.89 (m, 9H, ArH and 1H, CH CH). EI-MS
[M + H]+ m/z 480. 13C NMR (75 MHz, CDCl3) δ (ppm): 11.45 (C-6ꢀꢀ), 27.88 (C-2), 28.53 (C-
7), 52.30 (Ar–OCH3), 81.33 (C-5), 109.55 (C-4ꢀꢀ), 109.85 (C-7ꢀꢀ), 116.03 (C-8ꢀꢀ), 120.09 (Ar–C),
125.05 (Ar–C), 126.00 (Ar–C), 127.15 (Ar–C), 127.85 (Ar–C), 128.02 (Ar–C), 128.63 (Ar–C),
129.15 (Ar–C), 130.08 (Ar–C), 133.15 (Ar–C), 135.00 (Ar–C), 139.07 (Ar–C), 157.05 (C-5ꢀꢀ),
158.80 (C-3ꢀꢀ), 176.55 (C-3), 178.00 (C-8). Anal. Calcd. for C24H21N3O4S2 (479.58): C, 60.12;
H, 4.38; N, 8.76. Found: C, 60.09; H, 4.35; N, 8.74%.
5.13. 4-{5-[(E)-2-(4-Chlorophenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-9-phenyl-1,6-dithia-
4,9-diazaspiro[4.4]nonane-3,8-dione (4c)
White powder; Yield 70%, m.p. 215–216◦C; IR (KBr) cm−1: 1730 (C O), 1270 (C–S); H
1
=
NMR (300 MHz, CDCl3) δ: 2.30 (s, 3H, CH3), 4.03 (s, 2H, CH2), 4.25 (s, 2H, CH2), 6.9 (d,

Journal of Sulfur Chemistry 271
+
=
=
J = 15.2 Hz, 1H, CH CH), 7.25–7.96 (m, 9H, ArH and 1H, CH CH); EI-MS [M + H] m/z
484. Anal. Calcd. for C23H18N3O3S2Cl (484.00): C, 57.14; H, 3.72; N, 8.69. Found: C, 57.12; H,
3.70; N, 8.65%.
5.14. 4-{5-[(E)-2-(2-Chlorophenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-9-phenyl-1,6-dithia-
4,9-diazaspiro[4.4]nonane-3,8-dione (4d)
White powder; yield 70%, m.p. 212–213◦C; IR (KBr) cm−1: 1715 (C O), 1275 (C–S); H
1
=
NMR (300 MHz, CDCl3) δ: 2.23 (s, 3H, CH3), 4.20 (s, 2H, CH2), 4.43 (s, 2H, CH2), 7.06 (d,
+
=
=
J = 15.2 Hz, 1H, CH CH), 7.18–8.18 (m, 9H, ArH and 1H, CH CH); EI-MS [M + H] m/z
484. Anal. Calcd. for C23H18N3O3S2Cl (484.00): C, 57.14; H, 3.72; N, 8.69. Found: C, 57.09; H,
3.69; N, 8.63%.
5.15. 4-(4-Methoxyphenyl)-9-{3-methyl-5-[(E)-2-(4-methylphenyl)-1-ethenyl]-4-isoxazolyl}-
1,6-dithia-4,9-diazaspiro[4.4]nonane-3,8-dione (4e)
1
White powder; yield 68%, m.p. 235–236◦C; IR (KBr) cm−1: 1710 (C O), 1280 (C–S); H NMR
=
(300 MHz, CDCl3) δ: 2.25 (s, 3H, CH3), 2.51 (s, 3H, CH3), 3.80 (s, 3H, OCH3), 4.12 (s, 2H,
=
CH2), 4.32 (s, 2H, CH2), 7.05 (d, J = 15.2 Hz, 1H, CH CH), 7.26–8.17 (m, 8H, ArH and 1H,
+
=
CH CH). EI-MS [M + H] m/z 494. Anal. Calcd. for C25H23N3O4S2 (493.60): C, 60.85; H,
4.66; N, 8.51. Found: C, 60.82; H, 4.63; N, 8.48%.
5.16. 4-(4-Bromophenyl)-9-{3-methyl-5-[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-1,6-dithia-
4,9-diazaspiro[4.4]nonane-3,8-dione (4f)
1
White powder; yield 65%, m.p. 240–241◦C; IR (KBr) cm−1: 1715 (C O), 1270 (C–S); H NMR
=
(300 MHz, CDCl3) δ: 2.30 (s, 3H, CH3), 4.32 (s, 2H, CH2), 4.50 (s, 2H, CH2), 6.8 (d, J = 15.2 Hz,
+
=
=
1H, CH CH) and 7.2–8.0 (m, 9H, ArH and 1H, CH CH). EI-MS [M + H] m/z 528. Anal.
Calcd. for C23H18N3O3S2Br (528.45): C, 52.37; H, 3.41; N, 7.96. Found: C, 52.33; H, 3.38; N,
7.94%.
5.17. 4-(4-Methylphenyl)-9-{3-methyl-5-[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-1,6-dithia-
4,9-diazaspiro[4.4]nonane-3,8-dione (4g)
White powder; yield 70%, m.p. 232–233◦C; IR (KBr) cm−1: 1715 (C O), 1275 (C–S); H NMR
1
=
(300 MHz, CDCl3) δ: 2.32 (s, 3H, CH3), 2.45 (s, 3H, CH3), 4.15 (s, 2H, CH2), 4.30 (s, 2H,
=
CH2), 6.76 (d, J= 15.2 Hz, 1H, CH=CH) and 7.35–7.95 (m, 9H, ArH and 1H, CH CH). EI-MS
[M + H]+ m/z 464. Anal. Calcd. for C24H21N3O3S2 (463.58): C, 62.20; H, 4.53; N, 9.07. Found:
C, 62.18; H, 4.51; N, 9.05%.
5.18. 4-(4-Chlorophenyl)-9-{5-[(E)-2-(4-methoxyphenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-
1,6-dithia-4,9-diazaspiro[4.4]nonane-3,8-dione (4h)
1
White powder; yield 65%, m.p. 239–240◦C; IR (KBr) cm−1: 1720 (C O), 1275 (C–S); H NMR
=
(300 MHz, CDCl3) δ: 2.25 (s, 3H, CH3), 3.82 (s, 3H, OCH3), 4.12 (s, 2H, CH2), 4.40 (s, 2H,
=
=
CH2), 6.65 (d, J = 15.2 Hz, 1H, CH CH) and 7.32–7.85 (m, 8H, ArH and 1H, CH CH). EI-
MS [M + H]+ m/z 514. Anal. Calcd. for C24H20N3O4S2Cl (514.02): C, 56.14; H, 3.89; N, 8.18.
Found: C, 56.10; H, 3.86; N, 8.15%.

272 E. Rajanarendar et al.
5.19. Synthesis of 9-{3-methyl-5-[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-3,4-diphenyl-1-oxa-
6-thia-2,4,9-triazaspiro[4.4]non-2-en-8-one derivatives (5a-h) – general procedure
To a solution of 3-3-methyl-5-[(E)-2-aryl-1-ethenyl]-4-isoxazolyl-2-(arylimino)-1,3-thiazolan-
4-one (3) (0.01 mol) in dry chloroform (50 ml), cooled in ice-salt bath, a solution of benzhy-
droxamoyl chloride (0.01 mol) in chloroform (10 ml) was added. Triethyl amine (0.01 mol) in
chloroform (10 ml) was added to the reaction mixture at 0◦C for 15 min with constant stirring.
After the addition was completed, the stirring was continued for another 4 h at 0◦C. The chloro-
form layer was washed with water (2 × 25 ml) to make it free from triethyl amine hydrochloride
and the organic layer was dried (Na2SO4). The solvent was removed at ambient temperature and
the crude product triturated with light petrol repeatedly to obtain a residue. The residue on shaking
with methanol gave the solid, which was recrystallized from benzene.
5.20. 9-{3-Methyl-5-[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-3,4-diphenyl-1-oxa-6-thia-2,4,9-
triazaspiro[4.4]non-2-en-8-one (5a)
White powder; yield 78%, m.p. 202–204◦C; IR (KBr) cm−1: 1630 (C N), 1680 (C O); H NMR
1
=
=
=
(300 MHz, CDCl3) δ: 2.25 (s, 3H, CH3), 4.05 (s, 2H, CH2), 6.82 (d, J = 15.2 Hz, 1H, CH CH),
+
7.25–7.59 (m, 15H, ArH and 1H, CH CH). EI-MS [M + H] m/z 495. 13C NMR (75 MHz,
=
CDCl3) δ (ppm): 11.50 (C-6ꢀꢀ), 26.50 (C-7), 100.05 (C-5), 105.68 (C-4ꢀꢀ), 110.15 (C-7ꢀꢀ), 118.40
(C-8ꢀꢀ), 120.10 (Ar–C), 121.85 (Ar–C), 125.10 (Ar–C), 126.88 (Ar–C), 127.35 (Ar–C), 127.81
(Ar–C), 129.01 (Ar–C), 129.29 (Ar–C), 130.25 (Ar–C), 130.65 (Ar–C), 130.88 (Ar–C), 132.29
(Ar–C), 132.80 (Ar–C), 134.65 (Ar–C), 136.32 (Ar–C), 137.67 (Ar–C), 138.39 (Ar–C), 150.07
(C-3), 156.22 (C-3ꢀꢀ), 158.90 (C-5ꢀꢀ), 169.05 (C-8). Anal. Calcd. for C28H22N4O3S (494.57): C,
68.01; H, 4.45; N, 11.33. Found: C, 68.03; H, 4.48; N, 11.30%.
5.21. 9-{-5-[(E)-2-(4-Methoxyphenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-3,4-diphenyl-1-
oxa-6-thia-2,4,9-triazaspiro[4.4] non-2-en-8-one (5b)
1
White powder; yield 67%, m.p. 223–224◦C; IR (KBr) cm−1: 1640 (C N), 1680 (C O); H
=
=
NMR (300 MHz, CDCl3) δ: 2.32 (s, 3H, CH3), 3.78 (s, 3H, OCH3), 4.31 (s, 2H, CH2), 6.7+2
=
=
(d, J = 15.2 Hz, 1H, CH CH), 7.15–7.40 (m, 14H, ArH and 1H, CH CH); EI-MS [M + H]
m/z 525. 13C NMR (75 MHz, CDCl3) δ (ppm): 11.44 (C-6ꢀꢀ), 27.05 (C-7), 55.00 (Ar–OCH3),
100.08 (C-5), 106.01 (C-4ꢀꢀ), 111.20 (C-7ꢀꢀ), 119.50 (C-8ꢀꢀ), 120.05 (Ar–C), 120.98 (Ar–C), 124.85
(Ar–C), 126.05 (Ar–C), 127.55 (Ar–C), 128.00 (Ar–C), 129.33 (Ar–C), 129.55 (Ar–C), 130.08
(Ar–C), 131.04 (Ar–C), 133.00 (Ar–C), 132.60 (Ar–C), 135.02 (Ar–C), 136.65 (Ar–C), 137.80
(Ar–C), 138.44 (Ar–C), 140.05 (Ar–C), 141.08 (Ar–C), 151.05 (C-3), 155.23 (C-3ꢀꢀ), 158.88 (C-
5ꢀꢀ), 170.05 (C-8). Anal. Calcd. for C29H24N4O4S (524.60): C, 66.41; H, 4.58; N, 10.70. Found:
C, 66.39; H, 4.55; N, 10.67%.
5.22. 9-{-5-[(E)-2-(4-Chlorophenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-3,4-diphenyl-1-oxa-
6-thia-2,4,9-triazaspiro[4.4]non-2-en-8-one (5c)
1
White powder; yield 73%, m.p. 211–212◦C; IR (KBr) cm−1: 1635 (C N), 1685 (C O); H
=
=
NMR (300 MHz, CDCl3) δ: 2.25 (s, 3H, CH3), 4.22 (s, 2H, CH2), 6.80 (d, J = 15.2 Hz, 1H,
+
=
=
CH CH), 7.3–8.0 (m, 14H, ArH and 1H, CH CH); EI-MS [M + H] m/z 529. Anal. Calcd. for
C28H21N4O3SCl (529.02): C, 63.63; H, 3.97; N, 10.60. Found: C, 63.60; H, 3.95; N, 10.58%.

Journal of Sulfur Chemistry 273
5.23. Synthesis of 9-{-5-[(E)-2-(2-chlorophenyl)-1-ethenyl]-3-methyl-4-isoxazolyl}-3,4-
diphenyl-1-oxa-6-thia-2,4,9-triazaspiro[4.4]non-2-en-8-one (5d)
1
Yield 78%, m.p. 208–209◦C; IR (KBr) cm−1: 1640 (C N), 1670 (C O); H NMR (300 MHz,
=
=
=
CDCl3) δ: 2.25 (s, 3H, CH3), 4.30 (s, 2H, CH2), 6.70 (d, J = 15.2 Hz, 1H, CH CH), 7.22–7.80
+
=
(m, 14H, ArH and 1H, CH CH); EI-MS [M + H] m/z 529. Anal. Calcd. for C28H21N4O3SCl
(529.02): C, 63.63; H, 3.97; N, 10.60. Found: C, 63.66; H, 3.99; N, 10.63%.
5.24. 4-(4-Methoxyphenyl)-9-{-3-methyl-5[(E)-2-(4-methoxyphenyl)-1-ethenyl]-4-
isoxazolyl}-3-phenyl-1-oxa-6-thia-2,4,9-triazaspiro[4.4] non-2-en-8-one (5e)
1
White powder; yield 65%, m.p. 229–230◦C; IR (KBr) cm−1: 1635 (C N), 1675 (C O); HNMR
=
=
(300 MHz, CDCl3) δ: 2.23 (s, 3H, CH3), 2.55 (s, 3H, CH3), 3.80 (s, 3H, OCH3), 4.22 (s, 2H,
=
=
CH2), 6.80 (d, J = 15.2 Hz, 1H, CH CH), 7.15–7.93 (m, 13H, ArH and 1H, CH CH); EI-MS
[M + H]+ m/z 523. Anal. Calcd. for C30H26N4O3S (522.62): C, 68.91; H, 4.83; N, 10.60. Found:
C, 66.87; H, 4.86; N, 10.58%.
5.25. 3-(4-Chlorophenyl)-9-{3-methyl-5[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl-4-phenyl-1-
oxa-6-thia-2,4,9-triazaspiro[4.4]non-2-en-8-one (5f)
1
White powder; yield 68%, m.p. 232–233◦C; IR (KBr) cm−1: 1635 (C N), 1695 (C O); H NMR
=
=
=
(300 MHz, CDCl3) δ: 2.35 (s, 3H, CH3), 4.12 (s, 2H, CH2), 6.72 (d, J = 15.2 Hz 1H, CH CH)
+
=
and 7.22–8.14 (m, 14H, ArH and 1H, CH CH) EI-MS [M + H] m/z 529. Anal. Calcd. for
C28H21N4O3SCl (529.02): C, 63.63; H, 3.97; N, 10.60. Found: C, 63.58; H, 3.94; N, 10.58%.
5.26. 3-(4-Methoxyphenyl)-9-{-3-methyl-5[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-4-phenyl-
1-oxa-6-thia-2,4,9-triazaspiro[4.4]non-2-en-8-one (5g)
1
Yield 72%, m.p. 225–226◦C; IR (KBr) cm−1: 1640 (C N), 1680 (C O); H NMR (300 MHz,
=
=
CDCl3) δ: 2.20 (s, 3H, CH3), 3.80 (s, 3H, OCH3), 4.15 (s, 2H, CH2), 6.62 (d, J = 15.2 Hz,
+
=
=
1H, CH CH), 7.32–7.95 (m, 14H, ArH and 1H, CH CH); EI-MS [M + H] m/z 524. Anal.
Calcd. for C29H24N4O4S (524.60): C, 66.41; H, 4.58; N, 10.68. Found: C, 66.43; H, 4.55; N,
10.70%.
5.27. 9-{3-Methyl-5-[(E)-2-phenyl-1-ethenyl]-4-isoxazolyl}-3-(4-nitrophenyl)-4-phenyl-1-
oxa-6-Thia-2,4,9-triazaspiro[4.4] non-2-en-8-one (5h)
White powder; yield 70%, m.p. 235–236◦C; IR (KBr) cm−1: 1635 (C N), 1695 (C O); H
1
=
=
NMR (300 MHz, CDCl3) δ: 2.25 (s, 3H, CH3), 4.22 (s, 2H, CH2), 6.80 (d, J = 15.2 Hz,
+
=
=
1H, CH CH), 7.22–7.85 (m, 14H, ArH and 1H, CH CH); EI-MS [M + H] m/z 540. Anal.
Calcd. for C28H21N5O5S (539.57): C, 62.33; H, 3.89; N, 12.98. Found: C, 62.34; H, 3.86; N,
12.95%.
Acknowledgements
The authors are thankful to Head, Department of Chemistry, Kakatiya University, Warangal, India, for the facilities and
to Director, Indian Institute of Chemical Technology, Hyderabad, India, for the spectral facility.

274 E. Rajanarendar et al.
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