Synthesis of novel spirooxindole derivatives by one pot multicomponent reaction and their antimicrobial activity

Article abstract of DOI:10.1016/j.ejmech.2012.02.024

A series of novel spirooxindoles have been synthesized through 1,3-dipolar cycloaddition of an azomethine ylide generated from isatin and sarcosine or l-proline with the dipolarophile 1,4-naphthoquinone followed by spontaneous dehydrogenation. Synthesised compounds were evaluated for their antimicrobial activities against eight bacteria and three fungi. All the spirooxindole derivatives exhibited significant antibacterial activity against Staphylococcus aureus, S. aureus (MRSA), Enterobacter aerogens, Micrococcus luteus, Proteus vulgaris, Klebsiella pneumonia, Salmonella typhimurium, Salmonella paratyphi-B and anti-fungal activity against Malassesia pachydermatis, Candida albicans and Botyritis cinerea organisms. Among 23 compounds screened, 1′-acetyl-2, 5′-dimethyl-2,3-dihydrospiro[benzo[f]isoindole-1,3′-indoline] -2′,4,9-trione was found to be more active against tested bacteria and fungi.

Full text of DOI:10.1016/j.ejmech.2012.02.024

European Journal of Medicinal Chemistry 51 (2012) 79e91
Contents lists available at SciVerse ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis of novel spirooxindole derivatives by one pot multicomponent reaction
and their antimicrobial activity
Gangaru Bhaskar a, Yuvaraj Arun a, Chandrasekar Balachandran b, Chandrasekara Saikumar a,
,
Paramasivan T. Perumal a
*
a Organic Chemistry Division, Central Leather Research Institute (CSIR), Chennai 600 020, India
b Division of Microbiology, Entomology Research Institute, Loyola College, Chennai 600 034, India
a r t i c l e i n f o
a b s t r a c t
Article history:
A series of novel spirooxindoles have been synthesized through 1,3-dipolar cycloaddition of an azo-
Received 28 September 2011
Received in revised form
28 December 2011
Accepted 11 February 2012
Available online 17 February 2012
methine ylide generated from isatin and sarcosine or
L-proline with the dipolarophile 1,4-
naphthoquinone followed by spontaneous dehydrogenation. Synthesised compounds were evaluated
for their antimicrobial activities against eight bacteria and three fungi. All the spirooxindole derivatives
exhibited significant antibacterial activity against Staphylococcus aureus, S. aureus (MRSA), Enterobacter
aerogens, Micrococcus luteus, Proteus vulgaris, Klebsiella pneumonia, Salmonella typhimurium, Salmonella
paratyphi-B and anti-fungal activity against Malassesia pachydermatis, Candida albicans and Botyritis
cinerea organisms. Among 23 compounds screened, 10-acetyl-2,50-dimethyl-2,3-dihydrospiro[benzo[f]
isoindole-1,30-indoline]-20,4,9-trione was found to be more active against tested bacteria and fungi.
Ó 2012 Elsevier Masson SAS. All rights reserved.
Keywords:
Spirooxindole
Multicomponent reaction
Azomethine ylide cycloaddition
Antimicrobial activity
1. Introduction
BE (2) and malignant glioma GAMG [12]. Spirotryprostatins A 5 and
B 6 [13,14]foundinthe secondary metabolites ofAspergillusfugimatus
Heterocyclic compounds particularly spirooxindoles with
nitrogen-containing five membered ring have played an important
role in the field of medicinal chemistry (Fig. 1). Multicomponent
reactions (MCR’s) have emerged as a powerful tool for delivering
the molecular diversity needed in the combinatorial approaches for
the preparation of bioactive compounds [1]. 1,3-Dipolar cycload-
dition reaction is an efficient method for the construction of
heterocyclic units in a highly regio- and stereo- selective manner
[2]. In particular, the chemistry of azomethine ylides have gained
significance in recent years as it serves as an expedient precursor
for constructing spirooxindoles with nitrogen-containing five
membered heterocycles, which constitute the central skeleton of
numerous natural products [3].
Synthetic or natural heterocyclic compound containing spiroox-
indole framework is endowed with a wide range of pharmacological
activities [4]. Naturally occurring spirooxindole alkaloids (Fig.1), such
as horsfiline 1 [5e10] isolated from Horsfieldia superba and elacomine
2 [11] isolated from Eleagnus commutata find use as indigenous
medicine. Alstonisine 3 is a useful bioactive natural product. Mitra-
phylline 4 isolated from Uncaria tomentosa possesses anti-tumor
activity against human brain cancer cell lines, neuroblastoma SKN-
inhibit mammalian cell cycle at G2/M phase. Rhynchophylline 7
isolated from Uncaria rhynchophylla has been used as antipyretic,
anti-hypertensive and anticonvulsant medications for the treatment
of headache, vertigo and epilepsy [15] and as noncompetitive
antagonists of the NMDA receptor [16]. Generally, oxindole deriva-
tives possess anti-tumor [17], antimicrobial[18], antiproliferative [19]
and anti-fungal activities [20] and protein kinase B/Akt inhibitory
activity useful for the treatment of cancer [21].
2. Chemistry
The above mentioned biological importance of spirooxindole
alkaloids led us to synthesize novel spiro-heterocycles comprising
oxindole related ring systems [22]. In continuation of our studies in
the area of 1,3-dipolar cycloaddition reactions and with a view to
synthesis of novel spirooxindoles [23,24], we herein report the
novel one pot tandem/domino reaction of 1,3-dipolar cycloaddition
reaction followed by spontaneous dehydrogenation to form the
spirooxindole derivatives. To the best of our knowledge, there have
been no reports for the synthesis of spirooxindole derivatives
containing 1,4-naphthaquinone moiety.
In the present investigation, the 1,3-dipolar cycloaddition of
azomethine ylides, generated insitu via decarboxylative condensa-
* Corresponding author. Tel./fax: þ91 44 24913289.
E-mail address: ptperumal@gmail.com (P.T. Perumal).
tion of substituted isatins 8aen and sarcosine
9 to 1,4-
0223-5234/$ e see front matter Ó 2012 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2012.02.024

80
G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
Fig. 1. Naturally occurring spirooxindole compounds.
naphthoquinone 10 followed by spontaneous dehydrogenation in
ethanol afforded novel spirooxindoles 12aen in high yields
(Scheme 1).
To further explore the potential of this protocol for novel spi-
rooxindole synthesis, this reaction was further explored by heating
an equimolar mixture of substituted isatin 8aeh, 1,4-
This reaction was performed by heating an equimolar mixture of
substituted isatins 8aen, sarcosine 9 and 1,4-naphthoquinone 10 in
ethanol under reflux for 1.5 h. After completion of the reaction
(TLC), the reaction mixture was poured into ice-water, the resulting
solid was filtered off and purified by column chromatography to
obtain pure spirooxindole derivatives 12aen in 89e96% yields
(Table 1).
In fact we planned to synthesize 1,3-dipolar cycloaddition
product 11, but we obtained product 12 instead. In 1H NMR, we
found that there are no peaks for two CeH aliphatic protons for
desired product 11. 13C NMR exhibited the number of peaks equal to
the desired product 11 but two peaks were in the C]C region
instead of the aliphatic region. According to DEPT 135 and DEPT 90,
presence of two quaternary aromatic carbon signals instead of two
aliphatic eCH carbons also prove the formation of product 12. The
mass spectrum also confirmed the dehydrogenated product 12.
Single crystal XRD of compound 14b (Fig. 2) established the eluci-
dated dehydrogenated product 12 (Scheme 2).
We tried to isolate the 1,3-dipolar cycloaddition product 11l at
nitrogen atmosphere. We found this product 11l at nitrogen
atmosphere and isolated as a pure product and confirmed by
spectral data (see supporting information). But we obtained
dehydrogenated product in atmospheric reflux condition. So this
reaction can be a domino type reaction of 1,3-dipolar cycloaddition
followed by spontaneous air oxidation resulting in the novel spi-
rooxindole products 12l (Scheme 3).
naphthoquinone 10 and -proline 13 in ethanol under reflux for
1.5 h and obtained pure spirooxindole derivatives 14aeh in 87e93%
yields (Scheme 4). The results are summarized in Table 2.
L
The unexpected promising results prompted us to extend this
protocol to synthesize spirooxindole derivatives 17 and 18
involving acenaphthenequinone 15 and ninhydrin 16 under opti-
mized condition gave good yield of respective spirooxindole
derivatives in 91 and 93% of yield (Scheme 5).
The structure of unusual dehydrogenative spirooxindole deriv-
atives by 1,3-dipolar cycloaddition of azomethine ylide was eluci-
dated with the help of IR, 1H NMR, 13C NMR and Mass data as
illustrated for 12a. In the IR spectrum, the sharp peak appeared at
3425 cmꢀ1 corresponds to the spirooxindole NeH group and the
sharp peaks at 1721 and 1628 cmꢀ1 correspond to C]O functional
groups of the product 12a. In the 1H NMR spectrum, peaks in the
range of
in the region
proton and the signal at
the presence of eNCH3 group. The ABq peak at
J value 16.8 Hz for two protons shows the presence of pyrrolidine
ring eCH2 group. The 13C NMR, the peak at
: 77.6 ppm corresponds
to the spiro carbon and the peak at : 175.6 ppm shows amide
carbonyl carbon. The peaks at : 180.8 and 182.1 ppm confirmed the
presence of two carbonyl groups. In dept 135 and 90, the peaks at
d
: 6.85e8.00 ppm shows aromatic protons. A broad singlet
: 10.60 ppm signal confirmed the presence of eNH
: 2.12 ppm for three protons confirmed
: 4.21 ppm with the
d
d
d
d
d
d
d
:
57.7 & 34.7 ppm confirms the presence of eCH2 and eNCH3 group
respectively. A distinguishing peak was observed at m/z: 331 in the
Scheme 1. Synthesis of spirooxindole derivatives from substituted isatin, sarcosine and 1,4-napthoquinone.

G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
81
Table 1
Synthesis of spirooxindole derivatives from substituted isatin, sarcosine and 1,4-naphthoquinone.
Entry Isatin
Producta
Yield (%)b
1
91
89
94
2
3
4
92
5
92
6
94
(continued on next page)

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G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
Table 1 (continued )
Entry
Isatin
Producta
Yield (%)b
7
90
8
95
9
94
10
92
11
93
12
91
13
89

G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
83
Table 1 (continued )
Entry
Isatin
Producta
Yield (%)b
96
14
a
All the products were characterised by Mass, IR and NMR spectrum.
Isolated yield after purification by column chromatography.
b
mass spectrum for [M þ H]þ ion further conforms the product 12a.
The structure determined from an X-ray crystallographic study of
the single crystal of the derivative 14b confirms the structure that
deduced from spectroscopic data (Fig. 2). Table 3 gives crystal data,
data collection and refinement parameters of 14b.
bacteria. Compounds 12b, 12d, 12i and 12k showed good activity
against S. aureus. In comparison to compound 12a, the presence of
methyl group on the phenyl ring (12b) increased the potency
against S. aureus by one fold. Interestingly the presence of methyl
groups in both phenyl ring and nitrogen (12k) increased the
potency against S. aureus by two fold. Also compound 12d (N-ethyl)
increased the potency against S. aureus by two fold. In comparison
to compound 12a, the presence of acetyl group on the isatin
nitrogen (12n) increased the potency against bacteria S. aureus by
one fold, S. aureus (MRSA) by two fold, M. luteus, K. pneumonia and
S. typhimurium by three fold and E. aerogens, P. vulgaris and
S. paratyphi-B by four fold and also against fungi C. albicans by two
fold and M. pachydermatis by four fold. Compound 12n (N-acetyl)
showed good activity than standard drugs for most of the tested
bacteria and fungi.
3. Biology
In the present study, the antimicrobial activities of synthesized
compounds were screened against eight bacteria and three fungi
using in vitro disc diffusion method. The results revealed that most
of the synthesised compounds exhibited antimicrobial activities
against Staphylococcus aureus, S. aureus (MRSA), Enterobacter
aerogens, Micrococcus luteus, Proteus vulgaris, Klebsiella pneumonia,
Salmonella typhimurium, Salmonella paratyphi-B, Malassesia pachy-
dermatis, Candida albicans and Botyritis cinerea organisms. The
results are summarized in Table 4 and Fig. 3. Compounds 12d, 12i,
12k, 12l and 12n showed good activity more than standard drug
against S. aureus. Compound 12n showed good activity against both
Gram-positive and Gram-negative bacteria among all synthesized
compounds compare with standards. Compound 12k showed good
activity among N-alkyl substituted compounds 12d, 12l and 12m.
Halo substituted compounds 12i and 12j shows good activity
against tested bacteria. Compound 12n showed significant anti-
fungal activity against C. albicans, M. pachydermatis and B. cinerea.
The MIC values of active compounds (12aee & 12ien) against
bacteria and fungi are given in Table 5 and Fig. 4. Significant MIC
values were observed against Gram positive and Gram negative
4. Conclusion
In summary, we have reported the synthesis of novel spiroox-
indole derivatives through 1,3-dipolar cycloaddition of an azome-
thine ylide generated from isatin and sarcosine or L-proline with
the dipolarophile 1,4-naphthoquinone followed by dehydrogena-
tion and characterisation of synthesised spirooxindole derivatives.
These novel compounds were evaluated for their activities against
eight bacteria and three fungi. Compound 12n 10-acetyl-2,50-
dimethyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-indoline]-20,4,9-
trione was found to be more than 1.6 times active against S. aureus
(MRSA) bacteria than streptomycin and ciprofloxacin. Also more
than 6.4 times active against M. luteus and S. typhimurium bacteria
than ciprofloxacin. Also more than 3.2 times active against
C. albicans fungi than fluconazole. These results showed that
synthesized spirooxindole compound 12n might be a potential
antibacterial and anti-fungal agent.
5. Experimental
5.1. Chemistry
Melting points were determined in capillary tubes and are
uncorrected. IR spectra were taken as KBr pellets for solids on
a Perkin Elmer Spectrum RXI FT-IR. 1H NMR (500 MHz) and 13C
NMR (125 MHz) spectra were recorded in DMSO-d6 solutions with
TMS as an internal standard on a JEOL instrument. Mass spectra
were recorded on a Thermo Finnigan LCQ Advantage MAX 6000 ESI
spectrometer. Elemental analysis data were recorded using Thermo
Finnigan FLASH EA 1112 CHN analyzer.
Fig. 2. ORTEP diagram of synthesized compound 14b.

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G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
Scheme 2. Plausible mechanism for the formation of spirooxindoles.
5.1.1. Experimental procedure for the synthesis of spirooxindole
derivatives (12aen)
Anal. Calcd for C20H14N2O3: C, 72.72; H, 4.27; N, 8.48. Found: C,
72.59; H, 4.39; N, 8.37.
A mixture of substituted isatin 8aen (1 mmol), sarcosine 9
(1 mmol) and 1,4-naphthoquinone 10 (1 mmol) was refluxed in
ethanol (5 ml). Completion of the reaction was evidenced by TLC
analysis. After completion of the reaction, the reaction mixture was
poured into ice-water, the resulting solid was filtered off and
purified by column chromatography using ethyl acetate: petroleum
ether (3:7) as an eluent to afford pure spirooxindoles (12aen).
5.1.1.2. 2,50-Dimethyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-indo-
line]-20,4,9-trione (12b). Reddish brown solid; mp 210e211 ꢁC; IR
(cmꢀ1): 3774, 3419, 2369, 1945, 1713, 1588, 1182, 607; 1H NMR
(500 MHz, DMSO-d6):
d 2.14 (s, 3H), 2.15 (s, 3H), 4.21 (ABq, 2H,
J ¼ 16.1 Hz), 6.75 (d, 1H, J ¼ 8.4 Hz), 6.94e7.09 (m, 2H), 7.74e7.79
(m, 3H), 7.99 (d, 1H, J ¼ 6.8Hz), 10.4 (brs, 1H, -NH, D2O exchange-
able); 13C NMR (125 MHz, DMSO-d6):
d 21.0, 34.7, 57.7, 77.7, 110.2,
5.1.1.1. 2-Methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-indoline]-
20,4,9-trione (12a). Brown solid; mp 218e219 ꢁC; IR (cmꢀ1): 3425,
2926, 1969, 1721, 1628, 1594, 1383, 1221, 1048, 581; 1H NMR
126.3, 126.4, 126.5, 128.3, 130.4, 131.2, 132.5, 132.7, 134.9, 140.4,
147.6, 150.8, 162.9, 175.4, 180.8, 182.1; MS m/z ¼ 345 [MH]þ; Anal.
Calcd for C21H16N2O3: C, 73.24; H, 4.68; N, 8.13. Found: C, 73.34; H,
4.77; N, 8.01.
(500 MHz, DMSO-d6):
d
2.12 (s, 3H), 4.21 (ABq, 2H, J ¼ 16.8 Hz),
6.85e6.93 (m, 2H), 7.18 (d, 1H, J ¼ 6.9 Hz), 7.22 (t, 1H, J ¼ 7.6 Hz),
7.76e7.85 (m, 3H), 8.00 (d, 1H, J ¼ 6.9 Hz), 10.6 (brs, 1H, -NH, D2O
5.1.1.3. 10,2-Dimethyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-indo-
line]-20,4,9-trione (12c). Brown solid; mp 240e241 ꢁC; IR (cmꢀ1):
3415, 3054, 2931, 2832, 1719, 1662, 1601, 1462, 1336, 1250, 1216,
1085, 1018, 847, 790, 744, 713, 533, 473, 124; 1H NMR (500 MHz,
exchangeable); 13C NMR (125 MHz, DMSO-d6):
d 34.7, 57.7, 77.6,
110.5, 122.4, 125.7, 126.4, 126.6, 128.1, 130.2, 132.4, 132.7, 134.9,
135.0, 142.8, 147.5, 150.9, 175.6, 180.8, 182.1; MS m/z ¼ 331 [MH]þ;
Scheme 3. Synthesis of spirooxindole derivative 11l at nitrogen atmosphere.

G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
85
Scheme 4. Synthesis of spirooxindole derivatives from substituted isatin, L-proline and 1,4-napthoquinone.
Table 2
Synthesis of spirooxindole derivatives from substituted isatin,
L-proline and 1,4-naphthoquinone.
Entry Isatin
Producta
Yield (%)b
1
87
2
89
3
92
4
88
5
93
6
91
(continued on next page)

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G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
Table 2 (continued )
Entry
Isatin
Producta
Yield (%)b
7
90
8
92
a
All the products were characterised by Mass, IR and NMR spectrum.
Isolated yield after purification by column chromatography.
b
DMSO-d6):
d
2.09 (s, 3H), 3.16 (s, 3H), 4.23 (ABq, 2H, J ¼ 17.6 Hz),
(m, 2H), 7.78e7.86 (m, 3H), 8.01 (d, 1H, J ¼ 7.6 Hz); 13C NMR
6.98 (t,1H, J ¼ 6.8 Hz), 7.07 (d, 1H, J ¼ 8.4 Hz), 7.25 (d, 1H, J ¼ 7.6 Hz),
(125 MHz, DMSO-d6): d 34.7, 42.2, 57.8, 77.2, 110.1, 117.1, 123.0,
7.34 (t, 1H, J ¼ 7.6 Hz), 7.38e7.76 (m, 3H), 8.01 (d, 1H, J ¼ 7.6 Hz); 13C
125.6, 126.4, 126.6, 127.3, 130.2, 132.1, 132.4, 132.8, 134.8, 134.9,
143.4, 147.3, 151.2, 173.7, 180.8, 182.0; MS m/z ¼ 371 [MH]þ; Anal.
Calcd for C23H18N2O3: C, 74.58; H, 4.90; N, 7.56. Found: C, 74.67;
H, 4.82; N, 7.69.
NMR (125 MHz, DMSO-d6):
d 26.7, 34.8, 57.8, 77.2, 109.4, 122.9,
125.4, 126.4, 126.6, 127.3, 130.3, 132.4, 132.8, 134.8, 134.9, 144.3,
147.3, 151.0, 173.9, 180.7, 182.0; MS m/z ¼ 345[MH]þ; Anal. Calcd for
C21H16N2O3: C, 73.24; H, 4.68; N, 8.13. Found: C, 73.14; H, 4.61;
N, 8.27.
5.1.1.7. 10-Benzyl-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12g). Brown solid; mp 219e220 ꢁC; IR
(cmꢀ1): 3776, 3430, 3030, 2918, 2843, 1721, 1659, 1598, 1464, 1337,
1173, 969, 793, 698, 618, 550, 469; 1H NMR (500 MHz, DMSO-d6):
5.1.1.4. 1-Ethyl-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12d). Yellow solid; mp 247e248 ꢁC; IR
(cmꢀ1): 3412, 3051, 2931, 1719, 1662, 1462, 1332, 1216, 1084, 1018,
d
2.12 (s, 3H), 4.26 (ABq, 2H, J ¼ 16.8 Hz), 4.93 (s, 2H), 6.89 (d, 1H,
847, 713; 1H NMR (500 MHz, DMSO-d6):
d
1.33 (t, 3H, J ¼ 6.8 Hz),
J ¼ 7.6 Hz), 6.96 (t, 1H, J ¼ 6.9 Hz), 7.21e7.29 (m, 3H), 7.33 (t, 2H,
J ¼ 7.6 Hz), 7.40 (d, 2H, J ¼ 7.6 Hz), 7.78e7.86 (m, 3H), 8.01 (d, 1H,
2.13 (s, 3H), 3.38 (q, 2H, J ¼ 6.8 Hz), 4.25 (ABq, 2H, J ¼ 16.8 Hz),
6.97 (t, 1H, J ¼ 6.8 Hz), 7.07 (d, 1H, J ¼ 8.4 Hz), 7.24 (d, 1H,
J ¼ 7.6 Hz), 7.35 (t, 1H, J ¼ 7.6 Hz), 7. 36e7.74 (m, 3H), 8.01 (d, 1H,
J ¼ 6.9 Hz); 13C NMR (125 MHz, DMSO-d6):
d 34.8, 43.5, 57.8, 77.2,
110.1, 123.1, 125.6, 126.5, 126.6, 127.3, 127.6, 127.9, 129.1, 130.3, 132.4,
132.8, 134.9, 135.0, 136.6, 143.3, 147.2, 151.3, 174.2, 180.8, 182.0; MS
m/z ¼ 421 [MH]þ; Anal. Calcd for C27H20N2O3: C, 77.13; H, 4.79; N,
6.66. Found: C, 77.01; H, 4.86; N, 6.72.
J ¼ 7.6 Hz); 13C NMR (125 MHz, DMSO-d6):
d 10.9, 34.5, 39.5, 57.8,
77.2, 109.3, 122.9, 125.4, 126.3, 126.5, 127.3, 130.1, 132.4, 132.8,
134.8, 134.9, 144.4, 147.2, 151.1, 173.9, 180.7, 182.0; MS m/z ¼ 359
[MH]þ; Anal. Calcd for C22H18N2O3: C, 73.73; H, 5.06; N, 7.82.
Found: C, 73.84; H, 5.14; N, 7.72.
5.1.1.8. 50-Chloro-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12h). Yellow solid; mp 237e238 ꢁC; IR
(cmꢀ1): 3776, 3182,1869,1667,1598,1450,1277,1179, 959, 882, 712,
5.1.1.5. 2-Methyl-10-(prop-2-ynyl)-2,3-dihydrospiro[benzo[f]isoindole-
1,30-indoline]-20,4,9-trione (12e). Yellow solid; mp 223e224 ꢁC; IR
(cmꢀ1): 3779, 3656, 3238, 2934, 2365, 2208, 1724, 1678, 1602, 1460,
553; 1H NMR (500 MHz, DMSO-d6):
d 2.16 (s, 3H), 4.24 (ABq, 2H,
J ¼ 16.8 Hz), 6.88 (d, 1H, J ¼ 8.4 Hz), 7.27 (d, 1H, J ¼ 8.4 Hz),
7.31e7.33 (m, 1H), 7.78e7.84 (m, 3H), 8.01 (d, 1H, J ¼ 7.6 Hz), 10.7
(brs, 1H, eNH, D2O exchangeable); 13C NMR (125 MHz, DMSO-d6):
1338, 1185, 962, 720, 483; 1H NMR (500 MHz, DMSO-d6):
d 2.08 (s,
3H), 3.26 (t,1H, J ¼ 2.3 Hz), 4.23 (ABq, 2H, J ¼ 16.8 Hz), 4.57 (ABq, 2H,
J ¼ 17.6 Hz), 7.02 (t, 1H, J ¼ 7.6 Hz), 7.16 (d, 1H, J ¼ 8.4 Hz), 7.28 (d, 1H,
J ¼ 7.6 Hz), 7.36 (t, 1H, J ¼ 7.6 Hz), 7.77e7.85 (m, 3H), 7.99 (d, 1H,
d
34.7, 57.8, 77.6, 111.9, 126.0, 126.4, 126.5, 130.1, 130.3, 132.5, 132.8,
132.9, 134.8, 134.9, 141.8, 146.7, 151.2, 175.3, 180.8, 182.0; MS m/
z ¼ 364 [MH]þ; Anal. Calcd for C20H13ClN2O3: C, 65.85; H, 3.59; N,
7.68. Found: C, 65.74; H, 3.67; N, 7.74.
J ¼ 7.6 Hz); 13C NMR (125 MHz, DMSO-d6):
d 29.5, 34.6, 57.8, 74.9,
77.0, 78.3, 110.2, 123.3, 125.7, 126.4, 126.6, 127.2, 130.3, 132.4, 132.8,
134.8, 134.9, 142.4, 146.9, 151.2, 173.3, 180.6, 182.0; MS m/z ¼ 369
[MH]þ; Anal. Calcd for C23H16N2O3: C, 74.99; H, 4.38; N, 7.60. Found:
C, 75.09; H, 4.45; N, 7.49.
5.1.1.9. 50-Bromo-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12i). Brown solid; mp 172e173 ꢁC; IR
(cmꢀ1): 3249, 2868, 1720, 1596, 1459, 1282, 1203, 806, 708; 1H NMR
5.1.1.6. 10-allyl-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12f). Brown solid; mp 212e213 ꢁC; IR
(cmꢀ1): 3016, 2886, 1715, 1678, 1609, 1461, 1341, 1169, 962, 728,
(500 MHz, DMSO-d6):
d
2.14 (s, 3H), 4.24 (ABq, 2H, J ¼ 16.8 Hz), 6.86
(d, 1H, J ¼ 8.4 Hz), 7.33 (d, 1H, J ¼ 8.4 Hz), 7.31e7.34 (m, 1H),
7.77e7.85 (m, 3H), 8.01 (d, 1H, J ¼ 7.6 Hz), 10.7 (brs, 1H, -NH, D2O
483; 1H NMR (500 MHz, DMSO-d6):
d
2.12 (s, 3H), 4.26 (ABq, 2H,
exchangeable); 13C NMR (125 MHz, DMSO-d6):
d 34.7, 57.8, 77.6,
J ¼ 16.8 Hz), 4.29e4.39 (m, 2H), 5.17 (d, 1H, J ¼ 10.7 Hz), 5.31 (d,
126.1, 126.2, 126.6, 128.2, 130.2, 130.3, 132.7, 132.7, 132.9, 134.4,
1H, J ¼ 16.8 Hz), 5.81e5.95 (m, 1H), 6.96e7.01 (m, 2H), 7.26e7.33
134.9, 141.8, 146.6, 151.2, 175.2, 180.8, 182.1; MS m/z ¼ 409 [MH]þ;

G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
87
Scheme 5. Synthesis of spirooxindole derivatives from acenaphthenequinone or ninhydrin with sarcosine and 1,4-naphthoquinone.
Anal. Calcd for C20H13BrN2O3: C, 58.70; H, 3.20; N, 6.85. Found: C,
58.52; H, 3.13; N, 6.62.
DMSO-d6): d 21.0, 26.8, 34.7, 57.7, 77.8, 110.2, 126.2, 126.4, 126.5,
128.4, 130.4, 131.1, 132.5, 132.6, 134.9, 140.4, 147.6, 150.9, 162.9,
175.4, 180.8, 182.1; MS m/z ¼ 359 [MH]þ; Anal. Calcd for
C22H18N2O3: C, 73.73; H, 5.06; N, 7.82. Found: C, 73.59; H, 5.18;
N, 7.77.
5.1.1.10. 50-Iodo-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12j). Pale yellow solid; mp 174e175 ꢁC; IR
(cmꢀ1): 3259, 2862, 1711, 1596, 1459, 1281, 1204, 806, 708; 1H
NMR (500 MHz, DMSO-d6):
d
2.14 (s, 3H), 4.24 (ABq, 2H,
5.1.1.12. 10-Butyl-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12l). Brown solid; mp 148e149 ꢁC; IR
(cmꢀ1): 3054, 2933, 2872, 2802, 1707, 1667, 1602, 1459, 1342, 1218,
J ¼ 16.8 Hz), 6.84 (d, 1H, J ¼ 8.4 Hz), 7.31 (d, 1H, J ¼ 8.4 Hz),
7.32e7.35 (m, 1H), 7.79e7.86 (m, 3H), 8.01 (d, 1H, J ¼ 7.6 Hz), 10.7
(brs, 1H, -NH, D2O exchangeable); 13C NMR (125 MHz, DMSO-d6):
1177, 962, 752; 1H NMR (500 MHz, DMSO-d6):
d
0.92 (t, 3H, J ¼ 7.5),
d
34.7, 57.8, 77.6, 98.2, 126.1, 126.2, 126.6, 130.2, 130.3, 132.7, 132.7,
1.36 (sex, 2H, J ¼ 7.5 Hz), 1.63 (qt, 2H, J ¼ 6.9 Hz), 2.14 (s, 3H), 3.73 (t,
2H, J ¼ 6.9 Hz), 4.27 (ABq, 2H, J ¼ 16.8 Hz), 7.01 (t, 1H, J ¼ 7.2 Hz),
7.14 (d, 1H, J ¼ 7.8 Hz), 7.29 (d, 1H, J ¼ 7.8 Hz), 7.36 (td, 1H, J ¼ 7.8 Hz,
J ¼ 1.2 Hz), 7.81e7.86 (m, 3H), 8.05 (d, 1H, J ¼ 7.5 Hz); 13C NMR
132.9, 134.4, 134.9, 141.8, 146.6, 151.2, 175.2, 180.8, 182.1; MS m/
z ¼ 457 [MH]þ; Anal. Calcd for C20H13IN2O3: C, 52.65; H, 2.87; N,
6.14. Found: C, 52.53; H, 2.74; N, 6.27.
(125 MHz, DMSO-d6): d 13.6, 19.4, 28.9, 34.1, 57.2, 76.6, 109.0, 122.2,
5.1.1.11. 10,2,50-Trimethyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12k). Brown solid; mp 130e131 ꢁC. IR
(cmꢀ1): 3757, 3403, 2934, 2799, 2363, 1710, 1607, 1495, 1344,
1188, 1100, 806, 705; 1H NMR (500 MHz, DMSO-d6): 1H NMR
125.0, 125.9, 126.0, 127.0, 129.8, 131.9, 132.2, 134.3, 134.4, 143.2,
146.9, 150.5, 173.2, 180.2, 181.5; MS m/z ¼ 387 [MH]þ; Anal. Calcd
for C24H22N2O3: C, 74.59; H, 5.74; N, 7.25. Found: C, 74.43; H, 5.77;
N, 7.11.
(500 MHz, DMSO-d6):
d 2.12 (s, 3H), 2.15 (s, 3H), 3.16 (s, 3H), 4.22
(ABq, 2H, J ¼ 16.1 Hz), 6.75 (d, 1H, J ¼ 8.4 Hz), 6.92e7.12 (m, 2H),
5.1.1.13. 10-Hexyl-2-methyl-2,3-dihydrospiro[benzo[f]isoindole-1,30-
indoline]-20,4,9-trione (12m). Brown solid; mp 132e133 ꢁC; IR
(cmꢀ1): 3055, 2927, 2859, 1715, 1663, 1600, 1463, 1346, 1178, 1022,
7.71e7.74 (m, 3H), 7.78 (d, 1H, J ¼ 6.8Hz); 13C NMR (125 MHz,
965, 744; 1H NMR (500 MHz, DMSO-d6):
d
0.93 (t, 3H, J ¼ 7.5),
Table 3
Crystal data and structure refinement parameters of compounds 14b.
1.22e1.50 (m, 6H), 1.52e1.81 (m, 2H), 2.14 (s, 3H), 3.72 (t, 2H,
J ¼ 6.6 Hz), 4.28 (ABq, 2H, J ¼ 16.8 Hz), 7.01 (q, 1H, J ¼ 7.2 Hz), 7.09
(d, 1H, J ¼ 7.2 Hz), 7.29 (d, 1H, J ¼ 7.2 Hz), 7.36 (td, 1H, J ¼ 7.8 Hz,
J ¼ 1.2 Hz), 7.81e7.87 (m, 3H), 8.05 (d, 1H, J ¼ 7.5 Hz); 13C NMR
Empirical formula
Formula weight
Temperature
Wavelength
Crystal system, space group
Unit cell dimensions
C23 H18 N2 O3
370.39
298(2) K
0.71073 Å
(125 MHz, DMSO-d6):
d 13.5, 14.7, 19.4, 21.3, 28.8, 34.1, 57.1, 76.6,
Monoclinic, P2(1)/n
a ¼ 9.3713(4) Å; b ¼ 9.7977(4) Å;
c ¼ 20.3233(7) Å;
109.0, 122.3, 125.0, 125.8, 126.0, 127.1, 129.8, 131.8, 132.2, 134.4,
134.4, 143.1, 146.9, 150.5, 173.3, 180.2, 181.7; MS m/z ¼ 415 [MH]þ;
Anal. Calcd for C26H26N2O3: C, 75.34; H, 6.32; N, 6.76. Found: C,
75.43; H, 6.67; N, 6.89.
a
¼
g
¼ 90ꢁ;
b
¼ 99.495(2)ꢁ
Volume
1840.46 (13) Å3
Z, Density (calculated)
Absorption coefficient
F (000)
4, 1.337 mg mꢀ3
0.090 mmꢀ1
5.1.1.14. 10-Acetyl-2,50-dimethyl-2,3-dihydrospiro[benzo[f]isoindole-
1,30-indoline]-20,4,9-trione (12n). Brown solid; mp 167e168 ꢁC; IR
(cmꢀ1): 3340, 3082, 1711, 1664, 1600, 1468, 1371, 1267, 1072, 761; 1H
776
Crystal size
0.45 ꢂ 0.40 ꢂ 0.25 mm
q
range for data collection
2.03e32.43ꢁ
Limiting indices
ꢀ13 ꢃ h ꢃ 12, ꢀ14 ꢃ k ꢃ 14, ꢀ23 ꢃ l ꢃ 26
15,052/5097 [R(int) ¼ 0.0212]
Multi-scan
0.9779 and 0.9608
Full-matrix least-squares on F2
5097/0/258
NMR (500 MHz, DMSO-d6): d 2.11 (s, 3H), 2.25 (s, 3H), 4.21 (ABq, 2H,
Reflections collected/unique
Absorption correction
Max. and min. transmission
Refinement method
Data/restraints/parameters
Goodness-of-fit on F2
Largest diff. peak and hole
CCDC
J ¼ 16.8 Hz), 6.84e6.97 (m, 2H), 7.19 (d, 1H, J ¼ 6.9 Hz), 7.32 (t, 1H,
J ¼ 7.6 Hz), 7.77e7.84 (m, 3H), 8.00 (d, 1H, J ¼ 6.9 Hz); 13C NMR
(125 MHz, DMSO-d6):
d 26.4, 34.8, 57.7, 77.6, 110.5, 122.3, 125.5,
126.4, 126.5, 128.1, 130.7, 132.4, 132.7, 134.9, 135.0, 142.8, 147.5,
150.9, 173.1,175.6, 180.8, 182.1; MS m/z ¼ 373 [MH]þ; Anal. Calcd for
C22H16N2O4: C, 70.96; H, 4.33; N, 7.52. Found: C, 70.85; H, 4.39;
N, 7.34.
1.032
0.530 and ꢀ0.421 eÅꢀ3
852,448

88
G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
Table 4
In vitro antimicrobial activity of synthesized compounds.
Compounds
Zone of inhibition in mm
Gram positive bacteria
Gram negative bacteria
Fungi
S. aureus S. aureus (MRSA) M. luteus E. aerogens P. vulgaris K. pneumonia S. typhimurium S. Paratyphi-B M. pachydermatis C. albicans B. cinerea
12a
12b
12c
12d
12e
12f
12g
12h
12i
15
15
13
22
13
11
9
14
20
16
22
17
12
18
12
10
10
Ni
8
10
12
10
13
10
9
13
13
12
18
12
10
9
12
17
15
15
13
10
22
9
11
9
9
9
9
10
8
8
23
Na
10
13
10
12
11
9
11
9
10
15
9
13
13
18
12
11
9
10
13
10
11
13
13
10
24
9
10
9
Ni
Ni
8
11
9
9
12
12
14
19
15
11
9
11
19
14
22
15
16
24
10
11
11
9
10
11
11
10
10
Ni
8
13
8
7
11
12
11
11
10
9
10
8
9
10
10
9
10
9
8
8
10
8
10
8
11
12
11
9
10
15
Ni
9
8
9
Ni
Ni
Ni
9
9
Na
Ni
8
8
8
8
9
8
9
11
12
8
14
8
10
16
8
8
8
Ni
Ni
8
10
16
12
17
10
13
21
9
9
9
Ni
Ni
8
10
12
10
16
10
10
26
8
9
8
8
Ni
10
9
8
8
10
10
14
10
11
12
19
8
10
11
12
10
14
18
Ni
Ni
10
8
Ni
8
Ni
8
Ni
Na
22
12j
12k
12l
8
9
8
12m
12n
14a
14b
14c
14e
14f
14g
14h
17
28
Ni
8
Ni
8
Ni
Ni
Ni
Ni
9
9
10
Ni
Ni
9
9
8
Ni
Na
28
9
9
10
12
9
9
9
Ni
8
22
Na
10
10
10
23
Na
Ni
Ni
20
Na
18
9
Streptomycin 16
Ketoconazole Na
22
Na
20
Na
17
Na
NA e not applicable; NI e no inhibition.
5.1.2. Experimental procedure for the synthesis of spirooxindole
derivatives (14aeh)
5.1.2.1. 1,2,3,11b-Tetrahydrospiro[benzo[f]pyrrolo[2,1-a]isoindole-
5,30-indoline]-20,6,11-trione (14a). White solid; mp 227e228 ꢁC; IR
(cmꢀ1): 3378, 2878, 1735, 1606, 1466, 1304, 1156, 794, 727, 565; 1H
A mixture of substituted isatin 8aeh (1 mmol), -proline 13
L
(1 mmol) and 1,4-naphthoquinone 10 (1 mmol) was refluxed in
ethanol (5 ml). Completion of the reaction was evidenced by TLC
analysis. After completion of the reaction, the reaction mixture was
poured into ice-water, the resulting solid was filtered off and
purified by column chromatography using ethyl acetate: petroleum
ether (3:7) as an eluent to afford pure spirooxindoles (14aeh).
NMR (500 MHz, DMSO-d6): d 1.72e1.88 (m, 3H), 2.14e2.22 (m, 1H),
2.45e2.51 (m, 1H), 2.56e2.63 (m, 1H), 4.74 (t, 1H, J ¼ 7.6 Hz),
6.85e6.91 (m, 2H), 7.09 (d, 1H, J ¼ 7.6 Hz), 7.24 (t, 1H, J ¼ 7.6 Hz),
7.76e7.81 (m, 3H), 8.00 (d, 1H, J ¼ 7.6 Hz), 10.49 (brs, 1H, -NH, D2O
exchangeable); 13C NMR (125 MHz, DMSO-d6):
d 27.8, 30.9, 47.8,
70.7, 77.0, 110.7, 121.7, 126.1, 126.3, 126.5, 127.1, 130.4, 132.7, 133.1,
Fig. 3. Comparison of antimicrobial activity of synthesised compounds and standard drugs.

G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
89
Table 5
MIC ( g/ml) of compounds against tested bacteria and fungi.
m
Compounds
Minimum inhibitory concentration (
mg/ml)
Gram positive bacteria
Gram negative bacteria
Fungi
S. aureus S. aureus (MRSA) M. luteus E. aerogens P. vulgaris K. pneumonia S. typhimurium S. Paratyphi-B M. pachydermatis C. albicans B. cinerea
12a
12b
12c
12d
12e
12i
12j
12k
12l
62.5
31.25
62.5
15.62
125
15.62
62.5
15.62
62.5
250
250
125
250
250
125
250
500
125
500
500
62.5
>100
>100
NA
125
62.5
125
31.25
125
62.5
15.62
125
250
250
15.62
6.25
>100
NA
250
125
125
250
250
62.5
250
62.5
500
250
15.62
25
250
250
125
62.5
500
125
500
125
500
500
<15.62
6.25
<0.78
NA
125
62.5
31.25
250
250
500
250
250
250
500
15.62
6.25
<0.78
NA
125
250
125
250
125
250
500
250
500
500
500
500
500
<15.62
NI
250
125
125
250
250
500
125
500
250
250
15.62
NA
125
250
250
500
250
500
250
250
500
125
31.25
NA
125
250
250
500
250
250
250
250
500
500
125
NA
31.25
62.5
31.25
125
15.62
125
62.5
15.62
30
>100
NA
12m
12n
31.25
Streptomycin 6.25
Ciprofloxacin <0.78
Fluconazole
<0.78
NA
NA
6.25
NA
NA
NA
12.5
15
NA
>100
25
NA
NI
25
NA
Ketoconazole NA
NA
NA
NA
NA
NA
134.7, 134.8, 143.3, 146.1, 152.5, 177.7, 181.5, 182.6; MS m/z ¼ 357
[MH]þ; Anal. Calcd for C22H16N2O3: C, 74.15; H, 4.53; N, 7.86. Found:
C, 74.25; H, 4.47; N, 7.92.
1123,1000, 744, 480; 1H NMR (500 MHz, DMSO-d6):
d
1.71e1.88 (m,
3H), 2.13e2.23 (m, 1H), 2.44e2.50 (m, 1H), 2.57e2.62 (m, 1H), 3.12
(s, 3H), 4.75 (t, 1H, J ¼ 7.6 Hz), 6.84e6.92 (m, 2H), 7.10 (d, 1H,
J ¼ 7.6 Hz), 7.23 (t, 1H, J ¼ 7.6 Hz), 7.75e7.82 (m, 3H), 8.01 (d, 1H,
5.1.2.2. 50-Methyl-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo[2,1-a]
isoindole-5,30-indoline]-20,6,11-trione (14b). White solid; mp
239e240 ꢁC; IR (cmꢀ1): 3172, 2949, 1724, 1638, 1474, 1291, 1178,
J ¼ 7.6 Hz); 13C NMR (125 MHz, DMSO-d6):
d 26.8, 27.8, 31.0, 47.8,
70.8, 76.6, 109.6, 122.4, 125.3, 126.3, 126.5, 126.8, 130.6, 132.6, 133.1,
134.8, 134.9, 144.6, 146.0, 152.6, 176.1, 181.4, 182.6; MS m/z ¼ 371
[MH]þ; Anal. Calcd for C23H18N2O3: C, 74.58; H, 4.90; N, 7.56. Found:
C, 74.48; H, 4.78; N, 7.70.
716, 448; 1H NMR (500 MHz, DMSO-d6):
d 1.71e1.88 (m, 3H), 2.15
(s, 3H), 2.44e2.51 (m, 2H), 2.55e2.65 (m,1H), 4.74 (t,1H, J ¼ 6.8 Hz),
6.77 (d, 1H, J ¼ 7.6 Hz), 6.92 (s, 1H), 7.04 (d, 1H, J ¼ 7.6 Hz), 7.76e7.86
(m, 3H), 8.01 (d, 1H, J ¼ 6.9 Hz), 10.37 (brs, 1H, -NH, D2O
5.1.2.4. 10-Ethyl-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo[2,1-a]iso-
exchangeable); 13C NMR (125 MHz, DMSO-d6):
d
21.1, 27.8, 31.0,
indole-5,30-indoline]-20,6,11-trione
(14d). White
251e252 ꢁC; IR (cmꢀ1): 3416, 2932, 2359, 1702, 1602, 1472, 1342,
1119, 753; 1H NMR (500 MHz, DMSO-d6):
solid;
mp
47.8, 70.7, 77.1, 110.4, 126.2, 126.3, 126.4, 127.6, 130.6, 130.7, 132.7,
133.1, 134.7, 134.8, 140.8, 146.2, 152.4, 177.7, 181.5, 182.7; MS m/
z ¼ 371 [MH]þ; Anal. Calcd for C23H18N2O3: C, 74.58; H, 4.90; N,
7.56. Found: C, 74.49; H, 4.82; N, 7.68.
d
1.32 (t, 3H, J ¼ 6.9 Hz),
1.72e1.84 (m, 3H), 2.15e2.24 (m, 1H), 2.47e2.54 (m, 1H), 2.58e2.64
(m,1H), 3.39 (q, 3H, J ¼ 6.9 Hz), 4.74 (t,1H, J ¼ 7.6 Hz), 6.82e6.91 (m,
2H), 7.11 (d, 1H, J ¼ 7.6 Hz), 7.25 (t,1H, J ¼ 7.6 Hz), 7.74e7.81 (m, 3H),
5.1.2.3. 10-Methyl-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo[2,1-a]
isoindole-5,30-indoline]-20,6,11-trione (14c). White solid; mp
268e269 ꢁC; IR (cmꢀ1): 3413, 2934, 2356, 1708, 1609, 1474, 1335,
8.02 (d, 1H, J ¼ 7.6 Hz); 13C NMR (125 MHz, DMSO-d6):
d 11.1, 26.8,
27.8, 31.0, 39.3, 47.7, 70.6, 76.7, 109.5, 122.4, 125.4, 126.2, 126.6,
126.7, 130.6, 132.6, 133.1, 134.7, 134.9, 144.5, 146.1, 152.5, 176.1, 181.3,
Fig. 4. Comparison of MIC (mg/ml) values of synthesised compounds and standard drugs.

90
G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
182.6; MS m/z ¼ 385 [MH]þ; Anal. Calcd for C24H20N2O3: C, 74.98;
Completion of the reaction was evidenced by TLC analysis. After
completion of the reaction, the reaction mixture was poured into
ice-water, the resulting solid was filtered off and purified by column
chromatography using ethyl acetate: petroleum ether (3:7) as an
eluent to afford pure spirooxindole (17).
H, 5.24; N, 7.29. Found: C, 74.79; H, 5.34; N, 7.37.
5.1.2.5. 10-(Prop-2-ynyl)-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo
[2,1-a]isoindole-5,30-indoline]-20,6,11-trione (14e). White solid; mp
231e232 ꢁC; IR (cmꢀ1): 3778, 3433, 3246, 2892, 2365, 1724, 1664,
1605, 1468, 1341, 1180, 731, 486; 1H NMR (500 MHz, DMSO-d6):
5.1.3.1. 20-Methyl-20,30-dihydro-2H-spiro[acenaphthylene-1,10-benzo
[f]isoindole]-2,40,90-trione (17). Brown solid; mp 274e275 ꢁC; IR
(cmꢀ1): 3757, 3023, 2957, 2743, 1708, 1684, 1593, 1456, 1267, 1156,
d
1.72e1.79 (m, 1H), 1.82e1.89 (m, 2H), 2.15e2.23 (m, 1H),
2.42e2.45 (m, 1H), 2.53e2.55 (m, 1H), 3.27 (m, 1H), 4.53 (ABq, 2H,
J ¼ 17.5), 4.76 (t, 1H, J ¼ 7.7 Hz), 7.01 (t, 1H, J ¼ 7.6 Hz), 7.16 (d, 1H,
J ¼ 7.6 Hz), 7.20 (d, 1H, J ¼ 7.6 Hz), 7.38 (t, 1H, J ¼ 7.6 Hz), 7.74e7.81
(m, 2H) 7.84 (t, 1H, J ¼ 7.6 Hz), 8.01 (d, 1H, J ¼ 7.6 Hz); 13C NMR
972, 778, 698, 581; 1H NMR (500 MHz, DMSO-d6):
d 2.27 (s, 3H),
4.29 (ABq, 2H, J ¼ 16.8 Hz), 7.25e7.79 (m, 5H), 7.82 (t,1H, J ¼ 6.8 Hz),
7.99 (d, 1H, J ¼ 6.8 Hz), 8.01e8.12 (m, 3H); 13C NMR (125 MHz,
(125 MHz, DMSO-d6):
d
27.8, 29.6, 30.8, 47.8, 70.8, 75.1, 76.6, 78.3,
DMSO-d6): d 35.2, 58.8, 78.7, 123.6, 123.9, 126.1, 126.3, 128.4, 130.7,
110.3, 122.8, 125.2, 126.3, 126.5, 127.0, 130.6, 132.6, 133.2, 134.8,
134.9, 142.8, 145.6, 152.8, 175.3, 181.3, 182.6; MS m/z ¼ 395 [MH]þ;
Anal. Calcd for C25H18N2O3: C, 76.13; H, 4.60; N, 7.10. Found: C,
76.24; H, 4.51; N, 7.02.
131.9, 132.1, 133.4, 134.5, 135.3, 136.4, 138.3, 138.8, 140.1, 141.6, 146.7,
152.9, 181.0, 182.3, 194.4; MS m/z ¼ 366 [MH]þ; Anal. Calcd for
C24H15NO3: C, 78.89; H, 4.14; N, 3.83. Found: C, 78.79; H, 4.27;
N, 3.92.
5.1.2.6. 10-Allyl-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo[2,1-a]iso-
5.1.4. Experimental procedure for the synthesis of spirooxindole
derivative (18)
indole-5,30-indoline]-20,6,11-trione
(14f). White
204e205 ꢁC; IR (cmꢀ1): 3421, 2885, 1711, 1608, 1466, 1341, 1164,
962, 729, 481; 1H NMR (500 MHz, DMSO-d6):
1.73e1.79 (m, 1H),
solid;
mp
A mixture of isatin 8a (1 mmol), sarcosine 9 (1 mmol) and
ninhydrin 16 (1 mmol) was refluxed in ethanol (5 ml). Completion
of the reaction was evidenced by TLC analysis. After completion of
the reaction, the reaction mixture was poured into ice-water, the
resulting solid was filtered off and purified by column chroma-
tography using ethyl acetate: petroleum ether (3:7) as an eluent to
afford pure spirooxindole (18).
d
1.82e1.90 (m, 2H), 2.16e2.24 (1H, m), 2.48e2.52 (m, 1H), 2.55e2.62
(m, 1H), 4.31 (2H, m), 4.77 (t, 1H, J ¼ 7.6 Hz), 5.16 (d, 1H, J ¼ 10.7 Hz),
5.27 (d, 1H, J ¼ 16.0 Hz), 5.80 (m, 1H), 6.94e7.00 (m, 2H), 7.18 (d, 1H,
J ¼ 7.6 Hz), 7.31 (t, 1H, J ¼ 7.9 Hz), 7.76e7.80 (m, 2H), 7.81e7.87 (m,
1H), 8.02 (d, 1H, J ¼ 7.6 Hz); 13C NMR (125 MHz, DMSO-d6):
d 27.8,
30.9, 42.1, 47.7, 70.8, 76.6, 110.2, 116.8, 122.4, 125.3, 126.4, 126.5,
126.9, 130.5, 132.0, 132.6, 133.2, 134.8, 134.9, 143.7, 145.8, 152.8,
5.1.4.1. 2-Methyl-2,3-dihydrospiro[benzo[f]isoindole-1,20-indene]-
10,30,4,9-tetraone (18). Brown solid; mp 260e261 ꢁC; IR (cmꢀ1):
3766, 3023, 2943, 2743, 1711, 1656, 1573, 1464, 1237, 1156, 969,
175.9, 181.5, 182.6; MS m/z
¼
397 [MH]þ; Anal. Calcd for
C25H20N2O3: C, 75.74; H, 5.08; N, 7.07. Found: C, 75.62; H, 5.18;
N, 7.15.
778, 698, 618, 560, 469; 1H NMR (500 MHz, DMSO-d6):
d 2.29 (s,
3H), 3.89 (s, 1H), 4.28 (s, 2H), 7.22e7.78 (m, 2H), 7.83 (t, 1H,
5.1.2.7. 10-Benzyl-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo[2,1-a]
isoindole-5,30-indoline]-20,6,11-trione (14g). White solid; mp
241e242 ꢁC. IR (cmꢀ1): 3413, 2934, 2356, 1708, 1609, 1474, 1335,
J ¼ 6.9 Hz), 7.93 (d, 1H, J ¼ 6.7 Hz), 8.01e8.12 (m, 4H); 13C NMR
(125 MHz, DMSO-d6):
d 35.7, 59.3, 79.8, 123.3, 124.1, 126.5, 126.8,
131.0, 131.8, 132.8, 134.9, 135.3, 137.3, 138.0, 141.3, 146.2, 152.3,
178.5, 181.0, 182.3, 198.4; MS m/z ¼ 344 [MH]þ; Anal. Calcd for
C21H13NO4: C, 73.46; H, 3.82; N, 4.08. Found: C, 73.55; H, 3.71;
N, 4.20.
1123, 1000, 744, 480; 1H NMR (500 MHz, DMSO-d6):
d 1.71e1.92
(3H, m), 2.14e2.26 (1H, m), 2.48e2.53 (m, 1H), 2.54e2.63 (m,
1H), 4.77e4.84 (m, 1H), 4.92 (s, 2H), 6.88 (d, 1H, J ¼ 6.9 Hz), 6.95 (t,
1H, J ¼ 7.6), 7.20 (d, 1H, J ¼ 6.9 Hz), 7.29e7.41 (m, 6H), 7.74e7.89 (m,
3H), 8.02 (d, 1H, J ¼ 6.9 Hz); 13C NMR (125 MHz, DMSO-d6):
d
27.8,
5.2. Biological assays
30.9, 43.3, 47.7, 70.9, 76.6, 110.3, 122.6, 125.4, 126.4, 126.5, 127.0,
127.4, 127.8, 129.1, 130.5, 132.7, 133.2, 134.8, 134.9, 136.6, 143.6,
145.7,152.9,176.4,181.5,182.6; MS m/z ¼ 447 [MH]þ; Anal. Calcd for
C29H22N2O3: C, 78.01; H, 4.97; N, 6.27. Found: C, 77.89; H, 4.87;
N, 6.37.
5.2.1. Materials and methods for the antimicrobial activity
Streptomycin and ciprofloxacin (Sigma) were used as positive
controls against bacteria. Fluconazole and ketoconazole (Himedia,
Mumbai) were used as positive controls against fungi.
5.1.2.8. 50-Chloro-1,2,3,11b-tetrahydrospiro[benzo[f]pyrrolo[2,1-a]
isoindole-5,30-indoline]-20,6,11-trione (14h). White solid; mp
253e254 ꢁC; IR (cmꢀ1): 3367, 3199, 2954, 2869, 2360, 1737, 1647,
1462, 1282, 1184, 809, 721, 556; 1H NMR (500 MHz, DMSO-d6):
5.2.2. Tested microbes
The following gram positive bacteria were used for the experi-
ments; S. aureus MTCC 96, S. aureus (MRSA), E. aerogens MTCC111,
M. luteus. The Gram negative bacteria included: P. vulgaris MTCC
1771, K. pneumonia MTCC 109, S. typhimurium MTCC1251, S. para-
typhi-B. Fungi M. pachydermatis, C. albicans MTCC 227 and B. cin-
erea. All cultures were obtained from the Department of
Microbiology, Christian Medical College, Vellore, Tamil Nadu, India.
d
1.73e1.80 (m, 1H), 1.82e1.90 (m, 2H), 2.13e2.20 (m, 1H),
2.49e2.53 (m, 1H), 2.55e2.62 (m, 1H), 4.74 (t, 1H, J ¼ 6.9 Hz), 6.89
(d, 1H, J ¼ 7.6 Hz), 7.26e7.33 (m, 2H), 7.76e7.86 (m, 3H), 8.01 (d, 1H,
J ¼ 6.8 Hz), 10.60 (brs, 1H, -NH, D2O exchangeable). 13C NMR
(125 MHz, DMSO-d6):
d 27.8, 30.6, 47.9, 70.9, 76.9, 112.0, 125.9,
126.4, 127.3, 128.0, 128.8, 130.3, 132.7, 133.2, 134.7, 134.8, 142.2,
145.3,152.8,177.5,181.6,182.6; MS m/z ¼ 391 [MH]þ; Anal. Calcd for
C22H15ClN2O3: C, 67.61; H, 3.87; N, 7.17. Found: C, 67.71; H, 3.94;
N, 7.09.
5.2.3. Preparation of inoculums
Bacterial inoculums were prepared by growing cells in Mueller
Hinton Broth (MHA) (Himedia) for 24 h at 37 ꢁC. These cell
suspensions were diluted with sterile MHB to provide initial cell
counts of about 104 CFU/ml. The filamentous fungi were grown on
sabouraud dextrose agar (SDA) slants at 28 ꢁC for 10 days and the
spores were collected using sterile doubled distilled water and
homogenized. Yeast was grown on sabouraud dextrose broth (SDB)
at 28 ꢁC for 48 h.
5.1.3. Experimental procedure for the synthesis of spirooxindole
derivative (17)
A mixture of isatin 8a (1 mmol), sarcosine 9 (1 mmol) and
acenaphthenequinone 15 (1 mmol) was refluxed in ethanol (5 ml).

G. Bhaskar et al. / European Journal of Medicinal Chemistry 51 (2012) 79e91
91
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One of the authors G. Bhaskar thanks the Council of Scientific
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Appendix. Supplementary data
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