Expeditious synthesis, antibacterial activity evaluation and GQSAR studies of 3-bisoxindoles, 2-oxindolyl-2-hydroxyindan-1,3-diones and 2-oxindolyl-2-hydroxyacenaphthylen-1-ones

Article abstract of DOI:10.1039/c2ra01215d

The rhodium(ii)-catalyzed one-pot three-component reaction of 3-diazooxindole, benzyl alcohol and isatin or ninhydrin or acenaphthenequinone was explored to synthesize 3-bisoxindoles, 2-oxindolyl-2-hydroxyindan-1,3-diones and 2-oxindolyl-2-hydroxyacenaphthylen-1-ones with vicinal stereo centres, respectively through oxonium ylides. Their antibacterial activities against Staphylococcus aureus NCIM 5021, Escherichia coli NCIM 2931, Pseudomonas aeruginosa NCIM 5029, Salmonella typhimurium NCIM 2501 and Proteus vulgaris NCIM 2813 were evaluated by a microdilution method. Group-based quantitative structure activity relationships (GQSAR) for all antibacterial activities were developed. The statistical measures namely r² (0.66-0.85), r ² adj (0.63-0.84), q² (0.54-0.79) and F-ratio (11.63-34.17) were found to be satisfactory. All of the test and training set compounds were found to be within a 99% confidence level. Descriptors pertaining to hydrophobicity, topology and electrostatic interactions of the substitutions were found to contribute to their activities.

Full text of DOI:10.1039/c2ra01215d

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Expeditious synthesis, antibacterial activity evaluation
and GQSAR studies of 3-bisoxindoles, 2-oxindolyl-2-
hydroxyindan-1,3-diones and 2-oxindolyl-2-
hydroxyacenaphthylen-1-ones3
Cite this: DOI: 10.1039/c2ra01215d
Neelakandan Vidhya Lakshmi,a Ponnurengam Malliappan Sivakumar,b
Doraiswamy Muralidharan,a Mukesh Dobleb and Paramasivan T. Perumal*a
The rhodium(II)-catalyzed one-pot three-component reaction of 3-diazooxindole, benzyl alcohol and isatin
or ninhydrin or acenaphthenequinone was explored to synthesize 3-bisoxindoles, 2-oxindolyl-2-
hydroxyindan-1,3-diones and 2-oxindolyl-2-hydroxyacenaphthylen-1-ones with vicinal stereo centres,
respectively through oxonium ylides. Their antibacterial activities against Staphylococcus aureus NCIM
5021, Escherichia coli NCIM 2931, Pseudomonas aeruginosa NCIM 5029, Salmonella typhimurium NCIM
2501 and Proteus vulgaris NCIM 2813 were evaluated by a microdilution method. Group-based
quantitative structure activity relationships (GQSAR) for all antibacterial activities were developed. The
statistical measures namely r2 (0.66–0.85), r2 adj (0.63–0.84), q2 (0.54–0.79) and F-ratio (11.63–34.17) were
found to be satisfactory. All of the test and training set compounds were found to be within a 99%
confidence level. Descriptors pertaining to hydrophobicity, topology and electrostatic interactions of the
substitutions were found to contribute to their activities.
Received 1st December 2011,
Accepted 4th November 2012
DOI: 10.1039/c2ra01215d
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TMC-95s,3
Welwitindolinone
C,4
Celogentin
K,5
Introduction
Convolutamydines6 and SM-130686.7 There are many methods
known to synthesize 3-substituted 3-hydroxyindolin-2-ones via
nucleophilic addition to isatins, which generate only one
stereocentre.8
Multicomponent reactions (MCRs) are flexible, selective,
convergent and atom-efficient processes that facilitate the
construction of complex molecules in a single step.1 Oxindoles
incorporated with a quaternary stereogenic centre at C3 are
attractive targets in organic synthesis since they display a
fascinating array of biological applications.2 Oxindoles have
proven anti-infective properties. Strigacova et al. tested the
antifungal activity of these compounds against filamentous
fungi like R. oryzae, B. cinerea, F. moniliforme, A. flavus, T.
interdigitale, M. gypseurn.2c Chande et al. reported the
antitubercular activity of spirooxindoles against M. tuberculosis
H37Rv strain.2d They also possess antibacterial activity, which
was explored by several researchers.2e–h Moreover oxindoles
exhibit a plethora of activities, such as anticancer, anti-
protozoal, anti-inflammatory, spermicidal activities, and also
act against progesterone receptors.2i–2l Among the oxindole
derivatives, the 3-substituted 3-hydroxyindolin-2-one nucleus
represents an important structural motif present in an
extensive number of biologically active alkaloids, such as
Many studies of transition metal-catalyzed decomposition
of diazocarbonyl compounds have focused on cyclopropana-
tion, X–H (X = C, N, O, Si, S) insertion and ylide generation,9–11
and these reactions have also been used to prepare several
alkaloids, such as Aspidosperma, Kopsifoline and
Aspidophytine.12 The chemistry of oxonium ylides is an area
of continuing interest due to its wide synthetic utility.13–15
Several methods were known for the in situ generation of
oxonium ylides from rhodium carbenoids and for their
synthetic uses.14–16 To the best of our knowledge, this is the
first time 3-diazooxindole has been used for the one-pot
multicomponent synthesis of 3-bisoxindoles, which contain a
3-hydroxyindoline-2-one moiety with contiguous stereo carbon
centres with rhodium(II) acetate as a catalyst via in situ
generation and trapping of oxonium ylides. We herein disclose
a
method for the generation of oxonium ylides from
aOrganic Chemistry Division, Central Leather Research Institute (CSIR), Adyar,
Chennai-600 020, India. E-mail: ptperumal@gmail.com; Tel: +91 44 24913289
bDepartment of Biotechnology, Indian Institute of Technology Madras, Adyar,
Chennai-600036, India
3-diazooxindole and benzyl alcohols via rhodium carbenoids,
which undergo nucleophilic addition to isatin, ninhydrin and
acenaphthenequinone to synthesize 3-bisoxindoles, which
contain a 3-hydroxyindoline-2-one moiety, 2-oxindolyl-2-hydro-
xyindan-1,3-dione derivatives and 2-oxindolyl-2-hydroxyace-
Electronic supplementary information (ESI) available. See DOI: 10.1039/
3
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Encouraged by the results obtained in the above reaction,
we were further interested to carry out the reactions of
3-diazooxindole 1 with various substituted benzyl alcohols 3a–
e and isatin derivatives 5a–f. All of the reactions proceeded
smoothly in 90–100 min to give the respective 3-bisoxindoles
6a–j in good yields (82–90%). The reactions were diastereose-
lective, as only one isomer was obtained in all of the reactions
(Table 1).
Based on the above results,
a tentative mechanistic
interpretation to explain the formation of the product 6 is
proposed13,18,19 (Scheme 2). The reaction can be considered to
proceed via the initial formation of rhodium carbenoid 2,
which reacts with benzyl alcohol 3 to generate oxonium ylide
4. The oxonium ylide 4 generated is trapped by an electrophile,
such as isatin 5, to afford the product 6.
The O–H insertion side product 2a from 1 and 3 was not
formed due to the high electrophilicity of isatin 5.18,19 The
high electrophilicity of the carbonyl group of isatin 5 accounts
for the nucleophilic addition of oxonium ylide 4 to isatin 5.
The structures of products 6a–j were confirmed by spectral
studies and elemental analysis. The 1H NMR spectrum of
compound 6b exhibited characteristic singlets at d 6.16, 10.16
and 10.44 ppm for hydroxy proton and –NH protons (D2O
exchangeable) of two oxindole rings, respectively. The peaks at
d 10.16 and 10.44 ppm confirm the incorporation of two
oxindole rings in the structure. The 13C NMR spectral analysis
of compound 6b showed peaks at d 173.9 and 175.1 ppm for
the presence of amide carbonyl groups of two oxindole rings.
The mass spectrum of the compound displayed the molecular
ion peak at m/z 416.87. The stereochemistry of the products
6a–j were established through single-crystal X-ray analysis of
the compound 6a, which clearly illustrated the stereochemistry
in Fig. 1.20
Scheme 1 Synthesis of 3-bisoxindoles (6a–j).
naphthylen-1-ones with vicinal stereocentres, respectively.
Their antibacterial activity against five of the Gram +ve,
namely Staphylococcus aureus and Gram –ve bacterial strains,
namely Escherichia coli, Pseudomonas aeruginosa, Salmonella
typhimurium and Proteus vulgaris were studied.
Results and discussion
We started our initial study with rhodium(II) acetate (1 mol%)
catalyzed three component reaction of 3-diazooxindole 1,17
benzyl alcohol 3a and isatin 5a in dichloromethane, which
afforded highly functionalized bisoxindole 6a with adjacent
stereocentres in just a single step. Optimum yields were
obtained by slow addition of 3-diazooxindole 1 to the refluxing
solution of isatin 5a, benzyl alcohol 3a and rhodium(II) acetate
in dry dichloromethane under argon atmosphere. The solid
precipitated from the reaction was filtered and recrystallized
from ethanol to furnish the compound 6a as a single product.
The synthetic protocol is remarkably simple and requires no
further purification processes like column chromatography
(Scheme 1).
The promising results with isatin derivatives prompted us
to explore further with ninhydrin 7 and acenaphthenequinone
9 under the same optimized conditions to provide 2-oxindolyl-
2-hydroxyindan-1,3-diones 8a–e and 2-oxindolyl-2- hydroxyace-
naphthylen-1-ones 10a–e, respectively (Scheme
3
and
Scheme 4). The results are summarized in Table 2 and Table 3.
1
The H NMR spectrum of 8d exhibited a broad singlet at d
6.51 due to the –OH proton and signal at d 10.56 ppm for –NH
proton (D2O exchangeable).
Table 1 Synthesis of 3-bisoxindole deivatives 6a–j
Entry
Isatin (5)
R
R1
Alcohol (3)
R2
Product (6)a
Time (min)
Yield (%)b
1
2
3
4
5
6
7
8
9
5a
5a
5a
5a
5a
5b
5c
5d
5e
5f
H
H
H
H
H
Cl
Br
I
H
H
H
H
H
H
H
H
3a
3b
3c
3d
3e
3a
3a
3a
3a
3a
H
6a
6b
6c
6d
6e
6f
6g
6h
6i
90
90
90
100
90
90
90
90
90
90
87
90
89
87
83
83
82
82
87
86
OMe
Me
Cl
Br
H
H
H
H
H
allyl
methyl
H
H
10
6j
a
b
The products were characterized by IR, NMR, mass and elemental analysis. Isolated yield after recrystallization.
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Scheme 3 Synthesis of 2-oxindolyl-2-hydroxy-indan-1,3-diones 8a–e.
Scheme 2 A plausible mechanism for the synthesis of 3-bisoxindoles 6a–j.
the electron withdrawing substitutions present in the indoline
derivatives showed good antibacterial and antifungal activ-
ities.21 The electron withdrawing substituents reduce the
lipophilicity of the compound and make them soluble in
water. Thus, the bioavailability and antibacterial activity of the
compounds are increased. A bacterial cell membrane is also
comprised of lipopolysaccharide, which is hydrophilic in
nature, which further helps the penetration of these polar
compounds through this membrane. Other researchers and
research performed by us also found that polar substitutions
in compounds enhance the anti-infective activity.22–24
Moreover, hydrophilic/lipophilic balance (log P) of the
compounds plays an important role in the absorption,
distribution, metabolism and excretion of the compounds.
Resonances at d 173.2, 196.9 and 197.9 ppm (three carbonyl
groups) were observed in 13C NMR spectrum. The mass
spectrum of 8d displayed the molecular ion (M+ + H+) peak at
m/z 434.6.
Biological evaluation
Various oxindoles synthesized with different substitutions in
the R1 and R2 position, and their antibacterial activities in
micromolar concentrations against five of the organisms by
microdilution broth assay method21 are shown in Table 4. In
general compounds 6b, 6d and 6e are found to be more active,
while 6a, 6c and 10e are found to be less active than other
compounds. The most active compounds have an oxindole
group in their R1 position. Introduction of substitutions in the
oxindole ring (compounds 6f–6j) leads to a lowering of activity.
The replacement of an oxindole derivative with indan-1,3-
dione derivative (8a–8e) and acenaphthylen-1-ones (10a–10e)
leads to a reduction in the antibacterial activity. The least
active compounds, 6a and 6c, also have an oxindole group at
their R1 position, but the substitution at R2 seems to
determine the activity within 3-bisoxindoles. The highly active
compounds, 6b, 6d, 6e, have –OMe, –Cl and –Br substitution
(polar groups) in their structure, which belong to the electron
withdrawing substituents. Earlier researchers also found that
GQSAR studies
GQSAR is an ingenius method, which relates the biological
response variation to the molecular fragments of interest.
GQSAR also performs based on conformation and alignment,
which is absent in 3D QSAR, and it provides a site specific clue
for designing new chemical entities. It outperforms the 2D
QSAR where whole molecules are taken to consider the
biological activity variation, but not the molecular fragments.
Thus, the contribution of chemical variations against the
biological activity with common lead molecules are less
informative. Moreover the descriptor calculation is fast, as
well as allowing ease of interpretation of the results.
Fig. 1 ORTEP diagram of compound 6a.
Scheme 4 Synthesis of 2-oxindolyl-2-hydroxyacenapthylen-1-ones 10a–e.
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antibacterial activities against Escherichia coli for training and
test set compounds. All of the predictions lie within the 99%
confidence limits. Table 5 lists the numerical values of the
descriptors used in the training and test set of Escherichia coli.
In the case of Pseudomonas aeruginosa, compounds 6b, 6d
and 6e are the most active and compounds 6a, 6c and 10e are
the least active. The GQSAR equation for Pseudomonas
aeruginosa and the corresponding statistics are given below:
Table 2 Synthesis of 2-oxindolyl-2-hydroxyindan-1,3-diones 8a–e
Entry Alcohol (3) R2
Product (8)a Time (min) Yield (%)b
1
2
3
4
5
3a
3b
3c
3d
3e
H
8a
70
70
70
80
80
85
88
87
85
80
OMe 8b
Me
Cl
8c
8d
8e
Br
Activity
=
21.1765–1.1353(¡0.2523)*R1-QMDipoleY
+
a
The products were characterized by IR, NMR, mass and elemental
b
0.1912(¡0.0195)* R1-4PathCount
analysis. Isolated yield after recrystallization.
n = 15, r2 = 0.74, r2 adj = 0.72, q2 = 0.64, F-test = 17.27, r2 se =
0.22, q2 se = 0.27
R1-QMDipoleY descriptor contributes negatively to the
activity. As mentioned before, hydrophobic substitution at
the R1 position favours activity. R1-4PathCount belongs to the
topological descriptor and shows a positive contribution
towards the activity. It indicates the total number of fragments
of fourth order (four bond path) in a compound.
This group of descriptors describes the connectivity of the
atoms within the molecule and also explains its branching and
flexibility (or rigidity).29 An increase in branching increases the
hydrophobicity.30 Khan et al. also showed the positive
contribution of hydrophobicity of the molecule with the
antibacterial activity against Pseudomonas aeruginosa.31 Fig. 3
shows the parity plot between the observed and the predicted
antibacterial activities against Pseudomonas aeruginosa for
training and test set compounds. All of the predictions lie
within the 99% confidence limits. Table 6 lists the numerical
values of the descriptors used in the GQSAR.
In the case of Escherichia coli, compounds 6b, 6d and 6e are
the most active and 6a, 6c and 10e are the least active. The
GQSAR equation developed for E. coli and the corresponding
statistics are given below:
Activity
=
20.0228–0.4653(¡0.0227)*R1-QMDipoleY
+
54.9364(¡10.5006)*R1-Average +ve potential
n = 15, r2 = 0.76, r2 adj = 0.74, q2 = 0.69, F-test = 18.83, r2 se =
0.1164, q2 se = 0.131
The equation consists of two descriptors, namely
QMdipoleY and Average +ve potential. QMdipoleY contributes
negatively to the activity. This descriptor deals with the
strength and orientation of a molecule in an electrostatic
field and is a function of the polarity. So less polar compounds
will have higher activity. Similar results have been reported by
Tantitame et al., that less polar diastereomers exert better
antibacterial activity when compared to the more polar
diastereomers.25 Other researchers have also found the
contribution of the descriptor, QMDipoleZ, towards the
antibacterial activity.26 Average +ve potential descriptor
belongs to the electrostatic subclass and it is defined as the
average of the total +ve electrostatic potential on van der Waals
surface area of the molecule and it showed a positive
contribution towards the activity. The membranes of
Escherichia coli contain negatively charged phosphate groups,
which can attract more of the positively charged molecules.27
So the positively charged molecules can bind easily to the wall
of Escherichia coli and exhibit higher activity than negatively
charged molecules. Due to the positive contribution of the
average +ve potential and hydrophobicity of the molecules,
this group of compounds are called hydrophobic cations,
which have good permeability to the cell and organelles
reported by Lewis.28 Both the descriptors relate to the
properties of the functional groups in the R1 position. Fig. 2
shows the parity plot between the observed and the predicted
In the case of Proteus vulgaris, compounds 6b, 6d and 6e are
the most active and compounds 6a, 6c and 6f are the least
active. The GQSAR equation for Proteus vulgaris and the
corresponding statistics are given below:
Activity
=
21.2257–1.0943(¡0.1655)*R1-QMDipoleY
+
5.3711(¡1.3019)*R1-chiV5
n = 15, r2 = 0.66, r2 adj = 0.63, q2 = 0.54, F-test = 11.63, r2 se =
0.24, q2 se = 0.28
Once again we find R1-QMDipoleY as one of the descriptors
in the equation. ChiV5 belongs to the topological descriptor in
the sub-class of atomic valence connectivity index. It shows a
positive contribution towards the activity, which indicates that
hydrophobic substitution at R1 position favours activity. Fig. 4
shows the parity plot between the observed and the predicted
antibacterial activities against Proteus vulgaris for training and
test set compounds.
All of the predictions lie within the 99% confidence limits.
Table 7 lists the numerical values of the descriptors.
In the case of Salmonella typhimurium, compounds 6b, 6d,
6e and 8b are the most active and compounds 6a, 6c and 8e are
the least active. The GQSAR equation for Salmonella typhimur-
ium and the corresponding statistics are given below:
Table 3 Synthesis of 2-oxindolyl-2-hydroxyacenaphthylen-1-ones 10a–e
0.4156–0.7612(¡0.1424)*R1-QMDipoleY
+
Entry Alcohol (3) R2
Product (10)a Time (min) Yield (%)b
Activity
=
0.1119(¡0.0151)*R1-SssOE-index
1
2
3
4
5
3a
3b
3c
3d
3e
H
10a
75
75
75
80
80
81
84
82
81
83
N = 15, r2 = 0.85, r2 adj = 0.84, q2 = 0.79, F-test = 34.17, r2 se =
0.13, q2 se = 0.15
OMe 10b
Me
Cl
10c
10d
10e
R1-SssOE-index is the electrotopological index and is
defined as the number of oxygen atoms connected with two
single bonds.32 Among the active compounds, 6b and 8b have
Br
a
The products were characterized by IR, NMR, mass and elemental
b
analysis. Isolated yield after recrystallization.
a
SssOE-index of 4.19, which is relatively high (other
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Table 4 Oxindoles synthesized with different substitutions and their antibacterial activities
Substitutions used in GQSAR
R1 R2
Antibacterial activity (MIC in mM)
Compound no.
S. aureus
P. aeruginosa
E. coli
P. vulgaris
S. typhi
6a
0.647
1.294
0.323
1.294
0.647
6b
6c
0.075
1.249
0.038
1.249
0.038
0.312
0.075
0.038
0.624
2.497
6d
6e
0.149
0.067
0.074
0.067
0.074
0.034
0.149
0.067
0.074
0.034
6f
6g
6h
0.594
0.537
0.488
0.594
0.537
0.976
0.149
0.134
0.244
1.188
0.537
0.976
0.297
0.269
0.244
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Table 4 (Continued)
Substitutions used in GQSAR
R1 R2
Antibacterial activity (MIC in mM)
Compound no.
S. aureus
P. aeruginosa
E. coli
P. vulgaris
S. typhi
6i
0.586
0.586
0.147
0.586
0.293
6j
0.624
0.313
0.624
0.626
0.156
0.156
0.624
0.313
0.312
0.313
8a
8b
8c
0.291
0.605
0.291
0.605
0.146
0.076
0.291
0.605
0.146
0.302
8d
8e
0.288
0.523
0.288
0.523
0.144
0.131
0.288
0.523
0.288
0.523
10a
10b
1.186
0.277
1.186
0.554
0.148
0.138
1.186
1.107
0.297
0.277
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Table 4 (Continued)
Substitutions used in GQSAR
R1 R2
Antibacterial activity (MIC in mM)
Compound no.
S. aureus
P. aeruginosa
E. coli
P. vulgaris
S. typhi
10c
0.287
0.574
0.144
0.574
0.144
10d
10e
0.548
0.999
1.097
1.999
0.137
0.250
1.097
0.999
0.548
0.500
Chi5 belongs to the class of topological descriptors and it
describes the connectivity of the molecule. This descriptor
showed a positive contribution to the activity. An increase in
the connectivity will increase the hydrophobicity of the
molecule. Fig. 6 shows the parity plot between the observed
and the predicted antibacterial activities against Staphy-
lococcus aureus for training and test set compounds. All of
the predictions lie within the 99% confidence limits. Table 9
lists the numerical values of the descriptors used in the
training set and test set of Staphylococcus aureus.
Table 10 shows the percentage contribution of the descriptors
used in GQSARs for all of the microorganisms. A dipole of the
substituent group in the R1 position appears to contribute
negatively in all of the regression equations. Our previous research
also showed the contribution of dipole moment towards the
antibacterial activity of isoxazolidine derivatives against Staphylo-
coccus aureus, Escherichia coli and Pseudomonas aeruginosa.33
This method offers several advantages, such as a high yield
and simple experimental and isolation procedures, making it
an efficient route to the synthesis of 3-bisoxindoles, 2-oxindo-
lyl-2-hydroxyindan-1,3-diones and 2-oxindolyl-2-hydroxyace-
naphthylen-1-ones, which are important compounds in
organic and medicinal chemistry.
compounds has zero) compared to other compounds except
compound 10b. The latter also had a SssOE-index as 4.19 and
it showed moderate activity. Fig. 5 shows the parity plot
between the observed and the predicted antibacterial activities
against Salmonella typhimurium for training and test set
compounds. All of the predictions lie within the 99%
confidence limits. Table 8 lists the numerical values of these
descriptors.
In the case of Staphylococcus aureus, compounds 6b, 6d and
6e are the most active and 6c, 10a and 10e are the least active.
The GQSAR equation for Staphylococcus aureus and its
corresponding statistics are given below:
Activity
=
21.0378–0.6124(¡0.1928)*R1-QMDipoleY
+
1.5986(¡0.3018)*R1-chi5
N = 15, r2 = 0.73, r2 adj = 0.71, q2 = 0.62, F-test = 16.37, r2 se =
0.17, q2 se = 0.20
Conclusion
In conclusion, we have demonstrated the first example of
rhodium(II)-catalyzed one-pot, multicomponent reaction of 3-dia-
zooxindole, benzyl alcohols and isatin or ninhydrin or ace-
naphthenequinone to synthesize 3-bisoxindoles, 2-oxindolyl-2-
hydroxyindan-1,3-diones and 2-oxindolyl-2-hydroxyacenaphthy-
len-1-ones, respectively with adjacent tetra-substituted carbon
Fig. 2 A parity plot between the observed and the predicted antibacterial
activities against Escherichia coli for training and test set compounds with 99%
prediction bands (closed circles – training set; open circle – test set).
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Table 5 The numerical values of the observed and predicted activity and descriptors used in GQSAR for Escherichia coli
Compound
Observed activity
R1-QMDipoleY
R1-Average +ve potential
Predicted activity
Training set
6a
6b
6c
6f
6g
6h
6j
8a
8b
8d
8e
10a
10b
10d
10e
Test set
6d
0.490
1.426
0.506
0.828
0.872
0.613
0.807
0.806
0.837
0.842
0.884
0.829
0.859
0.863
0.602
20.118
20.585
0.167
20.119
20.119
20.119
20.117
20.115
0.250
0.160
0.143
20.112
0.435
0.157
0.013
0.021
0.011
0.013
0.013
0.013
0.013
0.013
0.018
0.017
0.016
0.013
0.020
0.017
0.016
0.745
1.428
0.500
0.747
0.746
0.746
0.751
0.749
0.840
0.827
0.782
0.742
0.852
0.828
0.780
0.141
1.129
1.474
0.834
1.122
0.843
0.164
0.148
20.118
0.164
0.017
0.016
0.013
0.011
0.011
0.825
0.779
0.753
0.505
0.508
6e
6i
8c
10c
0.157
centres. We suggest that the reaction occurs through a metal-
associated alcoholic oxonium ylide intermediate and is followed
by a nucleophilic addition to electrophiles, such as isatin,
ninhydrin and acenaphthenequinone. The high electrophilicity
and rigid cyclic conformation of isatin, ninhydrin and ace-
naphthenequinone are responsible for such a high chemo- and
diastereoselectivity. Antibacterial activities of the synthesised
oxindoles against five Gram +ve and Gram –ve organisms were
evaluated using microdilution method. Compounds 6b, 6d and
6e were found to be the most active against all of the five
organisms. Group-based QSAR has been developed for all five
antibacterial activities. The statistical parameters were found to
be in an accepted range. Among the groups in the R1 and R2
position, the descriptor on R1 is correlated with all of the
antibacterial activities, which suggests the importance of the R1
group. It shows that electrostatic (QMDipoleY, SssOE and average
+ve potential) (acts upon membrane integrity and permeability)
and, hydrophobic (chiV5, chi5 and 4PathCount) (for the penetra-
tion of the drug through the membrane and the retention of the
drug within bacterial cell) interactions are important contribu-
tions for the antibacterial activity. Other researchers have also
found the importance of electrostatic and lipophilicity towards
the antibacterial activities.34
Experimental section
General
All of the chemicals required for determining the biological
activities were purchased from Sigma-Aldrich (St Louis, MO,
USA), Himedia (Mumbai, India) and SRL (Mumbai, India) and
the five bacterial strains (S. aureus NCIM5021, E. coli NCIM
2931, P. vulgaris NCIM 2813, S. typhi NCIM 2501 and P.
aeruginosa NCIM 5029) were purchased from National
Chemical Laboratory, Pune, India. VLifemds3.5 is a software
package for computer aided drug discovery (VLife Sciences
Technologies Pvt. Ltd., Pune, India, http://www.vlifesciences.-
com) and it is used for molecular modelling. IR measurements
were done as KBr pellets for solids using a Perkin Elmer
Spectrum RXI FT-IR. 1H NMR (500 MHz) and 13C NMR (125
MHz) spectra were recorded as DMSO-d6 solution for all
compounds with TMS as an internal standard on a JEOL
instrument. Multiplicities are abbreviated as follows: singlet
(s), doublet (d), triplet (t), multiplet (m), and broad (br). The
mass of the compounds was analyzed using an Electrospray
Ionisation Method with Thermo Finnigan Mass spectrometer.
Melting points were determined in capillary tubes and are
uncorrected. Elemental analyses were recorded using
a
Thermo Finnigan FLASH EA 1112 CHN analyzer. Analytical
TLC was performed on precoated plastic sheets of silica gel
G/UV-254 of 0.2mm thickness (Macherey-Nagel). To facilitate
the QSAR, the observed activity (Minimum Inhibitory Concen-
tration against the three microorganisms calculated as mg
Fig. 3 A parity plot between the observed and the predicted antibacterial
activities against Pseudomonas aeruginosa for training and test set compounds
with 99% prediction bands (closed circles – training set; open circle – test set).
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Table 6 The numerical values of the observed and predicted activity and descriptors used in GQSAR for Pseudomonas aeruginosa
Compound
Observed activity
R1-QMDipoleY
R1-4PathCount
Predicted activity
Training set
6a
6b
6c
6f
6g
6h
6j
8a
8b
8d
8e
10a
10b
10d
10e
Test set
6d
20.112
1.426
20.096
0.226
0.270
0.011
0.205
0.203
0.536
0.540
0.282
20.074
0.257
20.040
20.301
20.118
20.585
0.167
20.119
20.119
20.119
20.117
20.115
0.250
0.160
0.143
20.112
0.435
0.157
6
10
6
6
6
6
6
6
10
8
8
6
10
8
8
0.105
1.400
20.219
0.106
0.106
0.106
0.104
0.102
0.452
0.172
0.191
0.098
0.242
0.175
0.193
0.141
1.129
1.173
0.232
0.218
0.241
0.164
0.148
20.118
0.164
8
8
6
6
6
0.167
0.186
0.105
20.215
20.207
6e
6i
8c
10c
0.157
ml21) was converted to 2log (mM). Spectral data is given in the
added and incubated for 2 h. The effect of ethanol on the
growth of the microorganism was also studied. The color
change was assessed visually, with the highest dilution
remaining blue (inhibition of growth) indicating the mini-
mum inhibitory concentration. A change in color from blue to
pink shows the growth of the organism.
supporting information .
3
Antibacterial evaluation
In vitro antibacterial activity (MIC) of the oxindole derivatives
were determined by microdilution broth assay method with
modifications, as reported by Sarker et al., using resazurin as
an indicator.35 Muller hinton broth was used to grow the
bacterial strains to a final inoculum size of 5 6 105 CFU
mL21. The compounds were dissolved in absolute ethanol to a
Modelling studies
Twenty compounds were synthesized, which have the oxindole
as the common template, while the functional groups at the R1
and R2 positions are different. The MIC (mM) data of oxindoles
concentration of 10 mg mL21 36
These serially diluted
.
solutions were added to successive wells in a 96-well microtiter
plate and incubated with the organisms for 18 h at 37 uC.
Growth and sterility controls were maintained during the
experiment. Compounds serially diluted with ethanol were
also kept in the 96-well microtiter plates (uninoculated
dilution) to determine if they precipitated out during the
course of the experiments. A blank array with ethanol alone
and its effect on the inhibition of the microorganism was also
studied. Ten microliters of 0.01% resazurin solution was
Table 7 The numerical values of the observed and predicted activity and
descriptors used in GQSAR for Proteus vulgaris
Observed
activity
Predicted
activity
Compound
R1-QMDipoleY
R1-chiV5
Training set
6j
8a
8b
8d
0.205
0.505
0.536
0.540
0.282
20.074
20.044
20.040
20.112
0.000
20.117
20.115
0.250
0.160
0.143
20.112
0.435
0.157
20.118
0.141
20.585
0.167
0.222
0.222
0.316
0.318
0.304
0.222
0.316
0.318
0.222
0.304
0.316
0.192
0.222
0.222
0.222
0.096
0.094
0.200
0.310
0.249
0.091
20.002
0.313
0.097
0.250
1.114
20.375
0.098
0.098
0.098
8e
10a
10b
10d
6a
10e
6b
6c
6f
6g
6h
1.125
20.397
20.075
0.270
20.119
20.119
20.119
0.011
Test set
8c
10c
6d
6e
0.218
0.241
0.828
1.173
0.232
0.164
0.157
0.164
0.148
20.118
0.192
0.192
0.318
0.304
0.222
20.371
20.363
0.305
0.243
0.097
Fig. 4 A parity plot between the observed and the predicted antibacterial
activities against Proteus vulgaris for training and test set compounds with 99%
prediction bands (closed circles – training set; open circle – test set).
6i
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Fig. 5 A parity plot between the observed and the predicted antibacterial
activities against Salmonella typhimurium for training and test set compounds
with 99% prediction bands (closed circles – training set; open circle – test set).
Fig. 6 A parity plot between the observed and the predicted antibacterial
activities against Staphylococcus aureus for training and test set compounds
with 99% prediction bands (closed circles – training set; open circle – test set).
listed in Table 4 was converted to 2log (mM) for the
development of the QSAR. Twenty of these oxindoles were
sketched using the Vlife mds 3.5 software and their minimum
energy conformations were determined using MMFF force
field.37 Three hundred and seventy four descriptors relating to
spatial, electronic, thermodynamic, conformational, topologi-
cal, information-content, quantum mechanical and structural
properties of the substituent groups in the R1 and R2 position
for all of the compounds were calculated. Several references
deal in detail with these descriptors.29,38,39 The total twenty
molecules were divided into training and test sets. The former
consists of fifteen molecules, which are used for the develop-
ment of the GQSAR equation and the latter consisting of five
molecules (compounds 6d, 6e, 6i, 8c and 10c), which were
selected at random were used for the validation of these
models. This technique is known as an external validation.
The developed GQSAR is also validated by calculating several
statistical parameters, such as r2, r2 adj, q2, F-ratio and
standard error.40 This procedure is known as the internal
validation method. A Genetic Algorithm technique (GFA) is
used to develop the best GQSAR equation relating the
descriptors with the activity. The GFA technique is useful
when one has a large pool of descriptors, but limited data,
where the challenge is to select the best set of descriptors.
Other researchers have also used the GFA technique effectively
to develop the QSAR models.41
Representative procedure for preparation of compounds 6a–j
To a refluxing dry CH2Cl2 solution of Rh2(OAc)4 (1 mol%),
benzyl alcohol 3 (0.60 mmol) and isatin 5 (0.50 mmol) under
argon atmosphere, 3-diazo-1,3-dihydro-indol-2-one 1 (0.60
mmol) in dry dichloromethane was added dropwise (approxi-
mately 10 ml in 60 min). After the addition was completed, the
reaction mixture was allowed to reflux for 30–40 min until the
disappearance of the starting materials, as determined by TLC,
and cooled to room temperature. The solid formed in the
reaction mixture was filtered, dried and recrystallized from
Table 8 The numerical values of the observed and predicted activity and descriptors used in GQSAR for Salmonella typhimurium
Compound
Observed activity
R1-QMDipoleY
R1-OxygensCount
Predicted activity
Training set
6j
8a
8b
8d
0.506
0.505
0.837
0.540
0.282
0.528
0.558
0.261
0.189
1.426
0.205
0.527
0.571
0.613
0.301
20.117
20.115
0.250
0.160
0.143
20.112
0.435
0.157
20.118
20.585
0.167
20.119
20.119
20.119
0.141
0
0
1
0
0
0
1
0
0
1
0
0
0
0
0
0.505
0.503
0.775
0.294
0.307
0.501
0.634
0.296
0.506
1.411
0.288
0.506
0.506
0.506
0.308
8e
10a
10b
10d
6a
6b
6c
6f
6g
6h
10e
Test set
6d
6e
6i
8c
10c
1.129
1.474
0.533
0.519
0.843
0.164
0.148
20.118
0.164
0
0
0
0
0
0.291
0.303
0.506
0.291
0.296
0.157
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was completed, the reaction mixture was allowed to reflux for
15–20 min until the disappearance of the starting materials, as
determined by TLC. The solid formed in the reaction mixture
was filtered, dried and recrystallized from ethanol to obtain
the pure product and the appropriate isolated yield is shown
in Table 3.
Table 9 The numerical values of the observed and predicted activity and
descriptors used in GQSAR for Staphylococcus aureus
Observed
activity
Predicted
activity
Compound
R1-chi5
R1-QMDipoleY
Training set
6a
6b
6c
6f
6g
6h
6j
8a
8b
0.189
1.125
20.096
0.226
0.270
0.312
0.205
0.505
0.536
0.540
0.282
20.074
0.558
0.261
0.000
0.75
1.11
0.61
0.75
0.75
0.75
0.75
0.75
1.11
0.90
0.90
0.75
1.11
0.90
0.90
20.12
20.58
0.17
20.12
20.12
20.12
20.12
20.12
0.25
0.16
0.14
20.11
0.43
0.16
0.234
1.087
20.161
0.234
0.234
0.234
0.233
0.232
0.576
0.305
0.315
0.230
0.463
0.306
0.316
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Representative procedure for the synthesis of 2-oxindolyl-2-
hydroxyindan-1,3-diones (8a–e)
To a refluxing dry CH2Cl2 solution of Rh2(OAc)4 (1 mol%),
benzyl alcohol 3 (0.60 mmol) and ninhydrin 7 (0.50 mmol)
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Table 10 The percentage contribution of the short-listed descriptors for developing GQSAR equation
Organism Name
Descriptor 1
% contribution of descriptor 1 Descriptor 2
% contribution of descriptor 2
Escherichia coli NCIM 2931
R1-QMDipoleY 240.72%
R1-Average+vePotential 59.28%
Pseudomonas aeruginosa NCIM 5029 R1-QMDipoleY 246.63%
R1-4PathCount
R1-chiV5
53.37%
50.39%
55.45%
62.60%
Proteus vulgaris NCIM 2813
Salmonella typhimurium NCIM 2501 R1-QMDipoleY 244.55%
Staphylococcus aureus NCIM5021
R1-QMDipoleY 237.40%
R1-QMDipoleY 249.61%
R1-SssOE
R1-chi5
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