Synthesis, antibacterial activity evaluation and QSAR studies of novel dispiropyrrolidines

Article abstract of DOI:10.1016/j.ejmech.2010.04.035

A series of novel dispiropyrrolidines have been synthesized through 1,3-dipolar cycloaddition of an azomethine ylide generated from sarcosine and isatin with the dipolarophile 3-benzylidene-1-methyl-pyrrolidine-2,5-dione. Their antibacterial activity was evaluated against Bacillus subtilis NCIM 2718,Staphylococcus aureus NCIM5021,Salmonella typhi NCIM2501,Pseudomonas aeruginosa NCIM 5029 and Proteus vulgaris NCIM2813 by two fold dilution method. Compound 6e exhibits reasonably good activity and compound 6c exhibits poor activity against all the organisms. The QSAR's were developed for all antibacterial activities. The models had either one or two descriptors (r ² 0.81-0.97,r_a²_dj 0:75-0:96,q ² 0.57-0.92,F-ratio 12.73-162.76). Topology,shape,charge distribution and hydrophobic nature of the molecules had pronounced effect on their antibacterial activity.

Full text of DOI:10.1016/j.ejmech.2010.04.035

European Journal of Medicinal Chemistry 45 (2010) 3446e3452
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis, antibacterial activity evaluation and QSAR studies
of novel dispiropyrrolidines
,
K. Karthikeyan a, P.M. Sivakumar b, M. Doble b, P.T. Perumal a
*
a Organic Chemistry Division, Central Leather Research Institute, Adyar, Chennai-600 020, India
b Department of Biotechnology, Indian Institute of Technology Madras, Chennai-600 036, 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 dispiropyrrolidines have been synthesized through 1,3-dipolar cycloaddition of an
azomethine ylide generated from sarcosine and isatin with the dipolarophile 3-benzylidene-1-methyl-
pyrrolidine-2,5-dione. Their antibacterial activity was evaluated against Bacillus subtilis NCIM 2718,
Staphylococcus aureus NCIM5021, Salmonella typhi NCIM2501, Pseudomonas aeruginosa NCIM 5029 and
Proteus vulgaris NCIM2813 by two fold dilution method. Compound 6e exhibits reasonably good activity
and compound 6c exhibits poor activity against all the organisms. The QSAR’s were developed for all
antibacterial activities. The models had either one or two descriptors (r2 ¼ 0.81e0.97, ra2dj ¼ 0:75e0:96,
q2 ¼ 0.57e0.92, F-ratio ¼ 12.73e162.76). Topology, shape, charge distribution and hydrophobic nature of
the molecules had pronounced effect on their antibacterial activity.
Received 10 February 2010
Received in revised form
28 April 2010
Accepted 30 April 2010
Available online 6 May 2010
Keywords:
Azomethine ylide cycloaddition
Dispiropyrrolidine
Antibacterial activity
QSAR studies
Ó 2010 Elsevier Masson SAS. All rights reserved.
1. Introduction
elevated levels of sorbitol [12] and behave as poliovirus and
rhinovirus 3C-proteinase inhibitors [13].
1,3-Dipolar cycloaddition reactions is an efficient method for the
construction of heterocyclic units in a highly regio- and stereo-
selective manner [1]. In particular, the chemistry of azomethine
ylides have gained significance in recent years as it serves as an
expedient precursor for the constructing nitrogen-containing five
membered heterocycles, which constitute the central skeleton of
numerous natural products [2]. The intermolecular [3 þ 2]-cyclo-
addition reaction of azomethine ylides with various alkenes
represents an efficient and convergent method for the synthesis of
pyrrolidine and pyrrolizidine units. Functionalised pyrrolidine
alkaloids constitute classes of compounds with significant biolog-
ical activities [3]. The 1,3-dipole (azomethine ylide) generated by
the decarboxylative condensation of isatin and secondary amino-
acid reacts with dipolarophiles containing an exocyclic double
bond to afford novel spiro pyrrolidine ring systems [4]. Spiro
compounds represent an important class of naturally occurring
substances characterized by their pronounced biological properties
[5e11]. The spirooxindole ring system forms the core structure of
many pharmacological agents and alkaloids. Spirooxindole deriv-
atives are found to be potent aldose reductase inhibitors (ARIs),
which help to treat and prevent diabetic complications arising from
Bacterial strains acquire resistance against commercial drugs
due to their regular usage and pose a threat to human kind.
Designing newer antibacterial agents to combat this resistance and
in the treatment of bacterial infections has forced researchers to
continuously search for new chemical entities with anti-infective
properties. Hence our studies have been focused towards the
synthesis and bio-evaluation of dispiropyrrolidine derivatives as
possible drug candidates.
2. Chemistry
In continuation of our studies in the area of cycloaddition
reactions [14] and with a view to synthesise of novel dis-
piroheterocyclic derivatives, we herein report the 1,3-dipolar
cycloaddition reaction of 3-benzylidene-1-methyl-pyrrolidine-
2,5-dione derivatives as 2
p components with various azomethine
ylides for the facile synthesis of biologically active dispiro
pyrrolidines.
In this reaction we prepared 3-benzylidene-1-methyl-pyrroli-
dine-2,5-dione (dipolarophile) 3aed from N-methyl maleimide
and various substituted benzaldehydes using PPh3 under neat
conditions at 70 ꢀC (Scheme 1, Table 1).
The 1,3-dipolar cycloaddition of azomethine ylide generated
insitu from the reaction of isatin 4aed and sarcosine 5 with
3-benzylidene-1-methyl-pyrrolidine-2,5-dione 3aed in methanol
* Corresponding author. Tel./fax: þ91 44 24913289.
E-mail address: ptperumal@gmail.com (P.T. Perumal).
0223-5234/$ e see front matter Ó 2010 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2010.04.035

K. Karthikeyan et al. / European Journal of Medicinal Chemistry 45 (2010) 3446e3452
3447
O
O
O
O
Ar
PPh3
Ar
+
+
N
CH NHCH COOH
ArCHO
+
O
N
3
2
N
Neat, 70 oC
N
R
5
O
O
2
O
1a-d
3a-d
3a-d
4a-d
reflux
MeOH
Hb
Scheme 1. Synthesis of 3-benzylidene-1-methyl-pyrrolidine-2,5-dione 3.
at reflux temperature yielded a series of dispiropyrrolidines 6aei in
good yield (Scheme 2, Table 2). The reaction afforded a single
product in all cases, as evidenced by thin layer chromatography
(TLC).
The structure of the dispiropyrrolidine derivatives was
confirmed by spectral analysis. IR spectrum of the pyrrolo-oxindole
derivative 6d, showed a peak at 1710 cmꢁ1 due to the carbonyl
group of the oxindole moiety. The 1H NMR spectrum exhibited
Hc
Ar
O
Ha
N
N
O
O
N
R
three triplet at
(J ¼ 9.2 Hz) were assigned to protons Ha, Hb and Hc of the pyrro-
lidine ring respectively. The 13C NMR, two peaks at
61.3 and
77.7 ppm corresponds to the two spiro carbons and the amide
d
3.39 (J ¼ 8.4 Hz), 3.74 (J ¼ 9.2 Hz) and 4.15
6a-i
Scheme 2. Synthesis of dispiropyrrolidines.
d
carbonyl carbons resonated at
d 174.4, 175.6 and 179.5 ppm. The
The QSAR equation developed for S. typhi and the statistics are
given below:
mass spectrum revealed the molecular ion peak (Mþ þ 1) at m/z
420. The structure was further confirmed through X-ray diffraction
studies [15] (Fig. 1).
Activity ¼ 0:8185 ꢁ 0:0492 ꢂ S ssO ꢁ 0:2135 ꢂ Jurs ꢁ RPCS
3. Biology
r ¼ 0:98; r2 ¼ 0:97; ra2dj ¼ 0:96; q2 ¼ 0:89;
F ꢁ ratio ¼ 86:43; error ¼ 0:03
Table 3 shows the observed antibacterial activities of dis-
piropyrrolidines given as micromolar concentration (mM) against
The biparametric equation consists of two descriptors namely,
S_ssO and Jurs-RPCS which are negatively correlated to activity.
Jurs-RPCS is a spatial descriptor and it is equal to the solvent
accessible area of the most positive atom divided by Jurs-RPCG,
where the later is calculated by considering the atomic partial
charges. It is a measure of the relative positive surface charge of the
molecule [17]. Outer membrane of the bacteria comprises of
glycolipid and lipopolysacchride (LPS), which are hydrophilic in
nature and it acts as a barrier for hydrophobic molecules. The inner
leaflet comprises of mixture of phospholipids, which are hydro-
phobic in nature and they limit the penetration of hydrophilic
compounds. Highly charged or polar low molecular weight
compounds will have more permeability through the outer
membrane than others [18]. Hence partial surface charge (Jurs-
RPCS) appears in the QSAR. Our earlier research showed the
contribution of other Jurs descriptors (Jurs-RPCG, Jurs-PPSA-3)
towards the antibacterial activity against S. typhi. [19e21].
five of the organisms evaluated by two fold dilution method.
Compound 6e is reasonably active against all the organisms and
compounds 6b, 6g and 6h are also active against majority of the
organisms and they have 2-OH or 4-Cl substitution in the aromatic
ring of the dipolarophile. Compound 6c, 6d and 6f with a 4-OCH3
substitution in the aromatic ring of the dipolarophile exhibited
poor activity. The highly active compounds have 2-OH or 4-Cl
substitution (polar groups) in their structure. Since these
compounds have more polar substitution, it increases the hydro-
philicity which increases their water solubility in the in vitro anti-
bacterial activity assay. Bacterial cell membrane also was
comprised of lipopolysaccharide which was hydrophilic in nature
which further helps in the penetration of these polar compounds
through this membrane. Other researchers and research performed
by us also found that polar substitution in compounds enhances the
anti-infective activity [16]. More over, hydrophilic/lipophilic
balance (log P) of the compounds plays an important role in the
absorption, distribution, metabolism and excretion of the
compounds.
S_ssO is an electrotopological descriptor that describes the
energy states of the O bonds, especially ether oxygen atoms in the
molecule [22]. The compounds 6c, 6d and 6f have ether linkage
4. QSAR studies
Table 2
In the case of Salmonella typhi, compounds 6b, 6e, 6g and 6h are
most active and compounds 6c, 6d and 6f are least active in this set.
Synthesis of dispiropyrrolidines.
Entry
Dipolarophiles (3)
R
Product
Time (h)
Yield %a
1
2
3
4
5
6
7
8
9
3a
3a
3b
3b
3a
3b
3c
3c
3d
H
Me
H
Me
Propargyl
Bn
H
Me
H
6a
6b
6c
6d
6e
6f
6g
6h
6i
3.0
4.0
4.5
3.0
6.0
3.5
4.0
4.5
5.0
75
71
74
72
64
76
66
65
69
Table 1
Synthesis of 3-benzylidene-1-methyl-pyrrolidine-2,5-dione 3.
Entry
Ar
Product
Time (min)
Yield %a
1
2
3
4
2-OHC6H4
4-OMeC6H4
4-ClC6H4
3a
3b
3c
3d
30
45
60
45
78
72
68
76
4-MeC6H4
a
a
Isolated yield after column chromatography.
Isolated yield after column chromatography.

3448
K. Karthikeyan et al. / European Journal of Medicinal Chemistry 45 (2010) 3446e3452
P. vulgaris. Fig. 3 shows the plot relating the predicted activity with
the experimental values for P. vulgaris.
In the case of Pseudomonas aeruginosa, compounds 6a, 6b, 6e, 6g
and 6h are the most active and 6d, 6f and 6i are the least active
ones. The QSAR equation developed for P. aeruginosa and the cor-
responding statistics is given below:
Activity ¼ 1:4142 ꢁ 1:5966 ꢂ S sssCH ꢁ 0:0197 ꢂ Shadow ꢁ YZ
r ¼ 0:90; r2 ¼ 0:81; ra2dj ¼ 0:75; q2 ¼ 0:61;
F ꢁ ratio ¼ 12:73; error ¼ 0:11
Both the descriptors in this biparametric model have negative
contribution towards activity. Shadow index is a molecular prop-
erty describing its shape, conformation and orientation. Shadow-YZ
denotes the area of the molecular shadow in the YZ plane [23]. Our
earlier research also showed the negative contribution of shadow
descriptors (shadow-Y length) towards the antibacterial activity of
isoxazolidine derivatives against P. aeruginosa [24]. S_sssCH, is an
electrotopological descriptor, that represents carbon atom which
has three single bonds. This has a negative contribution to activity
indicating that compounds with more S_sssCH will be less active.
Table 7 lists the numerical values of the descriptors used for
developing the QSAR equation for P. aeruginosa. Fig. 4 shows the
plot relating the predicted activity with the experimental values for
P. aeruginosa.
Fig. 1. ORTEP diagram of compound 6d.
In the case of Bacillus subtilis, compounds 6e, 6f and 6h are the
most active and compounds 6a and 6i are the least active. The QSAR
equation developed for antibacterial activity against B. subtilis and
the statistics are given below:
through their 4-OCH3 substitution, hence they are least active.
Table 4 lists the predicted and experimental activity (¼ ꢁlog (MIC)).
Table 5 lists the numerical values of the descriptors used for
developing the QSAR equation for S. typhi. Fig. 2 shows the plot
relating the predicted activity with the experimental values for
S. typhi.
Activity ¼ ꢁ3:1291 þ 0:5961 ꢂ IC þ 0:0164 ꢂ shadow ꢁ YZ
r ¼ 0:97; r2 ¼ 0:94; ra2dj ¼ 0:92; q2 ¼ 0:57;
F ꢁ ratio ¼ 48:33; error ¼ 0:03
IC belongs to the class of information e theoretic topological
indices which are calculated by multiplying average information
content with the total number of atoms. The average information
content is obtained based on Shannon information theory and is
calculated by the following formula [25,26].
In the case of Proteus vulgaris, compounds 6b, 6e, 6g and 6h are
the most active and compounds 6c, 6d and 6f are the least active.
The QSAR equation developed for P. vulgaris and the corresponding
statistics are given below:
Activity ¼ 0:8148 ꢁ 0:0508 ꢂ S ssO
r ¼ 0:98; r2 ¼ 0:96; ra2dj ¼ 0:95; q2 ¼ 0:92;
F ꢁ ratio ¼ 162:76; error ¼ 0:03
The electrotopological descriptor, S_ssO is described before and
once again it has a negative effect on activity. Compounds 6c, 6d
and 6f have ether linkage and have least activity. Their activities are
highly correlated with S_ssO (r ¼ 0.99). Table 6 lists the numerical
values of the descriptors used for developing the QSAR equation for
X
ni
n
ni
n
kIC ¼ ꢁ
log 2
Where n e total number of atoms in the molecule
ni e number of atoms in i-th class
IC is also related to the lipophilicity of the molecule [19]. This
descriptor shows a positive correlation with the activity. Increase in
the IC value leads to increase in the antibacterial activity. The
antibacterial activity against B. subtilis of the most active
compounds and the IC values are found to be highly correlated
(r ¼ 0.89). Shadow-YZ contributes positively denoting that increase
in area in YZ plane will lead to increase in activity. The antibacterial
activity against B. subtilis of most active compounds and shadow-YZ
are highly correlated (r ¼ 0.79). This descriptor also appeared in the
QSAR for P. aeruginosa. Table 8 lists the numerical values of the
descriptors used for developing the QSAR equation for B. subtilis.
Fig. 5 shows the plot relating the predicted activity with the
experimental values for B. subtilis.
Table 3
The observed antibacterial activities of dispiropyrrolidines.
Compounds
Antibacterial activity (MIC in mM)
B. subtilis
S. aureus
P. aeruginosa
S. typhi
P. vulgaris
6a
6b
6c
6d
6e
6f
6g
6h
0.639
0.617
0.617
0.596
0.291
0.504
0.610
0.590
0.642
0.168
0.001
0.639
0.617
0.617
0.298
0.291
0.252
0.305
0.295
0.321
0.084
0.002
0.080
0.077
0.154
0.298
0.146
0.252
0.152
0.147
0.321
0.168
0.004
0.160
0.154
0.308
0.298
0.146
0.252
0.152
0.147
0.160
0.084
0.004
0.160
0.154
0.308
0.298
0.146
0.252
0.152
0.147
0.160
0.168
0.001
In the case of Staphylococcus aureus, compounds 6d, 6e, 6f and 6h
are found to be the most active and compounds 6a, 6b and 6c are the
least active. The QSAR equation developed for the antibacterial
activity against S. aureus and the statistics are given below:
6i
Ampicillin
Tetracycline

K. Karthikeyan et al. / European Journal of Medicinal Chemistry 45 (2010) 3446e3452
3449
Table 4
Observed and predicted activities of dispiropyrrolidines.
Compounds Antibacterial activity (ꢁlog (MIC))
B. subtilis
S. aureus
P. aeruginosa
S. typhi
P. vulgaris
Observed
activity
Predicted
activity
Observed
activity
Predicted
activity
Observed
activity
Predicted
activity
Observed
activity
Predicted
activity
Observed
activity
Predicted
activity
6a
6b
6c
6d
6e
6f
6g
6h
6i
0.1947
0.2100
0.2100
0.2248
0.5360
0.2972
0.2147
0.2293
0.1925
0.1950
0.1594
0.2081
0.2675
0.5173
0.3009
0.2082
0.2507
0.2108
0.1947
0.2100
0.2100
0.5258
0.5360
0.5982
0.5157
0.5304
0.4935
0.2001
0.2078
0.2886
0.4729
0.5366
0.5500
0.4703
0.5937
0.4121
1.0978
1.1131
0.8121
0.5258
0.8371
0.5982
0.8168
0.8314
0.4935
1.1209
1.0554
0.8145
0.5011
0.8832
0.6670
0.7631
0.6527
0.6663
0.7968
0.8121
0.5110
0.5258
0.8371
0.5982
0.8168
0.8314
0.7946
0.8007
0.8185
0.5588
0.5214
0.8185
0.5552
0.8185
0.8185
0.8137
0.7968
0.8121
0.5110
0.5258
0.8371
0.5982
0.8168
0.8314
0.7946
0.8148
0.8148
0.5467
0.5462
0.8148
0.5429
0.8148
0.8148
0.8148
parameters are related to the solubility and hydrophobic/hydro-
philic nature of the compounds. The drug solubility affects its
penetration through bacterial cell membrane. Bacterial cell
membrane is a very complicated structure where the outer surface
is hydrophilic in nature and inner leaflet is lipophilic in nature.
Thus the former acts as a barrier for hydrophobic drugs and the
later acts as barrier for hydrophilic drugs. An ideal candidate for
antibacterial therapy should posses a good hydrophilic/lipophilic
balance. This QSAR study showed the contribution of the above
said descriptors.
Activity ¼ ꢁ10:2792 þ 1:6917 ꢂ IC þ 2:6087 ꢂ CHI ꢁ V ꢁ 3 C
r ¼ 0:94; r2 ¼ 0:89; ra2dj ¼ 0:85; q2 ¼ 0:71;
F ꢁ ratio ¼ 23:11; error ¼ 0:06
Information content shows a positive correlation and it also
appeared in the QSAR of B. subtilis. The antibacterial activity against
S. aureus of the most active compounds showed positive correla-
tion with IC (r ¼ 0.62). A relationship between IC and the anti-
bacterial activity against S. aureus is also observed by us earlier
[19]. CHI-V-3_C belongs to the Kier and Hall molecular connec-
tivity indices and the indices. It explains the branching, ring
structures and flexibility of the molecule. CHI-V-3_C has a positive
correlation with the activity. Table 4 shows the observed and
predicted activities of dispiropyrrolidines against all the five
organisms. Table 9 lists the numerical values of the descriptors
used for developing the QSAR equation for S. aureus. Fig. 6 shows
the plot relating the predicted activity with the experimental
values for S. aureus.
6. Experimental
Melting points were determined in capillary tubes and are
uncorrected. IR spectra were taken as neat for liquid compounds
and 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 CDCl3 and 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.
5. Conclusion
6.1. Experimental procedure for the synthesis of dispiropyrrolidines
derivatives (6aei)
In summary, we have reported the synthesis and character-
isation of nine dispiropyrrolidines. These compounds were evalu-
ated for their activities against five bacteria. Among these nine
compounds tested, compounds 6e and 6h exhibited reasonably
good activity and 6c exhibited least activity. QSAR was developed
by relating antibacterial activity of dispiropyrrolidines and their
physicochemical descriptors. Statistical measures obtained for all
five QSAR equations were found to be satisfactory. Electro-
topological descriptors, Jurs descriptors, topology, electrostatic
forces, shape and hydrophobic nature of the molecule were found
to contribute to the antibacterial activities. All the above said
A mixture of isatin 4aed (1 mmol), sarcosine 5 (1 mmol), and 3-
benzylidene-1-methyl-pyrrolidine-2,5-dione 3aed (1 mmol) was
refluxed in methanol (10 ml). Completion of the reaction was evi-
denced by TLC analysis. The solvent was removed under vacuo and
the crude product was subjected to column chromatography using
ethyl acetate: petroleum ether (2:8) as an eluent to afford pure
dispiropyrrolidines 6aei.
0.9
0.8
0.7
0.6
0.5
Table 5
Lists the numerical values of the descriptors used for developing the QSAR equation
for S. typhi.
Compounds
S_ssO
Jurs-RPCS
6a
6b
6c
6d
6e
6f
6g
6h
6i
0.0000
0.0000
5.2782
5.2882
0.0000
5.3522
0.0000
0.0000
0.0000
0.0833
0.0000
0.0000
0.1729
0.0000
0.0000
0.0000
0.0000
0.0226
0.5
0.6
0.7
0.8
0.9
Observed activity
Fig. 2. Shows the plot relating the predicted activity with the experimental values for
S. typhi.

3450
K. Karthikeyan et al. / European Journal of Medicinal Chemistry 45 (2010) 3446e3452
Table 6
Table 7
Lists the numerical values of the descriptors used for developing the QSAR equation
Lists the numerical values of the descriptors used for developing the QSAR equation
for P. vulgaris.
for P. aeruginosa.
Compounds
S_ssO
Compounds
S_sssCH
Shadow-YZ
6a
6b
6c
6d
6e
6f
6g
6h
6i
0.0000
0.0000
5.2782
5.2882
0.0000
5.3522
0.0000
0.0000
0.0000
6a
6b
6c
6d
6e
6f
6g
6h
6i
ꢁ0.5213
ꢁ0.5198
ꢁ0.3271
ꢁ0.2105
ꢁ0.5714
ꢁ0.3625
ꢁ0.3305
ꢁ0.3290
ꢁ0.2718
57.1392
60.3422
56.9542
63.4080
73.2639
67.3091
59.8376
65.3177
59.9963
6.1.1. 40-(2-Hydroxyphenyl)-10,100-dimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6a)
CHHCONMe), 2.48 (s, 3H, NCH3), 3.36 (t, 1H, J ¼ 8.4 Hz, Ha), 3.69 (s,
3H, OCH3), 3.71 (t,1H, J ¼ 9.2 Hz, Hb), 4.16 (t,1H, J ¼ 9.2 Hz, Hc), 6.76
(d, 1H, J ¼ 7.6 Hz, Ar-H), 6.88 (d, 1H, J ¼ 8.4 Hz, Ar-H), 6.95 (t, 1H,
J ¼ 7.6 Hz, Ar-H), 6.99 (d, 1H, J ¼ 8.4 Hz, Ar-H), 7.17e7.21 (m, 2H, Ar-
H), 7.30 (d, 1H, J ¼ 8.4 Hz, Ar-H), 7.57 (d, 1H, J ¼ 8.4 Hz, Ar-H), 10.65
Colourless solid; mp 236e238 ꢀC; IR (cmꢁ1): 3412, 2995, 1782,
1705, 1602, 1441, 1157; 1H NMR (500 MHz, DMSO-d6):
d 1.97 (d, 1H,
J ¼ 17.6 Hz, CHHCONMe), 2.02 (d, 1H, J ¼ 17.6 Hz, CHHCONMe), 2.04
(s, 3H, NCH3), 2.57 (s, 3H, NCH3), 3.25 (t, 1H, J ¼ 8.4 Hz, Ha), 4.01 (t,
1H, J ¼ 9.9 Hz, Hb), 4.34 (t, 1H, J ¼ 9.9 Hz, Hc), 6.73e6.76 (m, 2H, Ar-
H), 6.82 (t, 1H, J ¼ 6.9 Hz, Ar-H), 6.93 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.07 (t,
1H, J ¼ 6.9 Hz, Ar-H), 7.17e7.21 (m, 2H, Ar-H), 7.42 (d, 1H, J ¼ 7.6 Hz,
Ar-H), 9.59 (s, 1H, OH),10.56 (s, 1H, NH); 13C NMR (125 MHz, DMSO-
(s, 1H, NH); 13C NMR (125 MHz, DMSO-d6):
d 24.9, 35.1, 36.6, 48.3,
55.5, 58.8, 61.5, 77.9, 110.5, 114.6, 115.0, 122.4, 125.5, 126.5, 131.5,
132.7, 143.2, 159.0, 174.5, 177.6, 179.4; MS m/z ¼ 406 Mþ þ 1; Anal.
Calcd for C23H23N3O4 (405.17): C, 68.13; H, 5.72; N, 10.36. Found: C,
67.91; H, 5.69; N, 10.51.
d6): d 24.9, 35.4, 36.0, 41.9, 54.9, 59.6, 77.9, 110.4, 114.9, 119.8, 122.5,
124.7, 125.0, 126.7, 128.6, 128.9, 130.6, 143.7, 156.1, 175.4, 177.4,
180.2; MS m/z ¼ 392 Mþ þ 1; Anal. Calcd for C22H21N3O4 (391.15): C,
67.51; H, 5.41; N, 10.74. Found: C, 67.46; H, 5.45; N, 10.61.
6.1.4. 40-(4-Methoxyphenyl)-1,10,100-trimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6d)
Colourless solid; mp 174e176 ꢀC; IR (cmꢁ1): 3405, 2967, 1788,
1710, 1622, 1451, 1151; 1H NMR (500 MHz, DMSO-d6):
d 1.96 (s, 3H,
6.1.2. 40-(2-Hydroxyphenyl)-1,10,100-trimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6b)
NCH3), 2.03 (d, 1H, J ¼ 18.4 Hz, CHHCONMe), 2.28 (d, 1H, J ¼ 18.4 Hz,
CHHCONMe), 2.50 (s, 3H, NCH3), 3.07 (s, 3H, NCH3), 3.39 (t, 1H,
J ¼ 8.4 Hz, Ha), 3.70 (s, 3H, OCH3), 3.74 (t, 1H, J ¼ 9.2 Hz, Hb), 4.15 (t,
1H, J ¼ 9.2 Hz, Hc), 6.89 (d, 2H, J ¼ 8.4 Hz, Ar-H), 6.98 (d, 1H,
J ¼ 7.6 Hz, Ar-H), 7.04 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.26 (d, 1H, J ¼ 6.9 Hz,
Ar-H), 7.31e7.33 (m, 3H, Ar-H); 13C NMR (125 MHz, DMSO-d6):
Orange solid; mp 198e200 ꢀC; IR (cmꢁ1): 3421, 2994,1789,1702,
1601, 1444, 1167; 1H NMR (500 MHz, CDCl3):
d 2.10 (brs, 1H),
2.13e2.17 (m, 4H, NCH3, CHHCONMe), 2.33 (d, 1H, J ¼ 19.1 Hz,
CHHCONMe), 2.68 (s, 3H, NCH3), 3.14 (s, 3H, NCH3), 3.49 (t, 1H,
J ¼ 8.4 Hz, Ha), 4.16 (t, 1H, J ¼ 9.2 Hz, Hb), 4.59 (t, 1H, J ¼ 9.2 Hz, Hc),
6.79 (d, 2H, J ¼ 7.6 Hz, Ar-H), 6.92 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.07 (t, 1H,
J ¼ 7.6 Hz, Ar-H), 7.12 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.31 (t, 1H, J ¼ 7.6 Hz,
Ar-H), 7.43 (d, 1H, J ¼ 7.6 Hz, Ar-H), 7.47 (d, 1H, J ¼ 7.6 Hz, Ar-H); 13C
d
24.9, 26.2, 35.1, 36.4, 48.8, 55.6, 58.8, 61.3, 77.7, 109.5, 114.6, 123.1,
124.7, 126.3, 130.1, 130.6, 131.5, 144.7, 159.0, 174.4, 175.6, 179.5; MS
m/z ¼ 420 Mþ þ 1; Anal. Calcd for C24H25N3O4 (419.18): C, 68.72; H,
6.01; N, 10.02. Found: C, 68.78; H, 6.06; N, 10.14.
NMR (125 MHz, CDCl3): d 24.9, 25.9, 35.4, 35.9, 55.1, 60.1, 78.3,108.7,
116.0, 120.9, 123.3, 123.7, 123.9, 126.7, 128.9, 130.2, 130.5, 144.5,
154.9, 176.0, 176.1, 180.7; MS m/z ¼ 406 Mþ þ 1; Anal. Calcd for
C23H23N3O4 (405.17): C, 68.13; H, 5.72; N, 10.36. Found: C, 68.41; H,
5.71; N, 10.16.
6.1.5. 40-(2-Hydroxyphenyl)-10,100-dimethyl-1-propargldispiro
[indoline-3,20-pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6e)
Yellow solid; mp 178e180 ꢀC; IR (cmꢁ1): 3422, 2998, 1785, 1712,
1622, 1449, 1154; 1H NMR (500 MHz, DMSO-d6):
d 1.92 (s, 2H,
CH2CONMe), 2.01 (s, 3H, NCH3), 2.61 (s, 3H, NCH3), 3.17 (t, 1H,
J ¼ 2.3 Hz, C^CH), 3.31 (t, 1H, J ¼ 8.4 Hz, Ha), 4.02 (t, 1H, J ¼ 9.2 Hz,
Hb), 4.36 (t, 1H, J ¼ 9.2 Hz, Hc), 4.45 (s, 2H, NCH2), 6.76 (d, 1H,
J ¼ 7.6 Hz, Ar-H), 6.84 (t,1H, J ¼ 6.9 Hz, Ar-H), 7.05e7.07 (m, 3H, Ar-H),
7.30 (d, 1H, J ¼ 6.9 Hz, Ar-H), 7.35 (t,1H, J ¼ 8.4 Hz, Ar-H), 7.47 (d, 1H,
J ¼ 7.6 Hz, Ar-H), 9.62 (s, 1H, OH); 13C NMR (125 MHz, DMSO-d6):
6.1.3. 40-(4-Methoxyphenyl)-10,100-dimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6c)
Colourless solid; mp 232e234 ꢀC; IR (cmꢁ1): 3405, 2975, 1762,
1709, 1608, 1445, 1159; 1H NMR (500 MHz, DMSO-d6):
d 2.01 (s, 3H,
NCH3), 2.07 (d, 1H, J ¼ 18.4 Hz, CHHCONMe), 2.38 (d, 1H, J ¼ 18.3 Hz,
1.2
1.1
1.0
0.9
0.8
0.7
0.6
0.5
0.4
0.9
0.8
0.7
0.6
0.5
0.5
0.6
0.7
0.8
0.9
0.4
0.5
0.6
0.7
0.8
0.9
1.0
1.1
1.2
observed activity
Observed activity
Fig. 3. Shows the plot relating the predicted activity with the experimental values for
P. vulgaris.
Fig. 4. Shows the plot relating the predicted activity with the experimental values for
P. aeruginosa.

K. Karthikeyan et al. / European Journal of Medicinal Chemistry 45 (2010) 3446e3452
3451
Table 8
Table 9
Lists the numerical values of the descriptors used for developing the QSAR equation
Lists the numerical values of the descriptors used for developing the QSAR equation
for B. subtilis.
for S. aureus.
Compounds
IC
Shadow-YZ
Compounds
IC
CHI-V-3_C
6a
6b
6c
6d
6e
6f
6g
6h
6i
4.0044
3.8566
4.0314
3.9536
4.1014
3.9023
3.9523
3.8729
3.9523
57.1392
60.3422
56.9542
63.4080
73.2639
67.3091
59.8376
65.3177
59.9963
6a
6b
6c
6d
6e
6f
6g
6h
6i
4.0044
3.8566
4.0314
3.9536
4.1014
3.9023
3.9523
3.8729
3.9523
1.4203
1.5191
1.4367
1.5578
1.4864
1.6206
1.5576
1.6564
1.5353
NCH3), 2.02 (d, 1H, J ¼ 18.4 Hz, CHHCONMe), 2.27 (d, 1H, J ¼ 18.3 Hz,
CHHCONMe), 2.48 (s, 3H, NCH3), 3.09 (s, 3H, NCH3), 3.45 (t, 1H,
J ¼ 8.4 Hz, Ha), 3.72 (t, 1H, J ¼ 9.2 Hz, Hb), 4.20 (t, 1H, J ¼ 9.2 Hz, Hc),
7.00 (d, 1H, J ¼ 7.6 Hz, Ar-H), 7.04 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.22 (d, 1H,
J ¼ 7.6 Hz, Ar-H), 7.33 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.40 (d, 2H, J ¼ 8.4 Hz,
Ar-H), 7.46 (d, 2H, J ¼ 8.4 Hz, Ar-H); 13C NMR (125 MHz, DMSO-d6):
d
24.9, 28.8, 35.2, 35.4, 42.3, 54.9, 59.7, 74.6, 77.5, 78.4, 110.1, 114.9,
119.8,123.6,124.2,124.5,126.7,128.7,128.8,130.7,143.2,156.2,175.0,
175.2, 180.1; MS m/z ¼ 430 Mþ þ 1; Anal. Calcd for C25H23N3O4
(429.17): C, 69.92; H, 5.40; N, 9.78. Found: C, 69.74; H, 5.42; N, 9.84.
6.1.6. 1-Benzyl-40-(4-Methoxyphenyl)-10,100-dimethyldispiro
[indoline-3,20-pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6f)
Yellow solid; mp 104e106 ꢀC; IR (cmꢁ1): 3412, 2998, 1786, 1705,
d
24.9, 26.2, 35.1, 36.7, 48.4, 59.0, 61.3, 77.8, 109.8, 123.2, 124.5, 126.1,
129.1, 130.8, 132.5, 132.7, 137.6, 143.4, 174.2, 175.6, 179.2; MS m/
z ¼ 424 Mþ þ 1, 426 Mþ þ 3; Anal. Calcd for C23H22ClN3O3 (423.13):
C, 65.17; H, 5.23; N, 9.91. Found: C, 65.27; H, 5.16; N, 9.98.
1602, 1441, 1158; 1H NMR (500 MHz, DMSO-d6):
d 2.01 (s, 3H,
NCH3), 2.10 (d, 1H, J ¼ 18.3 Hz, CHHCONMe), 2.42 (d, 1H, J ¼ 18.4 Hz,
CHHCONMe), 2.49 (s, 3H, NCH3), 3.43 (t, 1H, J ¼ 8.4 Hz, Ha), 3.69 (s,
3H, OCH3), 3.79 (t,1H, J ¼ 9.2 Hz, Hb), 4.24 (t,1H, J ¼ 9.2 Hz, Hc), 4.88
(s, 2H, NCH2Ph), 6.88e6.92 (m, 3H, Ar-H), 6.99e7.03 (m, 1H, Ar-H),
6.1.9. 40-(4-Methylphenyl)-10,100-dimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6i)
Yellow solid; mp 216e218 ꢀC; IR (cmꢁ1): 3402, 2998, 1782, 1710,
7.22e7.33 (m, 9H, Ar-H); 13C NMR (125 MHz, DMSO-d6):
d 24.9,
1602, 1439, 1157; 1H NMR (500 MHz, DMSO-d6):
d 2.01 (s, 3H,
35.2, 36.4, 43.4, 48.4, 55.5, 58.7, 61.6, 77.6, 110.1, 114.6, 123.2, 124.8,
126.6,128.0,129.2,129.9,130.6,131.5,132.7,136.7,143.7,159.1,174.3,
175.7, 179.2; MS m/z ¼ 496 Mþ þ 1; Anal. Calcd for C30H29N3O4
(495.22): C, 72.71; H, 5.90; N, 8.48. Found: C, 72.67; H, 5.91; N, 8.64.
NCH3), 2.05 (d,1H, J ¼ 18.4 Hz, CHHCONMe), 2.24 (s, 3H, NCH3), 2.41
(d, 1H, J ¼ 18.3 Hz, CHHCONMe), 2.47 (s, 3H, NCH3), 3.35 (t, 1H,
J ¼ 8.4 Hz, Ha), 3.74 (t, 1H, J ¼ 9.2 Hz, Hb), 4.18 (t, 1H, J ¼ 8.4 Hz, Hc),
6.77 (d, 1H, J ¼ 7.6 Hz, Ar-H), 6.95 (t, 1H, J ¼ 7.6 Hz, Ar-H), 7.12e7.25
(m, 6H, Ar-H), 10.64 (s, 1H, NH); 13C NMR (125 MHz, DMSO-d6):
6.1.7. 40-(4-Chlorophenyl)-10,100-dimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6g)
d
21.2, 24.9, 35.1, 36.8, 48.6, 58.6, 61.5, 77.9,110.5, 122.4, 125.5, 126.6,
129.8, 130.3, 130.5, 135.4, 137.1, 143.2, 174.4, 177.5, 179.4; MS m/
z ¼ 390 Mþ þ 1; Anal. Calcd for C23H23N3O3 (389.17): C, 70.93; H,
5.95; N, 10.79. Found: C, 71.09; H, 5.94; N, 10.61.
Colourless solid; mp 220e222 ꢀC; IR (cmꢁ1): 3412, 2985, 1782,
1710, 1608, 1445, 1147; 1H NMR (500 MHz, DMSO-d6):
d 2.01 (s, 3H,
NCH3), 2.05 (d, 1H, J ¼ 18.4 Hz, CHHCONMe), 2.36 (d, 1H, J ¼ 17.6 Hz,
CHHCONMe), 2.46 (s, 3H, NCH3), 3.42 (t,1H, J ¼ 8.4 Hz, Ha), 3.70 (t,1H,
J ¼ 9.2 Hz, Hb), 4.22 (t,1H, J ¼ 8.4 Hz, Hc), 6.78 (d,1H, J ¼ 7.6 Hz, Ar-H),
6.95 (t, 1H, J ¼ 6.9 Hz, Ar-H), 7.14 (d, 1H, J ¼ 6.9 Hz, Ar-H), 7.22 (t, 1H,
J ¼ 7.6 Hz, Ar-H), 7.39e7.43 (m, 4H, Ar-H), 10.69 (s, 1H, NH); 13C NMR
6.2. Materials and methods for the antibacterial activity
B. subtilis NCIM 2718, S. aureus NCIM5021, S. typhi NCIM2501, P.
aeruginosa NCIM 5029 and P. vulgaris NCIM2813 were purchased
from National Chemical Laboratory, Pune, India and the reference
antibiotics were purchased from Hi-media labs (Mumbai, India).
(125 MHz, DMSO-d6):
d 24.9, 35.1, 36.8, 47.9, 59.1, 61.4, 78.1, 110.6,
122.5, 125.3, 126.3, 129.1, 130.6, 132.5, 132.6, 137.8, 143.2, 174.3, 177.6,
179.2; MS m/z ¼ 410 Mþ þ 1, 412 Mþ þ 3;Anal. Calcd forC22H20ClN3O3
(409.12): C, 64.47; H, 4.92; N,10.25. Found: C, 64.61; H, 4.97; N,10.17.
6.2.1. Antibacterial evaluation
Minimum Inhibitory Concentration (MIC) was determined by
microdilution broth assay method [27] as reported by Sarker et al.
6.1.8. 40-(4-Chlorophenyl)-1,10,100-trimethyldispiro[indoline-3,20-
pyrrolidine-30,300-pyrrolidine]-2,200,500-trione (6h)
Yellow solid; mp 226e228 ꢀC; IR (cmꢁ1): 3408, 2996, 1782, 1717,
0.7
0.6
0.5
0.4
0.3
0.2
0.1
1609, 1445, 1157; 1H NMR (500 MHz, DMSO-d6):
d
1.96 (s, 3H,
0.6
0.5
0.4
0.3
0.2
0.1
0.1
0.2
0.3
0.4
0.5
0.6
0.1
0.2
0.3
0.4
0.5
0.6
Observed activity
Observed activity
Fig. 5. Shows the plot relating the predicted activity with the experimental values for
B. subtilis.
Fig. 6. Shows the plot relating the predicted activity with the experimental values for
S. aureus.

3452
K. Karthikeyan et al. / European Journal of Medicinal Chemistry 45 (2010) 3446e3452
[28] with slight modifications, using resazurin dye as an indicator.
References
Bacterial strains were cultured in Muller hinton broth to reach
105 CFU/ml 10 mg of dis-
[1] (a) R. Huisgen, in: A. Padwa (Ed.), 1,3-Dipolar Cycloaddition Chemistry, vols.
1 and 2, Wiley, New York, 1984;
a
final inoculum size of
5
ꢂ
piropyrrolidines were dissolved in absolute ethanol to reach a final
volume of 1 ml [29]. This solution is serially diluted and added to
successive wells in a 96 well microtiter plate and incubated at 37 ꢀC
with respective micro-organisms for a period of 18 h. Growth and
sterility controls were maintained for all the experiment. Test
compounds serially diluted with ethanol were used (uninoculated
dilution) to determine if they precipitated out during the course of
(b) O. Tsuge, S. Kanemasa, in: A.R. Katritzky (Ed.), Advances in Heterocyclic
Chemistry, vol. 45, Academic Press, San Diego, 1989, pp. 231e252;
(c) R. Grigg, V. Sridharan, in: D.P. Curran (Ed.), Advances in Cycloaddition, vol.
3, Jai Press, London, 1993, pp. 161e180.
[2] (a) J.R. Liddell, Nat. Prod. Rep. 13 (1996) 187e193;
(b) J.R. Liddell, Nat. Prod. Rep. 15 (1998) 363e370.
[3] (a) T. Hartmann, L. Witte, in: S.W. Pelletier (Ed.), Chemistry, Biology and
Chemoecology of Pyrrolizidine Alkaloids, Alkaloids: Chemical and Biological
Perspectives, vol. 9, Pergamon Press, 1995;
(b) R.J. Molineux, in: S.W. Pelletier (Ed.), Alkaloids: Chemical and Biological
Perspective, Wiley, New York, 1987 (Chapter 1);
(c) S. Fujimori, Jpn. Patent Appl. 88-2912. Chem. Abstr. 112 (1990) 98409.
[4] (a) R.R. Kumar, S. Perumal, P. Senthilkumar, P. Yogeeswari, D. Sriram, Tetra-
hedron 64 (2008) 2962e2971;
the experiments. After 18 h, 10 mL of 0.01% resazurin solution was
added to the wells and incubated for 2 h. A blank assay with
ethanol was carried out. The color change was assessed visually.
Blue color showed that the compound inhibited the micro-
organism and a change in color from blue to pink showed the
growth of the organism.
(b) R.R. Kumar, S. Perumal, P. Senthilkumar, P. Yogeeswari, D. Sriram, Eur. J.
Med. Chem. 44 (2009) 3821e3829;
(c) R.R. Kumar, S.M. Rajesh, S. Perumal, D. Banerjee, P. Yogeeswari, D. Sriram,
Eur. J. Med. Chem. 45 (2010) 411e422.
[5] (a) A. Dell, D.H. Williams, H.R. Morris, G.A. Smith, J. Feeney, G.C.K. Roberts, J.
Am. Chem. Soc. 97 (1975) 2497e2502;
6.2.2. Modelling studies
The chemical structures of nine of the dispiropyrrolidines were
sketched and with the help of Cerius2 software (Accelrys, U.S.A),
their energies were minimized using consistent valence force field
(CVFF). This force field was parameterized using peptide and
protein structures and is suitable for small molecules [30]. To
facilitate the QSAR studies the antibacterial activity (minimum
(b) S. Sapo, O. Shiratori, K. Katagiri, J. Antibiot. 20 (1967) 270;
(c) C. Bailly, S. Echepare, F. Gago, M. Waring, Anti-Cancer Drug Des 14 (1999)
291e303.
[6] A.P. Kozikowski, Acc. Chem. Res. 17 (1984) 410e416.
[7] R.K. Howe, B.R. Shelton, J. Org. Chem. 55 (1990) 4603e4607.
[8] M. De Amici, C. De Michelli, V. Misani, Tetrahedron 46 (1990) 1975e1986.
[9] V.L. Cohen, E.E. Kleinmann, PCT. Int. Appl.. W.O. 24192. Chem. Abstr 123
(1995) 296610t.
[10] (a) W.A. Carroll, P.A. Grieco, J. Am. Chem. Soc. 115 (1993) 1164e1165;
(b) W.G. Earley, T. Oh, L.E. Overman, Tetrahedron Lett. 29 (1988) 3785e3788;
(c) Y. Ban, N. Taga, T. Oishi, Chem. Pharm. Bull. 24 (1976) 736e751;
(d) Y. Ban, M. Seto, T. Oishi, Chem. Pharm. Bull. 23 (1975) 2605e2613.
[11] (a) Y. Ban, N. Taga, T. Oishi, Tetrahedron Lett. 15 (1974) 187e190;
(b) E.E. Van Tamelen, J.P. Yardley, M. Miyano, W.B. Hinshaw Jr., J. Am. Chem.
Soc. 91 (1969) 7333e7341.
inhibitory concentration in
concentration.
m
M) is converted as ꢁlog (MIC)
Two hundred and forty-nine physicochemical properties
(descriptors), which include electronic, quantum mechanical, topo-
logical, spatial, structural, thermodynamic properties for these nine
structures, were determined. Several references deal with a detailed
description of these descriptors [31,32]. The Cerius2 software is
capable of developing the best QSAR using GFA (Genetic Function
Approximation) method. It can short list the descriptors from this
large pool. The quality of the regression models were tested using
various statistical parameters such as r2, ra2dj, cross-validated r2 (q2),
F-ratio, predicted residual sum of squares (PRESS) and standard error
of estimate. r2 measures the strength of the linear relationship
between the independent and the dependent variables. It is calcu-
[12] H. Kabat, E.F. Stohlman, M.I. Smith, J. Pharmacol. 80 (1994) 160e170.
[13] W.G. Rajeswaran, R.B. Labroo, L.A. Cohen, J. Org. Chem. 64 (1999) 1369e1371.
[14] (a) K. Karthikeyan, P.T. Perumal, S. Etti, G. Shanmugam, Tetrahedron 63 (2007)
10581e10586;
(b) K. Karthikeyan, T.V. Seelan, K.G. Lalitha, P.T. Perumal, Bioorg. Med. Chem.
Lett. 19 (2009) 3370e3373;
(c) K. Karthikeyan, R.S. Kumar, D. Muraidharan, P.T. Perumal, Tetrahedron
Lett. 50 (2009) 7175e7179;
(d) M. Damodiran, D. Muraidharan, P.T. Perumal, Bioorg. Med. Chem. Lett. 19
(2009) 3611e3614.
[15] S. Nirmala, K. Karthikeyan, E.T.S. Kamala, L. Sudha, P.T. Perumal, Acta Crys-
tallogr. E65 (2009) o1655.
[16] (a)Z. Zhong, R. Xing, S. Liu, L. Wang, S. Cai, P. Li, Carbohydr. Res. 343(2008)566e570;
(b) P.M. Sivakumar, S.P. Seenivasan, V. Kumar, M. Doble, Bioorg. Med. Chem. Lett.
(2010).doi:10.1016/j.bmcl.2010.03.083;
lated as r2 ¼ 1 ꢁ SSE/TSS, where SSE (sum of squares of the error) ¼ S
P
(Ydata ꢁ Ymodel)2. TSSðtotal sum of squaresÞ ¼ ðYdata ꢁ YÞ2. Y is
the average of all the Y data. Ydata and Ymodel are the observed and
model predicted activity values, respectively. Adjusted r2 is a modi-
fication to r2 that adjusts for the number of independent terms in
a model. The adjusted r2 will always be less than or equal to r2. It is
calculated as ra2dj ¼ 1 ꢁ ðn ꢁ 1=n ꢁ pÞð1 ꢁ r2Þ where n is total
number of observations and p is total number of terms in the model
(including the constant). Cross-validated r2(q2) ¼ 1 ꢁ PRESS/TSS.
Predicted sum of squares (PRESS) is the sum of the squared differ-
ences between the actual and predicted values for the dependent
variable using regression equation developed by leaving one data
point at a time. This LOO (leave one out) method is called internal
validation technique to determine the predictive capability of the
model. F-test is meant to find the significance of the regression
model and it is performed by comparing the ratio of the variances
between regression and the error (F-value) with the F-table value
for n ꢁ p ꢁ 1 degrees of freedom. An F-value greater than the F-table
value indicates that the regression relations is a not a chance fit but is
a statistically significant occurrence [33].
(c) P.M. Sivakumar, S.P. Seenivasan, V. Kumar, M. Doble, Bioorg. Med. Chem. Lett. 17
(2010) 1695e1700.
[17] D.T. Stanton, P.C. Jurs, Anal. Chem. 62 (1990) 2323e2329.
[18] A. Wiese, M. Munstermann, T. Gutsmann, B. Lindner, K. Kawahara,
U. Zahringer, U. Seydel, J. Membr. Biol. 162 (1998) 127e138.
[19] P.M. Sivakumar, G. Sheshayan, M. Doble, Chem. Biol. Drug Des. 71 (2008) 303e313.
[20] P.M. Sivakumar, S.K.G. Babu, M. Doble, Chem. Biol. Drug Des. 71 (2008) 447e463.
[21] P.M. Sivakumar, S. Priya, M. Doble, Chem. Biol. Drug Des. 73 (2009) 403e415.
[22] J.B. Bhonsle, A.K. Bhattacharjee, R.K. Gupta, J. Mol. Model. 13 (2007) 179e208.
[23] R.H. Rohrbaugh, P.C. Jurs, Anal. Chim. Acta 199 (1987) 99e109.
[24] M. Damodiran, P.M. Sivakumar, R. Senthil Kumar, D. Muralidharan, B.V.N. Phani
Kumar, M. Doble, P.T. Perumal, Chem. Biol. Drug Des. 74 (2009) 494e506.
[25] L.B. Kier, J. Pharm. Sci. 69 (1980) 807e810.
[26] D. Bonchev, Information Theoretic Indices for Characterization of Chemical
Structure. Wiley-Interscience, New York, 1983.
[27] F. Candan, M. Unlu, B. Tepe, D. Daferera, M. Polissiou, A. Sokmen, H.A. Akpulat,
J. Ethnopharmacol. 87 (2003) 215e220.
[28] S.D. Sarker, L. Nahar, Y. Kumarasamy, Methods 42 (2007) 321e324.
[29] E.S. Charles, V.K. Agrawal, S. Sharma, R.N. Iyer, O.P. Srivastava, Eur. J. Med.
Chem. 14 (1979) 435e438.
[30] W.D. Cornell, P. Cieplak, C.I. Bayly, I.R. Gould, K.M. Merz, D.M. Ferguson, D.
C. Spellmeyer, T. Fox, J.W. Caldwell, P.A. Kollman, J. Am. Chem. Soc. 117 (1995)
5179e5197.
[31] (a) R. Todeschini, M. Lasagni, E. Marengo, J. Chemom. 8 (1994) 263e272;
(b) R. Todeschini, V. Consonni, Handbook of Molecular Descriptors. Wiley-
VCH, Weinheim, 2000.
Acknowledgements
[32] M. Karelson, Molecular Descriptors in QSAR/QSPR. Wiley Interscience, New
York, 2000.
[33] A.R. Leach, Molecular Modelling Principles and Applications, second ed..
Pearson Education EMA, U.K, 2001, 698e702.
One of the authors K. Karthikeyan thanks the Council of Scien-
tific and Industrial Research, New Delhi, India for the research
fellowship.