Synthesis, chemical characterization and biological screening for cytotoxicity and antitumor activity of organotin (IV) derivatives of 3,4-methylenedioxy 6-nitrophenylpropenoic acid

Article abstract of DOI:10.3390/12102348

A series of mono-, di- and triorganotin compounds with general formulae [RSnL₂Cl], R = Bu (compound 3), [R₂SnL₂], where R = Me, Et, Bu, Oct (compounds 1, 2, 4 and 6) and [R3SnL], where R = Bu, Cy and Ph (compounds 5, 7 and 8) and where L = 3,4-methylenedioxy-6- nitrophenylpropenoic acid have been prepared and characterized by elemental analysis, multinuclear (¹H-, ¹³C- and ¹¹⁹Sn-) NMR and mass spectrometry. The ligand and its respective organotin complexes were screened for cytotoxicity using the brine shrimp lethality assay and for antitumor activity using the crown gall tumor inhibition (potato disc) assay. The bioassay results support the conclusion that the biological activities of these synthetic compounds are in the following order: [RSnL₂Cl] < [R₂SnL₂] < [R₃SnL].

Full text of DOI:10.3390/12102348

Molecules 2007, 12, 2348-2363
molecules
ISSN 1420-3049
© 2007 by MDPI
www.mdpi.org/molecules
Full Paper
Synthesis, Chemical Characterization and Biological Screening
for Cytotoxicity and Antitumor Activity of Organotin (IV)
Derivatives of 3,4-Methylenedioxy 6-nitrophenylpropenoic Acid
Muhammad Sheeraz Ahmad 1, Mukhtiar Hussain 2, Muhammad Hanif 2, Saqib Ali 2 and
Bushra Mirza 1,*
1
Department of Biochemistry, Faculty of Biological Sciences, Quaid-i-Azam University Islamabad
Pakistan; E-mail: sheeraz_qau@yahoo.com (Muhammad Sheeraz Ahmad)
2
Department of Chemistry, Quaid-i-Azam University, Islamabad-45320, Pakistan
* Author to whom correspondence should be addressed; E-mail: bushra_dr@yahoo.com
Received: 17 July 2007; in revised form: 9 August 2007 / Accepted: 10 August / Published: 25 October
2007
Abstract: A series of mono-, di- and triorganotin compounds with general formulae
[RSnL2Cl], R = Bu (compound 3), [R2SnL2], where R = Me, Et, Bu, Oct (compounds 1, 2,
4 and 6) and [R3SnL], where R = Bu, Cy and Ph (compounds 5, 7 and 8) and where L =
3,4-methylenedioxy-6-nitrophenylpropenoic acid have been prepared and characterized by
13
elemental analysis, multinuclear (1H-, C- and 119Sn-) NMR and mass spectrometry. The
ligand and its respective organotin complexes were screened for cytotoxicity using the
brine shrimp lethality assay and for antitumor activity using the crown gall tumor
inhibition (potato disc) assay. The bioassay results support the conclusion that the
biological activities of these synthetic compounds are in the following order: [RSnL2Cl] <
[R2SnL2] < [R3SnL].
Keywords: Antitumor; cytotoxicity; mass spectrometry; multinuclear NMR; organotin.
Introduction
Organotin (IV) complexes are extensively used as catalysts, stabilizers, biocides, antifouling agents
and as wood preservatives [1-5]. Organotin (IV) derivatives of carboxylic acids are of special interest

Molecules 2007, 12
2349
with regards to their methods of synthesis, structural elucidation and biological activity [6-9].
Generally these compounds can be well characterized by multinuclear NMR (1H-, 13C- and 119Sn-)
spectroscopy [10, 12]. In recent years organotin (IV) carboxylates have attracted much attention owing
to their potential biocidal activity and cytotoxicity [5, 12]. In addition, among organometallic
compounds there has been increased interest in organotin carboxylates due to their activity against
various types of cancer cells as many of the known di- and triorganotin (IV) carboxylates display
interesting antitumor activities [12-15].
Their structural diversity was also of interest as both diorganotin and triorganotin esters show rich
and diverse structural chemistry, as cited in a recent review [8]. Keeping in mind this structural and
biological diversity of organotin (IV) carboxylates, we have now synthesized some organotin (IV)
carboxylates of 3,4-methylenedioxy-6-nitrophenylpropenoic acid (L). The structures of these
compounds have been evaluated by various analytical techniques such as elemental analysis,
13
multinuclear NMR (1H-, C- and 119Sn-) and mass spectrometry. The ligand L and its organotin (IV)
derivatives have been screened for cytotoxicity using the brine shrimp lethality assay and for antitumor
activity with the crown gall tumor inhibition assay (potato disc assay).
Results and Discussion
Synthesis
Mono-, di- and triorganotin complexes were synthesized by refluxing stoichiometric amounts of
the ligand acid, R'COOH, with the corresponding oxides/hydroxides (Procedure I) or its sodium salt
with the corresponding organotin chlorides in toluene (Procedure II), as summarized in equations I-V.
RSn(OH)2Cl + 2HL ⎯→ RSn(Cl)L2 + 2H2O
R= n-Bu (3)
(I)
R2SnO + 2HL ⎯→ R2SnL2 + H2O
R = n-Bu (4), n-Oct (6)
(II)
R2SnCl2 + 2NaL ⎯→ R2SnL2 + 2NaCl
R = Me (1), Et (2)
(III)
(IV)
(V)
R3SnCl + NaL ⎯→ R3SnL + NaCl
R = n-Bu (5)
R3Sn(OH)+HL⎯→R3SnL+H2O
R= Cy (7), Ph (8)
All the newly synthesized compounds are air stable crystalline solids, soluble in common organic
solvents, and their physical data is summarized in Table 1. The structures of these compounds and
coordination behavior of metal have been addressed by different analytical techniques such as
elemental analysis, multinuclear NMR (1H, 13C and 119Sn) and mass spectrometry.

Molecules 2007, 12
2350
Table 1. Physical data of 3,4-methylenedioxy-6-nitrophenylpropenoic acid (L) and its
organotin (IV) derivatives.
Elemental Analysis
% Calculated (Found)
Compound Molecular
Molecular
Weight
M.P.
(˚C)
Yield
(%)
No.
Formula
C
H
N
42.51
(42.48)
44.35
2.90
(2.96)
3.46
4.51
1
2
C22H18O12N2Sn
C24H22O12N2Sn
622
650
237-240
176-178
86
67
(4.58)
4.31
(44.40)
(3.43)
(4.42)
42.13
3.07
4.10
3
4
C24H26O12N2SNCl
C28H30O12N2Sn
689.5
706
142-145
196-199
70
73
(42.24)
(3.15)
(4.04)
47.66
4.26
3.97
(47.74)
(4.16)
(4.01)
50.19
6.27
2.66
5
6
C22H33O6NSn
527
818
143-146
151-153
89
75
(50.23)
(6.30)
(2.70)
52.88
5.63
3.43
C36H46O12N2Sn
(52.96)
(5.60)
(3.50)
35.63
6.46
2.32
7
8
C28H39O6NSn
C28H21O6NSn
605
587
99-101
83
80
(35.69)
(6.50)
(2.36)
207-210
57.34
(54.43)
50.63
3.58
(3.60)
2.95
2.39
(2.45)
5.90
Ligand
C10H7O6N
237
278
80
(50.45)
(2.80)
(5.87)
1H-NMR Spectroscopy
1H-NMR spectra for the synthesized compounds and free acid have been recorded in CDCl3 and
1
DMSO solutions. They are reported in Table 2. H resonance signals of the protons attached to the
phenyl moieties of the ligand have been assigned by their distinct multiplicities, J-values and
comparison with the results obtained from the incremental method [9]. The methyl protons of
dimethyltin derivatives appear as sharp singlets at 0.89 ppm, with well-defined satellites; coupling
constants are included in Table 2. The α-CH2 protons of diethyltin (IV) compounds resonate as a
quartet at 1.52 ppm and have well-resolved coupling constants, while β-CH3 protons resonate as a
triplet at 1.34 ppm with 3J[1H,1H] = 8.2 Hz. The phenyl moieties of the triphenyl- and diphenyltin (IV)
derivatives mostly show a complex pattern and were assigned according to the literature [10]. Despite
the complex pattern of the 1H-NMR spectra of di- and tri-n-butyltin (IV) derivatives, assignments were
possible: a clear triplet due to the terminal methyl group appears in the 0.91-0.92 ppm range [11,12a].
The α-CH2 protons of the di-n-butyltin compound 4 appear as a triplet around 1.72 ppm, with
3J[1H,1H] = 8.0 Hz [12b]. The β-CH2 and γ-CH2 signals appear as multiplets and δ-CH3 as a triplet at
3
0.91 with J[1H,1H] = 10.0 Hz. In the tri-n-butyltin (IV) complex, δ-CH3 resonates as a triplet at 0.92
ppm with 3J[1H,1H] = 8.5 Hz, and γ-CH2 and β-CH2 protons appear as multiplets at 1.33-1.37 ppm and

Molecules 2007, 12
2351
1.61-1.65 ppm, respectively. The α-CH2 appears as a triplet at 1.60 ppm. The methylene protons (CH2)
of the n-octyltin (IV) moiety exhibit a somewhat different behavior, compared with the n-butyl groups
of the respective complexes. The α-CH2, β-CH2 protons give multiplet signals at 1.49-1.53 ppm,
respectively, which are consistent with the values calculated by the incremental method [13]. The γ-
CH2 and γ′-CH2 do not appear in the region. However, the δ′-CH3 protons resonate as a triplet in the
3
range of 0.65 ppm with J[1H,1H] = 7.0 Hz. The protons of the methylenedioxy moiety appear as
2
singlets at 6.00-6.29 ppm and were seen in almost all the compounds. The J[119Sn-1H] coupling
constants and the C-Sn-C angles (Table 3) calculated from 2J[119Sn-1H] values demonstrate that in the
diorganotin (IV) derivatives, tin shows a coordination number greater that four, probably five or six in
non coordinating solvents, while the triorganotin (IV) derivatives show distorted tetrahedral geometry
[13, 14].
1
Table 2. H-NMR data of 3,4-methylenedioxy-6-nitrophenylpropenoic acid and its
organotin (IV) derivatives a, b
Compound
1H No
Ligand
1
2
3
4
5
6
7
8
2
5
7
7.09 (s)
7.52 (s)
6.29 (s)
7.12 (s) 7.00 (s)
7.58(s) 7.54 (s)
6.24 (s) 6.12 (s)
7.96 8.17
(d,15.0) (d,16.5)
6.47 6.33
(d,15.0) (d,15.0)
7.10 (s)
7.58 (s)
7.09 (s)
7.52 (s)
7.01 (s)
7.51 (s)
7.02 (s)
7.33 (s)
7.03 (s)
7.51(s)
6.96 (s)
7.78 (s)
6.00 (s)
7.95
6.11 (s)
7.99
6.14 (s)
8.00
5.98 (s)
7.82
6.14 (s)
8.02
6.11 (s)
7.82
7.82
8
9
(d,15.9)
(d,15.0) (d, 15.0) (d,15.0)
6.47 6.30 6.31
(d,15.0) (d, 15.0) (d,15.0)
(d,16.0) (d, 15.5) (d,16.0)
6.50
6.17 6.31 6.30
(d,15.6)
(d,15.5) (d, 15.5) (d,15.0)
1.49-
0.89 (s)
1.52
1.72
1.6
α
−
−
1.53
(m)
−
−
(q, 10.0)
(t, 8.0)
1.34
1.34-
1.61-
1.49-
1.26-
7.35-
β
γ
−
−
−
−
−
−
−
−
−
−
−
−
−
−
−
(t, 8.2)
1.37 (m) 1.65 (m) 1.53 (m) 1.76 (m) 7.49 (m)
1.34-
1.33-
1.26-
1.76 (m) 7.49 (m)
1.26- 7.35-
1.76 (m) 7.49 (m)
7.35-
−
−
−
−
−
−
1.37 (m) 1.37 (m)
0.91
0.92
δ
(t, 10.0)
(t, 8.5)
γ-γ′
δ′
−
−
1.15 (bs)
−
−
−
0.65
−
−
−
(t, 7.0)
a Chemical shifts (δ) in ppm. 2J[119Sn, 1H]; n[1H, 1H] in Hz where n= 3,4 are listed in square brackets and
parenthesis, respectively. Multiplicity is given s=singlet, bs = broad signal, d = doublet, dd=doublet of doublet,
q = quartet, m = multiplet; b Numbering in accordance with Figures 1- 4.

Molecules 2007, 12
2352
Figure 1. 1H- and 13C-NMR numbering scheme of the ligand L.
O
8
2
3
1
6
9
O
10
OH
7
O
4
NO2
5
Table 3. (C−Sn−C) Angles (o) Based on NMR Parameters of organotin (IV) derivatives
of 3,4-methylenedioxy-6-nitrophenylpropenoic acid
Angle (o)
Compound No.
1J[119Sn, 13C] (Hz)
2J[119Sn, 1H] (Hz)
1J
2J
1
2
3
4
5
6
7
8
-
78
72
-
-
128.4
-
-
121.8
-
-
-
-
-
-
-
360
74
-
108.33
123.9
-
-
-
-
-
-
-
-
-
-
-
13C-NMR Spectroscopy
13C-NMR data recorded in CDCl3 and DMSO solutions of the ligand and its respective mono-, di-
13
and triorganotin (IV) derivatives are given in Table 4. The C-NMR spectral data for the R groups
attached to the tin atom where R = Me, Et, n-Bu, n-Oct, Ph and Cy were assigned by comparison with
n
related analogues as model compounds, combined with the J[119Sn,13C] coupling constants [15, 18].
The assignment of the 13C- resonance associated with the carboxylate ligands is based: (I) on
comparison with the results obtained from incremental methods [14], and (II) on comparison with the
resonance values available in the literature [19]. The positions of the phenyl and olefinic carbon
signals undergo minor variation in the complexes, compared to those observed in the free acid and its
sodium salt. The carboxylate carbon shifts to a lower field region almost in all the complexes,
indicating participation of the carboxyl group (COO) in coordination to tin (IV) [20].

Molecules 2007, 12
2353
Table 4. 13C- and 119Sn -NMR dataa,b of 3,4-methylenedioxy-6-nitrophenylpropenoic
acid and its organotin (IV) derivatives. a, b
13 C No Ligand
1
2
3
4
5
6
7
8
1
2
125.78
107.19
151.73
148.73
106.90
138.87
103.74
142.98
122.67
167.03
126.36
105.93
153.10
150.18
104.86
141.35
104.30
141.25
123.10
187.87
126.96
107.64
152.05
149.24
105.18
142.67
103.06
143.92
120.78
175.39
127.11
107.65
152.62
148.78
105.01
143.81
103.45
143.81
124.54
186.84
127.86
107.46
151.90
148.65
105.60
138.96
103.26
143.13
125.82
171.62
127.93
107.37
151.90
148..63
105.56
139.03
103.25
143.06
124.54
170.90
15.24
[340, 360]
26.79
27.04
13.64
-
127.25
107.68
153.09
150..29
105.75
141.52
104.68
144.34
122.49
174.75
128.07
107.39
151.87
148.55
105.56
138.78
103.20
143.07
124.85
170.70
127.53
107.40
151.94
148.87
105.63
140.88
103.20
143.10
122.77
171.96
3
4
5
6
7
8
9
10
α
-
4.34
19.17
22.70
22.70
25.43
14.11
130.22
β
γ
δ
α'
β'
γ'
δ'
-
-
-
-
-
-
-
-
-
-
-
-
-
-
14.11
25.57
26.37
23.27
33.56
29.72
31.34
30.29
32.51
25.90
29.37
137.13
-
-
-
-
-
-
29.19
26.83
31.95
128.97
14.11
13.67
28.85
128.97
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
-
119 Sn
-
-112.63 -150.68 -150.03 -156.43
+116.10
-175.00
45.64
-112.45
a Chemical shifts (δ) in ppm. nJ[117/119Sn, 13C]; nJ[119Sn, 13C]; in Hz are listed in square brackets; b
Numbering in accordance with Figures 1- 4.
Figure 2. Numbering scheme for groups attached to tin.
α
CH3
β
CH3
α
CH2
γ
CH2
α
CH2
Sn
β
CH2
δ
CH3
Sn
Sn
′
′
γ
CH2
α
CH2
α
CH2
γ
CH2
′
δ
CH3
′
CH2
β
CH2
β
δ
CH2
Sn
γ
γ
γ
β
β
α
α
Sn
δ
δ
Sn
β
γ
β

Molecules 2007, 12
2354
Figure 3. Structural diagrams of triorganotin (IV) Compounds.
R
R
R
R
R
O
O
O
Sn
O
O
Sn
R
O
R
Sn
R
R
R
O
R
R
n
Figure 4. Structural diagram of diorganotin (IV) Compounds.
R
R
O
O
O
Sn
R
R
O
119Sn-NMR Spectroscopy
The possibility of detecting the presence of coordinatively different organotin (IV) moieties was
explored by acquisition of 119Sn-NMR spectra for all the investigated compounds. The 119Sn- chemical
shifts usually cover a range, quoted relative to tetramethyltin, with increasing coordination number of
tin producing a large upfield shift for δ(119Sn). The 119Sn-NMR spectra were recorded in CDCl3
solutions (a non-coordinating solvent). It is reported [20, 21] that in alkyltin carboxylates, tetra-
coordinated tin has δ(119Sn) values ranging from about +200 to -60 ppm, and penta-coordinated tin
from -200 to -400 ppm. A single resonance peak of 119Sn chemical shift values obtained for the
triorganotin (IV) derivatives lies in the range expected for a tetrahedral geometry. 119Sn chemical shift
values of diorganotin compounds were observed in the -112.63 to -175.0 ppm range. It appears from
the shift values that a weakly hexacoordinated tin generates a deformed octahedron or a skewed
trapezoidal bipyramid with four strong and two somewhat weaker bonds, as indicated by the upfield
119Sn chemical shift values [14, 16, 20, 21].
In order to gain further information about the possible coordination geometries in solution, close
1
2
examination of the J[119Sn,13C] and J[119Sn,1H] coupling constants was undertaken as indicators of

Molecules 2007, 12
2355
2
1
structural details. As reported by Nadvornik et al. [13, 22a], J [119Sn, H] in n-Bu3Sn (IV)
carboxylates changes very little with a change in coordination number, being the smallest among such
2
couplings. In the case of diorganotin compounds 1 and 2, the J[119Sn,1H] values are 78 and 72 Hz,
respectively, which indicate that tin displays hypervalent behavior (valence higher than four) in these
compounds. .1J[119Sn,13C] is, on the other hand, quite useful for making predictions about the geometry
1
around the tin atom. In case of triorganotin (IV) carboxylates, J[119Sn,13C] values ranging between
327 and 387 Hz for tetra-coordinated compounds and between 442 and 509 Hz for penta-coordinated
1
ones were observed [22b]. For the tri-n-butyltin (IV) derivative the J[119Sn,13C] values were 360 Hz
and based on the Holeček and Lycka equation [20, 21], corresponds to a quasi-tetrahedral geometry in
CDCl3 solution. The geometric data calculated, as just described, are consistent with tetrahedral
geometries for the triorganotin (IV) species [20], i.e. monomers in solution. Unfortunately,
2
1J[119Sn,13C] values are not observed in any compound, but from the 119Sn values and J[119Sn,1H], it
can be predicted that diorganotin (IV) species, possess skew-trapezoidal bipyramidal geometries with
the lower apparent coordination number arising from the asymmetric coordination mode of the
carboxylate ligand [23, 24].
Mass Spectrometry
The mass spectra of all the synthesized compounds 1-8 were determined using the electrospray
ionization (ESI) technique. The molecular ion [M]+ peak is not usually observed in case of
organometallic compounds while using the conventional electron ionization (EI) or chemical
ionization (CI) mass spectral techniques. However in case of ESI, not only the molecular ion peak
[M]+, but also peaks due to aggregation of fragments with [M]+ or [M+H]+ are encountered [25]. These
fragment aggregates have molecular weights which is greater than [M] + (Table 5).
The 1st order +ve ESI-MS mode is more informative than the –ve mode because it can also provide
information about the polymeric species and hence it is the more preferred for organotin species [26].
The neutral loses observed in ESI-MS are attributed to loss of CO2, methane, ethane, butane and
butene. The presence of more than one tin isotope in a molecule causes a wide distribution of the total
signal among the many fragment aggregates, which decreases the relative intensity of the centroid
peak. The presence or absence of a tin atom in individual ion can easily be recognized on the basis of
the 10 characteristic natural tin isotopes, with the most abundant being the 120Sn isotope.
The base peak in these compounds whether di- or triorganotin (IV) compounds derives from the
different fragmentation patterns. All the investigated compounds were found to possess a base peak
resulting from the loss of the ligand acid (HL), the R group or the ligand moiety itself.

Molecules 2007, 12
2356
Table 5. Mass fragmentation patterns of organotin (IV) derivatives of
3,4-methylenedioxy-6-nitrophenylpropenoic acid.
Fragment, m/z (%)
1
2
3
4
5
6
7
8
[C23H20N2 [C22H16N2O [C24H19ClN2O [C32H37N2O [C30H50N [C40H54N2 [C28H39N
[C22H15N
O6Sn]+
O12Sn]+
12Sn]+
12Sn]+
12Sn]+
O6Sn]+
O12Sn]+
O6Sn]+
636 (8.5)
620 (3.5)
682.5 (2.5)
761(16.5) 640(7.3)
874 (9.5)
605 (2.5) 509(14.9)
[C22H16N2 [C15H12NO8 [C20H10ClN2O [C27H26N2O [C22H31N [C34H40N2 [C27H36N
[C21H13N
O4Sn]+
O12Sn]+
Sn]+
12Sn]+
12Sn]+
O6Sn]+
O12Sn]+
O4Sn]+
620 (17.3)
454 (41.6)
625.5 (11.3)
690 (7.6)
525 (6.4)
788 (16.2) 558 (7.3)
463 (3.3)
[C21H15N2
O10Sn]+
[C10H13O4S
n]+
[C14H14Cl
NO6Sn]+
[C24H19N2O
12Sn]+
[C18H23
NO6Sn]+
[C28H28N2 [C22H27N
[C15H8
NO4Sn]+
O10Sn]+
O6Sn]+
575 (3.4)
317 (17.1)
447.5 (32.7)
647 (19.9) 469 (23.6) 672 (26.3) 521 (33.4) 386 (27.1)
[C19H9N2
O10Sn]+
[Sn]+
[C5H11Sn]+
191 (41.5)
[C8H17N2O1 [C14H14N
2Sn]+ O6Sn]+
[C18H22N
O6Sn]+
[C16H16N [C9H6O2S
O6Sn]+ n]+
120 (3.7)
545 (37.7)
453 (25.2) 412 (12.7) 468 (8.7) 438 (17.1) 266 (72.4)
[C11H6NO
6Sn]+
[C4H9]+
[C4H8Sn]+
176 (6.9)
[C4H7Sn]+
175 (79.7)
[C7H10O2
Sn]+
[C9H16O2
Sn]+
[C9H12O2 [C11H6NO
Sn]+ 6Sn]+
57 (100)
368 (60.9)
[C4H4O2S
n]+
246 (40.7) 276 (100) 272 (63.4) 368 (60.9)
[C3H8Sn]+
164 (18.8)
[C2H3Sn]+
145 (100)
[C4H9]+
[C4H9]+
[C4H6O2S
n]+
[C6H5]+
-
-
57 (100)
57 (76.3)
77 (100)
204 (100)
206 (100)
[C4H9]+
-
-
-
-
-
-
57 (100)
Biological Activity
All the synthesized compounds and their acid ligand were screened for their antitumor activity and
cytotoxicity using standard in vitro biocidal screening tests.
Cytotoxicity
Bioactive compounds are often toxic to shrimp larvae. Hence, shrimp larvae have been extensively
used as rapid and simple preliminary test for cytotoxicity [11a, b]. All the synthesized compounds and
ligand were screened for cytotoxicity by brine-shrimp bioassay lethality method [11b] and the results
are summarized in Table 6. The LD50 data shows that all the compounds even the ligand acid are toxic
with LD50 values in the range 0.26 to 581.23 μg/mL for organotin derivatives and 975.24 μg/mL for
the ligand L Tri-organotin derivatives are more toxic than di-organotin derivatives, mono-organotin

Molecules 2007, 12
2357
derivatives and the ligand acid [27]. Compound 3 (a monoorganotin derivative) has been found to be
the least toxic and compound 5 (a triorganotin derivative) was the most toxic of all the synthesized
organotin derivatives [4, 5].
Table
6.
Cytotoxicity
data
of
organotin
(IV)
derivatives
of
3,4-methylenedioxy-6-nitrophenylpropenoic acid.
Compound No
LD50 µg/mL
1
2
-
130.98
3
581.23
33.39
0.26
4
5
6
191.64
0.40
7
8
0.56
Ligand
975.24
Against brine shrimps (in vitro); Data is based
on mean value of 3 replicates of each for 1000,
100 and 10 μg/mL concentrations; - = activity
not determined.
Antitumor Activity
For antitumor activity, the crown gall tumor inhibition assay (potato disc assay) [11b] was
performed for all the synthesized compounds using Agrobacterium tumefaciens (strain At10). Crown
gall is a neoplasmic disease of plants induced by Agrobacterium tumefaciens due to its Ti (tumor
inducing) plasmid [11b]. Since the mechanism of tumor induction is similar to that seen in animals,
this assay was used to evaluate the antitumor activity of these compounds. All the compounds showed
significant levels of tumor inhibition, as shown in Table 7. Activity observed for synthesized organotin
series was more than that of their parent acid, and triorganotin derivatives showed more tumor
inhibition than mono- and diorganotin derivatives. Furthermore, compounds 2, 5 and 8 showed 100%
tumor inhibition. The bioassays results reflect the conclusion that the activity are in the following
order: [RSnL2Cl] < [R2SnL2] < [R3SnL]. Further, it has been observed that the activities of these
mono-, di and triorganotin (IV) complexes are comparable with those reported earlier [2, 4, 5, 27].

Molecules 2007, 12
2358
Table 7. Antitumor activities of organotin (IV) derivatives of
3,4-methylenedioxy-6-nitrophenylpropenoic acid a, b, c, d
.
Average number of
tumors ± SE
2.8±0.89
% inhibition of
Compound No.
tumors
62.66
100
1
2
3
4
5
6
7
0.0±0.0
3.9±1.02
48
1.0±0.36
86.66
100
0.0±0.0
5.3±0.84
29.33
36
4.8±1.13
8
0.0±0.0
4.02±1.04
7.5±1.13
100
44
-
Ligand
(-)ive control
a) Potato disc antitumor assay, concentration: 1000 μg/mL in
DMSO; b) More than 20% tumor inhibition is deemed
c
significant; ) Percent Inhibition of tumors = [100 ─ (Number of
tumors for sample/Number of tumors for control)] × 100;
d) Data represents mean value of 15 replicates.
Experimental
General
Melting points were determined in a capillary tube using a MPD Mitamura Riken Kogyo (Japan)
electrothermal melting point apparatus. Multinuclear NMR (1H-, 13C- and 119Sn) spectra were recorded
1
on a Bruker ARX 300 MHz-FT-NMR spectrometer using CDCl3 as an internal reference for [δ H
(CDCl3) = 7.25 and δ 13C (CDCl3) = 77.0]. 119Sn-NMR spectra were obtained with Me4Sn as an
external reference [(119Sn) = 37.290665] [10]. Chemical shifts (δ) are given in ppm and coupling
constants J are given in Hz. The multiplicities of signals in 1H-NMR are given with chemical shifts; (s
= singlet, d = doublet, dd = doublet of doublet, t = triplet, q = quartet, m = multiplet). Mass spectral
data (ESI mode) were taken on a LCQ Finnigan MAT ion trap (USA). The m/z values were evaluated
assuming that H = 1, C = 12, N = 14, O = 16, Cl = 35.5 and Sn = 120.
Synthesis
Different mono-, di- and triorganotin (IV) carboxylates were prepared by treating the
corresponding ligand acid with organotin oxide/hydroxides or its sodium salt with organotin chlorides,
adopting various standard procedures [28].

Molecules 2007, 12
2359
Synthesis of the Ligand L
The ligand acid 3,4-methylenedioxy 6-nitrophenylpropenoic acid was synthesized by dissolving 6-
nitropiperonal (6.5 g, 0.033 mol) and malonic acid (7.5 g, 0.072 mol) in a mixture of pyridine (15 mL)
and piperidine (0.25 mL) in a 100 mL round bottom flask and heating under reflux for 1 hour in an oil
bath (Scheme 1). A rapid evolution of carbon dioxide took place. The reaction was completed by
boiling the reaction mixture for 5 minutes, then it was cooled and poured into an excess of water
containing enough hydrochloric acid to combine with the pyridine. The 3,4-methylendioxy-6-
nitrophenylpropenoic acid was filtered off, washed with a little water and dried. The acid obtained was
recrystallized from glacial acetic acid. The synthesized ligand acid is a crystalline solid, possessing a
sharp melting point. And the yield is typically in the 67-86 % range.
Scheme 1.
O
O
O
O
O
O
H
OH
i)C5H5N ii) trace of C5H11N
CH2(CO2H)2
+
Reflux at 95-100 ,Acidification
NO2
NO2
Synthesis of Complexes
Procedure-I
The sodium salt RCOONa, was prepared by adding dropwise an equimolar amount of sodium
hydrogen carbonate dissolved in distilled water to the ligand acid (RCOOH) dissolved in ethanol. The
clear solution obtained was concentrated under reduced pressure. The dry sodium salt R'COONa (8
mmol), was mixed with diorganotin dichloride (4 mmol) or triorganotin chloride (8 mmol) in 2:1 and
1:1 ratios, respectively, in dry toluene contained in a 250 mL two necked round bottom flask. The
mixture was refluxed for 9-10 hr. After refluxing a turbid solution was obtained, which was then left
overnight at room temperature. The sodium chloride collected at bottom was filtered off and the
solvent was evaporated by rotary evaporation. The resulting solid mass was recrystallized from
chloroform and n-hexane mixture (4:1). Compounds 1, 2 and 5 were prepared by this method in yields
of 86, 67 and 89%, respectively.
Procedure-II
The ligand acid, R'COOH (12 mmol) and diorganotin oxide (6 mmol), or triorganotin hydroxide
(12 mmol) in 2:1 and 1:1 ratios, respectively, were suspended in dry toluene (100 mL) in a single
necked round bottom flask (250 mL), equipped with a Dean-Stark apparatus. Anhydrous conditions are
required to avoid the oxidation of metal atom before the reaction. However, water is produced as a

Molecules 2007, 12
2360
secondary product and removed by the Dean-Stark apparatus. After the reaction is complete the
products (tin carboxylates) are stable to moisture and oxygen. The mixture was refluxed for 8-10 hours
and water formed during the condensation reaction was removed at regular intervals. It was then
cooled to room temperature and solvent was removed by rotary evaporator. The solid obtained was
recrystallized from chloroform containing a few drops of n-hexane. Compounds 3, 4, 6, 7, 8 were
prepared by this method. Yield of these compounds were 70, 73, 75, 83, and 80%, respectively.
Biological Activity
Cytotoxicity
The cytotoxity was studied by the brine-shrimp lethality assay method [11b]. Brine-shrimp
(Artemia salina) eggs (San Francisco Bay Brand, Inc, New York, CA94560, USA) were hatched in
artificial sea water (40 g sea salts/L) at room temperature (22-29 ºC). Sea salts were obtained from
from Instant Ocean, Inc, 8141, Tyler Boulevard, Mentor, OH44060, USA. After two days these
shrimps were transferred to vials (10 shrimps per vial) containing artificial sea water (5 mL) with
1000, 100 and 10 μg/mL final concentrations of each compound taken from their stock solutions of 20
mg/mL in DMSO. After 24 hours number of surviving shrimps was counted. Data was analyzed with a
finny computer programme (Probit analysis) to determine LD50 values.
Antitumor Activity
For antitumor activity crown gall tumor inhibition assay (Potato Disc Assay) [11b] was performed
for all these synthesized compounds. Potato discs (0.5 cm thickness) were obtained from surface
sterilized potatoes by using metallic cork borer (8 mm) and special cutter under complete aseptic
conditions. These potato discs were then transferred to petri plates each containing 1.5 % agar (25 mL,
1.5 g agar/100 mL distilled water). Five potato discs were placed on each plate and three plates were
used for each test sample along with same number of plates for control. Each compound (10 mg) was
dissolved in DMSO (1 mL) in separate test tubes as a stock solution. Then stock solution of the test
sample (0.5 mL, 10 mg/mL) was added to a broth culture of Agrobacterium tumefaciens (2 mL, At10,
a 48 hours culture containing 5×109 cells/mL) and autoclaved distilled water (2.5 mL) to give 1000
μg/mL final concentration. These cultures (50 μL) were poured onto each potato disc. The petri plates
were incubated at 28 ºC, the lids being taped down with Parafilm. After 21 days incubation, the
number of tumors was counted with the aid of dissecting microscope after staining with Lugol’s
solution (5% I2, 10% KI in distilled water).
Acknowledgements
The authors gratefully acknowledge financial grants from the Higher Education Commission,
Islamabad, Pakistan.

Molecules 2007, 12
2361
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