Solid-phase synthesis and biological evaluation of a parallel library of 2,3-dihydro-1,5-benzothiazepines

Article abstract of DOI:10.1016/j.bmc.2008.07.009

Solid-phase synthesis of a parallel library of 3′-hydroxy-2,3-dihydrobenzothiazepines has been carried out through [4+3] annulation of α,β-unsaturated ketones with aminothiophenol, using Wang resin as solid support. The synthesized compounds were evaluated for their potential as antibacterial, tumor inhibitors as well as acetyl- and butyrylcholinesterase inhibitors. None of the compounds showed any significant antibacterial activity. However, quite a few compounds showed significant potential as crown gall tumor inhibitors. These results reflect a strong exploratory potential in search of new benzothiazepines as source of anticancer agents. The results of the inhibition of cholinesterase revealed that benzothiazepines have a greater potential as butyrylcholinesterase inhibitors as compared to acetylcholinesterase. Moreover, the substitution of hydroxy group at C-3 in ring A led to increased activity when compared to unsubstituted- and 2′-OH substituted benzothiazepines.

Full text of DOI:10.1016/j.bmc.2008.07.009

Bioorganic & Medicinal Chemistry 16 (2008) 7691–7697
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Solid-phase synthesis and biological evaluation of a parallel library
of 2,3-dihydro-1,5-benzothiazepines
a
b
b
a
a
Farzana Latif Ansari a, , Fatima Iftikhar , Ihsan-ul-Haq , Bushra Mirza , Mohammad Baseer , Umer Rashid
*
a Department of Chemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan
b Department of Biochemistry, Quaid-i-Azam University, Islamabad 45320, Pakistan
a r t i c l e i n f o
a b s t r a c t
Solid-phase synthesis of a parallel library of 30-hydroxy-2,3-dihydrobenzothiazepines has been carried
out through [4+3] annulation of ,b-unsaturated ketones with aminothiophenol, using Wang resin as
solid support. The synthesized compounds were evaluated for their potential as antibacterial, tumor
inhibitors as well as acetyl- and butyrylcholinesterase inhibitors. None of the compounds showed any
significant antibacterial activity. However, quite a few compounds showed significant potential as crown
gall tumor inhibitors. These results reflect a strong exploratory potential in search of new benzothiaze-
pines as source of anticancer agents. The results of the inhibition of cholinesterase revealed that ben-
Article history:
Received 22 May 2008
Revised 2 July 2008
Accepted 3 July 2008
Available online 9 July 2008
a
Keywords:
Solid-phase synthesis
1,5-Benzothiazepines
Potato disk tumor inhibitors
Cholinesterase inhibitors
Antibacterial
zothiazepines have
a greater potential as butyrylcholinesterase inhibitors as compared to
acetylcholinesterase. Moreover, the substitution of hydroxy group at C-3 in ring A led to increased activ-
ity when compared to unsubstituted- and 20-OH substituted benzothiazepines.
Ó 2008 Elsevier Ltd. All rights reserved.
1. Introduction
dro-1,5-benzothiazepines were synthesized by using the most
widely accepted strategy that involved a [4+3] annulation of
For more than a century heterocycles have constituted one of
the largest areas of research in organic synthesis.1 Over the last
two decades, synthesis of N and S containing heterocyclic com-
pounds especially benzothiazepines retained the interest of
researchers due to the unique structural properties and broad
spectrum of biological activities of these compounds.2
a,b-unsaturated ketones (chalcones) with o-aminothiophenol
(o-ATP) under acidic conditions.17 In continuation to these stud-
ies, synthesis of a parallel library of 2,3-dihydro-1,5-benzothiaze-
pines with a 30-hydroxy substitution on ring A precursors has
been carried out with the objective of making a comparison with
the compounds previously synthesized.
Benzothiazepines have their role in the treatment of cardio-
vascular disorders,3 as Ca2+ channel blockers,4 inhibitors of
HIV-I integrase,5 antibiotics,6 muscle relaxants,7 and cytotoxic
agents.8 They are also known to have antimicrobial9–11 and anti-
hypertensive activities12 besides being used in the treatment of
diabetes.13 Keeping in view this broad spectrum of biological
activity associated with these compounds, we have reported ear-
lier the synthesis of some 2,3-dihydrobenzothiazepines with
unsubstituted ring A and 20-OH substitution on ring A. These
As observed earlier, classical or solution-phase synthesis of
these compounds involves various problems, related to their
yield and purity which in turn is affected by the substitution
on both rings A and B of chalcones. The decomposition of o-
ATP with the passage of time is another factor for low yield
and less purity of the desired products. In the recent years so-
lid-phase synthesis (SPS) has emerged as a powerful synthetic
tool that not only ensures greater purity but also results in im-
proved yield mainly due to a decrease in side products.10 Easier
work up, less time consumption, and easy purification are some
extra advantages of SPS that has attracted the attention of
researchers towards the use of this technique for the synthesis
of these heterocyclic compounds. Wang resin has usually been
employed when an OH group is available to anchor the solid
support. It is an acid-labile p-alkoxybenzylalcohol resin and its
applications to the synthesis of various types of compounds
via benzyl-ether linkage under Mitsunobu conditions18 are well
known. Bio-evaluation of the synthesized benzothiazepines in-
volved their screening as antibacterial, potato disk tumor inhib-
itors and as cholinesterase inhibitors.
compounds were evaluated against
a number of bioassays,
which revealed that the compounds have potential as enzyme
inhibitors such as acetyl- and butyryl cholinesterase, urease
and
a-glucosidase, and quite a few compounds were found to
be promising DPPH-free radical scavengers.14 Molecular docking
studies on the compounds that were identified as good inhibi-
tors of cholinesterase and urease inhibitors were also carried
out to investigate the ligand-enzyme interaction.15,16 2,3-Dihy-
* Corresponding author. Tel.: +92 512875587; fax: +92 512873869.
E-mail address: fla_qau@yahoo.com (F.L. Ansari).
0968-0896/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2008.07.009

7692
F. L. Ansari et al. / Bioorg. Med. Chem. 16 (2008) 7691–7697
(NMM) with Wang resin in the presence of triphenylphosphine
2. Results and discussion
(TPP) and diisopropylazodicarboxylate (DIAD) in reaction mixture
for 48 h. The resin-bound acetophenone was then reacted with
different aryl- and heteroaryl aldehydes under Claisen-Schmidt
condensations which led to resin-bound chalcones as shown in
Scheme 1. Without getting involved in the isolation and purifica-
tion of chalcones, they were treated directly with o-ATP in DMF
A parallel library of twenty-five 2,3-dihydro-1,5-benzothiaze-
pines (27–52) has been synthesized using chalcones (1–26) as pre-
cursors which in turn were synthesized through Claisen-Schmidt
condensation of 3-hydroxy acetophenone with 25 different aryl
aldehydes.
under
inert
atmosphere,
which
led
to
resin-bound
The generation of chalcone intermediates in good yield and high
purity was critical for the success of the synthesis of benzothiaze-
pines as the use of impure intermediates was detrimental to 1,5-
benzothiazepine synthesis.10 Analytical results showed that the
desired chalcones as well as 1,5-benzothiazepines were easily syn-
thesized in solution phase in high yield and acceptable purity as
long as the aromatic aldehyde did not contain any electron-donat-
ing group on ring B by reducing the electrophilicity of the carbonyl
and alkenic groups in chalcones toward nucleophilic attack by o-
ATP.
dihydrobenzothiazepines. The cleavage of the resin was done
through 50% TFA in CH2Cl2 which resulted in free 2,3-
dihydrobenzothiazepines (27–52). Table 1 shows a comparison
of the yield and purity of some selected compounds which were
synthesized both in solution and solid phase where a significant
difference in yield was observed.
3. Biological screening
Furthermore, the reactions with heteroaryl-substituted chal-
cones seem to undergo ring opening or complex decomposition
reactions thereby reducing the yield and purity of the products.
Moreover, attempt to catalyze these reactions by piperidine was
also not successful. Therefore, there existed a dire need to look
for an alternative synthetic technique. The problem of low yield
in the case of hydroxy-substituted aldehydes and ketones was cir-
cumvented by protecting the hydroxy group by tetrahydropyran;
however, the purity of the products could not be improved.
Although the protecting group strategy led to high yield at the ex-
pense of an additional synthetic step, the problem of purity re-
mained the same.
Therefore, solid-phase synthetic (SPS) strategy was chosen for
the synthesis of 2,3-dihydro-1,5-benzothiazepines using 30-hydro-
xy chalcones as precursors. The linking of 3-hydroxy group to
Wang resin has been carried out under Mitsunobu conditions18
by shaking acetophenone dissolved in N-methylmorpholine
The synthesized compounds were tested for the following bio-
logical activities.
Table 1
Comparison of yield and purity through solution- and solid-phase synthesis of some
selected benzothiazepines
Compound
R
Yield (%)
Solution phase
Puritya (%)
Solid phase
29
30
33
35
39
42
43
3-Cl C6H4
4-Cl C6H4
4-F C6H4
3-OH C6H4
4-OMe C6H4
3-NO2 C6H4
4-NO2 C6H4
65
80
81
51
47
78
83
80
87
93
70
58
83
90
85
83
91
75
80
>90
ꢀ90
a
Solid phase.
O
O
O
OH
'3
HO
'3
CH3
O
O
DIAD, TPP,NMM
48 hrs.
CH3
A
A
O
Ring B
CH
KOH 6 equiv.
SH
O
N
S
i)
'3
O
O
NH2
Ring B
HO '3
2
4
A
Ring B
3
ii)
A
50% TFA
Ring B
1, 27 = C6H5
Ring B
14, 40 = 3'',4''-OMe2C6H3
2, 28 = 2''-Cl C6H4
3, 29 = 3''-Cl C6H4
4, 30 = 4''-Cl C6H4
5, 31 = 2''-F C6H4
6, 32 = 3''-F C6H4
7, 33 = 4''-F C6H4
8, 34 = 4''-Br C6H4
9, 35 = 3''-OH C6H4
10, 36 = 4''-OH C6H4
11, 37 = 2''-OMe C6H4
12, 38 = 3''-OMe C6H4
13, 39 = 4''-OMe C6H4
15, 41 = 4''-OH-3''-OMe C6H3
16, 42 = 3''-NO2C6H4
17, 43 = 4''-NO2 C6H4
18, 44 = 4''-MeC6H4
19, 45 = 4''-NMe2 C6H4
20, 46 = Pyridin-2''-yl
21, 47 = Pyridin-3''-yl
22, 48 = Pyridin-4''-yl
23, 49 = Thien-2''-yl
24, 50 = 2''-Thienyl-5''-Me
25, 51 = 2''-Thienyl-5''-Br
26, 52 = Pyrrol-2''-yl
Scheme 1. Solid-phase synthesis of 2,3-dihydro-1,5-benzothiazepines (27–52).

F. L. Ansari et al. / Bioorg. Med. Chem. 16 (2008) 7691–7697
7693
Table 3
3.1. Antibacterial assay
Results of PDT bioassay conducted on 2,3-dihydro-1,5-benzothiazepines
All synthesized compounds have been tested for their potential
as antibacterial against five bacterial strains namely Bordetella
bronchiseptica, Micrococcus leuteus, Escherichia coli, Enterobac-
ter aerogenes, and Salmonella setubal by using Agar well diffu-
sion method. However, none of the tested compounds
showed any significant antibacterial activity against any bac-
terial strain.
Compound
R
% inhibition (ppm)
IC50
1000
100
97.8
10
27
28
29
30
31
33
34
35
36
37
38
39
40
41
42
43
44
45
47
48
49
51
C6H5
2-Cl C6H4
3-Cl C6H4
4-Cl C6H4
2-F C6H4
4-F C6H4
4-Br C6H4
3-OH C6H4
4-OH C6H4
2-OMe C6H4
3-OMe C6H4
4-OMe C6H4
3,4-OMe2 C6H3
3-OMe, 4-OH C6H3
3-NO2 C6H4
4-NO2 C6H4
4-Me C6H4
4-NMe2 C6H4
3-Pyridyl
100
95.7
93.6
74.4
91.4
40.4
85.1
100
100
98.5
100
100
100
95.7
100
65.9
87.2
100
51.0
100
100
92.8
—
72.3
74.4
68.0
51.93
31.9
25.5
21.2
36.1
27.6
81.42
67.142
57.4
70.2
36.1
42.5
12.7
65.9
38.2
29.7
55.1
87.2
80.3
—
5.41
6.94
14.01
9.0
295.5
1232
61.5
179.3
237.9
93.6
80.8
68.0
40.4
34.0
63.8
48.9
48.9
87.1
84.2
78.7
82.7
40.4
57.4
29.7
72.3
44.6
38.2
80.0
The benzothiazepines 29, 41, 43, and 45 showed weak activity
against a few bacterial strains as shown in Table 2.
7.06
3.2. Potato disk tumor inhibition assay (PDT)
3.28
4.69
8.23
The efforts made in minimizing the screening process to hasten
the pace of drug discovery are the development of bench top assays
which shortlist potential candidates that could be targeted for
more advanced screening. One of these bench top assays is the
crown gall tumor assay also known as potato disk tumor bioassay
as developed by McLaughlin. It is acceptable as a preliminary gen-
eral screen for antineoplastic activity of different compounds and
plant extracts regardless of the mode of inhibitory action on tumor
growth. Crown gall is a neoplastic plant disease caused by Agrobac-
terium tumefaciens.19 In this bioassay the ability of the test com-
pounds to inhibit the tumors induced by A. tumefaciens in model
systems such as potato tuber disks is evaluated. The rationale for
employing this bioassay rests on the fact that the tumorogenic
mechanism induced by A. tumefaciens in plants is in many ways
similar to that of animals.20 The use of this bioassay has resulted
in many shortlists of test compounds with anticancer activity
and has helped with the discovery of the novel anticancer com-
pounds. The synthesized compounds have been tested for their po-
tato disk tumor inhibition activity. The inhibition was studied at
1000, 100, and 10 ppm concentrations. These results along with
IC50 values (McLaughlin and Rogers, 1998) are shown in Table 3.
It is evident from the data in Table 3 that the OMe-substituted
compounds (37–40) were found to be the most promising in the
series with IC50 values varying from 3.28 to 8.23. Moreover, ben-
zothiazepines substituted with a 4-pyridyl and thiophene ring
(48, 49, and 51) were also detected to be strong inhibitors of crown
gall tumor with IC50 values 6.4, 4.8, and 11.2, respectively. The sub-
stitution of a chlorine atom on ring B (28–30) also resulted in an
increased tendency of tumor inhibition. These compounds could
therefore be targeted for more advanced screening. Studies on po-
247.3
26.2
331.35
30.7
192.5
926.2
6.1
4-Pyridyl
2-Thienyl
100
91.4
—
4.8
11.2
2-Thienyl-5-Br
Vincristinea
0.003
a
Concentrations at which standards is tested are 0.1, 0.01, 0.001, 0.000001, and
0.0000001.
Table 4
Results of PDT bioassay carried on some selected chalcones
Compound
R
% inhibition (ppm)
100
IC50
1000
10
2
3
7
2-Cl C6H4
3-Cl C6H4
4-F C6H4
3-OH C6H4
2-OMe C6H4
3.4-OMe2 C6H3
4-NMe2 C6H4
100
100
83.0
100
100
100
100
46.8
74.4
44.6
100
100
77.1
88.5
36.17
51.0
32.0
57.4
76.5
60.0
75.5
40
12.1
9.6
12.0
3.1
5.8
2.8
9
11
14
19
tato disk bioassay were also extended to some chalcones and it was
detected that they possessed better inhibitory potential than the
corresponding benzothiazepines as shown in Table 4.
IC50 values of chalcones varied from 2.8–40.0 ppm and chal-
cones 11, 14, and 19 were found to have significantly inhibited
the growth of tumors.
Table 2
Antibacterial activity of 2,3-dihydro-1,5-benzothiazepines (zone of inhibition, mm)
Compound
R
1
2
3
4
5
3.3. Cholinesterase inhibition assay
27
28
29
30
31
33
34
35
36
37
39
40
41
42
43
44
C6H5
2-Cl C6H4
3-Cl C6H4
4-Cl C6H4
2-F C6H4
4-F C6H4
4-Br C6H4
3-OH C6H4
4-OH C6H4
2-OMe C6H4
4-OMe C6H4
3,4-OMe2 C6H3
3-OMe, 4-OH C6H3
3-NO2 C6H4
4-NO2 C6H4
4-Me C6H4
4-NMe2 C6H4
3-Pyridyl
—
—
—
—
—
—
—
—
—
—
—
—
9.5
—
10.5
—
11.0
—
—
—
—
—
—
—
—
—
—
—
—
—
10.0
—
10.0
—
9.0
—
—
—
—
—
—
—
—
—
—
—
—
—
10.0
—
9.0
—
9.0
—
—
—
11.5
—
—
—
—
—
—
—
—
—
10.0
10.0
13.0
—
10.0
—
—
—
—
—
—
—
—
—
—
—
—
—
11.0
Nil
Nil
—
11.0
—
It is an established fact that many life-saving dugs perform their
function through enzyme inhibition pathways. The discovery of
new enzyme inhibition has led to many interesting discoveries in
drug development. We have reported earlier, a number of 2,3-
dihydro and 2,3,4,5-tetrahydrobenzothiazepines as inhibitors of
cholinesterase,14 having potential to develop into drugs against
Alzheimer’s disease.21 Continuing our on-going research for new
inhibitors of cholinesterases, some of the synthesized benzothiaze-
pines were screened for their potential as cholinesterase inhibitors
and quite a few compounds were found to have valuable butyr-
ylcholinesterase inhibitory characteristic as shown in Table 5. It
is noteworthy that the same set of compounds did not show signif-
icant activity against acetylcholinesterase with the exception of
compound 49.
45
47
49
Standard
2-Thienyl
Roxithromycin
—
14.0
—
13.0
—
12.0
—
15.0
—
12.0
We have reported earlier the cholinesterase inhibitory potential
of some benzothiazepines with ring A as unsubstituted and with a
hydroxyl group at C-2 position.14 However, these compounds were
(1) E. coli, (2) B. bronchiseptica, (3) S. setubal, (4) M. leuteus, (5) E. aerogenes.

7694
F. L. Ansari et al. / Bioorg. Med. Chem. 16 (2008) 7691–7697
Table 5
5.1. Solution-phase synthesis
In vitro quantitative inhibition of acetylcholinesterase (AChE) and butyrylcholinest-
erase (BChE)
A solution-phase synthesis of 2,3-dihydro-1,5-benzothiazepines
was carried out using chalcones as precursors which in turn were
synthesized through Claisen-Schmidt condensation of 3-hydroxy
acetophenone with different substituted aryl and heteroaryl
aldehydes.
Compound
R
IC50
(l
M)
BChE
AChE
27
28
Phenyl
2-Cl C6H4
40.0 0.7
—
1421.0 0.01
—
29
30
31
3-Cl C6H4
4-Cl C6H4
2-F C6H4
—
29.4 1.6
79.6 6.8
—
18.8 1.5
70.6 1.5
5.1.1. General method for the synthesis of chalcones (1–26)
All chalcones of the parallel library were synthesized by the fol-
lowing literature procedure22: A 4.0-M solution of sodium hydrox-
ide (30 ml, 4.8 mmol) was added to precooled rectified spirit
35
36
37
40
41
42
43
44
3-OH C6H4
4-OH C6H4
2-OMe C6H4
3,4-diMe C6H3
3-OMe, 4-OH C6H3
3-NO2 C6H4
4-NO2 C6H4
4-Me C6H4
4-NMe2 C6H4
3-Pyridyl
71.1 2.2
—
—
—
—
—
—
49.4 0.03
—
—
5.9 0.45
0.5 0.01
4.7 0.1
—
4.65 0.1
19.7 1.7
—
(17 ml) in
a
round-bottomed flask. 30-Hydroxyacetophenone
(5 mmol) and substituted aldehyde (5 mmol) were added to the
cold mixture with stirring. The reaction mixture was stirred for
3–5 h and monitored for its completion through TLC. When the
reaction was found to be complete, the reaction mixture was kept
at 0–4 °C overnight. The solid mass obtained, thereby, was cooled
and neutralized with ice-cooled dilute hydrochloric acid. The pre-
cipitates separated were filtered and recrystallized from distilled
ethanol.
—
—
75.2 2.1
34.4 0.7
—
45
47
49
2-Thienyl
—
3.97 0.02
Galanthamine
8
0.05
generally found to be inactive. It seems that substitution of a hy-
droxyl group at C-3 played an important role in increasing the
inhibitory potential of these compounds against butyrylcholinest-
erase. Molecular docking studies are being carried out to have a
better insight into their mechanism of inhibitory action.
5.1.2. General method for the synthesis of 2,3-dihydro-1,5-
benzothiazepines (27–52)
To a solution of chalcone (1 g/20 mmol) in dry acidic methanol
acidified by adding few drops of glacial acetic acid, was added o-
aminothiophenol (2.15 ml, 20 mmol). The mixture was refluxed
until a crystalline solid separated out. After cooling the solid prod-
uct was collected and washed with diethyl ether and cold metha-
nol. The crude solid was recrystallized from ethanol.
It may be seen from Table 5 that of the screened benzothiaze-
pines most of the compounds were found to be good inhibitors
of butyrylcholinesterase with IC50 varying from 3.97 to
1421 ppm. With IC50 of 5.9 and 3.97 ppm for AChE and BChE,
respectively, compound 49 was found to be the most active mem-
ber of the library. This is the only compound that showed activity
against both AChE and BChE. The other benzothiazepines 35 and
37 were found to inhibit BChE significantly with IC50 4.70 and
4.65 ppm, respectively. Moreover, 3-OH-, and 2-OMe-substituted
benzothiazepines also emerged as good inhibitors of butyrylcho-
linesterase. It is important to note that all three compounds can
be regarded as better inhibitors of butyrylcholinesterase than the
standard galanthamine (IC50 8 0.05). In contrast to butyrylcholin-
esterase inhibition, most of the compounds were found to be inac-
tive against acetylcholinesterase.
5.2. Solid-phase synthesis
Solid-phase synthesis of 2,3-dihydro-1,5-benzothiazepines in-
volved the following steps.
5.2.1. Synthesis of resin-bound acetophenone
3-Hydroxy acetophenone (7.35 mmol) were dissolved in 10 ml
of NMM followed by the addition of 0.66 g of TPP. After that
0.444 ml of DIAD was added dropwise for 15 min. The resulting
yellow solution was stirred for 5 h at room temperature. The re-
sin-bound acetophenone was then filtered and washed succes-
sively and extensively with CH2Cl2/THF (2:1), THF/H2O (1:1),
MeOH/THF (1:1), MeOH/CH2Cl2 (6 ml each) and dried under high
vacuum overnight (12 h). The substitution was confirmed by cleav-
age with 50% TFA in CH2Cl2 (1 h at room temperature) 21.
4. Conclusion
Solid-phase synthesis of 2,3-dihydro-1,5-benzothiazepines led
to an improved yield and better purity when compared to their
synthesis in solution phase. Compounds were tested for their po-
tential as antibacterial, cholinesterase inhibitors and potato disk
tumor inhibitors. Most of the tested compounds were found to
be good inhibitors of potato disk tumors and may be considered
as promising candidates for developing into antitumor drugs.
Benzothiazepines 35, 37 and 49 were found to be better butyr-
ylcholinesterase inhibitors than the standard galanthamine. None
of the compounds showed significant activity against any bacte-
rial strain.
5.2.2. Synthesis of resin-bound chalcones
The resin-bound acetophenone was added to a solution of aryl
aldehydes (3 equiv) in dry THF (25 ml). A solution of KOH (6 equiv)
in ethanol was added over a period of 15 min. The flask was shaken
for 24 h at room temperature. The solution was removed by filter-
ing the resin. The filtered resin-bound chalcone was washed suc-
cessively with 30% aqueous AcOH, H2O, THF, MeOH, THF, and
CH2Cl2 (all by 2ꢁ 40 ml), followed by drying under high vacuum
for 24 h.
5. Experimental
5.2.3. Synthesis ofresin-bound 2,3-dihydro-1,5-benzothiazepines
The resin-bound chalcone was suspended in a solution of o-
aminothiophenol in DMF and was refluxed for about 18 h. A few
drops of glacial acetic acid were added to the reaction mixture as
catalyst. The resin was washed successively with MeOH, CH2Cl2,
DMF, THF, diethyl ether, acetone, and ethyl acetate. It was then
dried under vacuum overnight. The product was cleaved with
50% TFA in CH2Cl2 (1 h at room temperature). The cleaved product
was directly submitted for GC–MS analysis.
1H NMR spectra were recorded on a Bruker 300 MHz NMR spec-
trometer in d6-acetone or d6-DMSO using tetramethylsilane as
internal reference. 1H NMR spectra were recorded at 300 MHz
while 13C NMR spectra were recorded at 75 MHz. Mass spectra
were measured on Agilent GC–MS gas chromatography mass spec-
trometer. Melting points were determined on a MEL-Temp appara-
tus and are un-corrected.

F. L. Ansari et al. / Bioorg. Med. Chem. 16 (2008) 7691–7697
7695
5.2.3.1. 2-(Phenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,5-ben-
zothiazepine (27). Yield 71%; mp 102 °C; 1H NMR (DMSO-d6): d
8.60 (s, 1H, OH), 6.93–7.70 (m, 13H, aryl H), 4.16 (dd,
J2,3a = 5.0 Hz, J2,3b = 12.0 Hz, 1H, H-2), 3.23 (dd, J3a,3b = 12.0 Hz,
5.2.3.9. 2-(300-Hydroxyphenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,
5-benzothiazepine (35). Yield 70%; mp 142 °C dec; 1H NMR
(DMSO-d6): d 8.89 and 8.63 (s, 2H, OH), 6.87–8.06 (m, 12H, aryl
H), 3.85 (d, J2,3b = 6.3 Hz, 1H, H-2), 3.34 (unresolved, 1H, H-3a),
J
3a,2 = 5.0 Hz, 1H, H-3a), 2.53 (t, J3b,3a = J3b,2 = 12.0 Hz, 1H, H-3b);
2.41 (d, J d 157.0 (C-4),
3b,2 = 6.3 Hz, 1H, H-3b); 13C NMR:
13C NMR: d 160.0 (C-4), 113.0–157.9 (aryl C), 45.5 (C-2), 28.2 (C-
3); EIMS (m/z, %): 330 [M+Å, 6].
113.0–157.9 (aryl C), 45.5 (C-2), 28.2 (C-3); EIMS (m/z, %): 347
[M+Å, 9].
5.2.3.10. 2-(400-Hydroxyphenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (36). Yield 53%; mp 170–172 °C; 1H NMR
(DMSO-d6): d 8.71 and 8.49 (s, 2H, OH), 6.76–8.00 (m, 12H, aryl
H), 3.58 (t, J2,3a = J2,3b = 7.2 Hz, 1H, H-2), 3.06 (unresolved, 1H, H-
3a), 2.87 (unresolved, 1H, H-3b); 13C NMR: d 157.1 (C-4), 113.0–
147.0 (aryl C), 40.2 (C-2), 28.0C-3; EIMS (m/z, %): 347 [M+Å, 8].
5.2.3.2. 2-(200-Chlorophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (28). Yield 81%; mp 45–47 °C; 1H NMR
(DMSO-d6): d 10.05 (s, 1H, OH), 7.09–8.43 (m, 12H, aryl H), 4.89
(dd, J2,3a = 5.1 Hz, J2,3b = 9.0 Hz, 1H, H-2), 3.43 (dd, J3b,3a = 14.0 Hz,
J
3b,2 = 9.0 Hz, 1H, H-3b), 3.36 (dd, J3a,3b = 14.0 Hz, J3a,2 = 5.1 Hz,
1H, H-3a); 13C NMR: d 167.5 (C-4), 115.2–162.2 (aryl C), 55.9 (C-
2), 27.2 (C-3); EIMS (m/z, %): 367 [M+Å+2, 3], 366 [M+Å+1, 1], 365
[M+Å, 9].
00
0
5.2.3.11. 2-(2 -Methoxyphenyl)-4-(3 -hydroxyphenyl)-2,3-dihydro-1,
5-benzothiazepine (37). Yield 61%; mp 62–64 °C; 1H NMR (d6-
acetone): d 8.11 (s, 1H, OH), 7.04–8.08 (m, 12H, aryl H), 3.58 (t,
5.2.3.3. 2-(300-Chlorophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (29). Yield 80%; mp 100–101 °C; 1H NMR
(DMSO-d6): d 10.0 (s, 1H, OH), 7.10–8.03 (m, 12H, aryl H), 4.32
J
2,3b = 6.9 Hz, 1H, H-2), 2.87 (unresolved, 1H, H-3a*), 2.50 (s, 3H,
O-CH3), 1.036 (t, J3b,2 = 6.9 Hz, 1H, H-3b*); 13C NMR: d 162.7 (C-
4), 114.0–158.3 (aryl C), 56.0 (O-CH3), 43.0 (C-2), 40.2 (C-3); EIMS
(m/z, %): 361[M+Å, 5].
(dd, J2,
J2,3a = 5.0 Hz, J3b = 12.9 Hz, 1H, H-2), 3.74 (t, J3b,3a =
J
3b,2 = 12.6 Hz, 1H, H-3b), 3.51 (dd, J3a,3b = 12.9 Hz, J3a,2 = 5.0 Hz,
1H, H-3a); 13C NMR: d 165.0 (C-4), 118.0–158.7 (aryl C), 56.3 (C-
2), 33.5 (C-3); EIMS (m/z, %): 367 [M+Å+2, 2], 366 [M+Å+1, 1], 365
[M+Å, 6].
5.2.3.12. 2-(300-Methoxyphenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,
5-benzothiaze-pine (38). Yield 59%; mp 58 °C; 1H NMR (d6-ace-
tone): d 8.63 (s, 1H, OH), 6.77–7.31 (m, 12H, aryl H), 3.41 (d,
5.2.3.4. 2-(400-Chlorophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (30). Yield 87%; mp 114 °C; 1H NMR (DMSO-
d6): d 10.05 (s, 1H, OH), 7.09–8.44 (m, 12H, aryl H), 5.0 (dd,
J
2,3b = 7.1 Hz, 1H, H-2), 3.0 (unresolved, 1H, H-3a), 2.76 (s, 3H, O-
CH3), 1.65 (d, J3b,2 = 7.1 Hz, 1H, H-3b); 13C NMR: d 161.6 (C-4),
115.0–158.0 (aryl C), 57.0 (O-CH3), 41.7 (C-2), 39.0 (C-3); EIMS
(m/z, %): 361 [M+Å, 8].
J
2,3a = 5.1 Hz, J2,3b = 12.9 Hz, 1H, H-2), 3.53 (t, J3b,3a = J3b,2 = 12.9 Hz,
1H, H-3b), 3.39 (dd, J3a,3b = 12.9 Hz, J3a,2 = 5.1 Hz, 1H, H-3a); 13C
NMR: d 160.0 (C-4), 113.0–157.9 (aryl C), 45.5 (C-2), 28.2 (C-3);
EIMS (m/z, %): 367 [M+Å+2, 3], 366 [M+Å+1, 1], 365 [M+Å, 10].
5.2.3.13. 2-(400-Methoxyphenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (39). Yield 56%; mp 80 °C; 1H NMR (d6-ace-
tone): d 9.10 (s, 1H, OH), 6.23–7.09 (m, 12H, aryl H), 3.83 (s, 3H,
O-CH3), 3.30 (d, J2,3b = J = 7.0 Hz, 1H, H-2), 2.9 (unresolved, 1H, H-
3a*), 1.0 (t, J3b,3a = J3b,2 = 6.9 Hz, 1H, H-3b*); 13C NMR: d 159.6 (C-
4), 114.7–158.0 (aryl C), 59.0 (O-CH3), 43.5 (C-2), 40.0 (C-3); EIMS
(m/z, %): 361 [M+Å, 10].
5.2.3.5. 2-(200-Fluorophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (31). Yield 87%; mp 68–70 °C; 1H NMR
(DMSO-d6): d 9.72 (s, 1H, OH), 6.96–7.59 (m, 12H, aryl H), 5.31
(dd, J2,3b = 12.6 Hz, J2,3a = 9.9 Hz, 1H, H-2), 3.36 (dd, J
=
3a,3b
12.6 Hz, J3a,2 = 9.9 Hz, 1H, H-3a), 2.87 (t, J3b,2 = J3b,3a = 12.6 Hz,
1H, H-3b); 13C NMR: d 168.9 (C-4), 114.0–160.6 (aryl C), 53.3 (C-
2), 35.3 (C-3); EIMS (m/z, %): 349 [M+Å, 7].
5.2.3.14. 2-(300,400-Dimethoxyphenyl)-4-(30-hydroxyphenyl)-2,3-
dihydro-1,5-benzo-thiazepine (40). Yield 61%; mp 96 °C; 1H NMR
(d6-acetone): d 9.7 (s, 1H, OH), 6.46–7.15 (m, 12H, aryl H), 3.80 and
3.77 (s, 3H, 300, 400O-CH3), 3.06 (d, J2,3b = 6.6 Hz, 1H, H-2), 2.83
(unresolved, 1H, H-3a), 2.0 (t, J3b,3a = J3b,2 = 6.6 Hz, 1H, H-3b); 13C
NMR: d 162.1 (C-4), 112.0–157.0 (aryl C), 61.0 and 59.0 ((300,400-
O-CH3), 49.5 (C-2), 39.7 (C-3); EIMS (m/z, %): 391 [M+Å, 6].
5.2.3.6. 2-(300-Fluorophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (32). Yield 73%; mp 73 °C; 1H NMR (DMSO-
d6): d 9.95 (s, 1H, OH), 7.90–6.59 (m, 12H, aryl H), 5.10 (dd,
J
J
2,3a = 9.9 Hz, J2,3b = 12.9 Hz, 1H, H-2), 3.27 (dd, J3a,3b = 12.9 Hz,
3a,2 = 9.9 Hz, 1H, H-3a), 2.66 (t, J3b,3a = J3b,2 = 12.9 Hz, 1H, H-3b);
13C NMR: d 162.0 (C-4), 113.0–164.7 (aryl C), 51.4 (C-2), 32.8 (C-
3); EIMS (m/z, %): 349 [M+Å, 7].
5.2.3.15. 2-(400-Hydroxy-300-methoxyphenyl)-4-(30-hydroxyphenyl)-
2,3-dihydro-1,5-benzothiazepine (41). Yield 55%; mp 108 °C; 1H
NMR (DMSO-d6): d 8.1 and 8.08 (s, 2H, OH), 7.04–8.04 (m, 11H, aryl
H), 3.83 (s, 3H,O-CH3), 3.43 (d, J2,3b = J = 14.1 Hz, 1H, H-2), 2.50
(unresolved, 1H H-3a), 1.03 (d, J3b,2 = 14.1 Hz, 1H, H-3b); 13C
NMR: d 158.3 (C-4), 113.0–157.9 (aryl C), 55.9 (O-CH3), 45.5 (C-
2), 28.2 C-3); EIMS (m/z, %): 376 [M+Å, 5].
5.2.3.7. 2-(400-Fluorophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (33). Yield 93%; mp 79–80 °C; 1H NMR
(DMSO-d6): d 8.60 (s, 1H, OH), 7.06–8.19 (m, 12H, aryl H), 5.27
(dd, J2,3b = 12.9 Hz, J2,3a = 5.0 Hz, 1H, H-2), 3.50 (dd, J3a,3b = 12.9 Hz,
J
3a,2 = 5.0, 1H, H-3a), 2.97 (t, J3b,3a = J3b,2 = 12.9 Hz, 1H, H-3b); 13C
NMR: d 168.3 (C-4), 113.9–163.3 (aryl C), 59.5 (C-2), 37.3 (C-3);
5.2.3.16. 2-(300-Nitrophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (42). Yield 83%; mp 82–84 °C; 1H NMR
(DMSO-d6): d 9.70 (s, 1H, OH), 6.96–8.22 (m, 12H, aryl H), 5.42
EIMS (m/z, %): 349 [M+Å, 8].
5.2.3.8. 2-(400-Bromophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (34). Yield 82%; mp 198 °C; 1H NMR (DMSO-
d6): d 9.78 (s, 1H, OH), 7.02–7.48 (m, 12H, aryl H), 4.94 (dd,
J2,3a = 10.0 Hz, J2,3b = 12.9 Hz, 1H, H-2), 3.25 (dd, J3a,3b = 12.9 Hz,
(dd, J2,3a = J = 9.0 Hz, J2,3b = 12.6 Hz, 1H, H-2), 3.38 (dd, J 3a,3b =
12.9 Hz, J3a,2 = 9.0 Hz, 1H, H-3a), 2.86 (t, J3b,2 = J3b,3a = 12.9 Hz, 1H,
H-3b); 13C NMR: d 169.0 (C-4), 114.1–158.1 (aryl C), 58.0 (C-2),
37.0 (C-3); EIMS (m/z, %): 376 [M+Å, 7].
J
3a,2 = 10.0 Hz, 1H, H-3a), 3.14 (t, J3b,2 = J3b,3a = 12.9 Hz, 1H, H-3b);
13C NMR: d 165.0 (C-4), 118.2–158.4 (aryl C), 50.1 (C-2), 40.0 (C-
3); EIMS (m/z, %): 411 [M+Å+2, 9], 409 [M+Å, 10].
5.2.3.17. 2-(400-Nitrophenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (43). Yield 90%; mp 73 °C; 1H NMR (DMSO-

7696
F. L. Ansari et al. / Bioorg. Med. Chem. 16 (2008) 7691–7697
d6): d 9.70 (s, 1H, OH), 6.96–8.22 (m, 12H, aryl H), 5.42 (dd,
5.07 (dd,
J
2,3a = 12.6 Hz,
J
2,3b = 13.2 Hz, 1H, H-2), 4.10 (dd,
1H, H-3a), 3.39 (t,
J2,3b = 12.6 Hz, J2,3a = 4.8 Hz, 1H, H-2), 3.47 (dd, J 3a,3b = 12.6 Hz,
3a,2 = 4.8 Hz, 1H, H-3a), 3.10 (t, J3b,2 = J3b,3a = 12.6 Hz, 1H, H-3b);
J
J
3a,3b = 13.2 Hz, J3a,2 = 12.6 Hz,
J
3b,3a = J3b,2 = 13.2 Hz, 1H, H-3b); 13C NMR: d 163.0 (C-4), 109.0–
13C NMR: d 179.8 (C-4), 115.1–167.3 (aryl C), 51.7 (C-2), 24.7 (C-
3); EIMS (m/z, %): 376 [M+Å, 5].
158.1 (aryl C), 53.8 (C-2), 40.0 (C-3); EIMS (m/z, %): 416 [M+Å+2,
7], 415 [M+Å+1, 1], 414 [M+Å, 8].
5.2.3.18. 2-(400-Methylphenyl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (44). Yield 71%; mp 97 °C; 1H NMR (d6-ace-
tone): d 9.70 (s, 1H, OH), 6.23–7.48 (m, 12H, aryl H), 4.94 (dd,
5.2.3.26. 2-(Pyrrol-200-yl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,
5-benzothiazepine (52). Yield 37%; mp 133 °C dec; 1H NMR
(DMSO-d6): d 8.13 (s, 1H, OH), 6.96–7.70 (m, 11H, aryl H), 7.26 (br
s, 1H, NH), 4.20 (dd, J2,3a = 6.3 Hz, J2,3b = 12.0 Hz, 1H, H-2), 3.41 (dd,
J2,3a = 4.8 Hz, J2,3b = 12.6 Hz, 1H, H-2), 3.25 (dd, J3a,3b = 12.6 Hz,
3a,2 = 4.8 Hz, 1H, H-3a), 3.04 (t, J3b,3a = J3b,2 = 12.6 Hz, 1H, H-3b),
J
J
3a,3b = 12.0 Hz, J3a,2 = 6.3 Hz, 1H, H-3a), 3.00 (t, J3b,3a = J3b,2 = 12.0 Hz,
2.35 (s, 3H, CH3); EIMS (m/z, %): 345 [M+Å, 8].
1H, H-3b); 13C NMR: d 160.3 (C-4), 109.0–131.2 (aryl C), 53.2 (C-2),
34.7 (C-3); EIMS (m/z, %): 321 [M+Å+1, 1], 320 [M+Å, 3].
5.2.3.19. 2-(400-N,N-Dimethylamino)-4-(30-hydroxyphenyl)-2,3-
dihydro-1,5-benzothiazepine (45). Yield 69%; mp 100 °C; 1H
NMR (DMSO-d6): d 8.45 (s, 1H, OH), 6.77–7.49 (m, 12H, aryl H),
4.96 (dd, J2,3a = J = 5.1 Hz, J2,3b = 12.0 Hz, 1H, H-2), 3.83 (s, 6H, N-
(CH3)2), 3.26 (dd, J 3a,3b = 12 Hz, J3a,2 = 5.1 Hz, 1H, H-3a), 3.04 (t,
J3b,3a = J3b,2 = 12 Hz, 1H, H-3b); 13C NMR: d 167.0 (C-4), 115.2–
162.2 (aryl C), 55.9 (C-2), 40.7 (N (CH3)2), 27.2 (C-3); EIMS (m/z,
%): 376 [M+Å+2, 1], 374 [M+Å, 10].
5.3. Biological screening
The synthesized compounds were screened for their potential
as antibacterials, potato disk tumor and as cholinesterase
inhibitors.
5.3.1. Antibacterial assay
All the synthesized compounds were tested against six bacterial
strains namely: B. bronchiseptica ATCC 4617, M. leuteus ATCC, P.
picketti ATCC 49129, E. coli ATCC 1522, E. aerogenes ATCC 13048,
and S. setubal ATCC 19196. Agar Well diffusion method was used for
these studies. 0.75 ml of the broth culture containing approx. 106
colony-forming units (CFU/ml) of testing strain was added to
75 ml of nutrient agar medium at 45 °C, mixed well, and poured into
a 14-cm sterile metallic Petri plate. The medium was allowed to
solidify and 8-mm wells were dug with sterile metallic borer. One
hundred microliters of the test sample (1 mg/ml in DMSO) was
poured in respective wells. In each plate DMSO and standard anti-
bacterial drug (Roxithromycin 1 mg/ml) served as negative and po-
sitive controls, respectively. Triplicate plates of each bacterial strain
were prepared. The plates were incubated at 37 °C for 24 h. The anti-
bacterial activity was determined by measuring the diameter of
zones showing complete inhibition (mm). Growth inhibition was
calculated with reference to positive control, Roxithromycin. All
tested compounds showed non-significant antibacterial activities.
5.2.3.20.
2-(Pyridin-200-yl)-4-(30-hydroxyphenyl)-2,3-dihydro-
1,5-benzothiazepine (46). Yield 51%; mp 110–114 °C; 1H NMR
(DMSO-d6): d 9.52 (s, 1H, OH), 6.40–8.61 (m, 12H, aryl H), 4.91
(dd,
J2,3a = 12.3 Hz, J2,3b = 14.4 Hz, 1H, H-2), 3.44 (dd,
J
3a,3b = 14.4 Hz,
J3a,2 = 12.30 Hz, 1H, H-3a), 1.05
(t,
J
3b,3a = J3b,2 = 14.4 Hz, 1H, H-3b); 13C NMR: d 162.3 (C-4), 115.0–
156.1 (aryl C), 50.2 (C-2), 37.9 (C-3); EIMS (m/z, %): 332 [M+Å, 1].
5.2.3.21. 2-(Pyridin-300-yl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,
5-benzothiazepine (47). Yield 50%; mp 88–91 °C; 1H NMR
(DMSO-d6): d 8.62 (s, 1H,OH), 6.42–8.30 (m, 12H, aryl H), 4.38
(dd,
J
2,3a = 9.9 Hz, J2,3b = J = 12.9 Hz, 1H, H-2), 3.37 (dd,
J
3a,3b = 12.9 Hz, J3a,2 = 9.9 Hz, 1H), 1.07 (t, J3b,3a = J3b,2 = 12.9 Hz, 1H,
H-3b); 13C NMR: d 161.7 (C-4), 115.0–159.1 (aryl C), 49.1 (C-2),
30.1 (C-3); EIMS (m/z, %): 332 [M+Å, 1].
5.2.3.22. 2-(Pyridin-400-yl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,
5-benzothiazepine (48). Yield. 48%; mp 119 °C dec; 1H NMR
(DMSO-d6): d 8.60 (s, 1H, OH), 7.65–7.91 (m, 12H, aryl H), 4.67
5.3.2. Potato disk tumor inhibition
(dd,
J
2,3a = 10.0 Hz, J2,3b = J = 14.0 Hz, 1H, H-2), 3.42 (dd,
The assay was performed according to a standard procedure23
,
J3a,3b = 14.0, J3a,2 = 10.0 Hz, 1H, H-3a), 2.50 (t, J3b,3a = J3b,2 = 14.0 Hz,
1H, H-3b); 13C NMR: d 160.0 (C-4), 113.0–157.9 (aryl C), 45.5 (C-
2), 28.2 (C-3); EIMS (m/z, %): 332 [M+Å, 1].
usinga48-h-oldbacterialcultureoftheAT-10strainofAgrobacterium
tumefaciens.Inoculumswiththreeconcentrationsofeachtestsample
(10, 100, and 1000 ppm) containing bacterial culture was prepared.
Red-skinned potatoes were surface-sterilized in 0.1% HgCl2 solution.
A bore of 8-mm diameter was used to create potato cylinders, which
were cut into 5-mm disks. Autoclaved agar solution (1.5%) (Petri
plates) was allowed to solidify. The potato disks were placed on agar,
5.2.3.23. 2-(Thien-200-yl)-4-(30-hydroxyphenyl)-2,3-dihydro-1,5-
benzothiazepine (49). Yield 70%; mp 55 °C; 1H NMR (DMSO-d6): d
8.41 (s, 1H, OH), 6.96–7.70 (m, 11H, aryl H), 5.50 (dd, J2,3a = 12.6 Hz,
J2,3b = J = 13.2 Hz, 1H, H-2), 3.53 (dd, J3a,3b = 13.2 Hz, J3a,2 = 12.6 Hz,
1H, H-3a), 2.90 (t, J3b,3a = J3b,2 = 13.2 Hz, 1H, H-3b); 13C NMR: d
168.2 (C-4), 113.9–157.7 (aryl C), 55.7 (C-2), 38.0 (C-3); EIMS
(m/z, %): 339 [M+Å+2, 12], 338 [M+Å+1, 3], 337 [M+Å, 9].
andinoculum(50 ll)waspouredoneachdisk.Theplatesweresealed
with parafilm to avoid contamination and moisture loss. The plates
were then incubated at 28 °C for 21 days, the potato disks were
stainedwithLugol’ssolution(10%KI/5%I2),andtumorswerecounted
under a dissecting microscope. All the tests were performed in repli-
cate, and thepercenttumor inhibitionwas calculatedby dividingthe
average number of tumors per sample by the average numbers of tu-
mors of a control, and then multiplied by a factor of 100. IC50 values
were calculated by the following literature procedure:
5.2.3.24. 2-(500-Methylthien-200-yl)-4-(30-hydroxyphenyl)-2,3-
dihydro-1,5-benzothiazepine (50). Yield 45%; mp 94–96 °C; 1H
NMR (DMSO-d6): d 8.30 (s, 1H, OH), 6.53–7.26 (m, 10H, aryl H),
4.61 (dd, J2,3a = 6.5 Hz, J2,3b = 12.3 Hz, 1H, H-2), 3.20 (dd, J
3a,3b = 12.3 Hz,
J3a,2 = 6.5 Hz,
1H,
H-3a),
2.78
(t,
J3b,3a = J3b,2 = 12.3 Hz, 1H, H-3b), 1.91 (s, 3H, CH3); 13C NMR: d
161.8 (C-4), 118.0–158.1 (aryl C), 50.6 (C-2), 38.9 (C-3), 16.0
(CH3); EIMS (m/z, %): 353 [M+Å+2, 10], 352 [M+Å+1, 1], 351 [M+Å, 9].
Average number of tumors of sample
Average number of tumors of control
% inhibition ¼ 100ꢂ
ꢁ100
2-(500-Bromothien-200-yl)-4-(30-hydroxyphenyl)-2,3-
5.3.3. Cholinesterase inhibition
The synthesized 2,3-dihydro-1,5-benzothiazepines were
screened for enzyme inhibition activity against acetylcholinester-
5.2.3.25.
dihydro-1,5-benzothiazepine (51). Yield 49%; mp 76 °C dec; 1H
NMR (DMSO-d6): d 9.35 (s, 1H, OH), 6.33–7.29 (m, 10H, aryl H),

F. L. Ansari et al. / Bioorg. Med. Chem. 16 (2008) 7691–7697
7697
ase (AChE) and butyrylcholine esterase. IC50 values were calculated
References
to show the efficiency of inhibition by these compounds. Acetylthi-
ocholine iodide and butyrylthiocholine chloride were used as sub-
strates to assay acetylcholinesterase and butyrylcholinesterase,
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respectively. Sodium phosphate buffer (pH 8.0) (150
5,50-dithio-bis(2-nitrobenzoic acid) DNTB (10
l), test compound
solution (10 l), and acetylcholinesterase (or butyrylcholinester-
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25 °C. The reaction was then initiated by the addition of 10 l acet-
ylthiocholine (or butyrylthiocholine) (10 l), respectively. The
ll, 100 mM),
l
l
l
l
l
hydrolysis of acetylthiocholine and butyrylthiocholine was moni-
tored by the formation of yellow 5-thio-2-nitrobenzoate anion as
a result of the reaction of DTNB with thiocholine released by the
enzymatic hydrolysis of acetylthiocholine and butyrylthiocholine,
respectively, at 412 nm. All test compounds and the control were
dissolved in EtOH. All the reactions were performed in triplicate
in a 96-well micro-plate in SpectraMax 340 (Molecular Devices,
USA). The percentage (%) inhibition was calculated as follows:
(E–S)/E ꢁ 100, where E is activity of enzyme without test com-
pound and S, the IC50 value were determined by monitoring the ef-
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on the inhibition values. The IC50 values were then calculated using
the EZ-Fit Enzyme Kinetics program (Perrella Scientific Inc., Am-
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Acknowledgment
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The authors thankfully acknowledge the financial support pro-
vided by the Higher Education Commission, Pakistan.