Syntheses and antibacterial activity of phendioxy substituted cyclic enediynes

Article abstract of DOI:10.1016/j.bmcl.2007.03.007

Syntheses and antibacterial activity of 13-membered 1,3-phendioxy substituted cyclic enediynes are reported. The compounds were screened against gram-positive and gram-negative strains and some of the compounds exhibit potent antibacterial activity.

Full text of DOI:10.1016/j.bmcl.2007.03.007

Bioorganic & Medicinal Chemistry Letters 17 (2007) 3226–3230
Syntheses and antibacterial activity of phendioxy substituted
cyclic enediynes
Mukesh Chandra Joshi, Gopal Singh Bisht and Diwan S. Rawat*
Department of Chemistry, University of Delhi, Delhi 110007, India
Received 15 November 2006; revised 6 February 2007; accepted 5 March 2007
Available online 12 March 2007
Abstract—Syntheses and antibacterial activity of 13-membered 1,3-phendioxy substituted cyclic enediynes are reported. The com-
pounds were screened against gram-positive and gram-negative strains and some of the compounds exhibit potent antibacterial
activity.
ꢀ 2007 Elsevier Ltd. All rights reserved.
Enediyne chemistry has captured the imagination of the
chemist and biologist throughout the world since the dis-
covery of natural product enediynes such as calicheami-
cin, esperamicin, maduroptin, dynamicin, and more
recently uncialamycin.1–4 Some of the natural product
enediynes are three order of magnitude more potent than
other anticancer drugs. The anticancer activity of these
compounds is due to the presence of highly unsaturated
1,5-diyne-3-ene unit that undergoes cycloaromatization
and generates benzene-1,4-diradical, which cleaves the
DNA.5,6 Unfortunately, due to modest selectivity for
cancer cells clinical use of this class of compounds has
been limited. In order to improve the biological activity
of the enediynes, efforts are being made to synthesize
analogous compounds with better efficacy.7–12 Apart
from anticancer activity, synthetic enediynes are known
to exhibit cytotoxicity against various cell lines,13,14 pro-
tein degradation activity15 and topoisomerase inhibitory
activity.16 Natural product enediynes are known to exhi-
bit potent antibacterial activity against gram-positive
and gram-negative strains.17 To the best of our knowl-
edge, none of the synthetic enediynes were evaluated
for antibacterial activity. To this end, we report syntheses
and antibacterial activity of substituted 13-membered
phendioxy cyclic enediynes.
Reaction of 2,4-dihydroxybenzaldehyde (2) with 1,8-
dibromo-oct-4-ene-1,6-diyne (5)18 in presence of
K2CO3 afforded desired 13-membered cyclic enediyne
(7) in 62% yield, which on reduction with NaBH4 in
MeOH affords corresponding alcohol (8). However, un-
der identical reaction conditions, reaction of resorcinol
(1) with 1,8-dibromo-oct-4-ene-1,6-diyne (5)18 results
in the decomposition of the dibromoenediyne precursor.
Alternatively, the desired 13-membered cyclic enediyne
(6) was synthesized by different approach, using
propargyl bromide and resorcinol as starting material
(Scheme 1).
Reaction of resorcinol (1) with propargyl bromide in dry
DMF in the presence of K2CO3 leads to the formation
of 3 in 92% yield. Sonogashira coupling of 1.0 equiva-
lents of 3 with cis-1,2-dichloroethylene over a Pd(PPh3)4
catalyst in the presence of CuI and BuNH2 generates 6
in 66% yield. The identity of these compounds was con-
firmed spectroscopically. Compounds 6, 7, and 8 were
tested against gram-positive and gram-negative strains
and these compounds exhibit potent antibacterial activ-
ity (Table 1). In order to study the effect of different sub-
stituents in the aromatic ring on antibacterial activity,
we planned to derivatise compound 7. Surprisingly,
reaction of cyclic enediyne 7 with primary amines did
not work even at elevated temperatures. Then desired
enediynes (13–20) were prepared by different approach
as shown in Scheme 3. Imines (9–12) used for the synthe-
sis of cyclic enediynes were prepared as shown in
Scheme 2. Reaction of equimolar amount of aldehyde
(2) with different amines, under anhydrous conditions,
yielded substituted imines in good yields. All imine
Scheme 1 illustrates two different strategies for the prep-
aration of the 13-membered cyclic enediynes.
Keywords: Enediyne; Anticancer activity; Antibacterial activity; Sono-
gashira coupling.
*
Corresponding author. Tel.: +91 11 27667465; fax: +91 11
27667501; e-mail: dsrawat@chemistry.du.ac.in
0960-894X/$ - see front matter ꢀ 2007 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2007.03.007

M. C. Joshi et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3226–3230
3227
OH
Propargyl bromide, K2CO3,
DMF
O
85- 92
OH
O
R
R
1, R = H
2, R = CHO
3, R = H
4, R = CHO
K2CO3,
3, cis-dichloroethylene
Pd(0), CuI, n-BuNH2
Benzene, 66
Br
Br
DMF
R = CHO
62
5
R
CH2OH
O
O
NaBH4, MeOH
80
O
O
6, R = H
7, R = CHO
8
Scheme 1.
Table 1. Antibacterial activity of cyclic enediynes
S. No.
MIC (lg/mL)
P. aeruginosa (ATCC, 11775)
E. coli (ATCC, 25668)
S. aureus (clinical isolate)
S. typhimurium (clinical isolate)
3
4
>1000
>1000
na
>1000
>1000
4
>1000
>1000
na
>1000
>1000
8
6
7
na
na
256
8
8
128
128
128
nda
128
64
8
4
13
14
15
16
17
18
19
20
21b
na
256
nda
na
na
nda
na
nda
na
128
32
128
4
128
16
128
512
na
na
na
64
na
na
128
4
16
1
8
16
0.5
2
2
na, MIC no activity up to 512 lg/mL.
a MIC not determined.
b Tetracycline was used as a reference compound.
derivatives were purified over SiO2 column and charac-
terized spectroscopically.
In vitro antibacterial activity: Antimicrobial susceptibil-
ity testing was carried out using National Committee
for Clinical Laboratory Standards (NCCLS) microdilu-
tion broth assay. Briefly, the bacterial strains were
grown in Mueller Hinton Broth (HIMEDIA) until expo-
nential growth was achieved. Test compounds were dis-
solved in DMSO/Water to make a series of two fold
dilution. 90 ll of 2–7 · 105 CFUs/mL of bacterial sam-
ple per mL of Mueller–Hinton broth (HIMEDIA) was
dispensed into 96-well polypropylene microtitre plate
(SIGMA). Then 10 ll of serially diluted compounds
was added. The microtitre plates were incubated over-
night at 37 ꢁC and the absorbance was read at 630 nm.
Uninoculated Mueller–Hinton broth was used as nega-
tive control. The tests were carried out in triplicate.
The minimum inhibitory concentration (MIC) is taken
Reaction of substituted imines (9–12) with 1,8-dib-
romo-oct-4-ene-1,6-diyne (5)18 in presence of K2CO3
in dry DMF leads to the formation of desired prod-
ucts in moderate yields as shown in Scheme 3. Reac-
tion completes in 8–10 h and excess of DMF from
the reaction mixture was removed at reduced pres-
sure. The residue thus obtained was washed with
water and crude product was purified over SiO2 col-
umn. The cyclic imine enediynes (13–16) were re-
duced to amine derivatives (17–20) using NaBH4 in
methanol. After usual work-up, these compounds
were purified by SiO2 column and characterized
spectroscopically.19

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M. C. Joshi et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3226–3230
R
OH
OH
Substituted amines,
MeOH, Mol. Sieves
CHO
N
65-
HO
HO
2
9, R= p-Methyl
10, R = p-Cl
11, R = p-Br
12, R = H
Scheme 2.
R
R
Br
Br
OH
O
O
N
N
K2CO3, DMF
45-
HO
13, R = p-Methyl
14, R = p-Cl
15, R = p-Br
16, R = H
9-12
R
R
O
O
O
O
N
H
N
NaBH4, MeOH
65-
17, R = p-Methyl
18, R = p-Cl
19, R = p-Br
20, R = H
13-16
Scheme 3.
as the lowest concentration of compound that inhibits
50% growth of microorganism. The compounds were
evaluated against gram-positive and gram-negative bac-
terial strains. Minimum inhibitory concentration (MIC)
of each compound is shown in Table 1.
For example, compound 17 possess much lower MIC
value than compound 13. In similar manner compound
20 has much higher antimicrobial activity than com-
pound 16. Moreover only compounds 17 and 20 have
potent activity against Pseudomonas aeruginosa. This
suggests unsaturation around nitrogen is not required
for antimicrobial activity of these synthetic compounds.
Tetracycline is used as standard drug for antimicrobial
activity. All tested compound showed varied activity
against different bacterial strains. The antimicrobial
activity data indicate that compounds 6, 8, 17, and 20
were most effective and others displayed moderate
activity while 15, 18, and 19 exhibited scarce activity.
Compound 20 was found to be the most active among
this series. In order to understand the origin of antibac-
terial activity of these cyclic enediynes, antibacterial
activity of control compounds 3, and 4 were deter-
mined. Compound 3 has been used as an intermediate
in the synthesis of cyclic enediyne (6), and simple
Compounds 6, 7 and 8 displayed varied activity against
Escherichia coli and Salmonella typhimurium but only
compounds 7 and 8 (in which R group is –CHO and
–CH2OH, respectively) exhibit potent activity against
Salmonella aureus. Higher activity of 8 as compared
to 7 confers that terminal hydroxyl group is somehow
playing a role in killing mechanism. In the series of cyc-
lic enediyne compounds (13–16) compounds 13 and 16
displayed weak activity against E. coli and S. typhimu-
rium but activity in case of compound 16 (where
R = H) increases twofold higher than compound 13.
All of the cyclic amine endiynes displayed higher MIC
values as compared to their cyclic imine counterparts.

M. C. Joshi et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3226–3230
3229
resorcinol ethers (3,4)20,21 were found inactive upto
1000 lg/mL. So, we believe that the antibacterial activ-
ity of these cyclic enediynes is due to the presence of
enediyne moiety, not due to the resorcinol ether. More-
over studies are currently underway to further optimize
these compounds against bacterial infection.
Combined organic layer was dried over anhydrous
Na2SO4 and filtered. The excess of solvent was evaporated
under vacuo. Residue was purified by column chroma-
tography (15% ethyl acetate in hexane). Yield 1.55 g
(92%); IR (KBr, cmÀ1): 2923, 2122, 1148; 1H NMR
(300 MHz, CDCl3): 2.62 (s, 2H), 4.75 (s, 4H), 6.62–6.78
(m, 3H), 7.19–7.23 (m, 1H); MS (m/z): 186 (M+), 161, 147.
Synthesis of 2,11-dioxa-bicyclo [10.3.1] hexadeca-
1(15),6,12(16),13-tetra-ene-4,8-diyne (6). To a suspension
of Pd(PPh3)4 (224 mg, 0.19 mmol), CuI (164 mg,
0.86 mmol) and n-butylamine (1.58 g, 21.5 mmol) in
anhydrous benzene (30 mL), solution of 3 (800 mg,
4.3 mmol) in 5 mL benzene was added dropwise and
reaction mixture was stirred for 20 min. Then cis-dichlo-
roethylene (420 mg, 4.3 mmol) was added by syringe
under nitrogen atmosphere at 40 ꢁC and reaction mixture
was stirred for 13 h at same temperature. Reaction
mixture was washed with cold hexanes and solvent was
removed under reduced pressure. The crude product thus
obtained was purified by SiO2 column (10% ethyl acetate
in hexane). Yield 600 mg (66%); IR (KBr, cmÀ1): 2922,
2211, 1148; 1H NMR (300 MHz, CDCl3): 4.86 (s, 4H),
5.90 (s, 2H), 6.43 (d, J = 7.4 Hz 1H), 6.63–6.66 (m, 2H),
7.18 (m, 1H); 13C NMR (75.5 MHz, CDCl3): 57.2 (OCH2),
81.49 (Cquart), 92.49 (Cquart), 103.06 (CH), 108.48 (CH),
111.80 (CH), 130.06 (CH), 130.34 (CH), 159.25 (Cquart);
MS (m/z): 210 (M+); Anal. calcd for C14H10O2: C,79.98;
H, 4.79. Found: C, 80.09; H, 4.99.
In conclusion, we have shown for the first time that syn-
thetic enediynes can have potential in the treatment of
microbial infections apart from their use in the antican-
cer drug discovery programme.
Acknowledgements
D.S.R. thanks Department of Science and Technology,
New Delhi, India, for financial support (DO No.: SR/
FTP/CSA-18/2003) and Dr. Santosh Pasha, Institute
of Genomics & Integrative Biology, Mall Road, Delhi,
for her help.
References and notes
1. Smith, A. L.; Nicolaou, K. C. J. Med. Chem. 1996, 39,
2103.
Synthesis
1(16),6,12,14-tetraene-4,8-di-yne-13 carboxaldehyde (7).
To solution of 2,4-dihydroxybenzaldehyde (2.0 g,
of
2,11-dioxabicyclo
[10.3.1]hexadeca-
2. Rawat, D. S.; Zaleski, J. M. Synlett 2004, 393.
3. Davies, J.; Wang, H.; Taylor, T.; Warabi, K.; Huang,
X.-H.; Andersen, R. J. Org. Lett. 2005, 7, 5233.
4. Joshi, M. C.; Joshi, P.; Rawat, D. S. Arkivoc 2006, 16, 65.
5. Shen, B.; Liu, W.; Nonaka, K. Curr. Med. Chem. 2003, 10,
2317.
6. Tuesuwan, B.; Kerwin, S. M. Biochemistry 2006, 45, 7265.
7. Suzuki, I.; Shigenaga, A.; Nemoto, H.; Shibuya, M.
Tetrahedron Lett. 2004, 45, 1955.
8. Banfi, L.; Basso, A.; Guanti, G.; Riva, R. Arkivoc 2006, 7,
261.
9. Benites, P. J.; Holmberg, R. C.; Rawat, D. S.; Kraft, B. J.;
Klein, L. J.; Peters, D. G.; Thorp, H. H.; Zaleski, J. M. J.
Am. Chem. Soc. 2003, 125, 6434.
10. Basak, A.; Bag, S. S.; Basak, A. Bioorg. Med. Chem. 2005,
13, 4096.
11. Lin, C.-F.; Lo, Y.-H.; Hsieh, M.-C.; Chen, Y.-H.; Wang,
J.-J.; Wu, M.-J. Bioorg. Med. Chem. 2005, 13, 3565.
12. Rawat, D. S.; Zaleski, J. M. J. Am. Chem. Soc. 2001, 123,
9675.
a
14.5 mmol), K2CO3 (10 g, 72.5 mmol) in 25 mL dry
DMF, solution of 1,8-dibromo-oct-4-ene-1,6-diyne18
(3.98 g, 15.2 mmol) in 5 mL DMF was added dropwise.
The reaction mixture was stirred at ambient temperature
for 8 h. The reaction mixture was extracted with CHCl3
(6 · 25 mL), and the combined organic layer was washed
with water (6 · 250 mL). The organic layer was dried over
anhydrous Na2SO4 and solvent was removed under
reduced pressure. The crude product was purified over
silica gel column using 10% ethyl acetate in hexanes as an
eluent. Yield 2.14 g (62%); mp. 138 ꢁC; IR (KBr, cmÀ1):
2932, 2225, 1690; 1H NMR (300 MHz, CDCl3): 5.00 (s,
2H), 5.07 (s, 2H), 5.95 (s, 2H), 6.64 (m, 1H), 7.36 (m, 1H),
7.83 (d, J = 6 Hz, 1H), 10.29 (s, 1H); 13C NMR
(75.5 MHz, CDCl3): 57.03 (OCH2), 57.45 (OCH2), 87.45
(Cquart), 87.72 (Cquart), 90.98 (Cquart), 91.11 (Cquart),
101.58 (CH), 112.41 (CH), 120.11 (Cquart), 123.41 (CH),
131.05 (CH), 160.89 (Cquart), 162.92 (Cquart), 188.48
(Cquart); MS (m/z): 238 (M+); Anal. calcd for C15H10O3:
C, 75.62; H, 4.23. Found: C, 75.91; H, 4.42.
13. Jones, G. B.; Fouad, F. S. Curr. Pharm. Des. 2002, 8,
2415.
14. Wang, Z.; He, Q.; Liang, Y.; Wang, D.; Li, Yi-Y.; Li, D.
Biochem. Pharm. 2003, 65, 1767.
15. Liu, X.; He, H.; Feng, Y.; Zhang, M.; Ren, K.; Shao, R.
Anti-cancer Drugs 2006, 17, 173.
16. Lin, C.-F.; Hsieh, P.-C.; Lu, W.-D.; Chiu, H.-F.; Wu,
M.-J. Bioorg. Med. Chem. 2001, 9, 1707.
17. Nicolaou, K. C.; Dia, W.-M. Angew. Chem., Int. Ed. Engl.
1991, 30, 1387.
Synthesis of (2,11-dioxabi-cyclo[10.3.1] hexadeca1(15),6,
12(16),13-tetra-ene-4,8-diyn-13 yl)methanol (8). To a solu-
tion of 7 (0.4 g, 1.6 mmol) in 20 mL dry methanol, sodium
borohydride (0.2 g, 7.1 mmol) was added at 20 ꢁC and
reaction mixture was stirred for 1 h. Solvent from the
reaction mixture was removed under reduced pressure and
crude product was purified by column chromatography
(30% ethyl acetate in hexanes). Yield: 322 mg (80%); mp.
135 ꢁC; IR (KBr, cmÀ1): 3433, 2208, 1590, 1447, 1268, 1099;
1H NMR (300 MHz, CDCl3): 1.98 (br s, 1H), 4.64 (s, 2H,
CH2OH), 4.90 (s, 2H, OCH2), 4.99 (s, 2H, OCH2), 5.90 (s,
2H), 6.62 (d, J = 6Hz, 1H), 7.22 (m, 1H), 7.38 (s, 1H); MS
(m/z): 240 (M+), 223; Anal. calcd for C15H12O3: C,74.99; H,
5.03. Found: C, 74.81; H, 5.22.
18. Konig, B.; Pitsch, W.; Dix, I.; Jones, P. G. Synthesis 1996,
446.
19. Synthesis of 1,3-bis-prop-2-ynyloxy-benzene (3). Mixture of
resorcinol (1.0 g, 9.1 mmol) and K2CO3 (12.5 g,
90.5 mmol) was stirred in 40 mL dry DMF at room
temperature for 30 min. Then propargyl bromide (2.32 g,
24.4 mmol) was added dropwise and reaction mixture was
stirred at room temperature for 7 h. The progress of
reaction was monitored by TLC, and reaction mixture was
extracted with CHCl3 (8 · 30 mL) and H2O (8 · 300 mL).
Synthesis of (2,11-Dioxa-bicyclo[10.3.1]hexadeca-1(16),6,
12,14-tetraene-4,8-diyn-13- ylmethylene)-p-tolyl-amine (13).
Mixture of p-tolylimino-methyl-benzene-1,3-diol (9)
(430 mg, 1.9 mmol) and K2CO3 (2.63 g, 19.1 mmol) in

3230
M. C. Joshi et al. / Bioorg. Med. Chem. Lett. 17 (2007) 3226–3230
15 mL dry DMF was stirred at room temperature for
Synthesis of 4-chlorophenyl-2,11-dioxa-bicyclo[10.3.1]
hexadeca-1(16),6,12,14-tetraene-4,8-diyn-13-yl-methyl)-amine
(18). To a solution of 4-chlorophenyl-2,11-dioxa-bicy-
clo[10.3.1]hexadeca-1(16),6,12,14-tetraene-4, 8-diyn-13-yl
methylene)-amine (14) (150 mg, 0.43 mmol) in 10 mL dry
methanol, NaBH4 (87 mg, 1.42 mmol) was added at room
temperature and reaction mixture was stirred for 2 h. The
excess of solvent was removed under reduced pressure and
crude product was purified by SiO2 (20% ethyl acetate in
hexanes). Yield: 100 mg (66%); mp. 185–187 ꢁC; 1H NMR
(300 MHz, CDCl3): 4.01 (br s, 1H, NH); 4.22 (s, 2H,
NCH2Ph), 4.89 (s, 2H, OCH2), 4.97 (s, 2H, OCH2), 5.90 (s,
2H), 6.54–6.61 (m, 2H), 7.11 (m, 2H), 7.22 (m, 2H), 7.38 (s,
1H); MS (m/z): 349 (M+), 224; Anal. calcd for
C21H16ClNO2: C, 72.10; H, 4.61; N, 4.00; Found: C,
72.03; H, 4.91; N, 4.25.
25 min under nitrogen atmosphere. To this suspension,
1,8-dibromooct-4-ene-2,6-diyne (5)18 (500 mg, 1.9 mmol)
in 10 mL dry DMF was added dropwise and reaction
mixture was stirred at room temperature under nitrogen
atmosphere for 13 h. After completion of the reaction, the
reaction mixture was poured in 50 mL water and product
was extracted with CHCl3 (6 · 20 mL). Combined organic
layer was washed with distilled water (8 · 75 mL) and
finally dried over anhydrous Na2SO4. After filtration,
excess of solvent was removed under reduced pressure. The
crude product was purified over SiO2 column using 8% ethyl
acetate in hexanes as an eluent. Yield: 297 mg(48%). mp
123–125 ꢁC. IR (KBr, cmÀ1): 2923, 2854, 2202, 1592, 1460,
1377, 1264, 1095; 1H NMR (300 MHz, CDCl3): 2.36 (s, 3H),
4.97 (s, 2H), 5.01 (s, 2H), 5.93 (s, 2H), 6.71 (dd, J = 8 Hz,
1.5 Hz, 1H), 7.10 (d, J = 6 Hz, 2H), 7.18 (d, J = 6 Hz, 2H),
7.36 (d, J = 1.5 Hz, 1H), 8.12 (d, J = 6.6 Hz, 1H), 8.78 (s,
1H); MS (m/z): 240 (M+); Anal. calcd for C22H17NO2: C,
80.71; H, 5.23; N, 4.28; Found: C, 81.02; H, 5.38; N, 4.49.
20. Srinivasan, M.; Sankaraman, S.; Hopf, H.; Dix, I.; Jones,
P. G. J. Org. Chem. 2001, 66, 4299.
21. Katritzky, A. R.; He, H.-Y.; Long, Q.; Cui, X.; Level, J.;
Wilcox, A. L. Arkivoc 2000, 3, 240.