Synthesis and antimicrobial studies on novel sulfonamides containing 4-azidomethyl coumarin

Article abstract of DOI:10.1016/j.ejmech.2009.12.022

A series of new and novel coumarin-6-sulfonamides with a free C4-azidomethyl group have been synthesized as antimicrobials in three steps starting from 7-methyl-4-bromomethylcoumarin 1. The reaction of 1 with chlorosulfonic acid was found to yield the corresponding 6-sulfonylchloride 2, which when treated with sodium azide led to intermediate 3. The title sulfonamides 5a-y were obtained from the reaction of 3 with various aromatic amines 4 in refluxing benzene. The chemical structures of the compounds were elucidated by IR, NMR and LC-MS spectral data. All the synthesized compounds have been screened for their in vitro anti-bacterial and anti-fungal activities. Some of the compounds have been found to be active against both bacterial species at a concentration of 1?μg/mL.

Full text of DOI:10.1016/j.ejmech.2009.12.022

European Journal of Medicinal Chemistry 45 (2010) 1151–1157
Contents lists available at ScienceDirect
European Journal of Medicinal Chemistry
journal homepage: http://www.elsevier.com/locate/ejmech
Original article
Synthesis and antimicrobial studies on novel sulfonamides containing
4-azidomethyl coumarin
,
c
Mahantesha Basanagouda a, K. Shivashankar b, Manohar V. Kulkarni a , Vijaykumar P. Rasal ,
*
Harishchandra Patel c, Sumit S. Mutha c, Ashwini A. Mohite c
a P.G. Department of Chemistry, Karnatak University, Dharwad 580 003, Karnataka, India
b P.G. Department of Chemistry, Central College, Bangalore University, Bangalore 560 001, Karnataka, India
c Department of Pharmacology, KLES’s College of Pharmacy, Belgaum-590 010, Karnataka, 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 new and novel coumarin-6-sulfonamides with a free C4-azidomethyl group have been
synthesized as antimicrobials in three steps starting from 7-methyl-4-bromomethylcoumarin 1. The
reaction of 1 with chlorosulfonic acid was found to yield the corresponding 6-sulfonylchloride 2, which
when treated with sodium azide led to intermediate 3. The title sulfonamides 5a–y were obtained from
the reaction of 3 with various aromatic amines 4 in refluxing benzene. The chemical structures of the
compounds were elucidated by IR, NMR and LC-MS spectral data. All the synthesized compounds have
been screened for their in vitro anti-bacterial and anti-fungal activities. Some of the compounds have
Received 5 January 2009
Received in revised form
9 October 2009
Accepted 10 December 2009
Available online 21 December 2009
Keywords:
Coumarin
Sulfonamide
Azide
been found to be active against both bacterial species at a concentration of 1 mg/mL.
Ó 2009 Elsevier Masson SAS. All rights reserved.
Antimicrobial activity
1. Introduction
dichloroacetamidomethyl coumarins [21] for their anti-bacterial
activity. Recent reports on the cytotoxic effects of coumarin sulfon-
amides [22,23], on leukemia cell lines have generated a renewed
interest in such class of compounds. Azido group has been recognized
as a bioorthogonal chemical reporter which is stable in physiological
conditions [24] and has been employed in the study of biosynthetic
pathways of biomolecules [25,26]. It is also expected that the polar
nature of the azide would play an important role in penetrating the
cell wall thus enhancing the bacterial potency of the compounds. In
continuation of our interest in designing oxygen heterocycle based
anti-bacterials [27], we have synthesized new sulfonamides with an
azido group at the allylic position in the coumarin ring and the results
of anti-bacterial and anti-fungal activity have been discussed in this
paper.
Coumarins are a class of lactones which are found widely in nature
and possess antiinflammatory [1,2], antiviral [3], antiHIV [4], anti-
fungal [5] and cytotoxic activities [6,7], which have been reviewed
[8]. In the development of newer antimicrobials, coumarins have
been identified as target specific plant anti-bacterial agents with
growth inhibitory potential particularly against Gram-positive
species [9]. This is based on the related antibiotic novobiocin which
targets the DNA gyrase [10]. Synthetic coumarins with chloram-
phenicol [11] side chain and cephalosporin amides [12] at C-3 posi-
tion, have been screened against a number of bacterial species. One of
the earliest attempts to enhance their antimicrobial potency led to 6-
sulphonamido coumarins [13–16], as anti-tubercular agents. Intro-
duction of a basic side chain at the allylic position with respect to the
biogenetically important C3-C4 double bond [17] was attempted to
reduce the acidic character of coumarin antibiotics. Amongst the
various alkyl amines and sulfides synthesized, the 4-piper-
azinomethyl and imidazomethyl sulfides [18] were found to be the
most active compounds. We have been studying structurally analo-
gous, 4-arylaminomethyl [19] 4-sulphonamidomethyl [20] and 4-
2. Chemistry
4-Bromoethylacetoacetate obtained from the bromination of
ethylacetoacetate, was treated with m-cresol under Pechmann
cyclization conditions using neat sulphuric acid as the condensing
agent. The reaction resulted in the formation of 4-bromomethyl-
7-methylcoumarin 1. The reactivity of this compound has been
explored in a cascaded manner via a regioselective electrophilic
substitution, allylic SN reaction and a nucleophilic displacement at
the oxidized sulfur. Compound 1 was refluxed with excess of
* Corresponding author. Tel.: þ91 836 2215286; fax: þ 91 836 2747884.
E-mail address: manohar274@gmail.com (M.V. Kulkarni).
0223-5234/$ – see front matter Ó 2009 Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.ejmech.2009.12.022

1152
M. Basanagouda et al. / European Journal of Medicinal Chemistry 45 (2010) 1151–1157
chlorosulfonic acid to give 4-bromomethyl-7-methyl-coumarin-6-
sulfonylchloride 2. The regioselective electrophilic substitution at
C-6 is probably because of the electron donating C7-CH3 group
and the spatial proximity of C-6 with the ring oxygen, which
resembles the para-position of phenols.
3. Results and discussion
Structure of C-6 sulfonyl chloride 2 was supported by the
appearance of two singlets in the 1H NMR at 7.40 and 8.45 ppm due
to C5-H and C8-H respectively. The C4-CH2 protons appeared as
singlet at 4.52 ppm where as C3-H observed as singlet at 6.63 ppm.
Formation of azide 3 was indicated by a strong band at 2135 cmꢀ1. It
was the change in the chemical shift of methylene protons to
5.00 ppm which supported the observed regiochemistry of the
nucleophilic displacement at the allylic carbon under the present
experimental conditions. Expectedly there was no significant
change in the aromatic region with singlets appearing at 7.69 and
8.24 ppm. Generation of sulfonamides 5a-y was indicated by a sharp
band around 3360 cmꢀ1 and the azide group remained in-tact
during the nucleophilic displacement at the sulfonyl chloride 3. In
the PMR spectrum of 4-azidomethyl-7-methyl-coumarin-6-
sulfonic acid p-tolylamide (5c) (R ¼ –C6H4–CH3-p) showed two
Treatment 2 with sodium azide in aqueous acetone at room
temperature resulted in the formation of 4-azidomethyl-7-meth-
ylcoumarin-6-sulfonyl chloride 3, the structure of which was sup-
ported by spectral data. Preferential attack of the linear azide
nucleophile at the allylic carbon is favorably due to the better
leaving group ability of bromine and slight steric hindrance caused
by the ortho-methyl group at C-7. However use of two equivalents
of sodium azide at room temperature leads to the formation of bis-
azide expected because of the higher reactivity of sulfonyl chloride
which has been recently reported in the reactions of 4-bromo-
methyl-benzenesulfonyl chloride [28]. Sulfonamides 5a–y were
obtained by refluxing aromatic amines and sulfonyl chloride
derivative 3 in dry benzene. Removal of solvent under reduced
pressure afforded the title compounds as solids which were puri-
fied by routine methods (Scheme 1).
singlets at 2.29 and 2.66 d ppm due to C7-CH3 and CH3 of phenyl ring,
respectively. The NH proton was observed as singlet at 4.49 ppm.
The methylene protons linked to azido group were observed as
Br
Br
Conc. H2SO4
O
H3C
O
O
0 - 5 °C
O
O
H3C
OH
1
ClSO3H
Br
N3
O
Cl
O
S
S
Cl
O
NaN3 / Acetone
Stirr
O
H3C
O
O
H3C
O
O
2
3
NH2
Benzene
Reflux
R
4
N3
H
N
O
S
R
O
H3C
O
O
5a-y
Scheme 1. Synthesis of 4-azidomethyl-7-methylcoumarin-6-sulfonamide 5.

M. Basanagouda et al. / European Journal of Medicinal Chemistry 45 (2010) 1151–1157
1153
singlet in the down field at 4.78 ppm. The C3-H of coumarin was
observed as singlet at 6.47 ppm. The aromatic protons resonated in
the region of 6.55–8.30 ppm. Further evidence for the formation of
sulfonamides was obtained from the mass spectral data. The Mass
spectrum of (5c) (R ¼ –C6H4–CH3-p) shows M þ H peak at m/z 385.
active compounds against C. albicans were 5c, 5b and 5u with 4-
methyl, 3-methyl and 2-chloro-5-nitro substituents exhibiting
MICs of 1, 4 and 4
mg/mL, respectively. The 2,6-dimethyl, 4-
methoxy, 2-chloro-5-nitro and naphthyl exhibited 2
mg/mL MIC
against A. fumigatus. A broad spectrum of anti-fungal activity was
observed in 5b, 5h, 5i, 5y for A. niger. Other compounds those were
effective at 2 mg/mL against P. chrysogenum were 5a, 5d, 5l, 5o, 5r
4. Biological screening
and 5x. The compounds 5a, 5d, 5h, 5l and 5x showed significant
activity against M. fuscus. It is interesting to note that all these
compounds exhibited a better activity than fluconazole.
4.1. Anti-bacterial screening
All the newly synthesized compounds were evaluated for their in
vitro anti-bacterial activity against (i) Gram-positive bacteria: Strep-
tococcus faecalis (MTCC 3382), Staphylococcus aureus (MTCC 3160),
Bacillus subtilis (MTCC 297) and (ii) Gram-negative bacteria: Pseudo-
monas aeruginosa (MTCC 1034), Klebsiella pneumoniae (MTCC 3384)
and Escherichia coli (MTCC 1089) by broth microdilution method [29].
The anti-bacterial data (Table 1) revealed that all the sulfonamides
exhibited moderate activity a few of them highly active, but majority of
them inactive against E. coli. Compounds with methoxy group 5g, 5h,
5. Experimental
5.1. Chemistry
The melting points were determined by open capillary method
and are uncorrected. The IR spectra (KBr disc) were recorded on
a Nicolet-5700 FT-IR spectrophotometer. 1H NMR spectra were
recorded on Bruker 300 MHz spectrometer using CDCl3 and DMSO-
d6 as solvents and TMS as an internal standard. The chemical shifts
5i, 5j, 5k were highly active against S. faecalis with MIC of 1
m-methoxy compound 5h was found to equally effective against P.
aeruginosa and K. pneumoniae with MIC of 1 and 2 g/mL, respectively.
mg/mL. The
are expressed in
d ppm. The mass spectra were recorded using
m
Agilent-single Quartz LC-MS. The elemental analysis was carried
out using Heraus CHN rapid analyzer. The purity of the compound
was checked by T.L.C. All the chemicals purchased were of analyt-
ical grade, and were used without further purification unless
otherwise stated.
The 2-chloro compound 5n and 4-bromo compound 5s were found to
be very active against S. faecalis, P. aeruginosa and K. pneumoniae. Many
compounds exhibited MIC similar to ciprofloxacin against three
bacterial species except E. coli. Highest activity was associated with
mono-nitro, chloro, bromo and methoxy substituents.
5.1.1. Synthesis of 4-bromomethyl-7-methylcoumarin (1)
The required 4-bromomethyl-7-methylcoumarin [19] have been
synthesized by the Pechmann cyclization of m-cresol with 4-bro-
moethylacetoacetate [31].
4.2. Anti-fungal screening
All the sulfonamides were screened for their anti-fungal activity
against Candida albicans, Aspergillus fumigatus, Aspergillus niger,
Penicillium chrysogenum, Mucor fuscus and Fusarium oxysporum in
DMSO by broth micro dilution method [30].
5.1.2. Synthesis of 4-bromomethyl-7-methyl-coumarin-6-sulfonyl
chloride (2)
The screening data (Table 2) gives a spectrum of their activity
and all of them were most active against F. oxysporum . The most
4-Bromomethyl-7-methylcoumarin 1 (2.53 g, 0.01 mol) was
taken in a two-necked R. B. flask with a dropping funnel and reflux
Table 1
Results of anti-bacterial activities of compounds 2, 3 and 5a–y MICs (mg/mL).
Compound
R
Gram-positive
Gram-negative
S. faecalis
S. aureus
B. subtilis
P. aeruginosa
K. pneumoniae
E. coli
2
3
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
5s
5t
5u
5v
–
–
16
31.25
31.25
250
16
8
62.5
>500
1
1
1
1
1
8
4
1
8
4
1
16
1
8
2
4
1
4
16
4
4
16
16
4
16
125
31.25
16
4
2
1
16
31.25
31.25
16
125
125
125
2
125
125
4
2
125
1
8
62.5
4
125
1
31.25
1
1
2
8
1
1
2
8
125
125
125
4
125
125
125
>500
>500
125
>500
>500
250
>500
250
62.5
>500
125
250
>500
>500
>500
>500
>500
>500
>500
>500
>500
250
H
3-CH3
4-CH3
8
1
3,4-CH3
2,4-CH3
2,6-CH3
2-OCH3
3-OCH3
4-OCH3
4-OC2H5
2-NO2
3-NO2
4-NO2
2-Cl
3-Cl
4-Cl
2,4-Cl
3,4-Cl
4-Br
3-Cl, 4-CH3
2-Cl, 5-NO2
2-NO2, 4-Cl
2-NO2, 4-CH3
2-CH3, 4-NO2
Naphthylamine
–
31.25
125
>500
>500
>500
>500
>500
62.5
8
125
125
16
16
250
2
125
62.5
1
62.5
2
1
16
16
125
2
1
4
1
2
1
1
4
2
1
1
8
31.25
4
16
8
2
4
16
16
16
1
>500
4
125
250
250
4
8
62.5
16
4
31.25
1
5w
5x
5y
16
125
8
125
>500
1
2
1
>500
1
Ciprofloxacin
1
1

1154
M. Basanagouda et al. / European Journal of Medicinal Chemistry 45 (2010) 1151–1157
Table 2
Results of anti-fungal activities of compounds 2, 3 and 5a–y MICs (
m
g/mL).
A. fumigatus
Compound
R
C. albicans
A. niger
P. chrysogenum
M. fuscus
F. oxysporum
2
3
–
–
8
8
4
8
4
8
1
4
1
2
1
2
5a
5b
5c
5d
5e
5f
5g
5h
5i
5j
5k
5l
5m
5n
5o
5p
5q
5r
5s
5t
5u
5v
5w
5x
H
3-CH3
4-CH3
16
4
1
8
8
8
8
16
8
16
2
62.5
31.25
2
8
1
4
4
4
8
16
2
2
2
8
16
2
31.25
31.25
>500
125
250
>500
4
4
4
125
1
1
1
1
1
1
1
1
1
2
1
4
1
1
1
1
1
1
1
1
1
2
1
62.5
62.5
1
16
16
31.25
4
250
62.5
8
3,4-CH3
2,4-CH3
2,6-CH3
2-OCH3
3-OCH3
4-OCH3
4-OC2H5
2-NO2
3-NO2
4-NO2
2-Cl
3-Cl
4-Cl
2,4-Cl
3,4-Cl
4-Br
3-Cl, 4-CH3
2-Cl, 5-NO2
2-NO2, 4-Cl
2-NO2, 4-CH3
2-CH3, 4-NO2
Naphthylamine
–
8
31.25
8
8
16
250
16
62.5
62.5
>500
8
>500
250
8
8
8
8
250
2
4
8
62.5
62.5
31.25
31.25
250
16
500
16
8
62.5
31.25
4
1
8
62.5
31.25
2
16
62.5
2
16
4
125
125
31.25
2
31.25
16
>500
16
16
125
250
8
16
16
125
31.25
4
16
16
16
8
31.25
31.25
250
16
62.5
8
62.5
2
31.25
8
500
16
2
500
125
8
5y
2
8
16
8
Fluconazole
8
8
condenser containing a CaCl2 guard tube. Chlorosulfonic acid (4 ml,
0.06 mol) was added through a dropping funnel in small portion
with occasional shaking. The reaction mixture was heated on
a water bath for 1 h (monitored by TLC), cooled and poured on to
crushed ice (100 g). The separated solid was filtered off, washed
with water, dried and recrystallized from chloroform to obtain
a colorless solid in 55% yield. m.p. 170–171 ꢁC; IR (KBr) cmꢀ1 1716
(KBr) cmꢀ1 1716 (C]O), 2124 (N3), 3367 (NH); 1H NMR (DMSO,
300 MHz, TMS): 2.67 (s, 3H, C7-CH3), 5.12 (s, 2H, C4-CH2), 6.37 (s,
1H, C3-H), 6.63–8.39 (m, 8H, Ar-H and NH); Anal. Calcd for
C17H14N4O4S (%): Calcd. C, 55.13; H, 3.81; N, 15.13. Found: C, 55.02;
H, 3.57; N, 15.01.
d
5.1.4.2. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid m-tolyla-
mide (5b). Yellow solid; yield 75%; m.p. 110 ꢁC; IR (KBr) cmꢀ1 1714
(C]O), 2120 (N3), 3345 (NH); 1H NMR (DMSO, 300 MHz, TMS):
(C]O); 1H NMR (CDCl3, 300 MHz, TMS):
d 2.89 (s, 3H, C7-CH3), 4.52
(s, 2H, C4-CH2), 6.63 (s, 1H, C3-H), 7.40 (s, 1H, C8-H), 8.45 (s, 1H, C5-
H); LC-MS: 353 (M þ 2). Anal. Calcd for C11H8BrClO4S (%): Calcd. C,
37.58; H, 2.29. Found: C, 37.21; H, 2.02.
d
2.66 (s, 3H, C7-CH3), 3.51 (s, 3H, 3-CH3 of phenyl), 4.59 (s, 1H, NH,
D2O exchangeable), 4.93 (s, 2H, C4-CH2), 6.34 (s, 1H, C3-H), 6.55–
8.37 (m, 6H, Ar-H); LC-MS 385 (M þ H). Anal. Calcd for C18H16N4O4S
(%): Calcd. C, 56.24; H, 4.20; N, 14.57. Found: C, 56.14; H, 4.13; N,
14.40.
5.1.3. Synthesis of 4-azidomethyl-7-methyl-coumarin-6-sulfonyl
chloride (3)
4-Bromomethyl-7-methyl-coumarin-6-sulfonyl
chloride
2
(3.51 g, 0.01 mol) was taken in acetone (20 ml) in R. B. flask. To this,
solution of sodium azide (0.78 g, 0.012 mol) in water (3 ml) was
added drop wise with stirring. The stirring was continued for 10 h.
Then, the reaction mixture was poured on to crushed ice (100 g).
The separated solid was filtered and recrystallized from benzene ꢀto1
obtain a colorless solid in 65% yield. m.p. 120–121 ꢁC; IR (KBr) cm
1728 (C]O), 2135 (N3, azido); 1H NMR (CDCl3, 300 MHz, TMS):
5.1.4.3. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid p-tolyla-
mide (5c). Brown solid; yield 72%; m.p. 188 ꢁC; IR (KBr) cmꢀ1 1710
(C]O), 2115 (N3), 3341 (NH); 1H NMR (DMSO, 300 MHz, TMS):
d
2.29 (s, 3H, 4-CH3 of phenyl), 2.66 (s, 3H, C7-CH3), 4.49 (s, 1H, NH,
D2O exchangeable), 4.78 (s, 2H, C4-CH2), 6.47 (s, 1H, C3-H), 6.55–
8.30 (m, 6H, Ar-H); LC-MS 385 (M þ H). Anal. Calcd for C18H16N4O4S
(%): Calcd. C, 56.24; H, 4.20; N, 14.57. Found: C, 56.04; H, 4.04; N,
14.39.
d
2.77 (s, 3H, C7-CH3), 4.66 (s, 2H, C4-CH2), 6.67 (s, 1H, C3-H), 7.39
(s, 1H, C8-H), 8.35 (s, 1H, C5-H); Anal. Calcd for C11H8ClN3O4S (%):
Calcd. C, 42.11; H, 2.57; N, 13.39. Found: C, 42.01; H, 2.31; N, 13.11.
5.1.4.4. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (3,4-
dimethyl-phenyl)-amide (5d). Colorless solid; yield 70%; m.p.
190 ꢁC; IR (KBr) cmꢀ1 1712 (C]O), 2115 (N3), 3352 (NH); 1H NMR
5.1.4. Synthesis of 4-azidomethyl-7-methyl-coumarin-6-
sulfonamides (5): general procedure
(DMSO, 300 MHz, TMS): d 2.00 (s, 3H, 4-CH3 of phenyl), 2.66 (s, 3H,
To a mixture of 4-azidomethyl-7-methyl-coumarin-6-sulfonyl
chloride 3 (3.13 g, 0.01 mol) in dry benzene (30 ml) the appropriate
aromatic amines 4 (0.015 mol) were added and heated under reflux
for 4 h (18–20 h for nitro amines) (monitored by TLC). The solvent
was removed under reduced pressure. The residue obtained was
filtered off and crystallized from benzene.
C7-CH3), 3.90 (s, 3H, 3-CH3 of phenyl), 5.00, (s, 2H, C4-CH2), 6.60 (s,
1H, C3-H), 6.70–8.40 (m, 5H, Ar-H), 10.0 (s, 1H, NH, D2O
exchangeable). Anal. Calcd for C19H18N4O4S (%): Calcd. C, 57.27; H,
4.55; N, 14.06. Found: C, 57.03; H, 4.32; N, 13.92.
5.1.4.5. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (2,4-
dimethyl-phenyl)-amide (5e). Brown solid; yield 69%; m.p. 161 ꢁC;
IR (KBr) cmꢀ1 1730 (C]O), 2134 (N3), 3440 (NH); 1H NMR (DMSO,
5.1.4.1. Synthesis of 4-azidomethyl-7-methyl-coumarin-6-sulfonic
acid phenylamide (5a). Colorless solid; yield 70%; m.p. 168 ꢁC; IR
300 MHz, TMS): d 2.13 (s, 6H, 2-CH3 and 4-CH3 of phenyl), 2.69 (s,

M. Basanagouda et al. / European Journal of Medicinal Chemistry 45 (2010) 1151–1157
1155
3H, C7-CH3), 5.16 (s, 2H, C4-CH2), 6.35 (s, 1H, C3-H), 6.70–8.25 (m,
5H, Ar-H), 8.51 (s, 1H, NH, D2O exchangeable). Anal. Calcd for
C19H18N4O4S (%): Calcd. C, 57.27; H, 4.55; N, 14.06. Found: C, 57.12;
H, 4.48; N, 14.01.
5.1.4.13. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (4-nitro-
phenyl)-amide (5m). Yellow solid; yield 61%; m.p. 165 ꢁC; IR (KBr)
cmꢀ1 1726 (C]O), 2123 (N3), 3431 (NH); 1H NMR (DMSO, 300 MHz,
TMS): d 2.68 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2), 6.58 (s, 1H, C3-H),
6.81–7.95 (m, 5H, Ar-H), 8.24 (s, 1H, C5-H), 8.40 (s, 1H, NH, D2O
exchangeable). Anal. Calcd for C17H13N5O6S (%): Calcd. C, 49.16; H,
3.15; N, 16.86. Found: C, 49.10; H, 3.08; N, 16.78.
5.1.4.6. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (2,6-
dimethyl-phenyl)-amide (5f). Green solid; yield 65%; m.p. 185 ꢁC; IR
(KBr) cmꢀ1 1730 (C]O), 2134 (N3), 3440 (NH); 1H NMR (DMSO,
300 MHz, TMS):
d
2.13 (s, 6H, 2-CH3 and 4-CH3 of phenyl), 2.69 (s,
5.1.4.14. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(2-
3H, C7-CH3), 5.16 (s, 2H, C4-CH2), 6.35 (s, 1H, C3-H), 6.70–8.25 (m,
5H, Ar-H), 8.51 (s, 1H, NH, D2O exchangeable). Anal. Calcd for
C19H18N4O4S (%): Calcd. C, 57.27; H, 4.55; N, 14.06. Found: C, 57.12;
H, 4.48; N, 13.97.
chloro-phenyl)-amide (5n). Brown solid; yield 65%; m.p. 184 ꢁC; IR
(KBr) cmꢀ1 1728 (C]O), 2125 (N3), 3409 (NH); 1H NMR (DMSO,
300 MHz, TMS): d 2.67 (s, 3H, C7-CH3), 4.97 (s, 2H, C4-CH2), 6.63 (s,
1H, C3-H), 6.80–7.68 (m, 5H, Ar-H), 8.23 (s, 1H, C5-H), 8.39 (s, 1H,
NH, D2O exchangeable). Anal. Calcd for C17H13ClN4O4S (%): Calcd. C,
50.44; H, 3.24; N, 13.84. Found: C, 50.32; H, 3.16; N, 13.78.
5.1.4.7. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(2-
methoxy-phenyl)-amide (5g). Black solid; yield 75%; m.p. 206 ꢁC; IR
(KBr) cmꢀ1 1739(C]O), 2143 (N3), 3433 (NH); 1H NMR (DMSO,
5.1.4.15. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(3-
300 MHz, TMS):
d
2.68 (s, 3H, C7-CH3), 3.82 (s, 3H, 2-OCH3 of
chloro-phenyl)-amide (5o). Grey solid; yield 61%; m.p. 196 ꢁC; IR
phenyl), 4.66 (s, 2H, C4-CH2), 6.27 (s, 1H, C3-H), 6.47–8.28 (m, 6H,
Ar-H), 8.41 (s, 1H, NH, D2O exchangeable). Anal. Calcd for
C18H16N4O5S (%): Calcd. C, 53.99; H, 4.03; N, 13.99. Found: C, 53.90;
H, 3.92; N, 13.94.
(KBr) cmꢀ1 1719 (C]O), 2128 (N3), 3395 (NH); 1H NMR (DMSO,
300 MHz, TMS): d 2.68 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2), 6.34 (s,
1H, C3-H), 6.58–7.69 (m, 5H, Ar-H), 8.24 (s, 1H, C5-H), 8.38 (s, 1H,
NH, D2O exchangeable); LC-MS 405 (M þ H). Anal. Calcd for
C17H13ClN4O4S (%): Calcd. C, 50.44; H, 3.24; N, 13.84. Found: C,
50.38; H, 3.20; N, 13.80.
5.1.4.8. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(3-
methoxy-phenyl)-amide (5h). Green solid; yield 77%; m.p. 119 ꢁC; IR
(KBr) cmꢀ1 1725 (C]O), 2123 (N3), 3372 (NH); 1H NMR (DMSO,
5.1.4.16. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(4-
300 MHz, TMS):
d
2.66 (s, 3H, C7-CH3), 3.58 (s, 3H, 3-OCH3 of
chloro-phenyl)-amide (5p). Colorless solid; yield 63%; m.p. 140 ꢁC;
phenyl), 4.55 (s, 1H, NH, D2O exchangeable), 4.93 (s, 2H, C4-CH2),
6.38 (s, 1H, C3-H), 7.65 (s, 1H, C8-H), 6.61–8.21 (m, 4H, Ar-H), 8.38
(s, 1H, C5-H); LC-MS 401 (M þ H). Anal. Calcd for C18H16N4O5S (%):
Calcd. C, 53.99; H, 4.03; N, 13.99. Found: C, 53.90; H, 3.95; N, 13.92.
IR (KBr) cmꢀ1 1725 (C]O), 2122 (N3), 3395 (NH); 1H NMR (DMSO,
300 MHz, TMS): d 2.66 (s, 3H, C7-CH3), 4.93 (s, 2H, C4-CH2), 6.62 (s,
1H, C3-H), 7.10–7.65 (m, 5H, Ar-H), 8.22 (s, 1H, C5-H), 8.36 (s, 1H,
NH, D2O exchangeable). Anal. Calcd for C17H13ClN4O4S (%): Calcd. C,
50.44; H, 3.24; N, 13.84. Found: C, 50.39; H, 3.20; N, 13.76.
5.1.4.9. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(4-
methoxy-phenyl)-amide (5i). Green solid; yield 74%; m.p. 200 ꢁC; IR
5.1.4.17. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (2,4-
dichloro-phenyl)-amide (5q). Black solid; yield 67%; m.p. 165 ꢁC; IR
(KBr) cmꢀ1 1726 (C]O), 2121 (N3), 3429 (NH); 1H NMR (DMSO,
(KBr) cmꢀ1 1714 (C]O), 2123 (N3), 3356 (NH); 1H NMR (DMSO,
300 MHz, TMS):
d 2.67 (s, 3H, C7-CH3), 3.39 (s, 3H, 4-OCH3 of
phenyl), 4.83 (s, 2H, C4-CH2), 6.58 (s, 1H, C3-H), 7.05–7.48 (m, 5H,
Ar-H), 7.57 (s, 1H, NH, D2O exchangeable), 8.14 (s, 1H, C5-H); LC-MS
401 (M þ H). Anal. Calcd for C18H16N4O5S (%): Calcd. C, 53.99; H,
4.03; N, 13.99. Found: C, 53.94; H, 3.99; N, 13.94.
300 MHz, TMS): d 2.68 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2), 6.63 (s,
1H, C3-H), 6.81–7.71 (m, 4H, Ar-H), 8.24 (s, 1H, C5-H), 8.40 (s, 1H,
NH, D2O exchangeable). Anal. Calcd for C17H12Cl2N4O4S (%): Calcd.
C, 46.48; H, 2.75; N, 12.75. Found: C, 46.41; H, 2.71; N, 12.70.
5.1.4.10. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(4-
5.1.4.18. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (3,4-
dichloro-phenyl)-amide (5r). Grey solid; yield 62%; m.p. 196 ꢁC; IR
(KBr) cmꢀ1 1726 (C]O), 2123 (N3), 3374 (NH); 1H NMR (DMSO,
ethoxy-phenyl)-amide (5j). Green solid; yield 70%; m.p. 191 ꢁC; IR
(KBr) cmꢀ1 1705 (C]O), 2115 (N3), 3325 (NH); 1H NMR (DMSO,
300 MHz, TMS):
d
1.31 (t, 3H, CH3 of C2H5), 2.64 (s, 3H, C7-CH3), 3.46
300 MHz, TMS): d 2.68 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2), 6.63 (s,
(q, 2H, CH2 of C2H5), 4.51 (s, 2H, C4-CH2), 6.37 (s, 1H, C3-H), 6.51–
7.34 (m, 5H, Ar-H), 7.57 (s, 1H, NH, D2O exchangeable), 8.35 (s, 1H,
C5-H); Anal. Calcd for C19H18N4O5S (%): Calcd. C, 55.06; H, 4.38; N,
13.52. Found: C, 55.00; H, 4.29; N, 13.48.
1H, C3-H), 6.81–7.68 (m, 4H, Ar-H), 8.23 (s, 1H, C5-H), 8.37 (s, 1H,
NH, D2O exchangeable). Anal. Calcd for C17H12Cl2N4O4S (%): Calcd.
C, 46.48; H, 2.75; N, 12.75. Found: C, 46.45; H, 2.69; N, 12.67.
5.1.4.19. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(4-
5.1.4.11. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (2-nitro-
phenyl)-amide (5k). Yellow solid; yield 64%; m.p. 101 ꢁC; IR (KBr)
cmꢀ1 1725 (C]O), 2122 (N3), 3351 (NH); 1H NMR (DMSO, 300 MHz,
bromo-phenyl)-amide (5s). Brown solid; yield 58%; m.p. 175 ꢁC; IR
(KBr) cmꢀ1 1709 (C]O), 2112 (N3), 3354 (NH); 1H NMR (DMSO,
300 MHz, TMS): d 2.65 (s, 3H, C7-CH3), 4.91 (s, 2H, C4-CH2), 6.38 (s,
TMS):
d
2.68 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2), 6.63 (s, 1H, C3-H),
1H, C3-H), 6.61–8.38 (m, 7H, Ar-H and NH). Anal. Calcd for
C17H13BrN4O4S (%): Calcd. C, 45.45; H, 2.92; N, 12.47. Found: C,
45.37; H, 2.88; N, 12.37.
6.91–8.09 (m, 5H, Ar-H), 8.24 (s, 1H, C5-H), 8.40 (s, 1H, NH, D2O
exchangeable). Anal. Calcd for C17H13N5O6S (%): Calcd. C, 49.16; H,
3.15; N, 16.86. Found: C, 49.09; H, 3.08; N, 16.73.
5.1.4.20. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(3-
5.1.4.12. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid (3-nitro-
phenyl)-amide (5l). Yellow solid; yield 62%; m.p. 101 ꢁC; IR (KBr)
cmꢀ11727 (C]O), 2099 (N3), 3344 (NH); 1H NMR (DMSO, 300 MHz,
chloro-4-methyl-phenyl)-amide (5t). Yellow solid; yield 60%; m.p.
154 ꢁC; IR (KBr) cmꢀ1 1726 (C]O), 2135 (N3), 3374 (NH); 1H NMR
(DMSO, 300 MHz, TMS): d 2.15 (s, 3H, 4-CH3 of phenyl), 2.68 (s, 3H,
TMS):
d
2.50 (s, 3H, C7-CH3), 4.80 (s, 2H, C4-CH2), 6.20 (s, 1H, C3-H),
C7-CH3), 4.98 (s, 2H, C4-CH2), 6.34 (s, 1H, C3-H), 6.56–7.75 (m, 4H,
Ar-H), 8.24 (s, 1H, C5-H), 8.39 (s, 1H, NH, D2O exchangeable). Anal.
Calcd for C18H15ClN4O4S (%): Calcd. C, 51.62; H, 3.61; N, 13.38.
Found: C, 51.58; H, 3.55; N, 13.29.
6.30–9.17 (m, 7H, Ar-H and NH). Anal. Calcd for C17H13N5O6S
(%): Calcd. C, 49.16; H, 3.15; N, 16.86. Found: C, 49.02; H, 3.01; N,
16.81.

1156
M. Basanagouda et al. / European Journal of Medicinal Chemistry 45 (2010) 1151–1157
5.1.4.21. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(2-
performed with the test medium supplemented with DMSO at the
same dilutions as used in the experiments.
chloro-5-nitro-phenyl)-amide (5u). Yellow solid; yield 58%; m.p.
153 ꢁC; IR (KBr) cmꢀ1 1729 (C]O), 2122 (N3), 3429 (NH); 1H NMR
(DMSO, 300 MHz, TMS):
d
2.68 (s, 3H, C7-CH3), 4.99 (s, 2H, C4-CH2),
5.3. Anti-fungal screening
6.06 (s, 1H, C3-H), 6.65–7.62 (m, 4H, Ar-H), 8.23 (s, 1H, C5-H), 8.41
(s, 1H, NH, D2O exchangeable). Anal. Calcd for C17H12ClN5O6S
(%): Calcd. C, 45.39; H, 2.69; N, 15.57. Found: C, 45.32; H, 2.61; N,
15.50.
The anti-fungal activity was performed against the following
standard fungal strains: C. albicans, A. fumigatus, A. niger, P. chrys-
ogenum, M. fuscus and F. oxysporum.
The MIC determination of the tested compounds was investi-
gated in comparison with Fluconazole by broth micro dilution
method [30]. Double dilutions of the test compounds and reference
drugs were prepared in Sabouraud’s dextrose broth. 10 mg of each
test compounds were dissolved in 1 ml of dimethyl sulfoxide
(DMSO) separately to prepare stock solution. Further progressive
dilutions with Sabouraud’s dextrose broth were performed to
obtain the required concentrations of 500, 250, 125, 62.5, 31.25, 16,
5.1.4.22. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(4-
chloro-2-nitro-phenyl)-amide (5v). Yellow solid; yield 56%; m.p.
189 ꢁC; IR (KBr) cmꢀ1 1726 (C]O), 2122 (N3), 3430 (NH); 1H NMR
(DMSO, 300 MHz, TMS): d 2.68 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2),
6.63 (s, 1H, C3-H), 6.81–7.69 (m, 4H, Ar-H), 7.93 (s, 1H, NH, D2O
exchangeable), 8.23 (s,1H, C5-H). Anal. Calcd for C17H12ClN5O6S (%):
Calcd. C, 45.39; H, 2.69; N, 15.57. Found: C, 45.35; H, 2.64; N, 15.54.
8, 4, 2, 1
m
g/ml. The Petri dishes were inoculated with 1–5 ꢂ 104
5.1.4.23. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(4-
colony forming units (cfu/ml) and incubated at 25 ꢁC for 48–72 h.
The minimum inhibitory concentration (MIC) was the lowest
concentration of the tested compound that yield no visible growth
on the plates. To ensure that the solvent had no effect on the fungal
growth, a control was performed with the test medium supple-
mented with DMSO at the same dilutions as used in the experi-
ments. The results are incorporated in Table 2.
methyl-2-nitro-phenyl)-amide (5w). Yellow solid; yield 59%; m.p.
179 ꢁC; IR (KBr) cmꢀ1 1725 (C]O), 2122 (N3), 3347 (NH); 1H NMR
(DMSO, 300 MHz, TMS): d 2.18 (s, 3H, 4-CH3 of phenyl), 2.67 (s, 3H,
C7-CH3), 4.97 (s, 2H, C4-CH2), 6.63 (s, 1H, C3-H), 6.80–7.68 (m, 4H,
Ar-H), 7.74 (s, 1H, NH, D2O exchangeable), 8.23 (s, 1H, C5-H). Anal.
Calcd for C18H15N5O6S (%): Calcd. C, 50.35; H, 3.52; N, 16.31. Found:
C, 50.29; H, 3.41; N, 16.27.
6. Conclusions
5.1.4.24. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic
acid
(2-
methyl-4-nitro-phenyl)-amide (5x). Yellow solid; yield 60%; m.p.
We have synthesized a series of coumarin sulfonamides bearing
an azido group. The investigation of antimicrobial screening data
reveals that the synthesized compounds showed significant anti-
bacterial activity to both Gram-positive and Gram-negative species.
In many cases these showed equivalent to Ciprofloxacin. On the
other hand these compounds showed fungicidal activity which was
higher than Fluconazole. As a result, we can say that these are
better fungicidal when their antimicrobial activity is compared.
Hence, it is concluded that there is ample scope for further study in
developing these as good lead compounds in developing new
antimicrobials.
110 ꢁC; IR (KBr) cmꢀ1 1720 (C]O), 2121 (N3), 3360 (NH); 1H NMR
(DMSO, 300 MHz, TMS): d 2.08 (s, 3H, 2-CH3 of phenyl), 2.65 (s, 3H,
C7-CH3), 5.39 (s, 2H, C4-CH2), 6.57 (s, 1H, C3-H), 6.72–8.27 (m, 6H,
Ar-H and NH). Anal. Calcd for C18H15N5O6S (%): Calcd. C, 50.35; H,
3.52; N, 16.31. Found: C, 50.11; H, 3.29; N, 16.12.
5.1.4.25. 4-Azidomethyl-7-methyl-coumarin-6-sulfonic acid naph-
thalen-1-ylamide (5y). Grey solid; yield 64%; m.p. 199 ꢁC; IR (KBr)
cmꢀ1 1729 (C]O), 2128 (N3), 3410 (NH); 1H NMR (DMSO, 300 MHz,
TMS): d 2.69 (s, 3H, C7-CH3), 4.98 (s, 2H, C4-CH2), 6.39–8.05 (m, 9H,
Ar-H), 8.24 (s, 1H, C5-H), 8.55 (s, 1H, NH, D2O exchangeable). Anal.
Calcd for C21H16N4O4S (%): Calcd. C, 59.99; H, 3.84; N, 13.33. Found:
C, 59.85; H, 3.78; N, 13.27.
Acknowledgements
The authors thank the University Sophisticated Instrumentation
Centre (USIC), Karnatak University, Dharwad for IR, NMR and
analytical data. Mr. Mahantesha Basanagouda is grateful to Karna-
tak University, Dharwad for providing University Research
Studentship.
5.2. Anti-bacterial screening
Standard strains were procured from the microbial type culture
collection and Gene Bank (MTCC), Institute of Microbial Tech-
nology, Chandigarh, India. The anti-bacterial activity of the
synthesized compounds 2, 3, 5a-y were performed in vitro against
(i) Gram-positive bacteria: S. faecalis (MTCC 3382), S. aureus (MTCC
3160), B. subtilis (MTCC 297) and (ii) Gram-negative bacteria: P.
aeruginosa (MTCC 1034), K. pneumoniae (MTCC 3384) and E. coli
(MTCC 1089) by broth micro dilution method [29].
The MIC determination of the tested compounds was investi-
gated in comparison with Ciprofloxacin. Double dilutions of the test
compounds and reference drugs were prepared in Muller-Hinton
agar. 10 mg of each test compounds were dissolved in 1 ml of
dimethyl sulfoxide (DMSO) separately to prepare stock solution.
Further, progressive dilutions with melted Mueller–Hinton agar
were performed to obtain the required concentrations of 500, 250,
References
[1] K.C. Fylaktakidou, D.J. Hadjipavlou-Litina, K.E. Litinas, D.N. Nicolaides, Curr.
Pharm. Design 10 (2004) 3813–3826.
[2] M.D. Ghate, M.V. Kulkarni, R. Shobha, S.Y. Kattimani, Eur. J. Med. Chem. 38
(2003) 297–304.
[3] J.R. Hwu, R. Singha, S.C. Hong, Y.H. Chang, A.R. Das, I. Vliegen, E. De Clercq,
J. Neyts, Antiviral. Res. 77 (2008) 157–162.
[4] V. Reutrakul, P. Leewanich, P. Tuchinda, M. Pohmakotr, T. Jaipetch, S. Sophasan,
T. Santisuk, Planta. Med. 69 (2003) 1048–1050.
[5] S. Sardari, Y. Mori, K. Horita, R.G. Micetich, S. Nishibe, M. Daneshtalab, Bioorg.
Med. Chem. 7 (1999) 1933–1940.
[6] D. Egan, P. James, D. Cooke, R. O’Kennedy, Cancer Lett. 118 (1997) 201–211.
[7] P. Valenti, A. Rampa, M. Recanatini, A. Bisi, F. Belluti, P. Da Re, M. Carrara,
L. Cima, Anticancer. Drug Des 12 (1997) 443–451.
[8] M.V. Kulkarni, G.M. Kulkarni, C.H. Lin, C.M. Sun, Curr. Med. Chem. 13 (2006)
2795–2818.
[9] K. Lewis, F.M. Ausubel, Natur. Biotech. 24 (2006) 1504–1507.
[10] M. Gellert, M.H. O’dea, T. Itoh, J.I. Tomizawa, Proc. Natl. Acad. Sci. USA 73
(1976) 4474–4478.
125, 62.5, 31.25, 16, 8, 4, 2, 1 mg/ml. The Petri dishes were inoculated
with 1–5 ꢂ104 colony forming units (cfu/ml) and incubated at 37 ꢁC
for 18 hours. The minimum inhibitory concentration (MIC) was the
lowest concentration of the tested compounds that yield no visible
growth on the plate was recorded in Table 1. To ensure that the
solvent had no effect on the bacterial growth, a control was
[11] H.S. Bevinakatti, V.V. Badiger, Arch. Pharm. (Weinheim) 314 (1981) 162–167.
[12] L. Bonsignore, F. Cottiglia, H. Elkhaili, F. Jehl, S.M. Lavagna, G. Loy, F. Manna,
H. Monteil, D. Pompei, D. Secci, Il. Farmaco 53 (1998) 425–430.

M. Basanagouda et al. / European Journal of Medicinal Chemistry 45 (2010) 1151–1157
1157
[13] K.A. Jensen, S.A.K. Christensen, Acta Chem. Scand. 3 (1947) 207.
[14] A.M. Islam, A.H. Bedair, A.A. El-Maghraby, F.M. Aly, H.A. Imam, Indian J. Chem.
21B (1982) 487.
[15] S.S. Hanmantgad, M.V. Kulkarni, V.D. Patil, Indian J. Chem. 24B (1985) 459–461.
[16] C.D. Lakkannavar, M.V. Kulkarni, V.D. Patil, J. Indian Chem. Soc. 69 (1992)
399–400.
[23] A. Purohit, L.W.L. Woo, D. Barrow, H.A.M. Hejaz, R.I. Nicholson, B.V.L. Potter,
M.J. Reed, Mol. Cell. Endocrinol. 171 (2001) 129–135.
[24] N.J. Agard, J.M. Baskin, J.A. Prescher, A. Lo, C.R. Bertozzi, ACS Chem. Biol. 1
(2006) 644–648.
[25] D.H. Dube, C.R. Bertozzi, Curr. Opin. Chem. Biol. 7 (2003) 616–625.
[26] J.W. Chin, T.A. Cropp, J.C. Anderson, M. Mukherjee, Z. Zhang, P.G. Schultz,
Science 301 (2003) 964–967.
[27] I.A. Khan, M.V. Kulkarni, C.M. Sun, Eur. J. Med. Chem. 40 (2005) 1168–1172.
[28] X. Hu, J. Sun, H.G. Wang, R. Manetsch, J. Am. Chem. Soc. 130 (2008)
13820–13821.
[29] E.W. Koneman, S.D. Allen, W.M. Janda, P.C. Schreckenberger, W.C. Winn. Jr.,
Color Atlas and Textbook of Diagnostic Microbiology, fifth ed. Lippincott-
Raven Publishers, Philadelphia, USA, 1997, pp. 785–856.
[30] Z.K. Khan, In vitro and vivo screening techniques for bioactivity screening and
evaluation. in: Proceedings of the International Workshop UNIDO-CDRI, 1997,
pp. 210–211.
[17] S.L. Born, A.S. Fix, D. Caudill, L.D. Lehmann-Mckeeman, Toxicol. Appl. Phar-
macol. 151 (1998) 45–56.
[18] P. Laurin, D. Ferroud, L. Schio, M. Klich, C.D. Hamelin, P. Mavais, P. Lassaigne,
A. Bonnefoy, B. Musicki, Bioorg. Med. Chem. Lett. 9 (1999) 2875–2880.
[19] M.V. Kulkarni, V.D. Patil, Arch. Pharm. (Weinheim) 314 (1981) 708–711.
[20] S.S. Hanmanthagad, M.V. Kulkarni, V.D. Patil, Ind. J. Chem. 25B (1986)
779–781.
[21] C.D. Lakkannavar, K.Y. Anklekar, G.M. Kulkarni, M.V. Kulkarni, Ind. J. Chem. 42B
(2003) 1548–1550.
[22] N.S. Reddy, M.R. Mallireddigari, S. Cosenza, K. Gumireddy, S.C. Bell, E.P. Reddy,
M.V. Ramana Reddy, Bioorg. Med. Chem. Lett. 14 (2004) 4093–4097.
[31] A. Burger, G.E. Ullyot, J. Org. Chem. 12 (1947) 346.