Preparative access to transformation products (TPs) of furosemide: A versatile application of anodic oxidation

Article abstract of DOI:10.1016/j.tet.2011.10.006

Furosemide, a pharmaceutical prescribed for the treatment of edema and hypertension, is a known contaminant of water. In this study, chemoselective anodic oxidation was implemented to assist in the identification and the preparation of furosemide transformation products (TPs), i.e., compounds deriving from furosemide and likely to appear in the environment. An aniline and a pyridinium are proposed as plausible TPs and an analytical study of the pyridinium is presented.

Full text of DOI:10.1016/j.tet.2011.10.006

Tetrahedron 67 (2011) 9518e9521
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Preparative access to transformation products (TPs) of furosemide: a versatile
application of anodic oxidation
*
ꢀ
ꢀ
Celine Laurence, Michael Rivard, Isabelle Lachaise, Julia Bensemhoun, Thierry Martens
ꢁ
ꢀ
ꢀ
ꢀ
Electrochimie et Synthese Organique, Institut de Chimie et des Materiaux Paris-Est (ICMPE), UMR 7182 CNRS, Universite Paris Est Creteil UPEC,
2-8 rue Henri Dunant, 94320 Thiais, France
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 15 August 2011
Received in revised form 3 October 2011
Accepted 3 October 2011
Available online 12 October 2011
Furosemide, a pharmaceutical prescribed for the treatment of edema and hypertension, is a known
contaminant of water. In this study, chemoselective anodic oxidation was implemented to assist in the
identification and the preparation of furosemide transformation products (TPs), i.e., compounds deriving
from furosemide and likely to appear in the environment. An aniline and a pyridinium are proposed as
plausible TPs and an analytical study of the pyridinium is presented.
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Anodic oxidation
Furosemide
Transformation products
Metabolites
1. Introduction
edematous states and hypertension. It is rapidly eliminated by renal
excretion and by biotransformation, especially conjugation.5 It has
The ubiquitous presence of drugs residues in aquatic systems
has justified a large number of studies over the last decade.1
Resulting from natural excretion (sometimes under unmeta-
bolised form) or inappropriate disposal of unused or expired
medication, these compounds contaminate surface-, ground-, and
drinking water thus exposing the environment and humans to their
potential effects.2 In this context, computational studies have pro-
posed models to assess the effects of contaminants on human
health.3 According to a prioritization methodology, lists have also
been established, identifying pharmaceuticals products (PPs) that
might pose a risk.4 One of them,4a based on the evaluation of
predicted environmental concentration (PEC), has classified forty
PPs (including furosemide 1) as highest risk compounds. However,
all of these computational studies are based on available data (such
as pharmacological- and environmental toxicological data), which
often have to be completed. In particular, a better risk assessment
would be obtained by taking into account the fate of PPs and more
precisely the impact of their (a)biotic transformation products (TPs)
susceptible to appear in the environment.
been unambiguously detected both in sewage treatment plants and
rivers.6 Some of its TPs have been described and all of them (except
conjugates) stem from an initial oxidation process (Scheme 1).
O
CHO
H
N
H
N
H
N
Cl
Cl
Cl
O
O
O
H2NO2S
CO2H
H2NO2S
CO2H
H2NO2S
CO2H
1
2
3
Cl
H2NO2S
NH2
CO2H
Cl
H2NO2S
N
Cl
N
O
OH
CO2H
H2NO2S
CO2
4
5
6
Scheme 1. Furosemide (1) and its TPs.
A first class of furosemide TPs highlighted the ability of the furan
moiety to subject oxidation. The metabolic oxidation of furosemide
by cytochrome P450 led either to the postulated epoxide (2) or to
its g
-ketoenal equivalent (3).7 Interestingly, these intermediates are
Furosemide (1), one of the forty highest risk compounds (with
PEC >100 ng/L) is a sulfonamide derived from anthranilic acid.
Classified as loop diuretic, it is prescribed for the treatment of
suspected to induce hepatotoxicity in mice by covalently binding to
biologic substrates. The chemical oxidation by dimethyldioxirane in
acetone leads also to the g-ketoenal intermediate (3), which rear-
ranges in the reaction mixture to give a zwitterionic pyridinium
(4).8
A second class of TPs revealed the ability of the nitrogen atom of
the amino group to undergo also oxidation. The electrochemical
* Corresponding author. Tel.: þ33 1 49 78 11 56; fax: þ33 1 49 78 11 48; e-mail
address: martens@icmpe.cnrs.fr (T. Martens).
0040-4020/$ e see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2011.10.006

ꢀ
C. Laurence et al. / Tetrahedron 67 (2011) 9518e9521
9519
oxidation at gold electrode of furosemide led to the imine (5).9
Aniline (6) was obtained either after metabolisation by the fun-
gus Cuninghamella elegans10 or in rat.7 Chemical oxidation of fu-
rosemide by diperiodatocuprate(III)11 or hydrogenolysis on
palladium12 also led to the aniline (6).
Except in the case of the aniline (6), no preparative access has
been described for these TPs, thus ruling out any toxicological study
or the development of any protocol able to detect them in envi-
ronmental matrices. In this context, the development of such pre-
parative access appeared as a prerequisite for any attempt to study
the environmental impact of furosemide. Considering previous
work9 and the well documented literature about application of
anodic oxidation to generate metabolites,13 the electrochemical
approach seemed to be indicated for this purpose.
curve). At the same time, the signal of bromides was also affected,
whereas the oxidation peak was increasing, the reduction peak
(0.43 V vs SCE) has totally disappeared.
2.2. Preparation of TPs
Preparative oxidation was firstly carried out in methanol at
room temperature in an undivided cell, using two graphite carbon
electrodes. The reaction was complete after 2.5 F/mol with a con-
stant current of 10 mA (current density: 1.25 mA/cm2). HPLC
analysis (MeOH/AcOH 0.1%, 50/50) of the reaction mixture revealed
the presence of furaldehyde (7) and aniline (6). 1% of diacetal (8)
was also detected, indicating the good chemoselectivity of the ox-
idation (Scheme 2).
H
2. Results
Cl
N
Cl
N
i
OMe
O
O
1
+
OMe
2.1. Cyclic voltametry
H2NO2S
CO2H
H2NO2S
CO2H
5
8 (1%, not isolated)
The cyclic voltametry of furosemide at glassy carbon electrode
in methanol revealed two oxidation peaks at 1.4 V and 1.6 V versus
SCE (Fig. 1, blue curve). In agreement with the literature14 they
were, respectively, attributed to the nitrogen atom of the aniline
function and to the furan ring. No peaks were observed in the
return scan, indicating an irreversible process.
H+
+
6 (45%, isolated)
OHC
O
7
i = carbon anode, MeOH, NEt4BF4, 10mA, 2.5F/mol, RT
Scheme 2. Direct oxidation of furosemide (1) in MeOH.
In an attempt to isolate the imine (5), size exclusion chroma-
tography (MeOH/CH2Cl2, 50/50) was conduced on the concentrated
reaction mixture. Unfortunately, no trace of imine could be isolated
and a pure fraction of aniline (6) was obtained with 45% yield.
The mediated oxidation was also tested. In the presence of
ammonium bromide (20 mol %) the reaction was done at constant
current of 10 mA (current density: 1.25 mA/cm2) in an undivided
cell, at room temperature. In such conditions, the reaction was
complete after 2.5 F/mol and appeared to be chemoselective in
favor of the diacetal (8), since no trace of aniline (6) was observed
by HPLC. Concentration of the reaction mixture gave a solid, which
was triturated with ethyl acetate. The mixture of dimethoxylated
diastereoisomers (8) was obtained after evaporation of the solvent
and without any further purification with 88% yield.
With the aim of both scaling up the process and keeping a rea-
sonable reaction time, the mediated oxidation was led at 30 and
50 mA. The influence of bromide concentration was studied in both
cases. The composition of the solution was analyzed after 2.5 F/mol
by HPLC (MeOH/AcOH 0.1%, 50/50) and quantification of aniline (6)
and diacetal (8) was done using authentic standard (Table 1).
Clearly, higher intensity revealed a negative effect on the chemo-
selectivity. In the presence of 20% of ammonium bromide,
900
800
700
600
500
400
300
200
100
0
Furosemide (5mM)
Furosemide (5mM) + 1 eq NEt4Br
NEt4Br (5mM)
0,6
0,8
1
1,2
1,4
1,6
1,8
2
Fig. 1. Cyclic voltametry in MeOH (50 mV/s); 0.1 M NEt4BF4; glassy carbon electrode.
It should be noted that the possibility of a mediated oxidation of
furosemide was demonstrated in the presence of 1 equiv of bro-
mides (1.2 V vs SCE). In such conditions, the intensity of the peak
attributed to the nitrogen atom was indeed decreasing (Fig. 1, red
Table 1
Mediated oxidation of furosemide (1) in MeOH (HPLC monitoring)
H
N
H
n eq NH4Br, I (mA)
carbon anode, 2.5F/mol
Cl
NH2
Cl
N
Cl
Cl
N
OH
OMe
O
O
+
+
OMe
H2NO2S
CO2H
MeOH, RT, NEt4BF4
H2NO2S
CO2
H2NO2S
CO2H
H2NO2S
CO2H
1
6
8
4 (insoluble)
Entrya
n (equiv NH4Br)
I¼30 mAb
I¼50 mAc
6
8
6
8
1
2
3
4
0.2
0.4
0.6
0.8
31
23
29
0
27
10
23
15
15
11d
11
31
42
27
38
91
a
Conditions: 0.26 mmol of (1), 10 mL MeOH, carbon electrodes, 0.13 mmol NEt4BF4, rt.
b
c
J¼3.75 mA/cm2.
J¼6.25 mA/cm2.
3% of (1) observed.
d

ꢀ
C. Laurence et al. / Tetrahedron 67 (2011) 9518e9521
9520
significant amount of aniline (6) was observed at 30 mA, whereas
no trace had been previously detected at 10 mA (vide supra). This is
consistent with the idea of a competition between mediated and
direct oxidation, the latter appearing when the capacity of the first
is overtaken.
To circumvent the production of aniline, the concentration of
bromide was increased. At 30 mA, 80% of bromide were necessary
to obtain a complete selectivity (entries 2e4). At 50 mA, the same
behavior is observed although the production of aniline could not
been totally prevented (even with 100% of bromide).
Besides the problem of chemoselectivity, the increase of current
intensity has also resulted in driving the formation of products other
than those expected. Among them, the pyridinium (4) resulting
from the acid decomposition of the desired diacetal (8) was ob-
tained. If the quantification of pyridinium has not been possible by
HPLC because of its very low solubility, it has been however ob-
served at 30 and 50 mA. The strong acidity locally generated at the
surface of the electrode and resulting from the oxidation might
explain this unwanted decomposition. The presence of inorganic
base in the mixture, such as cesium carbonate, did not improve the
yield of (8), other side-products appearing during the reaction.
Pyridinium (4), which revealed to be insoluble in most of the
organic solvents (except polar ones, such as DMSO), was easily
obtained in a quantitative yield after acidic hydrolysis of (8) at 40 ꢀC
in a (60/40) mixture of aqueous TFA (10 mM)/MeCN and filtration.
In order to initiate the study of pyridinium (4), a cyclic volta-
metry analysis was done in DMSO. The fact that no peak was ob-
served in oxidation, led us to suppose that this compound should
be a more persistent contaminant than the furosemide itself. On the
other hand, the reduction peak observed at ꢁ1.98 V versus SCE
(Fig. 2), highlighted its ability to behave as an electron acceptor. The
comparison with ferrocene indicated monoelectronic oxidant
properties (considering a charge transfer coefficient
a¼0.45).
200
150
pyridinium
ferrocene
100
50
0
-2,5
-2
-1,5
-1
-0,5
0
0,5
1
-50
-100
Fig. 2. Cyclic voltametry in DMSO (500 mV/s); 0.1 M NEt4BF4; glassy carbon electrode.
Unlike most of viologens,19 pyridinium (4) showed an irrevers-
ible reduction process presumably due to the recombination of the
resulting radicals. Interestingly, the reduction peak also appears at
much lower potential than those usually observed for dipyridinium
and pyridinium salts. This latter point seems to exclude the possi-
bility for pyridinium (4) to be reduced by NADPH and NADH.
However, the case of neurotoxic pyridinium salts also unable to
undergo bio-reduction20 should prompt further investigations.
3. Discussion
Anodic oxidation has demonstrated its ability to oxidize with an
excellent selectivity the amine function as well as the furan ring of
furosemide, depending on the presence or not of bromide in the
reaction mixture. Surprisingly, if the anodic oxidation of furan rings
in the presence of bromide ions15 has been abundantly described
and successfully applied to organic synthesis,16 the possibility to
oxidize the furan ring of furosemide remains somewhat un-
expected, given the potentials measured by cyclic voltametry. In the
case of furosemide, it might be suggested that a first oxidized form
(only obtained in the presence of bromide, such as a bromamine)
reacts in an intramolecular fashion to give the diacetal (8).
With this diacetal, our aim was to obtain a synthetic equivalent
of epoxide (2) or alternatively of ketoenal (3). Unfortunately, this
compound revealed an unexpected stability: surprisingly strong
acidic conditions (pH¼2.2) were indeed required to activate it.
More notably, attempts to obtain adducts from this diacetal in the
presence of N-acetyl cysteine or N-acetyl lysine,7 remained un-
successful: pyridinium (4) was invariably obtained.
4. Conclusions
It appears from this work that aniline (6) and pyridinium (4) are
susceptible to be present in the environment as TPs of furosemide.
Their access on preparative scale allows now to focus on them,
toxicological and environmental investigations.
The access to both compounds was easily led by anodic oxida-
tion. This reaction revealed to be highly chemoselective, the nature
of the oxidized moiety depending on the presence or not of bro-
mide in the reaction mixture. This versatility illustrated the rele-
vance of the electrochemical tool in the elaboration of new
heterocyclic structures.
For this reason, the preparation of more reactive species than (8)
was attempted. Anodic oxidation of furosemide in trifluoroethanol
or in a MeCN/AcOH mixture was done in order to obtain, re-
spectively, the fluorinated diacetal or the diacetoxylated de-
rivative.17 None of the expected products could be obtained.
It should be also noted that in all our reactions and in accordance
with the observation done on analytical scale,8 no significant
5. Experimental section
5.1. Chemistry
All chemicals were purchased from either Aldrich or Acros or-
ganic. Solvents were commercially available and used as received
without any further purification.
amount of g-ketoenal (3) could be isolated or has even been ob-
served. This led us to postulate that this is a species, whose presence
in the environment seems very unlikely, as that of the imine (5).
However, it is clear from this work that aniline (6) and pyr-
idinium (4) are two furosemide TPs whose presence (or even ac-
cumulation) in the environment is very likely. Concerning the
aniline (which do not enter in the synthesis of furosemide18), it
seems that no data refers to its toxicity, even though this com-
pound has long been known as a derivative of furosemide.12 Re-
garding the pyridinium, the synthesis presented here is the first on
preparative scale: this explains the absence of data (such toxico-
logical) relating to it.
5.2. Analysis procedures
The electrochemical experiments were performed with voltalab
Radiometer analytical instruments (PST006) and were carried out
in a 20 mL single compartment three-electrode glass cell with
a 1 mm diameter glassy carbon electrode (GCE) as working elec-
trode, a gold wire as counter electrode, and a saturated calomel
electrode as reference electrode. All experiments were carried out
at ambient temperature. The GCE was polished using 0.3
before each experiment.
m Al2O3

ꢀ
C. Laurence et al. / Tetrahedron 67 (2011) 9518e9521
9521
The GCE was placed in 0.1 M Et4NBF4 electrolyte and various
voltammograms were recorded from 25 to 500 mV/s. The cyclic
voltammograms of the investigated compounds were obtained in
methanol.
8.00e7.87 (m, 4H); 13C NMR (100 MHz, DMSO-d6):
d¼164.8, 158.0,
143.2, 142.7, 134.9, 134.4, 133.1, 133.1, 131.5, 131.4, 129.8, 127.4; IR:
3096, 1613, 1573, 1494, 1379, 1321, 1252, 1155, 1026, 963, 927, 865,
805, 694, 654, 603, 584 cmꢁ1; HRMS m/z calcd for C12H10ClN2O5S
[MþH]þ: 328.9999. Found: 328.9995. Anal. Calcd for
C12H9ClN2O5S$2H2O: C 39.51%, H 3.59%, N 7.68%. Found: C 39.47%, H
3.61%, N 7.46%.
Analysis of reaction mixtures was done by reverse phase chro-
matography using a VWR Hitachi Elite LaChrom HPLC chromato-
graph, controlled by an EZCHROM elite program. It was fitted with
a quaternary pump Hitachi L-2130, a Hitachi column oven L-2300
with thermostat at 40 ꢀC, and a Purospher RP-18 column (5
mm,
Acknowledgements
250 mmꢂ4.6 mm) coupled with a photodiode array detector
(L-2455) at 233 nm. The column was eluted at isocratic mode with
a mobile phase composed of MeOH/AcOH 0.1% (50/50 v/v) at a flow
ꢁ
C.L. is grateful to the Ministere de la Recherche for a grant. M.R.
ꢀ
and J.B. thank the Universite Paris-Est Creteil (UPEC), for financial
rate of 0.8 mL/min and sample volumes were 20 mL.
support.
5.2.1. 2-Amino-4-chloro-5-sulfamoyl-benzoic acid (6). Furosemide
(0.5 mmol) was dissolved in methanol (10 mL) and tetraethy-
lammonium tetrafluoroborate (0.4 mmol) was added and
employed as the supporting electrolyte. The furosemide was oxi-
dized at room temperature at a plate of carbon (I¼10 mA) in un-
divided cell under magnetic agitation. After consumption of 2.5
electrons per molecule, the electrolysis was stopped. The methanol
was removed under reduced pressure and the residue was purified
by size exclusion chromatography on Sephadex LH-20 (61 mg, 45%).
The expected product was obtained as a white solid. tR(HPLC)¼
2.83 min; mp 278 ꢀC (decomp.); 1H NMR (400 MHz, acetone-d6):
Supplementary data
Supplementary data associated with this article can be found in
the online version at doi:10.1016/j.tet.2011.10.006.
References and notes
1. (a) Zuccato, E.; Calamari, D.; Natangelo, M.; Fanelli, R. Lancet 2000, 355,
1789e1790; (b) Fent, K.; Weston, A. A.; Caminada, D. Aquat. Toxicol. 2006, 76,
122e159.
d
¼8.54 (s, 1H), 7.16 (br s, 2H), 7.02 (s, 1H), 6.50 (br s, 2H); 13C NMR
€
2. (a) Pharmaceuticals in the Environment, 3rd ed.; Kummerer, K., Ed.;
Springer
: Berlin Heidelberg, 2008; (b) Zuccato, E.; Castiglioni, S.;
(100 MHz, acetone-d6):
d
¼168.7, 155.4, 137.0, 134.9, 128.0, 118.7,
Fanelli, R. J. Hazard. Mater. 2005, 122, 205e209; (c) Richards, S. M.;
Cole, S. E. Ecotoxicology 2006, 15, 647e656; (d) Collier, A. C. Eco-
Health 2007, 4, 164e171; (e) Vulliet, E.; Cren-Olive, C.; Grenier-
108.0; IR: 3498, 3370, 3248, 1662, 1616, 1536, 1419, 1326, 1241, 1159,
1127, 968, 882, 842, 715, 686, 628, 597 cmꢁ1; HRMS m/z calcd for
C7H8ClN2O4S [MþH]þ: 250.9888. Found: 250.9831.
ꢀ
Loustalot, M.-F. Environ. Chem. Lett. 2011, 9, 103e114.
3. (a) Munoz, I.; Gomez, M. J.; Molina-Diaz, A.; Huijbregts, M. A. J.; Fernandez-
Alba, A. R.; Garcia-Calva, E. Chemosphere 2008, 74, 37e44; (b) Hursthouse, A.;
Kowalczyk, G. Environ. Geochem. Health 2009, 31, 165e187.
4. (a) Besse, J.-P.; Garric, J. Toxicol. Lett. 2008, 176, 104e123; (b) Isidori, M.; Bellotta,
M.; Cangiano, M.; Parella, A. Environ. Int. 2009, 35, 826e829; (c) Fick, J.; Lind-
berg, R. H.; Tysklind, M.; Larsson, D. G. J. Regul. Toxicol. Pharmacol. 2010, 58,
516e523.
5. Pichette, V.; du Souich, P. J. Am. Soc. Nephrol. 1996, 7, 345e349.
6. Castiglioni, S.; Bagnati, R.; Fanelli, R.; Pomati, F.; Calamari, D.; Zuccato, E. En-
viron. Sci. Technol. 2006, 40, 357e363.
7. Williams, D. P.; Antoine, D. J.; Butler, P. J.; Jones, R.; Randle, L.; Payne, A.; Ho-
ward, M.; Gardner, I.; Blagg, J.; Park, B. K. J. Pharmacol. Exp. Ther. 2007, 322,
1208e1220.
8. Chen, L.-J.; Burka, L. T. Chem. Res. Toxicol. 2007, 20, 1741e1744.
9. Shetti, N. P.; Sampangi, L. V.; Hegde, R. N.; Nandibewoor, S. T. Int. J. Electrochem.
Sci. 2009, 4, 104e121.
10. Hezari, M.; Davis, P. J. Drug Metab. Dispos. 1992, 20, 882e888.
11. Angadi, M. A.; Tuwar, S. M. J. Solution Chem. 2010, 39, 165e177.
12. Siedel, W.; Sturm, K.; Scheurich, W. Chem. Ber. 1966, 345e352.
13. (a) Madsen, K. G.; Olsen, J.; Skonberg, C.; Hansen, S. H.; Jurva, U. Chem. Res.
Toxicol. 2007, 20, 821e831; (b) Pitterl, F.; Chervet, J.-P.; Oberacher, H. Anal. Bi-
oanal. Chem. 2010, 397, 1203e1215.
14. Synthetic Organic Electrochemistry, 2nd ed.; Fry, A. J., Ed.; Wiley : New-York
Chichester Brisbane Toronto Singapore, 1989.
15. Clauson-Kaas, N.; Limborg, F.; Glens, K. Acta Chem. Scand. 1952, 6,
531e534.
5.2.2. 4-Chloro-2-[(2,5-dimethoxy-2,5-dihydro-furan-2-ylmethyl)-
amino]-5-sulfamoyl-benzoic acid (8). Into an undivided glass cell
equipped with two plates of carbon electrodes were dissolved fu-
rosemide (250 mg, 0.75 mmol), ammonium bromide (61 mg,
0.6 mmol), and tetraethylammonium tetrafluoroborate (81 mg,
0.4 mmol) in methanol (10 mL). Magnetic stirrer was used to get an
agitation of the reaction solution. A constant current of 30 mA passed
through reaction until 2.5 F/mol of charge was consumed. The so-
lution was transferred to a round bottom flask and methanol was
removed under reduced pressure. The residue was extracted with
ethyl acetate and theinorganic salt was eliminated by filtration. Ethyl
acetate was evaporated and a beige solid was obtained. Two di-
astereomers were present in a 50/50 ratio (268 mg, 91%). tR(HPLC)¼
4.78 min (dia 1)e5.52 min (dia 2); mp 136e140 ꢀC; 1H NMR
(400 MHz, acetone-d6):
d
¼8.55 (s, 2H), 7.07 (s, 1H), 7.05 (s, 1H),
6.22e6.15(m, 2H), 6.05(d, J¼13.4Hz,1H), 6.03(d, J¼13.5Hz,1H), 5.79
(br s,1H), 5.52 (br s,1H), 3.72e3.61 (m, 2H); 3.57e3.47 (m, 2H); 3.45
(s, 3H), 3.31 (s, 3H), 3.20 (s, 3H), 3.12 (s, 3H); 13C NMR (acetone-d6):
d
¼169.3, 169.1, 154.8, 154.5, 138.0, 137.8, 135.0, 135.0, 134.2, 134.2,
16. (a) Iwasaki, T.; Nishitani, T.; Horikawa, H.; Inoue, I. J. Org. Chem. 1982, 47,
3799e3802; (b) Shono, T.; Matsumura, Y.; Tsubata, Y.; Inoue, K.; Nishida, R.
Chem. Lett. 1983, 21e24; (c) Nieto-Mendoza, E.; Guevara-Salazar, J. A.; Ramirez-
Apan, M. T.; Frontana-Uribe, B. A.; Cogordan, J. A.; Cardenas, J. J. Org. Chem.
2005, 70, 4538e4541.
17. (a) Kolb, K. E.; Wilson, C. L. J. Chem. Soc. Chem. Commun. 1966, 271e272;
(b) Shono, T.; Matsumura, Y.; Yamane, S. Tetrahedron Lett. 1981, 34,
3269e3272.
132.4, 132.1, 127.4, 127.2, 114.9, 114.7, 114.6, 114.0, 109.9, 108.6, 108.5,
56.4, 56.0, 50.7, 50.4, 50.1, 49.9; IR: 3257,1674,1595,1503,1455,1331,
1229,1162,1043,1012, 957, 899, 832, 799, 686, 626, 579 cmꢁ1; HRMS
m/z calcd for C14H18ClN2O7S [MþH]þ: 393.0523. Found: 393.0445.
5.2.3. 1-(2-Carboxylate-5-chloro-4-sulfamoyl-phenyl)-3-hydroxy-
pyridinium (4). This compound was obtained by dissolving (8)
(136 mg, 0.3 mmol) in 2 mL of a 60/40 mixture of TFA (10 mM)/
MeCN at 40 ꢀC. The product 4 was obtained after filtration as
a white solid (126 mg, 100%). Mp 258e260 ꢀC; 1H NMR (100 MHz,
18. Perry, C. W.; Bader, G. J.; Liebman, A. A.; Barner, R.; Wuersch, J. J. Org. Chem.
1978, 43, 4391.
€
19. (a) Hunig, S.; Schenk, W. Liebigs Ann. Chem. 1979, 1523e1533; (b) Bird, C. L.;
Kuhn, A. T. Chem. Soc. Rev. 1981, 10, 49e82; (c) Deronzier, A.; Galland, B.; Viera,
M. Nouv. J. Chim. 1982, 6, 97e100.
20. Franck, D. M.; Arora, P. K.; Blumer, J. L.; Sayre, L. M. Biochem. Biophys. Res.
Commun. 1987, 147, 1095e1104.
DMSO-d6):
d¼8.78 (br s, 1H), 8.59 (br s,1H), 8.50 (s,1H), 8.09 (s,1H),