Towards calixarene-based prodrugs: Drug release and antibacterial behaviour of a water-soluble nalidixic acid/calix[4]arene ester adduct

Article abstract of DOI:10.1016/j.bmcl.2009.03.139

A water-soluble calixarene-based heterocyclic podand incorporating a quinolone antibiotic subunit, the nalidixic acid, was synthesised and fully characterised. Its prodrug behaviour was assessed in vitro by HPLC, demonstrating the release of the tethered

Full text of DOI:10.1016/j.bmcl.2009.03.139

Bioorganic & Medicinal Chemistry Letters 19 (2009) 2679–2682
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Towards calixarene-based prodrugs: Drug release and antibacterial behaviour
of a water-soluble nalidixic acid/calix[4]arene ester adduct
Hugues Massimba Dibama a, Igor Clarot a, Stéphane Fontanay a, Adel Ben Salem b, Maxime Mourer a,
Chantal Finance a, Raphaël E. Duval a, Jean-Bernard Regnouf-de-Vains a,
*
a SRSMC, UMR 7565 Nancy Université–CNRS; Groupe d’Etude des Vecteurs Supramoléculaires du Médicament, Faculté de Pharmacie, 5, rue Albert Lebrun, BP 80403,
54001 Nancy Cedex, France
b Laboratoire de Chimie Appliquée: Hétérocycles, Corps Gras et polymères. Département de Chimie, Faculté des Sciences de Sfax, 3000 Sfax, B.P. 1171, Tunisia
a r t i c l e i n f o
a b s t r a c t
Article history:
A water-soluble calixarene-based heterocyclic podand incorporating a quinolone antibiotic subunit, the
nalidixic acid, was synthesised and fully characterised. Its prodrug behaviour was assessed in vitro by
HPLC, demonstrating the release of the tethered quinolone in model biological conditions. Microbiolog-
ical studies performed on various Gram-positive and Gram-negative reference strains showed very inter-
esting antibacterial activities.
Received 3 February 2009
Revised 25 March 2009
Accepted 27 March 2009
Available online 1 April 2009
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Calixarene
Quinolone
Nalidixic acid
Prodrug
The highly interesting organisational and carrier properties of
the calixarenes have been modestly investigated in the medicinal
field; only very few reports, essentially patents, have been devoted
to their use as therapeutical agents.1–7 In order to develop calixa-
rene-based podands shaped as potent drug carriers and dispensers,
we have recently reported the synthesis and the characterisation of
drug-containing calix[4]arene species displaying two carboxy-pro-
tected 6-aminopenicillanic acids or two nalidixic acid residues,
tethered on distal phenol rings via amide or ester linkages, respec-
tively.8–10
As these compounds were designed as hydrophobic structures,
the lack of solubility in biological media made them uncompliant
for in vitro standard evaluation as antimicrobial agents. Gaining
hydrosolubility thus appeared the only way to access the biological
activities, notably to verify the prodrug behaviour of such struc-
tures, main aim of the project. Thus, we developed a synthetic
strategy leading to a water-soluble analogue of the previous com-
pounds, by means of introduction of hydrophilic groups at the
upper rim of the calixarene platform.
The synthetic strategy involves the grafting of the bromopropyl
ester of nalidixic acid at the lower rim of the calix[4]arene plat-
form. The latter was chosen in order to allow the main reaction
and purification processes in organic solvent, prior to releasing
the water-soluble form by a simple and non-destructive step.
Among the various water-soluble calixarene species available in
the literature and in our laboratory, the tetra-para-(amino-
ethyl)calix[4]arene 1 was first chosen. Its Boc-protected form al-
lows in fact functionalisation processes at the lower rim in
classical solvents, and it is easily deprotected in TFA/CH2Cl2 med-
ium, to give the corresponding water-soluble tetra ammonium
species.
The tetra-para-(aminoethyl)calix[4]arene
1 was prepared
according to Gutsche’s procedure.11 The four amino functions were
protected as Boc-derivatives, affording the tetra-para-(N-tert-
butoxycarbonyl-aminoethyl)calix[4]arene 2, with a yield of ca
70% after chromatography. The bromopropylnalidixate was re-
acted with an excess of 2 in refluxing MeCN, in the presence of
K2CO3 as base, and of a catalytic amount of KI, to give the mono-
nalidixic podand 3 with a yield of 60%. The four Boc-protective
groups of 3 were discarded by treatment with trifluoroacetic acid
in CH2Cl2, at room temperature, to give the tetra-ammonium salt
4 in a quantitative yield after lyophilisation.
We present here our preliminary synthetic, analytical and bio-
logical results, concerning a prodrug-like structure that involves
one nalidixic acid moiety tethered via a propyl linker to the lower
rim of the tetra-para-(aminoethyl)calix[4]arene.
The expected biological degradation process should give the
hydroxypropyl calixarene 6 as by-product of nalidixic acid. In order
to evaluate its own antibacterial activity, 6 was synthesised by a
two-step process involving basic degradation of 3 into the tetra-
* Corresponding author. Tel.: +33 383 682 315; fax: +33 383 682 345.
E-mail address: jean-bernard.regnouf@pharma.uhp-nancy.fr (J.-B. Regnouf-de-
Vains).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.03.139

2680
H. M. Dibama et al. / Bioorg. Med. Chem. Lett. 19 (2009) 2679–2682
Figure 1. Top: Typical chromatograms (k = 254 nm) obtained for nalidixic acid (Nal) standard (A), compound 4 in plasma medium at t = 22 h (B), compound 4 in plasma
medium at t = 0 h (C), compound 4 in mobile phase (D), alcohol 6 (E) and blank plasma (F). Bottom: UV spectrum of Nal (a), alcohol 6 (b) and compound 4 (c) extracted from
chromatograms A, E and D, respectively.
BOC alcohol 5, followed by deprotection of the amino groups in the
TFA/CH2Cl2 medium (Scheme 1).
tion (14,000 rpm, 5 min), 50 lL of supernatant were diluted five-
fold with the mobile phase before injection. As shown on Figure
1, the simultaneous release of nalidixic acid and the disappearance
of 4 were effectively observed (curve B vs C). All compounds under
study were identified with the help of the Diode Array Detector
and specific UV spectra were obtained for Nal, compounds 4 and
6 (Fig. 1, bottom). As illustrated by curve F, no interferences in
the chromatographic zone of nalidixic acid were observed, and
the major part of the void volume could be explained by residual
proteins from the rat plasma used.
All compounds were characterised by IR, UV–vis, 1H and 13C
NMR, elemental analysis and mass spectrometry. The two water-
soluble species 4 and 612 are in the cone conformation, as assessed
with 1H NMR by the presence of two AB-shaped Ar-CH2-Ar reso-
nance signals, and with 13C NMR, by their location at 30.9 ppm
for ester 4 and 30.96 and 31.12 ppm for alcohol 6. Elemental anal-
yses were consistent with the presence of the four expected trifluo-
roacetate groups for both compounds 4 and 6, four H2O associated
to ester 4, and three H2O associated to alcohol 6.
The reaction kinetic was evaluated from 0 to 90 h as illustrated
in Figure 2. The nalidixic acid quantity (release percentage) was
The expected prodrug behaviour of 4 was evaluated by HPLC
analysis of its decomposition into hydroxypropylcalixarene 6 and
nalidixic acid Nal in biological medium, that is, rat plasma. The
chromatographic conditions were optimised in order to focus on
Nal formation while allowing relatively short retention times for
all the components involved in the hydrolysis, that is, Nal, 4 and
alcohol 6. The chromatographic profile of lyophilised 4 remained
unchanged after a one-year storage period at 4 °C, revealing its
high stability in the solid form.
45.0
40.0
35.0
30.0
25.0
20.0
15.0
10.0
5.0
The chromatographic study was carried out with an RP18 sta-
tionary phase (125 Â 4 mm, 100 Å, 3
nium acetate buffer pH 2.0/MeCN/MeOH 65/30/5 (v/v/v) mobile
phase with a flow rate of 1.0 mL/min at 25 °C. A 20 L injection
lm), using a 50 mM ammo-
l
loop was used with diode array detection from 200 to 350 nm.
The hydrolyses were carried out on samples consisting in 1.0 of
4 mixed to 1.0 mL of plasma incubated at 37 °C. Aliquots of the
0.0
0
20
40
60
80
100
Time (hours)
reaction medium (50
MeCN to allow a sufficient protein precipitation. After centrifuga-
lL) were mixed to an equivalent volume of
Figure 2. Kinetic of nalidixic acid release at 37 °C in rat plasma (
Hinton broth (h).
D
) and in Mueller–

H. M. Dibama et al. / Bioorg. Med. Chem. Lett. 19 (2009) 2679–2682
2681
NH2
NHBoc
NHBoc
that probably limits its hydrolysis, explaining the plateau observed
for the formation of nalidixic acid.
NHBoc
NH2
NH2
O
H2N
BocHN
4 HCl
The Mueller–Hinthon broth employed in the in vitro antibacte-
rial assays should not display any enzymatic activities, due to ste-
rilisation process. Nevertheless, a chemical hydrolysis of the ester
junction could occur, and edge off biological results. Its hydrolytic
properties were thus evaluated according to the above-mentioned
procedure. In order to keep present all possible hydrolytic potency,
the protein precipitation step was discarded. No release of nalidixic
acid was observed at 24 h (approved standard time for MIC deter-
mination), and 48 h, confirming thus the neutrality of Mueller–
Hinthon broth in these evaluations (Fig. 2).
The antibacterial activities (minimum inhibitory concentration;
MIC) of prodrug 4, of its two expected metabolites nalidixic acid
(Nal) and 3-hydroxypropyl calix[4]arene 6, and of sodium trifluo-
roacetate were evaluated in liquid phase against various Gram-
negative (Escherichia coli ATCC 25922, Pseudomonas aeruginosa
ATCC 27853) and Gram-positive (Staphylococcus aureus ATCC
25923, and ATCC 29213, Enterococcus faecalis ATCC 29212) bacte-
ria, according to CA-SFM13 and NCCLS.14 Experiments were carried
out according to microdilution protocols performed in 96-well U
shape microtiter plates described by NCCLS.14
i
O
O
N
OH
OH
OH
Br(CH2)3O
OH
O
OH
OH
N
1
2
ii
NH2
NH2
NH2
4 CF3COOH
H2N
NHBoc
NHBoc
NHBoc
BocHN
a
iii
c
b
OH
O
O
OH
O
5
OH
4
9
6
7
OH
OH
O
O
O
O
3
O
OH
2
N
10
N
4
3
N
N
iv
4 CF3COOH
NH2
NH2
NHBoc
NHBoc
NH2
H2N
NHBoc
BocHN
O
O
iii
(K) EtO
+
OH
OH
OH
OH
O
N
N
OH
O
OH
Sodium trifluoroacetate did not exhibit any activity at concen-
OH
OH
trations inferior to 256 l
g mLÀ1).
6
5
The results presented in Table 1 show first that the activity of
nalidixic acid Nal is conform to the literature,15 close to the
Scheme 1. Synthetic pathway to podands 4 and 6. (i) NaOH, (Boc)2O, 1,4-dioxane,
H2O, 70%; (ii) MeCN, K2CO3, KI, rflx, 60%; (iii) CH2Cl2, TFA, 95%; (iv) EtOH, K2CO3,
61%. (atom numbering refers to NMR analysis12).
32 l
g mLÀ1 recommended by NCCLS.
The calixarene species 4 displays MIC values of 35
l
g mLÀ1 for
g mLÀ1
E. coli, 70
l
g mLÀ1 for the two S. aureus strains, and 140
l
for E. faecalis and P. aeruginosa, while the mono-alcohol 6 needs
from 2 (E. faecalis, P. aeruginosa) to 4 (E. coli, S. aureus) times higher
concentrations, in ranges considered as non-active.
calculated versus the maximum that could be obtained from com-
pound 4. A plateau was reached around 24 h indicating that the
reaction was terminated. The maximum percentage reached 40%;
this result was balanced as the sample treatment (protein defeca-
tion) could wash down significant quantities of active substances.
Analysis of blank assays (t0) showed that nalidixic acid was not
sequestered by the precipitate. The survey of residual concentra-
tion of 4 in solution showed, in the limits of the method, that the
latter was present in proportions lower than expected. The analysis
of the precipitate treated in acidic conditions (0.1 M HCl) showed
the release of residual 4 in an approximately complementary
amount. Thus 4 appears retained by seric proteins, but in a way
Considering in these conditions a 100% hydrolysis (best case),
the quantities of theoretically released nalidixic acid Nal and
monoalcohol 6 were calculated and compared to their MIC values.
The results given in Table 2 show that for the most susceptible
strains, that is, E. coli and S. aureus, the theoretical concentration
of released substance is 4 times less than the MIC for Nal, and ca
4 times less for mono-alcohol 6. For the less susceptible strains
(E. faecalis, P. aeruginosa), this ratio rises to 2 and 2.4, respectively.
In all cases, and even with the ideal hypothesis of 100% hydrolysis,
the MIC values obtained for the podand 4 are not consistent with a
simple release of nalidixic acid. Two hypotheses can be made at
this stage of the work, involving either a specific activity for com-
pound 4, or an additive or a multiplicative effect between 4 or 6
and nalidixic acid.
Table 1
MIC values in
l (l ) obtained by broth microdilution method,
mole.LÀ1 g mLÀ1
according to CLSI guideline, of compound 4 and its two hydrolysis by-products Nal
and alcool 6 over two Gram negative (E. coli and P. aeruginosa) and three Gram
positive (two S. aureus and E. faecalis) reference strains
These two important points are currently under investigation.
We have prepared a new water-soluble calixarene species
incorporating, via a labile bound, a quinolone antibiotic subunit.
This compound, designed as a nalidixic acid prodrug, effectively
displays this behaviour, that is, the release of quinolone in biolog-
ical medium, as assessed by analytical studies. In vitro antibacterial
evaluation showed that the title compound was efficient on one
Gram negative (E. coli) and two Gram positive (S. aureus) reference
strains, with a clear gain of activity when compared to its two
hydrolysis by-products. This observation calls for much deeper
4a
Nala
6a
E. coli ATCC 25922
25 (35)b
50 (70)
50 (70)
100 (140)
100 (140)
100 (23)
200 (46)
200 (46)
200 (46)
200 (46)
110 (128)
220 (256)
220 (256)
>220 (>256)
220 (256)
S. aureus ATCC 25923
S. aureus ATCC 29213
E. faecalis ATCC 29212
P. aeruginosa ATCC 27853
a
Molecular weights obtained from elemental analysis: 4: 1396.47; Nal: 232. 24;
6: 1164.98.
Values in brackets in l .
g/mLÀ1
b
Table 2
Comparison of MIC values (in
l
g mLÀ1) and theoretical amounts of Nal and 6 with 100% release
4
Theoretical released Nal in
l
g mLÀ1
Nal
Theoretical released 6 in
l
g mLÀ1
6
E. coli ATCC 25922
35
70
70
140
140
5.8
11.6
11.6
23.2
23.2
23
46
46
46
46
29.2
58.4
58.4
116.8
116.8
128
256
256
>256
256
S. aureus ATCC 25923
S. aureus ATCC 29213
E. faecalis ATCC 29212
P. aeruginosa ATCC 27853

2682
H. M. Dibama et al. / Bioorg. Med. Chem. Lett. 19 (2009) 2679–2682
and b, OCH2CH2CH2OCO); 40.50, 40.65 (ArCH2CH2NH3
c
a and c); 40.87
investigation, related to its proper antibacterial behaviour, as well
as to additive and/or multiplicative effects between the two re-
leased by-products or between title compound and nalidixic acid.
This investigation is already under way, in parallel to a wider anti-
bacterial screening involving multiresistant bacteria,16 bacterial
membrane targeting studies,10,17 and hydrolysis studies with bac-
terial esterases.
(ArCH2CH2NH3 b); 47.58 (CH3CH2N); 62.95 (OCH2CH2CH2OCO); 73.64
(OCH2CH2CH2OCO); 110.63 (C(3); 116.73 (CF3COO); 120.61 (C(9)); 122.60
(C(6)); 128.03, 128.46, 129.09, 134.07 (Co Ar a, c and b); 129.13, 129.24(Cm Ar
b); 129.38, 129.99 (Cm Ar a and c); 128.91, 130.62, 134.07, 134.61 (Cp Ar a, c
and b); 136.28 (C(5)); 147.45 (Ci Ar a or c); 148.18 (C(10)); 149.87 (C(2));
150.27 (Ci Ar b); 150.52 (Ci Ar a or c); 163.23 (CF3COO); 164.60 (C(7)); 166.78
(COO); 176.30 (C(4)). Anal. Calcd for C51H60N6O7, 4 CF3COOH, 4 H2O (1396.47):
C, 50.72; H, 5.19; N, 6.01. Found: C, 50.86; H, 5.20; N, 6.24. ES-MS (pos. mode):
435.47 [M–4 CF3COOH+2H+]2+/2. Compound 6: 1H NMR (400 MHz, D2O): 2.26 (t,
J = 5.6 Hz, 2H, ArOCH2CH2CH2OH); 2.66 (m, 4 H, ArCH2CH2NH a or c); 2.86 (t,
J = 6.8 Hz, 4 H, CH2CH2NH b); 3.05 (t, J = 6.8 Hz, 4H, ArCH2CH2NH a or c); 3.22
(d, J = 6.8 Hz, 4H, ArCH2CH2NH b); 3.54 (d, J = 13.8 Hz, 2H, ArCH2Ar); 3.62 (d,
J = 13.8 Hz, 2H, ArCH2Ar); 3.98 (t, 2 H, J = 5.6 Hz, ArOCH2CH2CH2OH); 4.11–4.33
(m, 6H, ArOCH2CH2CH2OH and 4H of ArCH2Ar); 6.91 (s, 2H, ArH a or c), 7.00 (s,
2 H, ArH a or c), 7.16 (br s AB, 4H, ArH b). 13C NMR (100 MHz, CDCl3): 30.96,
31.12 (Ar-CH2-Ar); 32.05, 32.24 (ArCH2CH2N and ArOCH2CH2CH2OH); 40.58,
40.73 (ArCH2CH2N a or c); 40.97 (ArCH2CH2N b); 59.27 (ArOCH2CH2CH2OH);
74.75 (ArOCH2CH2CH2OH); 116.67 (q, JC-F = 362 Hz, CF3); 129.43, 129.46 (Cm Ar
a, b and c); 129.56 (Cm Ar b); 130.06 (Cm Ar a or c); 129.53 (Cp Ar b); 130.73 (Cp
Ar a or c); 134.48 (Cp Ar a or c); 128.88, 129.34, 129.66, 134.71 (Co Ar a or c and
b); 147.68, 150.83 (Ci Ar a or c); 150.15 (Ci Ar b); 163.27 (q, JC-F = 53 Hz, COO).
Anal. Calcd for C39H50N4O5, 4 CF3COOH, 3 H2O (1164.98): C, 48.46; H, 5.19; N,
4.81. Found: C, 48.36; H, 4.84; N, 4.76. ES-MS (pos. mode): 655.40 [M–4
CF3COOH+H+]+, 328.24 [M–4 CF3COOH+2 H+]2+/2, 219.17 [M–4 CF3COOH+3
Acknowledgements
We thank the CNRS, the Ministère de la Recherche et de l’Ens-
eignement Supérieur, and the Région Lorraine for financial support.
We also thank Mr. Eric Dubs for synthesis of starting materials, Pr.
Isabelle Lartaud for the gift of rat serum, and Mrs. N. Marshall for
correcting the manuscript.
References and notes
1. Harris, Stephen J., WO Patent 95-IE8 19950124, 1995.
2. Hwang, Kou M.; Ki, You M.; Liu, Su Ying; Lee, T. C.; Choy, W.; Chen, J. WO Patent
9403165, 1985.
3. Hwang, Kou M.; Ki, You M.; Liu, Su Ying; Choy, W.; Chen, J. (Genelab
Technologies, Inc. USA) WO Patent 9403164, 1984.
4. Hulmes, D.; Coleman, A.; Aubert-Foucher, E. (CNRS, Fr) WO patent 0007585,
2000.
5. (a) Cornforth, J. W.; D’Arcy Hart, P.; Nicholls, G. A.; Rees, R. J. W.; Stock, J. A. Br. J.
Pharmacol. 1955, 10, 73; (b) D’Arcy Hart, P.; Armstrong, A. J.; Brodaty, E. Infect.
Immun. 1996, 64, 1491.
H+]3+/3
.
13. CA-SFM. Communiqué 2008 (Edition janvier 2008). http://www.sfm.asso.fr.
14. NCCLS. 2007. Methods for dilution antimicrobial susceptibility tests for
bacteria that grow aerobically; approved standard M7-A6, 6th ed. National
Committee for Clinical Laboratory Standards, Wayne, Pa, USA. Bacteria were
grown in Mueller-Hinton broth (Difco, 275730), or Mueller Hinton agar (Difco,
225250); purity of isolates was checked at the time of every test by
examination of colony morphology and Gram staining. For MIC
determination experiments, bacteria were inoculated in 96-well
U shape
6. (a) Colston, M. J.; Hailes, H. C.; Stavropoulos, E.; Herve, A. C.; Herve, G.;
Goodworth, K. J.; Hill, A. M.; Jenner, P.; Hart, P. D.; Tascon, R. E. Infect. Immun.
2007, 72, 6318; (b) Loiseau, F. A.; Hill, A. M.; Hii, K. K. Tetrahedron 2007, 63, 9947.
7. Casnati, A.; Fabbi, M.; Pelizzi, N.; Pochini, A.; Sansone, F.; Ungaro, R.; Di
Modugno, E.; Tarzia, G. Bioorg. Med. Chem. Lett. 1996, 6, 2699.
8. Ben Salem, A.; Regnouf-de-Vains, J.-B. Tetrahedron Lett. 2001, 42, 7033.
9. Ben Salem, A.; Regnouf-de-Vains, J.-B. Tetrahedron Lett. 2003, 44, 6769.
10. Korchowiec, B.; Ben Salem, A.; Corvis, Y.; Regnouf de Vains, J.-B.; Korchowiec,
J.; Rogalska, E. J. Phys. Chem. B 2007, 11146, 13231.
microtiter plates to yield a final inoculum of 1 Â 105 colony-forming units
(CFU)/mL. Then various concentrations of the drugs were added. The cultures
were grown for 18–24 h at 35 °C. The resulting bacterial growth was measured
with an ELISA plate reader (Multiskan EX, Thermo Electron Corporation,
France) at a wavelength of 540 nm.
15. see Orden, J. A.; Ruiz-Santa-Quiteria, J. A.; Garcia, S.; Cid, D.; De La Fuente, R.
Antimicrob. Agents Chemother. 1999, 43, 510.
16. (a) Following approaches described in: Grare, M.; Mourer, M.; Fontanay, S.;
Regnouf-de-Vains, J. -B.; Finance, C.; Duval, R. E. 47th Interscience Conference
on Antimicrobial Agents and Chemotherapy (ICAAC), Chicago, USA, 17–20
September 2007.; (b) Grare, M.; Massimba Dibama, H.; Lafosse, S.; Ribon, A.;
Mourer, M.; Regnouf-de-Vains, J.B.; Finance, C.; Duval, R. E. Clin. Microbiol.
Infect., in press.
11. Gutsche, C. D.; Nam, K. C. J. Am. Chem. Soc. 1988, 110, 6153.
12. Partial characterisation of compound 4: 1H NMR (400 MHz, D2O): 1.39 (t,
J = 6.5 Hz, 3H, CH3CH2N); 2.54 (m, 6H, 2 ArCH2CH2N+ArOCH2CH2CH2OCO);
2.66 (br s, 7 H, CH3 quinolone+2 ArCH2CH2N); 2.95 (t + t, 4H, ArCH2CH2NH3
a
and c); 3.09 (t, J = 7.6 Hz, 4H, ArCH2CH2NH3 b); 3.30,3.95 (‘q’, AB, JAB = 13.6 Hz,
4H, Ar-CH2-Ar); 3.41,4.13 (‘q’, AB, JAB = 13.6 Hz, 4H, Ar-CH2-Ar); 4.31 (br s, 4H,
ArOCH2CH2CH2OCO+CH3CH2N); 4.73 (br s, 2H, ArOCH2CH2CH2OCO); 6.72 (s,
2H, ArH a or c); 6.84 (s, 2H, ArH a or c); 6.88 (br d, 4H, ArH b); 7.39 (d,
J = 8.1 Hz, H(6) or H(5), quinolone); 8.33 (d, J = 8.3 Hz, H(6) or H(5), quinolone);
8.83 (s, 1 H, H(2), quinolone). 13C NMR (100 MHz, D2O): 14.63 (CH3CH2N);
24.51 (Me quinolone); 30.90 (Ar-CH2-Ar); 31.96, 32.15, 32.23 (ArCH2CH2NH3 a,
17. Following approaches described in: (a) Grare, M.; Dague, E.; Mourer, M.;
Regnouf-de-Vains, J.-B.; Finance, C.; Duval, J. F. L.; Duval, R. E.; Gaboriaud, F.
Pathol. Biol. 2007, 55, 465; b Grare, M.; Lafosse, S.; Regnouf-de-Vains, J. -B.;
Finance, C.; Duval, R. E. 48th annual Interscience Conference on Antimicrobial
Agents and Chemotherapy (ICAAC)/ Infectious Diseases Society of America
(IDSA), Washington, DC, USA, 25–28 October 2008. (ASM Fellows Travel Grant
to Grare M.).