Electrochemical and spectral characterization of the host-guest inclusion complex of the antiparasitic drug nitazoxanide with β-cyclodextrin

Article abstract of DOI:10.1007/s00706-013-1109-1

The formation of a host-guest inclusion complex of the antiparasitic drug nitazoxanide (NTZ) with β-cyclodextrin (β-CD) in aqueous solution has been characterized by electrochemical methods, cyclic and differential pulse voltammetry, and UV-Vis absorption spectroscopy. The nature of the host-guest inclusion complex between NTZ and β-CD has been elucidated. The experimental results confirmed the existence of 1:1 inclusion complex of NTZ with β-CD. The binding constants describing the extent of formation of the complexes have been determined, using Benesi-Hildebrand plots, using voltammetry and UV-Vis spectroscopy. NTZ exhibits an affinity for β-CD. A special configuration of the complex has been proposed. Graphical abstract: [Figure not available: see fulltext.].

Full text of DOI:10.1007/s00706-013-1109-1

Monatsh Chem (2014) 145:421–426
DOI 10.1007/s00706-013-1109-1
ORIGINAL PAPER
Electrochemical and spectral characterization of the host-guest
inclusion complex of the antiparasitic drug nitazoxanide
with b-cyclodextrin
•
Abd-Elgawad Radi Hossam M. Nassef
•
Abd-Elrahman El-Naggar
Received: 24 July 2013 / Accepted: 21 October 2013 / Published online: 15 November 2013
Ó Springer-Verlag Wien 2013
Abstract The formation of a host-guest inclusion com-
plex of the antiparasitic drug nitazoxanide (NTZ) with
b-cyclodextrin (b-CD) in aqueous solution has been char-
acterized by electrochemical methods, cyclic and
differential pulse voltammetry, and UV-Vis absorption
spectroscopy. The nature of the host-guest inclusion com-
plex between NTZ and b-CD has been elucidated. The
experimental results confirmed the existence of 1:1 inclu-
sion complex of NTZ with b-CD. The binding constants
describing the extent of formation of the complexes have
been determined, using Benesi-Hildebrand plots, using
voltammetry and UV-Vis spectroscopy. NTZ exhibits an
affinity for b-CD. A special configuration of the complex
has been proposed.
approved for treatment of diarrhea caused by G. intestinalis
in adults. It is the first new US FDA-approved drug for
treatment of Cryptosporidium and Giardia infections [4, 5].
NTZ is also active against parasitic and microbial patho-
gens, including Trypanosoma cruzi, the causative agent of
trypanosomiasis and leishmania mexicana [6], and the
ulcer-causing pathogen Helicobacter pylori [7]. NTZ is
also a potent inhibitor of hepatitis B and C virus replication
[8], being proposed for the treatment of chronic viral
hepatitis.
The anti-protozoal activity of NTZ is believed to be due
to interference with the pyruvate ferredoxin oxidoreductase
(PFOR) enzyme-dependent electron transfer reaction,
which is essential to anaerobic energy metabolism [9, 10].
NTZ is formulated in the solid state as the active phar-
maceutical ingredient of both suspensions and tablets. The
formulations that improve the solubility of this drug may,
therefore, provide improved therapeutic options for
patients. Interest has been devoted to improving the
physicochemical properties of several drugs by interaction
with cyclodextrins (CDs). CDs have widely proved their
usefulness as tools to generate aqueous drug solutions
without the use of organic cosolvents, surfactants, or lipids
as formulation adjuncts, which increase the dissolution
rates and oral bioavailability of solid drugs [11]. There are
no toxic effects associated with oral administration of CDs
because they are not absorbed across the gastrointestinal
tract [12]. The in vitro and in vivo efficacies of b-cyclo-
dextrin (b-CD) itself against C. parvum infection were
demonstrated [12, 13].
Keywords Nitazoxanide ꢀ b-Cyclodextrin ꢀ
Binding constant ꢀ UV-Vis spectroscopy ꢀ Voltammetry
Introduction
Nitazoxanide [2-acetyloxy-N-(5-nitro-2-thiazolyl)benzam-
ide, NTZ] is a nitrothiazole derivative notable for its
activity in treating both intestinal protozoal and helminthic
infections [1, 2]. It was patented in 1976 [3], and since
1996 NTZ has been marketed worldwide. The US Food
and Drug Administration (FDA) approved the oral sus-
pension of NTZ in December 2002 for the treatment of
diarrhea caused by Cryptosporidium species and Giardia
intestinalis in pediatric patients, and in July 2004, NTZ was
Inclusion phenomena do influence the electrochemistry
of an electroactive guest molecule. CD affects the elec-
trochemical behavior of the guest compound, and useful
information from the electrochemical measurements could
be obtained. It is a suitable method to study the inclusion
A.-E. Radi (&) ꢀ H. M. Nassef ꢀ A.-E. El-Naggar
Department of Chemistry, Faculty of Science,
Dumyat University, Dumyat 34517, Egypt
e-mail: abdradi@yahoo.com
123

422
A.-E. Radi et al.
complexation if electron distribution of a complexed
electroactive guest molecule in aqueous solution is differ-
ent from that in non-complexed state and if this change is
electrochemically detectable. The effect of b-CD on the
peak current and potential of voltammetric reduction/oxi-
dation of several drugs has been investigated [14–17].
Voltammetry is a useful technique not only for studying the
inclusion complexation phenomenon, but also for investi-
gating the mode of interaction of the host and guest
molecules.
great similarity with that observed for aryl amides [26], N-
phenylamide-containing compounds [27, 28], and 4- and
40-substituted benzanilides [29]. We postulate that the
anodic oxidation of NTZ is initiated by one-electron
transfer to form a cation radical at the nitrogen atom of the
amide function of the benzamide moiety. The cation radi-
cal generated at the electrode surface undergoes different
paths (e.g., rapid decomposition-rearrangement, fragmen-
tation, or addition of water) to form stable products.
Increasing the scan rate, the potential of the anodic peak
was slightly shifted to more positive values. Since for a
diffusion-controlled irreversible system Epa-Ep/2 = 47.7/
aana, where aa is the anodic charge transfer coefficient and
na the number of electrons in the rate determining step, it
can be calculated that aana = 1.32. Increasing the scan
rate, the peak current increased linearly with the square
root of the scan rate (m1/2), consistent with the diffusion-
limited oxidation of solution species.
Different spectroscopic techniques, such as UV-Vis
absorption, were employed for a thorough investigation of
the structural characteristics of such supramolecular
aggregates, which exhibited distinct spectroscopic features
and properties from both ‘‘guest’’ and ‘‘host’’ molecules
[18–21]. The stoichiometry and equilibrium constants for
complex formation, which permitted
a quantitative
description of the solubility behavior, were calculated [22].
Herein, the study of the formation of the host-guest
inclusion complex between NTZ and b-CD in aqueous
solution using two different techniques, spectral (UV-Vis
spectroscopy) and electrochemical (cyclic and differential
pulse voltammetry), was undertaken.
The addition of b-CD to the electrolyte solution causes a
decrease in the peak current, since the peak current is pro-
portional to the square root of the diffusion coefficient. CD
is a large macrocyclic compound; incorporation of a small
guest molecule in the bulky CD affords a lower apparent
diffusivity of the guest. The diffusion of the inclusion
complex to the electrode surface was very slow compared to
that of the NTZ, showing that the decrease in peak current is
due to the formation of a slowly diffusing NTZ-b-CD
complex. The diffusion coefficient of the drug with and
without b-CD could be determined using the following
relation for a diffusion-controlled irreversible system [30]:
Results and discussion
Electrochemical study
The voltammograms of NTZ yield pH-dependent irre-
versible cathodic peak/peaks corresponding to the
reduction of its NO2 group within the cathodic potential
windows at the glassy carbon electrode [23] or at the
mercury electrode [24]. The voltammetric response of O2
represents a common problem in cathodic voltammetry.
Then, deoxygenation with pure nitrogen gas before mea-
surements is needed. The voltammetry within the anodic
potential window, as presented here, results in a back-
ground current free from the voltammetric feature of O2,
with no need for nitrogen purging. The oxidation of NTZ
was initially studied by cyclic voltammetry (CV) in 0.2 M
phosphate buffer (pH 4.0 and 7.4) on a screen-printed
carbon electrode (Fig. 1). On the first scan, an oxidation
peak was observed. After changing the scan direction, no
reduction peak occurred, showing that the oxidation pro-
cess of NTZ is an irreversible process, in agreement with
the electrochemical oxidation behavior of NTZ at the
carbon paste electrode [25]. A study of the molecular
structure of NTZ together with the previous knowledge of
the voltammetric behaviour of the compounds with struc-
turally similar structures led us to attribute the
voltammetric peak of NTZ to the oxidation of the benz-
amide moiety. The voltammetric peak of NTZ showed a
1=2
ip ¼ 2:99 ꢁ 105ðanaÞ AD1=2Cm1=2
where ip/A is the peak current, A/cm2 is the surface area of
the electrode, C/mol cm-3 is the bulk concentration of the
electroactive species, D/cm2 s-1 is the diffusion coefficient,
m/V s-1 is the scan rate, na is the number of electrons in the
rate-determining step, and a is the transfer coefficient.
D values of NTZ were 5.1 9 10-7 and 2.5 9 10-8 cm2/s
for free drug and the NTZ–b-CD complex, respectively,
indicating the inclusion of NTZ in the cavity of b-CD,
which consequently reduced the diffusion of the bound
drug. The solution of the complex of NTZ with b-CD yields
the same electrochemical characteristics as NTZ itself, but
the peak potentials are displaced toward more positive
potentials. The voltammetric peaks are lower than those of
NTZ owing to the smaller diffusion coefficient of the bulky
inclusion complex. Such electrochemical behavior suggests
that NTZ is oxidized in its free form after dissociation of the
inclusion complex at the electrode surface [31].
The quantitative treatment of the dependence of the
relative peak current on the concentration of CD will
provide the value of the binding constant. The binding
constant K and stoichiometric ratios of the inclusion
123

Electrochemical and spectral characterization of the host-guest inclusion complex
423
Fig. 1 Cyclic voltammograms
of 5.0 9 10-5 M nitazoxanide
in the absence and presence of
different concentrations of b-
3.0
2.5
2.0
1.5
1.0
0.5
0.0
A
B
4
3
2
1
0
CD (1.0 9 10-5, 2.5 9 10-5
5.0 9 10-5, 1.0 9 10-4 M),
from top to bottom, at the
,
screen-printed carbon electrode
in phosphate buffer solution
(0.2 M); a pH 4.0, b pH 7.4,
scan rate 100 mV s-1. The
dotted lines represent the
background voltammograms
0.2 0.4 0.6 0.8 1.0 1.2
0.0
0.3
0.6
0.9
1.2
E/V vs. Ag-AgCl
E/V vs. Ag-AgCl
Fig. 2 Differential pulse
voltammograms of
1.0 9 10-5 M nitazoxanide in
0.2 M phosphate buffer; a pH
4.0, b pH 7.4 in the absence and
presence of different
A
B
concentrations of b-CD
(1.0 9 10-6, 2.5 9 10-6
,
1.0 9 10-5, 5.0 9 10-5 M),
from top to bottom, at the
40nA
20nA
screen-printed carbon electrode.
Pulse amplitude 50 mV, pulse
width 30 s, sample width 0.02 s.
The dotted lines represent the
background voltammograms
0.5
0.6
0.7
0.8
0.9
1.0
0.3
0.4
0.5
0.6
0.7
0.8
E/V vs. Ag-AgCl
E/V vs. Ag-AgCl
[b-CD]0 are the initial concentration of NTZ and b-CD,
respectively. A plot of [1/(iHG-iG)] versus 1/[b-CD] gives
a straight line with a good linear correlation (r = 0.98),
confirming the formation of a 1:1 inclusion complex. From
the intercept and slope values of this plot, the binding
complex of NTZ can be determined according to the
modified Benesi-Hildebrand relation (double reciprocal
plot) assuming the formation of a 1:1 host-guest complex:
1
1
1
¼
þ
iHG ꢂ iG Di K½NTZꢃ0Di½bꢂCDꢃ0
constant
K
was evaluated to be 0.32 9 104 and
0.49 9 104 M-1 for pH 4.0 and 7.4, respectively. The
interaction of NTZ with b-CD was also evaluated by the
differential pulse voltammetry. The peak current
decreased, and the peak potential shifted to the positive
side with an increase in the amount of b-CD as shown in
Fig. 2. This observation is in agreement with those for a
where iG is the oxidation peak current of the NTZ guest
molecule, and IHG is the oxidation peak current of inclusion
complex of NTZ–b-CD. iHG-iG means the peak current
difference between the inclusion complex and guest mol-
ecule. Di is the difference between the molar peak current
coefficient of the inclusion complex and NTZ; [NTZ]0 and
123

424
A.-E. Radi et al.
NTZ with b-CD previously described by CV. The binding
constant of NTZ with b-CD was calculated by treatment
similar to that described above for CV to afford 0.39 9 104
and 0.41 9 104 M-1 at pH 4.0 and 7.4, respectively.
The plot of solubilized drug against the CD concentration
showed good linearity, suggesting the formation of com-
plex with 1:1 composition, which is most likely formed
because of the inclusion of a thiazole ring in the hydro-
phobic cavities of b-CD molecules. The binding constant
for the formation of the NTZ–b-CD complex has been
determined by analyzing the changes in the intensity of
absorbance with the b-CD concentration. In the case of the
1:1 inclusion complex formed between NTZ and b-CD, the
equilibrium can be written as:
UV-Vis spectroscopy study
Among various methods proposed to characterize the
inclusion phenomena of CD systems, the spectrophoto-
metric techniques, using UV-Vis absorption [14, 17, 20],
are popular. Modification of UV spectra in the presence of
CDs provides evidence of the formation of an inclusion
complex. The inclusion CD complexes of various drugs
such as piroxicam [32], purine nucleosides [33], and the
anthracycline antibiotics doxorubicin [34] and gliclazide
[35] are studied by this technique.
b-CD þ NTZ ꢀ b-CD : NTZ
The binding constant (K) can be obtained by using the
modified Benesi-Hildebrand equation [37]:
1
1
1
¼
þ
A ꢂ A0 De K½NTZꢃ0De½b-CDꢃ0
Figure 3 shows the spectra for 1.0 9 10-4 M NTZ in
100 mM phosphate buffer solution (pH = 7.4, 6.0, and
4.0). The amino group existed as an anion above pH 6.0,
and NTZ is neutral (HNTZ) below pH 5.0. At pH 7.4, a
major absorbance peak at 416 nm is observed. At pH 6.0
(pKA), both the 416 and 348 nm absorbing species are
equally represented, suggesting that NTZ most likely exists
as an anion under physiological conditions [36]. The effect
of b-CD on the UV absorption spectra of the NTZ was
quantitatively investigated by holding the concentration of
the guest constant (i.e., 5.0 9 10-5 M NTZ) and varying
the host concentration between 0.0 and 1.0 9 10-4 M in
0.2 M phosphate buffer at pH 4.0 and 7.4. The absorption
intensities increased as a function of the b-CD concentra-
tion (Fig. 4). This behavior has been attributed to the
enhanced dissolution of the NTZ molecule through the
hydrophobic interaction between NTZ and b-CD. How-
where A-A0 is the difference between the absorbance of
NTZ in the presence and absence of b-CD, De is the dif-
ference between the molar absorption coefficient of NTZ
and the inclusion complex, and [NTZ]0 and [b-CD]0 are the
initial concentration of NTZ and b-CD, respectively. The
plot of 1/(A-A0) versus 1/[b-CD] is linear, confirming
the formation of a 1:1 inclusion complex. From the inter-
cept and slope values of this plot, K was calculated to be
0.29 9 104 and 0.51 9 104 M-1 at pH 4.0 and 7.4,
respectively, which is in a good agreement with that
determined by electrochemical methods. NTZ can exist as
one of three species (an anionic, zwitterionic, or cationic
species) because of its amphoteric property. Under physi-
ological conditions (pH = 7.4), the anionic form is the
predominant structure. Under acidic conditions the cationic
form is present. The formation constants are sensitive to
change of pH values, which reveals the selective inclusion
associated with the species form of NTZ. The inclusion
interaction of CDs with the anionic species of NTZ is
superior to that for its cationic species. This can be
attributed to a further stabilization caused by the formation
of extra hydrogen bonds between the NTZ anionic guest
and the secondary OH groups [38].
ever, a well-defined plateau was observed at 5.0 9 10-5
M
b-CD; at this point, the molecule may be fully complexed.
pH 7.4
0.6
0.5
Molecular modeling has become an interesting tool to
determine the host-guest interactions in CD technology.
Molecular mechanics calculation and modeling were per-
formed for the inclusion process using the Hyperchem
package. The minimum structure energy was calculated by
the MM ? force field. According to the molecular size, it
pH 4.0
0.4
pH 6.0
0.3
0.2
0.1
0.0
˚
is apparent that NTZ (ca. 6.5 A wide and 9.1 A long) is too
˚
large to fit entirely in the b-CD cavity (height of torus,
˚
˚
7.9 A; cavity diameter, 6.0–6.5 A). It is reasonable to
consider the partial inclusion of the NTZ in b-CD. Figure 5
shows the proposed structure of the inclusion complex of
NTZ with b-CD. This structure corresponds to the position
210
280
350
420
490
560
Fig. 3 UV-Vis absorption spectra of 1.0 9 10-4 M nitazoxanide in
0.2 M phosphate buffer solutions at pH 4.0, 6.0, and 7.4
˚
of the guest in which the thiazole ring (2.7 A) of the NTZ
123

Electrochemical and spectral characterization of the host-guest inclusion complex
425
Fig. 4 UV-Vis absorption
spectra of 5.0 9 10-5
nitazoxanide in the absence and
presence of different
0.4
B
A
M
0.75
0.50
0.25
0.00
concentrations of b-CD
(1.0 9 10-5, 1.5 9 10-5
,
0.3
0.2
0.1
3.5 9 10-5, 5.0 9 10-5 M),
from bottom upwards, in
phosphate buffer solution
(0.2 M); a pH 4.0, b pH 7.4
250
300
350
400
300
350
400
450
500
of b-CD is supplemented with the formation of hydrogen
bonds. The information obtained here may be used to
mimic the interactions of the NTZ molecule with proteins.
O
O
N
N
S
N
O
H
O
Experimental
O
CH3
Chemicals
Fig. 5 The proposed structure of the inclusion complex of nitazox-
anide with b-CD
Nitazoxanide, kindly provided by Sigma Pharmaceutical
Industries (Egypt), was used without further purification.
All experiments were performed with analytical reagent-
grade chemicals. b-CD, orthophosphoric acid (85 %),
potassium dihydrogen phosphate, disodium hydrogen
phosphate, and sodium phosphate were purchased from
Sigma Chemical Co. (St. Louis, USA). These chemicals
were used without further purification. Stock solutions of
NTZ (1 9 10-2 M) were prepared in methanol and stored
in a refrigerator at 4 °C. b-CD was purchased from Sigma-
Aldrich (St. Louis, MO) with the highest available purity.
Drying under a vacuum at 105 °C for 3 h has been shown
to be an effective method for removal of water from CD
[39]. Thus, this method was used for the determination of
moisture content of the b-CD samples. b-CD consists of
14 % water mass content, which was considered in the
calculations of solute concentrations. b-CD stock solution
was prepared by dissolution in double-distilled water.
Sample dilutions were carried out by taking the appropriate
aliquots from the stock solutions followed by dilution with
phosphate buffer. A series of 0.2 M phosphate buffer
solutions (H3PO4, KH2PO4, Na2HPO4, and Na3PO4) was
mixed in different amounts and diluted with distilled water
to obtain the required pH.
˚
is completely inside the cavity. The nitro group (3.6 A) is
located inside the b-CD cavity towards the narrow CD end
in the inclusion complex. Molecular modeling study also
˚
showed that the benzene ring (2.7 A) of NTZ appears to be
completely outside the CD cavity and the amide group
˚
(3.8 A) was not in cavity but close to the wider rim of
b-CD.
Conclusion
The present work demonstrated that the aqueous solubility
of NTZ was enhanced considerably by formation of an
inclusion complex with b-CD. Thus, the b-CD may be
useful in improving the dissolution and the bioavailability
of NTZ in pharmaceutical formulation. The electrochemi-
cal and spectral data showed good agreement, suggesting
the formation of a stable 1:1 stoichiometric complex of
b-CD–NTZ in solution. The data could suggest that the
thiazole moiety of the NTZ molecule is inserted into a
more hydrophobic environment inside the b-CD cavity.
The stabilization of insertion inside the toroidal molecules
123

426
A.-E. Radi et al.
10. Hemphill A, Mu¨ller N, Mu¨ller J (2007) Anti-Infect Agents Med
Chem 6:273
Voltammetric measurements
11. Rasheed A, Kumar CKA, Sravanthi VVNSS (2008) Sci Pharm
76:567
12. Irie T, Uekama K (1997) J Pharm Sci 86:147
Voltammetric measurements were carried out using a
CHI610C Electrochemical Analyzer controlled by CHI
version 9.09 software (CH Instruments, USA). A three-
electrode configuration was composed of a working SPCE
(4.0 mm diameter), printed from a carbon-based ink, a
silver–silver chloride pseudo-reference electrode made
from a silver-based ink, and the auxiliary electrode from a
carbon ink. All pH metric measurements were made on a
on a CG 808 Digital pH meter with an H61 pH glass
´
´
13. Castro-Hermida JA, Freire-Santos F, Oteiza-Lopez AM, Ares-
´
Mazas E (2000) J Parasitol 86:1118
14. Dang XJ, Nie MY, Tong J, Li HL (1998) J Electroanal Chem
448:61
15. Ferreira FDR, Da Silva EG, De Leo LPM, Calvo EJ, Bento EDS,
Goulart MOF, De Abreu FC (2010) Electrochim Acta 56:797
16. Radi AEM, Eissa SH (2011) Coll Czech Chem Commun 76:13
´
´
17. Yan˜ez C, Salazar R, Nun˜ez-Vergara LJ, Squella JA (2004) J
Pharm Biomed Anal 35:51
´ ´
18. Valero M, Costa SMB, Ascenso JR, Velazquez MM, Rodrıguez
¨
combination electrode (Schott Gerate, Mainz, Germany),
LJ (1999) J Inclusion Phenom 35:663
19. El-Kemary M, Organero JA, Santos L, Douhal A (2006) J Phys
Chem B 110:14128
20. Muthu Prabhu AA, Venkatesh G, Rajendiran N (2010) J Solution
Chem 39:1061
which was previously standardized with buffers of known
pHs. All measurements were performed at room
temperature.
21. Sankaranarayanan RK, Siva S, Venkatesh G, Muthu Prabhu AA,
Rajendiran N (2011) J Mol Liq 161:107
UV-Vis absorption measurements
22. Maupas B, Letellier S, Guyon F (1996) J Inclusion Phenom Mol
Recognit Chem 23:259
23. Jain R, Yadav RK, Rather JA (2013) J Pharm Anal. doi:10.1016/j.
jpha.2013.07.005
24. El-Desoky HS, Ghoneim MM, Abdel-Galeil MM (2010) J Braz
Chem Soc 21:669
25. Radi AE, El-Ghany NA, Wahdan T (2012) Monatsh Chem
143:697
26. Bard AJ (1984) Encyclopedia of electrochemistry of elements.
Marcel Dekker, New York
27. Garrido E, Lima J, Delerue-Matos C, Borges F, Silva A, Piedade
J, Oliveira Brett A (2003) J Agric Food Chem 51:876
28. Garrido E, Lima J, Delerue-Matos C, Borges F, Silva A, Brett A
(2001) Anal Chim Acta 434:35
29. Ikenoya S, Masui M, Ohmori H, Sayo H (1974) J Chem Soc
Perkin Trans 2:571
30. Bard AJ, Faulkner LR (1980) Electrochemical methods: funda-
mentals and applications. ??Wiley & Sons, New York
31. Strelets V, Mamedjarova I, Nefedova M, Pysnograeva N, Soko-
lov V, Pospisil L, Hanzlik J (1991) J Electroanal Chem 310:179
UV-Vis absorption spectra were recorded with a Perkin
Elmer UV-VIS double-beam spectrophotometer equipped
with a PC for data processing (UV WinLab ver. 2.80.03,
Perkin Elmer, USA). The solutions were scanned from 200
to 500 nm at a scan speed of 240 nm/min in a quartz cell
with a 1.0-cm path length. The UV-Vis spectra were
achieved for NTZ in the absence and presence of increas-
ing amounts of b-CD in 0.2 M phosphate buffer solution.
The baseline was established for each measurement by
placing an aqueous solution of b-CD in the reference
compartment at the same concentration of the sample. All
the data shown represent the average of at least three
determinations.
References
¨
32. Basan H, Goer NG, Ertas¸ N, Orbey MT (2001) J Pharm Biomed
Anal 26:171
1. Anderson VR, Curran MP (2007) Drugs 67:1947
2. Davila-Gutierrez CE, Vasquez C, Trujillo-Hernandez B, Huerta
M (2002) Am J Trop Med Hyg 66:251
3. Rossignol JF, Cavier R (1976) 2-Benzamido 5-nitrothiazoles. US
Patent 3,950,351, 13 April 1976 (1975) Chem Abstr 83:28216n
4. Cohen SA (2005) Curr Med Res Opin 21:999
33. Xiang TX, Anderson BD (1990) Int J Pharm 59:45
34. Bekers O, Beijnen JH, Otagiri M, Bult A, Underberg WJM
(1990) J Pharm Biomed Anal 8:671
35. Arias-Blanco MJ, Moyano JR, Perez-Martinez JI, Gines JM
(1998) J Pharm Biomed Anal 18:275
36. Hoffman PS, Sisson G, Croxen MA, Welch K, Harman WD,
Cremades N, Morash MG (2007) Antimicrob Agents Chemother
51:868
37. Benesi HA, Hildebrand JH (1949) J Am Chem Soc 71:2703
38. Snor W, Liedl E, Weiss-Greiler P, Viernstein H, Wolschann P
(2009) Int J Pharm 381:146
5. Fox LM, Saravolatz LD (2005) Clin Infect Dis 40:1173
´
´
´
6. Chan-Bacab MJ, Hernandez-Nun˜ez E, Navarrete-Vazquez G
(2009) J Antimicrob Chemother 63:1292
7. Guttner Y, Windsor HM, Viiala CH, Dusci L, Marshall BJ (2003)
Antimicrob Agents Chemother 47:3780
8. Keeffe EB, Rossignol JF (2009) World J Gastroenterol 15:1805
9. Gilles HM, Hoffman PS (2002) Trends Parasitol 18:95
39. Nakai Y, Yamamoto K, Terada K, Kajiyama A, Sasaki I (1986)
Chem Pharm Bull 34:2178
123