New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activities

Article abstract of DOI:10.1016/j.cbi.2014.04.003

This paper describes the synthesis of new enantiomerically pure 2-cyanoethyl-oxazolines in one step starting from a wide range of amino alcohols and 4-ethoxy-4-iminobutanenitrile with high to good yields (73-96%) via an appropriate procedure which can be used for a selective synthesis of mono-oxazolines. A simple operation as well as a practical separation is additional eco-friendly attributes of this method. All the synthesized compounds were identified and characterized with their physicochemical features and their spectral data (¹H NMR, ¹³C NMR and TOFMS ES⁺). Among the prepared mono-oxazolines, the mono-oxazoline (3a) [3-[(4S)-4-benzyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile] was tested to detect some biological activities. This compound was studied in vitro given the various types of pharmacological properties characterizing these compounds such as antioxidant, antimicrobial and analgesic activities. The antioxidant activity and mechanism of (3a) were identified using various in vitro antioxidant assays including 1,1-diphenyl-2-picryl-hydrazyl (DPPH), and superoxide anion radicals (O₂^-) scavenging activity. In addition, compared to Quercetin, the tested synthetic product reveals a relatively-strong antiradical activity towards the DPPH (activity percentage of 81.22%) free radicals and significantly decreased the reactive oxygen species such as (O₂ ^-) formation evaluated by the non-enzymatic (nitroblue tetrazolium/riboflavine) and the enzymatic (xanthine/xanthine oxidase) systems. Related activity values were, respectively, 66% and 60.30%. The oxazoline (3a) showed a high ability to reduce the O₂^- generation and proved to be a very potent radical scavenger. On the other hand, the analgesic property of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile (3a) was demonstrated. The subcutaneous administration of (3a) produced a significant reduction in the number of abdominal constrictions amounting to 73.81% in the acetic acid writhing test in mice. In addition to these advances, the oxazoline (3a) has been investigated as an antimicrobial agent. Our results showed that this molecule exhibited various levels of antibacterial effect against all the tested bacterial strains.

Full text of DOI:10.1016/j.cbi.2014.04.003

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Contents lists available at ScienceDirect
Chemico-Biological Interactions
journal homepage: www.elsevier.com/locate/chembioint
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New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis
and assessment of some biological activities
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Rym Hassani a, Yakdhan Kacem a, Hedi Ben Mansour b, Hamed Ben Ammar a, Béchir Ben Hassine a,
⇑
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a Laboratoire de Synthèse Organique Asymétrique et Catalyse Homogène (UR11ES56), Faculté des Sciences, Avenue de l’environnement, 5019 Monastir, Tunisia
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b Laboratoire de biotechnologie et Valorisation de Bio Géo Ressources (LBVBGR), Institut Supérieur de Biotechnologie, ISBST BioTechPole Sidi Thabet Université Manouba,
Ariana 2020, Tunisia
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a r t i c l e i n f o
a b s t r a c t
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Article history:
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This paper describes the synthesis of new enantiomerically pure 2-cyanoethyl-oxazolines in one step
starting from a wide range of amino alcohols and 4-ethoxy-4-iminobutanenitrile with high to good yields
(73–96%) via an appropriate procedure which can be used for a selective synthesis of mono-oxazolines. A
simple operation as well as a practical separation is additional eco-friendly attributes of this method. All
the synthesized compounds were identified and characterized with their physicochemical features and
their spectral data (1H NMR, 13C NMR and TOFMS ES+). Among the prepared mono-oxazolines, the
mono-oxazoline (3a) [3-[(4S)-4-benzyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile] was tested to detect
some biological activities. This compound was studied in vitro given the various types of pharmacological
properties characterizing these compounds such as antioxidant, antimicrobial and analgesic activities.
The antioxidant activity and mechanism of (3a) were identified using various in vitro antioxidant assays
including 1,1-diphenyl-2-picryl-hydrazyl (DPPHÅ), and superoxide anion radicals (OÅ2ꢀ) scavenging activ-
ity. In addition, compared to Quercetin, the tested synthetic product reveals a relatively-strong antirad-
ical activity towards the DPPH (activity percentage of 81.22%) free radicals and significantly decreased
the reactive oxygen species such as (OÅ2ꢀ) formation evaluated by the non-enzymatic (nitroblue tetrazo-
lium/riboflavine) and the enzymatic (xanthine/xanthine oxidase) systems. Related activity values were,
respectively, 66% and 60.30%. The oxazoline (3a) showed a high ability to reduce the OÅ2ꢀ generation
and proved to be a very potent radical scavenger. On the other hand, the analgesic property of the
3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile (3a) was demonstrated. The subcutaneous
administration of (3a) produced a significant reduction in the number of abdominal constrictions
amounting to 73.81% in the acetic acid writhing test in mice. In addition to these advances, the oxazoline
(3a) has been investigated as an antimicrobial agent. Our results showed that this molecule exhibited
various levels of antibacterial effect against all the tested bacterial strains.
Received 15 January 2014
Received in revised form 12 March 2014
Accepted 2 April 2014
Available online xxxx
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Keywords:
Oxazoline
Amino alcohol
Monohydrochloride
Iminoether
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Biological activities
Ó 2014 Published by Elsevier Ireland Ltd.
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1. Introduction
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Literature reveals that nitrogen containing heterocyclic mole-
cules constitutes the largest portion of chemical entities which
are part of many natural products, fine chemicals, biologically-
active pharmaceuticals and agrochemicals playing a vital role in
enhancing the quality of life.
Among a large variety of nitrogen-containing heterocyclic
compounds, the 2-oxazolines have impregnated numerous sub-
disciplines in the field of synthetic organic chemistry over
100 years since their discovery [1]. This versatile heterocycles
has served as a protecting group, a coordinating ligand, and an
activating moiety, often exhibiting all of these characteristics in a
single transformation. The well-defined reactivity of chiral
Abbreviations: ASL, lysine acetylsalicylate; COX, cyclooxygenases; DMSO,
dimethyl sulfoxide; DNA, deoxyribonucleic acid; DPPH, 1,1-diphenyl-2-pic-
rylhydrazyl radical; EDTA, ethylenediaminetetraacetic acid; EtOH, ethanol; HIV-1,
Human immunodeficiency virus; 1H NMR, proton nuclear magnetic resonance; IC50
,
concentration inhibiting a biological response by 50%; Me3SiCl, trimethylsilylchlo-
ride; MICs, Minimum Inhibitory Concentrations; MBCs, Minimum Bactericidal
Concentrations; NBT, nitroblue tetrazolium; NSADs, n-steroidal anti-inflammatory
drugs; PGE, prostaglandin E; TLC, thin layer chromatography; TOFMS ES+, time-of-
flight mass spectrometry; Zn(OAc)2, zinc acetate.
⇑
Corresponding author. Tel.: +216 73500279; fax: +216 73500278.
E-mail address: bechirbenhassine@yahoo.fr (B. Ben Hassine).
http://dx.doi.org/10.1016/j.cbi.2014.04.003
0009-2797/Ó 2014 Published by Elsevier Ireland Ltd.
Please cite this article in press as: R. Hassani et al., New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activ-
ities, Chemico-Biological Interactions (2014), http://dx.doi.org/10.1016/j.cbi.2014.04.003

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oxazolines has given rise to numerous highly-efficient strategies
for an asymmetric synthesis including their use as ligands in an
asymmetric catalysis.
have different biological activities. Based on the literature, we have
assessed their pharmacological properties such as antioxidant,
antimicrobial and analgesic activities.
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Various properties have imposed their use in a multitude of dis-
similar applications: as monomers in polymer production, as mod-
erators in analytical processes, and as conformationally rigid
peptide mimics in medicinal chemistry. Even natural systems have
chosen to incorporate 2-oxazolines into their chemical arsenal, as
evidenced by the rapidly growing number of identified natural
products and their attendant pharmacological properties (Fig. 1).
This could be explained also by the various applications of these
products in the synthesis of different therapeutic and biologi-
cally-active compounds [2] such as antidiabetic, antihypertensive,
antidepressive, antihypercholesterolemic, anticancer, anti HIV-1,
antitumor and antialzheimer agents [3]. Thanks to their structural
relationship with procaine, 2-oxazolines derivatives are expected
to have local anaesthetic properties [4]. In addition to these serious
matters, some 2-oxazoline derivatives act as enzyme inhibitors
[5,6]. Furthermore, the 2-oxazoline moieties are often found in var-
ious bioactive natural compounds and designed medicinal agents,
such as the Brasilibactin A [7]. These compounds exist as the key
fragment in numerous marine organisms, such as epi-oxazoline
halipeptin D [8].
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2. Materials and methods
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2.1. Analytical methods
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All reactions were monitored on thin-layer chromatographic
(TLC) Merck 60 F-254 silica-gel plates (0.25 mm layer thickness).
Column chromatography was performed on silica gel (70–230
mesh) using dichloromethane and methanol mixture as eluents.
Melting points were determined on an Electrothermal 9002 appa-
ratus and were uncorrected. The optical rotations were measured
by an Atago Polax-2L polarimeter. NMR spectra were recorded on
a Bruker AC 300 spectrometer [300 MHz (1H) and 75 MHz (13C)].
All chemical shifts were reported as d values (ppm) relative to
internal tetramethylsilane. Time-of-flight mass spectroscopy (TOF-
MS ES+) was carried out on Micromass, UK and Manchester. Ultra-
violet (UV) detector (Jasco 2075) and a data system processor was
used (Clarity Lite) to confirm the presence of the targeted products.
Detection was performed at 254 nm and 365 nm.
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On the other hand, 2-oxazolines are considered an important
class of heterocycles and are versatile intermediates in synthetic
organic chemistry [9,10]. Moreover, chiral oxazolines have been
widely used in asymmetric synthesis both as building blocks
[11], protecting groups for amino alcohols [12] and as auxiliaries,
metal entrapment ligands and ligands [13]. In this setting, a num-
ber of methods have been developed for the preparation of 2-oxaz-
olines from carboxylic acids [14], carboxylic esters [15], nitriles
[16], aldehydes [17], hydroxyamides [18] and olefins [19]. In spite
of the potential utility of aforementioned routes for the synthesis
of oxazoline derivatives, many of these procedures have demon-
strated various drawbacks including strong acidic conditions, long
reaction times, tedious work-up, low yields, use of expensive, toxic
or non-reusable catalysts, high temperatures, harsh reaction condi-
tions and use of toxic solvents such as CCl4, hexachloroethane or
chlorobenzene and/or co-occurrence of several side reactions. In
some cases, more than one step is required for the synthesis of
these heterocycles. Therefore, to avoid these limitations, there is
still a need to search for an efficient method with regard to toxicity,
solubility and reaction time.
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2.2. Chemicals
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(S)-2-Amino-3-methylbutanoic acid (
phenylacetic acid (( )-(+)-( )-phenylglycine), (S)-2-amino-3-
phenylpropanoic acid ( -phenylalanine), (S)-2-Amino-4-methyl-
pentanoic acid ( -leucine), (S)-2-Aminopropanoic acid ( -alanine),
(S)-2-amino-3-(1H-indol-3-yl)propanoic acid ( -tryptophan), suc-
cinonitrile and trimethylsilylchloride were purchased from Sig-
ma–Aldrich. The (R)-2-amino-1-butanol c was purchased from
Sigma–Aldrich while the a-amino alcohols a, b, d, e, f, g and e were
L-valine), S-(+)-a-amino-
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a
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L
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L
L
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L
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obtained by the reduction of the corresponding amino acids using
the method developed by Meyers [20]. 1,1-Diphenyl-2-pic-
rylhydrazyl radical (DPPH), ethylenediaminetetraacetic acid
(EDTA), potassium phosphate buffer,
a-tocopherol, nitro blue
tetrazolium (NBT) and riboflavin were purchased from Aldrich
(St. Louis, MO). All the other chemicals and solvents used in this
work were of an analytical quality and purchased from commercial
spots.
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In the present work, we describe the synthesis in one step of
2.3. Preparation of the synthetic compounds (general procedure)
some new chiral 4-substituted 2-cyanoethyl-oxazolines using
a-
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amino alcohols and the monohydrochloride of an iminoether as
starting materials. The results indicate that the present protocol
The synthesis of the 4-ethoxy-4-iminobutanenitrile monohy-
drochloride (2) was performed according to the modified method
described by Ben Ammar et al. [21]. The synthesis of 3-[4-ben-
zyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile (3a–g) was syn-
thesized as described by Meyers [22] (Scheme 1). The structures
is potentially applicable for the chemoselective conversion of
a-
amino alcohols to their corresponding 2-oxazolines in the presence
of iminoether. It is well known that the oxazolines’ derivatives
Epi-oxazoline halipeptin D
Brasilibactin A
Fig. 1. Examples of several biologically-active and pharmacological oxazolines.
Please cite this article in press as: R. Hassani et al., New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activ-
ities, Chemico-Biological Interactions (2014), http://dx.doi.org/10.1016/j.cbi.2014.04.003

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(CDCl3, 75 MHz): 14.15 (CH2), 24.20 (CH2), 41.58 (CH2), 67.26
(CH), 72.22 (CH2), 118.70 (CN), 126.67 (CH), 128.59 (CH), 129.35
(CH), 137.55 (C), 164.35 (C). TOFMS ES+ for C13H14N2O theoretical
[M+H]+: 215.1184; measured [M+H]+: 215.1184.
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2.3.2.2. 3-[(4S)-4,5-Dihydro-4-phenyl-1,3-oxazol-2-yl] propanenitrile
(3b). 93%; Yellow oil; [dichloromethane/methanol (99:01)].
[a]
D = [+15; c 1, CHCl3]. 1H NMR (CDCl3, 300 MHz): 2.63–2.77 (m,
4H), 4.14 (dd, 1H; J 16.8 Hz, J 8.4 Hz), 4.66 (dd, 1H; J 10.2 Hz, J
8.4 Hz), 5.22 (dd, 1H; J 18. 3 Hz, J 9.9 Hz), 7.21–7.33 (m, 5H). 13C
NMR (CDCl3, 75 MHz): 14.18 (CH2), 24.28 (CH2), 69.59 (CH),
75.30 (CH2), 118.73 (CN), 126.59–127.83 (CH), 128.65 (CH),
128.76–128.88 (CH), 141.73 (C), 165.31(C). TOFMS ES+ for
C12H12N2O theoretical [M+H]+: 201.1028; measured [M+H]+:
201.1030.
Scheme 1. Synthesis of new 3[4,5-dihydro-1,3-oxazol-2-yl] propanenitriles (3a–g).
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of the final products (3a–g), as well as the one of the intermediate
compounds were confirmed by 1H NMR, 13C NMR and TOFMS ES+.
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2.3.2.3. 3-[(4R)-4,5-Dihydro-4-ethyl-1,3-oxazol-2-yl] propanenitrile
(3c). 85%; Yellow oil; [dichloromethane/methanol (99:01)].
[a]
D = [+15; c 1, CHCl3]. 1H NMR (CDCl3, 300 MHz): 0.86–1.06 (m,
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169
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2.3.1. Synthesis of the 4-ethoxy-4-iminobutanenitrile
monohydrochloride
3H), 1.47–1.67 (m, 2H), 2.58–2.65 (t, 2H), 2.68–2.74 (t, 2H), 3.91
(dd, 1H; J 15.6 Hz, J 8.1 Hz), 4.02–4.03 (m, 1H), 4.05 (dd, 1H; J
1.8 Hz, J 1.2 Hz). 13C NMR (CDCl3, 75 MHz): 10.19 (CH2), 14.56
(CH3), 24.55 (CH2), 28.80 (CH2), 67.84 (CH), 72.88 (CH2), 116.71
(CN), 164.08 (C). TOFMS ES+ for C8H12N2O theoretical [M+H]+:
153.1028; measured [M+H]+: 153.1015.
Succinonitrile (3 g, 37.45 mmol), absolute ethanol (3.45 g,
74.91 mmol) and trimethylsilylchloride (TMSiCl) (4.07 g,
37.45 mmol) were respectively added in a Schlenk tube. The solu-
tion was cooled to ꢀ30 °C for 72 h until the precipitation of the salt
was achieved. Subsequently, the resulting salt was washed with
anhydrous ether and dried under vacuum and inert atmosphere.
The imidate chloride was obtained in 68% yield. 1H NMR (DMSO,
300 MHz): 1.01–1.09 (t, 2H), 2.38–2.42 (t, 2H), 4.06–4.11 (t, 2H),
4.42–4.49 (q, 3H), 7.23 (s, 1H), 7.40 (s, 1H). 13C NMR (DMSO,
75 MHz): 14.05 (–CH3), 28.64 (CH2), 30.50 (CH2), 60.85 (CH2),
119.04 (–CN), 171.52 (C@NH+2).
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2.3.2.4. 3-[(4S)-4,5-Dihydro-4-isopropyl-1,3-oxazol-2-yl] propaneni-
trile (3d). 89%; Yellow oil; [dichloromethane/methanol (99:01)].
[
a
]
D = [ꢀ20; c 1, CHCl3]. 1H NMR (CDCl3, 300 MHz): 0.78 (d, 3H; J
6.8 Hz), 0.85 (d, 3H; J 7 Hz), 1.12–1.22 (m, 1H), 1.57–1.73 (m,
1H), 2.47–2.68 (m, 4H), 3.75–3.94 (m, 1H), 4.04–4.22 (m, 1H) .13
C
NMR (CDCl3, 75 MHz): 14.23 (CH2), 18.10 (CH3), 18.67 (CH3),
24.24 (CH2), 32.56 (CH), 70.63 (CH2), 72.13(CH), 118.68 (CN),
163.66 (C). TOFMS ES+ for C9H14N2O theoretical [M+H]+:
167.1184; measured [M+H]+: 167.1180.
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2.3.2. Synthesis of the 4-substituted 2-cyanoethyl-oxazolines (3a–g)
The synthesis of 3-[(4S)-4-benzyl-4,5-dihydro-1,3-oxazol-2-yl]
propanenitrile (3a) is representative. In a Schlenk tube equipped
with a magnetic bar, the monohydrochloride of the 4-ethoxy-4-
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2.3.2.5. 3-[(4S)-4,5-Diyhdro-4-isobutyl-1,3-oxazol-2-yl] propaneni-
iminobutanenitrile (1.07 g, 6.61 mmol) and the (L)-phenylalaninol
trile (3e). 85%; Yellow oil; [dichloromethane/methanol (99:01)].
(1 g, 6.61 mmol) were introduced respectively in 50 mL of anhy-
drous methylene chloride. Then, the mixture was heated at reflux
for 10 h under an inert atmosphere. After the removal of NH4Cl by
filtration, the organic phase was evaporated and the obtained res-
idue was chromatographed on silica gel to afford 96% of the com-
pound (3a). The yield, optical rotation, and data for individual
compounds are given in (Table 1).
[a
]D=[+ 20; c 1, CHCl3]. 1H NMR (CDCl3, 300 MHz): 0.84–0.88 (m,
6H), 1.13–1.27 (m, 1H), 1.42–1.56 (m, 1H), 1.61–1.71 (m, 1H),
2.48–2.70 (m, 4H), 3.72–3.79 (m, 1H), 3.80–4.14 (m, 1H), 4.23–
4.33 (m, 1H). 13C NMR (CDCl3, 75 MHz): 13.33 (CH2), 14.55 (CH3),
15.06 (CH3), 23.06 (CH), 29.58 (CH2), 45.82 (CH2), 61.03 (CH),
72.34 (CH2), 118.17 (CN), 164.25 (C). TOFMS ES+ for C10H16N2O the-
oretical [M+H]+: 181.1341; measured [M+H]+: 181.1340.
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2.3.2.1. 3-[(4S)-4-Benzyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile
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240
241
242
(3a). 96%; Yellow oil; [dichloromethane/methanol (99:01)];
2.3.2.6. 3-[(S)-4,5-Dihydro-5-methyl-1,3-oxazol-2-yl] propanenitrile
(3f). 80%; Yellow oil; [dichloromethane/methanol (99:01)].
[
a
]
D = [ꢀ25; c 1, CHCl3]. 1H NMR (CDCl3, 300 MHz): 2.66–2.72
(m, 4H), 2.70–2.72 (dd, 1H; J 4.2 Hz, J 2.4 Hz), 3.05 (dd, 1H; J
13.8 Hz, J 5.4 Hz), 4.01 (dd, 1H; J 8.4 Hz, J 7.2 Hz), 4.22 (dd, 1H; J
9.3 Hz, J 5.7 Hz), 4.37–4.43 (m, 1H), 7.18–7.28 (m, 5H). 13C NMR
[
a
]
D = [ꢀ23; c 1, CHCl3]. 1H NMR (CDCl3, 300 MHz): 1.23 (m, 3H),
2.48–2.63 (m, 4H), 3.29 (dd, 1H; J 13.8 Hz, J 7.2 Hz), 3.83 (dd, 1H;
J 14.1 Hz, J 9.3 Hz), 4.55–4.65 (m, 1H). 13C NMR (CDCl3, 75 MHz):
14.04 (CH2), 20.41 (CH3), 30.82 (CH2), 60.40 (CH2), 66.11 (CH),
118.12 (CN), 169.53 (C). TOFMS ES+ for C8H12N2O theoretical
[M+H]+: 139.0827; measured [M+H]+: 139.0830.
Table 1
Preparation and physicochemical properties of new 2-cyanoethyl-oxazolines (3a–g)
according to Scheme 1.
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2.3.2.7. 3-[(4S)-((1H-Indol-3-yl)-methyl)-4,5-Diyhdro-1,3-oxazol-2-
Entrya
Config.
Yield%b
[
a
]
Physical aspect
D
yl] propanenitrile (3g). 73%; Yellow oil; [dichloromethane/metha-
D = [ꢀ17; c 1, MeOH]. 1H NMR (CDCl3, 300 MHz):
3a
3b
3c
3d
3e
3f
L
L
L
D
L
L
L
96
93
85
89
85
80
73
ꢀ25 (c 1, CHCl3)
+15 (c 1, CHCl3)
+15 (c 1, CHCl3)
ꢀ25 (c 1, CHCl3)
+20 (c 1, CHCl3)
ꢀ23 (c 1, CHCl3)
ꢀ17 (c 1, CHCl3)
Yellow oil
Yellow oil
Yellow oil
Yellow oil
Yellow oil
Yellow oil
Yellow oil
nol (99:01)]. [
a]
2.60–2.66 (m, 4H), 2.84 (dd, 1H; J 14.4 Hz, J 7.8 Hz), 3.15 (dd, 1H;
J 14.7 Hz, J 5.1 Hz), 4.02 (dd, 1H; J 8.4 Hz, J 7.2 Hz), 4.18 (dd, 1H;
J 8.7 Hz, J 7.2 Hz), 4.43–4.58 (m, 1H), 7.02 (s, 1H), 7.06–7.11
(td,1H; J 7.5 Hz, J 1.2 Hz), 7.13–7.19 (td, 1H; J 7.5 Hz, J 1.2 Hz),
7.33 (d, 1H; J 7.2 Hz), 7.59 (d, 1H; J 8.1 Hz), 8.35 (s, NH). 13C
NMR (CDCl3, 75 MHz): 14.07 (CH2), 29.45 (CH2), 60.95 (CH2),
65.95 (CH), 72.00 (CH2), 115.78 (CN), 110.75–135.79 (CIndol),
3g
a
Compounds (3a–g) was build-to-build distilled.
Yields of pure, isolated products (characterized by 1H, 13C NMR, TOFMS ES+).
b
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169.59 (C). TOFMS ES+ for C10H16N2O theoretical [M+H]+:
254.1293; measured [M+H]+: 254.1298.
(%) = [(ODcontrol ꢀ ODsample)/ODcontrol] ꢁ 100, where ODcontrol is the
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initial absorbance and ODsample is the value for the test sample
after incubation. IC50 was defined as the concentration (in
lg/
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2.4. Superoxide radical-scavenging activity
mL) of the substrate that causes as 50% loss of DPPH activity (color)
and was calculated by using the Litchfield and Wilcoxon test [25].
The results are expressed as the mean of data from at least three
independent experiments.
256
257
258
259
The ability of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl]
propanenitrile (3a) to ensnare superoxide anions was carried out
according two methods using the non-enzymatic and enzymatic
antioxidants assays.
320
2.6. Antimicrobial testing
260
261
262
263
264
265
266
267
268
269
270
271
272
273
2.4.1. A non-enzymatic antioxidant assay
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340
The antimicrobial activity of a synthetic molecule was tested on
the Gram-positive bacteria Staphylococcus aureus ATCC 25923 and
Enterococcus faecalis ATCC 29212 as well as the Gram negative bac-
teria Escherichia coli ATCC 25922 using the microdilution method
[26]. Overnight grown microbial suspensions were standardized
to approximately 105 cells/mL [27]. The microdilution method
was used to determine the Minimum Inhibitory Concentrations
(MICs) of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] propane-
The inhibition of NBT reduction by the photochemically-gener-
ated OÅ2ꢀ was used to determine the superoxide anion scavenging
activity of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] pro-
panenitrile (3a) according to the method described by Ben Man-
sour et al. [23]. In fact, the reaction mixture contained 6.5 mM
EDTA, 4 lM riboflavin, 96 lM NBT, and 51.5 mM potassium phos-
phate buffer (pH 7.4). Superoxide anions were measured by the in-
crease in the absorbance at 560 nm after 6 min of illumination at
room temperature. The synthetic product (3a) as well as the refer-
ence substance (Quercetin) were checked at different concentra-
tions with three repetitions. IC50 (concentration required to
inhibit NBT reduction by 50%) values were calculated from dose-
inhibition curves [24].
nitrile (3a). To do this, 100
approximately 105 cells/mL, was added to 100
(3a) dissolved in a minimum of EtOH (concentrations ranging from
10 g/mL to 500 g/mL in water). A set of tubes containing only
l
L of a microbial suspension containing,
l
L of the oxazoline
l
l
the microbial suspension served as the negative control. These
serially-diluted cultures were, then, incubated at 37 °C for 24 h.
Subsequently, 10 lL of each culture was placed on a substance-free
Mueller–Hinton agar plates and, further, incubated at 37 °C for
24 h. The MIC was defined as the lowest concentration of the tested
product that completely suppresses cell growth. A minimal Bacte-
ricidal Concentration (MBC) was defined as the lowest concentra-
tion of the extract that kills 99.99% of the tested bacteria [28].
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2.4.2. Inhibition of xanthine oxidase Activity
The inhibition of XOD-generated superoxide formation was
evaluated by measuring the UV absorbance of uric acid at
295 nm as proposed by Ben Mansour et al. [23b]. The assay mix-
ture consisted of a 100
centrations (100 g/mL; 200
solution of xanthine (X) (final concentration, 50
amine (final concentration, 0.2 mM), 200 L 0.1 mM EDTA, and
L distilled water. The reaction was initiated by adding
L XOD (final concentration, 11 mU/mL) dissolved in 0.2 M
l
L solution of test compound at each con-
g/mL and 300 g/mL), a 200
M) and hydroxyl-
l
l
l
l
lL
341
2.7. Analgesic testing
l
300
200
l
l
342
343
344
345
346
347
348
349
350
351
352
353
The analgesic activity can be performed according to the meth-
od of Koster et al. [29]. Swiss mice (20–30 g) were selected 1 day
prior to each test and were divided into three groups of six mice
each. The first group served as control (saline 10 mL/kg bw). The
second group was given the lysine acetylsalicylate (ASL)
(200 mg/kg bw) by the same route, as a reference drug. The third
group was treated with the oxazoline (3a) (100, 200, 300 and
400 mg/kg bw). All animals received 10 mL/kg (i.p.) of 1% acetic
acid, 30 min after treatment. The number of writhes was recorded
during 30 min, starting 5 min after the acetic acid injection. A
writhe is shown by an abdominal constriction and a stretching of
at least one hind limb.
phosphate buffer (pH 7.5). Following incubation at 37 °C for
30 min, the reaction was stopped by adding 100 L 0.58 M HCl.
l
The absorbance was then measured against a blank solution pre-
pared in the same way as described above, but replacing XOD by
buffer solution (no production of uric acid). A control solution
without the test compound was prepared in the same manner as
the assay mixture to measure the total uric acid production which
was calculated from the differential absorbance.
292
2.5. DPPH radical-scavenging activity
293
294
295
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297
298
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300
301
302
303
304
305
306
307
308
309
310
311
312
The nitrogen centered stable free radical 1,1-diphenyl-2-pic-
rylhydrazyl (DPPH) has often been used to characterize antioxi-
dants. It is reversibly reduced, and the odd electron in the DPPH
free radical gives a strong absorption maximum at 517 nm, (purple
in color). This property makes it suitable for spectrophotometric
studies. A radical scavenging antioxidant reacts with the DPPH sta-
ble-free radical and converts into the 1,1-diphenyl-2-picrylhydr-
azine. The resulting decolorization is steochiometric with respect
to the number of the captured electrons. The change in the absor-
bance produced in this reaction has been used to measure the anti-
oxidant properties. The free-radical scavenging capacity of the new
synthetic oxazoline was determined with DPPHÅ. Ethanol solutions
354
3. Results and discussion
355
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358
359
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362
363
364
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368
369
370
371
The 2-oxazoline ring system has been familiar for more than a
century. A number of reliable preparative methods, developed
before this heterocycles, knew a widespread utility and are still
valuable and in use today. The direct synthesis of 2-cyanometh-
yl-4-ethyl-oxazolines has been investigated by Elliot et al. [30],
starting from malononitrile and
conditions. Garcia et al. [31], has made possible the preparation
of the related 2-cyanomethyl-oxazolines from 2,2-dimethylmalo-
a-amino alcohols under acidic
nitrile and
a-amino alcohols ranging from high to good yields
were prepared containing 100, 30, 10, 3 and 1
lg/mL of the syn-
using the Zn(OAc)2 as catalyst, although the formation of bis-
oxazoline and methyl oxazoline that is not defined in this work,
was observed. However, this reported method for the synthesis
of 2-oxazolines suffer from some disadvantages such as long reac-
tion times, low yields, high temperatures with elimination of
ammonia, problems associated with the solubility of amino alcohol
in toluene and the lower stability of the unsubstituted oxazoline
ring which led to difficulties in scaling up reactions using amino
thetic product (3a) and 23.6 g/mL of DPPH. After incubation for
l
30 min at ambient temperature, the absorbance of the remaining
DPPH was determined colorimetrically at 517 nm. Radical scaveng-
ing activity was measured as the decrease in the absorbance of the
samples versus a DPPH standard solution. Results were expressed
as ‘‘percentage inhibition’’ (%) of the DPPH and the mean 50%
inhibiting concentration (IC50). % is defined by the formula:
Please cite this article in press as: R. Hassani et al., New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activ-
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alcohol as the starting material. Consequently, the design of a sim-
ple, inexpensive and environmentally-friendly method for the syn-
thesis of 2-oxazolines is a worthwhile.
Cl2 except for the compound (3g), the reaction was developed in a
solvent mixture CH2Cl2/EtOH as the (L)-tryptophanol is insoluble in
CH2Cl2.
The synthetic method of chiral 2-cyanoethyl-oxazolines is
In 13C NMR spectroscopy, these compounds showed character-
istic signals at 115.6–118.7 ppm for the cyano carbon atom, and in
the range 164.0–169.6 ppm for the sp2 C@N carbon atom. The
OCH2 protons of 4-monosubstituted oxazoline rings are diastereo-
topic. Consequently, each of the three hydrogens of the oxazoline
ring displayed a separate signal in the 1H NMR spectra of com-
pounds (3a–g). The assignment of these signals was done by ana-
lyzing 13C–[1H], DEPT135 and HMQC spectra. 1D NOESY
experiment was varied out to differentiate between the diastereo-
topic OCH2 protons.
based on the cyclocondensation of some
a-amino alcohols with
an iminoether such as the 4-ethoxy-4-iminobutanenitrile hydro-
chloride (2). It is worth noting that the reaction of succinonitrile
with ethanol in the presence of gaseous hydrogen chloride in anhy-
drous medium (diethyl ether) represents the most widespread ac-
cess to compound (2). Meanwhile, this method suffers from the use
of hydrogen chloride and strong acids. However, a new approach
based on the transformation of malononitrile in the iminoether
monohydrochloride has been reported [21]. In the present work,
the 4-ethoxy-4-iminobutanenitrile hydrochloride (2) was prepared
from succinonitrile (1) and trimethylsilylchloride using absolute
ethanol as a solvent at ꢀ30 °C for 72 h (Scheme 1).
We hypothesize that the oxazoline group with another func-
tional group (CN) could provide a strong chelating effect in order
to bind to the active site. We anticipated that the rigidity of the
molecule may also offer some advantages for a good drug. On the
other hand, the variation of the substituent 2-oxazoline has a sig-
nificant effect on the activity. In addition, aromatic groups having
oxazoline in position (4) and the cyano-group in position (2) fared
much better and more specifically increase the bioactive effect. To
prove our hypothesis, we have started by testing the antioxidant
activity.
Radical scavengers may directly react and quench the peroxide
radicals to terminate the peroxidation chain reactions and improve
the quality and stability of food products. Assays based upon the
use of DPPHÅ and OÅ2ꢀ radicals are among the most popular spectro-
photometric methods for the determination of the antioxidant
capacity of foods, beverages, vegetable extracts and chemical prod-
ucts. These chromogens and radical compounds can directly react
with antioxidants. Additionally, DPPHÅ and O2Åꢀ scavenging meth-
ods have been used to evaluate the antioxidant activity of com-
pounds due to the simple, rapid, sensitive, and reproducible
procedures [33]. First, it has been reported that antioxidant prop-
erties of some oxazolines are effective mainly via the scavenging
of superoxide anion radicals. The superoxide anion is the precursor
of hydrogen peroxide, hydroxyl radical, and singlet oxygen which
induce oxidative damage in lipids, proteins and DNA [34]. Superox-
ide radicals are normally formed first, and their effects can be mag-
nified because they produce other kinds of free radicals and
oxidizing agents. Superoxide anions derived from dissolved oxygen
by the nitroblue tetrazolium/riboflavine system will reduce NBT in
this system. The NBT assay is based on the capacity of the extracts
to inhibit the photochemical reduction of nitroblue tetrazolium
(NBT) in the presence of riboflavin.
Simple washing furnished the rather unstable crude 4-ethoxy-
4-iminobutanenitrile monohydrochloride (2) as a solid. A spectrum
of the crude free iminoether monohydrochloride (2) could also be
obtained but it was not possible to reach an accurate elemental
analysis of the crude product. Furthermore, the previously noted
instability of this class of salt compounds was then observed when
attempts were made to purify. Recrystallization and silica/alumina
chromatography all failed to improve the situation and actually
yielded a material of lower quality. The instability of the iminoe-
ther monohydrochloride (2) prompted us to form more stable
and easily characterizable derivatives in which the free imino-
group was protected. This simple method allowed us to obtain
an unstable product which can be degraded easily in the presence
of air, but with minimal impurities and without using strong acids.
However, despite this instability, the iminoether monohydrochlo-
ride (2) is a synthetic intermediate in the pathway leading to con-
densation systems and a precursor of the formation of the
oxazoline.
2-Oxazolines proved to be very sensitive to acidic conditions
undergoing the rearrangement leading to the opening cycle almost
quantitatively. Nevertheless, the rearrangement can be prevented
from working in inert conditions and in the absence of Lewis acids
and proton sources. We have demonstrated that the cycloconden-
sation proceeded smoothly at refluxing methylene chloride,
obtaining the expected product with 96% yield. Further, no side
products were detected in the reaction mixture as determined by
TLC analysis and 1H NMR spectra. This method avoids the multi-
step preparation of starting materials or the requirement of special
reagents. Hence, we designed a new type of oxazoline with a cya-
no-group to get some branched 2-cyanoethyl-oxazolines. In all
cases, the reaction affords the new 2-cyanoethyl-oxazolines (3a–
g) in high yields without any racemization of the asymmetric car-
bon (Table 1).
In the presence of an antioxidant that can donate an electron to
NBT, the purple color typical of the formazan decays, a change that
can be followed spectrophotometrically at 560 nm. The synthetic
product (3a) and the reference substance (Quercetin) were assayed
at different concentrations with three repetitions. The oxazoline
(3a) seems to be a potent antioxidant with an activity percentage
of 66% at the highest concentration (10 mg/mL) compared to the
positive control, Quercetin, and an IC50 of 4.31 mg/mL (Fig. 2).
This result confirms the ability of (3a) to decrease the XOD-
Recently, we have developed a synthesis method for easy and
efficient preparation of chiral 2-oxazolines based on cycloconden-
sation. We estimated that our methodology would be ready for the
preparation of a series of oxazoline with cyano group. The reaction
between the
a-amino alcohol and the monohydrochloride of an
iminoether allowed us to reach an unstable intermediate. Intramo-
lecular cyclization converts the final product after the release of
NH4Cl. It should be noted that the method is selective and the con-
version is complete for aromatic amino alcohols as for aliphatic
amino alcohols. It is also important to note that the synthesis of
mono-oxazoline is a chemoselective, but a time-dependent reac-
tion. Mono-oxazolines (3a–f) were satisfactorily produced after
10 h whereas the mono-oxazoline (3g) was exclusively obtained
generated superoxide radical in
manner with inhibition percentages up to 30.55%, 46.31%
and 60.30% at the concentrations 100 g/mL, 200 g/mL and
300 g/mL, respectively (Fig. 3).
a concentration-dependent
l
l
l
On the other hand, DPPH is a molecule containing a stable-free
radical. In the presence of an antioxidant that can donate an elec-
tron to DPPH, the purple color typical of the free DPPH radical de-
cays, a change that can be followed spectrophotometrically at
517 nm. This method is based on the reduction of DPPH in an alco-
holic solution in the presence of a hydrogen-donating antioxidant
due to the formation of the non-radical form of DPPH–H in the
reaction. DPPH is usually used as a reagent to evaluate the free
from the reaction of (
Our previous studies showed that the lowest yield of oxazoline is
often obtained with ( )-tryptophanol. However, in our original con-
ditions, the synthesis of the oxazoline series was obtained in CH2-
L)-tryptophanol with iminoether after 20 h.
L
Please cite this article in press as: R. Hassani et al., New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activ-
ities, Chemico-Biological Interactions (2014), http://dx.doi.org/10.1016/j.cbi.2014.04.003

CBI 7024
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R. Hassani et al. / Chemico-Biological Interactions xxx (2014) xxx–xxx
Fig. 2. Scavenging effects of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] pro-
panenitrile (3a) against photochemically-generated superoxide free radicals (OÅ2ꢀ).
Fig. 4. DPPH free-radical scavenging activity of the 3[(4S)-benzyl-4,5-dihydro-1,3-
oxazol-2-yl] propanenitrile (3a).
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530
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The activity of (3a) against the superoxide radical is a semi-
quantitative test [35], and the enzymatic assay (X/XOD) has more
relevance to physiological conditions than the photochemical-NBT
assay. The XOD catalyses the oxidation of hypoxanthine and xan-
thine to uric acid. During the oxidation of xanthine, the superoxide
radicals and the hydrogen peroxide are formed, this enzyme is con-
sidered as an important biological source of superoxide radicals
[36]. We can, then, conclude that (3a) is considered an antioxidant
not only because it acts as free-radical scavengers, but also because
it inhibits the XOD.Our results are very encouraging as far as the
superoxide radical (OÅ2ꢀ) is known to be a highly toxic species that
is generated by numerous biological and photochemical reactions.
Indeed, it can generates the hydroxyl radical which reacts with
DNA bases, amino acids, proteins, and polyunsaturated fatty acids,
and produces toxic effects.
In order to expand the library of oxazolines as a drug, analgesic
activity is of key interest. Most antiepileptic drugs are known to
have strong analgesic effects [37]. So far, the available analgesic
drugs exert a wide range of side effects and are either too potent
or too weak. The search for new analgesic compounds was a prior-
ity for pharmacologists and pharmaceutical industries [38]. Among
the several models of visceral pain, the writhing test has been
mostly used as a standard screening method [39]. This model in-
volves different nociceptive mechanisms, such as the sympathetic
system (Biogenic amines release), cyclooxygenases (COX) and their
metabolites and opioid mechanisms. Acetic acid acts indirectly by
inducing the release of an endogenous mediator which stimulates
the nociceptive neurons sensitive to NSAIDs (non-steroidal anti-
inflammatory drugs) and/or opioids. The subcutaneous administra-
tion of (3a) (100, 200, 300 and 400 mg/kg) produced a significant
reduction in the number of abdominal constrictions throughout
the entire period of observation in a dose- related manner with
respectively 61.9%, 64.29%, 71.43% and 73.81% in the acetic acid
writhing test (Table 2). The standard drug (ASL, 200 mg/kg) de-
creased the number of abdominal constrictions by 85.71% in the
acetic acid writhing test in mice (Table 2).
Fig. 3. Inhibition of xanthine oxidase activity of the 3[(4S)-benzyl-4,5-dihydro-1,3-
oxazol-2-yl] propanenitrile (3a).
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radical scavenging activity of antioxidants. This simple test can
provide information on the ability of a compound to donate an
electron and the mechanism of antioxidant action. The 3[(4S)-ben-
zyl-4,5-dihydro-1,3-oxazol-2-yl] propanenitrile (3a) was a very
potent radical scavenger, with a decrease percentage decrease vs.
the absorbance of the DPPH standard solution of 81.22%, at a con-
centration of 100
This value was slightly greater than that of the positive control,
g/mL -tocopherol.
Two different reactive species were used to evaluate the antiox-
lg/mL, and an IC50 values of 3.11 lg/mL (Fig. 4).
3
l
a
idant activity of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl]
propanenitrile (3a), the DPPH and superoxide radicals. Since the
DPPH radical is not biologically relevant, the DPPH assay was per-
formed as a preliminary study to estimate the direct free-radical
scavenging abilities of the test compound. The results obtained
with the tested product (3a) revealed a relatively strong antiradical
activity towards the DPPH free radical.
Results indicated that the 3[(4S)-benzyl-4,5-dihydro-1,3-oxa-
zol-2-yl] propanenitrile (3a) decreased significantly the NBT/ribo-
flavin generated superoxide radical in a concentration dependent
manner. The tested synthetic product shows a high ability to re-
duce the OÅ2ꢀ generation. This activity is dose-dependent and has
a similar profile to that of the Quercetin. In fact, no significant dif-
ference was observed when compared the IC50 of the tested com-
pound with Quercetin.
Using the conventional pharmacological model, we have dem-
onstrated the analgesic property of the product which can induce
an antinociception by a mechanism similar to nonnarcotics and/
or narcotic drugs, perhaps by blocking the receptor or the release
of endogenous substances that excite pain nerve endings [40].
NSAIDs such as ASL produce their antinociceptive and anti-inflam-
matory action via inhibiting cyclooxygenases in peripheral tissues,
Please cite this article in press as: R. Hassani et al., New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activ-
ities, Chemico-Biological Interactions (2014), http://dx.doi.org/10.1016/j.cbi.2014.04.003

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Table 2
609
610
611
612
613
614
615
616
617
618
619
620
621
of food contaminated with pathogenic bacteria, has been a real
concern to public health. E. coli accounted for the largest number
of epidemic cases and deaths.
Analgesic effect of the subcutaneous administration of 3[(4S)-benzyl-4,5-dihydro-1,3-
oxazol-2-yl] propanenitrile (3a) in the acetic acid 1% writhing test in mice.
Treatment
Dose
(mg/kg)
Number of
writhes s.e.m.
Inhibition of
writhing (%)
These observations show that the synthesized compounds are
capable of inhibiting the growth of Gram positive bacteria and
Gram negative bacteria. The compound (3a) possesses superior
an antimicrobial activity against all microorganisms under investi-
gation. There still exists a need for the development of new antimi-
crobial agents having superior activity and less side effects to
overcome resistant strains of microorganisms. Further studies to
puzzle out the mechanism of these compounds’ action and deter-
mine whether their activity is lethal or inhibiting to microorgan-
isms are underway.
Control (saline 10 mL/kg)
Synthetic product
ꢀ
42.50 2.00
16.50 2.50**
15.00 1.50**
12.00 3.00**
11.50 3.00
6.16 0.37**
ꢀ
100
200
300
400
200
61.90
64.29
71.43
73.81
85.71
Lysine acetylsalicylate
(reference drug)
Values are expressed as mean s.e.m. n = 6 animals.
**
P < 0.001.
622
4. Conclusion
Table 3
Antibacterial activity of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] propaneni-
trile, expressed as Minimum Inhibitory Concentration (MIC) and as Minimum
Bactericidal Concentration (MBC).
623
624
625
626
627
628
629
630
631
632
633
634
635
636
In this paper, we report an efficient one-pot synthesis of new
chiral 4-substituted 2-cyanoethyl-oxazolines with high yields by
condensation of optically-pure
a-amino alcohols and iminoether
S. aureus
E. faecalis
E. coli
monohydrochloride. The simple work-up, mild conditions and high
yields are features of this approach. This work falls within the
framework of valorization of these compounds as bioactive prod-
ucts. In this context, the antioxidant, analgesic and antimicrobial
activities of these compounds were assessed. These experiments
established that the investigated oxazolines had an excellent anti-
microbial, analgesic activity and a good antioxidant property. This
fact may be evidence for the potential of this class of compounds in
the search and development of new bioactive compounds. In terms
of their impacts, virtually all the activities studied revealed encour-
aging results.
ATCC25923 ATCC25922 ATCC 25922
MIC MBC MIC MBC MIC
MBC
411
3[(4S)-benzyl-oxazoline]
propanenitrilea
g/mL)
Ampicilinb
g/mL)
15
275
225
32.5 350
2.5 125
64
6
(l
(l
1.5
275
a
Values were expressed as means standard deviation of three experiments.
Positive control.
b
572
573
574
575
576
577
578
579
580
581
582
583
584
585
586
587
588
589
590
591
592
593
594
595
596
597
598
599
600
601
602
603
604
605
606
607
608
thereby reducing the PGE2 (prostaglandin E2) synthesis and inter-
fering with the mechanism of transduction in primary afferent
nociceptors [41]. This particular activity of the crude extract of
the defensive secretion and its semi-purified fractions is probably
related to their anti-inflammatory properties [42]. The analgesic
property of the 3[(4S)-benzyl-4,5-dihydro-1,3-oxazol-2-yl] pro-
panenitrile (3a) was demonstrated. Evaluating analgesic properties
in a single assay may not provide a full understanding of the ac-
tions of the synthetic product (3a) or it’s utility. Further works to
ascertain its mechanism of action are necessary.
We show continued interest in the exploration of the various
pharmacological activities of oxazoline (3a) as an active sites
source. Besides, in this context, another important feature of the
biological properties of (3a) was demonstrated by testing the anti-
microbial activity.
637
638
Conflict of Interest
The authors declare that there are no conflicts of interest.
639
Transparency document
640
641
642
The Transparency document associated with this article can be
found in the online version.
643
6. Uncited reference
The antibacterial activity of the 3[(4S)-benzyl-4,5-dihydro-1,3-
oxazol-2-yl] propanenitrile (3a) was evaluated on three pathogenic
bacteria. Our results revealed that this molecule exhibited various
levels of antibacterial effect against all the tested bacterial strains.
Minimum Inhibitory Concentration (MICs) values ranging from 15
Q2 644
[32].
645
Acknowledgments
to over 64
lg/mL, and Minimum Bactericidal Concentration
646
647
648
The authors are grateful to the DGRS (Direction Générale de la
Recherche Scientifique) of the Tunisian Ministry of Higher Educa-
tion and Scientific Research for the financial support.
(MBCs) values ranging from 275 to more than 411
l
g/mL (Table 3).
S. aureus was the most susceptible bacterial species, followed by E.
faecalis and finally Escherichia coli, with MIC values of 15, 32.5 and
64
control against S. aureus (225
exhibited the same activity with an MBC value of 275
l
g/mL respectively. Compared to ampicillin, used as a positive
g/mL), the tested oxazoline (3a)
g/mL.
649
References
l
l
650
651
652
653
654
655
656
657
658
659
660
661
662
663
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ities, Chemico-Biological Interactions (2014), http://dx.doi.org/10.1016/j.cbi.2014.04.003

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Please cite this article in press as: R. Hassani et al., New chiral 4-substituted 2-cyanoethyl-oxazolines: Synthesis and assessment of some biological activ-
ities, Chemico-Biological Interactions (2014), http://dx.doi.org/10.1016/j.cbi.2014.04.003