Synthesis, antibacterial and antifungal activities of new chiral 5-alkyl-3-(1′-benzenesulfonylpyrrolidin-2′-yl)-4,5-dihydro-1,2, 4-oxadiazin-6-ones

Article abstract of DOI:10.1016/j.crci.2009.11.006

Chiral α-bromoacid chlorides can be easily prepared from amino acids. Their condensation with new 1-benzenesulfonylpyrrolidin-2-carboxamidoxime derived from proline lead to the corresponding O-(bromoacyl) amidoximes. The latter afforded new chiral 5-alkyl-3-(1′-benzenesulfonylpyrrolidin- 2′-yl)-4,5-dihydro-1,2,4oxadiazin-6-ones in good chemical yields via intramolecular cyclization in the presence of one equivalent of NaH. These new compounds were evaluated for their antibacterial and antifungal activities using micro-dilution tests against some strains of bacteria and fungi. Our compounds showed an excellent antibacterial activity, which is better than the drug levofloxacin.

Full text of DOI:10.1016/j.crci.2009.11.006

C. R. Chimie 13 (2010) 1278–1283
Contents lists available at ScienceDirect
Comptes Rendus Chimie
www.sciencedirect.com
´
Full paper/Memoire
Synthesis, antibacterial and antifungal activities of new chiral 5-alkyl-3-
(10-benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-1,2,4-oxadiazin-6-ones
*
ˆ
´
Najeh Tka, Nafaa Jegham, Bechir Ben Hassine
Laboratoire de synthe`se organique asyme´trique et catalyse homoge`ne (01 UR 1201), de´partement de chimie, faculte´ des sciences de Monastir,
universite´ de Monastir, avenue de l’Environnement, 5019 Monastir, Tunisia
A R T I C L E I N F O
A B S T R A C T
Chiral -bromoacid chlorides can be easily prepared from amino acids. Their condensation
Article history:
a
Received 10 June 2009
with new 1-benzenesulfonylpyrrolidin-2-carboxamidoxime derived from proline lead to
the corresponding O-(bromoacyl) amidoximes. The latter afforded new chiral 5-alkyl-3-
(10-benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-1,2,4oxadiazin-6-ones in good chemi-
cal yields via intramolecular cyclization in the presence of one equivalent of NaH. These
new compounds were evaluated for their antibacterial and antifungal activities using
micro-dilution tests against some strains of bacteria and fungi. Our compounds showed an
excellent antibacterial activity, which is better than the drug levofloxacin.
Accepted after revision 10 November 2009
Available online 21 January 2010
Keywords:
Aminoacids
Amidoxime
a-bromoacids
´
ß 2009 Academie des sciences. Published by Elsevier Masson SAS. All rights reserved.
1,2,4-oxadiazin-6-ones
Antibacterial and antifungal activities
´
´
R E S U M E
Mots cle´s :
Des chlorures de
a-bromoacides chiraux pre´pare´s facilement au de´part d’acides amine´s
Aminoacides
Amidoxime
´
`
´
sont condenses avec le 1-benzenesulfonylpyrrolidin-2-carboxamidoxime derivant de la
proline et conduisent aux O-(bromoacyl) amidoximes correspondantes. Ces derniers
compose´s conduisent aux nouvelles 5-alkyl-3-(10-benze`nesulfonylpyrrolidin-20-yl)-4,5-
dihydro-1,2,4-oxadiazin-6-ones chirales avec de bons rendements chimiques via une
a-bromoacides
1,2,4-oxadiazin-6-ones
Activite´s antibacte´riennes et antifongiques
´
´
´
cyclisation intramoleculaire en presence d’un equivalent de NaH. Ces nouveaux produits
ont e´te´ e´value´s pour leurs activite´s antibacte´rienne et antifongique en utilisant des tests de
´
microdilution contre quelques souches de bacteries et de champignons. L’un de nos
´ ´ ´
a montre une excellente activite antibacterienne meilleure que celle du
produits
´
´
medicament levofloxacin.
ß 2009 Acade´mie des sciences. Publie´ par Elsevier Masson SAS. Tous droits re´serve´s.
1. Introduction
The high synthetic value of oxadiazinones has been
demonstrated specially in Diels–Alder cycloaddition [4,5]
Oxadiazinones are quite important biologically-active
compounds which have gained more interest thanks to
their utility as central nervous system stimulants [1,2] and
as oxidase B (MAO-B) inhibitor [3]. As far as we know, no
works reporting the antibacterial or antifungal activities of
this class of heterocycles have been reported.
and Favorski ring contraction [6]. Recently, fluorinated
oxadiazinones have been proposed as a rare example of
low molecular weight (LMW) hydrogelators [7].
On the other hand, the synthesis of optically-active
oxadiazinones has recently attracted considerable
interest in the asymmetric organic synthesis. Optical-
ly-active 1,3,4-oxadiazin-2-ones have been successfully
investigated by Hitchcock et al. and Vaughn and Hitch-
cock [8,9] as chiral auxiliaries in the aldol reaction.
Moreover, these compounds have been utilised by
* Corresponding author.
E-mail address: bechirbenhassine@yahoo.fr (B. Ben Hassine).
1631-0748/$ – see front matter ß 2009 Acade´mie des sciences. Published by Elsevier Masson SAS. All rights reserved.
doi:10.1016/j.crci.2009.11.006

N. Tka et al. / C. R. Chimie 13 (2010) 1278–1283
1279
Husson et al. as chiral auxiliaries to perform diaster-
eoselective alkylations [10].
easy method for the synthesis of 1,2,4-oxadiazoles [14,15].
As part of our work, directed towards the synthesis of new
heterocyclic compounds from amino acids [16,17], we
investigate chiral new (2S)-1-benzenesulfonylpyrrolidine-
Among oxadiazinones, 1,2,4-oxadiazin-6-ones are not
very broadly explored and little is known about these
compounds which were prepared only once by Hussein
et al. [11]. Hussein et al.’s method was not general and
gave 1,2,4-oxadiazin-6-ones with moderate yields rang-
ing from 12 to 73% and with no indication of the product’s
optical purity. Therefore, there is further scope to explore
more efficient synthetic methods. Herein, chemically
speaking, we report a simple and practical method to
achieve the synthesis of new optically pure 5-alkyl-3-(10-
benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-1,2,4-oxa-
diazin-6-ones by the condensation of chiral 1-benzene-
sulfonylpyrrolidin-2-carboxamidoxime with chiral 2-(S)-
2-bromoacid chlorides, followed by cyclization in the
presence of one equivalent of sodium hydride. Some tests
have been conducted in order to evaluate the antibacterial
and antifungal activities of our new 1,2,4-oxadiazin-6-
ones.
2-carboxamidoxime to cyclise with
a-bromoacid chlorides
yielding new 1,2,4-oxadiazin-6-ones.
The synthesis of the amidoxime group, mostly involve
the preparation of the cyano group, followed by the
conversion into an amidoxime group. During this study, we
describe the first synthesis of chiral (2S)-1-benzenesulfo-
nylpyrrolidine-2-carboxamidoxime 2. This compound was
easy prepared by treatment of (2S)-1-benzenesulfonyl-
pyrrolidine-2-carbonitrile 1 with a hydroxylamine alkaline
solution. However, unfortunately, the treatment of
a-
aminonitriles derived from other amino acids did not
provide the corresponding amidoxime with good yields
even after heating and prolonging the reaction time.
Work on condensation of amidoximes with acid
chlorides has been described before. It is well known that
acid chloride reacts with the O-part of the amidoxime
[18,19]. The desired esterification products 3a–f were
afforded with use of one equivalent of
a-bromoacid
2. Results and discussion
chloride and 1.2 equivalent of triethylamine. Reaction
was performed in an ice bath with short reaction times
(45–60 min). In the event of longer reaction times, the
reaction leads to a complex mixture.
2.1. Chemistry
The synthetic route to the target compounds 4a–f is
Finally, the intermediates 3a–f undergo an intermolec-
ular cyclization in the presence of one equivalent of NaH to
yield compounds 4a–f within 3 h at ꢀ25 8C with good
yields. Only one equivalent of NaH has been used in order
to avoid the racemisation of the initial stereocenter of the
proline by a possible elimination of the proton. We have
found that compounds 4a–f were formed without a
significant racemization of the asymmetric carbon 5 of
the oxadiazinone ring. This result is proved by NMR and by
HPLC studies.
outlined in Scheme 1. We have successfully converted
commercially available (L)-proline to the corresponding
optically pure (2S)-1-benzenesulfonylpyrrolidine-2-car-
bonitrile 1 [12]. The treatment of this compound with
hydroxylamine hydrochloride and two equivalents of
triethylamine overnight at room temperature led to the
corresponding chiral (2S)-1-benzenesulfonylpyrrolidine-
2-carboxamidoxime 2. The condensation of amidoxime 2
with different
the corresponding
procedure [13], gave the chiral (O-bromoacyl) (2S)-1-
benzenesulfonylpyrrolidine-2-carboxamidoximes 3a–f
a
-bromoacid chlorides, easily prepared from
a
-aminoacids following known
a
According to 1H NMR spectra of compound 4f prepared
from optically pure (20S)-1-benzenesulfonylpyrrolidine-20-
carboxamidoxime 2 and (D,L) valine, the signal relative to
with chemical yields ranging from 79 to 87%. The final
step of this sequence is the intramolecular cyclization of
3a–f in the presence of one equivalent of NaH affording the
corresponding 5-alkyl-3-(10-benzenesulfonylpyrrolidin-
20-yl)-4,5-dihydro-1,2,4-oxadiazin-6-ones 4a–f with good
chemical yields and excellent diastereomeric excesses.
The addition reaction of amidoxime to the activated
carboxylic acid derivatives has always been described as an
the proton linked to the asymmetric carbon of oxadiazi-
none ring resonates in different positions for each pair of
diasteromers. As shown in Fig. 1, the signal of the C*-H
proton of the mixture of the two diastereomers (20S,5R)
and (20S,5S) 4f appeared as two doublets. A first doublet at
3.95 ppm (J = 5.7 Hz) corresponding to the diastereomer
(20S,5S) and a second doublet at 3.99 ppm (J = 5.1 Hz)
Scheme 1. Synthesis of (20S, 5R), 5-alkyl-3-(1’-benzenesulfonyl-pyrrolidin-20-yl)-4,5-dihydro-1,2,4-oxadiazin-6-ones 4a–f. Reaction conditions: (i)
NH2OH.HCl/Et3N/CHCl3; (ii) 2-(S)-2-bromo-acid chlorides/Et3N/CH2Cl2, 0 8C ; (iii) NaH/THF, 3 h, ꢀ25 8C.

1280
N. Tka et al. / C. R. Chimie 13 (2010) 1278–1283
Fig. 1. The signal of the C*-H proton of (20S, 5R), (20S, 5S) 4f and (20S, 5R) 4b 5-isopropyl-3-(10-benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-
1,2,4oxadiazin-6-ones.
corresponding to the diastereomer (20S,5R). However, the
signal of the C*-H proton of the single diastereomer
(20S,5R) 4b prepared from optically-pure(2’S)-1-benzene-
25923, Escherichia coli ATCC 25922, Klepsiella pneumonie
(clinically isolated) and Enterococcus faecalis ATCC 29212.
The same compounds were also tested to assess their
activity against fungi. The yeasts were Candida glabrata
ATCC 90030, C. albicans ATCC 90028, C. kreusei ATCC 6258
and C. parapsilosis ATCC 22019.
Antibacterial activities of synthesized 1,2,4-oxadiazin-
6-ones 4a–f were studied along with an antibacterial drug
named levofloxacin. The obtained minimum inhibitory
concentrations (MIC) values of the tested compounds and
standard are shown in Table 1. The results revealed that all
compounds showed superior activity than levofloxacin.
Reported compounds 4a–f were found to be very effective
at low concentrations, in some cases, the value of
minimum inhibitory concentrations (MIC) of our com-
pounds was 10 times lower than levofloxacin. Compound
4c shows a good antibacterial activity against E. faecalis at a
concentration of 0.12 mg mLꢀ1. Compounds 4e against
S. aureus and 4b against E. coli exhibit a very significant
antibacterial activity at a very low concentration, respec-
sulfonylpyrrolidine-2’-carboxamidoxime 2 and (L)-valine,
appeared as one doublet at 3.99 ppm (J = 4.2 Hz) showing
the high diastereoselectivity of the intramolecular cycli-
zation (Fig. 1). In the same way, 1H NMR spectra of
compounds 4a, c–e indicated the absence of peaks
expected from racemisation.
The optical purity of 1,2,4-oxadiazin-6-ones 4a–e was
confirmed by HPLC. According to the HPLC studies,
compounds 4a–e are single isomers, and no peaks
corresponding to other diastereomers were detected.
Finally, based on 1H NMR and HPLC, we can conclude
that the nucleophelic intermolecular substitution of
bromine by the amino group affording the oxadiazinone
ring has occurred in a typical SN2 manner.
2.2. Biological activities
The synthesized compounds were tested at concentra-
tion ranging from 1 to 0.0018 mg mLꢀ1 for antibacterial
activity against bacterial strains. The strains of micro-
organisms employed were Staphylococcus aureus ATCC
tively, 0.06 and 0.03 mg mLꢀ1
.
Table 1 shows also the antifungal activities of the
synthesized 1,2,4-oxadiazin-6-ones 4a–f in comparison
with the antifungal amphotericin B. Based on the obtained
Table 1
Antibacterial and antifungal activities of 1,2,4-oxadiazin-6-ones 4a–f (MIC, mg mLꢀ1).
Bacteria
MIC 4a
MIC 4b
MIC 4c
MIC 4d
MIC 4e
MIC 4f
Amphotericin B
Levofloxacin
K. pneumonie (clinical isolated)
S. aureus ATCC 27950
E. faecalis ATCC 29212
E. coli ATCC 25922
0.06
0.25
0.25
0.25
0.25
0.12
0.25
0.03
0.25
0.25
0.12
0.06
0.12
0.25
0.25
0.25
0.12
0.06
0.25
0.25
0.25
0.25
0.5
nd
nd
nd
nd
0.3
0.61
1.22
0.3
0.25
Yeasts
C. parapsilosis ATCC 22019 5 45
C. kreusei ATCC 6258 2.5 45
C. albicans ATCC 90028
0.25
0.25
0.12
0.25
0.25
0.25
0.25
0.25
0.25
0.25
0.03
0.25
0.25
0.12
0.5
0.05
0.05
0.05
0.05
nd
nd
nd
nd
0.25
0.12
0.25
0.12
0.25
0.25
0.25
0.25
0.12
C. glabrata ATCC 90030
Levofloxacin: controlled antibiotic; amphotericin B: controlled antifungal. MIC: Minimum Inhibitory Concentrations, values given as mg mLꢀ1; nd: not
determined.

N. Tka et al. / C. R. Chimie 13 (2010) 1278–1283
1281
values, the synthesized compounds showed moderate
antifungal activities with MIC values ranging from 0.03 to
dichloromethane was stirred overnight at room tempera-
ture under N2. After the removal of the SOCl2 excess under
vacuum, the residue was dissolved in dry dichloromethane
and added slowly to a cold solution of (2S)-1-bzenesulfo-
nylpyrrolidin-2-carboxamidoxime (5 mmol) and 1.2
equivalent of triethylamine (6 mmol) in dry dichloro-
methane at (0 8C). The reaction mixture was stirred for
60 min in an ice bath, and then 20 mL of saturated
ammonium chloride solution was added. After separation,
the aqueous layer was extracted with chloroform
(3 ꢁ 50 mL). The chloroformic combined extracts were
dried over MgSO4, filtered and concentrated in vacuo and
the residue obtained was purified by column chromatog-
raphy on silica gel (40% ethylacetate: 60% cyclohexane) to
give (O-bromoacyl) (2S)-1-benzenesulfonylpyrrolidin-2-
carboxamidoxime 3a–f.
0.25 mg mLꢀ1
. In all cases, the minimum inhibitory
concentrations of the active compound are higher than
the standard used antibiotic, except for compound 4e
against C. glabrata which was found to show a good
antifungal activity at concentration (0.03 mg mLꢀ1) lower
than amphotericin B (0.05 mg mLꢀ1).
It should be pointed out that this evaluation of the
antibacterial and antifungal activities of 1,2,4-oxadiazin-6-
ones was performed for the first time during this study.
3. Conclusion
In summary, we have synthesized novel biologically
important 5-alkyl-3-(10-benzenesulfonylpyrrolidin-20-yl)-
4,5-dihydro-1,2,4-oxadiazin-6-ones 4a–f using chiral 2-
(S)-1-benzenesulfonylpyrrolidin-2-carboxamidoxime and
2-(S)-2-bromoacid chlorides, both derived from inexpen-
sive and readily available amino acids. The synthetic
method has been developed for the first time during this
work giving 4a–f in high chemical yields and occurring
without a racemisation as shown by the 1H NMR and HPLC.
The reported compounds were screened for their antibac-
terial and antifungal activities against a spectrum of
microbial organisms. These studies proved that all
compounds 4a–f showed excellent antibacterial activities,
much better than levofloxacin. As for compound 4e against
C. glabrata, it showed a beneficial antifungal activity at a
minimum concentration (0.03 mg mLꢀ1). The test dealing
with other biological activities of these new products is
under investigation in our laboratory.
4.1.2. (2S)-N-phenylsulfonylpyrrolidin-2-carbonitrile 1
Yield = 89 %; mp = 99–101 8C; [
c = 0.5); IR (cmꢀ1): CN = 2236 (CN); 1H NMR (300 MHz,
CDCl3) : 2.02–2.19 (m, 4H), 3.33 (m, 2H), 4.58 (dd, 1H,
J = 6.9 Hz, J = 2.7 Hz), 7.53–7.88 (m, 5H); 13C NMR (75 MHz,
CDCl3): 24.73, 31.97, 47.58, 48.66, 118.03 (CN), 127.55,
a
]
D
ꢀ1040; (CHCl3,
n
d
d
129.42, 133.59, 137.29, MS/EI m/z = 236 (M+.). Anal. Calcd
for C11H12N2O2S,236.29: C 55.91, H 5.12, N 11.86 . Found: C
56.13, H 4.99, N 11.75.
4.1.3. (2S)-N-phenylsulfonylpyrrolidin-2-carboxamidoxime
2
Yield = 94 %; mp = 54–56 8C; [
a
]
D ꢀ112; (CHCl3, c = 0.5);
1H NMR (300 MHz, CDCl3) :
d
1.62–1.78 (m, 2H), 1.86–1.95
(m, 1H), 2.12–2.19 (m, 1H), 3.16–3.24 (m, 1H), 3.55–3.69
(m, 1H), 4.13 (dd, 1H, J = 6.7 Hz, J = 3.1 Hz), 5.10 (l.b, 2H;
NH2); 7.54–7.88 (m, 5H); 13C NMR (75 MHz, CDCl3):
d
4. Experimental section
24.56, 30.23, 49.76, 59.35, 127.87, 129.41, 133.38,
135.94,153.93, MS/EI m/z = 269 (M+.). Anal. Calcd for
4.1. Chemistry
C11H15N3O3S,269.32: C 49.06, H 5.61, N 15.60. Found: C
49.13, H 5.69, N 15.65.
TLC was performed on Merck 60F-254 silica gel plates
(layer thickness 0.25 mm). Column chromatography was
performed on silica gel (70e230 mesh) using ethylacetate
and cyclohexane mixture. CH2Cl2 was distilled over CaH2
whereas THF was distilled over sodium. Melting points
were determined on an electrothermal 9002 apparatus and
are uncorrected. NMR spectra were recorded on AC-300
Bruker apparatus (1H at 300 MHz and 13C at 75 MHz). All
4.1.4. O-2-bromopropanoyle-1-benzenesulfonylpyrrolidin-
2’-carboxamidoxime 3a
Yield = 84 %; mp: 129–131 8C; [
a
]
D = ꢀ16.3, (CHCl3,
—
N
—
c = 0.16); IR (cmꢀ1); nC
O
= 1752; nC
= 1605; 1H NMR
—
—
(300 MHz, CDCl3):
d 1.09 (d, 3H, J = 6.9 Hz), 1.71 (m, 2H),
1.99 (m, 1H), 2.21 (m, 1H), 3.19 (m, 1H); 3.71 (m, 1H); 3.98
(q, 1H, J = 7.1 Hz); 4.33 (dd, 1H, J = 8.2 Hz, J = 3.9 Hz); 5.71
(s.l, NH), 7.65 (m, 3H); 7.91 (d, 2H, J = 8.1 Hz); 13C NMR
chemical shifts are reported as
d values (ppm) relative to
internal tetramethylsilane. IR spectra were recorded on
FTS-6000 BIO-RAD apparatus. Elemental analyses were
carried out by the Service of Microanalysis of the ‘‘Institut
national de recherche et d’analyse physicochimique de
Tunis’’. HPLC analyses were conducted on a heptane/
isopropanol system with a UV detector at 254 nm, using a
chirobiotic V column (250 ꢁ 46 mm) and a 0.6 mL/min
flow rate.
(75 MHz, CDCl3),
128.48, 129.65, 134.16, 136.15, 160.18, 166.98.
d; 19.91, 23.76, 31.54, 50.14, 53.45, 59.45,
0
0
´
4.1.5. O-2 -bromo-3 -methylbutanoyle-1-
benzenesulfonylpyrrolidin-2-carboxamidoxime 3b
Yield = 86%; mp: 140–142 8C;
[
a
]
D = ꢀ14.2, (CHCl3,
—
N
—
1
c = 0.16); IR (cmꢀ1); nC
O
= 1761; nC
= 1610; H NMR
—
—
(300 MHz, CDCl3):
d 1.09 (d, 3H, J = 6.5 Hz), 1.16 (d, 3H,
J = 7.3 Hz), 1.55 (m, 1H), 1.67 (m, 1H), 1.84 (m, 1H), 2.21 (m,
2H), 3.14 (m, 1H); 3.59 (m, 1H); 4.15 (d, 1H, J = 4.3 Hz); 4.31
(dd, 1H, J = 7.9 Hz, J = 4.1 Hz); 5.84 (s.l, NH), 7.63 (m, 3H);
7.84 (d, 2H, J = 8.1 Hz); 13C NMR (75 MHz, CDCl3),
d; 20.23,
4.1.1. General procedure for the preparation of 5-alkyl-3-(10-
benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-1,2,4-
oxadiazin-6-ones
A mixture of (2S),2-alkyl-2-bromoacid (5 mmol), SOCl2
(20 mmol) and three drops of dry DMF in anhydrous
20.59, 24.95, 31.92, 32.85, 50.42, 53.68, 59.10, 128.36,
129.93, 134.11, 135.50, 160.38, 166.77.

1282
N. Tka et al. / C. R. Chimie 13 (2010) 1278–1283
0
0
´
4.1.6. O-2 -bromo-4 -methylpentanoyle-1-
benzenesulfonylpyrrolidin-2-carboxamidoxime 3c
Yield = 81%; mp:146–148 8C;
D = ꢀ12.1, (CHCl3,
4.1.10. (20S, 5R)-5-methyl-3-(10-benzenesulfonylpyrrolidin-
20-yl)-4,5-dihydro-1,2,4-oxadiazin-6-one 4a
[
a
—
]
—
Yield = 91 %; mp:112–114 8C;
[
a
]
D = ꢀ6.0, (CHCl3,
c = 0.16); IR (cmꢀ1); nC
(300 MHz, CDCl3):
1.74 (m, 2H), 2.28 (m, 1H); 2.39 (m, 1H); 3.18 (m, 1H); 3.88
(m, 1H), 4.16 (dd, 1H J = 7.5 Hz, J = 6.2 Hz), 4.32 (dd, 1H,
J = 7.9 Hz, J = 3.1 Hz); 5.71 (s.l, NH), 7.71 (m, 3H); 7.81 (d,
2H, J = 8.1 Hz); 13C NMR (75 MHz, CDCl3),
O
= 1750; nC
N
= 1604; 1H NMR
c = 0.16); d.e > 99 % [HPLC analysis using a chirobiotic V
column 4.6 mm ꢁ 250 mm under the following conditions:
heptane/isopropanol (95/5) as mobile phase, rt,
—
—
d 1.08 (d, 6H, J = 5.6 Hz), 1.51 (m, 3H),
l
( 254 nm; flow rate = 0.6 mL/min. Retention time:
22.12 min]; IR (cmꢀ1):
nNH = 3254; nC
O
= 1750; nC
—
—
—
—
d
21.18, 23.84,
N d 1.17 (d, 3H,
= 1600; 1H NMR (300 MHz, CDCl3):
25.21, 26.02, 30.54, 33.54, 50.48, 52.21, 60.44, 128.64,
129.98, 134.04, 135.61, 161.12, 166.81.
J = 7.1 Hz), 1.64 (m, 2H), 1.88 (m, 1H), 2.15 (m, 1H), 3.17
(m, 1H); 3.59 (m, 1H); 3.95 (q, 1H, J = 7.2 Hz); 4.27 (dd, 1H,
J = 8.1 Hz, J = 3.6 Hz); 7.59 (m, 3H); 7.84 (d, 2H, J = 8.1 Hz),
8.88 (s.l, NH); 13C NMR (75 MHz, CDCl3),
d 17.82, 24.56,
0
0
´
4.1.7. O-2 -bromo-3 -methylpentanoyle-1-
benzenesulfonylpyrrolidin-2-carboxamidoxime 3d
30.23, 49.76, 59.35, 79.87, 127.87, 129.41, 133.38, 135.94,
150.25, 166.48. Anal. Calcd for C14H17N3O4S, 323.37: C,
52.00, H, 5.30, N, 12.99. Found: C, 52.06, H, 5.37, N, 13.03.
Yield = 85%; mp:132–134 8C;
[
a
]
D = ꢀ17.8, (CHCl3,
0.88 (t, 3H,
c = 0.16); 1H NMR (300 MHz, CDCl3):
d
J = 4.2 Hz), 0.98 (d, 3H, J = 7.1 Hz), 1.27 (m, 1H), 1.61 (m,
1H), 1.68 (m, 2H), 1.79 (m, 1H), 2.03 (m, 1H); 2.16 (m,
1H); 3.11 (m, 1H); 3.54 (m, 1H), 4.17 (d, 1H, J = 4.1 Hz);
4.23 (dd, 1H, J = 7.9 Hz, J = 3.2 Hz); 5.66 (s.l, NH); 7.58
(m, 3H); 7.80 (d, 2H, J = 8.1 Hz); 13C NMR (75 MHz,
4.1.11. (20S, 5R)-5-isopropyl-3-(10-
benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro1,2,4-
oxadiazin-6-one 4b
Yield = 93%; mp:125–127 8C;
[
a
]
D = ꢀ16.2, (CHCl3,
CDCl3),
d
11.07, 16.72, 24.94, 26.71, 31.82, 38.87, 50.41,
c = 0.16); d.e > 99% [HPLC analysis using a chirobiotic V
column 4.6 mm ꢁ 250 mm under the following conditions:
52.32, 59.21, 128.33, 129.89, 134.06, 135.57, 160.00,
166.87.
heptane/isopropanol (95/5) as mobile phase, rt,
l ( 254 nm,
flow rate = 0.6 mL/min. Retention time: 20.44 min];
1H NMR (300 MHz, CDCl3):
1.07 (d, 3H, J = 6.7 Hz), 1.13
0
0
´
4.1.8. O-2 -bromo-3 -phenylpropanoyle-1-
d
benzenesulfonylpyrrolidin-2-carboxamidoxime 3e
(d, 3H, J = 7.1 Hz), 1.68 (m, 2H), 1.93 (m, 1H), 2.38 (m, 2H),
3.22 (m, 1H); 3.57 (m, 1H); 3.99 (d, 1H, J = 4.2 Hz); 4.27 (dd,
1H, J = 7.8 Hz, J = 4.3 Hz); 7.65 (m, 3H); 7.85 (d, 2H,
J = 8.1 Hz), 8.89 (s.l, NH); 13C NMR (75 MHz, CDCl3),
d
Yield = 79%; mp:154–156 8C;
[
a
]
D = ꢀ21.6, (CHCl3,
—
N
—
c = 0.16); IR (cmꢀ1): nC
O
= 1751; nC
= 1611; 1H NMR
—
—
(300 MHz, CDCl3):
d
1.51 (m, 1H), 1.65 (m, 1H), 1.82
(m, 1H), 2.15 (m, 1H), 3.12 (m, 2H), 3.33 (m, 1H), 3.58 (m,
1H); 4.13 (m, 1H); 4.49 (m, 1H); 5.16 (s.l, NH),7.21 (m,
5H); 7.52 (m, 3H), 7.82 (m, 2H, J = 8.1 Hz); 13C NMR
17.51, 18.82, 24.51, 27.43, 29.44, 49.82, 58.58, 80.78,
127.88, 129.61, 133.80, 135.40, 150.81, 166.09. Anal. Calcd
for C16H21N3O4S,351.43: C, 54.69, H, 6.02, N, 11.96. Found:
C, 54.74, H, 6.09, N, 12.03.
(75 MHz, CDCl3),
d 24.89, 31.72, 41.67, 50.38, 57.07,
59.22, 127.80, 128.31, 129.09, 129.14, 134.01, 135.64,
136.24, 159.70, 166.82.
4.1.12. (20S, 5R)-5-isobutyl-3-(10-benzenesulfonylpyrrolidin-
20-yl)-4,5-dihydro-1,2,4-oxadiazin-6-one 4c
0
0
´
4.1.9. O-2 -bromo-3 -methylbutanoyle-1-
Yield = 90%; mp: 107–109 8C;
[
a
]
D = ꢀ22.8, (CHCl3,
benzenesulfonylpyrrolidin-2-carboxamidoxime 3f
c = 0.16); d.e > 99 % [HPLC analysis using a chirobiotic V
column 4.6 mm ꢁ 250 mm under the following conditions:
Yield = 87%; 1H NMR (300 MHz, CDCl3):
d 1.04 (m, 6H),
1.62 (m, 1H), 1.83 (m, 2H), 2.19 (m, 2H), 3.08 (m, 1H);
3.66 (m, 1H); 4.11, 4.16 (d, J = 5.6 Hz, d, J = 5.2 Hz, 1H);
4.29 (m, 1H); 5.86 (s, 2H, NH2),7.58 (m, 3H); 7.84 (d, 2H,
heptane/isopropanol (95/5) as mobile phase, rt, l ( 254 nm,
flow rate = 0.6 mL/min. Retention time: 25.09 min]; IR
(cmꢀ1):
n
NH = 3256; nC
O
= 1754; nC
N
= 1607; 1H NMR
—
—
—
—
J = 8.1 Hz); 13C NMR (75 MHz, CDCl3),
d
20.25 (20.33),
(300 MHz, CDCl3): d 0.93 (d, 6H, J = 5.8 Hz), 1.48 (m, 3H),
20.84, 24.52, 31.45 (31.76), 32.24 (32.37), 50.83, 52.57,
60.56 (60.88), 128.89, 129.62, 134.81, 135.58, 160.76,
166.15 (166.23).
1.67 (m, 2H), 2.15 (m, 1H); 2.33 (m, 1H); 3.14 (m, 1H); 3.46
(m, 1H), 4.04 (dd, 1H J = 7.7 Hz, J = 6.0 Hz), 4.25 (dd, 1H,
J = 7.9 Hz, J = 3.2 Hz); 7.63 (m, 3H); 7.83 (d, 2H, J = 8.1 Hz),
After purification, the products 3a–f were dissolved in
dry THF and added slowly to a solution of 1 equivalent of
NaH in dry THF cooled to ꢀ25 8C. The reaction mixture was
stirred for 3 h. After warming to room temperature, 10 mL
of saturated ammonium chloride solution and 50 mL of
chloroform were added. After separation, the aqueous
layer was extracted with chloroform (3 ꢁ 50 mL). The
chloroformic combined extracts were dried over MgSO4,
filtered and concentrated in vacuo and the residue
obtained was purified by column chromatography on
silica gel (20% ethylacetate: 80 cyclohexane) to give the 5-
alkyl-3-(10-benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-
1,2,4-oxadiazin-6-ones, which was recrystallized from
(20% ethylacetate: 80% hexane).
8.78 (s.l, NH); 13C NMR (75 MHz, CDCl3),
d 18.28, 19.84,
24.31, 25.02, 29.66, 33.45, 51.08, 59.15, 80.44, 128.83,
130.25, 133.84, 135.61, 151.15, 166.77.
4.1.13. (20S, 5R)-5-Sec-butyl-3-(10-
benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-1,2,4-
oxadiazin-6-one 4d
Yield = 92%; mp:102–104 8C;
[
a
]
D = ꢀ25.7, (CHCl3,
c = 0.16); d.e > 99% [HPLC analysis using a chirobiotic V
column 4.6 mm ꢁ 250 mm under the following conditions:
heptane/isopropanol (95/5) as mobile phase, rt,
l ( 254 nm,
flow rate = 0.6 mL/min. Retention time: 26.03 min]; IR
(cmꢀ1):
(300 MHz, CDCl3):
n
NH = 3252; nC
O
—
= 1749; nC
N
= 1596; 1H NMR
—
—
—
d
0.86 (t, 3H, J = 4.8 Hz), 0.96 (d, 3H,

N. Tka et al. / C. R. Chimie 13 (2010) 1278–1283
1283
J = 6.9 Hz), 1.38 (m, 1H), 1.57 (m, 3H), 1.86 (m, 1H), 2.12 (m,
1H); 2.28 (m, 1H); 3.16 (m, 1H); 3.50 (m, 1H), 4.00 (d, 1H,
J = 4.2 Hz); 4.21 (dd, 1H, J = 8.1 Hz, J = 3.4 Hz); 7.58 (m, 3H);
7.80 (d, 2H, J = 8.1 Hz), 8.75 (s.l, NH); 13C NMR (75 MHz,
initially prepared at the concentration of 1 mg mLꢀ1 was
added into the first well. Then, 100 L from their serial
dilutions was transferred into nine consecutive wells. The
last well containing 100 L of nutrient broth without
compound and 5 L of the inoculum on each strip was
m
m
CDCl3),
d
12.08, 14.43, 24.89, 26.02, 29.46, 34.33, 50.16,
m
58.95, 79.41, 128.23, 129.95, 134.14, 135.81, 151.06,
166.80. Anal. Calcd for C17H23N3O4S,365.45: C, 55.87, H,
6.34, N, 11.50. Found: C, 55.92, H, 6.40, N, 11.55.
used as negative control.
The plate was covered with a sterile plate sealer and
then incubated for 18 h at 37 8C. The MIC was defined as
the lowest concentration of the compound 4a–e to inhibit
the growth of microorganisms, after incubation. The
results were expressed in milligram per milliliter.
Each test was carried out in triplicate. The microorgan-
isms tested in this study were provided from European
Hospital of George-Pompidou (HEGP) (France).
4.1.14. (20S, 5R)-5-benzyl-3-(10-benzenesulfonylpyrrolidin-
20-yl)-4,5-dihydro-1,2,4-oxadiazin-6-one 4e
Yield = 77%; mp:120–122 8C;
[
a
]
D = ꢀ31.6, (CHCl3,
c = 0.16); d.e = 97% [HPLC analysis using a chirobiotic V
column 4.6 mm ꢁ 250 mm under the following conditions:
heptane/isopropanol (95/5) as mobile phase, rt,
l ( 254 nm,
flow rate = 0.6 mL/min. Retention time: 28.59 min]; IR
(cmꢀ1):
(300 MHz, CDCl3):
n
NH = 3250; nC
O
—
= 1753; nC
N
= 1601; 1H NMR
—
—
Acknowledgements
—
d
1.64 (m, 2H), 1.92 (m, 1H), 2.30 (m,
The authors would like to thank Hayet Edziri from
University of Monastir (laboratoire des maladies trans-
1H), 3.19 (m, 2H), 3.34 (m, 1H), 3.56 (m, 1H); 4.25 (m, 1H);
4.37 (m, 1H); 7.32 (m, 5H); 7.59 (m, 3H), 7.75 (m, 2H,
J = 8.1 Hz), 8.84 (s.l, NH); 13C NMR (75 MHz, CDCl3),
d24.83,
´
missibles et des substances biologiquement actives, faculte
de pharmacie, Monastir, Tunisia) for making biological
29.30, 34.64, 50.17, 58.92, 78.52, 127.21, 128.24, 128.84,
130.01, 134.16, 135.71, 136.75, 151.37, 166.40. Anal. Calcd
for C20H21N3O4S,399.47: C, 60.14, H, 5.30, N, 10.52. Found:
C, 60.18, H, 5.34, N, 10.58.
´
´
activities and the DGRSRT (Direction generale de la
´
recherche scientifique et de la renovation technologique)
of the Tunisian Ministry of Higher Education, Scientific
Research and Technology for the financial support of this
research.
4.1.15. (20S, 5R) and (20S, 5S)-5-isopropyl-3-(10-
benzenesulfonylpyrrolidin-20-yl)-4,5-dihydro-1,2,4-
oxadiazin-6-one 4f
References
Yield = 89%; two diastereomers (20S,5R) and (20S,5S)
[HPLC analysis using
4.6 mm ꢁ 250 mm under the following conditions: hep-
tane/isopropanol (95/5) as mobile phase, rt, ( 254 nm,
flow rate = 0.6 mL/min. Retention times: 20.44 min and
23.89 min]; 1H NMR (300 MHz, CDCl3):
1.07 (m, 6H), 1.72
(m, 2H), 1.93 (m, 1H), 2.36 (m, 2H), 3.23 (m, 1H); 3.56 (m,
1H); 3.95, 3.99 (d, J = 5.7 Hz, d, J = 5.1 Hz, 1H); 4.28 (m, 1H);
7.66 (m, 3H); 7.87 (d, 2H, J = 8.1 Hz), 8.87 (s.l, NH); 13C NMR
a
chirobiotic
V
column
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each well 100
m
L of nutrient broth and 5
mL of the
inoculum. A 200
m
L stock solution of the compound